Establishing a national reference laboratory for antimicrobial resistance using a whole-genome sequencing framework: Nigeria's experience.

Whole-genome sequencing (WGS) is finding important applications in the surveillance of antimicrobial resistance (AMR), providing the most granular data and broadening the scope of niches and locations that can be surveilled. A common but often overlooked application of WGS is to replace or augment reference laboratory services for AMR surveillance. WGS has supplanted traditional strain subtyping in many comprehensive reference laboratories and is now the gold standard for rapidly ruling isolates into or out of suspected outbreak clusters. These and other properties give WGS the potential to serve in AMR reference functioning where a reference laboratory did not hitherto exist. In this perspective, we describe how we have employed a WGS approach, and an academic-public health system collaboration, to provide AMR reference laboratory services in Nigeria, as a model for leapfrogging to national AMR surveillance.

Clones and Clusters of Antimicrobial-Resistant Klebsiella From Southwestern Nigeria.

BACKGROUND: Klebsiella pneumoniae is a World Health Organization high-priority antibiotic-resistant pathogen. However, little is known about Klebsiella lineages circulating in Nigeria. METHODS: We performed whole-genome sequencing (WGS) of 141 Klebsiella isolated between 2016 and 2018 from clinical specimens at 3 antimicrobial-resistance (AMR) sentinel surveillance tertiary hospitals in southwestern Nigeria. We conducted in silico multilocus sequence typing; AMR gene, virulence gene, plasmid, and K and O loci profiling; as well as phylogenetic analyses, using publicly available tools and Nextflow pipelines. RESULTS: Phylogenetic analysis revealed that the majority of the 134 K. pneumoniae and 5 K. quasipneumoniae isolates from Nigeria characterized are closely related to globally disseminated multidrug-resistant clones. Of the 39 K. pneumoniae sequence types (STs) identified, the most common were ST307 (15%), ST5241 (12%), ST15 (~9%), and ST25 (~6%). ST5241, 1 of 10 novel STs detected, is a single locus variant of ST636 carrying dfrA14, tetD, qnrS, and oqxAB resistance genes. The extended-spectrum ß-lactamase (ESBL) gene blaCTX_M-15 was seen in 72% of K. pneumoniae genomes, while 8% encoded a carbapenemase. No isolate carried a combination of carbapenemase-producing genes. Four likely outbreak clusters from 1 facility, within STs 17, 25, 307, and 5241, were ESBL but not carbapenemase-bearing clones. CONCLUSIONS: This study uncovered known and novel K. pneumoniae lineages circulating in 3 hospitals in Southwest Nigeria that include multidrug-resistant ESBL producers. Carbapenemase-producing isolates remain uncommon. WGS retrospectively identified outbreak clusters, pointing to the value of genomic approaches in AMR surveillance for improving infection prevention and control in Nigerian hospitals.

NDM-1- and OXA-23-producing Acinetobacter baumannii in wastewater of a Nigerian hospital.

IMPORTANCE: Acinetobacter baumannii is a leading cause of hospital-associated infections globally. A. baumannii reservoirs outside hospital settings are still unknown, and their occurrence in the environment is linked to clinical and anthropogenic activities. Although the risk of transmission of A. baumannii from environmental sources to humans is not fully understood, these sources pose significant risks for the continued dissemination of A. baumannii and their resistance traits. This study provides evidence that diverse and clinically relevant A. baumannii strains, many of which are resistant to carbapenems, are constantly being discharged into the environment through inadequately treated hospital wastewater. We further elucidate potential transmission routes between the environment and clinical infections and demonstrate the high prevalence of carbapenem resistance genes on highly mobile transposons among these strains. Our findings highlight the pressing need to address hospital wastewater as a crucial factor in curtailing the spread of carbapenem-resistant A. baumannii.

High Genetic Diversity of Carbapenem-Resistant Acinetobacter baumannii Isolates Recovered in Nigerian Hospitals in 2016 to 2020.

Acinetobacter baumannii causes difficult-to-treat infections mostly among immunocompromised patients. Clinically relevant A. baumannii lineages and their carbapenem resistance mechanisms are sparsely described in Nigeria. This study aimed to characterize the diversity and genetic mechanisms of carbapenem resistance among A. baumannii strains isolated from hospitals in southwestern Nigeria. We sequenced the genomes of all A. baumannii isolates submitted to Nigeria's antimicrobial resistance surveillance reference laboratory between 2016 and 2020 on an Illumina platform and performed in silico genomic characterization. Selected strains were sequenced using the Oxford Nanopore technology to characterize the genetic context of carbapenem resistance genes. The 86 A. baumannii isolates were phylogenetically diverse and belonged to 35 distinct Oxford sequence types (oxfSTs), 16 of which were novel, and 28 Institut Pasteur STs (pasSTs). Thirty-eight (44.2%) isolates belonged to none of the known international clones (ICs). Over 50% of the isolates were phenotypically resistant to 10 of 12 tested antimicrobials. The majority (n = 54) of the isolates were carbapenem resistant, particularly the IC7 (pasST25; 100%) and IC9 (pasST85; >91.7%) strains. blaOXA-23 (34.9%) and blaNDM-1 (27.9%) were the most common carbapenem resistance genes detected. All blaOXA-23 genes were carried on Tn2006 or Tn2006-like transposons. Our findings suggest that a 10-kb Tn125 composite transposon is the primary means of blaNDM-1 dissemination. Our findings highlight an increase in blaNDM-1 prevalence and the widespread transposon-facilitated dissemination of carbapenemase genes in diverse A. baumannii lineages in southwestern Nigeria. We make the case for improving surveillance of these pathogens in Nigeria and other understudied settings. IMPORTANCE Acinetobacter baumannii bacteria are increasingly clinically relevant due to their propensity to harbor genes conferring resistance to multiple antimicrobials, as well as their ability to persist and disseminate in hospital environments and cause difficult-to-treat nosocomial infections. Little is known about the molecular epidemiology and antimicrobial resistance profiles of these organisms in Nigeria, largely due to limited capacity for their isolation, identification, and antimicrobial susceptibility testing. Our study characterized the diversity and antimicrobial resistance profiles of clinical A. baumannii in southwestern Nigeria using whole-genome sequencing. We also identified the key genetic elements facilitating the dissemination of carbapenem resistance genes within this species. This study provides key insights into the clinical burden and population dynamics of A. baumannii in hospitals in Nigeria and highlights the importance of routine whole-genome sequencing-based surveillance of this and other previously understudied pathogens in Nigeria and other similar settings.

A bottom-up view of antimicrobial resistance transmission in developing countries.

Antimicrobial resistance (AMR) is tracked most closely in clinical settings and high-income countries. However, resistant organisms thrive globally and are transmitted to and from healthy humans, animals and the environment, particularly in many low- and middle-income settings. The overall public health and clinical significance of these transmission opportunities remain to be completely clarified. There is thus considerable global interest in promoting a One Health view of AMR to enable a more realistic understanding of its ecology. In reality, AMR surveillance outside hospitals remains insufficient and it has been very challenging to convincingly document transmission at the interfaces between clinical specimens and other niches. In this Review, we describe AMR and its transmission in low- and middle-income-country settings, emphasizing high-risk transmission points such as urban settings and food-animal handling. In urban and food production settings, top-down and infrastructure-dependent interventions against AMR that require strong regulatory oversight are less likely to curtail transmission when used alone and should be combined with bottom-up AMR-containment approaches. We observe that the power of genomics to expose transmission channels and hotspots is largely unharnessed, and that existing and upcoming technological innovations need to be exploited towards containing AMR in low- and middle-income settings.

Rectal Colonization and Nosocomial Transmission of Carbapenem-Resistant Acinetobacter baumannii in an Intensive Care Unit, Southwest Nigeria.

Background: Acinetobacter baumannii are of major human health importance because they cause life-threatening nosocomial infections and often are highly resistant to antimicrobials. Specific multidrug-resistant A. baumannii lineages are implicated in hospital outbreaks globally. We retrospectively investigated a suspected outbreak of carbapenem-resistant A. baumannii (CRAB) colonizing patients in an intensive care unit (ICU) of a tertiary hospital in Southwest Nigeria where genomic surveillance of Acinetobacter has hitherto not been conducted. Methods: A prospective observational study was conducted among all patients admitted to the ICU between August 2017 and June 2018. Acinetobacter species were isolated from rectal swabs and verified phenotypically with the Biomerieux Vitek 2 system. Whole genome sequencing (WGS) was performed on the Illumina platform to characterize isolates from a suspected outbreak during the study period. Phylogenetic analysis, multilocus sequence typing, and antimicrobial resistance gene prediction were carried out in silico. Results: Acinetobacter isolates belonging to the A. baumannii complex were recovered from 20 (18.5%) ICU patients. Single nucleotide polymorphism (SNP) analysis and epidemiological information revealed a putative outbreak clone comprising seven CRAB strains belonging to the globally disseminated international clone (IC) 2. These isolates had ≤2 SNP differences, identical antimicrobial resistance and virulence genes, and were all ST1114/1841. Conclusion: We report a carbapenem-resistant IC2 A. baumannii clone causing an outbreak in an ICU in Nigeria. The study findings underscore the need to strengthen the capacity to detect A. baumannii in human clinical samples in Nigeria and assess which interventions can effectively mitigate CRAB transmission in Nigerian hospital settings.

An ST131 clade and a phylogroup A clade bearing an O101-like O-antigen cluster predominate among bloodstream Escherichia coli isolates from South-West Nigeria hospitals.

Escherichia coli bloodstream infections are typically attributed to a limited number of lineages that carry virulence factors associated with invasiveness. In Nigeria, the identity of circulating clones is largely unknown and surveillance of their antimicrobial resistance has been limited. We verified and whole-genome sequenced 68 2016-2018 bloodstream E. coli isolates from three sentinel sites in South-Western Nigeria and susceptibility tested 67 of them. Resistance to antimicrobials commonly used in Nigeria was high, with 67 (100 %), 62 (92.5 %), 53 (79.1 %) and 37 (55.2 %) showing resistance to trimethoprim, ampicillin, ciprofloxacin and aminoglycosides, respectively. Thirty-five (51 %) isolates carried extended-spectrum ß-lactamase genes and 32 (91 %) of these were multidrug resistant. All the isolates were susceptible to carbapenems and colistin. The strain set included globally disseminated high-risk clones from sequence type (ST)12 (2), ST131 (12) and ST648 (4). Twenty-three (33.8 %) of the isolates clustered within two clades. The first of these consisted of ST131 strains, comprising O16:H5 and O25:H4 sub-lineages. The second was an ST10-ST167 complex clade comprising strains carrying O-antigen and capsular genes of likely Klebsiella origin, identical to those of avian pathogenic E. coli Sanji, and serotyped in silico as O89, O101 or ONovel32, depending on the tool used. Four temporally associated ST90 strains from one sentinel were closely related enough to suggest that at least some of them represented a retrospectively detected outbreak cluster. Our data implicate a broad repertoire of E. coli isolates associated with bloodstream infections in South-West Nigeria. Continued genomic surveillance is valuable for tracking clones of importance and for outbreak identification.

Genomic characterization of invasive typhoidal and non-typhoidal Salmonella in southwestern Nigeria.

BACKGROUND: Salmonellosis causes significant morbidity and mortality in Africa. Information on lineages of invasive Salmonella circulating in Nigeria is sparse. METHODS: Salmonella enterica isolated from blood (n = 60) and cerebrospinal fluid (CSF, n = 3) between 2016 and 2020 from five tertiary hospitals in southwest Nigeria were antimicrobial susceptibility-tested and Illumina-sequenced. Genomes were analysed using publicly-available bioinformatic tools. RESULTS: Isolates and sequence types (STs) from blood were S. Typhi [ST1, n = 1 and ST2, n = 43] and invasive non-typhoidal Salmonella (iNTS) (S. Enteritidis [ST11, n = 7], S. Durham [ST10, n = 2], S. Rissen [ST8756, n = 2], S. Chester [ST2063, n = 1], S. Dublin [ST10, n = 1], S. Infantis [ST603, n = 1], S. Telelkebir [ST8757, n = 1] and S. Typhimurium [ST313, n = 1]). S. Typhi ST2 (n = 2) and S. Adabraka ST8757 (n = 1) were recovered from CSF. Most S. Typhi belonged to genotype 3.1.1 (n = 44), carried an IncY plasmid, had several antibiotic resistance genes (ARGs) including blaTEM-1 (n = 38), aph(6)-Id (n = 32), tet(A) (n = 33), sul2 (n = 32), dfrA14 (n = 30) as well as quinolone resistance-conferring gyrA_S83Y single-nucleotide polymorphisms (n = 37). All S. Enteritidis harboured aph(3")-Ib, blaTEM-1, catA1, dfrA7, sul1, sul2, tet(B) genes, and a single ARG, qnrB19, was detected in S. Telelkebir. Typhoidal toxins cdtB, pltA and pltB were detected in S. Typhi, Rissen, Chester, and Telelkebir. CONCLUSION: Most invasive salmonelloses in southwest Nigeria are vaccine-preventable infections due to multidrug-resistant, West African dominant S. Typhi lineage 3.1.1. Invasive NTS serovars, including some harbouring typhoidal toxin or resistance genes, represented a third of the isolates emphasizing the need for better diagnosis and surveillance.