SECTION A
GENERAL MICROBIOLOGY RHIZOBIUM INTRODUCTION
The bacteria of the genus Rhizobium (family Rhizobiaceae)
are a genetically diverse and physiologically heterogeneous
group of microorganisms that are nevertheless classified
together by virtue of their ability to nodulate groups of
plants of the family Leguminosae.
This classification
scheme is usually referred to as “cross-inoculatin”
grouping.
A cross-inoculation group is a group of legumes
in which one species of Rhizobium nodulates all the legumes
within that group.
In the old system, species of Rhizobium
fall into two groups based on their growth characteristics.
Group I
Rhizobium leguminosarum – nodulates peas (pisum spp.),
vetch (Vicia spp.), lentils,
(Lens culinaris).
Rhizobium phaseoli
- nodulates beans (Phaseolus
Vulgaris and the scarlet
runner bean) (Phaseolus
coccineus).
Rhizobium trifolii
- nodulates the clovers, e.g.,
Trifolium subterraneum, T.
Semipilosum, T. repens, and
Other Trifolium spp.
Rhizobium meliloti
- nodulates alfalfa (Medicago
sativa) and other Medicago
spp., Melilotus spp.,
fenugreek (Trigonella).
Group II
Rhizobium lupini
- nodulates lupins (Lupinus
Spp.) and serradella
(Ornithopus spp.).
Rhizobium japonicum
- nodulates soybean (Glycine
max).
Rhizobium spp.
– nodulates members of the
“cowpea miscellany” group of
legumes, e.g., Vigna spp.,
peanut, Desmodium spp.,
Macroptilium spp., Lablab
sp., Lima bean, Stylosanthes
spp., etc.
In Group I are the fast-growing acid producers which
develop pronounced turbidity in liquid media within 2-3
days and have a mean doubling time of 2-4 hours.
The cells
are rod-shaped to pleomorphic, 0.5 to 0.9 microns in
diameter and 1.2 to 3.0 microns long, and are motile by 2-6
peritrichous flagella.
They can grow on a wide range of
carbohydrates, but usually grow best on glucose, mannitol,
or sucrose.
Rhizobia of this group are generally infective
on temperate legumes.
In group II are the slow-growing, alkali producing
rhizobia.
They require 3-5 days to produce moderate
turbidity in liquid media and have a mean doubling time of
6-7 hours.
Most strains in this group grow best with
pentoses as their carbon source.
The cells are
predominantly rod-shaped, and motile by a single polar or
subpolar flagellum.
This group nodulates tropical legume
species.
Rhizobia are characteristically Gram-negative and do not
form endospores.
Uneven Gram-staining is frequently
encountered with rhizobia depending on the age of the
culture.
Cells from a young culture and nodule bacteroids
usually show even Gram-staining while older cells and
longer cells show unstained areas along the cell giving a
banded appearance.
These unstained areas have been
identified to be large granules of polymeric-hydroxybutyric
acid (PHBA).
These granules are refractile under phase
contrast microscopy.
Though the host-dependent cross-inoculation group system of
classifying rhizobia has been subjected to much criticism,
because it is not a taxonomic one, it is the best practical
system currently available.
Classification of rhizobia is
becoming increasingly complex because of new findings, e.g.
some soybeans are now known to be nodulated by a distinct
group of fast-growing acid-producing rhizobia.
Thus, a new
system has been formulated to classify rhizobia.
This new
system recognizes three genera for the Rhizobiaceae.
In
this new system Genus I and Genus II include the rhizobia
while Genus III is for the agrobacteria.
All fast-growing
acid-producing rhizobia now fall under the new genus
Rhizobium (Genus I) and all slow-growing alkali-producing
rhizobia under the new genus Bradyrhizobium (Genus II).
Also, under this system, R. trifolii, R. phaseoli, and R.
leguminosarum are combined as one species, designated as
Rhizobium leguminosarum, comprising three biovars
(trifolii, phaseoli, and viceae).
R. meliloti remains as
before and R. loti has been assigned to the fast-growing
Lotus rhizobia.
Genetically related to R. loti are
rhizobia from Lotus corniculatus, Lotus tenuis, Cicer
arietinum, Leucaena leucocephala, Sophora microphylla etc.
The soybean rhizobia are now in two genera, i.e. R.
japonicum (fast-growing and acid-producing) and
Bradyrhizobium japonicum (slow-growing and alkaliproducing).
Rhizobia from Vigna, Arachis, Desmodium,
Macroptilium, Stylosanthes etc. are still unclassified, but
grouped as Bradyrhizobia spp.
The non-legume Parasponia
(called Trema previously) is also nodulated by
Bradyrhizobium sp.
Besides Leucaena, whose rhizobia are
now R. loti, there are other legumes (Sesbania, Neptunia,
Calliandra, Acacia) which are nodulated by fast-growing
acid-producing rhizobia and the taxonomic status of these
organisms may be resolved in the future.
As predominantly aerobic chemoorganotrophs, rhizobia are
relatively easy to culture.
They grow well in the presence
of oxygen and utilize relatively simple carbohydrates and
amino compounds.
With the exception of a few strains, they
have not been found to fix nitrogen away from their host
legume.
Some strains of Rhizobium require vitamins for
growth.
Optimal growth for most strains occurs in a
temperature range of 25-30oC, and at a pH of 6-7.
Despite
their usual aerobic metabolism, many strains are able to
grow well at oxygen tensions less than 0.01 atmosphere
(microaerophilic).
Rhizobia are somewhat unique among soil microorganisms I
their ability to form nitrogen-fixing symbioses with
legumes.
To enjoy the benefits of this partnership,
however, the rhizobia must not only exhibit saprophytic
competence among other soil microorganisms, but also outcompete other rhizobia for infection sites on legume roots.
Potential for physiological versatility is therefore an
important trait contributing to their adaptation to the
competitive and complex soil environment.