Anti-carbohydrate antibodies (Abs) play crucial roles in pathogen control, but their generation remains poorly understood. Here, we studied how B cell immunity against the Streptococcus pyogenes surface polysaccharide is influenced by age and antigen (Ag) exposure across human blood, spleen, and tonsils. The glycan-targeted response shifted from immunoglobulin M (IgM) biased in children toward IgG and IgA biased in adults. Both natural colonization and controlled human infection with S. pyogenes increased class-switched B cells, with evidence of within-clone switching. Glycan-specific B cells readily engaged in germinal center (GC) responses and underwent robust somatic hypermutation (SHM) but had reduced expression of T cell help-associated molecules, correlating with lower antibody-secreting cell (ASC) output. Thus, mucosal pathogen encounters elicit glycan responses that class switch, evolve, and diversify through the GC, with implications for the design and schedule of glycan-containing vaccines.

Background:The global health burden of Streptococcus pyogenes (Group A Streptococcus, GAS) disease has led the World Health Organization to declare GAS vaccine development an international priority. Severe acute GAS disease comprises hospitalised infections, toxin-mediated disease (scarlet fever and streptococcal toxic shock syndrome) and immune-mediated disease (acute rheumatic fever [ARF] and acute post-streptococcal glomerulonephritis). This study aimed to characterize the burden and spectrum of severe acute GAS disease in Auckland, New Zealand, to inform future GAS vaccine studies in this setting. Methods:This prospective population-based multicentre cross-sectional observational study utilized laboratory data and ARF notifications to capture all hospitalised GAS infections and toxin- and immune-mediated disease cases associated with hospitalisation in Auckland during 2023. The epidemiology, clinical features, incidence and hospitalisation costs of these cases are described. Findings:The complete clinical spectrum of severe acute GAS disease was observed, with 606 cases captured corresponding to 1:2000 people affected each year (52 cases/100,000 population/year). The burden is inequitably distributed across the population, greatest at the intersection of age, ethnicity and socioeconomic deprivation. In the most deprived areas, approximately 1:400 Pacific children <10-years and 1:200 Pacific adults ≥80-years are affected annually (261 and 483 cases/100,000 population/year, respectively). Direct hospitalisation costs exceeded $13·2 million New Zealand dollars. Interpretation:This study demonstrates that the complete clinical spectrum of severe acute GAS disease occurs in Auckland and is associated with a substantial health and economic burden. The high incidence and entire range of severe acute GAS disease present make Auckland an ideal location for future vaccine studies. Funding:Ministry of Health New Zealand, Health Research Council of New Zealand (HRC-NZ), Royal College of Pathologists of Australasia (RCPA) Foundation.

Introduction:The lack of specific diagnostic tests for acute rheumatic fever (ARF) and its consequence rheumatic heart disease (RHD) is a key barrier to effective control of these diseases in low resource settings. Methods:We used immunoassays to evaluate blood prothymosin alpha (PTA) and immunoglobulin G3 (IgG3) in patients with ARF, RHD and controls using serum samples from Pakistan and plasma samples from Uganda. Results:Across both sets of samples, we found total IgG3 was significantly elevated in definite ARF (n = 24) compared to controls (n = 36), possible ARF (n = 15) and chronic RHD (n = 11). Whether measured in serum or plasma, PTA levels were similar across these groups. Discussion:Our findings conflict with a previous report that found PTA was elevated in RHD compared to healthy controls. In contrast, while less marked than before, we found IgG3 was elevated in ARF, extending this observation to settings where the impact of RHD is greatest.

OBJECTIVES:To describe diagnostic and management characteristics of acute rheumatic fever (ARF) among participants in the 'Searching for a Technology-Driven Acute Rheumatic Fever Test' study, in order to answer clinical questions and determine epidemiological and practice differences in different settings. DESIGN:Multisite, prospective cohort study. SETTING:One hospital in northern Australia and two hospitals in New Zealand, 2018-2021. PARTICIPANTS:143 episodes of definite, probable or possible ARF among 141 participants (median age 10 years, range 5-23; 98% Indigenous). PRIMARY AND SECONDARY OUTCOME MEASURES:Participant characteristics, clinical, biochemical and echocardiographic data were explored using descriptive data. Associations with length of stay were determined using multivariable regression analysis. RESULTS:ARF presentations were heterogeneous with the most common ARF 'phenotype' in 19% of cases being carditis with joint manifestations (polyarthritis, monarthritis or polyarthralgia), fever and PR prolongation. The total proportion of children with carditis was 61%. Australian compared with New Zealand participants more commonly had ARF recurrence (22% vs 0%), underlying RHD (48% vs 0%), possible/probable ARF (23% vs 9%) and were underweight (64% vs 16%). Erythrocyte sedimentation rate (ESR) provided an incremental diagnostic yield of 21% compared with C reactive protein. No instances of RHD were diagnosed among participants in New Zealand. Positive throat Group A Streptococcus culture was more common in New Zealand than in Australian participants (69% vs 3%). Children often required prolonged hospitalisation, with median hospital length-of-stay being 7 days (range 2-66). Significant predictors for length of stay in a multivariable regression model were valve disease (adjusted OR (aOR) 1.56, 95% CI 1.23 to 1.98, p<0.001), requirement for corticosteroids (aOR 1.99, 95% CI 1.22 to 3.26, p=0.007) and higher ESR (aOR 1.4, 95% CI 1.17 to 1.67, p<0.001). CONCLUSIONS:This study provides new knowledge on ARF characteristics and management and highlights international variation in diagnostic and management practice. Differing approaches need to be aligned. Meanwhile, locally specific information can help guide patient expectations after ARF diagnosis.

The successive dominance of SARS-CoV-2 omicron sublineages presents challenges for vaccination strategies with respect to the antigenic content of boosters. New Zealand's COVID-19 elimination strategy (2020-2021) ensured the major vaccination campaign (Pfizer-BioNTech BNT162b2) was completed pre-omicron in an infection-naive population, providing a unique setting to explore the impact of omicron infection waves on vaccine responses. This study compared neutralising antibodies (NAb) to eight SARS-CoV-2 omicron sublineages 28-days and 11-months after a third dose. Participants (n = 219) were classified by antigen exposure in the intervening 10 months including additional vaccinations and/or infections. Both vaccination and infection boosted NAb levels to all sublineages. Antigenic maps showed infection had a major impact on NAb breadth, despite all participants being vaccinated with an ancestral-based vaccine. While vaccination remains an important tool to boost immunity, the breadth of NAbs observed suggest that attempts to match booster specificity with current circulating variants may not always be necessary.

Background Acute rheumatic fever (ARF) is a serious post-infectious sequala of Group A Streptococcus (GAS, Streptococcus pyogenes). In New Zealand (NZ) ARF is a major cause of health inequity. This study describes the genomic analysis of GAS isolates associated with childhood skin and throat infections in Auckland NZ. Methods Isolates (n = 469) collected between March 2018 and October 2019 from the throats and skin of children (5-14 years) underwent whole genomic sequencing. Equal representation across three ethnic groups was ensured through sample quotas with isolates obtained from Indigenous Maori (n = 157, 33%), NZ European/Other (n = 149, 32%) and Pacific Peoples children (n = 163, 35%). Using in silico techniques isolates were classified, assessed for diversity, and examined for distribution differences between groups. Comparisons were also made with GAS strains identified globally. Findings Genomic analysis revealed a diverse population consisting of 65 distinct sequence clusters. These sequence clusters spanned 49 emm-types, with 11 emm-types comprised of several, distinct sequence clusters. There is evidence of multiple global introductions of different lineages into the population, as well as local clonal expansion. The M1UK lineage comprised 35% of all emm1 isolates. Interpretation The GAS population was characterized by a high diversity of strains, resembling patterns observed in low-and middle-income countries. However, strains associated with outbreaks and antimicrobial resistance commonly found in high-income countries were also observed. This unique combination poses challenges for vaccine development, disease management and control.

Knowledge gaps regarding human immunity to Streptococcus pyogenes have impeded vaccine development. To address these gaps and evaluate vaccine candidates, we established a human challenge model of S. pyogenes pharyngitis. Here, we analyse antibody responses in serum and saliva against 19 antigens to identify characteristics distinguishing 19 participants who developed pharyngitis and 6 who did not. We show that pharyngitis elicits serum IgG responses to key vaccine antigens and a muted mucosal IgA response, whereas IgG responses are minimal and IgA responses more pronounced in participants without pharyngitis. Serum IgG responses to pharyngitis in adult participants resemble those in children and are inversely correlated with the magnitude of pre-existing responses. While a straightforward correlate of protection is not evident, baseline antibody signatures distinguish clinical and immunological outcomes following experimental challenge. This highlights the influence of a complex humoral imprint from previous exposure, relevant for interpreting immunogenicity in forthcoming vaccine trials.

Surges in infections caused by respiratory pathogens have been documented in multiple settings after relaxation of pandemic restrictions. Antibodies to major antigens from respiratory syncytial virus and group A Streptococcus waned significantly in a longitudinal adult cohort throughout the pandemic. This waning may have contributed to the pathogen-surges that followed. Antibodies to respiratory syncytial virus and group A Streptococcus waned significantly in a longitudinal cohort throughout the pandemic. This waning may have contributed to the infection surges caused by these pathogens when restrictions were relaxed.

Understanding the immune profile of acute rheumatic fever (ARF), a serious post-infectious sequelae of Streptococcal pyogenes (group A Streptococcus [GAS]), could inform disease pathogenesis and management. Circulating cytokines, immunoglobulins, and complement were analyzed in participants with first- episode ARF, swab-positive GAS pharyngitis and matched healthy controls. A striking elevation of total IgG3 was observed in ARF (90%> clinical reference range for normal). ARF was also associated with an inflammatory triad with significant correlations between interleukin-6, C-reactive protein, and complement C4 absent in controls. Quantification of GAS-specific antibody responses revealed that subclass polarization was remarkably consistent across the disease spectrum; conserved protein antigens polarized to IgG1, while M-protein responses polarized to IgG3 in all groups. However, the magnitude of responses was significantly higher in ARF. Taken together, these findings emphasize the association of exaggerated GAS antibody responses, IgG3, and inflammatory cytokines in ARF and suggest IgG3 testing could beneficially augment clinical diagnosis.

Exposure to Group A Streptococcus leads to a broad spectrum of disease and sequelae, as the bacterium employs a wide range of virulence factors to facilitate colonization of the host, propagation and onward transmission, disrupting both innate and adaptive immune responses. The protease SpyCEP has a crucial role in contributing to bacterial immune evasion by impairing neutrophil recruitment and killing of bacteria through the cleavage of interleukin-8 (IL-8). Given this critical function, SpyCEP represents a key vaccine antigen and quantifying functional anti-SpyCEP antibodies represents not only an important marker of vaccine efficacy, but also a tool to dissect the natural immune response. Here, we report the development and characterization of an IL-8 cleavage inhibition assay to measure the function of anti-SpyCEP antibodies in human sera. The assay was demonstrated to be sensitive, highly specific, linear and reproducible, and suitable for evaluating the function of anti-SpyCEP antibodies induced in humans in vaccine clinical trials and in observational studies of natural immunity.

The high burden of disease and the long-lasting sequelae following Streptococcus pyogenes (Strep A) infections make the development of an effective vaccine a global health priority. Streptolysin O (SLO), is a key toxin in the complex pathogenesis of Strep A infection. Antibodies are elicited against SLO after natural exposure and represent a key target for vaccine-induced immunity. Here we present the setup and characterization of a hemolysis assay to measure functionality of anti-SLO antibodies in human sera. Assay specificity, precision, linearity, reproducibility, and repeatability were determined. The assay was demonstrated to be highly sensitive, specific, reproducible, linear and performed well in assessing functionality of anti-SLO antibodies induced by exposed individuals. Moreover, different sources of critical reagents, in particular red- blood cells, have been compared and had minimal impact on assay performance. The assay presented here has throughput suitable for evaluating sera in vaccine clinical trials and sero-epidemiological studies to gain further insights into the functionality of infection- and vaccine-induced antibodies.

The COVID-19 pandemic has altered the epidemiology of many common childhood infections, including Group A streptococcal (GAS) disease. Acute post-streptococcal glomerulonephritis (APSGN) is a nonsuppurative complication of GAS pharyngitis and pyoderma. It remains the most common cause of pediatric acute glomerulonephritis globally. In Counties Manukau, New Zealand, APSGN rates have previously been shown to be the highest in the country, with marked ethnic and socioeconomic disparities. We performed a retrospective review of children aged 0-14 years who were discharged from Kidz First Hospital, Counties Manukau, between 2015 and 2023 and met the Strep A Vaccine Global Consortium consensus definition of APSGN. We describe a marked, sustained reduction in APSGN hospitalizations, temporally associated with the COVID-19 pandemic. This ongoing reduction in APSGN incidence is notable in light of contrasting reports of increasing incidence of rheumatic fever in New Zealand and invasive GAS disease internationally.

M1UK is associated with current surges in invasive infection globally, partly due to increased production of superantigen streptococcal pyrogenic exotoxin A. We show that M1UK is now the dominant invasive emm1 lineage in Aotearoa New Zealand and is genomically related to community infections, suggesting that measures that effectively prevent group A Streptococcus pharyngitis in children could reduce invasive disease.

This review summarises advances in research from Aotearoa, New Zealand (NZ) that have potential to reduce the inequitable distribution of acute rheumatic fever (ARF) and rheumatic heart disease (RHD). ARF incidence and RHD prevalence are unacceptably inequitable for Maori and Pacifica. Recent qualitative research has demonstrated mismatches between the lived experience of those with ARF/RHD and health service experience they encounter. NZ-led research has contributed knowledge to all stages of disease prevention (primordial, primary and secondary) and for tertiary management. Modifiable risk factors for ARF are racism across health sectors, household crowding, barriers to accessing primary health care, a high intake of sugar-sweetened beverages and preceding sore throat and skin infections. NZ research has evaluated the impact of a large-scale sore throat management programme and Streptococcal A vaccine development. This review highlights two programme domains of research by the authors that have the potential to reduce the burden of chronic RHD: firstly, effective immunomodulation of ARF to reduce the severity of carditis, with current clinical trials of hydroxychloroquine in NZ; secondly, the development of echocardiographic screening of previously undetected RHD. This now meets criteria for an effective screening test and has potential translation for disease control of RHD.

The ability of a third dose of the Pfizer-BioNTech BNT162b2 SARS-CoV-2 vaccine to stimulate immune responses against subvariants, including Omicron BA.1, has not been assessed in New Zealand populations. Unlike many overseas populations, New Zealanders were largely infection naïve at the time they were boosted. This adult cohort of 298 participants, oversampled for at-risk populations, was composed of 29% Māori and 28% Pacific peoples, with 40% of the population aged 55+. A significant proportion of the cohort was obese and presented with at least one comorbidity. Sera were collected 28 days and 6 months post second vaccination and 28 days post third vaccination. SARS-CoV-2 anti-S IgG titres and neutralising capacity using surrogate viral neutralisation assays against variants of concern, including Omicron BA.1, were investigated. The incidence of SARS-CoV-2 infection, within our cohort, prior to third vaccination was very low (<6%). This study found a third vaccine significantly increased the mean SARS-CoV-2 anti-S IgG titres, for every demographic subgroup, by a minimum of 1.5-fold compared to titres after two doses. Diabetic participants experienced a greater increase (∼4-fold) in antibody titres after their third vaccination, compared to non-diabetics (increase of ∼ 2-fold). This corrected for the deficiency in antibody titres within diabetic participants which was observed following two doses. A third dose also induced a neutralising response against Omicron variant BA.1, which was absent after two doses. This neutralising response improved regardless of age, BMI, ethnicity, or diabetes status. Participants aged ≥75 years consistently had the lowest SARS-CoV-2 anti-S IgG titres at each timepoint, however experienced the greatest improvement after three doses compared to younger participants. This study shows that in the absence of prior SARS-CoV-2 infection, a third Pfizer-BioNTech BNT162b2 vaccine enhances immunogenicity, including against Omicron BA.1, in a cohort representative of at-risk groups in the adult New Zealand population.

The data that support the findings of this study are available from the corresponding author upon reasonable request.