Pneumocystis jirovecii is a pathogen that causes Pneumocystis pneumonia (PCP), an infection in human immunodeficiency virus/acquired human immunodeficiency virus (HIV/AIDS) and other immunocompromised patients. The rare reports of P. jirovecii pneumonia in sub-Saharan Africa are controversial; there is a high concern in areas with high HIV/AIDS seropositivity. Sputum samples were analyzed by microscopy and polymerase chain reaction (PCR) after toluidine blue O. TB smear negative samples were analyzed and compared. We proposed to determine the prevalence and importance of P. jirovecii at the Coast General Teaching Referral Hospital smear negative and retreatment cases. A total of 350 participants were enrolled to the study; 100 participants were able to expectorate sputum sample. Median age was 38 years old, (IQR: 18-78) of whom 62% 62 were males and 38% 38 were females. Most of the patents were labelled as TB smear negative or and retreatment cases. The results showed that patients with pulmonary tuberculosis were co-infected with fungi; P. jirovecii as opportunistic fungal organisms. A total of 62 males (62%) and a total of females 38 (38%) participated in the study which involved a total of 100 samples. Males’ seemingly high prevalence susceptibility to TB could have been attributed to their poor health seeking behaviors. A total of 29 samples (11.0%) were positive for Pneumocystis jirovecii. Microscopy detected P. jirovecii cysts in 27.17% of the patients. The Nested PCR detected 38% positive for P. jirovecii. The sensitivity and specificity of nested PCR as compared to TBO staining was 4.27% and 20.6% respectively. Nested PCR test detected additional 12 more patients than toluidine O staining technique showing high sensitivity, TBO had an acceptable sensitivity and very high specificity. Keywords:

Keywords: Pneumocystis jirovecii; Nested PCR; Pulmonary tuberculosis; Smear negative