Characterization of the symbiont Rickettsia in the mirid bug Nesidiocoris tenuis (Reuter) (Heteroptera: Miridae).

Nesidiocoris tenuis (Reuter) (Heteroptera: Miridae) is an omnivorous insect used for biological control. Augmentative release and conservation of N. tenuis have been used for pest control in tomato crops. Intracellular bacterial symbionts of arthropods are common in nature and have diverse effects on their hosts; in some cases they can dramatically affect biological control. Fingerprinting methods showed that the symbiotic complex associated with N. tenuis includes Wolbachia and Rickettsia. Rickettsia of N. tenuis was further characterized by sequencing the 16S rRNA and gltA bacterial genes, measuring its amount in different developmental stages of the insect by real-time polymerase chain reaction, and localizing the bacteria in the insect's body by fluorescence in situ hybridization. The Rickettsia in N. tenuis exhibited 99 and 96% similarity of both sequenced genes to Rickettsia bellii and Rickettsia reported from Bemisia tabaci, respectively. The highest amount of Rickettsia was measured in the 5th instar and adult, and the symbionts could be detected in the host gut and ovaries. Although the role played by Rickettsia in the biology of N. tenuis is currently unknown, their high amount in the adults and localization in the gut suggest that they may have a nutritional role in this insect.

Synchrotron X-ray micro-computed tomography as a tool for in situ elucidation of insect bacteriomes.

Obligate bacterial endosymbionts are common, influential associates of arthropods, and are often found in specific organs termed bacteriomes. Three dimensional images of bacteriomes of the leafhopper Orosius albicinctus (Hemiptera: Cicadellidae) were reconstructed from synchrotron-based X-ray micro-computed tomography (CT). Results show that bilateral bacteriomes are located between the first and second abdominal tergites, are mushroom-shaped and consist two different types of tissue. Fluorescence in situ hybridization reveals that the primary bacterial symbiont Sulcia muelleri is in the 'cap' part of the of organ. The technique allows a noninvasive, in situ, means of visualizing bacteriomes and will facilitate understanding their form and function.

Biotype-dependent secondary symbiont communities in sympatric populations of Bemisia tabaci.

The sweet potato whitefly, Bemisia tabaci, harbors Portiera aleyrodidarum, an obligatory symbiotic bacterium, as well as several secondary symbionts including Rickettsia, Hamiltonella, Wolbachia, Arsenophonus, Cardinium and Fritschea, the function of which is unknown. Bemisia tabaci is a species complex composed of numerous biotypes, which may differ from each other both genetically and biologically. Only the B and Q biotypes have been reported from Israel. Secondary symbiont infection frequencies of Israeli laboratory and field populations of B. tabaci from various host plants were determined by PCR, in order to test for correlation between bacterial composition to biotype and host plant. Hamiltonella was detected only in populations of the B biotype, while Wolbachia and Arsenophonus were found only in the Q biotype (33% and 87% infection, respectively). Rickettsia was abundant in both biotypes. Cardinium and Fritschea were not found in any of the populations. No differences in secondary symbionts were found among host plants within the B biotype; but within the Q biotype, all whiteflies collected from sage harboured both Rickettsia and Arsenophonus, an infection frequency which was significantly higher than those found in association with all other host plants. The association found between whitefly biotypes and secondary symbionts suggests a possible contribution of these bacteria to host characteristics such as insecticide resistance, host range, virus transmission and speciation.

Negative evidence of Wolbachia in the predaceous mite Phytoseiulus persimilis.

The cytoplasmically inherited bacterium Wolbachia is widespread in arthropod species and has been repeatedly detected in the predaceous mite Phytoseiulus persimilis. Our original goal was to assess the prevalence of Wolbachia infection in P. persimilis and the potential fitness consequences for this host. To accomplish that goal, seven P. persimilis strains were obtained from Europe, Africa and the USA and reared on the phytophagous mite Tetranychus urticae as prey. After preliminary results showed that the T. urticae used was infected with Wolbachia, the minimum starvation time of the predators to prevent false positive results from undigested prey was determined. We tested DNA samples by PCR (polymerase chain reaction) after starving the predators or feeding them Wolbachia-free T. urticae for various periods. Those experiments showed that Wolbachia could not be detected after 16 h at 25 degrees C and 48 h at 20 degrees C. To verify the results of the PCR analyses, we furthermore conducted crossing experiments with antibiotic-treated and untreated individuals. No indications of Wolbachia effects were recorded. Additionally, we screened live eggs of four of the seven strains reared in our laboratory and alcohol samples of 10 other P. persimilis strains for the occurrence of Wolbachia by PCR, none of which tested positive. Synthesis of our study and previous reports suggests that infection of P. persimilis with Wolbachia is extremely rare and of minor importance. We discuss the significance of our findings for future studies on the presence of Wolbachia in predaceous arthropods.

Distribution of parthenogenesis-inducing symbionts in ovaries and eggs of Aphytis (Hymentoptera: Aphelinidae).

In parasitic Hymenoptera, severe sex ratio distortions are sometimes associated with the presence of symbiotic microorganisms. Symbiont-induced uniparental reproduction is found in several species of the genus Aphytis and is suspected in others. These intracellular bacteria (called Wolbachia) reside within the egg cytoplasm. In order to follow their vertical transmission, the distribution pattern of the symbionts in ovaries and eggs of two uniparental lines (A. lingnanensis and A. yanonensis) was studied by electron, confocal, and two-photon microscopy. The results obtained suggest that Wolbachia move from nurse cells to developing oocytes. In freshly laid eggs, the symbionts are concentrated in the posterior pole, away from the micropyle, and in later stages of embryogenesis they are found surrounding nuclei throughout the embryo.

Wolbachia density and host fitness components in Muscidifurax uniraptor (Hymenoptera: pteromalidae).

Intracellular bacteria of the genus Wolbachia are found in a variety of arthropod hosts, where they cause various reproductive disorders. Attempts to study the fitness advantages and disadvantages of carrying these symbionts have yielded contradicting results. Using various doses of the antibiotic rifampicin, we were able to manipulate the density of Wolbachia in the uniparental parasitoid Muscidifurax uniraptor (Hymenoptera: Pteromalidae). The effect of different titers of the symbiont on the fecundity, reproductive rate, longevity, survival rate, and sex ratio of the host was measured. The data gathered show that following antibiotic treatments, the percentage of males rises at low doses of rifampicin and then drops again. The total sex ratio of offspring produced by treated mothers was positively correlated with the numbers of Wolbachia found in eggs laid by these females. No significant effects were detected with regard to the other studied fitness components. It is concluded that in M. uniraptor, Wolbachia are not posing any burden on the life history trait studied.

Phylogenetic analysis of parthenogenesis-inducing Wolbachia in the genus Aphytis (Hymenoptera: Aphelinidae).

Parthenogenesis-inducing intracellular bacteria of the genus Wolbachia are found in a variety of parasitoid wasp genera. The presence of Wolbachia in the uniparental Aphytis species A. lingnanensis Compere, A. diaspidis (Howard), A. chilensis Howard, and A. chrysomphali (Mercet) was tested using primers specific for the ftsZ gene. The symbiont was detected in all of these species. Wolbachia ftsZ genes that were sequenced from the four hosts show a high degree of similarity. Both the PCR with specific primers for group 'A' and phylogenetic analysis place these Wolbachia in group 'A'. The fact that the tested Aphytis species belong to different phylogenetic groups and harbour what seem to be almost identical Wolbachia, lends credence to the horizontal transmission hypothesis.

Parthenogenesis-inducing microorganisms in Aphytis (Hymenoptera: Aphelinidae).

Production of males in uniparental lines of two species in the parasitic wasp genus Aphytis was induced by rifampicin, and male sexual functioning was determined. Wolbachia-specific 16S rDNA primers were used in a PCR in order to: (1) assess correlation between thelytokous reproduction and the presence of Wolbachia; (2) detect the loss of Wolbachia DNA in uniparental A. lingnanensis following antibiotic treatments, with or without the presence of a host; and (3) clone and sequence part of the Wolbachia 16S rDNA from the uniparental Aphytis species for phylogenetic studies. Males produced viable sperm that was transferred to the female spermatheca following mating. However, sperm failure to effect egg fertilization resulted in all-male progeny. Wolbachia were found in the two uniparental (A. lingnanensis and A. diaspidis) but not in the two biparental (A. lingnanensis and A. melinus) Aphytis lines tested. They can be detected in wasps up to 7 days following antibiotic treatments, regardless of the presence of host. The 16S rDNA for the symbionts in the two Aphytis species is virtually identical, and is most closely related to the Wolbachia found in Muscidifurax uniraptor (Pteromalidae).

A newly discovered bacterium associated with parthenogenesis and a change in host selection behavior in parasitoid wasps.

The symbiotic bacterium Wolbachia pipientis has been considered unique in its ability to cause multiple reproductive anomalies in its arthropod hosts. Here we report that an undescribed bacterium is vertically transmitted and associated with thelytokous parthenogenetic reproduction in Encarsia, a genus of parasitoid wasps. Although Wolbachia was found in only one of seven parthenogenetic Encarsia populations examined, the "Encarsia bacterium" (EB) was found in the other six. Among seven sexually reproducing populations screened, EB was present in one, and none harbored Wolbachia. Antibiotic treatment did not induce male production in Encarsia pergandiella but changed the oviposition behavior of females. Cured females accepted one host type at the same rate as control females but parasitized significantly fewer of the other host type. Phylogenetic analysis based on the 16S rDNA gene sequence places the EB in a unique clade within the Cytophaga-Flexibacter-Bacteroid group and shows EB is unrelated to the Proteobacteria, where Wolbachia and most other insect symbionts are found. These results imply evolution of the induction of parthenogenesis in a lineage other than Wolbachia. Importantly, these results also suggest that EB may modify the behavior of its wasp carrier in a way that enhances its transmission.