Low Acid Tolerance Research Articles

Global transcriptome analysis of Rhizobium favelukesii LPU83 in response to acid stress.

Acidic environments naturally occur worldwide and inappropriate agricultural management may also cause acidification of soils. Low soil pH values are an important barrier in the plant-rhizobia interaction. Acidic conditions disturb the establishment of the efficient rhizobia usually used as biofertilizer. This negative effect on the rhizobia-legume symbiosis is mainly due to the low acid tolerance of the bacteria. Here, we describe the identification of relevant factors in the acid tolerance of Rhizobium favelukesii using transcriptome sequencing. A total of 1924 genes were differentially expressed under acidic conditions, with ∼60% underexpressed. Rhizobium favelukesii acid response mainly includes changes in the energy metabolism and protein turnover, as well as a combination of mechanisms that may contribute to this phenotype, including GABA and histidine metabolism, cell envelope modifications and reverse proton efflux. We confirmed the acid-sensitive phenotype of a mutant in the braD gene, which showed higher expression under acid stress. Remarkably, 60% of the coding sequences encoded in the symbiotic plasmid were underexpressed and we evidenced that a strain cured for this plasmid featured an improved performance under acidic conditions. Hence, this work provides relevant information in the characterization of genes associated with tolerance or adaptation to acidic stress of R. favelukesii.

Enhanced Viability of Probiotics against Gastric Acid by One-Step Coating Process with Poly-L-Lysine: In Vitro and In Vivo Evaluation.

Due to their low acid tolerance, a majority of probiotics face difficulties with regard to surviving in the gastric environment long enough to reach the intestinal surfaces where they colonize and provide health benefits. We prepared a probiotic delivery system that can enhance their viability in acidic conditions by developing a one-step poly-L-lysine (PLL) coating process. We determined whether the coating process was successful by measuring the zeta potential and observing it with confocal scanning microscopy. PLL-coated L. plantarum (PLL-LP), incubated in a solution of pH 2 for 2 h, exhibited a higher viability (6.86 ± 0.12 log CFU/mL of viable cells) than non-coated L. plantarum (non-coated LP), which exhibited only 2.7 ± 1.23 log CFU/mL of viable cells. In addition, a higher amount of L. plantarum was detected in the feces of mice orally administered PLL-LP (6.2 ± 0.4 log CFU/g of feces) than in the feces of the control groups. In addition to enhancing probiotic viability in pH 2 solution, the PLL coating showed no effect on the probiotic growth pattern and the viability of either freeze-dried L. plantarum or L. plantarum, stored at −20 °C and 4 °C, respectively. Overall, these results indicated that the PLL coating is a promising potential probiotic delivery system.

Exploration of factors in response to low acid tolerance using random mutagenesis in Cronobacter malonaticus

Cronobacter species are associated with rare but severe infections in newborns, and their tolerance to environmental stress such as acid stress has been described. However, the factors involved in low acid tolerance in Cronobacter are poorly understood. Here, a transposon mutagenesis approach was used to explore the factors involved in acid tolerance in C. malonaticus. Eight mutants from mutant library (n = 215) were successfully screened through a comparison of growth with wild type (WT) strain under acid stress (pH 4.0). Eight mutating sites including glucosyltransferase MdoH, extracellular serine protease, sulfate transporter, phosphate transporter permease subunit PstC, lysine transporter, nitrogen regulation protein NR (II), D-alanine-D-alanine ligase, glucan biosynthesis protein G were successfully identified by arbitrary polymerase chain reaction and sequencing. The biomass of biofilm of eight mutants were significantly reduced using crystal violet staining (CVS) compared with that of WT. furthermore, the more compact biofilms of WT was observed than those of eight mutants through scanning electron microscopy (SEM), and confocal laser scanning microscopy (CLSM). Disassembly of biofilms appeared among mutants and WT strain from 48 h to 72 h through the increasing of dead cells and reduction of viable cells and exopolysaccharide. The study reveals the molecular basis involved in acid tolerance of C. malonaticus and a possible relationship between biofilm formation and acid tolerance, which provides valuable information for survival of C. malonaticus under acid stress.

Genomic Insights of Weissella jogaejeotgali FOL01T Reveals Its Food Fermentation Ability and Human Gut Adaptive Potential for Probiotic Applications in Food Industries.

Although the genus Leuconostoc, generally found in various fermented foods, has often been suggested to be a novel probiotic for food fermentation and health promotion, the strains in this genus showed low acid tolerance and low osmotic stress resistance activities, which are required for survival during food fermentation events. Recently, a novel species of Weissella, W. jogaejeotgali FOL01T (= KCCM 43128 = JCM 30580), was isolated from Korean fermented clams. To determine the genomic features of this new species, its genome was completely sequenced and analyzed. The genome consists of a circular chromosome of 2,114,163 bp of DNA with a G+C content of 38.8%, and the plasmid pFOL01 consists of 35,382 bp of DNA with a G+C content of 39.1%. The genome analysis showed its potential for use in food fermentation and osmotic stress resistance abilities for processing in food industries. In addition, this strain was predicted to have acid tolerance and adhesion to the mucosal layer for survival and colonization in the gut. Subsequent experiments substantiated these abilities, suggesting that W. jogaejeotgali may have probiotic potential and a high survival rate during food fermentation. Therefore, it may be suitable as a novel probiotic strain for various applications in food industries.

Genetic diversity of ATP synthase cab subunits amplified from Streptococcus mutans clinical isolates from Uyghur children with different caries susceptibility

To investigate the aciduricity and genetic diversity of ATP synthase subunit gene uncEBF derived from Uyghur children Streptococcus mutans (Sm) clinical isolates and the relationship between the genetic diversity of ATP synthase and Sm aciduric ability and caries susceptibility. Forty-one Sm strains derived from 24 caries-active individuals and 17 caries-free individuals, including 16 strains displaying high acid tolerance and 17 strains displaying low acid tolerance. Solutions of all isolated Sm with same density were made and cultured at pH 4.0 to 7.0 brain heart infusion (BHI) liquid. Terminal growth situation was compared. Gene uncEBF of these isolates were amplified with specific primers from Sm genomic DNA, and the polymerase chain reaction (PCR) products were analyzed by PCR-restriction fragment length polymorphism (RFLP) and sequenced. Aciduric ability of Sm isolated from the high caries-susceptible children were higher than that isolated from caries-free group (P = 0.023). Alu I digested fragments of uncEBF displayed two different patterns A and B. The distributions of A and B genotype strains with different acidurance were different (P = 0.039). A genotype included 7 strains displaying high acid tolerance and 2 strains displaying low acid tolerance;B genotype included 9 strains displaying high acid tolerance and 15 strains displaying low acid tolerance. The distributions of A and B genotype strains in different caries-sensitivity groups were different (P = 0.009). A genotype included 7 high caries-susceptible strains and 12 caries-free strains; B genotype included 17 high caries-susceptible strains and 5 caries-free strain. Some of these amplified uncEBF genes from different genotype were sequenced and testified that there existed variation of Alu I recognized sites. The high cariogenecity of Sm strains isolated from caries-active children shows a close relationship with the high aciduric ability of the isolated Sm strains. uncEBF gene of Sm F-ATPase obviously exhibits genetic diversity.

Natural and experimental Salmonella Typhimurium infections in foxes ( Vulpes vulpes)

The red fox ( Vulpes vulpes) can be considered as a relevant indicator species for Salmonella in the local environment and Salmonella faecal carriage was investigated in 215 red foxes in Norway shot during the winters 2002/2003 and 2003/2004. Fourteen (6.5%) of the foxes carried Salmonella. Four isolates were determined as serovars Kottbus ( n = 2) and Hessarek ( n = 2) of Salmonella enterica subspecies enterica, and one as S. enterica subspecies IIIb:61:k:1,5,(7). The remaining nine isolates were S. enterica subspecies enterica serovar Typhimurium 4,12:i:1,2 and all displayed the same pulsed-field gel electrophoresis (PFGE) profile designated A2. This serovar regularly causes disease outbreaks amongst small passerine birds during winter and most of these outbreaks are associated with the PFGE profile A2. The results strongly indicated that the Salmonella Typhimurium infections in red foxes were primarily acquired through ingestion of infected small passerines. To investigate the capability of the A2 strain to establish a true intestinal infection in the fox an inoculation experiment with an A2 isolate from small passerines was carried out in farmed silver foxes ( V. vulpes). The experiment also included one strain with an uncommonly occurring profile (X201) from small passerines. To highlight possible differences in capability of the two inoculation strains to pass the acid gastric juice in the fox, in vitro studies of their acid tolerance was carried out. Also their catalase activity and biofilm production were studied. All three foxes inoculated with the A2 strain developed sub-clinical intestinal infection of 2 weeks duration, whereas none of the three foxes inoculated with the X201 strain shed this bacterium. The X201 strain displayed a much lower capability, than the A2 strain, to survive at pH 3 in vitro. The low acid tolerance probably made it difficult for the X201 strain to pass the stomach and establish an intestinal infection in the experimental foxes. Reduced catalase activity and biofilm production were found for the X201 strain, indicating that the low acid tolerance was caused by a defect in the stationary-phase stress response system.

Effects of inner-phase components of water-in-oil-in-water emulsion on low-pH tolerance of Lactobacillus acidophilus incorporated into inner-water phase

The incorporation of Lactobacillus acidophilus, which has a low acid tolerance, into the inner-water phase of water-in-oil-in-water (W/O/W) emulsion improved the bacterial survival rate in a model gastric juice. The components that enhanced the acid tolerance of the bacterium in W/O/W emulsion were investigated. Although the acid tolerance enhancement induced by the tested components was low at pH 2, some components significantly improved the tolerance at pH 4. In particular, the acid tolerance enhancement induced by yeast extract and sodium caseinate was marked.

Genetic diversity of F-ATPase subunits gene uncA amplified from Streptococcus mutans clinical isolates

To study the genetic diversity of F-ATPase alpha subunit gene uncA derived from Streptococcus mutans (S. mutans) clinical isolates and to investigate the relationship between the genetic diversity of acidurance factor and S. mutans aciduric ability, also and the cariogenicity. Sixty-four S. mutans strains derived from 34 caries-active individuals and 30 caries-free individuals, including 18 strains displaying high acid tolerance and 20 strains displaying low acid tolerance. Gene uncA was amplified with specific primers from S. mutans genomic DNA, then the PCR products were analyzed by RFLP and sequenced. Two genotypes A and B of PCR-RFLP were revealed when digested with Hph I. Mbo II also produced two different pattern C and D. The distributions of A and B genotype strains with different caries-sensitivity groups were different (P < 0.05), and the proportion of A genotype strains from caries-activity group was higher than that from caries-free one. The distributions of C and D genotype strains with different acidurance strains were different (P < 0.05), and the proportion of C genotype strains from high acid tolerance group was higher than that from low acid tolerance group. These amplified uncA genes from different group were sequenced and there existed variation of Hph I and Mbo II recognized sites. This study indicates that uncA gene of S. mutans F-ATPase obviously displayed genetic diversity. The different Hph I-RFLP and Mbo II-RFLP genotypes could be related to the cariogenicity and acid tolerance of S. mutans strains.