Prevalence and distribution of acute gastrointestinal illness in the community of China: a population-based face-to-face survey, 2014-2015.

BACKGROUND: The true incidence of acute gastrointestinal illness in China is underrecognized by surveillance systems. The aims of this study were to estimate the incidence and prevalence of self-reported AGI in the community of China, and to investigate sociodemographic and epidemiological determinants of AGI. METHODS: We conducted a 12-months cross-sectional population-based survey in eight provinces of China during 2014-2015. The survey determined the prevalence and incidence of acute gastrointestinal illness (AGI) in the total permanent resident population in China according to the census of the population in 2010. The random multilevel population sample was stratified by geographic, population, and socioeconomic status. We used a recommended case definition of AGI, with diarrhea (three loose or watery stools) and/or any vomiting in a four-week recall. A face-to-face survey was conducted by selecting the member in the household with the most recent birthday. RESULTS: Among 56,704 sampled individuals, 948 (1,134 person-time) fulfilled the case definition; 98.5% reported diarrhea. This corresponds to 2.3% (95% CI:1.9%-2.8%) of an overall standardized four-week prevalence and 0.3 (95% CI: 0.23-0.34) episodes per person-year of annual adjusted incidence rate. There was no significant difference between males and females. The incidence rates were higher among urban residents, and in the spring and summer. In the whole study period, 50% of the cases sought medical care, of which 3.9% were hospitalized and 14.3% provided a biological sample for laboratory identification of the causative agent. Children aged 0-4 and young adults aged 15-24, people living in rural areas and people who traveled frequently had higher prevalence of AGI. CONCLUSION: Results showed that AGI represents a substantial burden in China, and will contribute to the estimation of the global burden of AGI. Complemented with data on the etiologies of AGI, these estimates will form the basis to estimate the burden of foodborne diseases in China.

Attribution Analysis of Household Foodborne Disease Outbreaks in China, 2010-2020.

Foodborne diseases have become a serious public health problem worldwide, and foodborne disease outbreaks have placed a heavy disease burden on China. Foodborne disease outbreaks occur most frequently among families in China. The objectives of this study were to analyze the cause of household foodborne disease outbreaks in China from 2010 to 2020 and to identify where preventive measures could be targeted. All data were obtained from the China Foodborne Disease Surveillance System Report. A total of 17,985 outbreaks, which resulted in 73,252 illnesses, 38,829 hospitalizations, and 1269 deaths, were reported in this period. Most household outbreaks of foodborne diseases occurred in May-October, and the highest number occurred in July (3620 outbreaks, 20%). The province with the highest number of outbreaks was Yunnan Province (4829 outbreaks), followed by Hunan Province (2264 outbreaks). The attribution analysis revealed that fungi (mainly poisonous mushrooms) were the most implicated food category, with 8873 (49.3%) cases. The second was poisonous plants and their products, with 1552 (8.6%) cases. Fungi were the primary etiologic agent, with 31,125 illnesses, accounting for 42.5% of the incidents. Inedibility and misuse (9423 outbreaks), unknown origin (2505 outbreaks), and improper processing (2365 outbreaks) were the main contributing factors causing outbreaks of foodborne diseases. The results show that southwest China was a high-risk area for household foodborne diseases. Therefore, public health institutions should strengthen supervision and food safety education of residents to reduce the outbreaks of household foodborne diseases.

Whole-Genome Sequencing-Based Characterization of Clinical Listeria monocytogenes Isolates in China, 2013-2019.

Invasive listeriosis is a rare but serious foodborne disease that causes maternal-neonatal, central nervous system, and bloodstream infections. The aim of this study was to assess the whole-genome sequencing (WGS)-based genetic diversity of clinical Listeria monocytogenes isolates over a 7-year period and prove the effect of WGS application in food vehicle investigation. A total of 360 isolates were recovered during 2013 and 2019 through the national listeriosis special surveillance program. Two hundred twenty-six isolates (62.8%) were associated with pregnancy. All isolates belonged to lineage I (214 isolates) or lineage II (146 isolates), with 4 serogroups (46.9% IIb, 39.7% IIa, 12.5% IVb, and 0.8% IIc). All isolates were in 25 clonal complexes (CCs) and 3 singletons, with CC87, CC8, and CC5 being the most common causes of human listeriosis. All clinical isolates were positive for Listeria pathogenicity island 1 (LIPI-1), LIPI-3 was present in 21.4% of isolates and LIPI-4 was detected in 29.2% of isolates. LIPI-4-positive isolates, including CC87, sequence type (ST)619, ST382, CC4, and CC2, have been shown to confer hypervirulence. Fifteen isolates harbored at least one antimicrobial encoding gene, including tet (M), mef (A), msr (D), and dfr (G). The sublineage designations were consistent with CC designations, and 215 distinct cgMLST types (CTs) were classified, the most abundant being CT58 and CT750. In summary, there is a high level of genetic diversity among the clinical isolates. WGS has strengthened listeriosis surveillance and will be implemented for other foodborne bacteria in the National Molecular Tracing Network for Foodborne Disease.

[Development of PCR rapid detection method for Listeria monocytogenes in oysters].

OBJECTIVE: To develop a polymerase chain reaction(PCR) method for rapid detection of Listeria monocytogenes in oysters without pre-enrichment. METHODS: The combination of ß-cyclodextrin and bentonite-coated activated carbon was used to remove PCR inhibitors from oyster samples, and the target gene inlB was used for the PCR subsequently. The specificity, sensitivity, and application of the developed method were verified, and the stability and application of the reagents stored under cryopreservation conditions were evaluated. RESULTS: The specificity of the developed PCR method was 100% for the detection of 130 target bacterial strains and 63 non-target bacterial strains. The method reduced the time required for Listeria monocytogenes detection to 4 h without pre-enrichment, and the detection limit was 10 CFU/25 g. The method was consistent with the conventional culture method on the detection rate and viable bacteria detection rate of Listeria monocytogenes in natural oyster samples(the coincidence rate was 100%). Additionally, the reagents could be used normally after storing at-20 ℃ for at least one year. CONCLUSION: The PCR method developed in this study has high specificity and sensitivity, and can be used for rapid, accurate detection of Listeria monocytogenes in oysters.

Mechanically Induced Highly Efficient Hydrogen Evolution from Water over Piezoelectric SnSe nanosheets.

Piezoelectric nanomaterials open new avenues in driving green catalysis processes (e.g., H2 evolution from water) through harvesting mechanical energy, but their catalytic efficiency is still limited. The predicted enormous piezoelectricity for 2D SnSe, together with its high charge mobility and excellent flexibility, renders it an ideal candidate for stimulating piezocatalysis redox reactions. In this work, few-layer piezoelectric SnSe nanosheets (NSs) are utilized for mechanically induced H2 evolution from water. The finite elemental method simulation demonstrates an unprecedent maximal piezoelectric potential of 44.1 V for a single SnSe NS under a pressure of 100 MPa. A record-breaking piezocurrent density of 0.3 mA cm-2 is obtained for SnSe NSs-based electrode under ultrasonic excitation (100 W, 45 kHz), which is about three orders of magnitude greater than that of reported piezocatalysts. Moreover, an exceptional H2 production rate of 948.4 µmol g-1 h-1 is achieved over the SnSe NSs without any cocatalyst, far exceeding most of the reported piezocatalysts and competitive with the current photocatalysis technology. The findings not only enrich the potential piezocatalysis materials, but also provide useful guidance toward high-efficiency mechanically driven chemical reactions such as H2 evolution from water.

Attribution Analysis of Foodborne Disease Outbreaks Related to Meat and Meat Products in China, 2002-2017.

This study aimed to understand the epidemiological characteristics of foodborne disease outbreaks related to meat and meat products in China from 2002 to 2017. Data collected from the National Foodborne Diseases Surveillance System and searched databases were analyzed. From 2002 to 2017, China reported 2815 outbreaks caused by foodborne diseases related to meat and meat products, resulting in 52,122 illnesses and 25,361 hospitalizations, and 96 deaths. Outbreaks were markedly seasonal and concentrated from May to September, accounting for 66.93%. Outbreaks were concentrated mainly in China's eastern coastal and southern regions. Unidimensional attribution analysis revealed that livestock meat was the most commonly implicated food category causing the outbreaks, accounting for 28.67%. Bacteria were the most common pathogenic cause of outbreaks, accounting for 51.94%. Clostridium botulinum was the most common pathogenic cause of death, accounting for 34.38%. Improper processing was the most common contributing factor, accounting for 27.89%. Households were the most common food preparation location causing the outbreak, accounting for 34.39%. Two-dimensional and multidimensional attribution analysis found that Salmonella contamination occurred in different locations and regions, mainly caused by various contributing factors and improper processing. Nitrite poisoning is caused by improper processing in households in East China. Bacterial causes were the commonest agents associated with foodborne diseases related to meat and meat products, and improving the safety and quality of meat and meat product should be a priority.

Application of Whole-Genome Sequencing in the National Molecular Tracing Network for Foodborne Disease Surveillance in China.

National Molecular Tracing Network for Foodborne Disease Surveillance (TraNet) was launched in 2013, which is the only real-time whole-genome sequencing (WGS)-based subtyping network in China for effective foodborne disease surveillance. TraNet covers three levels of public health laboratories, national, provincial, and municipal. The TraNet national databases have a total of more than 54,000 entries representing seven common foodborne bacteria from humans, food, and environments. Raw sequence data are uploaded to TraNet by Data Delivery Center. Assembled sequence data, pulsed-field gel electrophoresis (PFGE) profiles, antibiotic resistance patterns, and epidemiological data are submitted to national pathogen-specific databases managed by China National Center for Food Safety Risk Assessment. PFGE patterns and WGS-based subtyping are compared for rapid differentiation of clusters of geographically diverse foodborne infections. WGS-based TraNet has played significant roles in improving foodborne disease surveillance in China for rapid outbreak investigation, source tracking, and cluster analysis of particular pathogens across the country.

Sentinel Listeriosis Surveillance in Selected Hospitals, China, 2013-2017.

During 2013-2017, a total of 211 cases of listeriosis were reported by 64 sentinel hospitals in China to a national foodborne disease surveillance network. The average case-fatality rate was 31.2% for perinatal cases and 16.4% for nonperinatal cases. Sequence types 87 and 8 were the most prevalent types.

[Attribution analysis of food borne disease outbreaks in canteen from 2002 to 2016 in China].

OBJECTIVE: Based on the angle of the "pathogenic bacteria-food", to analyze the cause of events in canteen from 2002 to 2016 in China, and to provide scientific basis for prevention and control of food borne diseases. METHODS: Collect and finish the food borne disease outbreaks events in canteen from 2002 to 2016, do the descriptive analysis of the number of events, cases and deaths by the different cause food, pathogenic factors and link, in order to do the multi-dimensional attribution analysis. RESULTS: From 2002 to 2016, there were 2129 food poisoning events in canteen in China. By the analysis of single dimensional attribution, the main cause food were vegetables, meat and fungi, respectively, the total events of 46. 09%, 16. 91% and 10. 53%. Bio-contaminants was the primary cause of pathogenic factor, accounting for 34. 05%. Production and processing were the main trigger links, accounting for 43. 31%. By the multidimensional attribution analysis, the highest number of cases(426) caused by green beans contain saponi, there were 404 incidents caused by improper production and processing. Poisonous mushrooms caused 125 toxic incidents, among them, some incidents(75) were caused by accidental consumption and misuse. CONCLUSION: It is suggested to regard green bean as the main cause food, focus on the aspects of production and processing and strengthen supervision, in order to reduce the occurrence of disease.

The Epidemiology of Listeria monocytogenes in China.

Listeria monocytogenes, a ubiquitous bacterium in nature, can lead to human listeriosis through food consumption. Listeriosis is a rare, preventable, and treatable foodborne disease but can cause hospitalizations and fatalities. We reviewed the literature published in China to better understand the prevalence of L. monocytogenes in food products, incidence of human listeriosis, and characteristics of L. monocytogenes strains in China. The average prevalence of L. monocytogenes in Chinese food products in 28 provinces was 4.42%, with the highest prevalence of 8.91% in meat-poultry products, followed by aquatic animals, Chinese salad and salad, rice and flour products, and so on. Two hundred fifty-three invasive listeriosis cases were reported from 2011 to 2016 in 19 provinces, and the overall case-fatality rate was 25.7% with no deaths reported of pregnant women and children. L. monocytogenes strains were generally susceptible to most antibiotics, with ampicillin and penicillin G still effective in treatment. The predominant sequence types (STs) in food were ST9 and ST8, while clinically ST87 was most common ST in China. The national human listeriosis pilot surveillance started in 2013, and a total of 133 listeriosis cases have been collected until now. On the basis of the surveillance program, further research should be conducted to uncover the reason for the prevalence and pathogenic mechanism of the highly epidemiological hypervirulent ST87 strains in China.

[Analysis on 2011 quality control results on aerobic plate count of microbiology laboratories in China].

OBJECTIVE: To test the aerobic plate count examining capability of microbiology laboratories, to ensure the accuracy and comparability of quantitative bacteria examination results, and to improve the quality of monitoring. METHODS: The 4 different concentration aerobic plate count piece samples were prepared and noted as I, II, III and IV. After homogeneity and stability tests, the samples were delivered to monitoring institutions. The results of I, II, III samples were logarithmic transformed, and evaluated with Z-score method using the robust average and standard deviation. The results of IV samples were evaluated as "satisfactory" when reported as < 10 CFU/piece or as "not satisfactory" otherwise. Pearson χ2 test was used to analyze the ratio results. RESULTS: 309 monitoring institutions, which was 99.04% of the total number, reported their results. 271 institutions reported a satisfactory result, and the satisfactory rate was 87.70%. There was no statistical difference in satisfactory rates of I, II and III samples which were 81.52%, 88.30% and 91.40% respectively. The satisfactory rate of IV samples was 93.33%. There was no statistical difference in satisfactory rates between provincial and municipal CDC. CONCLUSION: The quality control program has provided scientific data that the aerobic plate count capability of the laboratories meets the requirements of monitoring tasks.

[Analysis of antimicrobial resistance results of Vibrio parahaemolyticus isolated from shellfish and its habitat in 2013].

OBJECTIVE: To evaluate the antimicrobial resistance of Vibrio parahaemolyticus isolated from shellfish and its habitat in Sichuan, Fujian and Guangxi. METHODS: The susceptibility of 186 isolates of Vibrio parahaemolyticus to 8 antibiotics were determined by broth microdilution susceptibility test. The antibiotics of ampicillin, cefotaxime, ceftazidime, gentamicin, tetracycline, chloramphenicol, ciprofloxacin and trimethoprim/sulfamethoxazole were used. RESULTS: The antibiotic resistance rate were 69.35% in which ampicillin resistance was the most prevalent. The geometric mean of ampicillin MIC value of all isolates was greater than the interpretation value of resistance. Among the 186 tested isolates, the multiple antibiotic resistance and/or intermediate resistance was 4. All strains were 100% sensitive to cefotaxime, ceftazidime, chloramphenicol and ciprofloxacin. The ampicillin susceptibility spectrum of aquaculture farm was the highest among the three sectors as 73.68% and Sichuan was the highest among the three provinces as 70.94% although there were no significant differences. There were 44 samples out of which 2 and above strains were isolated, and the susceptibility spectrum polymorphism rate of strains isolated from the same sample was 77.27% (34/44). CONCLUSION: The ampicillin resistance rate is relatively high, and shellfish habitat may be the main source of antimicrobial resistance of Vibrio parahaemolyticus. There is an urgent need to strengthen the surveillance and management of Vibrio parahaemolyticus in the shellfish breeding process.

[Molecular genetic makers for Vibrio parahaemolyticus--a review].

Vibrio parahaemolyticus is an important foodborne pathogen, of which the 03:K6 serotype caused many outbreaks in different countries since 1996. Based on the 10 years data (1992-2001) from China, gastroenteritis caused by Vibrio parahaemolyticus accounted for 31.1% of foodborne disease outbreaks that were resulted from microorganisms. Most environmental strains of Vibrio parahaemolyticus are non-pathogenic strains. However, clinical strains can producethermostable direct hemolysin (TDH), TDH-related hemolysin, and other virulence factors. Here we reviewed three commonly used molecular markers for Vibrio parahaemolyticus, including species-specific genes, the virulence genes and pandemic group-specific genes, so that to provide references for the rapid detection of Vibrio parahaemolyticus and the identification of its pathogenic factor.

[Study on detection of Salmonella in poultry samples by real-time PCR with Taqman probes].

OBJECTIVE: To develop a RT-PCR method for a rapid and sensitive detection of Salmonella in poultry samples. METHODS: The RT-PCR method was established and its specificity was testified on the basis of invA gene. Serial 10-fold diluted pure suspension culture of CMCC 50041 was detected by RT-PCR, the standard curve was constructed and the amplification efficiency was calculated. Artificially contaminated experiment was done, six artificially-inoculated samples containing final concentration of Salmonella CMCC 50041 (1, 10, 10(2), 10(3), 10(4), 10(5) and 10(6) CFU per 25 g poultry samples) were prepared respectively. All the samples were incubated for 0, 4, 8, 12 and 18 h and the DNA was extracted for RT-PCR detection, meanwhile by PCR detection and the traditional method. The sensitiveness and specificity were compared among the three methods. At the same time, 16 samples of retail whole poultry were collected from markets and detected by the above three methods as well, and thereby to further compare the positive detection among the three methods. RESULTS: The established RT-PCR method was specific for the detection of Salmonella. The sensitivity was 5.2×10(3) CFU/ml for pure Salmonella culture without enrichment. Correlation coefficients of standard curves constructed using the Ct versus log value of concentration of Salmonella showed good linearity over a 8-log dynamic range (5.2×10(3)-5.2×10(10) CFU/ml), with the R(2) at 0.999. RT-PCR detection limit for artificially contaminated samples after enriching for 12 h was 1 CFU/25 g sample, which was the same with the limit of PCR and 10 times more sensitive than the limit of traditional method. Standard curve of sample after enrichment for 12 h was established. Seven of 16 samples were detected positive by RT-PCR, which were also tested positive by PCR, while only five samples were positive by traditional method. The positive ones were quantitatively analyzed using standard curve of sample and determined the initial Salmonella numbers of CFU/25 g. CONCLUSION: The established RT-PCR technology was simple, rapid, sensitive and specific, which was suitable to quickly detect Salmonella in poultry samples.

[Epidemiological characteristics of Salmonella in animal source foods in Hunan].

OBJECTIVE: To study the molecular epidemiological characteristics of Salmonella in animal source foods in Hunan. METHODS: The fair trade markets and supermarkets of ten cities were chosen to sample animal source foods for isolating Salmonella in Hunan province in 2010. A total of 692 samples were collected by aseptic sampling, included 159 livestock meats, 152 poultry meats, and 381 aquatic products.Salmonella strains isolated were subjected to stereotyping, antimicrobial susceptibility testing and pulsed field gel electrophoresis (PFGE). RESULTS: Salmonella was detected in 93 of 692 animal food samples with the detection rate of 13.4%. The detection rates of Salmonella in poultry meats, livestock meats and aquatic products were 23.0% (35/152), 22.6% (36/159) and 5.8% (22/381) respectively. Therefore, the detection rate in aquatic products was lower than that of poultry meats and livestock meats (χ(2) = 33.86, P < 0.05; χ(2) = 33.29, P < 0.05, respectively). The serotypes of isolates showed diversity, and Salmonella Derby (33/94, 35.1%) was the predominant serotypes.79.8% (75/94) strains showed resistant to more than one antibiotic used in the test, 31.9% (30/94) strains showed resistant to more than 5 antibiotics. A significant difference was observed for multidrug resistance between Salmonella isolated from poultry (47.2%, 17/36) and livestock meats (22.2%, 8/36) (χ(2) = 4.96, P < 0.05). And the highest resistant rate was found in tetracycline, as high as 62.8% (59/94). All the strains were divided into 69 PFGE subtypes.Furthermore the dominating subtypes were type 7 (6 strains), type 15 (4 strains), type 22 (6 strains). CONCLUSION: Inspection results showed that Salmonella contamination in animal source foods were serious in Hunan province, and the isolates expressed high level resistance to the antibiotics.Furthermore the PFGE results indicated that there were epidemic strains of Salmonella in Hunan.

[Antimicrobial resistance profiles and genetic diversity of bovine staphylococcus aureus isolated in 5 provinces of China in 2013].

OBJECTIVE: To investigate antimicrobial resistance profiles and genetic diversity of staphylococcus aureus isolated from lactating cows of 5 provinces in China, 2013. METHODS: A total of 680 samples were collected from 15 herds (12 farms, 3 artels) in 5 provinces of China in 2013, including swabs of extramammary sites (bovine teat skin and milking machine liners) and quarter milk samples from lactating cows diagnosed with subclinical mastitis. The antimicrobial resistance of the isolates were tested by broth microdilution method and the genotypes were determined by PFGE (pulsed-field gel electrophoresis) method. RESULTS: A total of 111 isolates were isolated and identified as staphylococcus aureus. Resistance to penicillin (90.1% (100/111)), erythromycin (48.6% (54/111)), ciprofloxacin (36.9% (41/111)), clindamycin (27.9% (31/111)), gentamycin (18.9% (21/111)), chloramphenicol (9.0% (10/111)), tetracycline (7.2% (8/111)) of these strains were observed. All isolates were sensitive to oxacillin, vancomycin and selectrin. 92.8% (103/111) staphylococcus aureus isolates were resistant to at least one antimicrobial. 38.7% (43/111) strains were multi-drug resistant isolates. The resistance rate of isolates in artels (100% (48/48)) was higher than it in farms (87% (55/63)) and the difference was statistically significant (χ(2) = 4.80, P < 0.05). The multi-resistance rate of isolates in artels (54% (26/48)) was higher than it in farms (27% (17/63)) and the difference was also statistically significant (χ(2) = 8.48, P < 0.05). The 111 strains were clustered into 8 types, 6 out of which were consisted of 98% isolates (109/111), and were prevalent in 2 to 9 herds. Every herd had 1 to 4 types, and tend to be comprised by one major type. Most swab isolates were indistinguishable from isolates infecting the mammary gland. There were no relationship between antimicrobial resistance profiles and genotypes of these isolates. CONCLUSION: The drug resistance of staphylococcus aureus isolates associated with lactating cows of 5 provinces in China is serious, especially the isolates collected from artels. A few specialized clones were responsible for most of the cases of bovine mastitis in a single herd and some clones might have a broad geographic distribution.

[Analysis of molecular features of clinical Vibrio parahaemolyticus strains in China].

OBJECTIVE: To explore the phenetic and genetic features of clinical Vibrio parahaemolyticus strains from 2007-2009 in China. METHODS: A total of 135 clinical Vibrio parahaemolyticus strains, isolated from Zhejiang, Jiangsu, Sichuan, Guangxi, Liaoning Provinces during 2007 to 2009, were selected for the research. The occurrence of virulence genes thermostable direct hemolysin (tdh) and TDH-related hemolysin (trh), species-specific genes thermolabile hemolysin (tlh), toxR, VPM and gyrB, the pandemic clone gene markers(GS-PCR, PGS-PCR, orf8 and HU-α) in 135 Vibrio parahaemolyticus strains was detected by PCR. The antimicrobial susceptibilities to eight antimicrobial agents of the experimental strains were determined by the broth microdilution method. All strains were serotyped and underwent the cluster analysis with pulsed-field gel electrophoreses. RESULTS: The results of PCR methods claim that all experiment strains carry species-specific genes such as tlh, toxR, gyrB, VPM. Among clinical strains, 85.9% (116/135) carry tdh and/or trh. 85.2% (115/135) were positive for tdh, and 3.0% (4/135) were positive for trh; while 3 strains carried both.66.7% (90/135) , 80.7% (109/135) , 65.2% (88/135) , 66.7% (90/135) clinical strains carried the genes of GS-PCR, PGS-PCR, orf8, HU-α, respectively. The results of antibiotics susceptibility test showed that 8.1% (11/135) strains were resistant to at least one agent, including 9 strains were resistant to ampicillin, 2 strains were resistant to trimethoprim and sulphamethoxazole, and 1 strain were resistant to tetracycline. All clinical strains were sensitive to cefotaxime, ceftazidime, gentamicin, ciprofloxacin and chloromycetin.Serological analysis of the O and K antigens claimed that a total of 29 serotypes were identified for clinical strains, predominantly O3, O4 and O1 groups, accounting for 89.6% (121/135). O3: K6 was dominant serotype, accounting for 56.3% (76/135). The pandemic flora in China included O3: K6, O4: K68, O1: K36, O1: K25, O1: K5 and O3: K29 serotypes.Genomic DNAs of 135 clinical strains were digested with SfiI and NotI, the molecular size of PFGE restriction fragments used for analysis mainly ranged from 30-700 kb.When subjected to UPGMA clustering, 6 and 9 clusters were grouped by SfiI and NotI, and the minimal similarity was 52.6% and 58.7%, and pandemic flora were located in C groups and D group, respectively. CONCLUSION: Most of Vibrio parahaemolyticus strains isolated from clinical sources in China were pathogenic. The pandemic clone, especially O3: K6 was prevalent. The GS-PCR and HU-α genes were reliable markers to identify the pandemic flora. The serotype by PFGE was reliable to distinguish the pandemic flora and the sporadic strains.

[Identification and molecular subtyping of Campylobacter jejuni isolated from chicken carcass].

OBJECTIVE: To characterize and investigate the molecular types of Campylobacter jejuni isolated from slaughter chicken carcass, which would provide scientific data for campylobacter food poisoning traceability. METHODS: Biochemical and molecular biological methods were used for screening and identification of isolates from chicken special monitoring networks. Campylobacter Genus-specific primers 16S rRNA and species-specific primers MapA and CeuE were designed to perform a multiplex PCR to identify these isolated strains. Pulsed-field gel electrophoresis (PFGE) was employed to type Campylobacter jejuni isolates by digesting with restriction endonuclease Sma I and Kpn I respectively. Fingerprints of these isolates were analyzed by the software BioNumerics. RESULTS: 72 out of 81 isolates were confirmed as Campylobacter jejuni by biochemical test combined with PCR. 48 patterns were obtained from PFGE with Sma I and Kpn I. 72 isolated strains were divided into 13 clusters (A-M) according to 63.9% similarity by cluster analysis. Isolates from different provinces were distributed in 13 clusters and each cluster contained 1 to 11 patterns. The results showed that the 72 strain patterns distribution had complete regional homology, namely strains in the same pattern were from a single province. CONCLUSION: The comprehensive analysis of Sma I and Kpn I results may help improving the resolution of PFGE, increasing the accuracy of typing and reliability of traceability.

[Rapid detection of Listeria monocytogenes in pork samples by real-time PCR with Taqman probe].

OBJECTIVE: To develop a real-time PCR method for detection Listeria monocytogenes in pork samples. METHODS: Listeria monocytogenes specific primers and Taqman probe were chosen on the basis of hlyA gene. Real-time PCR method was developed and its specificity was proved. Serial 10-fold diluted pure suspension culture of CMCC 540004 were detected by real-time PCR, and standard curve was constructed. Artificially contaminated experiment was done, six artificially-inoculated samples containing final concentration of Listeria monocytogenes CMCC 540004 (1.3 x 10(0), 1.3 x 10(1), 1.3 x 10(2), 1.3 x 10(3), 1.3 x 10(4), 1.3 x 10(5) and 1.3 x 10(6) CFU per 25 g pork samples) were preparated respectively, meanwhile one sample without inoculation was as control of background value. All the samples were incubated in LB1 enrichment for 24 h and then take 0.1 ml culture solutions to 10 ml LB2 enrichment for 18 - 24 h. All the samples were incubated for 0, 4, 8, 12, 18, 24, 30, 36 and 46 h, and detected Listeria monocytogenes bacteria by PCR, respectively. Twenty-four samples of retail pork were collected from markets in Beijing and detected by the above three methods. RESULTS: Real-time PCR method established was specific for the detection of Listeria monocytogenes. The sensitivity was 1.3 x 10(3) CFU/ml for pure culture without enrichment. Real-time PCR detection limit for artificially contaminated samples after enriching for 24 h was 1.3 CFU/ 25 g, which is the same with the limit of PCR and traditional method after enrichment for 46 h. Standard curve of sample after enrichment for 24 h was established. The positive rate out of total 24 samples was 70.83% (17/24) by real-time PCR, which is the same with the result of PCR and traditional method. The positive ones were quantitative analyzed using standard curve of sample and determined the initial Listeria monocytogenes numbers of CFU/25 g. CONCLUSION; The established real-time PCR technology was simple, rapid, sensitive and specific, which was suitable to rapid detect Listeria monocytogenes in pork samples and the process was finished in 27 h.