Phenotypic and genetic characterization of Africanized Apis mellifera colonies with natural tolerance to Varroa destructor and contrasting defensive behavior.

Africanized Apis mellifera colonies with promising characteristics for beekeeping have been detected in northern Argentina (subtropical climate) and are considered of interest for breeding programs. Integral evaluation of this feral material revealed high colony strength and resistance/tolerance to brood diseases. However, these Africanized honeybees (AHB) also showed variable negative behavioral traits for beekeeping, such as defensiveness, tendency to swarm and avoidance behavior. We developed a protocol for the selection of AHB stocks based on defensive behavior and characterized contrasting colonies for this trait using NGS technologies. For this purpose, population and behavioral parameters were surveyed throughout a beekeeping season in nine daughter colonies obtained from a mother colony (A1 mitochondrial haplotype) with valuable characteristics (tolerance to the mite Varroa destructor, high colony strength and low defensiveness). A Defensive Behavior Index was developed and tested in the colonies under study. Mother and two daughter colonies displaying contrasting defensive behavior were analyzed by ddRADseq. High-quality DNA samples were obtained from 16 workers of each colony. Six pooled samples, including two replicates of each of the three colonies, were processed. A total of 12,971 SNPs were detected against the reference genome of A. mellifera, 142 of which showed significant differences between colonies. We detected SNPs in coding regions, lncRNA, miRNA, rRNA, tRNA, among others. From the original data set, we also identified 647 SNPs located in protein-coding regions, 128 of which are related to 21 genes previously associated with defensive behavior, such as dop3 and dopR2, CaMKII and ADAR, obp9 and obp10, and members of the 5-HT family. We discuss the obtained results by considering the influence of polyandry and paternal lineages on the defensive behavior in AHB and provide baseline information to use this innovative molecular approach, ddRADseq, to assist in the selection and evaluation of honey bee stocks showing low defensive behavior for commercial uses.

Transcriptome analysis of Anastrepha fraterculus sp. 1 males, females, and embryos: insights into development, courtship, and reproduction.

BACKGROUND: Anastrepha fraterculus sp. 1 is considered a quarantine pest in several American countries. Since chemical control applied in an integrated pest management program is the only strategy utilized against this pest, the development of pesticide-free methods, such as the Sterile Insect Technique, is being considered. The search for genes involved in sex-determination and differentiation, and in metabolic pathways associated with communication and mating behaviour, contributes with key information to the development of genetic control strategies. The aims of this work were to perform a comprehensive analysis of A. fraterculus sp. 1 transcriptome and to obtain an initial evaluation of genes associated with main metabolic pathways by the expression analysis of specific transcripts identified in embryos and adults. RESULTS: Sexually mature adults of both sexes and 72 h embryos were considered for transcriptome analysis. The de novo transcriptome assembly was fairly complete (62.9% complete BUSCO orthologs detected) with a total of 86,925 transcripts assembled and 28,756 GO annotated sequences. Paired-comparisons between libraries showed 319 transcripts differently expressed between embryos and females, 1242 between embryos and males, and 464 between sexes. Using this information and genes searches based on published studies from other tephritid species, we evaluated a set of transcripts involved in development, courtship and metabolic pathways. The qPCR analysis evidenced that the early genes serendipity alpha and transformer-2 displayed similar expression levels in the analyzed stages, while heat shock protein 27 is over-expressed in embryos and females in comparison to males. The expression of genes associated with courtship (takeout-like, odorant-binding protein 50a1) differed between males and females, independently of their reproductive status (virgin vs mated individuals). Genes associated with metabolic pathways (maltase 2-like, androgen-induced gene 1) showed differential expression between embryos and adults. Furthermore, 14,262 microsatellite motifs were identified, with 11,208 transcripts containing at least one simple sequence repeat, including 48% of di/trinucleotide motifs. CONCLUSION: Our results significantly expand the available gene space of A. fraterculus sp. 1, contributing with a fairly complete transcript database of embryos and adults. The expression analysis of the selected candidate genes, along with a set of microsatellite markers, provides a valuable resource for further genetic characterization of A. fraterculus sp. 1 and supports the development of specific genetic control strategies.

Geographic distribution of sex chromosome polymorphism in Anastrepha fraterculus sp. 1 from Argentina.

BACKGROUND: Anastrepha fraterculus is recognized as a quarantine pest in several American countries. This fruit fly species is native to the American continent and distributed throughout tropical and subtropical regions. It has been reported as a complex of cryptic species, and at least eight morphotypes have been described. Only one entity of this complex, formerly named Anastrepha fraterculus sp. 1, is present in Argentina. Previous cytogenetic studies on this morphotype described the presence of sex chromosome variation identified by chromosomal size and staining patterns. In this work, we expanded the cytological study of this morphotype by analyzing laboratory strains and wild populations to provide information about the frequency and geographic distribution of these sex chromosome variants. We analyzed the mitotic metaphases of individuals from four laboratory strains and five wild populations from the main fruit-producing areas of Argentina, including the northwest (Tucumán and La Rioja), northeast (Entre Ríos and Misiones), and center (Buenos Aires) of the country. RESULTS: In wild samples, we observed a high frequency of X1X1 (0.94) and X1Y5 (0.93) karyomorphs, whereas X1X2 and X1Y6 were exclusively found at a low frequency in Buenos Aires (0.07 and 0.13, respectively), Entre Ríos (0.16 and 0.14, respectively) and Tucumán (0.03 and 0.04, respectively). X2X2 and X2Y5 karyomorphs were not found in wild populations but were detected at a low frequency in laboratory strains. In fact, karyomorph frequencies differed between wild populations and laboratory strains. No significant differences among A. fraterculus wild populations were evidenced in either karyotypic or chromosomal frequencies. However, a significant correlation was observed between Y5 chromosomal frequency and latitude. CONCLUSIONS: We discuss the importance of cytogenetics to understand the possible route of invasion and dispersion of this pest in Argentina and the evolutionary forces acting under laboratory conditions, possibly driving changes in the chromosomal frequencies. Our findings provide deep and integral genetic knowledge of this species, which has become of relevance to the characterization and selection of valuable A. fraterculus sp. 1 strains for mass rearing production and SIT implementation.