Commenced in January 2007
Frequency: Monthly
Edition: International
Paper Count: 7

bacteriophage Related Abstracts

7 Viability and Sensitivity of SFN6B (Host-Specific Bacteriophage) towards Shigella Flexneri in Various Water Samples

Authors: Gimcheong Tan, Siewchuiang Sia

Abstract:

Bacteriophages are the most abundant and genetically diverse living entities on earth; they help in regulating and maintaining microbial diversity and balance in its natural ecosystem. In this study, the infectivity of SFN6B tailed phage was investigated in various water samples. Host bacteria (Shigella flexneri) were spiked in sterilized environmental and domestic water samples, followed by SFN6B treatment. Two incubation conditions were selected for this study, 37 oC and room temperature. S. flexneri and SFN6B viability were monitored hourly for consecutive 7 hours and extended viability study for consecutive 4 days. Absorbance of all bacteria spiked water samples were taken to monitor the bacteria count. Results showed reduction in the absorbance of the SFN6B treated water sample as compared to negative control, indicating reduction in bacterial count either due to negative growth or lysis by the lytic bacteriophage. Consistent with the result, SFN6B titer increases for first two days. However, prolong incubation of these cultures reaches equilibrium, between phage and bacteria. Temperature and water sample source also influence the interaction between S. flexneri and SFN6B. Stronger interaction was observed in 37oC as compared to room temperature, where higher bacteria count and phage titer increase were recorded. Availability of nutrient in water sample also plays a crucial role in the interaction between bacteria and phage. Higher nutrient level, such as lake and river waters were observed to give better infectivity and viability of both bacteria and phage as compared to tab water. It is believed that S. flexneri continue to remain viable and able to grow in the present of SFN6B bacteriophage, but the number was closely regulated by surrounding phages. This allows better understanding of the characteristics of SFN6B that could serve as the basis for future studies and applications.

Keywords: Infection, Microbial diversity, bacteriophage, Shigella flexneri

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6 Development of Two Phage Therapy-Based Strategies for the Treatment of American Foulbrood Disease Affecting Apis Mellifera capensis

Authors: Ridwaan N. Milase, Leonardo J. Van Zyl, Marla Trindade

Abstract:

American foulbrood (AFB) is the world’s most devastating honeybee disease that has drastically reduced the population of Apis mellifera capensis since 2009. The outbreak has jeopardized the South African bee keeping industry as well as the agricultural sector dependent on honeybees for honey production and pollination, leading to significant economic losses. AFB is caused by Paenibacillus larvae, a spore-forming, Gram positive facultative anaerobic and flagellated bacterium. The use of antibiotics within beehives has selected for resistant strains of P. larvae, while the current practice of burning spore contaminated beehives and equipment contributes to the economic losses in the honeybee-keeping industry. Therefore, phage therapy is proposed as a promising alternative to combat P. larvae strains affecting A. mellifera capensis. The genomes of two P. larvae strains isolated from infected combs in the Western Cape have been sequenced and annotated using bioinformatics tools. Genome analyses has revealed that these P. larvae strains are lysogens to more than 6 different prophages and possess different type of clustered regularly interspaced short palindromic repeat (CRISPRs) regions per strain. Active prophages from one of the two P. larvae strains were detected and identified using PCR. Electron microscopy was used to determine the family of the identified active prophages. Lytic bacteriophages that specifically target the two P. larvae strains were purified from sewage wastewater, beehive materials, and soil samples to investigate their potential development as anti-P. larvae agents. Another alternative treatment being investigated is the development of a prophage endolysin cocktail. Endolysin genes of the prophages have been targeted, cloned and expressed in Escherichia coli. The heterologously expressed endolysins have been purified and are currently being assessed for their lytic activity against P. larvae strains and other commensal microorganisms that compose the honeybee larvae microbiota. The study has shown that phage therapy and endolysins have a great potential as alternative control methods for AFB disease affecting A. mellifera capensis.

Keywords: Paenibacillus larvae, honeybee, bacteriophage, American foulbrood

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5 Application of Bacteriophages as Natural Antibiotics in Aquaculture

Authors: Jehee Lee, Chamilani Nikapitiya, Mahanama De Zoysa

Abstract:

Most of the bacterial diseases are associated with high mortalities in aquaculture species and causing huge economic losses. Different approaches have been taken to prevent or control of bacterial diseases including use of vaccines, probiotics, chemotherapy, water quality management, etc. Antibiotics are widely applying as chemotherapy to control bacterial diseases, however, it has been shown that frequent use of antibiotics is favored to develop multi-drug resistance bacteria. Therefore, phages and phage encoded lytic proteins are known to be one of the most promising alternatives for antibiotics to avoid the emergence of antibiotic-resistant bacteria. We isolated and characterized the two lytic phages, namely pAh-1 and pAs-1 against pathogenic Aeromonas hydrophila and Aeromonas salmonicida, respectively. Morphological characteristics were analyzed by Transmission electron microscopy (TEM) and host strain specificities were tested with Aeromonas and other closely related bacterial strains. TEM analysis revealed that both pAh-1 and pAsm-1 are composed of an icosahedral head and a segmented tail, and we suggest that, they are new members of Myoviridae family. Genome sizes of isolated phages were estimated by restriction enzyme digestion of genomic DNA using selected endonucleases followed by agarose gel electrophoresis. Estimated genome size of pAh-1 and pAs-1 were approximately 64 Kbp and 120 Kbp, respectively. Both pAh-1 and pAs-1 have shown narrow host specificity. Moreover, protective effects of phage therapy against fish pathogenic A. hydrophila were investigated in zebrafish model. The survival rate was 40% higher when zebrafish received intra-peritoneal injection (i.p.) of pAh-1 were simultaneously challenge A. hydrophila (2 x 106 CFU/fish) compared to that without phage treatment. Overall results suggest that both pAh-1 and pAs-1 can be used as a potential phage therapy to control Aeromonas infections in aquaculture.

Keywords: Antibiotic Resistance, bacteriophage, Aeromonas infections, bio-control, lytic phage

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4 Isolation and Molecular Characterization of Lytic Bacteriophage against Carbapenem Resistant Klebsiella pneumoniae

Authors: Guna Raj Dhungana, Roshan Nepal, Apshara Parajuli, Archana Maharjan, Shyam K. Mishra, Pramod Aryal, Rajani Malla

Abstract:

Introduction: Klebsiella pneumoniae is a well-known opportunistic human pathogen, primarily causing healthcare-associated infections. The global emergence of carbapenemase-producing K. pneumoniaeis a major public health burden, which is often extensively multidrug resistant.Thus, because of the difficulty to treat these ‘superbug’ and menace and some term as ‘apocalypse’ of post antibiotics era, an alternative approach to controlling this pathogen is prudent and one of the approaches is phage mediated control and/or treatment. Objective: In this study, we aimed to isolate novel bacteriophage against carbapenemase-producing K. pneumoniaeand characterize for potential use inphage therapy. Material and Methods: Twenty lytic phages were isolated from river water using double layer agar assay and purified. Biological features, physiochemical characters, burst size, host specificity and activity spectrum of phages were determined. One most potent phage: Phage TU_Kle10O was selected and characterized by electron microscopy. Whole genome sequences of the phage were analyzed for presence/absence of virulent factors, and other lysin genes. Results: Novel phage TU_Kle10O showed multiple host range within own genus and did not induce any BIM up to 5th generation of host’s life cycle. Electron microscopy confirmed that the phage was tailed and belonged to Caudovirales family. Next generation sequencing revealed its genome to be 166.2 Kb. bioinformatical analysis further confirmed that the phage genome ‘did not’ contain any ‘bacterial genes’ within phage genome, which ruled out the concern for transfer of virulent genes. Specific 'lysin’ enzyme was identified phages which could be used as 'antibiotics'. Conclusion: Extensively multidrug resistant bacteria like carbapenemase-producing K. pneumoniaecould be treated efficiently by phages.Absence of ‘virulent’ genes of bacterial origin and presence of lysin proteins within phage genome makes phages an excellent candidate for therapeutics.

Keywords: Phage Therapy, Klebsiella pneumoniae, bacteriophage, MDR, carbapenemase

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3 Preventive Effect of Three Kinds of Bacteriophages to Control Vibrio coralliilyticus Infection in Oyster Larvae

Authors: Jeong Woo Kang, Saekil Yun, Sib Sankar Giri, Jin Woo Jun, Hyoun Joong Kim, Sang Guen Kim, Sang Wha Kim, Se Jin Han, Se Chang Park, Cheng Chi

Abstract:

Vibrio corallilyticus is a well-known pathogen of coral. It is also infectious to a variety of shellfish species, including Pacific oyster (Crassostrea gigas) larvae. V. corallilyticus is remained to be a major constraint in marine bivalve aquaculture practice, especially in artificial seed production facility. Owing to the high mortality and contagious nature of the pathogen, large amount of antibiotics has been used for disease prevention and control. However, indiscriminate use of antibiotics may result in food and environmental pollution, and development of antibiotic resistant strains. Therefore, eco-friendly disease preventative measures are imperative for sustainable bivalve culture. The present investigation proposes the application of bacteriophage (phage) as an effective alternative method for controlling V. corallilyticus infection in marine bivalve hatcheries. Isolation of phages from sea water sample was carried out using drop or double layer agar methods. The host range, stability and morphology of the phage isolates were studied. In vivo phage efficacy to prevent V. corallilyticus infection in oyster larvae was also performed. The isolated phages, named pVco-5 and pVco-7 was classified as a podoviridae and pVco-14, was classified as a siphoviridae. Each phages were infective to four strains of seven V. corallilyticus strains tested. When oyster larvae were pre-treated with the phage before bacterial challenge, mortality of the treated oyster larvae was lower than that in the untreated control. This result suggests that each phages have the potential to be used as therapeutic agent for controlling V. corallilyticus infection in marine bivalve hatchery.

Keywords: Mortality, bacteriophage, Vibrio coralliilyticus, Oyster larvae

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2 Optimization the Multiplicity of Infection for Large Produce of Lytic Bacteriophage pAh6-C

Authors: Saekil Yun, Sib Sankar Giri, Jin Woo Jun, Hyoun Joong Kim, Sang Guen Kim, Sang Wha Kim, Jung Woo Kang, Se Jin Han, Se Chang Park

Abstract:

Emerging of the super bacteria, bacteriophages are considered to be as an alternative to antibiotics. As the demand of phage increased, economical and large production of phage is becoming one of the critical points. For the therapeutic use, what is important is to eradicate the pathogenic bacteria as fast as possible, so higher concentration of phages is generally needed for effective therapeutic function. On the contrary, for the maximum production, bacteria work as a phage producing factory. As a microbial cell factory, bacteria is needed to last longer producing the phages without eradication. Consequently, killing the bacteria fast has a negative effect on large production. In this study, Multiplicity of Infection (MOI) was manipulated based on initial bacterial inoculation and used phage pAh-6C which has therapeutic effect against Aeromonas hydrophila. 1, 5 and 10 percent of overnight bacterial culture was inoculated and each bacterial culture was co-cultured with the phage of which MOI of 0.01, 0.0001, and 0.000001 respectively. Simply changing the initial MOI as well as bacterial inoculation concentration has regulated the production quantity of the phage without any other changes to culture conditions. It is anticipated that this result can be used as a foundational data for mass production of lytic bacteriophages which can be used as the therapeutic bio-control agent.

Keywords: Optimization, bacteriophage, Aeromonas hydrophila, multiplicity of infection

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1 Characterization of Bacteriophage for Biocontrol of Pseudomonas syringae, Causative Agent of Canker in Prunus spp.

Authors: Mojgan Rabiey, Shyamali Roy, Billy Quilty, Ryan Creeth, George Sundin, Robert W. Jackson

Abstract:

Bacterial canker is a major disease of Prunus species such as cherry (Prunus avium). It is caused by Pseudomonas syringae species including P. syringae pv. syringae (Pss) and P. syringae pv. morsprunorum race 1 (Psm1) and race 2 (Psm2). Concerns over the environmental impact of, and developing resistance to, copper controls call for alternative approaches to disease management. One method of control could be achieved using naturally occurring bacteriophage (phage) infective to the bacterial pathogens. Phages were isolated from soil, leaf, and bark of cherry trees in five locations in the South East of England. The phages were assessed for their host range against strains of Pss, Psm1, and Psm2. The phages exhibited a differential ability to infect and lyse different Pss and Psm isolates as well as some other P. syringae pathovars. However, the phages were unable to infect beneficial bacteria such as Pseudomonas fluorescens. A subset of 18 of these phages were further characterised genetically (Random Amplification of Polymorphic DNA-PCR fingerprinting and sequencing) and using electron microscopy. The phages are tentatively identified as belonging to the order Caudovirales and the families Myoviridae, Podoviridae, and Siphoviridae, with genetic material being dsDNA. Future research will fully sequence the phage genomes. The efficacy of the phage, both individually and in cocktails, to reduce disease progression in vivo will be investigated to understand the potential for practical use of these phages as biocontrol agents.

Keywords: Pseudomonas, Biological Control, bacteriophage, bacterial cancker

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