APPLIED MICROBIOLOGY AND BIOTECHNOLOGY
·
2021
·Vol. 105
·期 7
·2841-2854
·
IF 4.813
Zhou Gang,Wang Ying-si,Peng Hong,Li Su-juan,Sun Ting-li,Shen Peng-fei,Xie Xiao-bao,Shi Qing-shan
Abstract The genus Citrobacter is commonly found in environmental and industrial settings, some members of which have been used for bioremediation of heavy metals owing to the absorption ability of their biofilms. Although our previous studies have found that the outer membrane protein A (OmpA) contributes to the process of Citrobacter werkmanii biofilm formation, the underlying mechanisms remain elusive. Therefore, we deleted ompA from the genome of C. werkmanii and investigated its phenotypes in comparison to the wild type strain (WT) and the complementary strain using biochemical and molecular techniques including RNA-Seq. Our results demonstrated that the deletion of ompA led to an increase in biofilm formation on both polystyrene and glass surfaces due to upregulation of some biofilm formation related genes. Meanwhile, swimming ability, which is mediated by activation of flagellar assembly genes, was increased on semi-solid plates in the ∆ompA strain when compared with WT. Additionally, inactivation of ompA also caused increased 1,2-benzisothiazolin-3-one (BIT) resistance, differential responses to Ca 2+ stress, curli protein expression and cellulose production. Finally, ∆ompA caused differential expression of a total of 1470 genes when compared with WT, of which 146 were upregulated and 1324 were downregulated. These genes were classified into different Gene Ontology (GO) and KEGG pathways. In summary, ompA in C. werkmanii contributes to a variety of biological functions and may act as a target site to modulate biofilm formation. Key points
• ompA is a negative regulator for biofilm formation by C. werkmanii.
• ompA inhibits swimming motility of C. werkmanii.
• ompA deletion causes different expression profiles in C. werkmanii.
研究领域:
微生物学
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分子生物学
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环境科学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
GENE
·
2022
·
IF 3.913
Gang Zhou,Ying-si Wang,Hong Peng,Su-juan Li,Ting-li Sun,Cai-ling Li,Qing-shan Shi,Xiao-bao Xie
Citrobacter werkmanii , an aerobe and mesophilic Proteobacterium, is universal in industrial putrefaction, coastal water, and human blood. Our previous studies have discovered that outer membrane protein X (OmpX) of C. werkmanii is involved in calcium response, but the underlying mechanisms and its molecular characteristics remain elusive. To that end, the ompX gene was deleted from the genome of C. werkmanii and its phenotypic variations were thoroughly investigated in conjunction with the wild type (WT) and complementary strains using biochemical and molecular techniques such as RNA-Seq, respectively. The results demonstrated that deleting ompX reduces biofilm formation on polystyrene and glass surfaces. Meanwhile, Δ ompX ’s swimming ability but not for its twitching or swarming abilities, was also reduced on semi-solid plates compared with WT, which was caused by inhibition of flagellar assembly genes, such as flgC , flhB , and fliE , etc. Furthermore, ompX inactivation altered susceptibility to various bactericide classes, as well as responses to Ca 2+ and Mg 2+ stress. In addition, when compared to WT, Δ ompX captures a total of 1,357 deferentially expressed genes (DEGs), of which 465 were up-regulated and 892 were down-regulated, which can be enriched into various GO ontology and KEGG pathway terms. Furthermore, ompX , as well as ompD and ompW , can be modulated at the transcriptional levels by rbsR and tdcA . Overall, the ompX gene contributed to a variety of biological functions in C. werkmanii and could be served as a targeted site for controlling biofilm formation and developing new bactericides.
研究领域:
微生物学
·
分子生物学
·
基因组学
·
生物信息学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Journal of Proteomics
·
2022
·
IF 3.855
Gang Zhou,Ying-si Wang,Hong Peng,Su-juan Li,Ting-li Sun,Qing-shan Shi,Jordi Garcia-Ojalvo,Xiao-bao Xie
Mounting evidence has shown that antimicrobial agents can interfere synergistically with bacterial viability and proliferation when acting together at both the planktonic and biofilm levels without clear underlying molecular mechanisms. Here, multiplexed proteomics by iTRAQ was used to study the interplay between two biocides, the isothiazolone 1,2-benzisothiazolin-3-one (BIT) and the chelating agent disodium ethylenediaminetetraacetic acid (EDTA-2Na), employing the Citrobacter werkmanii as a model system. We first confirmed that these two biocides act synergistically on this bacterial species and then extracted the proteomic profiles of C. werkmanii cells in the presence of BIT, EDTA-2Na, and their combinations. In particular, we identified 43 core proteins that are differentially expressed in all three conditions simultaneously. Meanwhile, we found that these core proteins are consistently up-regulated when these two biocides are present, but not for single biocides, where we found a balanced mix of up- and down-regulation. Meanwhile, most of the deletion mutants of the core DEPs exhibited biofilm growth inhibition under joint biocide action, while their response was very heterogenous, with respect to the wild-type strain. Together, our results show that while BIT and EDTA-2Na act on multiple protein targets, they interact synergistically at the protein level in a very consistent manner. Significance Our preliminary experiments have demonstrated that a combination of 1,2-benzisothiazolin-3-one (BIT) and EDTA-2Na shows higher inhibitory effects on planktonic growth and biofilm formation in both C. werkmanii and Staphylococcus aureus than when these two biocides act alone. However, the mechanistic basis of such synergistic interaction is still unknown. Therefore, the key proteins involved in the above-mentioned enhanced antimicrobial synergy were elucidated using multiplexed proteomics analysis by isobaric tags for relative and absolute quantification (iTRAQ). Our results reveal that the joint action of BIT and EDTA-2Na induces consistent protein expression alteration in a set of core proteins of C. werkmanii , which underlies a strong synergistic antimicrobial effect, which increase our understanding of the action modes of BIT and EDTA-2Na as well as their combinations.
研究领域:
微生物学
·
蛋白质组学
·
分子生物学
·
生物技术
✓ 已验证
产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）