Distribution and phylogeny of Wolbachia strains in wild mosquito populations in Sri Lanka

Abstract

Background: Wolbachia are a group of maternally inherited intracellular bacteria known to be widespread among arthropods. Infections with Wolbachia cause declines of host populations, and also induce host resistance to a wide range of pathogens. Over the past few decades, researchers were curious to use Wolbachia as a biological tool to control mosquito vectors. During the present study, assessment of the prevalence of Wolbachia infections among wild mosquito populations in Sri Lanka where mosquito-borne diseases are a major health concern, was carried out for the first time. DNA was extracted from the abdomens of mosquitoes, collected from seven provinces, and screened for the presence of Wolbachia by PCR using wsp and groE primers. Group-specific and strain-specific primers were used to classify Wolbachia into the supergroups A and B, and into the strains Mel, AlbA and Pip.

Results: A total of 330 individual mosquitoes belonging to 22 species and 7 genera were screened. Eighty-seven mosquitoes (26.36%) belonging to four species (i.e. Aedes albopictus, Culex quinquefasciatus, Armigeres subalbatus and Mansonia uniformis) were positive for Wolbachia infections. Primary vector of the dengue fever, Ae. aegypti was negative for Wolbachia infections while the secondary vector, Ae. albopictus, showed a very high infection rate. The filarial vector C. quinquefasciatus had a relatively high rate of infection. Japanese encephalitis vectors C. gelidus and C. triteaneorynchus, and the Anopheles vectors of malaria were negative for Wolbachia infections. Nine sequences of Wolbachia-positive PCR products were deposited in the GenBank and compared with other available data. Aedes albopictus was infected with both Wolbachia strains A (AlbA) and B (Pip) supergroups. Phylogenetic analysis of the wsp sequences showed two major branches confirming identities obtained from the PCR screening with strain-specific primers.

Conclusion: Wolbachia infections were found only among four mosquito species in Sri Lanka: Aedes albopictus, Culex quinquefasciatus, Armigeres subalbatus and Mansonia uniformis. Sequence data showed high haplotype diversity among the Wolbachia strains.

Keywords: Biological control; Mosquito control; Phylogeny; Sri Lanka; Wolbachia strains; Wsp.

Fig. 1

A map of Sri Lanka showing the provinces and the collection sites

Fig. 2

Results of the Wolbachia strain identification PCR assays for Aedes albopictus mosquitoes collected from the central province of Sri Lanka (n = 33), using different primer sets: Lane 1: Wolbachia confirmation general primers wsp81F and wsp691R (600 bp band); Lane 2: Group A primers wsp136 and wsp691R (556 bp); Lane 3: Mel strain-specific primers wsp308F and wsp691R (405 bp); Lane 4: AlbA strain-specific primers wsp328F and 691R (379 bp); Lane 5: Group B primers wsp81F and wsp522R (442 bp); Lane 6: Pip strain-specific primers wspp183F and wsp 691R (501 bp). PCR products were electrophoresed in 1.5% agarose gel, stained with ethidium bromide and visualized under UV light

Fig. 3

Neighbor-joining tree generated from aligned wsp sequences. Tree shown is midpoint rooted and bootstrap values (1,000 replicates) are labelled next to the branches. Taxa are labelled as the host names from which the Wolbachia strains were obtained. Wsp sequences from the present study are marked with black dots