Performance comparison: exome sequencing as a single test replacing Sanger sequencing.

Next generation sequencing tests are used routinely as first-choice tests in the clinic. However, systematic performance comparing the results of exome sequencing as a single test replacing Sanger sequencing of targeted gene(s) is still lacking. Performance comparison data are critically important for clinical case management. In this study, we compared Sanger-sequencing results of 258 genes to those obtained from next generation sequencing (NGS) using two exome-sequencing enrichment kits: Agilent-SureSelectQXT and Illumina-Nextera. Sequencing was performed on leukocytes and buccal-derived DNA from a single individual, and all 258 genes were sequenced a total of 11 times (using different sequencing methods and DNA sources). Sanger sequencing was completed for all exons, including flanking ± 8 bp regions. For the 258 genes, NGS mean coverage was > 20 × for > 98 and > 91% of the regions targeted by SureSelect and Nextera, respectively. Overall, 449 variants were identified in at least one experiment, and 407/449 (90.6%) were detected by all. Of the 42 discordant variants, 23 were determined as true calls, summing-up to a truth set of 430 variants. Sensitivity of true-variant detection was 99% for Sanger sequencing and 97-100% for the NGS experiments. Mean false-positive rates were 3.7E-6 for Sanger sequencing, 2.5E-6 for SureSelect-NGS and 5.2E-6 for Nextera-NGS. Our findings suggest a high overall concordance between Sanger sequencing and NGS performances. Both methods demonstrated false-positive and false-negative calls. High clinical suspicion for a specific diagnosis should, therefore, override negative results of either Sanger sequencing or NGS.

In Vitro Activity of Quaternary Ammonium in Prototheca Isolated from Clinical Bovine Mastitis Identified by Mass Spectrometry and PCR Sequencing of the cytb Gene Marker.

The in vitro algaecide activity of quaternary ammonium (QA) against Prototheca isolated from bovine clinical mastitis was investigated, in which the clinical severity was scored, milk samples were subjected to microbiological culture, and algal species were identified by molecular typing. A total of 4275 milk clinical samples of different cows from ten large dairy farms were used. Forty-four (1%) samples of cows from three dairy farms yielded growth of Prototheca, of which 88.6% (39/44) were identified as Prototheca bovis and 11.3% (5/44) as Prototheca sp. by MALDI-TOF MS, whereas 100% of the isolates were identified as P. bovis using PCR sequencing of the cytb gene. Among cows for which clinical severity scoring was available, 78.8% (26/33) and 21.2% (7/33) had mild and moderate infections, respectively, whereas no animal showed severe clinical signs. The algaecide activity of QA in Prototheca was observed in low concentrations among all isolates, in 20.4% (9/44) at 35 ppm, 36.4% (16/44) at 17 ppm, and 43.2% (19/44) at an 8 ppm, in addition to activity on three reference Prototheca strains. Overall, the study highlights the predominance of P. bovis as the causative agent of algal mastitis in bovines. Prototheca induced abnormalities preponderantly in the milk and mammary gland tissue of cows, and to our knowledge, our study is the first to apply clinical severity scoring in protothecal mastitis. In addition, the study underlines the activity of QA in low concentrations against Prototheca, indicating its potential use as an antiseptic/disinfectant in milking facilities and dairy environments.

Effect of Music Therapy on Parent-Infant Bonding Among Infants Born Preterm: A Randomized Clinical Trial.

Importance: Parent-infant bonding contributes to long-term infant health but may be disrupted by preterm birth. Objective: To determine if parent-led, infant-directed singing, supported by a music therapist and initiated in the neonatal intensive care unit (NICU), improves parent-infant bonding at 6 and 12 months. Design, Setting, and Participants: This randomized clinical trial was conducted in level III and IV NICUs in 5 countries between 2018 and 2022. Eligible participants were preterm infants (under 35 weeks' gestation) and their parents. Follow-up was conducted across 12 months (as part of the LongSTEP study) at home or in clinics. Final follow-up was conducted at 12 months' infant-corrected age. Data were analyzed from August 2022 to November 2022. Intervention: Participants randomized to music therapy (MT) plus standard care or standard care alone during NICU admission, or to MT plus standard care or standard care alone postdischarge, using computer-generated randomization (ratio 1:1, block sizes of 2 or 4 varying randomly), stratified by site (51 allocated to MT NICU, 53 to MT postdischarge, 52 to both, and 50 to neither). MT consisted of parent-led, infant-directed singing tailored to infant responses and supported by a music therapist 3 times per week throughout hospitalization or 7 sessions across 6 months' postdischarge. Main Outcome and Measure: Primary outcome was mother-infant bonding at 6 months' corrected age, measured by the Postpartum Bonding Questionnaire (PBQ), with follow-up at 12 months' corrected age, and analyzed intention-to-treat as group differences. Results: Of 206 enrolled infants with 206 mothers (mean [SD] age, 33 [6] years) and 194 fathers (mean [SD] age, 36 [6] years) randomized at discharge, 196 (95.1%) completed assessments at 6 months and were analyzed. Estimated group effects for PBQ at 6 months' corrected age were 0.55 (95% CI, -2.20 to 3.30; P = .70) for MT in the NICU, 1.02 (95% CI, -1.72 to 3.76; P = .47) for MT postdischarge, and -0.20 (95% CI, -4.03 to 3.63; P = .92) for the interaction (12 months: MT in NICU, 0.17; 95% CI, -2.71 to 3.05; P = .91; MT postdischarge, 1.78; 95% CI, -1.13 to 4.70; P = .24; interaction, -1.68; 95% CI, -5.77 to 2.41; P = .42). There were no clinically important between-group differences for secondary variables. Conclusions and Relevance: In this randomized clinical trial, parent-led, infant-directed singing did not have clinically important effects on mother-infant bonding, but was safe and well-accepted. Trial Registration: ClinicalTrials.gov Identifier: NCT03564184.

Klebsiella-induced infections in domestic species: a case-series study in 697 animals (1997-2019).

Klebsiella species, particularly K. pneumoniae, are well-known opportunistic enterobacteria related to complexity of clinical infections in humans and animals, commonly refractory to conventional therapy. The domestic animals may represent a source of the pathogenic and multidrug-resistant Klebsiella species to humans. Nevertheless, most studies involving Klebsiella-induced infections in domestic animals are restricted to case reports or outbreaks. We retrospectively investigated selected epidemiological data, clinical aspects, and in vitro susceptibility pattern of 697 non-repetitive Klebsiella infections in livestock and companion species (1997-2019). The isolates were obtained from different clinical disorders from dogs (n = 393), cattle (n = 149), horses (n = 98), cats (n = 27), pigs (n = 22), sheep (n = 5), goats (n = 2), and buffalo (n = 1), except four isolates from subclinical bovine mastitis. Urinary (223/697 = 32%), enteric (117/697 = 16.8%), mammary (85/697 = 12.2%), reproductive (85/697 = 12.2%), and respiratory disorders (67/697 = 9.6%) were the most common clinical manifestations. Other miscellaneous clinical pictures (116/697 = 16.6%) included abscesses, otitis, hepatitis, conjunctivitis, pyodermitis, sepsis, and encephalitis. Norfloxacin (183/245 = 74.7%) and gentamicin (226/330 = 68.5%) were the most effective antimicrobials. High in vitro resistance of the isolates was seen to ampicillin (326/355 = 91.8%), amoxicillin/clavulanic acid (25/62 = 40.3%), and trimethoprim/sulfamethoxazole (100/252 = 39.7), and multidrug resistance to ≥ 3 classes of antimicrobials was found in 20.4% (142/697) isolates. Wide variety of clinical manifestations of Klebsiella-induced infections was observed, with a predominance of urinary, enteric, mammary, reproductive, and respiratory tract disorders, reinforcing opportunistic behavior of agent. Poor in vitro efficacy was observed to some conventional antimicrobials and ~ 20% of isolates exhibited resistance pattern, reinforcing the need for proper use of drugs on therapy approaches in domestic animals to avoid multidrug-resistant bacteria, an emergent global concern.

Critical assessment of the latest classification of jaw cysts proposed by the World Health Organization (2017).

BACKGROUND: The fourth edition of the World Health Organization Classification of Head and Neck Tumors was published in January 2017, and includes a classification of odontogenic tumors and odontogenic cysts. The present review assesses the changes made in this new classification in relation to odontogenic and non-odontogenic jaw cysts. MATERIAL AND METHODS: An electronic search was conducted in the Cochrane Library, PubMed-MEDLINE and Scopus databases using the search terms: "odontogenic cyst" "WHO classification" "update". Studies written in English and published between January 2005 and April 2020 with a high level of scientific evidence were included, while studies not published in English, epidemiological studies, and studies with a low level of scientific evidence were excluded. RESULTS: The initial search identified 311 articles, and after the deletion of duplicates, 7 studies were selected for full-text assessment. After excluding two studies that failed to provide relevant information and had a low level of scientific evidence, 5 articles were finally included and stratified according to their level of scientific evidence based on the SORT (Strength of Recommendation Taxonomy) criteria. CONCLUSIONS: The incorporation of odontogenic and non-odontogenic cysts to the head and neck tumors classification underscores the recognition of the WHO of these important disorders of the jaws. Based on the current evidence, there is controversy as to whether odontogenic keratocysts should be regarded as cystic lesions or as neoplasms, though there is no such controversy in relation to calcifying odontogenic cysts. On the other hand, orthokeratinized odontogenic cysts have been included in the classification as a single entity differentiated from odontogenic keratocysts, while residual cysts have been removed from the classification. Key words:Odontogenic cyst, WHO classification, pseudocyst.

Phylogenetic history of patrilineages rare in northern and eastern Europe from large-scale re-sequencing of human Y-chromosomes.

The most frequent Y-chromosomal (chrY) haplogroups in northern and eastern Europe (NEE) are well-known and thoroughly characterised. Yet a considerable number of men in every population carry rare paternal lineages with estimated frequencies around 5%. So far, limited sample-sizes and insufficient resolution of genotyping have obstructed a truly comprehensive look into the variety of rare paternal lineages segregating within populations and potential signals of population history that such lineages might convey. Here we harness the power of massive re-sequencing of human Y chromosomes to identify previously unknown population-specific clusters among rare paternal lineages in NEE. We construct dated phylogenies for haplogroups E2-M215, J2-M172, G-M201 and Q-M242 on the basis of 421 (of them 282 novel) high-coverage chrY sequences collected from large-scale databases focusing on populations of NEE. Within these otherwise rare haplogroups we disclose lineages that began to radiate ~1-3 thousand years ago in Estonia and Sweden and reveal male phylogenetic patterns testifying of comparatively recent local demographic expansions. Conversely, haplogroup Q lineages bear evidence of ancient Siberian influence lingering in the modern paternal gene pool of northern Europe. We assess the possible direction of influx of ancestral carriers for some of these male lineages. In addition, we demonstrate the congruency of paternal haplogroup composition of our dataset with two independent population-based cohorts from Estonia and Sweden.

Utility of Ethyl-Glucuronide for Evaluation of Abstinence in Patients Presenting for Liver Transplantation.

BACKGROUND: Cirrhosis as a result of alcohol-related liver disease is one of the most common indications for liver transplantation (LT) in Spain. Patients presenting for LT should be checked for alcohol abuse in clinical interviews and use of laboratory tests to confirm abstinence. The ethyl-glucuronide (EtG) test is very sensitive and can be positive in urine up to 5 days after consumption. Our main objective is to know the rate of alcohol abstinence by using the urine EtG test in patients evaluated for LT and to assess its correlation with the clinical interviews and laboratory test. METHODS: We conducted a prospective analysis of the results of the EtG in urine of patients evaluated for LT from January 2017 to March 2019 and its correlation with the medical and psychiatric interviews and with the laboratory test. RESULTS: We included 160 patients who were referred to LT evaluation. Among all cases, 84.1% were men, with an average age of 57.8 years. Alcohol-related liver disease was the most frequent cause (64.1%). Urine-EtG was positive in 10 patients (6.2%), 9 of them in patients with ALD and 1 in a patient with hepatitis C virus. The alcohol consumption was recognized by 80% of the patients in the clinical interview. Cases with positive EtG had higher levels of analytical parameters than those with a negative test. CONCLUSIONS: In our series, 6.2% of patients referred for LT evaluation had recently consumed alcohol. The determination of EtG in urine is probably an effective and objective technique in the detection of alcohol consumption to ensure abstinence in the LT candidates.

The genetic variation in the R1a clade among the Ashkenazi Levites' Y chromosome.

Approximately 300,000 men around the globe self-identify as Ashkenazi Levites, of whom two thirds were previously shown to descend from a single male. The paucity of whole Y-chromosome sequences precluded conclusive identification of this ancestor's age, geographic origin and migration patterns. Here, we report the variation of 486 Y-chromosomes within the Ashkenazi and non-Ashkenazi Levite R1a clade, other Ashkenazi Jewish paternal lineages, as well as non-Levite Jewish and non-Jewish R1a samples. Cumulatively, the emerging profile is of a Middle Eastern ancestor, self-affiliating as Levite, and carrying the highly resolved R1a-Y2619 lineage, which was likely a minor haplogroup among the Hebrews. A star-like phylogeny, coalescing similarly to other Ashkenazi paternal lineages, ~1,743 ybp, suggests it to be one of the Ashkenazi paternal founders; to have expanded as part of the overall Ashkenazi demographic expansion, without special relation to the Levite affiliation; and to have subsequently spread to non-Ashkenazi Levites.