Frequencies of the pyrethroid resistance alleles of Vssc1 and CYP6D1 in house flies from the eastern United States.

House flies were collected from four dairies in Maine, New York, North Carolina, and Florida, where high levels of resistance to permethrin have been documented. Regions of two genes, CYP6D1 and Vssc1, having alleles that confer resistance to permethrin (and other pyrethroids) were analysed from individuals at each collection site. The combinations of resistance alleles for Vssc1 and CYP6D1 were highly variable between each state. The resistance allele CYP6D1v1 was found at a high frequency (0.63-0.91) at all sites. Individuals homozygous susceptible for CYP6D1 were very rare and detected only at the dairy in Maine. In addition to the typical Vssc1 mutation responsible for resistance, kdr (L1014F), we also identified individuals with a L1014H mutation. Although house flies homozygous for the L1014H mutation had a lower level of resistance to permethrin, compared to L1014F, the H1014 resistance allele was frequently detected. No individuals with the super-kdr allele (M918T + L1014F) were detected from the field collections. The intron 3 bp downstream of the kdr mutation was found to be extremely variable, providing an opportunity to reconstruct a phylogeny of Vssc1 alleles. Based on this analysis it appears the kdr-his mutation had multiple evolutionary origins, but that the kdr mutation may have had a single origin. The impacts of these findings on resistance management are discussed.

Molecular genetics of mosquito resistance to malaria parasites.

Malaria parasites are transmitted by the bite of an infected mosquito, but even efficient vector species possess multiple mechanisms that together destroy most of the parasites present in an infection. Variation between individual mosquitoes has allowed genetic analysis and mapping of loci controlling several resistance traits, and the underlying mechanisms of mosquito response to infection are being described using genomic tools such as transcriptional and proteomic analysis. Malaria infection imposes fitness costs on the vector, but various forms of resistance inflict their own costs, likely leading to an evolutionary tradeoff between infection and resistance. Plasmodium development can be successfully completed onlyin compatible mosquito-parasite species combinations, and resistance also appears to have parasite specificity. Studies of Drosophila, where genetic variation in immunocompetence is pervasive in wild populations, offer a comparative context for understanding coevolution of the mosquito-malaria relationship. More broadly, plants also possess systems of pathogen resistance with features that are structurally conserved in animal innate immunity, including insects, and genomic datasets now permit useful comparisons of resistance models even between such diverse organisms.

Evidence for recurrent paralogous gene conversion and exceptional allelic divergence in the Attacin genes of Drosophila melanogaster.

Insects produce a limited variety of antibacterial peptides to combat a wide diversity of pathogens. These peptides are often conserved across evolutionarily distant taxa, but little is known about the level and structure of polymorphism within species. We have surveyed naturally occurring genetic variation in the promoter and coding regions of three Attacin antibacterial peptide genes from 12 lines of Drosophila melanogaster. These genes exhibit high levels of silent nucleotide variations (1-3% per nucleotide heterozygosity), but are not excessively polymorphic at the amino acid level. There is extensive variation in the Attacin promoters, some of which may affect transcriptional efficiency, and one line carries a deletion in the Attacin A coding region that renders this gene nonfunctional. Two of the genes, Attacins A and B, are arranged in tandem and show evidence of repeated interlocus gene conversion. Attacin C, more divergent and located 1.3 Mbp upstream of Attacins A and B, does not appear to have been involved in such exchanges. All three genes are characterized by divergent haplotypes, and one Attacin AB allele appears to have recently increased rapidly in frequency in the population.

Linkage disequilibria and the site frequency spectra in the su(s) and su(w(a)) regions of the Drosophila melanogaster X chromosome.

Over the last decade, surveys of DNA sequence variation in natural populations of several Drosophila species and other taxa have established that polymorphism is reduced in genomic regions characterized by low rates of crossing over per physical length. Parallel studies have also established that divergence between species is not reduced in these same genomic regions, thus eliminating explanations that rely on a correlation between the rates of mutation and crossing over. Several theoretical models (directional hitchhiking, background selection, and random environment) have been proposed as population genetic explanations. In this study samples from an African population (n = 50) and a European population (n = 51) were surveyed at the su(s) (1955 bp) and su(w(a)) (3213 bp) loci for DNA sequence polymorphism, utilizing a stratified SSCP/DNA sequencing protocol. These loci are located near the telomere of the X chromosome, in a region of reduced crossing over per physical length, and exhibit a significant reduction in DNA sequence polymorphism. Unlike most previously surveyed, these loci reveal substantial skews toward rare site frequencies, consistent with the predictions of directional hitchhiking and random environment models and inconsistent with the general predictions of the background selection model (or neutral theory). No evidence for excess geographic differentiation at these loci is observed. Although linkage disequilibrium is observed between closely linked sites within these loci, many recombination events in the genealogy of the sampled alleles can be inferred and the genomic scale of linkage disequilibrium, measured in base pairs between sites, is the same as that observed for loci in regions of normal crossing over. We conclude that gene conversion must be high in these regions of low crossing over.

Y chromosomal fertility factors kl-2 and kl-3 of Drosophila melanogaster encode dynein heavy chain polypeptides.

The molecular identity and function of the Drosophila melanogaster Y-linked fertility factors have long eluded researchers. Although the D. melanogaster genome sequence was recently completed, the fertility factors still were not identified, in part because of low cloning efficiency of heterochromatic Y sequences. Here we report a method for iterative blast searching to assemble heterochromatic genes from shotgun assemblies, and we successfully identify kl-2 and kl-3 as 1beta- and gamma-dynein heavy chains, respectively. Our conclusions are supported by formal genetics with X-Y translocation lines. Reverse transcription-PCR was successful in linking together unmapped sequence fragments from the whole-genome shotgun assembly, although some sequences were missing altogether from the shotgun effort and had to be generated de novo. We also found a previously undescribed Y gene, polycystine-related (PRY). The closest paralogs of kl-2, kl-3, and PRY (and also of kl-5) are autosomal and not X-linked, suggesting that the evolution of the Drosophila Y chromosome has been driven by an accumulation of male-related genes arising de novo from the autosomes.

Contrasting patterns of nucleotide polymorphism at the alcohol dehydrogenase locus in the outcrossing Arabidopsis lyrata and the selfing Arabidopsis thaliana.

Nucleotide variation at the alcohol dehydrogenase locus (Adh) was studied in the outcrossing Arabidopsis lyrata, a close relative of the selfing Arabidopsis thaliana. Overall, estimated nucleotide diversity in the North American ssp. lyrata and two European ssp. petraea populations was 0.0038, lower than the corresponding specieswide estimate for A. thaliana at the same set of nucleotide sites. The distribution of segregating sites across the gene differed between the two species. Estimated sequence diversity within an A. lyrata population with a large sample size (0.0023) was much higher than has previously been observed for A. thaliana. This North American population has an excess of sites at intermediate frequencies compared with neutral expectation (Tajima's D = 2.3, P < 0.005), suggestive of linked balancing selection or a recent population bottleneck. In contrast, an excess of rare polymorphisms has been found in A. thaliana. Polymorphism within A. lyrata and divergence from A. thaliana appear to be correlated across the Adh gene sequence. The geographic distribution of polymorphism was quite different from that of A. thaliana, for which earlier studies of several genes found low within-population nucleotide site polymorphism and no overall continental differentiation of variation despite large differences in site frequencies between local populations. Differences between the outcrossing A. lyrata and the selfing A. thaliana reflect the impact of differences in mating system and the influence of bottlenecks in A. thaliana during rapid colonization on DNA sequence polymorphism. The influence of additional variability-reducing mechanisms, such as background selection or hitchhiking, may not be discernible.