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Biallelic pathogenic variants in CFTR manifest as cystic fibrosis (CF) or other CFTR-related disorders (CFTR-RDs). The 5T allele, causing alternative splicing and reduced protein activity, is modulated by the adjacent TG repeat element, though previous data have been limited to small, selective cohorts. Here, the risk and spectrum of phenotypes associated with the CFTR TG-T5 haplotype variants (TG11T5, TG12T5, and TG13T5) in the absence of the p.Arg117His variant are evaluated. Individuals who received physician-ordered next-generation sequencing of CFTR were included. TG[11-13]T5 variant frequencies (biallelic or with another CF-causing variant [CFvar]) were calculated. Clinical information reported by the ordering provider or the individual was examined. Among 548,300 individuals, the T5 minor allele frequency (MAF) was 4.2% (TG repeat distribution: TG11 = 68.1%, TG12 = 29.5%, TG13 = 2.4%). When present with a CFvar, each TG[11-13]T5 variant was significantly enriched in individuals with a high suspicion of CF or CFTR-RD (personal/family history of CF/CFTR-RD) compared to those with a low suspicion for CF or CFTR-RD (hereditary cancer screening, CFTR not requisitioned). Compared to CFvar/CFvar individuals, those with TG[11-13]T5/CFvar generally had single-organ involvement, milder symptoms, variable expressivity, and reduced penetrance. These data improve our understanding of disease risks associated with TG[11-13]T5 variants and have important implications for reproductive genetic counseling.

Research Question Is FAST-SeqS an accurate methodology for preimplantation genetic testing for whole-chromosome aneuploidy (PGT-A), what additional types of chromosomal abnormalities can be assessed, and what are the observed aneuploidy rates in a large clinical cohort? Design FAST-SeqS, a next-generation sequencing- (NGS-) based assay amplifying genome-wide LINE1 repetitive sequences, was validated using reference samples. Sensitivity and specificity were calculated. Clinically-derived trophectoderm biopsies submitted for PGT-A were assessed, and aneuploidy and mosaicism rates among biopsies were determined. Clinician-provided outcome rates were calculated. Results Sensitivity and specificity were >95% for all aneuploidy types tested in the validation. Comparison of FAST-SeqS to VeriSeq showed high concordance (98.5%). Among embryos with actionable results (n=182,827), 46.2% were aneuploid. Whole-chromosome aneuploidies were most commonly observed (72.9% without or 8.7% with a segmental aneuploidy [SA]), with rates increasing with egg age, while SA rates did not. SAs (n=20,557) were observed on all chromosomes (most commonly deletions), with frequencies roughly correlated with chromosome length. Mosaic-only abnormalities constituted 10.1% (n=3,862/38,145) of samples. Abnormal ploidy constituted 1.8% (n=2,370/128,991) of samples, triploidy being the most common (73.6%). Across 3,297 frozen embryo transfers, the mean clinical pregnancy rate was 62% (range 38-80%); the mean combined ongoing pregnancy and live birth rate was 57% (range 38-72%). Conclusion FAST-SeqS is a clinically reliable and scalable method for PGT-A, is comparable to whole genome amplification-based platforms, and detects additional information related to ploidy using SNP analysis. Clinical outcomes suggest an ongoing benefit of PGT-A using FAST-SeqS, consistent with other platforms.

PurposeTo evaluate the risk and spectrum of phenotypes associated in individuals with one or two of the CFTR T5 haplotype variants (TG11T5, TG12T5 and TG13T5) in the absence of the R117H variant.MethodsIndividuals who received testing with CFTR NGS results between 2014 and 2019 through Invitae at ordering provider discretion were included. TG-T repeats were detected using a custom-developed haplotype caller. Frequencies of the TG-T5 variants (biallelic or in combination with another CF-causing variant [CFvar]) were calculated. Clinical information reported by the ordering provider (via requisition form) or the individual (during genetic counseling appointments) was examined.ResultsAmong 548,300 individuals, the minor allele frequency of the T5 allele was 4.2% (TG repeat distribution: TG11=68.1%, TG12=29.5%, TG13=2.4%). When present with a CFvar, each of the TG[11-13]T5 variants were significantly enriched in individuals with a “high suspicion” of CF/CFTR-RD (personal/family history of CF/CFTR-RD) compared to those with very “low suspicion” for CF or CFTR-RD (hereditary cancer testing, CFTR not requisitioned). Compared to CFvar/CFvar individuals, TG[11-13]T5/CFvar individuals generally had single organ involvement, milder symptoms, variable expressivity, and reduced penetrance.DiscussionData from this study provides a better understanding of disease risks associated with inheriting TG[11-13]T5 variants and has important implications for reproductive genetic counseling.

Placental function impacts growth and development with lifelong consequences for performance and health. We provide novel insights into placental development in bovine, an important agricultural species and biomedical model.Concepti with defined genetics and sex were recovered from nulliparous dams managed under standardized conditions to study placental gross morphological and histomorphological parameters at the late embryo (Day48) and early accelerated fetal growth (Day153) stages.Placentome number increased 3-fold between Day48 and Day153. Placental barrier thickness was thinner, and volume of placental components, and surface areas and densities were higher at Day153 than Day48. We confirmed two placentome types, flat and convex. At Day48, there were more convex than flat placentomes, and convex placentomes had a lower proportion of maternal connective tissue (P 0.01). However, this was reversed at Day153, where convex placentomes were lower in number and had greater volume of placental components (P 0.01- P 0.001) and greater surface area (P 0.001) than flat placentomes. Importantly, embryo (r = 0.50) and fetal (r = 0.30) weight correlated with total number of convex but not flat placentomes.Extensive remodelling of the placenta increases capacity for nutrient exchange to support rapidly increasing embryo-fetal weight from Day48 to Day153. The cellular composition of convex placentomes, and exclusive relationships between convex placentome number and embryo-fetal weight, provide strong evidence for these placentomes as drivers of prenatal growth. The difference in proportion of maternal connective tissue between placentome types at Day48 suggests that this tissue plays a role in determining placentome shape, further highlighting the importance of early placental development.

Loss of function variants in the PCDH15 gene can cause Usher syndrome type 1F, an autosomal recessive disease associated with profound congenital hearing loss, vestibular dysfunction, and retinitis pigmentosa. The Ashkenazi Jewish population has an increased incidence of Usher syndrome type 1F (founder variant p.Arg245X accounts for 75% of alleles), yet the variant spectrum in a panethnic population remains undetermined. We sequenced the coding region and intron-exon borders of PCDH15 using next-generation DNA sequencing technology in approximately 14,000 patients from fertility clinics. More than 600 unique PCDH15 variants (single nucleotide changes and small indels) were identified, including previously described pathogenic variants p.Arg3X, p.Arg245X (five patients), p.Arg643X, p.Arg929X, and p.Arg1106X. Novel truncating variants were also found, including one in the N-terminal extracellular domain (p.Leu877X), but all other novel truncating variants clustered in the exon 33 encoded C-terminal cytoplasmic domain (52 patients, 14 variants). One variant was observed predominantly in African Americans (carrier frequency of 2.3%). The high incidence of truncating exon 33 variants indicates that they are unlikely to cause Usher syndrome type 1F even though many remove a large portion of the gene. They may be tolerated because PCDH15 has several alternate cytoplasmic domain exons and differentially spliced isoforms may function redundantly. Effects of some PCDH15 truncating variants were addressed by deep sequencing of a panethnic population.

This document summarizes laboratory guidelines for the detection, interpretation, and reporting of maternal cell contamination in prenatal analyses.

Chronic Myeloid Leukemia (CML) is diagnosed by the presence of the Philadelphia chromosome (Ph+). The number of cells containing the bcr/abl chromosomal translocation diminishes dramatically after current standard of care treatment. Patient progress is monitored quantitatively and long-term prognosis is favorable if the Ph+ cells are no longer detected by cytogenetic analysis or FISH (sensitivity of 20% and 2% respectively). Molecular monitoring of cDNA BCR/ABL transcripts is widely used to assess levels of minimal residual disease (MRD). In our laboratory, quantitative RT-PCR allows detection of BCR/ABL transcript down to 1 x 10−5 or one BCR/ABL transcript in 100,000 reference gene transcripts. Serial values from bone marrow (BM) and blood specimens are tracked over time and monitored for increases that may indicate unfavorable changes in disease status. Though the literature indicates that the levels of BCR/ABL transcript in BM and blood samples are comparable at the time of diagnosis, this may not be true in patients with MRD. To explore this question we compared BCR/ABL transcript levels in paired BM and blood samples that were either collected together or separately but within a short time frame. In 36% of these cases both specimen types had detectable low-level BCR/ABL transcript. The levels of transcript in bone marrow were consistently higher than the levels in the associated blood specimen, though there was a wide range of variability (3-50 fold). Additionally, when blood levels of transcript were near the lower limit of detection, we were more likely to observe a larger difference relative to bone marrow transcript levels than when blood levels of transcript were higher. These results indicate that while both BM and blood specimens are valuable for serial molecular monitoring, they may not be equivocal when sampled from patients with MRD. If a patient has been monitored using blood specimens, periodic concurrent testing of blood and bone marrow specimens can provide clinical utility.