Development of a real-time fluorescent reverse transcription loop-mediated isothermal amplification assay with quenching primers for rapid detection of rubella virus.

Rubella virus infection during early pregnancy sometimes causes severe birth defects termed congenital rubella syndrome. Although there are safe and effective live-attenuated vaccines, rubella has only been certified as eliminated in the Americas within the six World Health Organization regions. Rubella remains an endemic disease in many regions, and outbreaks occur wherever population immunity is insufficient. There are two main methods for diagnosis of rubella: detection of anti-rubella IgM antibodies by enzyme immunoassay and detection of the viral genome by real-time RT-PCR. Both of these methods require substantial time and effort. In the present study, a rapid rubella detection assay using real-time fluorescent reverse transcription loop-mediated isothermal amplification with quenching primers was developed. The time required for the new assay was one-half that required for a real-time RT-PCR assay. The assay had 93.6% positive percent agreement and 100% negative percent agreement for clinical specimens compared with the real-time RT-PCR assay. The new assay is considered useful for diagnosis of rubella in areas where rubella is endemic.

Case report: Rubella virus-associated cutaneous granuloma in an adult with TAP1 deficiency.

Rubella virus-associated granulomas commonly occur in immunocompromised individuals, exhibiting a diverse range of clinical presentations. These manifestations can vary from predominantly superficial cutaneous plaques or nonulcerative nodules to more severe deep ulcerative lesions, often accompanied by extensive necrosis and significant tissue destruction. TAP1 deficiency, an exceedingly rare primary immune-deficiency disorder, presents with severe chronic sino-pulmonary infection and cutaneous granulomas. This report highlights the occurrence of rubella virus-associated cutaneous granulomas in patients with TAP1 deficiency. Notably, the pathogenic mutation responsible for TAP1 deficiency stems from a novel genetic alteration that has not been previously reported. This novel observation holds potential significance for the field of diagnosis and investigative efforts in the context of immunodeficiency disorders.

HLA Associations of Intrathecal IgG Production against Specific Viruses in Multiple Sclerosis.

OBJECTIVE: Specific human leucocyte antigen (HLA) alleles are not only associated with higher risk to develop multiple sclerosis (MS) and other autoimmune diseases, but also with the severity of various viral and bacterial infections. Here, we analyzed the most specific biomarker for MS, that is, the polyspecific intrathecal IgG antibody production against measles, rubella, and varicella zoster virus (MRZ reaction), for possible HLA associations in MS. METHODS: We assessed MRZ reaction from 184 Swiss patients with MS and clinically isolated syndrome (CIS) and 89 Swiss non-MS/non-CIS control patients, and performed HLA sequence-based typing, to check for associations of positive MRZ reaction with the most prevalent HLA alleles. We used a cohort of 176 Swedish MS/CIS patients to replicate significant findings. RESULTS: Whereas positive MRZ reaction showed a prevalence of 38.0% in MS/CIS patients, it was highly specific (97.7%) for MS/CIS. We identified HLA-DRB1*15:01 and other tightly linked alleles of the HLA-DR15 haplotype as the strongest HLA-encoded risk factors for a positive MRZ reaction in Swiss MS/CIS (odds ratio [OR], 3.90, 95% confidence interval [CI] 2.05-7.46, padjusted = 0.0004) and replicated these findings in Swedish MS/CIS patients (OR 2.18, 95%-CI 1.16-4.02, padjusted = 0.028). In addition, female MS/CIS patients had a significantly higher probability for a positive MRZ reaction than male patients in both cohorts combined (padjusted <0.005). INTERPRETATION: HLA-DRB1*15:01, the strongest genetic risk factor for MS, and female sex, 1 of the most prominent demographic risk factors for developing MS, predispose in MS/CIS patients for a positive MRZ reaction, the most specific CSF biomarker for MS. ANN NEUROL 2024;95:1112-1126.

Crystal structure and biochemical activity of the macrodomain from rubella virus p150.

Rubella virus encodes a nonstructural polyprotein with RNA polymerase, methyltransferase, and papain-like cysteine protease activities, along with a putative macrodomain of unknown function. Macrodomains bind ADP-ribose adducts, a post-translational modification that plays a key role in host-virus conflicts. Some macrodomains can also remove the mono-ADP-ribose adduct or degrade poly-ADP-ribose chains. Here, we report high-resolution crystal structures of the macrodomain from rubella virus nonstructural protein p150, with and without ADP-ribose binding. The overall fold is most similar to macroD-type macrodomains from various nonviral species. The specific composition and structure of the residues that coordinate ADP-ribose in the rubella virus macrodomain are most similar to those of macrodomains from alphaviruses. Isothermal calorimetry shows that the rubella virus macrodomain binds ADP-ribose in solution. Enzyme assays show that the rubella virus macrodomain can hydrolyze both mono- and poly-ADP-ribose adducts. Site-directed mutagenesis identifies Asn39 and Cys49 required for mono-ADP-ribosylhydrolase (de-MARylation) activity.IMPORTANCERubella virus remains a global health threat. Rubella infections during pregnancy can cause serious congenital pathology, for which no antiviral treatments are available. Our work demonstrates that, like alpha- and coronaviruses, rubiviruses encode a mono-ADP-ribosylhydrolase with a structurally conserved macrodomain fold to counteract MARylation by poly (ADP-ribose) polymerases (PARPs) in the host innate immune response. Our structural data will guide future efforts to develop novel antiviral therapeutics against rubella or infections with related viruses.

Molecular analysis of rubella virus in travelers suspected of measles infection in São Paulo, Brazil / Análise molecular do vírus da rubéola em turistas suspeitos de infecção por sarampo em São Paulo, Brasil

OBJECTIVE: To identify measles virus genotypes in three cases of travelers suspected of measles infection. METHODS: Samples (blood and urine) were collected for serology, virus isolation, and genotyping. Sera were analyzed for IgM antibodies against measles virus and rubella virus by enzyme-linked immunosorbent assay (ELISA) (Siemens - Marburg, Germany). Clinical samples (lymphocytes and urine) were inoculated into Statens Serum Institute rabbit corneal epithelial cell line- ATCC CL 60 (SIRC) and Vero Slam cells. RNA was extracted from clinical samples and cell culture was inoculated and processed by polymerase chain reaction (PCR) with oligonucleotides specific for measles virus (MV) and rubella virus (RV). RESULTS: All patients showed IgM negative serology for MV and positive IgM for RV. RV belonging to genotypes 1B, 1C, and 1E were isolated from patients who came from Finland, Peru, and Germany, respectively. Genotype 1B has been found in Europe and on the East Coast of South America; 1C has been found in Peru and the West Coast of South America, and 1E, first identified in 1997, now appears to have worldwide distribution. CONCLUSION: Information about RV and MV genotypes circulating in São Paulo is essential for the control of measles, rubella, and congenital rubella syndrome (CRS) in Brazil.

OBJETIVO: Identificar o genótipo do vírus do sarampo em três viajantes suspeitos de infecção por sarampo. MÉTODOS: Amostras (sangue e urina) foram coletadas para sorologia, isolamento viral e genotipagem. As sorologias para pesquisa de IgM para o vírus do sarampo e da rubéola foi realizada utilizando-se o kit de ELISA (Siemens - Marburg, Alemanha). As amostras clínicas (linfócito e urina) foram inoculadas na SIRC (Statens Serum Institute rabbit corneal epithelial cell line-ATCC CL 60) e nas células Vero Slam. O RNA foi extraído das amostras clínicas e das células inoculadas e processadas por PCR, utilizando oligonucleotideos específicos para sarampo e rubéola. RESULTADOS: Todos os pacientes apresentaram sorologia IgM negativa para sarampo e positivo para rubéola. Os vírus da rubéola isolados dos pacientes que vieram da Finlândia, Peru e Alemanha pertencem aos genótipos 1B, 1C e 1E, respectivamente. O genótipo 1B foi encontrado na Europa e na costa oriental da América do Sul, o genótipo 1C foi encontrado no Peru e na costa oeste da América do Sul e o genótipo 1E, identificado pela primeira vez em 1997, agora aparenta ser um genótipo com distribuição mundial. CONCLUSÃO: O conhecimento dos genótipos de sarampo e rubéola que circulam em São Paulo é essencial para o controle do sarampo, rubéola e síndrome da rubéola congênita.

Campaña nacional de vacunación para la eliminación de la rubéola y del síndrome de rubéola congénita en Bolivia, 2006 / National vaccination campaign for the elimination of rubella and congenital rubella syndrome in Bolivia, 2006

En mayo 2006, Bolivia llevó a cabo una campaña nacional de vacunación en su población de 15 a 39 años para eliminar la rubéola y el síndrome de rubéola congénita del país. Durante seis semanas el país sumó los esfuerzos de los políticos, aliados nacionales e internacionales, trabajadores de salud y organizaciones sociales para alcanzar la meta. La cobertura nacional validada por una encuesta post campaña fue de 94%, sin diferencias significativasde edad, sexo o en la distribución rural urbana. La encuesta probó también un alto nivel de confianza de la población en los servicios de vacunación y el rol importante del personal de salud para informar a la población en el medio rural. Se demuestra que actividades de vacunación masivas en la población adulta y de ambos sexos pueden ser exitosas con una preparación minuciosa y un plan de comunicación estudiado.

Estudio de un mecanismo de persistencia del virus Rubéola en la infección congénita / Study of a mechanism of persistence of the virus Rubella in the congenital infection

Se investigó el rol de la apoptosis en un posible mecanismo de persistencia del virus Rubéola (Rub) en la infección congénita, utilizando cultivos primarios de fibroblastos fetales humanos (FFH) en activa proliferación. Se demostró que, en contraste con lo que se observa en células Vero, RK13, placenta humana a término, y fibroblastos diploides de adulto humano (Hs888Lu), Rub no induce apoptosis en FFH. Por inmunohistoquímica se detectó la actividad de JUN, gen que promueve la proliferación celular, en FFH pero no en Hs888Lu. El análisis de expresión de genes en FFH y Hs888Lu con microarreglos de ADN mostró que en FFH están sobre-expresados genes que estimulan la supervivencia y proliferación celular, como factores de crecimiento, PI3K, y oncogenes de la familia Ras. Las kinasas p-Akt y p-ERK se detectaron en FFH por western blotting. A continuación se demostró que Rub sí induce apoptosis en FFH, cuando se suprimen las vías PI3K–Akt y Ras–Raf–MEK–ERK. Significativamente, la infección de las células fetales no prospera en estas condiciones. Así, la persistencia de Rub en la infección congénita puede estar inversamente relacionada con la ocurrencia de apoptosis en las células hospedadoras. También se analizó la expresión de genes en células infectadas por Rub, identificándose más de 1500 genes celulares inducidos o suprimidos por el virus, muchos de ellos relacionados con la respuesta inmune, las vías metabólicas, y las cascadas de transmisión de señales. En FFH, Rub modifica la expresión de diferentes genes del desarrollo (genes Hox), que establecen el patrón corporal y promueven la proliferación y la diferenciación celular durante la morfogénesis. Estos datos representan nuevos puntos de partida para postular hipótesis sobre el modo en que el virus ejerce su efecto teratogénico, y brindan un amplio panorama para estudiar la relación entre Rub y la célula.