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Previous study has reported that Lactiplantibacillus plantarum ZDY2013 which was screened from traditional Chinese fermented soybeans has a strong acid resistance. The purpose of this study was to uncover the genes potentially related to its genetic adaptation and probiotic profiles, based on comparative genomic and comparative transcriptome analysis. We got the basic information about L. plantarum ZDY2013 and identified genes which are related to genetic adaptation and probiotic profiles, including carbohydrate transport and metabolism, cell wall/membrane/envelope biogenesis, proteolytic enzyme systems and amino acid biosynthesis, CRISPR adaptive immunity, stress responses, ability to adhere to the host intestinal wall, exopolysaccharide (EPS) biosynthesis, and bacteriocin biosynthesis. Comparative transcriptome showed CK group (normal MRS culture L. plantarum ZDY2013) and SCL group (pH 3.0 MRS culture L. plantarum ZDY2013) had 652 significant differentially expressed genes including 310 up-regulated genes and 342 down-regulated genes. Besides that, these genes had been classified through KEGG and GO functional annotation. In addition, we also found top 20 KEGG pathways adjusted to acid stress. Then, some genes were selected to verify the transcriptome analysis and explore the mechanism of how L. plantarum ZDY2013 tolerate acid stress. We found that some genes of ABC transporter, phosphotransferase system, oxidation reduction process, membrane transporter and phosphorylation metabolism process had a significant change. These results suggested that comparative characterization of the L. plantarum ZDY2013 genome and transcriptome provided the genetic basis for further elucidating the functional mechanisms of it.

A high-salt diet (HSD) is one of the key risk factors for hypertension and kidney injury. In this study, a HSD C57BL/6J mice model was established with 4% NaCl, and then different concentrations of Lactobacillus plantarum ZDY2013 were intragastrically administered for 2 weeks to alleviate HSD-induced renal injury. For the study, 16S rRNA gene sequencing, non-targeted metabonomics, real-time fluorescent quantitative PCR, and Masson’s staining were used to investigate the mechanism of L. plantarum ZDY2013 in alleviating renal damage. Results showed that HSD caused intestinal inflammation and changed the intestinal permeability of mice, disrupted the balance of intestinal flora, and increased toxic metabolites (tetrahydrocorticosteron (THB), 3-methyhistidine (3-MH), creatinine, urea, and L-kynurenine), resulting in serious kidney damage. Interestingly, L. plantarum ZDY2013 contributed to reconstructing the intestinal flora of mice by increasing the level of Lactobacillus and Bifidobacterium and decreasing that of Prevotella and Bacteroides. Moreover, the reconstructed intestinal microbiota significantly changed the concentration of the metabolites of hosts through metabolic pathways, including TCA cycle, ABC transport, purine metabolism, and histidine metabolism. The content of uremic toxins such as L-kynurenine, creatinine, and urea in the serum of mice was found to be decreased by L. plantarum ZDY2013, which resulted in renal injury alleviation. Our data suggest that L. plantarum ZDY2013 can indeed improve chronic kidney injury by regulating intestinal flora, strengthening the intestinal barrier, limiting inflammatory response, and reducing uremic toxins.

The key role of mitochondria in the integration of apoptosis and oxidative stress in mammals has been documented. In plants, mitochondria are implicated in cytoplasmic male sterility (CMS), programmed cell death (PCD), and oxidative stress. However, to date there has been no evidence presented of the interplay among CMS, PCD and oxidative stress. In this study, we demonstrate that the pollen abortion of Honglian CMS line of rice (Oryza sativa L.) displays a PCD phenotype. A premature loss of microspores was accompanied by several biochemical markers of apoptosis. Analysis of mitochondria revealed that during the PCD process there was disruption of the inner mitochondrial membrane potential in microspores. This disruption was correlated with excess production of reactive oxygen species and down-regulation of the activity of superoxide dismutase (SOD), ascorbate peroxidase (APX) and catalase in mitochondria. The excess accumulation of ROS spanned three stages from pollen mother cell to early-uninucleate stage, and was followed by the occurrence of PCD at meiosis. Our data suggest that the microspores suffered from severe oxidative stress during pollen development. It is likely the chronic oxidative stress triggered the tissue-specific PCD and consequently resulted in the abortion of microspores.

In this study, the anti-Helicobacterpylori activity of Lactobacillusplantarum ZDY2013 was investigated and Lactobacillusrhamnosus GG was used as a positive control. The anti-H. pylori mechanism in vitro was also examined. Results revealed that either the viable cells or supernatant of L. plantarum ZDY2013 could suppress the growth or urease activity of H. pylori. The inhibitory effects of L. plantarum ZDY2013 were relatively higher than those of L. rhamnosus GG (P 0.05), and such effects might be a result of their lactic acid production (e.g., 51.105 ± 0.097 mmol/L for L. plantarum ZDY2013 and 33.113 ± 0.063 mmol/L for L. rhamnosus GG). The anti-adhesion capacity of L. plantarum ZDY2013 against H. pylori was also stronger than that of L. rhamnosus GG in terms of inhibition, competition, and displacement. Among these inhibitory strategies, competition exhibited the best performance, with an inhibition ratio of 92.65%. Upon inhibition and anti-adhesion, the cells and supernatant of L. plantarum ZDY2013 significantly strengthened the expression of the anti-inflammatory cytokine IL-10, but attenuated the expression of the pro-inflammatory cytokine TNF-α in AGS cells induced by H. pylori SS1. Remarkably, the supernatant of ZDY2013 achieved a relatively higher anti-inflammatory effect than that exerted by its cells. With excellent lactic acid yield and antagonistic and anti-inflammatory effects against H. pylori SS1 infection, L. plantarum ZDY2013 shows potential to be used as a probiotics candidate.

Ulcerative colitis (UC) is an inflammatory bowel disease that is difficult to cure, with rising incidence in recent decades. Probiotics have become a new strategy for UC treatment. In this study, we chose 2 new multisource probiotics, Lactobacillus plantarum ZDY2013 from acid beans and Bifidobacterium bifidum WBIN03 from infant feces, and a mixture of both, to investigate the anti-inflammatory and antioxidant effect on H2O2-induced oxidative damage in a HT-29 cell model and dextran sodium sulfate (DSS)-induced UC in mice. Compared with the model group, the general relative indices results showed L. plantarum ZDY2013 and B. bifidum WBIN03 have a significant effect on DSS-induced UC in mice, by downregulating the pro-inflammatory cytokines (e.g., TNF-α) and upregulating antioxidant factors (e.g., SOD1, SOD2, GPX2) at the transcriptional level. By means of high-throughput sequencing (16S V3–V4) and systematical bioinformatics analyses, we found that colitis may be associated with the changes in intestinal flora, especially Firmicutes and Bacteroides. Administration of L. plantarum ZDY2013 increased the abundance of Lactobacillus animalis, whereas B. bifidum WBIN03 increased the abundance of Lachnospiraceae bacterium COE1. Our results revealed that a supplement of L. plantarum ZDY2013 and B. bifidum WBIN03 remit UC through modification of gut microbiota to regulate oxidative stress and inflammatory mediators.

This study aimed to disclose the acid tolerance mechanism of Lactobacillus plantarum by comparing L. plantarum ZDY 2013 with the type strain L. plantarum ATCC 8014 in terms of cell membrane, energy metabolism, and amino acid metabolism. L. plantarum ZDY 2013 had a superior growth performance under acidic condition with 100-fold higher survival rate than that of L. plantarum ATCC 8014 at pH 2.5. To determine the acid tolerance physiological mechanism, cell integrity was investigated through scanning electron microscopy. The study revealed that L. plantarum ZDY 2013 maintained cell morphology and integrity, which is much better than L. plantarum ATCC 8014 under acid stress. Analysis of energy metabolism showed that, at pH 5.0, L. plantarum ZDY 2013 enhanced the activity of Na+/K+-ATPase and decreased the ratio of NAD+/NADH in comparison with L. plantarum ATCC 8014. Similarly, amino acid metabolism of intracellular arginine, glutamate, and alanine was improved in L. plantarum ZDY 2013. Correspondingly, the activity of arginine deiminase and glutamate decarboxylase of L. plantarum ZDY 2013 increased by 1.2-fold and 1.3-fold compared with L. plantarum ATCC 8014 in acid stress. In summary, it is demonstrated that the special physiological behaviors (integrity of cell membrane, enhanced energy metabolism, increased amino acid and enzyme level) of L. plantarum ZDY 2013 can protect the cells from acid stress.

The small intestinal (SI) microbiota has an essential role in the maintenance of human health. However, data about the indigenous bacteria in SI as affected by probiotics are limited. In our study, the short-term and long-term effects of a probiotic candidate, Lactobacillus plantarum ZDY2013, on the SI microbiota of C57BL/6J mice were investigated by the Illumina HiSeq (Novogene Bioinformatics Technology Co., Ltd., Tianjin, China) platform targeting the V4 region of the 16S rDNA. A total of 858,011 sequences in 15 samples were read. The α diversity analysis revealed that oral administration with L. plantarum ZDY2013 for 3 wk led to a significant increase in the richness and diversity of the SI bacterial community. Principal coordinate analysis and unweighted pair-group method with arithmetic means analysis showed a clear alteration in the SI microbiota composition after 3 wk of L. plantarum ZDY2013 treatment, although these changes were not found 6 wk after ceasing L. plantarum ZDY2013 administration. Species annotation showed that the dominant phyla in SI microbiota were Firmicutes, Bacteroidetes, Proteobacteria, and Verrucomicrobia. Interestingly, operational taxonomic unit cluster analysis showed that administration with L. plantarum ZDY2013 for 3 wk significantly increased the abundance of Proteobacteria, but decreased that of Bacteroidetes. Linear discriminant analysis coupled with effect size identified 18 bacterial taxa (e.g., Ruminococcus spp. and Clostridium spp.) that overgrew in the SI microbiota of the mice administered with L. plantarum ZDY2013 for 3 wk, and most of them belonged to the phyla Bacteroidetes and Proteobacteria. However, only one bacterial taxon (e.g., Nocardioides spp.) was over-represented in the SI microbiota of mice 6 wk after L. plantarum ZDY2013 administration. Overall, this study shows that oral administration with probiotic results in an important but transient alteration in the microbiota of SI.

The aim of this study was to evaluate the safety of enterococci isolated from Chinese infants and screen out potential probiotic candidates. One hundred eight strains were isolated from feces of 34 healthy infants, and 38 strains of Enterococcus spp. were categorized as follows: E. faecalis (22), E. faecium (10), E. hirae (3), E. durans (2), and E. casseliflavus (1). Of these, 72.7% of E. faecalis came from infants delivered by cesarean and 62.5% of E. faecium from infants delivered vaginally. For safety evaluation of strains, we determined presence of virulence genes; production of hemolysin, gelatinase, and biofilm; and antimicrobial susceptibility of enterococci. Six out of 14 virulence genes were detected with a distribution of gelE (26.3%), cylA (39.4%), esp (15.8%), efaA (63.2%), asa1 (50.0%), and ace (50.0%). In phenotype analysis, 36.8% of the strains exhibited positive hemolytic activity and 17.5% were positive for production of gelatinase. Results of antimicrobial susceptibility showed that different percentages of the strains were resistant to ciprofloxacin (5.2%), vancomycin (7.8%), rifampicin (10.5%), erythromycin (52.6%), and gentamycin (52.6%); remarkably, none of the strains were resistant to ampicillin or chloramphenicol. In total, 10 strains, including 6 E. faecium, which are free of virulence determinants and sensitive to common antimicrobial agents (e.g., ampicillin and vancomycin), were further assessed for their probiotic properties. All strains survived well in simulated gastric fluid and intestinal tract, with maximum reductions of 0.600 and 0.887 log cfu/mL, respectively. Six strains of E. faecium could resist 0.3 to 1.0% bile salt, of which E. faecium WEFA23 presented the highest growth (75.06%) at 1.0% bile salt. All strains showed bile salt hydrolase activity on glycodeoxycholic acid, but only 3 of E. faecium showed activity on taurodeoxycholic acid. These results deliver useful information on the safety of enterococci in infants in China, and provide a protocol to screen probiotics for absence of virulence and antimicrobial susceptibility of enterococci.

Lactobacillus plantarum WLPL04, a specific strain isolated from human breast milk, was investigated for its survival capacity (acid and bile salt tolerance, survival in simulated gastrointestinal tract, inhibition of pathogens, antibiotic susceptibility, yield of exopolysaccharides) and probiotic properties (antiadhesion of pathogens, protection from harmful effect of sodium dodecyl sulfate, and antiinflammatory stress on Caco-2 cells). The results showed that Lb. plantarum WLPL04 had broad-spectrum activity against gram-positive strains (Listeria monocytogenes CMCC54007, Bacillus cereus ATCC14579, and Staphylococcus aureus CMCC26003) and gram-negative strains (Pseudomonas aeruginosa MCC10104, Shigella sonnei ATCC25931, Enterobacter sakazakii ATCC29544, Salmonella typhimurium ATCC13311, and Escherichia coli O157:H7). Antibiotic susceptibility tests showed that Lb. plantarum WLPL04 was susceptible to 8 of 14 antibiotics (e.g., erythromycin and nitrofurantoin) and resistant to 6 of 14 antibiotics (e.g., kanamycin and bacitracin). Lactobacillus plantarum WLPL04 was able to survive at pH 2.5 for 3h and at 0.45% bile salt for 12h, suggesting that it can survive well in the gastrointestinal tract. In addition, the exopolysaccharide yield of Lb. plantarum WLPL04 reached 426.73 ± 65.56 mg/L at 24h. With strategies of competition, inhibition, and displacement, Lb. plantarum WLPL04 reduced the adhesion of E. coli O157:H7 (35.51%), Sal. typhimurium ATCC 13311 (8.10%), and Staph. aureus CMCC 26003 (40.30%) on Caco-2 cells by competition, and subsequently by 59.80, 62.50, and 42.60%, respectively, for the 3 pathogens through inhibition, and by 75.23, 39.97, and 52.88%, respectively, through displacement. Lactobacillus plantarum WLPL04 attenuated the acute stress induced by sodium dodecyl sulfate on Caco-2 cells and significantly inhibited the expression of inflammatory cytokines (IL-6, IL-8 and tumor necrosis factor-α) on Caco-2 cells but increased IL-10 expression in vitro compared with the Salmonella-treated group. In summary, Lb. plantarum WLPL04 from breast milk could be considered as a probiotic candidate for dairy products to promote human health.

Helicobacter pylori is a gram-negative pathogen linked to gastric ulcers and stomach cancer. Gastric microbiota might play an essential role in the pathogenesis of these stomach diseases. In this study, we investigated the preventive effect of a probiotic candidate Lactobacillus plantarum ZDY 2013 as a protective agent against the gastric mucosal inflammation and alteration of gastric microbiota induced by H. pylori infection in a mouse model. Prior to infection, mice were pretreated with or without 400 µL of L. plantarum ZDY 2013 at a concentration of 10(9) cfu/mL per mouse. At 6 wk postinfection, gastric mucosal immune response and alteration in gastric microbiota mice were examined by quantitative real-time PCR and high-throughput 16S rRNA gene amplicon sequencing, respectively. The results showed that L. plantarum ZDY 2013 pretreatment prevented increase in inflammatory cytokines (e.g., IL-1β and IFN-γ) and inflammatory cell infiltration in gastric lamina propria induced by H. pylori infection. Weighted UniFrac principal coordinate analysis showed that L. plantarum ZDY 2013 pretreatment prevented the alteration in gastric microbiota post-H. pylori infection. Linear discriminant analysis coupled with effect size identified 22 bacterial taxa (e.g., Pasteurellaceae, Erysipelotrichaceae, Halomonadaceae, Helicobacteraceae, and Spirochaetaceae) that overgrew in the gastric microbiota of H. pylori-infected mice, and most of them belonged to the Proteobacteria phylum. Lactobacillus plantarum ZDY 2013 pretreatment prevented this alteration; only 6 taxa (e.g., Lachnospiraceae, Ruminococcaceae, and Clostridiaceae), mainly from the taxa of Firmicutes and Bacteroidetes, were dominant in the gastric microbiota of the L. plantarum ZDY 2013 pretreated mice. Administration of L. plantarum ZDY 2013 for 3 wk led to increase in several bacterial taxa (e.g., Rikenella, Staphylococcus, Bifidobacterium), although a nonsignificant alteration was found in the gastric microbiota. Overall, this study demonstrated that L. plantarum ZDY 2013 pretreatment played an important role in preventing gastric mucosal inflammation and gastric microbiota alteration induced by H. pylori infection, and the selective modulation in gastric microbiota posed by this intervention suggested that targeting gastric microbiota through oral administration of probiotics might be an alternative strategy to prevent H. pylori infection.

Acid tolerance responses (ATR) in Lactobacillus plantarum ZDY2013 were investigated at physiological and molecular levels. A comparison of composition of cell membrane fatty acids (CMFA) between acid-challenged and unchallenged cells showed that acid adaptation evoked a significantly higher percentage of saturated fatty acids and cyclopropane fatty acids in acid-challenged than in unchallenged cells. In addition, reverse transcription-quantitative PCR analysis in acid-adapted cells at different pH values (ranging from 3.0 to 4.0) indicated that several genes were differently regulated, including those related to proton pumps, amino acid metabolism, sugar metabolism, and class I and class III stress response pathways. Expression of genes involved in fatty acid synthesis and production of alkali was significantly upregulated. Upon exposure to pH 4.5 for 2 h, a higher survival rate (higher viable cell count) of Lactobacillus plantarum ZDY2013 was achieved following an additional challenge to 40 mM hydrogen peroxide for 60 min, but no difference in survival rate of cells was found with further challenge to heat, ethanol, or salt. Therefore, we concluded that the physiological and metabolic changes of acid-treated cells of Lactobacillus plantarum ZDY2013 help the cells resist damage caused by acid, and further initiated global response signals to bring the whole cell into a state of defense to other stress factors, especially hydrogen peroxide.

Milk is one of the earliest and most common allergen sources in the world, with β-lactoglobulin representing a major allergen protein. Numerous studies have reported that probiotics exert antiallergic and anti-inflammatory effects. Here, we examined the effects of 3 strains of Lactobacillus on immunomodulatory functions, intestinal barrier functions, and intestinal microbiota through a β-lactoglobulin-induced allergic mouse model. We found that the oral administration of Lactobacillus plantarum ZDY2013 and Lactobacillus rhamnosus GG suppressed allergic response, attenuating serum IgE and relieving anaphylaxis symptoms. The 3 strains of Lactobacillus could induce T helper (Th) 1 or T regulatory cells to differentiate to inhibit the Th2-biased response for regulating Th1/Th2 immune balance. Furthermore, L. plantarum ZDY2013 and L. rhamnosus GG enhanced intestinal barrier function through the regulation of tight junction. We also found that L. plantarum ZDY2013 and L. plantarum WLPL04 could regulate alterations in intestinal microbiota caused by allergies. In particular, Rikenella, Ruminiclostridium, and Lachnospiraceae UCG-006 were considerably reduced after treatment with L. plantarum ZDY2013 and L. plantarum WLPL04. These results suggested that 3 Lactobacillus strains may serve as an effective tool for the treatment of food allergies by regulating immune and gut microbiota.

Lactobacillus plantarum ZDY 2013, a novel strain isolated from Chinese traditional fermented acid beans, was systematically evaluated for its survival capacity under stress conditions (pH, bile salt, simulated gastrointestinal tract, and antibiotics), production of exopolysaccharide and antagonism against 8 pathogens. Its effect on mice gut microbiota was also investigated by quantitative PCR and PCR-denaturing gradient gel electrophoresis. The results showed that ZDY 2013 can grow at pH 3.5 and survive at pH 2.0 for 6 h and at 0.45% bile salt for 3 h. The exopolysaccharide yield was up to 204±7.68 mg/L. The survival rate of ZDY 2013 in a simulated gastrointestinal tract was as high as 65.84%. Antagonism test with a supernatant of ZDY 2013 showed maximum halo of 28 mm against Listeria monocytogenes . The inhibition order was as follows: Listeria monocytogenes , Salmonella typhimurium , Escherichia coli , Pseudomonas aeruginosa , Shigella sonnei , Enterobacter sakazakii , and Staphylococcus aureus . Lactobacillus plantarum ZDY 2013 was sensitive to some antibiotics (e.g., macrolide, sulfonamides, aminoglycoside, tetracyclines and β-lactams), whereas it was resistant to glycopeptides, quinolones, and cephalosporins antibiotics. Denaturing gradient gel electrophoresis profile demonstrated that ZDY 2013 administration altered the composition of the microbiota at various intestinal loci of the mice. Moreover, the quantitative PCR test showed that the administration of ZDY 2013 enhanced the populations of Bifidobacterium and Lactobacillus in either the colon or cecum, and reduced the potential enteropathogenic bacteria (e.g., Enterococcus , Enterobacterium , and Clostridium perfringens ). Lactobacillus plantarum ZDY 2013 exhibited high resistance against low pH, bile salt, and gastrointestinal fluid, and possessed antibacterial and gut microbiota modulation properties with a potential application in the development of dairy food and nutraceuticals.

In this study, fermentation of Allium chinense bulbs was carried out with Lactobacillus plantarum ZDY 2013. A decrease in pH from 6.8 to 3.5 and a stable lactic acid bacteria population were observed during 7-d fermentation. The total phenolic content increased by 2.7-fold in the aqueous and ethanol extracts of A. chinense bulbs after fermentation. Antioxidant capacity including 2,2-diphenyl-1-picrylhydrazyl radical-scavenging effect and reducing power of both extracts was significantly (P < 0.05) improved after fermentation. Antagonistic test against 6 pathogens showed that fermentation significantly (P < 0.05) enhanced the antimicrobial activity in both extracts of fermented bulbs, especially in the ethanol extracts of fermented bulbs against L. monocytogenes. Analysis of the free amino acid (FAA) profile by ion-exchange chromatography revealed that fermentation significantly (P < 0.05) increased total FAA content. In addition, among 27 kinds of volatile components analyzed by headspace-solid phase microextraction-gas chromatography-tandem mass spectrometry, sulfur-containing compounds accounted for 65.23%, but decreased to 43.65% after fermentation. Our results suggested that fermentation of A. chinense bulbs with L. plantarum could improve their biofunctionalities, and nutritional and chemical properties.

We report our investigation of the functions of PagN in Salmonella pathogenesis and its potential as a vaccine candidate. Further investigation conducted in this study indicates that the outer membrane protein PagN is important for Salmonella adhesion/invasion of epithelial cells as well as bacterial virulence. When pagN was deleted from Salmonella enterica serovar Typhimurium (S. Typhimurium), the adhesion and invasion of HT-29 epithelial cells was significantly decreased compared with the wild type strain. Mice infected with the pagN mutant strain exhibited less pathological signs in the intestine and survived longer than the wild-type-infected mice. PagN is widely distributed and conserved among clinical isolates of different Salmonella serovars, making PagN a potential vaccine candidate for Salmonella infection. To elucidate the potential of PagN as a vaccine, we expressed and purified recombinant PagN (rPagN). When rPagN was tested in mice, it provided significant protection against Salmonella infection in vivo. In vitro, anti-PagN serum enhanced clearance of Salmonella, indicating a contribution of PagN-specific antibodies to the killing process. This correlates well with the observed protection of mice immunized with rPagN. Our preliminary results indicate more functions of PagN in S. Typhimurium virulence as well as its potential as a protective vaccine.

Salmonella is the leading pathogenic bacteria in food and contaminated water. The aim of this study was to develop a rapid and reliable technique for simultaneous detection of the main three serotypes ( Salmonella enterica serovars Typhimurium, Paratyphi B and Typhi) of Salmonella . Primers were designed to amplify the genes specific to each of these three serotypes for simultaneous detection using polymerase chain reaction (PCR). To ensure the detection of only viable cells, propidium monoazide (PMA) was applied to selectively suppress the DNA signal from dead cells. Results showed that the PMA-multiplexed PCR (PMA-mPCR) assay always gave negative results for heat-killed Salmonella at concentrations up to 1 × 10 6 CFU/ml in pure culture or 1 × 10 6 CFU/g in spiked food products (tomato, chicken, beef and ham). Results showed that the detection limits of the PMA-mPCR assay were approximately 10 2 CFU/ml (4.3 × 10 2 CFU/ml for S. Typhimurium, 3.7 × 10 2 CFU/ml for S. Paratyphi B, 7.2 × 10 2 CFU/ml for S. Typhi) in pure culture and 10 3 CFU/g (4.3 × 10 3 CFU/g for S. Typhimurium, 3.7 × 10 3 CFU/g for S. Paratyphi B, 7.2 × 10 3 CFU/g for S. Typhi) in food produce. These results demonstrated that the PMA-mPCR assay can simultaneously detect and identify viable S . Typhimurium, Paratyphi B and Typhi in a short period of time, even in food produce.

Cytoplasmic male sterility (CMS) is a phenomenon widely observed in various plant species characterized with disrupted anther development caused by mitochondrial mutation. CMS is becoming a model system for the investigations of nucleus-cytoplasmic interaction. To reveal the possible effects of CMS genes on plant growth in adverse environment, plant development and biochemical characters of mitochondria from Honglian (HL)-CMS line Yuetai A and maintainer Yuetai B treated with H(2)O(2) were analyzed. Results showed that 40-60mM H(2)O(2) significantly inhibits rice seedling development and growth. When treated with H(2)O(2), ATP content and mitochondrial membrane potential in Yuetai A decreased significantly faster than those of Yuetai B. These biochemical changes were accompanied by the severe nuclear DNA fragmentation and the release of mitochondrial cytochrome c in the leaf cells of Yuetai A. In addition, the antioxidative enzyme activities and mitochondrial electron transfer chain complexes were significantly down-regulated. Disturbance of the biochemical indexes indicate that HL-CMS line is more susceptible to H(2)O(2) stress than the maintainer line, the deleterious effects caused by the CMS-related ORFH79 peptide compromises the adaptability of HL-CMS line to the adverse environment.

Objective To investigate the indigenous lactobacilli from the vagina of pregnant women and to screen the isolates with antagonistic potential against pathogenic microorganisms. Methods The strains were isolated from pregnant women's vagina and identified using the API50CH system. The ability of the isolates to produce hydrogen peroxide was analyzed semi-quantitatively using the TMB-HRP-MRS agar. The antagonistic effects of the isolates on pathogenic microorganisms were determined with a double layer agar plate. Results One hundred and three lactobacilli strains were isolated from 60 samples of vaginal secretion from healthy pregnant women. Among them, 78 strains could produce hydrogen peroxide, in which 68%, 80%, 80%, and 88% had antagonistic effects against Candida albicans CMCC98001, Staphylococcus aureus CMCC26003, Escherichia coli CMCC44113, and Pseudomonas aeruginosa CMCC10110, respectively. Conclusion The recovery of hydrogen peroxide-producing lactobacilli decreases with the increasing pregnant age and time. The most commonly isolated species from vagina of Chinese pregnant women are Lactobacillus acidophilus and Lactobacillus crispatus . Most of L. acidophilus and L. crispatus produce a high H 2 O 2 level.

Starch biosynthesis, a very critical event during pollen development, is not well understood in terms of gene regulation and intracellularcontrols. Using the rice HL-type cytoplasmic male sterile (HL-CMS) line with starch deficiency in pollen and two related male-fertile lines (Oryza sativa L.), we have studied two specific developmental stages: 'early', that is, before or during pollen mitosis I, without starch accumulation, and 'late', the actively starch-filling postpollen mitosis I phase. Real-time reverse transcription-PCR analysis showed temporal changes in the expression patterns of six metabolic genes involved in the sucrose-starch pathway, i.e. sucrose synthase (EC 2.4.1.13), Rsus3; acid invertase (EC 3.2.1.26), OSINV2; phosphoglucomutase (EC 5.4.2.2), OsPGM; uridine diphosphate-glucose pyrophosphorylase (EC 2.7.7.9), OsUGP, a subunit of adenosine diphosphate-glucose pyrophosphorylase (AGPase) (EC 2.2.7.2), OsAGPL3; soluble starch synthase (EC 2.4.1.21), OsSSI and three transporter genes [sucrose transporter, OsSUT3; hexose transporter, OSMST7 and plasma membrane H + -adenosine triphosphatase (ATPase) (EC 3.6.1.35), OSA2]. Regarding metabolic enzyme activities, lower activity of sucrose synthase was detected at 'late' compared with 'early' stage in the three rice lines; invertase and other downstream enzymes had higher activities at 'late' stage in male-fertile but not male-sterile lines. Meanwhile, significantly reduced sugars and ATP levels were detected at 'early' and 'late' stages in male-sterile compared with male-fertile tissues. Furthermore, transgenic data showed that AGPase gene can stimulate starch accumulation in pollen of male-sterile host lines, showing the importance of AGPase in the metabolic pathway. Taken together, these data suggest that the combined effects of the reduced energy supply and sugar levels in starch biosynthesis may lead to the observed temporal changes in gene expression, and eventually to pollen sterility.

Honglian (HL) is one of the three major types of cytoplasmic male sterility (CMS) of rice (Oryza sativa L) that has been commercially used in hybrid production. In previous studies, the CMS in HL is shown to be associated with a chimeric gene orfH79 that is cotranscribed with an extra atp6 in mitochondria. This study demonstrated that the intact F0F1-ATPase in HL CMS line was specifically reduced in both of its protein quantity and enzyme activity, whereas its F1 sector was not affected. It implies that the F0 sector presents a labile linkage with F1. In the presence of fertility restorer gene, F0F1-ATPase can be recovered. Furthermore, orfH79 transcripts were preferentially polyadenylated, and consequently degraded rapidly in florets of the restored hybrid plants, indicating that the atp6-orfH79 is involved in the sterile phenotype. With inhibition of cytochrome pathway of electron transfer chain, the biomass of the sterile plants grown in dark was significantly lower than that of the fertile lines. However, the respiration measurements showed an increase in the electron transferring capacity in the sterile plants, suggesting that the reduction of biomass in sterile line was caused by the disruption of F0F1-ATPase.

e-Poly-L-lysine (e-PL)(2) is widely used as an antibacterial agent because of its broad antimicrobial spectrum. However, the mechanism of e-PL against pathogens at the molecular level has not been elucidated. This study investigated the antibacterial activity and mechanism of e-PL against Escherichia coli O157:H7 CMCC44828. Propidium monoazide-PCR test results indicated that the threshold condition of e-PL for complete membrane lysis of E. coli O157:H7 was 10 μg/mL (90% mortality for 5 μg/mL). Further verification of the destructive effect of e-PL on cell structure was performed by atomic force microscopy and transmission electron microscopy. Results showed a positive correlation between reactive oxygen species (ROS)(3) levels and e-PL concentration in E. coli O157:H7 cells. Moreover, the mortality of E. coli O157:H7 was reduced when antioxidant N-acetylcysteine was added. Results from real-time quantitative PCR (RT-qPCR)(4) indicated that the expression levels of oxidative stress genes sodA and oxyR were up-regulated 4- and 16-fold, respectively, whereas virulence genes eaeA and espA were down-regulated after e-PL treatment. Expression of DNA damage response (SOS response)(5) regulon genes recA and lexA were also affected by e-PL. In conclusion, the antibacterial mechanism of e-PL against E. coli O157:H7 may be attributed to disturbance on membrane integrity, oxidative stress by ROS, and effects on various gene expressions, such as regulation of oxidative stress, SOS response, and changes in virulence.

We developed a rapid and reliable technique for simultaneous detection of Salmonella Typhimurium, Escherichia coli O157:H7 and Listeria monocytogenes that can be used in food products. Magnetic nano-beads (MNBs) based immunomagnetic separation (IMS) was used to separate the target bacterial cells while multiplex PCR (mPCR) was used to amplify the target genes. To detect only the viable bacteria, propidium monoazide (PMA) was applied to selectively suppress the DNA detection from dead cells. The results showed the detection limit of IMS-PMA-mPCR assay was about 10 2 CFU/ml (1.2 × 10 2 CFU/ml for S . Typhimurium, 4.0 × 10 2 CFU/ml for E . coli O157:H7 and 5.4 × 10 2 CFU/ml for L . monocytogenes ) in pure culture and 10 3 CFU/g (5.1 × 10 3 CFU/g for S . Typhimurium, 7.5 × 10 3 CFU/g for E . coli O157:H7 and 8.4 × 10 3 CFU/g for L . monocytogenes ) in spiking food products samples (lettuce, tomato and ground beef). This report has demonstrated for the first time, the effective use of rapid and reliable IMS combined with PMA treatment and mPCR assay for simultaneous detection of viable S . Typhimurium, E . coli O157:H7 and L . monocytogenes in spiked food samples. It is anticipated that the present approach will be applicable to simultaneous detection of the three target microorganisms for practical use.

Deoxynivalenol (DON) poses a health risk to animals and humans consuming contaminated food and feed. Currently, no effective method had been established to detoxify DON. Obtaining strains for the detoxification of DON from probiotic strains and ascertaining the mechanisms of detoxification will be helpful for assuring food safety in food processing. In this research, two Bacillus strains possessing the capability of detoxifying DON were picked out from 59 probiotics by using a two-run screening. The strategy of adjusting pH and eightfold dilution was successfully applied to eliminate the interferences caused by MRS and nutrition broth in DON determination in competitive indirect enzyme-linked immunosorbent assay (ELISA). The capability of detoxification reached a high level when the spent culture supernatant of these two Bacillus were incubated at 37C, 180 rpm for 12 h, the detoxification rate were >98% and 71.4%, respectively. The rate presented a sharp decrease when the heating temperature changed from 65–75C. The detoxification potential was completely lost, when the heating temperature was 100C, whereas the viable cells were demonstrated as having no detoxification potential. The mechanism for detoxification of DON is related to some thermosensitive substances in the SCS of these two Bacillus. PRACTICAL APPLICATIONS Twenty-five percent of the global grains were contaminated by mycotoxins, resulting in the problem of quality and safety in food and feeds and also great economic loss. As one of the most popular mycotoxins in maize and wheat, deoxynivalenol attracted few many detoxification approaches. In our study, we have found that two probiotic strains of Bacillus presented stronger detoxification ability of deoxynivalenol, therefore they could be applied in the fermentation process of feedstuffs and fermented food. Furthermore, our research provided a protocol for searching the potential bacteria, which could be used for the detoxification of other mytoxins, e.g., aflatoxin B1, ochratoxin A, zearalenone, etc.

Quantum dots (QDs) have received considerable attention due to their advantages and are widely used in biological studies, especially for multiplexed staining assays and immunological assays. Here we report an easy method for quantum dot synthesis and encapsulation, and use for efficient bioconjugation with secondary antibody. For the application of QDs-antibody conjugates, we approached a fast dot blotting immunological assay, and then we used a breast cancer cell line SKBR3 and anti-EpCAM antibody as the primary antibody for the cell imaging. The results shows that this conjugates can be successfully used in immunological assays such as dot blotting and cell imaging, and with the help of certain blocking buffers, specific and sensitive signals can be obtained.

To study the antibiotic susceptibility of eleven vaginal probiotic lactobacilli, the disc diffusion method in LAPTg agars was performed. The results had shown that all strains were resistance to tobramycin, lomefloxacin, nalidixic scid, ofloxacin, enoxacin, ciprofloxacin, norfloxacin and fleroxacin. Thereof eight were resistance to sparfloxacin, six were resistance to gentamicin and netilmicin. All the lactobacilli were sensitive to -lactamade inhibitors and macrolides whereas resistant to tinidazole. Therefore, the susceptibility to some antibiotics presented similar. On the other hand, the sensitivity to other antibiotics depended on strains. Our results indicated that resistant strains of lactobacilli against tinidazole could be combinated with tinidazole for the restoration of the normal urogenital flora and curing of urinary tract infections or bacterial vaginosis.

One of the cytoplasmic male sterility (CMS) types used for hybrid rice (Oryza sativa L.) production in China is the Honglian (HL)-CMS. Previous studies suggested that pollen abortion of the sterile plants was resulted from a special programmed cell death (PCD) program started at meiosis in the microspores. To elucidate the molecular basis of the pollen abortion, we compared the biochemical and physiological properties such as content of reactive oxygen species (ROS), ATP, NADH, total glutathione and ascorbate acid, the activities of dehydroascrbate reductase, glutathione reductase, ascorbate peroxides and superoxide dismutase, and the integrity of mitochondrial genome DNA isolated from an HL-CMS line, Yuetai A and its maintainer line, Yuetai B. Our results indicated that the mitochondria of the HL-CMS line suffered from a serious oxidative stress during microspores development. Oxidative stress induced by abnormal increased ROS at meiosis stage resulted in the depletion of ATP and NADH, and the degradation of mitochondrial genomic DNA. This suggests that the presence of redox signal originated in mitochondria affects the rest of the cell. Therefore, it is possible that the abortion of premature microspores in HL-CMS line is induced by the chronic oxidative stress in mitochondria in the early stage of pollen development.