Molecular epidemiology of Listeria monocytogenes isolated from ready-to-eat food in 2017 in China / 中华预防医学杂志

Objective@#To analyze the molecular characteristics of Listeria monocytogenes strains from ready-to eat food in China.@*Methods@#A total of 239 Listeria monocytogenes strains isolated from ready-to-eat food in 2017, all strains underwent whole-genome sequencing (WGS) , and comparisons uncovered population structure derived from lineages, clonal complex, serogroups, antimicrobial susceptibility and virulence, which were inferred in silico from the WGS data. Core genome multilocus sequence typing was used to subtype isolates.@*Results@#All strains were categorized into three different lineages, lineage Ⅱ was the predominant types in food, and IIa was the main serogroups. CC8, CC101 and CC87 were the first three prevalent CCs among 23 detected CCs, accounting for 49.4%. Only 4.6% (11 isolates) of tested strains harbored antibiotic resistance genes, which were mostly trimethoprim genes (7 isolates, 2.9%). All strains were positive for LIPI-1, and only a part of strains harbored LIPI-3 and LIPI-4, accounting for 13.8% (33 isolates) and 14.2% (34 isolates), respectively. ST619 carried both LIPI-3 and LIPI-4. 51.5% (123 isolates) of strains carried SSI-1, and all CC121 strains harbored SSI-2. Different lineages, serogroups and CCs can be separated obviously through cgMLST analysis, and 24 sublineages were highly concordant with CCs.@*Conclusion@#Ⅱa was the main serogroups in ready-to-eat food isolates in China; CC8, CC101 and CC87 were the prevalent CCs, and CC87 isolates was hypervirulent isolates, cgMLST method can be adopted for prospective foodborne disease surveillance and outbreaks detection.

Interpretation for the group standards of the Isolation and Identification of Campylobacter jejuni and Campylobacter coli / 中华流行病学杂志

Foodborne Campylobacter is recognized as the leading causes of the bacterial diarrheal illness in both developing and developed countries. C. jejuni and C. coli caused 95% of the human campylobacterisosis. Bacteria culture has been recognized as the "Gold standard" for the diagnosis of the Campylobacter infection. The National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention coordinated the experienced researchers from China National Center for Food Safety Risk Assessment and other local Centers for Disease Control and Prevention to write up the standards for entitled Isolation and Identification of Campylobacter jejuni and Campylobacter coli (T/CPMA 006-2019). The standard is drafted with principles of emphasizing the scientific, normative, applicability and feasible nature. This group standard recommended the procedures and steps for the isolation and identification of C. jejuni and C.coli from variant samples. The standard aims to improve the capacity for Campylobacter identification in China.

Prevalence and antimicrobial susceptibility of Salmonella isolated from broiler whole production process in four provinces of China / 中华预防医学杂志

Objective@#To determine the prevalence and antimicrobial susceptibility of Salmonella isolated from broiler production process in 4 provinces of China.@*Methods@#Using convenience sampling method, 238 sample sites from broiler whole production process were chosen in Henan, Jiangsu, Heilongjiang and Shandong provinces in 2012. A total of 11 592 samples were collected and detected to analyze prevalence baseline, including 2 090 samples from breeding chicken farms and hatcheries, 1 421 samples from broiler farms, 5 610 samples from slaughterhouses and 2 471 samples from distribution and retail stores. All Salmonella strains were isolated through selective enrichment, and were serotyped according to Kauffmann-White scheme. The antimicrobial susceptibilities of selected Salmonella strains were determined by the broth microdilution method and fourteen antimicrobial agents were examined.@*Results@#During incubation course, the average prevalence of Salmonella was 5.5% in feces of breeding hens, feces of chicks, and hatching eggs, 123 Salmonella strains were isolated. During cultivation course, the prevalence of Salmonella was 8.0% in feces from broiler farms, soil, feed, and workers, 114 Salmonella strains were isolated. During slaughter course, the prevalence of Salmonella was 24.9% in swabs pre-slaughter, dressed broiler carcasses, pre-cooled broiler carcasses, water from precooling pool, cutter and chipping boards, frozen chicken portions, and workers, 1 438 Salmonella strains were isolated. During distribution and sale course, the prevalence of Salmonella was 20.9% in transport carts, frozen chicken portions, retail chicken portions and workers, 551 Salmonella strains were isolated. The dominant Salmonella serotypes were Salmonella Enteritidis (n=1 229) and Salmonella Indiana (n=621). Among 1 231 examined strains, 97.2% Salmonella isolates were resistant to at least one antimicrobial, 69.9% Salmonella strains were multi-drug resistant isolates.@*Conclusion@#Our findings indicated that Salmonella contamination was common and serious in commercial broiler whole production process in China, especially in the course of defeathering, precooling and selling. The environment of broiler farm is the important source of Salmonella contamination. Additionally, antibiotic resistance of Salmonella was serious for common antimicrobials and multi-drug resistant strains existed widespread, which can pose potential risk on public health and clinical therapy.

Surveillance of contamination level and antimicrobial resistance analysis of Salmonella on broiler carcasses after chilling in 4 poultry slaughterhouses of Henan Province / 中华预防医学杂志

Objective@#Tests were carried out for obtaining contamination level and antimicrobial resistance of Salmonella on broiler carcasses after chilling in four poultry slaughterhouses in Henan.@*Methods@#Totally, two hundred sixty nine broiler carcasses after chilling were collected in four slaughterhouses with the daily slaughter amount around 15 000 to 50 000. For qualitative analysis of Salmonella EFSA method was used and for quantitative analysis of Salmonella modified Rappaport-Vassiliadis most probable number (MSRV-MPN) method was used. All of the isolates were subjected to antimicrobial susceptibility testing of 8 antibiotics by minimum inhibitory concentration (MIC).@*Results@#Overall, 48.7% (131/269) of the broiler carcasses after chilling were contaminated by Salmonella, and the average of contamination level is 1.32 most probable number MPN/g. Eight serotypes were detected. The dominant serotype is Salmonella enteritidis (93, 71.0%) followed by Salmonella Indiana (21, 16.0%). Only 2 (1.5%) Salmonella enteritidis strains were sensitive to all the tested antibiotics and the remaining 129 isolates were resistant to at least one kind of eight class antibiotics. Among them, resistant to NAL was the common (104, 79.4%) and 51 (38.9%) Salmonella isolates were multidrug-resistant.@*Conclusion@#The contamination rate and multiple antimicrobial resistant of Salmonella on broiler carcasses after chilling from slaughterhouses was very serious, while the isolates contained various serotypes.

Trends of foodborne diseases in China: lessons from laboratory-based surveillance since 2011 / 医学前沿

Foodborne disease is one of the most important public health issues worldwide. China faces various and unprecedented challenges in all aspects of the food chain. Data from laboratory-based foodborne disease surveillance systems from 2013 to 2016, as well as different regions and ages, can be found along with differences in the patterns of pathogens detected with diverse characteristics. Vibrio parahaemolyticus has been the leading cause of infectious diarrhea in China, especially among adults in coastal regions. Salmonella has been a serious and widely distributed pathogen responsible for substantial socioeconomic burden. Shigella was mostly identified in Northwest China and the inland province (Henan) with less-developed regions among children under 5 years. Data from foodborne disease outbreak reporting system from 2011 to 2016 showed that poisonous animals and plant factors responsible for most deaths were poisonous mushrooms (54.7%) in remote districts in southwest regions. The biological hazard that caused most cases reported (42.3%) was attributed to V. parahaemolyticus, the leading cause of foodborne outbreaks. In this review, we summarize the recent monitoring approach to foodborne diseases in China and compare the results with those in developed countries.

Epidemiological characteristics of Salmonella in animal source foods in Hunan / 中华预防医学杂志

<p><b>OBJECTIVE</b>To study the molecular epidemiological characteristics of Salmonella in animal source foods in Hunan.</p><p><b>METHODS</b>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).</p><p><b>RESULTS</b>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).</p><p><b>CONCLUSION</b>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.</p>

Study on detection of Salmonella in poultry samples by real-time PCR with Taqman probes / 中华预防医学杂志

<p><b>OBJECTIVE</b>To develop a RT-PCR method for a rapid and sensitive detection of Salmonella in poultry samples.</p><p><b>METHODS</b>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.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSION</b>The established RT-PCR technology was simple, rapid, sensitive and specific, which was suitable to quickly detect Salmonella in poultry samples.</p>

Analysis of molecular features of clinical Vibrio parahaemolyticus strains in China / 中华预防医学杂志

<p><b>OBJECTIVE</b>To explore the phenetic and genetic features of clinical Vibrio parahaemolyticus strains from 2007-2009 in China.</p><p><b>METHODS</b>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.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSION</b>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.</p>

Antimicrobial resistance profiles and genetic diversity of bovine staphylococcus aureus isolated in 5 provinces of China in 2013 / 中华预防医学杂志

<p><b>OBJECTIVE</b>To investigate antimicrobial resistance profiles and genetic diversity of staphylococcus aureus isolated from lactating cows of 5 provinces in China, 2013.</p><p><b>METHODS</b>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.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSION</b>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.</p>