一种用于艰难梭菌分离培养的新型显色培养基的性能评价

目的评价自主研发的艰难梭菌显色培养基(CDCA)性能。方法选用艰难梭菌及其他细菌的标准株进行CDCA显色的特异性评价;将艰难梭菌标准菌株梯度稀释后测定BD公司CDSA、生物梅里埃公司CDIF和CDCA的灵敏度。收集临床120份粪便样本分别采用3种培养基进行艰难梭菌分离培养,并对生长菌落进行质谱鉴定和tpi基因检测,以3种平板中任1种能培养出艰难梭菌的样本定义为真阳性。结果在CDCA平板上除了梭形梭菌、双酶梭菌、第三梭菌、脆弱拟杆菌等菌种能够少量生长并显色外,对其他实验菌种均有抑制作用。CDSA、CDIF和CDCA检测艰难梭菌的灵敏度分别为2.0×10~5 CFU/mL、8.0×10~1 CFU/mL和4.0×10~2 CFU/mL。120份标本中检出艰难梭菌31份(25.8%),CDSA、CDIF和CDCA培养阳性标本分别为25份(20.8%)、28份(23.3%)和26份(21.7%),3种显色培养基对临床腹泻标本检测阳性率差异无统计学意义(χ~2=0.418,P=0.811)。3种显色培养基的敏感性、特异性、阳性预测值、阴性预测值分别为:CDCA 83.8%、100%、100%、94.7%;CDIF 90.3%、98.9%、96.6%、96.7%;CDSA 80.6%、100%、100%、93.7%。结论本实验室研发的CDCA的特异性和灵敏度较高,可用于临床艰难梭菌的初步分离。     

难辩梭菌培养方法的比较

抗生素相关性腹泻或结肠炎,因抗生素的广泛应用而日益增多并逐渐为人们所注意。近年来对其病原学有了新的认识,一般认为难辨梭菌(Clostridiumdifficile)为其主要病原菌。选择性培养基甲氧头孢霉素—环丝氨酸—果糖琼脂(简称CCFA,下同)的出现,大大方便了从粪便中分离难辨梭菌。以后又有人倡用加有牛磺胆酸钠的CCFA(简称TCCFA),     

GeneXpert实时荧光定量PCR在诊断艰难梭菌感染中的临床应用

目的评价Gene Xpert实时荧光定量PCR法在诊断艰难梭菌感染中的应用价值。方法收集住院腹泻患者非重复粪便标本296份,同时进行Gene Xpert试验和产毒素培养(toxigenic culture,TC)。TC包括厌氧培养、菌株基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)鉴定、PCR扩增艰难梭菌毒素基因。以TC结果为参考,评价Gene Xpert试验的敏感性、特异性、阳性预测值、阴性预测值,并评价Gene Xpert试验和TC 2种方法结果的一致性。结果 Gene Xpert试验的敏感性、特异性、阳性预测值和阴性预测值分别为98.5%、92.6%、91.7%和98.7%。Gene Xpert试验与TC一致性好(Kappa=0.905)。Gene Xpert试验报告2株RT027型高毒力菌株,经核糖体分型确证为RT027型艰难梭菌。结论 Gene Xpert实时荧光定量PCR可快速、准确检测粪便标本中的艰难梭菌毒素基因,且可报告高毒力RT027型菌株,具有重要的临床应用价值。     

激光解析质谱联合CHROMagar显色培养技术鉴定环境表面MRSA的效果评价

摘要 目的 〖HT5"SS〗评价激光解析质谱联合CHROMagar MRSA显色培养基在鉴定环境表面耐甲氧西林金黄色葡萄球菌（MRSA）的应用价值。方法 采用CHROMagar MRSA显色培养技术,对5家医院重症监护病房（ICU）环境表面现场采集的标本中MRSA进行培养鉴定。结果 从CHROMagar MRSA显色培养基上获得573株可疑菌落,经MALDI-TOF-MS鉴定确认MRSA菌株147株,检出率为25.7%。同时用常规PCR方法和头孢西丁药敏试验验证确认MRSA菌株分别为145株和140株,符合率分别为98.6%和95.2%。结论 MALDI-TOF-MS技术联合CHROMagar MRSA显色培养基可作为更快速、更准确的MRSA筛查方法,适用于医院感染的快速诊断与防控。     

北京市某院难辨梭菌临床分离株耐药性及毒素基因携带情况分析

目的了解航天中心医院难辨梭菌临床分离株的耐药性及毒素基因携带情况。方法收集不明原因腹泻患者的粪便样本335例,采用TaqMan-MGB探针实时荧光定量聚合酶链反应(PCR)进行菌属鉴定及毒素基因检测。对tpi基因阳性样本进行厌氧培养、分离及药物敏感性试验。回顾性分析分离出A~+B~+菌株和A~-B~+菌株患者的临床资料。结果 335例粪便样本中,56例(16.7%)检出难辨梭菌tpi基因,其中10株(17.9%)A~-B~+菌株、46株(82.1%)A~+B~+菌株。检出菌株对阿莫西林、甲硝唑、克林霉素、万古霉素、莫西沙星、氨苄西林-舒巴坦及美罗培南的耐药率分别为7.1%、10.7%、75.0%、0、57.1%、5.4%、8.9%。有29株(51.8%)菌对克林霉素、莫西沙星双重耐药。分离出2种菌的患者病史、临床用药史、常规实验室检测指标差异均无统计学意义(P0.05)。结论航天中心医院难辨梭菌临床分离株以多重耐药模式多见,A~+B~+菌株和A~-B~+菌株感染患者临床特征无差异。     

婴儿艰难梭菌携带状况及菌株特征研究

目的调查婴儿艰难梭菌的携带状况及菌株特征。方法收集2015年8-11月在该院住院或门诊治疗的1岁内婴儿粪便标本238份,利用免疫层析法快速初筛艰难梭菌,阳性标本再利用CDIF平板进行厌氧培养以获得菌株,利用PCR方法检测艰难梭菌毒素A、B的编码基因tcdA、tcdB和二元毒素编码基因cdtA、cdtB,运用slpA测序分型(slpA ST)方法对菌株进行基因分型。结果 238份粪便标本共分离出50株艰难梭菌,3月、3~6月和6月至1岁三组婴儿艰难梭菌的分离率分别为9.3%,17.6%和27.3%,三组见比较差异有统计学意义(χ~2=6.940,P=0.0310.05)。52.0%(26/50)的菌株为产毒株,其中69.2%(18/26)的菌株产毒模式为tcdA+tcdB+cdtA-cdtB-。50株艰难梭菌可分为11种slpA ST型,产毒株最常见的基因型为slpA ST fr-02和kr-02,而非产毒株则为xr-03。结论 1岁内婴儿艰难梭菌携带率较高,且过半为产毒株,大多同时产毒素A和B。产毒株与非产毒株的基因型存在差别。     

艰难梭菌毒素基因检测及分子流行病学分析

目的了解腹泻患者艰难梭菌感染现状,并分析分离菌株的核糖体分型情况。方法收集161例住院腹泻患者粪便标本进行艰难梭菌毒素基因PCR检测,同时进行产毒培养法,并比较2种方法的一致性;对分离的艰难梭菌进行核糖体分型。结果艰难梭菌产毒培养法阳性率为9.94%(16/161),粪便艰难梭菌毒素基因阳性率为9.94%(16/161),2种方法结果差异无统计学(P0.05),一致性较好(Kappa0.75)。培养所得18株艰难梭菌中A、B毒素基因均阴性的2株(11.11%),tcdA~+tcdB~+产毒株15株(83.33%),tcdA~-tcdB~+1株(5.56%)。18株艰难梭菌核糖体分型分为16种型别(GS1～GS16),未发现核糖体分型027型菌株。结论艰难梭菌粪便毒素基因PCR检测可用于临床诊断,未发现本院艰难梭菌暴发流行。     

耐万古霉素肠球菌两种筛选方法的比较及耐药性分析

目的提高耐万古霉素肠球菌的检出率,为临床选药治疗提供依据。方法将粪便直接接种在含万古霉素琼脂平板(VAP)和含万古霉素肉汤(VB)培养基中,分离菌株用Vitek-AMS仪器鉴定,根据美国临床实验室标准化研究所(CLSI)指南做药敏试验。结果VAP培养基筛选出耐万古霉素肠球菌38株(13.1%),VB培养基筛选出49株(16.9%)。从住院病人粪便中分离出31株,其中22株VRE为屎肠球菌,9株VIE为鹑鸡肠球菌;从正常成人粪便分离出25株VIE,其中17为株鹑鸡肠球菌,8株为铅黄肠球菌;22株VRE对万古霉素的MIC>512μg/ml,36株VIE对万古霉素的MIC值为8~16μg/ml。56株(VRE VIE)对环丙沙星、红霉素耐药率均为75.0%,氨苄西林为80.4%,左氧氟沙星为71.4%,高水平庆大霉素为69.6%,呋喃妥因为57.1%,高水平链霉素为41.1%,氯霉素耐药率较低为3.6%。结论VB培养基对耐万古霉素肠球菌的检出率高于VAP培养基。耐万古霉素肠球菌对多种抗菌药物耐药。     

Xpert艰难梭菌检测系统的临床应用评价

目的评估Xpert艰难梭菌检测系统的临床应用价值。方法选取43份腹泻粪便标本,采用厌氧培养法、VIDAS检测A/B毒素法和Xpert艰难梭菌检测系统检测艰难梭菌,以产毒培养法作为金标准对检测方法进行评价,并比较不同检测方法对临床标本检测结果的一致性。通过自配027型标准菌株模拟粪便标本,验证Xpert艰难梭菌检测系统筛选027型流行株的能力。结果以产毒培养法为金标准,Xpert艰难梭菌检测系统的敏感性和特异性分别为90.9%和93.8%,阳性和阴性预测值分别为83.3%和96.8%。与产毒培养法结果一致性检验Kappa值为0.822(P0.05),与VIDAS检测A/B毒素法结果一致性检验Kappa值为0.419(P0.05);027型标准菌株模拟粪便标本检测结果阳性并报告为027型。结论 Xpert艰难梭菌检测系统能够快速准确地检测粪便标本中的艰难梭菌相关基因,且能准确报告027型高产毒力菌株。     

艰难梭菌实验室不同检测方法的比较

目的比较不同方法检测艰难梭菌(CD)的临床可行性。方法选取2016年疑似抗菌药物相关腹泻患者大便标本,采用酶联免疫吸附试验(ELISA)和鉴定培养基培养法进行检测,比较两种检测方法的灵敏性、特异性和一致性。结果 ELISA检出抗原阳性标本29例,其中毒素阳性25例,毒素阴性4例;鉴定培养基培养法检出可疑菌株29例,经质谱仪鉴定,其结果为CD28例,1例未检出种属;ELISA和鉴定培养基培养法的灵敏度和特异度分别为97%、100%和93%、95%,两者有极好的一致性(Kappa=0.92)。结论 ELISA具有快速、高效、省时、操作简单、判读容易等特点,可快速准确地筛检CD相关性腹泻病患;培养基培养法特异性好,可以快速得到感染株;二者联合使用可大大提高CD检出率,并对后期治疗有很大帮助。     

Treatment of Clostridioides (Clostridium) difficile infection

Abstract

Clostridioides (formerly: Clostridium) difficile infection (CDI) is a major cause of diarrhoea for inpatients as well as outpatients. Usually, CDI is healthcare-associated but the number of community-acquired infections is increasing. CDI is generally associated with changes in the normal intestinal microbiota caused by administration of antibiotics. Elderly and immunocompromised patients are at greater risk for CDI and CDI recurrence. Recently, the treatment options of CDI have undergone major changes: current recommendations speak against using metronidazole for primary CDI, fidaxomicin and bezlotoxumab have been added to the treatment armamentarium and microbial replacement therapies have emerged. Several other therapies are undergoing clinical trials. In this article, we review current treatment guidelines, present the most recent data on the options to treat CDI and glance towards future developments.

• KEY MESSAGES

• The cornerstones for the treatment of CDI are vancomycin and fidaxomicin. Metronidazole should be used only in mild-to-moderate disease in younger patients who have no or only few risk factors for recurrence.

• In recurrent CDI, bezlotoxumab infusion (a monoclonal antibody against C. difficile toxin B) may be considered as an adjunctive therapeutic strategy in addition to the standard care provided to patients with several risk factors for recurrence.

• Faecal microbiota transplantation (FMT) should be offered to patients with frequently recurring CDI.

     

进口培养基对卡他布拉汉菌分离鉴定的评价

目的了解呼吸道感染病人痰中卡他布拉汉菌的分离率方法采用法国生物梅里埃公司生产的哥伦比亚琼脂和DNA酶琼脂分离痰中的卡他布拉汉菌和做关键性的生化反应,鉴定结果与APINH试条进行对照。同时用国产培养基做对比试验结果卡他布拉汉菌在哥伦比亚琼脂配制的血平板上菌落明显大于国产血平板,与非致病性奈瑟氏菌有显著差别。关键性的生化反应在DNA酶琼脂平板上均显示阳性。鉴定结果的准确性与APINH试条完全相同。国产的DNA酶微量生化管仅4．4％为阳性。结论哥伦比亚琼脂和DNA酶琼脂是分离鉴定痰中卡他布拉汉菌良好的试剂。结合痰涂片镜检见到大量脓细胞吞噬G-双球菌的现象,我科在98年409例痰中共分离出45例卡他布拉汉菌,分离率为11％。占痰中各类病原菌的第4位。     

Prevalence of Clostridium difficile colonization at admission to rehabilitation

OBJECTIVES: To assess the prevalence of intestinal colonization with Clostridium difficile (C. difficile) at admission to acute rehabilitation and to identify risk factors associated with colonization. DESIGN: Case-control study. PARTICIPANTS: Consecutive admissions to 2 rehabilitation units (spinal cord injury, brain injury and stroke). SETTING: Free-standing acute rehabilitation facility. INTERVENTIONS: Rectal swabs for culture for C. difficile were obtained at admission and cytotoxin assay performed on all culture positive specimens. Rates of colonization with cytotoxic C. difficile were calculated. Charts were reviewed for medical and demographic factors that may have predisposed patients to colonization, and for possible symptoms at the time of admission. MAIN OUTCOME MEASURES: Percentage of patients with culture and cytotoxin assay positive for C. difficile. Frequency of specific patient characteristics that could predispose to C. difficile colonization. RESULTS: Of admission stool samples, 16.4% tested positive for C. difficile; none of these patients had been identified as colonized before admission. No patients were discordant for C. difficile positivity on culture and presence of a toxigenic strain. No medical or demographic factors were associated with increased risk of colonization, including age (t(52)=-.748, P=.458, not significant [NS]), diarrhea within 24 hours of admission (chi(1)(2) test=.001, P=.973 [NS]), or use of oral or intravenous antibiotics at admission (chi(1)(2) test=.044, P=.834 [NS]). CONCLUSIONS: Patients admitted to acute rehabilitation may have an elevated rate of intestinal colonization with C. difficile without having clinical symptoms. No medical or demographic characteristics were found to be predictive of colonization, however, most of the patients admitted had more than 1 factor that may have increased their susceptibility to infection with this organism. Inadvertent transfer of this organism within the rehabilitation setting may occur because asymptomatic colonization is not recognized.     

Clinical management of severe,fulminant, and refractory Clostridioides difficile infection

ABSTRACT

Introduction: Up to 15% of hospitalized patients with Clostridioides difficile infection (CDI) develop severe CDI (SCDI) or Fulminant CDI (FCDI). Due to high rates of mortality in medically-refractory CDI cases, 30% of patients with severe infection historically require surgical intervention. However, colectomy itself is an imperfect solution because it is difficult to predict who will fail medical therapy, patients with SCDI are more likely to have underlying medical conditions that make them poor surgical candidates, and post-surgical mortality still approaches 30–50%.

Areas covered: This review will serve as a clinically-based review of severe and fulminant CDI management including discussion of models to predict severe infection, emerging treatments, novel targets for therapy, and innovations in surgical management.

Expert opinion: Among the most promising studies to emerge in the last decade have involved fecal microbiota transplantation (FMT), which is already recommended by multiple society guidelines for recurrent CDI (RCDI). In the case of SCDI/FCDI, multiple studies have safely and successfully utilized FMT to produce rates of cure in the 70–90% range. Additionally, patients who have FCDI refractory to medical therapy and are poor candidates for colectomy may benefit from FMT as salvage therapy.     

消毒剂对淋球菌杀灭效果检测方法的研究

为选择消毒剂对淋球菌杀灭效果的较好检测方法,对不同培养基与接种方法进行了试验比较。结果,用平皿倾注(即将菌悬液接种平皿后倾注琼脂并混匀)接种,在培养基上生长的菌落<1 mm或不透明;用平板浸润法(即将菌悬液接种平板后轻轻旋转,让菌悬液均匀浸润平板)接种,在普通营养琼脂、酵母浸膏琼脂上生长的菌落<2 mm,菌落计数<2 515 000 cfu/ml,在酵母浸膏血琼脂、巧克力琼脂上者2～3 mm与>3 320 000 cfu/ml。结果表明,对淋球菌用平板浸润法接种比常用的平皿倾注法好,用酵母浸膏血琼脂培养结果与现用巧克力琼脂者无明显差别。     

绿脓杆菌污染调查用选择培养基的研究

新研制的改良NAC培养基,经医院拖布标本检测,其对绿脓杆菌的检出阳性率与NAC培养基比较,虽无显著差别。但改良NAC培养基上的绿脓杆菌菌落较大,绿色素亦较显著,分离时易于识别。     

Fidaxomicin: a minimally absorbed macrocyclic antibiotic for the treatment of Clostridium difficile infections

Fidaxomicin was approved for the treatment of Clostridium difficile infections in 2011. It has a novel mechanism of action and narrow spectrum of activity that makes it unique among the currently used therapies for this disease. Phase III clinical studies demonstrated a benefit of fidaxomicin over vancomycin for the outcomes of recurrence and global cure or sustained clinical response. This observation was confirmed within specific populations, including those of older age, immunocompromised due to active cancers, and patients taking concomitant antibiotics. Additionally, fidaxomicin significantly reduced recurrence rates compared to vancomycin among patients receiving treatment for recurrent C. difficile episodes. Fidaxomicin represents an advance in therapy for the treatment of C. difficile infections.     

Nutrient agar with sodium chloride supplementation for presumptive detection of Moraxella catarrhalis in clinical specimens

We previously reported that Nissui nutrient agar (N medium) promoted the growth of Moraxella catarrhalis but not commensal Neisseria spp. In the present study, we examined which constituent of N medium was responsible for the selective growth of M. catarrhalis using 209 M. catarrhalis and 100 commensal Neisseria spp. clinical strains. We found that peptone, but not meat extract or agar of N medium, had growth-promoting or growth-inhibiting ability with respect to M. catarrhalis and commensal Neisseria spp. Thus, we investigated the amino acid content of N peptone and found it had higher concentrations of amino acids than other commercial peptone products. On varying the sodium chloride concentration of reconstituted N medium, we noted that the concentration was an important factor in bacterial growth differences. Varying the sodium chloride concentration of other commercial nutrient agars achieved similar results to those for N medium. This is, to our knowledge, the first study observing that sodium chloride concentration is responsible for difference in growth between the two organisms. We also successfully isolated colonies of M. catarrhalis from respiratory specimens on N medium, whereas the growth of commensal Neisseria spp. was inhibited, and by adding bovine hematin and β-NAD we were able to isolate Haemophilus influenzae colonies as efficiently as with a chocolate agar. In conclusion, nutrient agar can be used as a medium for the preferential isolation of M. catarrhalis from upper respiratory tract specimens.