Association of a d -Alanyl- d -Alanine Carboxypeptidase Gene with the Formation of Aberrantly Shaped Cells during the Induction of Viable but Nonculturable Vibrio parahaemolyticus

Abstract

bVibrio parahaemolyticus is a halophilic Gram-negative bacterium that causes human gastroenteritis. When the viable but nonculturable (VBNC) state of this bacterium was induced by incubation at 4°C in Morita minimal salt solution containing 0.5% NaCl, the rod-shaped cells became coccoid, and various aberrantly shaped intermediates were formed in the initial stage. This study examined the factors that influence the formation of these aberrantly shaped cells. The proportion of aberrantly shaped cells was not affected in a medium containing D-cycloserine (50 g/ml) but was lower in a medium containing cephalosporin C (10 g/ml) than in the control medium without antibiotics. The proportion of aberrantly shaped cells was higher in a culture medium that contained 0.5% NaCl than in culture media containing 1.0 or 1.5% NaCl. The expression of 15 of 17 selected genes associated with cell wall synthesis was enhanced, and the expression of VP2468 (dacB), which encodes D-alanyl-D-alanine carboxypeptidase, was enhanced the most. The proportion of aberrantly shaped cells was significantly lower in the dacB mutant strain than in the parent strain, but the proportion was restored in the presence of the complementary dacB gene. This study suggests that disturbance of the dynamics of cell wall synthesis by enhanced expression of the VP2468 gene is associated with the formation of aberrantly shaped cells in the initial stage of induction of VBNC V. parahaemolyticus cells under specific conditions.

Current Perspectives on Viable but Non-Culturable (VBNC) Pathogenic Bacteria

“…It was also seen that in this pathogen, expression of VP2468 ( dacB ), which encodes d -alanyl- d -alanine carboxypeptidase, was more during the cold induction of VBNC. The process was accompanied by the disruption of cell wall synthesis resulting in aberrantly shaped cells (38). …”

Staying in Shape: the Impact of Cell Shape on Bacterial Survival in Diverse Environments

“…Quantitative PCR analysis of candidate cell wall synthesis and cell wall modification genes during the time window where irregular forms peaked revealed increased expression of DacB, a PG D-Ala-D-Ala carboxypeptidase. Disruption of dacB suppressed the accumulation of aberrant shapes during VBNC conversion, while complementation reversed this suppression (140). Comparison of the global PG compositions of H. pylori during exponential phase, during stationary phase, and in the coccoid VBNC form revealed a substantial increase in the dipeptide content within the sacculus, with a concomitant loss of tripeptides and tetrapeptides (141), in coccoid cells.…”

“…Recovery of E. coli L forms (bacteria lacking cell wall material through growth in the presence of cell wall inhibitors) similarly showed abundant morphological diversity and apparent budding (139). Studies of V. parahaemolyticus revealed that the proportion of irregular shapes observed during VBNC conversion increases in low-salt media, which leads to increased cellular turgor pressure (140). Computational modeling of the Gram-negative cell wall combined with limited antibiotic perturbation in E. coli suggests that localized disruption of cell wall integrity produces similar cell bulges and bends (20).…”

Alterations in the Cell Wall of Rhodococcus biphenylivorans Under Norfloxacin Stress

“…The expression of the four genes associated with cell wall was investigated by RT-qPCR according to Hung et al (2013) , with some modifications. The RNA samples were extracted by RNeasy Plus Mini Kit (Qiagen, Dusseldorf, Germany) according to the instructions provided by manufacturer.…”