AUTHOR=Zhong Huiyue , Zheng Nan , Wang Jiaqi , Zhao Shengguo 

TITLE=Isolation and pan-genome analysis of Enterobacter hormaechei Z129, a ureolytic bacterium, from the rumen of dairy cow

JOURNAL=Frontiers in Microbiology

VOLUME=14

YEAR=2023

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2023.1169973

DOI=10.3389/fmicb.2023.1169973

ISSN=1664-302X

ABSTRACT=<sec><title>Introduction</title><p>Urea is an important non-protein nitrogen source for ruminants. In the rumen, ureolytic bacteria play critical roles in urea-nitrogen metabolism, however, a few ureolytic strains have been isolated and genomically sequenced. The purpose of this study was to isolate a novel ureolytic bacterial strain from cattle rumen and characterize its genome and function.</p></sec><sec><title>Methods</title><p>The ureolytic bacterium was isolated using an anaerobic medium with urea and phenol red as a screening indicator from the rumen fluid of dairy cattle. The genome of isolates was sequenced, assembled, annotated, and comparatively analyzed. The pan-genome analysis was performed using IPGA and the biochemical activity was also analyzed by test kits.</p></sec><sec><title>Results</title><p>A gram-positive ureolytic strain was isolated. Its genome had a length of 4.52 Mbp and predicted genes of 4223. The 16S rRNA gene and genome GTDB-Tk taxonomic annotation showed that it was a novel strain of <italic>Enterobacter hormaechei</italic>, and it was named <italic>E. hormaechei</italic> Z129. The pan-genome analysis showed that Z129 had the highest identity to <italic>E. hormaechei</italic> ATCC 49162 with a genome average nucleotide identity of 98.69% and possessed 238 unique genes. Strain Z129 was the first <italic>E. hormaechei</italic> strain isolated from the rumen as we know. The functional annotation of the Z129 genome showed genes related to urea metabolism, including urea transport (<italic>urt</italic>A<italic>-urt</italic>E), nickel ion transport (<italic>ure</italic>J, <italic>ton</italic>B, <italic>nix</italic>A, <italic>exb</italic>B, <italic>exb</italic>D, and <italic>rcn</italic>A), urease activation (<italic>ure</italic>A-ureG) and ammonia assimilation (<italic>gdh</italic>A, <italic>gln</italic>A, <italic>gln</italic>B, <italic>gln</italic>E, <italic>gln</italic>L, <italic>gls</italic>A, <italic>glt</italic>B, and <italic>glt</italic>D) were present. Genes involved in carbohydrate metabolism were also present, including starch hydrolysis (<italic>amy</italic>E), cellulose hydrolysis (<italic>cel</italic>B and <italic>bgl</italic>X), xylose transport (<italic>xyl</italic>F<italic>-xyl</italic>H) and glycolysis (<italic>pgi</italic>, <italic>pgk</italic>, <italic>fba</italic>A, <italic>eno</italic>, <italic>pfk</italic>A, <italic>gap</italic>, <italic>pyk</italic>, <italic>gpm</italic>L). Biochemical activity analysis showed that Z129 was positive for alkaline phosphatase, leucine arylamidase, acid phosphatase, naphthol-AS-BI-phosphohydrolase, α-glucosidase, β-glucosidase, and pyrrolidone arylaminase, and had the ability to use D-ribose, L-arabinose, and D-lactose. Urea-nitrogen hydrolysis rate of Z129 reached 55.37% at 48 h of incubation.</p></sec><sec><title>Discussion</title><p>Therefore, the isolated novel ureolytic strain <italic>E. hormaechei</italic> Z129 had diverse nitrogen and carbon metabolisms, and is a preferred model to study the urea hydrolysis mechanism in the rumen.</p></sec>