The present investigationaimed to evaluate the prevalence of some microorganisms of public health importance (Escherichia coli, Salmonella, Staphylococcus aureus and Listeria monocytogenes) in fresh beef and study the growth and survival behavior of these pathogens when subjected to different types of cooking. Our findings showed that out of 100 fresh beef samples analyzed for microbial quality 90 (90%) were contaminated with different kinds of microorganisms namely E.coli (48%), Salmonella spp. (18%), Staphylococcous spp. (16%) and Listeria spp. (8%). The most E.coli isolated was E.coli O₁₁₁: H₄ (43.75%) followed by E.coli O₁₂₆:H₁₁ (27.08%), E.coli O₁₂₈:H₁₁ (22.92%) and E.coli O₁₅₇:H₇ (6.25%).Three species of Salmonella were isolated. The highest prevalence of them was S.typhimurium (44.4%), while S.enetertiidis and S. anatum rank as a second (27.8%) for each. Staphylococcous spp was isolated from (16%) of tested samples whereas (75%) of them recorded as Staph. aureus and (18.75%) recorded as Staph. epidermis, while (6.25%) recorded as Staph. gallinarum, all of them were coagulase–positive. Fifty percent of isolated Listeria spp. were characterized as L. monocytogenes, (25%) as L.innocua while the presence of L.welshimeri and L.invanovii was (12.5%) for each. Thermal inactivation of inoculated E.coli O₁₅₇:H₇, S.enetertiidis, Staph. aureus and L.monocytogenes inoculated in fresh beef were evaluated by boiling, frying and roasting treatments. At internal temperature of 65°C using boiling, the log cycles reduction were 1.3, 2.1, 2.2 and 2.2 for aforementioned microorganisms, respectively. By frying the reduction values were 1.5, 2.1, 2.3 and 2.1, respectively. The corresponding values by roasting were 2.6, 2, 2.3 and 1.4, respectively. E.coli O₁₅₇:H₇ couldn't be detected at internal temperature of 80, 83 and 74°C by boiling, frying and roasting, respectively. Both S. enetertiidis and Staph aureus couldn't be detected at internal temperature of 80, 80 ad 71°C by the treatments, respectively, while L. monocytogenes couldn't be detected at internal temperature of 80, 80 and 78°C respectively. The sensitivity of the isolated pathogens to heat inactivation was measured by assessing the D-values. These values were calculated from the survival curves. For E.coli O₁₅₇:H₇, they were 1.1, 1.1 and 1.2 minutes by boiling, frying and roasting treatments, respectively. Those recorded for S. enteritidis were 1.1, 1.0 and 1.2 minutes, respectively. In case of Staph aureus they were 1.1, 0.9 and 1.1 minutes, respectively while in case of L. monocytogenes the recorded values were 1.1, 0.8 and 1.1, minutes, respectively. Cooking fresh beef by boiling resulted in cooking weight loss (CWL) ranged from 8.1 to 17.47% according to time of exposure. By roasting the CWL ranged from 4.77 to 23.5% while by frying it was 15 to 23.53%. The increase in the pH value was directly proportional to time of exposure to boiling but not clearly demarked by other cooking methods. This study cleared that fresh beef from fresh beef shops at Assiut City, Egypt can acts as a source of major human pathogens. For safe consumption, such meat must cooked to internal temperature of 83°C when using traditional cooking methods. The D-values recorded in this study may be helpful guide for thermal processing of meat.