Genome sequencing of human in vitro fertilisation embryos for pathogenic variation screening.

Whole-genome sequencing of preimplantation human embryos to detect and screen for genetic diseases is a technically challenging extension to preconception screening. Combining preconception genetic screening with preimplantation testing of human embryos facilitates the detection of de novo mutations and self-validates transmitted variant detection in both the reproductive couple and the embryo's samples. Here we describe a trio testing workflow that involves whole-genome sequencing of amplified DNA from biopsied embryo trophectoderm cells and genomic DNA from both parents. Variant prediction software and annotation databases were used to assess variants of unknown significance and previously not described de novo variants in five single-gene preimplantation genetic testing couples and eleven of their embryos. Pathogenic variation, tandem repeat, copy number and structural variations were examined against variant calls for compound heterozygosity and predicted disease status was ascertained. Multiple trio testing showed complete concordance with known variants ascertained by single-nucleotide polymorphism array and uncovered de novo and transmitted pathogenic variants. This pilot study describes a method of whole-genome sequencing and analysis for embryo selection in high-risk couples to prevent early life fatal genetic conditions that adversely affect the quality of life of the individual and families.

Hypermutability in environmental Pseudomonas aeruginosa and in populations causing pulmonary infection in individuals with cystic fibrosis.

Pseudomonas aeruginosa is the pathogen most commonly associated with morbidity and mortality in cystic fibrosis (CF) patients. The host-pathogen interactions responsible for progressive CF lung diseases are complex. However, there is growing interest in the role of hypermutable P. aeruginosa (that is, those strains with an increased mutation frequency due to mutations in mismatch repair and error prevention genes), in terms of both bacterial adaptation and antimicrobial resistance. The prevalence of hypermutable P. aeruginosa in chronic CF infection has been established, and at 37 % is surprisingly high. To the authors' knowledge, there are no reports of prevalence during the early stages of infection, in environmental pseudomonas, which are believed to be the primary source of infection, and in epidemic strains, which have emerged as a major challenge. The aim of this study was to establish the prevalence of hypermutable P. aeruginosa in these pseudomonas populations. The hypothesis was that hypermutability would be rare in early and in environmental P. aeruginosa but in contrast would explain the relatively recent emergence of epidemic strains. It was found that 10/100 (10 %) of early isolates were strong or weak mutators, suggesting that the CF lung is not the only factor influencing the existence of mutators in this group of patients. Two weak mutators (6 %) were found in 32 environmental isolates. Only two of 15 (13 %) epidemic P. aeruginosa strains were hypermutable, and although closer analysis revealed this issue to be complex, on the whole the data suggested that the atypical characteristics of these highly transmissible strains cannot solely be explained by this phenomenon. The higher than predicted prevalence of mutators in early infection, and in environmental isolates, reinforces the importance of early and aggressive treatment for P. aeruginosa infection in CF.