Bacterial meningitis poses enormous healthcare challenges due to a high mortality, morbidity and sequelae. Neisseria (N.) meningitidis, Haemophilus (H.) influenzae, Streptococcus (S.) pneumoniae and S. agalactiae remain among the most prevalent infectious agents that cause bacterial meningitis in children. The objective of this study was the simultaneous detection of these pathogens in suspected cerebrospinal fluid (CSF) by using multiplex polymerase chain reaction (mPCR) and compare PCR results with standard diagnostics currently used in clinical practice. CSF specimens were obtained from 515 children (<5 years) clinically suspected of having acute bacterial meningitis. Based on bacterial culture, four isolates of salmonella sp and one Citrobacter freundii isolate were identified. The remaining 510 CSF specimens, having negative culture, were subjected to mPCR. Twenty-three (4.51%) CSF samples yielded a PCR positive signal. The pathogens identified were: S. pneumoniae (n=13), H. influenzae (n=7) and N. meningitidis (n=3). S. agalactiae was not detected. Using sequential multiplex PCR, serogrouping of S. pneumoniae revealed 3 different serotypes: serotype 19A (n=6), 19F (n=4) and serotype 23F (n=3). Only the serotype A was identified for the 3N. meningitidis isolates. Despite vaccination, S. pneumoniae remains a leading cause of pediatric invasive disease. Detecting causative organism remains the most critical aspect for management of children with suspected meningitis. PCR method is more sensitive and rapid than culture for detecting the infectious agents. Institution of PCR diagnostics is recommended for early and appropriate therapy.