TRIBOLIUM INFORMATION BULLETIN
NUMBER 10 - March, 1967
Foreword
v
Editor’s Random Notes
vi
Announcements
x
Stock Lists
1
New Mutants
65
Notes - Research
Quantitative differences between the “aureate” and normal
Phenotypes in T. castaneum, Mary Ackermann
86
Acarophenax tribolii – a parasitoid of Tribolium,
William W. Allen
87
Further simplification of a bioassay method with adults
Of T. confusum Duv., B. Berck
88
Sex differences in recombination values for linkage
Group V of T. castaneum, A. A. Dewees
89
Population performance of single – and mixed – species
of flour beetles, H. E. Erdman
91
Ontogeny and X-radiation sensitivity of the flour beetle,
Tribolium castaneum, mutant – sooty, H. E. Erdman
91
Metamorphic and adult life span modifications of flour
Beetles Tribolium confusum, X-irradiated as pupae, H.E.Erdan
92
Comparative studies with Tribolium. III. The productivity
of T. castaneum and T. confusum on synthetic diets,
I.R. Franklin, J. Chandra and A. Sokoloff
93
Egg-laying rate of virgin females of Tribolium castaneum
at different temperatures, Ma. C. Fuentes and R.G. Ruano
96
Algae flour as nutrition for Tribolium confusum, P. Geisert
101
The use of vital dyes for marking Tribolium eggs in fresh
And aged flour, Frank K. Ho
103
Identification of eye mutants in Tribolium larvae,
Frank K. Ho
105
Susceptibility of Sitophilus granaries to moderate cold,
R. W. Howe and B. D. Hole
107
Quinoid secretions in Tribolium confusum,
R. K. Ladisch and S. K. Ladisch
108
Sexual dimorphism in the pupal setae of Tribolium,
E. L. Lange
111
Regression of the sex ratio on maternal grandfather’s age,
I.M. Lerner and N. Inouye
113
Evaluation of an anti-feeding compound as a protectant
Against stored-products insects, S. R. Loschiavo
115
The distribution of Oryzaephilus Mercator Fauvel in
Canada, S. R. Loschiavo and L.B. Smith
115
Factors involved in the survival of Tribolium confusum
Populations, D. J. McDonald and Mrs. L. Stoner
116
Cytochemical studies of Sitophilus granarius mycetomes,
A.J. Musgrave and S. B. Singh
119
Mycetomes as possible obligate symbiotes of Sitophilus,
A.J. Musgrave and I. Grinyer
119
A quantitative approach to the feeding of Hymenolepis
Diminuta eggs to the flour beetle, Tribolium confusum,
D.M. O’Brian, A.Levine and F.F. Katz
120
The behavior and biology of Flour beetles, genus
Tribolium, as studied in laboratory gradients of
Temperature and humidity, A.K. Onyearu
121
The scanning electron microscope, R. F. W. Pease,
T.L. Hayes and A. Sokoloff
122
The response of larvae of Trogoderma granarium Everts to
-100 C, Elisabeth M. Reynolds and Barbara M. Rundle
131
Allelism of “mottled” (mt) and “melanotic stink glands” (msg)
In Tribolium castaneum, T.H. Schmitz and D.C. Englert
132
The chromosome numbers of some stored product
Coleoptera, D. D. Shaw
133
Sex chromosome variation in D. maculates and
D. frischii, D. D. Shaw
134
The effects of synchrony of egg batches on fitness
Characters and competition in Tribolium castaneum,
R. R. Sokal
135
A comparison of fitness characters and their responses to
Density in stock and selected cultures of wild type and black
Tribolium castaneum, R. R. Sokal
142
Preliminary population studies with mutants of Tribolium
Castaneum Herbst. I. The paddle gene, A. Sokoloff
147
Preliminary population studies with mutants of Tribolium
Castaneum Herbst. II. The black gene, A. Sokoloff
149
Preliminary population studies with mutats of Tribolium
Castaneum Herbst. III. The jet gene, A. Sokoloff
152
Preliminary population studies with mutants of
Tribolium castaneum Herbst. IV. The pearl gene,
A.Sokoloff
153
Preliminary population studies with mutants of Latheticus
Oryzae Waterh. I. The pearl gene, A. Sokoloff
161
Additions to established linkage groups, A. Sokoloff,
M. Ackermann and B. Heinze
169
Additional sex-linked lethals in Tribolium castaneum Herbst,
A.Sokolof, N. Inouye and R.S. St. Hilaire
169
Further studies of productivity of Tribolium cstaneum and
Tribolium confusum in homo- and heterospecific matings,
A.Sokoloff and J. Lanier
173
Influence of collecting frequency on egg-laying rate of fecundated
And virgin females in Tribolium castaneum,
Ma. P. Tagarro and F. Orozco
175
Sex pheromones and defensive secretions from Tenebrionid
Beetles, W. Tschinkel
179
Some observations on mating frequencies in Tribolium
Castaneum strains, D. Wool
182
Selection for 13-day larval size in Tribolium under two
Nutritional levels, Y. Yamada and A. E. Bell
186
Relative fitness of selected strains under different
Environments, Y. Yamada and A. E. Bell
189
A method for isolating C. turcicus to obtain virgin females,
M. Ackermann
191
A method for enhancing propagation of poorly viable flour
beetles, J. Lauck
191
Technique to determine the contents of cocoons of Cryptolestes
Turcicus, J. T. Robertson and L.B. Smith
192
New method for weighing Tribolium pupae, W. Rumball
192
A method for rearing Eleodes (longicollis?) (Coleoptera:
Tenebrionidae) in the laboratory, A. Sokoloff
194
Directory – Geographical
196
Directory – Alphabetical
219
NEW MUTANTS
A. Dermestes maculates (Dermestidae)
*REPORT OF D. D. SHAW
1. Deformed antennae (def). A mutant reported in TIB 9:58, now proved to be
controlled by an autosomal recessive gene of-variable penetrance and giving
apparently normal viability. The mutant is similar in appearance to the
phenoldeviant “branched” in Tribolium castaneum. Linkage tests with “rufous”
and “fuscous” are being carried out.
Deformed antenna
Normal
2. Fuscous (fu) (Mali). This mutant was isolated from a culture from Mali and is
phenotypically similar to fuscous already described in TIB 8:42. Tests of
allelism are under way.
3. Muddy. An eye-color mutant isolated from a culture of Indian origin. The eyes
appear grey-brown compared with the normal brown black. Genetics not yet
studied.
4. Sexpitles. Originated in a single male offspring from a “rufous” x “white tip”
cross. The male has no sex-pit. Genetics not yet studied.
5. White tip (Wt). This mutant was described in TIB 9 and has proved to be
controlled by a dominant gene giving normal viability. The mutant is
phenotypically similar to that described by Philip (1940) which was controlled by
an autosomal recessive gene. Linkage tests with rufous and fuscous are being
carried out.
B. Tribolium castaneum (Tenebrionidae)
*REPORT OF CONSTANTINO, DITTMAR, HANCOCK, LANGE AND
SHIDELER.
1. Corn oil sensitive (cos) – Costantino, 1964. An autosomal recessive gene
identified during an investigation of the genetic basis of selection response for
small 13-day larval weight in a population of Tribolium castaneum. A preliminary
report on this gene was given by Constantino, Bell, and Rogler (Nature 210:221,
1966. The properties of the cos gene are as follows:
a. Larval growth of the cos homozygote, as measured by 13-day larval weight,
varies inversely to the concentration of corn oil in the diet.
b. Viability also varies inversely to the concentration of corn oil in the diet (83%
viable, at 13 days, in the diet with no supplemental corn oil decreasing to
51% in the diet with 5% corn oil).
c. The degree of dominance, with respect to 13-day larval weight when grown
in a diet with 5% corn oil, is essentially complete.
d. .Gas-liquid chromatographic analyses of normal (+/+) and sensitive
(cos/cos) individuals grown on diets with and without supplemental corn oil
suggest that excessive quantities of linoleic acid in the cos homozygote may
be the basis for the observed sensitivity to corn oil.
Linkage studies and further biochemical analyses are presently underway.
2. Grossly deformed (gdf) – Lange, 1965. Autosomal recessive of good expression
isolated from Purdue Foundation. The pupae have the following appearance:
a. Elytra shortened and barely extending to the abdomen.
b. Tibia is narrower in width and curved in a manner similar to bowleg.
c. Urogomphi are reduced in size.
d. Genital lobes are absent.
e. Eyes are similar to squint or bar eye.
f. Club of the antennae is fused.
g. Gin traps are reduced in size.
h. Setae are reduced to a large but variable amount.
The adults have the following appearance:
a.
b.
c.
d.
e.
Elytra shortened and cover about half of the abdomen.
Flight wings normal.
Tibia has a form similar to bowleg.
Eyes appear similar to squint but some facets are present.
Antennal club is fused with a variable number of stalk segments, with generally the
first five segments of the stalk unfused.
The homozygous recessive is lethal in the adult stage probably because of
desiccation brought about by the reduced elytra. It is not allelic to squint and is not
linked to Bar eye. Further linkage studies are in progress.
3. Bumpy (by) – Hancock. Autosomal recessive found in sub-population of Purdue +
Foundation, has variable expression and incomplete penetrance, similar in
phenotype to Sokoloff’s ble, yields 3% recombination with Sa. Bumpy is being
forwarded to Sokoloff’s laboratory for allelism tests with ble.
4. Miniature appendaged (maD) – Dittmar. Spontaneous sex-linked recessive found
in a multiple recombination study, allelic to Sokoloff’s ma and similar in expression.
5. Multi-urogomphi (Mu) – Dittmar. Autosomal dominant found in Purdue b
Foundation, incomplete penetrance with expression varied giving one or two
additional urogomphi in larvae and pupae. Preliminary recombination studies for
Mu suggest independence of linkage groups 2, 3, 4, 5, 7 and 8. Similar in
expression to eu described by Lasley and Sokoloff in TIB 3 and has been
forwarded to that laboratory for further comparisons.
6. Reduced elytra (re) – Shideler. Autosomal recessive of reduced viability found in
sub-population of the Purdue black Foundation, incomplete penetrance and
variable expression, elytra are severely reduced and sometimes missing.
7. split-back (sb) – Shideler. Autosomal recessive found in a sub=population of the
Purdue perarl Foundation, has expression varying from shortened elytra to severe
splitting or diverging of elytra. Similar in phenotype to the description of short
elytra (Ho and Dawson, shH & D, Sokoloff, shS, TIB 1962) and has been
forwarded to them for tests of allelism.
*REPORT OF R.W. LEMON AND D. G. BLACKMAN
1. mahogany (my). Autosomal recessive (TIB 9:59), not allelic with black or sooty.
Linked with sooty and Bar eye in linkage Group IV (cross over values of 43% and
21.5% respectively). Results of three point crosses not yet known.
2. Pearl-like (TIB 9:59). Linkage tests for groups III, IV, V, VII and X proved
negative. Crosses with ptl in group IX were inconclusive due to incomplete
penetrance of ptl. The tests with Mo in group VI showed possible linkage, but
difficulty in classifying Mo beetles made it necessary to repeat the tests which are
not yet concluded. However, a very recent cross between “pearl-like” and “ivory”
produced an F1 all mutant beetles; it is therefore possible that “pearl-like” is a
reoccurrence of “ivory.”.
*REPORT OF A. SOKOLOFF
1. Charcoal (Chr) – Ackermann, 1966. Found during the course of determining
whether “prothoracic margin irregular,” an abnormality involving the anterior margin
of the prothorax was heritable (it was probably a phenodeviant). Autosomal
dominant with recessive lethal effects. Charcoal beetles resemble tawny and
sooty. No synthetic lethal effect has been observed with Spa, Be, Fta and Bamp.
2. diminutive appendages (dim) – Heinze, 1966. Spontaneous in a stock of Bamp.
An autosomal recessive with good penetrance and fairly good viability. The elytra
are very short, exposing two or three abdominal segmens. The membranous
wings fail to fold and protrude beyond the elytral tips. Many adults display a split of
the abbreviated elytra, beginning about one-third of the length of the elytra from the
scutellum. The legs are also much shorter than normal but maintain proper
proportions as in miniature appendaged (see figure below). The mutant can
readily be identified in the pupa stage by the short elytra which do not completely
cover the membranous wings (as in the normal) and the legs are noticeably
shorter. A more careful examination reveals fusions of antenna- and tarsomeres.
These are summarized in Tables 1 and 2. This mutant is not allelic with ov or aa..
Table 1.
Antennal fusions in dim. (15 beetles of each sex scored)
Males
0
4-5
6-7
6-7-8
7-8
7-8-9
8-9
8-11
10-11
Females_____
Right
Left
Right
Left
11
4
----1
---
4
3
-1
--1
---
5
4
1
-2
-4
---
7
2
--1
1
3
1
1
Table 2.
Number of tarsal segments involved in fusions or missing in Dim.
(15 beetles of each sex scored)
Pair of Legs
1st pair
2nd pair
3rd pair
0 1 2 3 4
0 1 2 3 4
0 1 2 3
Females Right
Left
5 1 7 1 1
5 3 4 2 1
7 4 2 3 5 4 4 1 -
5 3 7 5 6 3 1
Males
8 0 6 2 8 2 3 1 1
6 1 7 1 11 3 2 - -
8 2 5 8 1 5 1
Right
Left
3. fused antennal segments-3b (fas-3b) – Heinze, 1966. Spontaneous, in a
population of black and pearl scored for these genes for many generations in
studies of natural selection. (Originally called “aggregate antennae” but found to
be allelic with fas-3, therefore renamed.) Incompletely recessive, with fusions of
antennameres as shown in the accompanying table. It may be noted that male
antennae exhibit more fusions of the club. Female antennae show more fusions
involving both funicular and club segments. Also, male antennae exhibit more
fusions which involve fewer segments, while the fusions in female antennae are
more extensive (involving more segments).
4. Knobby prothorax (knp) – Sokoloff, 1966. Spontaneous in a stock of Mo mas.
Possibly a phenodeviant, but more likely autosomal recessive of incomplete
penetrance. Penetrance is greater when knp females rather than males are used
in backcrosses, and greater in backcrosses in general than in F1 x F1 crosses
(expression in the latter being very weak) characterized by the fact that the
prothorax has more or less symmetrical “knobs” extending lateroposteriorly. These
are as large as ¼ - 1/3 the length of the prothorax. The prothoracic margin
continues around the knobs, but many of these have a “suture” more or less
continuous with the posterior like the prothorax but the ventral part may be
sclerotized. This mutation is similar to knp in T. confusum. (TIB 9:62, see
illustration on front cover, TIB 9).
5. Overhung split (ohs) – Heinze, 1966. Common in au stocks, but does not appear
to be linked to au. Its appearance is like that shown in the figure below, with the
elytra longer than the abdomen and exhibiting a pronounced split and sometimes
tips curling down. Some may extend beyond the abdomen without splitting. Not
allelic with we, spl, te, dve, bal, or cye. Probably a recessive with poor penetrance.
Table 3.
Segments involved in fusions in the fas-3b mutant
(50 males and 50 females scored)
Males
Right
0
4-5
4-6
4-8
4-10
4-11
5-6
5-7
5-8
5-9
5-11
6-7
6-8
6-9
6-10
6-11
7-8
7-9
7-11
8-9
8-11
9-10
9-11
10-11
9
-----3
-1
1
2
1
3
1
-5
3
1
2
5
-1
4
13
Females
Left
7
--1
-1
4
-1
-3
5
-2
-4
3
-3
2
1
3
7
13
Right
7
1
1
--1
3
1
--3
1
3
1
-6
2
1
5
4
2
1
4
11
Left
6
1
1
-1
3
2
1
--1
5
2
-1
8
2
-3
4
2
3
6
5
6. Prothoracic margin irregular (pmi) – Hoy, 1965. Spontaneous in a population
started two years previously with 12 Kr irradiated male mated with anon-irradiated
female. The anterior ventral margin of the prothorax appears as in the figure on
page 77. A phenodeviant. From a cross of two pmi beetles only 2/578 were pmi.
7. Red-2 (r-2) – Heinze, 1966. One adult was found in a test cross between au s
and Be which had chestnut-colored eyes; this male was mated to produce
offspring with pale eyes as young beetles, and darker red eyes as older adults. An
allelism test with red showed the mutation to be its allele; mode of inheritance tests
showed it to also be an x-linked recessive gene, as is red.
8. Reduced tarsi and antennae (rta) – Heinze, 1966. In a “scalloped prothorax” stock
(initially derived from an irradiation experiment which had been performed several
years before) were found 10 adults and two pupae with antennae and legs reduced
in segment number.
Table 4 shows the antennameres, Table 5 the tarsomeres involved in fusions and Table 6
the total number of tarsomeres seen in those legs with no apparent fusions but with 0-5
tarsomeres.
It is evident that the modal number of fusions is in antennameres 4-5, but also there
appears to be a sex-difference, males having the club segments affected while that in
females does not exhibit fusions. The tarsomeres were reduced to 1-2 per leg in most
cases, some legs having no tarsomeres at all. The mutation is readily identifiable in the
pupa stage, the legs appearing noticeably shorter than normal, as in Fta (see Plate 5, H,
I, J in Sokoloff, 1966). Preliminary crosses suggest an autosomal recessive mode of
inheritance.
Table 4.
Antennal fusions found in a sample of “reduced tarsi and antennae” (rta) in T.
castaneum (N = 21 females and 21 males)
Males
Right
0
2-3
3-5
4-5
5-6
6-7
7-8
9-10
10-11
11
--8
-1
-4
1
Females
Left
Right
Left
12
--6
2
-1
2
1
11
--9
1
-----
13
1
1
5
--1
---
Table 5.
Tarsomeres involved in fusions in “reduced tarsi and antennae” (rta)
In T. castaneum (N = 21 females and 21 males)
Pair of Legs
1st Pair
2nd pair
3rd pair
0
1
0
1
0
1
Males
Right
Left
20
16
1
5
19
21
2
--
21
20
-1
Females
Right
Left
20
21
1
--
20
21
1
--
21
20
-1
Table 6.
Total number of segments observed in the tarsi of “reduced tarsi
And antennae” (rta) in T. castaneum (N = 21 beetles of each sex)
Leg and Tarsus Number___________________________
1
0 1 2 3 4
Male
5
2
0 1 2 3 4
5
3
0 1 2 3
4
Right
Left
3 4 1 0 1 12
4 3 0 1 0 3
-
2 3 6 1 -
- 16
- 14
1 1 4 - 15
- 4 3 - 14
Female Right
Left
2 4 - - 3 12
3 3 1 - - 14
-
2 1 - - 18
3 1 - 1 16
1 7 2 1 10
2 8 2 - 9
9. Rugose elytra (rue) – Heinze, 1966. A spontaneous mutation commonly seen in
populations derived from aureate. The adult has very finely corrugated elytra,
giving them an almost granular appearance. Mode of inheritance tests show that
rue can be separated from au but the expression is a little less remarkable in nonaureate adults. The rue mutation is an autosomal recessive with good penetrance
and viability, and not allelic to rough.
10. Short antenna-3 (sa-3) – Heinze and Hoy, 1966. In a cye fas stock were found
several adults and pupae with very short and stubby legs, as well as short
antennae and elytra. In allelism tests with aa-1 47/102 adults were seen to have
weakly affected legs; with stock aa ov, 15/233 adults had very weakly affected
legs; with stock sa/sa all adults had strongly shortened appendages. The new
mutant is, therefore, considered to be allelic with sa and will be called sa-3. A
mode of inheritance test shows sa-3 to be an autosomal, incompletely recessive
with 100% penetrance and high viability.
11. Short median abdominal projection (smp) – Ackermann, 1966. Autosomal
recessive of apparently uniform expression and complete penetrance. A mutation
somewhat resembling but not as pronounced as Bamp (Blunt abdominal and
metathoracic projection, Hoy, 1965). (See TIB 9:60 for illustration of Bamp). It
may be most readily identified by a shorter median abdominal projection of the
apparent first abdominal segment, with the result that the tip does not extend
between the hind coxae and is not accommodated by the inverted-V-shaped
metasternellum of the metathorax (see figure on p.77). The mesosternellum and
the intercoxal process of the metathorax barely join each other between
themesothoracic legs. The mesosternellum appears somewhat ventrally convex.
In crosses of smp x
the F1 x F1 gave 187+:63 smp F2 and the backcross
196+:123 smp. In the reciprocal cross the ratios were 257+:58 smp for F2 and the
backcross 109+:59 smp. Also noted was a large number of matings which failed
and failed again, indicating a possible sterility factor.
12. Truncated elytra-2 (te-2) – Heinze, 1966. In a cross between r Mr x sy. Wild type
several male adults and pupae were seen with short, folded te-like elytra. A good
stock was built up and an allelism test with te showed the new mutant to be allelic.
Since the new te has very strong expression and seems to be healthier than the
old, a new stock shall be maintained.
Sex-linked Lethals
All the sex-linked lethal described below were found in the Berkeley synthetic strain in
experiments designed to test the Cavalli-Sforza’s model of mutation frequency as a
regression on the age of the maternal grandfather (see note by Lerner and Inouye
elsewhere in this issue of TIB).
1. Lethal-5 (1-5_ -- Sokoloff and Inouye, 1964. Located about 25 units to the left of
py (away from r) (see note by Sokoloff, Inouye and St. Hilaire). Allelic with 12 and
14.
2. Lethal-6 (1-6) – Sokoloff and Inouye, 1964. Located about 12 units to the left of
py (away from r).
3. Lethal-7 (1-7) – Sokoloff and Inouye, 1964. Located about 20 units to the left of
py (away from r).
Figure Legend
Upper Row, Center. – The “prothoracic margin irregular” phenodeviant in T.castaneum.
Note that the probasisternum is indented on both sides between the
Midline and the tergosternal suture.
Upper Right. -- The “incomplete metathoracic projections” mutant in T.confusum.
Note the posterior projections of the metathorax are short, failing to form an
inverted V to accommodate the medial abdominal projection of the first
apparent abdominal segment.
Lower Left – The prosternumless (psl) mutation in T.confusum. Both the probasisTernum and mesobasisternum sternella are reduced. The prosternellum
Forms a small ridge and as a result of its reduction in size the coxae of the
Forelegs are freer, and not as widely separated. The mesosternellum fails
to fuse with the intercoxal process of the metabasisternum, again freeing the
coxae of the middle pair of legs and the mesobasisternum acquired a suturelike appearance (represented by stipples).
Lower Middle. – incomplete meso (basi) sternum (ims). Illustration of the mutant in
T. castaneum described in TIB 6:25 and in T. confusum in TIB 9:68.
Lower Right. – “short median abdominal projection” mutant in T. castaneum. Note
that the “point” of the first apparent abdominal segment is blunt.
C. Tribolium confusum
*REPORT OF A. SOKOLOFF
Autosomal
1. Black (b) – Ho and Ackermann, 1966. Spontaneous in a mixed population
XXIII-12 of T. anaphe and T. confusum. Not allelic with ebony nor with ebony2. A recessive allele of black.
2. Engraved meta (basi) sternum dots (ems dt) – Heinze, 1966. Spontaneous.
Found in a “frosted” (allele of dpe) stock. In adults, characterized by a very
small indentation or hole just above the middle of one or both antecoxal pieces
(see sketch) but not connected to the latter as in jac, jagged antecoxal piece
(see Plate 7F in Sokoloff, 1966). There may be 1-3 dots per basisternum, and
not necessarily the same number of dots on each side. Allelic to ems, and like
ems autosomal recessive with incomplete penetrance.
3. Legless (lgl) – Hoy and Sokoloff, 1966. Spontaneous in a Reduced eye stock,
five males and three females were found with a body color resembling
mahogany (much lighter than Park’s ebony), and both forelegs missing down
to the coxa. Outcrossed to wild type and scoring F2 no or few beetles have
legs missing or partly so. But, mated to ech other, they produce progeny
without one or both prothoracic legs. The elytra in all beetles are rugose, they
may be a little shorter than the abdomen in a few beetles, and the proximal
third of the elytra may fail to meet at the midline. Those beetles with both legs
missing, taken out of the flour, are often seen moving with the lower part of the
head on the glass container, being pushed as the beetles attempt to walk.
Others can hold the body off the container as they walk. Tentatively this
condition has been designated as a phenodeviant because beetles with the
legs absent may not be seen at all in F2. However, since there is some
evidence that the legs may develop and turn white, it is possible that this may
be an aging effect young beetles exhibiting normal legs which, a few weeks
later, fall off. Attempts are therefore being made to work out the mode of
inheritance by examining body color and/or the wrinkled condition of the elytra.
4. Pointed elytra-like (pel) – Sokoloff, 1963. Elytra mostly divergent and pointed,
somewhat shorter than normal resembling dve and/or pointed elytra of T.
castaneum. (See Plate 9, i, j in Sokoloff, 1966). Occasionally, there may be
beetles with elytra only short (not divergent) resembling sh in T. castaneum.
Another form of expression: elytra longer than abdomen and drooping at the
sides. Autosomal recessive, good viability.
5. Prosternumless (psl) – Sokoloff, Ackermann and Daly, 1966. Autosomal,
incompletely recessive (in reciprocal crosses of psl x + about 10 per cent of the
F1 beetles showed the character). (Found during tests of allelism between sep
and dep.) This mutation is characterized by a reduction of the sternellum of
the prothorax and mestothorax to a vestigial, keel-like structure (see figure on
p.77). As a result, the globose coxae of the fore- and middle-legs lie ventral to
the prosternum and mesosternum (“above” as one examines the ventral
aspect of the beetle as it lies on its back). Since the mesobasisternum fails to
reach the intercoxal process of the metasternum, the mesobasisternum
appears to “sink in” above the coxae (i.e., it is directed postero-dorsally). The
mesobasisternum acquires a shiny ridge along the midline resembling a
suture. The probasisternum also may develop a less pronounced (and not
shiny) suture along the midline. In some beetles the tergosternal suture
becomes less pronounced.
6. Stunted (stt) – Sokoloff and Lauck, 1966. Spontaneous. Autosomal recessive
mutation found in some crosses involving pointed-elytra-like (pel, q.v.). A gene
with pleiotropic effects producing, like the sex-linked ma, miniature
appendaged, individuals with the whole body shorter than normal, but the
effect being most pronounced on the appendages as follows:
a. Antennael Thesre is considerabke, often asymmetrical, fustion of the
antennameres. The antennae appear elbowed, and because of fusion and
reduction in size of segments 7-8 the antennae may appear clubbed resembling
that in T. castaneum. The distribution of fusions in the two sexes is shown in Table
1 which also indicates that the antennameres most often fused are 7 and 8. It is
also evident that the gene often fails to express itself on the side of the body,
producing one antenna normal and another abnormal. In the affected antennae
there may be as few as two and as many as eight antennameres in fustions, some
of which may involve more than two antennameres. When four consecutive
segments are fused, usually they will form a kidney-shaped structure pointing the
other way. Summarized in Table 2 are the numbers of antennameres which may
be affected in one antenna. Table 3 shows how funicular and club segments or
both may be affected.
b. Legs. All the podomeres are reduced (the femur, in particular, becoming
globase), resulting in a beetle with all legs considerably shorter (in a manner
similar to that illustrated for ma in T. castaneum in Sokoloff, 1960, Can. J. Genet.
Cytol 2:28-33). The tarsomeres are affected, the distribution of fusions being as
shown in Table 4. Table 5, summarizing the fusions in the antennae and the tarsi
of this mutant, appears to indicate that the chief difference between the two sexes
is in the tarsi, the males exhibiting a greater number of fusions than females.
c.
Mouth parts. Although this point was not investigated thoroughly there seems to
be a partial fusion at least of some maxillary palpomeres.
d. Elytra and membranous wings. These are considerably reduced: the elytra are
about half as long as the abdomen, often divergent, and often they exhibit a blister
on the lateral margin. The veins and striae on the caudal half of the elytron
become less pronounced and often hardly visible. The membranous wings trail
(the tips of the wings fail to fold under the proximal portion), but they extend
beyond the elytra, almost as far as the tip of the abdomen. All the veins appear to
be present, but the cells are considerably reduced.
e. Other parts of the body. The prosternal process and the post-coxal bridges are
reduced, with the result that a fairly extensive area of the prosternum in front of
the mesobasisternum appears unsclerotized. The lateral parts of the coxae
sockets appear to be more open than in the normal, so that at least the coxae of
the fore- and hind-legs have a greater freedom to rotate within them. The
intercoxal process of the apparent first abdominal segment appears somewhat
shorter so it does not fit as well between the metasternellal. The tergal aspect of
the abdomen is, of course, exposed. This circumstance, as well as the fact that
the hind-legs are too short to enable the beetle to free its hind-end and the forelegs are too short to free
Table 1.
Distribution of fused antennameres in the T. confusum mutant
“stunted” (N = 20 males and 20 females)
Segments
Involved
1-4
3-4
3-5
3-8
4-5
Males
Right
Left
-3
1
-2
-1
1
-1
Females
Right
Left
-2
2
1
2
1
-1
1
1
4-8
4-11
5-6
5-7
5-8
5-11
6-7
6-8
6-11
7-8
7-9
7-11
8-9
8-11
9-10
9-11
10-11
2
1
4
3
1
1
2
1
-11
----5
1
2
1
-5
---1
3
-9
2
---3
1
1
-1
3
1
1
1
2
1
1
8
--2
-2
-1
---1
--2
2
-10
1
1
2
1
4
-1
Table 2.
Numbers of segments involved in antennal fusions of “stunted”
(N = 20 males and 20 females)
N
Segments
Involved
2
3
4
5
6
7
8
Males
Right
Left
32
4
1
-2
1
--
21
8
1
1
1
-1
Females
Right
Left
20
4
2
2
-1
--
22
4
1
1
----
Table 3.
Part of antennae where fusions were found in “stunted”
(N = 20 males and 20 Females)
Males
Funicle
Club
Both
Right
Left
30
8
1
25
6
2
Females
Right
Left
20
3
5
21
4
5
Table 4.
Distribution of fused (or missing) tarsomeres in the T. confusum
Mutant “stunted” (N = 20 males and 20 females)
Leg
Pair
1
2
3
N
Tarsomeres
in Fusion
or Missing
Right
Left
0
1
2
3
4
5
1
2
15
3
---
-5
12
2
1
--
1
11
3
3
2
--
4
-6
6
3
1
--
0
1
2
3
4
5
1
5
10
3
---
2
6
13
1
---
6
6
7
-1
1
9
5
5
1
---
0
1
2
3
4
-7
13
2
--
-7
18
1
--
4
4
13
-1
2
1
18
---
Males
Females
Right
Left
Table 5.
Number of fused segments in antennae and tarsi of male and female
stunted T. confusum (N = 20 males and 20 females)
Antennae
Sex
Males
Side
Total Ave. No. Segments
Fusions
of
involved
N
Fusions
N
Tarsi
Ave. of
Segments Ave. of
Segments/ Fused
Segments/
Fusions
N
Fusions
R
L
36
33
1.8
1.6
97
75
4.8
3.7
114
120
5.7
6.0
Females R
L
42
28
1.4
2.1
79
79
3.9
3.9
97
86
4.8
4.3
the head from the pupal skin, results in a great mortality in the imago attempts to eclose
from the pupa. It is quite probable that because of such major changes in the
appendages and the overall size reduction of the body that this mutant might be
recognized in the larval stage.
It is clear that both in the case of antennamere and tarsomere fusions (or deletions) occur
more frequently in females than in males. In both sexes the first three segments appear
to escape fusion. In males the modal group appears to be in the fusion of segments 4-5;
in females these segments as well as segments 8-9 appear equally susceptible to fusion.
Also, if club segments are involved, they appear to fuse more often in females than in
males. The females showed somewhat more fusions of tarsomeres than did the males.
7. Wide split-1 (wspl-1) – Lauck, 1966. A mutation involving the leytra was found
in the legless mutation. It appears as the figure on Plate 11 of Sokoloff, 1966,
and is allelic with a similar mutation found, but apparently not reported, by P.S.
Dawson.
Sex-linked
1. Alate prothorax (apt) – Strong, 1966. Originally found in a T. confusum synthetic
strain maintained in soy plus yeast for many generations. This recessive, sexlinked, homeotic mutant is apparent in both pupae and adults. Both show large
elytra-like growths on the dorso-lateral aspect of the prothorax. Although
somewhat distorted, these growths show many of the characteristics of elytra (see
illustration on front cover). The normal expression is similar to the strongest
expression found in T. castaneum, apt, and rarely is as weak as the normal
expression of T. castaneum, apt (i.e., small knobs on the dorso-lateral aspect of
the prothorax). The expression is fairly constant and the penetrance appears to be
complete. The viability, about 75%, is surprisingly high for such a drastic mutation.
Most deaths take place during the molt from the pupa to the adult stage. Linkage
data with eslt and lp are available (see note elsewhere inTIB) and experiments with
red eye are in progress.
Tests of Allelism
1. Black (b) – Ho and Ackermann, 1966. Black (b) in T. confusum has proved to be
a recessive allele of the semidominant black. It has been redesignated as b-4, Ho
and Ackermann, 1966.
2. Frosted, reported in T. confusum (TIB 8:62) is allelic with dirty pearl eye (dpe) and
it is therefore renamed dpe-1, Hoy and Heinze, 1966.
Addenda and Corrigenda
Incomplete metathoracic projections (imp) (TIB 8:52) in T. castaneum is an
autosomal recessive with slightly variable expression and incomplete penetrance (about
15% of individuals fail to show the character).
ACKERMANN, MARY
Department of Genetics
University of California
Berkeley, California
*Quantitative differences between the “aureate” and Normal phenotypes in T. castaneum
The “aureate” mutation produces beetles whose bodies appear more pubescent
than the wild type. Counts of body hairs were made on typical adult beetles shosen at
random from the au and normal stocks. Counts were made through a compound
microscope at 150X magnification for sternites and 660X magnification for wings, using
an ocular reticle calibrated for a square approximately 0.1 mm to a side for the sternite,
and 0.024 mm for the wing counts. One of the membranous wings and the abdomen of
ten beetles of each type had been temporarily mounted in paraffin oil. One square area
from the central portion of each of the five visible sternites, and five contiguous areas
along the main axis of the wing (near the distal tip to avoid veins) were chosen, and in
each the number of bristles or hairs was determined.
The results are shown in Table 1. From the data in the table it can be seen that,
although the difference is not significant for the membranous wings, it is significant for
each of the abdominal sternites, the number of bristles being increased two- or threefold
in the mutant.
Table 1.
Numbers of setae in normal and aureate abdominal
Sternites and wings in T. castaneum (N = 10)
Apparent abdominal
sternite
Normal
aureate_____
1
2
3
4
5
9.4 + 0.2
10.7 + 0.4
9.7 + 0.4
10.6 + 0.3
9.9 + 0.4
30.6 + 1.4
27.4 + 1.7
28.3 + 1.5
27.4 + 1.1
23.7 + 2.1
Total
10.6 + 0.17
27.25 + .69
10.84 + 2.36
11.10 + 1.66
Membranous wings
Thus, from our observations, the aureate gene seems to affect only the sclerotized
portions of the exoskeleton. Scanning electron micrographs (see research note
elsewhere in this bulletin) show that there is a definite increase of cervical setae (at the
anterior margin of the prothorax, particularly at the gular border) and on the antennae.
Most remarkable is the fact that the interommatidial bristles, which are single in normal
beetles, are doubled under the influence of au.
This work was supported by USPHS grant GM-08942.
ALLEN, WILLIAM W.
Division of Entomology and Acarology
University of California
Berkeley, California
*Acarophenax tribolii – a parasitoid of Tribolium
In 1964 small mites were found infesting some of the Tribolium cultures at
Berkeley. Since a general decline in the vigor of the cultures was associated with these
infestations, it was necessary that some remedial action be taken. A review of the
literature revealed that these mites, belonging to the family Pymotidae, had been
described by Newstead and Duvall (1918) as Acarophenax tribolii. In addition to the
taxonomic description they gave a generalized account of the life history and habits of
these mites. More recently Cross (1965) reviewed the family Pymotidae and summarized
what was known about the life history of Acarophenax tribolii by referring back to the
paper by Newstead and Duvall (1918).
The adult mites are found attached to the bodies of both adult and larval beetles.
Commonly 10 to 12 adult mites may be found on the bodies of the beetles. There is
some question whether the mites feed on the adults and larvae of Tribolium, or whether
the relationship is merely phoretic.
After several days the adult mites drop from the beetles and seek out a Tribolium
egg. The mites attach themselves to the eggs, and in a day or two suck out the contents.
During feeding there is marked enlargement of the body of the female (physogastry), and
all power of locomotion is lost. Complete development of the immature mites takes place
within the body of their mother. There is good evidence that mating takes place before
the adult females leave the body of the mother. Since development takes only three or
four days and commonly there may be 12 to 14 progeny per mother, it is understandable
why infestations can build up rapidly.
Since establishment of new Tribolium cultures from small numbers of adults
observed to be free of mites was unsuccessful in eliminating infestations, it was obvious
that more drastic action would be necessary to clean up the cultures. A number of tests
were conducted to determine if certain chemicals could be found that would control the
mites without having an adverse effect on the beetles. These tests revealed that there
was selectivity in the insecticide endosulfan (Thiodan) and the acaricide Morestan when
used in the flour at a rate of 300 ppm of the actual material. Because Morestan generally
is regarded to be of lower toxicity to insects it was selected for use in cleaning up the
cultures.
In actual practice it was found that Morestan did have a detrimental effect on the
beetles particularly when they were held at higher temperatures. This difference can be
explained by the fact that the initial tests were run with adults and mature larvae rather
than the early larval instars. Nevertheless, it was possible to use Morestan to help free
the cultures of the mites. This was done by placing beetles in flour treated with 300 ppm
Morestan and holding them at a temperature of 200C. After 10 days the beetles were
removed from the treated flour and used to establish new cultures in clean flour. Through
use of this treatment and greater care with the cultures it has been possible to hold the
mites to a very low level.
Literature Cited
Cross, A. E. 1965. The generic relationships of the family Pymotidae (Acarina:
Trombidiformes). University of Kansas Science Bull. 45 (2):275 pp.
Newstead, R. and H. M. Duvall. 1918. Bionomic, morphological, and economic report on
the acarids of stored grain and flour. Royal Soc. Rep. of Grain Pests (War) Committee,
2:19-25.
BERCK, B.
Canada Department of Agriculture
Winnipeg 19, Manitoba, Canada
Further simplification of a bioassay method with Adults of T. confusum Duv.
A simplified bioassay method used by the author for screening and validation tests
with fumigant gases was described in TIB 9 (2). The method employed 100 cc. all-glass
syringes (1) as combined test cages and micro fumigation chambers.
In the current work with phosphine (3), it was found that not all the syringes that
had been tested were uniformly gas-tight at all positions of the syringe piston. In a search
for alternative leak-proof test chambers, 125 ml. Erlenmeyer flasks and 4 oz. medicine
bottles fitted with gas-tight stoppers and septums were much cheaper, more durable and
easier to clean and use than the syringes. They were used in a simplified bioassay
method for the simultaneous determination of LD50 and LT50 values (4). Changes in
CO2-02-N2 relationships in these miniaturized test chambers are readily monitored. The
main advantages are that a wise range of gas concentrations and temperatures can be
tested in a relatively small space, and insect mortalities can be readily assessed over a
wide time span. Thus, it was possible to determine the LT50 of 0.002 mg. phosphine per
litre air during a 14-day test with adults of T. confusum Duv.
Literature Cited
1. Berck, B. 1965. Sorption of ethylene dbromide, ethylene dichloride and carbon
tetrachloride by cereal products. J. Agr. Food Chem. 13:248-254.
2. Berck, B. 1966. Simplified bioassay method with adults of T. confusum Duv.
Tribolium Information Bull. 9:74-75.
3. Berck, B. 1967. Paper in preparation.
4. Berck, B. 1967. Paper in preparation.
DEWEES, A. A.
Population Genetics Institute
Purdue University
Lafayette, Indiana
*Sex differences in recombination values for Linkage group V of T. castaneum
In a study to establish the linkage relationships among the eye color mutants, ruby
(rb) and maroon (m), and the jet (j) body color mutant of linkage group V in T. castaneum,
it was noted that the recombination values were not equal for the two sexes. However,
Lasley (1960, TIB, p. 14) reported evidence for equality of recombination in both sexes
between jet and split in the same linkage group.
The phenomenon of unequal crossing over in reciprocal crosses was first pointed
out by Sokoloff (1963, TIB 6; 1964) for markers of the seventh linkage group of T.
castaneum (male rates higher than female rates) and was later studied by Johnson
(1966) with reference to linkage groups IV and VII.
Preliminary tests (Dewees, 1965, TIB 8:72) indicated that rb was located about 27
units to the left of j near m. Due to the difficulties in classifying adult maroon beetles, a
regular three-point linkage test of m, rb, and j ws not undertaken. However, a modified
two-point linkage test of m and rb in the trans phase was set up with the jet gene included
as a marker on the ruby chromosome. The progeny were scored in the late pupal stage
as either wild type or mutant with regard to eye color and the recombinants (wild type)
were then test mated to rb j/ rb j beetles to determine the most probable sequence of the
three loci. From 87 single pair matings of m rb +/m rb + x + rb j/m + + , 60 wild type
recombinants were recovered from a total of 8,085 progeny. In the reciprocal cross from
96 matings only eight out of 13,246 progeny had wild type eyes. The calculated
recombination rates and their corresponding 95% fiducial limits are shown below.
Heterozygous
Parent
Recombination rate
between m and rb
95% fiducial limit
Male
Female
.0148
.0012
( .0113, . 0191)
( .0005, .0024)
Clearly, these rates are statistically different. The gene sequence was determined
to be m – rb – j.
To further investigate this sex difference, regula two-point linkage tests of rb and j
were conducted. Both cis and trans phases were examined in reciprocal crosses. Within
each sex the recombination rates observed for the cis and trans phases were not
significantly different at the 1% probability level, therefore the results presented below
represent pooled estimates.
Heterozygous
Parent
Male
Female
No. of
matings
19
20
Total No. of
progeny
Recombination rates
between rb and j + s.e.
1974
3196
.3283 + .0212
.2118 + .0145
Here also it can be seen that the recombination rate in the males is significantly
greater than the rate observed in females. Whether or not these two instances represent
a general phenomenon for the fifth linkage group is not known, however further tests
involving markers of this linkage group are being planned.
As for the possible mechanism responsible for sex differences in recombination
rates, Sokoloff (1964) discusses evidence from other organisms that seems to indicate
that such a phenomenon could result when the two sexes differ in the distribution of a
single chiasma. However, an equally plausible explanation might be one in which there is
a differential segregation of chromosomes in one sex, probably the female, such that the
recombinant chromosome is more likely to be incorporated in the polar body during
second division segregation thus lowering the number of recombinant progeny produced.
This type of mechanism could very well account for the several cases in which male
recombination rates are observed to be higher than female rates.
ERDMAN, H. E.
Biology Department
Battelle-Northwest
Richland, Washington 99352
Population performance of single—and mixed—species of flour beetles
Knowledge on the interaction(s) among environment, population structure and
inter-species competition is beeded to understand better species survival and population
performance. Single—andmixed—species population of flour beetles were cultured at
250C, 290C and 350C and censused monthly for eight months prior to chronic gamma
irradiation. Results of the radiation will be reported later.
Tribolium confusum Duval (“Chicago Standard”) and Tribolium castaneum Herbst (sooty)
were sexed as pupae and held until sexually mature. Fifteen replicates of single species
populations (begun with 20 pairs) and of mixed-species populations (begun with 10 pairs
of both species) were censused as to numbers of larvae, pupae and adults. All live forms
and eggs were returned to new food. Eggs were not counted and adults were counted as
to species because of their color difference.
Population stability was achieved only at the high temperature. Adults’ forms were most
abundant and showed least variability, probably due to their longer life relative to the
other life cycle stages.
Fitness, measured on total number of life forms, indicated that Tribolium confusum was
better adapted when not in competition at 290C; whereas Tribolium castaneum was
better adapted at the temperature extremes. In mixed-species population, Tribolium
castaneum was progressively less fit than Tribolium confusum. Numbers of total life
forms per population replicate after eight months were greatest at 250C and least at
320C, indicating the temperature effect on population performance.
ERDMAN, H. E.
Biology Department
Battelle-Northwest
Richland, Washington 99352
Ontogeny and X-radiation sensitivity of the flour beetle, Tribolium castaneum, mutant :
sooty
Nonlinearity of radiation sensitivity during the life cycle reflects the differential
sensitivities of dividing cells compared to nondividing cells and the degree of
differentiation. This report defines the radiation sensitivities of the flour beetle, Tribolium
castaneum (mutant: sooty) throughout its life cycle for specific stages which show
developmental changes in somatic and gonadic tissues.
Approximately 100 individuals at each of nine different stages during the life cycle
were X-rayed. X-rays were delivered at 1 kR/min by a generator operated at 30 ma, 250
kvp, 0.25 mm cooper + 1.0 mm aluminum filtration, and a 2.5 inch target-subject distance.
Culture conditions were 30 + 10C and 70 + 10 per cent relative humidity.
Virgin adults were crossed after radiation. An exposure was considered as
sterilizing if no progeny developed with 2 – 3 months and as lethal if individuals failed to
mature or died within 3 – 4 weeks post eclosion. Exposures were performed at 1000 R
increments, or in some cases, at 500 R increments.
The weakest link in the life cyle was the 1 – 3 hour old egg in which nuclei are
rapidly dividing and undifferentiated.
The differential radiosensitivity between somatic cells and gonad anlagen diverged
markedly during post-larval stages because:
(1) Germ cells remained at a radiosensitive stage in cell division, and
(2) The increased differentiation of somatic tissue required greater exposures for
lethality.
ERDMAN, H. E.
Biology Department
Battelle-Northwest
Richland, Washington 99352
Metamorphic and adult life span modifications of flour beetles. Tribolium confusum, Xirradiated as pupae
Adult life span in mammals and insects was decreased when immature stages
were irradiated supposedly by induced mutations in somatic cells. When pupae are
radiated one would expect any lethal effect on genetic material would be eliminated
during metamorphosis into the adult, instead of occurring after the adult stage
successfully developed. This investigation determined whether metamorphosis and
length of life could be altered when flour beetle pupae were X-irradiated.
T. confusum Duval (“Chicago Standard” stock) pupae, age 25-26 days, were sexed
and irradiated. The X-ray unit operated at 250 kv, 0.86 mm copper half value layer, 0.25
mm copper and 1.0 mm aluminum filtration. The anode was 2.5 in. from the subject and
the apparatus delivered approximately 1 kr/min. Doses were 0, 1, 2.5, 5, 7.5, 10, 20, 30,
40, 50, 60 and 70 kr. Each of the five replicates at eah dose contained ten pupae of each
sex in 10 g food. Deaths were noted twice weekly. Culture conditions were 29 + 10 C
and 60-75 per cent relative humidity.
Lethality data for males and females were combined, because sex made no
difference in flour beetles.
Emergence of adults from pupae given 1 or 2.5 kR was comparable to that of
control. Metamorphic incompletion increased progressively for doses from 5 to 70 kR, but
metamorphosis was not delayed. Partial metamorphosis occurred in some individuals
given 5 kR and greater doses—that is, thorax and head were adult but the elytra,
although pigmented, was pupal in size and position and the abdomen retained pupal
characteristics. Some individuals given 30 kR and greater were unable to cast off their
meconium. Death probably resulted from intestinal obstruction and not from starvation as
heavily radiated adults ingest.
Reduction in length of life of adults radiated as pupae was a sensitive indication of
radiation damage. Mortality increased progressively with increased exposure.
Exact cause(s) of death remain unknown because nuclear effects were difficult to
differentiate from cytoplasmic effects. Death was probably due to chemical alteration of
the cytoplasm because pupae developed adult structures. Antimetabolites could result
from radiation-induced alterations in the chemistry of pigmentation. Other biochemical
processes continue from juvenile stages to the adult and interruption of such processes
could result in adverse effects on metamorphosis and adult life span when pupae are
irradiated.
FRANKLIN, I. R., J. CHANDRA AND A. SOKOLOFF
Department of Genetics
University of California
Berkeley, California 94720
Comparative studies with Tribolium. III. The productivity of T. castaneum and T.
confusum on synthetic diet.
A synthetic medium described by Naylor (1964) will allow development of
T.castaneum and T. confusum, but has been shown to be a poor diet, in terms of
productivity, compared to “natural” diets, such as wheat and corn flours (Sokoloff, et al.,
1966b). However, the productivity of both species is raised approximately tenfold in
Naylor”s medium to which brewer’s yeast has been added, and is comparable to
productivity in commonly used diets, such as wheat + yeast and corn+ yeast. Since yeast
supplements are generally considered to be an important source of vitamins, it was
considered desirable to examine the effect on productivity of varying the vitamin content
of the Naylor medium. The vitamin mixture described by Naylor contains varying
amounts of micotinic acid, thiamine, pantothenate, riboflavin, pyridoxine, carnitine, folic
acid and biotin, and the level of this mixture is 134 ug/fram of medium. Five media were
tested containing different concentrations of vitamin mixture, and for comparison a sixth
medium containing 5% brewer’s yeast was used. Fifteen females and 10 males of each
species were transferred eah day for four days into five grams of the test diets, and the
adult progeny were counted during the following two months. Each species was
replicated three times, and the average productivity replicate for the two species and the
six diets as shown in Table 1.
Table 1.
Supplement to Naylor’s medium vitamin mixture ug/gram diet
Species
0
50
100
200
400
5%
Yeast
T. castaneum
T. confusum
1
-
39
86
70
91
47
102
40
79
605
391
While it is apparent that the vitamin mixture is necessary for development in each
species, there is little difference between the four diets in which the vitamin supplement
was added. The differences in productivity between the two species is comparable with
the conclusions of Sokoloff, et al. (1966b) that T. confusum performs better than T.
castaneum on “poor” diets, and worse on “good” diets.
It was suggested by Sokoloff, et al. (1966a) that the two species T. castaneum and
T.confusum may differ in their requirements for certain amino acids. Various
combinations of eight amino acids, including those suggested by the above authors, were
added to the Naylor medium and the productivity of both species was measured. The
following concentrations of finely ground crystalline amino acies were used (Table 2).
Table 2.
Amino acid concentrations used in enriching the Naylor medium
Amino acid
Concentration
alanine
2.5 mg/gram medium
aspartic acid
3.5
“
“
“
cystine
1.5
“
“
“
glycine
4.0
“
“
“
leucine
4.5
“
“
“
methionine
1.0
“
“
“
threonine
2.0
“
“
“
tyrosine
3.0
“
“
“
As before, 15 females and 10 males were transferred four times into five grams of
fresh medium, and the total number of adults emerging in the subsequent two months
were counted. Thirty-two diets were used, representing a 1l8 replicate of a 28 factorial
design. None of the main effects were significant for either species, as illustrated by the
analysis of variance table (Table 3).
Table 3.
Analysis of variance
Source
Main effects
Two factor interactions
Residual
Mean Square
CF
d.f.
CS
8
20
3
31
197.8 n.s.
203.9 n.s.
61.8
586.0 n.s.
1,096.1 n.s.
218.2
These experiments show that the vitamins and amino acids which were tested are
not limiting the productivity of either species in Naylor’s medium, and the poor
performance of T. castaeum and T. confusum is due to absence of other factors which
are present in brewer’s yeast.
This investigation was supported by USPHS grant GM-08942.
Literature Cited
Naylor, A.F. 1964. Possible value of casein, gluten, egg albumin or fibrin as whole
proteins in the diet of two strains of the flour beetle Tribolium confusum (Tenebrionidae).
Can. J. Zool. 42:1-9.
Sokoloff, A., I. R. Franklin, L. F. Overton and F. K. Ho. 1966a. Comparative studies with
Tribolium I. Productivity of T. castaneum and T. confusum in several commercially
available diets. J. Stored Prod. Res. 1:295-311.
Sokoloff, A., I. R. Franklin and R. K. Lakhanpal. 1966b. Comparative studies with
Tribolium II. Productivity of T. castaneum and T. confusum in natural, semisynthetic and
synthetic diets. J. Stored Prod. Res. 1:313-324.
FUENTES, Ma. C. AND R.G. RUANO
Laboratorio de Genetica de Poblaciones
Instituto Nacional de Investigaciones Agronomicas
Madrid, Spain
*Egg-laying rate of virgin females of Tribolium castaneum at different temperatures
To start quantitative genetics studies in connection with the genotype x
environment interation, some tests were run to determine the variation of the egg-laying
rate in virgin females of Tribolium castaneum when they are exposed to different
temperatures.
From some previous studies testing the egg-laying at 280 C, 320 C and 360 C during a
four day period comprised between the seventh and the eleventh days after adult
emergence, we observed that both 280 C and 360 C were not as favorable environments
as 320 C when egg-laying is the character studied.
We ran two experiments testing egg-laying during consecutive periods of 48 hours.
The first test started at 240 C going two by two degrees up to 420C and afterwards going
in the same way down to 240C again. The second one started at 420C going down to
240C and upto 420C, also increasing or decreasing by two degrees in each 48 hour
period. Temperatures greater than 420C begin to be lethal.
At the same time some controls were maintained at 330 C in both tests to evaluate
changes due to microenvironment ariations through the testing periods, other
uncontrolled effects and to take into account the decreasing effect of the females age
after the first days of a greater lay. Such controls were females of the same age as the
experimental ones.
Enough pupae from the “Consejo” strain of Tribolium castaneum were sexed to
have 200 females emerging on the same day, at which time they were placed in individual
vials. During the larval and pupal stage and until day 7 after adult emergence, they were
maintained at a temperature of 330C and a humidity of 70% RH; humidity was not
changed in any case during the experiments. At day 7 after emergence fresh medium
was put in each vial and at that time 100 females were shifted to 240C conditions and 100
remained at 330 C. At day 9 eggs were collected by suction-sifting and counted, and the
experimental females were changed to 260 C, leaving the controls at 330 C. The same
procedure was followed every two days going up with the experimental temperatures until
day 25, when experimental females were subjected to 420 C conditions. At day 27 and
after egg collection, they were shifted to 400 C again and so on going down arriving at
day 43 when they were placed in the 240 C conditions. The test finished at day 45 after
egg collection and count.
The second test was run in the same way but starting with the 100 experimental
females at 420 C and going down to 240 C and up to 420 C.
Table 1 gives for both tests the average egg-laying figures obtained.from the
experimental and ontrol females at specified temperatures. Deviations from controls and
corrected figures with the average for all the controls in both tests are also given. Table 2
includes the figures averaged for every temperature disregarding any possible influence
or effects of the previous temperatures.
Table 1
Mean Number of eggs produced by T. castaneum at different
temperatures (oC) in consecutive period of 48 hours
Table 2.
Figures for each temperature, averaging data in Table 1
Testing
Temperature
0 C
m
24
26
28
30
32
34
36
5.71
7.84
8.68
10.85
12.80
14.73
12.91
Control
at 330 C
m
13.21
12.79
11.50
13.57
12.73
12.34
13.06
Deviation
from
control
- 7,50
- 4.95
- 2.82
- 2.72
+ 0.07
+ 2.39
- 0.15
Data
adjusted to
controls*_______
5.13
7.68
9.81
9.91
12.70
15.02
12.48
38
40
42
10.17
5.52
1.58
12.53
12.98
11.43
- 2.36
- 7,46
- 9.85
10.27
5.17
2.78
*Standard error for corrected figures: + 0.92
In order to see the results more clearly, the corrected figures for both tests are plotted in
gaph 1. Graph 2 presents four second order para-bolas adjusted to the points of graph 1
by the least squares method. Equations of such parabolas are:
Test 1, increasing temperatures: y = - 0.18 + 0.25 x -- 0.36 x2
Test 1, decreasing temperatures: y = 0.60 + 0.16 x – 0.51 x2
Test 2, increasing temperatures: y = 1.45 + 0.36 x – 0.62 x2
Test 2, decreasing temperatures: y = 0.70 + 0.58 x – 0.44 x2
Using the corrected figures it is concluded:
(a) In our strain, temperatures around 33 or 340 C are the optimum for egg-laying.
Temperatures above or below those seem to be worse; the greater the distance
from the optimum the smaller is the average egg-laying rate.
(b) Response to the same temperature does not seem to be the same when
progressive temperatures are increasing or decreasing, having gotten an average
of 8.90 and 9.29 eggs respectively. Test 1 (starting at 240C) has an average
figure of 9.04 and test 2 (starting at 420 C) 9.15 eggs. In any case these
differences do not appear to be significant, nor is the interaction of those possible
effects. We reach the tentative conclusion when we see both data and graphics,
that in general laying is greater with decreasing temperatures.
(c) It is interesting to note that even though the differences are not significant we are
tempted to conclude that after the females have been submitted to a very high
temperature the laying rate afterwards is larger, because the combination of test 2
(starting at 420 C) and decreasing temperatures (coming from 420 C) has the best
effect. This could be explained by assuming that at 420 C the females experience
greater stress retaining eggs which, after some days, they lay in excess.
GEISERT, P.
Oak Park River Forest High School
Oak Park, Illinois
Algae flour as nutrition for Tribolium confusum
Algae can be cultivated in heavy water resulting in fully deuterated species. In an
attempt to determine the effect of deuteration on higher organisms I planned to use
deuterated algae as nutrition for Tribolium confusum. As a preliminary study
Scenedesmu obliquus grown in water culture was processed into a flour and fed to
Tribolium to determine the feasibility of using this algae as deuterated nutrition for the
beetle.
The preliminary survey used 10 adults/vial containing two grams of algae flour
prepared by desiccating and grinding water grown algae. The amount of materials used
was limited by the ability to produce the algae. Various concentrations of the algae flour
were prepared using flour media to dilute the algae.
The populations were surveyed with the following results (see Table 1).
Algae as presently processed would not make a good “carrier” of deuterium for
studies with Tribolium. The flour beetle should be able to support itself in a high
concentration of algae before an evaluation of the effect of deuterium is attempted. A
more carefully controlled evaluation of the effect of algae on Tribolium is presently being
planned, and modifications are being made in the preparation of the algae flour.
I feel bacterial toxins were probably present in the previous algal preparations and
future techniques will attempt to separate the algae from other microorganisms and their
metabolic products.
Table 1
Algal concentration_______________________
Date
100%
60%
50%
40%
30%
20%
15%
10%
0%
ADULTS
5-4
5-12
5-19
6-2
6-14
7-5
10
8
1
0
0
0
10
9
8
7
6
4
10
10
8
6
5
5
10
10
9
8
8
5
10
10
9
8
8
7
10
10
10
10
9
13
10
10
9
9
7
11
10
10
10
10
10
11
10
9
8
7
8
9
LARVAE
5-4
5-12
5-19
6-2
6-14
7-5
0
0
0
0
0
0
0
0
0
0
0
5
0
0
0
0
1
0
0
0
1
4
1
10
0
0
0
3
2
12
0
0
0
5
11
8
0
0
0
6
7
6
0
0
0
6
8
14
0
0
2
15
6
10
EGGS
5-4
5-12
5-19
6-2
0
0
0
0
0
0
0
0
0
0
3
0
0
8
20
25
0
16
4
0
37
30
32
0
83
52
56
0
45
36
21
0
93
80
13
PUPAE
Total count of pupae from 5-4 to 7-5
1
2
5
6
12
0
0
0
0
HO, FRANK K.
Department of Genetics
University of California
Berkeley, California 94720
*The use of vital dyes for marking Tribolium eggs In fresh and aged flour
The development of a technique for marking eggs with dyed flour (Rich 1956) has
facilitated the measurement of oviposition and cannibalism rates in a population. This
technique has been used advantageously by Sonleitner (1961), and Ho and Dawson
(1966). In the studies of Rich, and Ho and Dawson, 0.5 per cent neutral red dye
concentration was used. Sonleitner preferred to use only0.2 to 0.3 per cent dye
concentration. For coloring the eggs, commercial lour is considered the most ready-touse choice. If whole wheat flour is used, however, it should be sifted through a number 5
bolting cloth sieve. This insures the eggs to be thoroughly coated with the fine-colored
flour particles. To prepare the flour, dye should be mixed and ground into the flour, then
sifted through the number 5 bolting cloth sieve and stored in a high humidity condition for
at least a week before using. Marked eggs are obtained by allowing beetles to oviposit in
the dyed medium. Eggs are removed from such egg farms using the number 2 or
number 5 bolting cloth sieves.
In some population studies, more than two or three strains or species of eggs may
be used in an experiment. The dyes for marking egg, other than the neutral red
mentioned may be needed. For this purpose, the safety range of Bismarck brown y,
methylene blue and methyl green in Tribolium studies has been observed. All these dyes
are well-known stains which have been used in histology and bacteriology for many
years.
In addition, the aged-colored flour and the fresh-colored flour were tested for a
nutritive comparison. The fresh-colored flour in this study was prepared and stored in the
incubator at 290 C, 60% R.H. with a cheese cloth cover over the bottles one week before
using; the aged-colored flour was prepared and stored in the same manner, after which
the cheese cloth was replaced with an air tight cover and stored in room condition for
more than three years. T. cstaneum and T. confusum synthetic strain beetles, two weeks
old, were used as the parents of the egg farms. In the colored flour testing series, five
jars included one control and four colored flour for each species. After three days, 500
eggs were collected from each jar; 100 eggs were placed in each vial containing eight
grams of colored or white flour. A total of 25 vials for each species were used. The
cultures were introduced into an incubator maintained at 29 0 C and 60% relative humidity.
The adults were censused 45 days later. These experiments were carried out in the
same manner for both fresh and aged flour. The results are summarized in Table 1.
It is evident that no appreciable effect is produced when Tribolium are reared in the
colored flour from egg through adult stages. The beetle emerging rate from five replicates
of each colored flour was satisfactory. It is very interesting to know that flour aged more
than three years can be used almost as effectively as the fresh-colored flour for both
species.
Table 1
Per cent of Tribolium adults emerged from 500 eggs in fresh colored,
aged colored and commercial white flour (five replicates for each test)
Neutral red
Bismarck brown y
Methyl green
Methylene blue
Control
T. castaneum
Fresh
Aged
Flour
flour
T. confusum
Fresh
Aged
flour
flour
81.00
86.60
93.20
74.40
79.00
85.00
83.60
82.00
84.00
78.60
62.20
85.00
81.00
80.50
82.10
82.00
80.00
67.50
76.00
80.50
It should be pointed out that the emerging rate of beetles in vials containing neutral
red in the aged flour for T. castaneum, methyl green in the aged flour for T. confusum,
methylene blue in the fresh flour for T. castaneum, and methylene blue in the aged flour
for T. confusum, was slightly lower than the emerging rate of the controls.
The results indicate that the vital dyes used are not poisonous to Tribolium when
being used for egg coloring at a 0.5 per cent or less concentration in the flour.
The observation also suggests that all the dyes used in this study give brightly
colored, easily distinguishable eggs.
This work was supported in part by USPHS grant GM-08942.
Literature Cited
Ho, F. K. and P. S. Dawson. 1966. Egg cannibalism by Tribolium larvae.
Ecology 47:318-322.
Rich, E. R. 1956. Egg cannibalism and fecundity in Tribolium. Ecology
37:109-120.
Sonleitner, F. J. 1961. Factors affecting egg cannibalism and fecundity in populations
of adult Tribolium castaneum. Herbst. Physiol. Zool. 34:233-255.
HO, FRANK K.
Department of Genetics
University of California
Berkeley, California 94720
*Identification of eye mutants in Tribolium larvae
The previous studies (Good, 1936 and Ho, 1960) indicated that the arrangement of
the ommatidia (facets), the shape and size of the compound eyes, and the distance
between eyes in the pupal and the adult stages can be used to identify the species of
Tribolium castaneum and T. confusum. In the larval stage no compound eyes have
developed. However, the setal map of the larva has been used advantageously for their
species identification (Ho, 1967 in press).
Tribolium has been used for genetic studies only recently. At present, over 150
mutants are described, including 11 eye mutants in T. castaneum and nine in T.
confusum (Sokoloff, 1966). To distinguish the wild type and eye mutant larvae,
examination of their stemmatal characteristics has been considered a practical technique.
In general, three types of stemmata (eyespot) are present in Tribolium larvae. In wild
type and in some mutants with deforemed eyes (microcephalic, and Bar eye) the
stemmata appear black; in the eye mutants red, pink, eyespot, light eyespot, ruby spot
and dirty pearl eye of T. confusum, they are brown. There are no visible stemmata in
pearl, platinum eye, Microphthalmic, frosted, and chestnut eye mutants, probably
because the lack of pigment renders these stemmatal structures invisible or because the
stemmata are too small or absent. In the squint (sq) beetle no ommatidia form, with the
result that the ocular diaphragm, unaffected by sq, forms in the pupa and lies exposed in
its normal position (Sokoloff and Dawson, 1963: Sokolof, 1966). The sq gene apparently
suppresses the formation of the stemmata as well, since these structures are not visible
in the larva. In microcephalic adults, the eyes are variably reduced and they bear very
few facets. In some cases, the eye on the side may be absent. If these beetles are
selected, they produce produce progeny with both eyes missing (Sokoloff, 1966).
Therefore sometimes, in a microcephalic stock, three types of larvae may be found:
those with stemmata missing on either side, those with stemmata missing on both sides,
and those with stemmata present on both sides. If the stemma is present, it is always
black. The color of the stemmata in larvae usually corresponds with the color of the
compound eyes of the resulting pupae and adults. The data are summarized in Table 1.
The stemmata are located near the antennae from the first instar through the early
part of the last larval instar. They are paired and more or less fused, and are located on
each side of the head. In the later part of the last larval instar, the stemmata migrate
along the head margin to a position near the vertex, and gradually become lighter, finally
disappearing when the larva nears pupation (Ho, 1961). In these last larval stages the
mutant may not be distinguishable from wild type.
This work was supported in part by USPHS grant GM-08942.
Table 1.
The type of stemmata of Tribolium larvae
Species
T. castaneum
Wild type or
eye mutants
wild type
Bar
Red
Pink
Pearl
Ivory
Squint
Microphthalmic
Chestnut
platinum eye
microcephalic
glas
Symbol
+
Be
r
pPK
P
i
sq
Mo
c
pte
mc
gl
Type of
stemmata
black
black
brown
Invisible
invisible
invisible
invisible
invisible
invisible
invisible
usually invisible;
if present, black
undetermined
(specimen not available)
T. confusum
wild type
red
pink
eyespot
light eyespot
ruby spot
dirty pearl eye
pearl
frosted
ruby
+
r
pK
es
es lt
rus
dpe
P
fro
rby
black
brown
brown
brown
brown
brown
brown
invisible
invisible
undetermined
(specimen not available)
Literature Cited
Good, N. E. 1936. The flour beetles of the genus Tribolium. U. S. Dept. Agric.
Tech. Bull. 498.
Ho, F. K. 1960. Discrimination between the pupae of Tribolium confusum.
Duv. And T. castaneum (Hbst.). Ann. Entomol. Soc. Amer. 53:280-281.
Ho, F. K. 1961. Optic organs of Tribolium confusum and T. castaneum and
Their usefulness in age determination. Ann. Entomol. Soc. Amer. 54: 921-925.
Ho, F. K. 1967. Identification of Tribolium larvae by their setal characteristics.
Ann. Entomol. Soc. Amer. (in press).
Sokoloff, A. 1966. The genetics of Tribolium and related species. Academic Press In.,
New York, 212p.
Sokoloff, A. and P. S. Dawson. 1963. Linkage studies in Tribolium castaneum Herbst.
IX. The map position of antennapedia, squint, short elytra and elbowed antenna.
Can. J. Genet. Cytol. 5:450-458.
HOWE, R. W. AND B. D. HOLE
Pest Infestation Laboratory
Slough, England
*Susceptibility of Sitophilus granaries to moderate cold.
This experiment was performed following the failure in several successive years to
establish populations of granary weevil in freshly-harvested wheat and barley stored in
farm bins. Cultures were prepared by groups of 800 adult weevils on successive lots of
320 g of Manitoba wheat for one day at 250 C and 70% R. H. Each culture was split into
32 samples and stored at 250 C and at a predetermined age each sample was moved
into a room controlled at 150 C and left there for eight weeks. In all, 24 samples of every
age from 1 to 39 days old at 250 C were exposed at 150 C and a similar number kept as
controls. On return to 250 C the samples were sieved regularly and the fecundity of a
sample of the emerged adults was measured for one week. Very young eggs and the
pre-pupal stage seem to be susceptible to exposure at 150 C, a temperature close to the
developmental threshold.
Age at
% alive
Exposure
No.
after
Days
emerged 6 weeks
1
2
3
4
5
6
7
8
9
10
119
187
360
463
651
669
717
662
608
625
98
99
99
98
94
99
100
99
99
99
Age at
% alive
Exposure
No.
after
Days
emerged 6 weeks
21
22
23
24
25
26
27
28
29
30
316
272
244
221
238
297
353
404
499
497
37
45
27
33
26
20
30
35
50
58
Eggs per
beetle
8.1
9.9
8.1
9.4
7.8
9.0
9.4
8.1
8.3
7.9
Eggs per
beetle
0.7
1.4
1.0
0.4
0.8
1.9
3.1
3.0
4.3
5.6
Age at
% alive
exposure No.
after
Eggs per
days emerged 6 week beetle
11
12
13
14
15
16
17
18
19
20
687
754
691
607
622
592
568
512
462
404
98
98
99
98
91
89
70
63
64
44
6.8
8.3
8.7
7.5
6.3
6.8
3.6
3.9
2.4
1.5
Age at
% alive
exposure No.
after
Eggs per
days emerged 6 week beetle
31
32
33
34
35
36
37
38
39
Control
551
640
728
707
728
751
731
708
723
804
56
72
84
84
90
----100
4.3
5.6
4.8
6.8
5.8
----8.4
Predominant stage in each group
1 -- 6 eggs
7 – 10 larvae
11 -- 14
15 -- 19
20 -- 25
1
2
3
4
26 – 28 prepupa
29 – 34 pupa
35 – 37 adults in grain
38 – 39 adults emerged
LADISCH, ROLF K.
Cancer Research Unit
Immaculata College
Immaculata, Pennsylvania
LADISCH, STEPHAN K.
Chemistry Department
The Johns Hopkins University
Baltimore, Maryland
Quinoid secretions in Tribolium confusum
We have most recently perfected a micro-polarographic method for the quantitative
determination of simple alkyl-parabenzo-quinones in the odorous secretion of the flour
beetle T. confusum. The method is capable of measuring quantities of quinine a small as
1 x 10-4 mgs, making it an excellent tool for the study of quinines in individual specimens
of insects—even in melanotic stink gland mutants msg (Sokoloff, 1964) which bear the
chemicals at as low as the submicrogram level. The procedure also furnishes conclusive
information on the minor amounts of hydroquinone that invariably appear to accompany
the quinine in the insect secretion. Furthermore, data of electrochemical significance
concerning the reversibility of the redox process and the extent of electron transfer can
readily be obtained by evaluating the analytical evidence from these extremely small
amounts of test substance. These special aspects, and details of our new method will be
discussed elsewhere.
In essence, the beetle to be studied is crushed in 500 microliters phosphate buffer
solution, pH 7, and the mixture is subjected to conventional direct current polarographic
electrolysis. Light is carefully excluded from the sample to minimize possible photoreduction. The resultant polarogram shows the amount of quinine (cathodic wave height),
the amount of hydroquinone (anodic wave height), and the half-wave potential. The latter
provides evidence on the identity of the insect chemicals in comparison with synthetic
alkyl-parabenzoquinones, i.e., in this instance with 2-ethyl-1, 4-benzoquinone, and 2methyl-1, 4-benzoquinone. We have used a Leeds and Northrup Polarotron P-40 and a
Sargent Model III Polarograph. The sample holder for the Polarotron was redesigned with
an electrolysis chamber of 250 microliters minimum capacity, and with a bridge containing
de-aerated electrolyte.
Typical polarograms of the very small amounts of quinine and hydroquinone
determined in mutants msg are depicted in Fig. 1. Computed quinone and hydroquinone
values for both the wild-type and the mutant insects are listed in Table 1, for ten individual
beetles in each case. Am average amount of 52.6 micrograms quinine per beetle was
thus found for the wild-type culture which was several months old. This result is
somewhat higher than that previously arrived at with a more indirect analytical procedure.
The latter has disclosed progressive biosynthesis of quinine in T. confusum as the beetles
grow to maturity, from zero in newborn adults to approximately 35 micrograms per beetle
at the age of four weeks (Ladisch, 1965).
Fig. 1. Polarograms of quinoid secretions in melanotic stink gland mutants
T. confusum msg. Each curve refers to an individual insect from
The culture. Data obtained in supporting electrolyte .05M alkali phosphate
buffer pH 7,0, at 250 C. Drop time, 3.58 sec; mass flow Hg, 2.33 mgs/sec.
Corrected for residual current.
Table 1.
Quinoid compounds in T. confusum, micrograms/insect.
Tenspecimens each of wild-type and mutant cultures.
(Q, Quinone; HQ, Hydroquinone) Wild-type culture mutant msg
Q
57.9
69.0
53.1
36.6
62.1
31.2
35.1
40.2
64.8
75.6
HQ
1.2
2.4
1.5
2.4
3.0
2.1
2.7
6,0
5.4
3.3
Total
59.1
71.4
54.6
39.0
65.1
33.3
37.8
46.2
70.2
78.9
Average Q: 52.6
Average HQ: 3.0
Q
HQ
Total
None
1.1
None
2.2
None
0.3
None
None
1.8
2.8
1.1
1.9
1.8
2.3
1.3
1.4
0.5
0.1
2.9
2.9
1.1
3.0
1.8
4.5
1.3
1.7
0.5
0.1
4.7
5.7
Average Q: 0.8
Average HQ: 1.6
The present data for mutants msg confirm the findings of Engelhardt, Rapoport,
and Sokoloff (19650 with respect to the greatly reduced amount of quinine in these
beetles. In contrast, hydroquinone was not detected in the mutants by these authors,
who extracted the volatile compounds by passing a stream of air over the insects and
condensing the sublimed matter in cold traps. This method would appear unsuitable to
demonstrate hydroquinone in insects as this compound will not sublime under these
conditions due to its low vapor pressure of a few microns Hg at the temperatures
employed. Information on the vapor pressure of insect quinines may be found in a recent
article by Ladisch and Suter (1965). Evidently, as is now shown, there is more
hydroquinone than quinine in the glands of these beetles (Fig. 1, Table I).
Hydroquinone has not been previously detected in wild-type insects by Loconti and
Roth (1953), Ladisch and McQue (1953), and Ladisch (1963; 1965), all of whom have
used sublimation methods in the isolation of the beetle quinines. Sixteen hundredths of a
microgram of hydroquinone per beetle was reported by Engelhardt et al. (1965) for wildtype insects. Yet, this compound was most likely a degradation product formed in vitro
from the quinine after collection, and is probably not related to hydroquinone existing in
the glands of the beetles. We have found inect quinines from T. confusum to convert
photochemically with ease to hydroquinone. In aqueous solution at pH 7, and exposed to
light of 750 foot-candles intensity for one hour, these quinines undergo complete
reduction. The solution is stable when stored in darkness. We shall discuss this
phenomenon in detail elsewhere. Employing the described direct method of analysis, we
have now found an average of 3.0 micrograms of glandular hydroquinone per insect of
the wild-type strain (Table 1).
The presently described analytical procedure is easy to perform and most reliable.
It has proved to be of special benefit in our current study of quinoid agents which we have
been able to detect in several genera and numerous species of the family Tenebrionidae.
Literature Cited
Engelhardt, M., H. Rapoport and A. Sokoloff. 1965. Science 150:632/
Ladisch, R. K. 1963. Penna. Acad. Sci. Proc. 37:127; ibid. 1965. 39:3
Ladisch, R. K. and B. McQue. 1953. Science 118:324.
Ladisch, R. K. and Sister St. A. Suter. 1965. Penna. Acad. Sci. Proc. 39:42.
Loconti, D. J. and L. M. Roth. 1953. Ann. Entom. Soc. Am. 46:281.
Sokoloff, A. 1964. Can. J. Genet. Cytol. 6:259.
We thank Dr. Alexander Sokoloff, University of California, for a culture of mutants
T. confusum msg. This study was supported in part by a grant from the Damon Runyon
Memorial Ffund for Cancer Research. We shall preset a more comprehensive treatment
of this subject, including potential environmental health aspects of quinoid toxicants from
stored-food insects at the April 1967 annual meeting of the Pennsylvania Academy of
Sciences.
LANGE, EUGENE L.
Population Genetics Institute
Purdue University
Lafayette, Indiana
*Sexual dimorphism in the pupal setae of Tribolium
The setae of Tribolium larvae and pupae have been examined by several people.
The variation that has been noted has dealt with specific differences (Mertz, 1961; Ho,
1960, 1964). No known intraspecific difference has been reported previously. This is a bit
surprising considering the mutants affecting seta number and shape in Drosophila. The
following is a description of a pupal sex difference in the position and number of setae in
Tribolium castaneum Herbst.
On the posterior margin of the eighth sternite of the pupae are a series of setae,
the pregenital setae (Fig. 1). The most lateral pairs are the long first lateral and the short
second lateral. They are slightly lateral and cephalad of the genital lobes and are present
in both sexes in this position. The second lateral is sometimes absent. Medially to these
pairs of setae are the first and second medial setae. The second medial is the smaller and
more sephalad of them. In males (left half of Fig. 1) both of the medials may be present
although usually only the first medial is found. They are approximately equally spaced
between the laterals. In females only the first medial is present, and it is about as far from
the first lateral as the second lateral is.
Fig .1.
Composite Male-female tribolium Castaneum showing pupal sex differences in setae
LERNER, I. MICHAEL AND NOBUO INOUYE
Department of Genetics
University of California
Berkeley, California 94720
*Regression of the sex ratio on maternal grandfather’s age
1961 Cavalli-Sforza proposed a method of estimating the spontaneous mutation
rate for sex-linked lethal by the regression of the sex-ratio of still (a) and live-born (b)
children on the age of the maternal grandfather at the time of the mother’s birth. The
rationale of these methods was given by him in the original paper as well as in a 1962
publication. Both he and Krebhiel (1966) found significant positive regressions on
samples of humans by method (a). For method (b) Cavalli-Sforza (1961) obtained, on
material involving some 67,000 women, a negative regression which was not significantly
different from zero. He demonstrated that (b) would be a less sensitive measure of the
effect, and proposed a massive study on a scale sufficiently large to obtain significant
results, The results have so far not been published.
Meanwhile, we have undertaken a similar study of our synthetic stock Tribolium
castaneum and T. confusum with method (b). Two separate experiments with the first of
the species, and a single with the second, containing ten replicates each (five males
mated to 10 stock females per replicate) were performed. In them, daughters were
obtained from males at monthly intervals (with some minor exceptions), and ten daughter
per replicate were mated to ten stock males and permitted to produce pupae, which were
then sexed. The age of the grandfathers spanned the period of 2 to 24 months, and any
of those that died were replaced by males of the same age. All the stocks were
maintained at 290C and 70% relative humidity. The total number of pupae examined was
over 150,000.
The pooled data for the different experiments are presented in Table 1 separately
for the two species. It is apparent from mere inspection that no linear regression is
detectible in T. confusum. The data for T. castaneum are not as unequivocal, and hence a
chi-square test was carried out:
Regression chi-sq.
Residual chi-sq.
Total chi-sq.
0.70 for 1 d.f.
20.23 for 20 d.f.
20.91 for 21 d.f.
The discrepancy in the total is likely due to rounding off figures in the calculation.
There does not seem to be a significant negative regression, although the possibility of
deviations from chance in some other manner is not excluded.
Thus it seems that the extent of our material bears out Cavalli-Sforza’s suggestion
that method (b) is not very sensitive, so that on the basis of our data, no estimate of
mutation rate is possible, They are, nevertheless, presented here, since if other attempts
at studying the same problem are made it may be possible to use them in combination
with other material.
Table 1.
Sex ratio and age of grandfather
Age of maternal
Grandfather at the
Time of mother’s
Eclosion
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
T. castaneum
No. of
pupae
%
sexed
males
3341
Total
5663
5808
5839
2628
6049
6305
6761
5961
5659
6946
7803
6423
5238
5619
2152
4100
1750
1710
1488
666
1307
99216
T. confusum
No. of
pupae
%
sexed
males
51.39
3883
49.70
49.87
4750
50.57
49.69
5658
48.30
48.36
3899
49.81
50.02
4067
49.79
50.24
5252
49.95
48.96
5371
50.12
50.39
4718
50.59
50.39
4876
51.35
49.48
4983
50.41
49.55
4587
49.86
49.97
2204
49.09
49.76
49.77
49.47
50.65
49.02
48.34
49.53
47.51
48.65
48.13______________________________
49.61
54248
49.99
Literature Cited
Cavalli-Sforza, L. L. 1961. Un metodo per la stima della frequenza di mutazione
Nell’uomo: Risultati preliminari. Atti Assoc. Genet. Ital., 6:151-162.
Cavalli-Sforza, L.L. 1962. Demographic attack on genetic problems. The use of
Vital and Health Stat. for Genet. And Rad. Studies (UN-WHO, N.Y.): 221-233.
Krehbiel, E.L. 1966. An estimation of the cumulative mutation rate for sex-linked
Lethal in man which produce fetal deaths. Am. J. Human Genet. 18:127-143.
This work was supported in part by USPHS grant GM-08942.
LOSCHIAVO, S. R.
Canada Department of Agriculture Research Station
25 Dafoe Road
Winnipeg 19, Manitoba, Canada
Evaluation of an anti-feeding compound as a protectant against stored-products insects
An anti-feeding compound, 24055 discovered by American Cyanamid Company
was initially reported to inhibit feeding by certain phytophagous chewing insects. This
interesting property suggested that the compound might be exploited as a protectant
against stored-products insects. Laboratory experiments were conducted to determine
whether 24055 applied to the outer surface of cotton sacks prevented penetration and
oviposition by flour beetles and grain beetles. The compound was used as a 25% wettable powder in water and applied to empty sacks at different rates of application.
Adults of Tribolium confusum, Cryptolestes turcicus and Oryzaephilus
surinamensis did not penetrate or lay eggs through the mesh of treated sacks filled with
flour. They died or became moribund within one week of exposure. This compound was
shown to cause mortality of flour beetles by volatile action and by contact action, the
former having the greater effect. Its ability to protect packaged cereal products is currently
being investigated. Its anti-feeding and insecticidal properties are being examined in
relation to reproduction and mortality of treated adults.
LOSCHIAVO, S. R. AND L. B. SMITH
Canada Department of Agriculture Research Station
25 Dafoe Road
Winnipeg 19, Manitoba, Canada
The distribution of Oryzaephilus Mercator Fauvel in Canada
Since late summer 1966, we have received an unusually large number of reports
of the merchant grain beetle, Oryzaephilus Mercator Fauvel occurring in packaged cereal
products. This species had not been reported in Canada before 1952. It was recorded
from Ottawa in 1952 and Vancouver in 1954.
This beetle superficially resembles the saw-toothed grain beetle, O. surinamensis
and was considered by many workers to be a variant of the latter species. However,
Howe (1953, 1956) and Slow (1956) confirmed Fauvel’s earlier finding (1889) that O.
Mercator was a separate species. We know that the merchant grain beetle has frequently
been erroneously identified as the saw-toothed grain beetle. Consequently, we suspect
that the former species is distributed in Canada far more widely and for a longer time than
indicated by the two reported records mentioned above.
The increasing frequency of reports of the merchant grain beetle suggests that it is
becoming firmly established as a pest of stored products in households, retail stores, and
storage warehouses in Canada. It has been found in flour, rolled oats, biscuit products,
peanuts, walnuts, puffed rice, cake mix, chocolate bars, dried fruits and vegetables, flax
seed, Oriental foods, and copra.
A survey is being conducted to determine whether this species enters the country
on certin imported products from which it may spread to other food commodities via
normal handling, shipping, and storing operations.
MCONALD, D. J. AND L. STONER (MRS.)
Department of Biology
Dickinson College
Carlisle, Pennsylvania
*Factors involved in the survival of Tribolium confusum populations
Flour beetle populations can persist for long periods of time in small volumes of
unrenewed flour. One factor that may influence the longevity of a population under these
conditions is the effectiveness of the self-limiting mechanisms that restrict population size,
for it seems likely that a smaller population would be able to persist longer than a larger
one. However, there may be other factors influencing longevity and these might be
revealed by a close study of populations from genetically distinct strains that differ in
several population characteristics. In these first experiments two strains were used, McGill
black and a wild type (red) strain. One hundred single pair populations of each strain and
another hundred populations of mixed pairs, one red adult and one black, were started in 5
gms of flour medium. These three population types are referred to hereafter as the R. B.
AND H. (heterozygous) populations. The populations were censused at two-week
intervals and since the medium was never renewed they eventually became extinct. Table
1 shows some of the data obtained during weeks 10 through 28 inclusive. Since the adult
and egg numbers of the H populations fall between the R and B populations it is possible
that these characteristics are determined by the body color genes which exhibit no
dominance.
Table 1.
Mean characteristics of red, black, and heterozygous populations
Population type
Live
Adults
Egg
Red
Heterozygous Black
97.6 + 19.5 81.2 + 17.2 71.1 + 13.8
20.5 + 7.3
28.7 + 10.0 42.6 + 13.6
Tests of
Significance between
R and B R and H H and B
t = 11.0 t = 6.3
t = 4.5
P .001 P .001 P .001
t = 14.2
t = 6.6
t = 8.2
Numbers
P .001 P
Weeks
37.4 + 6.5
Duration
45.1 + 5.4
39.9 + 5.6
.001 P
t = 2.9
t = 9.1
P .005 P .001
.001
t = 6.7
P .001
Since the H populations, although bigger than the B populations, survive several weeks
longer, it is clear that factors other than the adult population size can influence survival in a
depleted environment. However, within each population type there is a negative
correlation between adult population size and duration. Table 2 shows this as well as the
Table 2.
Correlations between adult numbers (A), egg numbers
(E), and weeks duration (D)
Red
Population type
Heterozygous
Black
Total
Correlation
coefficient
rAE
rAD
rED
-.70 P .001
-.72 P .001
.49 P .001
-.41 P .001
-.88 P .001
.37 P .001
-.49 P .001
-.67 P .001
.32 P .001
-.97 P .001
-.02 P .10
-.94 P .001
-1.01 P .001
.06 P .10
-.42 P .001
-.76 P .001
-.02 P .10
-1.46 P .001
Partial
Correlation
Coefficient
rAD.E
rED.A
rAE.D
fact that the egg number is apparently not a factor in determining duration. When the
effects of the negative correlations between eggs and duration are not significant. Since
the level of adult numbers is established early in the population’s life, it is possible that this
level can influence the population’s chance of surviving a period of stress, generated by
depleted resources. Since, in the present instance, a smaller population has a greater
chance of surviving, the results suggest a way in which natural selection acting on the
whole population as a unit might establish self-limiting population mechanisms. In another
experiment, using the same strain, when the population reaches its peak number of adults
at eight weeks, all the adults are discarded then and every fourth week thereafter. Table 3
shows some of the data. Again, for reasons not yet known, the H populations survive
longest although not significantly longer than the B populations. Within each population
type the total number of adults produced is positively correlated with duration. This can be
interpreted to mean that populations differ in their efficiency in converting flour into beetles,
Table 3.
Correlation between the total number of live adults produced (T),
The duration of the population in weeks (D), ad the mean number
of adults at the peak period, weeks six and eight (P)
Red
Means
Heterozygous Black
Total live
Adults produced
166
157
148
Weeks duration
41
51
Peak adults
x 6-8 weeks
92
76
Total
Correlation
coefficient
rTD
rPD
rTP
Red
.
43
P .01
P .01
T tests
R and H H and B
.0001
.01
47
.005
.0001
74
.0001
.0001 P = .38
Heterozygous
- .043 P .1
.39
R and B
.005
P = .11
Black
.31
P .01
.54
P .01
-.24
P .02
.05
P .1
P
.34
P .01
.06
.1
Partial
Correlation
Coefficient
rTD.P
.528 P .01
.345 P .01
.593 P .01
rTP.D
.519 P .01
.157 P .1
.443 P .01
rPD.T
-.305 P .01
-.287 P .01
-.214 P .05
and the more efficient populations have an increased chance of surviving. However, the
rate at which this conversion takes place should have some effect on duration. Those
populations producing more beetles early in the population’s life should exhaust the
environment sooner. Consequently, a negative correlation might be expected between the
peak number produced at weeks 6 ad 8 and duration. However, since the peak number is
part of the total number, and the total is positively correlated to duration, the negative
relationship between peak number and duration only emerges in the partial correlation
coefficients. Other characteristics, such as the mean weight of the individuals in the
population, measured as the weight of pupae, appeared to have no effect on population
survival.
MUSGRAVE, A. J. AND S. B. SINGH
Department of Zoology
University of Guelph
Guelph, Canada
*Cytochemical studies of Sitophilus granaries mycetomes
Cytochemical studies of the outer membranes and chromatinic material in the
inherited, supposedly symbiotic mycetomal micro-organisms of Sitophilus granaries have
revealed the presence of considerable amounts of DNA occurring as either consolidated
dots or a spongy network. There is also evidence that, at times, the organisms are
surrounded by cell walls.
MUSGRAVE, A. J. AND I. GRINYER
Department of Soology
University of Guelph
Guelph, Canada
*Mycetomes as possible obligate symbiotes of Sitophilus
The peripheral membranes of the micro-organisms of the mycetocytes of adult
midgut caecae and of larval mycetomes of Sitophilus granaries (L.). GG strain, have been
examined with an electron microscope. The majority of the mycetocytes were depleted of
intracellular organelles but contained large numbers of mycetomal micro-organisms, most of
which exhibited only one peripheral membrane. Some mycetocytes, however, had welldeveloped ultrastructure and harbored mycetomal micro-organisms which showed two
peripheral membranes, namely a cell wall and plasma membrane. Intermediate conditions
also occurred.
It is suggested that the absence of host-provided membranes around the microorganisms categorizes them as obligate symbiotes.
O’BRIAN, DENNIS M., ANDREA LEVINE AND
FRANK F. KATZ
Department of Biology
Seton Hall University
South Orange, New Jersey 07079
A quantitative approach to the feeding of Hymenolepis diminuta Eggs to the flour beetle,
Tribolium confusum
All stock cultures were maintained at 300 C, 92 to 96% relative humidity and
12L:12D light conditions. The medium consisted of 95% bleached Gold Medal Wondra®
flour and 5% National Active Dry ® yeast and was changed at least every two weeks. Eggs
collected from apopulation of beetles four to six weeks following their eclosion were used to
set up the experimental groups which consisted of virgin adults maintained in individual vials
throughout the duration of the experiment.
At six to eight weeks of age, the experimental beetles were starved for a period of
five to six days prior to being allowed to feed for a 48-hour period on a known number of
eggs. The eggs were obtained from five to ten freshly obtained terminal gravid segments of
H. diminuta and embedded in a 17% gelatin, 2% sucrose medium. The solvent for
preparing the gelatin and sucrose consisted of three parts distilled water: one part
supernatant of a mammalian saline extract of freshly-obtained homogenized gravid
segments. This extract was centrifuged for a period of five minutes at 1800 rpm and the
supernatant was removed. The eggs and diluted supernatant was added to the gelatin
sucrose mixture just prior to gellation. After examination of the surrounding feeding area
and feculae for eggs or egg shells, it appeared that no eggs or embryos were passed
through the digestive tract during the 48- hour feeding period, and the 24-hour period after
replacement on the regular diet. Thus, it was possible to determine the number of eggs
consumed per beetle by subtracting the number remaining at the end of the 48 hour feeding
period from the initial number. Selective staining with Trypan Blue (0.02 to 0.04% in
mammalian saline) in addition to observational studies indicated over 90% viability of the
eggs freshly obtained from gravid segments and those kept for the 48-hour period in the
gelatin medium. In addition, eggs maintained for 48 hours in the gelatin medium did not
take up the stain at a faster rate than freshly obtained eggs, indicating no decrease in the
relative viability of the eggs. The majority of beetles that fed consumed between one and 60
eggs, 21 to 29% becoming infected, with a mean of 1.5 to 3.0 cysticercoids per infected
beetle.
ONYEARU, A. K.
University of St. Andrews
Dundee, Scotland
The behavior and biology of flour beetles, genus Tribolium as studied in laboratory
gradients of temperature and humidity
Under the supervision of Dr. F. L. Waterhouse, Natural History Department of
Queens College, University of St. Andrews, Dundee, Scotland, Dr. A. K. Onyearu has
recently completed a three-year research program on the behavior and biology of Tribolium
spp. including some genetic strains and geographical populations of Tribolium castaneum,
Hbst.
The laboratory studies were mainly conducted in a gradient environment. The
equipment for this has been described earlier (Graham, Onyearu and Waterhouse, 1965).
Tribolium populations employed in these investigations include:
1. Seven geographical stocks of Tribolium castaneum obtained from:
Umuahia
Onadam
Kano
Kenya
Kingston
Rangoon
Tokyo
-
(Eastern Nigeria)
(Western Nigeria)
(Northern Nigeria)
(East Africa)
(Jamaica, West Indies)
(Burma)
(Japan)
2. Three genetic strains of Tribolium castaneum namely:
Black, Pearl and Mahogany. The last is a recent find described elsewhere.
(Onyearu and Graham, in press).
3. Laboratory strains of :
Tribolium confusum duVal
Tribolium anaphe Hint.
Tribolium madens Charp.
Tribolium destructor Uytt.
The following aspects were covered in these investigations:
1. Adult distributions in the gradient environments.
2. Oviposition as a functional aspect of adult orientation on the gradients of
temperature and humidity.
3. Developmental periods in gradient environments as well as under the uniformly
controlled conditions of the C. T. Room.
Results of these investigations have served to emphasize the need for relating
experimental conditions and experimental animals to those occurring in the field to which the
original ecological problems relate. Details of these studies formed the theme of a Doctoral
thesis recently submitted to the University of St. Andrews (Onyearu, 1966). Further
publications now under preparation are to be made elsewhere.
Literature Cited
Graham, W. M., A. K. ONYEARU AND f. l. Waterhouse. 1965. Temperature and
moisture gradient equipment. Can. Ent. 97 (8):880-996.
Onyearu, A. K. 1966. Laboratory studies on the behavior and biology of flour
Beetles (Genus Tribolium), in gradient environments, with special
Reference to species and strain comparisons. Ph. D. Thesis, University
Of St. Andrews, Scotland.
Onyearu, A. K. and W. M. Graham. 1967. Life history studies of two body color
Mutants of Tribolium castaneum Hbst. (Coleoptera Tenebrionidae). (In press).
PEASE, R. F. W., T. L. HAYES AND A. SOKOLOFF
Electron Research Lab., Donner Lab.,
Lawrence Radiation Lab., Department of Genetics
University of California, Berkeley
The scanning electron microscope
The scanning electron microscope utilizes a fine, moving beam of electron
(diameter as small as 50 A) to probe the specimen for a variety of kinds of physical,
chemical and electrical information. As the scanning beam is swept across the specimen,
the electron beam of a standard cathode ray tube is driven in synchrony with it and the
brightness of the cathode ray beam is modulated by the signal from one of several
detectors. The secondary radiation produced by the interaction of the scanning electron
beam is not imaged but is only used to convey information about the particular point being
bombarded. The image is a result of the synchrony between the scanning beam in the
microscope column and the beam of the cathode ray tube. The resulting 1:1
correspondence of points on the specimen with points on the face of the cathode ray tube
serves to identify the location of the information seen by the detectors. The separation of
information and localization allows many different kinds of information to be gained while
utilizing the localization possible with an electron beam. Different kinds of secondary
radiation are produced by the interaction of the electron beam with the specimen and each
carries a particular kind of information. For example, visible light may be produced and give
information about the molecular structure of the biological material itself or the location of
light producing stains within the specimen. Operated in this mode, the scanning electron
microscope could be described as a high resolution fluorescence microscope (Pease et al.
1966a). Secondary electrons are another form of radiation produced by the interaction of
the scanning beam with the specimen. Since production of these secondary electrons is a
function of the angle between the scanning beam and the surface of the specimen,
stereoscopic information can be obtained. The scanning electron microscope operated in
this mode can be described as a high resolution stereoscopic microscope (Hayes et al.
1966). In addition, ultraviolet radiation, characteristic X-rays, backscattered electrons,
energy loss electrons and specimen current are all produced in the specimen and might be
used to build up the scanning electron microscope image. The principles of the instrument
and several of its modes of operation have been recently reviewed (Oatley et al. 1965).
The present paper deals with an attempt to visualize living Tribolium by using the
scanning electron microscope in the secondary electron mode of operation. There are
several advantages both physiologically and morphologically if the specimen can be viewed
while it is living. Morphologically, the reduction in the possibility of artifacts and the
simplicity of sample preparation are important. Physiologically, the possibility of observing
“on-going” processes would be most valuable, as well as utilizing the tiny electron beam as
a micro-radiation source in radiobiological studies (Pease et al. 1966b).
Literature Cited
Pease, R. F. W. and T. L. Hayes. 1966a. Nature 210:1049.
Hayes, T. L., R. F. W. Pease and L. W. McDonald. 1966.
Lab. Invest. 15:1320-1326.
Oatley, C. W., W. C. Nixon and R. F. W. Pease. 1965. In Advances in
Electronics and Electron Physics, 21:181-247, Academic Press Inc.,
New York.
Pease, R. F. W., T. L. Hayes, A. S. Camp and N. M. Amer. 1966.
Science 154:1185-1186.
REYNOLDS, ELISABETH M. AND BARBARA M. RUNDLE
Pest Infestation Laboratory
Slough, Bucks, England
The response of larvae of Trogoderma granarium Everts to – 100 C
Some preliminary observations of the response of fully grown larvae of Trogoderma
granarium Everts to low temperatures have been made by placing samples of approximately
50 larvae in a deep freeze cabinet at – 100 C for various periods of time. The larae were
taken directly to the cabinet from cultures maintained at 300 C and after exposure were
placed on food and returned to 300 C. The larvae were then examined, initially at weekly
intervals and then fortnightly or three weekly, for six months.
Periods of exposure to -100 C of from two hours to 14 days were tested. Exposures of up to
3.5 hours appeared to have no lasting effect on the larvae. From 7.5 to 72 hours larval
mortality increased to 96%. No larvae survived exposures of five days or longer.
All the exposures tested immobilized the larvae. On their return to 300 C some larvae
remained immobile for a long time. Such larvae displayed no sign of damage and it was
impossible to tell from their appearance whether they would eventually die or resume
development. Dead larvae became desiccated and dark in color.
After short exposures some larvae were still immobile at the end of the observation period.
Increase in exposure not only increased mortality but also appeared to decrease the period
of immobility before the larvae could be seen to be dead.
These findings are demonstrated in Table 1 below. In this summary which combines the
results of three separate tests some slight approximations in dates have been made in order
to divide the observation period into the five intervals shown. Results for exposures giving
less than 100% kill have been combined into groups.
Table 1.
Numbers of survivors and dead larvae found during the observation
Period at 300 C following exposure to – 100 C.
Length of
Exposure
To -100 C
0, 1.5
3.5 h
7.5-41 h
48-72 h
5 days
7 days
10 days
12 days
14 days
Observation period, days from start
of exposure
Stages
found
0-30
31-60
61-90 91-125 126-166
Total insects found
Adults
at
end
dead
134
1
4
47
7
178
adults
dead
Larvae
129
3
2
0
0
2
1
0
1
0
adults
dead
larvae
33
5
0
1
8
91
65
13
5
4
adults
dead
larvae
8
3
1
4
1
170
30
20
2
0
17
4
222
dead
larvae
47
2
0
1
0
0
0
50
36
37
38
43
8
17
10
7
2
1
1
0
1
0
0
0
0
0
0
0
0
0
0
0
0
0
0
0
47
55
49
50
“
“
“
“
SCHMITZ, THOMAS H. 1 AND DUWAYNE C. ENGLERT
Department of Zoology
Southern Illinois University
Carbondale, Illinois
Allelism of “mottled” (mt) and “melanotic stink glands” (msg) in Tribolium castaneum
Two mutations of similar phenotype have been reported, “melanotic stink glands,
“msg (Sokoloff and Hoy, TIB 8:55-56), and “mottled”. Mt (Englert, TIB 9:59-60). Both of
these mutations are affected by apparent deposition of polymerized darkly pigmented
secretions composed of ethylquinones in the region of the stink glands.
1 Present address: Population Genetics Institute, Purdue University, Lafayette,
Indiana.
Both mutations are inherited as autosomal recessives exhibiting incomplete
penetrance and variable expressivity. Under ordinary laboratory conditios “mottled” exhibits
an average penetrance of 63 per cent. In the stock received from the Berkeley Stock
Center, msg was found to exhibit a penetrance of approximately 60-70 per cent under our
laboratory conditions. However, the viability of the stock ws less than good. Linkage tests of
the two had indicated a relationship with linkage group III (includes “black”, b). Therefore, a
test for allelism utilizing single pair matings between the two stocks was conducted. From
29 single pair matings, only 16 of 178 beetles (8.9 per cent) exhibited what ould be called
the mutant phenotype.
A second test was conducted, this time utilizing mass matings in an attempt to
increase productivity. A total of 464 progeny were examined, 17 of which exhibited the
mutant phenotype (3.7 per cent). Neither of these tests could be considered conclusive, so
to further establish allelism, the msg stock was crossed to the Purdue wild foundation stock
to give a similar genetic background to that of mt. Mutant progeny from F2 segregants were
used to start the “new” msg stock.
Beetles from the reconstituted msg stock were crossed with mt beetles to again
check for allelism. The results revealed only 3 of 132 beetles which were of the “mutant”
phenotype (0.8 per cent), indicating that similar genetic background did not increase the
penetrance of the “mutant” phenotype. Thus, in the classical sense of allelism the two
mutants cannot be considered to be allelic.
SHAW, DAVID D.
Pest Infestation Laboratory
Slough, Bucks, England
The chromosome numbers of some stored product Coleoptera
Species
Karyotype
DERMESTIDAE
Dernestes nacykatys
“
“
“
“
D frischii
D frischii
D ater
D haemorrhoidalis
D lardarius
D peruvianus
8 II
8 II
8 II
8 II
8 II
8 II
8 II
8 II
8 II
+
+
+
+
+
+
+
+
+
Xyp
Xy1y2
Xy1y2y3
Xyp
Xy1y2
Xyp
XY
Xyp
Xyp
Trogoderma parabile
T. glabrum
Anthrenus verbasci
A. Flavipes
9 II
9 II
8 II
8 II
+
+
+
+
Xyp
Xyp
Xyp
Xyp
JOHN 7 SHAW 1966
OSTOMATIDAE
Tenebroides mauritanicus
11 II + Xyp
SHAW (Unpublished)
TENEBRIONIDAE
Alphitobius diaperinus
Tenebrio molitor
T.obscurus
Gnathocerus cornutus
9 II
9 II
9 II
9 II
+
+
+
+
XO
Xyp
Xyp
Xyp
SHAW, DAVID D.
Pest Infestation Laboratory
Slough, Bucks, England
Sex chromosome variation in D. maculates and D. frischii
Five cultures of Dermestes maculates were available for cytological examination.
The individuals from stocks originating in Australia, South Africa, Nigeria and Sudan all
showed a standard Xy parachute sex bivalent but all the individuals examined from a
Indian culture were characterized by multiple y’s. In 32 out of 33 individuals examined,
two y chromosomes were present leading to the formation of a sex multiple of three at
meiosis. This multiple shows regular meiotic behavior giving a consistent X-2y
segregation with no observed deviation.
In a single individual of the Indian strain, three y chromosomes were found and
here the meiotic behavior was less regular. At first metaphase, two principal patterns of
orientation were observed.
First, and most frequently (35 out of 40 cells scored), the three y’s co-orientated
with the single X giving a regular X-3y segregation. In five other cells one of the y’s was
orientated to the same pole as the X giving an Xy-2y pattern of segregation. The
consequences of this irregular behavior were seen at second division for in addition to (8
+ X) and (8 : 3y) cell types, (8 + X + y) and (8 + 2y) cells were also seen.
Five of the 25 males of D. frischii examined also turned out to be the Xy1y2 type.
Two possible explanations can be offered to account for these multiple y variants.
Either they represent a polymorphism which exists in nature or, alternatively, we are
dealing with a system of supernumerary y-chromosomes which have arisen in culture
presumably as a consequence of inbreeding coupled with reduced competition.
A 3y strain of D. maculates has now been selected and it is hoped to isolate
females which may possess a y-chromosome due to the irregular behavior of the 3 y’s at
meiosis. (In conjunction with Dr. B. John, Department of Genetics, Birmingham
University, England,)
SOKAL, ROBERT R.
Department of Entomology
The University of Kansas
Lawrence, Kansas
*The effects of synchrony of egg batches on fitness characters and competition in
Tribolium castaneum.
In a previous study (Sokal and Karte, 1964) the effects of density on three
characters affecting fitness (survival to adulthood, dry weight of adults, and length of
developmental period) were described for ++, +b and bb individuals in pure culture and in
mixed cultures at varying proportions. It was believed that these findings would help
explain the results of a selection experiment (Sokal and Sonleitner, 1965; a more
extensive manuscript is in preparation) in which by 10 generations the frequency of b
increased from 0.25 to 0.53, other cultures started at gene frequency 0.50 had increased
to 0.58, while yet othes started at 0,75 had maintained themselves approximately at this
value. Analysis of the data revealed, however, that the results of the earlier study differed
in two important details from those of the selection experiment. In the study of Sokal and
Karten (1964) black had consistently higher or at least equal survival to adulthood when
compared to ++, while in the selection experiment the wild type strain invariably had
appreciably higher survival than bb. Also developmental period of bb was less than that
of the wild type in the experiment of Sokal and Karten (at least at the lower densities),
while in the selection experiment, the black strain developed considerably slower than the
wild type strain (bY as much as 19 days).
Study of these results and another experiment reported later in this issue (Sokal,
1967) ruled out genetic differences between the original stocks and the selected strains
as responsible for the differences in survival and developmental period. Another possible
cause of the divergent results of the two sets of experiments is that Sokal and Karten
used egg batches obtained from 4-hr egg collections while the selection study was based
on 3-day egg yields. It is assumed that the latter egg batches are les synchronous and
may therefore be responsible for differences in survival and developmental period. The
present paper tests this hypothesis.
The materials and techniques employed are identical to those reported in Sokal
and Karten (1964). The design differed in that only two densities, 20/g and 100/g were
set up with pure cultures of ++, +b and bb, and with mixed cultures representing HardyWeinberg proportions for gene frequencies 0.1, 0.5 and 0.9 of black. For each
combination of conditions 4-hr as well as 3-day eggs were tested. Replication for each
experimental condition was four vials for the pure cultures and 0.5qb, and eight vials for
the other gene frequencies at density 20/g, while at density 100/g the respective
replications were three and six vials. The entire experiment was repeated four times.
Survival to adulthood: The overall survival not broken down by genotype is shown
in Table 1. The 4-hr data compare well with previous results by Sokal and Karten (1964)
and Sokal and Huber (1963). Percentages of survival at densities 20/g are higher than at
100/g, even for the ++ cultures which previously did not show this trend. At density 20/g
the 3-day egg batches did not differ from the 4-hr vials in per cent survival, but at 100/g
they are significantly lower than the same eggs at density 20/g and also the 4=hr cultures
at density 100/g. The lowest single per cent survival is in the 3-day cultures of ++ which
dip to 49.61%.
Table 1. Overall adult survival at different gene frequencies, and two densities
For 4-hr and 3-day egg batches
Gene frequency
Egg batch Density
0.00
0.10
0.50
+b
0.90
1.00
4 hrs
20/g
100/g
81.98
71.01
79.84
64.39
85.78
76.65
87.97
60.24
83.20
77.68
85.31
71.21
3 days
20/g
100/g
83.90
49.61
83.33
55.43
80.26
60.75
86.72
62.85
78.25 86.25
63.76 66.79
Figure 1 illustrates the survivorships as percentages of input (averaged over the
four experiments) for pure and mixed cultures at the three gene frequencies, two
densities and for the two types of egg batches. Although the details of this figure differ
somewhat from corresponding Figures 1 and 4 in Sokal and Karten (1964), the general
trends are identical, showing heterozygous superiority under most conditions and a trend
toward higher survival of wild type with increasing gene frequency of black at the highest
density. Some instances of genetic facilitation are again demonstrated. However, with
respect to the main feature of the experiment, the effect of synchrony of the egg batches
on survival, no apparent difference can be illustrated. This is borne out by a factorial
analysis of variance which shows only density and replication as significant main effects.
Most significant interactions involve replication as one of the factors, the outcomes of the
several experiments having fluctuated considerably.
Dry weight of adults: These results (not illustrated here) are quite comparable to
the findings of Sokal and Karten (1964). Again, weights are in the relation +b ++ bb.
This relation is maintained at both densities and for both types of egg batches. Weight is
not affected by synchrony of egg batches. Beetles at density 20/g are considerably
heavier than those at 100/g.
Length of developmental period: This variable is affected by density and differs
considerably among genotypes. Table 2 shows the relations among developmental
periods of the three genotypes in this experiment and in that of Sokal and Karten. Note
the reversal in relationship between ++ and bb for the two densities of the 4-hr cultures.
However, in the 3-day egg batches, patterned after the selection experiment, this reversal
does not occur. Thus, the 4-hr cultures actually come closer to the bb ++ pattern of the
selection experiment (based on 3-day egg yields at high densities) than do the 3-day
cultures. Mean developmental period was apparently affected markedly by the synchrony
of the egg hatch, yet when mean hatching period, the time from the start of the
experiment until the hatching of each individual egg, was calculated it was seen that the
increase in developmental period in the 3-day eggs could be accounted for by their
greater mean hatch time.
Table 2.
Relation among developmental periods of the three genotypes.
Egg batch
Density
4 hours
3 days
20/g
This study
Sokal & Karten “64
++ bb = +b
++ bb +b
++ = bb = +b
100/g
This study
Sokal & Karten “64
+b bb ++
bb +b ++
++ +b bb
More informative than means are cumulative frequency distributions of emerging
adults of the three genotypes shown in Figure 2. Figures 2a and 2b are representative of
the general findings. They illustrate density 100/g of the 3-day batches in pure culture
and at gene frequency 0.9, respectively. It is clear from these graphs that the ++ strain
lags consistently behind the other two, which for most of the experimental conditions
produced coincident curves, or if they did separate, showed the bb to be slightly ahead of
the +b. This relationship (of ++ lagging behind bb) was also found in separate,
unpublished experiments carried out by F. J. Sonleitner in our laboratory. Notice how in
the pure strains (Figure 2a) the emergence patterns are much more diffuse and
development takes longer than in the mixed strins at gene frequency 0.9. This culture
exhibits genetic facilitation, with all three strains developing faster and closer together as
shown by the steeper slope of the curves. Under only one condition (100/g of the 4-hr
egg batches in pure culture; see Figure 2c) was ++ not the slowest strain.
Conclusion: While the relations described above have considerable interest, they
do not demonstrate any major effects due to greater or lesser synchronization in egg
batches and are thus not able to explain the differences between the early results of
Sokal and Karten (and now also the present results) and those observed in the selection
experiment of Sokal and Sonleitner.
Contribution No.1351 from the Department of Entomology, The University of
Kansas. This is paper No. 5 in a series on the ecological genetics of Tribolium. Numbers
1 through 4 are listed at the end of paper No. 6 (Sokal, 1967, in this issue of TIB). This
research was supported by the National Science Foundation under grant GB-2170 and by
a Public Health Research Career Program Award (No.3-K3-GM-22, 021-O1S1) from the
National Institute of General Medical Sciences. The technical assistance of Miss Yu-Jen
Chen, Mrs. Maxine L. Howe and Mrs. Cornella B. Tollefson are very much appreciated.
Literature Cited
Sokal, R. R. 1967. A comparison of fitness characters and their response to density in
Stock and selected cultures of wild type and black Tribolium castaneum,
TIB 10: 142-147.
Sokal, R. R. and I. Huber. 1963. Competition among genotypes in Tribolium castaneum
At varying densities and gene frequencies (the black locus). Genetics 49:195-211
Sokal, R. R. and F. J. Sonleitner. 1965. Components of selection in Tribolium
(Coleoptera) AND HOUSEFLIES. Proceedings of the XII International
Congress of Entomology. 274-275.
Figure 1. Adult survival expressed as percentages of egg input averaged over the
replicates and experiments of the study and shown for the two types of
egg batches, two densities and three gene frequencies employed.
Leftmost column with noncontiguous bars represents the results of rearing
the beetles in pure culture. Hollow bars represent the ++ genotype,
hatched bars +b and black bb.
SOKAL, ROBERT R.
Department of Entomology
The University of Kansas
Lawrence, Kansas
*A comparison of fitness characters and their responses to density in stock and selected
cultures of wild type and black Tribolium castaneum.
The wild type and black control strains of a selection experiment (Sokal and
Sonleitner, 1965; a more extensive manuscript is in preparation) differed in two important
respects from the stock cultures from which they were derived. Invariably the wild type
strain had appreciably higher adult survival than the bb cultures, this difference increasing
at the higher densities. By contrast, experiments with the stock cultures carried out by
Sokal and Karten (1964) and Sokal (1967) had shown that black had consistently higher,
or at least equal, survival to adulthood when compared with ++. However, these
experiments were carried out in 8 g of flour in 6-dram shell vials, while the observations in
the selection experiment had been made in 40g of flour contained in half-pint Mason jars.
A second discrepancy between the two types of experiments is that the developmental
period of bb was less than that of the ++ strains under all but one set of conditions in the
stock cultures, while in the selection experiment bb took consistently longer to develop
than ++.
The purpose of the present experiments is to delineate more sharply the nature of
the differences in survival to adulthood, dry weight of adults, and length of developmental
period between the controls of the selection experiment and the stock cultures from
which they were derived. These comparisons were carried out under conditions of the
earlier experiments by Sokal and Karten (1964) and Sokal (1967), i.e., in 8 g of flour in
shell vials. While an analysis of these differences is of primary importance for an understanding of the results of the selection experiment, the findings are here interpreted in
terms of the treatment to which the controls in the selection experiment had been
subjected and are of some general interest from this point of view.
The materials and techniques are identical to those reported in Sokal and Karten
(1964). However, only pure strains ++ and bb were tested at four densities, 5/g, 20/g,
50/g, and 100/g. The two strains were the standard UPF wild type and black stock
cultures employed in work in our laboratory (see stock list) and the ++ and bb controls
from the RSE selection experiment (Sokal and Sonleitner, 1965). The controls were
taken from generations 25 and 26, respectively, of the second replicate for these two
strains. The eggs for these experiments were obtained during a 4-hour period.
Replication for each experimental condition was 10 vials at density 5/g, 4 vials at density
20/g, and 3 vials each at densities 50/g and 100/g. The entire experiment was repeated
two times.
Survival to adulthood: Survival as percentage of egg input is graphed in Figure 1,
which shows that survival decreases with an increase in density for all tested cultures. In
both the stock and selection controls, the black strain ad a consistently higher survival,
quite in contrast with our findings under the conditions of the selection experiment. Thus,
the genetically determined differences in survival between the black and wild type stocks
have not changed in the selection controls, or if they have, these differences are not
expressed under the conditions of these experiments (8 g of flour in 6-dram shell vials). I
may therefore be that the reverse relationship observed in the selection experiment is due
to the environment peculiar thereto (40 g of flour in half-pint jars) or is only expressed in
that environment. An overall decrease in survival of the selection controls is noticeable in
Figure 1. The survival is expressed in degrees (because of the angular transformation);
on the average the selection controls have five degrees lower survival than the
corresponding stock cultures. Investigation of per cent hatchability of eggs revealed that
these differences in survival are due to differences in larval or pupal survival.
Dry weight of adults: These relations (not illustrated) are generally consistent with
previous findings in various experiments (e. g., Sokal and Karten, 1964). The ++ strain is
heavier than the bb strain and there are no differences between the stock cultures and
the selection controls.
Length of developmental period: Here, relations in the stock cultures are as
described in earlier studies, with the ++ strain having a longer developmental period than
bb at the low densities. While even at density 100/g the average developmental period of
the ++ is half a day longer than that of the bb, the steeper increase of the developmental
period of bb in response to density is evident. Studies by Sokal ad Karten (1964) and
Sokal (1967) have shown that under these conditions of high density bb has a longer
developmental period than ++. This relationship is illustrated in the selection controls in
Figure 2. Since the actual selection experiment was run at asymptotic densities, between
82/g and 90/g, it may simply be that differences in developmental period observed in that
experiment are reactions to the density in the cultures. Notice, however, that the selected
strains hve on the average a developmental period two days longer than their
counterparts from stock cultures. These findings can be corroborated by unpublished
data by F. J. Sonleither who, using the earlier generations 12 and 13 of the selection
controls at density 12.5/g, found that the ++ control had a mean of 30.5 days while the bb
controls had a mean of 29.3 days. Comparable figures obtained by him for stock cultures
were 29.8 days versus 28.5 days.
Conclusions: This experiment is unable to explain the reversion of survival of ++
and bb in the selection experiment. This may be due to the difference in environmental
conditions in the jars as contrasted with the vials of this experiment. This point is now
being investigated. As for length of developmental period, the genotype-density interation
evident for the selection controls in Figure 2 and matching experiences in stock cultures
by Sokal and Karten (1964) and Sokal (1967) may be able to explain the longer
developmental period of bb in the selection experiment, although the differences
observed there are far greater than the difference of about two days noted in this
experiment.
Of general interest are the overall differences in survival and developmental period
between the stock cultures and the selection controls. The selection controls had been
subjected to a pattern of stock-keeping in which adults from a culture were permitted to
oviposit for three days, then removed from the culture and the eggs reared until almost all
of the adults had emerged. This has apparently resulted in inadvertent selection for long
developmental periods. No effort was made to use only the earliest emerging beetles as
progenitors, such as might be done in a Drosophila experiment where the investigator is
eager to carry out as many generations as possible. Such selection might have led to
short developmental period (see Hunter, 1959, for a striking example). In our selection
experiment, the most successful beetles were those which remained as relatively small
larvae for a considerable period of time, not pupating until most other pupae had already
done so. Thus, their chances of being cannibalized were minimized. Selection for such
slow-developing larvae would, of course, retard the mean developmental period of the
entire strain, which appears to have taken place both in the wild type ad bb selection
controls (also in hybrid strains not reported on here). The lower overall survival of the
selection controls vis0a0vis the stock cultures may simply be due to their longer
developmental period, during which they are exposed to more vicissitudes of the
environment or may reflect that more of the earlier pupae are cannibalized by the larger
numbers of remaining larvae in the selection controls.
Contribution No.1352 from the Department of Entomology, The University of
Kansas. This is paper No. 6 in a series on the ecological genetics of Tribolium. Nos. 1
through 5 are Schlager (1963), Sokal and Huber (1963), Sokal and Karten (1964), Karten
(1965), and Sokal (1967), respectively. This research was supported by the National
Science Foundation under grant GB-2170 and by a Public Health Research Career
Program Award (No. 3-K3-GM-22, 021-O1S1) of the National Institute of General Medical
Sciences. The technical assistance of M. Young-chen Chang, Mrs. Maxine L. Howe, and
Mrs. Cornella B. Tollefson are very much appreciated.
Literature Cited
Hunter, P. E. 1959. Selection of Drosophila melanogaster for length of larval period.
Zeitschrift fur Vererbungslehre 90:7028.
Karten, I. 1965. Genetic differences and conditioning in Tribolium castaneum.
Physiological Zoology 38:69-7S.
Schlager, G. 1963. The ecological genetics of the mutant sooty in populations of
Tribolium castaneum. Evolution 17:254-273.
Sokal, R.R. 1967. The effects of synchrony of egg batches on fitness characters and
Competition in Tribolium castaneum TIB 10:135-141.
Sokal, R. R. ad I. Huber. 1963. Competition among genotypes in Tribolium castaneum
At varying densities and gene frequencies (the sooty locus). American
Naturalist 97:169-184.
Figure 1. Adult survival expressed as degrees (angular transformation of percentage of
Egg input) averaged over the replicates and experiments of the study.
Results are shown for the four strains tested at the four densities.
Figure 2. Mean developmental period in days averaged over the replicates and
Experiments of the study. Results are shown for the four strains tested
At the four densities.
Sokal, R. R. and I. Karten. 1964. Competition among genotypes in Tribolium castaneum
at varying densities and gene frequencies (the black locus).
Genetics 49:195-211.
Sokal, R. R. and F. J. Sonleitner. 1965. Components of selection in Tribolium
(Coleoptera) And houseflies. Proceedings of the XII International Congress of
Entomology, 274-275.
SOKOLOFF, A
Department of Genetics
University of California, Berkeley
And Natural Sciences Division,
California State College,
San Bernardino
*Preliminary population studies with mutants of Tribolium Castaneum Herbst. I. The
paddle gene.
Note: This and the following experiments in this series were performed a few
years back. They had to be discontinued either because the incubator became an
incinerator; the populations became infected with protozoan parasites; or because while
moving from one part of the country to another constant conditions could not be
maintained. In all the series of experiments reported below the beetles were introduced
into four ounce glass jars containing 50 gras of standard medium (whole wheat flour
enriched with brewer’s yeast in a proportion 19:1). The lids had punched out centers
replaced with silk bolting cloth to allow free transpiration and thus prevent accumulation of
moisture and development of mold. All jars were kept in a 290 C, 70 per cent R. H.
incubator. The medium was renewed every two weeks. Censuses were taken several
months apart. Live beetles of all stages were returned to their jars after census. Both
dead and live beetles were scored, the latter after being etherized for 5-10 minutes. The
live beetles were returned to their respective jars and the dead ones discarded after
counting. Thus, it is possible that adults represented cohorts of several overlapping
generations.
It may be pointed out that where populations are large, the computations of
gene frequency give essentially the same results when taking into consideration only the
live beetles as when both live and dead beetles are summed to produce a larger sample.
This conclusion is particularly valid for the pearl gene, which is peculiar in that its adaptive
value is about equal (or sometimes higher) than its wild type allele.
Since these experiments were carried out some of these mutants have been used
in populations where artificially discrete generations of a month have been created. (See,
for example, Sokoloff, 1964, Proc. Ent. Congress, London, 1964). This is a much more
satisfactory method permitting the accumulation of 12 generations per year, which is
about what would be obtained in the case of Drosophila melanogaster populations. The
data are presented with little comment, in the hopes that they may help others planning
similar experiments.
The founding population of each of the six replicates involving pd consisted of 25 +
, 25 + , 25 pd , 25 pd . The data are given in Table 1, where wild type and paddle
progeny, whether living or dead, are given according to sex. The gene frequencies,
obtained directly from the males, and estimated for the females are also given. It is clear
that for the short period of observation there is no consistent trend, male’s frequencies
sometimes exceed those of females and vice versa. It is clear, however, that the gene
frequencies obtained for the two sexes are clearly different.
The pd gene is not very useful if one wants to identify the genotype or phenotype
of dead males and females, because the antennae and/or tarsi often break off.
This work was supported in part by USPHS grant GM 08942.
SOKOLOFF, A.
Department of Genetics
University of California, Berkeley
And Natural Sciences Division
California State College, San Bernardino
Preliminary population studies with mutants of Tribolium castaneum Herbst. II. The black
gene
Two allelic mutations were available: the black mutant derived from the Chicago
wild type strain, and the black mutant derived from the McGill wild type strain. Eight
replicate populations were set up with 25 +/+ ; 25 +/+ (Chicago wild) and 25 b/b ; 25
b/b (derived from Chicago +/+). Eight replicate populations were set up with 25 +/+ ,
25 +/+ (Chicago wild) and 25 b/b ; 25 b/b derived from the much more productive
McGill wild type strain (Sokoloff, Shrode and Bywaters, 1965? Phys. Zool). The data as
well as estimated and real frequencies are given in Tables 1 and 2, respectively. Half of
the replicates in each had to be discontinued at the end of three months because of
incubator failure, and the last two observations 17 and 23 months after the experiments
were begun, were made after the populations had been taken out of the incubator and
subjected to room and lower temperatures while they were being transported across the
country. The gene frequencies (estimated and real) of the two populations are, however,
initially different, the McGill black being greater than the Chicago black. At the end of two
years, however, the Chicago black populations consist of black at a gene frequency
between 0.40 and 0.50 while the McGill black populations ended up with a gene
frequency between 0.10 and 0.40.
Aside from these differences it is clear that the two populations differ in:
(1) Density (i.e. number of adults observed at census).
(2) Mortality.
This work was supported in part by USPHS grant GM 08942.
SOKOLOF, A.
Department of Genetics
University of California, Berkeley
And Natural Sciences Division
California State College, San Bernardino
Preliminary population studies with mutants of Tribolium castaneum Herbst. III. The jet
gene.
The autosomal recessive body color gene jet, and the Chicago wild type (from
which it was originally derived) were introduced in equal numbers in regard to sex and
genotype (25 + ; 25 + ; 25 j ; 25 j ). Half of the cultures were continued for 23 months
in the same manner as the other populations. The data and gene frequencies are
summarized in
Table 1.
This work was supported in part by USPHS grant GM 08942.
Table 1.
Phenotype and changes in gene frequency of jet. (The founding population
Consisted of 25 + , 25 + : 25 j , 25 j .)
Adults found
Replicate
Months
later
live
+/+
dead
j/j
live
dead
Estimated
gene frequency
1
3
9
17
23
923
699
565
321
95
383
741
359
164
143
121
34
12
58
153
37
.3883
.4121
.4200
.3952
2
2
10
1411
1071
98
1242
333
258
28
238
.4369
.4406
3
3
9
17
928
754
576
87
505
840
187
166
107
12
89
165
.4095
.4247
.3959
23
589
815
72
75
.3300
2
2
8
1341
1042
1234
180
43
429
321
388
384
83
12
90
.4893
.4209
.4871
5
3
9
17
23
754
547
520
376
80
380
658
448
215
166
136
45
27
85
165
71
.4711
.4825
.4553
.3270
6
3
1277
120
367
15
.4724
7
3
9
17
23
871
676
544
380
79
471
696
948
247
215
106
33
15
134
211
85
.4700
.4912
.4039
.2826
8
2
10
1642
1316
72
1393
475
291
22
360
.4703
.4702
4
4 repeat
SOKOLOFF, A.
Department of Genetics
University of California, Berkeley
And Natural Sciences Division
California State College, San Bernardino
Preliminary population studies with mutants of Tribolium castaneum Herbst.IV. The pearl
gene.
In these experiments the gene ws introduced as homozygotes and as
heterozygotes according to the following scheme:
Set 1
+ /+
M
F
50
50
50
50
50
--
50
49
45
25
--50
Set 2___________
p/p
M
F
+/p
M
F
M
+/+
-----50
-1
5
25
50
--
---50
25
--
1
10
50
50
-25
50
50
50
--50
F
p/p
M
F
49
40
--50
--
----25
--
-----25
-----
25
5
1
--
50
50
50
50
25
45
49
50
---
10
1
50
--
---
-50
40
49
The results are summarized in Tables 1 and 2. Essentially the results show that in
most of the replicates set up the frequency of the pearl gene remains as that introduced
almost ad infinitum.
Table 1.
Phenotype and gene grequency of pearl in populations with varying
Initial gene frequencies of pearl introduced as homozygotes.
Adults found
Founders
Months
elapsed
+/+
live
dead
p/p
live
dead
A 50 + ; 50 +
1
3
849
136
---
---
2
2 repeat
3
3
1367
1343
47
78
-----
-----
3
3
844
161
---
---
4
4 repeat
3
3
1665
1051
71
70
-----
-----
Gene
frequency
B 49 +/+ ; 1 p/p ; 50 +/+
1
3
807
165
---
---
2
3
4
6
1653
1419
1255
56
354
643
1
-----
1
1
4
. .02408
.02366
.04584
3
3
883
142
1
---
.03122
4
3
6
1425
941
40
81
--1
-----
.03125
942
1508
101
55
3
3
--1
.05355
.05053
C 45 +/+ ; 5 p/p ; 50 +/+
1
2
3
3
4
1262
124
2
2
.05364
3
3
847
148
4
---
.06328
4
3
4
1565
1386
47
205
5
3
--3
.05561
.06129
D 25 +/+ ; 25 p/p; 50 +/+
E
1
3
981
138
24
---
.1449
2
2
4
1229
1615
50
146
24
49
1
13
.1385
.1844
3
3
997
124
25
.1477
4
2
5
1271
1512
43
189
25
40
13
.1366
.1738
25 + ; 25 p ; 25 + ; 25 p
1
2
3
3
5
6
14
954
1016
1194
1365
1058
79
76
135
345
1832
135
298
346
391
297
16
35
43
76
326
.3571
.4834
.4758
.4631
.4211
3
3
3 repeat 3
7
893
846
1177
107
--318
109
208
214
12
--85
.3285
.4442
.4095
4
3
4
4 repeat 3
9
1122
1164
946
796
62
97
128
855
367
378
255
173
16
44
53
191
.4917
.5007
.4730
.4250
5
747
102
135
12
.3841
6
2 1111
4 1151
6 1348
9 1139
14 760
43
128
507
774
872
304
310
361
340
220
15
40
124
177
194
.4653
.4636
.4553
.4613
.4498
7
3
9
73
428
124
121
11
49
.3723
.3723
3
766
740
17
22
7 repeat
8
8 repeat
632
240
814
---
105
40
175
---
.3955
.3793
3
7
736
1352
--294
206
288
--54
.4677
.4147
2
4
2
5
10
961
915
884
1184
1032
66
106
57
456
858
329
279
266
277
238
21
43
19
130
178
.5041
.4897
.4822
.4459
,4247
F 25 +/+ ; 25 p/p ; 50 p/p
1
3
652
53
244
25
.5255
2
2
4
6
642
736
675
35
59
520
489
553
495
29
59
509
.6584
.6595
.6767
3
3
653
54
257
29
.5367
4
3
4
6
819
360
1252
51
457
89
551
114
543
32
380
35
.7080
.6139
.5488
G 5 +/+ ; 45 p/p ; 50 +/+
1
3
208
26
558
82
.8557
2
2
4
129
139
27
13
978
53
1015 132
.9320
.9397
3
3
124
11
438
84
.8913
4
2
4
106
151
3
30
709
929
52
184
.9353
.9274
H 1 +/+ ; 49 p/p ; 50 p/p
1
3
11
4
694
96
.9814
2
2
4
17
24
--- 1049
8 1083
39
148
.9821
.9873
3
3
28
7
128
.9789
674
4
2
4
10
20
29
41
--- 989
3 1122
17
972
25
227
916
I 50 p/p ; 50 p/p
.9903
.9883
.9850
Pearl frequency
1
3
---
---
610
148
1.0
2
2
4
-----
-----
1034
1122
44
219
1.0
1.0
3
3
---
---
469
105
1.0
4
2
4
-----
-----
1072
869
54
262
Table 2.
Phenotype and gene frequency of pearl in populations with arious
Initial gene frequencies of pearl introduced as heterozygotes.
Founders
Months
later
live
Adults found
+/+
p/p
dead
Live
dead
Pearl
frequency
J 1 +/p ; 49 +/+ ; 50 +/+
1
3
926
150
0
0
2
3
4
6
1342
1285
1321
45
228
485
0
0
0
0
0
0
3
3
989
173
0
0
4
2
4
1111
1562
38
248
0
0
0
0
136
0
0
K 1 +/p ; 49 +/+ : 1 +/p ; 49 +/+
1
3
988
?
2
3
5
7
1170
1568
1531
33
245
93
2
3
1
1
0
.04074
.04691
,02481
3
3
921
140
0
0
?
4
3
4
6
1148
1553
1296
64
191
673
0
0
1
0
0
0
?
?
.02254
L 5 +/p ; 45 +/+ ; 5 +/p ; 45 +/+
Live
mutant
dead
Gene
frequency
1
3
1038
117
0
0
?
2
2
4
6
1082
1331
1358
48
298
425
2
4
5
0
5
3
.04204
.07412
.06684
3
3
903
132
0
0
?
4
3
3
6
1398
1736
1488
41
218
659
8
13
12
0
2
4
.07439
.08728
.08801
M. 25 +/p ; 25 +/+ ; 25 +/p ; 25 +/+
1
3
1000
85
69
4
.2511
2
3
4
1406
1543
69
186
90
104
8
10
.2516
.2487
3
3
976
114
81
5
.2708
4
2
5
6
1060
1431
986
28
215
547
70
103
78
2
5
29
.2626
.2481
.2554
N 25 +/p ; 25 p/p ; 25 +/p ; 25 p/p
1
3
741
77
360
39
.5719
2
3
5
599
590
37
146
493
414
33
131
.6728
.6522
7
858
426
517
238
.6085
3
3
531
82
364
36
.6284
4
3
5
6
708
599
632
29
117
266
637
532
522
29
150
212
.6889
.6984
.6780
O 5 +/p ; 45 p/p ; 5 +/p ; 45 p/p
1
3
112
13
427
85
.8965
2
2
4
177
153
6
23
1033
1025
43
188
.9244
.9345
3
3
181
22
497
86
.8612
4
3
5
181
195
11
31
994
1071
72
147
,9205
.9184
P 1 +/p ; 49 p/p ; 1 +/p ; 49 p/p
1
3
16
3
555
146
.9867
2
2
4
24
24
2
8
979
1119
49
343
.9876
.9892
3
3
65
6
562
106
.9507
4
2
4
6
10
18
25
2
3
6
872
1028
1062
43
208
508
.9935
.9916
.9903
Q 1 +/p ; 49 p/p ; 50 p/p
1
3
8
0
631
150
.9949
2
2
4
12
22
0
6
841
996
38
378
.9932
.9899
3
3
5
2
656
168
.9958
4
2
4
8
7
1
1
1077
1062
47
198
.9960
.9968
SOKOLOFF, A.
Department of Genetics
University of California, Berkeley
And Natural Sciences Division
California State College, San Bernardino
Preliminary population studies with mutants of Latheticus Oryzae Watern. I. The pearl
gene.
These experiments were designed to test, in a comparative way, the performance
of pearl in populations. For this reason the founders in the various sets were introduced
in a manner similar to the experiment reported above for Tribolium castaneum. The first
notable difference between the twospecies is that the number of adults produced by
Latheticus is far lower than that obtained in Tribolium cultures. Furthermore, the
developmental period of Latheticus in these cultures is astonishingly slow. Cultures set
up six to eight months before may contain only the original adults, the progeny being in
the late larva or in the pupa stage. The cultures appear as if the larvae are all of the
same age, and they remain thus for a long period of time. Eggs and small larvae
apparently are eaten by the older larvae as they are produced. At the other end, the first
few pupae forming are destroyed by the younger larvae. The small size of the
populations as well as the long developmental period makes it impractical to pursue
population studies with this organism.
The data, summarized in Tables 1 and 2, insofar as they go, appear to indicate
that when pearl is frequent and wild type infrequent the results in the two species are
comparable.
Table 1.
Frequency changes of pearl in Latheticus oryzae introduced initially
At frequencies from 0-1.0 into each of six replicates as homozygotes.
Adults found
Founders
A
Months
later
+/+
p/p
live
dead
live
dead
Gene
frequency
50 + ; 50 :
1
8
114
29
---
---
0
2
6
11
124
218
44
176
-----
-----
0
0
3
4
8
7
11
93
188
241
7
92
179
-------
-------
0
0
0
5
8
93
7
---
---
0
6
7
11
136
190
91
178
-----
-----
0
0
B 49 + ; 1 p/p ; 50 +
1
8
111
40
1
---
2
5
7
74
231
31
39
1
---
-----
3
8
102
18
1
---
4
7
11
186
166
66
162
1
0
--2
5
8
96
4
2
---
6
7
11
124
256
55
105
1
---
--1
C 45 +/+ ; 5 p/p ; 50 +
1
8
99
11
6
---
2
5
7
72
204
23
68
5
9
--2
3
8
101
18
4
1
4
7
11
112
209
79
152
4
2
--3
5
8
89
7
5
---
6
7
11
142
174
92
177
4
3
1
2
D 25 +/+ ; 25 p/p ; 50 +/+
1
8
85
32
25
---
2
5
67
6
25
---
.1972
.1189
..1185
7
131
131
21
12
.3345
3
8
70
20
26
---
4
7
11
66
175
43
95
21
22
4
8
5
8
73
9
26
---
6
7
11
200
178
96
97
21
21
2
13
.3317
.3162
E 25 +/+ ; 25 p/p ; 25 +/+ ; 25 p/p
1
8
20
26
55
47
85
17
19
56
49
34
33
--5
36
.6364
.6094
.5732
2
5
7
32
194
18
61
41
37
9
5
.3761
3
8
20
26
55
19
0
7
33
15
50
23
13
--7
27
.6681
.6984
.8528
4
7
11
34
74
16
76
47
37
2
25
.5408
5
8
20
26
59
23
54
11
18
37
48
20
32
2
5
19
.6455
.6155
.5993
6
7
11
113
163
98
74
41
29
5
20
.4231
.4139
F 50 +/+ ; 50 p/p
1
8
61
20
47
1
2
5
7
46
145
4
65
44
35
6
9
3
8
68
21
48
2
4
7
11
87
222
43
105
35
11
11
33
.3444
5
8
56
10
45
4
6
7
11
94
221
79
111
41
31
10
25
.3799
G 25 +/+ ; 25 p/p ; 50 p/p
1
8
30
5
82
10
2
6
8
21
102
4
14
63
164
12
32
3
8
29
4
71
7
4
7
11
49
102
42
65
71
144
51
62
5
8
25
4
75
7
6
7
11
54
46
27
56
102
75
37
80
.7926
.7432
.7766
H 5 +/+ ; 45 p/p ; 50 p/p
1
8
4
4
97
7
2
5
7
3
10
2
2
75
194
20
60
3
8
10
4
92
8
4
7
11
13
15
6
10
121
204
52
85
5
8
5
---
90
6
6
7
11
6
6
2
5
84
145
27
86
I 1 +/+ ; 49 p/p ; 50 p/p
1
8
1
---
96
15
2
5
1
---
86
13
,9772
.9594
.9770
7
3
2
150
76
3
8
1
---
95
11
4
7
11
-----
1
---
88
282
15
133
5
8
2
---
98
10
6
7
11
1
2
--- 79
1 108
20
85
.9891
.9923
J 50 p/p ; 50 p/p
1
8
---
---
101
18
2
7
11
-----
-----
131
229
40
116
3
contaminated with wild type
4
7
11
-----
-----
85
202
23
82
5
8
---
---
96
5
6
7
11
-----
-----
92
216
48
128
Table 2.
Phenotype and gene frequency of pearl in populations with various
Gene frequencies of pearl introduced as heterozygotes
Adults found
Founders
Months
later
+/+
live
p/p
dead
live
dead
K 1 +/p ; 49 +/+ ; 50 +/+
1
8
101
10
---
---
2
7
173
92
2
---
Gene
frequency
11
257
130
---
1
3
8
100
9
---
---
4
5
9
104
278
19
101
-----
-----
5
8
101
5
---
---
6
5
9
116
207
24
152
-----
-----
.05075
L 10 +/p ; 40 +/+ ; 50 +/+
1
8
93
7
1
---
2
7
11
94
206
11
164
--2
-----
3
8
102
15
---
---
4
5
9
87
181
13
97
-----
-----
5
8
97
8
---
---
6
5
9
90
207
9
130
1
1
-----
.07332
.05439
M 50 +/p ; 50 +/+
1
4
97
4
1
---
2
4
8
94
152
5
90
1
6
--1
5
97
2
1
---
3
.1676
Q 25 +/p ; 25 p/p ; 50 +/+
1
6
75
6
26
2
5
9
64
91
9
55
25
19
6
80
8
25
3
6
.3824
4
5
9
69
129
4
108
23
14
1
8
,2914
5
6
66
6
25
6
5
9
73
178
2
91
20
19
5
9
.3071
S 10 +/p ; 40 p/p ; 50 p/p
1
6
10
---
85
7
2
5
9
9
12
1
8
82
96
7
62
3
6
10
---
90
5
4
5
9
9
17
--10
83
83
7
130
5
6
10
2
86
6
6
5
9
10
18
--6
86
118
4
89
.9421
.9421
.9466
T 1 +/p ; 49 p/p : 50 p/p
1
6
1
3
97
---
2
5
9
1
2
--1
97
151
22
109
3
6
2
---
97
3
4
5
9
1
2
-----
90
176
8
75
5
6
2
---
96
3
6
5
1
---
87
66
97
59
3
39
-----
-----
M 50 +/p ; 50 +/+
4
5
9
.9943
.9960
.9967
5
5
98
3
---
---
6
4
8
95
172
3
106
2
9
-----
.1771
N 50 +/p ; 50 +/p
1
5
98
2
---
---
2
4
8
95
181
5
90
--47
--9
3
5
98
2
---
---
4
4
8
95
114
3
69
1
16
--6
5
5
99
3
---
1
6
4
8
99
71
--1
1
28
-----
.4139
.3491
P 50 +/+ ; 25 +/p ; 25 p/p
1
6
74
8
24
1
2
5
9
72
184
4
123
23
10
1
15
3
6
76
4
25
1
4
5
9
59
150
16
113
19
5
6
14
5
6
77
5
24
1
6
5
9
70
163
5
137
22
6
3
23
.2744
.2858
.2967
SOKOLOFF, A., M. ACKERMANN AND B. HEINZE
Department of Genetics
University of California,
Berkeley, California
*Additions to established linkage groups
I.
T. castaneum
A. X-chromosome
1. Lethal-5 (1-5), located about 25 units to the left of py (away from r ).
Allelic with 12 and 14.
2. Letal-6 (1-6), located about 12 units to the left of py (away from r ).
3. Lethal-7 (1-7), located about 20 units to the left of py (away from r ).
B. Autosomes
Aureate is located about 42 units away from black. Three-point crosses
To locate it in respect to other gees are now in progress.
II.
T. confusum
A. X-chromosome
1. Alate prothorax, apt, is between es lt and lp, about four units to the
left of lp.
This work was supported by USPHS grant GM 08942.
SOKOLOFF, A., N. INOUYE AND R. S. ST. HITAIRE
Department of Genetics
University of California, Berkeley
And Natural Sciences Division,
California State College , San Bernardino
*Additional sex-linked lethal in Tribolium castaneum Herbst.
In an experiment designed to determine the frequency of lethal as a regression on
the age of the maternal grandfather (see Lerner and Inouye in the present issue of TIB)
several females gave aberrant sex-ratios and two of these, on retesting, proved to be
heterozygous for lethal. The material was originally derived from the Berkeley synthetic
strain marked with sooty (for details on its construction and maintenance see Lerner and
Ho. 1961, Am. Nat. 95:329).
In order to locate these lethal and determine possible allelism, four virgin females
from stock 29c and eight from stock 68a were mated with py r males. Because of lack of
time it was not possible to set up the sequential matings immediately with a consequent
over lap in generations. Female virgins were tan mated with py r males, four from 29c
and eight from 68a. In the former, one female proved to be heterozygous for the lethal
producing 1+ and 4 py r males, and 6 + and 4 py r females. The “+” virgin females (py r
+/+ + 1x) were placed in individual creamers and remated with py r males, allowed to lay
eggs for a week and transferred to fresh medium four times at intervals of a week to
increase the number of progeny. From 68a two females designated as 68a-1 and 68a-6
and producing 9 py r : 14+, 1 py, 6 py r , and 3+, 1 py, 13 py r : 18 +, 1 r, 1 py, 10 py
r , respectively, were the source of the carriers of the other lethal. In 68a-1 eight “+”
females and in 68a-6 twelve “+” females were heterozygous for the lethal. These females
were separated in individual creamers and allowed to remain with their progeny until the
latter emerged as adults.
Results and Conclusion
Experiment 29c
The various broods in experiment 29 c have been tested and found homogeneous.
Therefore, the data have been pooled and they are shown in Table 1. They make it
possible to determine that the crossover frequency for the three genes involved is :
Males
r - py
44/425 = 10.35
r – 1 29c
136/425 = 32.00
py – 1 29c
104/425 = 24.47
Females
78/917 = 8.51
Therefore, the order is 1 s9c – py – r. Previous studies have located 1s at 22.78
units from py (from 90/395 crossovers detected between py and 12) and 14 at 25.45 units
from the same gene (from 70/275 crossovers between py- and 14). 12 and 14 have been
found allelic (Sokoloff and Dawson, 1963, Can. J. Genet. Cytol. 5:138). Chi square tests
of homogeneity between 1s, 14 and 129c indicate the data are homogeneous. Therefore,
129c must be considered as a recurrence of 12 and it is designated as 1-5. (cf. section on
New Mutants.)
Tables 2 and 3 summarize the data for experiment 68.
The data for 68a-1 indicate the crossover frequencies are:
py - r
Males
Females
47/398 = 11.81
92/613 = 15.01
r – lethal
87/398 = 21.86
py – lethal
46/398 = 11.56
so the order is : lethal – py – r.
The data for 68a-6 give the following crossover values:
Males
py - r
30/416 = 7.21
r - lethal
99/416 = 23.80
py - lethal
81/416 = 19.47
Females
84/822 = 10.22
and the order of the three genes is lethal - py - r.
68a-1 and 68a-6 have been tested for allelism by the Chi square that for homogeneity
with 12, 14 and 15. The statistical tests suggest these lethal are not allelic with them nor
with each other. Neither are they allelic with 13. Therefore, 68a-1 is redesignated 16 and
68a-6 17.
We do not know whether the original female whose progeny indicated the
presence of a sex-linked lethal carried both lethal or whether a second lethal occurred
during the course of these experiments.
This work was supported by USPHS grant GM 08942.
Table 1. Progeny of py r +/++ 29c x py r +/.
Phenotype
Male
Female
py r
283
266
++
98
573
py
38
38
6
40
425
917
r
Total
Table 2. Progeny of py r +/++ 68a-1 x py r + (nine successful creamers)
Phenotype
Male
Female
308
216
++
43
305
py
44
56
3
36
398
613
py r
r
Total
Table 3. Progeny of py r +/++ 68a-6 x py r +/. (12 successful creamers)
Phenotype
Male
Female
py r
311
268
++
py
r
75
24
6
470
36
48
416
822
Total
SOKOLOFF, A AND J. LANTER
Department of Genetics
University of California, Berkeley
And Natural Sciences Division
California State College, San Bernardino
Further studies of productivity of Tribolium castaneum and Tribolium confusum in homoand heterospecific matings
In a previous paper (Sokoloff, Shrode and Bywaters, 1965, Phys. Zool. 38:165), it
was shown that productivity (defined as the number of adult progeny per female within a
specific interval or the number of fertile eggs per female surviving to the adult stage) is a
reliable genetic character. It was used in a previous study by Sokoloff and Inouye (TIB
6:61) to determine whether females of T. castaneum (CS) or T. confusum (CF) are
affected in their reproductive capacities by the presence of males of the other species. It
was found that when CF females are in association with CS males their productivity drops
somewhat, but not as much as when CS females their productivity drops somewhat, but
not as much as when CS females are in creamers together with CF males. This previous
experiment left unanswered the question whether this drop in productivity, observed
particularly for CS, was not the result of depriving the females from their mating partners.
Tagarro and Rico (TIB 9:12) showed that when males are removed from cultures where
females are already fecundated, they continue to lay fertile eggs at a slightly lower rate
than when males were present, but that this difference is not significant. The period they
tested was between days 7 and 11 after the females had eclosed. Since in the Sokoloff
and Inouye experiments the females were left without males for a much longer period, the
present experiments were performed.
For each of the species or species combination the experimental set up was as
follows: into each of 10 creamers were introduced four pairs of beetles 10 days old
(females and males had been isolated as pupae) on a Thursday. At the same time on
Monday the imagoes were transferred into new containers, the old medium with eggs
being returned to the original creamers after counting the eggs. The adults were moved
to new quarters every twenty-four hours for three days, and the eggs were counted before
returning them to their respective creamers. Every week the beetles were manipulated in
the following way: the first week the females in all 10 creamers had males of the same
species as partners. The second week the set was broken into two subsets. From
subset A the males were removed altogether, and these females are referred as
“widows”. From subset B half of the males were removed and replaced by males of the
other species. We refer to these females as “bigamists”. On the third week the females
of subset A are supplied with males of the other species, and refer to them as
“miscegynists, type I”. From subset B the remaining two males of the same species are
removed and replaced by males of the other species, and the females referred to as
“miscegynists, type II”. Finally, on the fourth week, the foreign males are removed from
both subsets and males of the same species re-introduced. (The two subsets will be
referred to as “repurified I” and “repurified II”, respectively.)
If any dead beetles were found, they were sexed and replaced by beetles of the
same age, sex and species.
Fertility of the eggs was determined by counting larvae when they were three
weeks old.
The data have been analyzed to determine any significant difference between the
three successive 24-hour periods. Since no significant difference has been found, the
values of the three successive days have been pooled providing 30 observations for the
pure species (10 replicates x 3 days) and 15 for the subsets (5 replicates x 3 days). The
results are summarized in Table 1.
Comparisons of the means by t-test indicates that none of the differences
observed for CF are significant. For CS significant differences can be shown at the .02
level for the means of egs produced by “pure species” and “widows” and at the .01 level
for “pure species” vs. “miscegynists-I”, “miscegynists-II”, and “repurified”. Significant
differences are obtained at the .01 level for fertility of eggs of “pure species” vs.
“bigamists”, “miscegynists-I” and “miscegynists-II”. The values obtained for “pure
species” and “re-purified II” are significantly different at the .02 level.
This experiment appears to indicate that when inseminated CF females are
introduced with CS males either they reject them, or if copulation takes place between
them, the foreign sperm play no role in fertilization of CF eggs. On the other hand, if CS
females are confined with CF males (and no CS males are present) they inseminate them
, and the foreign sperm may fertilize a fairly large number or all of CS eggs. The result is
that females exhibit partial or complete sterility. These sterile or semisterile CS females
recover almost immediately from any “damage” the foreign males may have produced if
CS males are reintroduced.
Therefore, it is concluded that in mixed species cultures heterospecific matings
should not influence greatly the outcome of competition if the numbers of the two species
are large. Where the numbers of CS are small (and CF large), the absence of sufficient
CS males to service the CS females present might result in the loss of CS from those
cultures.
This work was supported by USPHS grant GM-08942.
TAGARRO, MA. P. AD F. OROZCO
Laboratorio de Genetica de Poblaciones
Instituto Nacional de Investigaciones Agronomicas
Madrid, Spain
Influence of collecting frequency on egg-laying rate of fecundated and virgin females in
Tribolium castaneum
Egg-laying rate is the quantitative character we are using more in our genetic
research with Tribolium castaneum. We have defined a period comprised between the 7 th
and the 11th days after adult emergence to measure that character.
Egg collecting and counting is a time-consuming task, so we were interested to
reduce as much as possible the number of collections in such testing period. At 330 C
and 70% RH larvae hatch 66 + 6 hours after the eggs are laid. So with fecundated
females it is necessary to collect no later than every other day (two collections in the fourday period). With virgin females this problem does not exist, so it is possible to collect
once at the end of the testing period.
However, we wondered whether collecting every day, every other day, or at the
end of the four-day period could have some influence in the total number of eggs laid or
counted. Favorable effect of fresh medium after a collection, disturbing effect of the
suction sifting, cannibalism, etc., could contribute to find different figures in the total lay of
the four days.
Four trials were run comparing 24 vs 48 hours collecting frequencies both with
fecundated and with virgin females. Two more trials were run to compare 48 vs. 96 hours
collecting frequencies only with virgin females. We used our “Consejo” strain of Tribolium
castaneum. Fecundated females were mated at adult emergence and the males (one per
female) were maintained during the testing period. At day 7th after emergence the
medium in each vial was changed. Egg collecting was made by suction sifting every day,
every other day or at day 11, according to the corresponding treatment, and afterwards
fresh medium was added. The figure analyzed was the total number of eggs laid per
female during the four-day testing period.
Table 1 includes the analysis of each of the four experiments run with fecundated
females comparing 24 vs. 48 hours collecting frequencies, together with a pooled analysis
of the four as a whole. We can see that in no case there exist differences between
treatments.
Table 2 includes similar information as in Table 1 but with the data obtained with
virgin females. The results in this case are not as clear as with fecundated females
because in the A trial we find significant differences to the .05 level in favor of 48 h. and in
the B one slightly significant (only to .10 level) but in favor of the 24 h. frequency.
However, when considering the pooled analysis we do not find significant differences to
any level between treatments and the significance for the interaction reflects the
difference of results obtained in A and B. The greater influence of uncontrolled effects in
egg laying of virgin females as compared with the lay of fecundated ones, widely
observed by us in many experiments, could explain the anomalous results of A. In a
practical sense we can consider the non-significant differences found for treatments in the
pooled analysis as favorable.
Table 3 contains the analysis of the two experiments comparing 48 vs. 96 hours collecting
frequencies with virgin females and the pooled analysis of both.
Table 1. Means, standard errors, df, MS and F values of the analysis of
Four experiments studying the influence of 24 vs. 48 hours
Collecting frequencies on the laying rate of fecundated females.
Experiment
Means and S. E.
A
24 h. 63.5 + 2.7
48 h. 65.5 + 2.7
B
Sources
df
MS
F
Treatments
Error
1
45
46.04
173.50
0.27
24 h. 76,9 + 3.6
48 h. 72.5 + 4.0
Treatments
Error
1
43
218.11
317.11
0.69
C
24 h. 34.4 + 2.9
48 h. 38.3 + 2.8
Treatments
Error
1
37
154.47
161.00
0.96
D
24 h. 61.0 + 3.3
48 h. 58.0 + 3.5
Treatments
Error
1
45
108.15
273.09
0.40
Pool
24 h. 60,4 + 1.6
48 h. 58.9 + 1.6
Experiments
Treatments
Exp. X Treat.
Error
3
1
3
170
10,881.46
110.06
138.90
233.47
46.61*
0.47
0.59
*Significant to .005 level.
Table 2. Means, standard errors, df, MS and F values of the analysis of
Four experiment studying the influence of 24 vs. 48 hours
Collecting frequencies on the laying rate of virgin females.
Experiment Means and S. E.
Sources
df
MS
F
252.01
38,04
6.62
A’
24 h. 9.8 + 1.1
48 h. 13.7 + 1.1
Treatments
Error
1
62
B’
24 h. 19.4 + 1.7
Treatments
1
243.95
3.77*
48 h. 14.9 + 1.6
Error
46
64.65
C’
24 h..
48 h.
9.3 + 1.5
7.9 + 1.5
Treatments
Error
1
38
21.03
44.17
0.48
D’
24 h.
48 h.
14.1 + 1.9
16.8 + 1.9
Treatments
Error
1
48
89.78
89.05
1.01
Pool
24 h.
48 h.
12.9 + 0.8
13.6 + 0.8
Experiments
Treatments
Exp. X Treat.
Error
3
1
3
194
635.57
25.59
193.73
58.17
10.93
0.44
3.33
*Significant to .10 level;
to 0.04 level; to .005 level.
Table 3. Means, standard errors, df, MS and F values of the analysis of
Two experiments studying the influence of 48 vs. 96 hours
Collecting frequencies on the laying rate of virgin females.
Experiments
Means and S. E.
Sources
df
MS
F
E
48 h. 12.8 + 1.2
96 h. 13.0 + 1.2
Treatments
Error
1
96
0.66
69.09
F
48 h. 28.4 + 2.3
96 h. 28.3 + 2.3
Treatments
Error
1
96
0.66
250.33
Pool
48 h. 20.6 + 1.3
96 h. 20.6 + 1.3
Experiments
1
Treatments
1
Exp. X Treat. 1
Error
192
11.694.88
0.00
1.30
159.71
0.01
0.00
73.23*
0.00
0.01
*Significant to .005 level.
No differences at all are found between treatments in any experiment nor in the
pooled analysis. This result also helps to interpret the figures in Table 2 in the sense that
the significant differences observed in A and B must be due to chance effects.
In all sets of trials differences between experiments are always significant. This is
not surprising because many uncontrolled effects influence the number of eggs laid by
females of Tribolium castaneum. Pooling the analysis of different trials is quite fair
because we have proportional or nearly proportional subclass numbers; analysis with
means of treatments x experiments cells give the same results reported here.
Therefore, we can conclude tht under our experimental conditions, when we are
interested to measure the egg-laying rate from day 7 to day 11 after adult emergence, it
does not make any difference whether we collect eggs every day or every other day in
both fecundated and virgin females and even in doing only one collection at the end of
the four days with virgin females. The longest “between collections” period with
fecundated females is 48 hours because larvae may hatch before three days.
This is a partial and preliminary study included in Grant No. FG-Sp-137 of P.L.
480, contract with the USDA.
TSCHINKEL, W.
Department of Biochemistry
University of California
Berkeley, California
*Sex pheromones and defensive secretions from Tenebrionid beetles
Studies on the sex pheromones and defensive secretions of Tenerbrionid beetles
are under way in this laboratory. Sex pheromones have been demonstrated in Tenebrio
molitor and Zophobas rugipes (a large Central American species) and a biological assay
for the pheromone of Tenebrio molitor has been devised (Tschinkel, Willson and
Bern,1967). Attempts to isolate the sex pheromone of Tenebrio molitor are in progress.
The secretion of the prothoracic defensive glands of Zophobas rugipes is being
studied by various chromatographic and spectral techniques. These glands secrete
phenols, but are otherwise homologous to the quinine-secreting prothoracic stink glands
of Tribolium (Roth, 1943).
Disturbed larvae of Zophobas rugipes frequently squirt hemolymph along with an
acrid odor. This phenomenon and an effect of larval crowding on pupation are under
study.
Attempts are being made to establish a number of Tenebrionid beetles (especially
subfamily Tenebrioninae) in laboratory cultures for comparative studies on their sex
pheromones and defensive secretions. Success has been limited. The results are
summarized in Table 1.
Literature Cited
Roth, L. M. 1943. Studies on the gaseous secretion of Tribolium confusum Duval. II. The
odoriferous glands of Tribolium confusum. Ann. Ent. Soc. Am. 36:397-424.
Tschinkel, W. R., C.D. Willson and H. A. Bern. 1967. Sex pheromone of the mealworm
beetle (Tenebrio molitor). J. Exp. Zool. 164:81-85.
WOOL, DAVID
Department of Entomology
The University of Kansas
Lawrence, Kansas
*Some observations on mating frequencies in Tribolium
Castaneum strains
Introduction. In a series of papers, Ehrman and associates (Ehrman, 1965, 1966;
Ehrman et al., 1965) described a phenomenon observed earlier by Petit (1958) with
important evolutionary implications. When two strains of Drosophila are mated in unequal
proportions, in some cases the rarer strain, regardless of genotype, will mate more often
than expected on the basis of its frequency in the observation chamber. The series of
experiments reported here was designed to look for a similar phenomenon in two strains
of Tribolium. In the course of the experiment, some interesting observations on the
mating habits of these strains were made, which are also reported here.
Materials and Methods. The standard UPF wild type and black strains of T.
castaneum employed in work in this laboratory (see stock list) served as experimental
material. Large numbers of pupae were recovered from multiple cultures of the two
stocks, sexed and then put, in unisexual groups of 10, in 6-dram holding vials containing
about 4g of flour. The bb beetles emerged two days later than the ++ adults and
therefore were two days younger throughout the experiment. The first experiment was
set up four days after the emergence of bb adults, and the last one two weeks later.
Mating was observed in two-inch Syracuse dishes with 20 and 20 in varying
genotypic proportions. All the males were marked by paint at least three days before the
experiment to distinguish true mating pairs from the frequent x mounts (see results).
The genotype of each sex and time of mating was recorded. Each observation dish was
watched constantly for two consecutive 30-minute periods, during which a considerable
number of copulations was observed. Two series of observations were made. In the first,
after the end of the fixed observation period, all the males were removed, and the females
were transferred individually to vials with about 4g of flour, where they were left to oviposit
for three days, after which they were discarded. The genotype of the was recorded on
the vial, and the resulting brood was classified by genotype to infer the type or types of
males that had sired it.
In the second series, the FF and MM were separated after the first 30 minutes’
observation period. The “old” MM were given a new group of virgin FF and the “old” FF,
a fresh group of (the previous genotypic proportions being maintained). The two
dishes were observed simultaneously for another 30 minutes, after which all the beetles
were discarded. No brood analysis was carried out in this series.
The first series was set up at proportions 10:90 and 25:75, with both wild type and
black being rare in turn. The second series was run at 10:90 only. Two 50:50
observations were carried out as controls.
Results. Previous to the main experiment, couples (in copula) were recovered
from large wild type and black populations and the sex of the participants determined by
microscopic examination. It appeared that wild type or black will mate with any
Tribolium available, regardless of sex. Mounting of MM on other MM occurred in 30.5%
of the cases among the wild type and 34.5% among the blacks (based on 127 pairs); a
test of independence yielded X2 = 0.277 and fails to show any difference in behavior
between the strains in this respect. In all subsequent experiments the males were
marked, and only true ( M x F) matings are considered in the following sections.
The observations in the Syracuse dishes may be summarized in the following form;
the asterisks refer to total X2 for all replicates.
The symbols (++) or (bb) following asterisks indicate the genotype that mated more
often. Significant heterogeneity among the replicates is shown by the superscript h.
All new x new cases were observed in the first 30-minute periods. All other combinations
were in the second 30-minute periods.
The mean number of copulations in 30 minutes for 10:90 proportions differed
greatly among the four combinations: new FF x new MM (21.7), new FF x old MM
(12.5), old FF x new MM (15.0), and old FF x old MM (6.75). Analyzing the variance
of the 26 replicates, the following table shows a highly significant mean square among
combinations.
Source of variation
Genotype of rare strains
Combinations
Interaction
Error
df
1
3
3
18
MS
1.78
248.26***
28.74
17.35
Single degree of freedom comparisons showed a highly significant difference
between the mean number of copulations between new FF x new MM and the mean of
the other three taken as a whole (P << .001), and a significant difference between old FF
x old MM and the two (old FF x new MM and new FF x old MM) combinations taken
together (P < .01). No significant difference was found between the mean of new FF x
old MM versus old FF x new MM
Similar results were obtained in the eight 25:75 replicates, which contained only
old FF x old MM and new FF x new MM combinations.
Surprisingly, the results of the brood analysis of the 10:90 proportions showed that
very few of the in the observation dish were fertilized although, based on the average
mating frequency, every one of the 20 could have been fertilized. (Similar results were
obtained in the 25:75 series.)
Rare strain
Number of matings
observed
Number of fertilized
females
++
25
29
6
7
bb
24
27
12
13
Control (50:50)
23
5
In a few cases, a F was mated in sequence to a heterotypic M and then a
homotypic M. Only 5 of 10 FF were fertilized by any of the males: 3 bb FF and 2 ++ FF.
The brood of the bb FF contained only the offspring of the first (++) MM. The brood of the
++FF contained mainly the first (bb) male’s offspring, although 3.5 – 8.5% of the offspring
were from the second mating.
Discussion. As can be seen from the results presented above, the Petit-Ehrman
phenomenon does not occur in these two strains of Tribolium under our test conditions.
However, it was found that black FF, when rare, consistently mated much more
frequently than expected, mostly with wild type males. In all cases where a significant
deviation from expected mating frequencies for MM or FF was present, the frequency of
++ MM x bb FF matings was very much higher than expected (P << .001 by the G-test).
This trend in the bb FF was consistent for all replicates. The only case in which
heterogeneity was significant for the FF was caused by an exceptionally large deviation
from expectation in one of the six replicates. No clear trend could be shown for the MM.
Such a behavior, if reflected in the offspring, could result in an increasing gene
frequency of black.
Virgin pairs showed the highest mating frequency per half hour. When both and
had mated in the half hour preceding the experiment, they showed the lowest number of
mating. In cases where one sex was virgin, while the other had mated in the half hour
preceding the experiment, the mating frequency ws in-between, with no significant
difference between the reciprocal crosses. Possibly mated may resist a subsequent
mating shortly after the first one, and the may need a period of rest between matings.
It is interesting to note that even in the old x new and new x old
combinations, the black females, when rare, mated significantly more often than
expected. Presumably they are unable to resist the males as effectively as the wild-type
females.
Why the number of fertilized females should be so much lower than the number of
observed copulations is obscure. When b was rare (wild-type common) more were
fertilized tan when wild type was rare. Some of the matings could have been infertile
because of the unusual conditions of light and substrate in the observation dish. By
contrast, when in another experiment 20 and 20 at proportions 10:90 and 50:50 were
left in flour for 12 hours, 18, 18 and 20 females were fertilized in the cases where + was
rare, the two alleles equally frequent, and b rare, respectively.
The brood analysis data are not helpful in testing the Petit-Ehrman phenomenon
because the evidence shows a considerable number of double matings (mating of one
female with more than one male). Regrettably, this can be demonstrated only if the
female mated with both types of males. Ignoring all evident double matings, no significant
higher mating frequency of the rare form can be shown.
Contribution No.1350 from the Department of Entomology, the University of
Kansas. This is paper No. 7 in a series on the ecological genetics of Tribolium. Numbers
1 through 5 are listed at the end of paper No.6 (Sokal, 1967) in this issue of TIB. This
research was supported by a National Science Foundation Grant GB-2170 (Robert R.
Sokal, principal investigator).
Literature Cited
Ehrman, L. 1965. Direct observation of sexual isolation between allopatric and
Sympatric strains of the different Drosophila paulistorum races.
Evolution 19:459-464.
Ehrman, L. 1966. Mating success and genotype frequency in Drosophila.
Animal Behavior 14:332-339.
Ehrman, L., B. Spassky, O. Pavlovsky, T. Dobzhansky. 1965. Sexual selection,
geotaxis, and chromosomal polymorphism in experimental populations of
Drosophila pseudoobscura. Evolution 19:337-346.
Petit, C. 1958. Le determinisme genetique et psycho-physiologique de la
Competition sexuelle chez Drosophila melanogaster. Bull. Biol.
France Belgique 92:248-329.
Sokal, R. R. 1967. A comparison of fitness characters and their response to density in
stock and selected cultures of wild-type and black Tribolium castaneum.
Tribolium Information Bulletin 10:142-147.
YAMADA, YUKIO AND A. E. BELL
Population Genetics Institute
Purdue University
Lafayette, Indiana
*Selection for 13-day larval size in Tribolium
Under two mutritional levels
Selection experiments for larval growth under two nutritional levels were conducted for
sixteen generations so as to evaluate the effectiveness of various selection methods in
the presence of genetic-environmental inter-action.
A strain of Tribolium castaneum, Purdue “+” Foundation stock was used as the
experimental organism. The two levels of nutrition, so-called Good and Poor rations,
which were originally formulated by R. H. Hardin, were used as the environments. The
primary difference between the two rations depends on the content of brewer’s dry yeast
and corn oil. The Good contains 10% of dry yeast and 5% corn oil but none at all in the
Poor.
The character for selection is the 13-day larval weight in two directions. Genetic
parameters of the initial population for the character were: 0.40 in heritability under both
environments and the genetic correlation between Good and Poor performance was 0.80.
The experimental populations which were originated from the base population by random
sampling are listed in Table 1.
Table 1. Symbols of experimental populations
GL
= Selected for large under the Good level every generation.
PL
= Selected for large under the Poor level every generation.
GPL = Selected for large on average performance under both levels every
generation.
GPL = Selected for large under Good and Poor in alternating generation.
GS
= Selected for small under Good every generation.
PS
= Selected for small under Poor every generation.
GPS = Selected for small on average performance under both levels every
generation.
GPS = Selected for small under Good and Poor in alternating generations.
C
= Unselected controls consisted of 20 pair matings, each contributes one male and
one female to the next generation.
The experiment was repeated twice but one week apart. Each set of replicated
experimental populations was sampled from two different sublines derived from the base
population a few generations prior to the initiation of selection.
The mating and selection were made in such a way that each pair produces eggs in the
creamer which contains standard wheat medium for 48 hours, and then the parents are
transferred to Good medium for 24 hours, Poor ration for 24 hours and additional Good or
Poor ration for 24 hours, so as to have two creamers of the same ration for selection
purpose depending on the population, until the 7th generation. For instance, GL had two
Goods and one Poor, while PL had towo Poors and one Good. The GPL and GPS had
only one Good and One Poor. Each mating randomly sampled five larvae for each
creamer weighed and the sum of two creamers was used as the selection criterion. The
measurement taken under the opposite environment rather than for selection was used
as the measure of correlated trait. Since the 8th generation inclusively, two Goods and
two Poors were measured for all populations in the same manner. Once the families for
selection were decided, the full sibs raised under the standard medium were picked up
randomly and sexed for mating to produce the next generation. Therefore, the parental
individuals in all lines were never exposed to either Good or Poor but standard medium.
The individuals measured were discarded after weighing. This technique eliminates any
carry-over environmental effect from parental generation.
Collect the desired pupae for selection. The chips are then collected, their wights
recorded, and returned to the pool.
The pupae show no tendency to roll about on the pan provided the weighing room is kept
warm but, if necessary, the pan grid can be constructed of a wire mesh to prevent such
movement. The number of pupae which can be weighed together is then limited only by
the size of pan or size of scale.
SOKOLOFF, A.
Department of Genetics
University of California, Berkeley
And Natural Sciences Division
California State College, San Bernardino
A method for rearing Eleodes (longicollis?) (Coleoptera:Tenebrionidae) in the laboratory
The stink beetle Eleodes (longicollis?), an inhabitant of the desert areas of the
Southwestern United States and Mexico, when irritated does a “headstand” (lowering its
head and raising the posterior end of the abdomen) and discharges quinines (see
illustrations in Eisner, T. and J. Meinwald, 1966. Defensive secretions of Arthropods.
Science 153:1341-1350). Because of repeated encounters with this beetle outdoors as
well as indoors (janitors often place specimens on my desk should I want to keep them),
attempts have been made to maintain the adults in captivity and to find a medium in
which these beetles can be maintained in stock in the laboratory.
The adult specimens were initially kept in baby food jars provided with grapes as
sources of food and moisture. This food appeared to be adequate since the beetles lived
on it for about a month and the female began to lay eggs in clusters. The latter were
transferred to another jar containing standard flour beetle medium (whole wheat flour +
brewer’s yeast in a proportion of 19:1). The larvae were seen eating this medium, but a
few days later they had died, still in the first larvae instar.
In order to stimulate oviposition, the adult female was placed in flour beetle
medium, and this proved to be a highly satisfactory food for Eleodes, provided that a
source of moisture was available. This was supplied by adding a piece of apple or some
other fruit. The increase in moisture induces the growth of mold, so adult beetles have to
be transferred rather frequently to fresh medium. Nonetheless, the effort pays off, for at
this writing, the beetles have lived in this medium for about five moths, and the female
continues to lay eggs. The larvae also require transferring to fresh medium every so
often because of the tendency of the medium to cake and eventually become moldy when
pieces of fruit are added as a source of moisture. The eggs, somewhat larger than those
in Tribolium but alike in this genus of a white color, are laid in clutches of about 20-30.
The larvae were only about 1-1/2 to 2 mm long in the first instar. At about 240C, the
largest larvae have reached a size of almost three centimeters in three months, and they
resemble the larvae of Tenebrio molitor. Out of 110 small larvae introduced in this jar,
only about 10 per cent survive because the immature stages of Eleodes are highly
cannibalistic. It was possible to observe the cannibalistic activities of these organisms
when the larvae were about 1 cm long. At least two tunnels within which the larvae
travelled were evident. They were against the glass wall of the container, and a larva
from the supper gallery had made its way to the lower gallery where it had encountered a
larva of about the same size; it apparently had dealt the latter a bite on the tergite of the
mesothorax which largely immobilized -- at least there was no apparent effort on the part
of the larva preyed upon to escape--and the predator larva had half of its head within the
body of the larva it was eating. When the larvae are removed from the medium one does
not find any pieces of dead larvae, unless the larvae was recently killed.
The medium, insofar as it has been possible to observe, provides larvae the
necessary nutritional requirements for good health and development. It is possible,
therefore, that the tendency toward cannibalism is a trait characteristic of the family.
It is evident from the above that beetles in this family other than Tribolium and
other flour beetles have potentialities of being raised in the laboratory and thus become
potentially useful organisms in research population genetics and population ecology.
The writer has not tried it, but it comes to mind that Tenebrio molitor, the
mealworm, and Zophobus, another Tenebrionid which normally lives on bat guano, can
be reared on bran provided a source of water such as a slice of potato, is provided to the
beetles every so often. The survival of Eleodes could also be enhanced by rearing this
species in bran medium distributed in a container in layers between paper toweling and to
which wood shavings are provided so the smaller larvae can escape the predatory
activities of larger larvae. The lack of an incubator at San Bernardino for this particular
purpose has prevented the exploration of the question of the optimal conditions of
temperature and R.H. for the development of this beetle from egg to adult.
P.S. A visit to the laboratory of Dr. Clyde Willson, Biochemistry Department,
University of California, Berkeley, after the above note was written, revealed that Mr.
Walter Tschinkel is extremely interested in developing techniques for rearing a wide
variety of Tenebrionidae (see Research Note elsewhere in this issue). Mr. Tschinkel
informs me that according to Dr. Eisner, Eleodes can be kept on bran for as long as six or
seven years in the laboratory (if supplied with water). During this time they produce
numerous progeny, but they fail to reach the adult stage. Whether the present diet will
prove to be more suitable so Eleodes will undergo metamorphosis remains to be seen.