Distribution of Vibrio vulnificus along the coastal area of Shimane Prefecture and contamination of retail fish and shellfish with V. vulnificus in Shimane Prefecture, Japan

We studied distribution of Vibrio vulnificus in sea water, sediment, and shellfish along the coast and contamination of retail fish and shellfish with V. vulnificus in Shimane Prefecture, Japan between 2001 and 2004. V. vulnificus was isolated from 77 (68.1%) of 113 sea water samples with < or =10(5) MPN/L, 88 (70.4%) of 125 sediment samples with < or = 10(7) MPN/100g, 8 (17.2%) of 46 fish samples with < or = 10(6) CFU/10 g and 68 (43.8%) of 156 shellfish samples with < or = 10(6) MPN/10g. This organism was isolated from 64 (70.3%) of 91 shellfish samples, (short-necked clam, clam, hen clam, oyster and scallop, etc) with < or =10(6) MPN/10g and from 7 (43.7%) of 16 prawn samples with <10(2) MPN/10g. These isolates were divided into 11 serotypes with the most prevalent serotype being 04 followed by 01, 03 and 06. Serotype 012 strain (isolated from a patient from interior Shimane Prefecture in August 2004) was isolated from sea water and a sediment sample and from 3 samples of short-necked clam produced in Kumamoto Prefecture. These findings suggested potential transmission of V vulnificus through the water environment along the coast of the Japan Sea, that V. vulnificus is easily spread by coldtransport of fresh marine product, and that the occurrence of V. vulnificus infection is possible regardless of one's nearness to coastal areas where V. vulnificus is prevalent.     

Basic studies on Vibrio vulnificus infection: isolation of V. vulnificus from sea water,sea mud,and oysters

To clarify the environmental distribution of Vibrio vulnificus, sea water, sea mud, and oysters were examined at 13 sites, i.e. 4 sites in the Tokyo Bay (eastern Japan) and 9 sites (5 sites for oysters) in Tokushima Prefecture (western Japan). 1. V. vulnificus was isolated from 80 (54.8%) of the 146 samples of sea water examined. It was isolated from 19 (41.3%) of the 46 samples from western Japan and 61 (61.0%) of the 100 samples from eastern Japan. 2. It was isolated from 40 (40.8%) of the 98 samples of sea mud obtained in eastern Japan. 3. It was isolated from 655 (30.3%) of the 2,165 samples of oysters. They were 30 (9.7%) of 309 samples from western Japan and 625 (33.7%) of 1,856 samples from eastern Japan. 4. The density of V. vulnificus was 0.3-1.1 x 10(6) MPN/L in seawater, 0.3-1.1 x 10(5) MPN/100 g in sea mud, and 0.3-1.1 x 10(7) MPN/100 g in oysters. 5. Seasonally, V. vulnificus was isolated from 44 (6.2%) of the 713 samples in spring, 450 (72.6%) of the 620 samples in summer, 264 (51.8%) of the 510 samples in fall, and 17 (3.0%) of the 56 samples in winter. Thus, the isolation rates of V. vulnificus from sea water and oysters tended to be higher in eastern Japan than in western Japan and to be highest in summer, then, in fall.     

鳗鲡创伤弧菌的分子鉴定

目的对分离自发病欧洲鳗鲡的创伤弧菌疑似菌株进行准确的鉴定。方法首先尝试利用一对16SrRNA基因特异性通用引物PCR扩增一株鳗源创伤弧菌疑似株的基因组DNA，得到一个约500bp的DNA片段，将该DNA片段亚克隆至pMDl8-T载体，鉴定克隆化成功之后，送专业公司进行测序，得到一个502bp的DNA产物，NCBI上同源性比较表明，该片段与GeneBank上注册的创伤弧菌的16SrDNA序列同源性最高（100％），同时排除了哈维氏弧菌的可能。设计一对创伤弧菌溶血素特异性引物，实现了对鳗鲡创伤弧菌的分子鉴定。结果建立一种简洁的鳗鲡创伤弧菌的分子鉴定方法。结论国内首次自发病欧鳗分离到创伤弧菌，并给出分子鉴定证据。     

Identification of Vibrio vulnificus from patients with otitis media and septicemia by PCR method

Sucrose-negative colonies were isolated on TCBS agar plate from otorrhea specimens from otitis media patient in the Adult Diseases Examination Center, Hirosaki Medical Association. The isolate was identified as Vibrio vulnificus by nested PCR method which amplify V. vulnificus-specific sequences directed against 23S rRNA genes. The PCR method was applied to identify 6 strains originally isolated from septicemia patients in Kurashiki Central Hospital and formerly identified as V. vulnificus by phenotypic characteristics. When examined by the API20E system, the above PCR confirmed-V. vulnificus isolates were correctly identified as V. vulnificus with % ID 99.8, though this gave 3 different profiles. Cytotoxin-hemolysin gene was detected in otorrhea strain as well as septicemia strains by PCR method. The above results suggest that PCR method targeted cytotoxin-hemolysin gene is suitable for rapid and accurate identification of the isolate, because the result is obtained in less than 4 h. To our knowledge this is the first report on the V. vulnificus infection in Aomori Prefecture and the isolation from otorrhea.     

Vibrio vulnificus as a health hazard for shrimp consumers.

Over the last 30 years, a number of Vibrio species found in the aquatic environment have been indicated as cause of disease in human beings. Vibrio vulnificus is an emergent pathogen, an invasive and lethal marine bacterium related to wound infection and held accountable for gastroenteritis and primary septicemia. It occurs quite frequently in marine organisms, mainly in mollusks. This study aimed at isolating and identifying strains of V. vulnificus based upon the analysis of twenty samples of seabob shrimp, Xiphopenaeus kroyeri (Heller), purchased at the Mucuripe fish market (Fortaleza, Brazil). TCBS agar was used to isolate suspect strains. Seven of twenty-nine strains isolated from six different samples were confirmed as such by means of biochemical evidence and thus submitted to biological assays to determine their virulence. The susceptibility of the V. vulnificus strains to a number of antibiotics was tested. None of the V. vulnificus strains showed signs of virulence during a 24-hour observation period, possibly due to the shedding of the capsules by the cells. As to the results of the antimicrobial susceptibility tests, the seven above-mentioned V. vulnificus strains were found to be sensitive to nitrofurantoin (NT), ciprofloxacin (CIP), gentamicin (GN) and chloramphenicol (CO) and resistant to clindamycin (CI), penicillin (PN) and ampicillin (AP).     

Vibrio vulnificus infection: clinical and bacteriological analysis of four cases

Vibrio vulnificus (V. vulnificus) infection has recently been drawing attention as a high mortality disease especially in a patient with the preexisting chronic liver disease. The illness caused by V. vulnificus is divided into three groups such as primary septicemia, wound infection and gastrointestinal illness. Primary septicemia, which is the most common in Japan, is defined as a systemic illness presenting fever or hypotension with recovery of V. vulnificus from blood or tissue without the apparent primary focus of infection. We diagnosed four cases as infectious diseases of V. vulnificus by isolating it from each clinical material in Kurashiki Central Hospital from 1984 through 1997. We investigated clinical manifestations of the four cases including season of the onset, presence of drinking habit, underlying diseases, consumption of raw seafood, symptoms, presence of disseminated intravascular coagulation (DIC) or shock, laboratory data, administered antibiotics and the outcomes of the treatment. And for each strain, we also performed in vitro drug susceptibility tests. The age of the patients ranged from 49 to 61 years old (mean 56), and all of the patients were male. Each of them had a chronic liver disease as an underlying disease. Two of them had a history of raw seafood consumption prior to the onset of the illness. Skin manifestations appeared in two of the four patients. All the patients complicated septic shock and DIC. V. vulnificus was isolated from the venous blood cultures of them. Three of the four were given a diagnosis of primary septicemia and one was made a diagnosis of acute cholecystitis which has never been reported previously. Three of the four patients died and only the rest was alive as a result of antimicrobial therapy. In the sensitivity tests, the four strains were revealed to be very sensitive to the antimicrobials such as minocycline, cephalosporins of the third generation and carbapenems. Once patients with a chronic liver disease are infected with V. vulnificus, their prognosis is poor. Every effort should be made to advise not to have uncooked seafood. Physicians should be informed about the characteristics of the disease caused by this bacteria and treat any suspicious case promptly and appropriately.     

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.     

Prevalence of Listeria spp and Listeria monocytogenes in meat and fermented fish in Malaysia

Fermented fish and meat samples were purchased from supermarket and wet market for microbiological analysis of Listeria species and Listeria monocytogenes contamination. Listeria species were isolated from 17 (73.9%) of 23 samples of imported frozen beef, 10 (43.5%) of the 23 samples of local beef and 14 (56%) of the 25 samples of fermented fish from wet market. Listeria monocytogenes occurred in 15 (75%) of the frozen beef samples, 6 (30.4%) of the 23 samples of local meat and 3 (12%) of the 25 samples from fermented fish. Listeria species was not isolated from any of the 23 samples of imported frozen beef from supermarket and from the 5 samples of buffalo meat examined. This highlights the possibility of Listeria spp or L. monocytogenes to persist in meat and fermented fish in wet market and raises the problem of illness due to the handling and consumption of Listeria-contaminated meat or fermented fish are likely as evidence by the high contamination rates of samples sold at the wet market.     

A collective review of Vibrio vulnificus infection in Japan

Vibrio vulnificus is found globally in marine coastal waters. Infection with this organism, via ingestion of raw shellfish or exposure to marine water, can cause necrotizing fasciitis and sepsis, which have high mortality and short latency. In Japan, many cases have been reported since 1980, mainly from hospitals in western prefectures. However, because of the sporadic nature of infection outbreaks, a thorough epidemiologic survey has not been done. We studied the epidemiological and clinical characteristics of Vibrio vulnificus infections reported in Japan from 1975 to 2005. We identified 185 cases using the medical article search engines Ichushi (Japan Medical Abstracts Society), CiNii (Citation Information by National Institute of Informatics), and PubMed over 30 years. The median age of patients was 59. The number of male patients was eight times the number of female patients; however, no significant difference in mortality was found between genders. In yearly distributions of patients, 20 cases were reported in 2001, a year in which we experienced more rainfall and a longer rainy season in Northern Kyushu. In monthly distribution, about 80% of cases were reported from July to September when sea water temperatures rise. About 40% of cases were reported in four prefectures around the Ariake Sea. The underlying disease indicated liver dysfunction in 90% of patients, but mortality was the same regardless of the infection pathway (oral ingestion or wound). Because of its rapid aggravation and high mortality rate, public education is important to prevent new cases. It is also highly recommended that patients with preexisting liver dysfunction avoid raw fish and limit exposure to marine water during the summer.     

Vibrio vulnificus bacteremia associated with chronic lymphocytic leukemia, hypogammaglobulinemia, and hepatic cirrhosis: relation to host and exposure factors in 252 V. vulnificus infections reported in Louisiana

BACKGROUND: Vibrio vulnificus infection in persons with B-chronic lymphocytic leukemia (B-CLL) or hypogammaglobulinemia has been reported infrequently. PATIENT AND METHODS: A woman with B-CLL, hypogammaglobulinemia, and hepatic cirrhosis died of V. vulnificus bacteremia after eating cooked shrimp and crabs. We reviewed host and exposure data in 252 cases of V. vulnificus infection reported in Louisiana during the interval of 1980 through 2004. RESULTS: V. vulnificus was isolated from blood in 122 cases (48.8%). Preexisting conditions in 138 cases included liver disease (41.3%), malignancy (13.8%), and immunosuppression (9.4%). The prevalence of preexisting conditions was significantly greater in cases with positive blood cultures than in cases with positive wound or stool cultures. Exposure data in 116 cases revealed crab consumption without raw oyster consumption or seawater exposure in 3.4%. CONCLUSION: The present patient had several conditions associated with increased risk of V. vulnificus infection and bacteremia, especially hepatic cirrhosis, but her route of exposure to V. vulnificus was unusual.     

Achilles tendinitis in Vibrio vulnificus sepsis: a rare presentation

Vibrio vulnificus infection commonly presents as sepsis or wound infection. A case of alcoholic liver cirrhosis developed acute left heel pain and swelling 2 days after ingesting raw fish. Two sets of blood cultures indicated V. vulnificus. HLA-B27 was negative. Ultrasonography revealed left Achilles tendinitis. Fluid aspirated from the left retrocalcaneal bursa was sterile. The patient recovered completely 3 weeks after treatment. To our knowledge, V. vulnificus septicaemia with Achilles tendinitis, which may be the only indication of reactive arthritis (ReA), has not been reported previously. Prompt recognition of this life-threatening infection and adequate treatment with antibiotics is vital.     

Increasing drug resistance in Vibrio cholerae O1 and non-O1 strains isolated from diarrheal cases in Japan

Drug resistance trends were investigated for 271 Vibrio cholerae O1 (V.c O1) and 401 V. cholerae non-O1 (V.c non-O1) strains isolated from mainly imported diarrheal cases during 1981-2001 in Japan. The results of drug resistance test using 8 drugs (CP, TC, SM, KM, ABPC, ST, NA, and NFLX) showed that 34.7% of the V. c O1 strains and 15.7% of V.c non-O1 strains were multi-drug or mono-drug resistant. The incidence of drug resistant strains has increased since 1991, and it has been remarkable in V.c O1 strains that increased from 1.2% in 1981-1985 to 70.8% in 1996-2001. The drug resistance patterns of the resistant strains classified into 6 types in V.c O1 and 21 types in V.c non-O1. The prevalent patterns recognized were SM (75.5%), CP.TC.SM.ST (10.6%) and CP.SM.ST (8.5%) in V.c O1, and SM (25.4%) and ABPC (25.4%) in V.c non-O1. Ten V.c O1 strains (3.7%) and 10 V.c non-O1 strains (2.5%) were multi-drug resistant including TC. Among those, 13 strains were isolated from travelers who returned to Japan from Thailand. One V.c O1 strain (0.4%) and 6 V.c non-O1 strains (1.5%) were NA high-resistant and fluoroquinolones low-sensitive. Among those, 4 strains were isolated from travelers who returned to Japan from India.     

Molecular typing of vibrio vulnificus isolated from clinical specimens by pulsed-field gel electrophoresis and random amplified polymorphic DNA analysis

Primary V. vulnificus septicemia has been continuously reported in Korea. We analyzed the molecular diversity of V. vulnificus strains isolated from clinical specimens using pulsed-field gel electrophoresis (PFGE) and random amplified polymorphic DNA (RAPD) analysis. We analyzed a total of 23 V. vulnificus strains isolated from 22 patients between 1992 and 1997. RAPD analysis was performed using five random primers, and we obtained chromosomal DNA restriction patterns with NotI and SfiI by PFGE. Two isolates from one patient disclosed similarity value 1.0 by RAPD and had an indistinguishable pattern when analyzed with PFGE, which indicated that they were the same strains. The remaining 22 isolates from 22 patients could be separated into 5 different molecular types in RAPD analysis. The range of similarity values among the isolates was wide (0.29-0.81). Of 22 strains, four strains could not be typed in repeated trials by PFGE with NotI and SfiI. The PFGE patterns of 18 strains showed a remarkable polymorphism consisting 12-19 fragments (20-870 kb). These results show that V. vulnificus strains isolated from Korea are genetically diversified for the most part.     

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。     

创伤弧菌基因组学研究进展

创伤弧菌是人类三大致病弧菌之一,能够通过食源性传播或者伤口感染导致发病,近年来全球发病率逐步升高。随着新一代测序技术的发展,已有上百株创伤弧菌全基因组序列在国际公共数据库公布。这些数据为创伤弧菌基因组学研究提供了重要基础,并促进了对该病原菌遗传多样性、传播和致病机制的认识。本文从基因分型、种群结构和重要毒力因子3个方面对创伤弧菌基因组学研究进展进行归纳总结,以期为创伤弧菌感染、进化和防控研究提供借鉴。     

Characteristics of Listeria monocytogenes isolated from imported meat in Japan

The genomic structure of the iap region in Listeria monocytogenes (serovar 4b), isolated from chicken imported into Japan, was compared with those from Japanese strains. The isolate was similar to the Japanese strains in a comparatively new, rare group. Such strains might be imported from foreign countries.     

青岛市售养殖海水虾中副溶血性弧菌的分离及耐药性分析

目的 对青岛市售养殖海水虾中副溶血性弧菌进行分离鉴定,并对分离菌株进行耐药性分析。方法 以常规培养法分离,全自动细菌鉴定仪进行鉴定,PCR扩增法检测毒力基因,K-B法进行药敏试验。结果 样品的副溶血性弧菌检出率为78.1% (50/64),所有菌株均不含tdh基因,其中3株菌含trh基因,所有菌株对头孢拉啶耐受,96%菌株对氨苄西林和阿莫西林耐受,部分菌株对头孢呋新钠、链霉素、四环素、土霉素和复方新诺明耐受,少量菌株出现耐受3类抗生素以上的多重耐药性。结论 青岛市售养殖海水虾中副溶血性弧菌存在较严重的污染和一定程度的耐药情况。     

Species and serovar-distribution, and drug-resistance of Shigella strains isolated from imported and domestic cases during 1995-1999 in Tokyo

A total of 290 Shigella strains consisting of 180 imported strains and 110 domestic strains isolated during 1995-1999 in Tokyo were examined regarding their species and serovar-distribution and their drug-resistance. In both groups, S. sonnei (70.0% in the imported strains, 80.9% in the domestic strains) was found to be the most prevalent species, followed by S. flexneri (20.0% in the imported strains, 19.1% in the domestic strains). S. dysenteriae and S. boydii were only isolated in the imported cases. Among the S. flexneri serovar, 1b, 2a, 6, 2b, and 3a were predominant in the imported strains, whereas 1b and 2a were predominant in the domestic strains. Provisional new serovar Shigella strains were isolated from 11 imported cases and 2 domestic cases. The drug-resistance test using 9 drugs (CP, TC, SM, KM, ABPC, ST, NA, FOM, and NFLX) showed that 92.2% of the imported strains and 94.5% of the domestic strains were resistant to some of the drugs tested. Drug-resistance patterns of the resistant strains varied in 25 types. Among those, a triple drug-resistance type with TC.SM.ST was found as the most frequent pattern in both groups. None of the strains were resistant to NFLX.     

Epidemiological and microbiological analysis of community-associated methicillin-resistant Staphylococcus aureus strains isolated from a Japanese hospital

Reports of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections have recently increased in Japan; however, these studies contain limited information on their epidemiology. We performed a single-center study in the Tokyo Medical University Hospital located in Shinjuku, a central area of Tokyo, Japan. From 2,099 MRSA isolates obtained during July 2007 to March 2009, we selected 44 MRSA isolates with a MIC of <2 μg/mL for imipenem. Among 44 isolates, 28 strains had type IV or type V SCCmec, and we classified them as CA-MRSA. We identified only 1 Panton-Valentine leukocidin (PVL)-positive MRSA strain, which belonged to SCCmec type V. The PVL-positive CA-MRSA strain was isolated from a patient with multiple subcutaneous abscesses. The patient had returned to Japan from India; thus, the strain may have been contracted from outside of Japan. Thirteen (46.4%) and 15 strains (53.6%) were isolated from outpatients and inpatients, respectively. The major sites of infection included the respiratory tract (8 strains, 28.6%), skin/soft tissue (4 strains, 14.3%), and nasal cavity (4 strains, 14.3%). It is important to note that the most common site of CA-MRSA infection in inpatients was the respiratory tract; respiratory infections with CA-MRSA frequently cause severe infectious diseases.