Abstract：
Many anuran amphibians deposit their eggs in foam nests, biostructures that help protect the eggs and tadpoles from predators. Currently, there are no other identification and description studies of the cultivable microbiota role in the nests of the Leptodactylid frogs such as Physalaemus cuvieri, Leptodactylus vastus and Adenomera hylaedactyla. This study aimed to isolate and identify the culturable bacteria from these three anuran species\' nests, as well as to prospect enzymes produced by this microbiota. Foam nests samples and environmental samples were diluted and viable cell count was determined. Bacterial morphotypes from foam nest samples were isolated through spread plate technique. Isolates\' DNAs were extracted followed by rRNA 16S gene amplification and Sanger sequencing. To evaluate their enzymatic potential, the isolates were cultured in ATGE medium supplemented with starch (0.1% w/v), gelatin (3% w/v) and skimmed milk (1% w/v), to verify amylase and protease activity. A total of 183 bacterial morphotypes were isolated, comprising 33 bacterial genera. Proteobacteria phylum was the most abundant in all the three nests (79%). The genera Pseudomonas and Aeromonas were the most abundant taxon in P. cuvieri and L. vastus. In A. Hylaedactyla, were Enterobacter and Bacillus. Regarding enzymatic activities, 130 isolates displayed protease activity and 45 isolates were positive for amylase activity. Our results provide unprecedented information concerning culturable bacterial microbiota of the foam nests of the Leptodactylid frogs, as well as their potential for biomolecules of biotechnological interest.
摘要：
许多无双两栖动物将卵存放在泡沫巢中，有助于保护卵和t免受捕食者侵害的生物结构。目前,没有其他鉴定和描述研究可培养的微生物群在Leptodactylid青蛙的巢中的作用，例如Physalaemuscuvieri，浮萍和腺毛。本研究旨在从这三种无性系巢中分离和鉴定可培养细菌，以及这种微生物群产生的酶的前景。将泡沫巢样品和环境样品稀释并测定活细胞计数。通过铺板技术从泡沫巢样品中分离出细菌形态型。提取分离株的DNA，然后进行rRNA16S基因扩增和Sanger测序。为了评估它们的酶潜力，分离物在补充有淀粉(0.1%w/v)的ATGE培养基中培养,明胶（3％w/v）和脱脂奶（1％w/v），验证淀粉酶和蛋白酶的活性。共分离出183种细菌形态,包括33个细菌属。在所有三个巢中，变形杆菌门最丰富（79％）。假单胞菌属和气单胞菌属是库维埃氏疟原虫中最丰富的分类群。在A.Hylaedactyla中，肠杆菌和芽孢杆菌。关于酶活性，130个分离株显示蛋白酶活性，45个分离株对淀粉酶活性呈阳性。我们的结果提供了有关Leptodactylid青蛙泡沫巢的可培养细菌微生物群的前所未有的信息，以及它们对生物技术感兴趣的生物分子的潜力。
