Diversity and evolution of the Wolbachia endosymbionts of Bemisia (Hemiptera: Aleyrodidae) whiteflies

Wolbachia is the most prevalent symbiont described in arthropods to date. Wolbachia can manipulate host reproduction, provide nutrition to insect hosts and protect insect hosts from pathogenic viruses. So far, 13 supergroups of Wolbachia have been identified. The whitefly Bemisia tabaci is a complex containing more than 28 morphologically indistinguishable cryptic species. Some cryptic species of this complex are invasive. In this study, we report a comprehensive survey of Wolbachia in B. tabaci and its relative B. afer from 1658 insects representing 54 populations across 13 provinces of China and one state of Australia. Based on the results of PCR or sequencing of the 16S rRNA gene, the overall rates of Wolbachia infection were 79.6% and 0.96% in the indigenous and invasive Bemisia whiteflies, respectively. We detected a new Wolbachia supergroup by sequencing five molecular marker genes including 16S rRNA, groEL, gltA, hcpA, and fbpA genes. Data showed that many protein‐coding genes have limitations in detecting and classifying newly identified Wolbachia supergroups and thus raise a challenge to the known Wolbachia MLST standard analysis system. Besides, the other Wolbachia strains detected from whiteflies were clustered into supergroup B. Phylogenetic trees of whitefly mitochondrial cytochrome oxidase subunit I and Wolbachia multiple sequencing typing genes were not congruent. In addition, Wolbachia was also detected outside the special bacteriocytes in two cryptic species by fluorescence in situ hybridization, indicating the horizontal transmission of Wolbachia. Our results indicate that members of Wolbachia are far from well explored.     

Alternative evolutionary outcomes following endosymbiont‐mediated selection on male mating preference alleles

In many arthropods, intracellular bacteria, such as those of the genus Wolbachia, may spread through host populations as a result of cytoplasmic incompatibility (CI). Here, there is sterility or reduced fertility in crosses between infected males and uninfected females. As the bacterium is maternally inherited, the reduced fertility of uninfected females increases the frequency of the infection. If the transmission fidelity of the bacterium is less than 100%, the bacterium cannot invade from a low frequency, but if its frequency exceeds a threshold, it increases to a high, stable, equilibrium frequency. We explore the expected evolutionary dynamics of mutant alleles that cause their male bearers to avoid mating with uninfected females. For alleles which create this avoidance behaviour conditional upon the male being infected, there is a wide zone of parameter space that allows the preference allele to drive Wolbachia from the population when it would otherwise stably persist. There is also a wide zone of parameter space that allows a joint stable equilibrium for the Wolbachia and a polymorphism for the preference allele. When the male's avoidance of uninfected females is unconditional, the preference allele's effect on Wolbachia frequency is reduced, but there is a narrow range of values for the transmission rate and CI fertility that allow an unconditional preference allele to drive Wolbachia from the population, in a process driven by positive linkage disequilibrium between Wolbachia and the preference allele. The possibility of the evolution of preference could hamper attempts to manipulate wild populations through Wolbachia introductions.     

Melanic mutation causes a fitness decline in bean beetles infected by Wolbachia

Wolbachia cannot live outside a host, which is thought to be the reason for host‐Wolbachia coevolution toward benign parasitism, especially because the fitness of Wolbachia is traded against its host's fitness. Insect melanism has been reported to have a positive effect on pathogen resistance, but melanic mutants of Callosobruchus analis (Fabricius) and Callosobruchus chinensis (L.) (Coleoptera: Chrysomelidae) are infected with Wolbachia. Callosobruchus chinensis is infected with CI‐inducing Wolbachia, and melanic mutants exhibit fitness decline. Interestingly, this decline is not observed in C. analis melanic mutants that are infected with CI‐free Wolbachia. Our research question is whether the infection of CI‐inducing Wolbachia causes fitness decline of melanic hosts in C. analis. We examined fecundity, fertility, and longevity of C. analis melanic mutants and compared them between uninfected and infected hosts with CI‐inducing Wolbachia. Infected melanic mutants of C. analis exhibited fitness decline leading to reduced hatch rates even when parental combinations were compatible. Wolbachia can invade a host population by causing CI to decrease the fraction of uninfected hosts, but melanic mutant hosts decrease the number of infected hosts through fitness decline. Nevertheless, the melanism in hosts is not able to stop Wolbachia invasion in C. analis.     

Wolbachia have made it twice: Hybrid introgression between two sister species of Eurema butterflies

Wolbachia, cytoplasmically inherited endosymbionts of arthropods, are known to hijack their host reproduction in various ways to increase their own vertical transmission. This may lead to the selective sweep of associated mitochondria, which can have a large impact on the evolution of mitochondrial lineages. In Japan, two different Wolbacahia strains (wCI and wFem) are found in two sister species of pierid butterflies, Eurema mandarina and Eurema hecabe. In both species, females infected with wCI (C females) produce offspring with a nearly 1:1 sex ratio, while females infected with both wCI and wFem (CF females) produce all‐female offspring. Previous studies have suggested the historical occurrence of hybrid introgression in C individuals between the two species. Furthermore, hybrid introgression in CF individuals is suggested by the distinct mitochondrial lineages between C females and CF females of E. mandarina. In this study, we performed phylogenetic analyses based on nuclear DNA and mitochondrial DNA markers of E. hecabe with previously published data on E. mandarina. We found that the nuclear DNA of this species significantly diverged from that of E. mandarina. By contrast, mitochondrial DNA haplotypes comprised two clades, mostly reflecting Wolbachia infection status rather than the individual species. Collectively, our results support the previously suggested occurrence of two independent historical events wherein the cytoplasms of CF females and C females moved between E. hecabe and E. mandarina through hybrid introgression.     

Combined effects of elevated CO2 concentration and Wolbachia on Hylyphantes graminicola (Araneae: Linyphiidae)

The increasing concentration of carbon dioxide in atmosphere is not only a major cause of global warming, but it also adversely affects the ecological diversity of invertebrates. This study was conducted to evaluate the effect of elevated CO₂ concentration (ambient, 400 ppm and high, 800 ppm) and Wolbachia (Wolbachia‐infected, W⁺ and Wolbachia‐uninfected, W^?) on Hylyphantes graminicola. The total survival rate, developmental duration, carapace width and length, body weight, sex ratio, net reproductive rate, nutrition content, and enzyme activity in H. graminicola were examined under four treatments: W^? 400 ppm, W^? 800 ppm, W⁺ 400 ppm, and W⁺ 800 ppm. Results showed that Wolbachia‐infected spiders had significantly decreased the total developmental duration. Different instars showed variations up to some extent, but no obvious effect was found under elevated CO₂ concentration. Total survival rate, sex ratio, and net reproductive rate were not affected by elevated CO₂ concentration or Wolbachia infection. The carapace width of Wolbachia‐uninfected spiders decreased significantly under elevated CO₂ concentration, while the width, length and weight were not significantly affected in Wolbachia‐infected spiders reared at ambient CO₂ concentration. The levels of protein, specific activities of peroxidase, and amylase were significantly increased under elevated CO₂ concentration or Wolbachia‐infected spiders, while the total amino content was only increased in Wolbachia‐infected spiders. Thus, our current finding suggested that elevated CO₂ concentration and Wolbachia enhance nutrient contents and enzyme activity of H. graminicola and decrease development duration hence explore the interactive effects of factors which were responsible for reproduction regulation, but it also gives a theoretical direction for spider's protection in such a dynamic environment. Increased activities of enzymes and nutrients caused by Wolbachia infection aids for better survival of H. graminicola under stress.     

Experimental evolution reveals habitat‐specific fitness dynamics among Wolbachia clades in Drosophila melanogaster

The diversity and infection dynamics of the endosymbiont Wolbachia can be influenced by many factors, such as transmission rate, cytoplasmic incompatibility, environment, selection and genetic drift. The interplay of these factors in natural populations can result in heterogeneous infection patterns with substantial differences between populations and strains. The causes of these heterogeneities are not yet understood, partly due to the complexity of natural environments. We present experimental evolution as a new approach to study Wolbachia infection dynamics in replicate populations exposed to a controlled environment. A natural Drosophila melanogaster population infected with strains of Wolbachia belonging to different clades evolved in two laboratory environments (hot and cold) for 1.5 years. In both treatments, the rate of Wolbachia infection increased until fixation. In the hot environment, the relative frequency of different Wolbachia clades remained stable over 37 generations. In the cold environment, however, we observed marked changes in the composition of the Wolbachia population: within 15 generations, one Wolbachia clade increased more than 50% in frequency, whereas the other two clades decreased in frequency, resulting in the loss of one clade. The frequency change was highly reproducible not only among replicates, but also when flies that evolved for 42 generations in the hot environment were transferred to the cold environment. These results document how environmental factors can affect the composition of Wolbachia in D. melanogaster. The high reproducibility of the pattern suggests that experimental evolution studies can efficiently determine the functional basis of habitat‐specific fitness among Wolbachia strains.     

Multiple paternity in a wild population of Armadillidium vulgare: influence of infection with Wolbachia?

Female multiple mating has been extensively studied to understand how nonobvious benefits, generally thought to be of genetic nature, could overcome heavy costs such as an increased risk of infection during mating. However, the impact of infection itself on multiple mating has rarely been addressed. The interaction between the bacterium Wolbachia and its terrestrial crustacean host, Armadillidium vulgare, is a relevant model to investigate this question. In this association, Wolbachia is able to turn genetic males into functional females (i.e. feminization), thereby distorting the sex ratio and decreasing the number of available males at the population scale. Moreover, in A. vulgare, females have been shown to mate multiply under laboratory conditions and males prefer uninfected females over infected ones. Additionally, different Wolbachia strains are known to infect A. vulgare and these strains differ in their transmission rate and virulence. All these elements suggest a potential impact of different Wolbachia strains on multiple mating. To investigate this assumption, we collected gravid females in a wild A. vulgare population harbouring both uninfected females and females infected with one of two different Wolbachia strains (wVulM and wVulC) and performed paternity analyses on the obtained broods using microsatellite markers. We demonstrate that (i) multiple paternity is common in this wild population of A. vulgare, with a mean number of fathers of 4.48 ± 1.24 per brood and (ii) females infected with wVulC produced broods with a lower multiple paternity level compared with females infected with wVulM and uninfected ones. This work improves our knowledge of the impact of infections on reproductive strategies.     

Variation in Wolbachia cidB gene,but not cidA,is associated with cytoplasmic incompatibility mod phenotype diversity in Culex pipiens

Endosymbiotic Wolbachia bacteria are, to date, considered the most widespread symbionts in arthropods and are the cornerstone of major biological control strategies. Such a high prevalence is based on the ability of Wolbachia to manipulate their hosts' reproduction. One manipulation called cytoplasmic incompatibility (CI) is based on the death of the embryos generated by crosses between infected males and uninfected females or between individuals infected with incompatible Wolbachia strains. CI can be seen as a modification‐rescue system (or mod‐resc) in which paternal Wolbachia produce mod factors, inducing embryonic defects, unless the maternal Wolbachia produce compatible resc factors. Transgenic experiments in Drosophila melanogaster and Saccharomyces cerevisiae converged towards a model where the cidB Wolbachia gene is involved in the mod function while cidA is involved in the resc function. However, as cidA expression in Drosophila males was required to observe CI, it has been proposed that cidA could be involved in both resc and mod functions. A recent correlative study in natural Culex pipiens mosquito populations has revealed an association between specific cidA and cidB variations and changes in mod phenotype, also suggesting a role for both these genes in mod diversity. Here, by studying cidA and cidB genomic repertoires of individuals from newly sampled natural C. pipiens populations harbouring wPipIV strains from North Italy, we reinforce the link between cidB variation and mod phenotype variation fostering the involvement of cidB in the mod phenotype diversity. However, no association between any cidA variants or combination of cidA variants and mod phenotype variation was observed. Taken together our results in natural C. pipiens populations do not support the involvement of cidA in mod phenotype variation.     

Effects of host interaction with Wolbachia on cytoplasmic incompatibility in the two‐spotted spider mite Tetranychus urticae

The endosymbiotic bacterium Wolbachia pipientis infects a wide range of arthropods and induces a variety of reproductive anomalies, including cytoplasmic incompatibility (CI). Three populations of the two‐spotted spider mite, Tetranychus urticae, were investigated in the present study. Based on gene sequencing, they had different host genetic backgrounds but were infected with the same Wolbachia strain. We also examined the CI level relative to host background, male age, Wolbachia density, and Ankyrin (ANK) gene expression in T. urticae. The results of the present study suggest that: (1) CI differences between populations appear to be a result of host genetic background; (2) male age is not a factor determining intensity of CI; (3) Wolbachia density in males may serve as threshold factor necessary for the CI to occur in T. urticae, after which other factors become important in determining the strength of CI; and (4) hosts may modulate CI intensity through modulation of ANK gene expression in males. Our results describe a new type of interaction between Wolbachia and its hosts, and the effect of the interactions on CI. Further investigations on the functions of Wolbachia ankyrin gene products and their host targets, particularly with respect to host reproductive manipulation, are also imperative.     

Hybridization in natural sympatric populations of Dermacentor ticks in northwestern North America

Hybridization in ticks has been described in a handful of species and mostly as a result of laboratory experiments. We used 148 AFLP loci to describe putative hybridization events between D. andersoni and D. variabilis in sympatric populations from northwestern North America. Recently, D. variabilis has expanded its range westward into the natural range of D. andersoni. Using a sample of 235 D. andersoni and 62 D. variabilis, we identified 31 individuals as putative hybrids: four F₂ individuals and 27 backcrosses to D. andersoni (as defined by NewHybrids ). We found no evidence of hybrids backcrossing into D. variabilis. Furthermore, all hybrids presented 16S mtDNA signatures characteristic of D. andersoni, which indicates the directionality of the hybrid crosses: female D. andersoni × male D. variabilis. We also discovered 13 species‐specific AFLP fragments for D. andersoni. These loci were found to have a decreased occurrence in the putative hybrids and were absent altogether in D. variabilis samples. AFLP profiles were also used to determine the levels of genetic population structure and gene flow among nine populations of D. andersoni and three of D. variabilis. Genetic structure exists in both species (D. andersoni, Φ_ST = 0.110; D. variabilis, Φ_ST = 0.304) as well as significant estimates of isolation by distance (D. andersoni, ρ = 0.066, P = 0.001; D. variabilis, ρ = 0.729, P = 0.001).     

Symbiont‐conferred reproduction and fitness benefits can favour their host occurrence

Double infections of Wolbachia and Spiroplasma are frequent in natural populations of Tetranychus truncatus, a polyphagous mite species that has been a dominant species in China since 2009. However, little is known about the causes and ecological importance of such coexistences. In this study, we established T. truncatus strains with different infection types and then inferred the impact of the two endosymbionts on host reproduction and fitness. Double infection induced cytoplasmic incompatibility, which was demonstrated by reduction in egg hatchability of incompatible crosses. However, doubly infected females produced more eggs relative to other strains. Wolbachia and Spiroplasma did not affect host survival, whereas doubly infected females and males developed faster than other strains. Such reproduction and fitness benefits provided by double infections may be associated with the lower densities of each symbiont, and the quantitative results also confirmed competition between Wolbachia and Spiroplasma in doubly infected females. These symbiont‐conferred beneficial effects maintain stable prevalence of the symbionts and also help drive T. truncatus outbreaks in combination with other environmental factors.     

Wolbachia infection alters the relative abundance of resident bacteria in adult Aedes aegypti mosquitoes,but not larvae

Insect–symbiont interactions are known to play key roles in host functions and fitness. The common insect endosymbiont Wolbachia can reduce the ability of several human pathogens, including arboviruses and the malaria parasite, to replicate in insect hosts. Wolbachia does not naturally infect Aedes aegypti, the primary vector of dengue virus, but transinfected Ae. aegypti have antidengue virus properties and are currently being trialled as a dengue biocontrol strategy. Here, we assess the impact of Wolbachia infection of Ae. aegypti on the microbiome of wild mosquito populations (adults and larvae) collected from release sites in Cairns, Australia, by profiling the 16S rRNA gene using next‐generation sequencing. Our data indicate that Wolbachia reduces the relative abundance of a large proportion of bacterial taxa in Ae. aegypti adults, that is in accordance with the known pathogen‐blocking effects of Wolbachia on a variety of bacteria and viruses. In adults, several of the most abundant bacterial genera were found to undergo significant shifts in relative abundance. However, the genera showing the greatest changes in relative abundance in Wolbachia‐infected adults represented a low proportion of the total microbiome. In addition, there was little effect of Wolbachia infection on the relative abundance of bacterial taxa in larvae, or on species diversity (accounting for species richness and evenness together) detected in adults or larvae. These results offer insight into the effects of Wolbachia on the Ae. aegypti microbiome in a native setting, an important consideration for field releases of Wolbachia into the population.     

Segregation of male‐sterility alleles across a species boundary

Hybrid zones may serve as bridges permitting gene flow between species, including alleles influencing the evolution of breeding systems. Using greenhouse crosses, we assessed the likelihood that a hybrid zone could serve as a conduit for transfer of nuclear male‐sterility alleles between a gynodioecious species and a hermaphroditic species with very rare females in some populations. Segregation patterns in progeny of crosses between rare females of hermaphroditic Schiedea menziesii and hermaphroditic plants of gynodioecious Schiedea salicaria heterozygous at the male‐sterility locus, and between female S. salicaria and hermaphroditic plants from the hybrid zone, were used to determine whether male‐sterility was controlled at the same locus in the parental species and the hybrid zone. Segregations of females and hermaphrodites in approximately equal ratios from many of the crosses indicate that the same nuclear male‐sterility allele occurs in the parent species and the hybrid zone. These rare male‐sterility alleles in S. menziesii may result from gene flow from S. salicaria through the hybrid zone, presumably facilitated by wind pollination in S. salicaria. Alternatively, rare male‐sterility alleles might result from a reversal from gynodioecy to hermaphroditism in S. menziesii, or possibly de novo evolution of male sterility. Phylogenetic analysis indicates that some species of Schiedea have probably evolved separate sexes independently, but not in the lineage containing S. salicaria and S. menziesii. High levels of selfing and expression of strong inbreeding depression in S. menziesii, which together should favour females in populations, argue against a reversal from gynodioecy to hermaphroditism in S. menziesii.     

Pleistocene climate change and phylogeographic structure of the Gymnocarpos przewalskii (Caryophyllaceae) in the northwest China: Evidence from plastid DNA,ITS sequences,and Microsatellite

Northwestern China has a wealth of endemic species, which has been hypothesized to be affected by the complex paleoclimatic and paleogeographic history during Quaternary. In this paper, we used Gymnocarpos przewalskii as a model to address the evolutionary history and current population genetic structure of species in northwestern China. We employed two chloroplast DNA fragments (rps16 and psbB‐psbI), one nuclear DNA fragment (ITS), and simple sequence repeat (SSRs) to investigate the spatial genetic pattern of G. przewalskii. High genetic diversity (cpDNA: h_S = 0.330, h_T = 0.866; ITS: h_S = 0.458, h_T = 0.872) was identified in almost all populations, and most of the population have private haplotypes. Moreover, multimodal mismatch distributions were observed and estimates of Tajima's D and Fu's FS tests did not identify significantly departures from neutrality, indicating that recent expansion of G. przewalskii was rejected. Thus, we inferred that G. przewalskii survived generally in northwestern China during the Pleistocene. All data together support the genotypes of G. przewalskii into three groups, consistent with their respective geographical distributions in the western regions—Tarim Basin, the central regions—Hami Basin and Hexi Corridor, and the eastern regions—Alxa Desert and Wulate Prairie. Divergence among most lineages of G. przewalskii occurred in the Pleistocene, and the range of potential distributions is associated with glacial cycles. We concluded that climate oscillation during Pleistocene significantly affected the distribution of the species.     

Comparative performance of contact plates,electrostatic wipes,swabs and a novel sampling device for the detection of Staphylococcus aureus on environmental surfaces

Aims

To evaluate the performance of four sampling methods [contact plates, electrostatic wipes (wipe), swabs and a novel roller sampler] for recovery of Staphylococcus aureus from a stainless steel surface.

Methods and Results

Stainless steel test plates were inoculated with Staph. aureus, dried for 24 h and sampled using each of the four methods. Samples were either incubated directly (roller, contact plate) or processed using elution and membrane filtration (swab, wipe). Performance was assessed by calculating the apparent sampling efficiency (ASE), analytical sensitivity (Sn) and percentage of replications with positive growth. The wipe demonstrated the best performance across all inoculating concentrations (ASE_48 h = 18%; Sn_48 h = 7 CFU per 100 cm²). The swab performed well when corrected for area actually sampled (ASE_48 h = 24%; Sn_48 h = 76 CFU per 100 cm²). Of the contact‐based methods, the newly developed roller sampler outperformed the contact plate (roller: ASE_48 h = 10%; Sn_48 h = 17 CFU per 100 cm²; contact plate: ASE_48 h = 0·04%; Sn_48 h = 1412 CFU per 100 cm²); both contact samplers performed better at higher inoculating concentrations (6E3 CFU per 100 cm² for the roller and 6E6 CFU per 100 cm² for the contact plate). Overall, the electrostatic wipe produced the highest number of replications resulting in positive growth (74%_24 h, 91%_48 h).

Conclusions

This study demonstrates that selection of the sampling method must be carefully considered, given that different methods have varying performance.

Significance and Impact of the Study

This is the first study assessing static wipes for sampling and one that uses a more real‐world‐relevant 24‐h drying time. The results help with infection control, and environmental health professionals choose better sampling methodologies.     

F supergroup <Emphasis Type="Italic">Wolbachia</Emphasis> in bush crickets: what do patterns of sequence variation reveal about this supergroup and horizontal transfer between nematodes and arthropods?

Wolbachia pipientis, an intracellular, α-proteobacterium, is commonly found in arthropods and filarial nematodes. Most infected insects are known to harbor strains of Wolbachia from supergroups A or B, whereas supergroups C and D occur only in filarial nematodes. Here, we present molecular evidence from two genes (ftsZ and 16S rDNA) that 2 Orthopterans (the bush cricket species Orocharis saltator and Hapithus agitator; Gryllidae: Eneopterinae) are infected with Wolbachia from the F supergroup. Additionally, a series of PCR tests revealed that these bush cricket specimens did not harbor nematodes, thus indicating that our positive results were not a by-product of nematodes being present in these cricket samples. Patterns of molecular variation suggest that (1) strains of F supergroup Wolbachia exhibit less genetic variation than the nematode-specific C and D supergroups but more than the A and B supergroups found in arthropods and (2) that there is no evidence of recombination within F supergroup strains. The above data support previous findings that F supergroup Wolbachia is not only harbored in both nematodes and arthropods, but that horizontal transfer has likely occurred recently between these diverse taxonomic groups (although the exact details of such horizontal transmissions remain unclear). Moreover, the limited genetic variation and lack of recombination in the F supergroup suggest that this clade of Wolbachia has radiated relatively rapidly with either (1) little time for recombination to occur or (2) selection against recombination as occurs in the mutualistic C and D strains of Wolbachia – both of which remain to be explored further.     

Maintenance of adaptive differentiation by Wolbachia induced bidirectional cytoplasmic incompatibility: the importance of sib-mating and genetic systems

Background  

Bacteria of the genus Wolbachia are reproductive parasites widespread among arthropods. The most common effect arising from the presence of Wolbachia in a population is Cytoplasmic Incompatibility (CI), whereby postmating reproductive isolation occurs in crosses between an infected male and an uninfected female, or when a male is infected with a different strain of Wolbachia to that of the female (bidirectional CI). Previous theoretical models have demonstrated that bidirectional CI can contribute to the genetic divergence of populations in haploid and diploid organisms. However, haplodiploid organisms were not considered in these models even though they include Nasonia parasitoid wasps – the best example of the implication of Wolbachia in ongoing speciation. Moreover, previous work did not investigate inbreeding mating systems, which are frequently observed in arthropod species.     

Wolbachia pipientis occurs in Aedes aegypti populations in New Mexico and Florida,USA

The mosquitoes Aedes aegypti (L.) and Ae. albopictus Skuse are the major vectors of dengue, Zika, yellow fever, and chikungunya viruses worldwide. Wolbachia, an endosymbiotic bacterium present in many insects, is being utilized in novel vector control strategies to manipulate mosquito life history and vector competence to curb virus transmission. Earlier studies have found that Wolbachia is commonly detected in Ae. albopictus but rarely detected in Ae. aegypti. In this study, we used a two‐step PCR assay to detect Wolbachia in wild‐collected samples of Ae. aegypti. The PCR products were sequenced to validate amplicons and identify Wolbachia strains. A loop‐mediated isothermal amplification (LAMP) assay was developed and used for detecting Wolbachia in selected mosquito specimens as well. We found Wolbachia in 85/148 (57.4%) wild Ae. aegypti specimens from various cities in New Mexico, and in 2/46 (4.3%) from St. Augustine, Florida. Wolbachia was not detected in 94 samples of Ae. aegypti from Deer Park, Harris County, Texas. Wolbachia detected in Ae. aegypti from both New Mexico and Florida was the wAlbB strain of Wolbachia pipientis. A Wolbachia‐positive colony of Ae. aegypti was established from pupae collected in Las Cruces, New Mexico, in 2018. The infected females of this strain transmitted Wolbachia to their progeny when crossed with males of Rockefeller strain of Ae. aegypti, which does not carry Wolbachia. In contrast, none of the progeny of Las Cruces males mated to Rockefeller females were infected with Wolbachia.     

High prevalence of Wolbachia infection does not explain unidirectional cytoplasmic incompatibility of Altica flea beetles

1. Wolbachia are intracellular bacteria found in a wide range of arthropods that can impact reproductive isolation of their hosts. Previous studies showed unidirectional cytoplasmic incompatibility (CI) and high infection rates by Wolbachia in the closely related leaf beetles Altica fragariae and Altica viridicyanea; however, whether this reproductive isolation was induced by Wolbachia remains unclear. 2. This study estimated the prevalence of Wolbachia in Altica beetles, assessed genetic diversity of Wolbachia strains infecting these beetles, and tested whether Wolbachia-induced CI explains reproductive isolation of A. fragariae and A. viridicyanea. 3. The results show that all of the 11 tested Altica species were infected by Wolbachia, and the infection rate was as high as 97.0% (n = 235). Multi-strain infections were common, being found in 10 of the 11 species tested and accounting for 23.0% of all screened specimens. In total, 35 Wolbachia strains were identified based on 208 wsp sequences obtained. Although a majority of A. fragariae and A. viridicyanea individuals from the Beijing population were infected with only one strain each, multi-strain infections did occur in both species. Antibiotic curing experiments did not change hatching success in either inter- or intraspecific crosses of A. fragariae and A. viridicyanea, indicating that these Wolbachia strains do not induce CI. These results were further corroborated by the lack of the Wolbachia cifB gene, which is responsible for causing CI. 4. These findings suggest that high prevalence of Wolbachia infection is unlikely to contribute to reproductive isolation and speciation in this system.     

Evidence for a recent horizontal transmission and spatial spread of Wolbachia from endemic Rhagoletis cerasi (Diptera: Tephritidae) to invasive Rhagoletis cingulata in Europe

The widespread occurrence of Wolbachia in arthropods and nematodes suggests that this intracellular, maternally inherited endosymbiont has the ability to cross species boundaries. However, direct evidence for such a horizontal transmission of Wolbachia in nature is scarce. Here, we compare the well‐characterized Wolbachia infection of the European cherry fruit fly, Rhagoletis cerasi, with that of the North American eastern cherry fruit fly, Rhagoletis cingulata, recently introduced to Europe. Molecular genetic analysis of Wolbachia based on multilocus sequence typing and the Wolbachia surface protein wsp showed that all R. cingulata individuals are infected with wCin2 identical to wCer2 in R. cerasi. In contrast, wCin1, a strain identical to wCer1 in R. cerasi, was present in several European populations of R. cingulata, but not in any individual from the United States. Surveys of R. cingulata from Germany and Hungary indicated that in some populations, the frequency of wCin1 increased significantly in just a few years with at least two independent horizontal transmission events. This is corroborated by the analysis of the mitochondrial cytochrome oxidase II gene that showed association of wCin1 with two distinct haplotypes in Germany, one of which is also infected with wCin1 in Hungary. In summary, our study provides strong evidence for a very recent inter‐specific Wolbachia transmission with a subsequent spatial spread in field populations.