Wolbachia infection negatively impacts Drosophila simulans heat tolerance in a strain- and trait-specific manner.

The susceptibility of insects to rising temperatures has largely been measured by their ability to survive thermal extremes. However, the capacity for maternally inherited endosymbionts to influence insect heat tolerance has been overlooked. Further, while some studies have addressed the impact of heat on traits like fertility, which can decline at temperatures below lethal thermal limits, none have considered the impact of endosymbionts. Here, we assess the impact of three Wolbachia strains (wRi, wAu and wNo) on the survival and fertility of Drosophila simulans exposed to heat stress during development or as adults. The effect of Wolbachia infection on heat tolerance was generally small and trait/strain specific. Only the wNo infection significantly reduced the survival of adult males after a heat shock. When exposed to fluctuating heat stress during development, the wRi and wAu strains reduced egg-to-adult survival but only the wNo infection reduced male fertility. Wolbachia densities of all three strains decreased under developmental heat stress, but reductions occurred at temperatures above those that reduced host fertility. These findings emphasize the necessity to account for endosymbionts and their effect on both survival and fertility when investigating insect responses to heat stress.

How often are male mosquitoes attracted to humans?

Many mosquito species live close to humans where females feed on human blood. While male mosquitoes do not feed on blood, it has long been recognized that males of some species can be attracted to human hosts. To investigate the frequency of male mosquito attraction to humans, we conducted a literature review and human-baited field trials, as well as laboratory experiments involving males and females of three common Aedes species. Our literature review indicated that male attraction to humans is limited to a small number of species, including Ae. aegypti and Ae. albopictus. In our human-baited field collections, only 4 out of 13 species captured included males. In laboratory experiments, we found that male Ae. notoscriptus and Ae. vigilax showed no attraction to humans, while male Ae. aegypti exhibited persistent attraction for up to 30 min. Both male and female Ae. aegypti displayed similar preferences for different human subjects, suggesting that male Ae. aegypti respond to similar cues as females. Additionally, we found that mosquito repellents applied to human skin effectively repelled male mosquitoes. These findings shed light on mosquito behaviour and have implications for mosquito control programmes, particularly those involving the release or monitoring of the male mosquito population.

A diversity of endosymbionts across Australian aphids and their persistence in aphid cultures.

There is increasing interest in the use of endosymbionts in pest control, which will benefit from the identification of endosymbionts from potential donor species for transfer to pest species. Here, we screened for endosymbionts in 123 Australian aphid samples across 32 species using 16S DNA metabarcoding. We then developed a qPCR method to validate the metabarcoding data set and to monitor endosymbiont persistence in aphid cultures. Pea aphids (Acyrthosiphon pisum) were frequently coinfected with Rickettsiella and Serratia, and glasshouse potato aphids (Aulacorthum solani) were coinfected with Regiella and Spiroplasma; other secondary endosymbionts detected in samples occurred by themselves. Hamiltonella, Rickettsia and Wolbachia were restricted to a single aphid species, whereas Regiella was found in multiple species. Rickettsiella, Hamiltonella and Serratia were stably maintained in laboratory cultures, although others were lost rapidly. The overall incidence of secondary endosymbionts in Australian samples tended to be lower than recorded from aphids overseas. These results indicate that aphid endosymbionts probably exhibit different levels of infectivity and vertical transmission efficiency across hosts, which may contribute to natural infection patterns. The rapid loss of some endosymbionts in cultures raises questions about factors that maintain them under field conditions, while endosymbionts that persisted in laboratory culture provide candidates for interspecific transfers.

A rapidly spreading deleterious aphid endosymbiont that uses horizontal as well as vertical transmission.

Endosymbiotic bacteria that live inside the cells of insects are typically only transmitted maternally and can spread by increasing host fitness and/or modifying reproduction in sexual hosts. Transinfections of Wolbachia endosymbionts are now being used to introduce useful phenotypes into sexual host populations, but there has been limited progress on applications using other endosymbionts and in asexual populations. Here, we develop a unique pathway to application in aphids by transferring the endosymbiont Rickettsiella viridis to the major crop pest Myzus persicae. Rickettsiella infection greatly reduced aphid fecundity, decreased heat tolerance, and modified aphid body color, from light to dark green. Despite inducing host fitness costs, Rickettsiella spread rapidly through caged aphid populations via plant-mediated horizontal transmission. The phenotypic effects of Rickettsiella were sensitive to temperature, with spread only occurring at 19 °C and not 25 °C. Body color modification was also lost at high temperatures despite Rickettsiella maintaining a high density. Rickettsiella shows the potential to spread through natural M. persicae populations by horizontal transmission and subsequent vertical transmission. Establishment of Rickettsiella in natural populations could reduce crop damage by modifying population age structure, reducing population growth and providing context-dependent effects on host fitness. Our results highlight the importance of plant-mediated horizontal transmission and interactions with temperature as drivers of endosymbiont spread in asexual insect populations.

Differences in gene expression in field populations of Wolbachia-infected Aedes aegypti mosquitoes with varying release histories in northern Australia.

Aedes aegypti is the principal mosquito vector of dengue, yellow fever, Zika and chikungunya viruses. The wMel strain of the endosymbiotic bacteria Wolbachia pipientis was introduced into the vector as a novel biocontrol strategy to stop transmission of these viruses. Mosquitoes with Wolbachia have been released in the field in Northern Queensland, Australia since 2011, at various locations and over several years, with populations remaining stably infected. Wolbachia infection is known to alter gene expression in its mosquito host, but whether (and how) this changes over the long-term in the context of field releases remains unknown. We sampled mosquitoes from Wolbachia-infected populations with three different release histories along a time gradient and performed RNA-seq to investigate gene expression changes in the insect host. We observed a significant impact on gene expression in Wolbachia-infected mosquitoes versus uninfected controls. Fewer genes had significantly upregulated expression in mosquitoes from the older releases (512 and 486 from the 2011 and 2013/14 release years, respectively) versus the more recent releases (1154 from the 2017 release year). Nonetheless, a fundamental signature of Wolbachia infection on host gene expression was observed across all releases, comprising upregulation of immunity (e.g. leucine-rich repeats, CLIPs) and metabolism (e.g. lipid metabolism, iron transport) genes. There was limited downregulation of gene expression in mosquitoes from the older releases (84 and 71 genes from the 2011 and 2013/14 release years, respectively), but significantly more in the most recent release (509 from the 2017 release year). Our findings indicate that at > 8 years post-introgression into field populations, Wolbachia continues to profoundly impact expression of host genes, such as those involved in insect immune response and metabolism. If Wolbachia-mediated virus blocking is underpinned by these differential gene expression changes, our results suggest it may remain stable long-term.

A male-killing Wolbachia endosymbiont is concealed by another endosymbiont and a nuclear suppressor.

Bacteria that live inside the cells of insect hosts (endosymbionts) can alter the reproduction of their hosts, including the killing of male offspring (male killing, MK). MK has only been described in a few insects, but this may reflect challenges in detecting MK rather than its rarity. Here, we identify MK Wolbachia at a low frequency (around 4%) in natural populations of Drosophila pseudotakahashii. MK Wolbachia had a stable density and maternal transmission during laboratory culture, but the MK phenotype which manifested mainly at the larval stage was lost rapidly. MK Wolbachia occurred alongside a second Wolbachia strain expressing a different reproductive manipulation, cytoplasmic incompatibility (CI). A genomic analysis highlighted Wolbachia regions diverged between the 2 strains involving 17 genes, and homologs of the wmk and cif genes implicated in MK and CI were identified in the Wolbachia assembly. Doubly infected males induced CI with uninfected females but not females singly infected with CI-causing Wolbachia. A rapidly spreading dominant nuclear suppressor genetic element affecting MK was identified through backcrossing and subsequent analysis with ddRAD SNPs of the D. pseudotakahashii genome. These findings highlight the complexity of nuclear and microbial components affecting MK endosymbiont detection and dynamics in populations and the challenges of making connections between endosymbionts and the host phenotypes affected by them.

Developing Wolbachia-based disease interventions for an extreme environment.

Aedes aegypti mosquitoes carrying self-spreading, virus-blocking Wolbachia bacteria are being deployed to suppress dengue transmission. However, there are challenges in applying this technology in extreme environments. We introduced two Wolbachia strains into Ae. aegypti from Saudi Arabia for a release program in the hot coastal city of Jeddah. Wolbachia reduced infection and dissemination of dengue virus (DENV2) in Saudi Arabian mosquitoes and showed complete maternal transmission and cytoplasmic incompatibility. Wolbachia reduced egg hatch under a range of environmental conditions, with the Wolbachia strains showing differential thermal stability. Wolbachia effects were similar across mosquito genetic backgrounds but we found evidence of local adaptation, with Saudi Arabian mosquitoes having lower egg viability but higher adult desiccation tolerance than Australian mosquitoes. Genetic background effects will influence Wolbachia invasion dynamics, reinforcing the need to use local genotypes for mosquito release programs, particularly in extreme environments like Jeddah. Our comprehensive characterization of Wolbachia strains provides a foundation for Wolbachia-based disease interventions in harsh climates.

Barley Yellow Dwarf Virus Influences Its Vector's Endosymbionts but Not Its Thermotolerance.

The barley yellow dwarf virus (BYDV) of cereals is thought to substantially increase the high-temperature tolerance of its aphid vector, Rhopalosiphum padi, which may enhance its transmission efficiency. This is based on experiments with North American strains of BYDV and R. padi. Here, we independently test these by measuring the temperature tolerance, via Critical Thermal Maximum (CTmax) and knockdown time, of Australian R. padi infected with a local BYDV isolate. We further consider the interaction between BYDV transmission, the primary endosymbiont of R. padi (Buchnera aphidicola), and a transinfected secondary endosymbiont (Rickettsiella viridis) which reduces the thermotolerance of other aphid species. We failed to find an increase in tolerance to high temperatures in BYDV-infected aphids or an impact of Rickettsiella on thermotolerance. However, BYDV interacted with R. padi endosymbionts in unexpected ways, suppressing the density of Buchnera and Rickettsiella. BYDV density was also fourfold higher in Rickettsiella-infected aphids. Our findings indicate that BYDV does not necessarily increase the temperature tolerance of the aphid transmission vector to increase its transmission potential, at least for the genotype combinations tested here. The interactions between BYDV and Rickettsiella suggest new ways in which aphid endosymbionts may influence how BYDV spreads, which needs further testing in a field context.

Genomic and Phenotypic Comparisons Reveal Distinct Variants of Wolbachia Strain wAlbB.

The intracellular bacterium Wolbachia inhibits virus replication and is being harnessed around the world to fight mosquito-borne diseases through releases of mosquitoes carrying the symbiont. Wolbachia strains vary in their ability to invade mosquito populations and suppress viruses in part due to differences in their density within the insect and associated fitness costs. Using whole-genome sequencing, we demonstrate the existence of two variants in wAlbB, a Wolbachia strain being released in natural populations of Aedes aegypti mosquitoes. The two variants display striking differences in genome architecture and gene content. Differences in the presence/absence of 52 genes between variants include genes located in prophage regions and others potentially involved in controlling the symbiont's density. Importantly, we show that these genetic differences correlate with variation in wAlbB density and its tolerance to heat stress, suggesting that different wAlbB variants may be better suited for field deployment depending on local environmental conditions. Finally, we found that the wAlbB genome remained stable following its introduction in a Malaysian mosquito population. Our results highlight the need for further genomic and phenotypic characterization of Wolbachia strains in order to inform ongoing Wolbachia-based programs and improve the selection of optimal strains in future field interventions. IMPORTANCE Dengue is a viral disease transmitted by Aedes mosquitoes that threatens around half of the world population. Recent advances in dengue control involve the introduction of Wolbachia bacterial symbionts with antiviral properties into mosquito populations, which can lead to dramatic decreases in the incidence of the disease. In light of these promising results, there is a crucial need to better understand the factors affecting the success of such strategies, in particular the choice of Wolbachia strain for field releases and the potential for evolutionary changes. Here, we characterized two variants of a Wolbachia strain used for dengue control that differ at the genomic level and in their ability to replicate within the mosquito. We also found no evidence for the evolution of the symbiont within the 2 years following its deployment in Malaysia. Our results have implications for current and future Wolbachia-based health interventions.

Wolbachia inhibits ovarian formation and increases blood feeding rate in female Aedes aegypti.

Wolbachia, a gram-negative endosymbiotic bacterium widespread in arthropods, is well-known for changing the reproduction of its host in ways that increase its rate of spread, but there are also costs to hosts that can reduce this. Here we investigated a novel reproductive alteration of Wolbachia wAlbB on its host Aedes aegypti, using studies on mosquito life history traits, ovarian dissection, as well as gene expression assays. We found that an extended period of the larval stage as well as the egg stage (as previously shown) can increase the proportion of Wolbachia-infected females that become infertile; an effect which was not observed in uninfected females. Infertile females had incomplete ovarian formation and also showed a higher frequency of blood feeding following a prior blood meal, indicating that they do not enter a complete gonotrophic cycle. Treatments leading to infertility also decreased the expression of genes related to reproduction, especially the vitellogenin receptor gene whose product regulates the uptake of vitellogenin (Vg) into ovaries. Our results demonstrate effects associated with the development of infertility in wAlbB-infected Ae. aegypti females with implications for Wolbachia releases. The results also have implications for the evolution of Wolbachia infections in novel hosts.

Fitness costs of Wolbachia shift in locally-adapted Aedes aegypti mosquitoes.

Aedes aegypti mosquito eggs can remain quiescent for many months before hatching, allowing populations to persist through unfavourable conditions. A. aegypti infected with the Wolbachia strain wMel have been released in tropical and subtropical regions for dengue control. wMel reduces the viability of quiescent eggs, but this physiological cost might be expected to evolve in natural mosquito populations that frequently experience stressful conditions. We found that the cost of wMel infection differed consistently between mosquitoes collected from different locations and became weaker across laboratory generations, suggesting environment-specific adaptation of mosquitoes to the wMel infection. Reciprocal crossing experiments show that differences in the cost of wMel to quiescent egg viability were mainly due to mosquito genetic background and not Wolbachia origin. wMel-infected mosquitoes hatching from long-term quiescent eggs showed partial loss of cytoplasmic incompatibility and female infertility, highlighting additional costs of long-term quiescence. Our study provides the first evidence for a shift in Wolbachia phenotypic effects following deliberate field release and establishment and it highlights interactions between Wolbachia infections and mosquito genetic backgrounds. The unexpected changes in fitness costs observed here suggest potential tradeoffs with undescribed fitness benefits of the wMel infection.

Sex-specific distribution and classification of Wolbachia infections and mitochondrial DNA haplogroups in Aedes albopictus from the Indo-Pacific.

The arbovirus vector Aedes albopictus (Asian tiger mosquito) is common throughout the Indo-Pacific region, where most global dengue transmission occurs. We analysed population genomic data and tested for cryptic species in 160 Ae. albopictus sampled from 16 locations across this region. We found no evidence of cryptic Ae. albopictus but found multiple intraspecific COI haplotypes partitioned into groups representing three Asian lineages: East Asia, Southeast Asia and Indonesia. Papua New Guinea (PNG), Vanuatu and Christmas Island shared recent coancestry, and Indonesia and Timor-Leste were likely invaded from East Asia. We used a machine learning trained on morphologically sexed samples to classify sexes using multiple genetic features and then characterized the wAlbA and wAlbB Wolbachia infections in 664 other samples. The wAlbA and wAlbB infections as detected by qPCR showed markedly different patterns in the sexes. For females, most populations had a very high double infection incidence, with 67% being the lowest value (from Timor-Leste). For males, the incidence of double infections ranged from 100% (PNG) to 0% (Vanuatu). Only 6 females were infected solely by the wAlbA infection, while rare uninfected mosquitoes were found in both sexes. The wAlbA and wAlbB densities varied significantly among populations. For mosquitoes from Torres Strait and Vietnam, the wAlbB density was similar in single-infected and superinfected (wAlbA and wAlbB) mosquitoes. There was a positive association between wAlbA and wAlbB infection densities in superinfected Ae. albopictus. Our findings provide no evidence of cryptic species of Ae. albopictus in the region and suggest site-specific factors influencing the incidence of Wolbachia infections and their densities. We also demonstrate the usefulness of ddRAD tag depths as sex-specific mosquito markers. The results provide baseline data for the exploitation of Wolbachia-induced cytoplasmic incompatibility (CI) in dengue control.

A wMel Wolbachia variant in Aedes aegypti from field-collected Drosophila melanogaster with increased phenotypic stability under heat stress.

Mosquito-borne diseases remain a major cause of morbidity and mortality. Population replacement strategies involving the wMel strain of Wolbachia are being used widely to control mosquito-borne diseases. However, these strategies may be influenced by temperature because wMel is vulnerable to heat. wMel infections in Drosophila melanogaster are genetically diverse, but few transinfections of wMel variants have been generated in Aedes aegypti. Here, we successfully transferred a wMel variant (termed wMelM) originating from a field-collected D. melanogaster into Ae. aegypti. The new wMelM variant (clade I) is genetically distinct from the original wMel transinfection (clade III), and there are no genomic differences between wMelM in its original and transinfected host. We compared wMelM with wMel in its effects on host fitness, temperature tolerance, Wolbachia density, vector competence, cytoplasmic incompatibility and maternal transmission under heat stress in a controlled background. wMelM showed a higher heat tolerance than wMel, likely due to higher overall densities within the mosquito. Both wMel variants had minimal host fitness costs, complete cytoplasmic incompatibility and maternal transmission, and dengue virus blocking under laboratory conditions. Our results highlight phenotypic differences between Wolbachia variants and wMelM shows potential as an alternative strain in areas with strong seasonal temperature fluctuations.

A decade of stability for wMel Wolbachia in natural Aedes aegypti populations.

Mosquitoes carrying Wolbachia endosymbionts are being released in many countries for arbovirus control. The wMel strain of Wolbachia blocks Aedes-borne virus transmission and can spread throughout mosquito populations by inducing cytoplasmic incompatibility. Aedes aegypti mosquitoes carrying wMel were first released into the field in Cairns, Australia, over a decade ago, and with wider releases have resulted in the near elimination of local dengue transmission. The long-term stability of Wolbachia effects is critical for ongoing disease suppression, requiring tracking of phenotypic and genomic changes in Wolbachia infections following releases. We used a combination of field surveys, phenotypic assessments, and Wolbachia genome sequencing to show that wMel has remained stable in its effects for up to a decade in Australian Ae. aegypti populations. Phenotypic comparisons of wMel-infected and uninfected mosquitoes from near-field and long-term laboratory populations suggest limited changes in the effects of wMel on mosquito fitness. Treating mosquitoes with antibiotics used to cure the wMel infection had limited effects on fitness in the next generation, supporting the use of tetracycline for generating uninfected mosquitoes without off-target effects. wMel has a temporally stable within-host density and continues to induce complete cytoplasmic incompatibility. A comparison of wMel genomes from pre-release (2010) and nine years post-release (2020) populations show few genomic differences and little divergence between release locations, consistent with the lack of phenotypic changes. These results indicate that releases of Wolbachia-infected mosquitoes for population replacement are likely to be effective for many years, but ongoing monitoring remains important to track potential evolutionary changes.

Characterization of the first Wolbachia from the genus Scaptodrosophila, a male-killer from the rainforest species S. claytoni.

The Scaptodrosophila genus represents a large group of drosophilids with a worldwide distribution and a predominance of species in Australia, but there is little information on the presence and impacts of Wolbachia endosymbionts in this group. Here we describe the first Wolbachia infection from this group, wClay isolated from Scaptodrosophila claytoni (van Klinken), a species from the east coast of Australia. The infection is polymorphic in natural populations, occurring at a frequency of around 6%-10%. wClay causes male killing, producing female-biased lines; most lines showed 100% male killing, though in 1 line it was <80%. The lines need to be maintained through the introduction of males unless the infection is removed by tetracycline treatment. wClay is transmitted at a high fidelity (98.6%) through the maternal lineage and has been stable in 2 laboratory lines across 24 generations, suggesting it is likely to persist in populations. The infection has not been previously described but is closely related to the male-killing Wolbachia recently described from Drosophila pandora based on multilocus sequence typing and the wsp gene. Male-killing Wolbachia are likely to be common in drosophilids but remain difficult to detect because the infections can often be at a low frequency.

Long-Range But Not Short-Range Attraction of Male Aedes aegypti (Diptera: Culicidae) Mosquitoes to Humans.

Female Aedes aegypti (Linnaeus) mosquitoes integrate multiple sensory cues to locate human hosts for blood meals. Although male Ae. aegypti swarm around and land on humans in nature to mate, direct evidence of attraction to humans is limited. Male mosquito attraction to human host cues is often undetectable in confined laboratory assays, leading to a misconception that male mosquitoes are not attracted to humans. We used semifield experiments to demonstrate robust attraction of male Ae. aegypti to humans. Human-baited traps captured up to 25% of released males within 15 min, whereas control traps without humans as bait failed to capture males. Rapid attraction to humans was further demonstrated through videography. Males swarmed around and landed on human subjects, with no activity recorded in paired unbaited controls. Finally, we confirm the lack of discernible male attraction to humans in small laboratory cages. Our experiments demonstrate that both male and female Ae. aegypti show attraction to humans, but with clear sex-specific behavioral differences at short-range. Male mosquito attraction to humans is likely to be important for mating success in wild populations and its basis should be further explored. Our results highlight the importance of arena size and assay design for mosquito behavioral research. A better understanding of host cues that attract males could help us to improve mosquito surveillance and control.

Genetic stability of Aedes aegypti populations following invasion by wMel Wolbachia.

BACKGROUND: Wolbachia wMel is the most commonly used strain in rear and release strategies for Aedes aegypti mosquitoes that aim to inhibit the transmission of arboviruses such as dengue, Zika, Chikungunya and yellow fever. However, the long-term establishment of wMel in natural Ae. aegypti populations raises concerns that interactions between Wolbachia wMel and Ae. aegypti may lead to changes in the host genome, which could affect useful attributes of Wolbachia that allow it to invade and suppress disease transmission. RESULTS: We applied an evolve-and-resequence approach to study genome-wide genetic changes in Ae. aegypti from the Cairns region, Australia, where Wolbachia wMel was first introduced more than 10 years ago. Mosquito samples were collected at three different time points in Gordonvale, Australia, covering the phase before (2010) and after (2013 and 2018) Wolbachia releases. An additional three locations where Wolbachia replacement happened at different times across the last decade were also sampled in 2018. We found that the genomes of mosquito populations mostly remained stable after Wolbachia release, with population differences tending to reflect the geographic location of the populations rather than Wolbachia infection status. However, outlier analysis suggests that Wolbachia may have had an influence on some genes related to immune response, development, recognition and behavior. CONCLUSIONS: Ae. aegypti populations remained geographically distinct after Wolbachia wMel releases in North Australia despite their Wolbachia infection status. At some specific genomic loci, we found signs of selection associated with Wolbachia, suggesting potential evolutionary impacts can happen in the future and further monitoring is warranted.

Limonene Emissions: Do Different Types Have Different Biological Effects?

Limonene is one of the most abundant pollutants indoors, and it contributes to the formation of additional pollutants, such as formaldehyde and photochemical smog. Limonene is commonly used in fragranced consumer products, such as cleaning supplies and air fresheners, which have also been associated with health problems. Limonene can exist in different enantiomeric forms (R-limonene and S-limonene) and be derived from different sources. However, little is known about whether different forms and sources of limonene may have different effects. This research explored whether different types of limonene, at the same concentrations, could elicit different biological effects. To investigate this question, the study employed Aedes aegypti mosquitoes, which have sophisticated olfactory abilities, in olfactometer tests of repellency/attraction. The results indicate that a synthetic source of R-limonene is more repellent than a natural source of R-limonene. In addition, synthetic sources of both R-limonene and S-limonene are not significantly different in repellency. These findings can contribute to our understanding and further exploration of the effects of a common fragrance compound on air quality and health.

High Incidence of Related Wolbachia across Unrelated Leaf-Mining Diptera.

The maternally inherited endosymbiont, Wolbachia pipientis, plays an important role in the ecology and evolution of many of its hosts by affecting host reproduction and fitness. Here, we investigated 13 dipteran leaf-mining species to characterize Wolbachia infections and the potential for this endosymbiont in biocontrol. Wolbachia infections were present in 12 species, including 10 species where the Wolbachia infection was at or near fixation. A comparison of Wolbachia relatedness based on the wsp/MLST gene set showed that unrelated leaf-mining species often shared similar Wolbachia, suggesting common horizontal transfer. We established a colony of Liriomyza brassicae and found adult Wolbachia density was stable; although Wolbachia density differed between the sexes, with females having a 20-fold higher density than males. Wolbachia density increased during L. brassicae development, with higher densities in pupae than larvae. We removed Wolbachia using tetracycline and performed reciprocal crosses between Wolbachia-infected and uninfected individuals. Cured females crossed with infected males failed to produce offspring, indicating that Wolbachia induced complete cytoplasmic incompatibility in L. brassicae. The results highlight the potential of Wolbachia to suppress Liriomyza pests based on approaches such as the incompatible insect technique, where infected males are released into populations lacking Wolbachia or with a different incompatible infection.

A wAlbB Wolbachia Transinfection Displays Stable Phenotypic Effects across Divergent Aedes aegypti Mosquito Backgrounds.

Aedes mosquitoes harboring intracellular Wolbachia bacteria are being released in arbovirus and mosquito control programs. With releases taking place around the world, understanding the contribution of host variation to Wolbachia phenotype is crucial. We generated a Wolbachia transinfection (wAlbBQ) in Aedes aegypti and performed backcrossing to introduce the infection into Australian or Malaysian nuclear backgrounds. Whole Wolbachia genome sequencing shows that the wAlbBQ transinfection is nearly identical to the reference wAlbB genome, suggesting few changes since the infection was first introduced to A. aegypti over 15 years ago. However, these sequences were distinct from other available wAlbB genome sequences, highlighting the potential diversity of wAlbB in natural Aedes albopictus populations. Phenotypic comparisons demonstrate the effects of wAlbB infection on egg hatching and nuclear background on fecundity and body size but no interactions between wAlbB infection and nuclear background for any trait. The wAlbB infection was stable at high temperatures and showed perfect maternal transmission and cytoplasmic incompatibility regardless of the host background. Our results demonstrate the stability of wAlbB across host backgrounds and point to its long-term effectiveness for controlling arbovirus transmission and mosquito populations. IMPORTANCEWolbachia bacteria are being used to control the transmission of dengue virus and other arboviruses by mosquitoes. For Wolbachia release programs to be effective globally, Wolbachia infections must be stable across mosquito populations from different locations. In this study, we transferred Wolbachia (strain wAlbB) to Aedes aegypti mosquitoes with an Australian genotype and introduced the infection to Malaysian mosquitoes through backcrossing. We found that the phenotypic effects of Wolbachia are stable across both mosquito backgrounds. We sequenced the genome of wAlbB and found very few genetic changes despite spending over 15 years in a novel mosquito host. Our results suggest that the effects of Wolbachia infections are likely to remain stable across time and host genotype.