An investigation on the prevalence of bovine tuberculosis (BTB) in randomly selected cattle (n= 160) was carried out in Hyderabad and Tando Allahyar districts. The animals were first screened through single intradermal tuberculin test (SITT) and both positive and negative reactors of SITT were further investigated through rapid test. Attempts were also made to isolate the Mycobacterium bovis organism from the milk and nasal secretions of cattle using the Lowenstein-Jensen media. An overall prevalence of 34.38% was recorded by rapid test that was higher (P< 0.001) than SITT (3.13%) and culture test (2.50%). A somewhat higher prevalence was recorded in Hyderabad district (SITT 3.75%, Rapid 36%, culture 3.75%) as compared to Tando Allahyar district (SITT 2.5%, Rapid 32%, culture 1.25%). In Hyderabad district, rapid test showed a significantly higher (P< 0.05) prevalence in male than females. Similarly, a higher (P< 0.05) prevalence was observed for 5-8 years age than > 8 years, in non-pregnant animals than pregnant and in 2-4 and >4 parities than 1 parity. However, in Tando Allahyar district, rapid test declared a significantly higher prevalence (P< 0.05) in female than male cattle, in 5-8 years age than > 8 years, in non-pregnant than pregnant, having 2-4 liters/d milk production than more than 4 liters/d, and in late stage of lactation than early or mid-stage. The prevalence in Tando Allahyar district revealed by culture technique showed a significantly (P< 0.05) higher percentage in >4 parity than 1 parity or 2-4 parity, having 4-8 liters/d milk production than those having 2-4 or >8 liters/d, and in late stage of lactation than early or mid-stage. In brief, BTB is prevailing in both Hyderabad and Tando Allahyar districts; however it is relatively higher in Hyderabad district than Tando Allahyar district. Infected animals shed more M. bovis in nasal secretions (3/4; 75%) as compared to milk (1/4; 25%). Rapid test showed the highest prevalence as compared to other techniques hence could be regarded as the most sensitive technique for BTB.