Date of Award

11-10-2022

Degree Type

Thesis

Degree Name

MPhil in Biological and Biomedical Sciences

First Advisor

Dr. Junaid Iqbal

Second Advisor

Dr. Farah Naz Qamar

Third Advisor

Dr. Najeeha Talat Iqbal

Department

Biological and Biomedical Sciences

Abstract

Cholera is an intestinal infection caused by bacterium Vibrio cholerae which is Gram negative, motile, facultative anaerobic organism and is usually found in brackish and salt water. Every year 1.3 to 4 million people suffer from cholera. Not all strains of Vibrio cholerae are pathogenic, only some can cause infection in humans. Almost all strains which are pathogenic to humans express and secretes a potent enterotoxin called cholera toxin. The toxin enters the intestinal epithelial cell and in turn causessecretion of huge amount of water and minerals from these cells which clinically appears as rice water like diarrheal stool. The infection spread through fecal oral route due to the consumption of fecal contaminated food and water. In recent years, environmental surveillance has been slowly replacing traditional community and hospital & laboratory-based surveillance of disease especially gut diseases. For polio eradication program environmental surveillance of sewage played a vital role by informing about circulating polio virus in different countries specially after vaccination drives. This has helped the agencies targeting polio eradication effort to use their resources where the pathogenic strains of polio virus still exist including Pakistan. Recently, similar sewage surveillance strategies were also used to know about circulating strains and community burden of SARS-CoV-2 virus in different countries. Using environmental surveillance for bacterial disease like cholera could be very challenging as the causative agent Vibrio cholerae known to inhabit fresh and brackish water bodies. To understand which pathogenic and nonpathogenic strains are circulating in environmental and clinical samples, we have designed this study to where, we have isolated V. cholerae strains from different suspected cholera patients and sewers. These strains were subjected to two different PCR based typing assays to differentiate pathogenic strains from non-pathogenic strains. Lastly all these strains were subjected to whole genome sequencing using NextGen sequencing technique to know the genomic architecture of these strains. Our results showed that all the clinical strains isolated from symptomatic patients belong to the O1 serotype which is commonly reported from this part of the world. All these pathogenic strains also carried cholera toxin gene on an integrated prophage as well as other virulence markers. Because of time constraints we could not sequence a whole lot of environmental strains after molecular typing exercise but the sequenced, five sewage[1]isolated strains did not belong to O1 or O139 serotypes. All of them showed the absence of cholera toxin gene and absence of most of virulence factors otherwise detected in clinical strains. Surprisingly some of sewage strains also showed presence of antimicrobial genes against different classes of antibiotics which may be because of the results of horizontal gene transfer between same or different species within their natural habitat. We are hopeful that our and similar future studies will pave the way to identify markers which could be used to accurately identify Vibrio cholerae strains of human origin (pathogenic) from environmental samples.

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1

Last Page

133

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