创伤弧菌基于基因组学的毒力因子及耐药基因的研究进展

创伤弧菌是一种致命的机会致病菌,在目前食源性疾病中致死率最高。创伤弧菌主要通过摄入未煮熟的海产品以及破损皮肤接触海水引起感染,病情恶化后导致危及生命的败血症。基因组学测序技术的发展迅速,在美国国家生物技术信息中心（NCBI）数据库中已公布的创伤弧菌基因组草图序列285株,完成图序列24株。创伤弧菌基因组学的研究,不仅有利于精确分型、进化关系和致病机制的研究,还利于对毒力因子、耐药基因进行预测,在创伤弧菌引起的食物中毒后进行快速检测、流行病学调查和预防发挥重要作用。目前针对创伤弧菌耐药性分析的研究较少,本文从创伤弧菌的基因组学角度出发,对重要毒力因子、抗生素耐药情况、耐药机制及耐药基因予以概述。     

海洋致病菌致病机制及其感染的治疗研究进展

海洋致病菌多为广泛分布于海洋的革兰氏阴性菌,是造成开放性伤口继发性感染、食用海产品发生食物中毒的主要致病菌,如治疗不及时病死率较高。海洋致病菌主要有弧菌属的创伤弧菌、副溶血性弧菌、霍乱弧菌及铜绿假单胞菌等。创伤弧菌分布于（亚）热带气候水域,可通过生物分型法、传统培养法和PCR法分型鉴定;致病机制研究集中在黏附相关蛋白、细胞毒性、宿主防御、摄铁系统、分泌系统和毒力调控等方面;全身性感染或开放伤口感染,通常用强力霉素和第三代头孢菌素进行抗生素治疗。副溶血性弧菌在亚洲、欧洲、美洲分布广泛,通过PCR法进行分型鉴定;致病机制与创伤弧菌相似;可引起急性肠胃炎及其他组织病变,轻症感染补充水分即可,重症感染使用强力霉素和第三代头孢菌素治疗。霍乱弧菌分布广泛,夏季浓度较高,通过玻片凝集试验、PCR法进行分型鉴定;致病机制与创伤弧菌相似;可引发霍乱,通过补液或抗生素治疗。铜绿假单胞菌分布于海水表层,通过玻片凝集实验、抗菌谱等进行分型鉴定;致病机制与弧菌属不同;可感染身体各处,早期治疗应用抗生素,重症患者应选择广谱的强力抗菌药。     

黄姑鱼创伤弧菌的分离和鉴定

目的从患病黄姑鱼体内分离出致病力较强的菌株L1,人工感染实验证实该菌株为黄姑鱼的病原菌。对该细菌进行了形态、生理生化特性测定和16SrRNA分子鉴定,测定16SrRNA基因序列,分析相关细菌相应序列的同源性,构建了系统进化树。结果表明菌株L1与创伤弧菌(Vibrio vulnificus)的亲缘关系最近,具有98.7%的同源性。结合该菌株的生理生化特性,可鉴定菌株L1为创伤弧菌。     

日本鳗鲡混合感染迟缓爱德华氏菌与创伤弧菌的分离与鉴定

目的对日本鳗鲡混合感染迟缓爱德华氏菌(Edwardsiella tarda)和创伤弧菌(Vibrio vulnificus)的病原进行了分类鉴定。方法对两株病原菌进行了形态、API-ID32E鉴定系统和分子生物学鉴定,分别测定了菌株AnGH080301和An-GH080302的16SrRNA和HSP60基因序列,构建了系统进化树。结果按形态特征和API-ID32E鉴定系统分别初步鉴定菌株AnGH080301和AnGH080302为迟缓爱德华氏菌和创伤弧菌。菌株AnGH080301的16SrRNA基因部分序列(登录号FJ646618)与迟缓爱德华氏菌的16S rRNA基因(登录号AB050832)同源性最高,达99.7%;菌株AnGH080302的HSP60基因部分序列(登录号FJ646619)与创伤弧菌HSP60基因(登录号BA000037)的同源性最高,达99.8%。结论综合菌株的生理生化特性和分子生物学鉴定结果,可将菌株AnGH080301和AnGH080302分别鉴定为迟缓爱德华氏菌和创伤弧菌,迟缓爱德华氏菌与创伤弧菌均为人兽共患病病原,其混合感染日本鳗鲡致病的报道尚属首次。     

德尔卑沙门菌的研究进展

德尔卑沙门菌是重要的人兽共患病原菌,可引起人食物中毒和败血症等症状,而且对牲畜的繁殖及健康带来严重威胁。本文就德尔卑沙门菌的流行现状、基因组学及致病机制等方面的研究进行总结,以期为该菌的防治提供指导。     

交叉引物恒温扩增技术检测创伤弧菌的应用研究

目的 建立一种快速检测创伤弧菌(Vibrio vulnificus)的交叉引物恒温扩增(cross priming amplification,CPA)技术。方法 针对创伤弧菌vvhA基因序列保守区域设计CPA引物和探针,并优化反应体系和反应条件,同时验证方法的灵敏度和特异性。结果 对创伤弧菌的检测具有高度特异性;对创伤弧菌菌液检测的灵敏度达到了1.28×10³ CFU /mL,此法 40～60 min 内即可完成检测。利用CPA法针对本实验室前期研究的CB型、EA型、CAB型、CA型和EB型等亚型创伤弧菌毒株计算特异性和敏感性,5种亚型中除了CB亚型特异性和敏感性为95.65%和100%,其余亚型特异性和敏感性均为100%。结论 本研究建立的CPA法具有较高的特异性和灵敏性,可有效缩短检验时间提高检验效率。     

济宁市淡水产品养殖链条中致病性弧菌分布调查

目的调查济宁市淡水产品养殖链条中致病性弧菌分布情况。方法 2016-2018年采集济宁市淡水产品及养殖链条环境样品488份,依据GB4789.7-2013《食品安全国家标准食品微生物学检验副溶血性弧菌检验》、《国家食品微生物风险监测工作手册》和《淡水动物性水产品养殖环节中常见弧菌专项监测工作手册》等进行霍乱弧菌、副溶血性弧菌、创伤弧菌、溶藻弧菌和河流弧菌的检测。结果济宁市淡水产品及养殖链条环境产品霍乱弧菌、副溶血性弧菌和创伤弧菌总检出率分别为23.98%(117/488)、14.96%(73/488)和0.20%(1/488),未检出溶藻弧菌和河流弧菌;养殖链条中的鱼类、螃蟹类、环境(饲料、水藻、鱼苗、粪便、手)、水体、水底沉积物弧菌检出率分别为52.08%(75/144)、13.64%(6/44)、29.17%(7/24)、43.29%(71/164)和28.57%(32/112)。霍乱弧菌在鱼类样品中的检出率较高,为37.50%(54/144),117株霍乱弧菌血清型鉴定均为非O1/非O139型霍乱弧菌,均未检出ctxAB毒力基因。副溶血性弧菌在水体中的检出率较高,为21.95%(36/164),对73份样品分离的副溶血性弧菌进行tlh基因、tdh和trh毒力基因检测,tlh+tdh-trh+和tlh+tdh-trh-携带率较高,分别为46.58(34/73)、42.47%(31/73)。结论济宁市淡水产品养殖链条中致病性弧菌主要分布在水体及鱼类中,且均为非O1/非O139型霍乱弧菌,致病风险较低;副溶血性弧菌trh毒力基因携带率较高,有潜在的致病风险。     

溶藻弧菌急性胃肠炎3例报告

溶藻弧菌（Vibrio alginolyticus）是一种致病性弧菌,广泛存在于自然界,特别是在海水和海水养殖动物中常见。大量研究资料表明,它是鱼、虾、贝类等多种海水养殖动物致病和死亡的常见致病菌。但是,溶藻弧菌对人类的致病性尚未肯定。过去认为,溶藻弧菌对人类不致病,或仅能引起局部伤口感染。直至80年代以后,溶藻弧菌所致的人眼结膜炎、中耳炎、败血症等病例相继在国外报告。     

布鲁氏菌基因组学研究进展北大核心CSCD

布鲁氏菌是一类革兰氏阴性、胞内寄生菌,缺乏经典的毒力因子,致病性不仅依靠侵袭力和内毒素,还有赖于较强的环境适应能力。目前,布鲁氏菌入侵机体和在胞内持续存在的机制尚未明确。布鲁氏菌基因组学研究不仅可全面了解布鲁氏菌的基因组成、分子进化、毒力因子以及致病机制等特点,还可以对一些致病相关基因和重要的蛋白进行预测。该研究为开发研究疫苗和新型抗生素提供分子基础,从而为构建新的有效的布病治疗和防控策略提供理论依据。本文就布鲁氏菌的基因组组成、特征,全基因组测序分析、比较基因组研究进展予以概述。     

布鲁氏菌基因组学研究进展

布鲁氏菌是一类革兰氏阴性、胞内寄生菌,缺乏经典的毒力因子,致病性不仅依靠侵袭力和内毒素,还有赖于较强的环境适应能力。目前,布鲁氏菌入侵机体和在胞内持续存在的机制尚未明确。布鲁氏菌基因组学研究不仅可全面了解布鲁氏菌的基因组成、分子进化、毒力因子以及致病机制等特点,还可以对一些致病相关基因和重要的蛋白进行预测。该研究为开发研究疫苗和新型抗生素提供分子基础,从而为构建新的有效的布病治疗和防控策略提供理论依据。本文就布鲁氏菌的基因组组成、特征,全基因组测序分析、比较基因组研究进展予以概述。     

A case of septicemia type Vibrio vulnificus infection with necrotizing fasciitis rescued by lower extremity amputation]

We report a case of septicemia type Vibrio vulnificus infection. The patient was a 74-year-old man who had liver cirrhosis and hepatocelluler carcinoma. He felt a pain in the right femoral lesion after eating raw shellfish (Japanese "Umitake") two days ago. He was admitted to our emergency center due to his shock status and thrombocytopenia two days after the onset. We diagnosed necrotizing fasciitis due to Vibrio vulnificus infection, his life was saved by emergency amputation of the right lower extremity. The culture of the blood and vesicle fluid showed Vibrio vulnificus. There are some reports that the debridement was effective to necrotizing fasciitis due to Vibrio vulnificus infection, but these reports are all about single upper extremity lesion. As far as we know, this is the second report of lower extremity necrotiaong fasciitis due to septicemia type Vibrio vulnificus infection rescued by extremity amputation in Japan. The mortality of septicemia type Vibrio vulnificus infection with necrotizing fasciitis is very high, this is quite a valuable report in making a decision for therapy of septicemia type Vibrio vulnificus infection.     

Rapid detection of Vibrio vulnificus using a loop-mediated isothermal amplification method

Vibrio vulnificus is found in marine waters near the coast around the world. Infection with this gram-negative rod, via ingestion of raw seafood or via a skin wound following contact with contaminated estuarine or marine water, can cause necrotizing fasciitis and sepsis. Most of patients with Vibrio vulnificus infection have underlying liver dysfunction or diabetes mellitus. Due to the high mortality and short latent periods, control of this infection depends on early identification of the bacterial species and prompt initiation of intensive care. Accordingly, the development of a technique that can identify this microbe quickly and accurately is of great importance. Loop-mediated isothermal amplification (LAMP) is a novel nucleic acid amplification method to detect specific genes with rapidity and high sensitivity. In this study, we developed LAMP for the detection of Vibrio vulnificus. Using 28 Vibrio vulnificus strains and 53 other bacterial strains, we confirmed the high specificity of this method. Moreover, our LAMP method also showed high sensitivity, with a minimum detection level of one colony-forming unit per test. Furthermore, we developed simplified and conventional pretreatments for the method using experimental animal models. All of these attempts have lod to our non being able to detect Vibrio vulnificus within 1 hour.     

Vibrio vulnificus septicemia in Spain

Vibrio vulnificus is a virulent marine organism, able to contaminate sea-food. It usually produces bacteremia associated with secondary skin lesions in patients with underlying conditions, such as hepatic cirrhosis. We report a case of septic shock and characteristic skin lesions, due to Vibrio vulnificus in a patient with cirrhosis, who had eaten raw oysters. The patient survived in spite of the severity of the clinical picture. We conclude that Vibrio vulnificus infection must be considered in the differential diagnosis of sepsis and skin lesions.     

Vibrio vulnificus rtxA1 gene recombination generates toxin variants with altered potency during intestinal infection

Vibrio vulnificus is a food-borne bacterial pathogen associated with 1% of all food-related deaths, predominantly because of consumption of contaminated seafood. The ability of V. vulnificus to cause disease is linked to the production of a large cytotoxin called the "multifunctional-autoprocessing RTX" (MARTX(Vv)) toxin, a factor shown here to be an important virulence factor by the intragastric route of infection in mice. In this study, we examined genetic variation of the rtxA1 gene that encodes MARTX(Vv) in 40 V. vulnificus Biotype 1 strains and found four distinct variants of rtxA1 that encode toxins with different arrangements of effector domains. We provide evidence that these variants arose by recombination either with rtxA genes carried on plasmids or with the rtxA gene of Vibrio anguillarum. Contrary to expected results, the most common rtxA1 gene variant in clinical-type V. vulnificus encodes a toxin with reduced potency and is distinct from the toxin produced by strains isolated from market oysters. These results indicate that an important virulence factor of V. vulnificus is undergoing significant genetic rearrangement and may be subject to selection for reduced virulence in the environment. This finding would imply further that in the future on-going genetic variation of the MARTX(Vv) toxins could result in the emergence of novel strains with altered virulence in humans.     

Vibrio vulnificus pollution of imported frozen Black Tiger shrimps in Japan

The Ariake Sea area of Japan is endemic for Vibrio vulnificus infection. V vulnificus was isolated from slime from tidal flats, seawater, and fish Sea year-round as we reported previously. To identify new routes and factors of V vulnificus infection, we studied V. vulnificus pollution of imported frozen Black Tiger shrimps purchased from a fish market in Kurume, Fukuoka, Japan. V. vulnificus was isolated from 9 of 100 tails (9%) of Philippines products, 3 of 100 tails (3%) of Indonesia products, and 0 out of 100 tails (0%) of Madagascar products. Cytotoxin-hemolysin genes were identified in 7 V. vulnificus strains isolated from patients with V vulnificus septicemia, 9 strains from Philippine products, and 3 strains from Indonesian products. These results suggest that imported frozen Black Tiger shrimps are a new sources of V. vulnificus infection.     

Two cases of sucrose-fermenting Vibrio vulnificus infection in which 16S rDNA sequencing was useful for diagnosis

Vibrio vulnificus is a Gram-negative bacterium which is associated with severe infections in humans. We experienced two cases of sucrose-fermenting V. vulnificus infection. The causative agents in both cases were unidentifiable by conventional identification systems because of their unique characteristics, and sequencing of 16S rDNA was found to be useful for diagnosis.     

A fatal case of Vibrio vulnificus presenting as septic arthritis.

Vibrio vulnificus is an invasive gram-negative bacillus that may cause necrotizing cellulitis, bacteremia, and/or sepsis. Although V vulnificus infection is uncommon, it is frequently fatal and is usually attributed to ingestion of raw shellfish or traumatic exposure to a marine environment; patients are also often found to have a hepatic disorder (cirrhosis, alcohol abuse, or hemochromatosis) or an immunocompromised health status, and most commonly present with septicemia or a wound infection. We describe a patient who presented with septic arthritis as the first clinical manifestation of a V vulnificus infection. The organism was subsequently identified in a synovial fluid aspirate.     

A life saving case of Vibrio vulnificus (septic type)]

After eating raw shellfish, 62-year-old man suffered from watery diarrhea and leg pain. The disease developed to septicemia and necrotizing fasciitis of bilateral lower extremities. Vibrio vulnificus was isolated from wound. He was treated with antibiotics, surgical debridement and hyperbaric oxygen therapy, and consequently was saved. Effect of hyperbaric oxygen therapy on V. vulnificus infection was discussed.     

Clinical features and an epidemiological study of Vibrio vulnificus infections

Vibrio vulnificus, a recently described halophilic Vibrio species, has been isolated from the blood, wounds, and other skin lesions of patients with primary sepsis or wound infections. Because no study of risk factors for infections with V vulnificus has been reported, a case-control study was performed with the 30 patients from whom V vulnificus isolates were recently submitted to the Centers for Disease Control (Atlanta, Georgia). Patients with primary sepsis were more likely than controls to have eaten raw oysters recently (P less than .01) and to have a history of liver disease (P less than .02). Persons with liver disease should be warned that raw oysters are an important source of this life-threatening infection. Patients with wound infections were more likely than controls to have had recent exposure of the skin to salt water or shellfish (P less than .05). Physicians should therefore consider V vulnificus in the differential diagnosis of severe wound infections with these exposures.