AUTHOR=Yang Ruifu , Lai Bipeng , Liao Kang , Liu Baomo , Huang Lixia , Li Shaoli , Gu Jincui , Lin Ziying , Chen Yili , Wang Shuaishuai , Qiu Yanli , Deng Jiating , Chen Simin , Zhuo Chao , Zhou Yanbin 

TITLE=Overexpression of BIT33_RS14560 Enhances the Biofilm Formation and Virulence of Acinetobacter baumannii

JOURNAL=Frontiers in Microbiology

VOLUME=13

YEAR=2022

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2022.867770

DOI=10.3389/fmicb.2022.867770

ISSN=1664-302X

ABSTRACT=<p><italic>Acinetobacter baumannii</italic>, a strictly aerobic, non-lactose fermented Gram-negative bacteria, is one of the important pathogens of nosocomial infection. Major facilitator superfamily (MFS) transporter membrane proteins are a class of proteins that widely exists in microbial genomes and have been revealed to be related to biofilm formation in a variety of microorganisms. However, as one of the MFS transporter membrane proteins, little is known about the role of BIT33_RS14560 in <italic>A. baumannii</italic>. To explore the effects of BIT33_RS14560 on biofilm formation of <italic>A. baumannii</italic>, the biofilm formation abilities of 62 isolates were firstly investigated and compared with their transcript levels of <italic>BIT33_RS14560</italic>. Then, this specific gene was over-expressed in a standard <italic>A. baumannii</italic> strain (ATCC 19606) and two isolates of extensively drug-resistant <italic>A. baumannii</italic> (XDR-Ab). Bacterial virulence was observed using a <italic>Galleria mellonella</italic> infection model. High-throughput transcriptome sequencing (RNA seq) was performed on ATCC 19606 over-expressed strain and its corresponding empty plasmid control strain. <italic>Spearman’s</italic> correlation analysis indicated a significant negative correlation (<italic>R = −0.569</italic>, <italic>p = 0.000</italic>) between the △CT levels of BIT33_RS1456 and biofilm grading of <italic>A. baumannii</italic> isolates. The amount of <italic>A. baumannii</italic> biofilm was relatively high within 12–48 h. Regardless of standard or clinical strains; the biofilm biomass in the <italic>BIT33_RS14560</italic> overexpression group was significantly higher than that in the control group (<italic> p &lt; 0.0001</italic>). Kaplan–Meier survival curve analysis showed that the mortality of <italic>G. mellonella</italic> was significantly higher when infected with the <italic>BIT33_RS14560</italic> overexpression strain (<italic>χ<sup>2</sup> = 8.462, p = 0.004</italic>). RNA-Seq showed that the mRNA expression levels of three genes annotated as OprD family outer membrane porin, glycosyltransferase family 39 protein, and glycosyltransferase family 2 protein, which were related to bacterial adhesion, biofilm formation, and virulence, were significantly upregulated when <italic>BIT33_RS14560</italic> was over-expressed. Our findings provided new insights in identifying potential drug targets for the inhibition of biofilm formation. We also developed a practical method to construct an over-expressed vector that can stably replicate in XDR-Ab isolates.</p>