Measles Foci Reduction Neutralization Test (FRNT).

The plaque reduction neutralization test (PRNT) and the enzyme-linked immunosorbent assay (ELISA) are both widely used to assess immunity to infectious diseases such as measles, but they use two different measurement principles: ELISA measures the ability of antibodies to bind to virus components, while the PRNT detects the aptitude of antibodies to prevent the infection of a susceptible cell. As a result, detection of measles virus (MV) neutralizing antibodies is the gold standard for assessing immunity to measles. However, the assay is laborious and requires experience and excellent technical skills. In addition, the result is only available after several days. Therefore, the classical PRNT is not suitable for high-throughput testing. By using an immunocolorimetric assay (ICA) to detect MV-infected cells, the standard PRNT has been developed into a focus reduction neutralization test (FRNT). This assay is faster and has improved specificity. The FRNT described here is extremely useful when immunity to measles virus needs to be assessed in patients with a specific medical condition, such as immunocompromised individuals in whom presumed residual immunity needs to be assessed. The FRNT is not generally recommended for use with large numbers of specimens, such as in a seroprevalence study.

Measles vaccination - An underestimated prevention measure: Analyzing a fatal case in Hildesheim, Germany.

Measles and rubella are targeted for elimination in the WHO region Europe. To reach the elimination goal, vaccination coverage of 95% must be achieved and sustained, the genotype information has to be provided for 80% of all outbreaks and transmission chains of a certain variant must not be detected for >12 months. The latter information is collected at Germany's National Reference Center Measles, Mumps, Rubella (NRC MMR). We describe here an outbreak of measles occurring in Hildesheim. The outbreak comprised 43 cases and lasted 14 weeks. Surprisingly, a high number of vaccination failures was observed since 11 cases had received two doses of the MMR vaccine and 4 additional cases were vaccinated once. A 33-year-old woman passed away during the outbreak. She was the mother of 5 children between 4 and 16 years of age. Two schoolchildren contracted measles and passed it on to the rest of the family. Due to delivery bottlenecks, the vaccination of the mother was delayed. She developed measles-like symptoms 3 days after vaccination and was found dead on the morning of day 8 after vaccination. A post-mortem examination was done to identify the cause of death. Moreover, molecular characterization of the virus was performed to analyze whether she was infected by the wildtype virus circulating at that time in Hildesheim or whether the vaccine may have been a concomitant and aggravating feature of her death. The result showed that the samples taken from her at the time of death and during necropsy contained the wildtype measles virus variant corresponding to MVs/Gir Somnath.IND/42.16 (WHO Seq-ID D8-4683) that fueled the Hildesheim outbreak and circulated in Germany from March 2018 to March 2020. The vaccine virus was not detected. Moreover, two aspects uncovered by the post-mortem examination were remarkable; the woman died from giant cell pneumonia, which is a complication seen in immune-suppressed individuals and she was actively using cannabis. THC is known to influence the immune system, but literature reports describing the effects are limited.

Vaccine-induced neutralizing antibodies bind to the H protein of a historical measles virus.

Measles is a highly contagious airborne viral disease. It can lead to serious complications and death and is preventable by vaccination. The live-attenuated measles vaccine (LAMV) derived from a measles virus (MV) isolated in 1954 has been in use globally for six decades and protects effectively by providing a durable humoral and cell-mediated immunity. Our study addresses the temporal stability of epitopes on the viral surface glycoprotein hemagglutinin (H) which is the major target of MV-neutralizing antibodies. We investigated the binding of seven vaccine-induced MV-H-specific monoclonal antibodies (mAbs) to cell-free synthesized MV-H proteins derived from the H gene sequences obtained from a lung specimen of a fatal case of measles pneumonia in 1912 and an isolate from a current case. The binding of four out of seven mAbs to the H protein of both MV strains provides evidence of epitopes that are stable for more than 100 years. The binding of the universally neutralizing mAbs RKI-MV-12b and RKI-MV-34c to the H protein of the 1912 MV suggests the long-term stability of highly conserved epitopes on the MV surface.

A bead-based multiplex assay covering all coronaviruses pathogenic for humans for sensitive and specific surveillance of SARS-CoV-2 humoral immunity.

Serological assays measuring antibodies against SARS-CoV-2 are key to describe the epidemiology, pathobiology or induction of immunity after infection or vaccination. Of those, multiplex assays targeting multiple antigens are especially helpful as closely related coronaviruses or other antigens can be analysed simultaneously from small sample volumes, hereby shedding light on patterns in the immune response that would otherwise remain undetected. We established a bead-based 17-plex assay detecting antibodies targeting antigens from all coronaviruses pathogenic for humans: SARS-CoV-2, SARS-CoV, MERS-CoV, HCoV strains 229E, OC43, HKU1, and NL63. The assay was validated against five commercial serological immunoassays, a commercial surrogate virus neutralisation test, and a virus neutralisation assay, all targeting SARS-CoV-2. It was found to be highly versatile as shown by antibody detection from both serum and dried blot spots and as shown in three case studies. First, we followed seroconversion for all four endemic HCoV strains and SARS-CoV-2 in an outbreak study in day-care centres for children. Second, we were able to link a more severe clinical course to a stronger IgG response with this 17-plex-assay, which was IgG1 and IgG3 dominated. Finally, our assay was able to discriminate recent from previous SARS-CoV-2 infections by calculating the IgG/IgM ratio on the N antigen targeting antibodies. In conclusion, due to the comprehensive method comparison, thorough validation, and the proven versatility, our multiplex assay is a valuable tool for studies on coronavirus serology.

Acute orchitis deciphered: Coxsackievirus B strains are the main etiology and their presence in semen is associated with acute inflammation and risk of persistent oligozoospermia.

Although various viruses are considered to be the clinical cause for acute orchitis, it is completely unclear to what extent and which viruses are etiologically involved in acute orchitis and what the clinic and course of these patients are like. Therefore, a prospective study was set up to decipher acute isolated orchitis. Between July 2007 and February 2023, a total of 26 patients with isolated orchitis were recruited and compared with 530 patients with acute epididymitis. We were able to show for isolated orchitis, that (1) orchitis is usually of viral origin (20/26, 77%) and enteroviruses with coxsackievirus B strains (16/26, 62%) are predominant, (2) virus isolates could be received from semen indicating the presence of replication-competent virus particles, (3) a polymerase chain reaction (PCR) for enteroviruses should be conducted using semen provided at the onset of disease, because the virus is not detectable in serum/urine, (4) there is a circannual occurrence with the maximum in summer, (5) orchitis is associated with a characteristic inflammatory cytokine panel in the semen and systemic inflammation, (6) orchitis is usually rapidly self-limiting, and (7) about 30% of patients (6/20) suffer ongoing oligozoospermia. These seven emerging aspects are likely to fundamentally change thinking and clinical practice regarding acute isolated orchitis.

Detection of measles virus in Bulgaria from 2012 to 2018.

AIM: To determine the circulation patterns of measles virus in Bulgaria from 2012 to 2018 after a large measles outbreak in the country (2009-2011). METHODS: Three types of clinical material were collected: serum samples, urine samples, and nasal swabs. Enzyme-linked immunosorbent assay (ELISA) was used to detect specific viral immunoglobulin (Ig) M/IgG antibodies. Viral RNA was extracted from all urine and nasal swabs. RESULTS: In the investigated period, 102 patients were confirmed to have measles (age range: two months to 55 years). A total of 101 samples (99%) were measles-IgM positive. Most of them were detected in 2017 (73%, 74/101), when a measles outbreak in the country was reported. The majority of patients were unvaccinated children aged under 13 months. Out of 101 measles serum samples confirmed by ELISA, 18 (20.45%) were measles-IgG positive and 15 (17.05%) were borderline. Thirty-three positive PCR products were sequenced and genotyped. In 2013, 2016, 2017, and 2018, three different measles viral genotypes were detected: D8, H1, and B3. Most patients were unvaccinated or insufficiently vaccinated. CONCLUSION: Preventive measures are indispensable to limit the infection in different regions of Bulgaria and its spread to other countries. As vaccination coverage against measles and other vaccine-preventable infections, including SARS-Co2, is low, it is necessary to perform molecular identification of viruses to monitor their circulation and pathogenicity.

Corrigendum: Specifically increased rate of infections in children post measles in a high resource setting.

[This corrects the article DOI: 10.3389/fped.2022.896086.].

Specifically Increased Rate of Infections in Children Post Measles in a High Resource Setting.

Objectives: Post-measles increased susceptibility to subsequent infections seems particularly relevant in low-resource settings. We tested the hypothesis that measles causes a specifically increased rate of infections in children, also in a high-resource setting. Methods: We conducted a retrospective cohort study on a large measles outbreak in Berlin, Germany. All children with measles who presented to hospitals in Berlin were included as cases, children with non-infectious and children with non-measles infectious diseases as controls. Repeat visits within 3 years after the outbreak were recorded. Results: We included 250 cases, 502 non-infectious, and 498 infectious disease controls. The relative risk for cases for the diagnosis of an infectious disease upon a repeat visit was 1.6 (95% CI 1.4-2.0, p < 0.001) vs. non-infectious and 1.3 (95% CI 1.1-1.6, p = 0.002) vs. infectious disease controls. 33 cases (27%), 35 non-infectious (12%) and 57 (18%) infectious disease controls presented more than three times due to an infectious disease (p = 0.01, and p = 0.02, respectively). This results in a relative risk of more than three repeat visits due to an infection for measles cases of 1.8 (95% CI 1.3-2.4, p = 0.01), and 1.4 (95% CI 1.0-1.9, p = 0.04), respectively. Conclusion: Our study demonstrates for the first time in a high-resource setting, that increased post-measles susceptibility to subsequent infections in children is measles-specific-even compared to controls with previous non-measles infections.

ICTV Virus Taxonomy Profile: Matonaviridae 2022.

The family Matonaviridae comprises enveloped viruses with positive-sense RNA genomes of 9.6-10 kb. The genus Rubivirus includes rubella virus (species Rubivirus rubellae) infecting humans, ruhugu virus (species Rubivirus ruteetense) infecting bats and rustrela virus (species Rubivirus strelense) infecting rodents and zoo animals. Rubella virus is spread via droplets. Postnatal infection leads to benign disease with rash and fever. Infection of seronegative women with rubella virus during the first trimester of pregnancy will often result in severe foetal malformations, known as congenital rubella syndrome. Vaccines are globally available. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Matonaviridae, which is available at ictv.global/report/matonaviridae.

Seroprevalence of Measles-, Mumps-, and Rubella-specific antibodies in the German adult population - cross-sectional analysis of the German Health Interview and Examination Survey for Adults (DEGS1).

BACKGROUND: The WHO European Region targets the elimination of measles, rubella, and the congenital rubella syndrome and welcomes mumps elimination via the joint MMR vaccine. In a push towards this elimination goal, Germany introduced a recommendation on MMR vaccination for adults in 2010 to prevent increasing numbers of measles cases among adults and to strengthen herd immunity. METHODS: The prevalence of anti-measles, -mumps, and -rubella IgG antibodies was analysed in 7,115 participants between the ages of 18 and 79 years in the German Health Interview and Examination Survey. Risk factors of seronegativity of adults born 1970 or later were determined. FINDINGS: The seroprevalence of anti-measles IgG antibodies was more than 97% in adults born before 1965 and less than 90% in adults born afterwards. Prevalence and GMTs declined with later years of birth. Seronegativity was associated with two-sided migration background and region of residence in East Germany. For anti-mumps IgG antibodies, the seroprevalence was less than 90% in almost all age groups. Prevalence and GMTs declined with later years of birth. Seronegativity was not associated with any socio-demographic factor. Anti-rubella IgG seropositivity was found in more than 90% of adults born before 1985. GMTs declined in younger age groups. Seronegativity was associated with birth between 1980 and 1993 and male gender. High socio-economic status lowered the odds of being seronegative. INTERPRETATION: These data reinforce the implementation of the vaccination recommendation for adults and provide the basis for further evaluation of this measure. FUNDING: The Federal Ministry of Health, Germany.

Small-molecule polymerase inhibitor protects non-human primates from measles and reduces shedding.

Measles virus (MeV) is a highly contagious pathogen that enters the human host via the respiratory route. Besides acute pathologies including fever, cough and the characteristic measles rash, the infection of lymphocytes leads to substantial immunosuppression that can exacerbate the outcome of infections with additional pathogens. Despite the availability of effective vaccine prophylaxis, measles outbreaks continue to occur worldwide. We demonstrate that prophylactic and post-exposure therapeutic treatment with an orally bioavailable small-molecule polymerase inhibitor, ERDRP-0519, prevents measles disease in squirrel monkeys (Saimiri sciureus). Treatment initiation at the onset of clinical signs reduced virus shedding, which may support outbreak control. Results show that this clinical candidate has the potential to alleviate clinical measles and augment measles virus eradication.

Measles virus and rinderpest virus divergence dated to the sixth century BCE.

Many infectious diseases are thought to have emerged in humans after the Neolithic revolution. Although it is broadly accepted that this also applies to measles, the exact date of emergence for this disease is controversial. We sequenced the genome of a 1912 measles virus and used selection-aware molecular clock modeling to determine the divergence date of measles virus and rinderpest virus. This divergence date represents the earliest possible date for the establishment of measles in human populations. Our analyses show that the measles virus potentially arose as early as the sixth century BCE, possibly coinciding with the rise of large cities.

A Placenta Derived C-Terminal Fragment of ß-Hemoglobin With Combined Antibacterial and Antiviral Activity.

The placenta acts as physical and immunological barrier against the transmission of viruses and bacteria from mother to fetus. However, the specific mechanisms by which the placenta protects the developing fetus from viral and bacterial pathogens are poorly understood. To identify placental peptides and small proteins protecting from viral and bacterial infections, we generated a peptide library from 10 kg placenta by chromatographic means. Screening the resulting 250 fractions against Herpes-Simplex-Virus 2 (HSV-2), which is rarely transmitted through the placenta, in a cell-based system identified two adjacent fractions with significant antiviral activity. Further rounds of chromatographic purification and anti-HSV-2 testing allowed to purify the bioactive peptide. Mass spectrometry revealed the presence of a 36-mer derived from the C-terminal region of the hemoglobin ß subunit. The purified and corresponding chemically synthesized peptide, termed HBB(112-147), inhibited HSV-2 infection in a dose-dependent manner, with a mean IC50 in the median µg/ml range. Full-length hemoglobin tetramer had no antiviral activity. HBB(112-147) did not impair infectivity by direct targeting of the virions but prevented HSV-2 infection at the cell entry level. The peptide was inactive against Human Immunodeficiency Virus Type 1, Rubella and Zika virus infection, suggesting a specific anti-HSV-2 mechanism. Notably, HBB(112-147) has previously been identified as broad-spectrum antibacterial agent. It is abundant in placenta, reaching concentrations between 280 and 740 µg/ml, that are well sufficient to inhibit HSV-2 and prototype Gram-positive and -negative bacteria. We here additionally show, that HBB(112-147) also acts potently against Pseudomonas aeruginosa strains (including a multi-drug resistant strain) in a dose dependent manner, while full-length hemoglobin is inactive. Interestingly, the antibacterial activity of HBB(112-147) was increased under acidic conditions, a hallmark of infection and inflammatory conditions. Indeed, we found that HBB(112-147) is released from the hemoglobin precursor by Cathepsin D and Napsin A, acidic proteases highly expressed in placental and other tissues. We propose that upon viral or bacterial infection, the abundant hemoglobin precursor is proteolytically processed to release HBB(112-147), a broadly active antimicrobial innate immune defense peptide.

Hospital outbreak of measles - Evaluation and costs of 10 occupational cases among healthcare worker in Germany, February to March 2017.

After treatment of an inpatient with measles, an outbreak occurred within the unprotected healthcare workers (HCW) of a regional hospital in Hesse, Germany in February and March 2017. Overall, 10 HCW contracted measles. Remarkably, none of the affected HCW had direct contact to the index patient. One nosocomial transmission to a patient occurred. The economic impact of the outbreak is estimated to approximately 700,000€. Medical institutions play a major role in the management of measles outbreaks, since the risk of exposure as well as nosocomial transmission to vulnerable patients and HCW is very high. To avoid outbreaks it is essential to have an easily accessible documentation of the immune-status of all HCW. The role of occupational medicine in identifying and closing vaccination gaps is of particular importance.

Post-exposure prophylaxis for measles with immunoglobulins revised recommendations of the standing committee on vaccination in Germany.

Passive immunisation with immunoglobulins as post-exposure prophylaxis after contact with measles is recommended by the German Standing Committee on Vaccination (STIKO) particularly for unprotected individuals at high risk of complications for whom active immunization is contraindicated, such as infants <6 months of age, immunocompromised patients and pregnant women. The efficacy of passive immunisation in preventing measles depends on how soon after exposure it is administered, the concentration of measles antibodies in the immunoglobulin products and dosage. Since the global introduction of standard active immunisation against measles and the concomitant reduction in virus circulation, the levels of measles virus (MV)-specific IgG antibodies in the population have dropped. Thus, the concentration of MV-specific antibodies in immunoglobulin products derived from human plasma donors has declined as the proportion of vaccinated donors has increased. The MV-neutralizing capacity of immunoglobulin products is not routinely tested in Germany. No official data exist on the concentrations of MV-specific IgG antibodies in individual batches of immunoglobulins available in Germany and the required minimum level for MV-specific IgG is not stipulated. The STIKO re-evaluated available data and measurements of MV-neutralizing capacities of available immunoglobulin (IgG) products in Germany at the National Reference Centre Measles, Mumps, Rubella at the Robert Koch Institute. Based on the findings, STIKO modified its previous recommendations on the post-exposure use of immunoglobulins (2001), especially with respect to risk groups, application and dosage. STIKO now recommends a single intravenous administration of immunoglobulins (400 mg/kg body weight) as soon as possible, preferably within six days, after exposure to measles, specifically for infants aged <6 months, susceptible pregnant women and immunocompromised patients.

Rubella Viruses Shift Cellular Bioenergetics to a More Oxidative and Glycolytic Phenotype with a Strain-Specific Requirement for Glutamine.

The flexible regulation of cellular metabolic pathways enables cellular adaptation to changes in energy demand under conditions of stress such as posed by a virus infection. To analyze such an impact on cellular metabolism, rubella virus (RV) was used in this study. RV replication under selected substrate supplementation with glucose, pyruvate, and glutamine as essential nutrients for mammalian cells revealed its requirement for glutamine. The assessment of the mitochondrial respiratory (based on the oxygen consumption rate) and glycolytic (based on the extracellular acidification rate) rate and capacity by respective stress tests through Seahorse technology enabled determination of the bioenergetic phenotype of RV-infected cells. Irrespective of the cellular metabolic background, RV infection induced a shift of the bioenergetic state of epithelial cells (Vero and A549) and human umbilical vein endothelial cells to a higher oxidative and glycolytic level. Interestingly there was a RV strain-specific, but genotype-independent demand for glutamine to induce a significant increase in metabolic activity. While glutaminolysis appeared to be rather negligible for RV replication, glutamine could serve as donor of its amide nitrogen in biosynthesis pathways for important metabolites. This study suggests that the capacity of RVs to induce metabolic alterations could evolve differently during natural infection. Thus, changes in cellular bioenergetics represent an important component of virus-host interactions and could complement our understanding of the viral preference for a distinct host cell population.IMPORTANCE RV pathologies, especially during embryonal development, could be connected with its impact on mitochondrial metabolism. With bioenergetic phenotyping we pursued a rather novel approach in virology. For the first time it was shown that a virus infection could shift the bioenergetics of its infected host cell to a higher energetic state. Notably, the capacity to induce such alterations varied among different RV isolates. Thus, our data add viral adaptation of cellular metabolic activity to its specific needs as a novel aspect to virus-host evolution. In addition, this study emphasizes the implementation of different viral strains in the study of virus-host interactions and the use of bioenergetic phenotyping of infected cells as a biomarker for virus-induced pathological alterations.

Vaccination titres pre- and post-transplant in paediatric renal transplant recipients and the impact of immunosuppressive therapy.

BACKGROUND: Avoidance of vaccine-preventable infections in paediatric renal allograft recipients is of utmost importance. However, the development and maintenance of protective vaccination titres may be impaired in this patient population owing to their need for immunosuppressive medication. METHODS: In the framework of the Cooperative European Paediatric Renal Transplant Initiative (CERTAIN), we therefore performed a multi-centre, multi-national study and analysed vaccination titres pre- and post-transplant in 155 patients with serial titre measurements in comparison with published data in healthy children. RESULTS: The percentage of patients with positive vaccination titres before renal transplantation (RTx) was low, especially for diphtheria (38.5%, control 75%) and pertussis (21.3%, control 96.3%). As few as 58.1% of patients had a hepatitis B antibody (HBsAb) titre >100 IU/L before RTx. 38.1% of patients showed a vaccination titre loss post-transplant. Patients with an HBsAb titre between 10 and 100 IU/L before RTx experienced a significantly (p < 0.05) more frequent hepatitis B vaccination titre loss post-transplant than patients with an HBsAb titre >100 IU/L. The revaccination rate post-transplant was low and revaccination failed to induce positive titres in a considerable number of patients (27.3 to 83.3%). Treatment with rituximab was associated with a significantly increased risk of a vaccination titre loss post-transplant (odds ratio 4.26, p = 0.033). CONCLUSIONS: These data show a low percentage of patients with positive vaccination titres pre-transplant, a low revaccination rate post-transplant with limited antibody response, and a high rate of vaccination titre losses.

Large measles outbreak introduced by asylum seekers and spread among the insufficiently vaccinated resident population, Berlin, October 2014 to August 2015.

The largest measles outbreak in Berlin since 2001 occurred from October 2014 to August 2015. Overall, 1,344 cases were ascertained, 86% (with available information) unvaccinated, including 146 (12%) asylum seekers. Median age was 17 years (interquartile range: 4-29 years), 26% were hospitalised and a 1-year-old child died. Measles virus genotyping uniformly revealed the variant 'D8-Rostov-Don' and descendants. The virus was likely introduced by and initially spread among asylum seekers before affecting Berlin's resident population. Among Berlin residents, the highest incidence was in children aged < 2 years, yet most cases (52%) were adults. Post-exposure vaccinations in homes for asylum seekers, not always conducted, occurred later (median: 7.5 days) than the recommended 72 hours after onset of the first case and reached only half of potential contacts. Asylum seekers should not only have non-discriminatory, equitable access to vaccination, they also need to be offered measles vaccination in a timely fashion, i.e. immediately upon arrival in the receiving country. Supplementary immunisation activities targeting the resident population, particularly adults, are urgently needed in Berlin.