Biocontrol of V. cholerae using bacteriophage

Cholera is a persistent threat to public health and is endemic in many countries. Of late, there is an emergence of antibiotic resistance in Vibrio cholerae and treatment is effective only if given early, thus there is a need for rapid and more effective treatment of cholera. One such treatment could be the use of bacteriophages. During infection, V. cholerae adheres to the surface of enterocytes but does not invade the host. They are therefore not protected from bacteriophage infection. The study presented in this dissertation evaluates the potential of bacteriophage being used as a biocontrol for V. cholerae. The aim of this project was isolation and in vitro characterisation of bacteriophages, selection of a candidate bacteriophage for biocontrol and its use in an infant rabbit model to assess its therapeutic efficacy. Seven phages were isolated in China, attempts to isolate in the UK environments were unsuccessful and five more phages were obtained from various sources. In total twelve phages were characterised for the one step growth curves following their host strain growth curves, their lytic spectra, electron microscopy, PFGE, restriction analysis and annotation of sequenced genomes. These in vitro characterisations could help in selecting the candidate bacteriophage for in vivo phage therapy trials. Amongst the phages studied, the phage Φ1 most nearly fitted the selection criteria. Its burst size was 43 ± 5.5; while the latent period was 12 ± 0.0 and it had broad host range as it could lyse 67 % of the total 91 strains; while its genome did not show any undesirable genes associated with lysogeny/antibacterial resistance or any cholera toxin genes upon genome annotation. In therapeutic trials using an infant rabbit model, Φ1 reduced the bacterial numbers significantly (4.7 log10 reduction with P < 0.001) and treated animals showed no symptoms of disease

Biocontrol of V. cholerae using bacteriophage

Cholera is a persistent threat to public health and is endemic in many countries. Of late, there is an emergence of antibiotic resistance in Vibrio cholerae and treatment is effective only if given early, thus there is a need for rapid and more effective treatment of cholera. One such treatment could be the use of bacteriophages. During infection, V. cholerae adheres to the surface of enterocytes but does not invade the host. They are therefore not protected from bacteriophage infection. The study presented in this dissertation evaluates the potential of bacteriophage being used as a biocontrol for V. cholerae. The aim of this project was isolation and in vitro characterisation of bacteriophages, selection of a candidate bacteriophage for biocontrol and its use in an infant rabbit model to assess its therapeutic efficacy. Seven phages were isolated in China, attempts to isolate in the UK environments were unsuccessful and five more phages were obtained from various sources. In total twelve phages were characterised for the one step growth curves following their host strain growth curves, their lytic spectra, electron microscopy, PFGE, restriction analysis and annotation of sequenced genomes. These in vitro characterisations could help in selecting the candidate bacteriophage for in vivo phage therapy trials. Amongst the phages studied, the phage Φ1 most nearly fitted the selection criteria. Its burst size was 43 ± 5.5; while the latent period was 12 ± 0.0 and it had broad host range as it could lyse 67 % of the total 91 strains; while its genome did not show any undesirable genes associated with lysogeny/antibacterial resistance or any cholera toxin genes upon genome annotation. In therapeutic trials using an infant rabbit model, Φ1 reduced the bacterial numbers significantly (4.7 log10 reduction with P < 0.001) and treated animals showed no symptoms of disease

Complete genome sequences of seven Vibrio cholerae phages isolated in China

The complete genome sequences of seven closely related Vibrio cholerae phages isolated from environmental sites in southeastern China are reported here. Phages QH, CJY, H1, H2, H3, J2, and J3 are members of the Podoviridae family and are highly similar to the previously sequenced Vibrio phages VP2, VP5, and phiVC8

Prevalence of microorganisms of hygienic interest in an organized abattoir in Mumbai, India

Background: The magnitude of food-borne illnesses in India is unknown because of lack of surveillance networks. Monitoring the prevalence of food-borne pathogens and indicators of contamination in primary production at abattoirs is imperative for creating a data bank and for effective control of such pathogens before they enter the food chain. Methodology: Microorganisms of hygienic interest were screened for their prevalence at Deonar Abattoir, Mumbai. Swab samples from 96 sheep/goat carcass sites were collected and analyzed for Staphylococcus spp., Bacillaceae, Clostridiaceae and Enterobacteriaceae. Results: Average Staphylococcus aureus and Staphylococcus epidermidis counts were 3.15 ± 0.18 and 3.46 ± 0.17 log10 CFU/cm2, respectively. Bacillus cereus, Bacillus subtilis and Clostridium spp. counts were 3.10 ± 0.08, 3.41 ± 0.19 and 0.76 ± 0.06 log10 CFU/cm2, respectively. The Escherichia coli count was 3.54 ± 0.06 and the Klebsiella aerogenes count was 3.22 ± 0.22 log10 CFU/cm2. Counts for Proteus vulgaris and Proteus mirabilis were 3.44 ± 0.14 log10 CFU/cm2 and 3.71 ± 0.12 log10 CFU/cm2, respectively. S. epidermidis had the highest percentage prevalence at (41.6%), followed by K. aerogenes (31.9%), B. subtilis (28.2%) and P. vulgaris (23.6%). Salmonella spp. were not isolated. Conclusions: The data demonstrate high prevalence and diversity of micro flora on carcasses in the primary Indian production facility, which might be attributed to either human handling or improper dressing especially during evisceration process. Appropriate training for personal and production hygiene is essential for workers in Indian meat production facilities. © 2010 Bhandare et al

Bacteriophage-induced lipopolysaccharide mutations in escherichia coli lead to hypersensitivity to food grade surfactant sodium dodecyl sulfate

Bacteriophages (phages) are considered as one of the most promising antibiotic alternatives in combatting bacterial infectious diseases. However, one concern of employing phage application is the emergence of bacteriophage-insensitive mutants (BIMs). Here, we isolated six BIMs from E. coli B in the presence of phage T4 and characterized them using genomic and phenotypic methods. Of all six BIMs, a six-amino acid deletion in glucosyltransferase WaaG likely conferred phage resistance by deactivating the addition of T4 receptor glucose to the lipopolysaccharide (LPS). This finding was further supported by the impaired phage adsorption to BIMs and glycosyl composition analysis which quantitatively confirmed the absence of glucose in the LPS of BIMs. Since LPSs actively maintain outer membrane (OM) permeability, phage-induced truncations of LPSs destabilized the OM and sensitized BIMs to various substrates, especially to the food-grade surfactant sodium dodecyl sulfate (SDS). This hypersensitivity to SDS was exploited to design a T4–SDS combination which successfully prevented the generation of BIMs and eliminated the inoculated bacteria. Collectively, phage-driven modifications of LPSs immunized BIMs from T4 predation but increased their susceptibilities as a fitness cost. The findings of this study suggest a novel strategy to enhance the effectiveness of phage-based food safety interventions

Reviving phage therapy for the treatment of cholera

Cholera remains a major risk in developing countries, particularly after natural or man-made disasters. Vibrio cholerae El Tor is the most important cause of these outbreaks, and is becoming increasingly resistant to antibiotics, so alternative therapies are urgently needed. In this study, a single bacteriophage, Phi_1, was used prophylactically and therapeutically to control cholera in an infant rabbit model. In both cases, phage-treated animals showed no clinical signs of disease, compared with 69% of untreated control animals. Bacterial counts in the intestines of phage-treated animals were reduced by up to 4 Log10 CFU/g. There was evidence of phage multiplication only in animals which received a V. cholerae challenge. No phage-resistant bacterial mutants were isolated from the animals, despite extensive searching. This is the first evidence that a single phage could be effective in the treatment of cholera, without detectable levels of resistance. Clinical trials in human patients should be considered

Genomic and Phenotypic Analysis of Salmonella enterica Bacteriophages Identifies Two Novel Phage Species

Bacteriophages (phages) are potential alternatives to chemical antimicrobials against pathogens of public health significance. Understanding the diversity and host specificity of phages is important for developing effective phage biocontrol approaches. Here, we assessed the host range, morphology, and genetic diversity of eight Salmonella enterica phages isolated from a wastewater treatment plant. The host range analysis revealed that six out of eight phages lysed more than 81% of the 43 Salmonella enterica isolates tested. The genomic sequences of all phages were determined. Whole-genome sequencing (WGS) data revealed that phage genome sizes ranged from 41 to 114 kb, with GC contents between 39.9 and 50.0%. Two of the phages SB13 and SB28 represent new species, Epseptimavirus SB13 and genera Macdonaldcampvirus, respectively, as designated by the International Committee for the Taxonomy of Viruses (ICTV) using genome-based taxonomic classification. One phage (SB18) belonged to the Myoviridae morphotype while the remaining phages belonged to the Siphoviridae morphotype. The gene content analyses showed that none of the phages possessed virulence, toxin, antibiotic resistance, type I–VI toxin–antitoxin modules, or lysogeny genes. Three (SB3, SB15, and SB18) out of the eight phages possessed tailspike proteins. Whole-genome-based phylogeny of the eight phages with their 113 homologs revealed three clusters A, B, and C and seven subclusters (A1, A2, A3, B1, B2, C1, and C2). While cluster C1 phages were predominantly isolated from animal sources, cluster B contained phages from both wastewater and animal sources. The broad host range of these phages highlights their potential use for controlling the presence of S. enterica in foods

The microbiological testing regulations for cannabis products in Canada: Are they enough from food safety and public health point of view?

Canada has recently legalised cannabis for its production, sale and usage in various ways viz. medicinal, recreational and edible. The Access to Cannabis for Medical Purposes Regulations (ACMPR) suggests that the microbial and chemical contaminants of fresh or dried marihuana or cannabis must be as per Schedule B of the Food and Drugs Act. It prescribes the testing for total aerobic count (TAC), yeast and mould count (YMC), Salmonella counts, Escherichia coli counts and total coliforms. The recent study revealed that there are 13 endophytic bacterial isolates and 30 endophytic fungal isolates which are harboured by cannabis. Though all phytopathogens may not infect humans, except perhaps immunocompromised individuals; it is obvious that present microbiological testing may not be enough to detect the plethora of microorganisms. Also, the testing for viruses like hepatitis A and hepatitis B are not provisioned by regulation though there are reports of their outbreaks and there is no provision for testing bacteria like Clostridium botulinum and Listeria monocytogenes. The present traditional culture-based testing methods may not be adequate in identifying food safety risks

Chronic Hematuria in a Camel

[Vet. World 2009; 2(4.000): 148-148