Study subjects

A two stage (i.e. the discovery stage & the validation stage) case-control study have been designed. First Whole Exome Sequencing (WES) was performed to identify and suggest T2DM pathogenic variants in the target population. In the second stage the WES suggested pathogenic variants were confirmed for its association with T2DM using massARRAY genotyping. A total of 1000 individuals (healthy volunteers = 500 and T2DM cases = 500) of Pashtun ethnicity were recruited from seven districts (Peshawar, Mardan, Charsadda, Bannu, Kohat, Dir and swat) of Khyber Pakhtunkhwa, Pakistan for analysis. The cases were matched up with age, gender and ethnicity. Written informed consent was obtained from all the participants. For illiterate/un-educated patient’s understanding the informed consent form was read and explained in local Pashtu language and after patient’s agreement signed on his/her behalf by any of his/her relative/attendant. Patient’s detailed demographics and clinical parameter were noted on carefully designed proforma. The inclusion criteria for case group were (i) Diabetes diagnosed according to Internal Diabetes Federation (IDF) standard protocols, i.e., Fasting Blood Glucose (FBS) level greater than 126 mg/dL and random blood glucose (RBS) level greater than 200 mg/dL; (ii) Age between 30–80 years and (iii) Belonging from Pakistani Pashtun population. Exclusion criteria for case group were (i) Patients with chronic illness (presence of malignancies), recent severe infection (Hepatitis/Corona virus infection); and (ii) Patients of age not in range of 30–80 years. Whereas Controls were healthy volunteers from general population; age and gender wise matched with case group and fasting blood sugar level less than 100 mg/dL.