Background: An estimated 350 million persons worldwide are chronically infected with Hepatitis B virus (HBV). These are at increased risk of developing liver cirrhosis and hepatocellular carcinoma. The aim of the study was to determine the prevalence of Hepatitis B and co-existing Hepatitis D and to investigate the pattern of lab parameters among patients positive for hepatitis B and/or D. Methods: This is an observational cross-sectional prospective study conducted from 2009 to 2014 in Lyari General Hospital Karachi Pakistan. A total of 10270 patients were registered in the liver clinic from June 2009 to December 2014 for hepatitis B vaccination. All patients who had visited the Liver clinic of Lyari General Hospital for hepatitis B vaccination were first screened for hepatitis B, and D viruses by HBsAg, and Anti HDV antibodies and anti HCV by ELISA method. Patients who had HBsAg negative were then vaccinated. If HBsAg was found to be positive the patients have been underwent through further tests for the confirmation of hepatitis B and D (Hepatitis B Virus DNA polymerase chain reaction (PCR), hepatitis B e-antigen (HBeAg), hepatitis Be antibody (HBeAb), and if anti HDV positive then HDV RNA PCR was performed). Patients found to have a positive HBVDNA PCR with positive or negative HBeAg were treated. Patients with positive HBeAb were seroconverted. Patients with a positive anti HDV antibody underwent an HDVRNA PCR and if positive were treated. Statistical analysis was performed using SPSS (IBM SPSS Statistics 24.0). Results: A total of 10270 patients were registered in the liver clinic. Overall 206 (2%) were found to be HBV positive, in which 26.2% had Hepatitis D co-infection. More than half (57.3%) were male and 70.4% were married with the mean age of 33.2 ± 10.9 years. About one-fourth of the total were HbeAg positive (25.2%) and hepatitis D antibody positive (26.2%), while 21% had HDV PCR positive and 6.3% had hepatitis C antibody positive. The mean hemoglobin was 12.2, white cell count 6052.3, viral load of HBV DNA was 44100856.97, SGPT was 63.47, albumin was 4.24 and mean INR was 1.09. Conclusion: Prevalence of Hepatitis B infection was 2% and Hepatitis D infection was present in 26.2% of hepatitis B infected patients.