Oral colonisation by antimicrobial-resistant Gram-negative bacteria among long-term care facility residents: prevalence, risk factors, and molecular epidemiology

Abstract

Background: For residents of long-term care facilities (LTCFs), antimicrobial-resistant bacteria (ARB) are a risk factor, yet their oral colonisation, potentially leading to aspiration pneumonia, remains unclear. This study was undertaken to survey the prevalence, phenotypic characteristics, and molecular epidemiology of antimicrobial-resistant Gram-negative bacteria in the oral cavity of LTCF residents, and to analyse the risk factors for such carriers.

Methods: This study involved 98 residents of a LTCF in Hiroshima City, Japan, aged between 55 and 101 years. Oropharyngeal swabs were collected and plated on screening media for ESBL-producing and carbapenem-resistant bacteria; isolates were identified and tested for antibiotic susceptibility; biofilm formation was tested in vitro; identification of epidemic clones were pre-determined by PCR; resistance genes, sequence types, and whole-genome comparison of strains were conducted using draft genome sequences. Demographic data and clinical characterisations were collected and risk factors analysed.

Results: Fifty-four strains from 38% of the residents grew on screening media and comprised predominantly of Acinetobacter spp. (35%), Enterobacteriaceae spp. (22%), and Pseudomonas spp. (19%). All Escherichia coli isolates carried CTX-M-9 group and belonged to the phylogroup B2, O25:H4 ST131 fimH30 lineage. Six Acinetobacter baumannii isolates presented identical molecular characteristics and revealed more biofilm production than the others, strongly suggesting their clonal lineage. One Acinetobacter ursingii isolate displayed extensive resistance to various ß-lactams due to multiple acquired resistance genes. One Pseudomonas aeruginosa isolate showed exceptional resistance to all ß-lactams including carbapenems, aminoglycosides, and a new quinolone, showing a multidrug-resistant Pseudomonas aeruginosa (MDRP) phenotype and remarkable biofilm formation. Genome sequence analysis revealed this isolate was the bla_IMP-1-positive clone ST235 in Japan. Strokes (cerebral infarction or cerebral haemorrhage) and percutaneous endoscopic gastrostomy tubes were recognised as risk factors for oral colonisation by ARB in the LTCF residents.

Conclusions: ARB, as defined by growth on screening agar plates, which carried mobile resistance genes or elements or conferred high biofilm formation, were already prevalent in the oral cavity of LTCF residents. Health-care workers involved in oral care should be aware of antimicrobial resistance and pay special attention to transmission prevention and infection control measures to diminish ARB or mobile resistance elements dissemination in LTCFs.

Keywords: Acinetobacter baumannii; Acinetobacter ursingii; Antimicrobial resistant bacteria; Carbapenemase; Escherichia coli; Extended-spectrum ß-lactamases; Oral colonisation; Pseudomonas aeruginosa; Risk factor analysis.

Fig. 1

The percentage of antimicrobial-resistant bacteria isolated from oral pharyngeal samples

Fig. 2

Biofilm production by Acinetobacter spp. isolates (a) and Pseudomonas spp. isolates (b). Bacteria were grown in LB medium in the absence or presence of 1% glucose. Biofilm formation was measured at OD_590nm using a microtiter plate biofilm-formation assay and the crystal violet dye. The average and standard error of the mean for each sample are shown

Fig. 3

Comparison of genome sequences of Escherichia coli isolates (A), Acinetobacter baumannii isolates (B) and Pseudomonas aeruginosa isolates (C). (a) Comparison of genome sequences of four E. coli isolates. The unassembled sequences of E. coli isolates 43E, 73E, 77E, and 95E were aligned and compared to the complete genome sequence of E. coli K-12.MG1655 (accession number: NC_000913), E. coli EC958 (accession number: HG941718) and E. coli H105 (accession number: CP021454) strains. (b) Comparison of genome sequences of six A. baumannii isolates with the same POT code of 104–12-10 and A. ursingii isolate 56C. The unassembled sequences of these A. baumannii isolates were aligned and compared to the complete genome sequence of A. baumannii ATCC17987 strain (accession number: NZ_CP018664). (c) Comparison of genome sequences of P. aeruginosa NCGM2.S1 (type E), PA058447 (type F), 71E, and PAO1 strains. The unassembled sequences of P. aeruginosa isolate 71E and PA058447 strain were aligned and compared to the complete genome sequence of P. aeruginosa NCGM2.S1 (accession number: AP012280) and PAO1 (accession number: NC_002516) strains