Isolation of Previously Uncultured Slow-Growing Bacteria by Using a Simple Modification in the Preparation of Agar Media

Applied and Environmental Microbiology
Souichiro KatoYoichi Kamagata

Abstract

Most microorganisms living in the environment have yet to be cultured, owing at least in part to their slow and poor propagation properties and susceptibility to oxidative stress. Our previous studies demonstrated that a simple modification in the preparation of agar media, i.e., autoclaving the phosphate and agar separately (termed "PS" medium), can greatly improve the culturability of microorganisms by mitigating oxidative stress compared with the use of "PT" medium (autoclaving the phosphate and agar together). Here, we attempted to isolate phylogenetically novel bacteria by combining PS medium with prolonged cultivation. After inoculation with forest soil or pond sediment samples, significantly more colonies appeared on PS medium than on PT medium. A total of 98 and 74 colonies that emerged after more than 7 days of cultivation were isolated as slow growers from PS and PT media, respectively. Sequencing analysis of their 16S rRNA genes revealed that the slow growers recovered from PS medium included more phylogenetically novel bacteria than those from PT medium, including a strain that could be classified into a novel order in the class Alphaproteobacteria Further physiological analysis of representative strains showed that...Continue Reading

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Citations

Jan 9, 2019·Ecology and Evolution·Anna M AlessiJames P J Chong
Aug 8, 2019·The ISME Journal·Andrew D SteenJ Cameron Thrash
Dec 18, 2020·Frontiers in Cellular and Infection Microbiology·Sabrina NaudDidier Raoult
Jul 23, 2021·Microbiology Resource Announcements·Motoyuki WatanabeWataru Kitagawa

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