聚（乙烯醇）水凝胶包埋的红球菌属在暴露于石油烃后进行种特异性同时 PCR 检测和活力测试。

Simultaneous species-specific PCR detection and viability testing of poly(vinyl alcohol) cryogel-entrapped Rhodococcus spp. after their exposure to petroleum hydrocarbons.

机构信息

Institute of Ecology and Genetics of Microorganisms, Russian Academy of Sciences, 13 Golev Street, 614081 Perm, Russia; Microbiology and Immunology Department, Perm State National Research University, 15 Bukirev Street, 614900 Perm, Russia.

出版信息

J Microbiol Methods. 2013 Aug;94(2):135-140. doi: 10.1016/j.mimet.2013.05.016. Epub 2013 Jun 5.

DOI:10.1016/j.mimet.2013.05.016

PMID:23747656

Abstract

A method of simultaneous species-specific PCR detection and viability testing of poly(vinyl alcohol) cryogel-entrapped Rhodococcus spp. was developed that allowed the estimation of immobilized Rhodococcus opacus and Rhodococcus ruber survival after their exposure to petroleum hydrocarbon mixture. Spectrophotometric INT assay revealed high tolerance of gel-immobilized rhodococci to petroleum hydrocarbons, while among two Rhodococcus strains studied, R. ruber tolerated better to hydrocarbons compared to R. opacus. These findings were confirmed by respirometry results that showed increased respiratory activity of gel-immobilized Rhodococcus strains after 10-day incubation with 3% (v/v) petroleum hydrocarbon mixture. Moreover, jointly incubated rhodococcal strains demonstrated higher oxidative activities toward petroleum hydrocarbons than individual strains. Both Rhodococcus species were recovered successfully in cryogel granules using 16S rDNA-targeted PCR, even though the granules were previously stained with INT and extracted with ethanol. The method developed can be used for rapid detection and monitoring of gel-immobilized bacterial inocula in bioreactors or contaminated soil systems.

摘要

开发了一种同时检测聚乙烯醇(PVA)包埋罗克索菌属的种特异性 PCR 和活力的方法，该方法允许在罗克索菌属暴露于石油烃混合物后估计固定化罗克索菌属和红球菌属的存活。分光光度 INT 测定法显示凝胶固定化罗克氏菌对石油烃具有高耐受性，而在所研究的两种罗克氏菌菌株中，与罗克索菌属相比，红球菌属对烃类的耐受性更好。这些发现通过呼吸计结果得到了证实，该结果表明，在与 3%（v/v）石油烃混合物孵育 10 天后，凝胶固定化罗克氏菌的呼吸活性增加。此外，共同孵育的罗克氏菌菌株对石油烃的氧化活性高于单个菌株。即使颗粒先前用 INT 染色并用乙醇提取，16S rDNA 靶向 PCR 也能成功地从 PVA 包埋的罗克索菌属颗粒中回收。该方法可用于生物反应器或污染土壤系统中凝胶固定化细菌接种物的快速检测和监测。