The molecular characters and antibiotic resistance of Clostridioides difficile from economic animals in China.

BACKGROUND: It has been performed worldwidely to explore the potential of animals that might be a reservoir for community associated human infections of Clostridioides difficile. Several genetically undistinguished PCR ribotypes of C. difficile from animals and human have been reported, illustrating potential transmission of C. difficile between them. Pig and calf were considered as the main origins of C. difficile with predominant RT078 and RT033, respectively. As more investigations involved, great diversity of molecular types from pig and calf were reported in Europe, North American and Australia. However, there were quite limited research on C. difficile isolates from meat animals in China, leading to non-comprehensive understanding of molecular epidemiology of C. difficile in China. RESULTS: A total of 55 C. difficile were isolated from 953 animal stool samples, within which 51 strains were from newborn dairy calf less than 7 days in Shandong Province. These isolates were divided into 3 STs and 6 RTs, of which ST11/RT126 was predominant type, and responsible for majority antibiotic resistance isolates. All the isolates were resistant to at least one tested antibiotics, however, only two multidrug resistant (MDR) isolates were identified. Furthermore, erythromycin (ERY) and clindamycin (CLI) were the two main resistant antibiotics. None of the isolates were resistant to vancomycin (VAN), metronidazole (MTZ), tetracycline (TET), and rifampin (RIF). CONCLUSIONS: In this study, we analyzed the prevalence, molecular characters and antibiotic resistance of C. difficile from calf, sheep, chicken, and pig in China. Some unique features were found here: first, RT126 not RT078 were the dominant type from baby calf, and none isolates were got from pig; second, on the whole, isolates from animals display relative lower resistant rate to these 11 tested antibiotics, compared with isolates from human in China in our previous report. Our study helps to deep understanding the situation of C. difficile from economic animals in China, and to further study the potential transmission of C. difficile between meat animals and human.

Antibiotic resistance and molecular characterization of diarrheagenic Escherichia coli and non-typhoidal Salmonella strains isolated from infections in Southwest China.

BACKGROUND: Bacterial diarrhea is one of the most common causes for medical consultations, mortality and morbidity in the world. Diarrheagenic Escherichia coli (DEC) and non-typhoidal Salmonella (NTS) are major intestinal pathogens in developing countries, and the indiscriminate use of antibiotics has greatly contributed to resistant strains. Hence, the aim of the present study is to identify the antimicrobial resistance patterns and the molecular characteristics of DEC and NTS in southwest, China. METHODS: 1121 diarrheal patients and 319 non-diarrheal subjects across all age groups were recruited from four sentinel hospitals from June 2014 to July 2015 in Kunming City, Yunnan Province. Each stool specimen was collected to detect DEC and NTS with standard microbiological and molecular methods. Antimicrobial resistance testing was performed with the Kirby-Bauer disk diffusion method, and the standards for antimicrobial susceptibility testing complied with the Clinical and Laboratory Standards Institute (CLSI). Molecular characterization of strains was carried out using pulsed-field gel electrophoresis (PFGE). A structured questionnaire was used to record basic epidemiological data (e.g. sex, age, residence, season, etc.). Data were analyzed using Chi-square or Fisher's exact test. RESULTS: DEC was detected in 127 (11.33%) diarrhea cases and 9 (2.82%) non-diarrheal cases (χ2 = 20.69, P < 0.001, OR = 4.36, 95% CI: 2.19-8.65), and the prevalence of NTS isolated from diarrhea cases was higher than that of non-diarrheal cases across all age groups (n = 42, 3.75%, n = 1, 0.31%, χ2 = 10.10, P = 0.002, OR = 12.38, 95% CI: 1.70-90.29). The rates of resistance to ten antibiotics of DEC and NTS showed significant differences (χ 2 = 386.77, P < 0.001; χ2 = 191.16, P < 0.001). The rates of resistance to Amoxicillin and Clavulafiate (AMC), Cephalothin (CEP), Gentamicin (GEN) and Sulfamethoxazole-Trimethoprim (SXT) of DEC isolated from diarrhea cases were higher than those of NTS isolated from diarrhea patients (37.01% vs 14.29%, χ2 = 7.57, P = 0.006; 29.92% vs 11.90%, χ2 = 5.40, P = 0.02; 37.01% vs 11.90%, χ2 = 5.80, P = 0.016; 62.20% vs 26.19%, χ2 = 16.44, P < 0.001; respectively). Ciprofloxacin (CIP) was the most sensitive antibiotic for DEC and NTS strains isolated from diarrhea cases. Resistance rates of DEC isolates from cases and controls to more than three kinds antimicrobials (multidrug resistance, MDR) showed no significant differences (81.10% vs 88.89%, P = 0.33). Pulsotype patterns of DEC strains were highly diverse; however, the pulsotype pattern of NTS strains was closely related to the serotype. The pattern of S. enteritidis was highly similar, but the S. enterica Typhimurium strain was discrete. CONCLUSIONS: Antibiotic resistance of Enterobacteriaceae is of great concern. The societal effects of antibiotic use justify strict monitoring to combat increases in antimicrobial resistance. Molecular epidemiology and systematic epidemiological investigation can provide accurate evidence for tracking the infection source.

Case-control study of diarrheal disease etiology in individuals over 5 years in southwest China.

BACKGROUND: Acute diarrhea is one of the major public health problems worldwide. Most of studies on acute diarrhea have been made on infants aged below 5 years and few efforts have been made to identify the etiological agents of acute diarrhea in people over five, especially in China. METHODS: 271 diarrhea cases and 149 healthy controls over 5 years were recruited from four participating hospitals between June 2014 and July 2015. Each stool specimen was collected to detect a series of enteric pathogens, involving five viruses (Rotavirus group A, RVA; Norovirus, NoV; Sapovirus, SaV; Astrovirus, As; and Adenovirus, Ad), seven bacteria (diarrheagenic Escherichia coli, DEC; non-typhoidal Salmonella, NTS; Shigella spp.; Vibrio cholera; Vibrio parahaemolyticus; Aeromonas spp.; and Plesiomonas spp.) and three protozoa (Cryptosporidium spp., Giardia lamblia, G. lamblia, and Blastocystis hominis, B. hominis). Standard microbiological and molecular methods were applied to detect these pathogens. Data was analyzed using Chi square, Fisher-exact tests and logistic regressions. RESULTS: The prevalence of at least one enteric pathogen was detected in 29.2% (79/271) acute diarrhea cases and in 12.1% (18/149) in healthy controls (p < 0.0001). Enteric viral infections (14.4%) were the most common in patients suffering from acute diarrhea, followed by bacteria (13.7%) and intestinal protozoa (4.8%). DEC (12.5%) was the most common causative agent in diarrhea cases, followed by NoV GII (10.0%), RVA (7.4%) and B. hominis (4.8%). The prevalence of co-infection was statistically higher (p = 0.0059) in the case group (7.7%) than in the healthy control (1.3%). RVA-NoV GII (3.0%) was the most common co-infection in symptomatic cases. CONCLUSIONS: DEC was the most predominant pathogen in diarrhea cases, but it was largely overlooked because the lack of laboratory capacities. Because of the high prevalence of co-infections, it is recommended the urgent development of alternative laboratory methods to assess polymicrobial infections. Such methodological improvements will result in a better prevention and treatment strategies to control diarrhea illness in China.

Impact of co-infections with enteric pathogens on children suffering from acute diarrhea in southwest China.

BACKGROUND: Acute diarrhea is a global health problem, resulting in high morbidity and mortality in children. It has been suggested that enteric pathogen co-infections play an important role in gastroenteritis, but most research efforts have only focused on a small range of species belonging to a few pathogen groups. This study aimed to assess the impact of co-infections with a broad range of enteric pathogens on children aged below five years who suffer from acute diarrhea in southwest China. METHOD: A total of 1020 subjects (850 diarrhea cases and 170 healthy controls) were selected from four sentinel hospitals in Kunming, Yunnan province, southwest China, from June 2014 to July 2015. Stool specimens were collected to detect five virus (rotavirus group A, RVA; norovirus, NoV; Sapovirus, SaV; astrovirus, As; and adenovirus, Ad), seven bacterial (diarrheagenic Escherichia coli, DEC; non-typhoidal Salmonella, NTS; Shigella spp.; Vibrio cholera; Vibrio parahaemolyticus; Aeromonas spp.; and Plesiomonas spp.), and three protozoan (Cryptosporidium spp., Giardia lamblia, and Blastocystis hominis, B. hominis) species using standard microbiologic and molecular methods. Data were analyzed using the partial least square regression technique and chi-square test. RESULTS: At least one enteric pathogen was detected in 46.7 % (n = 397) of acute gastroenteritis cases and 13.5 % (n = 23) of healthy controls (χ(2) = 64.4, P < 0.05). Single infection with RVA was associated with acute diarrhea (26.5 % vs. 5.8 %, P < 0.05). The prevalence of a single infection with B. hominis in diarrhea cases was higher than in healthy controls (3.1 % vs. 0.5 %, OR = 4.7, 95 % CI: 1.01-112.0). Single infection with NoV GII was not associated with diarrhea (4.4 % vs. 3.5 %, OR = 1.2, 95 % CI: 0.5-3.3). Single infections with bacterial species were not observed. The prevalence of co-infections with two enteric pathogens in diarrhea cases was higher than in asymptomatic children (20.1 % vs. 5.3 %, P < 0.05). RVA-NoV GII was the most common co-infection in symptomatic children (4.4 %), with it aggravating the severity of diarrhea. CONCLUSIONS: Although it is clear that RVA has an overwhelming impact on diarrhea illnesses in children, co-infection with other enteric pathogens appears to also aggravate diarrhea severity. These findings should serve as evidence for public health services when planning and developing intervention programs.

Comparison of lipopolysaccharide and protein immunogens from pathogenic Yersinia enterocolitica bio-serotype 1B/O:8 and 2/O:9 using SDS-PAGE.

OBJECTIVE: Yersinia enterocolitica is an extracellular pathogen and its related antigens interact with the host immune system. We investigated the difference in immunological characteristics between a highly pathogenic and poorly pathogenic strain of Y. enterocolitica. METHODS: We used SDS-PAGE and western blotting to characterize lipopolysaccharide (LPS), Yersinia outer membrane proteins (Yops), membrane proteins, and whole-cell proteins from poorly pathogenic Y. enterocolitica bio-serotype 2/O:9, isolated from China, and highly pathogenic bio-serotype 1B/O:8, isolated from Japan. RESULTS: These two strains of Y. enterocolitica had different LPS immune response patterns. Comparison of their Yops also showed differences that could have accounted for their differences in pathogenicity. The membrane and whole-cell proteins of both strains were similar; immunoblottting showed that the 35 kD and perhaps the 10 kD proteins were immunogens in both strains. CONCLUSION: The major antigens of the two strains eliciting the host immune response were the LPS and membrane proteins, as shown by comparing protein samples with reference and purified preparations.