Dataset Information

Molecular Detection and Characterization of Wolbachia pipientis from Culex quinquefasciatus Collected from Lahore, Pakistan.

ABSTRACT: The gram-negative, pleomorphic endosymbiont Wolbachia is known to infect a large number of insects and other arthropods naturally. This bacterium modifies the host biology, mainly causing reproductive alterations including feminization, death of male, parthenogenesis, and importantly cytoplasmic incompatibility. Wolbachia-induced cytoplasmic incompatibility results in nonviable offspring and vector population suppression. In addition, this bacterium rapidly spreads and propagates within the host population. This study is the first report on Wolbachia detection and characterization from Culex quinquefasciatus collected from Lahore, Pakistan. For this purpose, mosquito adults were collected from different localities of Lahore and identified at the species level. A total of 145 pairs of ovaries were individually subjected to DNA isolation, and polymerase chain reaction amplification of three (wsp, 16S rRNA, and ftsZ) genes were investigated. In all, 128 females were found positive, representing 82.3% infection rate. The phylogenetic analysis indicated that the detected endosymbiont had 100% homology with Wolbachia pipientis wPip strain and supergroup B. The detection of the local strain of Wolbachia (wPip) will be useful in investigating its potential for the control of dengue vector (Aedes aegypti) and reducing dengue transmission in Pakistan.

SUBMITTER: Sarwar MS

PROVIDER: S-EPMC5928706 | biostudies-literature | 2018 Jan

REPOSITORIES: biostudies-literature

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Molecular Detection and Characterization of Wolbachia pipientis from Culex quinquefasciatus Collected from Lahore, Pakistan.

Sarwar Muhammad Sajjad MS Jahan Nusrat N Shahbaz Fatima F

The American journal of tropical medicine and hygiene 20180101 1

The gram-negative, pleomorphic endosymbiont Wolbachia is known to infect a large number of insects and other arthropods naturally. This bacterium modifies the host biology, mainly causing reproductive alterations including feminization, death of male, parthenogenesis, and importantly cytoplasmic incompatibility. Wolbachia-induced cytoplasmic incompatibility results in nonviable offspring and vector population suppression. In addition, this bacterium rapidly spreads and propagates w ...[more]

PMID: 29141766

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Project description:Polymerase chain reaction (PCR) is often used to detect microorganisms, pathogens, or both, including the reproductive parasite Wolbachia pipientis (Rickettsiales: Anaplasmataceae), in mosquitoes. Natural populations of Culex pipiens L. (Diptera: Culicidae) mosquitoes are infected with one or more strains of W. pipientis, and crosses between mosquitoes harboring different Wolbachia strains provide one of the best-known examples of cytoplasmic incompatibililty (CI). When we used PCR to monitor Wolbachia in the Buckeye strain of Culex pipiens, and in a Wolbachia-cured sister colony obtained by tetracycline treatment, we noted false negative PCR reactions with DNA samples from infected mosquitoes; these results were inconsistent with direct microscopic observation of Wolbachia-like particles in gonads dissected from mosquitoes in the same population. Assays with diluted template often improved detection of positive samples, suggesting that DNA prepared from whole mosquitoes contained an inhibitor of the PCR reaction. We reconciled discrepancies between PCR and microscopy by systematic measurement of the PCR reaction in the presence of an internal standard. Mosquito decapitation before DNA extraction restored the reliability of the PCR reaction, allowing accurate determination of Wolbachia infection status in infected and tetracycline-cured mosquito populations, consistent with microscopic examination. Using PCR primers based on the Tr1 gene, we confirmed that the Wolbachia infection in the Buckeye strain of Culex pipiens belongs to the genotype designated wPip1. Finally, to explore more widely the distribution of PCR inhibitors, we demonstrated that DNA isolated from the cricket, Acheta domesticus (L.); the beetle, Tenebrio molitor L.; the honey bee, Apis mellifera L.; and the mosquito, Anopheles punctipennis Say also contained PCR inhibitors. These results underscore the importance of measuring the presence of inhibitors in PCR templates by using a known positive standard, and provide an approach that will facilitate use of PCR to monitor environmental samples of mosquitoes that harbor endosymbionts or pathogenic organisms.

Project description:Following an outbreak of chikungunya virus (CHIKV) infections in Thailand in 2019, numerous cases of CHIKV infection have been diagnosed in Bangkok, the capital of the country. In our previous investigation of the vectors for disease transmission, we found natural infection of CHIKV in both male and female Aedes aegypti mosquitoes collected from the outbreak areas in Bangkok. Some reports mentioned the detection of CHIKV in Culex mosquitoes. In Thailand, the Culex quinquefasciatus Say mosquito is a common species found in urban and rural settings that coexists with Ae. aegypti. However, the role of Cx. quinquefasciatus mosquitoes in the spread of the Indian Ocean Lineage (IOL) of CHIKV in Thailand has never been investigated. In this study, Cx. quinquefasciatus were collected (16 males and 27 females) from an outbreak area in Bangkok. Eight of the 27 in field-caught female Cx. quinquefasciatus were positive for IOL CHIKV RNA, and 99-100% identity and full 100% coverage of sequences similar to CHIKV isolated from female Ae. aegypti in Bangkok, Thailand, whereas viral RNA was not detected in male samples using nested-RT-PCR. To determine whether CHIKV is able to replicate in Cx. quinquefasciatus, the laboratory strain of Cx. quinquefasciatus was allowed to feed on blood containing IOL CHIKV isolated from patient serum. The nested-RT-PCR, virus isolation, and immunofluorescence assay (IFA) were performed for CHIKV detection and replication. The results showed that CHIKV RNA was detected in Cx. quinquefasciatus until day 4 post infection. CHIKV did not produce any remarkable signs of infection, dissemination, or transmission in Cx. quinquefasciatus, and cytopathic effect (CPE) was not observed in C6/36 cells when infected with supernatant obtained from Cx. quinquefasciatus at days 7, 10, 14, and 21 post infection when compared to Ae. aegypti. The data from this study infer that CHIKV may be detected in Cx. quinquefasciatus but that the mosquito is not able to transmit CHIKV in Thailand.

Dataset Information

Molecular Detection and Characterization of Wolbachia pipientis from Culex quinquefasciatus Collected from Lahore, Pakistan.

Publications

Molecular Detection and Characterization of <i>Wolbachia pipientis</i> from <i>Culex quinquefasciatus</i> Collected from Lahore, Pakistan.

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