NPM promotes hepatotoxin-induced fibrosis by inhibiting ROS-induced apoptosis of hepatic stellate cells and upregulating lncMIAT-induced TGF-ß2.

Liver fibrosis is caused by a variety of chronic liver injuries and has caused significant morbidity and mortality in the world with increasing tendency. Elucidation of the molecular mechanism of liver fibrosis is the basis for intervention of this pathological process and drug development. Nucleophosmin (NPM) is a widely expressed nucleolar phosphorylated protein, which is particularly important for cell proliferation, differentiation and survival. The biological role of NPM in liver fibrosis remains unknown. Here we show that NPM promotes liver fibrosis through multiple pathways. Our study found that NPM was up-regulated in cirrhosis tissues and activated in hepatic stellate cells (HSCs). NPM inhibition reduced liver fibrosis markers expression in HSCs and inhibited the HSCs proliferation and migration. In mice model, NPM knockdown in HSCs or application of specific NPM inhibitor can remarkably attenuate hepatic fibrosis. Mechanistic analysis showed that NPM promotes hepatic fibrosis by inhibiting HSCs apoptosis through Akt/ROS pathway and by upregulating TGF-ß2 through Akt-induced lncMIAT. LncMIAT up-regulated TGF-ß2 mRNA by competitively sponging miR-16-5p. In response to liver injury, hepatocytes, Kupffer cells and HSCs up-regulated NPM to increase TGF-ß2 secretion to activate HSCs in a paracrine or autocrine manner, leading to increased liver fibrosis. Our study demonstrated that NPM regulated hepatotoxin-induced fibrosis through Akt/ROS-induced apoptosis of HSCs and via the Akt/lncMIAT-up-regulated TGF-ß2. Inhibition of NPM or application of NPM inhibitor CIGB300 remarkably attenuated liver fibrosis. NPM serves a potential new drug target for liver fibrosis.

Genomic Epidemiology of Imported Cases of COVID-19 in Guangdong Province, China, October 2020 - May 2021.

Objective: The pandemic of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been engendering enormous hazards to the world. We obtained the complete genome sequences of SARS-CoV-2 from imported cases admitted to the Guangzhou Eighth People's Hospital, which was appointed by the Guangdong provincial government to treat coronavirus disease 2019 (COVID-19). The SARS-CoV-2 diversity was analyzed, and the mutation characteristics, time, and regional trend of variant emergence were evaluated. Methods: In total, 177 throat swab samples were obtained from COVID-19 patients (from October 2020 to May 2021). High-throughput sequencing technology was used to detect the viral sequences of patients infected with SARS-CoV-2. Phylogenetic and molecular evolutionary analyses were used to evaluate the mutation characteristics and the time and regional trends of variants. Results: We observed that the imported cases mainly occurred after January 2021, peaking in May 2021, with the highest proportion observed from cases originating from the United States. The main lineages were found in Europe, Africa, and North America, and B.1.1.7 and B.1.351 were the two major sublineages. Sublineage B.1.618 was the Asian lineage (Indian) found in this study, and B.1.1.228 was not included in the lineage list of the Pangolin web. A reasonably high homology was observed among all samples. The total frequency of mutations showed that the open reading frame 1a (ORF1a) protein had the highest mutation density at the nucleotide level, and the D614G mutation in the spike protein was the commonest at the amino acid level. Most importantly, we identified some amino acid mutations in positions S, ORF7b, and ORF9b, and they have neither been reported on the Global Initiative of Sharing All Influenza Data nor published in PubMed among all missense mutations. Conclusion: These results suggested the diversity of lineages and sublineages and the high homology at the amino acid level among imported cases infected with SARS-CoV-2 in Guangdong Province, China.

Occurrence and dietary exposure assessment of pentachlorophenol in livestock, poultry, and aquatic foods marketed in Guangdong Province, China: Based on food monitoring data from 2015 to 2018.

As a persistent organic pollutant, pentachlorophenol (PCP) has serious impacts on human health. However, its presence in animal source food products sold in the Guangdong Province (GD) of China, and the resultant dietary exposure have not been elucidated. To address this gap, 3,100 samples from seven food categories, including beef, pork, mutton, offals, broilers, hen eggs, and farmed freshwater fish, marketed throughout four geographical regions of GD, were collected from 2015 to 2018. Gas chromatography coupled with mass spectrometry was employed to detect PCP levels in these food matrices. PCP was found in all food categories, but the average contamination levels were low, ranging from 0.40 µg/kg wet weight (ww) (hen eggs) to 5.85 µg/kg ww (offals). However, higher concentrations of PCP were detected (P < 0.05) in animal source food from the North region. Additionally, a temporal declining trend was observed in this four-year consecutive survey. The estimated human dietary exposure of PCP to population groups, including the general population and subgroups (male and female, children, and adults), was found to be far below the permissible daily intake (3 µg/kg body weight). Therefore, the health impacts of PCP should be correspondingly low for local residents, based on current toxicological knowledge. Regional exposure patterns varied due to different extents of contamination in the four areas, and pork, broilers, and freshwater fish were the major sources of dietary PCP exposure. PRACTICAL APPLICATION: As a persistent organic pollutant, pentachlorophenol (PCP) has serious impacts on human health. However, its presence in animal source food products sold in Guangdong Province of China, and the resultant dietary exposure have not been elucidated. In this study, we conducted an in-depth investigation on the occurrence of PCP in major foodstuff categories, including beef, pork, mutton, broilers, offals, hen eggs, and farmed freshwater fish, marketed in all 21 prefecture-level divisions of Guangdong Province, in order to provide integral insights for regulatory authorities.

Heterogeneous nuclear ribonucleoprotein A2/B1 is a negative regulator of human breast cancer metastasis by maintaining the balance of multiple genes and pathways.

BACKGROUND: Heterogeneous nuclear ribonucleoprotein (hnRNP) A2/B1 is an important RNA-binding protein that affects the RNA processing, splicing, transport and stability of many genes. hnRNPA2/B1 is expressed during proliferation and metastasis of various cancer types and promotes such processes. However, the precise role and mechanism of hnRNPA2/B1 in breast cancer remain unclear. METHODS: The association of hnRNPA2/B1 with breast cancer metastasis was assessed using tissue chips, mouse models and publicly available data. The role and mechanism of hnRNPA2/B1 in breast cancer metastasis were studied in cell lines and mouse models. FINDINGS: In contrast to other cancer research findings, hnRNPA2/B1 expression was negatively correlated with breast cancer metastasis. hnRNPA2/B1 inhibited MDA-MB-231 triple-negative breast cancer (TNBC) cell metastasis in vitro and in vivo. hnRNPA2/B1 knockout activated ERK-MAPK/Twist and GR-beta/TCF4 pathways but inhibited STAT3 and WNT/TCF4 signalling pathways. Profilin 2 (PFN2) promoted breast cancer cell migration and invasion, whereas hnRNPA2/B1 bound directly to the UAGGG locus in the 3'-untranslated region of PFN2 mRNA and reduced the stability of PFN2 mRNA. INTERPRETATION: Our data supported the role of hnRNPA2/B1 in tumour metastasis risk and survival prediction in patients with breast cancer. The inhibitory role of hnRNPA2/B1 in metastasis was a balance of downstream multiple genes and signalling pathways. PFN2 downregulation by hnRNPA2/B1 might partly explain the inhibitory mechanism of hnRNPA2/B1 in breast cancer metastasis. Therefore, hnRNPA2/B1 might be used as a new prognostic biomarker and valuable molecular target for breast cancer treatments.

Levels, congener profiles, and dietary intake assessment of polychlorinated dibenzo-p-dioxins/dibenzofurans and dioxin-like polychlorinated biphenyls in beef, freshwater fish, and pork marketed in Guangdong Province, China.

Persistent organic pollutants such as polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and dioxin-like polychlorinated biphenyls (DL-PCBs) consisting of non-ortho and mono-ortho PCBs are suggested to be very hazardous and have adverse effects on human health. However, their levels and congener profiles in retail foods marketed in Guangdong Province of China have not been elucidated thus far. Thus, in this study, 226 individual samples of beef, freshwater fish, and pork marketed across four regions of Guangdong Province were randomly collected during 2013-2015 to determine their levels of PCDD/Fs and DL-PCBs. The results showed that the total toxic equivalency quantities (TEQs) of most samples were below the maximum limits except for the 26 samples collected from the vicinities of pollution areas. The median total TEQs of these three categories were 0.174, 0.488, and 0.113pgTEQ/g fw, respectively, which indicated that the contamination status of the studied foods was not serious. For congener profiles, significantly different patterns were observed in three food groups, but with the same major TEQ contributors being 2,3,4,7,8-PeCDF in beef, freshwater fish, and pork. Regional differences of congener profiles in each food group were also found in this study, which might be attributed to the regionally different distributions of PCDD/Fs and DL-PCBs in environment media. The dietary exposures of four population subgroups (girls, boys, male adults, and female adults) to PCDD/Fs and DL-PCBs via three food groups were estimated to assessed the potential risks. They were all lower than the provisional tolerable monthly intake (PTMI, 70pgTEQ/kgbw/month) established by Joint FAO/WHO Expert Committee on Food Additive. In these food categories, the exposure to PCDD/Fs and DL-PCBs via freshwater fish was the highest one, which accounted for about 20% of PTMI, indicating that it was the major route to expose dioxin compounds.

Insulin resistance in ischemic stroke.

Insulin resistance often refers to a pathological condition in which cells fail to respond to the normal actions of insulin. Increasing literature has noted a critical role of insulin resistance in the pathogenesis of ischemic stroke. Insulin resistance plays an important role in the pathogenesis of ischemic stroke via enhancing advanced changes of atherosclerosis. A variety of literature indicates that insulin resistance enhances platelet adhesion, activation and aggregation which are conducive to the occurrence of ischemic stroke. Insulin resistance also induces hemodynamic disturbances and contributes to the onset of ischemic stroke. In addition, insulin resistance may augment the role of the modifiable risk factors in ischemic stroke and induce the occurrence of ischemic stroke. Preclinical and clinical studies have supported that improving insulin resistance may be an effective measure to prevent or delay ischemic stroke.

Ephrin-B2/Fc promotes proliferation and migration, and suppresses apoptosis in human umbilical vein endothelial cells.

Tumor growth and metastasis are angiogenesis dependent. Angiogenic growth involves endothelial cell proliferation, migration, and invasion. Ephrin-B2 is a ligand for Eph receptor tyrosine kinases and is an important mediator in vascular endothelial growth factor-mediated angiogenesis. However, research offer controversial information regarding effects of ephrin-B2 on vascular endothelial cells. In this paper, proteome analyses showed that ephrin-B2/Fc significantly activates multiple signaling pathways related to cell proliferation, survival, and migration and suppresses apoptosis and cell death. Cytological experiments further confirm that ephrin-B2/Fc stimulates endothelial cell proliferation, triggers dose-dependent migration, and suppresses cell apoptosis. Results demonstrate that soluble dose-dependent ephrinB2 can promote proliferation and migration and inhibit apoptosis of human umbilical vein endothelial cells. These results also suggest that ephrinB2 prevents ischemic disease and can potentially be a new therapeutic target for treating angiogenesis-related diseases and tumors.

Nucleophosmin Regulates Intracellular Oxidative Stress Homeostasis via Antioxidant PRDX6.

Reactive oxygen species (ROS) play both deleterious and beneficial roles in cancer cells. Nucleophosmin (NPM) is heavily implicated in cancers of diverse origins, being its gene over-expression in solid tumors or frequent mutations in hematological malignancies. However, the role and regulatory mechanism of NPM in oxidative stress are unclear. Here, we found that NPM regulated the expression of peroxiredoxin 6 (PRDX6), a member of thiol-specific antioxidant protein family, consequently affected the level and distribution of ROS. Our data indicated that NPM knockdown caused the increase of ROS and its relocation from cytoplasm to nucleoplasm. In contrast, overexpression or cytoplasmic localization of NPM upregulated PRDX6, and decreased ROS. In addition, NPM knockdown decreased peroxiredoxin family proteins, including PRDX1, PRDX4, and PRDX6. Co-immunoprecipitation further confirmed the interaction between PRDX6 and NPM. Moreover, NSC348884, an inhibitor specifically targeting NPM oligomerization, decreased PRDX6 and significantly upregulated ROS. These observations demonstrated that the expression and localization of NPM affected the homeostatic balance of oxidative stress in tumor cells via PRDX6 protein. The regulation axis of NPM/PRDX/ROS may provide a novel therapeutic target for cancer treatment. J. Cell. Biochem. 118: 4697-4707, 2017. © 2017 Wiley Periodicals, Inc.

Relocation of NPM Affects the Malignant Phenotypes of Hepatoma SMMC-7721 Cells.

Nucleophosmin(NPM), heavily implicated in diverse solid tumors, is an important multifunctional protein mainly located in the nucleolus. Our previous study confirmed that NPM can also localize and accumulate in the cytoplasm of liver cancer cells. However, the role of cytoplasmic NPM (NPMc +) is unclear. Here, we showed that both nucleolar NPM and NPMc+ could promote cell proliferation, although the effect of NPMc+ was weaker than that of NPM. Cell adhesion ability of hepatoma cells was significantly reduced to a greater extent by NPMc+ expression. Nucleolar NPM enhanced cell migration and invasion, whereas NPMc+ impeded cell migration and invasion. The investigation of NPM interactional proteins by proteomic method demonstrated that the NPM was involved in multiple biological processes. By contrast, the interactional proteins of NPMc+ were mainly implicated in tRNA amino acylation regulation. The interactional network of NPMc+ was significantly small and simple. These results suggested that relocation of NPM altered its interactional network and consequently disturbed the primary functions, including cell proliferation, adhesion, migration, and invasion. NPM plays a promotional role in cancer and the reducing relocation may be a potential therapeutic target for hepatocellular carcinoma. J. Cell. Biochem. 118: 3225-3236, 2017. © 2017 Wiley Periodicals, Inc.

Gastroenteritis outbreaks caused by Norovirus genotype II.7 in a college in China (Zhuhai, Guangdong) in 2011.

BACKGROUND: Noroviruses are the most common cause of epidemic gastroenteritis worldwide. Noroviruses are comprised of at least five genogroups (GI-GV) and >35 genotypes. GII.7 is a nonpredominant genotype associated with Norovirus outbreaks. On November 17, 2011, Zhuhai Center for Disease Control monitored an increasing number of gastroenteritis cases at a local college. To determine the causes and control the outbreak effectively, we carried out an epidemiologic investigation. METHODS: Suspected cases were defined as those with one of the following symptoms: nausea, vomiting, abdominal pain, or diarrhea presenting on or after November 15 among the people who lived at the college. Probable cases were defined as cases with vomiting or diarrhea over three times per day on or after November 15. Confirmed cases were suspected or probable cases positive for Norovirus (nucleic acid). We conducted a case-control study to identify risk factors of the outbreak. Norovirus was tested by real-time reverse transcription-polymerase chain reaction, and Norovirus polymerase chain reaction products were further sequenced. RESULTS: In total, 63 cases were identified, which were scattered in all 14 departments of the college. The outbreak lasted for 84 h. Time distribution mode indicated a point-source outbreak. Fifty-one cases and 94 controls were contacted. Seventy-five percent of the cases compared to 19% of the controls were exposed to delicatessens (various salad and meat products) from the "Y" convenience store (odds ratio=12, 95% confidence interval 5.4-28). Laboratory tests showed 14 of the 15 cases and two asymptomatic food handlers were positive for Norovirus nucleic acid. There was 100% similarity between the cases and the food handlers when we compared the nucleotide sequences of Norovirus, which clustered with GII.7 genotype. CONCLUSIONS: Delicatessens from the "Y" convenience store were associated with the GII.7 Norovirus outbreak. We strongly recommend food supervision and quality control in convenience stores to decrease the risk of future Norovirus outbreaks.

[Surveillance and analysis on the pathogenic features of Salmonella in Guangdong province in 2010].

OBJECTIVE: In order to better understand the nature of Salmonella infection in diarrheal patients in Guangdong province, the study analyzed the serum types, antibiotic resistance and molecular determinants of the isolated Salmonella strains. METHODS: In year 2010, 8405 diarrhea patients from 16 surveillant hospital in Guangzhou, Zhongshan, Dongguan, Zhuhai, Maoming, Yangjiang and Jiangmen cities in Guangdong province, were recruited in the study. A total of 8405 fecal specimen were collected and subjected to Salmonella isolation and culture. The isolated Salmonella strains were further analyzed via serotyping, antimicrobial susceptibility testing, and PFGE. The χ(2) test was applied to compare the differences between the isolated Salmonella strains in different seasons and districts. BioNumerics software was used to analyze the PFGE results in order to determine the correlation between different Salmonella strains. RESULTS: The positive rate of the surveillant Salmonella in Guangdong province was 3.58% (301/8405) in 2010; with the gender ratio at 1.34:1 (166/124). Salmonella infection was found in all age groups, and most in infants, accounting for 57.48% (173/301). The isolated rates of Salmonella were separately 3.48% (61/1751), 4.97% (134/2695), 3.08% (73/2370) and 2.08% (33/1589) in the four seasons; and the difference was statistically significant (χ(2) = 27.29, P < 0.01). The isolated rates of Salmonella in different regions were as follows: Zhuhai 15.43% (25/162), Maoming 7.53% (18/239), Dongguan 6.51% (39/599), Yangjiang 3.64% (14/385), Zhongshan 3.03% (70/2309), Guangzhou 2.90% (126/4349) and Jiangmen 2.49% (9/362). The difference between regions was statistically significant (χ(2) = 100.75, P < 0.01). Except one strain of the isolated Salmonella cannot be serotyped, the other 300 strains were divided into 42 serotypes, of which Salmonella typhimurium and Salmonella enteritidis were dominant, account for 45.18% (136/301) and 10.96% (33/301) respectively. Although over 85% of Salmonella were sensitive to cephalosporin, ACSSuT resistance patterns (defined as resistance to at least ampicillin, chloramphenicol, streptomycin, sulfamethoxazole and tetracycline) reached 34.88% (105/301), the highest resistant rate was found in serotype Salmonella typhimurium, as high as 65.44% (89/136). 136 strains of Salmonella typhimurium were divided into 51 PFGE types, showed great genetic diversity. 33 strains of Salmonella enteritidis were divided into 18 PFGE types. The strains with same PFGE pattern may have different drug-resistant patterns, and vice versa. CONCLUSION: Salmonella typhimurium and Salmonella enteritidis were the dominant serotypes causing infectious diarrhea in Guangdong province. Cephalosporin was the primary choice in clinical medicine. However, Salmonella typhimurium was resistant to drug most seriously in Guangdong province. There was no significant correlation between Salmonella resistance patterns and PFGE type.

Case-control and family-based association studies of novel susceptibility locus 8q24 in nonsyndromic cleft lip with or without cleft palate in a Southern Han Chinese population located in Guangdong Province.

Nonsyndromic cleft lip with or without cleft palate (NSCL/P) is one of the most common congenital malformations and a susceptibility locus on chromosome 8q24 has been replicated as a genetic risk factor for NSCL/P in patients of European and Asian descent. However, given considerable variations in allele frequencies across geographical regions studied, the aim of this study was to investigate the association of rs987525 located at 8q24 with NSCL/P only among the southern Han Chinese population from Guangdong province. We recruited 216 NSCL/P cases, their parents, and 200 controls to conduct case-control analysis and family-based association studies. Genotyping of rs987525 was carried out by the matrix assisted laser desorption ionization-time of flight mass spectrometry method. Case-control analysis showed allele and genotype distributions for rs987525 were not significantly associated with the risk of NSCL/P in our study population. Similar results were found when all cases were stratified into cleft lip only and cleft lip with cleft palate. A transmission disequilibrium test showed no statistically significant transmission of A nor C alleles and family-based association test (FBAT) analysis provided no evidence of NSCL/P risk with single markers. These results do not provide evidence for an association between rs987525 at 8q24 and the risk of NSCL/P in the southern Han Chinese population from Guangdong province.

[Surveillance on Salmonella infection in Guangdong province, 2008 - 2009].

OBJECTIVE: To understand the infection of Salmonella (S.) in patients with diarrhea and outbreaks caused by Salmonella to identify the serotypes, resistance to antibiotics and PFGE types of the strains from the surveillance program in Guangdong province. METHODS: S. strains from patients with diarrhea were detected, and all the positive strains collected in routine and outbreak surveillance programs, were tested by serum agglutination, antibiotic susceptibility and PFGE. RESULTS: 71 S. strains were isolated from 1922 stool samples in 2008, with positive rate as 3.7%. 85 S. strains were isolated from 2110 stool samples in 2009, with positive rate as 4.0%. All the 156 strains were divided into 37 serotypes, with S. serotype typhimurium and enteritidis as the most common serotypes. 10 incidents of food poisoning were detected, of which 4 were caused by enteritidis and 3 by typhimurium. A suspected outbreak by enteritidis was discovered and under epidemiological investigation. The findings indicated that 2 of the 4 patients from this outbreak were infected with identical enteritidis isolates. 80% of the 229 isolates were found susceptible to cephalosporins and quinolone and 59.3% of them were multiresistant to the antibiotics. CONCLUSION: S. enteritidis and S. typhimurium were the most common serotypes that caused infectious diarrhoea and food poisoning in Guangdong province.

[Automated ribotyping of Salmonella and Staphylococcus aureus in food poisoning of Guangdong province].

OBJECTIVE: To understand the genetic polymorphism of Salmonella and Staphylococcus aureus in Guangdong province, as well as to explore methods for identifying and tracing the source of these two foodborne pathogens. METHODS: Using the automated ribotyping system, two foodborne pathogens were tested with either EcoR I or Pvu II restriction enzymes. BioNumerics software was then applied for image analysis, database establishment and other corresponding analysis. RESULTS: Digestion of 32 Salmonella isolates with Pvu II yielded 19 different ribotypes, and digestion of 14 Salmonella isolates with EcoR I yielded 2 different ribotypes. Staphylococcus aureus isolates showed greater genetic diversity, whereas EcoR I digestion of 49 different isolates yielded 31 different ribotypes. CONCLUSION: Unique Salmonella and Staphylococcus aureus isolates could be identified through ribotyping. Although Salmonella serotyping and ribotyping were not strongly correlated, the combination of both restriction enzymes could be used to more effectively identify the genetic relationship among different strains as well as the source of food poisoning. Thus, not only could the genetic relationships amongst the different strains be inferred through ribotyping skills, the source of food poisoning and mode of transmission could also be determined under the use of this method.

[Surveillance program on and the distribution related to the virulence-associated genes of Vibrio cholerae in estuary of Pearl River].

OBJECTIVE: To understand the distribution, molecular characteristics and virulence genes of the O1 and O139 Vibrio cholerae isolates from the Pearl River Estuary water. METHODS: Vibrio cholerae isolates collected from the Pearl River estuary waters from January 2009 to December 2010, were tested by PCR for eight virulence-related genes, including cholera toxin (ctxA), zonula occludens toxin (zot), accessory cholera enterotoxin (ace), hemolysin (hlyA), toxin-coregulated pilus (tcpA), outer membrane protein (ompU), and the regulatory protein genes (tcpI, toxR). Genetic relation was assessed by pulsed-field gel electrophoresis (PFGE) and the patterns were clustered by BioNumerics. RESULTS: From 1152 aquatic samples, 69 isolates were identified, including 41 Inaba, 18 Ogawa and 10 O139. All the isolates showed ctxA negative, while the hlyA and toxR genes were positive in all the isolates. 34.15% (14/41) of the Inaba strains were hlyA(+) toxR(+) ompU(+) ace(+) zot(+) tcpI(+), while 66.67% (12/18) belonged to Ogawa strains and 70% (7/10) of the O139 strains were hlyA(+) toxR(+). Through PFGE analysis, the O1 isolates formed three clusters in this study. The patterns of O1 isolates differed widely, with the similarity as 72.8% - 100.0%, while the patterns of O139 isolates having the similarity of 69.9% - 95.5%. CONCLUSION: The non-toxigenic O1 and O139 V. cholerae had a wide distribution in the environment of Pearl River estuary water during the non-epidemic period of cholera. All the aquatic isolates presented diversities on the related virulent genes.

[Comparative study on the phenotypic characteristics and molecular typing of foodborne Vibrio parahaemolyticus in Guangdong province].

OBJECTIVE: To understand the phenotypic characteristics of foodborne Vibrio parahaemolyticus in Guangdong province through carrying out a comprehensive comparison including pulse field gel electrophoresis, ribotyping and serotyping. METHODS: 74 different Vibrio parahaemolyticus strains isolated from seafood and cases due to food poisoning in Guangdong province were under serotyping and susceptibility testing, in addition to the testing of direct heat hemolysin (tdh) and the heat hemolysin-related hemolysin hormone (trh) via PCR. Ribosomal genotyping (ribotyping) with EcoR I restriction enzyme was utilized on 74 different Vibrio parahaemolyticus isolates, whereas pulsed-field gel electrophoresis (PFGE) with the Not I restriction enzyme was used on 74 different Vibrio parahaemolyticus isolates. BioNumerics software was used to compare the isolates from different sources, times and places in order to elicit the correlation between different strains. RESULTS: Although Vibrio parahaemolyticus was 100.00% sensitive to chloramphenicol, it still presented different levels of resistance against 13 other antibiotics. Among the 74 different strains of Vibrio parahaemolyticus under testing, 24.32% showed positive for the tdh virulence gene, whereas 4.05% positive for trh. 74 different Vibrio parahaemolyticus strains were found to belong to 26 serotypes, where the O5:K17 and O2:K28 serotypes were dominant in those isolates that causing seafood-poisoning. The O3:K6 serotype was found to be the dominant of those isolates that causing food-poisoning. Based on ribosomal genotyping, the 74 Vibrio parahaemolyticus isolates were divided into 62 different ribotypes, whereas the 74 strains of Vibrio parahaemolyticus were divided into 67 different PFGE types, thus exhibiting considerable genetic diversities of the strains. CONCLUSION: Majority of the isolates causing food-poisoning carried tdh virulence gene. PFGE was shown to have the highest resolution, followed by ribotyping with serotyping being the lowest, where the combination of the three could improve the resolution.

Phenotypic and genotypic characterization Vibrio cholerae O139 of clinical and aquatic isolates in China.

To enhance the understanding of epidemiological impact of environmental Vibrio cholerae O139 strains, we characterized 10 clinical and 20 environmental isolates collected from human clinical samples and Pear River estuary during 2006 to 2008. Isolates were tested by PCR for eight virulence genes: cholera toxin (ctxA), zonula occludens toxin (zot), accessory cholera enterotoxin (ace), hemolysin (hlyA), NAG-specific heat-stable toxin (st), toxin-coregulated pilus (tcpA), outer membrane protein (ompU), and regulatory protein genes (tcpI). Genetic relatedness was assessed by pulsed-field gel electrophoresis (PFGE), and antibiotic susceptibility was determined using disk diffusion. Seven of eight virulence markers were detected in six clinical isolates and one environmental isolate. One clinical and one environmental isolate were positive for six virulence markers. 60% clinical isolates showed multi-drug resistance to tetracycline (TET), Nalidixic acid (NAL), chloramphenicol (CHL), and ampicillin (AMP), 70% were resistant to Trimethoprim + Sulfamethoxazole (SXT), while only 35% environmental strains were resistant to SXT. PFGE analysis revealed that the isolates in this study were formed three clusters. Cluster III was more related to strains from diarrheal patients than the strains in other clusters. Different from the clinical strains, most environmental strains lacked CTX and TCP gene clusters. Most environmental strains possess a single resistance profile, while most clinical isolates show multidrug resistant. PFGE analysis indicated the cluster III has more possibility to become a potential pathogenic clonal cluster.

[Development of a DNA microarray for detecting 8 common species of food-borne bacterial pathogens in south China].

OBJECTIVE: To prepare a DNA Microarray that can detect 8 common species of food borne bacterial pathogens in south China. METHODS: All the 70mer oligo probes were designed on the characteristic genome loci of the 8 species of food borne bacterial pathogens. Eight subarrays corresponding to the 8 food borne bacterial pathogens were spotted onto the slide and integrated into a pan-array on the chip. A number of identified and known bacterial samples from the storage bank were selected for the validation test. RESULTS: Based on the PPR ranking, for LM sub-array, the PPR of the 3 Listeria bacteria LM, Lin and Liv was 68.8%, 51.8% and 59.6%, respectively, while that of the non-Listeria bacterial samples was all below 43%. For VC sub-array, the PPR of VC sample was 54.1% and that of the non-VC bacterial samples was lower than 17.2%. For VP sub-array, the PPR was 66.7% for VP sample and below 24.2% for non-VP bacterial samples. For Sal sub-array, the PPR was 55.9% for Sal sample and below 50.5% for non-Sal bacterial samples. For Shi sub-array, the PPR of Shi sample and the non-Shi bacterial samples was 53.8% and below 36.6%, respectively. For SA sub-array, the PPR of SA sample and non-SA bacterial samples was 65.2% and below 22.7%, respectively. For CJ sub-array, the PPR of the 2 Campylobacter bacteria CJ and CC were 88.2% and 58.8%, respectively, and that of the non-Campylobacter bacterial samples was lower than 35.3%. For EC sub-array, the PPR of EC sample was 47.9%, and that of the non-EC bacterial samples was lower than 41.6%. Evaluation of the Biosafood-8 chip developed in this study by 18 biological samples from different origins demonstrated its good specificity and accuracy in the identification of the pathogens. CONCLUSION: The chip we developed can clearly differentiate the target food borne pathogenic bacteria and non-target bacteria and allows specific and accurate identification of the species of the tested bacteria isolates.

[Construction of recombinant adenovirus containing TK gene and its effect against human liver cancer cells].

OBJECTIVE: To construct a replication-defective adenovirus containing TK gene and investigate the killing effects of TK gene against human liver cancer cells SMMC-7721. METHODS: The recombinant adenovirus ADV-TK was constructed using homologous recombination in the cells. SMMC-7721 cells transfected with recombined adenovirus were exposed to GCV, and the cell viability was measured by MTT assays. RESULTS: The recombinant adenovirus containing TK gene was successfully constructed. Transfection by the recombinant adenovirus ADV-TK and GCV exposure significantly suppressed the growth of SMMC-7721 cells. CONCLUSION: A replication-defective adenovirus containing TK gene has been successfully constructed, and in combination with GCV, the recombinant adenovirus produces significant killing effect against SMMC-7721 cells in vitro.