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Facultative bacterial endosymbionts are associated with many arthropods and are primarily transmitted vertically from mother to offspring. However, phylogenetic affiliations suggest that horizontal transmission must also occur. Such horizontal transfer can have important biological and agricultural consequences when endosymbionts increase host fitness. So far horizontal transmission is considered rare and has been difficult to document. Here, we use fluorescence in situ hybridization (FISH) and multi locus sequence typing (MLST) to reveal a potentially common pathway of horizontal transmission of endosymbionts via parasitoids of insects. We illustrate that the mouthparts and ovipositors of an aphelinid parasitoid become contaminated with Wolbachia when this wasp feeds on or probes Wolbachia-infected Bemisia tabaci AsiaII7, and non-lethal probing of uninfected B. tabaci AsiaII7 nymphs by parasitoids carrying Wolbachia resulted in newly and stably infected B. tabaci matrilines. After they were exposed to infected whitefly, the parasitoids were able to transmit Wolbachia efficiently for the following 48 h. Whitefly infected with Wolbachia by parasitoids had increased survival and reduced development times. Overall, our study provides evidence for the horizontal transmission of Wolbachia between insect hosts by parasitic wasps, and the enhanced survival and reproductive abilities of insect hosts may adversely affect biological control programs.

Bemisia tabaci (Hemiptera: Aleyrodidae) is a globally distributed pest composed of at least 34 morphologically indistinguishable cryptic species. At least seven species of endosymbiont have been found infecting some or all members of the complex. The origin(s) of the associations between specific endosymbionts and their whitefly hosts is unknown. Infection is normally vertical, but horizontal transmission does occur and is one way for new infections to be introduced into individuals. The relationships between the different members of the cryptic species complex and the endosymbionts have not been well explored. In this study, the phylogenies of different cryptic species of the host with those of their endosymbionts were compared. Of particular interest was whether there was evidence for both coevolution and horizontal transmission. Congruence was observed for the primary endosymbiont, Portiera aleyrodidarum, and partial incongruence in the case of two secondary endosymbionts, Arsenophonus and Cardinium and incongruence for a third, Wolbachia. The patterns observed for the primary endosymbiont supported cospeciation with the host while the patterns for the secondary endosymbionts, and especially Wolbachia showed evidence of host shifts and extinctions through horizontal transmission rather than cospeciation. Of particular note is the observation of several very recent host shift events in China between exotic invader and indigenous members of the complex. These shifts were from indigenous members of the complex to the invader as well as from the invader to indigenous relatives.

BACKGROUND: The whitefly Bemisia tabaci is cryptic species complex composed of numerous species. Individual species from the complex harbor a diversity of bacterial endosymbionts including Wolbachia. However, while Wolbachia is known to have a number of different roles, its role in B. tabaci is unclear. Here, the antibiotic rifampicin is used to selectively eliminate Wolbachia from B. tabaci so as to enable its roles in whitefly development and reproduction to be explored. The indirect effects of Wolbachia elimination on the biology of Encarsia bimaculata, a dominant parasitoid of B. tabaci in South China, were also investigated. METHODOLOGY/PRINCIPAL FINDING: qRT-PCR and FISH were used to show that after 48 h exposure to 1.0 mg/ml rifampicin, Wolbachia was completely inactivated from B. tabaci Mediterranean (MED) without any significant impact on either the primary symbiont, Portiera aleyrodidarum or any of the other secondary endosymbionts present. For B. tabaci MED, Wolbachia was shown to be associated with decreased juvenile development time, increased likelihood that nymphs completed development, increased adult life span and increased percentage of female progeny. Inactivation was associated with a significant decrease in the body size of the 4th instar which leads us to speculate as to whether Wolbachia may have a nutrient supplementation role. The reduction in nymph body size has consequences for its parasitoid, E. bimaculata. The elimination of Wolbachia lead to a marked increase in the proportion of parasitoid eggs that completed their development, but the reduced size of the whitefly host was also associated with a significant reduction in the size of the emerging parasitoid adult and this was in turn associated with a marked reduction in adult parasitoid longevity. CONCLUSIONS/SIGNIFICANCE: Wolbachia increases the fitness of the whitefly host and provides some protection against parasitization. These observations add to our understanding of the roles played by bacterial endosymbionts.

The whitefly Bemisia tabaci is a cosmopolitan insect species complex that harbors the obligate primary symbiont Portiera aleyrodidarum and several facultative secondary symbionts including Wolbachia, which have diverse influences on the host biology. Here, for the first time, we revealed two different localization patterns of Wolbachia present in the immature and adult stages of B. tabaci AsiaII7 cryptic species. In the confined pattern, Wolbachia was restricted to the bacteriocytes, while in the scattered pattern Wolbachia localized in the bacteriocytes, haemolymph and other organs simultaneously. Our results further indicated that, the proportion of B. tabaci AsiaII7 individuals with scattered Wolbachia were significantly lower than that of confined Wolbachia, and the distribution patterns of Wolbachia were not associated with the developmental stage or sex of whitefly host. This study will provide a new insight into the various transmission routes of Wolbachia in different whitefly species.

The ladybird Propylaea japonica (Thunberg) is one of most important natural enemies of aphids in China. This species is threatened by the extensive use of insecticides but genomics-based information on the molecular mechanisms underlying insecticide resistance is limited. Hence, we analyzed the transcriptome and expression profile data of P. japonica in order to gain a deeper understanding of insecticide resistance in ladybirds. We performed de novo assembly of a transcriptome using Illumina's Solexa sequencing technology and short reads. A total of 27,243,552 reads were generated. These were assembled into 81,458 contigs and 33,647 unigenes (6,862 clusters and 26,785 singletons). Of the unigenes, 23,965 (71.22%) have putative homologues in the non-redundant (nr) protein database from NCBI, using BLASTX, with a cut-off E-value of 10(-5). We examined COG, GO and KEGG annotations to better understand the functions of these unigenes. Digital gene expression (DGE) libraries showed differences in gene expression profiles between two insecticide resistant strains. When compared with an insecticide susceptible profile, a total of 4,692 genes were significantly up- or down- regulated in a moderately resistant strain. Among these genes, 125 putative insecticide resistance genes were identified. To confirm the DGE results, 16 selected genes were validated using quantitative real time PCR (qRT-PCR). This study is the first to report genetic information on P. japonica and has greatly enriched the sequence data for ladybirds. The large number of gene sequences produced from the transcriptome and DGE sequencing will greatly improve our understanding of this important insect, at the molecular level, and could contribute to the in-depth research into insecticide resistance mechanisms.

The sweetpotato whitefly, Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) is a widely distributed and destructive agricultural pest on various host plants. The biology of two biotypes of B. tabaci : the invasive B and a new defined Cv biotype, on a range of host plants (hibiscus, laurel, poinsettia, collard, cucumber and tomato) were studied in the laboratory. Results revealed that the developmental periods of the B biotype immatures were not significantly different on the tested host plants except those between laurel and collard. The Cv biotype immatures developed significantly slower on cucumber and tomato than on the other plants. B. tabaci B biotype had the highest survivorship on collard (68.55%), and the lowest on laurel (33.24%), while the Cv biotype had the highest and lowest survivorships on laurel (61.63%) and tomato (36.74%). Host plants did not significantly affect the pre-ovipostion period regardless of biotype. The longest averaged longevity and highest fecundity of B biotype were both recorded on collard: 25.15 days and 143.0 eggs. The highest fecundity of Cv biotype was 196.49 eggs on laurel and its longest longevity was on hibiscus (19.62 days). The intrinsic rate of natural increase ( r m ) of B biotype on the three vegetables were all higher than those on the three ornamentals whereas the r m of Cv biotype on the three ornamentals were all higher that those on the three vegetables. Our research indicates that B. tabaci B and Cv biotypes have different host plant suitabilities. The three tested vegetables were more suitable for B biotype while the three tested ornamental plants were more suitable for Cv biotype. The potential mechanism for the different suitability of B and Cv biotypes on various host plants is also discussed.

Aim: Flt4 plays a key role in promoting tumor metastasis by stimulating solid tumor lymphangiogenesis. In this study, mouse Flt4 (mFlt4) was displayed on T4 phage in order to explore the feasibility of breaking immune tolerance to “self-antigens” and to evaluate the phage's antitumor activity. Methods: A T4 phage nanometer particle expressing mFlt4 on the surface was constructed for evaluation as a recombinant vaccine. The presence of the mFlt4 gene in the T4-mFlt4 recombinant vaccine was verified by PCR and Western blot analysis. The immunotherapeutic potential of T4-mFlt4 was tested in mice injected with Lewis lung carcinoma (LLC) cells. Anti-Flt4 antibody producing B cells were detected by ELISPOT. The effects of T4-mFlt4 on lymphatic metastasis and lymphangiogenesis were investigated in a mouse antimetastasis assay and by Flt4 and CD105 immunohistochemistry. Results: The T4-mFlt4 recombinant vaccine demonstrated antitumor activity and elicited autoantibodies against mFlt4. Mice carrying LLC-derived tumors exhibited prolonged survival when given the vaccine compared with control-treated animals. The vaccine also inhibited lymphangiogenesis and tumor metastasis in the mouse models. However, T4-mFlt4 was not observed to inhibit tumor growth. Conclusion: The T4-mFlt4 recombinant vaccine induced protective antitumor immunity and antimetastasis against LLC. Induction of an autoimmune response directed against tumor progression merits further study as a new strategy for immunotherapy in cancer.

Aphis craccivora Koch is one of the most important pests of legumes, and Rhopalosiphum maidis (Fitch), is a serious pest of Gramineous crops. Susceptible strains of these two species were obtained and the baseline toxicity of 15 commonly used insecticides was established by a standard glass tube residual film method. The results showed that LC 50 values of 15 tested insecticides against A. craccivora and R. maidis varied from 0.79 to 52.23 mg/L and 1.03 to 39.20 mg/L, respectively. Chlorpyrifos was proved to be the most toxic insecticide against both the species whereas Abamectin was least toxic. Susceptible toxicity baselines data of both A. craccivora and R. maidis to 15 insecticides established in this study could be used as a reference for resistance monitoring or other related researches.

Bemisia tabaci is a species complex, of which the B and Q biotypes are globally distributed. B. tabaci feeds on more than 600 plant species including cotton, which is one of the main crops in Pakistan. In this study, the biotypes/haplotypes of B. tabaci populations collected from cotton plants in Pakistan, China, and Egypt were identified, and their phylogenetic relationships were investigated. None of the populations from Pakistan or Egypt belonged to the B or Q biotype: all 16 samples from Pakistan belonged to haplotype “PCG-1” and it is a group of unresolved populations all from Pakistan, while all three populations from Egypt belonged to unresolved haplotype “ECG.” The three populations from China belonged to the B biotype. Phylogenetic analysis indicated that the PCG-1, ECG, and B biotypes clustered into different clades even though they share the same cotton plant species as their host. The association between the outbreaks of cotton leaf curl disease and the occurrence of the PCG haplotype of B. tabaci in Pakistan is discussed.

The entomopathogenic fungus Metarhizium anisopliae is currently used as an efficient biological control agent against different insects. The prevalence and genetic variability of the entomopathogenic fungus Metarhizium anisopliae var. anisopliae in Asian countries (China, Laos, Singapore and South Korea) and a European country (The Netherlands) was examined. The fungus was found to be widespread in agricultural and forest soils throughout China especially in the south and south western regions, with a maximum recovery percentage of 81.6, while in Laos it was found to be abundant in the forest soils only, the soil of Netherlands also seemed to be an excellent source of entomopathogens with a significant recovery of insect pathogenic fungi. Simple sequence repeats (SSR) and ITS-rDNA sequence data showed that the isolates are closely linked to each other. The gene diversity (He) and polymorphic information content values of sixty two isolates showed mean values of 0.37 and 0.63. The majority of the isolates belonged to one of the closely related genotypes and these were found to be dominant in the agricultural as well as forest ecosystem. Genetic distances among the isolates according to locations and different sources showed minimal variation ranging from 0.23 to 0.34%, with maximum genetic distance of 0.34% among the isolates from Laos and The Netherlands. The reason for the limited variation is uncertain, however even the similarity index among the isolates can endow with sufficient knowledge for the implications of the methods to develop this fungus as a microbial control agent.