巢式聚合酶链反应快速检测血液标本中伤寒沙门菌的实验研究

目的 建立一种检测血液中伤寒沙门菌的方法。方法 根据伤寒沙门菌特异鞭毛基因设计两对引物,通过巢式聚合酶链反应（PCR）扩增伤寒沙门DNA片断,扩增产物通过凝胶电泳进行分析。结果 用伤寒沙门菌DNA系列稀释进行试验,巢式PCR能够检出10个伤寒沙门菌。36份培养阳性标本,33份PCR阳性;6份培养阴性但临床高度可疑的伤寒病人标本PCR阳性;10份其它原因引起发热的临床标本均为阴性。结论 巢式PCR能够快速、特异、准确地检出血液中的伤寒沙门菌。     

伤寒、甲、乙、丙型副伤寒沙门菌多重荧光PCR方法的评估及临床应用

目的建立和评估多重荧光PCR方法快速鉴定伤寒、甲、乙、丙型副伤寒沙门菌的方法,并应用于临床血培养样品的检测。方法收集留取医院的可疑伤寒患者的血培养标本,用多重实时荧光PCR法和传统分离培养法同时进行分离鉴定,分析比较双盲实验结果,评价多重实时荧光PCR方法的灵敏度、特异度等检测性能指标。结果共检测临床样本538例,多重荧光PCR法检出阳性218例,比传统培养法多检出6例;阴性320例,与传统培养法一致。以传统检测方法为金标准,多重荧光PCR方法的灵敏度为100%,特异度为98.2%。结论建立的多重荧光PCR检测方法操作简便,可快速、特异、灵敏地检测出伤寒、甲、乙、丙型副伤寒沙门菌,可以应用于临床标本的早期快速分型诊断,提升重大传染病的应急处置能力和监测能力。     

应用荧光定量PCR检测食品中沙门菌

目的应用荧光定量PCR技术检测食品中沙门菌,为病原监测和快速诊断提供实验依据。方法选择沙门菌invA基因设计引物,应用SYBRGreen I染料建立荧光定量PCR法。分别对61株沙门菌、14株非沙门菌以及食品模拟标本、市售禽蛋制品进行检测,观察方法的特异性、敏感性和可行性。结果建立的荧光定量PCR法检测所有沙门菌均出现了特异的熔解曲线,扩增产物的熔点值79.6℃左右;目标菌DNA的扩增产物循环数（Ct值）为20,空白对照及非沙门菌的Ct值大于30,非沙门菌均未出现特异的熔解曲线。每反应最低检测的沙门菌数是31 CFU;对食品中人工污染的沙门菌的检测,最低检测量是1 CFU,检测时间为7～8 h,与培养法比较,阳性符合率100%;对市售的369份禽蛋制品进行检测,阳性结果 19份,而细菌培养法检测阳性结果为12份。结论基于SYBRGreen I染料建立的荧光定量PCR检测沙门菌是敏感、特异、省时、省力的方法,适用于沙门菌快速检测。     

荧光PCR法与细菌培养法在鼠伤寒沙门菌食物中毒检测中的应用比较

目的把荧光PCR法与细菌培养法在鼠伤寒沙门菌食物中毒检测中的应用进行比较。方法使用以往的细菌培养方式和荧光PCR方式对10份样本实施检验。结果这10份样本经过检验之后,PVR检验出沙门菌5株,阳性概率相对较高为50%,以往的方式检验出3株沙门菌,阳性概率为30%相对较低。结论与以往的培养方式进行对比,使用PCR方式能够增强检验出鼠伤寒沙门菌的概率,节约检验时长,合适用在食物中毒这一方面的检验。     

CAS、SS、HE培养基检测沙门菌敏感性分析

目的：通过比对试验了解显色培养基CAS和普通培养基SS、HE对沙门菌检测的敏感性。方法：对320份SS、HE培养阴性的腹泻便,用CAS培养基再次复检,确定阳性者再用SS、HE做对应检测。结果：320份SS、HE阴性标本,经CAS复检,得阳性标本6份,漏检率1.87%,6份阳性标本SS检出2份,HE检出1份。结论：SS、HE对沙门菌的敏感度低于CAS。     

16S rRNA及18S rRNA同时检测脑脊液中的病原菌

目的 提高脑脊液中病原菌的快速、准确鉴定率。方法 以细菌的保守序列16SrRNA和真菌的18SrRNA为通用引物，用多重PCR技术对标准菌株及46例中枢神经系统感染患者的脑脊液进行扩增，同时与脑脊液的培养结果进行比较。结果 标准菌株金黄色葡萄球菌、大肠埃希菌、脑膜炎奈瑟菌、结核分枝杆菌、化脓性链球菌及白色念球菌均出现特异条带，人白细胞DNA为阴性。46份脑脊液标本培养阳性39例，另有2例PCR阳性而培养阴性，后经诱导传代培养证实为L型金黄色葡萄球菌。PCR阳性41例，与培养分离鉴定结果符合率为100％。结论 多重PCR可快速鉴定脑脊液中病原菌，为临床及时治疗提供必要的诊断信息。     

628例食源性腹泻婴幼儿粪便与肛拭子标本中病原菌的培养及主要致病菌谱的分析及其防治对策

目的:分析食源性腹泻婴幼儿粪便与肛拭子标本中病原菌培养及主要致病菌构成情况与防治对策.方法:选取医院诊疗的食源性腹泻婴幼儿(2016年4月-2020年2月)628例临床资料,分析其粪便与肛拭子标本中病原菌培养结果及其主要致病菌构成情况,并提出干预措施.结果:628例患者标本中,经细菌培养结果显示阳性301例(检出病原菌301株),其阳性率为47.93％;食源性腹泻婴幼儿感染致病菌主要为沙门菌属占41.86％;对沙门菌属分布情况分析发现,其中肠炎沙门菌、鼠伤寒沙门菌、斯坦利沙门菌为主要致病菌.结论:食源性腹泻婴幼儿感染病原菌主要为沙门菌属、副溶血弧菌、致病性大肠埃希菌、志贺菌属;而肠炎沙门菌、鼠伤寒沙门菌、斯坦利沙门菌为沙门菌属中主要致病菌,临床治疗过程中应加强对目标病原菌的监测,积极做好干预措施,以提高其治疗效果与遏制其传播.     

628例食源性腹泻婴幼儿粪便与肛拭子标本中病原菌的培养及主要致病菌谱的分析及其防治对策

目的:分析食源性腹泻婴幼儿粪便与肛拭子标本中病原菌培养及主要致病菌构成情况与防治对策.方法:选取医院诊疗的食源性腹泻婴幼儿(2016年4月-2020年2月)628例临床资料,分析其粪便与肛拭子标本中病原菌培养结果及其主要致病菌构成情况,并提出干预措施.结果:628例患者标本中,经细菌培养结果显示阳性301例(检出病原菌301株),其阳性率为47.93％;食源性腹泻婴幼儿感染致病菌主要为沙门菌属占41.86％;对沙门菌属分布情况分析发现,其中肠炎沙门菌、鼠伤寒沙门菌、斯坦利沙门菌为主要致病菌.结论:食源性腹泻婴幼儿感染病原菌主要为沙门菌属、副溶血弧菌、致病性大肠埃希菌、志贺菌属;而肠炎沙门菌、鼠伤寒沙门菌、斯坦利沙门菌为沙门菌属中主要致病菌,临床治疗过程中应加强对目标病原菌的监测,积极做好干预措施,以提高其治疗效果与遏制其传播.     

PCR检测细菌16s rRNA基因在临床感染性疾病诊断中的应用

目的 建立检测临床常见细菌16 s rRNA基因的PCR方法.方法 分析细菌16 s rRNA基因保守区,设计通用引物对7种常见细菌16 s rRNA基因进行PCR扩增;并用该方法检测临床血液标本中的16 s rRNA基因表达,与传统培养方法结果进行比较.结果 7种标准菌株均出现特异阳性条带;感染性疾病血液标本16 s rRNA基因阳性表达.结论 16 s rRNA基因PCR法可用于快速检测临床细菌感染.     

Salmonella typhi infection in adults is not limited to travellers returning from the tropics

We reviewed the case notes of 23 adult patients infected with Salmonella typhi and admitted to the infectious disease unit, Auckland Hospital between January 1977 and December 1984. Fifteen had typhoid fever and eight were chronic carriers of S typhi. All isolates were sensitive to amoxycillin, chloramphenicol and cotrimoxazole. Ten of those with typhoid fever had recently been in tropical countries, predominantly Pacific Islands. The remaining five all lived in South Auckland and had not travelled out of New Zealand: we suspect that contaminated shellfish collected from the Manukau Harbour in South Auckland were the source. Typhoid fever should be suspected in young travellers returning to New Zealand with fever, diarrhoea, abdominal pain and headache. Similarly this diagnosis should be suspected in Polynesians and Maoris from South Auckland who have not travelled. All but one patient with typhoid fever responded clinically to the initial regimen which was usually oral amoxycillin given for a median 18 days. One other patient relapsed. Cholescystectomy and subsequent oral antibacterials eradicated S typhi from five biliary carriers with abnormal gallbladders. Prolonged high dose oral amoxycillin alone was effective in one of two carrier patients with normal gallbladders. The role of the Department of Health in identifying carriers of S typhi remains important.     

Clinical Importance of Salmonella Paratyphi A Infection to Enteric Fever in Nepal

Background : Enteric fever in Nepal is caused by infection with Salmonella typhi or Salmonella paratyphi A. The clinical presentation of these two illnesses has never been compared in a population of travelers and expatriates. If the illnesses are clinically comparable, and if S. paratyphi A infection is sufficiently common, the choice of typhoid vaccine for Nepal may have to take into account the vaccine's efficacy in preventing infection with S. paratyphi A.
Methods : NonNepalese patients presenting to the CIWEC Clinic with a history of 3 days of fever or greater were considered eligible for the study. Patients with positive blood or stool cultures for S. typhi or S. paratyphi A were entered into the study (along with three patients who had positive Widal titers only). A questionnaire was administered by a physician to determine signs and symptoms. Treatment with oral chloramphenicol was openly compared to treatment with oral ciprofloxacin.
Results : Forty-five cases of enteric fever were diagnosed during the 2 years of the study. Infection with S. typhi accounted for 20 cases, and S. paratyphi A was isolated in 22 cases. The illnesses were clinically indistinguishable. Treatment with chloramphenicol and ciprofloxacin was clinically comparable.
Conclusions : Infection with S. paratyphi A accounts for a significant percentage of enteric fever presentations among tourists in Nepal, and the illness is comparable to infection with S. typhi. Therefore, the choice of typhoid vaccine for long-term travelers or expatriates in Nepal should take into account the vaccine's potential ability to also prevent S. paratyphi A infection. The only typhoid vaccine that can currently offer this type of cross protection is the whole-cell killed preparation.     

Epidemiology of typhoid fever in Mauritius

BACKGROUND: The epidemiology of typhoid fever in Mauritius was studied to determine whether there was any need for tourists visiting Mauritius to be vaccinated against the disease, and where Mauritians with typhoid fever had been infected. Data on antibiotic susceptibility of Salmonella typhi isolates from Mauritius were also analyzed. METHODS: Since 1997 every time S. typhi is isolated from blood cultures at our laboratory, an epidemiologic inquiry is conducted to determine the likely origin of the infection and the outcome of treatment, and the information collected is recorded. Results of antibiotic susceptibility testing are also noted. Data recorded on cases between 1997 and 2004 were reviewed and analyzed. RESULTS: S. typhi was isolated on 25 occasions during the 8-year period. The infection was likely to have been acquired in Mauritius in only 6 cases (24%). Another 6 cases (24%) occurred in expatriate workers from the Indian subcontinent. Of the 13 Mauritians (52%) who probably acquired the infection abroad, 11 had a history of recent travel to India. Thirteen of 14 S. typhi isolates from cases acquired in India were resistant to nalidixic acid. Of the 6 cases acquired in Mauritius, 4 occurred in children under 12 years and 1 was caused by a multiply resistant strain. Twenty-two patients made an uneventful recovery. One death was indirectly caused by typhoid fever, and there was 1 case each of intestinal perforation and relapse. CONCLUSIONS: In Mauritius typhoid fever is mainly an imported disease, but indigenous cases of the illness occur rarely and sporadically. Travelers to Mauritius need not be vaccinated against typhoid fever as the risk of acquiring the disease in the country is negligible. Mauritians traveling to India must be made aware of the risk of typhoid fever and of preventive measures. Ceftriaxone should be used as the initial first-line treatment of infection acquired in India.     

伤寒的临床特点与耐药性研究

目的　探讨近年伤寒的临床特点和常用抗菌药物的耐药性及疗效。方法　收集经血培养和(或 )骨髓培养确诊的伤寒住院病人 97例 ,分析其临床特征、药敏试验和治疗效果。结果　本组伤寒的临床特征是热程长 (平均为 2 5 6d) ,病情重 ,并发症多。稽留热仍较多见 (5 5 7% ) ,脾肿大是主要体征。肥达反应阳性率为 6 7 1%。药敏试验呈多重耐药性 ,抗菌药物以喹诺酮类和三代头孢菌素疗效显著 ,其他药物单用效果普遍不佳。结论　多重耐药性的伤寒临床症状重 ,对常用治疗药物耐药性大 ,抗菌治疗应以氟啶酸或氟嗪酸等喹诺酮类为首选 ,联合用药效果更佳。     

常规PCR与套式PCR的敏感性比较研究

建立了套式引物聚合酶链式反应用于Ⅰ型艾滋病毒ｐｏｌ基因的检测，并比较了常规单对引物ＰＣＲ和套式引物ＰＣＲ的敏感性。结果表明，当模板ＤＮＡ分子的数量低于１０００个时，常规单对引物ＰＣＲ反应不能检出，而套式引物ＰＣＲ反应则可检出低至１０个分子的模板ＤＮＡ。提示采用套式引物可以大大提高聚合酶链式反应的敏感性。     

全血DNA滤纸片中HIV1前病毒基因的PCR检测

建立套式引物聚合酶链式反应检测全血ＤＮＡ滤纸片中的ＨＩＶ１前病毒基因ｐｏ１ＤＮＡ片段。分别采集１２例ＨＩＶ１感染者的外周血约５０μｌ，经与ＥＤＴＡ－蛋白酶ｋ孵育后滴在无菌滤纸片上，３７℃下干燥，将滤纸片密封于塑料袋中，在４℃及室温下保存１周～６０周后，分别将滤纸片置０．５ｍｌ试管中直接进行ＨＩＶ１ｐｏ１基因的外侧引物ＰＣＲ检测，然后进行内侧引物的ＰＣＲ检测。结果表明，全血ＤＮＡ滤纸片在室温下保存２８周，在４℃下保存３４周仍可检出ＨＩＶ１目的基因。根据ＰＣＲ产物的琼脂糖凝胶电泳溴化乙啶染色带形并参比实验设立的标准对照可直接判断结果。该方法具有快速、特异、敏感的特点，敏感性可以达到检出１０个靶ＤＮＡ分子，可作为抗体确证试验的补充方法用于ＨＩＶ１感染的诊断     

氧氟沙星与哌拉西林治疗伤寒疗效比较

目的：观察氧氟沙星与哌拉西林治疗伤寒疗效。方法：本院经血、骨髓、粪培养有伤寒杆菌生长的102例住院患者随机分成2组,分别给予氧氟沙星0．2g或哌拉西林4g,加入5％GS250ml ,iv gtt,q12h,疗程14d。观察2组总有效率、细菌清除率、体温恢复正常时间及不良反应。结果：2组总有效率细菌清除率均为100％,体温恢复正常时间2组差异无显著性,不良反应发生率分别为21．2％及16％。结论：氧氟沙星与哌拉西林均为治疗伤寒的有效药物,均有治愈率高、退热快的特点,但哌拉西林的不良反应明显低于氧氟沙星。     

Typhoid and Paratyphoid Fever: A 10-Year Retrospective Study of 41 Cases in a Parisian Hospital

BACKGROUND: Enteric fever remains a major cause of fever in travelers. We evaluated new trends in enteric fever. METHODS: We reviewed the epidemiological, clinical, biological, bacteriological data, and outcome of all cases of typhoid and paratyphoid fever seen in our department over the last decade. The inclusion criteria were the presence of signs compatible with enteric fever and isolation of Salmonella typhi or Salmonella paratyphi A, B, or C from blood or stool cultures or any other site. RESULTS: Among the 41 patients, 38 (93%) had travel-associated enteric fever. The main geographic source of contamination was the Indian subcontinent. One patient had been vaccinated with parenteral Vi vaccine 1 year previously. Fever and headaches were the only signs which were present in more than 80% of patients. The Widal test at inclusion was positive in 27%, and a second serological test was found to be positive in 50% of evaluated cases. Blood cultures and stool cultures were positive in 34 cases and 10 cases, respectively. Salmonellae spp were isolated in both hemocultures and stool cultures in 4 cases and in urine in 1 case. Two strains of S. typhi were resistant to ampicillin, chloramphenicol, and trimethoprim-sulfamethoxazole. One strain of S. typhi and one of S. paratyphi B were nalidixic acid resistant. All evaluable patients were cured with the exception of 2 patients (1 failure, 1 relapse). We observed 3 toxic reactions. No patients died. CONCLUSION: The diagnosis and outcome of enteric fever are hampered by the lack of specificity of clinical and biological signs, the increasing rates of antimicrobial resistance, and the occurrence of toxic reactions during treatment.     

鼻咽癌中EB病毒LMP1基因的序列变异发生机制探讨

目的检测贵州省鼻咽癌中EB病毒LMP1基因N-和C-末端区序列变异的热点,并探讨其产生的机制。方法收集贵州省贵阳医学院有明确病理诊断的鼻咽癌患者鼻咽新鲜活检标本30例。采用巢式聚合酶链反应（PCR）扩增EB病毒LMP1基因N-和C-末端区,用Xho I对N-末端区扩增产物进行酶切分析,从中选取2例患者N-末端区的扩增产物进行序列分析。同时检测C-末端区扩增产物30bp缺失的情况。结果30例鼻咽癌组织EB病毒LMP1基因C-末端区存在两种基因型：缺失型（即30bp缺失）和野生型,其中病例组有40％的突变率,对照组有10％的突变率,差异有显著性（P〈O．05）。在本次研究中的30例鼻咽癌及随机收集贵阳医学院体格检查健康成人30例的外周血（EDTA抗凝）2mL作对照组中EB病毒LMP1基因N-末端区都发生了突变,都存在原有酶切位点的缺失。结论鼻咽癌中EB病毒LMP1基因突变在鼻咽癌的发生发展中起重要的作用。