Arginine deiminase pathway is far more important than urease for acid resistance and intracellular survival in Laribacter hongkongensis: a possible result of arc gene cassette duplication

Abstract

BackgroundLaribacter hongkongensis is a Gram-negative, urease-positive bacillus associated with invasive bacteremic infections in liver cirrhosis patients and fish-borne community-acquired gastroenteritis and traveler’s diarrhea. Its mechanisms of adaptation to various environmental niches and host defense evasion are largely unknown. During the process of analyzing the L. hongkongensis genome, a complete urease cassette and two adjacent arc gene cassettes were found. We hypothesize that the urease cassette and/or the arc gene cassettes are important for L. hongkongensis to survive in acidic environment and macrophages. In this study, we tested this hypothesis by constructing single, double and triple non-polar deletion mutants of the urease and two arc gene cassettes of L. hongkongensis using the conjugation-mediated gene deletion system and examining their effects in acidic environment in vitro, in macrophages and in a mouse model.ResultsHLHK9∆ureA, HLHK9∆ureC, HLHK9∆ureD and HLHK9∆ureE all exhibited no urease activity. HLHK9∆arcA1 and HLHK9∆arcA2 both exhibited arginine deiminase (ADI) activities, but HLHK9∆arcA1/arcA2 double deletion mutant exhibited no ADI activity. At pH 2 and 3, survival of HLHK9∆arcA1/arcA2 and HLHK9∆ureA/arcA1/arcA2 were markedly decreased (p < 0.001) but that of HLHK9∆ureA was slightly decreased (p < 0.05), compared to wild type L. hongkongensis HLHK9. Survival of HLHK9∆ureA/arcA1/arcA2 and HLHK9∆arcA1/arcA2 in macrophages were also markedly decreased (p < 0.001 and p < 0.01 respectively) but that of HLHK9∆ureA was slightly decreased (p < 0.05), compared to HLHK9, although expression of arcA1, arcA2 and ureA genes were all upregulated. Using a mouse model, HLHK9∆ureA exhibited similar survival compared to HLHK9 after passing through the murine stomach, but survival of HLHK9∆arcA1/arcA2 and HLHK9∆ureA/arcA1/arcA2 were markedly reduced (p < 0.01).ConclusionsIn contrast to other important gastrointestinal tract pathogens, ADI pathway is far more important than urease for acid resistance and intracellular survival in L. hongkongensis. The gene duplication of the arc gene cassettes could be a result of their functional importance in L. hongkongensis.

Transcriptomic Analysis of Laribacter hongkongensis Reveals Adaptive Response Coupled with Temperature

“…hongkongensis , which is different from other gastrointestinal tract pathogens [56]. In our transcriptomic analysis, intriguingly, the mRNA levels of urease and urea transporter (LHK_01035-LHK_01037, LHK_01040- LHK_01044) were enhanced significantly (6 to 12-fold) at 37°C.…”

“…Moreover, proteomic and gene regulatory studies of two arc gene cassettes as well as two copies of the argA and argB genes in L . hongkongensis at two physiologically relevant temperatures (37°C for human body temperature and 20°C for freshwater habitat temperature) revealed its acid resistance mechanisms under anaerobic and acidic conditions [19–22]. Although a number of evidence displayed the complex and versatile adaptation to different body temperatures of niches, systematic transcriptomic studies have not yet been carried out.…”

Molecular characterization of arginine deiminase pathway in Laribacter hongkongensis and unique regulation of arginine catabolism and anabolism by multiple environmental stresses

“…Arginine catabolism via the arginine deiminase (ADI) pathway has been shown to be widely distributed in bacterial species, enabling them to survive within harsh acidic environments and evade host defence mechanisms (Cunin et al ., ; Degnan et al ., ; Dong et al ., ; Ryan et al ., ; Choi et al ., ; Xiong et al ., ). The ADI pathway consists of three enzymes: arginine deiminase, ornithine carbamoyltransferase and carbamate kinase; which are encoded by arcA , arcB and arcC respectively .…”

Identification and analysis of genomic islands in Burkholderia cenocepacia AU 1054 with emphasis on pathogenicity islands

“…The unmarked, non-polar mutant strains were constructed as described by Flannagan et al [37], with slight modifications. Briefly, 5’- and 3’-flanking regions of target genes ( copR and lipA ) were amplified by PCR from chromosomal DNA, respectively, and the individual PCR products were mixed to generate an in-frame deletion pattern of target genes by an overlapping PCR method, as described previously [38, 39]. Then, the overlapping amplicon containing the in-frame deletion pattern of target genes was sub-cloned into pGPI-SceI, resulting in recombinant plasmids pGPI- copR and pGPI- lipA respectively.…”