Comparison of 16S ribosomal RNA gene sequence analysis and conventional culture in the environmental survey of a hospital.

BACKGROUND

Nosocomial infection is one of the most common complications within health care facilities. Certain studies have reported outbreaks resulting from contaminated hospital environments. Although the identification of bacteria in the environment can readily be achieved using culturing methods, these methods detect live bacteria. Sequencing of the 16S ribosomal RNA (16S rRNA) gene is recognized to be effective for bacterial identification. In this study, we surveyed wards where drug-resistant bacteria had been isolated and compared conventional culture methods with 16S rRNA gene sequencing methods.

METHODS

Samples were collected using sterile swabs from two wards (northern and southern) at Gunma University Hospital contaminated by sp.. We extracted DNA directly from the swabs. Following extraction, the DNA was amplified using polymerase chain reaction (PCR). The PCR products were cloned using the plasmid vector. The plasmid DNA were sequenced, and identification were performed using database. 16S rRNA gene sequence analyses were compared conventional culture methods.

RESULTS

In the northern ward, sp. was detected from only two of 14 samples using the culture method. In contrast, 16S rRNA gene sequencing analysis detected sp. from seven of 14 samples. Drug-resistant sp. was isolated from bathrooms of the southern ward and was detected from four of seven samples using the culture method in comparison with six of seven samples by 16S rRNA gene sequencing analysis.

CONCLUSIONS

Molecular biological analysis showed a higher sensitivity to detect specific bacteria and detected a greater number of species than the culture method. Our results suggest that 16S rRNA gene sequencing analysis is useful to identify range of contamination which were not found in conventional culture method. When a nosocomial outbreak cannot be adequately controlled, molecular biological analysis may serve as a useful tool for environmental surveys in hospitals.