RESUMEN

The expression of two major virulence factors of Vibrio cholerae, cholera toxin (CT) and toxin coregulated pilus (TCP), is induced by environmental stimuli through a cascade of interactions among regulatory proteins known as the ToxR regulon when the bacteria reach the human small intestine. ToxT is produced via the ToxR regulon and acts as the direct transcriptional activator of CT (ctxAB), TCP (tcp gene cluster), and other virulence genes. Unsaturated fatty acids (UFAs) and several smallmolecule inhibitors of ToxT have been developed as antivirulence agents against V. cholerae. This study reports the inhibitory effects of fatty acids and virstatin (a small-molecule inhibitor of ToxT) on the transcriptional activation functions of ToxT in isogenic derivatives of V. cholerae strains containing various toxT alleles. The fatty acids and virstatin had discrete effects depending on the ToxT allele (different by 2 amino acids), V. cholerae strain, and culture conditions, indicating that V. cholerae strains could overcome the effects of UFAs and small-molecule inhibitors by acquiring point mutations in toxT. Our results suggest that small-molecule inhibitors should be examined thoroughly against various V. cholerae strains and toxT alleles during development.

RESUMEN

Vibrio cholerae is a cholera-causing pathogen known to instigate severe contagious diarrhea that affects millions globally. Survival of vibrios depend on a combination of multicellular responses and adapt to changes that prevail in the environment. This process is achieved through a strong communication at the cellular level, the process has been recognized as quorum sensing (QS). The severity of infection is highly dependent on the QS of vibrios in the gut milieu. The quorum may exist in a low/high cell density (LCD/HCD) state to exert a positive or negative response to control the regulatory pathogenic networks. The impact of this regulation reflects on the transition of pathogenic V. cholerae from the environment to infect humans and cause outbreaks or epidemics of cholera. In this context, the review portrays various regulatory processes and associated virulent pathways, which maneuver and control LCD and HCD states for their survival in the host. Although several treatment options are existing, promotion of therapeutics by exploiting the virulence network may potentiate ineffective antibiotics to manage cholera. In addition, this approach is also useful in resource-limited settings, where the accessibility to antibiotics or conventional therapeutic options is limited.

RESUMEN

The expression of the two major virulence genes of Vibrio cholerae-tcpA (the major subunit of the toxin co-regulated pilus) and ctxAB (cholera toxin)-is regulated by the ToxR regulon, which is triggered by environmental stimuli during infection within the human small intestine. Special culture methods are required to induce the expression of virulence genes in V. cholerae in the laboratory setting. In the present study, induction of the expression of virulence genes by two point mutations (65th and 139th amino acids) in toxT, which is produced by the ToxR regulon and activates the transcription of the virulence genes in V. cholerae, under laboratory culture conditions has been investigated. Each of the four toxT alleles assessed displayed different transcriptional activator functions in a given V. cholerae strain. Although the ToxR regulon has been known to not be expressed by El Tor biotype V. cholerae strains cultured under standard laboratory conditions, the variant toxT alleles that we assessed in this study enabled the expression virulence genes in El Tor biotype strains grown under simple culture conditions comprising shake culture in LB medium, suggesting that the regulation of virulence gene expression may be regulated more complexly than previously thought and may involve additional factors beyond the production of ToxT by the ToxR regulon.

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RESUMEN

Cholera is an acute secretory diarrhoeal disease caused by the bacterium Vibrio cholerae. The key determinants of cholera pathogenicity, cholera toxin (CT), and toxin co-regulated pilus (TCP) are part of the genome of two horizontally acquired Mobile Genetic Elements (MGEs), CTXΦ, and Vibrio pathogenicity island 1 (VPI-1), respectively. Besides, V. cholerae genome harbors several others MGEs that provide antimicrobial resistance, metabolic functions, and other fitness traits. VPI-1, one of the most well characterized genomic island (GI), deserved a special attention, because (i) it encodes many of the virulence factors that facilitate development of cholera (ii) it is essential for the acquisition of CTXΦ and production of CT, and (iii) it is crucial for colonization of V. cholerae in the host intestine. Nevertheless, VPI-1 is ubiquitously present in all the epidemic V. cholerae strains. Therefore, to understand the role of MGEs in the evolution of cholera pathogen from a natural aquatic habitat, it is important to understand the VPI-1 encoded functions, their acquisition and possible mode of dissemination. In this review, we have therefore discussed our present understanding of the different functions of VPI-1 those are associated with virulence, important for toxin production and essential for the disease development.

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RESUMEN

Efforts to develop vaccines for prevention of acute diarrhea have been going on for more than 40 y with partial success. The myriad of pathogens, more than 20, that have been identified as a cause of acute diarrhea throughout the years pose a significant challenge for selecting and further developing the most relevant vaccine candidates. Based on pathogen distribution as identified in epidemiological studies performed mostly in low-resource countries, rotavirus, Cryptosporidium, Shigella, diarrheogenic E. coli and V. cholerae are predominant, and thus the main targets for vaccine development and implementation. Vaccination against norovirus is most relevant in middle/high-income countries and possibly in resource-deprived countries, pending a more precise characterization of disease impact. Only a few licensed vaccines are currently available, of which rotavirus vaccines have been the most outstanding in demonstrating a significant impact in a short time period. This is a comprehensive review, divided into 2 articles, of nearly 50 vaccine candidates against the most relevant viral and bacterial pathogens that cause acute gastroenteritis. In order to facilitate reading, sections for each pathogen are organized as follows: i) a discussion of the main epidemiological and pathogenic features; and ii) a discussion of vaccines based on their stage of development, moving from current licensed vaccines to vaccines in advanced stage of development (in phase IIb or III trials) to vaccines in early stages of clinical development (in phase I/II) or preclinical development in animal models. In this first article we discuss rotavirus, norovirus and Vibrio cholerae. In the following article we will discuss Shigella, Salmonella (non-typhoidal), diarrheogenic E. coli (enterotoxigenic and enterohemorragic), and Campylobacter jejuni.

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BACKGROUND AND OBJECTIVES: The toxin co-regulated pilus A (TcpA) has been described as a critical pathogenicity factor of Vibrio cholerae. TcpA is a candidate for making subunit vaccine against cholera. The aim of this study was to produce a candidate vaccine by expressing recombinant TcpA in E. coli. MATERIALS AND METHODS: In this study, the toxin co-regulated pilus A gene from EL-Tor, V. cholerae subspecies, was amplified by PCR and sub-cloned into prokaryotic expression vector pGEX4T1. E. coli BL21 (DE3) was transformed with pGEX4T1- TcpA and gene expression was induced by IPTG and purified by GST resin. The integrity of the product was confirmed by Western blot analysis using a standard rabbit anti-V. cholerae antibody. Sera reactivity of infected individuals was further analyzed against the recombinant TcpA protein. RESULTS: The concentration of purified recombinant protein was calculated to be 8 mg/L of initial culture. The integrity of product was confirmed by Western blot analysis using a standard rabbit anti V. cholerae antibody. Sera reactivity of infected individual was further analyzed against the recombinant TcpA protein. The obtained data indicated that recombinant TcpA protein from V. cholerae was recognized by patient serum and animal sera. CONCLUSION: These results show that the recombinant TcpA is antigenic and could be used in a carrier host as an oral vaccine against cholera.