ISRAEL JOURNAL OF
VETERINARY MEDICINE
ABSTRACTS OF PAPERS PRESENTED AT
THE 27TH VETERINARY SYMPOSIUM,APRIL 2,
2003
Vol.
58 (2-3)
2003
DEDICATED TO THE MEMORY AND IN HONOR OF
MR. ROMAN LUBETZKY
HELD AT THE VETERINARY TEACHING HOSPITAL,
AGRICULTURAL CAMPUS, RISHON LE-ZION
Symposium chairperson: Prof. K. Perk
DOWNREGULATION OF MHC CLASS-II RECEPTORS OF DH82
CELLS
FOLLOWING INFECTION WITH EHRLICHIA CANIS.
S. Harrus1, T. Waner2, D. Friedman-Morvinski3, Z. Fishman1, H.
Bark1 and A. Harmelin4.
1. Koret School of Veterinary Medicine, The Hebrew University of
Jerusalem, Israel,
2. Israel Institute for Biological Research, Ness Ziona, Israel,
3. Department of Immunology, The Weizmann Institute of Science,
Rehovot, Israel,
4. Department of Veterinary Resources, The Weizmann Institute of Science,
Rehovot, Israel
Ehrlichia canis, the etiologic agent of canine monocytic ehrlichiosis, is an
obligatory intracellular bacterium, infecting monocytes and macrophages of the
monocytic phagocytic system. Its intracellular location coupled with a number of
other mechanisms allows the rickettsia to evade the host’s immune system.
In order to evaluate whether infection with E. canis alters the expression of
MHC class I and/or MHC class II receptors, flow cytometry was performed on
DH82 cells infected with Ehrlichia canis (90% infection) and on uninfected
DH82 cells of the same passage, using anti-canine MHC class I and II antibodies.
MHC class II expression was evident in 47.6% and 46.2% of uninfected
DH82 cells using 2 different anti-MHC class II antibodies, no MHC class II
expression was evident in DH82cells infected with E. canis.
The results of our study indicate that infection of DH82 macrophages with
E. canis down regulates their MHC class II receptors. These results suggest a
possible mechanism by which E. canis evades the immune system.
PROGRAMMED CELL DEATH IN BACTERIA
G. Glaser, H. Engelberg - Kulka and I. Marianovski
Hebrew University - Hadassah Medical School Jerusalem, Israel.
In bacteria, programmed cell death is mediated by “addiction modules,”
unique genetic systems located in certain extra-chromosomal elements that in E.
coli are known to be low copy plasmids and prophages. An “addiction module”
consists of a pair of genes: the downstream gene specifies for a stable toxic
protein while the upstream gene encodes a short-lived antidote that antagonizes
the toxin. When the plasmid(s) or other extrachromosomal element(s) is (are)
lost, the cured cells are selectively killed because the unstable antitoxin is
degraded faster than is the stable toxin. Thus, the cells are said to be addicted to
the continued presence of the non-chromosomal genetic element. Along with
plasmid partition and other very precise mechanisms for preventing plasmid loss,
addiction modules assist in maintaining stability in the host of the extrachromosomal elements on which they are borne.
Gene pairs have been found on the E. coli chromosome that are
homologous to some of the extra-chromosomal “addiction modules”. On the
chromosome downstream from the relA gene, responsible for the “stringent
response,” we found the gene pair mazE and mazF. More recently, we have
reported that mazEF is the first described chromosomal regulable “addiction
module” responsible for programmed cell death in bacteria. The chromosomallyborne mazEF system has all the properties of an extra-chromosomal “addiction
module”. Specifically: (i) MazF is a toxic protein that is antagonized by the
antitoxic protein, MazE; (ii) MazF is long lived, while MazE is a labile protein
degraded by the protease ClpPA; (iii) MazE and MazF interact; (iv) MazE and
MazF are co-expressed; (v) MazE, the antitoxic protein, is synthesized in excess
over the toxic protein MazF; and (vi) mazEF is weakly auto-regulated by MazE
and efficiently auto-regulated by the combined action of MazE and MazF. In
addition to these characteristics, the mazEF system has several unique properties:
its promoter has an unusual DNA structure called the “alternating palindrome”,
and also carries a binding site for the factor for inversion stimulation (FIS).
Moreover, mazEF-mediated cell death can also be triggered by several stressful
conditions that inhibit mazEF expression: (a) Under conditions of amino acid
starvation, high cellular concentrations of 3’ 5’ guanosine-bis-pyrophosphate
(ppGpp), the product of the RelA protein, and (b) Antibiotics that are general
inhibitors of transcription and/or translation. We have therefore proposed that the
chromosomally borne mazEF system plays a role in programmed cell death
under stressful conditions.
CLINICAL MANIFESTATION OF FELINE INFECTIOUS ANEMIA
IN ISRAEL AND MOLECULAR CHARACTERIZATION OF ISRAELI
MYCOPLASMA HAEMOFELIS AND CANDIDATUS M.
HAEMOMINUTUM (PREVIOUSLY HAEMOBARTONELLA) ISOLATES
S. Harrus1, S. Tasker2, S. E. Shaw2, Z. Fishman1, I. Aroch1, E. Lavy1 and
G. Baneth1.
1. School of Veterinary Medicine, The Hebrew University of Jerusalem,
Israel
2. Department of Clinical Veterinary Science, University of Bristol, UK
In order to characterize Israeli isolates of feline haemotropic Mycoplasma
spp., blood samples from infected cats were collected. Mycoplasma (previously
Haemobartonella) DNA was amplified and genetically characterized. Near
complete 16S rRNA and RNase P genes of 4 isolates were sequenced. Both
Mycoplasma haemofelis (the large form) and Candidatus M. hemominutum (the
small form) were detected in Israeli cats.
Previously reported†signs of naturally occurring clinical feline
haemoplasmosis in Israeli cats included tachypnea, lethargy, depression,
anorexia, infestation with fleas, pale mucous membranes, icterus, emaciation,
dehydration, splenomegaly, anemia and leucocytosis. Thirty eight % and 22% of
cats, tested for FeLV antigen and FIV antibodies respectively, were found to be
positive. The prevalence of FeLV and FIV in the study was much higher than
that of FeLV and FIV in the general Israeli cat population.
Mycoplasma haemofelis is considered to be pathogenic, while Candidatus
M. hemominutum is considered non-pathogenic. Therefore, reported signs are
predominately attributed to M. haemofelis. In this study, two cats were found to
be infected with Candidatus M. haemominutum (the small form). Both cats were
FIV positive and showed clinical disease with severe anemia (PCV of 9% and
16%). This finding suggests that Candidatus M. haemominutum may cause
clinical disease in immunosuppressed cats.
EVALUATION OF A RAPID DOT ELISA KIT FOR CANINE
PARVOVIRUS AND
CANINE DISTEMPER VIRUS IMMUNOGLOBULIN M (IgM)
ANTIBODIES
T. Waner1, S. Mazar2, E. Nachmias3, E. Keren-Kornblatt2 and S. Harrus4.
1. Veterinary Clinic, 9 Meginay Hagalil St., Rehovot.
2. Biogal Galed Laboratories, Kibbutz Galed.
3. Rupin Veterinary Hospital.
4. Koret School of Veterinary Medicine, Hebrew University of Jerusalem.
A dot enzyme-linked immunosorbent assay (ELISA) for the detection of
IgM antibodies against canine distemper virus (CDV) and canine parvovirus
(CPV) was assessed. The titers (n=100) of CDV and CPV IgM antibodies by the
Immunocomb® ELISA kit were compared with results of an
immunofluorescence assay (IFA). The correlation between dot ELISA technique
and IFA was found to be highly significant (p<0.001). The kinetics of the
antibody response of IgG and IgM CPV and CDV antibodies following
vaccination was followed using Immunocomb® ELISA. High levels of CPV IgM
antibodies were first detected at 7 days post-vaccination (PV), and from day 9 PV
all pups had high titers of CPV IgG antibodies. High concentrations of CDV
IgM were detected at 9 days PV, with the highest average titers present on day 12
PV; CDV IgG antibodies were present from 9 days PV. It was concluded that
using an IgM-ELISA kit under clinic conditions, on a single serum sample, might
enable the clinician to strengthen his diagnosis of CPV or CDV at an early stage
of the disease, however this has to be evaluated during natural CPV and CDV
disease rather than post-vaccination
RIEMERELLA ANATIPESTIFER-LIKE ORGANISM CAUSING
MENINGITIS IN TURKEYS:
PATHOLOGICAL FINDINGS AND BIOLOGICAL FEATURES
I. Lublin1, S. Mechani1, S. Perl2 and H. Yuval3
1. Division of Avian & Aquatic Diseases
2. Division of Pathology, Kimron Veterinary Institute, POB 12, 50250 Bet
Dagan, Israel.
3. Poultry Veterinarian, Herzlya
In June 2002 a syndrome involving the central nervous system occurred in
several relatively young meat-type turkey farms in Israel, and in some cases was
accompanied by a mild respiratory disease. The clinical signs were recumbency,
incoordination, head tremors and torticollis. Morbidity and mortality rates were
usually low, except for the cases in which another disease, such as TRT, was
involved. Since its first appearance, the syndrome was diagnosed on about six
different farms, in about 15 outbreaks (in two of the farms the syndrome occurred
over several generations in the same buildings). The age range of the turkeys was
from 9 days to 20 weeks. To determine the etiology many agents were examined,
all of them were negative except for a bacterium that resembled Riemerella
anatipestifer from which it differed in many properties. The negative agents
were:
Viruses
Newcastle and other paramyxoviruses, avian encephalomyelitis,
flaviviruses (turkey meningoencephalitis, West Nile Fever)
Bacteria
Fungi
other
factors
Pasteurella multocida, Ornithobacterium rhinotracheale,
Chlamydia psittaci, botulinum
Aspergillus spp., other fungi
mechanical injury
The main pathological findings were hemorrhages in the meninges, usually
without prominent findings in other organs. Purulent meningitis was observed on
histopathological examination of the cerebrum and cerebellum.
In all these outbreaks we isolated, mostly from brain but sometimes also
from lungs and liver, a gram-negative coccobacillus, non-motile, growing in
aerobic conditions but with slight improvement in micro-aerophilic conditions,
routinely diagnosed as Riemerella anatipestifer-like. It had small colonies
resembling Pasteurella spp. and ORT after 24 h (370C) but much bigger,
yellowish-colored and different from the former after 48 h. The organism was
isolated in most of the cases from the brain, and in part of them also from the
lungs. Biochemical analysis of the organism (API 20 NE, BD BBL Crystal
Microbial ID System), attributed it to the Flavobacteriaceae, like ORT and
Riemerella anatipestifer, with a few biochemical properties different from these
two. The viability time of this organism on culture media was much longer when
compared to Riemerella (in contrast to Riemerella, it could be refreshed and recultured even after a week at 370C or at room temperature). The resemblances
and differences between the two bacteria are summarized in the table.
Resemblances
Differences
both are coccobacilles
quite different microscopic morphology
both are gram-negatives
colonies much bigger in the “new” organism
both are non-motile
both are aerobicmicroaerophilic
appearance of a yellowish pigment in the “new”
organism
few different biochemical properties longer
viability
time of the “new” organism
USE OF INSERTION SEQUENCE ELEMENTS ISMB-A1 AND
ISMB-B1 FOR MOLECULAR TYPING
AND EPIDEMIOLOGY OF MYCOPLASMA BOVIS
I. Lysnyansky1, D. Yogev2 and S. Levisohn1
1. Department of Poultry Diseases, Kimron Veterinary Institute, Bet Dagan,
Israel.
2. Department of Membrane and Ultrastructure Research, The Hebrew
University-Hadassah Medical School, Jerusalem 91120, Israel
Bacterial insertion sequences (IS) are small (800-2500bp) mobile genetic
elements with simple genetic organization capable of inserting at multiple sites in
the target molecule and mediating gene activation, deletion, rearrangement,
recombination and transfer. More than 500 IS have been described, widely
distributed in bacterial genera, including several Mollicutes species.
Recently, four new IS have been identified in Mycoplasma bovis PG45, the
most important etiological agent of bovine mycoplasmosis. They were designated
ISMB-A1, ISMB-A2, ISMB-B1 and ISMB-B2 and revealed high homology with
IS4 and IS30 family of insertion sequences of E. coli.
Southern blot analysis showed that international reference strains of M.
bovis (11, mainly from Germany) contain multiple copies of each IS family, but
the number of copies and genomic position varied among the strains. Preliminary
analysis of a cross-section of 22 M. bovis isolates from Israel (1995-2002)
showed a variety of molecular fingerprints. Clusters of strains with identical
patterns were identified. Potentially this method could be used to identify foci of
infection and to differentiate local strains from those introduced by imported
animals.
SEROPREVALENCE OF ANAPLASMA (EHRLICHIA)
PHAGOCYTOPHILUM
ANTIBODIES AMONG HEALTHY DOGS AND HORSES IN
ISRAEL.
O. Levi1, T. Waner2, G. Baneth1, A. Keysary2, Y. Bruchim1, J.
Silverman1 and S. Harrus1.
1. Koret School of Veterinary Medicine, Hebrew University of Jerusalem,
Rehovot, Israel
2. Israel Institute for Biological Research, Ness Ziona, Israel.
Previous studies have demonstrated the presence of antibodies to
A.phagocytophilum in Israel, in both the golden jackal (Canis aureus syriacus)
and humans. This study was undertaken to investigate whether domestic dogs
and horses in Israel have been exposed to A. phagocytophilum. Of the 195 dogs
tested, 9% were seropositive for A. phagocytophilum and 30% were seropositive
to E. canis. Twenty nine percent of the dogs seropositive for E. canis were also
seropositive to A. phagocytophilum. Two dogs had A. phagocytophilum- IFA
titers greater than E. canis. The equine serological survey revealed no
seropositive horses among the 300 horses tested (titer<1:40).
The results presented in this study suggest that dogs in Israel could be
exposed to A.phagocytophilum, however cross-reaction with E. canis is more
likely. In spite of the high prevalence of ticks on horses in Israel during the
summer months, no evidence for exposure to A. phagocytophilum was apparent.
RABIES VIRUS DETECTION BY RT- PCR IN DECOMPOSED
NATURALLY INFECTED BRAINS
David, D.1, Yakobson, B.1, Rotenberg, D.1, Dveres, N.1, Davidson, I.2 and
Stram, Y.3
1. Rabies Laboratory, Pathology Division
2. Division of Avian Diseases and
3. Division of Virology, Kimron Veterinary Institute, 50250 Bet Dagan,
Israel.
The warm climate of Israel and mishandling of the cadavers during transit
to the laboratory requires an accurate method for diagnosis of rabies in
decomposed tissues. By using the reverse transcriptase polymerase chain reaction
(RT-PCR) ten decomposed brain samples collected between 1998-2000 that were
diagnosed as negative by direct fluorescent antibody test (FAT), were found
positive. Three of the 10 decomposed brains were confirmed as positive by
isolation of rabies virus in tissue culture and by mouse inoculation (MIT), while
the other seven decomposed samples were found positive only by RT-PCR.
Direct sequencing and molecular analysis of a 328-bp fragment of the N gene of
all the rabies sequences confirmed their geographical origin. These results
demonstrated the importance of the RT-PCR in the detection of rabies virus in
decomposed naturally infected brains, especially in cases when the sample is not
suitable for other laboratory assays. Thus, the RT-PCR can provide a positive
diagnosis; however, when a negative result is obtained due to the nature of the
decomposed tissue that can be caused by technical reasons a false negative might
be the case.
AN EPIDEMIOLOGICAL STUDY OF CULICOIDES
HYPERSENSITIVITY AMONG HORSES IN ISRAEL
A. Steinman1, G. Peer1 and E. Klement2
1. Koret School of Veterinary Medicine, The Hebrew University of
Jerusalem, P.O.Box 12, 76100 Rehovot, Israel.
2. Center for Vaccine Development and Evaluation, IDF Medical Corps,
M.P. 02149, Israel.
Culicoides hypersensitivity (sweet itch) is a chronic, recurrent, seasonal
dermatitis of horses. Genetic predisposition, gender, age and color were
suggested previously as possible risk factors for Culicoides hypersensitivity. The
present study was undertaken to establish the effect of the above factors on the
prevalence of Culicoides hypersensitivity among horses in Israel. During summer
2001, a cross-sectional study was conducted which involved 408 horses from 18
farms. Horses with suspected Culicoides hypersensitivity (based on a primary
questionnaire) were examined. Diagnosis was based on the presence of typical
clinical signs and the seasonality of the disease. The Mantel-Haenszel method
was used to estimate odds ratios following stratification on potential confounders
(age, breed, and farm). A total of 114 out of 408 horses (28 per cent) were
affected. A significant difference (P<0.001) was found between horses kept 300
m below sea level and 800 m above sea level. When compared to local breed,
Arab horses (ORMH=0.175), Warmblood horses (ORMH=0.45) and
Thoroughbreds (ORMH=0.45) were less affected, while ponies (ORMH=2.2)
were most affected. Horses older then 10 years of age were more affected
(ORMH=1.86), when compared to younger horses. The results of this study
imply that the most important factors affecting the prevalence of Culicoides
hypersensitivity are the altitude, breed and age. Gender and color were not
significantly correlated with the prevalence of this disease. Since a genetic basis
of Culicoides hypersensitivity exists, it is concluded that environmental factors,
including management regimes are at least as important in determining the
prevalence of the disease. It is suggested that in future studies of insect-borne
diseases the farm, age and breed should be regarded as potential confounders and
controlled in order to assess properly other risk factors.