Breakpoint structure of the Anopheles gambiae 2Rb chromosomal inversion

<p>Abstract</p> <p>Background</p> <p>Alternative arrangements of chromosome 2 inversions in <it>Anopheles gambiae </it>are important sources of population structure, and are associated with adaptation to environmental heterogeneity. The forces responsible for their origin and maintenance are incompletely understood. Molecular characterization of inversion breakpoints provides insight into how they arose, and provides the basis for development of molecular karyotyping methods useful in future studies.</p> <p>Methods</p> <p>Sequence comparison of regions near the cytological breakpoints of 2Rb allowed the molecular delineation of breakpoint boundaries. Comparisons were made between the standard 2R<it>+</it>^{<it>b </it>}arrangement in the <it>An. gambiae </it>PEST reference genome and the inverted 2R<it>b </it>arrangements in the <it>An. gambiae </it>M and S genome assemblies. Sequence differences between alternative 2R<it>b </it>arrangements were exploited in the design of a PCR diagnostic assay, which was evaluated against the known chromosomal banding pattern of laboratory colonies and field-collected samples from Mali and Cameroon.</p> <p>Results</p> <p>The breakpoints of the 7.55 Mb 2R<it>b </it>inversion are flanked by extensive runs of the same short (72 bp) tandemly organized sequence, which was likely responsible for chromosomal breakage and rearrangement. Application of the molecular diagnostic assay suggested that 2R<it>b </it>has a single common origin in <it>An. gambiae </it>and its sibling species, <it>Anopheles arabiensis</it>, and also that the standard arrangement (2R<it>+</it>^<it>b</it>) may have arisen twice through breakpoint reuse. The molecular diagnostic was reliable when applied to laboratory colonies, but its accuracy was lower in natural populations.</p> <p>Conclusions</p> <p>The complex repetitive sequence flanking the 2R<it>b </it>breakpoint region may be prone to structural and sequence-level instability. The 2R<it>b </it>molecular diagnostic has immediate application in studies based on laboratory colonies, but its usefulness in natural populations awaits development of complementary molecular tools.</p

Phylogenomics revealed migration routes and adaptive radiation timing of holarctic malaria mosquito species of the Maculipennis group

BackgroundPhylogenetic analyses of closely related species of mosquitoes are important for better understanding the evolution of traits contributing to transmission of vector-borne diseases. Six out of 41 dominant malaria vectors of the genus Anopheles in the world belong to the Maculipennis Group, which is subdivided into two Nearctic subgroups (Freeborni and Quadrimaculatus) and one Palearctic (Maculipennis) subgroup. Although previous studies considered the Nearctic subgroups as ancestral, details about their relationship with the Palearctic subgroup, and their migration times and routes from North America to Eurasia remain controversial. The Palearctic species An. beklemishevi is currently included in the Nearctic Quadrimaculatus subgroup adding to the uncertainties in mosquito systematics.ResultsTo reconstruct historic relationships in the Maculipennis Group, we conducted a phylogenomic analysis of 11 Palearctic and 2 Nearctic species based on sequences of 1271 orthologous genes. The analysis indicated that the Palearctic species An. beklemishevi clusters together with other Eurasian species and represents a basal lineage among them. Also, An. beklemishevi is related more closely to An. freeborni, which inhabits the Western United States, rather than to An. quadrimaculatus, a species from the Eastern United States. The time-calibrated tree suggests a migration of mosquitoes in the Maculipennis Group from North America to Eurasia about 20-25 million years ago through the Bering Land Bridge. A Hybridcheck analysis demonstrated highly significant signatures of introgression events between allopatric species An. labranchiae and An. beklemishevi. The analysis also identified ancestral introgression events between An. sacharovi and its Nearctic relative An. freeborni despite their current geographic isolation. The reconstructed phylogeny suggests that vector competence and the ability to enter complete diapause during winter evolved independently in different lineages of the Maculipennis Group.ConclusionsOur phylogenomic analyses reveal migration routes and adaptive radiation timing of Holarctic malaria vectors and strongly support the inclusion of An. beklemishevi into the Maculipennis Subgroup. Detailed knowledge of the evolutionary history of the Maculipennis Subgroup provides a framework for examining the genomic changes related to ecological adaptation and susceptibility to human pathogens. These genomic variations may inform researchers about similar changes in the future providing insights into the patterns of disease transmission in Eurasia

Analysis of the Aedes albopictus C6/36 genome provides insight into cell line utility for viral propagation

BACKGROUND: The 50-year-old Aedes albopictus C6/36 cell line is a resource for the detection, amplification, and analysis of mosquito-borne viruses including Zika, dengue, and chikungunya. The cell line is derived from an unknown number of larvae from an unspecified strain of Aedes albopictus mosquitoes. Toward improved utility of the cell line for research in virus transmission, we present an annotated assembly of the C6/36 genome. RESULTS: The C6/36 genome assembly has the largest contig N50 (3.3 Mbp) of any mosquito assembly, presents the sequences of both haplotypes for most of the diploid genome, reveals independent null mutations in both alleles of the Dicer locus, and indicates a male-specific genome. Gene annotation was computed with publicly available mosquito transcript sequences. Gene expression data from cell line RNA sequence identified enrichment of growth-related pathways and conspicuous deficiency in aquaporins and inward rectifier K+ channels. As a test of utility, RNA sequence data from Zika-infected cells were mapped to the C6/36 genome and transcriptome assemblies. Host subtraction reduced the data set by 89%, enabling faster characterization of nonhost reads. CONCLUSIONS: The C6/36 genome sequence and annotation should enable additional uses of the cell line to study arbovirus vector interactions and interventions aimed at restricting the spread of human disease

Radical remodeling of the Y chromosome in a recent radiation of malaria mosquitoes

open28openHall A.B.; Papathanos P.-A.; Sharma A.; Cheng C.; Akbari O.S.; Assour L.; Bergman N.H.; Cagnetti A.; Crisanti A.; Dottorini T.; Fiorentini E.; Galizi R.; Hnath J.; Jiang X.; Koren S.; Nolan T.; Radune D.; Sharakhova M.V.; Steele A.; Timoshevskiy V.A.; Windbichler N.; Zhang S.; Hahn M.W.; Phillippy A.M.; Emrich S.J.; Sharakhov I.V.; Tu Z.J.; Besansky N.J.Hall, A. B.; Papathanos, P. -A.; SHARMA DHAKAL, Apsara; Cheng, C.; Akbari, O. S.; Assour, L.; Bergman, N. H.; Cagnetti, A.; Crisanti, A.; Dottorini, T.; Fiorentini, E.; Galizi, R.; Hnath, J.; Jiang, X.; Koren, S.; Nolan, T.; Radune, D.; Sharakhova, M. V.; Steele, A.; Timoshevskiy, V. A.; Windbichler, N.; Zhang, Shangu; Hahn, M. W.; Phillippy, A. M.; Emrich, S. J.; Sharakhov, I. V.; Tu, Z. J.; Besansky, N. J

Evolutionary Dynamics of the Ty3/Gypsy LTR Retrotransposons in the Genome of Anopheles gambiae

Ty3/gypsy elements represent one of the most abundant and diverse LTR-retrotransposon (LTRr) groups in the Anopheles gambiae genome, but their evolutionary dynamics have not been explored in detail. Here, we conduct an in silico analysis of the distribution and abundance of the full complement of 1045 copies in the updated AgamP3 assembly. Chromosomal distribution of Ty3/gypsy elements is inversely related to arm length, with densities being greatest on the X, and greater on the short versus long arms of both autosomes. Taking into account the different heterochromatic and euchromatic compartments of the genome, our data suggest that the relative abundance of Ty3/gypsy LTRrs along each chromosome arm is determined mainly by the different proportions of heterochromatin, particularly pericentric heterochromatin, relative to total arm length. Additionally, the breakpoint regions of chromosomal inversion 2La appears to be a haven for LTRrs. These elements are underrepresented more than 7-fold in euchromatin, where 33% of the Ty3/gypsy copies are associated with genes. The euchromatin on chromosome 3R shows a faster turnover rate of Ty3/gypsy elements, characterized by a deficit of proviral sequences and the lowest average sequence divergence of any autosomal region analyzed in this study. This probably reflects a principal role of purifying selection against insertion for the preservation of longer conserved syntenyc blocks with adaptive importance located in 3R. Although some Ty3/gypsy LTRrs show evidence of recent activity, an important fraction are inactive remnants of relatively ancient insertions apparently subject to genetic drift. Consistent with these computational predictions, an analysis of the occupancy rate of putatively older insertions in natural populations suggested that the degenerate copies have been fixed across the species range in this mosquito, and also are shared with the sibling species Anopheles arabiensis

Imaginal Discs – A New Source of Chromosomes for Genome Mapping of the Yellow Fever Mosquito Aedes aegypti

Dengue fever is an emerging health threat to as much as half of the human population around the world. No vaccines or drug treatments are currently available. Thus, disease prevention is largely based on efforts to control its major mosquito vector Ae. aegypti. Novel vector control strategies, such as population replacement with pathogen-incompetent transgenic mosquitoes, rely on detailed knowledge of the genome organization for the mosquito. However, the current genome assembly of Ae. aegypti is highly fragmented and requires additional physical mapping onto chromosomes. The absence of readable polytene chromosomes makes genome mapping for this mosquito extremely challenging. In this study, we discovered and investigated a new source of chromosomes useful for the cytogenetic analysis in Ae. aegypti – mitotic chromosomes from imaginal discs of 4th instar larvae. Using natural banding patterns of these chromosomes, we developed a new band-based approach for physical mapping of DNA probes to the precise chromosomal positions. Further application of this approach for genome mapping will greatly enhance the utility of the existing draft genome sequence assembly for Ae. aegypti and thereby facilitate application of advanced genome technologies for investigating and developing novel genetic control strategies for dengue transmission

Arm-specific dynamics of chromosome evolution in malaria mosquitoes

<p>Abstract</p> <p>Background</p> <p>The malaria mosquito species of subgenus <it>Cellia </it>have rich inversion polymorphisms that correlate with environmental variables. Polymorphic inversions tend to cluster on the chromosomal arms 2R and 2L but not on X, 3R and 3L in <it>Anopheles gambiae </it>and homologous arms in other species. However, it is unknown whether polymorphic inversions on homologous chromosomal arms of distantly related species from subgenus <it>Cellia </it>nonrandomly share similar sets of genes. It is also unclear if the evolutionary breakage of inversion-poor chromosomal arms is under constraints.</p> <p>Results</p> <p>To gain a better understanding of the arm-specific differences in the rates of genome rearrangements, we compared gene orders and established syntenic relationships among <it>Anopheles gambiae, Anopheles funestus</it>, and <it>Anopheles stephensi</it>. We provided evidence that polymorphic inversions on the 2R arms in these three species nonrandomly captured similar sets of genes. This nonrandom distribution of genes was not only a result of preservation of ancestral gene order but also an outcome of extensive reshuffling of gene orders that created new combinations of homologous genes within independently originated polymorphic inversions. The statistical analysis of distribution of conserved gene orders demonstrated that the autosomal arms differ in their tolerance to generating evolutionary breakpoints. The fastest evolving 2R autosomal arm was enriched with gene blocks conserved between only a pair of species. In contrast, all identified syntenic blocks were preserved on the slowly evolving 3R arm of <it>An. gambiae </it>and on the homologous arms of <it>An. funestus </it>and <it>An. stephensi</it>.</p> <p>Conclusions</p> <p>Our results suggest that natural selection favors specific gene combinations within polymorphic inversions when distant species are exposed to similar environmental pressures. This knowledge could be useful for the discovery of genes responsible for an association of inversion polymorphisms with phenotypic variations in multiple species. Our data support the chromosomal arm specificity in rates of gene order disruption during mosquito evolution. We conclude that the distribution of breakpoint regions is evolutionary conserved on slowly evolving arms and tends to be lineage-specific on rapidly evolving arms.</p

Molecular characterization and evolution of a gene family encoding male-specific reproductive proteins in the African malaria vector Anopheles gambiae

<p>Abstract</p> <p>Background</p> <p>During copulation, the major Afro-tropical malaria vector <it>Anopheles gambiae </it>s.s. transfers male accessory gland (MAG) proteins to females as a solid mass (i.e. the "mating plug"). These proteins are postulated to function as important modulators of female post-mating responses. To understand the role of selective forces underlying the evolution of these proteins in the <it>A. gambiae </it>complex, we carried out an evolutionary analysis of gene sequence and expression divergence on a pair of paralog genes called <it>AgAcp34A-1 </it>and <it>AgAcp34A-2</it>. These encode MAG-specific proteins which, based on homology with <it>Drosophila</it>, have been hypothesized to play a role in sperm viability and function.</p> <p>Results</p> <p>Genetic analysis of 6 species of the <it>A. gambiae </it>complex revealed the existence of a third paralog (68-78% of identity), that we named <it>AgAcp34A-3</it>. FISH assays showed that this gene maps in the same division (34A) of chromosome-3R as the other two paralogs. In particular, immuno-fluorescence assays targeting the C-terminals of <it>AgAcp34A-2 </it>and <it>AgAcp34A-3 </it>revealed that these two proteins are localized in the posterior part of the MAG and concentrated at the apical portion of the mating plug. When transferred to females, this part of the plug lies in proximity to the duct connecting the spermatheca to the uterus, suggesting a potential role for these proteins in regulating sperm motility. <it>AgAcp34A-3 </it>is more polymorphic than the other two paralogs, possibly because of relaxation of purifying selection. Since both unequal crossing-over and gene conversion likely homogenized the members of this gene family, the interpretation of the evolutionary patterns is not straightforward. Although several haplotypes of the three paralogs are shared by most <it>A. gambiae </it>s.l. species, some fixed species-specific replacements (mainly placed in the N- and C-terminal portions of the secreted peptides) were also observed, suggesting some lineage-specific adaptation.</p> <p>Conclusions</p> <p>Progress in understanding the signaling cascade in the <it>A. gambiae </it>reproductive pathway will elucidate the interaction of this MAG-specific protein family with their female counterparts. This knowledge will allow a better evaluation of the relative importance of genes involved in the reproductive isolation and fertility of <it>A. gambiae </it>species and could help the interpretation of the observed evolutionary patterns.</p