Seye Abimbola and co-authors argue for a transformation in global health research and practice in the post-COVID-19 world.

With the rise in fluoroquinolone-resistant Salmonella Typhi and the recent emergence of ceftriaxone resistance, azithromycin is one of the last oral drugs available against typhoid for which resistance is uncommon. Its increasing use, specifically in light of the ongoing outbreak of extensively drug-resistant (XDR) Salmonella Typhi (resistant to chloramphenicol, ampicillin, cotrimoxazole, streptomycin, fluoroquinolones and third-generation cephalosporins) in Pakistan, places selective pressure for the emergence and spread of azithromycin-resistant isolates. However, little is known about azithromycin resistance in Salmonella, and no molecular data are available on its mechanism. We conducted typhoid surveillance in the two largest pediatric hospitals of Bangladesh from 2009–2016. All typhoidal Salmonella strains were screened for azithromycin resistance using disc diffusion and resistance was confirmed using E-tests. In total, we identified 1,082 Salmonella Typhi and Paratyphi A strains; among these, 13 strains (12 Typhi, 1 Paratyphi A) were azithromycin-resistant (MIC range: 32–64 μg/ml) with the first case observed in 2013. We sequenced the resistant strains, but no molecular basis of macrolide resistance was identified by the currently available antimicrobial resistance prediction tools. A whole genome SNP tree, made using RAxML, showed that the 12 Typhi resistant strains clustered together within the 4.3.1.1 sub-clade (H58 lineage 1). We found a non-synonymous single-point mutation exclusively in these 12 strains in the gene encoding AcrB, an efflux pump that removes small molecules from bacterial cells. The mutation changed the conserved amino acid arginine (R) at position 717 to a glutamine (Q). To test the role of R717Q present in azithromycin-resistant strains, we cloned acrB from azithromycin-resistant and sensitive strains, expressed them in E. coli, Typhi and Paratyphi A strains and tested their azithromycin susceptibility. Expression of AcrB-R717Q in E. coli and Typhi strains increased the minimum inhibitory concentration (MIC) for azithromycin by 11- and 3-fold respectively. The azithromycin-resistant Paratyphi A strain also contained a mutation at R717 (R717L), whose introduction in E. coli and Paratyphi A strains increased MIC by 7- and 3-fold respectively, confirming the role of R717 mutations in conferring azithromycin resistance. This report confirms 12 azithromycin-resistant Salmonella Typhi strains and one Paratyphi A strain. The molecular basis of this resistance is one mutation in the AcrB protein at position 717. This is the first report demonstrating the impact of this non-synonymous mutation in conferring macrolide resistance in a clinical setting. With increasing azithromycin use, strains with R717 mutations may spread and be acquired by XDR strains. An azithromycin-resistant XDR strain would shift enteric fever treatment from outpatient departments, where patients are currently treated with oral azithromycin, to inpatient departments to be treated with injectable antibiotics like carbapenems, thereby further burdening already struggling health systems in endemic regions. Moreover, with the dearth of novel antimicrobials in the horizon, we risk losing our primary defense against widespread mortality from typhoid. In addition to rolling out the WHO prequalified typhoid conjugate vaccine in endemic areas to decrease the risk of pan-resistant Salmonella Typhi strains, it is also imperative to implement antimicrobial stewardship and water sanitation and hygiene intervention to decrease the overall burden of enteric fever. With the outbreak of extensively resistant (XDR) typhoid fever in Pakistan, azithromycin has become the last oral drug to treat typhoid. Although no azithromycin resistant XDR isolate has been reported to date, the increasing use of azithromycin and the clear historical record of widespread dissemination of resistance to all previously widely used antimicrobials by typhoidal Salmonella, suggest we will soon face strains resistant to all oral antibiotics. This makes it imperative to elucidate the mechanism of azithromycin resistance in typhoidal Salmonella. We tested 1,082 typhoidal Salmonella isolates from the two largest pediatric hospitals of Bangladesh and identified 13 azithromycin-resistant isolates. Using comparative genomics, we identified a mutation in a specific protein called AcrB that makes these isolates resistant to azithromycin. All azithromycin-resistant strains were susceptible to cephalosporin but resistant to all other oral antibiotics. The Pakistan outbreak strain is resistant to all common oral antibiotics and only susceptible to azithromycin. Acquisition of the plasmid that confers cephalosporin resistance in XDR strains by the Bangladeshi azithromycin-resistant strains or rise of the AcrB mutation in the XDR strains could be the end of oral treatment for typhoid. This poses serious threats to the health system of LMICs where typhoid is endemic. Currently, the majority of typhoid patients are prescribed oral treatment in the outpatient department, but an azithromycin-resistant XDR strain would shift enteric fever treatment from outpatient departments to inpatient departments to be treated with injectable antibiotics like carbapenems, further burdening already struggling health systems in endemic regions. Moreover, with the dearth of novel antimicrobials in the horizon, we risk losing our primary defense against widespread mortality from enteric fever.

Background: The Global Typhoid Genomics Consortium was established to bring together the typhoid research community to aggregate and analyse Salmonella enterica serovar Typhi (Typhi) genomic data to inform public health action. This analysis, which marks 22 years since the publication of the first Typhi genome, represents the largest Typhi genome sequence collection to date (n=13,000).Methods: This is a meta-analysis of global genotype and antimicrobial resistance (AMR) determinants extracted from previously sequenced genome data and analysed using consistent methods implemented in open analysis platforms GenoTyphi and Pathogenwatch.Results: Compared with previous global snapshots, the data highlight that genotype 4.3.1 (H58) has not spread beyond Asia and Eastern/Southern Africa; in other regions, distinct genotypes dominate and have independently evolved AMR. Data gaps remain in many parts of the world, and we show the potential of travel-associated sequences to provide informal ‘sentinel’ surveillance for such locations. The data indicate that ciprofloxacin non-susceptibility (>1 resistance determinant) is widespread across geographies and genotypes, with high-level ciprofloxacin resistance (≥3 determinants) reaching 20% prevalence in South Asia. Extensively drug-resistant (XDR) typhoid has become dominant in Pakistan (70% in 2020) but has not yet become established elsewhere. Ceftriaxone resistance has emerged in eight non-XDR genotypes, including a ciprofloxacin-resistant lineage (4.3.1.2.1) in India. Azithromycin resistance mutations were detected at low prevalence in South Asia, including in two common ciprofloxacin-resistant genotypes.Conclusions: The consortium’s aim is to encourage continued data sharing and collaboration to monitor the emergence and global spread of AMR Typhi, and to inform decision-making around the introduction of typhoid conjugate vaccines (TCVs) and other prevention and control strategies.Funding: No specific funding was awarded for this meta-analysis. Coordinators were supported by fellowships from the European Union (ZAD received funding from the European Union’s Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No 845681), the Wellcome Trust (SB, Wellcome Trust Senior Fellowship), and the National Health and Medical Research Council (DJI is supported by an NHMRC Investigator Grant [GNT1195210]).

The burden of meningitis in low-and-middle-income countries remains significant, but the infectious causes remain largely unknown, impeding institution of evidence-based treatment and prevention decisions. We conducted a validation and application study of unbiased metagenomic next-generation sequencing (mNGS) to elucidate etiologies of meningitis in Bangladesh. This RNA mNGS study was performed on cerebrospinal fluid (CSF) specimens from patients admitted in the largest pediatric hospital, a World Health Organization sentinel site, with known neurologic infections (n = 36), with idiopathic meningitis (n = 25), and with no infection (n = 30), and six environmental samples, collected between 2012 and 2018. We used the IDseq bioinformatics pipeline and machine learning to identify potentially pathogenic microbes, which we then confirmed orthogonally and followed up through phone/home visits. In samples with known etiology and without infections, there was 83% concordance between mNGS and conventional testing. In idiopathic cases, mNGS identified a potential bacterial or viral etiology in 40%. There were three instances of neuroinvasive Chikungunya virus (CHIKV), whose genomes were >99% identical to each other and to a Bangladeshi strain only previously recognized to cause febrile illness in 2017. CHIKV-specific qPCR of all remaining stored CSF samples from children who presented with idiopathic meningitis in 2017 (n = 472) revealed 17 additional CHIKV meningitis cases, exposing an unrecognized meningitis outbreak. Orthogonal molecular confirmation, case-based clinical data, and patient follow-up substantiated the findings. Case-control CSF mNGS surveys can complement conventional diagnostic methods to identify etiologies of meningitis, conduct surveillance, and predict outbreaks. The improved patient- and population-level data can inform evidence-based policy decisions. IMPORTANCE Globally, there are an estimated 10.6 million cases of meningitis and 288,000 deaths every year, with the vast majority occurring in low- and middle-income countries. In addition, many survivors suffer from long-term neurological sequelae. Most laboratories assay only for common bacterial etiologies using culture and directed PCR, and the majority of meningitis cases lack microbiological diagnoses, impeding institution of evidence-based treatment and prevention strategies. We report here the results of a validation and application study of using unbiased metagenomic sequencing to determine etiologies of idiopathic (of unknown cause) cases. This included CSF from patients with known neurologic infections, with idiopathic meningitis, and without infection admitted in the largest children’s hospital of Bangladesh and environmental samples. Using mNGS and machine learning, we identified and confirmed an etiology (viral or bacterial) in 40% of idiopathic cases. We detected three instances of Chikungunya virus (CHIKV) that were >99% identical to each other and to a strain previously recognized to cause systemic illness only in 2017. CHIKV qPCR of all remaining stored 472 CSF samples from children who presented with idiopathic meningitis in 2017 at the same hospital uncovered an unrecognized CHIKV meningitis outbreak. CSF mNGS can complement conventional diagnostic methods to identify etiologies of meningitis, and the improved patient- and population-level data can inform better policy decisions.

In the early 1900s, with mortality of ∼30%, typhoid and paratyphoid (caused by Salmonella Typhi and Paratyphi A) ravaged parts of the world; with improved water, sanitation, and hygiene in resource-rich countries and the advent of antimicrobials, mortality dwindled to <1%. Today, the burden rests disproportionately on South Asia, where the primary means for combatting the disease is antimicrobials.