Symbiotic bacteria affect competence for pathogen transmission in insect vectors, including mosquitoes. However, knowledge on mosquito-microbiome-pathogen interactions remains limited, largely due to methodological reasons. The current, cost-effective practice of sample pooling used in mosquito surveillance and epidemiology prevents correlation of individual traits (i.e., microbiome profile) and infection status. Moreover, many mosquito studies employ laboratory-reared colonies that do not necessarily reflect the natural microbiome composition and variation in wild populations. As a consequence, epidemiological and microbiome studies in mosquitoes are to some extent uncoupled, and the interactions among pathogens, microbiomes, and natural mosquito populations remain poorly understood. This study focuses on the effect the pooling practice poses on mosquito microbiome profiles, and tests different approaches to find an optimized low-cost methodology for extensive sampling while allowing for accurate, individual-level microbiome studies. We tested the effect of pooling by comparing wild-caught, individually processed mosquitoes with pooled samples. With individual mosquitoes, we also tested two methodological aspects that directly affect the cost and feasibility of broad-scale molecular studies: sample preservation and tissue dissection. Pooling affected both alpha- and beta-diversity measures of the microbiome, highlighting the importance of using individual samples when possible. Both RNA and DNA yields were higher when using inexpensive reagents such as NAP (nucleic acid preservation) buffer or absolute ethanol, without freezing for short-term storage. Microbiome alpha- and beta-diversity did not show overall significant differences between the tested treatments compared to the controls (freshly extracted samples or dissected guts). However, the use of standardized protocols is highly recommended to avoid methodological bias in the data.
- MeSH
- Aedes mikrobiologie MeSH
- Bacteria klasifikace genetika MeSH
- Culex mikrobiologie MeSH
- komáří přenašeči mikrobiologie MeSH
- mikrobiologické techniky MeSH
- mikrobiota * MeSH
- ochrana biologická MeSH
- střevní mikroflóra MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- antibakteriální látky normy terapeutické užití MeSH
- Bordetella pertussis izolace a purifikace účinky léků MeSH
- chemické látky - účinky a užití MeSH
- Culex klasifikace mikrobiologie růst a vývoj MeSH
- dezinfekce * trendy zákonodárství a právo MeSH
- dezinficiencia klasifikace terapeutické užití MeSH
- dezinsekce metody MeSH
- klíště MeSH
- lidé MeSH
- moskyti - kontrola metody statistika a číselné údaje MeSH
- repelenty proti hmyzu klasifikace terapeutické užití MeSH
- štěnice MeSH
- sterilizace * přístrojové vybavení trendy zákonodárství a právo MeSH
- virus Ebola MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
Growth temperature range, resistance to selective antibiotics, activities of 23 enzymes, protein fingerprints and fatty acids composition of the spirochaetal strain BR91, isolated from the Culex pipiens mosquito, were tested. The spirochaetes were grown in BSK-H Complete liquid medium. The optimal in vitro growth temperature of the strain was 33 °C. Strain BR91 was sensitive to trimethoprim, nalidixic acid, 5-fluorouracil, and tolerated phosphomycin. The strain produced acid and alkaline phosphatase, esterase (C4), esterase-lipase (C8), leucine arylamidase, naphthol-AS-BI-phosphohydrolase and α-fucosidase. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) assay revealed several major proteins in the size range of 13-16 kDa, 22-30 kDa and 37-131 kDa. Fatty acid methyl ester (FAME) analysis showed that C₁₄:₀, C₁₆:₀, C₁₈:₁ ω9c and summed feature 5 (C₁₈:₂ ω6,9c and/or C₁₈:₀ anteiso) are major fatty acids. This study highlights certain phenotypic differences between strain BR91 and the Lyme disease spirochaete Borrelia burgdorferi, and supports the hypothesis that strain BR91 represents a unique taxonomical entity in a system of spirochaetal species.
- MeSH
- antibakteriální látky farmakologie MeSH
- bakteriální proteiny analýza MeSH
- Borrelia burgdorferi MeSH
- Culex mikrobiologie MeSH
- Culicidae MeSH
- enzymy analýza MeSH
- kultivační média chemie MeSH
- mastné kyseliny analýza MeSH
- mikrobiální testy citlivosti MeSH
- Spirochaetaceae chemie klasifikace izolace a purifikace fyziologie MeSH
- teplota MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
In this study, larvae (1,179 ex.) of mosquito genera Culex were examined for the presence of spirochaetes by Dark Field Microscopy (DFM) at the locality of Blansko (Czech Republic) in of 2004-2008. DFM spirochaete positive samples (25.4 %) were investigated by nested PCR; only 4 samples were positive for the presence of Borrelia burgdorferi sensu lato, which is 0.3 % of the total examined samples. We can conclude that only a low percentage of pathogenic borreliae are presented in mosquito larvae, while the spirochaete of undefined genera infect larvae in high amounts.
- MeSH
- Aedes mikrobiologie MeSH
- Borrelia burgdorferi izolace a purifikace MeSH
- časové faktory MeSH
- Culex mikrobiologie MeSH
- larva mikrobiologie MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- práce podpořená grantem MeSH
- MeSH
- Borrelia burgdorferi genetika izolace a purifikace klasifikace MeSH
- Culex mikrobiologie MeSH
- genom bakteriální MeSH
- klíšťata mikrobiologie MeSH
- lidé MeSH
- Siphonaptera mikrobiologie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- přehledy MeSH
- Geografické názvy
- Evropa MeSH
- MeSH
- Culex mikrobiologie parazitologie MeSH
- epidemiologické faktory MeSH
- savci patologie MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Geografické názvy
- Rakousko MeSH
