Chromosomal microarray detects genetic risks of neurodevelopmental disorders in newborns with congenital heart disease

Objective: To compare the genetic testing results of neonates with CHD by chromosomal microarray to karyotyping and fluorescence in situ hybridisation analysis. Methods: This was a single-centre retrospective comparative study of patients with CHD and available genetic testing results admitted to the cardiac ICU between January, 2004 and December, 2017. Patients from 2004 to 2010 were tested by karyotyping and fluorescence in situ hybridisation analysis, while patients from 2012 to 2017 were analysed by chromosomal microarray. Results: Eight-hundred and forty-nine neonates with CHD underwent genetic testing, 482 by karyotyping and fluorescence in situ hybridization, and 367 by chromosomal microarray. In the karyotyping and fluorescence in situ hybridisation analysis group, 86/482 (17.8%) had genetic abnormalities detected, while in the chromosomal microarray group, 135/367 (36.8%) had genetic abnormalities detected (p < 0.00001). Of patients with abnormal chromosomal microarray results, 41/135 (30.4%) had genetic abnormality associated with neurodevelopmental disorders that were exclusively identified by chromosomal microarray. Conotruncal abnormalities were the most common diagnosis in both groups, with karyotyping and fluorescence in situ hybridisation analysis detecting genetic abnormalities in 26/160 (16.3%) patients and chromosomal microarray detecting abnormalities in 41/135 (30.4%) patients (p = 0.004). In patients with d-transposition of the great arteries, 0/68 (0%) were found to have genetic abnormalities by karyotyping and fluorescence in situ hybridisation compared to 7/54 (13.0%) by chromosomal microarray. Conclusions: Chromosomal microarray identified patients with CHD at genetic risk of neurodevelopmental disorders, allowing earlier intervention with multidisciplinary care and more accurate pre-surgical prognostic counselling.

The human gut archaeome: identification of diverse haloarchaea in Korean subjects

Background: Archaea are one of the least-studied members of the gut-dwelling autochthonous microbiota. Few studies have reported the dominance of methanogens in the archaeal microbiome (archaeome) of the human gut, although limited information regarding the diversity and abundance of other archaeal phylotypes is available.Results: We surveyed the archaeome of faecal samples collected from 897 East Asian subjects living in South Korea. In total, 42.47% faecal samples were positive for archaeal colonisation; these were subsequently subjected to archaeal 16S rRNA gene deep sequencing and real-time quantitative polymerase chain reaction-based abundance estimation. The mean archaeal relative abundance was 10.24% ± 4.58% of the total bacterial and archaeal abundance. We observed extensive colonisation of haloarchaea (95.54%) in the archaea-positive faecal samples, with 9.63% mean relative abundance in archaeal communities. Haloarchaea were relatively more abundant than methanogens in some samples. The presence of haloarchaea was also verified by fluorescence in situ hybridisation analysis. Owing to large inter-individual variations, we categorised the human gut archaeome into four archaeal enterotypes.Conclusions: The study demonstrated that the human gut archaeome is indigenous, responsive, and functional, expanding our understanding of the archaeal signature in the gut of human individuals.

Sterile osteomyelitis in the ulnar diaphysis of a young indoor cat

Case summary A 3-year-old neutered male indoor British Shorthair cat was referred for a 2-week history of intermittent right forelimb lameness. Radiographic examination showed a diaphyseal monostotic, expansile, fusiform, lytic lesion in the right ulna. CT further defined the lesion and also demonstrated ipsilateral pulmonary consolidation. Histology was conclusive of osteomyelitis, and microbiology and fluorescence in situ hybridisation analysis (FISH) were negative on aerobic and anaerobic bacterial culture, as well as fungal culture. Clinical and radiographic improvement was seen after anti-inflammatory treatment and a short initial period of antibiosis. Relevance and novel information This is an unusual monostotic diaphyseal cortical location for osteomyelitis in cats and, moreover, may represent a rare case of sterile osteomyelitis. To our knowledge, non-traumatic osteomyelitis in this location in cats has not been reported in the veterinary literature.

How common is germinal mosaicism that leads to premeiotic aneuploidy in the female?

Purpose Molecular cytogenetic analysis has confirmed that a proportion of apparently meiotic aneuploidy may be present in the germ cells prior to the onset of meiosis, but there is no clear perception of its frequency. The aim of this review is to assess the evidence for premeiotic aneuploidy from a variety of sources to arrive at an estimate of its overall contribution to oocyte aneuploidy in humans. Methods Relevant scientific literature was covered from 1985 to 2018 by searching PubMed databases with search terms: gonadal/germinal mosaicism, ovarian mosaicism, premeiotic aneuploidy, meiosis and trisomy 21. Additionally, a key reference from 1966 was included. Results Data from over 9000 cases of Down syndrome showed a bimodal maternal age distribution curve, indicating two overlapping distributions. One of these matched the pattern for the control population, with a peak at about 28 years and included all cases that had occurred independently of maternal age, including those due to germinal mosaicism, about 40% of the cohort. The first cytological proof of germinal mosaicism was obtained by fluorescence in situ hybridisation analysis. Comparative genomic hybridisation analysis of oocyte chromosomes suggests an incidence of up to 15% in premeiotic oocytes. Direct investigation of fetal ovarian cells led to variable results for chromosome 21 mosaicism. Conclusions Oocytes with premeiotic errors will significantly contribute to the high level of preimplantation and prenatal death. Data so far available suggests that, depending upon the maternal age, up to 40% of aneuploidy that is present in oocytes at the end of meiosis I may be due to germinal mosaicism.

Agar pre-embedding of small skin biopsies: real-life benefits and challenges in high throughput pathology laboratories

Paraffin embedding of small, thin tissue samples requires specific expertise for optimal orientation before tissue sectioning. This study evaluates the real-life utility of the agar pre-embedding technique for small skin biopsies with regards to lengthening of work times, problems in orientation (re-embedding) and ancillary techniques (immunohistochemistry and in situ hybridisation) between two high work flow pathology laboratories, one of which routinely uses the agar pre-embedding technique and one which does not. The mean time required for pre-embedding in agar was 30.4 s, but time for paraffin embedding for agar pre-embedded samples was shorter than the traditional method (177 vs 296 s; p<0.005). The number of skin samples requiring re-embedding was significantly higher with the traditional embedding method (p<0.005). No problems in immunoreactivity were observed in all 1900 reactions performed with 17 different antibodies. Fluorescence in situ hybridisation analysis was optimised with a prolonged protease K incubation time (21 vs 18 min).

Influence of the method of whole wheat inclusion on performance and caecal microbiota profile of broiler chickens

SummaryA study was conducted to investigate the effect of method of whole wheat inclusion on performance and caecal microbiota profile of broiler chickens. Fluorescencein situhybridisation analysis was used to characterise the microbiota by using genus-specific probes. Three treatments, namely, ground wheat (GW) or 200 g/kg whole wheat (WW) replacing GW before or after pelleting were evaluated. A total of 144, one-day-old male broilers (Ross 308) were allocated to 18 cages (eight broilers per cage) based on body weight and six cages were randomly assigned to each treatment. The diets were offeredad libitumfrom day 11 to 35 post-hatch. The WW fed birds, regardless of the method of inclusion, resulted in poorer weight gain (P < 0.05) and reduced feed intake (P < 0.001), but a similar feed per gain (P > 0.05) compared to those fed the GW diet. The WW diet, regardless to the method of inclusion, had no effect (P > 0.05) on the populations ofLactobacillusandBacteroides spp.compared with the GW diet. TheBifidobacterium spp.population was higher (P < 0.05) in birds fed the GW diet compared with WW feeding, regardless of the method of inclusion. A reduction (P < 0.05) in the numbers of pathogenicClostridiumandCampylobacter spp.were observed in caecal samples from birds fed WW diets, regardless of method of inclusion, compared with those fed the GW diet, which was attributed to increased gizzard activity. Birds fed WW diets, regardless to the method of inclusion, showed a reduction in gizzard pH (P < 0.05), microbial gas production (P < 0.05), and an increase in gizzard weight (P < 0.05) relative to the GW treatment. The results indicated that the gizzard has an important function as a barrier organ, one that prevents pathogenic bacteria from entering the distal digestive tract.

Achondroplasia with SRY-positive 46, XX disorder of sex development: an extremely rare association

Summary A 40-year-old man with achondroplasia presented with symptoms of hypogonadism, low libido and gynaecomastia. He was found to have hypergonadotropic hypogonadism, and karyotype and fluorescent in situ hybridisation analysis showed SRY-positive 46, XX disorder of sex development (DSD). He was tested to have the common activating mutation of the FGFR3 gene implicated in achondroplasia, indicating that he had the two rare conditions independently, with an extremely low incidence of 1 in 400 million. This, to the best of our knowledge, is the first report of an individual having these two rare conditions concurrently. This case highlights that individuals with achondroplasia should have normal sexual development, and in those presenting with incomplete sexual maturation or symptoms of hypogonadism should prompt further evaluation. We also propose a plausible link between achondroplasia and 46, XX DSD through the intricate interactions between the SRY, SOX9 and FGFR9 gene pathways. Learning points: The SOX9 and FGF9 genes, which are upregulated by the SRY gene, are important in both sex determination in the embryo, as well as endochondral bone growth. Patients with achondroplasia should have normal sexual development and function in the absence of other confounding factors. Patients with achondroplasia who present with symptoms and signs of abnormal sexual development and/or hypogonadism should be appropriately investigated for other causes.

Antenatal microbial colonisation of mammalian gut

ABSTRACTThe widely accepted dogma of intrauterine sterility and initial colonisation of the newborn during birth has been blurred by recent observations of microbial presence in meconium, placenta and amniotic fluid. Given the importance of a maternal-derived in utero infant seeding, it is crucial to exclude potential environmental or procedural contaminations, and to assess foetal colonisation before parturition. To ascertain antenatal microbial colonisation in mammals, we analysed sterilely collected intestinal tissues from rodent foetuses in parallel with experimental controls, and tissues from autoptic human foetuses. Next generation sequencing (NGS) showed the presence of pioneer microbes in both rat and human intestines, as well as in rodent placentas and amniotic fluids. Live microbes were isolated from culture-dependent analyses from homogenized rat foetal intestines.Microbial communities showed foetus- and dam-dependent clustering, confirming the high interindividual variability of microbiota even in the antenatal period. Fluorescent in situ hybridisation analysis confirmed the microbes’ existence in the lumen of the developing gut.These findings have vast implications for an emerging field of enhancing the management of healthy pregnancies, and for understanding how the infant microbiome starts and it is thus shaped.