Rubella virus seropositivity after infection or vaccination as a risk factor for multiple sclerosis.

BACKGROUND: Multiple sclerosis (MS) is a demyelinating disease affecting millions of people worldwide. Hereditary susceptibility and environmental factors contribute to disease risk. Infection with Epstein-Barr virus (EBV) and human herpesvirus 6A (HHV-6A) have previously been associated with MS risk. Other neurotropic viruses, such as rubella virus (RV), are possible candidates in MS aetiopathogenesis, but previous results are limited and conflicting. METHODS: In this nested case-control study of biobank samples in a Swedish cohort, we analysed the serological response towards RV before the clinical onset of MS with a bead-based multiplex assay in subjects vaccinated and unvaccinated towards RV. The association between RV seropositivity and MS risk was analysed with conditional logistic regression. RESULTS: Seropositivity towards RV was associated with an increased risk of MS for unvaccinated subjects, even when adjusting for plausible confounders including EBV, HHV-6A, cytomegalovirus and vitamin D (adjusted odds ratio [AOR] = 4.0, 95% confidence interval [CI] 1.8-8.8). Cases also had stronger antibody reactivity towards rubella than controls, which was not seen for other neurotropic viruses such as herpes simplex or varicella zoster. Furthermore, we observed an association between RV seropositivity and MS in vaccinated subjects. However, this association was not significant when adjusting for the aforementioned confounders (AOR = 1.7, 95% CI 1.0-2.9). CONCLUSIONS: To our knowledge, these are the first reported associations between early RV seropositivity and later MS development. This suggests a broadening of the virus hypothesis in MS aetiology, where molecular mimicry between rubella epitopes and human central nervous system molecules could be an attractive possible mechanism.

Prospective evaluation of the relevance of Epstein-Barr virus antibodies for early detection of nasopharyngeal carcinoma in Chinese adults.

BACKGROUND: Epstein-Barr virus (EBV) is a major cause of nasopharyngeal carcinoma (NPC) and measurement of different EBV antibodies in blood may improve early detection of NPC. Prospective studies can help assess the roles of different EBV antibodies in predicting NPC risk over time. METHODS: A case-cohort study within the prospective China Kadoorie Biobank of 512 715 adults from 10 (including two NPC endemic) areas included 295 incident NPC cases and 745 subcohort participants. A multiplex serology assay was used to quantify IgA and IgG antibodies against 16 EBV antigens in stored baseline plasma samples. Cox regression was used to estimate adjusted hazard ratios (HRs) for NPC and C-statistics to assess the discriminatory ability of EBV-markers, including two previously identified EBV-marker combinations, for predicting NPC. RESULTS: Sero-positivity for 15 out of 16 EBV-markers was significantly associated with higher NPC risk. Both IgA and IgG antibodies against the same three EBV-markers showed the most extreme HRs, i.e. BGLF2 (IgA: 124.2 (95% CI: 63.3-243.9); IgG: 8.6 (5.5-13.5); LF2: [67.8 (30.0-153.1), 10.9 (7.2-16.4)]); and BFRF1: 26.1 (10.1-67.5), 6.1 (2.7-13.6). Use of a two-marker (i.e. LF2/BGLF2 IgG) and a four-marker (i.e. LF2/BGLF2 IgG and LF2/EA-D IgA) combinations yielded C-statistics of 0.85 and 0.84, respectively, which persisted for at least 5 years after sample collection in both endemic and non-endemic areas. CONCLUSIONS: In Chinese adults, plasma EBV markers strongly predict NPC occurrence many years before clinical diagnosis. LF2 and BGLF2 IgG could identify NPC high-risk individuals to improve NPC early detection in community and clinical settings.

Human papillomavirus-driven head and neck cancers in Japan during 2008-2009 and 2018-2019: The BROADEN study.

There is limited understanding of epidemiology and time trends of human papilloma virus (HPV)-driven head and neck cancers (HNC) in Japan, especially outside of the oropharynx. To assess HPV-driven HNC, a non-interventional study (BROADEN) of HNC patients diagnosed in 2008-2009 and 2018-2019 was conducted in Japan. Adult patients with oropharyngeal, nasopharyngeal, laryngeal, hypopharyngeal or oral cavity cancers were included in this study. HPV was centrally tested using p16INK4a immunohistochemistry, HPV-DNA PCR and HPV E6*I mRNA. HPV attributability required positivity in at least two tests (p16INK4a immunohistochemistry, HPV-DNA PCR, HPV E6*I mRNA) in the oropharynx, and HPV-DNA and HPV E6*I mRNA positivity for non-oropharynx sites. Nineteen hospitals included a total of 1108 patients, of whom 981 had valid samples. Men accounted for 82% of HNC diagnoses. Patients in the earlier cohort were younger and included a higher percentage of smokers. There was an increasing trend of HPV-driven oropharyngeal cancer over the last decade, from 44.2% to 51.7%. HPV attribution in nasopharyngeal cancers was 3.2% in 2008-2009 and 7.5% in 2018-2019; and 4.4% and 0% for larynx respectively. In total, 95.2% of HPV-driven HNC were attributed to HPV genotypes included in the 9-valent HPV vaccine being HPV16 the most prominent genotype. These results suggest that an epidemiologic shift is happening in Japan, with a decrease in smoking and alcohol use and an increase in HPV-driven HNC. The increasing trend of HPV-driven HNC in Japan highlights the need for preventive strategies to mitigate the rise of HPV-driven HNC.

Development and external validation of a head and neck cancer risk prediction model.

BACKGROUND: Head and neck cancer (HNC) incidence is on the rise, often diagnosed at late stage and associated with poor prognoses. Risk prediction tools have a potential role in prevention and early detection. METHODS: The IARC-ARCAGE European case-control study was used as the model development dataset. A clinical HNC risk prediction model using behavioral and demographic predictors was developed via multivariable logistic regression analyses. The model was then externally validated in the UK Biobank cohort. Model performance was tested using discrimination and calibration metrics. RESULTS: 1926 HNC cases and 2043 controls were used for the development of the model. The development dataset model including sociodemographic, smoking, and alcohol variables had moderate discrimination, with an area under curve (AUC) value of 0.75 (95% CI, 0.74-0.77); the calibration slope (0.75) and tests were suggestive of good calibration. 384 616 UK Biobank participants (with 1177 HNC cases) were available for external validation of the model. Upon external validation, the model had an AUC of 0.62 (95% CI, 0.61-0.64). CONCLUSION: We developed and externally validated a HNC risk prediction model using the ARCAGE and UK Biobank studies, respectively. This model had moderate performance in the development population and acceptable performance in the validation dataset. Demographics and risk behaviors are strong predictors of HNC, and this model may be a helpful tool in primary dental care settings to promote prevention and determine recall intervals for dental examination. Future addition of HPV serology or genetic factors could further enhance individual risk prediction.

COVID-19 and long-term impact on symptoms and Health-Related Quality of Life in Costa Rica: the RESPIRA cohort study.

BACKGROUND: Evidence continues to accumulate regarding the potential long-term health consequences of COVID-19 in the population. To distinguish between COVID-19-related symptoms and health limitations from those caused by other conditions, it is essential to compare cases with community controls using prospective data ensuring case-control status. The RESPIRA study addresses this need by investigating the lasting impact of COVID-19 on Health-related Quality of Life (HRQoL) and symptomatology in a population-based cohort in Costa Rica, thereby providing a robust framework for controlling HRQoL and symptoms. METHODS: The study comprised 641 PCR-confirmed, unvaccinated cases of COVID-19 and 947 matched population-based controls. Infection was confirmed using antibody tests on enrollment serum samples and symptoms were monitored monthly for 6 months post-enrolment. Administered at the 6-month visit (occurring between 6- and 2-months post-diagnosis for cases and 6 months after enrollment for controls), HRQoL and Self-Perceived Health Change were assessed using the SF-36, while brain fog, using three items from the Mental Health Inventory (MHI). Regression models were utilized to analyze SF-36, MHI scores, and Self-Perceived Health Change, adjusted for case/control status, severity (mild case, moderate case, hospitalized) and additional independent variables. Sensitivity analyses confirmed the robustness of the findings. RESULTS: Cases showed significantly higher prevalences of joint pain, chest tightness, and skin manifestations, that stabilized at higher frequencies from the fourth month post-diagnosis onwards (2.0%, 1.2%, and 0.8% respectively) compared to controls (0.9%, 0.4%, 0.2% respectively). Cases also exhibited significantly lower HRQoL than controls across all dimensions in the fully adjusted model, with a 12.4 percentage-point difference [95%CI: 9.4-14.6], in self-reported health compared to one year prior. Cases reported 8.0% [95%CI: 4.2, 11.5] more physical limitations, 7.3% [95%CI: 3.5, 10.5] increased lack of vitality, and 6.0% [95%CI: 2.4, 9.0] more brain fog compared to controls with similar characteristics. Undiagnosed cases detected with antibody tests among controls had HRQoL comparable to antibody negative controls. Differences were more pronounced in individuals with moderate or severe disease and among women. CONCLUSIONS: PCR-confirmed unvaccinated cases experienced prolonged HRQoL reductions 6 months to 2 years after diagnosis, this was particularly the case in severe cases and among women. Mildly symptomatic cases showed no significant long-term sequelae.

Kinetics of EBV antibody-based NPC risk scores in Taiwan NPC multiplex families.

Nasopharyngeal carcinoma (NPC) risk prediction models based on Epstein-Barr virus (EBV)-antibody testing have shown potential for screening of NPC; however, the long-term stability is unclear. Here, we investigated the kinetics of two EBV-antibody NPC risk scores within the Taiwan NPC Multiplex Family Study. Among 545 participants with multiple blood samples, we evaluated the stability of a 2-marker enzyme-linked immunosorbent assay score and 13-marker multiplex serology score using the intra-class correlation coefficient (ICC) by fitting a linear mixed model that accounted for the clustering effect of multiple measurements per subject and age. We also estimated the clustering of positive tests using Fleiss's kappa statistic. Over an average 20-year follow-up, the 2-marker score showed high stability over time, whereas the 13-marker score was more variable (p < .05). Case-control status is associated with the kinetics of the antibody response, with higher ICCs among cases. Positive tests were more likely to cluster within the same individual for the 2-marker score than the 13-marker score (p < .05). The 2-marker score had an increase in specificity from ~90% for single measurement to ~96% with repeat testing. The 13-marker score had a specificity of ~73% for a single measurement that increased to ~92% with repeat testing. Among individuals who developed NPC, none experienced score reversion. Our findings suggest that repeated testing could improve the specificity of NPC screening in high-risk NPC multiplex families. Further studies are required to determine the impact on sensitivity, establish optimal screening intervals, and generalize these findings to general population settings in high-risk regions.

Quantification of human papillomavirus cell-free DNA from low-volume blood plasma samples by digital PCR.

The incidence rate of human papillomavirus-driven oropharyngeal cancer (HPV-OPC) is increasing in countries with high human development index. HPV cell-free DNA (cfDNA) isolated from 3 to 4 mL blood plasma has been successfully used for therapy surveillance. A highly discussed application of HPV-cfDNA is early detection of HPV-OPC. This requires sensitive and specific cfDNA detection as cfDNA levels can be very low. To study the predictive power of pre-diagnostic HPV-cfDNA, archived samples from epidemiological cohorts with limited plasma volume are an important source. To establish a cfDNA detection workflow for low plasma volumes, we compared cfDNA purification methods [MagNA Pure 96 (MP96) and QIAamp ccfDNA/RNA] and digital PCR systems (Biorad QX200 and QIAGEN QIAcuity One). Final assay validation included 65 low-volume plasma samples from oropharyngeal cancer (OPC) patients with defined HPV status stored for 2-9 years. MP96 yielded a 28% higher cfDNA isolation efficiency in comparison to QIAamp. Both digital PCR systems showed comparable analytical sensitivity (6-17 copies for HPV16 and HPV33), but QIAcuity detected both types in the same assay. In the validation set, the assay had 80% sensitivity (n = 28/35) for HPV16 and HPV33 and a specificity of 97% (n = 29/30). In samples with ≥750 µL plasma, the sensitivity was 85% (n = 17/20), while in samples with <750 µL plasma, it was 73% (n = 11/15). Despite the expected drop in sensitivity with decreased plasma volume, the assay is sensitive and highly specific even in low-volume samples and thus suited for studies exploring HPV-cfDNA as an early HPV-OPC detection marker in low-volume archival material.IMPORTANCEHPV-OPC has a favorable prognosis compared to HPV-negative OPC. However, the majority of tumors is diagnosed after regional spread, thus making intensive treatment necessary. This can cause lasting morbidity with a large impact on quality of life. One potential method to decrease treatment-related morbidity is early detection of the cancer. HPV cfDNA has been successfully used for therapy surveillance and has also been detected in pre-diagnostic samples of HPV-OPC patients. These pre-diagnostic samples are only commonly available from biobanks, which usually only have small volumes of blood plasma available. Hence, we have developed a workflow optimized for small-volume archival samples. With this method, a high sensitivity can be achieved despite sample limitations, making it suitable to conduct further large-scale biobank studies of HPV-cfDNA as a prognostic biomarker for HPV-OPC.

Interactions between High Seroreactivity to Human Herpes Virus 6A and Epstein-Barr Virus in MS Development: A Presymptomatic Case-Control Study.

Synergistic interactions between human herpesvirus 6A (HHV-6A) and Epstein-Barr virus (EBV) are hypothesized in the etiopathogenesis of multiple sclerosis (MS). This study investigated if HHV-6A and EBV seroreactivities interact regarding the risk of developing MS. Antibodies against viral antigens were analyzed in biobank samples from 670 individuals who later developed MS and matched controls. Additive interactions were analyzed. A significant interaction between HHV-6A and EBNA-1 seroreactivities was observed in study participants above the median age of 24.9 years (attributable proportion due to interaction = 0.45). This finding supports the hypothesis that HHV-6A and EBV infections interact in MS development. ANN NEUROL 2024;96:302-305.

Serum IgG antibodies to HPV6 L1, E2, E4, E6, and E7 proteins among children prospectively followed-up for three years.

BACKGROUND: Current knowledge implicates that human papillomavirus (HPV) infection can be acquired at early age. However, the role of HPV-specific passive immunization from mother to neonate is nearly unexplored, especially against the HPV early proteins. We analyzed IgG antibodies against HPV6 early (E2, E4, E6, E7) and late (L1) proteins in children prospectively followed-up for three years. METHODS: A total of 272 children and their mothers from the Finnish Family HPV Study were included in these analyses. Serum samples were obtained from pregnant mothers at their third trimester and from newborn/infants at 1-, 2-, 6-, 12-, 24-, and 36-month visits after birth. Antibodies to the early and late proteins were analyzed by multiplex serology based on glutathione S-transferase fusion protein capture to fluorescent beads. RESULTS: Maternal antibodies to all tested HPV6 proteins were transferred to neonates, concordance between maternal and neonates' antibody levels being highly significant (p<0.001). Seropositivity of HPV6 L1 in the neonates declined during the first six months of life, whereas changes in the E-protein antibodies were less obvious. After the maternal antibodies have vanished, seroconversion to HPV6 L1 at 12 months (median) and to the HPV6 E-proteins between 23-35 months was observed. CONCLUSION: IgG antibodies against HPV6 E- and L-proteins are transferred from mothers to their children. Seroconversion against HPV6 L1, E2, E4, E6, and E7 does occur in early childhood, as a sign of acquired HPV6 infection by vertical or horizontal transmission starting at 12 months of age.

Empirical sample-specific approaches to define HPV16 and HPV18 seropositivity in unvaccinated, young, sexually active women.

Given low seroconversion rates following human papillomavirus (HPV) infection, fixed external cutoffs may lead to errors in estimating HPV seroprevalence. We evaluated finite mixture modeling (FMM) and group-based trajectory modeling (GBTM) among unvaccinated, sexually active, HPV-exposed women to determine study-specific HPV16 and HPV18 seropositivity thresholds. We included 399 women (aged 18-24 years) enrolled in the HPV Infection and Transmission Among Couples Through Heterosexual Activity (HITCH) cohort study between 2005 and 2011 in Montreal, Canada. Participants' blood samples from up to six visits spanning 2 years were tested by multiplex serology for antibodies [median fluorescence intensity (MFI)] specific to bacterially expressed HPV16 and HPV18 L1 glutathione S-transferase fusion proteins. We applied FMM and GBTM to baseline and longitudinal antibody titer measurements, respectively, to define HPV type-specific seronegative and seropositive distributions. Study-specific thresholds were generated as five standard deviations above the mean seronegative antibody titers, mimicking cutoffs (HPV16: 422 MFI; HPV18: 394 MFI) derived from an external population of sexually inactive, HPV DNA-negative Korean women (aged 15-29 years). Agreement (kappa) of study-specific thresholds was evaluated against external cutoffs. Seroprevalence estimates using FMM (HPV16: 27.5%-43.2%; HPV18: 21.7%-49.5%) and GBTM (HPV16: 11.8%-11.8%; HPV18: 9.9%-13.4%) thresholds exceeded those of external cutoffs (HPV: 10.2%; HPV18: 9.7%). FMM thresholds showed slight-to-moderate agreement with external cutoffs (HPV16: 0.26%-0.46%; HPV18: 0.20%-0.56%), while GBTM thresholds exhibited high agreement (HPV16: 0.92%-0.92%; HPV18: 0.82%-0.99%). Kappa values suggest that GBTM, used for longitudinal serological data, and otherwise FMM, for cross-sectional data, are robust methods for determining the HPV serostatus without prior classification rules.IMPORTANCEWhile human papillomavirus (HPV) seropositivity has been employed as an epidemiologic determinant of the natural history of genital HPV infections, only a fraction of women incidentally infected with HPV respond by developing significant antibody levels. HPV seropositivity is often determined by a dichotomous fixed cutoff based on the seroreactivity of an external population of women presumed as seronegative, given the lack of evidence of HPV exposure. However, considering the variable nature of seroreactivity upon HPV infection, which arguably varies across populations, such externally defined cutoffs may lack specificity to the population of interest, causing inappropriate assessment of HPV seroprevalence and related epidemiologic uses of that information. This study demonstrates that finite mixture modeling (FMM) and group-based trajectory modeling (GBTM) can be used to independently estimate seroprevalence or serve as the basis for defining study-specific seropositivity thresholds without requiring prior subjective assumptions, consequently providing a more apt internally valid discrimination of seropositive from seronegative individuals.

Antibodies against high-risk human papillomavirus proteins as markers for noncervical HPV-related cancers in a Black South African population, according to HIV status.

Human papillomavirus (HPV) proteins may elicit antibody responses in the process toward HPV-related malignancy. However, HPV seroepidemiology in noncervical HPV-related cancers remains poorly understood, particularly in populations with a high prevalence of human immunodeficiency virus (HIV). Using a glutathione S-transferase-based multiplex serology assay, antibodies against E6, E7 and L1 proteins of HPV16 and HPV18 were measured in sera of 535 cases of noncervical HPV-related cancers (anal (n = 104), vulval (n = 211), vaginal (n = 49), penile (n = 37) and oropharyngeal (n = 134)) and 6651 non-infection-related cancer controls, from the Johannesburg Cancer Study that recruited Black South African with newly diagnosed cancer between 1995 and 2016. Logistic and Poisson regression models were used to calculate adjusted odds ratios (aOR) and prevalence ratios (aPR) and 95% confidence intervals (CI) in cases versus controls. HPV16 E6 was more strongly associated with noncervical HPV-related cancers than HPV16 L1 or E7, or HPV18 proteins: anal (females (HPV16 E6 aOR = 11.50;95%CI:6.0-22.2), males (aOR = 10.12;95%CI:4.9-20.8), vulval (aOR = 11.69;95%CI:7.9-17.2), vaginal (aOR = 10.26;95%CI:5.0-21), penile (aOR = 18.95;95%CI:8.9-40), and oropharyngeal (females (aOR = 8.95;95%CI:2.9-27.5), males (aOR = 3.49;95%CI:1.8-7.0)) cancers. HPV16-E6 seropositivity ranged from 24.0% to 35.1% in anal, vulval, vaginal and penile cancer but was significantly lower (11.2%) in oropharyngeal cancer. After adjustment for HIV, prevalence of which increased from 22.2% in 1995-2005 to 54.1% in 2010-2016, HPV16 E6 seropositivity increased by period of diagnosis (aPR for 2010-2016 vs. 1995-2006 = 1.84;95%CI:1.1-3.0). Assuming HPV16 E6 seroprevalence reflects HPV attributable fraction, the proportion of certain noncervical-HPV-related cancers caused by HPV is increasing over time in South Africa. This is expected to be driven by the increasing influence of HIV.

Genetics of immune response to Epstein-Barr virus: prospects for multiple sclerosis pathogenesis.

Epstein-Barr virus (EBV) infection has been advocated as a prerequisite for developing multiple sclerosis (MS) and possibly the propagation of the disease. However, the precise mechanisms for such influences are still unclear. A large-scale study investigating the host genetics of EBV serology and related clinical manifestations, such as infectious mononucleosis (IM), may help us better understand the role of EBV in MS pathogenesis. This study evaluates the host genetic factors that influence serological response against EBV and history of IM and cross-evaluates them with MS risk and genetic susceptibility in the Swedish population. Plasma IgG antibody levels against EBV nuclear antigen-1 (EBNA-1, truncated=aa[325-641], peptide=aa[385-420]) and viral capsid antigen p18 (VCAp18) were measured using bead-based multiplex serology for 8744 MS cases and 7229 population-matched controls. The MS risk association for high/low EBV antibody levels and history of IM was compared to relevant clinical measures along with sex, age at sampling, and associated HLA allele variants. Genome-wide and HLA allele association analyses were also performed to identify genetic risk factors for EBV antibody response and IM history. Higher antibody levels against VCAp18 (OR=1.74, 95% CI=1.60-1.88) and EBNA-1, particularly the peptide (OR=3.13, 95% CI=2.93-3.35), were associated with an increased risk for MS. The risk increased with higher anti-EBNA-1 IgG levels up to twelve times the reference risk. We also identified several independent HLA haplotypes associated with EBV serology overlapping with known MS risk alleles (e.g., DRB1*15:01). Although there were several candidates, no variants outside the HLA region reached genome-wide significance. Cumulative HLA risk for anti-EBNA-1 IgG levels, particularly the peptide fragment, was strongly associated with MS. In contrast, the genetic risk for high anti-VCAp18 IgG levels was not as strongly associated with MS risk. IM history was not associated with class II HLA genes but negatively associated with A*02:01, which is protective against MS. Our findings emphasize that the risk association between anti-EBNA-1 IgG levels and MS may be partly due to overlapping HLA associations. Additionally, the increasing MS risk with increasing anti-EBNA-1 levels would be consistent with a pathogenic role of the EBNA-1 immune response, perhaps through molecular mimicry. Given that high anti-EBNA-1 antibodies may reflect a poorly controlled T-cell defense against the virus, our findings would be consistent with DRB1*15:01 being a poor class II antigen in the immune defense against EBV. Lastly, the difference in genetic control of IM supports the independent roles of EBNA-1 and IM in MS susceptibility.

Seroprevalence of sexually transmitted infections over 44 years - A cross-sectional study in Sweden.

BACKGROUND: Sexually transmitted infections (STIs) may cause substantial individual suffering and a large economic burden for society. This study examined the seroprevalence of Chlamydia trachomatis, Mycoplasma genitalium, herpes simplex virus (HSV) types 1 and 2, and several human papillomaviruses (HPV) in the Swedish population over time. METHODS: The study population consisted of 30-year-old women attending maternity care, and 50 year-old men and women attending health check-ups, from 1975 to 2018. Antibody status was determined by multiplex serology and quantified using median reporter fluorescence intensity (MFI). RESULTS: A total of 891 samples were analysed (519 from 30-year-old women, 186 from 50 year-old women and 186 from 50 year-old men). Of these, 41.5% showed seropositivity for Chlamydia trachomatis, 16.7% for Mycoplasma genitalium, 70.5% for HSV-1, 14.9% for HSV-2, 13.2% for high-risk HPV, and 8.3% for low-risk HPV. Seropositivity for Mycoplasma genitalium, HSV-1 and especially Chlamydia trachomatis decreased over time. CONCLUSIONS: There was a decrease over time in Chlamydia trachomatis seroprevalence, probably due to contact tracing, testing and early treatment; this might also have affected Mycoplasma genitalium seroprevalence. Despite the reduction, seroprevalences are still high, so continued and new efforts to reduce STI incidence are essential.

[Update: Epidemiology and Prevention of Oropharyngeal Cancer]. / Update: Epidemiologie und Prävention des Oropharynxkarzinoms.

Due to the association with the causal HPV-16 infection, the oropharyngeal carcinoma spreads into two separate entities depending on HPV-16 positivity. More recent data show a diversified picture of the importance and prevalence of the surrogate parameter p16 (discordance) for a definitive HPV-16 association, which varies worldwide. In the context of prevention options, vaccination is of major and HPV screening of healthy people only of little importance.

Helper T cell immunity in humans with inherited CD4 deficiency.

CD4+ T cells are vital for host defense and immune regulation. However, the fundamental role of CD4 itself remains enigmatic. We report seven patients aged 5-61 years from five families of four ancestries with autosomal recessive CD4 deficiency and a range of infections, including recalcitrant warts and Whipple's disease. All patients are homozygous for rare deleterious CD4 variants impacting expression of the canonical CD4 isoform. A shorter expressed isoform that interacts with LCK, but not HLA class II, is affected by only one variant. All patients lack CD4+ T cells and have increased numbers of TCRαß+CD4-CD8- T cells, which phenotypically and transcriptionally resemble conventional Th cells. Finally, patient CD4-CD8- αß T cells exhibit intact responses to HLA class II-restricted antigens and promote B cell differentiation in vitro. Thus, compensatory development of Th cells enables patients with inherited CD4 deficiency to acquire effective cellular and humoral immunity against an unexpectedly large range of pathogens. Nevertheless, CD4 is indispensable for protective immunity against at least human papillomaviruses and Trophyrema whipplei.

Assessing the Potential of HPV16 E6 Seroprevalence as a Biomarker for Anal Dysplasia and Cancer Screening-A Systematic Review and Meta-Analysis.

Elevated rates of human papillomavirus (HPV)-related anal high-grade squamous intraepithelial lesions (HSIL) and anal cancer (AC) in populations like men who have sex with men (MSM) living with HIV underscore the need for effective screening. While high-resolution anoscopy-guided biopsy is the gold standard, limited provider availability poses a challenge. This has spurred interest in identifying biomarkers for improved AC prevention. Antibodies against HPV16 oncoprotein E6, known as markers for cervical and oropharyngeal cancers, are the focus of the current study. The systematic review and meta-analysis included six studies meeting inclusion criteria, assessing HPV16 E6 seroprevalence in individuals with anal HSIL or AC. A two-step meta-analysis estimated pooled odds ratios and 95% confidence intervals (CI) for HPV16 E6 seroprevalence and HSIL or AC. Pooled prevalence, sensitivity, specificity, and diagnostic odds ratios were also calculated. This meta-analysis revealed a 3.6-fold increased risk of HSIL for HPV16 E6 seropositive individuals, escalating to a 26.1-fold risk increase for AC. Pooled specificity and sensitivity indicated a high specificity (0.99; 95%CI: 0.99, 0.99) but lower sensitivity (0.19; 95%CI: 0.10, 0.34) for HPV16 E6 serostatus as an AC biomarker. In conclusion, while HPV16 E6 seroprevalence demonstrates specificity as a potential biomarker for HPV-related AC, its utility as a standalone screening tool may be limited. Instead, it could serve effectively as a confirmation test, particularly in high-risk populations, alongside other diagnostic methods. Further research is imperative to explore HPV16 E6 seroconversion dynamics and alternative screening algorithms.

Impact of Different Cofactors on Naturally Acquired Human Papillomavirus Antibody Levels Among Unvaccinated Pregnant Women.

Human papillomavirus (HPV) infections are common, transmitted by sexual and nonsexual routes. The present case-control setting was designed to examine potential cofactors associated with either persistently low or high HPV-antibody levels. The study subjects were from the Finnish HPV Family cohort of 329 baseline pregnant, non-HPV-vaccinated women, who were sampled for genital and oral HPV-DNA and HPV serology at baseline, and at 12, 24, and 36 months. Antibodies to the L1 major capsid protein of HPV 6, 11, 16, 18, and 45 were analyzed by multiplex HPV serology and HPV genotyping was performed. This study included 59 women, 23 women with persistently low (<200 median fluorescence intensity [MFI]) and 36 women with persistently high and always positive (>200 MFI) levels of these antibodies for all five HPV genotypes. Potential HPV-associated covariates were derived from detailed questionnaires. Only cofactors other than detected HPV genotype significantly impact on the levels of natural HPV antibodies. A higher number of past sexual partners or a history of diagnosed genital warts were significant covariates of high HPV antibody levels (p = 0.023 and p = 0.043, respectively). Of interest, women with a history of allergies presented with low levels of HPV antibodies (p = 0.03), potentially exposing these women to an increased risk of future HPV-related diseases that merit closer surveillance.

Advancing Toxoplasma gondii multiplex serology.

Toxoplasma gondii is a highly prevalent pathogen causing zoonotic infections with significant public health implications. Yet, our understanding of long-term consequences, associated risk factors, and the potential role of co-infections is still limited. Seroepidemiological studies are a valuable approach to address open questions and enhance our insights into T. gondii across human populations. Here, we present substantial advancements to our previously developed T. gondii multiplex serology assay, which is based on the immunodominant antigens SAG1 and P22. While our previous bead-based assay quantified antibody levels against multiple targets in a high-throughput fashion requiring only a small sample volume, impaired assay characteristics emerged in sample dilutions beyond 1:100 and when being transferred to magnetic beads. Both are now critical for inclusion in large-scale seroprevalence studies. Using the truncated versions, SAG1D1 and P22trunc, significantly enhanced signal-to-noise ratios were achieved with almost perfect concordance with the gold-standard Sabin-Feldman dye test. In sample dilutions of 1:100, the diagnostic accuracy of SAG1D1 and P22trunc reached sensitivities (true positive rates) of 98% and 94% and specificities (true negative rates) of 93% and 95%, respectively. Importantly, performance metrics were reproducible in a 1:1,000 sample dilution, using both magnetic and nonmagnetic beads. Thresholds for seropositivity were derived from finite mixture models and performed equally well as thresholds by receiver operating characteristic analysis. Our improved multiplex serology assay is therefore able to generate robust and reproducible performance metrics under various assay conditions. Inclusion of T. gondii antibody measurements with other pathogens, in multiplex serology panels will allow for large-scale seroepidemiological research. IMPORTANCE: Toxoplasma gondii is a pathogen of significant public health concern due to its widespread prevalence and zoonotic potential. However, our understanding of key aspects, such as risk factors for infection and disease, potential outcomes, and their trends, remains limited. Seroepidemiological studies in large cohorts are invaluable for addressing these questions but remain scarce. Our revised multiplex serology assay equips researchers with a powerful tool capable of delivering T. gondii serum antibody measurements with high sensitivity and specificity under diverse assay conditions. This advancement paves the way for the integration of T. gondii antibody measurements into multi-pathogen multiplex serology panels, promising valuable insights into public health and pathogen interactions.

Assessing the feasibility of a multimodal liquid biopsy for the diagnosis of HPV-associated oropharyngeal squamous cell carcinoma.

OBJECTIVES: In this feasibility study, we explored the combined use of circulating tumor human papillomavirus (HPV) DNA (ctHPVDNA) and HPV serology as diagnostic tests for HPV-associated oropharyngeal squamous cell carcinoma (OPSCC). METHODS: Among patients with research-banked serum or plasma at diagnosis, IgG antibodies to oncoproteins from HPV types 16, 18, 31, 33, 35, 45, 52, and 58 were detected with multiplex serology. Positivity for HPV 16 was defined based on detection of combinations of anti-E6, E1, E2, and E7 and for other high-risk types on detection of anti-E6 and anti-E7. Circulating tumor HPV DNA was detected by custom digital droplet polymerase chain reaction (ddPCR) assays for HPV types 16, 18, 33, 35, and 45. p16 immunohistochemistry and high-risk HPV RNA in situ hybridization (ISH) using a cocktail of 18 high-risk HPV types were performed on tissue. RESULTS: Of 75 patients, 67 (89.3%) were HPV-associated (p16 and HPV RNA ISH positive) and 8 (10.7%) were HPV-independent. All 8 HPV-independent patients were seronegative and negative for ctHPVDNA (100% specificity). Serology was positive in 53 (79.1%) of 67 HPV-associated patients, while ddPCR was positive for ctHPVDNA in 59 (88.6%) of 67 HPV-associated patients. Requiring both tests to be positive resulted in a sensitivity of 50 (74.6%) of 67 while combining assays (either positive) improved sensitivity to 62 (92.6%) of 67. CONCLUSIONS: Compared to HPV RNA ISH, HPV serology and ctHPVDNA are sensitive and highly specific biomarkers for HPV-associated OPSCC at the time of presentation.

Blood-based screening for HPV-associated cancers.

Background: HPV-associated oropharyngeal cancer (HPV+OPSCC) is the most common HPV-associated cancer in the United States yet unlike cervical cancer lacks a screening test. HPV+OPSCCs are presumed to start developing 10-15 years prior to clinical diagnosis. Circulating tumor HPV DNA (ctHPVDNA) is a sensitive and specific biomarker for HPV+OPSCC. Taken together, blood-based screening for HPV+OPSCC may be feasible years prior to diagnosis. Methods: We developed an HPV whole genome sequencing assay, HPV-DeepSeek, with 99% sensitivity and specificity at clinical diagnosis. 28 plasma samples from HPV+OPSCC patients collected 1.3-10.8 years prior to diagnosis along with 1:1 age and gender-matched controls were run on HPV-DeepSeek and an HPV serology assay. Results: 22/28 (79%) of cases and 0/28 controls screened positive for HPV+OPSCC with 100% detection within four years of diagnosis and a maximum lead time of 7.8 years. We next applied a machine learning model classifying 27/28 cases (96%) with 100% detection within 10 years. Plasma-based PIK3CA gene mutations, viral genome integration events and HPV serology were used to orthogonally validate cancer detection with 68% (19/28) of the cohort having multiple cancer signals detected. Molecular fingerprinting of HPV genomes was performed across patients demonstrating that each viral genome was unique, ruling out contamination. In patients with tumor blocks from diagnosis (15/28), molecular fingerprinting was performed within patients confirming the same viral genome across time. Conclusions: We demonstrate accurate blood-based detection of HPV-associated cancers with lead times up to 10 years before clinical cancer diagnosis and in doing so, highlight the enormous potential of ctDNA-based cancer screening.