Wolbachia, sex ratio bias and apparent male killing in the harlequin beetle riding pseudoscorpion

Bacterial endosymbionts that manipulate host reproduction are now known to be widespread in insects and other arthropods. Since they inhabit the cytoplasm and are maternally inherited, these microorganisms can enhance their fitness by biasing host sex ratio in favour of females. At its most extreme, sex ratio manipulation may be achieved by killing male embryos, as occurs in a number of insect species. Here, we provide evidence for the first case of male killing by a tetracycline-sensitive microbe in pseudoscorpions. Using a combination of inheritance studies, antibiotic treatment and molecular assays, we show that a new strain of Wolbachia is associated with extreme female bias in the pseudoscorpion, Cordylochernes scorpioides. In a highly female-biased line, sex ratio distortion was maternally inherited, and occurred in conjunction with a high rate of spontaneous abortion and low reproductive success. Antibiotic treatment cured females of the Wolbachia infection, restored offspring sex ratio to 1:1, and significantly enhanced female reproductive success. The discovery of apparent male-killing in C. scorpioides is of interest because pseudoscorpions are viviparous. Theory predicts that male killing should be favoured, if male death enhances the fitness of infected female siblings. In a live-bearing host, reallocation of maternal resources from dead male embryos to their sisters provides a direct, physiological mechanism through which fitness compensation could favour male killing by cellular endosymbionts. Our results suggest, however, that fitness compensation and the spread of male-killing endosymbionts may be undermined by a high rate of spontaneous abortion in infected females of this viviparous arthropod.     

Reproductive compensation favours male-killing Wolbachia in a live-bearing host

Wolbachia are maternally inherited, cellular endosymbionts that can enhance their fitness by biasing host sex ratio in favour of females. Male killing (MK) is an extreme form of sex-ratio manipulation that is selectively advantageous if the self-sacrifice of Wolbachia in males increases transmission through females. In live-bearing hosts, females typically produce more embryos than can be carried to term, and reproductive compensation through maternal resource reallocation from dead males to female embryos could increase the number of daughters born to infected females. Here, we report a new strain of MK Wolbachia (wCsc2) in the pseudoscorpion, Cordylochernes scorpioides, and present the first empirical evidence that reproductive compensation favours the killing of males in a viviparous host. Females infected with the wCsc2 strain produced 26 per cent more and significantly larger daughters than tetracycline-cured females. In contrast to the previously described wCsc1 MK Wolbachia strain in C. scorpioides, wCsc2 infection was not accompanied by an increase in the rate of spontaneous brood abortion. Characterization of the wCsc1 and wCsc2 strains by multi-locus sequence typing and by Wolbachia surface protein (wsp) gene sequencing indicates that the marked divergence between these two MK strains in their impact on host reproductive success, and hence in their potential to spread, has occurred in association with homologous recombination in the wsp gene.     

Maintenance of a male-killing Wolbachia in Drosophila innubila by male-killing dependent and male-killing independent mechanisms

Many maternally inherited endosymbionts manipulate their host's reproduction in various ways to enhance their own fitness. One such mechanism is male killing (MK), in which sons of infected mothers are killed by the endosymbiont during development. Several hypotheses have been proposed to explain the advantages of MK, including resource reallocation from sons to daughters of infected females, avoidance of inbreeding by infected females, and, if transmission is not purely maternal, the facilitation of horizontal transmission to uninfected females. We tested these hypotheses in Drosophila innubila, a mycophagous species infected with MK Wolbachia. There was no evidence of horizontal transmission in the wild and no evidence Wolbachia reduced levels of inbreeding. Resource reallocation does appear to be operative, as Wolbachia-infected females are slightly larger, on average, than uninfected females, although the selective advantage of larger size is insufficient to account for the frequency of infection in natural populations. Wolbachia-infected females from the wild-although not those from the laboratory-were more fecund than uninfected females. Experimental studies revealed that Wolbachia can boost the fecundity of nutrient-deprived flies and reduce the adverse effect of RNA virus infection. Thus, this MK endosymbiont can provide direct, MK-independent fitness benefits to infected female hosts in addition to possible benefits mediated via MK.     

Occasional males in parthenogenetic populations of Asobara japonica (Hymenoptera: Braconidae): low Wolbachia titer or incomplete coadaptation?

Wolbachia are endosymbiotic bacteria known to manipulate the reproduction of their hosts. Some populations of the parasitoid wasp Asobara japonica are infected with Wolbachia and reproduce parthenogenetically, while other populations are not infected and reproduce sexually. Wolbachia-infected A. japonica females regularly produce small numbers of male offspring. Because all females in the field are infected and infected females are not capable of sexual reproduction, male production seems to be maladaptive. We investigated why these females nevertheless produce males. We tested three hypotheses: high rearing temperatures could result in higher offspring sex ratios (more males), low Wolbachia titer of the mother could lead to higher offspring sex ratios and/or the Wolbachia infection is of relatively recent origin and not enough time has passed to allow complete coadaptation between Wolbachia and host. In all, 33% of the Wolbachia-infected females produced males and 56% of these males were also infected with Wolbachia. Neither offspring sex ratio nor male infection frequency was significantly affected by rearing temperature or Wolbachia concentration of the mother. The mitochondrial DNA sequence of one of the uninfected populations was identical to that of two of the infected populations. Therefore, the initial Wolbachia infection of A. japonica must have occurred recently. Mitochondrial sequence variation among the infected populations suggests that the spread of Wolbachia through the host populations involved horizontal transmission. We conclude that the occasional male production by Wolbachia-infected females is most likely a maladaptive side effect of incomplete coevolution between symbiont and host in this relatively young infection.     

Opposite sex-specific effects of Wolbachia and interference with the sex determination of its host Ostrinia scapulalis

In the adzuki bean borer, Ostrinia scapulalis, the sex ratio in most progenies is 1 : 1. Females from Wolbachia-infected matrilines, however, give rise to all-female broods when infected and to all-male broods when cured of the infection. These observations had been interpreted as Wolbachia-induced feminization of genetic males into functional females. Here, we show that the interpretation is incorrect. Females from both lines have a female karyotype with a WZ sex-chromosome constitution while males are ZZ. At the time of hatching from eggs, WZ and ZZ individuals are present at a 1 : 1 ratio in broods from uninfected, infected and cured females. In broods from Wolbachia-infected females, ZZ individuals die during larval development, whereas in those from cured females, WZ individuals die. Hence, development of ZZ individuals is impaired by Wolbachia but development of WZ females may require the presence of Wolbachia in infected matrilines. Sexual mosaics generated (i) by transfection of uninfected eggs and (ii) by tetracycline treatment of Wolbachia-infected mothers prior to oviposition were ZZ in all tissues, including typically female organs. We conclude that: (i) Wolbachia acts by manipulating the sex determination of its host; and (ii) although sexual mosaics can survive, development of a normal female is incompatible with a ZZ genotype.     

Evolution of early male-killing in horizontally transmitted parasites

Early male-killing (MK) bacteria are vertically transmitted reproductive parasites which kill male offspring that inherit them. Whereas their incidence is well documented, characteristics allowing originally non-MK bacteria to gradually evolve MK ability remain unclear. We show that horizontal transmission is a mechanism enabling vertically transmitted bacteria to evolve fully efficient MK under a wide range of host and parasite characteristics, especially when the efficacy of vertical transmission is high. We also show that an almost 100% vertically transmitted and 100% effective male-killer may evolve from a purely horizontally transmitted non-MK ancestor, and that a 100% efficient male-killer can form a stable coexistence only with a non-MK bacterial strain. Our findings are in line with the empirical evidence on current MK bacteria, explain their high efficacy in killing infected male embryos and their variability within and across insect taxa, and suggest that they may have evolved independently in phylogenetically distinct species.     

No variation for Wolbachia-induced hybrid breakdown in two populations of a spider mite

Wolbachia are cytoplasmically transmitted bacteria that infect several species of mites. In the two-spotted spider mite Tetranychus urticae Koch this symbiont can induce reproductive incompatibility. Wolbachia-induced reproductive incompatibility is observed in crosses between Wolbachia-infected (W) males and uninfected (U) females. This incompatibility is expressed in F1 broods as male-biased sex ratios, an effect called cytoplasmic incompatibility (CI). However, in the two-spotted spider mite, Wolbachia-induced reproductive incompatibility may extend to the F2: broods of virgin F1 females from U×W crosses sometimes suffer increased mortality rates. This F2 effect is called hybrid breakdown (HB). Several isofemale lines derived from mites collected from rose and cucumber plants had been previously tested for CI. Here we report on the results obtained for HB.     

VARIATION IN OFFSPRING SIZE WITH BIRTH ORDER IN PLACENTAL FISH: A ROLE FOR ASYMMETRIC SIBLING COMPETITION?

Asymmetric sibling competition arises when siblings with different competitive abilities share a limited resource. Such competition occurs in species with postnatal parental care and may also occur when mothers provision embryos between fertilization and birth (matrotrophy). We hypothesized that the combination of matrotrophy and the simultaneous provisioning of embryos in different stages of development (superfetation) leads to asymmetric competition between sibling embryos. Moreover, we expect the intensity of this competition to increase with the level of superfetation as high levels of superfetation result in greater temporal overlap between broods. This hypothesis predicts that offspring from early broods, which predominantly compete with less‐developed siblings, will be larger at birth than offspring from later broods, which experience competition from more and less‐developed siblings. Data on offspring size at birth from two populations of the highly matrotrophic fish, Heterandria formosa, and similar studies of poeciliid fish spanning a range of life histories are consistent with our hypothesis. Together these results suggest that sibling competition is a direct consequence of the evolution of matrotrophy and superfetation in poeciliid fish.     

Wolbachia与昆虫精卵细胞质不亲和

Wolbachia是广泛分布在昆虫体内的一类共生菌,能通过多种机制调节宿主的生殖方式,包括诱导宿主精卵细胞质不亲和(CI)、孤雌生殖、雌性化、杀雄等,其中细胞质不亲和为最普遍的表型,即感染Wolbachia的雄性和未感染或感染不同品系Wolbachia的雌性宿主交配后,受精卵不能正常发育,在胚胎期死亡。多数CI胚胎在第1次分裂时,来自父本的染色质浓缩缺陷,导致父本遗传物质无法正常分配到子细胞中,因而引起胚胎死亡。守门员模型认为,产生CI可能需要有两种因子,其中之一使得精子发生修饰改变,导致受精后雄性原核发育滞后。第2种因子可能与Wolbachia的原噬菌体有关,在胚胎发育后期导致胚胎死亡。近期的研究已发现,在Wolbachia感染的宿主中,一些与生殖细胞发生和繁殖相关基因的表达发生了显著改变,Wolbachia可能因此对宿主的生殖产生重大影响,进而导致CI的产生。本文主要综述了CI的细胞学表型、解释CI的模型及其分子机理,向读者展示一个小小的细菌是如何通过精妙的策略影响昆虫宿主的繁殖,从而实现其自身的生存和传播的。     

Sexual mosaics induced by tetracycline treatment in the Wolbachia-infected adzuki bean borer, Ostrinia scapulalis.

Wolbachia-infected Ostrinia scapulalis (Lepidoptera: Crambidae) females generate all-female or nearly all-female broods. Curing the infection by tetracycline treatment during larval stages results in the generation of all-male broods in the next generation. Here we show that sexually mosaic offspring are produced by Wolbachia-infected females treated with tetracycline at the adult stage. The sexual mosaics had wings that were composed of distinctive female and male sectors. Besides wings, the sexually dimorphic mid tibiae displayed an intermediate morphology in some of the mosaics. Many of the mosaic individuals had an abnormal structure of the external genitalia as well, a combination of the male uncus and the female ovipositor. We assume that Wolbachia has a feminizing effect on O. scapulalis genetic males and, hence, incomplete curing of the Wolbachia infection results in the generation of sexually mosaic individuals.     

Investigation of venereal, transplacental, and contact transmission of the Lyme disease spirochete, Borrelia burgdorferi, in Syrian hamsters.

A hamster was inoculated with the SI-1 strain of Borrelia burgdorferi and subsequently served as a host to larval Ixodes scapularis Say. Approximately 68% of the nymphs resulting from the fed larvae were infected. Nymphs from this group were fed on uninfected hamsters, and 3 of 4 males and 6 of 6 females became infected. The infected hamsters were allowed to mate with uninfected partners to test for venereal transmission. Six infected females were mated with 6 uninfected males, whereas 3 infected males were mated with 6 uninfected females. None of the uninfected hamsters became infected after mating. Two protocols were used to determine if transplacental transmission of B. burgdorferi occurred. One group included 6 nonpregnant infected females that were subsequently mated and became pregnant. Three of the females were allowed to carry to full term, whereas the other 3 were killed prior to parturition. All fetuses and offspring were negative for B. burgdorferi based on cultures and monoclonal antibody assays. Another group of 6 females was infected via tick bite after becoming pregnant; those females were allowed to carry fetuses to birth and all were negative. Attempts at contact transmission of B. burgdorferi from 2 infected females to 2 uninfected male and 2 uninfected female hamsters and from 2 infected males to 2 uninfected male and uninfected female hamsters via urine or feces failed.     

THE EFFECT OF SIBMATING ON THE INFECTION DYNAMICS OF MALE-KILLING BACTERIA

Male-killing (MK) bacteria are vertically transmitted endosymbionts that selectively kill the male offspring of their hosts. Simple mathematical models describe the infection dynamics using two parameters, the bacterial transmission rate and a fitness compensation for siblings of killed males. These models cannot explain two phenomena that have been observed in nature: the persistence of extreme MK causing all-female broods, and the coexistence of two different strains of MK bacteria in the same host population. In the present study, we extend the simple MK models and investigate theoretically the effects of sibmating on the infection dynamics. We demonstrate analytically that, in general, sibmating reduces MK prevalence, and can even cause its extinction. As a special case of this finding, we show that sibmating allows a stable coexistence between no infection and extreme MK. Furthermore, we performed computer simulations and showed that, depending on male mating capacity, a stable coexistence of two strains is possible if sibmating occurs but is below a threshold. The results suggest that sibmating might be an important factor for understanding the infection dynamics of MK bacteria.     

Effects of two acanthocephalan parasites on the fecundity and pairing status of female Gammarus pulex (Crustacea: Amphipoda)

Acanthocephalan parasites are known to alter the reproductive biology and physiology of their hosts in various ways. In this study we investigated the influence of two acanthocephalan parasites, Pomphorhynchus laevis and Polymorphus minutus, on the fecundity and pairing success of female Gammarus pulex. The results show that P. laevis and P. minutus affect female intermediate host reproduction in different ways. Females infected with P. minutus were totally castrated, whereas those infected with P. laevis only showed reduced fecundity. The oocytes of P. laevis-infected females showed a similar structure to those of uninfected females, although infected females had a higher proportion of oocytes that had failed to reach complete maturity. In comparison, the oocytes of P. minutus-infected females demonstrate a clearly altered structure that suggests a major disruption to the process of vitellogenesis. In the field, males paired more frequently with uninfected females than with infected ones, and is a stronger effect for P. minutus-infected females than P. laevis-infected females. We suggest that the difference in pairing success of P. minutus-infected and P. laevis-infected females is a direct result of the different effects that the two parasites have on female fecundity.     

Offspring dynamics affect food provisioning,growth and mortality in a brood-caring spider

In brood-caring species, family members are faced with a conflict over resource distribution. While parents are selected to adapt the amount of care according to their offspring''s needs, offspring might be selected to demand more care than optimal for parents. Recent studies on birds have shown that the social network structure of offspring affects the amount of care and thus the fitness of families. Such a network structure of repeated interactions is probably influenced by within-brood relatedness. We experimentally manipulated the group composition in a brood-caring spider to test how the presence of unrelated spiderlings affects the dynamics between female and brood as well as within broods. Broods consisting of siblings grew better and had a lower mortality compared with mixed broods, no matter whether the caring female was a genetic or foster mother. Interestingly, we found that foster mothers lost weight when caring for sibling broods, whereas females caring for mixed broods gained weight. This indicates that females may be willing to share more prey when the brood contains exclusively siblings even if the entire brood is unrelated to the female. Resource distribution may thus be negotiated by offspring dynamics that could have a signalling function to females.     

Vertical transmission of Wolbachia in Tetranychus kanzawai Kishida and Panonychus mori Yokoyama (Acari: Tetranychidae)

The vertical transmission of Wolbachia in two species of spider mite was investigated and compared. One species, Tetranychus kanzawai Kishida, was infected with a modification negative strain of Wolbachia while the other species, Panonychus mori Yokoyama, was infected with a modification positive strain. The infection showed perfect maternal transmission in the laboratory population of T. kanzawai in which Wolbachia-infected females produced infected offspring regardless of whether they mated with infected or uninfected males, and uninfected females produced Wolbachia-free progenies without regard to the infection status of their mating partners. In artificial P. mori populations initiated with 50% infected and 50% uninfected female adults, the infection frequencies among progenies increased with each generation, reaching 100% at the sixth generation in the Sendai population and after the sixth generation in the Toyama population. In another experiment, in which an artificial T. kanzawai population was composed of 50% infected and 50% uninfected female adults, the infection frequency in progeny populations increased very slowly, reaching 62.5% at the 15th generation. The difference in infection frequency in the two spider mites may be due to the different strains of Wolbachia.     

Offspring sex ratio produced by female guppies in the wild correlates with sexual ornaments of their sons

The attractiveness hypothesis predicts that females produce broods with male-biased sex ratios when they mate with attractive males. This hypothesis presumes that sons in broods with male-biased sex ratios sired by attractive males have high reproductive success, whereas the reproductive success of daughters is relatively constant, regardless of the attractiveness of their sires. However, there is little direct evidence for this assumption. We have examined the relationships between offspring sex ratios and (1) sexual ornamentation of sons and (2) body size of daughters in broods from wild female guppies Poecilia reticulata. Wild pregnant females were collected and allowed to give birth in the laboratory. Body size and sexual ornamentation of offspring were measured at maturity. Our analysis revealed a significant positive correlation between offspring sex ratios (the proportion of sons per brood) and the total length as well as the area of orange spots of sons, two attributes that influence female mating preferences in guppies. The sex ratio was not associated with the body size of daughters. These results suggest that by performing adaptive sex allocation according to the expected reproductive success of sons and daughters, female guppies can enhance the overall fitness of their offspring.     

Extra‐pair mating in a socially monogamous and paternal mouth‐brooding cardinalfish

Many vertebrates form monogamous pairs to mate and care for their offspring. However, genetic tools have increasingly shown that offspring often arise from matings outside of the monogamous pair bond. Social monogamy is relatively common in coral reef fishes, but there have been few studies that have confirmed monogamy or extra‐pair reproduction, either for males or for females. Here, long‐term observations and genetic tools were applied to examine the parentage of embryos in a paternally mouth‐brooding cardinalfish, Sphaeramia nematoptera. Paternal care in fishes, such as mouth‐brooding, is thought to be associated with a high degree of confidence in paternity. Two years of observations confirmed that S. nematoptera form long‐term pair bonds within larger groups. However, genetic parentage revealed extra‐pair mating by both sexes. Of 105 broods analysed from 64 males, 30.1% were mothered by a female that was not the partner and 11.5% of broods included eggs from two females. Despite the high paternal investment associated with mouth‐brooding, 7.6% of broods were fertilized by two males. Extra‐pair matings appeared to be opportunistic encounters with individuals from outside the immediate group. We argue that while pair formation contributes to group cohesion, both males and females can maximize lifetime reproductive success by taking advantage of extra‐pair mating opportunities.     

Effects of mating order and male size on embryo survival in a pipefish

In species that provide parental care, individuals should invest adaptively in their offspring in relation to the pre‐ and post‐zygotic care provided by their partners. In the broad‐nosed pipefish, Syngnathus typhle L., females transfer large, nutrient‐rich eggs into the male brood pouch during mating. The male broods and nourishes the embryos for several weeks before independent juveniles emerge at parturition. Given a choice, females clearly prefer large partners. Yet, females provide protein‐richer eggs when the same individual mates with a smaller than a larger male. In the present study, we allowed each female to mate with one small and one large male, in alternated order. We found a strong effect of female mating order, with larger clutches and higher embryo mortality in first‐ than second‐laid broods, which may suggest that eggs over‐ripen in the ovaries or reflect the negative effects of high embryo density in the brood pouch. In either case, this effect should put constraints on the possibility of a female being selective in mate choice. We also found that small and large males produced embryos of similar size and survival, consistent with the reproductive compensation hypothesis, suggesting that, in this species, larger males provide better nourishment to the embryos than smaller males. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2015, 114 , 639–645.     

Wolbachia affects oviposition and mating behaviour of its spider mite host

Wolbachia bacteria are transmitted from mother to offspring via the cytoplasm of the egg. When mated to males infected with Wolbachia bacteria, uninfected females produce unviable offspring, a phenomenon called cytoplasmic incompatibility (CI). Current theory predicts that ‘sterilization’ of uninfected females by infected males confers a fitness advantage to Wolbachia in infected females. When the infection is above a threshold frequency in a panmictic population, CI reduces the fitness of uninfected females below that of infected females and, consequently, the proportion of infected hosts increases. CI is a mechanism that benefits the bacteria but, apparently, not the host. The host could benefit from avoiding incompatible mates. Parasite load and disease resistance are known to be involved in mate choice. Can Wolbachia also be implicated in reproductive behaviour? We used the two‐spotted spider mite – Wolbachia symbiosis to address this question. Our results suggest that uninfected females preferably mate to uninfected males while infected females aggregate their offspring, thereby promoting sib mating. Our data agrees with other results that hosts of Wolbachia do not necessarily behave as innocent bystanders – host mechanisms that avoid CI can evolve.