Characterisation and relative abundance of bacteria present in the urine of a frog

# Urinary bacteria ID and relative abundance [https://doi.org/10.5061/dryad.vt4b8gv0b](https://doi.org/10.5061/dryad.vt4b8gv0b) Bacterial species composition and relative abundance present in urine samples collected from individual male Common eastern froglets, *Crinia signifera* (n=14). All frogs were collected from a single, large population, located adjacent to the Ecological Research Centre (ERC) at the University of Wollongong (34.4048° S, 150.8717° E). Collections occurred on October 19 and 26, 2022. Each row represents a species of bacteria identified to the finest scale that was able to be identified according to the SILVA rRNA database by the Australian Genome Research Facility. Unassigned species are species that were not able to be identified to taxonomic order. Total absolute bacterial abundance is the sum of all absolute abundance raw reads counts for each species within each urine sample; data for each species is the relative abundance of each species within the sample, displayed as a percentage (total number of reads assigned to a species divided by the total number of reads in the sample * 100). ## Data sets included 1. Relative abundance of bacteria in the urine of each male individual 1. Phylum level relative abundance 2. Species level relative abundance 2. Species identification data and ID confidence ## Description of the data and file structure To identify the composition and abundance of bacterial species within urine, urine samples were collected from 14 individual male frogs within 4-5 days of collection from the field. Urine was collected from 14 male frogs according to protocols previously developed (Silla & Roberts 2012), whereby a fire-polished microcapillary tube was inserted into the cloaca to stimulate urination. Each urine sample was absorbed onto a sterile dry rayon-tip swabs (CLASSIQ swabs 167KS01, Copan Diagnostics, Inc., USA), placed in a 1.5 mL Eppendorf tube, and stored in a -80 °C freezer until sample processing (microbiome extraction and profiling). Frozen samples were stored for 23 weeks before being transported on dry ice to the Australian Genome Research Facility in Urrbrae, South Australia for genome sequencing and species identification. DNA extraction was then performed using the DNeasy® 96 PowerSoil® Pro Kit (QIAGEN) as per the manufacturer’s instructions, for manual high-throughput isolation of microbial genomic DNA. PCR amplification was then performed, and PCR amplicons were generated using the primers and conditions; the first stage PCR was cleaned using magnetic beads, and samples were visualised on 2% Sybr Egel (Thermo-Fisher). A secondary PCR to index the amplicons was performed with Platinum SuperFi II mastermix (Life Technologies, Australia). The resulting amplicons were cleaned again using magnetic beads, quantified by fluorometry (Promega Quantifluor), and normalised. The eqimolar pool was cleaned a final time using magnetic beads to concentrate the pool and then measured using a High-Sensitivity D1000 Tape on an Agilent 2200 TapeStation. The pool was diluted to 5nM and molarity was confirmed again using a Qubit High Sensitivity dsDNA assay (ThermoFisher). This was followed by sequencing on an Illumina MiSeq (San Diego, CA, USA) with a V3, 600 cycle kit (2 x 300 base pairs paired-end). Output generated included absolute abundance and relative abundance of bacterial species in each sample, identified using the SILVA rRNA database. ## Data sharing/Access This data set is to be used with permission from the corresponding author: Aimee Silla [[e-mail]](mailto:asilla@uow.e) ## Funding This study was funded by a grant from the Australian Research Council, Discovery Early Career Researcher Award (DE210100812) awarded to A.Silla

# 尿液细菌鉴定与相对丰度 [https://doi.org/10.5061/dryad.vt4b8gv0b]
本数据集包含从14只雄性东部普通蛙（*Crinia signifera*）的尿液样本中鉴定得到的细菌物种组成与相对丰度信息。所有青蛙采集自澳大利亚卧龙岗大学生态研究中心（ERC）周边的单一大型种群，采样坐标为南纬34.4048°、东经150.8717°。样本采集时间为2022年10月19日与26日。
每一行代表依据澳大利亚基因组研究中心采用的SILVA核糖体RNA（rRNA）数据库可鉴定至最精细分类阶元的细菌物种。未分类物种指无法鉴定至目级分类单元的物种。总绝对细菌丰度为每份尿液样本中各物种的绝对丰度原始读长计数之和；单物种数据为其在样本中的相对丰度，以百分比形式呈现（分配至某一物种的读长总数除以样本总读长数再乘以100）。

本数据集包含以下内容：
1. 各雄性个体尿液中的细菌相对丰度
1. 门水平相对丰度
2. 物种水平相对丰度
2. 物种鉴定数据与鉴定置信度

## 数据与文件结构说明
为鉴定尿液内细菌物种的组成与丰度，研究人员在野外采样后的4-5天内，从14只雄性青蛙体内收集尿液样本。尿液采集遵循此前开发的实验方案（Silla & Roberts 2012）：将经火焰抛光的毛细管插入泄殖腔以刺激排尿。每份尿液样本被吸附至无菌干燥的粘胶纤维头拭子（CLASSIQ swabs 167KS01，美国Copan Diagnostics公司），随后置于1.5 mL Eppendorf离心管中，于-80℃冰箱保存直至样本处理（微生物组提取与谱型分析）。冰冻样本在储存23周后，通过干冰运输至南澳大利亚乌鲁布雷的澳大利亚基因组研究中心进行基因组测序与物种鉴定。
DNA提取严格按照制造商说明书，使用DNeasy® 96 PowerSoil® Pro试剂盒（QIAGEN）完成，以实现微生物基因组DNA的手动高通量分离。随后进行PCR扩增，使用指定引物与反应条件生成PCR扩增子；第一轮PCR产物经磁珠纯化后，通过2% Sybr Egel凝胶（Thermo-Fisher）进行电泳可视化。第二轮PCR用于为扩增子添加索引，使用Platinum SuperFi II预混液（澳大利亚Life Technologies公司）。所得扩增子再次经磁珠纯化，通过荧光定量法（Promega Quantifluor）进行定量并均一化。等摩尔混合的扩增子文库最终经磁珠纯化以浓缩文库，随后使用安捷伦2200 TapeStation的High-Sensitivity D1000 Tape进行质检。文库被稀释至5nM，再次通过Qubit高灵敏度双链DNA定量试剂盒（ThermoFisher）确认摩尔浓度。随后使用Illumina MiSeq测序仪（美国加利福尼亚州圣地亚哥）搭配V3 600循环试剂盒（2×300 bp双端测序）进行测序。
测序产出数据包含各样本中细菌物种的绝对丰度与相对丰度，均通过SILVA rRNA数据库完成鉴定。

## 数据共享与获取
本数据集需经通讯作者Aimee Silla许可后方可使用（邮箱：[e-mail]）。

## 资助信息
本研究由澳大利亚研究委员会的发现早期职业研究员奖（DE210100812）资助给A.Silla。