Temperature affects the tripartite interactions between bacteriophage WO, Wolbachia, and cytoplasmic incompatibility

Wolbachia infections are a model for understanding intracellular, bacterial symbioses. While the symbiosis is often studied from a binary perspective of host and bacteria, it is increasingly apparent that additional trophic levels can influence the symbiosis. For example, Wolbachia in arthropods harbor a widespread temperate bacteriophage, termed WO, that forms virions and rampantly transfers between coinfections. Here we test the hypothesis that temperatures at the extreme edges of an insect's habitable range alter bacteriophage WO inducibility and in turn, Wolbachia densities and the penetrance of cytoplasmic incompatibility. We report four key findings using the model wasp, Nasonia vitripennis: First, both cold treatment at 18 C and heat treatment at 30 C reduce Wolbachia densities by as much as 74% relative to wasps reared at 25 C. Second, in all cases where Wolbachia densities decline due to temperature changes, phage WO densities increase and inversely associate with Wolbachia densities. Heat has a marked effect on phage WO, yielding phage densities that are 552% higher than the room temperature control. Third, there is a significant affect of insect family on phage WO and endoysmbiont densities. Fourth, at extreme temperatures, there was a temperature-mediated adjustment to the density threshold at which Wolbachia cause complete cytoplasmic incompatibility. Taken together, these results demonstrate that temperature simultaneously affects phage WO densities, endosymbiont densities, and the penetrance of cytoplasmic incompatibility. While temperature shock enhances bacteriophage inducibility and the ensuing bacterial mortality in a wide range of medically and industrially-important bacteria, this is the first investigation of the associations in an obligate intracellular bacteria. Implications to a SOS global sensing feedback mechanism in Wolbachia are discussed.     

Solving the Wolbachia paradox: modeling the tripartite interaction between host, Wolbachia, and a natural enemy

Wolbachia is one of the most common symbionts of arthropods. Its establishment requires lateral transfer to and successful transmission within novel host species. However, Wolbachia performs poorly when introduced into new host species, and models predict that Wolbachia should seldom be able to establish from low initial frequencies. Recently, various symbionts, including Wolbachia, have been shown to protect their hosts from natural enemies. Hence, Wolbachia invasion may be facilitated by the dynamic interaction between it, its host, and a natural enemy. We model such an interaction whereby Wolbachia induces either complete resistance, partial resistance, or tolerance to a host-specific pathogen and also induces the common manipulation phenotype of cytoplasmic incompatibility (CI). We show that the presence of the pathogen greatly facilitates Wolbachia invasion from rare and widens the parameter space in which "imperfect" Wolbachia strains can invade. Furthermore, positive frequency-dependent selection through CI can drive Wolbachia to very high frequencies, potentially excluding the pathogen. These results may explain a poorly understood aspect of Wolbachia biology: it is widespread, despite performing poorly after transfer to new host species. They also support the intriguing possibility that Wolbachia strains that encode both CI and natural-enemy resistance could potentially rid insects, including human disease vectors, of important pathogens.     

Tripartite associations among bacteriophage WO, Wolbachia, and host affected by temperature and age in Tetranychus urticae

A phage density model of cytoplasmic incompatibility (CI), which means lytic phages reduce bacterial density associated with CI, significantly enhances our understanding of the tripartite associations among bacteriophage WO, Wolbachia and host. However, WO may alternate between lytic and lysogenic life cycles or change phage production under certain conditions including temperature, host age and host species background. Here, extreme temperatures can induce an alteration in the life cycle of WO and change the tripartite associations among WO, Wolbachia and CI. Based on the accumulation of the WO load, WO can transform into the lytic life cycle with increasing age. These findings confirmed that the environment plays an important role in the associations among WO, Wolbachia and host.     

Phylogenetic relationships of the Wolbachia of nematodes and arthropods

Wolbachia are well known as bacterial symbionts of arthropods, where they are reproductive parasites, but have also been described from nematode hosts, where the symbiotic interaction has features of mutualism. The majority of arthropod Wolbachia belong to clades A and B, while nematode Wolbachia mostly belong to clades C and D, but these relationships have been based on analysis of a small number of genes. To investigate the evolution and relationships of Wolbachia symbionts we have sequenced over 70 kb of the genome of wOvo, a Wolbachia from the human-parasitic nematode Onchocerca volvulus, and compared the genes identified to orthologues in other sequenced Wolbachia genomes. In comparisons of conserved local synteny, we find that wBm, from the nematode Brugia malayi, and wMel, from Drosophila melanogaster, are more similar to each other than either is to wOvo. Phylogenetic analysis of the protein-coding and ribosomal RNA genes on the sequenced fragments supports reciprocal monophyly of nematode and arthropod Wolbachia. The nematode Wolbachia did not arise from within the A clade of arthropod Wolbachia, and the root of the Wolbachia clade lies between the nematode and arthropod symbionts. Using the wOvo sequence, we identified a lateral transfer event whereby segments of the Wolbachia genome were inserted into the Onchocerca nuclear genome. This event predated the separation of the human parasite O. volvulus from its cattle-parasitic sister species, O. ochengi. The long association between filarial nematodes and Wolbachia symbionts may permit more frequent genetic exchange between their genomes.     

Review of treatment methods to remove Wolbachia bacteria from arthropods

Wolbachia are intracellular bacteria that infect numerous and diverse arthropod species including economic pests of crops and disease vectors affecting livestock and humans. Research on these symbionts has identified profound effects of Wolbachia on their hosts with possible application in pest control. Such research often requires methods to cure infections. To facilitate future research on these bacteria, we reviewed the Wolbachia literature to summarize results of 110 studies spanning 62 taxa that report on treatment methods and outcomes. Application of tetracycline in diet is the most common method and is typically successful. Rifampicin is secondarily used, and may be successful when tetracycline is not. Elevated temperatures can be used to eliminate infections, but is not often used. Rearing hosts under crowded conditions or starvation has been shown to reduce Wolbachia titre which affects maternal transmission. Application of treatment methods has a number of considerations with possible implications for the interpretation of data. This review is intended to alert the reader to treatment options and potential non-target effects.     

Isolation and characterization of the bacteriophage WO from Wolbachia, an arthropod endosymbiont

Wolbachia is a group of obligate symbiotic bacteria found in many insects and other arthropods. The presence of Wolbachia alters reproduction in the host, but the mechanisms are unknown. Molecular biological studies of Wolbachia have delayed significantly, and one of the reasons is the lack of transformation techniques of this bacterium. In the present study, bacteriophage particles were isolated from Wolbachia for the first time. The purified phage had an isometric head that was approximately 40 nm in diameter and contained linear double-stranded DNA of approximately 20 kbp. Partial sequence information (total of 20,484 bp) revealed that there were 24 open reading frames including a structural gene module, and genes for replication and lysogenic conversion. This bacteriophage is the only known mobile genetic element potentially used for transformation of Wolbachia.     

Wolbachia endosymbionts responsible for various alterations of sexuality in arthropods.

Rickettsia-like maternally inherited bacteria have been shown to be involved in a variety of alterations of arthropod sexuality, such as female-biased sex ratios, parthenogenesis, and sterility of crosses either between infected males and uninfected females or between infected individuals (cytoplasmic incompatibility). We have characterized several of these microorganisms through partial sequences of the small (16S) and large (23S) subunit ribosomal DNA. All the symbionts identified, which include several cytoplasmic incompatibility microorganisms, several endosymbionts of terrestrial isopods, and symbionts of two thelytokous Trichogramma wasp species, belong to a monophyletic group of related symbionts, some of which have previously been detected in several insects exhibiting cytoplasmic incompatibility. Three molecular lineages can be identified on the basis of 16S as well as 23S sequences. Although they are only known as endocellular symbionts, Wolbachia spread by horizontal transfer across host lineages as evidenced by their diversification which occurred long after that of their hosts, and by the non-congruence of the phylogenetic relationships of symbionts and their hosts. Indeed, symbionts of two different lineages have been found in the same host species, whereas closely related endosymbionts are found in distinct insect orders. Isopod endosymbionts form a separate lineage, and they can determine feminization as well as cytoplasmic incompatibility. The ability to determine cytoplasmic incompatibility, found in all lineages, is probably ancestral to this group.     

Wolbachia prophage DNA adenine methyltransferase genes in different Drosophila-Wolbachia associations

Wolbachia is an obligatory intracellular bacterium which often manipulates the reproduction of its insect and isopod hosts. In contrast, Wolbachia is an essential symbiont in filarial nematodes. Lately, Wolbachia has been implicated in genomic imprinting of host DNA through cytosine methylation. The importance of DNA methylation in cell fate and biology calls for in depth studying of putative methylation-related genes. We present a molecular and phylogenetic analysis of a putative DNA adenine methyltransferase encoded by a prophage in the Wolbachia genome. Two slightly different copies of the gene, met1 and met2, exhibit a different distribution over various Wolbachia strains. The met2 gene is present in the majority of strains, in wAu, however, it contains a frameshift caused by a 2 bp deletion. Phylogenetic analysis of the met2 DNA sequences suggests a long association of the gene with the Wolbachia host strains. In addition, our analysis provides evidence for previously unnoticed multiple infections, the detection of which is critical for the molecular elucidation of modification and/or rescue mechanism of cytoplasmic incompatibility.     

Phylogenetic evidence for horizontal transmission of Wolbachia in host-parasitoid associations

Endosymbiotic Wolbachia infect a number of arthropod species in which they can affect the reproductive system. While maternally transmitted, unlike mitochondria their molecular phylogeny does not parallel that of their hosts. This strongly suggests horizontal transmission among species, the mechanisms of which remain unknown. Such transfers require intimate between-species relationships, and thus host-parasite associations are outstandingly appropriate for study. Here, we demonstrate that hymenopteran parasitoids of frugivorous Drosophila species are especially susceptible to Wolbachia infection. Of the five common European species, four proved to be infected; furthermore, multiple infections are common, with one species being doubly infected and two triply infected (first report). Phylogenetic statuses of the Wolbachia infecting the different species of the community have been studied using the gene wsp, a highly variable gene recently described. This study reveals exciting similarities between the Wolbachia variants found in parasitoids and their hosts. These arguments strongly support the hypothesis of frequent natural Wolbachia transfers into other species and open a new field for genetic exchanges among species, especially in host-parasitoid associations.     

The associations between chronotype,a healthy diet and obesity

Unhealthy diet has been associated with obesity. Evening type has been associated with unhealthier food and nutrient intake that could predict a higher risk of obesity among them as compared to morning type. However, thus far no study has examined the interrelationships between chronotype, a healthy diet and obesity. We examined whether a healthy diet mediates the association between chronotype and obesity and whether chronotype modifies the association between a healthy and obesity. The National FINRISK 2007 Study included 4421 subjects aged 25–74 years. Diet was assessed using a validated food frequency questionnaire. Baltic Sea diet score (BSDS), including nine dietary components, was used as a measure of adherence to a healthy Nordic diet. Weight, height, body fat percentage and waist circumference were measured, and body mass index values were calculated. Chronotype was assessed using a shortened version of Horne and Östberg’s morningness–eveningness questionnaire (MEQ). The sum score calculated from MEQ was either used as a continuous variable or divided into tertiles of which the lowest tertile demonstrated evening preference and the highest tertile demonstrated morning preference. A series of regression analyses were conducted to determine whether the BSDS mediates the association between chronotype and obesity. Likelihood ratio test was used to determine whether chronotype modifies the association between the BSDS and the obesity measures. After testing the interaction, chronotype-stratified analysis for the association between the BSDS and obesity measures was determined by linear regression. Generally, the evening types had lower adherence to the BSDS and were more often smokers (men), physically inactive and had lower perceived health than the other chronotypes (p < 0.05). The poorer health behavior of this group, however, was not manifested in their obesity measures, and no evidence that the BSDS would mediate the association between chronotype and obesity was found (p > 0.05). No evidence that chronotype would modify the association between the BSDS and obesity was found either (p > 0.05).     

Emerging systems: between vertebrates and arthropods,the Lophotrochozoa

Novel molecular model organisms for the study of development and regeneration are emerging among the Lophotrochozoa, the third major branch of bilaterian animals. The polychaete Platynereis, the leech Helobdella, the snail Ilyanassa, and several planarians are efficiently accessed for molecular techniques including large-scale whole-mount in situ hybridization screening, RNA interference or morpholino knock-down. Joint efforts include the generation of genomic resources in the form of expressed sequence tag collections and bacterial artificial chromosome libraries. Current research focuses on early pattern formation during cleavage, the emergence and diversification of body segments, and the formation of photoreceptor cells and eyes. Several lophotrochozoan groups (in particular nereid polychaetes) exhibit modes of development, organ design, or body plans that are considered ancestral in many respects. This is also reflected in the level of genes, making these groups ideally suited for developmental comparative studies.     

A Survey of the bacteriophage WO in the endosymbiotic bacteria Wolbachia

Bacteriophages are common viruses infecting prokaryotes. In addition to their deadly effect, phages are also involved in several evolutionary processes of bacteria, such as coding functional proteins potentially beneficial to them, or favoring horizontal gene transfer through transduction. The particular lifestyle of obligatory intracellular bacteria usually protects them from phage infection. However, Wolbachia, an intracellular alpha-proteobacterium, infecting diverse arthropod and nematode species and best known for the reproductive alterations it induces, harbors a phage named WO, which has recently been proven to be lytic. Here, phage infection was checked in 31 Wolbachia strains, which induce 5 different effects in their hosts and infect 25 insect species and 3 nematodes. Only the Wolbachia infecting nematodes and Trichogramma were found devoid of phage infection. All the 25 detected phages were characterized by the DNA sequence of a minor capsid protein gene. Based on all data currently available, phylogenetic analyses show a lack of congruency between Wolbachia or insect and phage WO phylogenies, indicating numerous horizontal transfers of phage among the different Wolbachia strains. The absence of relation between phage phylogeny and the effects induced by Wolbachia suggests that WO is not directly involved in these effects. Implications on phage WO evolution are discussed.     

Genes for the type IV secretion system in an intracellular symbiont, Wolbachia, a causative agent of various sexual alterations in arthropods

Wolbachia species are intracellular bacteria known to cause reproductive abnormalities in their hosts. In this study, we identified Wolbachia genes encoding homologs to the type IV secretion system by which many pathogenic bacteria secrete macromolecules. The genes identified encoded most of the essential components of the secretion system and were cotranscribed as an operon.     

Evolutionary genomics of a temperate bacteriophage in an obligate intracellular bacteria (Wolbachia)

Genome evolution of bacteria is usually influenced by ecology, such that bacteria with a free-living stage have large genomes and high rates of horizontal gene transfer, while obligate intracellular bacteria have small genomes with typically low amounts of gene exchange. However, recent studies indicate that obligate intracellular species that host-switch frequently harbor agents of horizontal transfer such as mobile elements. For example, the temperate double-stranded DNA bacteriophage WO in Wolbachia persistently transfers between bacterial coinfections in the same host. Here we show that despite the phage's rampant mobility between coinfections, the prophage's genome displays features of constraint related to its intracellular niche. First, there is always at least one intact prophage WO and usually several degenerate, independently-acquired WO prophages in each Wolbachia genome. Second, while the prophage genomes are modular in composition with genes of similar function grouping together, the modules are generally not interchangeable with other unrelated phages and thus do not evolve by the Modular Theory. Third, there is an unusual core genome that strictly consists of head and baseplate genes; other gene modules are frequently deleted. Fourth, the prophage recombinases are diverse and there is no conserved integration sequence. Finally, the molecular evolutionary forces acting on prophage WO are point mutation, intragenic recombination, deletion, and purifying selection. Taken together, these analyses indicate that while lateral transfer of phage WO is pervasive between Wolbachia with occasional new gene uptake, constraints of the intracellular niche obstruct extensive mixture between WO and the global phage population. Although the Modular Theory has long been considered the paradigm of temperate bacteriophage evolution in free-living bacteria, it appears irrelevant in phages of obligate intracellular bacteria.     

Shift-specific associations between age,chronotype and sleep duration

The objective of this study was to examine the association of age with chronotype and sleep duration in day workers and rotating shift workers, including night shift work. Between October 2012 and February 2015, a cross-sectional study was conducted in a German chemical company. Using the “Munich ChronoType Questionnaire” (MCTQ), data about sleep onset and sleep offset during workdays and work-free days were retrieved and the chronotype was computed during regular voluntary occupational health check-ups. Associations between age and chronotype, as well as sleep duration, were assessed using linear regression analyses. Potential effect modification by the working time system was examined. Within the study period, 4,040 employees (82.3% and 17.7% were engaged in day work and rotating shift work, respectively) completed the questionnaire. Study participants were on average 41.8 years old (Min = 18.0, Max = 65.0, SD = 10.2) and predominantly male (75.4%). Mean chronotype and overall sleep duration was 03:22 (SD = 54 min) and 7.2 h (SD = 1.0 h) respectively. Older age was associated with earlier chronotype and reduced overall sleep duration in both day workers and rotating shift workers (p < 0.001 for all models). Compared to day workers, employees whom engaged in rotating shift work were later chronotypes and had overall a longer sleep duration. With older age, the difference between day and rotating shift workers regarding chronotype increased, while the difference regarding overall sleep duration decreased (p_interaction<0.005 for both models). This finding could indicate that both changes in circadian physiology and exposure to certain work schedules contribute to the age-related changes. Older rotating shift workers, with early chronotypes may have issues with night shifts, while day work and morning shifts may be best compatible to earlier chronotypes. Differences in sleep timing across age groups, might indicate that the same work hours will affect shift workers differently, dependent on their age, suggesting that more flexible and chronotype-adapted work hours could provide useful; especially for older employees. Sleep education in the form of courses and health campaigns could be a way to raise awareness of the importance of a healthy sleep pattern. This could be achieved by learning strategies to better adjust individual sleep patterns to work hours.     

Lateral transfers of insertion sequences between Wolbachia,Cardinium and Rickettsia bacterial endosymbionts

Various bacteria live exclusively within arthropod cells and collectively act as an important driver of arthropod evolutionary ecology. Whereas rampant intra-generic DNA transfers were recently shown to have a pivotal role in the evolution of the most common of these endosymbionts, Wolbachia, the present study show that inter-generic DNA transfers also commonly take place, constituting a potent source of rapid genomic change. Bioinformatic, molecular and phylogenetic data provide evidence that a selfish genetic element, the insertion sequence ISRpe1, is widespread in the Wolbachia, Cardinium and Rickettsia endosymbionts and experiences recent (and likely ongoing) transfers over long evolutionary distances. Although many ISRpe1 copies were clearly expanding and leading to rapid endosymbiont diversification, degraded copies are also frequently found, constituting an unusual genomic fossil record suggestive of ancient ISRpe1 expansions. Overall, the present data highlight how ecological connections within the arthropod intracellular environment facilitate lateral DNA transfers between distantly related bacterial lineages.     

节肢动物的分类和演化

节肢动物门是动物界中最大的1个门。按新的分类系统,本门分三叶虫亚门(已灭绝)、螯肢亚门、甲壳亚门、六足亚门和多足亚门等5亚门。六足亚门相当于以前分类系统中的昆虫纲(广义的), 是最重要的一类节肢动物;此亚门分原尾纲、弹尾纲、双尾纲和昆虫纲(狭义的)等4纲;昆虫纲分3亚纲30目,包括了前昆虫纲“有翅亚纲”中的各目。为便于读者了解新、旧系统的异同,文中列举前人的代表性系统加以对照。同时,对学术界关于节肢动物的起源和演化的一些新观点予以必要的说明,对现时流行的“泛节肢动物”、“泛甲壳动物”等概念作了简要的介绍。     

Specificity of host-parasite relations between arthropods and terrestrial vertebrates

Specificity of partners in host-parasite system is one of its main characteristics. Unfortunately this term has different senses in scientific literature. In everyday practice one judges an extent of host specificity of a parasite mainly by indices of its occurrence and abundance on different host species. An occurrence of parasites in nature reflects general result of complex eco-physiological interrelationships between partners in hostparasite system. Specificity of parasites in a choice of hosts may depend on a belonging of the latter to certain taxa (phylogenetic specificity), or on biotic and abiotic factors (ecological specificity). In arthropods, the phylogentic specificity and coevolution are characteristic to a greater extent for permanent hosts (lice, Mallophaga, cheyletoid and feather mites). A coevolutionaryphylogenesis is disturbed by transfers of parasites onto new hosts, by different rates of speciation in filial lines or by an extinction of several parasite taxa. In temporary parasites different forms of ecological specificity are prevalent. A host specificity is expressed to the lesser extent in mosquitoes, horseflies and in other blood-sucking Diptera. In temporary parasites with a long-term feeding (ticks) coevolutionary sequences are relatively rare, because this parasites had to adapt not only to a life on host, but also to a lesser stable environment. In some nest-burrow bloodsuckers (fleas, gamasid mites and argasid ticks) the ecological specificity is shown no by their relations with certain host species, but by an associations with habitats occupied by hosts (burrow, nests, caves). In relation with a high dynamics of host-parasite system, a specificity of its partners is comparative and it is kept up only under specific ecological conditions.     

Spacer 2 region and 5S rDNA variation of Wolbachia strains involved in cytoplasmic incompatibility or sex-ratio distortion in arthropods

Bacteria in the genus Wolbachia are widespread in arthropods and can induce sex-ratio distortion or cytoplasmic incompatibility in their hosts. The phylogeny of Wolbachia has been studied using 16S rDNA and the cell cycle gene ftsZ, but sequence variation of those genes is limited. The spacer 2 region (SR2) was amplified to determine whether this region would improve phylogenetic resolution. The SR2 of Wolbachia is 66 bp long, shows higher variation than ftsZ and has very low homology with closely related bacteria. Due to the small length of SR2 of Wolbachia, little phylogenetic information could be retrieved.