Viral Transfer and Inactivation through Zooplankton Trophic Interactions

Waterborne viruses are responsible for numerous diseases and are abundant in aquatic systems. Understanding the fate of viruses in natural systems has important implications for human health. This research quantifies the uptake of the bacteriophage T4 and the enteric virus echovirus 11 when exposed to the filter feeders Tetrahymena pyriformis and Daphnia magna, and also examines the potential of viral transfer due to trophic interactions. Experiments co-incubating each species with the viruses over 72-96 h showed up to a 4 log virus removal for T. pyriformis, while direct viral uptake by D. magna was not observed. However, viral uptake by D. magna occurred indirectly by viral transfer from prey to predator, through D. magna feeding on virus-loaded T. pyriformis. This prey-predator interaction resulted in a 1 log additional virus removal compared to removal by T. pyriformis alone. Incomplete viral inactivation by D. magna was observed through recovery of infective viruses from the daphnid tissue. This research furthers our understanding of the impacts of zooplankton filter feeding on viral inactivation and shows the potential for viral transfer through the food chain. The viral-zooplankton interactions observed in these studies indicate that zooplankton may improve water quality through viral uptake or may serve as vectors for infection by accumulating viruses

Increased efficacy for in-house validation of real-time PCR GMO detection methods

To improve the efficacy of the in-house validation of GMO detection methods (DNA isolation and real-time PCR, polymerase chain reaction), a study was performed to gain insight in the contribution of the different steps of the GMO detection method to the repeatability and in-house reproducibility. In the present study, 19 methods for (GM) soy, maize canola and potato were validated in-house of which 14 on the basis of an 8-day validation scheme using eight different samples and five on the basis of a more concise validation protocol. In this way, data was obtained with respect to the detection limit, accuracy and precision. Also, decision limits were calculated for declaring non-conformance (>0.9%) with 95% reliability. In order to estimate the contribution of the different steps in the GMO analysis to the total variation variance components were estimated using REML (residual maximum likelihood method). From these components, relative standard deviations for repeatability and reproducibility (RSDr and RSDR) were calculated. The results showed that not only the PCR reaction but also the factors ‘DNA isolation’ and ‘PCR day’ are important factors for the total variance and should therefore be included in the in-house validation. It is proposed to use a statistical model to estimate these factors from a large dataset of initial validations so that for similar GMO methods in the future, only the PCR step needs to be validated. The resulting data are discussed in the light of agreed European criteria for qualified GMO detection methods

Adaptation and validation of the ACMG/AMP variant classification framework for MYH7-associated inherited cardiomyopathies: recommendations by ClinGen’s Inherited Cardiomyopathy Expert Panel

Purpose Integrating genomic sequencing in clinical care requires standardization of variant interpretation practices. The Clinical Genome Resource has established expert panels to adapt the American College of Medical Genetics and Genomics/Association for Molecular Pathology classification framework for specific genes and diseases. The Cardiomyopathy Expert Panel selected MYH7, a key contributor to inherited cardiomyopathies, as a pilot gene to develop a broadly applicable approach. Methods: Expert revisions were tested with 60 variants using a structured double review by pairs of clinical and diagnostic laboratory experts. Final consensus rules were established via iterative discussions. Results: Adjustments represented disease-/gene-informed specifications (12) or strength adjustments of existing rules (5). Nine rules were deemed not applicable. Key specifications included quantitative frameworks for minor allele frequency thresholds, the use of segregation data, and a semiquantitative approach to counting multiple independent variant occurrences where fully controlled case-control studies are lacking. Initial inter-expert classification concordance was 93%. Internal data from participating diagnostic laboratories changed the classification of 20% of the variants (n = 12), highlighting the critical importance of data sharing. Conclusion: These adapted rules provide increased specificity for use in MYH7-associated disorders in combination with expert review and clinical judgment and serve as a stepping stone for genes and disorders with similar genetic and clinical characteristics