疾病监测  2015, Vol. 30 Issue (4): 337-341

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文章信息

郑文龙, 王卓, 马洁, 赵帅, 江国虹
ZHENG Wen-long, WANG Zhuo, MA Jie, ZHAO Shuai, JIANG Guo-hong
副溶血性弧菌病原学和分子流行病学流行特征研究进展
Progress in research of etiological and molecular epidemiological characteristics of Vibrio parahaemolyticus
疾病监测, 2015, 30(4): 337-341
Disease Surveillance, 2015, 30(4): 337-341
10.3784/j.issn.1003-9961.2015.04.022

文章历史

收稿日期:2014-11-18
副溶血性弧菌病原学和分子流行病学流行特征研究进展
郑文龙, 王卓, 马洁, 赵帅, 江国虹     
天津市疾病预防控制中心, 天津 河东 300011
摘要:副溶血性弧菌是导致我国细菌性食物中毒事件的首要原因.临床分离株和环境分离株的病原学特征有着明显的差异.临床分离株以O3:K6血清型为主,tdh和(或)trh基因携带率较高,一般在80%以上,脉冲场凝胶电泳(PFGE)基因图谱具有明显的优势图谱,并且与血清分型具有一致性.而环境分离株(包括食品)多无优势血清型和优势PFGE图谱,tdh和(或)trh基因携带率远低于临床分离株,多在6%以下.各地临床分离株的耐药性差异较大,但对氨苄西林等早期药物耐药率均较高.环境分离株较临床分离株的耐药性更为严重和复杂.
关键词副溶血性弧菌    血清分型    毒力基因    脉冲场凝胶电泳分型    耐药性    
Progress in research of etiological and molecular epidemiological characteristics of Vibrio parahaemolyticus
ZHENG Wen-long, WANG Zhuo, MA Jie, ZHAO Shuai, JIANG Guo-hong     
Tianjin Municipal Center for Disease Control and Prevention, Tianjin 300011, China
Abstract:Vibrio Parahaemolyticus is the major pathogen causing food-borne disease in China. This paper summarizes the progress in research of etiological and molecular epidemiological characteristics of V. parahaemolyticus in recent 5 years. The research indicated that there was obvious difference in the etiological characteristics between clinical isolates and environmental isolates. The clinical isolates mainly belonged to serotype O3:K6, and 80% of the clinical isolates carried tdh gene and(or) trh gene. The predominant PFGE pattern of the clinical isolate was obvious, consistent with the serotype. But the environmental isolates had no predominant serotype and PFGE pattern, their carriage rate of tdh gene or trh gene(<6%) was lower than that of clinical isolates. The drug resistance of the clinical isolates varied with area, and the resistance to ampicillin was high. While the drug resistance of environmental isolates was more serious and complex.
Key words: Vibrio parahaemolyticus    Serotype    Virulence gene    PFGE typing    Drug resistance    

副溶血性弧菌(Vibrio parahaemolyticus,VP)是一种革兰阴性嗜盐杆菌,是引发食源性疾病的主要致病菌之一。1988年以来的报道显示,VP引发的食物中毒的发生规模及人群暴露规模呈明显上升趋势,均已超过沙门菌,跃居首位[1, 2]。我国卫生部疫情直报系统(http://www.moh.gov.cn/publicfiles//business /htmlfiles/mohwsyjbgs/index.htm)监测显示,在 2009-2011 年期间VP是引起细菌性食物中毒事件的首要病因。掌握VP病原学特征、耐药性以及基因同源等流行特征是预防控制VP性食源性疾病的重要基础,本研究综述了近年来关于VP病原特征流行状况的研究工作,并对其流行特征进行讨论。

1 VP血清分型流行特点

血清分型鉴定的依据是不同的菌株其表面抗原不同,这些表面抗原可以被抗体或抗血清所检测。VP的鞭毛H抗原特异性低,所有的菌株均相同,故仅以菌体O抗原和荚膜K抗原进行血清分型。现报道有13个热稳定O抗原群(O1~O13),71种热不稳定K抗原(K1~K71)。自1996年,世界流行的VP血清型发生了较大变化,O3 ∶ K6型取代O4 ∶ K8型成为主要流行血清型。随着基因检测技术在病原学分析中的应用,O3 ∶ K6的克隆衍生株被越来越多研究发现。这些衍生株在血清分型中虽然与O3 ∶ K6不同,但却具有与O3 ∶ K6相同的基因标识ORF8,并且其分子形态与O3 ∶ K6几乎一致,主要包括有O4 ∶ K68、O1 ∶ K25和 O1 ∶ KUT(未分型),通常也被称作O3 ∶ K6系列[3]。Yamazaki 等[4]对日本1988 2001年由VP引起的食源性暴发事件和散在病例进行研究显示:1988 1995年,由O3 ∶ K6型引起的暴发事件为3%,而1996-2001年上升为75%。此后有很多关于泰国、印度等亚洲国家、美国、欧洲、南美和非洲的报道显示O3 ∶ K6型成为世界范围内流行的优势血清型[5, 6, 7]。但是,也有部分地区如美洲的部分国家从未发生过O3 ∶ K6的流行[3]。García等[8]关于智利南部VP流行的研究显示O3 ∶ K6的流行程度影响着人群VP的流行程度,当O3 ∶ K6流行时,临床患者明显增加,反之临床患者人数明显减少。

我国近几年VP的病原学研究显示,O3 ∶ K6仍是主要流行血清型。李薇薇等[9]报道了2007-2009年中国VP临床分离株分子特征分析的研究成果,对来自浙江、江苏、四川、广西和辽宁5省(自治区)的临床分离株分析发现,135株临床株检出29种血清型,以O3、O4、O1群为主,占89.6%(121/135),O3 ∶ K6为优势血清型,占56.3%(76/135)。陈洪友等[10]对上海市2010-2012年分离自患者和食品中共1136株VP进行血清学和分子特征进行研究,结果显示76.8%的为O3 ∶ K6,其次为O4 ∶ K68(9.4%)、O1 ∶ K25(6.8%)、O1 ∶ K36(4.5%)。曲梅等[11]研究了来自北京市2118例腹泻散发患者分离出的114株VP,O3 ∶ K6为优势血清型,占63.16%。

有研究提示临床分离株和环境分离株在血清分型上存在一定差别,环境分离株O3 ∶ K6占比明显低于临床分离株[12, 13]

2 毒力基因携带特征

VP的主要致病作用是其可产生溶血毒素。主要的溶血毒素有耐热性溶血毒素(TDH)和耐热性溶血毒素相关的溶血毒素(TRH),此外VP还可产生不耐热溶血毒素(TLH)。TDH和TRH是VP致病的主要因素,关于美国、欧洲和亚洲的研究显示接近或超过90%的VP临床分离株携带tdh和(或)trh基因[14, 15, 16],而环境分离株仅有0~6%携带上述毒素[14, 17, 18, 19]。李薇薇等[9]报道了2007-2009年来自我国5个省份的临床分离株,85.9%的菌株tdh和(或)trh阳性,tdh、trh的携带率分别为85.2%和3.0%,远高于环境分离株的2.1%[20]。曲梅等[11]报道了北京临床分离株的结果,tdh存在于大部分菌株中,所占比例为93.86%;O3 ∶ K6型菌株tdh基因阳性率(98.61%)明显高于非O3 ∶ K6型菌株(85.71%)。

但是,Wootipoom等[21]报道了泰国南部Hat Yai医院2000 2005年VP分离株的特征,2000-2003年tdh(+) trh(-)菌株占比较为稳定,分别为64.1%、67.5%、69.7%和 67.7%,2004年和2005年出现下降,分别为56.1%和55.5%。FAO/WHO[22] 2011年关于海产品VP风险评估报告显示,不携带tdh和(或) trh基因的临床分离株比例在增加,并且多与重症病例相关。另外,美国[23]、法国[24]和墨西哥[15]也有部分地区环境和食品分离株tdh和(或)trh基因的阳性率甚至达到52%。Flores-Primo 等[25]关于墨西哥产生蚝中分离VP的研究显示冬季tdh的携带率明显高于夏季。说明VP临床分离株和环境分离株(食品)的毒力基因也在随着时间和环境的改变发生改变。

3 耐药性

对VP进行耐药监测有利于及时掌握VP的耐药变化趋势,以便指导临床用药。随着我国水产养殖业的兴起,在养殖过程中使用抗生素也会影响VP的耐药情况。因此,对VP进行耐药监测也可用于对水产养殖业管理政策的制定和调整。美国临床实验室标准化协会(CLSI)就耐药的实验方法、抗生素的选择原则及结果判定标准制定了系列规定[26],为多数研究所采纳。各地分离的VP菌株耐药性出现多样化的特点。北京地区113株临床分离株12种药的实验结果显示,对氨苄西林的耐药率高达96.26%,对庆大霉素为33.64%,对其他抗生素的耐药率均低于4%[11]。江苏临床分离株的研究结果则显示95株VP对四环素耐药率为2.1%,复方磺胺甲恶唑耐药率为3.2%[27]。上海地区318株临床分离株对氨苄西林的耐药率在95%以上,对氯霉素、头孢噻肟、四环素等也存在耐药现象,对头孢他啶、头孢吡肟、庆大霉素等8种抗生素则完全敏感[28]。李薇薇等[9]的研究显示:四川、江苏、浙江、广西4省(自治区)均检出耐药株。8.1%(11/135)的VP临床分离株耐药,其中9株耐氨苄西林,1株耐复方磺胺甲恶唑,1株同时耐复方磺胺甲恶唑和四环素。

环境分离株与临床分离株耐药性差异也比较大。青岛地区养殖虾中分离的50株VP对18种抗生素的耐药性结果显示所有菌株对头孢拉啶100%耐受,对氨苄西林和阿莫西林高度耐药,耐药率均为96%。所有菌株对头孢曲松、头孢吡肟、庆大霉素、 诺氟沙星、奈啶酸100%敏感,部分菌株对头孢呋新钠、链霉素、四环素、土霉素、复方新诺明耐受,少量菌株出现耐受3类抗生素以上的多重耐药性,分离到的3株trh基因阳性菌株同时耐受氨苄西林、阿莫西林和头孢拉啶[29]。Jiang 等[30]报道了养殖海参中VP的耐药性,87株菌株对氨苄西林和头孢唑啉100%耐药,对链霉素、头孢呋辛钠、四环素、磺胺甲恶唑和喹诺酮的耐药率分别为43.7%、18.4%、4.6%、 2.3%和2.3%。56.2%的菌株至少耐受3种以上抗生素。陈永红等[31]的研究结果显示,南京地区海产品中分离的60株VP对30种药物的耐药率由高至低依次为洁霉素(90.3%)、红霉素(72.3%)、氯霉素(66.7%)、多黏菌素(51.7%)、青霉素(33%)、强力霉素(28.3%)、四环素(26.7%)、卡那霉素(25%)、氨苄西林(23.3%)、新霉素(21.7%)、氟哌酸(16.7%)、复方新诺明(15%)、痢特灵(13.3%)、庆大霉素(10%)、痢特灵(5%)、先锋霉素Ⅳ(5%),而头孢曲松、头孢呋肟、万古霉素、美满霉素、奥复星、麦迪霉素、先锋霉素V、丁胺卡那霉素、诺氟沙星、丙氟哌酸、复达欣均不耐药。国外一些关于VP环境分离株的报道显示耐药其耐药性也较为严重。Kitiyodom等[32]从养殖斑节对虾中分离到25株VP其中17株耐氨苄西林,11株耐红霉素,6株耐磺胺甲基异恶唑,4株耐4环素,3株耐链霉素和甲氧苄氨嘧啶。Sahilah等[33]从马来西亚4个地区收集了37株VP,对青霉素、氨苄西林、头孢呋辛、阿米卡星和头孢他啶的耐药率分别为89.0%、68.0%、38.0%、6.0%和14.0%。Oh等[34]研究了2005 2007年养殖鱼中VP分离株的耐药性,65.1%的菌株至少耐受1种抗生素,对氨苄西林、利福平、链霉素、甲氧苄氨嘧啶的耐药率分别为57.8%、11.9%、8.7%和6.4%。

对VP临床分离株和环境分离株的研究发现,各地分离株的耐药性差异较大,但对氨苄西林等早期药物耐药率均较高。环境分离株较临床分离株的耐药性更为严重和复杂。

4 脉冲场凝胶电泳分子分型特征

脉冲场凝胶电泳(pulsed-field gelelectrophoresis,PFGE) 具有分型分辨力强、结果稳定性重复性好、易于解释等特点,被推崇为同源性分析的“金标准”[35]。1996年,美国疾病预防控制中心以该技术为核心建立了食源性疾病分子分型监测网络实验室体系(PulseNet),随后加拿大、欧盟、亚太等国家和地区先后加入了该网络,2003年起中国也加入了这个全球性的体系。多数研究显示,食物中毒暴发株和临床分离株往往具有优势带型,说明其遗传基因较相近;而环境和食品分离株多无优势带型,呈现遗传多样性。例如,翁琴云等[36]对厦门市2005-2011年25起食物中毒事件的108株VP研究显示,具有明显的优势带型,占63.9%,对应的血清型分别为O3 ∶ K6、O4 ∶ K68 和O1 ∶ K25。深圳、江苏、沈阳、浙江等[37, 38, 39, 40]均显示出相同的规律,即食物中毒暴发株和临床分离株均具有优势带型。章红红等[41]报道了上海地区505株VP的PFGE研究结果,食物中毒暴发株和临床散发病例分离株具有相同的优势带型,食品分离株无优势带型,且与上两者差别较大。李薇薇等[9]报道VP临床株PFGE分型与血清型密切相关,O3 ∶ K6及其“亲属”血型具有较相近的基因图谱。

5 结语

VP是引起细菌性食源性疾病的重要致病菌。临床分离株和环境分离株的病原学特征有着明显的差异。临床分离株以O3 ∶ K6血清型为主,tdh和(或)trh基因携带率较高,一般在80%以上,PFGE基因图谱具有明显的优势图谱,并且与血清分型具有一致性。而环境分离株(包括食品)多无优势血清型和优势PFGE图谱,tdh和(或)trh基因携带率远低于临床分离株,多在2%左右。各地分离株的耐药性差异较大,但对氨苄西林等早期药物耐药率均较高。环境分离株较临床分离株的耐药性更为严重和复杂。

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