A multiplex PCR/LDR assay for simultaneous detection and identification of the NIAID category B bacterial food and water-borne pathogens

Enteric pathogens that cause gastroenteritis remain a major global health concern. The goal of this study was to develop a multiplex PCR/ligation detection reaction (LDR) assay for the detection of all NIAID category B bacterial food and water-borne pathogens directly from stool specimens. To validate the PCR/LDR assay, clinical isolates of Campylobacter spp., Vibrio spp., Shigella spp., Salmonella spp., Listeria monocytogenes, Yersinia enterocolitica, and diarrheagenic Escherichia coli were tested. The sensitivity and specificity of the assay were assessed using a large number of seeded culture-negative stool specimens and a smaller set of clinical specimens from Haiti. The overall sensitivity ranged from 91% to 100% (median 100%) depending on the species. For the majority of organisms, the sensitivity was 100%. The overall specificity based on initial testing ranged from 98% to 100% depending on the species. After additional testing of discordant samples, the lowest specificity was 99.4%. PCR/LDR detected additional category B agents (particularly diarrheagenic E. coli) in 11/40 specimens from Haiti that were culture-positive for V. cholerae and in approximately 1% of routine culture-negative stool specimens from a hospital in New York. This study demonstrated the ability of the PCR/LDR assay to detect a large comprehensive panel of category B enteric bacterial pathogens as well as mixed infections. This type of assay has the potential to provide earlier warnings of possible public health threats and more accurate surveillance of food and water-borne pathogens.     

Superantigens produced by infectious pathogens: molecular mechanism of action and biological significance

Summary “Superantigens” have in common an extremely potent stimulatory activity for CD4⁺, CD8⁺, and some γδ⁺ T lymphocytes. Superantigens use a unique mechanism: they crosslink variable parts of the T cell receptor with MHC class II molecules on accessory or target cells. The interaction site on the T cell receptor is the variable part of the β-chain (V_β). There are several reasons why these molecules have aroused such tremendous interest in recent years. First, they have provided key information on tolerance mechanisms, both on the deletion of T cells in the thymus and on the induction of peripheral tolerance by anergy and apoptosis. Second, of all polyclonal T cell stimulators they are the ones that most closely mimic the recognition of specific antigen. Finally, they have been recognized as important factors in the pathogenicity of the producing pathogens, inducing shock and immunosuppression. Moreover, it has been postulated that superantigens could be involved in the pathogenesis of certain human diseases.     

Acute infectious aortitis presenting as pyelonephritis

A 65-years-old man with poorly controlled type 2 diabetes presented to the emergency department in sepsis with a 2-week history of new-onset fever, abdominal pain and pyuria. A Computed Tomography without contrast ruled out nephrolithiasis and hydronephrosis, but showed infiltration around the infra-renal aorta (5 x 1 cm) and several retroperitoneal lymph nodes. The periaortic infiltration raised suspicion for acute infectious aortitis. Aortic CT angiography confirmed an aortic pseudo-aneurysm in the infra-renal aorta without signs of impending aortic rupture. A Positron Emission Tomography-Computed Tomography confirmed abnormal 18F-FDG uptake adjacent to the right posterolateral aortic wall. The patient underwent abdominal aortic reconstruction with cryopreserved arterial allograft. Infectious aortitis is a rare but lethal cause of sepsis. Males over 50 years old with diabetes mellitus and peripheral vascular atherosclerotic disease are considered at higher risk. The diagnosis should be considered when a septic patient has vascular risk factors, abdominal pain, bacteremia and suggestive imaging. A CT without contrast showing infiltration around the aorta should, in some context, be promptly followed by Aortic CTA to search for acute aortitis among other less urgent differentials. Antibiotic therapy and involvement of vascular surgeons should be initiated in the emergency department.     

电解质紊乱与小儿感染性腹泻病原学的相关性

目的探讨电解质紊乱与小儿感染性腹泻病原学的相关性。方法用ISE法测定258例感染性腹泻患儿血清钾钠氯,用偶氮砷III法测定患儿血清钙,经细菌培养和快速轮状病毒抗原酶标诊断试剂鉴定感染性腹泻的致病原。结果病毒性腹泻患儿并发电解质紊乱80.7%(75/93);低钠血症的发生率在细菌感染性腹泻与病毒性腹泻组之间的差异具有显著性(p<0.01);虽然各种小儿感染性腹泻均可引起低钾血症,但在细菌组与病毒组之间的差异并不具有显著意义;高氯血症可见于各种感染性腹泻,以病毒性腹泻多见。结论小儿感染性腹泻为临床常见病、多发病,大多数感染性腹泻患儿伴有血清电解质紊乱,了解电解质紊乱与病原学的相关性,及时纠正各种电解质紊乱,有助于小儿感染性腹泻的及时治愈。     

婴幼儿感染性腹泻的致病菌及肠道病毒监测结果分析

目的了解导致深圳市东部地区婴幼儿感染性腹泻的主要致病茵及肠道病毒,获得该地区婴幼儿感染性腹泻病原谱,为预防控制疾病及临床用药提供参考依据。方法收集深圳市东部地区2010年1月至2012年12月腹泻婴幼儿的粪便标本1095例,进行致病菌常规培养,同时进行肠道病毒（包括轮状病毒、诺如病毒、星状病毒和肠道腺病毒）的核酸检测。结果1095例腹泻粪便标本中,检出致病菌215株,阳性率为19．63％,以致泻性大肠埃希菌为主,共135株,其中致病性大肠埃希菌最多．为57．04％,产毒性大肠埃希茵为37．04％,侵袭性大肠埃希菌为5．19％,检出1例肠出血性大肠埃希菌O157：H7;其次是沙门菌（4．11％）;再次是金黄色葡萄球茵（2．28％）。肠道病毒的总阳性率为23．83％,其中轮状病毒、诺如病毒、星状病毒和肠道腺病毒的检出率分别为9．04％、13．97％、0．73％和0．64％。结论每年6～12月为细菌感染腹泻的高峰期,深圳市东部地区婴幼儿腹泻病原菌以致泻性大肠埃希茵、沙门茵及金黄色葡萄球菌为主要病原菌,〈2岁的低龄儿童应警惕感染金黄色葡萄球菌的可能性。病毒性腹泻发病率呈逐年上升趋势．其中诺如病毒和轮状病毒是主要的病原体。每年秋、冬季为病毒感染发病高峰期。针对深圳市东部地区婴幼儿感染性腹泻的特点,应及时做好监测工作,应高度重视对防治工作的管理并加大人力、物力投入。     

Acute acalculous cholecystitis due to infectious causes

Acute acalculous cholecystitis (AAC) is an inflammation of the gallbladder not associated with the presence of gallstones. It usually occurs in critically ill patients but it has also been implicated as a cause of cholecystitis in previously healthy individuals. In this subgroup of patients, infectious causes comprise the primary etiology. We, herein, discuss the pathophysiological mechanisms involved in AAC, focusing on the infectious causes. AAC associated with critical medical conditions is caused by bile stasis and gallbladder ischemia. Several mechanisms are reported to be involved in AAC in patients without underlying critical illness including direct invasion of the gallbladder epithelial cells, gallbladder vasculitis, obstruction of the biliary tree, and sequestration. We emphasize that multiple pathogenic mechanisms may concurrently contribute to the development of AAC in varying degrees. Awareness of the implicated pathogens is essential since it will allow a more focused examination of the histopathological specimens. In conclusion, additional research and a high degree of clinical suspicion are needed to clarify the complex spectrum of mechanisms that are involved in the pathogenesis of AAC.     

2012年5—12月上海市感染性腹泻细菌和病毒病原学监测结果分析

目的了解2012年5—12月上海市感染性腹泻的病原谱和流行病学特征。方法随机抽取6所不同级别医院,对在其肠道门诊就诊的感染性腹泻病例按一定的抽样间隔进行调查,并采集粪便标本,进行细菌培养和病毒检测。结果从1001例病例粪便标本中检出阳性465例,检出率为46.45%。检出细菌7种、病毒5种。细菌检出率为12.09%,主要为副溶血性弧菌(61例)、沙门菌(35例)。病毒检出率为32.07%,主要为诺如病毒(262例)、轮状病毒(40例)。阳性例数呈较明显的夏季高峰和秋冬季高峰。465例阳性病例的平均年龄为(44.20±17.56)岁,病原阳性组和阴性组的年龄构成差异无统计学意义(P0.05)。细菌阳性组中有腹痛和发热症状的构成比均高于病毒阳性组,有呕吐和恶心症状的构成比均低于病毒阳性组;细菌阳性组发病前5天有聚餐史和外出史的构成比均高于病毒阳性组,二组有宠物接触史的比例均高于阴性组,上述差异均有统计学意义(P0.05)。结论上海地区腹泻病原谱较为广泛,季节高峰较为明显,应针对各病原检出率的差异在不同季节开展相应的防控措施。     

精神病患者医院感染性腹泻的发病状况调查及护理干预

目的 :探讨精神病患者并发医院感染性腹泻的发病状况与控制措施。方法 :对本院 1997年 6月至 2 0 0 1年 8月住院治疗的精神病患者 80 91例中并发医院感染性腹泻患者的发病状况进行调查分析。于 1999年下半年开始对 410 7例患者进行护理干预 ,并研究干预对患者医院感染性腹泻感染率的影响。结果 :干预前精神病患者并发医院感染性腹泻的感染率为 3 .87%。临床表现 :83例伴消化不良与胃肠道症状 (3 6.73 % ) ,3 9例有肠胀气、肠麻痹、肠梗阻、水与电解质紊乱及休克 (17.2 6% ) ,1例死亡 (0 .44 % ) ,10 3例腹泻无其他症状 (45 .83 % )。实验室检查结果 :共检出病原微生物 14种 ,除福氏痢疾杆菌外 ,其余均为条件致病菌 ,且伴多重抗生素耐药。对医院感染性腹泻采取护理干预措施后 ,患者医院感染性腹泻感染率由 3 .87%下降至 1.75 % ,感染率差异有统计学意义 (P <0 .0 1)。结论 :精神病患者医院感染性腹泻的感染率高 ,应引起重视 ,但采取适当的护理干预后 ,患者的医院感染性腹泻感染率可得到控制     

Hektoen肠道琼脂用于粪便中病原菌的首次分离

目的：推荐Hektoen肠道琼脂（HE琼脂）为选择性平皿,从粪便样品中分离沙门菌和志贺菌。方法：介绍了HE琼脂的制备和使用,并与SS琼脂同时对119份临床粪便样品作比较测定。结果：HE琼脂分离出21株沙门菌和志贺菌,而SS琼脂上仅分离出15株。结论：HE可以代替SS琼脂用于粪便样品中的病原体分离。     

北京市西城区引起感染性腹泻主要肠道病原对头孢菌素耐药状况分析

目的结合流行病学资料了解和分析北京市西城区主要肠道病原对头孢类抗生素的耐药状况。方法收集2008-2011年本地区引起感染性腹泻主要肠道病原187株,聚合酶链反应(polymerase chain reaction,PCR)检测超广谱β-内酰胺酶类(extended spectrumβ-lactamaseas,ESBLs)耐药基因,明确基因型,阳性产物在GenBank上进行比对确定亚型;按照Kirby-Bauer法进行药物敏感性试验,测定对8种头孢类抗生素耐药情况。结果 187株肠道病原中,34株检出携带ESBLs耐药基因,其中志贺菌中检出33株,沙门菌中检出1株。18株病原携带OXA-1亚型耐药基因,11株病原携带CTX-M-14亚型,另有5株病原携带有2种ESBLs耐药基因;药敏结果显示,肠道病原对头孢噻吩耐药率最高,为40%。对1种以上头孢类抗生素耐药的肠道病原中,志贺菌耐药率为81%,副溶血性弧菌耐药率为46%,沙门菌耐药率为33%;结合流行病学资料发现,20~29岁年龄组耐药菌株检出率最高,为27%,其次为≥40岁组和10岁组,检出率分别为23%和21%。结论本地区主要肠道病原中志贺菌耐头孢类抗生素水平最高,检出携带ESBLs耐药基因菌株最多;ESBLs耐药基因以OXA-1亚型和CTX-M-14亚型为主;耐药菌株以20~29岁年龄组检出率最高。     

一次食源性致病菌质控样的检验结果分析

为了保证全国食源性致病菌监测检验结果的可靠性,提高检验能力,本中心对中国疾控中心发放的2015年食源性致病菌考核盲样进行检测。结果检出单增李氏特菌、蜡样芽孢杆菌、阪崎杆菌。检测结果为优秀。     

腹泻患儿粪便中轮状病毒与肠道腺病毒检测结果分析

目的 了解本地区婴幼儿腹泻轮状病毒及肠道腺病毒感染情况.方法 采用胶体金免疫层析法检测7 705例腹泻婴幼儿粪便轮状病毒及肠道腺病毒抗原.结果 检出轮状病毒抗原阳性1 094例,RV阳性率为14.2％,其中RV-Ag和ADV-Ag阳性33例,占轮状病毒阳性样本的3％.在≤28 d,29 d～≤6个月,6个月～3岁,＞3岁四个年龄组中,RV阳性率分别为4.7％、8.1％、19.1％、7.8％.女性患儿轮状病毒感染率高于男性;10～12月份RV感染率明显高于其他月份;ADV感染率则以4～9月份较高,6个月～3岁婴幼儿腺病毒感染率明显高于其他年龄组患儿,女性患儿腺病毒阳性率也高于男性患儿.结论 轮状病毒和肠道腺病毒感染可以发生在儿童的各个年龄段,建议小儿出现腹泻症状时,应检测轮状病毒及腺病毒抗原.     

条件致病菌所致腹泻的病原学分析

目的条件致病菌所致腹泻的病原学分析。方法对近三年来临床粪便培养标本分离的有氧条件致病菌共336株进行菌种分布的分析和对三种临床腹泻类型进行菌种分布的比较。结果引致腹泻的条件致病菌中金黄色葡萄球菌占11．01％,肠杆菌料占56．83％,假单胞菌属占15．19％,弧菌科的气单胞菌属占9．53％,耶尔森氏菌占1．76％,真菌占5．65％。结论条件致病菌所致腹泻,急性者以金黄色葡萄球菌所致为主,迁延性和慢性者以肠杆菌科为主。应严密监测实验室中无病原菌生长而条件致病菌优势生长的粪便培养情况,及时向临床报告。     

Drug clearance by diarrhea induction

Perfusion of the gastrointestinal tract with electrolyte solution has been used as an equivalent to peritoneal dialysis in patients with renal failure. It was hypothesized that the back-diffusion into the gastrointestinal tract induced by this procedure could affect systemically circulating drugs as well. Nine dogs were given intravenous phenobarbital and subjected to gastrointestinal perfusion in attempt to lower serum levels. All dogs served as their own controls. Decline in serum phenobarbital level was used to calculate the amount of drug removed from circulation. Five hours after initiation of gastrointestinal perfusion, the experimental dogs had cleared 24.8% of the initial dose of phenobarbital, while controls cleared 3.13% of the initial dose of phenobarbital. This procedure could be of potential benefit to patients in emergency departments who have ingested toxic substances.     

Bacterial, yeast, parasitic, and viral enteropathogens in HIV-infected children from São Paulo State, Southeastern Brazil

We present here the frequency of enteropathogens in an HIV-infected children group and investigate their correlation with clinical and sociodemographic characteristics by collecting 100 stool samples from 55 HIV-seropositive Brazilian children. All specimens were processed according to standard methods for bacterial and yeast detection. A commercially available enzyme-linked immunosorbent assay was used to detect protozoan, and to perform virus detection, molecular tests were applied. Consumption of raw vegetables and fruits and severe immunosuppression were significantly associated with diarrhea. Cryptosporidium parvum was the commonest enteropathogen, followed by Candida albicans, enteropathogenic Escherichia coli, and astrovirus. The number of potential pathogenic agents identified in fecal specimens in asymptomatic HIV-seropositive infants is high, which raises the need for additional investigation in this area as well as in other Brazilian regions.     

Evaluation of a multiplex PCR assay for simultaneous detection of bacterial and viral enteropathogens in stool samples of paediatric patients

We evaluated a multiplex PCR assay, the Seeplex Diarrhoea ACE detection, that simultaneously detects 15 enteric pathogens, including Salmonella spp., Shigella spp., Vibrio spp., toxin B producer Clostridium difficile, Campylobacter spp., Clostridium perfringens, Yersinia enterocolitica, Aeromonas spp., Escherichia coli O157:H7, verocytotoxin-producing Escherichia coli, adenovirus, Group A rotavirus, norovirus GI and GII, and astrovirus. We compared this assay with clinical methods routinely used in our laboratory, for detecting enteropathogens in stool samples collected from 245 paediatric patients with suspected infectious gastroenteritis. We recovered 61 bacterial pathogens and 121 enteric viruses with our laboratory assays, while we detected 78 bacteria and 167 viruses with the molecular assay. We calculated specificity and sensitivity for both methods after analysis of discordant results and demonstrated greater sensitivity for multiplex PCR than for our routine methods, with the exception of Salmonella spp. and toxigenic C. difficile detection. The multiplex PCR assay proved to be a reliable tool to directly detect the most common enteropathogens in stool samples but with some limitations.     

新发再发病原体与血液安全

乙肝病毒、丙肝病毒及人类免疫缺陷病毒等输血相关病原体经输血传播风险逐渐减低,目前维持在较低水平,新发再发病原体越来越多被认识到其对输血安全构成威胁,其中一些已知的,如疟原虫、人细小病毒B19、戊型肝炎病毒、登革热病毒、人类T淋巴细胞白血病病毒及西尼罗病毒等已经被证实可经输血传播.其中,登革病毒曾经在中国爆发或流行过,将...     

季节性求和自回归移动平均模型在北京市房山区感染性腹泻发病趋势预测中的应用

目的 构建北京市房山区感染性腹泻发病的季节性求和自回归移动平均(seasonal autoregressive integrated moving average,SARIMA)模型并进行预测。方法 应用R 3.0.1软件程序包中的TSA对2004-2013年房山区感染性腹泻月发病率构建模型,并对2014年各月感染性腹泻月发病率进行预测和评价。结果 SARIMA(0, 0, 2)(0, 1, 1)12模型较好地拟合既往时间段月发病率,对2014年发病趋势拟合平均相对误差为19.164%,对年发病率拟合平均相对误差为2.303%。结论 SARIMA(0, 0, 2)(0, 1, 1)12模型能够很好拟合感染性腹泻月发病率数据,可用于房山区感染性腹泻发病趋势的短期预测,为下一步采取针对性防控措施提供科学依据。