Current Microbiology

Ye, Julian; Luo, Yun; Fang, Weijia; Pan, Junhang; Zhang, Zheng; Zhang, Yanjun; Chen, Zhiping; Jin, Dazhi

doi: 10.1007/s00284-014-0752-zpmid: 25510171

The pathogenic mechanism of Vibrio cholerae manifests as diarrhea and causes life-threatening dehydration. Here, we observe the human intestinal epithelialcells (HIEC) response to Cholera toxin (CT) by a real-time cell analysis (RTCA) platform, and disclose the difference from CT-induced cytotoxicity and others in HIEC. An HIEC cell of 1.0 × 105 cells/mL was characterized as the suitable concentration for each well. For experimentation, the assay requires an inoculation of CT dissolved in Dulbecco’s phosphate-buffered saline with 0.1 % gelatin for a period of 18–25 h. The dimensionless impedance cell index curve presented characteristic dose- and time-dependent drop responses at the first stage, and the CT-induced cytotoxicity was the most remarkable following exposure for 18–25 h (P = 0.0002). Following the obvious cytotoxic reaction, the CI curve gradually increased over time until the original CI value, indicating that self-recovery occurred. The CT-induced CI curve for HIEC was different from that induced by other toxins, including diphtheria and Clostridium difficile toxin. Collectively, these results suggest that the CT-induced cytotoxicity in HIEC was absolutely different from that induced by C. difficile and other toxins because of the different pathogeneses that were correlated with the specific CI curve generated by the RTCA system. In summary, our data show that the assay described here is a convenient and rapid high-throughput tool for real-time monitoring of host cellular responses to CT on the basis of the characteristic CI curve.

Houda, Nasr; Abdelwaheb, Chatti; Asma, Ben; Ines, Mehri; Ahmed, Landoulsi; Abdennaceur, Hassen

doi: 10.1007/s00284-014-0756-8pmid: 25561403

In this study, the effect of operational conditions on biofilm development and nitrification in moving-bed biofilm reactor (MBBR) was investigated. The reactor was operated in a continuously fed regime during 170 days and with theoretical hydraulic retention time of 7 h, respectively. The presence of chemical oxygen demand (COD) increased the time required to form stable nitrifying. Subsequent stepwise increase of influent COD caused an increment in total polysaccharide (PS) and protein (PN) content, which was accompanied by an attachment of the biofilm, as shown by atomic force microscope (AFM). PS and PN concentrations proved to be good indicators of biomass development and attachment in MBBR system. Reactor was operated and water quality was characterized before and after treatment. Parameters including pH, 5-day biochemical oxygen demand (BOD5), total suspended solids (TSS) (COD), PN, PS, and fecal bacteria in both raw and treated wastewater were monitored during the treatment. The removal rates of ammonium-nitrogen (NH4 +-N), BOD5, COD, and TSS are 95, 67.5, 69.2, and 73.33 %, respectively. The average bacterial reduction between the inlet and the outlet was of the order of 5 ± 1 logarithmic units for fecal coliforms. AFM showed that distinct biofilm and extracellular polymeric substances were formed in biofilm was thicker in the 70 days than in the 30 days. These results showed that the consumption rate for each substrate increased parabolically with biofilm thickness due to the increased amount of biomass Thus, MBBR can serve as a promising technology for wastewater treatment and can be scaled up for small communities in the developing countries.

Paul, Sangeeta; Aggarwal, Chetana; Thakur, Jyoti; Bandeppa, G.; Khan, Md.; Pearson, Lauren; Babnigg, Gyorgy; Giometti, Carol; Joachimiak, Andrzej

doi: 10.1007/s00284-014-0761-ypmid: 25561404

Bacillus licheniformis strain SSA 61, originally isolated from Sambhar salt lake, was observed to grow even in the presence of 25 % salt stress. Osmoadaptive mechanisms of this halotolerant B. licheniformis strain SSA 61, for long-term survival and growth under salt stress, were determined. Proline was the preferentially accumulated compatible osmolyte. There was also increased accumulation of antioxidants ascorbic acid and glutathione. Among the different antioxidative enzymes assayed, superoxide dismutase played the most crucial role in defense against salt-induced stress in the organism. Adaptation to stress by the organism involved modulation of cellular physiology at various levels. There was enhanced expression of known proteins playing essential roles in stress adaptation, such as chaperones DnaK and GroEL, and general stress protein YfkM and polynucleotide phosphorylase/polyadenylase. Proteins involved in amino acid biosynthetic pathway, ribosome structure, and peptide elongation were also overexpressed. Salt stress-induced modulation of expression of enzymes involved in carbon metabolism was observed. There was up-regulation of a number of enzymes involved in generation of NADH and NADPH, indicating increased cellular demand for both energy and reducing power.

Yu, Yang; Yang, Zijun; Guo, Kai; Li, Zhe; Zhou, Hongzi; Wei, Yanli; Li, Jishun; Zhang, Xinjian; Harvey, Paul; Yang, Hetong

doi: 10.1007/s00284-014-0764-8pmid: 25561405

Trichoderma harzianum is an important commercial biocontrol fungal agent. The temperature has been shown to be an important parameter and strain-specific to the mycelia growth of fungi, but less report makes the known of the mechanisms in T. harzianum. In our study, a 6-h treatment of heat increased the thiobarbituric acid reactive substances (TBARS) and nitric oxide (NO) concentration in mycelia to 212 and 230 % the level of the control, respectively. The exogenous NO donor sodium nitroprusside (150 μM) reduced the TBARS concentration to 53 % of that under heat stress (HS). At the same time, the NO-specific scavenger at 250 μM, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-1-oxyl-3-oxide, prevented the exogenous NO-relieved TBARS accumulation under HS. The increased NO concentration under HS was reduced 41 % by the NO synthase (NOS) inhibitor L-NG-nitroarginine methyl ester, but not the nitrate reductase (NR) inhibitor tungstate. Our study exhibited that NO can protect the mycelia of T. harzianum from HS and reduce the oxidative damage by enhancing the activity of NOS and NR.

Li, Guohui; Zhou, Qian; Hu, Zhaoyang; Wang, Peng; Tang, Qi; Chen, Keping; Yao, Qin

doi: 10.1007/s00284-014-0765-7pmid: 25561406

Bombyx mori bidensovirus (BmBDV) VD1-ORF4 consists of 3,318 nucleotides, which codes for a predicted protein with molecular weight of about 127 kDa. However, the authentic proteins encoded by VD1-ORF4 in silkworm midguts infected with BmBDV and their interacting proteins are still unclear. In this study, Western blot analysis revealed that a 127-kDa protein was confirmed to be translated from the VD1-ORF4 transcript using polyclonal antibodies and monoclonal antibodies against VD1-ORF4 deduced amino acid. Moreover, four smaller proteins with molecular weight of about 70, 60, 53, and 42 kDa were also examined in the infected midguts. Transient expression assay indicated that the expression amount of VD1-ORF4 fused with egfp was at least 30-fold lower than that of egfp gene, and immunofluorescence staining result indicated that these proteins encoded by VD1-ORF4 were located in both the cytoplasm and nucleus. Co-immunoprecipitation result showed that Aminopeptidase and Heat shock protein 90 can be captured by these proteins encoded by VD1-ORF4. In conclusion, multiple proteins were produced from the transcripts of VD1-ORF4 gene by an uncertain expression strategy, which may play important roles in viral replication and assembly.