Neonatal sepsis is a leading cause of morbidity and mortality in developing countries. This study aimed to assess the proportion of culture-confirmed sepsis, bacteriological pathogen profile, culture report turnaround times, antibiotic susceptibility patterns, and treatment outcomes of all with neonatal sepsis admitted in two tertiary care hospitals in Yangon, Myanmar, 2017–2019. This was a cross sectional study utilizing a standardized electronic database and paper-based records. Bacteriological profiles and associated factors were analyzed with descriptive statistics and Poisson Regression. Of those with suspected sepsis, 42% were bacteriologically confirmed and 74% of confirmed sepsis was resistant to at least first-line antibiotics. Neonates with late onset sepsis (LOS) (aPR: 1.2 (95% CI: 1.1–1.4, p = 0.008)) were more likely to have bacteriologically confirmed sepsis (45%) versus early onset sepsis (38%). Gram-negative organisms were most commonly isolated (63%), associated with multidrug-resistant organisms and with a high case-fatality rate (64%). These findings suggest that enhanced national guidance regarding infection control and prevention, antibiotic stewardship, and first-line antibiotic choices need to be provided. The link between LOS with infection and prevention protocols needs to be further explored in this context to decrease sepsis risk, neonatal mortality, and reduce further antimicrobial resistance.