Detecting heterozygosity in shotgun genome assemblies: Lessons from obligately outcrossing nematodes.

The majority of nematodes are gonochoristic (dioecious) with distinct male and female sexes, but the best-studied species, Caenorhabditis elegans, is a self-fertile hermaphrodite. The sequencing of the genomes of C. elegans and a second hermaphrodite, C. briggsae, was facilitated in part by the low amount of natural heterozygosity, which typifies selfing species. Ongoing genome projects for gonochoristic Caenorhabditis species seek to approximate this condition by intense inbreeding prior to sequencing. Here we show that despite this inbreeding, the heterozygous fraction of the whole genome shotgun assemblies of three gonochoristic Caenorhabditis species, C. brenneri, C. remanei, and C. japonica, is considerable. We first demonstrate experimentally that independently assembled sequence variants in C. remanei and C. brenneri are allelic. We then present gene-based approaches for recognizing heterozygous regions of WGS assemblies. We also develop a simple method for quantifying heterozygosity that can be applied to assemblies lacking gene annotations. Consistently we find that approximately 10% and 30% of the C. remanei and C. brenneri genomes, respectively, are represented by two alleles in the assemblies. Heterozygosity is restricted to autosomes and its retention is accompanied by substantial inbreeding depression, suggesting that it is caused by multiple recessive deleterious alleles and not merely by chance. Both the overall amount and chromosomal distribution of heterozygous DNA is highly variable between assemblies of close relatives produced by identical methodologies, and allele frequencies have continued to change after strains were sequenced. Our results highlight the impact of mating systems on genome sequencing projects.

High levels of genetic variation exist in Aspergillus niger populations infecting Welwitschia mirabilis hook.

Aspergillus niger is an asexual, haploid fungus which infects the seeds of Namibia's national plant, Welwitschia mirabilis, severely affecting plant viability. We used 31 randomly amplified polymorphic DNA markers to assess genetic variation among 89 A. niger isolates collected from three W. mirabilis populations in the Namib Desert. While all isolates belonged to the same vegetative compatibility group, 84% were unique genotypes, and estimates of genotypic evenness and Simpson's index of diversity approached 1.0 in the three populations. Analysis of molecular variance revealed that 78% of the total variation sampled was among isolates from individual W. mirabilis plants. Lower, but significant, amounts of variation detected among isolates from different plants (12%) and different sites (10%) also indicated some site- and plant-level genetic differentiation. Total gene diversity (H(T) = 0.264) was mostly attributable to diversity within populations (H(S) = 0.217); the relatively low level of genetic differentiation among the sites (G(ST) = 0.141) suggests that gene flow is occurring among the three distant sites. Although sexual reproduction has never been observed in this fungus, parasexuality is a well-known phenomenon in laboratory strains. We thus attribute the high levels of genetic variation to parasexuality and/or wind-facilitated gene flow from an as of yet undocumented broader host range of the fungus on other desert vegetation. Given the apparent ease of transmission, high levels of genetic diversity, and potentially broad host range, A. niger infections of W. mirabilis may be extremely difficult to control or prevent.