Host-dependent resistance of Group A Streptococcus to sulfamethoxazole mediated by a horizontally-acquired reduced folate transporter.

Described antimicrobial resistance mechanisms enable bacteria to avoid the direct effects of antibiotics and can be monitored by in vitro susceptibility testing and genetic methods. Here we describe a mechanism of sulfamethoxazole resistance that requires a host metabolite for activity. Using a combination of in vitro evolution and metabolic rescue experiments, we identify an energy-coupling factor (ECF) transporter S component gene (thfT) that enables Group A Streptococcus to acquire extracellular reduced folate compounds. ThfT likely expands the substrate specificity of an endogenous ECF transporter to acquire reduced folate compounds directly from the host, thereby bypassing the inhibition of folate biosynthesis by sulfamethoxazole. As such, ThfT is a functional equivalent of eukaryotic folate uptake pathways that confers very high levels of resistance to sulfamethoxazole, yet remains undetectable when Group A Streptococcus is grown in the absence of reduced folates. Our study highlights the need to understand how antibiotic susceptibility of pathogens might function during infections to identify additional mechanisms of resistance and reduce ineffective antibiotic use and treatment failures, which in turn further contribute to the spread of antimicrobial resistance genes amongst bacterial pathogens.

Molecular Characterization and Antibiotic Susceptibility of Non-PCV13 Pneumococcal Serotypes among Vaccinated Children in Cape Coast, Ghana.

Preventive strategies involving the use of pneumococcal conjugate vaccines (PCVs) are known to drastically reduce pneumococcal disease. However, PCV vaccination has been plagued with serotype replacement by non-PCV serotypes. In this study, we describe the prevalence and molecular characteristics of non-PCV13 serotypes (non-vaccine serotypes, NVTs) from pneumococcal carriage isolates obtained from children < 5 years old in Cape Coast, Ghana, after PCV introduction. The isolates were subjected to antibiotic susceptibility testing and multilocus sequence typing (MLST), and molecular techniques were used to detect the presence of virulence genes. Serotypes 11A, 13, 15B, 23B, and 34 formed the top five of the 93 NVT isolates. As such, 20 (21.5%), 49 (48.4%), and 70 (74.3%) isolates were non-susceptible to penicillin, tetracycline, and cotrimoxazole, respectively. Sixteen (17.2%) multidrug-resistant isolates were identified. However, non-susceptibility to ceftriaxone and erythromycin was low and all isolates were fully susceptible to levofloxacin, linezolid, and vancomycin. Whereas pcpA, pavB, lytA, and psrP genes were detected in nearly all serotypes, pilus islet genes were limited to serotypes 11A, 13, and 23B. MLST for predominant serotype 23B isolates revealed three known and seven novel sequence types (STs). ST172 and novel ST15111 were the most dominant and both STs were related to PMEN clone Columbia23F-26 (ST338). In conclusion, non-PCV13 serotype 23B was the most prevalent, with characteristics of rapid clonal expansion of ST172 and ST15111, which are related to international clones of the pneumococcus. Continuous monitoring of NVTs in Ghana is, therefore, essential, as they have the potential to cause invasive disease, show high antibiotic resistance, and attenuate the effects of PCV vaccination.

Changes in the incidence of invasive disease due to Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis during the COVID-19 pandemic in 26 countries and territories in the Invasive Respiratory Infection Surveillance Initiative: a prospective analysis of surveillance data.

BACKGROUND: Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis, which are typically transmitted via respiratory droplets, are leading causes of invasive diseases, including bacteraemic pneumonia and meningitis, and of secondary infections subsequent to post-viral respiratory disease. The aim of this study was to investigate the incidence of invasive disease due to these pathogens during the early months of the COVID-19 pandemic. METHODS: In this prospective analysis of surveillance data, laboratories in 26 countries and territories across six continents submitted data on cases of invasive disease due to S pneumoniae, H influenzae, and N meningitidis from Jan 1, 2018, to May, 31, 2020, as part of the Invasive Respiratory Infection Surveillance (IRIS) Initiative. Numbers of weekly cases in 2020 were compared with corresponding data for 2018 and 2019. Data for invasive disease due to Streptococcus agalactiae, a non-respiratory pathogen, were collected from nine laboratories for comparison. The stringency of COVID-19 containment measures was quantified using the Oxford COVID-19 Government Response Tracker. Changes in population movements were assessed using Google COVID-19 Community Mobility Reports. Interrupted time-series modelling quantified changes in the incidence of invasive disease due to S pneumoniae, H influenzae, and N meningitidis in 2020 relative to when containment measures were imposed. FINDINGS: 27 laboratories from 26 countries and territories submitted data to the IRIS Initiative for S pneumoniae (62 837 total cases), 24 laboratories from 24 countries submitted data for H influenzae (7796 total cases), and 21 laboratories from 21 countries submitted data for N meningitidis (5877 total cases). All countries and territories had experienced a significant and sustained reduction in invasive diseases due to S pneumoniae, H influenzae, and N meningitidis in early 2020 (Jan 1 to May 31, 2020), coinciding with the introduction of COVID-19 containment measures in each country. By contrast, no significant changes in the incidence of invasive S agalactiae infections were observed. Similar trends were observed across most countries and territories despite differing stringency in COVID-19 control policies. The incidence of reported S pneumoniae infections decreased by 68% at 4 weeks (incidence rate ratio 0·32 [95% CI 0·27-0·37]) and 82% at 8 weeks (0·18 [0·14-0·23]) following the week in which significant changes in population movements were recorded. INTERPRETATION: The introduction of COVID-19 containment policies and public information campaigns likely reduced transmission of S pneumoniae, H influenzae, and N meningitidis, leading to a significant reduction in life-threatening invasive diseases in many countries worldwide. FUNDING: Wellcome Trust (UK), Robert Koch Institute (Germany), Federal Ministry of Health (Germany), Pfizer, Merck, Health Protection Surveillance Centre (Ireland), SpID-Net project (Ireland), European Centre for Disease Prevention and Control (European Union), Horizon 2020 (European Commission), Ministry of Health (Poland), National Programme of Antibiotic Protection (Poland), Ministry of Science and Higher Education (Poland), Agencia de Salut Pública de Catalunya (Spain), Sant Joan de Deu Foundation (Spain), Knut and Alice Wallenberg Foundation (Sweden), Swedish Research Council (Sweden), Region Stockholm (Sweden), Federal Office of Public Health of Switzerland (Switzerland), and French Public Health Agency (France).

Application of a Pneumococcal Serotype-specific Urinary Antigen Detection Test for Identification of Pediatric Pneumonia in Burkina Faso.

BACKGROUND: Serotype-specific diagnosis of pneumococcal community-acquired pneumonia in children under age 5 years would mark a major advancement for understanding pneumococcal epidemiology and supporting vaccine decision-making. METHODS: A Luminex technology-based multiplex urinary antigen detection (UAD) diagnostic assay was developed and subsequently validated in adults, but its applicability to children is unknown. This study aimed to set appropriate cutoffs for use of the UAD in a healthy pediatric population and apply these cutoffs in children with pneumonia in sub-Saharan Africa. The cutoffs were determined by assessing 379 urines obtained from healthy children under age 5 years from the Bobo-Dioulasso area for serotypes included in 13-valent pneumococcal conjugate vaccine (UAD-1) and the 11 other serotypes unique to 23-valent pneumococcal polysaccharide vaccine (UAD-2). RESULTS: Based on the assigned cutoff values, among 108 children who met the World Health Organization consolidation endpoint criteria, UAD-1 and UAD-2 were positive in 23.3% and 8.3%, respectively; among 364 children with clinically suspected pneumonia who did not meet the World Health Organization criteria, UAD-1 and UAD-2 were positive for 6.6% and 3.6%, respectively. Pneumococcal carriage prevalence was similar among pneumonia cases (30%) versus controls (35%) as was semiquantitative carriage density. CONCLUSIONS: UAD-1 and UAD-2 were able to distinguish community controls from children with pneumonia, particularly pneumonia with consolidation. Future studies are needed to confirm these results and more fully assess the contribution of pneumococcal carriage and concurrent viral infection.

Post-Vaccination Streptococcus pneumoniae Carriage and Virulence Gene Distribution among Children Less Than Five Years of Age, Cape Coast, Ghana.

In 2012, Ghana introduced PCV13 into its childhood immunization program. To monitor the pneumococcus after PCV13 vaccination, we analyzed serotypes, antibiotic resistance, and virulence genes of pneumococcal carriage isolates among children under five years of age. We obtained nasopharyngeal swabs from 513 children from kindergartens and immunization centers in Cape Coast, Ghana. Pneumococcal serotypes were determined by multiplex-PCR and Quellung reaction. Antibiotic resistance and virulence genes prevalence were determined by disc diffusion and PCR respectively. Overall, carriage prevalence was 29.4% and PCV13 coverage was 38.4%. Over 60% of the isolates were non-PCV13 serotypes and serotype 23B was the most prevalent. One isolate showed full resistance to penicillin, while 35% showed intermediate resistance. Resistance to erythromycin and clindamycin remained low, while susceptibility to ceftriaxone, levofloxacin and vancomycin remained high. Penicillin resistance was associated with PCV13 serotypes. Forty-three (28.5%) strains were multidrug-resistant. Virulence genes pavB, pcpA, psrP, pilus-1, and pilus-2 were detected in 100%, 87%, 62.9%, 11.9%, and 6.6% of the strains, respectively. The pilus islets were associated with PCV13 and multidrug-resistant serotypes. PCV13 vaccination had impacted on pneumococcal carriage with a significant increase in non-PCV13 serotypes and lower penicillin resistance. Including PcpA and PsrP in pneumococcal protein-based vaccines could be beneficial to Ghanaian children.

Colonization dynamics of Streptococcus pneumoniae in a remote African population: A prospective cohort study.

The aim of this study was to analyze the population dynamics of Streptococcus pneumoniae in a remotely living African Pygmy population. The same pygmy population (Gabon) was prospectively screened for nasopharyngeal S. pneumoniae colonization in 2011 (n = 103), 2013 (n = 104) and 2017 (n = 107). Non-duplicate isolates (n = 126) were serotyped and tested for antimicrobial resistance (broth microdilution). At the three sampling time points, resistance rates were highest for tetracycline (36-58%), followed by penicillin (parenteral, meningitis-breakpoints, 6-39%) and chloramphenicol (3-15%). The majority of isolates was non-typeable (NT, n = 18/126, 14.3%) followed by serotype 6B (n = 17/126, 13.5%), 21 and 15A (n = 9/126, 7.1%, each). The distribution of serotypes was highly dynamic as only three serotypes (14, 17F, NT) were detected during all three visits. Resistance rates and serotypes of nasopharyngeal S. pneumoniae markedly changed in the remote Babongo population. This rapid change in serotypes could challenge the selection of pneumococcal vaccine.

Identification of Streptococcus suis Meningitis through Population-Based Surveillance, Togo, 2010-2014.

During 2010-2014, we enrolled 511 patients with suspected bacterial meningitis into surveillance in 2 districts of northern Togo. We identified 15 persons with Streptococcus suis infection; 10 had occupational contact with pigs, and 12 suffered neurologic sequelae. S. suis testing should be considered in rural areas of the African meningitis belt.

The Relevance of a Novel Quantitative Assay to Detect up to 40 Major Streptococcus pneumoniae Serotypes Directly in Clinical Nasopharyngeal and Blood Specimens.

For epidemiological and surveillance purposes, it is relevant to monitor the distribution and dynamics of Streptococcus pneumoniae serotypes. Conventional serotyping methods do not provide rapid or quantitative information on serotype loads. Quantitative serotyping may enable prediction of the invasiveness of a specific serotype compared to other serotypes carried. Here, we describe a novel, rapid multiplex real-time PCR assay for identification and quantification of the 40 most prevalent pneumococcal serotypes and the assay impacts in pneumonia specimens from emerging and developing countries. Eleven multiplex PCR to detect 40 serotypes or serogroups were optimized. Quantification was enabled by reference to standard dilutions of known bacterial load. Performance of the assay was evaluated to specifically type and quantify S. pneumoniae in nasopharyngeal and blood samples from adult and pediatric patients hospitalized with pneumonia (n = 664) from five different countries. Serogroup 6 was widely represented in nasopharyngeal specimens from all five cohorts. The most frequent serotypes in the French, South African, and Brazilian cohorts were 1 and 7A/F, 3 and 19F, and 14, respectively. When both samples were available, the serotype in blood was always present as carriage with other serotypes in the nasopharynx. Moreover, the ability of a serotype to invade the bloodstream may be linked to its nasopharyngeal load. The mean nasopharyngeal concentration of the serotypes that moved to the blood was 3 log-fold higher than the ones only found in the nasopharynx. This novel, rapid, quantitative assay may potentially predict some of the S. pneumoniae serotypes invasiveness and assessment of pneumococcal serotype distribution.

Discovery of Streptococcus pneumoniae serotype 6 variants with glycosyltransferases synthesizing two differing repeating units.

Streptococcus pneumoniae is a persistent, opportunistic commensal of the human nasopharynx and is the leading cause of community-acquired pneumonia. It expresses an anti-phagocytic capsular polysaccharide (PS). Genetic variation of the capsular PS synthesis (cps) locus is the molecular basis for structural and antigenic heterogeneity of capsule types (serotypes). Serogroup 6 has four known members (6A-6D) with distinct serologic properties, homologous cps loci, and structurally similar PSs. cps of serotypes 6A/6B have wciNα, encoding α-1,3-galactosyltransferase, whereas serotypes 6C/6D have wciNß encoding α-1,3-glucosyltransferase. Two atypical serogroup 6 isolates (named 6X11 and 6X12) have been discovered recently in Germany. Flow cytometric studies using monoclonal antibodies show that 6X11 has serologic properties of 6B/6D, whereas 6X12 has 6A/6C. NMR studies of their capsular PSs revealed that 6X11 and 6X12 have two different repeating units with a distribution of ~40:60 6B:6D and 75:25 6A:6C PS, respectively. Sequencing of the wciNα gene in 6X12 and 6X11 revealed single and double nucleotide substitutions, respectively, resulting in the amino acid changes A150T and D38N. Substitution of alanine with threonine at position 150 in a 6A strain was associated with hybrid serologic and chemical profiles like 6X12. The hybrid serotypes represented by 6X12 and 6X11 strains are now named serotypes 6F and 6G. Single amino acid changes in cps genes encoding glycosyltransferases can alter substrate specificities, permit biosynthesis of heterogeneous capsule repeating units, and result in new hybrid capsule types that may differ in their interaction with the immune system of the host.