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1.
Front Microbiol ; 14: 1178538, 2023.
Article in English | MEDLINE | ID: mdl-37283925

ABSTRACT

The aquatic bacterium Vibrio cholerae is the etiological agent of the diarrheal disease cholera, which has plagued the world for centuries. This pathogen has been the subject of studies in a vast array of fields, from molecular biology to animal models for virulence activity to epidemiological disease transmission modeling. V. cholerae genetics and the activity of virulence genes determine the pathogenic potential of different strains, as well as provide a model for genomic evolution in the natural environment. While animal models for V. cholerae infection have been used for decades, recent advances in this area provide a well-rounded picture of nearly all aspects of V. cholerae interaction with both mammalian and non-mammalian hosts, encompassing colonization dynamics, pathogenesis, immunological responses, and transmission to naïve populations. Microbiome studies have become increasingly common as access and affordability of sequencing has improved, and these studies have revealed key factors in V. cholerae communication and competition with members of the gut microbiota. Despite a wealth of knowledge surrounding V. cholerae, the pathogen remains endemic in numerous countries and causes sporadic outbreaks elsewhere. Public health initiatives aim to prevent cholera outbreaks and provide prompt, effective relief in cases where prevention is not feasible. In this review, we describe recent advancements in cholera research in these areas to provide a more complete illustration of V. cholerae evolution as a microbe and significant global health threat, as well as how researchers are working to improve understanding and minimize impact of this pathogen on vulnerable populations.

2.
STAR Protoc ; 3(4): 101810, 2022 12 16.
Article in English | MEDLINE | ID: mdl-36386878

ABSTRACT

A zebrafish model was developed to study AIEC colonization, invasion, and inflammation. This model can also be used to study the beneficial effects of a probiotic on AIEC infection of adult zebrafish. Bacteria are grown in vitro and then fish are infected with AIEC by immersion. Subsequently, colonization and inflammation can be assessed. Exposing fish to probiotic at different time points relative to AIEC can determine beneficial effects of probiotics as prophylactics or therapeutics against AIEC. For complete details on the use and execution of this protocol, please refer to Nag et al. (2022).


Subject(s)
Escherichia coli Infections , Zebrafish , Animals , Escherichia coli , Bacterial Adhesion , Escherichia coli Infections/therapy , Inflammation/microbiology
3.
iScience ; 25(7): 104572, 2022 Jul 15.
Article in English | MEDLINE | ID: mdl-35769878

ABSTRACT

Adherent-invasive Escherichia coli (AIEC) is an opportunistic pathogen associated with major inflammatory bowel disease, Crohn disease, and ulcerative colitis. Unfavorable conditions push commensal AIEC to induce gut inflammation, sometimes progressing to inflammation-induced colon cancer. Recently, zebrafish have emerged as a useful model to study human intestinal pathogens. Here, a zebrafish model to study AIEC infection was developed. Bath inoculation with AIEC resulted in colonization and tissue disruption in the zebrafish intestine. Gene expression of pro-inflammatory markers including interleukin-1ß (IL-1ß), tumor necrosis factor alpha (TNFα), interferon-γ (IFNγ), and S100A-10b (akin to human calprotectin) in the zebrafish intestine was significantly induced by AIEC infection. The probiotic E. coli Nissle 1917 (EcN) was tested as a therapeutic and prophylactic against AIEC infection and reduced AIEC colonization, tissue damage, and pro-inflammatory responses in zebrafish. Furthermore, EcN diminished the propionic-acid-augmented hyperinfection of AIEC in zebrafish. Thus, this study shows the efficacy of EcN against AIEC in an AIEC-zebrafish model.

4.
Infect Immun ; 90(3): e0046621, 2022 03 17.
Article in English | MEDLINE | ID: mdl-35072520

ABSTRACT

Vibrio cholerae, the cause of human cholera, is an aquatic bacterium found in association with a variety of animals in the environment, including many teleost fish species. V. cholerae infection induces a proinflammatory response followed by a noninflammatory convalescent phase. Neutrophils are integral to this early immune response. However, the relationship between the neutrophil-associated protein calprotectin and V. cholerae has not been investigated, nor have the effects of limiting transition metals on V. cholerae growth. Zebrafish are useful as a natural V. cholerae model as the entire infectious cycle can be recapitulated in the presence of an intact intestinal microbiome and mature immune responses. Here, we demonstrate that zebrafish produce a significant neutrophil, interleukin 8 (IL-8), and calprotectin response following V. cholerae infection. Bacterial growth was completely inhibited by purified calprotectin protein or the chemical chelator N,N,N',N'-tetrakis(2-pyridinylmethyl)-1,2-ethanediamine (TPEN), but growth was recovered by the addition of the transition metals zinc and manganese. The expression of downstream calprotectin targets was also significantly increased in the zebrafish. These findings illuminate the role of host calprotectin in combating V. cholerae infection. Inhibition of V. cholerae growth through metal limitation may provide new approaches in the development of anti-V. cholerae therapeutics. This study also establishes a major role for calprotectin in combating infectious diseases in zebrafish.


Subject(s)
Cholera , Vibrio cholerae , Animals , Cholera/microbiology , Leukocyte L1 Antigen Complex , Neutrophils , Vibrio cholerae/physiology , Zebrafish
5.
Front Cell Infect Microbiol ; 11: 722520, 2021.
Article in English | MEDLINE | ID: mdl-34888255

ABSTRACT

The gram-negative bacterium Vibrio cholerae causes the life-threatening diarrheal disease cholera, which is spread through the ingestion of contaminated food or water. Cholera epidemics occur largely in developing countries that lack proper infrastructure to treat sewage and provide clean water. Numerous vertebrate fish species have been found to be natural V. cholerae hosts. Based on these findings, zebrafish (Danio rerio) have been developed as a natural host model for V. cholerae. Diarrheal symptoms similar to those seen in humans are seen in zebrafish as early as 6 hours after exposure. Our understanding of basic zebrafish immunology is currently rudimentary, and no research has been done to date exploring the immune response of zebrafish to V. cholerae infection. In the present study, zebrafish were infected with either pandemic El Tor or non-pandemic, environmental V. cholerae strains and select immunological markers were assessed to determine cellular immunity and humoral immunity. Significant increases in the gene expression of two transcription factors, T-bet and GATA3, were observed in response to infection with both V. cholerae strains, as were levels of mucosal related antibodies. Additionally, the cytokine IL-13 was shown to be significantly elevated and paralleled the mucin output in zebrafish excretions, strengthening our knowledge of IL-13 induced mucin production in cholera. The data presented here further solidify the relevancy of the zebrafish model in studying V. cholerae, as well as expanding its utility in the field of cholera immunology.


Subject(s)
Cholera , Vibrio cholerae , Animals , Diarrhea , Humans , Immunity , Zebrafish
6.
J Bacteriol ; 202(24)2020 11 19.
Article in English | MEDLINE | ID: mdl-32778562

ABSTRACT

Vibrio is a large and diverse genus of bacteria, of which most are nonpathogenic species found in the aquatic environment. However, a subset of the Vibrio genus includes several species that are highly pathogenic, either to humans or to aquatic animals. In recent years, Danio rerio, commonly known as the zebrafish, has emerged as a major animal model used for studying nearly every aspect of biology, including infectious diseases. Zebrafish are especially useful because the embryos are transparent, larvae are small and facilitate imaging studies, and numerous transgenic fish strains have been constructed. Zebrafish models for several pathogenic Vibrio species have been described, and indeed a fish model is highly relevant for the study of aquatic bacterial pathogens. Here, we summarize the zebrafish models that have been used to study pathogenic Vibrio species to date.


Subject(s)
Disease Models, Animal , Vibrio Infections/microbiology , Vibrio/physiology , Zebrafish , Animals , Humans , Vibrio/classification , Vibrio/genetics , Vibrio/pathogenicity , Virulence , Zebrafish/microbiology
7.
Microb Pathog ; 140: 103927, 2020 Mar.
Article in English | MEDLINE | ID: mdl-31846743

ABSTRACT

The use of traditional foods and beverages or their bioactive compounds as anti-virulence agents is a new alternative method to overcome the increased global emergence of antimicrobial resistance in enteric pathogens. In the present study, we investigated the anti-virulence activity of a polyphenolic fraction previously isolated from Kombucha, a 14-day fermented beverage of sugared black tea, against Vibrio cholerae O1. The isolated fraction was mainly composed of the polyphenols catechin and isorhamnetin. The fraction, the individual polyphenols and the combination of the individual polyphenols significantly inhibited bacterial swarming motility and expression of flagellar regulatory genes motY and flaC, even at sub-inhibitory concentrations. The polyphenolic compounds also decreased bacterial protease secretion and mucin penetration in vitro. In vivo study revealed that the polyphenolic fraction significantly inhibited V. cholerae induced fluid accumulation in the rabbit ileal loop model and intestinal colonization in suckling mice model. Therefore, the anti-virulence activity of the Kombucha polyphenolic fraction involved inhibition of motility and protease secretion of V. cholerae, thus preventing bacterial penetration through the mucin layer as well as fluid accumulation and bacterial colonization in the intestinal epithelial cells. The overall results implied that Kombucha might be considered as a potential alternative source of anti-virulence polyphenols against V. cholerae. To the best of our knowledge, this is the first report on the anti-virulence activity of Kombucha, mostly attributed to its polyphenolic content.


Subject(s)
Kombucha Tea , Polyphenols/pharmacology , Vibrio cholerae/drug effects , Animals , Anti-Bacterial Agents/pharmacology , Bacterial Outer Membrane Proteins/genetics , Bacterial Outer Membrane Proteins/metabolism , Bacterial Proteins/drug effects , Catechin/pharmacology , Cell Movement/drug effects , Cholera/drug therapy , Gene Expression/drug effects , Intestine, Small/drug effects , Intestine, Small/microbiology , Mice , Peptide Hydrolases/drug effects , Plant Extracts/pharmacology , Quercetin/analogs & derivatives , Quercetin/pharmacology , Rabbits , Vibrio cholerae/pathogenicity , Virulence/drug effects , Virulence/genetics , Virulence Factors/genetics , Virulence Factors/metabolism
8.
Zebrafish ; 16(4): 388-400, 2019 08.
Article in English | MEDLINE | ID: mdl-31145047

ABSTRACT

Zebrafish (Danio rerio) are an attractive model organism for scientific studies, including host-microbe interactions. The organism is particularly useful for studying aquatic microbes that can colonize vertebrate hosts, including Vibrio cholerae. Previous studies have established the presence of a core zebrafish intestinal microbiome, and tracked the development of the zebrafish intestinal microbiome from the larval stage to adulthood. An unexplored matter in this host-microbe relationship is the effect of the housing system on the zebrafish intestinal and tank water microbiomes. In this study, we used 16S rRNA gene sequencing to investigate the response of zebrafish intestinal and tank water microbiomes to a change in housing conditions. Zebrafish in the separated fish tanks showed no initial differences in the structures of their intestinal microbial profiles; the same prominent bacteria were present and abundant across tanks. Immediately after the housing switch, the zebrafish intestinal microbial profiles changed in composition and structure. Within 5 days of the housing switch, the intestinal microbiome had stabilized, and changed significantly from the prehousing switch profile. This study demonstrates that although external factors can significantly perturb and alter the zebrafish intestinal microbiome, the microbiome displays a large level of selective resilience whose primary members (namely Vibrio) persist.


Subject(s)
Gastrointestinal Microbiome , Housing, Animal , Water Microbiology , Zebrafish/microbiology , Animals , RNA, Bacterial/analysis , RNA, Ribosomal, 16S/analysis
9.
Clin Cancer Res ; 25(15): 4791-4807, 2019 08 01.
Article in English | MEDLINE | ID: mdl-30940656

ABSTRACT

PURPOSE: The radiosensitivity of the normal intestinal epithelium is the major limiting factor for definitive radiotherapy against abdominal malignancies. Radiosensitizers, which can be used without augmenting radiation toxicity to normal tissue, are still an unmet need. Inhibition of proteosomal degradation is being developed as a major therapeutic strategy for anticancer therapy as cancer cells are more susceptible to proteasomal inhibition-induced cytotoxicity compared with normal cells. Auranofin, a gold-containing antirheumatoid drug, blocks proteosomal degradation by inhibiting deubiquitinase inhibitors. In this study, we have examined whether auranofin selectively radiosensitizes colon tumors without promoting radiation toxicity in normal intestine. EXPERIMENTAL DESIGN: The effect of auranofin (10 mg/kg i.p.) on the radiation response of subcutaneous CT26 colon tumors and the normal gastrointestinal epithelium was determined using a mouse model of abdominal radiation. The effect of auranofin was also examined in a paired human colonic organoid system using malignant and nonmalignant tissues from the same patient. RESULTS: Both in the mouse model of intestinal injury and in the human nonmalignant colon organoid culture, auranofin pretreatment prevented radiation toxicity and improved survival with the activation of p53/p21-mediated reversible cell-cycle arrest. However, in a mouse model of abdominal tumor and in human malignant colonic organoids, auranofin inhibited malignant tissue growth with inhibition of proteosomal degradation, induction of endoplasmic reticulum stress/unfolded protein response, and apoptosis. CONCLUSIONS: Our data suggest that auranofin is a potential candidate to be considered as a combination therapy with radiation to improve therapeutic efficacy against abdominal malignancies.


Subject(s)
Auranofin/pharmacology , Colonic Neoplasms/radiotherapy , Intestinal Mucosa/drug effects , Protective Agents/pharmacology , Radiation Injuries/prevention & control , Radiation-Sensitizing Agents/pharmacology , Tumor Suppressor Protein p53/metabolism , Animals , Antirheumatic Agents/pharmacology , Apoptosis , Cell Line, Tumor , Cell Survival/drug effects , Colonic Neoplasms/metabolism , Colonic Neoplasms/pathology , Cyclin-Dependent Kinase Inhibitor p21/metabolism , Endoplasmic Reticulum Stress/drug effects , Humans , Intestinal Mucosa/injuries , Intestinal Mucosa/metabolism , Intestinal Mucosa/pathology , Male , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Radiation Injuries/pathology , Unfolded Protein Response/drug effects , Xenograft Model Antitumor Assays
10.
J Bacteriol ; 201(14)2019 07 15.
Article in English | MEDLINE | ID: mdl-30858296

ABSTRACT

The phage shock protein (Psp) system is a stress response pathway that senses and responds to inner membrane damage. The genetic components of the Psp system are present in several clinically relevant Gram-negative bacteria, including Vibrio cholerae However, most of the current knowledge about the Psp response stems from in vitro studies in Escherichia coli and Yersinia enterocolitica In fact, the Psp response in V. cholerae has remained completely uncharacterized. In this study, we demonstrate that V. cholerae does have a functional Psp response system. We found that overexpression of GspD (EpsD), the type II secretion system secretin, induces the Psp response, whereas other V. cholerae secretins do not. In addition, we have identified several environmental conditions that induce this stress response. Our studies on the genetic regulation and induction of the Psp system in V. cholerae suggest that the key regulatory elements are conserved with those of other Gram-negative bacteria. While a psp null strain is fully capable of colonizing the infant mouse intestine, it exhibits a colonization defect in a zebrafish model, indicating that this response may be important for disease transmission in the environment. Overall, these studies provide an initial understanding of a stress response pathway that has not been previously investigated in V. choleraeIMPORTANCEVibrio cholerae leads a dual life cycle, as it can exist in the aquatic environment and colonize the human small intestine. In both life cycles, V. cholerae encounters a variety of stressful conditions, including fluctuating pH and temperature and exposure to other agents that may negatively affect cell envelope homeostasis. The phage shock protein (Psp) response is required to sense and respond to such insults in other bacteria but has remained unstudied in V. cholerae Interestingly, the Psp system has protein homologs, principally, PspA, in a number of bacterial clades as well as in archaea and plants. Therefore, our findings not only fill a gap in knowledge about an unstudied extracytoplasmic stress response in V. cholerae, but also may have far-reaching implications.


Subject(s)
Bacterial Proteins/metabolism , Cholera Toxin/metabolism , Heat-Shock Proteins/metabolism , Type II Secretion Systems/metabolism , Vibrio cholerae/genetics , Animals , Bacterial Proteins/genetics , Cholera Toxin/genetics , Gene Expression Regulation, Bacterial , Heat-Shock Proteins/genetics , Mice , Mucins/analysis , Type II Secretion Systems/genetics , Vibrio cholerae/metabolism , Virulence , Zebrafish/microbiology
12.
Infect Immun ; 86(12)2018 12.
Article in English | MEDLINE | ID: mdl-30249745

ABSTRACT

The Vibrio cholerae O1 serogroup is responsible for pandemic cholera and is divided into the classical and El Tor biotypes. Classical V. cholerae produces acid when using glucose as a carbon source, whereas El Tor V. cholerae produces the neutral product acetoin when using glucose as a carbon source. An earlier study demonstrated that Escherichia coli strains that metabolize glucose to acidic by-products drastically reduced the survival of V. cholerae strains in vitro In the present study, zebrafish were fed 1% glucose and either inoculated with single V. cholerae or E. coli strains or coinfected with both V. cholerae and E. coli A significant decrease in classical biotype colonization was observed after glucose feeding due to acid production in the zebrafish intestine. El Tor colonization was unaffected by glucose alone. However, the El Tor strain exhibited significantly lower colonization of the zebrafish when either of the acid-producing E. coli strains was coinoculated in the presence of glucose. An E. coli sugar transport mutant had no effect on V. cholerae colonization even in presence of glucose. Glucose and E. coli produced a prophylactic effect on El Tor colonization in zebrafish when E. coli was inoculated before V. cholerae infection. Thus, the probiotic feeding of E. coli inhibits V. cholerae colonization in a natural host. This suggests that a similar inhibitory effect could be seen in cholera patients, especially if a glucose-based oral rehydration solution (ORS) is administered in combination with probiotic E. coli during cholera treatment.


Subject(s)
Cholera/prevention & control , Escherichia coli/metabolism , Glucose/metabolism , Intestines/microbiology , Vibrio cholerae O1/pathogenicity , Acids/metabolism , Animals , Antibiosis , Bacterial Load , Biological Transport , Cholera/microbiology , Escherichia coli/physiology , Probiotics/pharmacology , Zebrafish/microbiology
13.
J Vis Exp ; (137)2018 07 12.
Article in English | MEDLINE | ID: mdl-30059022

ABSTRACT

Vibrio cholerae is best known as the infectious agent that causes the human disease cholera. Outside the human host, V. cholerae primarily exists in the aquatic environment, where it interacts with a variety of higher aquatic species. Vertebrate fish are known to be an environmental host and are a potential V. cholerae reservoir in nature. Both V. cholerae and the teleost fish species Danio rerio, commonly known as zebrafish, originate from the Indian subcontinent, suggesting a long-standing interaction in aquatic environments. Zebrafish are an ideal model organism for studying many aspects of biology, including infectious diseases. Zebrafish can be easily and rapidly colonized by V. cholerae after exposure in water. Intestinal colonization by V. cholerae leads to the production of diarrhea and the excretion of replicated V. cholerae. These excreted bacteria can then go on to colonize new fish hosts. Here, we demonstrate how to assess V. cholerae-intestinal colonization in zebrafish and how to quantify V. cholerae-induced zebrafish diarrhea. The colonization model should be useful to researchers who are studying whether genes of interest may be important for host colonization and/or for environmental survival. The quantification of zebrafish diarrhea should be useful to researchers studying any intestinal pathogen who are interested in exploring zebrafish as a model system.


Subject(s)
Diarrhea/metabolism , Vibrio cholerae/genetics , Animals , Disease Models, Animal , Humans , Zebrafish
14.
PLoS Negl Trop Dis ; 11(7): e0005728, 2017 Jul.
Article in English | MEDLINE | ID: mdl-28727722

ABSTRACT

Few live attenuated vaccines protect against multiple serotypes of bacterial pathogen because host serotype-specific immune responses are limited to the serotype present in the vaccine strain. Here, immunization with a mutant of Shigella flexneri 2a protected guinea pigs against subsequent infection by S. dysenteriae type 1 and S. sonnei strains. This deletion mutant lacked the RNA-binding protein Hfq leading to increased expression of the type III secretion system via loss of regulation, resulting in attenuation of cell viability through repression of stress response sigma factors. Such increased antigen production and simultaneous attenuation were expected to elicit protective immunity against Shigella strains of heterologous serotypes. Thus, the vaccine potential of this mutant was tested in two guinea pig models of shigellosis. Animals vaccinated in the left eye showed fewer symptoms upon subsequent challenge via the right eye, and even survived subsequent intestinal challenge. In addition, oral vaccination effectively induced production of immunoglobulins without severe side effects, again protecting all animals against subsequent intestinal challenge with S. dysenteriae type 1 or S. sonnei strains. Antibodies against common virulence proteins and the O-antigen of S. flexneri 2a were detected by immunofluorescence microscopy. Reaction of antibodies with various strains, including enteroinvasive Escherichia coli, suggested that common virulence proteins induced protective immunity against a range of serotypes. Therefore, vaccination is expected to cover not only the most prevalent serotypes of S. sonnei and S. flexneri 2a, but also various Shigella strains, including S. dysenteriae type 1, which produces Shiga toxin.


Subject(s)
Cross Protection , Dysentery, Bacillary/prevention & control , Gene Deletion , Host Factor 1 Protein/deficiency , Shigella Vaccines/immunology , Shigella/genetics , Shigella/immunology , Animals , Disease Models, Animal , Dysentery, Bacillary/immunology , Dysentery, Bacillary/pathology , Guinea Pigs , Male , Microbial Viability , Serogroup , Shigella Vaccines/administration & dosage , Shigella Vaccines/genetics , Survival Analysis , Vaccines, Attenuated/administration & dosage , Vaccines, Attenuated/genetics , Vaccines, Attenuated/immunology , Virulence
15.
Jpn J Infect Dis ; 70(1): 111-114, 2017 Jan 24.
Article in English | MEDLINE | ID: mdl-27169940

ABSTRACT

Diabetes mellitus and diarrhea are becoming increasingly burdensome worldwide, particularly in developing countries such as India. Diabetic patients are susceptible to infection with pathogenic bacteria, particularly those causing invasive enteric infections. In this study, we observed changes in the pathophysiological features of mice with streptozotocin-induced hyperglycemia. In our experiments, both hyperglycemic and control mice were infected with pathogenic enteric bacteria-non-typhoidal Salmonella, Shigella flexneri, or Vibrio parahaemolyticus. Morbidity, mortality, and bacterial load were all higher in the diabetic mice than in the control mice, and the phagocytic and bactericidal activities of peritoneal macrophages isolated from hyperglycemic mice were lower than they were in the controls. We hypothesize that hyperglycemia leads to a downregulation of the innate immune response, which in turn increases vulnerability to enteric bacterial infection.


Subject(s)
Diabetes Mellitus, Experimental/complications , Diarrhea/epidemiology , Dysentery, Bacillary/epidemiology , Hyperglycemia/chemically induced , Hyperglycemia/complications , Salmonella Infections, Animal/epidemiology , Vibrio Infections/epidemiology , Animals , Antibiotics, Antineoplastic , Bacterial Load , Diarrhea/immunology , Disease Models, Animal , Disease Susceptibility , Dysentery, Bacillary/immunology , Macrophages, Peritoneal/immunology , Mice , Mice, Obese , Salmonella Infections, Animal/immunology , Streptozocin/administration & dosage , Survival Analysis , Vibrio Infections/immunology
16.
Vaccine ; 34(42): 5099-5106, 2016 09 30.
Article in English | MEDLINE | ID: mdl-27614779

ABSTRACT

Salmonella-induced gastroenteritis causes massive morbidity and mortality in both adults and children of developing countries. However, it is difficult to study the mode of infection and vaccine efficacy due to inadequacies of current animal models. For this reason, we have explored using zebrafish as an improved model for non-typhoidal Salmonella (NTS) infection, including Salmonella enterica Typhimurium, Salmonella enterica Enteritidis and Salmonella enterica Weltevreden. In this study, we found that after infection of zebrafish with NTS, severe diarrhea like symptoms were observed and NTS significantly colonized the zebrafish intestine without any manipulation of the normal intestinal microbiota of the fish. Furthermore, these strains can colonize for longer than 72h and induce severe inflammation in the intestine, which may induce fish death. We also found that infected fish can transmit the pathogen into naïve fish. Moreover, we have established that zebrafish is an excellent model for vaccine study. Successive triple bath vaccination with heat-killed single serotype S. Typhimurium and S. Enteritidis immunogen induced protective efficacy against a high dose (10(8)CFU/ml) of infection with these pathogens. This study provides a natural infection model for the study of NTS infection, transmission and vaccine efficacy.


Subject(s)
Disease Models, Animal , Immunogenicity, Vaccine , Salmonella Infections/prevention & control , Salmonella Vaccines/immunology , Salmonella typhimurium/pathogenicity , Zebrafish , Animals , Diarrhea/microbiology , Intestines/immunology , Intestines/microbiology , Intestines/pathology , Salmonella Infections/microbiology , Salmonella Infections/transmission , Salmonella Infections, Animal/microbiology , Salmonella typhimurium/immunology , Typhoid-Paratyphoid Vaccines/immunology
17.
Immunobiology ; 221(8): 918-26, 2016 08.
Article in English | MEDLINE | ID: mdl-27039316

ABSTRACT

Diarrhea is a very common health problem in both developing and developed countries. Among the major entero-invasive bacteria, Shigella, Salmonella and Campylobacter cause serious problems in different geographic regions. Recently we have shown immunogenicity and protective efficacy of heat killed multi-serotype Shigella immunogen in different animal models. In our present study, we have advanced our research by preparing a combination heat-killed immunogen of three different entero-invasive bacteria, Shigella, Salmonella and Campylobacter. After three doses on 0th, 14th and 28th day of oral immunization with tri-valent heat-killed (TVHK) immunogen in rabbit model, the immunogenicity was determined by differential count of white blood cells and immunoglobulin assay at various time points. During oral immunization differential count of lymphocytes increased where as polymorphonuclear leucocytes (PMN) count decreased. Serum IgG and IgA showed significant elevation during oral immunization and remained at a detectable value upto 120 days. Protection study was performed in both, in vitro and in vivo conditions, using bacteriocidal assay and rabbit ligated ileal loop model, respectively, which conferred protection against homologous bacteria. Moreover, immunoblot assay against whole cell lysate and lipopolysaccharide exhibited significant amount of antigen-specific immunoglobulins raised against three different bacteria which proved that proteins along with lipopolysaccharides played a pivotal role in immunogenicity and protective efficacy. This trivalent heat-killed immunogen could be a low-cost, simple, oral, non-living vaccine candidate for future use against invasive diarrhea.


Subject(s)
Campylobacter jejuni/immunology , Gram-Negative Bacterial Infections/immunology , Immunogenicity, Vaccine , Salmonella typhimurium/immunology , Shigella flexneri/immunology , Animals , Antibodies, Bacterial/immunology , Disease Models, Animal , Gram-Negative Bacterial Infections/prevention & control , Immunoglobulin A/immunology , Immunoglobulin G/immunology , Rabbits
18.
Immunobiology ; 221(7): 839-44, 2016 07.
Article in English | MEDLINE | ID: mdl-26858186

ABSTRACT

Gastroenteritis is a global burden; it's the major cause of morbidity and mortality both in adults and children of developing countries. Salmonella is one of the leading causes of bacteria-mediated gastroenteritis and due to its increasing multidrug antibiotic resistance; Salmonella-mediated gastroenteritis is difficult to control. Retinoic acid, the biologically active agent of vitamin A has an anti-inflammatory effect on experimental colitis. In this study we have shown All trans retinoic acid (ATRA) treatment down regulates Salmonella-mediated colitis in a murine model. Macroscopic signs of inflammation such as decrease in body weight and cecum weight, shorter length of proximal colon and pathological score of colitis were observed less in ATRA treated mice than in a vehicle control group. ATRA treatment not only reduced pro-inflammatory cytokine responses, such as TNF-α, IL-6, IL-1ß, IFN-γ and IL-17 production but also increased IL-10 response in the supernatant of intestinal tissue. Results also suggested that ATRA treatment enhances the number of FoxP3-expressing T regulatory cells in MLN and also decreases bacterial load in systemic organs. We concluded that ATRA treatment indeed reduces Salmonella Typhimurium-mediated gastroenteritis in mice, suggesting it could be an important part of an alternative therapeutic approach to combat the disease.


Subject(s)
Anti-Inflammatory Agents/therapeutic use , Gastroenteritis/drug therapy , Salmonella typhi/immunology , T-Lymphocytes, Regulatory/immunology , Tretinoin/therapeutic use , Animals , Cells, Cultured , Cytokines/blood , Disease Models, Animal , Forkhead Transcription Factors/metabolism , Gastroenteritis/immunology , Humans , Inflammation Mediators/blood , Male , Mice , Mice, Inbred BALB C
19.
Antimicrob Agents Chemother ; 59(12): 7471-6, 2015 12.
Article in English | MEDLINE | ID: mdl-26392502

ABSTRACT

The severe diarrheal disease cholera is endemic in over 50 countries. Current therapies for cholera patients involve oral and/or intravenous rehydration, often combined with the use of antibiotics to shorten the duration and intensity of the disease. However, as antibiotic resistance increases, treatment options will become limited. Linoleic acid has been shown to be a potent negative effector of V. cholerae virulence that acts on the major virulence transcription regulator protein, ToxT, to inhibit virulence gene expression. ToxT activates transcription of the two major virulence factors required for disease, cholera toxin (CT) and toxin-coregulated pilus (TCP). A conjugated form of linoleic acid (CLA) is currently sold over the counter as a dietary supplement and is generally recognized as safe by the U.S. Food and Drug Administration. This study examined whether CLA could be used as a new therapy to reduce CT production, which, in turn, would decrease disease duration and intensity in cholera patients. CLA could be used in place of traditional antibiotics and would be very unlikely to generate resistance, as it affects only virulence factor production and not bacterial growth or survival.


Subject(s)
Anti-Bacterial Agents/pharmacology , Bacterial Proteins/antagonists & inhibitors , Cholera Toxin/biosynthesis , Linoleic Acids, Conjugated/pharmacology , Transcription Factors/antagonists & inhibitors , Vibrio cholerae/drug effects , Animals , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Cholera/drug therapy , Cholera/physiopathology , Cholera Toxin/genetics , DNA, Bacterial/metabolism , Disease Models, Animal , Gene Expression Regulation, Bacterial , Rabbits , Transcription Factors/genetics , Transcription Factors/metabolism , Vibrio cholerae/metabolism , Vibrio cholerae/pathogenicity , Virulence/genetics , Virulence Factors/genetics , Virulence Factors/metabolism
20.
Immunobiology ; 220(11): 1275-83, 2015 Nov.
Article in English | MEDLINE | ID: mdl-26210044

ABSTRACT

Recently we have shown the homologous protective efficacy of heat killed multi-serotype Shigella (HKMS) immunogens in a guinea pig colitis model. In our present study, we have advanced our research by immunizing rabbits with a reduced number of oral doses and evaluating the host's adaptive immune responses. The duration of immunogenicity and subsequently protective efficacy was determined against wild type heterologous Shigella strains in a rabbit luminal model. After three successive oral immunizations with HKMS immunogens, serum and lymphocyte supernatant antibody titer against the heterologous shigellae were reciprocally increased and remained at an elevated level up to 180 days. Serogroup and serotype specific O-antigen of lipopolysaccharide and immunogenic proteins of heterologous challenge strains were detected by immunoblot assay. Up-regulation of IL-12p35, IFN-γ and IL-10 mRNA expression was detected in immunized rabbit peripheral blood mononuclear cells (PBMC) after stimulation with HKMS in vitro. HKMS-specific plasma cell response was confirmed by production of a relatively higher level of HKMS-specific IgG in immunized PBMC supernatant compared to control group. Furthermore, the immunized groups of rabbits exhibited complete protection against wild type heterologous shigellae challenge. Thus HKMS immunogens induced humoral and Th1-mediated adaptive immunity and provided complete protection in a rabbit model. These immunogens could be a broad spectrum non-living vaccine candidate for human use in the near future.


Subject(s)
Antigens, Bacterial/immunology , Dysentery, Bacillary/immunology , Dysentery, Bacillary/prevention & control , Immunity, Humoral , Immunization , Shigella/immunology , Adaptive Immunity , Administration, Oral , Animals , Antibodies, Bacterial/blood , Antibodies, Bacterial/immunology , Antibody Formation , Antigens, Bacterial/administration & dosage , Disease Models, Animal , Female , Immunity, Cellular , Immunoglobulin A, Secretory/blood , Immunoglobulin A, Secretory/immunology , Immunoglobulin Isotypes/blood , Immunoglobulin Isotypes/immunology , Male , Plasma Cells/immunology , Plasma Cells/metabolism , Rabbits , Serogroup , Shigella/classification
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