Fundação Oswaldo Cruz, FIOCRUZ RONDÔNIA, Porto Velho, RO, Brazil.
Instituto Federal de Rondônia - IFRO, Porto Velho, RO, Brazil.
J Med Entomol. 2021 Jul 16;58(4):1900-1907. doi: 10.1093/jme/tjab028.
Microorganisms living in the midgut of Anopheles mosquitoes have been studied to fight vector-borne diseases, such as malaria. Studies on the microbiota of the Neotropical Anopheles darlingi, the most important Brazilian vector for malaria, have been reported for the same purpose. Our aims were to isolate and identify culturable bacteria from An. darlingi mosquito guts through their feces and to estimate the species richness and the frequency distribution of the sampled bacteria. Sixty wild females of An. darlingi mosquitoes were captured at two rural locations, near Porto Velho, Rondônia, Brazil. Bacteria were isolated from mosquito feces, which were collected using cages which permit the collection of feces on LB nutrient agar plates. Sixty bacterial colonies were isolated and stored in glycerol at -80°C. Bacteria were identified by sequencing their 16S rRNA gene obtained using PCR and Sanger sequencing. To aid in species identification, MALDI-TOF, VITEK2, and BBL Crystal were used as complementary protocols. The sequences obtained from the 60 bacterial isolates were compared to sequences deposited in GenBank (NCBI) using BLAST. Homology greater than 97% between the query and the subject was used as the criteria for assigning the identity of each isolate. Fourteen species from eight different genera were identified among the 60 isolates. The most frequent species were Serratia liquefaciens (20%) and Serratia marcescens (15%). Due to their established apathogenicity and according to previous studies, we suggest Serratia and Pantoea species as suitable for paratransgenesis development to fight malaria in Brazilian Amazon.
已对生活在疟疾病媒按蚊中肠内的微生物进行研究，以对抗蚊媒传染病，如疟疾。为了同样的目的，也有研究报告了对新热带按蚊属（Anopheles darlingi）肠道微生物群的研究，该蚊种是巴西最重要的疟疾传播媒介。我们的目的是通过粪便从按蚊属（Anopheles darlingi）蚊的肠道中分离和鉴定可培养细菌，并估计所采样细菌的物种丰富度和频率分布。在巴西朗多尼亚州波多韦柳市附近的两个农村地区，共捕获了 60 只野生雌性按蚊属（An. darlingi）蚊子。使用允许在 LB 营养琼脂板上收集粪便的笼子收集蚊子的粪便，以从蚊子粪便中分离细菌。从 60 个细菌菌落中分离出来，并储存在-80°C 的甘油中。通过使用 PCR 和 Sanger 测序获得 16S rRNA 基因来对细菌进行测序，从而对其进行鉴定。为了帮助进行物种鉴定，使用 MALDI-TOF、VITEK2 和 BBL Crystal 作为补充方案。使用 BLAST 将从 60 个细菌分离株中获得的序列与 GenBank（NCBI）中保存的序列进行比较。查询与主题之间的同源性大于 97%，用作分配每个分离株身份的标准。在 60 个分离株中鉴定出来自八个不同属的 14 个种。最常见的物种是液化沙雷氏菌（20%）和粘质沙雷氏菌（15%）。由于它们已被证实无致病性，并且根据先前的研究，我们建议使用沙雷氏菌属和泛菌属的物种来开发共生转化，以在巴西亚马逊地区对抗疟疾。
