|Abstract or Summary
- Although much has been learned about the comparative
nodulating behavior of simple mixtures of rhizobial strains
under non-soil situations, it is unclear how these findings
relate to the factors influencing nodulation success by the
complex mixtures of strains found within soil-borne
rhizobial populations. Information on the structure and
physiological behavior of soil populations is almost nonexistent.
To achieve a better understanding of the situation
in soil, studies were carried out with the following
objectives. (i) To delineate by serological analysis the
population composition of nodule occupants of Rhizobium
leguminosarum bv.trifolii recovered from a variety of annual
and perennial clover (Trifolium) species planted into Abiqua
soil. (ii) To further the development of an assay to
evaluate the substrate responsiveness of specific indigenous
serotypes of R. leguminosarum bv. trifolii while they reside
within the soil microbial community. Immunodiffusional
analysis of isolates recovered from nodules of five annual
(T. subterraneum, T. incarnatum, T. vesiculosum, T.
parviflorum, T. patens) and three perennial (T. pratense, T.
repens, T. hybridum) species of clover revealed that the
serotypic composition of the natural population of R.
leguminosarum bv. trifolii in Abiqua soil is almost
completely known. With antisera to 14 antigenically distinct
serotypes at our disposal, only 19 of 272 isolates recovered
from these eight clover species were antigenically unknown.
While the perennial species showed no pronounced preference
for particular serotypes, a substantial proportion (37-75%)
of nodule occupants from each of the annual clovers (with
the exception of T. vesiculosum) reacted with antiserum AS6.
These isolates could be subdivided by their serological
reactions of non-identity with either antisera AS6, AS27, or
both antisera AS21 and AS27. Using multi-locus allozyme
electrophoresis (MLAE) to analyze population structure
within serotypes, isolates representing serocluster AS6 were
found to be rooted at a similarity of 0.82 and clustered
with the other three serotypes (AG4, AS21, and AS27) only at
a similarity of 0.37. In contrast to AS6, MLAE analysis
revealed that "genotypic distances" between the 7 ETs
representing AG4 could be large. The chapter on the
nalidixic acid cell-elongation assay only represents the
second report of its use on soil microbial populations.
Nalidixic acid was found to be the most suitable DNA gyrase
inhibitor for rhizobial studies since norfloxacin and
ciprofloxacin at extremely low concentrations (2.0 and 0.5
mg/l, respectively) reduced the proportion of elongating
cells significantly. In contrast to other indigenous
serotypes, the majority of members of serotype AR23 did not
elongate in response to yeast extract (YE). Regardless of
nutrient type, or concentration, the percentage of elongated
cells of AR23 remained low (<16%) even after 24 h of
incubation. While the cell elongation response of serotype
AS6 occured more rapidly to YE than did AR23, a less
vigorous response by AS6 was observed when other nutrient
sources were used. The appearance of elongated cells was
delayed and the final percentage of elongated cells was