Pertussis, or whooping cough, is an acute respiratory disease mainly affecting infants and children and is caused by Bordetella pertussis and Bordetella parapertussis. The aim of this study was to investigate the share of Bordetella species from potential whooping cough cases during 2005-2009. Eight hundred and two samples from suspected pertussis cases were collected, mainly from 2 provinces of Pakistan. Bacterial culture, identification, DNA extraction and routinely used polymerase chain reaction (PCR) methods using IS1001, IS1002 and IS481 were used to identify the Bordetella species. The results were unexpected, because all of the isolates collected from the different cities were identified as B. parapertussis (7.4%); B. pertussis was not isolated from any sample. However, PCR results indicated the presence of a small percentage (0.6%) of B. pertussis among the total cases studied. This study suggests that vaccines to protect against both B. pertussis and B. parapertussis should be considered.

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Although a whole-cell pertussis vaccine was introduced in Pakistan in 1980, little is known about the pertussis prevalence and circulating strains in Pakistan. The aim of this study was to analyze Bordetella parapertussis isolates circulating between 2005 and 2009 in Pakistan and to compare them with those found in other countries during different periods. A total of 59 (7.35%) B. parapertussis isolates from 802 subjects (median age, 3 years) from Pakistan, with pertussis-like symptoms were investigated. We carried out genotyping and DNA microarray analyses on these isolates and compared them with some international isolates of B. parapertussis. We found that the allele for pertactin (prn) found in strains studied from Pakistan was identical to the predominant type found in Europe. We showed that B. parapertussis isolates circulating in Pakistan are part of the same pulsed-field gel electrophoresis group to those circulating in Finland during the period of 1982-2007. Finally, microarray analysis confirmed that the isolates collected in Pakistan, were quite similar to international strains. Overall, these results confirm that B. parapertussis is extremely monomorphic. The high isolation rate of B. parapertussis (7.35%) compared to Bordetella pertussis (0.5%) may suggest that the whole-cell vaccine used in Pakistan is effective against B. pertussis (0.5% infections detected), but much less so against B. parapertussis.

Objective: To investigate the existence of genetically diverse vibrio cholerae variant strains in a rural Sindh district, and to find out the phylogenetic relationship of indigenous vibrio cholerae strains. Methods: The cross-sectional study was conducted from April 2014 to May 2016 in Khairpur, Pakistan, and comprised stool samples/rectal swabs collected from the main and city branches of the Khairpur Medical College Teaching Hospital, and the Pir Abdul Qadir Shah Jeelani Institute of Medical Sciences, Gambat. The samples were identified using standard microbiological, biochemical, serological techniques and polymerase chain reaction targeting the ompW gene. Whole genome sequencing and bioinformatics tool MUMmer 3.2.3 was used to compare indigenous and contemporary vibrio cholerae strains circulating in the province of Sindh. Neighbour-joining tree method was used to construct the phylogenic tree. Results: Of the 360 samples, 76(21.11%) were found positive for vibrio cholera strains. The species-specific ompW gene was amplified at the correct size of 588bp. The isolates belonged to serogroup Inaba, O1, biotype El Tor.  Unique sequences with same genomic coordinates showed that test strains were not similar to the reference sequence. Conserved genome sequences showed that 12 Out of 16 (75%) of the test strains were similar to each Other except the 3 strains isolated from Khairpur and 1 from Karachi. Multiple sequence alignment of the regions translated into protein showed that 13 out of 16 (81.25%) test strains were similar except 2 strains from Khairpur and 1 From Karachi. The phylogenetic tree showed that all isolated strains descended from the same ancestor along with the reference strain. Conclusions: Vibrio cholerae O1 El Tor variant existed in Khairpur. Key Words: Cholera, Vibrio cholerae, Variant, Khairpur, Phylogeny, MUMmer.

Background: Cholera is a diarrheal disease that is caused by the bacterium Vibrio cholerae that transmits through contaminated water, food, oral-fecal route and spread through poor sanitation system. This study aims to investigate the reports of clinical and drinking water samples during an epidemic in Pakistan. Methodology: This study was conducted in District Khairpur including 660 samples (360 from clinical and 300 from drinking water) from 2014-2016. All samples were enriched in Alkaline peptone water for 6 hours and then streaked on TCBS agar, incubated at 35ºC for 24 hours. The next day standard microbiological, biochemical, serological techniques were used for the identification of V. cholerae and further identification was performed using PCR. Results: Out of 360 clinical samples, 76(21.11%) were positive for V. cholerae. The species-specific Outer membrane protein W precursor (ompW) gene was amplified and shown at the correct size (588 bp) through agarose gel electrophoresis. The serotyping revealed that the isolates belonged to serogroup Inaba, O1. cholerae O1 strains shown typical El Tor phenotype similar to V. cholerae El Tor strain N16961 (PBR VP+) used as the reference strain in this study. All age groups were affected where the highest onset was seen among those aged 19 years and above. The culture of drinking water (n=300), observed negative on TCBS agar. Conclusion: As far as our knowledge, this is the first time when the presence of V. cholerae El Tor peak has been reported for causing cholera outbreaks in Khairpur Sindh, Pakistan. However, these findings can be used for further investigations and recognizing control measures.