Identification of Vibrio vulnificus from Patients with Otitis Media and Septicemia by PCR Method

Background: Due to frequent exposure to surface water and contact with animals, children represent a group susceptible to zoonotic diseases. Objectives: The present study aims to determine the presence and prevalence of the main zoonotic agents in R. norvegicus populations in Tehran, Iran. Methods: In the present study, 100 R. norvegicus were captured within a time span of one year from five districts of Tehran, Iran. Fecal and blood samples were collected from rodents and serum was recovered after centrifugation. The presence of specific IgG antibodies against Leptospira spp. and Rabies virus was detected using a commercial qualitative rat ELISA kit. A conventional PCR assay was employed to detect the presence of Vibrio vulnificus in the commensal R. norvegicus population. Results: In general, 80% (n = 80/100) and 20% (n = 20/100) of rats were males and females, respectively. The results of the ELSA assay showed that of the 100 R. norvegicus captured in Tehran, 7% (n = 7/100) and 1% (n = 1/100) were positive for Leptospira spp. and Rabies virus, respectively. Leptospira spp. revealed the highest frequency (20%; 4/20) among R. norvegicus collected from the eastern part of Tehran. Rabies virus was detected only from the southern (5%; 1/20) part of Tehran. Results of the PCR method showed that the percentage of the rats tested positive for V. vulnificus was 5%. Overall, the surveyed zoonotic microorganisms had the highest (n = 5/20; 25%) and lowest (n = 1/20; 5%) frequency rates in the eastern and northern parts of Tehran, respectively. Conclusions: The results accentuate the necessity of implementing rodent control programs and regular disinfection as well as avoiding contact with rodent populations in urban environments.

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Simultaneous isolation and enumeration of virulent Vibrio cholerae and Vibrio vulnificus using an advanced MPN-PCR method

Archives of Microbiology ◽

10.1007/s00203-021-02613-y ◽

2021 ◽

Vol 204 (1) ◽

Author(s):

Jae-Hwa Lee ◽

Seul-Ki Park ◽

Fazlurrahman Khan ◽

Du-Min Jo ◽

Do-ha Lee ◽

...

Keyword(s):

Vibrio Cholerae ◽

Vibrio Vulnificus ◽

Pcr Method

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Simultaneous Isolation and Enumeration of Virulent Vibrio Cholerae and Vibrio Vulnificus using an Advanced MPN-PCR Method

10.21203/rs.3.rs-528186/v1 ◽

2021 ◽

Author(s):

Jae-Hwa Lee ◽

Seul-Ki Park ◽

Fazlurrahman Khan ◽

Du-Min Jo ◽

Do-ha Lee ◽

...

Keyword(s):

Vibrio Cholerae ◽

Vibrio Vulnificus ◽

Detection Method ◽

Low Cost ◽

Luria Bertani ◽

Most Probable Number ◽

Probable Number ◽

Pcr Method ◽

Cell Enumeration ◽

Differential Medium

Abstract Vibrio cholerae and Vibrio vulnificus are one of the critical foodborne pathogens that need to be intensively controlled their infection as a result of the intake and distribution of seafood, especially raw oysters. For this reason, various methods have already been developed for the detection and enumeration of these bacteria. The most probable number (MPN)-PCR (polymerase chain reaction) method is commonly used with the selective-differential medium for the efficiency and convenience of cell enumeration. One of the most frequently used for the detection of Vibrio spp. is Thiosulfate-Citrate-Bile salts-Sucrose (TCBS) agar. But this selective-differential medium can fail to distinguish between V. cholerae, V. vulnificus, and Vibrio alginolyticus. For this reason, the conventional MPN-PCR method with TCBS medium for the detection of Vibrio spp. has a problem that processing PCR to the two-times. This study suggests a simple and minimized detection method using one-time PCR and non-NaCl Luria-Bertani (LB-0) medium culture. This detection method is based on the difference in salt requirement between V. cholerae and V. vulnificus. Employing the developed methodology, the simultaneous cell enumeration of V. cholerae and V. vulnificus can be possible at a low cost. Furthermore, this study proposes a new specific primer to detect virulence-related genes from V. cholerae and V. vulnificus. This advanced MPN-PCR method was verified using bioaccumulated pacific oysters (Crassostrea gigas) by V. cholerae and V. vulnificus.

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Method for the specific identification of the emerging zoonotic pathogen Vibrio vulnificus Lineage 3 (formerly Biotype 3)

Journal of Clinical Microbiology ◽

10.1128/jcm.01763-20 ◽

2020 ◽

Author(s):

Hector Carmona-Salido ◽

Naiel Bisharat ◽

Carmen Amaro

Keyword(s):

Vibrio Vulnificus ◽

Genomic Analysis ◽

Epidemiological Studies ◽

Rapid Identification ◽

Comparative Genomic ◽

Zoonotic Pathogen ◽

Genomic Study ◽

Comparative Genomic Study ◽

Pcr Method ◽

Unique Ability

Vibrio vulnificus is a zoonotic pathogen that is spreading worldwide due to global warming. Lineage 3 (L3; formerly biotype 3) includes the strains of the species with the unique ability to cause fish farm-linked outbreaks of septicaemia. The L3-strains emerged recently and are particularly virulent and difficult to identify. Here we describe a new developed PCR method based on a comparative genomic study useful for both rapid identification and epidemiological studies of this interesting emerging group. The comparative genomic analysis also revealed the presence of a genetic duplication in the L3 strains that could be related to the unique ability of this lineage to produce septicemia outbreaks.

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Evaluation of the Polymerase Chain Reaction Method for Identification of Vibrio vulnificus Isolated from Marine Environments

Journal of Food Protection ◽

10.4315/0362-028x-60.1.81 ◽

1997 ◽

Vol 60 (1) ◽

pp. 81-83 ◽

Cited By ~ 23

Author(s):

EIJI AONO ◽

HARUO SUGITA ◽

JUNJIRO KAWASAKI ◽

HIROSHI SAKAKIBARA ◽

TAKUMI TAKAHASHI ◽

...

Keyword(s):

Polymerase Chain Reaction ◽

Dna Hybridization ◽

Vibrio Vulnificus ◽

Polymerase Chain Reaction Method ◽

Tokyo Bay ◽

Phenotypic Characteristics ◽

Chain Reaction ◽

Hemolysin Gene ◽

Pcr Method ◽

Polymerase Chain

The polymerase chain reaction (PCR) method for identification of Vibrio vulnificus in the marine environment was evaluated by comparing it to both the conventional and DNA-DNA hybridization methods. Of 13,325 isolates obtained from seawater and sediment samples, and oyster and goby specimens collected from the coastal waters of Tokyo Bay, Japan, only 61 isolates were identified as V. vulnificus on the basis of phenotypic characteristics and the amplification of the cytotoxin-hemolysin gene by the PCR method. All 61 isolates were further confirmed to be V.vulnificus by a DNA-DNA hybridization method and the API 20E system although they were divided into 13 groups on the basis of their API 20E profiles. These results strongly suggest that the PCR method is useful for identification of this organism.

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Application of a novel pathogenicity marker in a multiplex real-time PCR method to assess total and pathogenic Vibrio vulnificus in food and environmental samples

Food Control ◽

10.1016/j.foodcont.2013.07.007 ◽

2014 ◽

Vol 35 (1) ◽

pp. 274-283 ◽

Cited By ~ 5

Author(s):

Alejandro Garrido-Maestu ◽

María-José Chapela ◽

Belén Román ◽

Juan M. Vieites ◽

Ana G. Cabado

Keyword(s):

Real Time ◽

Real Time Pcr ◽

Environmental Samples ◽

Vibrio Vulnificus ◽

Pathogenic Vibrio ◽

Pcr Method

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Detection of vvha gene of Vibrio vulnificus and trh gene of Vibrio parahaemolyticus from Black Sea oysters using RT-PCR method

Journal of Biotechnology ◽

10.1016/j.jbiotec.2017.06.1032 ◽

2017 ◽

Vol 256 ◽

pp. S68 ◽

Cited By ~ 1

Author(s):

Tudor Laurentiu ◽

Mitranescu Elena ◽

Simion Violeta ◽

Pirvu Monica ◽

Tudor Aneta Laura

Keyword(s):

Black Sea ◽

Vibrio Parahaemolyticus ◽

Vibrio Vulnificus ◽

Rt Pcr ◽

Pcr Method

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Nested PCR method for rapid and sensitive detection of Vibrio vulnificus in fish, sediments, and water.

Applied and Environmental Microbiology ◽

10.1128/aem.61.9.3476-3478.1995 ◽

1995 ◽

Vol 61 (9) ◽

pp. 3476-3478 ◽

Cited By ~ 58

Author(s):

C R Arias ◽

E Garay ◽

R Aznar

Keyword(s):

Nested Pcr ◽

Vibrio Vulnificus ◽

Sensitive Detection ◽

Pcr Method

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Multiplex PCR detection of clinical and environmental strains ofVibrio vulnificusin shellfish

Canadian Journal of Microbiology ◽

10.1139/w04-085 ◽

2004 ◽

Vol 50 (11) ◽

pp. 911-922 ◽

Cited By ~ 34

Author(s):

Gitika Panicker ◽

Michael C.L Vickery ◽

Asim K Bej

Keyword(s):

Multiplex Pcr ◽

Real Time ◽

Real Time Pcr ◽

Vibrio Vulnificus ◽

High Specificity ◽

Sybr Green ◽

Sybr Green I ◽

Tissue Homogenate ◽

Oyster Tissue ◽

Pcr Method

In this study, we developed a PCR-based rapid detection method for clinically important pathogenic strains of Vibrio vulnificus. Positive amplification of the 504-bp viuB fragment was seen in all 22 clinical isolates tested but only in 8 out of 33 environmental isolates. The combination of the species-specific 205-bp vvh fragment along with viuB in a multiplexed PCR enabled us to confirm the presence of potentially pathogenic strains of V. vulnificus. No amplification of other Vibrio spp. or non-Vibrio bacteria was evidenced, suggesting a high specificity of detection by this method. The sensitivity of detection for both targeted genes was 10 pg of purified DNA, which correlated with 103V. vulnificus CFU in 1 mL of pure culture or 1 g un-enriched seeded oyster tissue homogenate. This sensitivity was improved to 1 CFU per gram of oyster tissue homogenate in overnight-enriched samples. A SYBR Green I based real-time PCR method was also developed that was shown to produce results consistent with the conventional PCR method. Application of the multiplexed real-time PCR to natural oyster tissue homogenates exhibited positive detection of vvh in 51% of the samples collected primarily during the summer months; however, only 15% of vvh positive samples exhibited viuB amplicons. The rapid, sensitive, and specific detection of clinically important pathogenic V. vulnificus in shellfish would be beneficial in reducing illnesses and deaths caused by this pathogen.Key words: Vibrio, multiplex PCR, shellfish, SYBR Green I, real-time PCR.

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