Function Of RmlB In The Listeria Monocytogenes

Listeria monocytogenes is a gram-positive foodborme pathogen that could form biofilms on the most solid surfaces.The biofilm can be highly resistance to antibiotics,and difficult to be cleaned away.Therefore,they can cause severe problems for public hygiene and human health.So far,L.monocytogenes is widely susceptible to most common antibiotic,we used antibiotics to treat listeriosis clinically,but more and more studies have shown that resistance of L.monocytogenes has gradually increased.lt is well known that the morphology and structure of bacteria are correlation with their physiological function,immunity and pathogenicity.L-rhamnose is widely exists in the cell wall and capsule of many pathogenic bacteria.The gene rmlB encodes the key enzyme in the synthesis of L-rhamnose.This paper explores the function of rmlB on physiological characteristics,antibiotic resistance,biofilm formation,and virulence in L.monocytogenes.1)The rmlB gene was successfully deleted through homologous recombination,the mutant is EGDeArmlB.2)The bacterial growth,morphology,motility and stress resistance of wild-type and the rmlB deletion mutant were compared.The data show that the deletion of rmlB had no significant effect on the growth,stress resistance,cell morphology,cell wall structure and motility of L.monocytogenes,which suggests that the synthesis of rhamnose is not a necessary cellular functional pathway for the morphology,growth and movement in L.monocytogenes.3)The antibiotic resistance between the wild-type and the rmlB deletion mutant was compared by minimal inhibitory concentration,disc agar diffusion method and the tolerance assay.The transcription of PBPs genes of the wild type and mutant stains were investigated.The results show that compared with the wild-type strain,the deletion of rmlB resulted in an increased susceptibility of L.monocytogenes to envelope-acting antibiotics such as several cephalosporins and bacitracin(p≤0.01),which is possible related to the reduction of the expression of lmo2229(the encoding gene of PBP4).4)The biofilm formation ability of the wild type and mutant strains were measured using the microtiter plate and the fluorescence microscopic assay.The data show that compared with the wild-type strain,the deletion of rmlB lead to a significant decrease in bacterial biofilm formation(p≤0.01),but no significant effect on the initial attachment process of biofilm formation.5)The transcription of major virulence genes in L.monocytogenes(prfA,p1cA,plcB,actA A mpl,hly,gtcA)and hemolytic activities of the wild type and mutant stains were investigated.The results show that compared with the wild-type strain,the transcription expression of the well-known virulent gene hly and gtcA in L.monocytogenes and bacterial haemolytic activity were significantly reduced in the rmlB deletion mutant(p≤0.01),but there is no significant effect on the expression of other major virulence factors,including virulence gene transcription regulatory protein PrfA.In conclusion,rmlB plays a crucial role in L.monocytogenes against the envelope-acting antibiotics,in biofilm formation and virulence.