Regulation of the sperm-to-oocyte transition in Caenorhabditis briggsae hermaphrodites by the Cbr-met-2 and Cbr-fem-3 genes.

In Caenorhabditis briggsae hermaphrodites, spermatogenesis begins in the L4 larval stage and persists into early adulthood. Oogenesis begins after spermatogenesis; the sperm-to-oocyte transition is irreversible. The timing of this transition is believed to have evolved in response to selection to maximize the intrinsic growth rate. Sperm-to-oocyte transitions occurred early in Cbr-met-2 and Cbr-fem-3 mutants. These early transitions resulted in reduced brood sizes, but had little or no impact on the intrinsic growth rate. In Cbr-met-2; Cbr-fem-3 doubly mutant hermaphrodites, the transition to oogenesis occurred even earlier and brood size was further reduced, indicating that Cbr-met-2 and Cbr-fem-3 regulate the sperm-to-oocyte transition through separate pathways. Mutations in Cbr-met-2 also resulted in an increase in the frequency of males in mutant populations. These increased male frequencies were not caused by increased rates of X nondisjunction during oogenesis in mutant hermaphrodites. Rather, increases in the rates of outcrossing in mutant populations likely were an indirect effect of reduced brood sizes derived from self-fertilization. Based on these observations, it is possible that the timing of the sperm-to-oocyte transition in C. briggsae evolved in response to sexual selection on hermaphrodites to limit rates of outcrossing. Mutations in the orthologous Caenorhabditis elegans gene, Cel-met-2, did not impact the timing of the sperm-to-oocyte transition, consistent with the independent evolution of hermaphroditic reproduction in these species. Although brood sizes were reduced in Cel-met-2 mutant strains, increased male frequencies were not observed. Cbr- and Cel-met-2 mutations also differed in terms of germline mortality, observed in C. elegans, but not in C. briggsae.

Caenorhabditis briggsae recombinant inbred line genotypes reveal inter-strain incompatibility and the evolution of recombination.

The nematode Caenorhabditis briggsae is an emerging model organism that allows evolutionary comparisons with C. elegans and exploration of its own unique biological attributes. To produce a high-resolution C. briggsae recombination map, recombinant inbred lines were generated from reciprocal crosses between two strains and genotyped at over 1,000 loci. A second set of recombinant inbred lines involving a third strain was also genotyped at lower resolution. The resulting recombination maps exhibit discrete domains of high and low recombination, as in C. elegans, indicating these are a general feature of Caenorhabditis species. The proportion of a chromosome's physical size occupied by the central, low-recombination domain is highly correlated between species. However, the C. briggsae intra-species comparison reveals striking variation in the distribution of recombination between domains. Hybrid lines made with the more divergent pair of strains also exhibit pervasive marker transmission ratio distortion, evidence of selection acting on hybrid genotypes. The strongest effect, on chromosome III, is explained by a developmental delay phenotype exhibited by some hybrid F2 animals. In addition, on chromosomes IV and V, cross direction-specific biases towards one parental genotype suggest the existence of cytonuclear epistatic interactions. These interactions are discussed in relation to surprising mitochondrial genome polymorphism in C. briggsae, evidence that the two strains diverged in allopatry, the potential for local adaptation, and the evolution of Dobzhansky-Muller incompatibilities. The genetic and genomic resources resulting from this work will support future efforts to understand inter-strain divergence as well as facilitate studies of gene function, natural variation, and the evolution of recombination in Caenorhabditis nematodes.

A toolkit for rapid gene mapping in the nematode Caenorhabditis briggsae.

BACKGROUND: The nematode C. briggsae serves as a useful model organism for comparative analysis of developmental and behavioral processes. The amenability of C. briggsae to genetic manipulations and the availability of its genome sequence have prompted researchers to study evolutionary changes in gene function and signaling pathways. These studies rely on the availability of forward genetic tools such as mutants and mapping markers. RESULTS: We have computationally identified more than 30,000 polymorphisms (SNPs and indels) in C. briggsae strains AF16 and HK104. These include 1,363 SNPs that change restriction enzyme recognition sites (snip-SNPs) and 638 indels that range between 7 bp and 2 kb. We established bulk segregant and single animal-based PCR assay conditions and used these to test 107 polymorphisms. A total of 75 polymorphisms, consisting of 14 snip-SNPs and 61 indels, were experimentally confirmed with an overall success rate of 83%. The utility of polymorphisms in genetic studies was demonstrated by successful mapping of 12 mutations, including 5 that were localized to sub-chromosomal regions. Our mapping experiments have also revealed one case of a misassembled contig on chromosome 3. CONCLUSIONS: We report a comprehensive set of polymorphisms in C. briggsae wild-type strains and demonstrate their use in mapping mutations. We also show that molecular markers can be useful tools to improve the C. briggsae genome sequence assembly. Our polymorphism resource promises to accelerate genetic and functional studies of C. briggsae genes.

Comparison of C. elegans and C. briggsae genome sequences reveals extensive conservation of chromosome organization and synteny.

To determine whether the distinctive features of Caenorhabditis elegans chromosomal organization are shared with the C. briggsae genome, we constructed a single nucleotide polymorphism-based genetic map to order and orient the whole genome shotgun assembly along the six C. briggsae chromosomes. Although these species are of the same genus, their most recent common ancestor existed 80-110 million years ago, and thus they are more evolutionarily distant than, for example, human and mouse. We found that, like C. elegans chromosomes, C. briggsae chromosomes exhibit high levels of recombination on the arms along with higher repeat density, a higher fraction of intronic sequence, and a lower fraction of exonic sequence compared with chromosome centers. Despite extensive intrachromosomal rearrangements, 1:1 orthologs tend to remain in the same region of the chromosome, and colinear blocks of orthologs tend to be longer in chromosome centers compared with arms. More strikingly, the two species show an almost complete conservation of synteny, with 1:1 orthologs present on a single chromosome in one species also found on a single chromosome in the other. The conservation of both chromosomal organization and synteny between these two distantly related species suggests roles for chromosome organization in the fitness of an organism that are only poorly understood presently.

Genetically Distinct Behavioral Modules Underlie Natural Variation in Thermal Performance Curves.

Thermal reaction norms pervade organismal traits as stereotyped responses to temperature, a fundamental environmental input into sensory and physiological systems. Locomotory behavior represents an especially plastic read-out of animal response, with its dynamic dependence on environmental stimuli presenting a challenge for analysis and for understanding the genomic architecture of heritable variation. Here we characterize behavioral reaction norms as thermal performance curves for the nematode Caenorhabditis briggsae, using a collection of 23 wild isolate genotypes and 153 recombinant inbred lines to quantify the extent of genetic and plastic variation in locomotory behavior to temperature changes. By reducing the dimensionality of the multivariate phenotypic response with a function-valued trait framework, we identified genetically distinct behavioral modules that contribute to the heritable variation in the emergent overall behavioral thermal performance curve. Quantitative trait locus mapping isolated regions on Chromosome II associated with locomotory activity at benign temperatures and Chromosome V loci related to distinct aspects of sensitivity to high temperatures, with each quantitative trait locus explaining up to 28% of trait variation. These findings highlight how behavioral responses to environmental inputs as thermal reaction norms can evolve through independent changes to genetically distinct modular components of such complex phenotypes.

Inbreeding and outbreeding depression in Caenorhabditis nematodes.

The nematode Caenorhabditis elegans reproduces primarily by self-fertilization of hermaphrodites, yet males are present at low frequencies in natural populations (androdioecy). The ancestral state of C. elegans was probably gonochorism (separate males and females), as in its relative C. remanei. Males may be maintained in C. elegans because outcrossed individuals escape inbreeding depression. The level of inbreeding depression is, however, expected to be low in such a highly selfing species, compared with an outcrosser like C. remanei. To investigate these issues, we measured life-history traits in the progeny of inbred versus outcrossed C. elegans and C. remanei individuals derived from recently isolated natural populations. In addition, we maintained inbred lines of C. remanei through 13 generations of full-sibling mating. Highly inbred C. remanei showed dramatic reductions in brood size and relative fitness compared to outcrossed individuals, with evidence of both direct genetic and maternal-effect inbreeding depression. This decline in fitness accumulated over time, causing extinction of nearly 90% of inbred lines, with no evidence of purging of deleterious mutations from the remaining lines. In contrast, pure strains of C. elegans performed better than crosses between strains, indicating outbreeding depression. The results are discussed in relation to the evolution of androdioecy and the effect of mating system on the level of inbreeding depression.

High nucleotide polymorphism and rapid decay of linkage disequilibrium in wild populations of Caenorhabditis remanei.

The common ancestor of the self-fertilizing nematodes Caenorhabditis elegans and C. briggsae must have reproduced by obligate outcrossing, like most species in this genus. However, we have only a limited understanding about how genetic variation is patterned in such male-female (gonochoristic) Caenorhabditis species. Here, we report results from surveying nucleotide variation of six nuclear loci in a broad geographic sample of wild isolates of the gonochoristic C. remanei. We find high levels of diversity in this species, with silent-site diversity averaging 4.7%, implying an effective population size close to 1 million. Additionally, the pattern of polymorphisms reveals little evidence for population structure or deviation from neutral expectations, suggesting that the sampled C. remanei populations approximate panmixis and demographic equilibrium. Combined with the observation that linkage disequilibrium between pairs of polymorphic sites decays rapidly with distance, this suggests that C. remanei will provide an excellent system for identifying the genetic targets of natural selection from deviant patterns of polymorphism and linkage disequilibrium. The patterns revealed in this obligately outcrossing species may provide a useful model of the evolutionary circumstances in C. elegans' gonochoristic progenitor. This will be especially important if self-fertilization evolved recently in C. elegans history, because most of the evolutionary time separating C. elegans from its known relatives would have occurred in a state of obligate outcrossing.

Insights into species divergence and the evolution of hermaphroditism from fertile interspecies hybrids of Caenorhabditis nematodes.

The architecture of both phenotypic variation and reproductive isolation are important problems in evolutionary genetics. The nematode genus Caenorhabditis includes both gonochoristic (male/female) and androdioecious (male/hermaprodite) species. However, the natural genetic variants distinguishing reproductive mode remain unknown, and nothing is known about the genetic basis of postzygotic isolation in the genus. Here we describe the hybrid genetics of the first Caenorhabditis species pair capable of producing fertile hybrid progeny, the gonochoristic Caenorhabditis sp. 9 and the androdioecious C. briggsae. Though many interspecies F(1) arrest during embryogenesis, a viable subset develops into fertile females and sterile males. Reciprocal parental crosses reveal asymmetry in male-specific viability, female fertility, and backcross viability. Selfing and spermatogenesis are extremely rare in XX F(1), and almost all hybrid self-progeny are inviable. Consistent with this, F(1) females do not express male-specific molecular germline markers. We also investigated three approaches to producing hybrid hermaphrodites. A dominant mutagenesis screen for self-fertile F(1) hybrids was unsuccessful. Polyploid F(1) hybrids with increased C. briggsae genomic material did show elevated rates of selfing, but selfed progeny were mostly inviable. Finally, the use of backcrosses to render the hybrid genome partial homozygous for C. briggsae alleles did not increase the incidence of selfing or spermatogenesis relative to the F(1) generation. These hybrid animals were genotyped at 23 loci, and significant segregation distortion (biased against C. briggsae) was detected at 13 loci. This, combined with an absence of productive hybrid selfing, prevents formulation of simple hypotheses about the genetic architecture of hermaphroditism. In the near future, this hybrid system will likely be fruitful for understanding the genetics of reproductive isolation in Caenorhabditis.

REPRODUCTIVE ISOLATION IN RHABDITIDAE (NEMATODA: SECERNENTEA); MECHANISMS THAT ISOLATE SIX SPECIES OF THREE GENERA.

We have attempted interspecific hybridizations among six species of rhabditid nematodes: Caenorhabditis elegans, Caenorhabditis briggsae, Caenorhabditis remanei, Caenorhabditis sp. v, Rhabditis sp., and Pelodera teres. Copulation was observed in all crosses between Caenorhabditis species; however, none resulted in the generation of stable hybrid populations. No copulation was observed in crosses between Caenorhabditis males and Rhabditis or Pelodera females, even when congeneric females were present, suggesting that Caenorhabditis males are able to selectively recognize congeneric females by a short-range stimulus. All pairwise combinations of Caenorhabditis species were isolated to some degree by gametic mechanisms; 7 of 12 combinations were cross infertile and 5 of 12 were cross-fertile but had low brood sizes. In cross-fertile combinations, most hybrid embryos were inviable and arrested prior to gastrulation. Only in crosses of C. briggsae males to C. sp. v females did any hybrids survive embryogenesis. Most of these C. briggsae/C. sp. v hybrids arrested during larval development, and the few that reached adulthood invariably were female. These results are consistent with the presence of at least two lethal factors in the C. briggsae-C. sp. v combination: a maternal lethal factor in the cytoplasm of C. briggsae and a recessive lethal factor on the X chromosome of C. sp. v.