Collagen prolyl 4-hydroxylases (C-P4Hs) play a central role in the formation and stabilization of the triple helical domain of collagens. P4HA1 encodes the catalytic α(I) subunit of the main C-P4H isoenzyme (C-P4H-I). We now report human bi-allelic P4HA1 mutations in a family with a congenital-onset disorder of connective tissue, manifesting as early-onset joint hypermobility, joint contractures, muscle weakness and bone dysplasia as well as high myopia, with evidence of clinical improvement of motor function over time in the surviving patient. Similar to P4ha1 null mice, which die prenatally, the muscle tissue from P1 and P2 was found to have reduced collagen IV immunoreactivity at the muscle basement membrane. Patients were compound heterozygous for frameshift and splice site mutations leading to reduced, but not absent, P4HA1 protein level and C-P4H activity in dermal fibroblasts compared to age-matched control samples. Differential scanning calorimetry revealed reduced thermal stability of collagen in patient-derived dermal fibroblasts versus age-matched control samples. Mutations affecting the family of C-P4Hs, and in particular C-P4H-I, should be considered in patients presenting with congenital connective tissue/myopathy overlap disorders with joint hypermobility, contractures, mild skeletal dysplasia and high myopia.

OBJECTIVE: To assess the causes of reported discordance between noninvasive prenatal testing (NIPT) and ultrasound or other clinical information. METHODS: In this retrospective, observational study, all cases in which single-nucleotide polymorphism (SNP)–based NIPT reported normal sex chromosomes and the laboratory was notified by the patient or health care provider of discordance between NIPT and observed or expected fetal sex from clinical information were reviewed. When discordances were unresolved after internal and external laboratory clerical data review or repeat ultrasound imaging, additional clinical records, genetic testing results and pregnancy outcomes were reviewed. RESULTS: Of the 1,301,117 eligible NIPT cases, fetal sex discordances were reported in 91 (0.007%; 1:14,300; 95% CI 1:11,600–1:17,800); partial or complete outcome information was available for 83 of 91 cases. In 30 of 83 (36%) cases, karyotyping was performed, and sufficient clinical information was provided to establish the diagnosis of disorders of sexual development. The disorders of sexual development were classified into three categories: 46,XY disorders of sexual development (n=19), 46,XX disorders of sexual development (n=4), and sex chromosome disorders of sexual development (n=7). In 28 of 83 (34%) cases, the cause of the apparent discrepancy was attributable to human error, predominantly phlebotomy labeling or ultrasound misassignment. In 25 of 83 cases, a diagnosis was not possible; the outcome reported was either abnormal (18/83, 22%) or no abnormalities were reported (7/83, 8%). When normal sex chromosomes were predicted by SNP-based NIPT and clinical information was discordant, disorders of sexual development were common. Internal laboratory clerical data review and re-imaging confirmed the NIPT fetal sex reports in 34% cases, providing reassurance that no further evaluation was necessary. CONCLUSION: Identification of apparent fetal sex discordances with NIPT results, and reporting this suspicion to the laboratory, provides an opportunity for further evaluation to identify the cause of apparent discordances and the involvement of a multi-disciplinary team, as necessary to prepare for postnatal care. We propose a protocol for evaluation of these cases. FUNDING SOURCE: This study was funded by Natera, Inc.

Objective: An extra haplotype is infrequently encountered in single nucleotide polymorphism(SNP)‐based non‐invasive prenatal testing (NIPT) and is usually attributed to an undetected twin or triploidy. We reviewed a large series to establish relative frequencies of these outcomes and identify alternative causes.Methods: In 515,804 women receiving NIPT from September 2017 through March 2019, all results with an extra haplotype were reviewed. Known viable and vanished twin pregnancies were excluded. For positive cases, pregnancy outcome information was sought.Results: Of 1005 results with an extra haplotype (1 in 513), pregnancy outcome was available for 773 cases: 11% were confirmed or suspected triploidy; 65% to vanished twin; 10% with pregnancy loss. Rare explanations included complete mole, chimera, undisclosed donor egg pregnancy, maternal organ transplant and one instance of maternal neoplasm. Among triploid cases that were detected and independently confirmed, 23/27 (85%) were diandric.Conclusion: SNP‐based NIPT, with detection of an extra haplotype, is 11% predictive of triploidy. For results with an extra haplotype, ultrasound is recommended to establish viability, evaluate for twins (viable or vanished), and detect findings consistent with triploidy. Review of patient history, serum screening, and ultrasound will reduce the number of CVS or amniocenteses necessary to confirm a diagnosis of triploidy.

Non-invasive prenatal screening (NIPS) based on the analysis of cell-free DNA in maternal plasma has been shown to have high sensitivity and specificity. We gathered follow-up information for pregnancies in women with test-positive NIPS results from 2014–2017 with quarterly assessments of positive predictive values (PPVs). A non-inferiority analysis with a minimum requirement of 70%/80% of expected performance for trisomy 21 and 18 was used to ensure testing met expectations. PPVs were evaluated in the context of changes in the population receiving testing. For all quarters, PPVs for trisomies 21 and 18 exceeded the requirement of > 70% of the reference PPV. Overall observed PPVs for trisomy 21, 18, 13 and monosomy X were similar for women aged <35 (90.9%, 95% Confidence Interval (CI) 88.6–92.7%) compared to women with advanced maternal age (94.5%, 95% CI 93.1–95.6%). Despite significant declines in test-positive rates from 1.18% to 0.62% for trisomy 21, and from 0.75% to 0.48% for trisomies 18, 13 and monosomy X combined, PPVs remained stable through the four-year interval. We conclude that quarterly evaluation of PPV provides an overview of past testing and helps demonstrate long-term consistency in test performance, even in the setting of increasing use by women with lower a priori risks.

Objective: Twins account for approximately 1 in 30 live births in the United States. However, there are limited clinical experience studies published in noninvasive prenatal testing (NIPT) for detecting aneuploidies in twins. This study reports the performance of an SNP‐based NIPT in the largest cohort with known outcomes for high‐risk aneuploidy results.Method: This is a retrospective analysis of 18,984 results from commercial single‐nucleotide polymorphism (SNP)‐based NIPT tests performed in twins between October 2, 2017 and December 31, 2019. Follow‐up for all 211 high‐risk cases was solicited.Results: Follow‐up outcomes were obtained in 105 cases. Positive predictive values (PPVs) for high‐risk results were 88.7% (63/71, 95% Confidence Interval [CI]: 79.0%–95.0%) for trisomy 21% and 72.7% (8/11, 95% CI: 39.0%–94.0%) for trisomy 18. The results were stratified into monozygotic (MZ) and dizygotic (DZ). The PPVs in MZ were 100% for both trisomy 21 (4/4, 95% CI: 40%–100%) and trisomy 18 (1/1, 95% CI: 2.5%–100%). No trisomy 13 cases were detected in the MZ group. The PPVs in DZ were 88.1% (59/67, 95% CI: 77.8%–94.7%), 70.0% (7/10, 95% CI: 34.8%–93.3%), and 66.7% (2/3, 95% CI: 9.4%–99.2%) for trisomy 21, trisomy 18, and trisomy 13, respectively.Conclusion: The performance of SNP‐based NIPT in this large twin cohort was comparable to previously reported twin NIPT studies. SNP‐based NIPT allows for zygosity‐based PPV assessment.