Oliveira Letícia de C, Silveira Aline M M, Monteiro Andréa de S, Dos Santos Vera L, Nicoli Jacques R, Azevedo Vasco A de C, Soares Siomar de C, Dias-Souza Marcus V, Nardi Regina M D
Departamento de Biologia Geral, Instituto de Ciências Biológicas, Universidade Federal de Minas GeraisBelo Horizonte, Brazil.
Departamento de Microbiologia, Instituto de Ciências Biológicas, Universidade Federal de Minas GeraisBelo Horizonte, Brazil.
Front Microbiol. 2017 May 19;8:876. doi: 10.3389/fmicb.2017.00876. eCollection 2017.
A bacteriocinogenic L156.4 strain isolated from the feces of NIH mice was identified by 16S rRNA gene sequencing and MALDI-TOF mass spectrometry. The entire genome was sequenced using Illumina, annotated in the PGAAP, and RAST servers, and deposited. Conserved genes associated with bacteriocin synthesis were predicted using BAGEL3, leading to the identification of an open reading frame (ORF) that shows homology with the GG (ATCC 53103) prebacteriocin gene. The encoded protein contains a conserved protein motif associated a structural gene of the Enterocin A superfamily. We found ORFs related to the prebacteriocin, immunity protein, ABC transporter proteins, and regulatory genes with 100% identity to those of HN001. In this study, we provide evidence of a putative bacteriocin produced by L156.4 that was further confirmed by assays. The antibacterial activity of the substances produced by this strain was evaluated using the deferred agar-spot and spot-on-the lawn assays, and a wide antimicrobial activity spectrum against human and foodborne pathogens was observed. The physicochemical characterization of the putative bacteriocin indicated that it was sensitive to proteolytic enzymes, heat stable and maintained its antibacterial activity in a pH ranging from 3 to 9. The activity against , which was used as an indicator strain, was detected during bacterial logarithmic growth phase, and a positive correlation was confirmed between bacterial growth and production of the putative bacteriocin. After a partial purification from cell-free supernatant by salt precipitation, the putative bacteriocin migrated as a diffuse band of approximately 1.0-3.0 kDa by SDS-PAGE. Additional studies are being conducted to explore its use in the food industry for controlling bacterial growth and for probiotic applications.
从NIH小鼠粪便中分离出的产细菌素L156.4菌株,通过16S rRNA基因测序和基质辅助激光解吸电离飞行时间质谱法进行鉴定。使用Illumina对其全基因组进行测序,在PGAAP和RAST服务器中进行注释并保存。利用BAGEL3预测与细菌素合成相关的保守基因,从而鉴定出一个与GG(ATCC 53103)前细菌素基因具有同源性的开放阅读框(ORF)。编码的蛋白质包含一个与肠球菌素A超家族结构基因相关的保守蛋白质基序。我们发现与前细菌素、免疫蛋白、ABC转运蛋白和调控基因相关的ORF与HN001的ORF具有100%的同一性。在本研究中,我们提供了L156.4产生推定细菌素的证据,并通过实验进一步证实。使用延迟琼脂斑点法和琼脂平板点样法评估该菌株产生的物质的抗菌活性,观察到其对人类和食源性病原体具有广泛的抗菌活性谱。推定细菌素的理化特性表明,它对蛋白水解酶敏感,热稳定,并且在pH值为3至9的范围内保持其抗菌活性。在细菌对数生长期检测到对用作指示菌株的抗菌活性,并且证实细菌生长与推定细菌素的产生之间存在正相关。通过盐沉淀从无细胞上清液中部分纯化后,推定细菌素在SDS-PAGE上迁移为一条约1.0 - 3.0 kDa的弥散带。正在进行进一步的研究,以探索其在食品工业中用于控制细菌生长和益生菌应用的用途。