Prenatal diagnosis of PORCN-related developmental syndrome in a fetus: A novel phenotype.

We report a case of a female fetus born to an unrelated couple with a complex fetal phenotype of a pleural effusion, a cardiac malformation, and syndactyly of the toes. Prenatal exome sequencing identified a variant of uncertain significance in the PORCN gene that was upgraded to likely pathogenic following postnatal clinical examination. The phenotype described in cases with variants in the PORCN gene is often associated with findings that cannot be prospectively diagnosed by ultrasonography. This is the first report of a prenatal phenotype involving a fetal effusion associated with variants in the PORCN gene, with skeletal findings identified later in gestation on ultrasonography. The diagnosis was confirmed on neonatal examination.

Diagnostic and perinatal outcomes in consanguineous couples with a structural fetal anomaly: A cohort study.

INTRODUCTION: Consanguineous unions occur when a couple are related outside marriage and is associated with adverse genetic and perinatal outcomes for affected offspring. The objectives of this study were to evaluate: (i) background characteristics, (ii) uptake of prenatal and postnatal investigation and (iii) diagnostic outcomes of UK consanguineous couples presenting with a fetal structural anomaly. MATERIAL AND METHODS: This was a retrospective and partly prospective cohort study comparing consanguineous (n = 62) and non-consanguineous (n = 218) pregnancies with current or previous fetal structural anomalies reviewed in a UK prenatal genetic clinic from 2008 to 2019. Outcomes were compared using odds ratios (OR). RESULTS: Most consanguineous couples were of Pakistani ethnicity (odds ratio [OR] 29, 95% confidence interval [95% CI] 13-62) and required use of an interpreter [OR 9, 95% CI 4-20). In the consanguineous group, the uptake of prenatal invasive testing was lower (OR 0.4, 95% CI 0.2-0.7) and the number declining follow up was greater (OR 10, 95% CI 3-34) than in the non-consanguineous group. This likely explained the lower proportion of consanguineous couples where a final definitive unifying diagnosis to explain the fetal structural anomalies was reached (OR 0.3, 95% CI 0.2-0.6). When a diagnosis was obtained in this group, it was always postnatal and most often using genomic sequencing technologies (OR 6, 95% CI 1-27). The risk of perinatal death was greater (OR 3, 95% CI 1-6) in the consanguineous group, as was the risk of fetal structural anomaly recurrence in a subsequent pregnancy (OR 4, 95% CI 1-13). There was no difference in the uptake of perinatal autopsy or termination of pregnancy between groups. CONCLUSIONS: Consanguineous couples are a vulnerable group in the prenatal setting. Although adverse perinatal outcomes in this group are more common secondary to congenital anomalies, despite the evolution of genomic sequencing technologies, due to a lower uptake of prenatal testing it is less likely that a unifying diagnosis is obtained and recurrence can occur. There is a need for proactive genetic counseling and education from the multidisciplinary team, addressing language barriers as well as religious and cultural beliefs in an attempt to optimize reproductive options.

Refining the phenotype associated with GNB1 mutations: Clinical data on 18 newly identified patients and review of the literature.

De novo germline mutations in GNB1 have been associated with a neurodevelopmental phenotype. To date, 28 patients with variants classified as pathogenic have been reported. We add 18 patients with de novo mutations to this cohort, including a patient with mosaicism for a GNB1 mutation who presented with a milder phenotype. Consistent with previous reports, developmental delay in these patients was moderate to severe, and more than half of the patients were non-ambulatory and nonverbal. The most observed substitution affects the p.Ile80 residue encoded in exon 6, with 28% of patients carrying a variant at this residue. Dystonia and growth delay were observed more frequently in patients carrying variants in this residue, suggesting a potential genotype-phenotype correlation. In the new cohort of 18 patients, 50% of males had genitourinary anomalies and 61% of patients had gastrointestinal anomalies, suggesting a possible association of these findings with variants in GNB1. In addition, cutaneous mastocytosis, reported once before in a patient with a GNB1 variant, was observed in three additional patients, providing further evidence for an association to GNB1. We will review clinical and molecular data of these new cases and all previously reported cases to further define the phenotype and establish possible genotype-phenotype correlations.

Dye Encapsulation in Polynorbornene Micelles.

The encapsulation efficiency of high-Tg polynorbornene micelles was probed with a hydrophobic dye 2,6-diiodoboron-dipyrromethene (BODIPY). Changes in the visible absorption spectra of aggregated versus monomeric dye molecules provided a probe for assessing encapsulation. Polynorbornene micelles are found to be capable of loading up to one BODIPY dye per ten polymers. As the hydrophilic block size increased in the polymeric amphiphiles, more of the dye was incorporated within the micelles. This result is consistent with the dye associating with the polymer backbone in the shell of the micelles. The encapsulation rate varied significantly with temperature, and a slight dependence on micellar morphology was also noted. Additionally, we report a 740 µs triplet lifetime for the encapsulated BODIPY dye. The lifetime is the longest ever recorded for a BODIPY triplet excited state at room temperature and is attributed to hindered triplet-triplet annihilation in the high-viscosity micellar shell.

Chapter 2: Non-invasive prenatal diagnosis.

Non-invasive prenatal diagnosis of monogenic disorders is becoming integrated into routine clinical care for many indications. This is carried out by testing cell-free DNA extracted from the plasma portion of a maternal blood sample. The cell-free DNA is low in concentration, and consists of a mixture of maternal and fetally-derived DNA which are not easy to separate. Methods used therefore need to be rapid, sensitive and specific, including real-time PCR, digital PCR and next generation sequencing with complex algorithms. Testing may be required for pregnancies with an increased chance of a monogenic disorder due to family history or carrier status, or where there are specific abnormalities identified by ultrasound scan. In these situations, testing is considered to be diagnostic and therefore does not require confirmation by invasive testing. With increased access to genomic technologies, and more diagnoses for rare disease patients, future demand for NIPD and possibilities during pregnancy will continue.

Börjeson-Forssman-Lehmann syndrome: delineating the clinical and allelic spectrum in 14 new families.

Börjeson-Forssman-Lehmann syndrome (BFLS) is an X-linked intellectual disability syndrome caused by variants in the PHF6 gene. We ascertained 19 individuals from 15 families with likely pathogenic or pathogenic PHF6 variants (11 males and 8 females). One family had previously been reported. Six variants were novel. We analysed the clinical and genetic findings in our series and compared them with reported BFLS patients. Affected males had classic features of BFLS including intellectual disability, distinctive facies, large ears, gynaecomastia, hypogonadism and truncal obesity. Carrier female relatives of affected males were unaffected or had only mild symptoms. The phenotype of affected females with de novo variants overlapped with the males but included linear skin hyperpigmentation and a higher frequency of dental, retinal and cortical brain anomalies. Complications observed in our series included keloid scarring, digital fibromas, absent vaginal orifice, neuropathy, umbilical hernias, and talipes. Our analysis highlighted sex-specific differences in PHF6 variant types and locations. Affected males often have missense variants or small in-frame deletions while affected females tend to have truncating variants or large deletions/duplications. Missense variants were found in a minority of affected females and clustered in the highly constrained PHD2 domain of PHF6. We propose recommendations for the evaluation and management of BFLS patients. These results further delineate and extend the genetic and phenotypic spectrum of BFLS.

A pilot study investigating feasibility of mainstreaming germline BRCA1 and BRCA2 testing in high-risk patients with breast and/or ovarian cancer in three tertiary Cancer Centres in Ireland.

In the Republic of Ireland (ROI), BRCA1/BRCA2 genetic testing has been traditionally undertaken in eligible individuals, after pre-test counselling by a Clinical Geneticist/Genetic Counsellor. Clinical Genetics services in ROI are poorly resourced, with routine waiting times for appointments at the time of this pilot often extending beyond a year. The consequent prolonged waiting times are unacceptable where therapeutic decision-making depends on the patient's BRCA status. "Mainstreaming" BRCA1/BRCA2 testing through routine oncology/surgical clinics has been implemented successfully in other centres in the UK and internationally. We aimed to pilot this pathway in three Irish tertiary centres. A service evaluation project was undertaken over a 6-month period between January and July 2017. Eligible patients, fulfilling pathology and age-based inclusion criteria defined by TGL clinical, were identified, and offered constitutional BRCA1/BRCA2 testing after pre-test counselling by treating clinicians. Tests were undertaken by TGL Clinical. Results were returned to clinicians by secure email. Onward referrals of patients with uncertain/pathogenic results, or suspicious family histories, to Clinical Genetics were made by the treating team. Surveys assessing patient and clinician satisfaction were sent to participating clinicians and a sample of participating patients. Data was collected with respect to diagnostic yield, turnaround time, onward referral rates, and patient and clinician feedback. A total of 101  patients underwent diagnostic germline BRCA1/BRCA2 tests through this pathway. Pathogenic variants were identified in 12 patients (12%). All patients in whom variants were identified were appropriately referred to Clinical Genetics. At least 12 additional patients with uninformative BRCA1/BRCA2 tests were also referred for formal assessment by Clinical Geneticist or Genetic Counsellor. Issues were noted in terms of time pressures and communication of results to patients. Results from a representative sample of participants completing the satisfaction survey indicated that the pathway was acceptable to patients and clinicians. Mainstreaming of constitutional BRCA1/BRCA2 testing guided by age- and pathology-based criteria is potentially feasible for patients with breast cancer as well as patients with ovarian cancer in Ireland.

Effects of Social Housing on Dairy Calf Social Bonding.

Social housing for dairy calves has a range of benefits for social development, yet there is limited understanding of how social bonds form early in life. We characterized effects of early life social contact on the development of social preference for calves varying in familiarity. A total of 40 calves were tested in a social preference test at 4 weeks of age to assess the formation of social bonds and preference for their peers. Within an open-field social preference test, focal calves were presented with two stimulus calves, one 'more familiar' and one 'less familiar'. We found that pair-housed calves spent more time in close proximity with either stimulus calf and had a greater preference for their pen-mate, compared to another calf reared within visual contact. Individually housed calves exhibited no preference for calves reared within visual but not physical contact compared to calves that were completely unfamiliar. Of the calves that approached both stimulus calves, individually housed calves that approached the 'less familiar' calf first spent less time near the 'more familiar' calf, whereas behavior of pair-housed calves was not affected by the first calf approached. These results suggest that physical contact is necessary for the development of social bonds in young dairy calves, and early life social housing may support the development of normal social behavior in dairy cattle.

Prognostic Value of Left and right ventricular deformation strain analysis on Acute Cellular rejection in Heart Transplant recipients: A 6-year outcome study.

PURPOSE: Two-dimensional (2D) strain analysis is a sensitive method for detecting myocardial dysfunction in acute cellular rejection (ACR) from post-transplant complications. This study aims to evaluate the utility of novel left (LV) and right ventricular (RV) strain parameters for prognostic risk stratification associated with ACR burden at 1-year post transplantation. METHODS: 128 Heart transplant patients, assessed between 2012 and 2018, underwent transthoracic echocardiography and endomyocardial biopsy. 2D strain analysis was performed and history of rejection burden was assessed and grouped according to ACR burden at 1-year post transplantation. The primary endpoint was all-cause mortality at 6-years follow up. RESULTS: 21 patients met primary the endpoint. Multivariate analysis of 6-year all-cause mortality showed LV global longitudinal strain (LV GLS) (Hazard Ratio [HR] = 1.21, CI = 1.06-1.49), LV early diastolic strain rate (LV ESr) (HR = 1.31, CI = 1.12-1.54), RV GLS (HR = 1.12, CI = 1.02-1.25) and RV ESr (HR = 1.26, CI = 1.12-1.47) were significant predictors of outcome. Univariate analysis also showed LV GLS, LV ESr, RV GLS and RV ESr were significant predictors of outcome. Optimal cut-off for predicting 6-year mortality for LV GLS by receive operator characteristic was 15.5% (sensitivity: 92%, specificity: 79%). Significant reductions (p < 0.05) in LV GLS, RV GLS and LV and RV ESr between rejection groups were seen. CONCLUSIONS: Non-invasive LV and RV strain parameters are predictors of mortality in post-transplant patient with ACR. LV GLS and LV ESr are superior to other strain and conventional echo parameters.

The prenatal exome - a door to prenatal diagnostics?

Introduction: Prenatal exome sequencing (ES) allows parents the opportunity to obtain arapid molecular diagnosis of monogenic etiology when their fetus is found to have structural anomalies detected on prenatal ultrasound. Such information can improve antenatal and neonatal counseling, decision-making and management, and expand reproductive options in subsequent pregnancies.Areas covered: This review appraises the evidence, from acomprehensive search of bibliographic databases, for the introduction of ES into the fetal medicine care pathway when investigating congenital malformations. The perspectives of clinical geneticists, clinical scientists, fetal medicine specialists, and patients are explored in relation to the novel investigation and the benefits and challenges of its use in ongoing pregnancies with particular reference to UK medical practice.Expert opinion: ES provides agenetic diagnosis for more than 1 in 10 fetuses with structural differences on ultrasound and normal conventional tests (karyotype or chromosomal microarray) in carefully selected cases. The diagnostic rate increases for certain phenotypes and can range between 6% and 80% where conventional cytogenetics have not detected adiagnosis. Expert oversight is required to ensure that patients receive high-quality, evidence-based care and accurate counseling, supported by amultidisciplinary team familiar with the test and its implications.

Multi-Organ Dysfunction in Cerebral Palsy.

Cerebral Palsy (CP) describes a heterogenous group of non-progressive disorders of posture or movement, causing activity limitation, due to a lesion in the developing brain. CP is an umbrella term for a heterogenous condition and is, therefore, descriptive rather than a diagnosis. Each case requires detailed consideration of etiology. Our understanding of the underlying cause of CP has developed significantly, with areas such as inflammation, epigenetics and genetic susceptibility to subsequent insults providing new insights. Alongside this, there has been increasing recognition of the multi-organ dysfunction (MOD) associated with CP, in particular in children with higher levels of motor impairment. Therefore, CP should not be seen as an unchanging disorder caused by a solitary insult but rather, as a condition which evolves over time. Assessment of multi-organ function may help to prevent complications in later childhood or adulthood. It may also contribute to an improved understanding of the etiology and thus may have an implication in prevention, interventional methods and therapies. MOD in CP has not yet been quantified and a scoring system may prove useful in allowing advanced clinical planning and follow-up of children with CP. Additionally, several biomarkers hold promise in assisting with long-term monitoring. Clinicians should be aware of the multi-system complications that are associated with CP and which may present significant diagnostic challenges given that many children with CP communicate non-verbally. A step-wise, logical, multi-system approach is required to ensure that the best care is provided to these children. This review summarizes multi-organ dysfunction in children with CP whilst highlighting emerging research and gaps in our knowledge. We identify some potential organ-specific biomarkers which may prove useful in developing guidelines for follow-up and management of these children throughout their lifespan.

The genetic and biochemical basis of trimethylaminuria in an Irish cohort.

BACKGROUND: Inherited trimethylaminuria (TMAU), a rare genetic disorder of hepatic metabolism of trimethylamine (TMA) causing excessive accumulation of malodorous trimethylamine (TMA), is a socially distressing disorder. Diagnosis is made by biochemical analysis of urine, with the calculation of flavin monooxygenase trimethylamine conversion capacity. Genetic testing, sequencing the entire coding region of the FMO3 gene has been recommended for affected individuals who convert less than 90% of the total TMA load to TMAO. METHODS: Genetic analysis was undertaken for 13 Irish patients with TMAU of varying phenotypic severity (three severe, six moderate, and four mild). RESULTS: A genetic diagnosis was made for seven patients, including for five of the nine moderate to severely affected cases. We noted the c.913G>T;p.(Glu305*) and c.458C>T;p.(Pro153Leu) mutations in this Irish population with severe TMAU which is consistent with our earlier findings in Australian and North American families of Irish and British descent.Three individuals were noted to be homozygous for the common variant haplotype c.472G>A;923A>G;p.(Glu158Lys);(Glu308Gly). We also identified three novel variants in this population, which are likely to be pathogenic: c.682G>A;p(Gly228Ser), c.694G>T:p(Asp232Tyr), and c.989G>A;p.(Gly330Glu). CONCLUSION: Urinary biochemical analysis probably remains the first line diagnostic approach to classify the various types of TMAU. FMO3 gene analysis is likely only to be informative for certain presentations of TMAU.

Resonance Raman Spectroscopy of the T1 Triplet Excited State of Oligothiophenes.

The characterization of triplet excited states is essential for research on organic photovoltaics and singlet fission. We report resonance Raman spectra of two triplet oligothiophenes with n-alkyl substituents, a tetramer and hexamer. The spectra of the triplets are more complex than the ground state, and we find that density functional theory calculations are a useful starting point for characterizing the bands. The spectra of triplet tetrathiophene and hexathiophene differ significantly from one another. This observation is consistent with a T1 excitation that is delocalized over at least five rings in long oligomers. Bands in the 500-800 cm(-1) region are greatly diminished for an aggregated sample of hexathiophene, likely caused by fast electronic dephasing. These experiments highlight the potential of resonance Raman spectroscopy to unequivocally detect and characterize triplets in thiophene materials. The vibrational spectra can also serve as rigorous standards for evaluating computational methods for excited-state molecules.