[
International Worm Meeting,
2005]
The free living soil nematode Caenorhabditis elegans has been cultured in the lab for over four decades on agar plates feeding almost exclusively on E. coli OP50, which served as their source of nutrition as well as their immediate environment. Almost all we know about gene function in C. elegans has been determined in this environment. Yet, many of the genes in C. elegans genome have not been assigned a function. It seems reasonable that at least some proportion of the genes of unknown function evolved to allow C. elegans to respond to its natural environment. As part of a larger project to discover gene functions required for ecological interactions, we are conducting parallel studies examining changes in nematode diversity in response to environmental perturbations in the field, (Jones et al., unpublished) and using C. elegans to model those perturbations in the lab. As a start, we have identified several bacterial isolates from Konza Prairie (near Manhattan, KS) soils, of which Micrococcus luteus was the most abundant of the culterable bacterial taxa. We have used cDNA microarrays to identify genes involved in nematode responses to different bacterial diet or environments by comparing genes expressed by wild-type C. elegans grown on plates seeded with OP50 those expressed when they are grown on M. luteus. We have identified 142 genes with significant changes in expression levels in response to growth on M. luteus. Quantitative real time PCR analysis of eight randomly selected genes showed nearly identical expression levels as the microarrays, validating our results. Mutant alleles were available for five genes that were upregulated in response to growth on M. luteus. In order to examine the contribution of these genes to nematode fitness in different bacterial environments, we have examined brood sizes. The brood size of wild-type hermaphrodites grown on M. luteus is only slightly depressed as compared to those grown on OP50. Four mutants had a statistically significant reduction in brood sizes, when grown on M. luteus. Furthermore, mutant phenotypes have not been reported for two of these. Thus it seems that, not only are the expression of specific genes induced in response to a change in environment, but some of these genes contribute to fitness traits. These results demonstrate that assessing C. elegans gene functions in more natural environments can allow new functions to be assigned to genes of unknown function. Experiments are currently underway to determine whether these same genes or others will be induced in response to growth on other soil bacteria.
[
Mid-west Worm Meeting,
2004]
Nematode communities at the Konza Tallgrass Prairie Biological Station near Manhattan, KS are known to respond to nitrogen addition and burning. Using a combination of morphometric identification and molecular methods, nematode 'species' have been identified and preliminary results show differential responses to environmental perturbations at the genus level. (see abstract by Jones et al .) We are interested in linking the responses of organisms to environmental change at the genetic level. Little is known about how the environment affects organisms at the level of expression of individual genes. We are addressing this using laboratory soil cultures and the genetic model nematode Caenorhabditis elegans . We aim to use C. elegans to discover genes that are induced or repressed in response to changes in soil nitrogen and water by using laboratory soil cultures to model nematode environments on Konza. Homologs of the C. elegans genes exhibiting the greatest changes in expression will be identified in resident nematode 'species' and their expression examined. As a first step we have developed a C. elegans soil culture system. We collected 13 soil bacterial isolates from native Kansas soils. N2 grew and developed normally on plates seeded with each of the isolates. Using the most abundant soil bacterial isolate, Micrococcus luteus , we have developed soil cultures that support soil bacterial and nematode growth. We have optimized growth conditions and found that water addition every six days maximizes nematode growth in soil culture. We also found that bacterial addition every three days increases nematode abundance in soil culture. We have been able to quickly extract nematodes from our soil cultures, which is essential for isolation of mRNAs for microarray studies. As a control for our future work, we are using cDNA microarrays to discover genes that are specifically expressed or repressed during soil growth. Microarray experiments comparing E. coli (OP50) fed N2 populations to M. luteus fed N2 populations grown on plates, as well as to those grown in soil culture, are currently under way.
[
Mid-west Worm Meeting,
2004]
While much is known about the roles genes play in development and physiology, little is known about how the environment affects organisms at the level of expression of individual genes. Using resident nematode populations sampled from the Konza Tallgrass Prairie Biological Station near Manhattan, Kansas, we are attempting to link the responses of organisms to environmental change at the genetic level. We hypothesize that different species may have varying genetic capacities to respond to changes in the environment; either by differences in the genes they possess or in how those genes are regulated. We are currently testing these possibilities by looking at the responses of bacterial feeding nematodes to changes in soil chemistry. Although the Konza nematode community is known to respond strongly to nitrogen addition and burning at the family level, it is not known if there are differential responses to treatment effects at the genus level or below. As an initial step towards the identification of taxa that are sensitive to these treatment effects, the objective of this portion of the study was to define taxonomic diversity across treatment plots that differed in nitrogen addition and burning regime. In addition to morphometric identification to genus, we are using molecular methods to identify taxonomic diversity at the 'species' level. We sequenced the 3' 500 bases of the 18S ribosomal RNA gene along with the entire ITS1 sequence. From 900bp of sequence, we have currently identified 17 'species' of nematodes across 4 different families. Preliminary results based on morphometric designations show differential effects of nitrogen and burning at the genus level. Using molecular information, further research will address treatment effects at lower taxonomic levels. Once treatment sensitivity has been identified at the lowest taxonomic scale possible, we will then attempt to identify the genes responsible for the observed responses.