Phenotypic and molecular detection of Acinetobacter baumannii retrieved from the environmental surfaces

Abstract

Background: Hospital Associated Infections (HAI) pose a great deal of risk to those with impaired immune system, especially Intensive Care Unit (ICU) and Neonatal Intensive Care Unit (NICU) patients. Among the important nosocomial pathogens, Acinetobacter baumannii has been marked as the “red alert” pathogen due to its resistance to major antibiotics including the last-resort antibiotics like carbapenems. Acinetobacter baumannii causes sepsis, pneumonia, urinary tract infection, surgical site infections etc. Here we focused on the identification, characterization and investigation of antibiotic susceptibility of A.baumannii isolated from the environmental surface of the neonatal intensive care unit of a charitable child hospital from rural Bangladesh. Methodology: A total of 23 samples were collected from different neonatal wards in amies media and transported to Dhaka at 2-8°C. The samples were inoculated in the Chromagar Acinetobacter media. Phenotypic characterization was done via methods such as colony morphology, Gram’s staining, oxidase test, catalase test. Identification and antimicrobial susceptibility testing was done using the VITEK-2 instrument. Finally, 16s rRNA sequencing was performed (PCR, Gel Electrophoresis, Sanger Sequencing) and a Maximum Likelihood phylogenetic tree was formed using BioEdit, Chromas, Blast,Mega, IQ-tree. Result: A total of 23 environmental swab samples were collected. Bacterial growth was observed on 43% (10/23) environmental swab samples from which A.baumannii like colonies were observed on the Chromagar Acinetobacter media. VITEK-2 identification and antimicrobial susceptibility test on the suspected Acinetobacter spp. confirmed four of them to be A.baumannii (25% of 16) while confirming two (2/4) of the A.baumannii (50%) as MDR isolates. Both the MDR isolates were resistant against Cephalosporins, Carbapenems and Fluoroquinolones; intermediately resistant to Colistin and sensitive to Tigecycline and Cefoperazone/ sulbactam. The 16s rRNA sequencing supported the VITEK-2 identification of 75% (3/4) A.baumannii correctly while identifying 25% (1/4) as A.nosocomialis with percentage identity above 97%. The phylogenetic inference revealed that the A.nosocomialis didn’t share much similarity with its comparison group whereas the three A.baumannii isolates were found to be more closely related to one another (bootstrap value 99.7) than other global comparison strains, indicating that the same region of isolation might have a connection with sharing similarity as well as spreading of these isolates across the hospital.