RESUMO
Rabies diagnosis proficiency tests on animal specimens using four techniques (FAT, RTCIT, conventional RT-PCR and real-time RT-PCR) were organised over 10 years (2009-2019). Seventy-three laboratories, of which 59% were from Europe, took part. As the panels were prepared with experimentally-infected samples, the error rate of laboratories on positive and negative samples was accurately estimated. Based on fitted values produced by mixed modelling including the variable "laboratory" as a random variable to take into account the longitudinal design of our dataset, the technique that provided the most concordant results was conventional RT-PCR (99.3%; 95% CI 99.0-99.6), closely followed by FAT (99.1%; 95% CI 98.7-99.4), real-time RT-PCR (98.7%; 95% CI 98.1-99.3) and then RTCIT (96.8%; 95% CI 95.8-97.7). We also found that conventional RT-PCR provided a better diagnostic sensitivity level (99.3% ±4.4%) than FAT (98.7% ±1.6%), real-time RT-PCR (97.9% ±0.8%) and RTCIT (95.3% ±5.1%). Regarding diagnostic specificity, RTCIT was the most specific technique (96.4% ±3.9%) followed closely by FAT (95.6% ±3.8%), real-time RT-PCR (95.0% ±1.8%) and conventional RT-PCR (92.9% ±0.5%). Due to multiple testing of the samples with different techniques, the overall diagnostic conclusion was also evaluated, and found to reach an inter-laboratory concordance level of 99.3%. The concordance for diagnostic sensitivity was 99.6% ±2.0% and for diagnostic specificity, 98.0% ±8.5%. Molecular biology techniques were, however, found to be less specific than expected. The potential reasons for such findings are discussed herein. The regular organisation of performance tests has contributed to an increase in the performance of participating laboratories over time, demonstrating the benefits of such testing. Maintaining a high-quality rabies diagnosis capability on a global scale is key to achieving the goal of eliminating dog-mediated human rabies deaths. The regular organisation of exercises on each continent using selected local strains to be tested according to the local epidemiological situation is one factor that could help increase reliable diagnosis worldwide. Rabies diagnosis capabilities could indeed be enhanced by providing adequate and sustainable proficiency testing on a large scale and in the long term.
Assuntos
Testes Diagnósticos de Rotina/métodos , Técnicas de Diagnóstico Molecular/métodos , Vírus da Raiva/isolamento & purificação , Raiva/diagnóstico , Animais , Cães , Europa (Continente) , Humanos , Laboratórios , RNA Viral , Vírus da Raiva/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
Tetracycline and rhodamine are bait uptake biomarkers commonly used for decades in oral rabies vaccination campaigns. They require tooth collection and the capture or death of animals. Here, we considered the use of marked baits with plastic beads, a physical biomarker allowing noninvasive scat survey by direct observation in the field. Such methodology would be in compliance with animal welfare concerns. The development of a relative bait uptake estimation on the basis of observation marked scat could indeed be beneficial for the monitoring of oral vaccination programs, especially in programs dedicated to dogs ( Canis familiaris) or protected species.
Assuntos
Doenças do Cão/prevenção & controle , Vacina Antirrábica/imunologia , Raiva/prevenção & controle , Rodaminas/química , Tetraciclina/química , Administração Oral , Animais , Biomarcadores , Cães , Fezes/química , Corantes Fluorescentes/química , Plásticos , Vacina Antirrábica/administração & dosagem , Vacinação/veterináriaRESUMO
Zika virus (ZIKV) recently emerged in the Western Hemisphere with previously unrecognized or unreported clinical presentations. Here, we identify two putative binding mechanisms of ancestral and emergent ZIKV strains featuring the envelope (E) protein residue asparagine 154 (ASN154) and viral phosphatidylserine (PS). Synthetic peptides representing the region containing ASN154 from strains PRVABC59 (Puerto Rico 2015) and MR_766 (Uganda 1947) were exposed to neuronal cells and fibroblasts to model ZIKV E protein/cell interactions and bound MDCK or Vero cells and primary neurons significantly. Peptides significantly inhibited Vero cell infectivity by ZIKV strains MR_766 and PRVABC59, indicating that this region represents a putative binding mechanism of ancestral African ZIKV strains and emergent Western Hemisphere strains. Pretreatment of ZIKV strains MR_766 and PRVABC59 with the PS-binding protein annexin V significantly inhibited replication of PRVABC59 but not MR_766, suggesting that Western hemisphere strains may additionally be capable of utilizing PS-mediated entry to infect host cells. These data indicate that the region surrounding E protein ASN154 is capable of binding fibroblasts and primary neuronal cells and that PS-mediated entry may be a secondary mechanism for infectivity utilized by Western Hemisphere strains.