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Molecular Characterization of Culturable Aerobic Bacteria in the Midgut of Field-Caught Culex tritaeniorhynchus, Culex gelidus, and Mansonia annulifera Mosquitoes in the Gampaha District of Sri Lanka.

ABSTRACT: Background:Larval and adult mosquito stages harbor different extracellular microbes exhibiting various functions in their digestive tract including host-parasite interactions. Midgut symbiotic bacteria can be genetically exploited to express molecules within the vectors, altering vector competency and potential for disease transmission. Therefore, identification of mosquito gut inhabiting microbiota is of ample importance before developing novel vector control strategies that involve modification of vectors. Method:Adult mosquitoes of Culex tritaeniorhynchus, Culex gelidus, and Mansonia annulifera were collected from selected Medical Officer of Health (MOH) areas in the Gampaha district of Sri Lanka. Midgut lysates of the field-caught non-blood-fed female mosquitoes were cultured in Plate Count Agar medium, and Prokaryotic 16S ribosomal RNA partial genes of the isolated bacteria colonies were amplified followed by DNA sequencing. Diversity indices were used to assess the diversity and richness of the bacterial isolates in three mosquito species. The distribution pattern of bacterial isolates between different mosquito species was assessed by Distance-Based Redundancy Analysis (dbRDA). Results:A total of 20 bacterial species (Staphylococcus pasteuri, Bacillus megaterium, Staphylococcus cohnii, Pantoea dispersa, Staphylococcus chromogenes, Bacillus aquimaris, Staphylococcus arlettae, Staphylococcus sciuri, Staphylococcus warneri, Moraxella osloensis, Enterobacter sp., Klebsiella michiganensis, Staphylococcus hominis, Staphylococcus saprophyticus, Streptomyces sp., Bacillus niacin, Cedecea neteri, Micrococcus luteus, Lysinibacillus sphaericus, and Bacillus licheniformis) were identified. All of these species belonged to three phyla, Proteobacteria, Firmicutes, and Actinobacteria, out of which phylum Firmicutes (71.1%) was the most prominent. The least number of species was recorded from Actinobacteria. The relative distribution of midgut microbes in different mosquito species differed significantly among mosquito species (Chi-square, ? 2 = 486.091; df = 36; P ? 0.001). Midgut microbiota of Cx. tritaeniorhynchus and Cx. gelidus indicated a similarity of 21.51%, while Ma. annulifera shared a similarity of 6.92% with the cluster of above two species. The gut microbiota of Cx. tritaeniorhynchus was also significantly more diverse and more evenly distributed compared to Ma. annulifera. Simpson's diversity, Margalef's diversity, and Menhinick's diversity indices were higher in Cx. gelidus. Of the recorded species, P. dispersa and strains of nonpathogenic species in Bacillaceae family (B. megaterium, B. niacini, B. licheniformis, and L. sphaericus) can be recommended as potential candidates for paratransgenesis. Conclusion:The relative distribution of midgut microbes in different mosquito species differed significantly among the three studied adult mosquito species. The present data strongly encourage further investigations to explore the potential usage of these microbes through paratransgenic approach for novel eco-friendly vector control strategies.

SUBMITTER: Gunathilaka N

PROVIDER: S-EPMC7556092 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Publications

Molecular Characterization of Culturable Aerobic Bacteria in the Midgut of Field-Caught <i>Culex tritaeniorhynchus</i>, <i>Culex gelidus</i>, and <i>Mansonia annulifera</i> Mosquitoes in the Gampaha District of Sri Lanka.

Gunathilaka Nayana N Ranasinghe Koshila K Amarasinghe Deepika D Rodrigo Wasana W Mallawarachchi Harendra H Chandrasena Nilmini N

BioMed research international 20201001

<h4>Background</h4>Larval and adult mosquito stages harbor different extracellular microbes exhibiting various functions in their digestive tract including host-parasite interactions. Midgut symbiotic bacteria can be genetically exploited to express molecules within the vectors, altering vector competency and potential for disease transmission. Therefore, identification of mosquito gut inhabiting microbiota is of ample importance before developing novel vector control strategies that involve mod ...[more]

PMID: 33083488

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Project description:BackgroundThe midgut microbiota of mosquitoes maintain basal immune activity and immune priming. In recent years, scientists have focused on the use of microbial communities for vector control interventions. In the present study, the midgut bacteria of larvae and adults of Aedes aegypti and Ae. albopictus were assessed using both field-collected and laboratory-reared mosquitoes from Sri Lanka.MethodsAdults and larvae of Ae. aegypti and Ae. albopictus were collected from three selected areas in Gampaha Medical Officer of Health area, Gampaha District, Western Province, Sri Lanka. Bacterial colonies isolated from mosquito midgut dissections were identified by PCR amplification and sequencing of partial 16S rRNA gene fragments.ResultsAdults and larvae of Ae. aegypti and Ae. albopictus harbored 25 bacterial species. Bacillus endophyticus and Pantoea dispersa were found more frequently in field-collected Ae. aegypti and Ae. albopictus adults, respectively. The midgut bacteria of Ae. aegypti and Ae. albopictus adults (X2 = 556.167, df = 72, P < 0.001) and larvae (X2 = 633.11, df = 66, P < 0.001) were significantly different. There was a significant difference among the bacterial communities between field-collected adults (X2 = 48.974, df = 10, P < 0.001) and larvae (X2 = 84.981, df = 10, P < 0.001). Lysinibacillus sphaericus was a common species in adults and larvae of laboratory-reared Ae. aegypti. Only P. dispersa occurred in the field-collected adults of Ae. aegypti and Ae. albopictus. Species belonging to genera Terribacillus, Lysinibacillus, Agromyces and Kocuria were recorded from Aedes mosquitoes, in accordance with previously reported results.ConclusionsThis study generated a comprehensive database on the culturable bacterial community found in the midgut of field-collected (Ae. aegypti and Ae. albopictus) and laboratory-reared (Ae. aegypti) mosquito larvae and adults from Sri Lanka. Data confirm that the midgut bacterial diversity in the studied mosquitoes varies according to species, developmental stage and strain (field vs laboratory).

Project description:Dengue is the most important mosquito-borne viral infection in Sri Lanka causing an enormous social and economic burden in the country. In the absence of therapeutic drugs and the developed vaccines are under investigation, vector control is the best strategy to reduce the disease transmission. Therefore, the development of novel tools to control dengue vector mosquitoes has become the need of the hour. Novaluron is a recently developed Insect Growth Regulator (IGR) which inhibits chitin synthesis in immature stages of insects. The aim of the study was to identify the efficacy of a simple and cost-effective Autocidal Gravid Ovitrap (AGO) developed using Novaluron to control dengue outbreaks in the District of Gampaha, Sri Lanka. Laboratory and semifield experiments were performed to identify the activity range, optimum field dosage, and residual effects of Novaluron following the World Health Organization guidelines, and field experiments were performed in the Ragama Medical Officer of Health (MOH) area. Two study areas 800?m apart were selected and assigned as treated and control areas randomly. In each study area, 30 households were selected randomly. Each household was given two ovitraps, one placed indoors and the other placed outdoors. Mortality and survival counts were recorded separately for one-year time period and data were analyzed using a two-way repeated measures analysis of variance model. During the laboratory experiments, the adult emerging inhibition was 100% in all tested concentrations. The optimum field dosage was 2?ppm and the residual effect was 28 days. In the field experiments, significantly higher mortality counts were recorded in treated areas both indoor- and outdoor-placed AGOs. Two-factor repeated measures ANOVA followed by Tukey's test confirmed that the mean mortality count is high for the developed AGOs both indoor and outdoor settings. The developed AGO can be deployed to control both indoor and outdoor dengue vector mosquito populations, and in dengue-risk areas, the ovitrap will be supportive to local health authorities to enhance the efficiency of future vector control programs.

Project description:Some arbovirus infections, especially dengue, have increased rapidly over the last few decades in Sri Lanka. Prevalence and distribution of different mosquito species have been limitedly documented, which remains grossly inadequate in providing evidence for potential health risks. In this study, the diversity and species composition of mosquitoes in four selected districts in Sri Lanka (Kurunegala, Gampaha, Kegalle, and Kandy) were investigated. Entomological surveys were conducted from a total of 160 temporary and permanent mosquito breeding habitats identified in the study area from June 2017 to October 2018. Mosquito immature stages were sampled using standard dipping, siphoning, or pipetting methods and identified up to the species level. Percentage relative abundance and habitat characteristics such as species richness, dominance, and Shannon-Weiner diversity were calculated for each surveyed habitat type. Associations between co-occurring species were estimated by Hulbert's coefficient of interspecific association (C8). A total of 4663 mosquito larvae belonging to seven genera and fifteen species of mosquitoes were collected. The relative distribution of mosquito species differed significantly among the four studied districts (X 2 = 143.248; df = 33; P < 0.001). According to Kruskal-Wallis statistics (P < 0.05 at 95% of significance), all diversity indices for immature stages of medically important mosquitoes varied significantly across different breeding sites. Paddy fields had the significantly highest species richness of 4.0 ± 2.82. The coefficients of interspecific association among all the recorded medically important vector mosquitoes were found negative during the present study. The findings of the current study would be useful to identify the entomological potential for disease transmission and facilitate the implementation of appropriate vector control interventions. This would ultimately provide an avenue to improve the personal skills of health staff rather than limiting their knowledge to specified disease vectors, under which the control program is concerned.

Project description:BackgroundOrthohantaviruses and leptospira are emerging zoonotic pathogens of high public health significance. The epidemiology of orthohantavirus infections and leptospirosis is similar and presents related clinical pictures in humans. However, a paucity of data on actual reservoir hosts for orthohantaviruses and leptospira exists. Therefore, this study aimed at determining the occurrence of orthohantaviruses and leptospira in small mammals captured in an endemic region of Sri Lanka.MethodsRodents and shrews were morphologically and/or genetically identified using morphological keys and DNA barcoding techniques targeting the cytochrome oxidase b subunit gene (Cytb). Lung tissues and sera were subsequently analyzed for the presence of orthohantavirus RNA using qRT-PCR. Sera of rats were tested for IgG antibodies against orthohantaviruses and leptospira.ResultsForty-three (43) small mammals representing: Rattus (R.) rattus (black rat) or R. tanezumi (Asian rat), Suncus murinus (Asian house shrew), R. norvegicus (brown rat) and Mus musculus (house mouse) were investigated. No orthohantavirus RNA was detected from the lung tissue or serum samples of these animals. Elevated levels of IgG antibodies against Puumala orthohantavirus (PUUV) and/or Seoul orthohantavirus (SEOV) antigens were detected in sera of 28 (72%) out of the 39 rats analysed. Interestingly, 36 (92%) of the 39 rats also showed presence of anti leptospira-IgG antibodies in their serum, representing dual infection or dual exposure in 26/39 (66.7%) of examined rats.ConclusionsThis project targets important public health questions concerning the occupational risk of orthohantavirus infections and/or leptospirosis in an endemic region of Sri Lanka. Most rats (72%) in our study displayed antibodies reacting to orthohantavirus NP antigens, related to PUUV and/or SEOV. No correlation between the orthohantavirus and leptospira IgG antibody levels were noticed. Finally, a combination of both morphological and DNA barcoding approaches revealed that several species of rats may play a role in the maintenance and transmission of orthohantavirus and leptospira in Sri Lanka.

Dataset Information

Molecular Characterization of Culturable Aerobic Bacteria in the Midgut of Field-Caught Culex tritaeniorhynchus, Culex gelidus, and Mansonia annulifera Mosquitoes in the Gampaha District of Sri Lanka.

Publications

Molecular Characterization of Culturable Aerobic Bacteria in the Midgut of Field-Caught <i>Culex tritaeniorhynchus</i>, <i>Culex gelidus</i>, and <i>Mansonia annulifera</i> Mosquitoes in the Gampaha District of Sri Lanka.

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