Diarrhea pathogens in rotavirus-negative infants in Hangzhou / 中华检验医学杂志

Objective To investigate the distribution of diarrhea pathogens in infants without rotavirus-detection in Hangzhou. Methods 605 stool samples of children with rotavirus-negative diarrhea were collected from Hangzhou First People's Hospital of Zhejiang University, Children's Hospital of Zhejiang University and Hangzhou Children's Hospital from March 2017 to June 2018. The routine test results were analyzed retrospectively and Bristol score was used for the characteristics of stool samples. DNA and/or RNA were extracted from fecal samples with DNA and RNA extraction kit. The extracted DNA and RNA-reversed cDNA were used as templates. 7 common pathogens DNA and/or RNA were amplified by polymerase chain reaction (PCR). The amplified products were detected by agarose gel electrophoresis. The positive rates of pathogens were analyzed by chi-square test. Results Among 605 children, 375 were male (28±11) months and 230 were female (29±10) months. Bristol score of stool samples was mainly in type 6 (496, 82％), followed by type 7 (85, 14％) and type 5 (24, 4％). Among 605 results 97 cases were occult blood positive (positive rate 16％) and 170 cases were white blood cell positive (positive rate 28％).452 of 605 stool samples were positive for pathogen target genes. The positive rate was 74.7％. 319 cases detected single pathogen gene fragments. 127 cases detected two pathogen genes and 6 cases detected three pathogen gene fragments. The positive rate of Clostridium difficile toxin B (48.9％, 296/605)was the highest than the others, followed by Salmonella (20.0％, 121/605) and Norovirus (10.9％, 66/605). The positive rate of Clostridium difficile toxin A was 1.0％ (6/605). The positive rates of pathogens in male and female children were 86.7％(325 / 375) and 86.5％ (199 / 230) respectively, with (χ2 =0.002, P=0.959). Conclusions Salmonella and Norovirus were the main pathogens in children with diarrhea who were negative for rotavirus detection in Hangzhou. The high positive rate of Clostridium difficile toxin B may be related to the colonization of Clostridium difficile in the gastrointestinal tract of infants rather than the pathogen of diarrhea because of the low positive rate of Clostridium difficile toxin A. There was no gender difference in the detection rate of diarrhea pathogens.

Prevalence, toxin gene profile, antibiotic resistance, and molecular characterization of Clostridium perfringens from diarrheic and non-diarrheic dogs in Korea

Clostridium perfringens causes diarrhea and other diseases in animals and humans. We investigated the prevalence, toxin gene profiles, and antibiotic resistance of C. perfringens isolated from diarrheic dogs (DD) and non-diarrheic dogs (ND) in two animal hospitals in Seoul, Korea. Fecal samples were collected from clinically DD (n = 49) and ND (n = 34). C. perfringens was isolated from 31 of 49 DD (63.3%) and 21 of 34 ND dogs (61.8%). All C. perfringens strains were positive for the α toxin gene, but not for the β, ε, or ι toxin genes; therefore, all strains were identified as type A C. perfringens. All isolates were cpe-negative, whereas the β2 toxin gene was identified in 83.9% and 61.9% of isolates from DD and ND, respectively. Most isolates were susceptible to ampicillin (94%), chloramphenicol (92%), metronidazole (100%), moxifloxacin (96%), and imipenem (100%). However, 25.0% and 21.2% of isolates were resistant to tetracycline and clindamycin, respectively. Molecular subtyping of the isolated strains was performed by using pulsed-field gel electrophoresis. Fifty-two isolates were classified into 48 pulsotypes based on more than 90% similarity of banding patterns. No notable differences were observed among the isolates from DD and ND.

Clostridium difficile Infection: Is Clinical Phenotype Associated with Bacterial Toxin? / 胃肠病学

Clostridium difficile (C.difficile) infection is the leading cause of hospital-acquired diarrhea and pseudomembranous colitis.C.difficile-associated disease (CDAD) is mediated mainly by two bacterial toxins, TcdA and TcdB, which cause the diarrhea, as well as colitis and even intestinal necrosis.It has been indicated that level of C.difficile toxin is an important factor influencing the clinical phenotype of CDAD, however, the exact association between toxin and clinical phenotype remains unclear.In this article, we summarized the clinical phenotype of CDAD, the structure, function and regulatory mechanism of C.difficile toxin and discussed the relationship between C.difficile toxin and clinical phenotype, which may help to understand the pathogenic mechanism and provide possible therapeutic target for C.difficile infection.

Regulatory Mechanism of Clostridium difficile Toxin-associated Pathogenic Gene and Anti-toxin Treatment / 胃肠病学

Clostridium difficile( C. difficile)is a major nosocomial infection pathogen and the principal causative agent of antibiotic-associated diarrhea. The toxigenic C. difficile strains cause colonic injury and inflammation mainly by secreting enterotoxin A( TcdA)and cytotoxin B( TcdB). The severity of C. difficile associated disease( CDAD)is correlated to the toxin level during host infection. However,the toxigenic capacity of C. difficile varies widely among strains,which correlates with the gene regulation involved during toxin production. This article reviewed the regulatory mechanism of C. difficile toxin-associated pathogenic gene and anti-toxin treatment.

Evaluation of a real-time PCR assay for the qualitative detection of Clostridium difficile toxin gene and its clinical application / 中华检验医学杂志

Objective In comparison of the performances for the detection of Clostridium difficile toxin B genes from stool between BD MAX Cdiff assay and a laboratory-developed (LD) assay.The LD assay was evaluated in clinical application.Methods This study was a clinical application research.A total of 147 stool specimens from patients with diarrhea in Hangzhou First Hospital affiliated with Zhejiang Chinese Medical University were detected by the two assays from 1 July to 30 September 2014.DNA extraction and amplification of the tcdB gene were performed automatically on the BD MAX platform.Meanwhile, the tcdA and tcdB gene were detected by the LD real-time PCR assay after DNA extraction.Then, the results were analyzed by use of SPSS 10.0.Results A total of 147 stool samples were collected.There were 33 C.difficile positive cases and 114 negative cases detected by both of two assays.However, there were four stool samples had incongruent results.In comparison with BD MAX, the LD assay had a sensitivity of 93.94% (31/33), a specificity of 98.25% (112/114), a positive predictive value of 93.94% (31/33), and negative predictive value 98.25% (112/114).Furthermore, the results of the LD assay were statistically coherent with that of the BD assay (Kappa=0.922, P<0.01).Conclusions The LD assay was highly sensitive and accurate as BD MAX Cdiff assay in the detection of toxigenic Clostridium difficile.Furthermore, this LD assay could be also applied to detection of clinical stool samples directly with low cost.The assay will be more promising in diagnosis of toxigenic C.difficile in clinical application in China due to no additional instrument needed.

Influência da ativação ultrassônica do gel de hipoclorito de sódio 3 por cento, e da medicação intracanal com hidróxido de cálcio sobre micro-organismos e ácido lipoteicóico / Influence of activation by ultrasound on sodium hypochlorite 3% gel and an intracanal medication of calcium hydroxide on microrganisms and lipotheicoic acid

O objetivo deste estudo foi avaliar in vitro a ação do gel de hipoclorito de sódio (NaOCl) 3 % e da medicação intracanal (MIC) de hidróxido de cálcio (CaOH2), agitados ou não por ultrassom sobre Enterococcus faecalis, Escherichia coli e ácido lipoteicóico (LTA). Para isso 80 dentes humanos unirradiculares tiveram suas coroas removidas padronizando seu comprimento em 16 mm +- 0,5 mm, sendo seus canais instrumentados inicialmente até instrumento R25 (Reciproc). Os canais foram contaminados com suspensões de E. faecalis e E. coli. Os canais foram instrumentados utilizando-se instrumento R40, sob irrigação com 2 ml de NaOCl 3% gel seguida de irrigação com 10 ml de solução salina estéril e apirogênica. Após os canais foram irrigados com EDTA 17%, seguido de irrigação com 5 ml de solução salina estéril e apirogênica, sendo por último preenchidos com medicação intracanal de hidróxido de cálcio mantida durante 7 ou 14 dias. Os espécimes foram divididos inicialmente em 2 grupos (n=40) de acordo com a agitação ultrassônica (US) ou não da substância química auxiliar. Sendo novamente divididos de acordo com a agitação ultrassônica ou não da medicação intracanal (MIC) e o tempo de ação desta (n=10): 1) NaOCl + Ca(OH)2 (7 dias). 2) NaOCl + Ca(OH)2 (14 dias). 3) NaOCl + Ca(OH)2 com US (7 dias) 4) NaOCl + Ca(OH)2 com US (14 dias). 5) NaOCl com US + Ca(OH)2 (7 dias). 6) NaOCl com US + Ca(OH)2 (14dias) 7) NaOCl com US + Ca(OH)2 com US (7dias). 8) NaOCl com US + Ca(OH)2 com US (14 dias). Foram realizadas coletas do canal radicular 28 dias após o início da contaminação dos espécimes (1ª coleta), logo após o preparo biomecânico (PBM) (2ª coleta), logo após o preenchimento com EDTA (3ª coleta) e após o tempo de ação da MIC (4ª coleta). Para todas as coletas foram avaliadas a atividade antimicrobiana (UFC/ml) e quantificação de LTA pelo teste de Elisa. Os resultados foram submetidos aos testes estatísticos Kruskal-Wallis e Dunn (5%). Verificou-se pela análise microbiana e quantificação de LTA, que o NaOCl 3% gel foi capaz de eliminar quase completamente os micro-organismos dos canais radiculares, mas não o ácido lipoteicóico, independente da ativação ultrassônica. A ativação da MIC de Ca(OH)2 não exerceu efeito sobre micro-organismos. Sobre LTA, a ativação da MIC de Ca(OH)2 não foi eficaz, sendo os grupos com maior percentual de redução os que não sofreram agitação da MIC, nem do NaOCl. Conclui-se que o NaOCl 3% gel boa tem capacidade antimicrobina, independente da ativação ultrassônica. O PBM não foi capaz de detoxificar o LTA dos canais radiculares. A MIC com ativação ultrassônica não foi eficiente na redução de LTA(AU)

The aim of this study was to evaluate in vitro the action of sodium hypochlorite (NaOCl) 3% gel and calcium hydroxide as an intracanal medication, both activated by ultrasound on the reduction of E. faecalis e E. coli and lipotheicoic acid (LTA). Eight human single-rooted teeth with standardized size of 16 mm were prepared initially to R25 instrument (Reciproc) and distributed in microplate (n = 10). After sterilization (Co60 gamma radiation), the infection was carried out 8 microlitres of E. coli suspension, and after 7 days 8 microlitres of E. faecalis suspension, maintained for 21 days. Following the collection confirmation was performed (1st collection), then the root were instrumented using R40 instrument, irrigation with 2 ml of NaOCl 3% gel followed by washing with 10 ml of saline. The specimens were divided into 8 groups (n = 10) according to different ultrasonic irrigation protocols and different periods exposed to intracanal medication: 1) NaOCl + Ca(OH)2(7 days). 2) NaOCl + Ca(OH)2 (14 days). 3) NaOCl + Ca(OH)2 with PUI (7 dias) 4) NaOCl + Ca(OH)2 with PUI (14 days). 5) NaOCl with PUI + Ca(OH)2 (7 days). 6) NaOCl with PUI + Ca(OH)2 (14 days) 7) NaOCl with PUI + Ca(OH)2 (7 days). 8) NaOCl with PUI + Ca(OH)2 with PUI (14 days). Were made to sample colections of content of root canals, after instrumentation (2nd sample), after the use of EDTA (3rd sample) and after the Ca(OH)2 (4th Collection) . Microbiological culture and LTA quantification revealed that 3 % NaOCl gel was capable to eliminate E. faecalis and E. coli almost completely from root canals, but not lipotheicoic acid, regardless the use of ultrasonic activation. The ultrasonic activation of intracanal medication (Ca(OH)2) was not effective in removing neither microorganisms nor LTA. In addition, the groups with greatest reduction were the ones with no ultrasonic activation either of intracanal medication or NaOCl. It was concluded that 3% NaOCl gel is effective on antimicrobial activity regardless the use of ultrasonic activation. Biomechanical preparation was not capable to detoxify LTA from root canals. Ultrasonic activation of intracanal medication was not effetive on LTA reduction(AU)

Staphylococcal superantigen-specific IgE antibodies: degree of sensitization and association with severity of asthma / Anticorpos IgE específicos para superantígenos estafilocócicos: grau de sensibilização e associação com a gravidade da asma

ABSTRACT Objective: To determine the presence of staphylococcal superantigen-specific IgE antibodies and degree of IgE-mediated sensitization, as well as whether or not those are associated with the severity of asthma in adult patients. Methods: This was a cross-sectional study involving outpatients with asthma under treatment at a tertiary care university hospital in the city of Rio de Janeiro, Brazil. Consecutive patients were divided into two groups according to the severity of asthma based on the Global Initiative for Asthma criteria: mild asthma (MA), comprising patients with mild intermittent or persistent asthma; and moderate or severe asthma (MSA). We determined the serum levels of staphylococcal toxin-specific IgE antibodies, comparing the results and performing a statistical analysis. Results: The study included 142 patients: 72 in the MA group (median age = 46 years; 59 females) and 70 in the MSA group (median age = 56 years; 60 females). In the sample as a whole, 62 patients (43.7%) presented positive results for staphylococcal toxin-specific IgE antibodies: staphylococcal enterotoxin A (SEA), in 29 (20.4%); SEB, in 35 (24.6%); SEC, in 33 (23.2%); and toxic shock syndrome toxin (TSST), in 45 (31.7%). The mean serum levels of IgE antibodies to SEA, SEB, SEC, and TSST were 0.96 U/L, 1.09 U/L, 1.21 U/L, and 1.18 U/L, respectively. There were no statistically significant differences between the two groups in terms of the qualitative or quantitative results. Conclusions: Serum IgE antibodies to SEA, SEB, SEC, and TSST were detected in 43.7% of the patients in our sample. However, neither the qualitative nor quantitative results showed a statistically significant association with the clinical severity of asthma.

RESUMO Objetivo: Determinar a presença de anticorpos IgE específicos para superantígenos estafilocócicos e o grau de sensibilização mediada por esses, assim como se esses estão associados à gravidade da asma em pacientes adultos. Métodos: Estudo transversal incluindo asmáticos adultos em acompanhamento ambulatorial em um hospital universitário terciário no Rio de Janeiro (RJ). Os pacientes foram alocados consecutivamente em dois grupos de gravidade da asma segundo critérios da Global Initiative for Asthma: asma leve (AL), com asmáticos leves intermitentes ou persistentes, e asma moderada ou grave (AMG). Foram determinados os níveis séricos de anticorpos IgE antitoxinas estafilocócicas, e os resultados foram comparados por análise estatística. Resultados: Foram incluídos 142 pacientes no estudo: 72 no grupo AL (mediana de idade = 46 anos; 59 do sexo feminino) e 70 do grupo AMG (mediana de idade = 56 anos; 60 do sexo feminino). Na amostra geral, 62 pacientes (43,7%) apresentaram resultados positivos para dosagens de anticorpos IgE antitoxinas estafilocócicas: enterotoxina (TX) A, em 29 (20,4%); TXB, em 35 (24,6%); TXC, em 33 (23,2%); e toxic shock syndrome toxin (TSST), em 45 (31,7%). As médias das dosagens séricas de anticorpos IgE específicos anti-TXA, TXB, TXC e TSST foram, respectivamente, de 0,96 U/l, 1,09 U/l, 1,21 U/l, e 1,18 U/l. Não houve diferença estatisticamente significativa dos resultados qualitativos ou quantitativos entre os grupos. Conclusões: A presença de anticorpos IgE séricos anti-TXA, TXB, TXC e TSST, foi detectada em 43,7% nessa amostra de pacientes, mas não houve associação estatisticamente significativa entre seus resultados qualitativos ou quantitativos e gravidade clínica da asma.

Detection of PSM-mec by Vitek mass spectrometry for rapid identification of nosocomial-acquired methicillin-resistant Staphylococcus aureus / 中华检验医学杂志

Objective To evaluate the method of PSM-mec detection by Vitek MS for nosocomialacquired methicillin-resistant Staphylococcus aureus (MRSA) identification.Methods Totally 167 isolates of MRSA and 100 isolates of methicillin-sensitive Staphylococcus aureus (MSSA) used in this research were non-repetitively and prospectively collected between June 2012 and December 2013,two different SCCmec genotyping methods were applied for the MRSA strains,Vitek MS was used for identification of the isolates,the acquisition mass-spectrogram and the result mass-spectrogram at Myla system were analyzed among the different SCCmec type of MRSA.Results The 167 isolates of MRSA were classified into 5 major SCCmec types,among which SCCmec Ⅰ accounting for 3.6％ (6 isolates);SCCmec Ⅱ 6.0％ (10 isolates);SCCmec Ⅲ and Ⅲa 84.4％ (141 isolates);SCCmec Ⅳand Ⅳ a 4.8％ (8 isolates);SCCmec Ⅴ 1.2％ (2 isolates),respectively.The peak adjacent to the horizontal axis of a m/z 2 500 could be visually identified between the SCCmec Ⅱ and Ⅲ MRSA,of which the delta toxin peak were presented at m/z 3 005-3 009 or m/z 3 037-3 056,while the strains without delta toxin peak and the other types of MRSA or MSSA had no characteristic peak at the same position.Conclusions Nosocomial-acquired MRSA of the drug-resistant condition could be rapidly differentiated and forecasted by Vitek MS.Vitek MS could serve as a routine clinical assistance for epidemiological investigations of nosocomial-acquired MRSA in local area.

The etiology, molecular epidemiology and clinical characteristics of Clostridium difficile infections in Xiangya Hospital / 中华检验医学杂志

Objective A preliminary study on the etiology , the gene typing , the PCR-ribotyping and the clinical features of Clostridium difficile from clinical isolates at Xiangya Hospital could improve the isolation rate and provide the basis for effectively prevention of C.difficile.Methods A prospective observational study was performed.A total of 452 stool samples were collected during June to December 2012 at Xiangya Hospital.All stools were anaerobic cultured by selective medium and identified by API 20A for C.difficile.The positive isolates were detected the toxin genes ( tcdA, tcdB, cdtA, cdtB ) and ribotyping (16S-23S internal spacer region ) by PCR.The clinical data of all patients were collected and analyzed through Logistic regression to discover the risk factors for the development of C.difficile infection ( CDI ) . Results The rate of CDI occurrence was 13.94%(63/452), among them, 42.86%(36/63) were A-B+strains and only 14.29%(9/63) were obtained from community acquired-CDI.No binary toxin was detected in any of the isolates.Eleven different PCR ribotypes were identified , the dominant ribotype CD017 accounted for 22.22%(14/63).Logistic regression analysis showed that the risk factors for CDI included age>55(P=0.016;OR=4.45;95%CI:1.33-14.91), diarrhea frequency(P=0.007, OR=0.03;95%CI:0.002 -0.38 ) and the duration of diarrhea ( P =0.015; OR =7.86; 95%CI: 1.50 -41.16 ) . Conclusions C.difficile is the main pathogens of diarrhea patients and is mainly from hospital infections with higher detection rate of A -B+ in Xiangya Hospital.Ribotyping exist comparative advantages type CD017.No evidence suggests outbreak of C.difficile infection.

Modo de ação do biolarvicida Lysinibacillus sphaericus: identificação do epitopo da toxina Binária no receptor Cqm1 e base molecular da seletividade / Mode of action of biolarvicide Lysinibacillus sphaericus: identification of the epitope of the CQM1 Binary toxin receptor and molecular basis of selectivity

A toxina Binária (Bin) é o principal fator tóxico da bactéria entomopatógena Lysinibacillus sphaericus e sua ação em Culex quinquefasciatus depende da ligação com receptores no intestino das larvas. Os receptores são as a-glicosidases Cqm1, localizadas no epitélio, ligadas por uma âncora de glicosil-fosfatidilinositol. Larvas de Aedes aegypti são refratárias à toxina, pois, não apresentam receptores funcionais, apesar de apresentarem um gene que codifica a proteína Aam1, com alta similaridade à Cqm1. Devido às lacunas a respeito do espectro de ação da toxina Bin, o objetivo deste estudo foi identificar epitopos de ligação da toxina no receptor Cqm1 e determinar a base molecular da sua ação para estas espécies de vetores. Os resultados obtidos a partir da análise comparativa das proteínas Cqm1 e Aam1 levaram à identificação de um epitopo da toxina Bin no receptor Cqm1, situado uma alça na região N-terminal S129-A312. Este epitopo é composto pelos aminoácidos 155PATGGG160, não conservados em Aam1 (158AETGKL163), e os resíduos 159GG160 são críticos para a ligação com a Bin...

The Bin toxin is the main toxic factor of the bacterium Lysinibacillus sphaericuswhose action in Culex quinquefasciatuslarvae depends on its binding to the midgut epithelial receptors...