The spreading of multidrug resistance (MDR) strains in the hospital settings via contaminated surfaces have been increasingly reported where Gram-negative bacteria have been implicated in causing most nosocomial infections. This study aimed to determine the rate of contamination with multi-resistant gram-negative bacteria in the hospital environment. A cross-sectional study was conducted at Muhimbili National Hospital paediatric department, between July and August 2020. Non-repetitive surface swab samples were collected from predefined surfaces and medical device surfaces, and cultured on MacConkey agar with and without antibiotics. Isolates were identified using biochemical test and tested for antibiotic susceptibility using the Kirby-Bauer disk diffusion method. The rate of hospital contamination with Gram-negative bacteria across the Pediatrics units was 30%, with a high rate observed in oncology units (34.8%) and the malnutrition/diarrhoea ward (32.1%). Sink/washing basin had the highest frequency of bacterial contamination (74.2%). We observed a high rate of ESBL (32.5%), with Acinetobacter baumannii, Klebsiella pneumoniae, and E. coli being the predominant ESBL-producing Gram-negative bacteria, while carbapenemase-producing Gram-negative bacteria was detected at 22.8%. Highest resistance rates (63-100%) were observed against ceftriaxone and trimethoprim-sulfamethoxazole. Up to 51% of the Gram-negative bacteria showed resistant to meropenem. MDR strains were detected in 61.4% of Gram-negative bacteria isolated. In conclusion, we observed a high rate of MDR bacteria contaminating hospital surfaces. The higher rate of MDR calls for a need to strengthen infectious prevention control measures, including cleaning practices in the hospital environment, to reduce the risk of transmission of resistant strains to patients and healthcare workers.