Molecular and clinical epidemiology of carbapenem resistant Acinetobacter baumannii, Pseudomonas aeruginosa and Enterobacterales in Fiji: a multicentre prospective observational study

Abstract

Background: Carbapenem resistant organisms (CROs) such as Acinetobacter baumannii (CRAb), Pseudomonas aeruginosa (CRPa), Escherichia coli (CREc), and Klebsiella pneumoniae (CRKp) have been identified by the World Health Organization (WHO) as global priority pathogens. The dissemination of these pathogens and clonal outbreaks within healthcare facilities are of serious concern, particularly in regions with limited resources. In Fiji, where healthcare services are primarily provided by public hospitals, understanding the extent and nature of this problem is essential for the development of effective patient management, prevention interventions and control strategies.

Methods: CROs isolated from 211 (77.3%) non-sterile (urinary catheters, urine, sputum, wound swab, and endotracheal tube) and 62 (22.7%) normally sterile (blood, cerebrospinal fluid, intravascular catheter, and aspirates) body sites of 272 patients treated at the three major hospitals in Fiji, the Colonial War Memorial Hospital (CWMH), Lautoka Hospital (LTKH), and Labasa Hospital (LBSH), and outer peripheral health centres around Fiji, were analysed. Clinical and demographic patient data such as age, sex, admission diagnosis, admission and discharge dates, patient outcomes, date of death, start and end date of meropenem and colistin treatment were reviewed. These CRO isolates comprised A. baumannii, P. aeruginosa, E. coli, and K. pneumoniae, that were prospectively collected at the microbiology laboratory of CWMH and LBSH from January 2020 through August 2021 and at the LTKH from January 2020 to December 2021. In addition, 10 retrospectively stored CRPa isolates collected from patients at the CWMH from January through December 2019, were also included in the study. All isolates were characterised using mass spectrometry, antimicrobial susceptibility testing, and whole genome sequencing. Phylogenetic relationships among the CROs were assessed through core genome single nucleotide polymorphism (SNP) analysis. The CRAb isolates were also compared to the CRAb isolates from CWMH isolated in 2016/2017 and 2019, along with CRAb isolates obtained from Fijian patients admitted to New Zealand hospitals in 2020 and 2021 from our retrospective study.

Findings: Of 272 patients, 140 (51.5%) were male, the median (range) age of patients was 45 (<1-89) years, 161 (59.2%) were I-Taukei, 104 (38.2%) Fijians of Indian descent, and 7 (2.6%) were from other ethnic backgrounds. 234 (86.0%) of these 272 patients, had their first positive CRO sample collected ≥72 h following admission and the remaining 38 (14.0%) were isolated within 72 h following admission. Of the 273 CROs, 146 (53.5%) were collected at the CWMH, 66 (24.2%) LTKH, and 61 (22.3%) LBSH, while 62 (22.7%) were isolated from normally sterile sites and 211 (77.3%) from sites that are not sterile. Of 273 isolates, 131 (48.0%) were CRAb, 90 (33.0%) CRPa, 46 (16.8%) CREc, and 6 (2.2%) CRKp. Of 131 CRAb, 108 (82.4%) were ST2, with three distinct clones, all encoding bla _OXA-23 and bla _{OXA - 66}, while clone 3 also encoded bla _NDM-1; bla _OXA-23 was associated with two copies of ISAba1 insertion element, forming the composite transposon Tn2006. The first two CRAb ST2 clones were genetically linked to those isolated at CMWH 2016 through 2019, while the third was genetically linked to isolates from Fijian patients admitted to New Zealand hospitals in 2020 and 2021. Of CRPa, 65 (72.2%) were ST773 and carried β-lactamase genes bla _NDM-1, bla _OXA-50, and bla _OXA-395. Of 10 retrospective CRPa isolates, all belonged to CRPa ST773 and carried bla _NDM-1, bla _OXA-50, and bla _OXA-395. Of 46 CREc, 44 (95.7%) were ST410 and encoded bla _NDM-7 on an IncX3 plasmid. Of 6 CRKp, 4 (66.7%) were ST16 and carried bla _NDM-5 on an IncX3 plasmid. Other sequence types of CRPa (ST9, ST357, ST654, ST664), CRAb (ST25, ST374, ST499), CREc (ST167), and CRKp (ST45, ST336) were also detected. Of those receiving meropenem treatment in the prospective study, 30 (57.7%) received it inappropriately. Of 272 patients, 65 (23.9%) died within the 30 days after first positive CRO isolation.

Interpretation: We identified nosocomial transmission of distinct clones of CRAb ST2, CRPa ST773, CREc ST410, and CRKp ST16 within and between the three major hospitals in Fiji. Moreover, community onset infections associated with CRPa, CREc, and CRAb were also detected. Of note, cross-border transmission of CRAb ST2 clone 3 strain between Fiji and New Zealand was also detected. These clones encoded an array of carbapenem resistance genes associated with mobile genetic elements, including plasmids, transposons, and integrative and conjugative elements, signifying their potential for increased mobility, further acquisition of resistance genes, and spread. Inappropriate use of meropenem was common. Of note, the majority of patients who died had acquired CRO during their hospital stay. These findings highlight the need for stringent IPC strategies focusing on catheter and ventilator management, meticulous wound care, rigorous sepsis control, consistent hand hygiene, effective use of disinfectants, and thorough sanitisation of both hospital environments and medical equipment in the three major hospitals in Fiji. Additionally, diligent surveillance of AMR and robust antimicrobial stewardship are crucial for effectively managing nosocomial infections.

Funding: This project was funded by the Otago Medical School Foundations Trust (Dean's Bequest Fund) and a Fiji National University seed grant. The funders of the study had no role in the study design, data collection, data analysis, data interpretation, or writing of the report.

Keywords: Acinetobacter baumannii; Carbapenem resistant; Escherichia coli; Klebsiella pneumoniae; Pseudomonas aeruginosa.

Fig. 1

Phylogenetic analysis, transmission networks, and mobile genetic elements of carbapenem resistant A. baumannii ST2 isolates from Fiji (2020–2021). a) Core genome SNP phylogeny of carbapenem resistant A. baumannii ST2 isolates from Fiji (2020–2021). Phylogenetic tree inferred from core genome SNPs of 108 CRAb ST2 isolates. The core genome was generated by aligning at least 92% of the available sequence data for all isolates. The core SNP density, representing the average number of SNPs per base pair in the core genome, was 163 SNPs across 4,062,284 base pairs in the reference genome. The reference genome used was from CRAb ST2 (SAMN37733145) isolated from CWMH in 2019. The tree was rooted using the earliest isolate (reference) from CWMH. Isolation dates, patient admission locations, and AMR phenotypic and genotypic profiles are annotated on the tree. The scale bar on the tree represents the frequency of mutations per site, providing a measure of genetic distance between isolates. Clusters were determined by fastbaps. b) Putative transmission trees of CRAb ST2 clones 1–3 among CWMH, LTKH, and LBSH, 2020–2021. Transmission trees showing inter-hospital transmission directions of CRAb ST2 clones 1 (i), 2 (ii), and 3 (iii) based on sample SNP distance and collection timeline. Numbers in red signify the SNP distance. Transmission tree generated and visualised using GraphSNP. The black arrows represent the earliest positive isolate. c) Comparative distribution of mobile genetic elements and AMR genes in CRAb ST2 isolates (clone 1–3) from Fiji, 2020–2021. The figure presents a circular depiction of the completed genomes of Fiji's CRAb ST2 clone 1 (FJ567_LT), clone 2 (FJ437_LB), and clone 3 (FJ104_CW) isolates, alongside the dispersion of MGEs carrying AMR genes, visualised using BRIG. The innermost circle corresponds to CRAb ST2 clone 1, the middle circle to CRAb ST2 clone 2, and the outermost circle to CRAb ST2 clone 3. Arrows indicate the existence of an MGE at that particular genome location. The detailed representation of MGEs can be found in the boxes (a–n). AMR abbreviation: aph, aminoglycoside phosphotransferase; arm, aminoglycoside resistance methylase; ant, aminoglycoside nucleotidyltransferase; aadA, aminoglycoside adenylyltransferase; abaQ, A. baumannii quinolone resistance transporter; ade, adenine deaminase; ampC, AmpC beta-lactamase; amvA, Acinetobacter baumannii AmvA efflux pump; ble-MBL, bleomycin resistance gene; bla_NDM, New Delhi metallo-beta-lactamase; bla_OXA, OXA-type beta-lactamase; bla_PER, PER beta-lactamases; cat, chloramphenicol acetyltransferase; gyr, DNA gyrase; par, Type IV topoisomerase; qnrS1, quinolone resistance gene; qacE, quaternary ammonium compounds; MFS, major facilitator superfamily; mph, macrolide phosphotransferase; msr(E), macrolide resistance gene; RND, resistance nodulation-division; sul, sulfonamide resistance gene.

Fig. 1

Phylogenetic analysis, transmission networks, and mobile genetic elements of carbapenem resistant A. baumannii ST2 isolates from Fiji (2020–2021). a) Core genome SNP phylogeny of carbapenem resistant A. baumannii ST2 isolates from Fiji (2020–2021). Phylogenetic tree inferred from core genome SNPs of 108 CRAb ST2 isolates. The core genome was generated by aligning at least 92% of the available sequence data for all isolates. The core SNP density, representing the average number of SNPs per base pair in the core genome, was 163 SNPs across 4,062,284 base pairs in the reference genome. The reference genome used was from CRAb ST2 (SAMN37733145) isolated from CWMH in 2019. The tree was rooted using the earliest isolate (reference) from CWMH. Isolation dates, patient admission locations, and AMR phenotypic and genotypic profiles are annotated on the tree. The scale bar on the tree represents the frequency of mutations per site, providing a measure of genetic distance between isolates. Clusters were determined by fastbaps. b) Putative transmission trees of CRAb ST2 clones 1–3 among CWMH, LTKH, and LBSH, 2020–2021. Transmission trees showing inter-hospital transmission directions of CRAb ST2 clones 1 (i), 2 (ii), and 3 (iii) based on sample SNP distance and collection timeline. Numbers in red signify the SNP distance. Transmission tree generated and visualised using GraphSNP. The black arrows represent the earliest positive isolate. c) Comparative distribution of mobile genetic elements and AMR genes in CRAb ST2 isolates (clone 1–3) from Fiji, 2020–2021. The figure presents a circular depiction of the completed genomes of Fiji's CRAb ST2 clone 1 (FJ567_LT), clone 2 (FJ437_LB), and clone 3 (FJ104_CW) isolates, alongside the dispersion of MGEs carrying AMR genes, visualised using BRIG. The innermost circle corresponds to CRAb ST2 clone 1, the middle circle to CRAb ST2 clone 2, and the outermost circle to CRAb ST2 clone 3. Arrows indicate the existence of an MGE at that particular genome location. The detailed representation of MGEs can be found in the boxes (a–n). AMR abbreviation: aph, aminoglycoside phosphotransferase; arm, aminoglycoside resistance methylase; ant, aminoglycoside nucleotidyltransferase; aadA, aminoglycoside adenylyltransferase; abaQ, A. baumannii quinolone resistance transporter; ade, adenine deaminase; ampC, AmpC beta-lactamase; amvA, Acinetobacter baumannii AmvA efflux pump; ble-MBL, bleomycin resistance gene; bla_NDM, New Delhi metallo-beta-lactamase; bla_OXA, OXA-type beta-lactamase; bla_PER, PER beta-lactamases; cat, chloramphenicol acetyltransferase; gyr, DNA gyrase; par, Type IV topoisomerase; qnrS1, quinolone resistance gene; qacE, quaternary ammonium compounds; MFS, major facilitator superfamily; mph, macrolide phosphotransferase; msr(E), macrolide resistance gene; RND, resistance nodulation-division; sul, sulfonamide resistance gene.

Fig. 2

Phylogenetic analysis, transmission networks, and mobile genetic elements of carbapenem resistant P. aeruginosa ST773 isolates from Fiji, 2020–2021. a) Core genome SNP phylogeny of carbapenem resistant P. aeruginosa ST773 isolates from Fiji (2020–2021). Phylogenetic tree inferred from core genome SNPs of 65 CRPa ST773 prospective isolates. The core genome was generated by aligning at least 97% of the available sequence data for all isolates. The core SNP density, representing the average number of SNPs per base pair in the core genome, was 84 SNPs across 6,825,462 base pairs in the reference genome. The reference genome used was CRPa ST773 (FJ453_LB), which was isolated from LBSH in 2020. The tree was rooted using the earliest isolate from Fiji (FJ03_CW) isolated from CWMH on 8th January 2020. Isolation dates, patient admission locations, and AMR phenotypic and genotypic profiles are annotated on the tree. The scale bar on the tree represents the frequency of mutations per site, providing a measure of genetic distance between isolates. b) Putative transmission tree of CRPa ST773 among CWMH, LTKH and LBSH, 2020–2021. A transmission tree showing inter-hospital transmission directions of CRPa ST773 based on sample SNP distance and collection timeline. Red numbers between nodes signify the SNP distance. Transmission tree generated and visualised using GraphSNP. The black arrow represents the earliest positive isolate, FJ03_CW (isolated 8th January 2020). c) Distribution of MGE on the representative CRPa ST773 (FJ453_LB) genome. AMR abbreviation: aph, aminoglycoside phosphotransferase; aadA, aminoglycoside adenylyltransferase; bcr, bicyclomycin; bla_NDM, New Delhi metallo-beta-lactamase; bla_OXA, OXA-type beta-lactamase; floR, florfenicol resistance gene; fos, fosfomycin resistance gene; gyr, DNA gyrase; MFS, major facilitator superfamily; mex, multidrug efflux system; muxABC-opmB, an aminocourmarin-specific efflux pump; opr and opm, outer membrane factors that form part of efflux pump complexes; parR/S, a two-component regulatory system; qacE, quaternary ammonium compounds; qnrv, quinolone resistance gene; rmtB, 16S rRNA methyltransferase; RND, Resistance-Nodulation-Division; sul, sulfonamide resistance gene; tet, tetracycline resistane gene; triABC-opmH, triclosan efflux pump.

Fig. 2

Phylogenetic analysis, transmission networks, and mobile genetic elements of carbapenem resistant P. aeruginosa ST773 isolates from Fiji, 2020–2021. a) Core genome SNP phylogeny of carbapenem resistant P. aeruginosa ST773 isolates from Fiji (2020–2021). Phylogenetic tree inferred from core genome SNPs of 65 CRPa ST773 prospective isolates. The core genome was generated by aligning at least 97% of the available sequence data for all isolates. The core SNP density, representing the average number of SNPs per base pair in the core genome, was 84 SNPs across 6,825,462 base pairs in the reference genome. The reference genome used was CRPa ST773 (FJ453_LB), which was isolated from LBSH in 2020. The tree was rooted using the earliest isolate from Fiji (FJ03_CW) isolated from CWMH on 8th January 2020. Isolation dates, patient admission locations, and AMR phenotypic and genotypic profiles are annotated on the tree. The scale bar on the tree represents the frequency of mutations per site, providing a measure of genetic distance between isolates. b) Putative transmission tree of CRPa ST773 among CWMH, LTKH and LBSH, 2020–2021. A transmission tree showing inter-hospital transmission directions of CRPa ST773 based on sample SNP distance and collection timeline. Red numbers between nodes signify the SNP distance. Transmission tree generated and visualised using GraphSNP. The black arrow represents the earliest positive isolate, FJ03_CW (isolated 8th January 2020). c) Distribution of MGE on the representative CRPa ST773 (FJ453_LB) genome. AMR abbreviation: aph, aminoglycoside phosphotransferase; aadA, aminoglycoside adenylyltransferase; bcr, bicyclomycin; bla_NDM, New Delhi metallo-beta-lactamase; bla_OXA, OXA-type beta-lactamase; floR, florfenicol resistance gene; fos, fosfomycin resistance gene; gyr, DNA gyrase; MFS, major facilitator superfamily; mex, multidrug efflux system; muxABC-opmB, an aminocourmarin-specific efflux pump; opr and opm, outer membrane factors that form part of efflux pump complexes; parR/S, a two-component regulatory system; qacE, quaternary ammonium compounds; qnrv, quinolone resistance gene; rmtB, 16S rRNA methyltransferase; RND, Resistance-Nodulation-Division; sul, sulfonamide resistance gene; tet, tetracycline resistane gene; triABC-opmH, triclosan efflux pump.

Fig. 3

Phylogenetic analysis, transmission networks, and mobile genetic elements of carbapenem resistant Escherichia coli ST410 isolated in Fiji (2020–2021). a) Core genome SNP phylogeny of carbapenem resistant Escherichia coli ST410 isolated in Fiji (2020–2021). Phylogenetic tree inferred from core genome SNPs of 44 CREc ST410 isolates. The core genome was generated by aligning at least 95% of the available sequence data for all isolates. The core SNP density, representing the average number of SNPs per base pair in the core genome, was 53 SNPs across 5,121,089 bp in the reference genome. The reference genome used was CREc ST410 (FJ431_LB), isolated from LBSH in 2020. The tree was rooted using the earliest isolate (reference). Isolation dates, patient admission locations, and AMR phenotypic and genotypic profiles are annotated on the tree. The scale bar on the tree represents the frequency of mutations per site, providing a measure of genetic distance between isolates. b) Putative transmission tree of CREc ST410 in CWMH, LTKH and LBSH, 2020–2021. A transmission tree shows inter-hospital transmission directions of CREc ST410 based on sample SNP distance and collection timeline. Numbers in red signify the SNP distance. Transmission tree generated and visualised using GraphSNP. The black arrow represents the earliest positive isolate. c) Plasmids harbouring insertion sequence (IS) and AMR genes in CREc ST410. i) The CREc ST410 representative isolate carried an IncX3 plasmid that contained bla_NDM-7 and multiple insertion elements. An IS5 element was detected upstream of the bla_NDM-7 in the IncX3 plasmid. ii) IncFII carried multiple AMR genes and IS. Plasmids were visualised using SnapGene (v6.2.1). AMR abbreviation: aph, aminoglycoside phosphotransferase; aadA, aminoglycoside adenylyltransferase; aac, aminoglycoside acetyltransferase; bla_NDM, New Delhi metallo-beta-lactamase; bla_OXA, OXA-type beta-lactamase; bla_TEM, temoneira beta-lactamase; bla_CMY, cephamycinase beta-lactamase; bla_CTX-M, cefotaximase-munich extended-spectrum beta-lactamase; bla_Ec, beta-lactamase Escherichia coli; ble-MBL, bleomycin resistance gene; cat chloramphenicol acetyltransferase; gyr, DNA gyrase; dfrA, dihydrofolate reductase; florR, florfenicol ressitance gene;mph, macrolide phosphotransferase; par, Type IV topoisomerase; qnrS1, quinolone resistance S1; acrAB-tolC, multidrug efflux pump; qacE, quaternary ammonium compounds; RND, resistance nodulation-division; sul, sulfonamide resistance gene; tet, tetracycline resistance gene.

Fig. 3

Phylogenetic analysis, transmission networks, and mobile genetic elements of carbapenem resistant Escherichia coli ST410 isolated in Fiji (2020–2021). a) Core genome SNP phylogeny of carbapenem resistant Escherichia coli ST410 isolated in Fiji (2020–2021). Phylogenetic tree inferred from core genome SNPs of 44 CREc ST410 isolates. The core genome was generated by aligning at least 95% of the available sequence data for all isolates. The core SNP density, representing the average number of SNPs per base pair in the core genome, was 53 SNPs across 5,121,089 bp in the reference genome. The reference genome used was CREc ST410 (FJ431_LB), isolated from LBSH in 2020. The tree was rooted using the earliest isolate (reference). Isolation dates, patient admission locations, and AMR phenotypic and genotypic profiles are annotated on the tree. The scale bar on the tree represents the frequency of mutations per site, providing a measure of genetic distance between isolates. b) Putative transmission tree of CREc ST410 in CWMH, LTKH and LBSH, 2020–2021. A transmission tree shows inter-hospital transmission directions of CREc ST410 based on sample SNP distance and collection timeline. Numbers in red signify the SNP distance. Transmission tree generated and visualised using GraphSNP. The black arrow represents the earliest positive isolate. c) Plasmids harbouring insertion sequence (IS) and AMR genes in CREc ST410. i) The CREc ST410 representative isolate carried an IncX3 plasmid that contained bla_NDM-7 and multiple insertion elements. An IS5 element was detected upstream of the bla_NDM-7 in the IncX3 plasmid. ii) IncFII carried multiple AMR genes and IS. Plasmids were visualised using SnapGene (v6.2.1). AMR abbreviation: aph, aminoglycoside phosphotransferase; aadA, aminoglycoside adenylyltransferase; aac, aminoglycoside acetyltransferase; bla_NDM, New Delhi metallo-beta-lactamase; bla_OXA, OXA-type beta-lactamase; bla_TEM, temoneira beta-lactamase; bla_CMY, cephamycinase beta-lactamase; bla_CTX-M, cefotaximase-munich extended-spectrum beta-lactamase; bla_Ec, beta-lactamase Escherichia coli; ble-MBL, bleomycin resistance gene; cat chloramphenicol acetyltransferase; gyr, DNA gyrase; dfrA, dihydrofolate reductase; florR, florfenicol ressitance gene;mph, macrolide phosphotransferase; par, Type IV topoisomerase; qnrS1, quinolone resistance S1; acrAB-tolC, multidrug efflux pump; qacE, quaternary ammonium compounds; RND, resistance nodulation-division; sul, sulfonamide resistance gene; tet, tetracycline resistance gene.

Fig. 4

Phylogenetic analysis and mobile genetic elements of carbapenem resistant Klebsiella pneumoniae isolates from Fiji (2020–2021). a) Core genome SNP phylogeny of carbapenem resistant Klebsiella pneumoniae ST16 isolates from Fiji (2020–2021). Phylogenetic tree inferred from core genome SNPs of four CRKp ST16 isolates. The core genome was generated by aligning at least 97% of the available sequence data for all isolates. The core SNP density, representing the average number of SNPs per base pair in the core genome, was 3 SNPs across 5,758,482 bp in the reference genome. The reference genome used was CRKp ST16 (FJ515_LT), isolated from LTKH in 2020. Sequence types, isolation dates, patient admission locations, and AMR phenotypic and genotypic profiles are annotated on the tree. The scale bar on the tree represents the frequency of mutations per site, providing a measure of genetic distance between isolates. b) IncX3 plasmids in CRKp ST16, CRKp ST45, and CRKp ST336: mapping of prominent IS and AMR genes. IncX3 plasmids from representative (i) CRKp ST16 (FJ515_LT), (ii) CRKp ST45 (FJ538_LT), and (iii) CRKp ST336 (FJ542_LT). The IncX3 plasmid detected in CRKp ST336 (FJ542_LT) exhibited a 100% similarity to the plasmid in CREc ST410 (FJ555_LT), when analysed using BLAST database comparsion (Appendix Fig. 8). Plasmids were visualised using SnapGene (v6.2.1). AMR abbreviation: aadA, aminoglycoside adenylyltransferase; bla_NDM, New Delhi metallo-beta-lactamase; bla_TEM, temoneira beta-lactamase; bla_CTX-M, cefotaximase-munich extended-spectrum beta-lactamase; bla_SHV, sulfhydryl variable beta-lactamase; ble-MBL, bleomycin resistance gene; gyr, DNA gyrase; dfrA, dihydrofolate reductase; kpnF, K. pneumoniae efflux pump; mph, macrolide phosphotransferase; qnr, quinolone resistance; qacE, quaternary ammonium compounds; rmtB, 16S rRNA methyltransferase; SMR, small multidrug resistance; sul, sulfonamide resistance gene; tet, tetracycline resistance gene.

Fig. 5

Geographical distribution and transfer of patients with CROs across health facilities in Fiji, 2020–2021. A total of 273 CRO strains were present across the three major hospitals and outer centers in Fiji. Arrows denote the transfer of patients across various hospitals and outer centers. Total number of CROs of each ST are shown in parentheses. Map created with National Geographic MapMaker.