Wolbachia and DNA barcoding insects: patterns, potential and problems

Wolbachia is a genus of bacterial endosymbionts that impacts the breeding systems of their hosts. Wolbachia can confuse the patterns of mitochondrial variation, including DNA barcodes, because it influences the pathways through which mitochondria are inherited. We examined the extent to which these endosymbionts are detected in routine DNA barcoding, assessed their impact upon the insect sequence divergence and identification accuracy, and considered the variation present in Wolbachia COI. Using both standard PCR assays (Wolbachia surface coding protein – wsp), and bacterial COI fragments we found evidence of Wolbachia in insect total genomic extracts created for DNA barcoding library construction. When >2 million insect COI trace files were examined on the Barcode of Life Datasystem (BOLD) Wolbachia COI was present in 0.16% of the cases. It is possible to generate Wolbachia COI using standard insect primers; however, that amplicon was never confused with the COI of the host. Wolbachia alleles recovered were predominantly Supergroup A and were broadly distributed geographically and phylogenetically. We conclude that the presence of the Wolbachia DNA in total genomic extracts made from insects is unlikely to compromise the accuracy of the DNA barcode library; in fact, the ability to query this DNA library (the database and the extracts) for endosymbionts is one of the ancillary benefits of such a large scale endeavor – for which we provide several examples. It is our conclusion that regular assays for Wolbachia presence and type can, and should, be adopted by large scale insect barcoding initiatives. While COI is one of the five multi-locus sequence typing (MLST) genes used for categorizing Wolbachia, there is limited overlap with the eukaryotic DNA barcode region

Wolbachia in the flesh: symbiont intensities in germ-line and somatic tissues challenge the conventional view of Wolbachia transmission routes

Symbionts can substantially affect the evolution and ecology of their hosts. The investigation of the tissue-specific distribution of symbionts (tissue tropism) can provide important insight into host-symbiont interactions. Among other things, it can help to discern the importance of specific transmission routes and potential phenotypic effects. The intracellular bacterial symbiont Wolbachia has been described as the greatest ever panzootic, due to the wide array of arthropods that it infects. Being primarily vertically transmitted, it is expected that the transmission of Wolbachia would be enhanced by focusing infection in the reproductive tissues. In social insect hosts, this tropism would logically extend to reproductive rather than sterile castes, since the latter constitute a dead-end for vertically transmission. Here, we show that Wolbachia are not focused on reproductive tissues of eusocial insects, and that non-reproductive tissues of queens and workers of the ant Acromyrmex echinatior, harbour substantial infections. In particular, the comparatively high intensities of Wolbachia in the haemolymph, fat body, and faeces, suggest potential for horizontal transmission via parasitoids and the faecal-oral route, or a role for Wolbachia modulating the immune response of this host. It may be that somatic tissues and castes are not the evolutionary dead-end for Wolbachia that is commonly thought

Detection of Wolbachia in the Tick Ixodes ricinus is Due to the Presence of the Hymenoptera Endoparasitoid Ixodiphagus hookeri

The identification of micro-organisms carried by ticks is an important issue for human and animal health. In addition to their role as pathogen vectors, ticks are also the hosts for symbiotic bacteria whose impact on tick biology is poorly known. Among these, the bacterium Wolbachia pipientis has already been reported associated with Ixodes ricinus and other tick species. However, the origins of Wolbachia in ticks and their consequences on tick biology (known to be very diverse in invertebrates, ranging from nutritional symbionts in nematodes to reproductive manipulators in insects) are unknown. Here we report that the endoparasitoid wasp Ixodiphagus hookeri (Hymenoptera, Chalcidoidea, Encyrtidae) – strictly associated with ticks for their development - is infested at almost 100% prevalence by a W. pipientis strain belonging to a Wolbachia supergroup that has already been reported as associated with other hymenopteran parasitoids. In a natural population of I. ricinus that suffers high parasitism rates due to I. hookeri, we used specific PCR primers for both hymenopteran and W. pipientis gene fragments to show that all unfed tick nymphs parasitized by I. hookeri also harbored Wolbachia, while unparasitized ticks were Wolbachia-free. We demonstrated experimentally that unfed nymphs obtained from larvae exposed to I. hookeri while gorging on their vertebrate host also harbor Wolbachia. We hypothesize that previous studies that have reported W. pipientis in ticks are due to the cryptic presence of the endoparasitoid wasp I. hookeri. This association has remained hidden until now because parasitoids within ticks cannot be detected until engorgement of the nymphs brings the wasp eggs out of diapause. Finally, we discuss the consequences of this finding for our understanding of the tick microbiome, and their possible role in horizontal gene transfer among pathogenic and symbiotic bacteria

Behavioral and genetic characteristics of a new species of Nasonia

Nasonia (Hymenoptera: Pteromalidae) is a genus of parasitoid wasps, which is fast emerging as a model system for evolutionary, genetic, developmental and host–endosymbiont interaction studies. In this study, we report a new species, Nasonia oneida, distinguish its behavioral, genetic and morphological features, and characterize its pre-mating and post-mating isolation with the other Nasonia species. Phylogenetic analyses indicate that N. oneida is the sister species to Nasonia giraulti with its own uniquely distinct cuticular hydrocarbon profiles, behavioral characteristics and subtle morphological differences. An important characteristic of N. oneida is the strong mate discrimination shown by the females against all the other Nasonia species. A genetic analysis of this phenotype by interspecies hybridization indicates that this strong discriminating phenotype is recessive. A formal species description of N. oneida Raychoudhury & Desjardins is also provided.

Data from: Cuticular hydrocarbon divergence in the jewel wasp Nasonia: Evolutionary shifts in chemical communication channels?

The data package contains eight datasets: - CSV files containing information on the GC-MS data (GC: 7890A; MS: 5975C; Agilent Technologies, Waldbronn, Germany) for all Nasonia samples, obtained with “Enhanced Chemstation”, G1701AA, Version A.03.00. Columns are identified as follows: Peak number, average retention time, peak type, peak width, peak area (integrated absolute value), start retention time, end retention time. Further description of the data generation see Materials and Methods section in the paper. - CSV files containing information on the GC-MS data (GC: 7890A; MS: 5975C; Agilent Technologies, Waldbronn, Germany) for the Trichomalopsis outgroup samples, obtained with “Enhanced Chemstation”, G1701AA, Version A.03.00. Columns are identified as follows: Peak number, average retention time, peak type, peak width, peak area (integrated absolute value), start retention time, end retention time. Further description of the data generation see Materials and Methods section in the paper. - CHC peak areas standardized with the normalization method of the function "decostand" of the community ecology R package “vegan” (Dixon, 2003), for a more detailed description see Materials and Methods section in the paper. - Functions of the linear discriminant analysis (DA) calculated from the normalized CHC data on the Nasonia samples, first three functions plotted simultaneously in Figure 1 in the paper, further details on the DA see Materials and Methods and Results, respectively. - Phylogenetic distance matrix, used in comparison with the average manhattan chemical distances between the Nasonia species based on the relative differences between their normalized CHC peak areas, performed separately for males and females (Fig. 2 and 3, respectively). The molecular distances are based on the Nasonia phylogeny, which are themselves based on the genetic divergence of a mitochondrial gene, COI sequences retrieved from NCBI, pairwise p-distances calculated with MEGA 3.0 (Werren et al., 2010) For further information, please see Materials and Methods section in the paper as well as the Tables S2 and S3, which give the average manhattan distances the phylogenetic distances were compared to. - Single results of the Mantel tests, performed five times for both male CHC divergence vs. Nasonia phylogeny and female CHC divergence versus Nasonia phylogeny, all five p values and their respective averages are given for both combinations, as well as the r value. Each single test was performed with 9999 permutations. - Raw data on the behavioral assays, focussing on N. oneida and N. giraulti males (Fig. 4). Whether a copulation has occurred or not is given in the second column for each pairing in a binary coding (1 = yes, 0 = no), and the average number of occurred copulations is given for each pairing with untreated and treated female dummies. Species affiliation and treatment of female dummies indicated in the caption of each pairing. Male species affiliation can be found in the main caption on each spread sheet. Furthermore, the male arrestment time has been recorded (third column), but those data have not been used in this study. Mating results on two further N. giraulti strains are given as well. - Relative (percentages) distribution of CHC compound classes among the species and sexes of Nasonia and the outgroup Trichomalopsis, data underlying Fig. S2

Budapesti Szemle

Here we report evidence of a mitochondrial–Wolbachia sweep in North American populations of the parasitoid wasp Nasonia vitripennis, a cosmopolitan species and emerging model organism for evolutionary and genetic studies. Analysis of the genetic variation of 89 N. vitripennis specimens from Europe and North America was performed using four types of genetic markers: a portion of the mitochondrial cytochrome oxidase I gene, nine polymorphic nuclear microsatellites, sequences from 11 single-copy nuclear markers and six Wolbachia genes. The results show that the European populations have a sevenfold higher mitochondrial sequence variation than North American populations, but similar levels of microsatellite and nuclear gene sequence variation. Variation in the North American mitochondria is extremely low (π=0.31%), despite a highly elevated mutation rate (~35–40 times higher than the nuclear genes) in the mitochondria of Nasonia. The data are indicative of a mitochondrial sweep in the North American population, possibly due to Wolbachia infections that are maternally co-inherited with the mitochondria. Owing to similar levels of nuclear variation, the data could not resolve whether N. vitripennis originated in the New or the Old World.