Sepsis is considered a race to death between pathogens and host immune system, associated with a high mortality worldwide. Until now, blood cultures (BCs) are still the “gold standard" for identification of pathogens causing sepsis. Its major drawback is long turn around time (TAT) of 24–48 h up to several days, which
is not compatible with the need for early sepsis diagnosis. Molecular assays
as matrix-assisted laser- desorption ionization time-of-flight mass spectrometry(MALDI-TOF MS) reduce the TAT. Herein, the main aim of the study is to develop rapid and highly sensitive molecular proteomic non-nucleic acid assay for identification (ID) of the most common bacteria causing sepsis and pyogenic infections in human and animals. Eighty clinical samples, including 50 animal pus samples and 30 neonatal whole blood samples were subjected for conventional and molecular identification using VITEK-MS. About 70 % of samples were BC positive, including 33/50(66 %) of animal pus samples and 23/30 (76.7 %) of neonatal blood samples. The identified bacterial species were only three conventionally versus twelve using VITEK-MS, with variable confidence value (C.V) that was 99.9 % for 55/58 (94.8 %) isolates, 50 % for 2/58 (3.4 %) isolates and 26.5 % with 1/58 (1.74 %) isolate. The species-level identification for Streptococci was more challenging than for Staphylococci and Enterococci with only 50 % C.V for S. dysagalactiaesubsp.dysagalactiae. Culture provided concordant species- and genus-level identification with VITEK-MS for 24/58(41.3 %) and 34/58(58.6 %) of isolates, respectively. The overall concordance was 100 % for all isolates except P. aeruginosawas 87.5 %.