Genome report: chromosome-scale genome assembly of the West Indian fruit fly Anastrepha obliqua (Diptera: Tephritidae).

The West Indian fruit fly, Anastrepha obliqua, is a major pest of mango in Central and South America and attacks more than 60 species of host fruits. To support current genetic and genomic research on A. obliqua, we sequenced the genome using high-fidelity long-read sequencing. This resulted in a highly contiguous contig assembly with 90% of the genome in 10 contigs. The contig assembly was placed in a chromosomal context using synteny with a closely related species, Anastrepha ludens, as both are members of the Anastrepha fraterculus group. The resulting assembly represents the five autosomes and the X chromosome which represents 95.9% of the genome, and 199 unplaced contigs representing the remaining 4.1%. Orthology analysis across the structural annotation sets of high quality tephritid genomes demonstrates the gene annotations are robust, and identified genes unique to Anastrepha species that may help define their pestiferous nature that can be used as a starting point for comparative genomics. This genome assembly represents the first of this species and will serve as a foundation for future genetic and genomic research in support of its management as an agricultural pest.

Genomic Tools in Biological Invasions: Current State and Future Frontiers.

Human activities are accelerating rates of biological invasions and climate-driven range expansions globally, yet we understand little of how genomic processes facilitate the invasion process. Although most of the literature has focused on underlying phenotypic correlates of invasiveness, advances in genomic technologies are showing a strong link between genomic variation and invasion success. Here, we consider the ability of genomic tools and technologies to (i) inform mechanistic understanding of biological invasions and (ii) solve real-world issues in predicting and managing biological invasions. For both, we examine the current state of the field and discuss how genomics can be leveraged in the future. In addition, we make recommendations pertinent to broader research issues, such as data sovereignty, metadata standards, collaboration, and science communication best practices that will require concerted efforts from the global invasion genomics community.

Comparative Mitogenome Analysis of Two Native Apple Snail Species (Ampullariidae, Pomacea) from Peruvian Amazon.

Apple snails of the genus Pomacea Perry, 1810 (Mollusca: Caenogastropoda: Ampullariidae) are native to the Neotropics and exhibit high species diversity, holding cultural and ecological significance as an important protein source in Peru. However, most genetic studies in Pomacea have focused mostly on invasive species, especially in Southeast Asia, where they are considered important pests. In this study, we assembled and annotated the mitochondrial genomes of two Pomacea species native to the Peruvian Amazon: Pomacea reevei Ampuero & Ramírez, 2023 and Pomacea aulanieri (Deville & Hupé, 1850). The mitogenomes of P. reevei and P. aulanieri comprise 15,660 and 16,096 bp, respectively, and contain the typical 37 genes of the animal mitochondria with a large control region of 292 bp in P. reevei and 524 bp in P. aulanieri-which fall within the range of what is currently known in Pomacea. Comparisons with previously published mitogenomes in Pomacea revealed differences in the overlapping of adjacent genes, the size of certain protein-coding genes (PCGs) and the secondary structure of some tRNAs that are consistent with the phylogenetic relationships between these species. These findings provide valuable insights into the systematics and genomics of the genus Pomacea.

Phylogenomic analysis provides diagnostic tools for the identification of Anastrepha fraterculus (Diptera: Tephritidae) species complex.

Insect pests cause tremendous impact to agriculture worldwide. Species identification is crucial for implementing appropriate measures of pest control but can be challenging in closely related species. True fruit flies of the genus Anastrepha Schiner (Diptera: Tephritidae) include some of the most serious agricultural pests in the Americas, with the Anastrepha fraterculus (Wiedemann) complex being one of the most important due to its extreme polyphagy and wide distribution across most of the New World tropics and subtropics. The eight morphotypes described for this complex as well as other closely related species are classified in the fraterculus species group, whose evolutionary relationships are unresolved due to incomplete lineage sorting and introgression. We performed multifaceted phylogenomic approaches using thousands of genes to unravel the evolutionary relationships within the A. fraterculus complex to provide a baseline for molecular diagnosis of these pests. We used a methodology that accommodates variable sources of data (transcriptome, genome, and whole-genome shotgun sequencing) and developed a tool to align and filter orthologs, generating reliable datasets for phylogenetic studies. We inferred 3031 gene trees that displayed high levels of discordance. Nevertheless, the topologies of the inferred coalescent species trees were consistent across methods and datasets, except for one lineage in the A. fraterculus complex. Furthermore, network analysis indicated introgression across lineages in the fraterculus group. We present a robust phylogeny of the group that provides insights into the intricate patterns of evolution of the A. fraterculus complex supporting the hypothesis that this complex is an assemblage of closely related cryptic lineages that have evolved under interspecific gene flow. Despite this complex evolutionary scenario, our subsampling analysis revealed that a set of as few as 80 loci has a similar phylogenetic resolution as the genome-scale dataset, offering a foundation to develop more efficient diagnostic tools in this species group.

HiMAP2: Identifying phylogenetically informative genetic markers from diverse genomic resources.

Multiplexed amplicon sequencing offers a cost-effective and rapid solution for phylogenomic studies that include a large number of individuals. Selecting informative genetic markers is a critical initial step in designing such multiplexed amplicon panels, but screening various genomic data and selecting markers that are informative for the question at hand can be laborious. Here, we present a flexible and user-friendly tool, HiMAP2, for identifying, visualizing and filtering phylogenetically informative loci from diverse genomic and transcriptomic resources. This bioinformatics pipeline includes orthology prediction, exon extraction and filtering of aligned exon sequences according to user-defined specifications. Additionally, HiMAP2 facilitates exploration of the final filtered exons by incorporating phylogenetic inference of individual exon trees with raxml-ng as well as the estimation of a species tree using astral. Finally, results of the marker selection can be visualized and refined with an interactive Bokeh application that can be used to generate publication-quality figures. Source code and user instructions for HiMAP2 are available at https://github.com/popphylotools/HiMAP_v2.

Phylogenomic approach reveals strong signatures of introgression in the rapid diversification of neotropical true fruit flies (Anastrepha: Tephritidae).

New sequencing techniques have allowed us to explore the variation on thousands of genes and elucidate evolutionary relationships of lineages even in complex scenarios, such as when there is rapid diversification. That seems to be the case of species in the genus Anastrepha, which shows great species diversity that has been divided into 21 species groups, several of which show wide geographical distribution. The fraterculus group has several economically important species and it is also an outstanding model for speciation studies, since it includes several lineages that have diverged recently possibly in the presence of interspecific gene flow. Our main goal is to test whether we can infer phylogenetic relationships of recently diverged taxa with gene flow, such as what is expected for the fraterculus group and determine whether certain genes remain informative even in this complex scenario. An analysis of thousands of orthologous genes derived from transcriptome datasets of 10 different lineages across the genus, including some of the economically most important pests, revealed signals of incomplete lineage sorting, vestiges of ancestral introgression between more distant lineages and ongoing gene flow between closely related lineages. Though these patterns affect the phylogenetic signal, the phylogenomic inferences consistently show that the morphologically identified species here investigated are in different evolutionary lineages, with the sole exception involving Brazilian lineages of A. fraterculus, which has been suggested to be a complex assembly of cryptic species. A tree space analysis suggested that genes with greater phylogenetic resolution have evolved under similar selection pressures and are more resilient to intraspecific gene flow, which would make it more likely that these genomic regions may be useful for identifying fraterculus group lineages. Our findings help establish relationships among the most important Anastrepha species groups, as well as bring further data to indicate that the diversification of fraterculus group lineages, and even other lineages in the genus Anastrepha, has been strongly influenced by interspecific gene flow.

Identificación molecular y relaciones evolutivas de Pomacea nobilis, base para la autenticación específica del churo negro de la Amazonia peruana / Molecular identification and evolutionary relationships of Pomacea nobilis, basis for the specific authentication of the black apple snail from Peruvian Amazonia

Resumen En la Amazonia Peruana los caracoles dulceacuícolas de la familia Ampullariidae son conocidos como churos y originalmente han sido descritas para Perú alrededor de 20 especies. Aunque son muy usadas para alimentación, medicina tradicional y objeto de muchos estudios para su cultivo e industrialización, solamente es mencionada en la literatura la especie Pomacea maculata. Se llevó a cabo la identificación molecular sobre la base del marcador mitocondrial COI, de individuos de churos negros (Pomacea) comercializados en los mercados de Iquitos, así como los usados en platos a la carta en la ciudad de Lima, contrastados con otros individuos de procedencia de su hábitat natural. Se encontró que estos especímenes expendidos corresponden a la especie Pomacea nobilis (Reeve, 1856). El análisis filogenético molecular mostró que P. nobilis es especie hermana de P. guyanensis, en el grupo de P. glauca, distantemente relacionada de P. maculata. Las distancias no corregidas encontradas entre ellas, para el marcador mitocondrial COI, fueron de 11.33% a 13.17%, mientras que con P. maculata fueron de 13.67% a 15.33%. Estos resultados demostraron la eficacia del código de barras de ADN para la identificación y autenticación de la especie, lo que le da un valor agregado para su eventual comercio de exportación.

Abstract In the Peruvian Amazon, freshwater snails of the Ampullariidae family are known as churos, and around 20 species have originally been described for Peru. Although they are widely used for food, traditional medicine and the object of many studies for their cultivation and industrialization, only the species Pomacea maculata is mentioned in the literature. Molecular identification was carried out based on the mitochondrial marker COI of individuals of "churo negro" apple snails (Pomacea) commercialized in the markets of Iquitos, as well as those used in restaurant dishes in the city of Lima, and contrasted with specimens from their natural habitat. It was found that these specimens, correspond to the species Pomacea nobilis (Reeve, 1856). The molecular phylogenetic analysis showed P. nobilis as the sister species of P. guyanensis, in the P. glauca group, distantly related to P. maculata. The uncorrected distances found between them, for the mitochondrial marker COI, were from 11.33% to 13.17%, while with P. maculate were from 13.67% to 15.33%. These results demonstrated the effectiveness of the DNA barcode for the identification and authentication of the species, which gives it added value for its eventual export trade.

Avian haemosporidians in the cattle egret (Bubulcus ibis) from central-western and southern Africa: High diversity and prevalence.

We described the geographic distribution of 82 haemosporidian lineages (Plasmodium, Haemoproteus, and Leucocytozoon) in the cattle egret sampled in five countries in central-western and southern Africa. Seventy-three lineages have not previously been reported. We determined the prevalence of three haemosporidians in the samples. We investigated the influence of the internal environment of the host and environmental variables on the Plasmodium diversity and whether environmental variables may explain spatial variations in the prevalence of Plasmodium. We screened DNA from 509 blood samples from nestlings in 15 African colonies for infection by sequencing the cytochrome b gene of parasites. The molecular phylogenetic analysis was performed using Bayesian methods and including sequences from the MalAvi and GeneBank databases. We found 62 new Plasmodium lineages in a clade with MYCAME02, which is a lineage described in waterbirds and recently identified in birds of prey as Plasmodium paranucleophilum. Two Haemoproteus lineages identified in cattle egret formed a distinct group with Haemoproteus catharti and MYCAMH1 (Haemoproteus spp.). Seven Leucocytozoon lineages found in the cattle egret clustered with Leucocytozoon californicus. We found different Plasmodium diversities among the colonies sampled, demonstrating that the internal environment of the host is not the primary determinant of diversity. A linear mixed-effects multivariate model showed that precipitation was positively associated with Plasmodium diversity when controlling for the effects of temperature, colony composition (mixed and non-mixed species) and country. Moreover, a generalized mixed model showed that temperature was positively associated with the prevalence of Plasmodium when controlling for precipitation, elevation and country. We conclude that the cattle egret is a good model for future haemosporidian studies, as we found a significant number of new lineages in this host, which occupies regions with different climate characteristics where environmental variables exert an influence on the diversity and prevalence of Plasmodium.

Evidence of Adaptive Evolution and Relaxed Constraints in Sex-Biased Genes of South American and West Indies Fruit Flies (Diptera: Tephritidae).

Several studies have demonstrated that genes differentially expressed between sexes (sex-biased genes) tend to evolve faster than unbiased genes, particularly in males. The reason for this accelerated evolution is not clear, but several explanations have involved adaptive and nonadaptive mechanisms. Furthermore, the differences of sex-biased expression patterns of closely related species are also little explored out of Drosophila. To address the evolutionary processes involved with sex-biased expression in species with incipient differentiation, we analyzed male and female transcriptomes of Anastrepha fraterculus and Anastrepha obliqua, a pair of species that have diverged recently, likely in the presence of gene flow. Using these data, we inferred differentiation indexes and evolutionary rates and tested for signals of selection in thousands of genes expressed in head and reproductive transcriptomes from both species. Our results indicate that sex-biased and reproductive-biased genes evolve faster than unbiased genes in both species, which is due to both adaptive pressure and relaxed constraints. Furthermore, among male-biased genes evolving under positive selection, we identified some related to sexual functions such as courtship behavior and fertility. These findings suggest that sex-biased genes may have played important roles in the establishment of reproductive isolation between these species, due to a combination of selection and drift, and unveil a plethora of genetic markers useful for more studies in these species and their differentiation.

Odorant-binding proteins expression patterns in recently diverged species of Anastrepha fruit flies.

We studied two species of closely related South American fruit flies, Anastrepha fraterculus and Anastrepha obliqua which, despite being able to interbreed, still show some ecological and reproductive differences. Because part of these differences, such as host and mate preferences, may be related to olfactory perception, we focused our investigation on the differential expression of Odorant-binding protein (OBP) gene family, which participate in initial steps of the olfactory signal transduction cascade. We investigated patterns of expression of eight OBP genes by qPCR in male and female head tissues of both species. The expression patterns of these OBPs suggest that some OBP genes are more likely involved with the location of food resources, while others seem to be associated with mate and pheromone perception. Furthermore, the expression patterns obtained at different reproductive stages indicate that OBP expression levels changed significantly after mating in males and females of both species. All eight OBP genes analyzed here showed significant levels of differential expression between A. fraterculus and A. obliqua, suggesting that they may hold important roles in their olfactory perception differences, and consequently, may potentially be involved in their differentiation.

Differential transcriptome analysis supports Rhodnius montenegrensis and Rhodnius robustus (Hemiptera, Reduviidae, Triatominae) as distinct species.

Chagas disease is one of the main parasitic diseases found in Latin America and it is estimated that between six and seven million people are infected worldwide. Its etiologic agent, the protozoan Trypanosoma cruzi, is transmitted by triatomines, some of which from the genus Rhodnius. Twenty species are currently recognized in this genus, including some closely related species with low levels of morphological differentiation, such as Rhodnius montenegrensis and Rhodnius robustus. In order to investigate genetic differences between these two species, we generated large-scale RNA-sequencing data (consisting of four RNA-seq libraries) from the heads and salivary glands of males of R. montenegrensis and R. robustus. Transcriptome assemblies produced for each species resulted in 64,952 contigs for R. montenegrensis and 70,894 contigs for R. robustus, with N50 of approximately 2,100 for both species. SNP calling based on the more complete R. robustus assembly revealed 3,055 fixed interspecific differences and 216 transcripts with high levels of divergence which contained only fixed differences between the two species. A gene ontology enrichment analysis revealed that these highly differentiated transcripts were enriched for eight GO terms related to AP-2 adaptor complex, as well as other interesting genes that could be involved in their differentiation. The results show that R. montenegrensis and R. robustus have a substantial quantity of fixed interspecific polymorphisms, which suggests a high degree of genetic divergence between the two species and likely corroborates the species status of R. montenegrensis.

Phylogeography of Partamona rustica (Hymenoptera, Apidae), an Endemic Stingless Bee from the Neotropical Dry Forest Diagonal.

The South America encompasses the highest levels of biodiversity found anywhere in the world and its rich biota is distributed among many different biogeographical regions. However, many regions of South America are still poorly studied, including its xeric environments, such as the threatened Caatinga and Cerrado phytogeographical domains. In particular, the effects of Quaternary climatic events on the demography of endemic species from xeric habitats are poorly understood. The present study uses an integrative approach to reconstruct the evolutionary history of Partamona rustica, an endemic stingless bee from dry forest diagonal in Brazil, in a spatial-temporal framework. In this sense, we sequenced four mitochondrial genes and genotyped eight microsatellite loci. Our results identified two population groups: one to the west and the other to the east of the São Francisco River Valley (SFRV). These groups split in the late Pleistocene, and the Approximate Bayesian Computation approach and phylogenetic reconstruction indicated that P. rustica originated in the west of the SFRV, subsequently colonising eastern region. Our tests of migration detected reduced gene flow between these groups. Finally, our results also indicated that the inferences both from the genetic data analyses and from the spatial distribution modelling are compatible with historical demographic stability.

Head Transcriptomes of Two Closely Related Species of Fruit Flies of the Anastrepha fraterculus Group Reveals Divergent Genes in Species with Extensive Gene Flow.

Several fruit flies species of the Anastrepha fraterculus group are of great economic importance for the damage they cause to a variety of fleshy fruits. Some species in this group have diverged recently, with evidence of introgression, showing similar morphological attributes that render their identification difficult, reinforcing the relevance of identifying new molecular markers that may differentiate species. We investigated genes expressed in head tissues from two closely related species: A. obliqua and A. fraterculus, aiming to identify fixed single nucleotide polymorphisms (SNPs) and highly differentiated transcripts, which, considering that these species still experience some level of gene flow, could indicate potential candidate genes involved in their differentiation process. We generated multiple libraries from head tissues of these two species, at different reproductive stages, for both sexes. Our analyses indicate that the de novo transcriptome assemblies are fairly complete. We also produced a hybrid assembly to map each species' reads, and identified 67,470 SNPs in A. fraterculus, 39,252 in A. obliqua, and 6386 that were common to both species. We identified 164 highly differentiated unigenes that had a mean interspecific index ([Formula: see text]) of at least 0.94. We selected unigenes that had Ka/Ks higher than 0.5, or had at least three or more highly differentiated SNPs as potential candidate genes for species differentiation. Among these candidates, we identified proteases, regulators of redox homeostasis, and an odorant-binding protein (Obp99c), among other genes. The head transcriptomes described here enabled the identification of thousands of genes hitherto unavailable for these species, and generated a set of candidate genes that are potentially important to genetically identify species and understand the speciation process in the presence of gene flow of A. obliqua and A. fraterculus.

Blood parasites in nestlings of wood stork populations from three regions of the American continent.

This study documents the prevalences and lineages of hemoparasites in wood stork nestlings from 3 regions of the American continent: southeastern United States (n = 90), northern Brazil (n = 74), and central-western Brazil (n = 125). Identification was based on PCR amplification of a mitochondrial small subunit ribosomal RNA gene. A fragment of the hemoparasite cytochrome B gene in infected individuals was utilized for Bayesian phylogenetic analysis. Four wood stork nestlings were infected by Haemoproteus, 1 from northern Brazil and 3 from the United States, and all shared the same haplotype. Morphological analysis confirmed the infection of the U.S. birds by Haemoproteus. Infection by Plasmodium was found in wood stork nestlings from northern (6) and central-western Brazil (14). Five Plasmodium lineages (MYCAMP1-2, and MYCAMP4-6) were found in the Brazilian central-western region and 3 Plasmodium lineages (MYCAMP2-3, and MYCAMP7) were found in the northern region. The most prevalent haplotype (MYCAMP2) differs from the others by 1 mutation, and the less prevalent haplotypes are derived from MYCAMP2. We did not find Plasmodium or Haemoproteus in nestlings younger than 15 and 30 days old, respectively. This is the first documentation of Plasmodium and Haemoproteus infection in wood storks in Brazilian breeding populations. Potential connectivity among wood stork populations was indirectly supported by the presence of identical Haemoproteus lineages in U.S. and northern Brazilian populations, and by the presence of identical Plasmodium haplotypes in the northern and central-western Brazilian populations.