Neonatal rhesus monkeys as an animal model for rotavirus infection

AIM To establish a rotavirus(RV)-induced diarrhea model using RV SA11 in neonatal rhesus monkeys for the study of the pathogenic and immune mechanisms of RV infection and evaluation of candidate vaccines.METHODS Neonatal rhesus monkeys with an average age of 15-20 d and an average weight of 500 g ± 150 g received intragastric administration of varying doses of SA11 RV( 107 PFUs/mL, 106 PFUs/mL, or 105 PFUs/mL, 10 mL/animal) to determine whether the SA11 strain can effectively infect these animals by observing their clinical symptoms, fecal shedding of virus antigen by ELISA, distribution of RV antigen in the organs by immunofluorescence, variations of viral RNA load in the organs by qRT-PCR, histopathological changes in the small intestine by HE staining, and apoptosis of small intestinal epithelial cells by TUNEL assay.RESULTS The RV monkey model showed typical clinical diarrhea symptoms in the 108 PFUs SA11 group, where we observed diarrhea 1-4 d post infection(dpi) and viral antigen shed in the feces from 1-7 dpi. RV was found in jejunal epithelial cells. We observed a viral load of approximately 5.85 × 103 copies per 100 mg in the jejunum at 2 dpi, which was increased to 1.09 × 105 copies per 100 mg at 3 dpi. A relatively high viral load was also seen in mesenteric lymph nodes at 2 dpi and 3 dpi. The following histopathological changes were observed in the small intestine following intragastric administration of SA11 RV: vacuolization, edema, and atrophy. Apoptosis in the jejunal villus epithelium was also detectable at 3 dpi.CONCLUSION Our results indicate that we have successfully established a RV SA11 strain diarrhea model in neonatal rhesus monkeys. Future studies will elucidate the mechanisms underlying the pathogenesis of RV infection, and we will use the model to evaluate the protective effect of candidate vaccines.

Etiology,clinical features,and epidemiological analysis of diarrhea patients visiting a gastrointestinal clinic in a comprehensive hospital in Beijing,China,in 2023

Objective:To investigate the clinical features and epidemiology of diarrhea patients and analyze the current distribution of enteropathogens causing diarrhea in a comprehensive hospital in Beijing,China,in 2023.Materials and Methods:From April to October 2023,we enrolled patients with diarrheal diseases who visited the gastrointestinal Clinic in our hospital.The patients'demographic,epidemiological,and clinical features were obtained via a questionnaire.Stool samples were examined for 20 enteropathogens by multiplex polymerase chain reaction testing.Results:We enrolled 260 patients;men and adults accounted for 55.77%and 95.77%of the patients,respectively.The median age was 37 years.Eighty-four enteropathogens,72 bacteria and 12 viruses,were identified in 74 patients.Enteroaggregative Escherichia coli was the predominant agent.Patients with and without pathogens detected in stool samples showed no significant differences in age,sex,gastrointestinal symptoms,and stool characteristics.Possible food-related events were recorded in 57.31%of the patients.Leukocyte counts in patients with bacterial infections were higher than those of patients with viral infections and those with no detected pathogens(p<0.05).Seasonality of bacterial distribution was observed(p<0.05).Conclusion:Bacteria were predominant pathogens among the diarrhea patients.The incidence of diarrhea was related to hot weather and foodborne illness.Bacterial diarrhea may cause systemic infection.The clinical symptoms of infectious diarrhea were usually non-specific and unrelated to the type of infection.Timely and comprehensive multi-pathogen surveillance might be helpful to detect suspected pathogens and promote epidemic prevention and control.

Comparative study on epidemiological and etiological characteristics of patients with acute diarrhea with febrile or non-febrile symptoms in China

Background Acute diarrhea with fever can potentially represent a more severe form of the disease compared to non-febrile diarrhea. This study was to investigate the epidemiological characteristics and enteric pathogen composition of febrile-diarrheal patients, and to explore factors including pathogens associated with fever by age group.Methods A nationwide surveillance study of acute diarrheal patients of all ages was conducted in 217 sentinel hospitals from 31 provinces (autonomous regions or municipalities) in China between 2011 and 2020. Seventeen diarrhea-related pathogens, including seven viruses and ten bacteria, were investigated and their association with occurrence of fever symptoms was assessed using multivariate logistic analysis.Results A total of 146,296 patients with acute diarrhea (18.6% with fever) were tested. Th diarrheal children below 5 years had the highest frequency of fever (24.2%), and related to significantly higher prevalence of viral enteropathogens (40.2%) as compared with other age groups (P < 0.001). Within each age group, the febrile-diarrheal patients were associated with a significantly higher prevalence of bacterial pathogens than afebrile-diarrheal patients (allP < 0.01). There was discrepancy when each pathogen was compared, i.e., nontyphoidalSalmonella (NTS) was over-represented in febrile vs non-febrile patients of all age groups, while the febrile vs non-febrile difference for diarrheagenicEscherichia coli (DEC) was only significant for adult groups. The multivariate analysis revealed significant association between fever and infection with rotavirus A among children [odds ratio (OR) = 1.60], for DEC in adult groups (OR = 1.64), for NTS in both children (OR = 2.95) and adults (OR = 3.59).Conclusions There are significant discrepancy of the infected enteric pathogens in patients with acute diarrhea with fever between age groups, and it is valuable for priority detection of NTS and rotavirus A in patients with children < 5 years old and NTS and DEC in adult patients. The results may be useful in identifying dominant pathogen candidates for the application of diagnostic assays and prevention control.

Effects of Polygonum hydropiper on the expressions of inflammatory cytokines and cytochrome P450 enzymes in mice with E.coli-induced diarrhea

In the present study,we aimed to investigate the effects of Polygonum hydropiper aqueous extract(PHE)on the expression levels of inflammatory cytokines and cytochrome P450(CYPs)in mice with E.coli-induced diarrhea.BALB/c mice were randomly divided into the control group,model group,enrofloxacin-treated group,and two PHE-treated groups with different doses.The diarrhea model was established by intraperitoneal injection of enteropathogenic E.coli(EPEC)in mice.The enrofloxacin-treated group was given enrofloxacin at 5 mg/kg by intragastric gavage(i.g.)for 11 d.PHE-treated groups were given PHE at 5 g/kg and 10 g/kg by i.g.for 11 d.The histopathological characteristics of the duodenum and liver were observed by HE staining.The levels of inflammatory cytokines and CYPs in the duodenum and liver of mice were determined by ELISA.The m RNA and protein expressions of CYPs were determined by q RT-PCR and Western blotting analysis,respectively.The results showed that PHE could significantly alleviate the injury of the duodenum and liver induced by EPEC infection,reduce the contents and m RNA expressions of inflammatory cytokines,and regulate the m RNA and protein expressions of the major subtypes of CYPs.These findings indicated that PHE had an apparent therapeutic effect on EPEC-induced diarrhea,and its mechanism might be related to inhibition of inflammatory cytokines and regulation of CYPs.

江苏省某监测点家畜来源肠致病性大肠埃希菌分离株分子特征分析

目的分析江苏省某监测点家畜来源肠致病性大肠埃希菌分离株分子特征,进而评估其潜在的致病性,为感染性腹泻的监测与防控提供依据。方法采用全基因组测序技术对江苏省某监测点分离的家畜来源的37株肠致病性大肠埃希菌(EPEC)菌株进行特征性分析。结果本地家畜来源的EPEC均为非典型EPEC(aEPEC),其血清型多样,携带多种与腹泻相关的毒力基因,有9种ST型且具有区域性流行特征,eae基因包含有5种亚型,β1为优势亚型,其在腹泻病人中也最为常见。结论aEPEC对公众健康构成潜在威胁,应在感染性腹泻的监测工作中,加强对家畜粪便及养殖环境的监测。

猕猴肠道致病菌的检测

本文用分离培养的方法,对来自5个繁殖群的268只猕猴进行了沙门氏菌、痢疾杆菌、空肠弯曲菌和致病性大肠杆菌的检查,并观察了这些病菌对猕猴的致泻率。结果表明,26.5％(71/268)的猕猴带有肠道致病菌,其中36％的带菌动物在一个月内发生胰泻。

肠致病性大肠埃希菌的直接基因鉴定

目的 测定腹泻儿童的肠致病性大肠埃希菌 (EPEC)的bfpA和eaeA基因鉴定EPEC。 方法 用聚合酶链反应直接从粪便标本中和传统方法分离后获得的菌株中测定bfpA和eaeA基因。 结果 主要的血清型为O12 4K72 ,bfpA和eaeA基因存在于血清学凝集的 2 3株EPEC菌株中 ;直接从粪便标本中和传统方法分离后获得的菌株中测定bfpA和eaeA基因的结果有显著性差异 (P <0 0 1) ,两种基因的检出率有显著性差异 (P <0 0 5 )。

致婴幼儿腹泻的肠致病性大肠埃希菌血清分型和毒力基因检测

目的了解引起婴幼儿腹泻的肠致病性大肠埃希菌血清分型情况及其毒力基因携带状况。方法采集2011年6月至2012年2月医院儿科婴幼儿腹泻患者的粪便标本进行大肠埃希菌的分离培养,通过血清凝集试验和细菌生化反应筛选出肠致病性大肠埃希菌,采用多重PCR检测毒力基因escV、bfpA、Stx1和Stx2。结果 503例粪便标本中共分离出47株肠致病性大肠埃希菌,检出率是9.34%。血清分型有9种,最多的血清型是O86:k61,占了阳性菌株的27.66%。毒力基因escV阳性合计22株,bfpB阳性合计17株,检出率分别是46.81%和36.17%,其中,(escV+bfpB)阳性14株,escV阳性8株,bfpB阳性3株,携带毒力基因的菌株阳性率为53.19%,未检出毒力基因Stx1和Stx2。结论肠致病性大肠埃希菌血清学分型与毒力基因携带有差异,有条件的实验室可以直接检测大肠埃希菌的毒力基因,为临床提供更准确的诊断依据。

悬浮芯片法调查广州腹泻病原体分布及临床特征分析

目的:了解引起腹泻病原体种类随季节、年龄的分布,病原体种类与症状、实验检查结果的关系。方法:收集2014-2015年腹泻者粪便326例,使用液相悬浮芯片法检测15种病原体。结果:诺如病毒GⅠ/GⅡ型(15.6%,51/326)、A组轮状病毒(14.4%,47/326)和弯曲菌(12.6%,41/326)检出最多。寄生虫20例(6.13%),合并感染51例(15.6%)。秋冬季以A组轮状病毒(13.2%,43/326)、诺如病毒GⅡ型(9.8%,32/326)、弯曲菌(9.5%,31/326)为主。儿童(<14岁)A组轮状病毒(11.9%,39/326)常见,病原检出率较成人高。病毒与细菌感染引起隐血(P<0.001)、白细胞升高(P<0.001)、呕吐(P<0.01)比率有差异。结论:研究提供了腹泻病原体种类随季节、年龄分布的情况;细菌性与病毒性腹泻临床表现和实验检查有差异。

三峡库区仔猪传染性腹泻病原调查

为了弄清三峡库区仔猪传染性腹泻病原的感染及流行规律,本试验于2013-2015年从三峡库区的18个规模化猪场采集了180份发生腹泻的病料进行猪致病性大肠杆菌(EPEC)、猪流行性腹泻病病毒(FEDV)、猪传染性胃肠炎病毒(TGEV)和猪轮状病病毒(RDV)4种病原的检测,结果表明,三峡库区6个地区4种仔猪腹泻传染性病原检出率由高至低顺序为PEDV、RDV、TGEV和EPEC,依次为22%、12.7%.5.5%和3.8%,表明三峡库区规模化猪场PEDV和RDV感染广泛,且三峡库区规模化猪场存在多种病原混合感染情况。

2010～2011年北京市大兴区感染性腹泻监测

目的探讨北京市大兴区腹泻病例中肠道致病菌感染状况,分析其流行病学特征,为预防控制及临床治疗提供科学依据。方法采集监测点医院就诊的腹泻病例粪便标本,进行9种肠道致病菌的检测并对检出的志贺菌进行的药敏试验。结果 2年共采集样本760份,检出肠道致病菌167株,阳性率为21.97%,检出以志贺菌为主,其次为沙门菌、副溶血性弧菌。志贺菌对部分药物耐药性加强,对三代喹诺酮类药物保持敏感,但有所下降。结论近2年肠道致病菌阳性率上升,志贺菌菌型发生变化,目前以D群为主。夏季副溶血性弧菌感染增多,志贺菌对抗生素的耐药性发生一定程度改变,敏感性呈现整体下滑的趋势。

苦参提取液对肠致病性大肠杆菌感染小鼠的疗效研究

目的研究苦参提取液对肠致病性大肠杆菌标准株O10(EPECO10)感染小鼠肠道的保护作用。方法将50只小鼠平均分为五组,每组10只,分别为模型组、治疗组Ⅰ(高浓度,含生药浓度为0.500 g/m L)、治疗组Ⅱ(中浓度,含生药浓度为0.250 g/m L)、治疗组Ⅲ(低浓度,含生药浓度为0.125 g/m L)及空白组。空白组小鼠腹腔注射生理盐水,其余四组小鼠腹腔注射EPECO10菌液建立腹泻模型,治疗组Ⅰ、Ⅱ、Ⅲ给予不同浓度的苦参灌胃,模型组和空白组用生理盐水灌胃。比较五组小鼠腹泻指数、炎症评分及肠道中超氧化物歧化酶(SOD)的含量。结果模型组小鼠的腹泻指数、炎症评分及肠道SOD的表达与治疗组Ⅰ、Ⅱ、Ⅲ比较,差异均有统计学意义(P<0.01)。结论苦参提取液可以明显减轻小鼠的腹泻症状,改善其肠道的病理表现,具有良好的治疗效果。

腹泻患者粪便病原微生物培养方法的探讨

目的探讨腹泻患者粪便中肠道致病菌培养的方法。方法粪便标本由微生物检验人员常规做粪便性状观察和盐水涂片革兰染色或抗酸染色镜检,用5%血琼脂平板、中国蓝琼脂平板、SS琼脂平板、碱性蛋白胨水增菌液作一般细菌培养。结果 200例腹泻患者粪便病原微生物检验诊断回顾性分析,传统培养方法分离的沙门菌属和志贺菌属细菌已不是主要的肠道病原菌。结论传统的单一使用一种SS琼脂平板已经难以适应肠道致病菌发生的重大变迁,用5%血琼脂平板、中国兰平板、SS琼脂平板、碱性蛋白胨水增菌液对腹泻患者粪便培养,并在培养之前,由微生物检验人员常规做大便性状观察和盐水涂片革兰染色或抗酸染色镜检,以提高病原菌的检出率。

四川5个规模化养猪场仔猪腹泻的病原学研究

为了弄清2015年10月份四川5个规模化养猪场仔猪腹泻的病因,试验采用RT-PCR、PCR和常规细菌学的方法,参考Gen Bank登录序列合成8对PCR引物,分别用于扩增猪传染性胃肠炎病毒(TGEV)的S基因、猪流行性腹泻病毒(PEDV)的M基因、猪轮状病毒(PRV)的VP4基因、博卡病毒(PBo V)的VP1/VP2基因、猪库布病毒(PKo V)的3D基因,以及肠致病性大肠杆菌(EPEC)K88的fae G基因、K99的fan C基因、987P的fas G基因,其目标DNA片段大小分别为1 062 bp、467 bp、750 bp、496 bp、323 bp、792 bp、500 bp和551 bp。结果表明:SC1猪场病例样本扩增出PKo V 3D基因,大小为300 bp;SC2、SC3、SC4和SC5猪场病例样本均扩增出PBo V VP1/VP2基因,大小为500 bp;其他病原的检测结果均为阴性。经测序和遗传多样性分析表明,PKo V 3D基因、PBo V VP1/VP2基因与Gen Bank登录序列同源性分别在96%和98%以上。说明此次调查的5个猪场存在PBo V和PKo V的感染,但其感染来源和途径还不清楚。

实时荧光定量PCR法与常规细菌鉴定法在肠道致病菌检测中的应用效果对比分析

目的分析实时荧光定量聚合酶链反应(PCR)法与常规细菌鉴定法在肠道致病菌检测中的应用效果。方法选取2018年9月至2019年10月于我院就诊的360例腹泻患者为研究对象,采集所有患者的粪便标本并实施检测,同时进行实时荧光定量PCR法和常规细菌鉴定法检测,对比两种不同检测方法对肠道致病菌的检测阳性情况。结果360份粪便标本中,常规细菌鉴定法检测出14株沙门菌,阳性率为3.89%,6株志贺菌,阳性率为1.67%;实时荧光定量PCR法检测出15株沙门菌经,阳性率为4.17%,6株志贺菌,阳性率为1.67%。两种检测方法对沙门菌及志贺菌的检测阳性率比较,组间差异无统计学意义(P>0.05)。将实时荧光定量PCR法检测阳性患者的粪便标本再行常规细菌鉴定法检测,阳性率为100.00%。结论作为能够将DNA直接从粪便标本中提取的方法,实施荧光定量PCR法可对肠道疾病患者致病菌予以准确检测,同时操作简便,能够缩短诊断时间。

Molecular Identification of Shiga-Toxin Producing and Enteropathogenic <i>Escherichia coli</i>(STEC and EPEC) in Diarrheic and Healthy Young Alpacas

Isolation and biochemical and molecular identification of 303 strains of Escherichia coli obtained from diarrheic and healthy young alpacas of Puno-Peru, were realized. PCR amplification for 7 virulence factor genes associated with STEC, STEC O157:H7, EPEC: sxt1, sxt2, rfbO157, fliCH7, hlyA, eae y bfp were determined. A total of 39 strains (12.88%) showed amplification for one or more of these genes. Twenty three strains (59%) were classified as STEC and 16 strains (41%) as EPEC. An 88.18% (34/39) of STEC and EPEC strains were obtained from healthy alpacas and only 11.82% (5/39) from diarrheic alpacas considering this specie as potential zoonotic reservoir of STEC and EPEC.

北京地区腹泻就诊患者中肠致病性大肠埃希菌分离株特征分析

目的分析北京市通州区腹泻就诊患者中肠致病性大肠埃希菌(EPEC)分离株的特征。方法 eaeA基因聚合酶链反应(PCR)扩增阳性的菌株,进行系统生化鉴定、EPEC诊断血清分型、bfpA基因PCR扩增以及脉冲场凝胶电泳(PFGE)分析。结果 20株成年人腹泻患者EPEC分离株中,仅1株鉴定为O126血清群;4株儿童腹泻患者分离株中,仅1株鉴定为O26血清群。儿童与成年人腹泻患者EPEC分离株bfpA基因均为阴性。23株EPEC菌株的PFGE带型呈高度多态性,聚类图显示与患者年龄及性别无明显相关。结论北京市通州区腹泻患者EPEC分离株以非典型为主,单纯通过血清群来诊断EPEC的方法,将会造成极大的漏诊。

2013-2014年十堰城区儿童感染性腹泻病原学分析

目的：了解湖北十堰5岁以下儿童感染性腹泻流行特征和病原监测情况，为制定预防控制措施提供依据。方法：收集2013年1月至2014年12月湖北省十堰市太和医院5岁以下儿童临床诊断为腹泻病的粪便标本进行致病菌和病毒的检测。结果：从622份粪便标本共检阳性标本413例，阳性率为66．39％（413／622），其中轮状病毒246例、诺如病毒90例、札如病毒19例、星状病毒11例，致腹泻大肠埃希菌29例。轮状病毒发病高峰在10月至次年的1月，感染对象主要为6月-2岁婴幼儿；诺如病毒发病高峰在11月至次年1月，感染病毒的对象主要为6—12月婴幼儿，札如病毒、星状病毒感染无明显季节特征；细菌性腹泻发病高峰在5—9月，致腹泻大肠埃希菌为细菌性腹泻的最主要病原，感染对象主要为3～5岁婴幼儿。结论：2013—2014年湖北十堰城区感染性腹泻以2岁以下婴幼儿为主，轮状病毒为2岁以下儿童病毒性腹泻的最主要病原，致腹泻大肠埃希菌为3—5岁儿童细菌性腹泻的最主要病原菌。

北京市门头沟区感染性腹泻细菌病原谱构成及耐药性分析

目的了解2018—2019年北京市门头沟区感染性腹泻细菌病原体的构成及主要致病菌的耐药情况,指导抗生素的正确使用。方法收集2018—2019年北京市门头沟区感染性腹泻病例粪便标本进行沙门菌、志贺菌、致泻性大肠埃希菌(diarrheagenic Escherichia coli,DEC)、副溶血性弧菌、弯曲菌等病原菌的检测,应用微量肉汤稀释法进行沙门菌和DEC的药敏试验,并对病例资料进行统计学分析。结果745份粪便标本检出致病菌218株,其中DEC检出率最高,检出率为12.48%(93/745),其次为沙门菌,检出率为7.11%(53/745)。沙门菌对氨苄西林和萘啶酸的耐药率最高,为66.04%,其次分别为磺胺异噁唑、链霉素和氨苄西林-舒巴坦;DEC对氨苄西林耐药率最高,为65.59%,其次分别为萘啶酸、四环素和磺胺异噁唑。结论北京市门头沟区感染性腹泻病例DEC检出率最高,沙门菌和DEC耐药情况严重,应持续进行监测以掌握耐药谱变化趋势,指导临床正确用药。

2014-2015年广州地区腹泻患儿和健康儿童致泻性大肠埃希菌流行特征及耐药分析

目的 了解广州地区致泻性大肠埃希菌感染情况、流行病学特征及耐药情况。方法 2014年7月～2015年7月分别收集广州地区某综合医院腹泻患儿及健康儿童粪便标本143份、83份,进行细菌分离培养,采用聚合酶链反应检测肠致病性大肠埃希菌、聚集性大肠埃希菌、产毒性大肠埃希菌和肠侵袭性大肠埃希菌,Kirby Bauer法（K-B纸片扩散法）进行药敏试验。结果 四种致泻性大肠埃希菌在腹泻患儿与健康儿童中的检出率分别为1.4%、3.6%;2.8%、2.4%;1.4%、1.2%;0.7%、1.2%;腹泻患儿3～7岁年龄段检出率最高,占26.7%;然而,健康儿童中5～7岁年龄段检出率最高,占8.4%。药敏试验结果显示,大肠埃希菌对四环素、氨苄西林耐药率为52.6%～63.2%;头孢类耐药率为5.3%～21.1%;多重耐药率50.5%。结论 致泻性大肠埃希菌在腹泻患儿及健康儿童均有检出,并存在多重感染,提示致泻性大肠埃希菌是机会感染菌;除此之外,耐药及多重耐药高,应加强致泻性大肠埃希菌监测及管理抗生素使用。