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Background: Advances in molecular technologies and in-silico variant prediction tools offer wide-ranging opportunities in diagnostic settings, yet they also present with significant limitations., Objective: Here, we contextualise the limitations of next-generation sequencing (NGS), multiplex ligation-dependent probe amplification (MLPA) and in-silico prediction tools routinely used by diagnostic laboratories by reviewing specific experiences from our diagnostic laboratory., Methods: We investigated discordant annotations and/or incorrect variant 'callings' in exons of 56 genes constituting our cardiomyopathy and connective tissue disorder NGS panels. Discordant variants and segmental duplications (SD) were queried using the National Center for Biotechnology Information (NCBI) Basic Local Alignment Search Tool and the University of California Santa Cruz genome browser, respectively, to identify regions of high homology. Discrepant variant analyses by in-silico models were re-evaluated using updated file entries., Results: We observed a 5% error rate in MYH7 variant 'calling' using MLPA, which resulted from >90% homology of the MYH7 probe-binding site to MYH6 . SDs were detected in TTN , PKP2 and MYLK . SDs in MYLK presented the highest risk (15.7%) of incorrect variant 'calling'. The inaccurate 'callings' and discrepant in-silico predictions were resolved following detailed investigation into the source of error., Conclusion: Recognising the limitations described here may help avoid incorrect diagnoses and leverage the power of new molecular technologies in diagnostic settings., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2019. No commercial re-use. See rights and permissions. Published by BMJ.)

Inherited cardiomyopathies (ICs) are a major cause of heart disease. Given their marked clinical and genetic heterogeneity, the content and clinical utility of IC multi-gene panels has been the topic of continuous debate. Our genetics diagnostic laboratory has been providing clinical diagnostic testing for ICs since 2012. We began by testing nine genes and expanded our panel by fivefold in 2015. Here, we describe the implementation of a cost-effective next-generation sequencing (NGS)-based assay for testing of IC genes, including a protocol that minimizes the amount of Sanger sequencing required to confirm variants identified by NGS, which reduces the cost and time of testing. The NGS assay was developed for the simultaneous analysis of 45 IC genes and was assessed for the impact of panel expansion on variant detection, turnaround time, and cost of testing in a cohort of 993 patients. The assay led to a considerable reduction in test cost and turnaround time. However, only a marginal increase was observed in the diagnostic yield, whereas the rate of inconclusive findings increased considerably. These findings suggest that the ongoing evaluation of gene content and monitoring of clinical utility for multi-gene tests are essential to achieve maximum clinical utility of multi-gene tests in a publicly funded health care setting., (Copyright © 2019 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.)

We calculated the amniotic fluid indexes (AFIs) of 310 women on 459 occasions. Normative data were analyzed and compared with data in several high-risk groups. In the normal gestations there was a progressive increase in AFI with advancing gestation until 32 weeks, after which there was a decline. The mean AFIs in abnormal gestations varied with the clinical diagnoses. These values were compared to those obtained by assessing amniotic fluid volume (AFV), that is a pocket more than 2 cm. There were 51 patients with abnormal AFVs. Forty-two had decreased fluid, six also had decreased AFIs; nine had increased AFVs and five (all with diabetes) also had increased AFIs. Thus, AFIs in normal pregnancies showed an orderly pattern of change with gestational age, and there was no accurate correlation between AFI and AFV. Thus, using AFV alone may lead to false interpretations of amniotic fluid status.

Objective: To assess the activity of systemic lupus erythematosus (SLE) during pregnancy and to distinguish it from preeclampsia., Methods: We prospectively measured the complement activation products Ba, Bb, SC5b-9, and C4d, as well as the conventional complement determinants C3, C4, and CH50, during pregnancy in 14 patients with SLE and 10 women with preeclampsia., Results: Four of the 14 SLE patients were considered to have disease flares, 3 occurring in the second trimester and 1 postpartum. In these patients, significant abnormalities of Ba, Bb, SC5b-9, and CH50 were noted. In contrast, measures of C4d did not distinguish between pregnant patients who had flares and those whose SLE remained stable. Although decreased values of C3 were rarely seen in the patients with stable disease, normal values of C3 during lupus pregnancy were not reliably associated with stable disease. Three of 10 non-SLE patients with preeclampsia had elevated levels of Ba; however, in each case, the CH50 level was close to or within the normal range. This was in sharp contrast to the findings observed in the 4 patients with active SLE, in whom high levels of plasma Ba were always associated with low CH50 values. Moreover, the ratio of CH50 to Ba was significantly lower in the patients with lupus flares than in the non-SLE patients with preeclampsia., Conclusion: While a decline in the CH50 level alone could otherwise be attributed to decreased synthesis of complement components, these data demonstrate that ongoing activation of the alternative complement pathway can accompany disease flares in pregnant women with SLE.

Serum amylase activity and the amylase:creatinine clearance ratio (Cam:Ccr%) are two of the most commonly used indicators for the diagnosis of pancreatitis. However, published data on the effect of pregnancy on these indicators are conflicting. Furthermore, there are no published data on the effect of pregnancy on serum lipase activity, which is considered one of the most sensitive and specific indicators of pancreatitis. A study was undertaken to determine the effect of pregnancy and gestational age on serum amylase, serum lipase and Cam:Ccr% levels and to establish a baseline of normal values for use in the diagnosis of pancreatitis in pregnant women. Serum amylase, serum lipase and Cam:Ccr% levels were determined on a sample population consisting of 175 pregnant women with gestational ages ranging from 5 to 40 weeks and on a control group of 44 reproductive-age, nonpregnant women. The study results indicated that there is no significant difference in serum amylase, serum lipase and Cam:Ccr% levels between pregnant and nonpregnant women. Cam:Ccr% showed a small but statistically significant increase in the third trimester of pregnancy.

Although stapling techniques have gained wide acceptance in general surgery, they are still not commonly used in obstetrics. U.S. Surgical Corporation has introduced a stapling device suitable for use in cesarean sections. The copolymer staples (a blend of polylactic and polyglycolic acids) maintain their tensile strength until healing occurs and absorb without producing granulation tissue. The benefits include minimal trauma to tissue and reduced operating time, blood loss and postoperative morbidity. From July 1988 to February 1989, all patients undergoing low transverse cervical cesarean sections were randomized to either group 1 with the uterine incision performed in a routine manner or group 2 with the uterine incision cut and stapled using the Stapler. The preoperative management, intraoperative technique and postoperative surveillance were similar for both groups. The uterine incision was assessed by pelvic sonography during the postpartum period. Statistical analysis was performed using Fisher's exact test and chi-square analysis. Both groups were comparable for age, race, parity, gestational age and primary diagnosis. The length of the operative procedure was significantly shorter (p less than 0.05) in the stapled group. These patients also had a statistically significantly decreased incidence of uterine incisions and lacerations. All other parameters were not significantly different in the two groups. The stapled uterine incisions were visible by ultrasonography in more patients in the stapled group throughout the postpartum period than in the sutured group. Thus, stapling of the uterine incision was an acceptable alternative to traditional suturing techniques and it was possible to visualize clearly these incisions during the postpartum period.

A prospective study was undertaken to determine the effect of twin birthweight discordancy on Apgar scores and umbilical blood acid-base parameters. Using the paired t-test, small but statistically significant differences were seen in these parameters favoring the heavier twin over its lighter sibling. These differences were also affected by birth order, with the first-born being favored.

A prospective study was undertaken which examined 179 sets of twins, 68 premature (less than 36 weeks of gestation) and 111 term. The purpose of this study was to assess differences in the acid-base status between twins related to gestational age, birth order and the time interval between twin births. Although the twin blood-gas data is within the range considered normal, statistically significant differences favoring the first-born were noted for both preterm and term twins. These differences do not depend on gestational age, route of delivery or presentation, and become evident when the interval between twin births exceeds one minute. We postulate that after delivery of the first twin, the reduced uterine size causes a decrease in the intervillous blood flow and consequently a reduction in the respiratory exchange between the second fetus, still in utero, and its placenta.

Assessment of leukocyte esterase activity in amniotic fluid for the rapid and reliable diagnosis of chorioamnionitis has been demonstrated previously. We compared in vitro inhibition of esterase activity in amniotic fluid with bacterial cultures to identify the origins of the specific esterases released by the infecting organisms. One hundred forty-one samples were tested (90 uninfected, 51 infected). Each sample was evaluated for Gram stain, cultures, and an in vitro esterase assay followed by ebelactone inhibition. Forty-two patients had positive amniotic fluid cultures. Ebelactone produced varying degrees of inhibition of esterase activity (range, 20% to 60%) in the uninfected samples and in those infected with gram-negative organisms. There was no inhibition in the samples infected with gram-positive organisms. Thus different groups of bacteria may elicit the production of different and specific esterases in infected amniotic fluid, as shown by the differences in in vitro inhibition.

Sixty-six cerclages were performed by one surgeon in 46 patients with documented cervical incompetence. The mean operative blood loss was 25 ml, and the mean operating time was 18 minutes with no postoperative morbidity. Fifty-five of the pregnancies were carried for at least 37 weeks, eight were delivered before 37 weeks, and three are continuing. Six of the eight preterm deliveries were a result of factors unrelated to cervical incompetence. The two remaining preterm births were probably also unrelated because one occurred 8 days after emergent cerclage placement and the other occurred 12 weeks after the procedure, both as a result of premature rupture of membranes. There were no cesarean deliveries related to the Shirodkar suture. The efficacy of the procedure was demonstrated by a corrected perinatal survival rate of 100% and term delivery of 88%. (Since this article was written all three of the undelivered patients were delivered of infants after 37 weeks' gestation for a term delivery rate of 88% with 100% neonatal survival.

Ballantyne syndrome is a condition of multiple causes and is characterized by maternal, placental, and fetal edema. We describe the first case of Ballantyne syndrome in a patient whose fetus had ultrasonographic demonstration of an aneurysm of Galen's vein.

Chorioamnionitis during pregnancy is a common diagnostic dilemma for the obstetrician. Fast and accurate diagnosis poses the most significant problem. Since leukocytes are known to be released in response to infections, examination of amniotic fluid for their presence is an important part of the evaluation for chorioamnionitis. These neutrophils contain several esterases that are not present in serum, urine, or vaginal secretions. The esterases are not influenced by bacteria or by commonly used drugs. We have previously described the reliability of leukocyte esterase (LE) activity for the detection of chorioamnionitis in term pregnancies uncomplicated by other diseases. A prospective study was performed to assess the presence or absence of LE activity to establish normal values in amniotic fluid at various gestational ages prior to term before its use as a possible predictor for chorioamnionitis. Sterile amniotic fluid specimens were obtained from 13 patients undergoing second trimester genetic amniocentesis (with gestational ages varying from 15 to 19 weeks) and from 11 patients with a wide variety of medical problems, undergoing amniocentesis for Rh sensitization or lung maturation studies (with gestational ages ranging from 25 to 27 weeks). All patients with ruptured membranes or preterm labor were excluded. Each amniotic fluid sample was divided into two parts, one of which was transported to the laboratory for aerobic and anaerobic cultures and the other used for LE activity detection as measured by dipstick. The LE results were retrospectively compared with the results of the cultures. LE activity was always found to be negative when the culture results were negative. negative.(ABSTRACT TRUNCATED AT 250 WORDS)

Isolated herpes simplex virus encephalitis in pregnancy is a rare illness with an elusive diagnosis. We describe the second patient to survive this disease and the first to have no sequelae, because of prompt diagnosis and treatment with acyclovir.

Congenital hereditary fructose intolerance is associated with the inability to tolerate fructose and carbohydrates, which are converted into fructose. We describe management of a pregnancy complicated by this disease in the mother and its implications for the neonate.

Accurate and rapid diagnosis of chorioamnionitis poses a major diagnostic dilemma. We previously reported that leukocyte esterase activity in amniotic fluid, as measured by dipstick assay, could be used as an aid in the diagnosis of chorioamnionitis. This study examines the effectiveness of an in vitro spectrophotometric assay of esterase activity in amniotic fluid. We define baseline levels of esterase activity in uninfected amniotic fluid and demonstrate a quantitative increase when infection is present. Fifty-seven amniotic fluid samples obtained at second- and third-trimester amniocenteses were divided into three parts, one for culture and two for a comparison of esterase activities by the dipstick and spectrophotometric methods. In this study, the spectrophotometric assay, because of its higher specificity and sensitivity in the determination of elevated esterase activity, was shown to be more reliable for predicting chorioamnionitis than either the dipstick or culture method.