• Journal of Microbiology and Biotechnology
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• 제21권9호
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• pp.914-920
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• 2011

Vibrio cholerae utilizes mannitol through an operon of the phosphoenolpyruvate-dependent phosphotransferase (PTS) type. A gene, mtlD, encoding mannitol-1-phosphate dehydrogenase was identified within the 3.9 kb mannitol operon of V. cholerae. The mtlD gene was cloned from V. cholerae O395, and the recombinant enzyme was functionally expressed in E. coli as a $6{\times}$His-tagged protein and purified to homogeneity. The recombinant protein is a monomer with a molecular mass of 42.35 kDa. The purified recombinant MtlD reduced fructose 6-phosphate (F6P) using NADH as a cofactor with a $K_m$ of $1.54{\pm}0.1$ mM and $V_{max}$ of $320.8{\pm}7.81\;{\mu}mol$/min/mg protein. The pH and temperature optima for F6P reduction were determined to be 7.5 and $37^{\circ}C$, respectively. Using quantitative real-time PCR analysis, mtlD was found to be constitutively expressed in V. cholerae, but the expression was up-regulated when grown in the presence of mannitol. The MtlD expression levels were not significantly different between V. cholerae O1 and non-O1 strains.

• 한국식품위생안전성학회지
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• 제35권1호
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• pp.37-44
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• 2020

패류 중 소비량이 높은 굴에서의 고병원성 Vibrio균(V. vulnificus와 V. cholerae)의 식중독 발생 가능성을 분석하기 위하여 위해평가를 실시하였다. 남해권역, 서해권역, 수도권에서 유통되고 있는 굴 88개를 채취하여 V. vulnificus와 V. cholerae의 오염실태를 조사하고, 생굴에서의 V. vulnificus와 V. cholerae의 생장 경향을 관찰하였다. 2017년의 국민건강영양조사 데이터를 활용하여 생굴의 섭취자 비율 및 섭취량을 조사하였으며, 생굴 섭취로 인한 V. vulnificus와 V. cholerae의 식중독 발생 가능성을 분석하기 위해 @RISK를 통해 위해평가를 실시하였다. 88개의 생굴에서 V. vulnificus와 V. cholerae는 검출되지 않아 Beta distribution과 자연로그를 이용한 식을 통해 초기오염수준을 추정하였다. 그 결과 두 세균 모두 -3.6 Log CFU/g으로 생굴에 오염되어 있는 것으로 추정되었다. 생굴에 bioaccumulation된 V. vulnificus와 V. cholerae는 생장하지 않고 초기 접종 수준을 유지하는 것으로 확인되었다. 생굴을 섭취했다고 응답한 사람은 7,167명 중 25명이었으며, 따라서 섭취자 비율은 0.35%로 나타났다. 섭취량에 대한 최적확률분포는 exponential distribution으로 나타났으며 생굴의 평균 섭취량은 66.8 g으로 확인되었다. V. vulnificus와 V. cholerae의 용량-반응 모델은 Beta-Poisson model을 사용하였다. 이상의 데이터를 이용하여 위해평가 시뮬레이션을 개발하고 분석하였다. 초기오염수준으로 오염된 생굴을 바로 섭취함으로써 발생할 수 있는 식중독발생 가능성은 V. vulnificus의 경우 평균 9.08×10^-15, V. cholerae는 8.16×10^-13이며, 섭취자 비율이 식중독 발생 가능성에 가장 큰 영향을 미치는 것으로 확인되었다.

• 한국수산과학회지
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• 제28권1호
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• pp.60-66
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• 1995

새로운 식중독 원인균으로 주목을 받고 있고 수권에서 잘 검출된다고 알려져 있는 V cholerae non-O1과 V. mimicus를 대상으로 민물, 기수 및 해수에서 염도와 온도의 변화에 따른 균수의 증감을 조사한 결과는 다음과 같다. 1. 민물에서 V. cholerae non-O1은 $26\pm1^{\circ}C$보다 4와 $15^{\circ}C$에서 생잔기간이 길었으며 V. mimius는 4와 $26\pm1^{\circ}C$보다 $15^{\circ}C$에서 생잔기간이 길었다. 2. 기수에서는 두 균주 모두 15, 4, $26\pm1^{\circ}C$으로 생잔기간이 길었다. 3. 해수에서는 두 균주 공히 온도가 높을수록 생잔기간이 짧았으며, $4^{\circ}C$에서 실험조건 중 생잔기간이 가장 길었다. 4. 민물에서는 V. mimicus보다 V. cholerae non-O1의 안정성이 높았고, 기수에서는 V. cholerae non-O1보다 V. mimicus의 안정성이 높았으며 해수에서는 두 균 사이에 큰 차이가 없었다. 5. 염도와 온도의 변화는 V. cholerae non-O1과 V. mimicus의 생잔에 중요한 인자이다.

• Advances in Traditional Medicine
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• 제8권2호
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• pp.171-177
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• 2008

Epigallocatechin 3-gallate (EGCG), one of the major catechins of tea, was isolated from the decaffeinated, crude methanolic extract of Indian green tea (Camellia sinensis L. O. Kuntze) using chromatographic techniques. EGCG was then screened for antidiarrhoeal activity against 30 strains (clinical isolates) of V. cholerae, which is a well known Gram negative bacillus functioning as the pathogen of cholera. V. cholerae strains like V. cholerae 69, 71, 83, 214, 978, 1021, 1315, 1347, 1348, 569B and ATCC 14033 were inhibited by EGCG at a concentration of $25\;{\mu}g/ml$ whereas V. cholerae 10, 522, 976 were even more sensitive, being inhibited at $10\;{\mu}g/ml$ level. However, V. cholerae DN 16, DN 26, 30, 42, 56, 58, 113, 117, 564, 593, 972 and ATCC 14035 were inhibited at $50\;{\mu}g/ml$ level of EGCG. Only four strains were inhibited at $100\;{\mu}g/ml$. In this study the isolated compound was found to be bacteriostatic in its mechanism of action. In the in vivo experiment using the rabbit ileal loop model two different dosages of EGCG ($500\;{\mu}g/ml$ and $1,000\;{\mu}g/ml$) were able to protect the animals when they were challenged with V. cholerae 569B in the ileum.

• 한국자원식물학회:학술대회논문집
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• 한국자원식물학회 2019년도 추계학술대회
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• pp.8-8
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• 2019

The stringent response (SR) is characterized as a bacterial defense mechanism in response to various growth-inhibiting stresses. It is activated by accumulation of a small nucleotide regulator, (p)ppGpp, and induces global changes in bacterial transcription and translation. Recent work from our group has shown that (p)ppGpp plays a critical role in virulence and survival in Vibrio cholerae. The genes, relA and relV, are involved in the production of (p)ppGpp, while the spoT gene encodes an enzyme that hydrolyzes it in V. cholerae. A mutant strain defective in (p)ppGpp production (i.e. ${\Delta}relA{\Delta}relV{\Delta}spoT$ mutant) lost the ability to produce cholera toxin (CT) and lost their viability due to uncontrolled production of organic acids, when grown with extra glucose. In contrast, the ${\Delta}relA{\Delta}spoT$ mutant, a (p)ppGpp overproducer strain, produced enhanced level of CT and exhibited better growth in glucose supplemented media via glucose metabolic switch from organic fermentation to acetoin, a neutral fermentation end product, fermentation. These findings indicates that (p)ppGpp, in addition to its well-known role as a SR mediator, positively regulates CT production and maintenance of growth fitness in V. cholerae. This implicates SR as a promising drug target, inhibition of which may possibly downregulate V. cholerae virulence and survival fitness. Therefore, we screened a chemical library and identified a compound that induces medium acidification (termed iMAC) and thereby loss of wild type V. cholerae viability under glucose-rich conditions. Further, we present a potential mechanism by which the compound inhibits (p)ppGpp accumulation. Together, these results indicate that iMAC treatment causes V. cholerae cells to produce significantly less (p)ppGpp, an important regulator of the bacterial virulence and survival response, and further suggesting that it has a therapeutic potential to be developed as a novel antibacterial agent against cholera.

• 한국수산과학회지
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• 제56권5호
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• pp.626-633
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• 2023

Vibrio spp. are aquatic bacteria that are ubiquitous in warm estuarine and marine environments. Especially, V. vulnificus and V. cholerae are currently known to cause potentially fatal infections in humans. This study investigated the distribution and antibiotic resistance of V. vulnificus and V. cholerae isolated from coastal areas of Gyeongsangnam-do in 2022. A total of 252 samples of water, shellfish and coastal sediment were collected from 7 locations along the coast, and 124 samples of fishery products were collected from markets. Among the 252 samples, forty-four V. vulnificus (11.7%) and fourteen V. cholerae non-O1/non-O139 (3.7%), none of which carried the ctx gene, were isolated. Out of the 124 samples, 6 (4.8%) tested positive for V. vulnificus and V. cholerae was not detected. The isolation rates of V. vulnificus and V. cholerae showed a significant correlation with environmental factors such as seawater temperature and salinity. In an antibiotic resistance test, V. vulnificus was susceptible to amikacin, gentamicin, imipenem trimethoprim/sulfamethoxazole, and ciprofloxacin, but resistant to cefoxitin (100.0%), followed by tetracycline (9.1%). Multidrug resistance was also observed. Continuous monitoring of Vibrio pathogens with water temperature and salinity is expected to help reduce the outbreaks, and rational use of antibiotic agents is needed to prevent the accession of antibiotic-resistant microorganisms in aquatic ecosystems.

• Genomics & Informatics
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• 제13권2호
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• pp.53-59
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• 2015

In developing countries threat of cholera is a significant health concern whenever water purification and sewage disposal systems are inadequate. Vibrio cholerae is one of the responsible bacteria involved in cholera disease. The complete genome sequence of V. cholerae deciphers the presence of various genes and hypothetical proteins whose function are not yet understood. Hence analyzing and annotating the structure and function of hypothetical proteins is important for understanding the V. cholerae. V. cholerae O139 is the most common and pathogenic bacterial strain among various V. cholerae strains. In this study sequence of six hypothetical proteins of V. cholerae O139 has been annotated from NCBI. Various computational tools and databases have been used to determine domain family, protein-protein interaction, solubility of protein, ligand binding sites etc. The three dimensional structure of two proteins were modeled and their ligand binding sites were identified. We have found domains and families of only one protein. The analysis revealed that these proteins might have antibiotic resistance activity, DNA breaking-rejoining activity, integrase enzyme activity, restriction endonuclease, etc. Structural prediction of these proteins and detection of binding sites from this study would indicate a potential target aiding docking studies for therapeutic designing against cholera.

• Journal of Microbiology and Biotechnology
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• 제32권11호
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• pp.1396-1405
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• 2022

Cholera remains a major global public health problem, for which oral cholera vaccines (OCVs) being a valuable strategy. Patients, who have recovered from cholera, develop antibody responses against LPS, cholera toxin (CT), toxin-coregulated pilus (TCP) major subunit A (TcpA) and other antigens; thus, these responses are potentially important contributors to immunity against Vibrio cholerae infection. However, assessments of the efficacy of current OCVs, especially inactivated OCVs, have focused primarily on O-antigen-specific antibody responses, suggesting that more sophisticated strategies are required for inactivated OCVs to induce immune responses against TCP, CT, and other antigens. Previously, we have shown that the toxT-139F allele enables V. cholerae strains to produce CT and TCP under simple laboratory culture conditions. Thus, we hypothesized that V. cholerae strains that express TCP via the toxT-139F allele induce TCP-specific antibody responses. As anticipated, V. cholerae strains that expressed TCP through the toxT-139F allele elicited antibody responses against TCP when the inactivated bacteria were delivered via a mouse model. We have further developed TCP-expressing V. cholerae strains that have been used in inactivated OCVs and shown that they effect an antibody response against TcpA in vivo, suggesting that V. cholerae strains with the toxT-139F allele are excellent candidates for cholera vaccines.

• Journal of Microbiology and Biotechnology
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• 제33권6호
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• pp.736-744
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• 2023

The introduction of the toxT-139F allele triggers the expression of TCP (toxin co-regulated pilus) and CT (cholera toxin) under simple laboratory culture conditions in most Vibrio cholerae strains. Such V. cholerae strains, especially strains that have been used in OCVs (oral cholera vaccines), can induce antibody responses against TCP in animal models. However, CT produced in these V. cholerae strains is secreted into the culture medium. In this study, V. cholerae strains that can express intracellular CTB under the control of the toxT-139F allele have been constructed for potential application in OCVs. First, we constructed a recombinant plasmid directly linking the ctxAB promoter to ctxB without ctxA and confirmed CTB expression from the plasmid in V. cholerae containing the toxT-139F allele. We constructed another recombinant plasmid to express NtrCTB, from which 14 internal amino acids-from the 7^th to the 20^th amino acid-of the leader peptide of CTB have been omitted, and we found that NtrCTB remained in the cells. Based on those results, we constructed V. cholerae strains in which chromosomal ctxAB is replaced by ntrctxB or ntrctxB-dimer. Both NtrCTB and NtrCTB-dimer remained in the bacterial cells, and 60% of the NtrCTB-dimer in the bacterial cells was maintained in a soluble form. To develop improved OCVs, these strains could be tested to see whether they induce immune responses against CTB in animal models.