Wolbachia Infection Research Articles

Global Dynamics for Competition between Two Wolbachia Strains with Bidirectional Cytoplasmic Incompatibility

Releasing Wolbachia-infected mosquitoes to suppress or replace wild vector mosquitoes has been carried out in 24 countries worldwide, showing great promise in controlling mosquitoes and mosquito-borne diseases. To face the instability of Wolbachia infection in different environments during the area-wide application, we should consider the overlapping of two Wolbachia strains. In this case, bidirectional cytoplasmic incompatibility occurs, which results in mating partners infected with exclusive Wolbachia strains producing inviable offspring. To determine the better Wolbachia candidate for release, we develop an ordinary differential equation model to study the global dynamics for competition between two Wolbachia strains. Our theoretical results on the sharp estimate of stable curves completely determine the fate of the two Wolbachia strains, which help choose appropriate strains for release.

When does the female bias arise? Insights from the sex determination cascade of a flea beetle with a strongly skewed sex ratio

Reproduction-manipulating bacteria like Wolbachia can shift sex ratios in insects towards females, but skewed sex ratios may also arise from genetic conflicts. The flea beetle Altica lythri harbors three main mtDNA strains that are coupled to three different Wolbachia infections. Depending on the mtDNA types, the females produce either offspring with a balanced sex ratio or exclusively daughters. To obtain markers that can monitor when sex bias arises in the beetle’s ontogeny, we elucidated the sex determination cascade of A. lythri. We established a RT-PCR method based on length variants of dsx (doublesex) transcripts to determine the sex of morphologically indistinguishable eggs and larvae. In females of one mtDNA type (HT1/HT1*) known to produce only daughters, male offspring were already missing at the egg stage while for females of another type (HT2), the dsx splice variants revealed a balanced sex ratio among eggs and larvae. Our data suggest that the sex determination cascade in A. lythri is initiated by maternally transmitted female-specific tra (transformer) mRNA as primary signal. This tra mRNA seems to be involved in a positive feedback loop that maintains the production of the female splice variant, as known for female offspring in Tribolium castaneum. The translation of the maternally transmitted female tra mRNA must be inhibited in male offspring, but the underlying primary genetic signal remains to be identified. We discuss which differences between the mtDNA types can influence sex determination and lead to the skewed sex ratio of HT1.

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.

Characterization of the Bacterial Microbiome in Natural Populations of Barley Stem Gall Midge, Mayetiola hordei, in Morocco

Mayetiola hordei (Kieffer), known as barley stem gall midge, is one of the most destructive barley pests in many areas around the world, inflicting significant qualitative and quantitative damage to crop production. In this study, we investigate the presence of reproductive symbionts, the effect of geographical origin on the bacterial microbiome's structure, and the diversity associated with natural populations of M. hordei located in four barley-producing areas in Morocco. Wolbachia infection was discovered in 9% of the natural populations using a precise 16S rDNA PCR assay. High-throughput sequencing of the V3-V4 region of the bacterial 16S rRNA gene indicated that the native environments of samples had a substantial environmental impact on the microbiota taxonomic assortment. Briefly, 5 phyla, 7 classes, and 42 genera were identified across all the samples. To our knowledge, this is the first report on the bacterial composition of M. hordei natural populations. The presence of Wolbachia infection may assist in the diagnosis of ideal natural populations, providing a new insight into the employment of Wolbachia in the control of barley midge populations, in the context of the sterile insect technique or other biological control methods.

The effects of three release strategies on Wolbachia infection frequency in mosquito populations

ABSTRACT In control of wild mosquitoes to fight mosquito-borne diseases, release of mosquitoes with Wolbachia is one of the effective biological control methods. There are three release strategies, namely releasing both Wolbachia-infected females and males, only Wolbachia-infected females and only Wolbachia-infected males. All these three strategies have been confirmed to be capable of speeding up the Wolbachia persistence in mosquito populations. In this paper, we investigate how supplementary releases affect the Wolbachia spread dynamics in mosquito populations. Our aim is to compare the effectiveness among these three release strategies. We obtain theoretical results and provide numerical simulations that show that the first two strategies are more effective than the last strategy. For the first two strategies, the former strategy is either less effective than the latter strategy in each generation, or more effective than the latter strategy in previous generations, and then becomes less effective in late generations.

Native Wolbachia infection and larval competition stress shape fitness and West Nile virus infection in Culex quinquefasciatus mosquitoes.

Wolbachia transinfections established in key mosquito vectors, including Aedes aegypti are typically associated with pathogen blocking-reduced susceptibility to infection with key pathogens and reduced likelihood those pathogens are transmitted to new hosts. Host-symbiont-virus interactions are less well understood in mosquitoes like Culex quinquefasciatus, which naturally harbor Wolbachia, with pathogen blocking observed in some populations but not others, potentially due to innate differences in their Wolbachia load. In nature, mosquito larvae are often subject to developmental stresses associated with larval competition, which can lead to reduced body size and differential susceptibility to arbovirus infection. In this study, we sought to understand whether competition stress and Wolbachia infection in Cx. quinquefasciatus combine to impact host fitness and susceptibility to infection with West Nile virus. We reared Wolbachia-infected and uninfected Cx. quinquefasciatus larvae under three competition stress levels, increasing larval density without increasing the amount of food supplied. We then monitored larval development and survival, measured wing length and quantified Wolbachia density in adults, and then challenged mosquitoes from each treatment group orally with West Nile virus. We observed that high competition stress extended development time, decreased the likelihood of eclosion, decreased body size, and increased susceptibility to West Nile virus (WNV) infection. We also observed that Wolbachia infection reduced WNV load under low competition stress, and significantly improved the rate of survival for larval reared under higher competition stress. Consequently, our data suggest that native Wolbachia infection in Cx. quinquefasciatus has differential consequences for host fitness and susceptibility to WNV infection depending on competition stress.

First report of natural Wolbachia infections in mosquitoes from Cuba

Mosquitoes are extensively responsible for the transmission of pathogens. Novel strategies using Wolbachia could transform that scenario, since these bacteria manipulate mosquito reproduction, and can confer a pathogen transmission-blocking phenotype in culicids. Here, we screened the Wolbachia surface protein region by PCR in eight Cuban mosquito species. We confirmed the natural infections by sequencing and assessed the phylogenetic relationships among the Wolbachia strains detected. We identified four Wolbachia hosts: Aedes albopictus, Culex quinquefasciatus, Mansonia titillans, and Aedes mediovittatus (first report worldwide). Knowledge of Wolbachia strains and their natural hosts is essential for future operationalization of this vector control strategy in Cuba.

Modeling Wolbachia infection frequency in mosquito populations via a continuous periodic switching model

Abstract In this article, we develop a continuous periodic switching model depicting Wolbachia infection frequency dynamics in mosquito populations by releasing Wolbachia-infected mosquitoes, which is different from the discrete modeling efforts in the literature. We obtain sufficient conditions on the existence of a unique and exactly two periodic solutions and analyze the stability of each periodic solution, respectively. We also provide a brief discussion and several numerical examples to illustrate our theoretical results.

Modeling Sustained Transmission of Wolbachia among Anopheles Mosquitoes: Implications for Malaria Control in Haiti.

Wolbachia infection in Anopheles albimanus mosquitoes can render mosquitoes less capable of spreading malaria. We developed and analyzed a mechanistic compartmental ordinary differential equation model to evaluate the effectiveness of Wolbachia-based vector control strategies among wild Anopheles mosquitoes in Haiti. The model tracks the mosquito life stages, including egg, larva, and adult (male and female). It also accounts for critical biological effects, such as the maternal transmission of Wolbachia through infected females and cytoplasmic incompatibility, which effectively sterilizes uninfected females when they mate with infected males. We derive and interpret dimensionless numbers, including the basic reproductive number and next-generation numbers. The proposed system presents a backward bifurcation, which indicates a threshold infection that needs to be exceeded to establish a stable Wolbachia infection. The sensitivity analysis ranks the relative importance of the epidemiological parameters at baseline. We simulate different intervention scenarios, including prerelease mitigation using larviciding and thermal fogging before the release, multiple releases of infected populations, and different release times of the year. Our simulations show that the most efficient approach to establishing Wolbachia is to release all the infected mosquitoes immediately after the prerelease mitigation process. Moreover, the model predicts that it is more efficient to release during the dry season than the wet season.

NDUFA8 potentially rescues Wolbachia-induced cytoplasmic incompatibility in Laodelphax striatellus.

The endosymbiont Wolbachia manipulates host reproduction by several strategies, one of the most important of which is cytoplasmic incompatibility (CI). CI can be rescued when Wolbachia-infected males mate with females infected with the same Wolbachia strain. However, the potential rescue mechanism of CI in the small brown planthopper Laodelphax striatellus is unclear. In this study, comparative transcriptome analysis was applied to explore the effect of Wolbachia on L. striatellus eggs. A total of 1387 differentially expressed genes were identified. RNA interference of 7 Wolbachia-upregulated key planthopper genes reduced egg reproduction, suggesting that Wolbachia might improve fecundity in L. striatellus by affecting these 7 genes. Suppressing the expression of another upregulated gene, NDUFA8 (encoding NADH dehydrogenase [ubiquinone] 1 α subcomplex subunit 8-like) by RNA interference significantly increased the mortality of early embryos without affecting the number of deposited eggs. Wolbachia infection upregulated the mRNA level of NDUFA8, and dsNDUFA8 treatment of Wolbachia-infected females recreated CI-like symptoms, suggesting that NDUFA8 is associated with the rescue phenotype. Because all L. striatellus populations worldwide are infected with Wolbachia, NDUFA8 is a potential pest control target.

The association of host and vector characteristics with Ctenocephalides felis pathogen and endosymbiont infection.

Surveillance of the fleas and flea-borne pathogens infecting cats is important for both human and animal health. Multiple zoonotic Bartonella and Rickettsia species are known to infect the most common flea infesting cats and dogs worldwide: Ctenocephalides felis, the cat flea. The ability of other flea species to transmit pathogens is relatively unexplored. We aimed to determine cat host and flea factors independently associated with flea Bartonella and Rickettsia infection. We also assessed flea and cat infection by flea-host pair and location. To accomplish these aims, we performed qPCR for the detection of Bartonella, hemotropic Mycoplasma, Rickettsia, and Wolbachia DNA using paired cat and flea samples obtained from free-roaming cats presenting for spay or neuter across four locations in the United States. A logistic regression model was employed to identify the effect of cat (sex, body weight, geographic location, and Bartonella, hemotropic Mycoplasma, and Rickettsia spp., infection) and flea (clade and Rickettsia and Wolbachia infection) factors on C. felis Bartonella clarridgeiae infection. From 189 free roaming cats, we collected 84 fleas: Ctenocephalides felis (78/84), Cediopsylla simplex (4/84), Orchopeas howardi (1/84), and Nosopsyllus fasciatus (1/84). Ctenocephalides felis were phylogenetically assigned to Clades 1, 4, and 6 by cox1 gene amplification. Rickettsia asembonensis (52/84) and B. clarridgeiae (16/84) were the most common pathogenic bacteria detected in fleas. Our model identified host cat sex and weight as independently associated with B. clarridgeiae infection in fleas. Rickettsia asembonensis (52/84), Rickettsia felis (7/84) and Bartonella henselae (7/84) were detected in specific clades: R. felis was detected only in Clades 1 and 6 while B. henselae and R. asembonensis were detected only in Clade 4. Wolbachia spp., also displayed clade specificity with strains other than Wolbachia wCfeT only infecting fleas from Clade 6. There was poor flea and host agreement for Bartonella spp., infection; however, there was agreement in the Bartonella species detected in cats and fleas by geographic location. These findings reinforce the importance of considering reservoir host attributes and vector phylogenetic diversity in epidemiological studies of flea-borne pathogens. Widespread sampling is necessary to identify the factors driving flea-borne pathogen presence and transmission.

Diversity of Wolbachia infection and its influence on mitochondrial DNA variation in the diamondback moth, Plutella xylostella

Plutella xylostella is a pest that severely damages cruciferous vegetables worldwide and has been shown to be infected with the maternally inherited bacteria Wolbachia, with the main infected strain was plutWB1. In this study, we performed a large-scale global sampling of P. xylostella and amplified 3 mtDNA genes of P. xylostella and 6 Wolbachia genes to analyze the infection status, diversity of Wolbachia in P. xylostella, and its effect on mtDNA variation in P. xylostella. This study provides a conservative estimate of Wolbachia infection rates in P. xylostella, which was found to be 7% (104/1440). The ST 108 (plutWB1) was shared among butterfly species and the moth species P. xylostella, revealing that Wolbachia strain plutWB1 acquisition in P. xylostella may be through horizontal transmission. The Parafit analyses indicated a significant association between Wolbachia and Wolbachia-infected P. xylostella individuals, and individuals infected with plutWB1 tended to cluster in the basal positions of the phylogenetic tree based on the mtDNA data. Additionally, Wolbachia infections were associated with increased mtDNA polymorphism in the infected P. xylostella population. These data suggest that Wolbachia endosymbionts may have a potential effect on mtDNA variation of P. xylostella.

Investigating the Diversity of Wolbachia across the Spiny Ants (Polyrhachis)

Among insects, Wolbachia is an exceedingly common bacterial endosymbiont with a range of consequences of infection. Despite the frequency of Wolbachia infection, very little is known about this bacteria’s diversity and role within hosts, especially within ant hosts. In this study, we analyze the occurrence and diversity of Wolbachia across the spiny ants (Polyrhachis), a large and geographically diverse genus. Polyrhachis samples from throughout the host genus’ phylogenetic and biogeographical range were first screened for single infections of Wolbachia using the wsp gene and Sanger sequencing. The multilocus sequence typing (MLST) scheme was then used on these singly infected samples to identify the Wolbachia strains. A Wolbachia phylogeny was inferred from the Polyrhachis samples analyzed in this study as well as other Formicidae MLST profiles from the MLST online database. We hypothesized that three key host factors were impacting Wolbachia diversity within the Polyrhachis genus: biogeography, phylogeny, and species level. The results suggest that the phylogeny and biogeography of Polyrhachis hosts have no impact on Wolbachia diversity; however, species level may have some limited influence. Additionally, Wolbachia strains appear to group according to being either Old World or New World strains. Among the taxa able to form complete MLST allelic profiles, all twenty are seemingly new strains.

Ovarian transcriptional response to Wolbachia infection in D. melanogaster in the context of between-genotype variation in gene expression.

Wolbachia is a maternally transmitted endosymbiotic bacteria that infects a wide variety of arthropod and nematode hosts. The effects of Wolbachia on host biology are far-reaching and include changes in host gene expression. However, previous work on the host transcriptional response has generally been investigated in the context of a single host genotype. Thus, the relative effect of Wolbachia infection versus vs. host genotype on gene expression is unknown. Here, we explicitly test the relative roles of Wolbachia infection and host genotype on host gene expression by comparing the ovarian transcriptomes of 4 strains of Drosophila melanogaster (D. melanogaster) infected and uninfected with Wolbachia. Our data suggest that infection explains a small amount of transcriptional variation, particularly in comparison to variation in gene expression among strains. However, infection specifically affects genes related to cell cycle, translation, and metabolism. We also find enrichment of cell division and recombination processes among genes with infection-associated differential expression. Broadly, the transcriptomic changes identified in this study provide novel understanding of the relative magnitude of the effect of Wolbachia infection on gene expression in the context of host genetic variation and also point to genes that are consistently differentially expressed in response to infection among multiple genotypes.

Effects of Wolbachia on Transposable Element Expression Vary Between Drosophila melanogaster Host Genotypes.

Transposable elements (TEs) are repetitive DNA sequences capable of changing position in host genomes, thereby causing mutations. TE insertions typically have deleterious effects but they can also be beneficial. Increasing evidence of the contribution of TEs to adaptive evolution further raises interest in understanding what factors impact TE activity. Based on previous studies associating the bacterial endosymbiont Wolbachia with changes in the abundance of piRNAs, a mechanism for TE repression, and to transposition of specific TEs, we hypothesized that Wolbachia infection would interfere with TE activity. We tested this hypothesis by studying the expression of 14 TEs in a panel of 25 Drosophila melanogaster host genotypes, naturally infected with Wolbachia and annotated for TE insertions. The host genotypes differed significantly in Wolbachia titers inside individual flies, with broad-sense heritability around 20%, and in the number of TE insertions, which depended greatly on TE identity. By removing Wolbachia from the target host genotypes, we generated a panel of 25 pairs of Wolbachia-positive and Wolbachia-negative lines in which we quantified transcription levels for our target TEs. We found variation in TE expression that was dependent on Wolbachia status, TE identity, and host genotype. Comparing between pairs of Wolbachia-positive and Wolbachia-negative flies, we found that Wolbachia removal affected TE expression in 21.1% of the TE-genotype combinations tested, with up to 2.3 times differences in the median level of transcript. Our data show that Wolbachia can impact TE activity in host genomes, underscoring the importance this endosymbiont can have in the generation of genetic novelty in hosts.

Two Complete Genomes of Male-Killing Wolbachia Infecting Ostrinia Moth Species Illuminate Their Evolutionary Dynamics and Association with Hosts.

Wolbachia is an extremely widespread intracellular symbiont which causes reproductive manipulation on various arthropod hosts. Male progenies are killed in Wolbachia-infected lineages of the Japanese Ostrinia moth population. While the mechanism of male killing and the evolutionary interaction between host and symbiont are significant concerns for this system, the absence of Wolbachia genomic information has limited approaches to these issues. We determined the complete genome sequences of wFur and wSca, the male-killing Wolbachia of Ostrinia furnacalis and Ostrinia scapulalis. The two genomes shared an extremely high degree of homology, with over 95% of the predicted protein sequences being identical. A comparison of these two genomes revealed nearly minimal genome evolution, with a strong emphasis on the frequent genome rearrangements and the rapid evolution of ankyrin repeat-containing proteins. Additionally, we determined the mitochondrial genomes of both species' infected lineages and performed phylogenetic analyses to deduce the evolutionary dynamics of Wolbachia infection in the Ostrinia clade. According to the inferred phylogenetic relationship, two possible scenarios were proposed: (1) Wolbachia infection was established in the Ostrinia clade prior to the speciation of related species such as O. furnacalis and O. scapulalis, or (2) Wolbachia infection in these species was introgressively transferred from a currently unidentified relative. Simultaneously, the relatively high homology of mitochondrial genomes suggested recent Wolbachia introgression between infected Ostrinia species. The findings of this study collectively shed light on the host-symbiont interaction from an evolutionary standpoint.

In vitro maintenance of the endosymbiont Wolbachia of Dirofilaria immitis.

Wolbachia has an obligatory mutualistic relationship with many onchocercid nematodes of the subfamilies Dirofilariinae and Onchocercinae. Till date, no attempts have been made for the in vitro cultivation of this intracellular bacteriumfrom thefilarioid host. Hence, the current study attempted cell co-culture method using embryonic Drosophila S2 and the LD cell lines to cultivate Wolbachia from Dirofilaria immitis microfilariae (mfs) harvested from infected dogs. Microfilariae (mfs = 1500) were inoculated in shell vials supplemented with Schneider medium using both cell lines. The establishment and multiplication of the bacterium were observed during the initial inoculation, at day 0 and before every medium change (from days 14 to 115). An aliquot (50µl) from each time point was tested by quantitative real-time PCR (qPCR). Comparing the average of Ct values, obtained by the tested parameters (i.e., LD/S2 cell lines and mfs with/without treatment), the S2 cell line without mechanical disruption of mfs provided the highest Wolbachia cell count by qPCR. Despite the maintenance of Wolbachia within both S2 and LD-based cell co-culture models for up to 115days, a definitive conclusion is still far. Further trials using fluorescent microscopy and viable staining will help to demonstrate the cell line infection and viability of Wolbachia. Use of considerable amount of untreated mfs to inoculate the Drosophilia S2 cell lines, as well as the supplementation of the culture media with growth stimulants or pre-treated cells to increase their susceptibility for the infection and development of a filarioid-based cell line system are recommended for the future trials.

Distribution of Wolbachia infection in butterflies (Lepidoptera): First systematic report from Thailand

ABSTRACT Wolbachia is a genus of Rickettsia-like bacteria that infects a broad range of insect species throughout the world. It often influences host reproduction to facilitate its transmission to their offspring and spread through populations. The occurrence and ecology of Wolbachia has not been surveyed in detail in Thai butterflies. In this study, we conduct the first systematic survey of Wolbachia infection by sampling 623 specimens from 46 butterfly species from 5 families obtained from tropical forests in 3 geographic regions of Thailand. Wolbachia infections were detected using three PCR primer sets: 16S rRNA, ftsZ and wsp. The results showed evidence for widespread Wolbachia infection in Thai butterflies. The Wolbachia presence was confirmed in 291 individuals from 17 butterfly species of 2 families, comprising 86 individuals from the Central Region (32 males and 54 females), 72 individuals from the Northeast Region (25 males and 47 females), and 133 individuals from the West Region (46 males and 87 females). This report will be useful for understanding the distribution of Wolbachia in butterflies from Thailand.

A Systematic Review of Mathematical Models of Dengue Transmission and Vector Control: 2010-2020.

Vector control methods are considered effective in averting dengue transmission. However, several factors may modify their impact. Of these controls, chemical methods, in the long run, may increase mosquitoes' resistance to chemicides, thereby decreasing control efficacy. The biological methods, which may be self-sustaining and very effective, could be hampered by seasonality or heatwaves (resulting in, e.g., loss of Wolbachia infection). The environmental methods that could be more effective than the chemical methods are under-investigated. In this study, a systematic review is conducted to explore the present understanding of the effectiveness of vector control approaches via dengue transmission models.

Species composition of the parasitoid genus Eurytoma (Hymenoptera: Eurytomidae) varies at a local scale in Diplolepis variabilis (Hymenoptera: Cynipidae) galls

Abstract We evaluated the diversity and spatiotemporal emergence patterns of parasitoid wasp species in the genus Eurytoma Illiger (Hymenoptera: Eurytomidae) associated with leaf galls induced by Diplolepis variabilis Bassett (Hymenoptera: Cynipidae) throughout the Okanagan Valley, British Columbia, Canada. We collected D. variabilis galls throughout the region, recorded adult Eurytoma emergence times, extracted mitochondria DNA from a subset of the emergents, and used cytochrome c oxidase 1 and cytochrome b amplificons to assign sampled individuals to species. We also assayed for the bacterial endosymbiont Wolbachia to evaluate the incidence of thelytoky in Eurytoma. Eurytoma showed an extended emergence period and a significant right skew in male emergence. We recorded six Eurytoma species, one of which appears to be previously undescribed. The greatest diversity of Eurytoma species occurs in the north of the sampled area. Five of the six species showed Wolbachia infection in at least some individuals, and the most abundant and widespread species, Eurytoma imminuta Bugbee, showed high Wolbachia infection levels and an all-female population, which strongly indicates endosymbiont-induced thelytoky. Our results demonstrate that gall communities are locally variable, and we suggest that future studies be undertaken to quantify variation in community composition at more local scales than has been past practice.