Application of Bacteriophage in Nosocomial infection control, A new concept to disinfect Hospital environment

Abstract

Introduction: Health care associated infections (HAIs) are one of the important public health problems which might result in significant rise in mortality and morbidity, predominantly in immune compromised patients of Intensive Care Unit (ICU). A study showed that in the Asian region, risks of HAIs have been estimated to be 2-20 times higher than in developed countries up to 25% of hospitalized patients having acquired infection (Ling et al., 2015) and also a report shows that in United States, roughly 9.2 out of every 100 patients acquire a nosocomial infection, according to Healthline and among them also some HAIs is quite serious and potentially life-threatening. The common bacteria causing nosocomial infections include Pseudomonas aeruginosa, klebsiella pneumonia, Acinetobacter baumanii, Enterococci etc. (Khan, Baig, & Mehboob, 2017). Therefore, disinfectants like alcohols, hypochlorite, hydrogen peroxide, amyl phenol, glutaraldehyde etc. are widely used in hospital to kill these organisms which cause NAs but sometimes these chemical disinfectants might be hazardous, irritants and toxic to us and environment. Among many alternatives, bacteriophage mediated bio-control of the pathogenic bacteria is considering as one of the best options. Our objective of the study is to isolate, identify the pathogenic bacteria from the hospital environment and evaluate the efficacy of newly isolated lytic bacteriophage to minimize the bacterial load on hospital fomites. Methodology: The study time period of this research was 6 months and, in this study, we isolated phage against Pseudomonas aeruginosa bacteria collected from ICU of Teku hospital. Then most potent phage was characterized morphologically and Physiochemically. Burst size was obtained from one step growth curve. Intraspecific and interspecific host range was assessed by spot assay. During this research work, we used sterile fomite cloth pieces and marble tile as they are most potent sources were bacteria reside. Firstly, we contaminate the fomites and tiles with Pseudomonas aeruginosa bacteria then after we used Pseudomonas aeruginosa phage to decontaminate those cloths and tile. Result: Altogether 16 bacterial strain, 4 Pseudomonas aeruginosa and 12 Staphylococcus aureus were confirmed by Gram staining and Biochemical test and among them 2 Pseudomonas aeruginosa strains and 5 Staphylococcus aureus strains were found to be Carbapenem resistance and Multi Drug Resistant (MDR) respectively. Total six bacteriophages against Pseudomonas aeruginosa were isolated from different sewage samples. And one of the most potent phage P4 was characterized morphologically and physiochemically. Burst size of the phage was found to be 28 virions per bacterium. Protein profiling was done by SDS-PAGE where protein band between 20-250Kda were found and Phage P4 belongs to order Caudovirales and family Siphoviridae. Similarly, Phage was found to tolerate temperature of 70°C for 20 minutes, pH 3-12, exhibiting multiple host range as well. Decontamination assay was done on the sterile fabric cloth which showed that the P4 phage having MOI value 1 showed higher rate of decontamination with log reduction of 1 and p-value (0.002) i.e., significant. Further Comparison was done between P4 phage and normal disinfectant Phenol where we found that single phage has more disinfectant rate than phenol. xi

Similarly, time period up to which bacteriophage can show their effectivity as disinfectant was also done and it showed highest level of effectivity up to 6 hours at MOI 1 with the CFU/ml log reduction of 1. Conclusion: The result from the present thesis reveals several characteristics of the bacteriophage (P4), for instance, effective lytic capability, multiple host range, and stability in wide range of pH and temperature. Higher rate of decontamination with the log reduction of 1 CFU/ml and p-value (0.002) was showed by P4 phage having MOI value of 1. While doing comparison single phage have more disinfectant rate than normal disinfectant & P4 phage can show its highest level of effectivity up to 6 hrs at MOI 1. Keywords: Nosocomial Infection, disinfectant, Pseudomonas aeruginosa, bacteriophage, decontamination