Background: Wolbachia,one of the most abundant taxa of intracellular Alphaproteobacteria, is widespread among arthropods and filarial nematodes. The presence of these maternally inherited bacteria is associated with modifications of host fitness, including a variety of reproductive abnormalities, such as cytoplasmic incompatibility, thelytokous parthenogenesis, host feminization and male-killing.Wolbachiahas attracted much interest for its role in biological, ecological and evolutionary processes as well as for its potential use in novel and environmentally-friendly strategies for the control of insect pests and disease vectors including a major agricultural pest, the South American fruit fly,Anastrepha fraterculusWiedemann (Diptera: Tephritidae).Results: We usedwsp,16S rRNAand a multilocus sequence typing (MLST) scheme includinggatB,coxA,hcpA,fbpA, andftsZ genes to detect and characterize theWolbachiainfection in laboratory strains and wild populations ofA. fraterculusfrom Argentina. Wolbachiawas found in allA. fraterculusindividuals studied.Nucleotide sequences analysis ofwspgene allowed the identification of twoWolbachianucleotide variants (namedwAfraCast1_A andwAfraCast2_A). After the analysis of 76 individuals, a high prevalence of thewAfraCast2_A variant was found both, in laboratory (82%) and wild populations (95%). MLST analysis identified bothWolbachiagenetic variants as sequence type 13. Phylogenetic analysis of concatenated MLST datasets clusteredwAfraCast1/2_A in the supergroup A. Paired-crossing experiments among single infected laboratory strains showed a phenotype specifically associated towAfraCast1_A that includes slight detrimental effects on larval survival, a female-biased sex ratio; suggesting the induction of male-killing phenomena, and a decreased proportion of females producing descendants that appears attributable to the lack of sperm in their spermathecae.Conclusions: We detected and characterized at the molecular level twowspgene sequence variants ofWolbachiaboth in laboratory and wild populations ofA. fraterculus sp.1 from Argentina. Crossing experiments on singly-infectedA. fraterculusstrains showed evidence of a male killing-like mechanism potentially associated to thewAfraCast1_A -A. fraterculusinteractions. Further mating experiments including antibiotic treatments and the analysis of early and late immature stages of descendants will contribute to our understanding of the phenotypes elicited by theWolbachiavariantwAfraCast1_A inA. fraterculus sp.1.

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. 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. 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.

Background: Wolbachia, one of the most abundant taxa of intracellular Alphaproteobacteria, is widespread among arthropods and filarial nematodes. The presence of these maternally inherited bacteria is associated with modifications of host fitness, including a variety of reproductive abnormalities, such as cytoplasmic incompatibility, thelytokous parthenogenesis, host feminization and male-killing. Wolbachia has attracted much interest for its role in biological, ecological and evolutionary processes as well as for its potential use in novel and environmentally-friendly strategies for the control of insect pests and disease vectors including a major agricultural pest, the South American fruit fly, Anastrepha fraterculus Wiedemann (Diptera: Tephritidae). Results: We used wsp , 16S rRNA and a multilocus sequence typing (MLST) scheme including gat B, cox A, hcp A, fbp A, and fts Z genes to detect and characterize the Wolbachia infection in laboratory strains and wild populations of A. fraterculus from Argentina . Wolbachia was found in all A. fraterculus individuals studied. Nucleotide sequences analysis of wsp gene allowed the identification of two Wolbachia nucleotide variants (named w AfraCast1_A and w AfraCast2_A). After the analysis of 76 individuals, a high prevalence of the w AfraCast2_A variant was found both, in laboratory (82%) and wild populations (95%). MLST analysis identified both Wolbachia genetic variants as sequence type 13. Phylogenetic analysis of concatenated MLST datasets clustered w AfraCast1/2_A in the supergroup A. Paired-crossing experiments among single infected laboratory strains showed a phenotype specifically associated to w AfraCast1_A that includes slight detrimental effects on larval survival, a female-biased sex ratio; suggesting the induction of male-killing phenomena, and a decreased proportion of females producing descendants that appears attributable to the lack of sperm in their spermathecae. Conclusions: We detected and characterized at the molecular level two wsp gene sequence variants of Wolbachia both in laboratory and wild populations of A. fraterculus sp. 1 from Argentina. Crossing experiments on singly-infected A. fraterculus strains showed evidence of a male killing-like mechanism potentially associated to the w AfraCast1_A - A. fraterculus interactions. Further mating experiments including antibiotic treatments and the analysis of early and late immature stages of descendants will contribute to our understanding of the phenotypes elicited by the Wolbachia variant w AfraCast1_A in A. fraterculus sp. 1.

A whole-genome assembly and annotation of Wolbachia sp. infecting Anastrepha fraterculus sp. 1 were generated by a metagenomic analysis of sequencing reads from a host genome project. This study contributes to the characterization of this endosymbiotic bacterium and provides valuable insights for research on host-symbiont interactions and pest management strategies.