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In current sensorimotor theories pertaining to speech perception, there is a notable emphasis on the involvement of the articulatory-motor system in the processing of speech sounds. Using ultra-high field diffusion-weighted imaging at 7 Tesla, we visualized the white matter tracts connected to areas activated during a simple speech-sound production task in 18 healthy right-handed adults. Regions of interest for white matter tractography were individually determined through 7T functional MRI (fMRI) analyses, based on activations during silent vocalization tasks. These precentral seed regions, activated during the silent production of a lip-vowel sound, demonstrated anatomical connectivity with posterior superior temporal gyrus areas linked to the auditory perception of phonetic sounds. Our study provides a macrostructural foundation for understanding connections in speech production and underscores the central role of the articulatory motor system in speech perception. These findings highlight the value of ultra-high field 7T MR acquisition in unraveling the neural underpinnings of speech., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Introduction: X-linked dystonia-parkinsonism (XDP) is a neurodegenerative disorder that may result in severe speech impairment. The literature suggests that there are differences in the speech of individuals with XDP and healthy controls. This study aims to examine the motor speech characteristics of the mixed dystonia-parkinsonism phase of XDP., Method: We extracted acoustic features representing coordination, consistency, speed, precision, and rate from 26 individuals with XDP and 26 controls using Praat, MATLAB, and R software. Group demographics were compared using descriptive statistics. A one-way analysis of variance (ANOVA) with Tukey's post hoc test was used to test for acoustic differences between the two groups., Results: The XDP group had significantly lower consistency, speed, precision, and rate than controls (p < 0.05). For coordination, the XDP group had a smaller ratio of pause duration during transitions when compared to controls., Discussion: To our knowledge, this study is the first to describe the motor speech characteristics of the mixed dystonia-parkinsonism phase of XDP. The motor speech of mixed dystonia-parkinsonism XDP is similar to prior characterizations of mixed hyperkinetic-hypokinetic dysarthria with noted differences in articulatory coordination, consistency, speed, precision, and rate from healthy controls. Identifying the motor speech components of all three phenotypes of XDP (i.e., dystonia-dominant phase, parkinsonism-dominant phase, and mixed dystonia-parkinsonism phase) is needed to establish markers of speech impairment to track disease progression., Competing Interests: Declaration of competing interest This work was supported by the Massachusetts General Hospital Collaborative Center for X-linked Dystonia-Parkinsonism, Boston, MA; the National Institutes of Health (NIH)National Institute on Deafness and Other Communication Disorders [grant number K24DC0016312]; and the NIHNational Institute of Neurological Disorders and Stroke [grant number K23NS123369]. The funding sources did not participate in the study design; data collection, analysis, and interpretation; report writing; or decision to submit the article for publication. The authors declare that there are no conflicts of interest relevant to this work., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Horizontal gene transfer by plasmids can confer metabolic capabilities that expand a host cell's niche. Yet, it is less understood whether the coalescence of specialized catabolic functions, antibiotic resistances and metal resistances on plasmids provides synergistic benefits. In this study, we report whole-genome assembly and phenotypic analysis of five Salmonella enterica strains isolated in the 1980s from milk powder in Munich, Germany. All strains exhibited the unusual phenotype of lactose-fermentation and encoded either of two variants of the lac operon. Surprisingly, all strains encoded the mobilized colistin resistance gene 9 ( mcr-9 ), long before the first report of this gene in the literature. In two cases, the mcr-9 gene and the lac locus were linked within a large gene island that formed an IncHI2A-type plasmid in one strain but was chromosomally integrated in the other strain. In two other strains, the mcr-9 gene was found on a large IncHI1B/IncP-type plasmid, whereas the lac locus was encoded on a separate chromosomally integrated plasmidic island. The mcr-9 sequences were identical and genomic contexts could not explain the wide range of colistin resistances exhibited by the Salmonella strains. Nucleotide variants did explain phenotypic differences in motility and exopolysaccharide production. The observed linkage of mcr-9 to lactose metabolism, an array of heavy-metal detoxification systems, and other antibiotic resistance genes may reflect a coalescence of specialized phenotypes that improve the spread of colistin resistance in dairy facilities, much earlier than previously suspected.

Background: Assessment of motor function restoration following face transplant (FT) is difficult, as standardized, bilateral tests are lacking. This study aims to bolster support for software-based analysis through international collaboration., Methods: FaceReader (Noldus, Wageningen, The Netherlands), a facial expression analysis software, was used to analyze posttransplant videos of eight FT patients from Boston, Massachusetts (range, 1 to 9 years after transplant), two FT patients from Helsinki, Finland (range, 3 to 4 years after transplant), and three FT patients from Antalya, Turkey (range, 6.5 to 8.5 years after transplant). Age-matched healthy controls from respective countries had no history of prior facial procedures. Videos contained patients and controls performing facial expressions evaluated by software analysis using the Facial Action Coding System. Facial movements were assigned intensity score values between 0 (absent) and 1 (fully present). Maximum values were compared with respective healthy controls to calculate percentage restoration., Results: Of 13 FT patients, eight patients were full FT, five patients were partial FT, and two patients were female patients. Compared with healthy controls, the median restoration of motor function was 36.9% (interquartile range, 28.8% to 52.9%) for all patients with FT ( P = 0.151). The median restoration of smile was 37.2% (interquartile range, 31.5% to 52.7%) for all patients with FT ( P = 0.065). When facial nerve coaptation was performed at the distal branch level, average motor function restoration was 42.7% ± 3.61% compared with 27.9% ± 6.71% at the proximal trunk coaptation level ( P = 0.032). Use of interpositional nerve grafts had no influence on motor outcomes., Conclusions: Software-based analysis is suitable to assess motor function after FT. International collaboration strengthens outcome data for FT., Clinical Question/level of Evidence: Therapeutic, IV., (Copyright © 2023 by the American Society of Plastic Surgeons.)

Objectives: Although facial transplantation is considered effective for restoring facial appearance, research on speech outcomes following surgery is limited. More research is critically needed to inform patients of expected rates and extent of recovery, and to develop interventions aimed at improving speech outcomes., Methods: Four patients in early recovery (3 weeks-24 months postsurgery) and three patients in late recovery (36-60 months postsurgery) were included. Clinical measures of speech recovery, including speech intelligibility measured using the Sentence Intelligibility Test, a lip strength testing device (Iowa Oral Performance Instrument), and kinematic measures of lip and jaw function measured using high-resolution 3D optical motion capture were used to describe the rate and extent of functional speech and lip recovery, describe and compare the rate of functional speech recovery and kinematic lip and jaw changes in early and late stages of recovery, and explore the association between kinematic measures and functional speech., Results: Speech intelligibility, speaking rate, and lip strength were below normative values in the first 2 years of postsurgery. Participants in the first 2 years of recovery demonstrated steeper slopes of improvement in clinical and kinematic measures than participants in the later stages of recovery (36-64 months). Gains in jaw range of movement and gains in lip speed and range of movement were significantly correlated with rates of sentence intelligibility improvement. Gains in lip strength were not associated with functional speech improvement., Conclusions: These findings motivate ongoing work aimed at developing interventions for improving motor speech function in this population., Level of Evidence: 3 Laryngoscope, 132:2359-2367, 2022., (© 2022 The Authors. The Laryngoscope published by Wiley Periodicals LLC on behalf of The American Laryngological, Rhinological and Otological Society, Inc.)

Tuatara (Sphenodon punctatus) are one of the most phylogenetically isolated vertebrate species and provide a unique host system to study virus evolution. While the tuatara genome, sequenced in 2020, revealed many endogenous viral elements, we know little of the exogenous viruses that infect tuatara. We performed a metatranscriptomics study of tuatara cloaca samples from a wild population on Takapourewa (Stephens Island), Aotearoa New Zealand. From these data we identified 49 potentially novel viral species that spanned 19 RNA viral families and/or orders, the vast majority (48) of which were likely dietary-related. Notably, using a protein structure homology search, we identified a highly divergent novel virus within the Picornaviridae which may directly infect tuatara. Additionally, two endogenous tuatara adintoviruses were characterised that exhibited long-term viral-host co-divergence. Overall, our results indicate that the tuatara cloacal virome is highly diverse, likely due to a large number of dietary-related viruses., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Although speech declines rapidly in some individuals with amyotrophic lateral sclerosis (ALS), longitudinal changes in speech have rarely been characterized. The study objectives were to model the rate of decline in speaking rate and speech intelligibility as a function of disease onset site, sex, and age at onset in 166 individuals with ALS; and estimate time to speech loss from symptom onset. We also examined the association between clinical (speaking rate/intelligibility) measures and patient-reported measures of ALS progression (ALSFRS-R). Speech measures declined faster in the bulbar-onset group than in the spinal-onset group. The rate of decline was not significantly affected by sex and age. Functional speech was still maintained at 60 months since disease onset for most patients with spinal onset. However, the time to speech loss was 23 months based on speaking rate &lt; 120 (w/m) and 32 months based on speech intelligibility &lt; 85% in individuals with ALS-bulbar onset. Speech measures were more responsive to functional decline than were the patient-reported measures. The findings of this study will inform future work directed toward improving speech prognosis in ALS, which is critical for determining the appropriate timing of interventions, providing appropriate counseling for patients, and evaluating functional changes during clinical trials., (© 2022. The Author(s).)

Purpose : For patients diagnosed with ALS, dysphagia can result in aspiration, malnutrition, and mortality. The purpose of this study was to develop a clinical prediction model capable of identifying patients with ALS at imminent risk for developing swallowing complications. Methods : A retrospective cohort study using the Pooled Resource Open-Access ALS Clinical Trials Database (PRO-ACT) was conducted. After dividing the PRO-ACT database into development and validation cohorts with dysphagia defined from the ALS Functional Rating Scale (ALSFRS), a multivariable Cox proportional hazards regression model estimated the probability of dysphagia at 3, 6, and 12-months with subsequent evaluation of model discrimination and calibration. Results : With 2057 participants in the development cohort and 1891 in the validation cohort, the Cox model included 7 clinical variables: spinal-onset; bulbar, fine and gross motor ALSFRS subscale scores; respiratory impairment; functional progression rate; and time from diagnosis. The cumulative incidence of dysphagia was 18% at 3-months, 29% at 6-months, and 45% at 12-months. The mean predicted probability of dysphagia development ranged from 4.5% in the bottommost risk decile to 40% in the topmost decile at 3 months, 10%-72% at 6 months, and 25%-93% at 12 months. In the validation cohort, the model had good discrimination and calibration with an optimism corrected c-statistic of 0.70 and calibration slope of 0.96. Conclusions : The ALS dysphagia risk score can be used to identify patients with ALS at high risk for self-reported dysphagia development who would benefit from a comprehensive swallowing assessment and proactive dysphagia management strategies.

Viruses that infect fish are understudied, yet they provide important evolutionary context to the viruses that infect terrestrial vertebrates. We surveyed gill tissue meta-transcriptomes collected from two species of native freshwater fish from Aotearoa New Zealand- Retropinna retropinna and Gobiomorphus cotidianus . A total of 64 fish were used for gill tissue meta-transcriptomic sequencing, from populations with contrasting life histories-landlocked (i.e., lacustrine) and diadromous-on the South Island and Chatham Islands. We observed that both viral richness and taxonomic diversity were significantly associated with life history and host species, with lacustrine R. retropinna characterised by higher viral alpha diversity than diadromous R. retropinna . Additionally, we observed transcripts of fish viruses from 12 vertebrate host-associated virus families, and phylogenetically placed eight novel RNA viruses and three novel DNA viruses in the Astroviridae , Paramyxoviridae , Orthomyxoviridae , Rhabdoviridae , Totiviridae , Poxviridae , Alloherpesviridae, and Adintoviridae in their evolutionary contexts. These results represent an important survey of the viruses that infect two widespread native fish species in New Zealand, and provide insight useful for future fish virus surveys.

Horizontal transfer of the integrative and conjugative element ICEMlSymR7A converts non-symbiotic Mesorhizobium spp. into nitrogen-fixing legume symbionts. Here, we discover subpopulations of Mesorhizobium japonicum R7A become epigenetically primed for quorum-sensing (QS) and QS-activated horizontal transfer. Isolated populations in this state termed R7A* maintained these phenotypes in laboratory culture but did not transfer the R7A* state to recipients of ICEMlSymR7A following conjugation. We previously demonstrated ICEMlSymR7A transfer and QS are repressed by the antiactivator QseM in R7A populations and that the adjacently-coded DNA-binding protein QseC represses qseM transcription. Here RNA-sequencing revealed qseM expression was repressed in R7A* cells and that RNA antisense to qseC was abundant in R7A but not R7A*. Deletion of the antisense-qseC promoter converted cells into an R7A*-like state. An adjacently coded QseC2 protein bound two operator sites and repressed antisense-qseC transcription. Plasmid overexpression of QseC2 stimulated the R7A* state, which persisted following curing of this plasmid. The epigenetic maintenance of the R7A* state required ICEMlSymR7A-encoded copies of both qseC and qseC2. Therefore, QseC and QseC2, together with their DNA-binding sites and overlapping promoters, form a stable epigenetic switch that establishes binary control over qseM transcription and primes a subpopulation of R7A cells for QS and horizontal transfer., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

To provide protection against viral infection and limit the uptake of mobile genetic elements, bacteria and archaea have evolved many diverse defence systems. The discovery and application of CRISPR-Cas adaptive immune systems has spurred recent interest in the identification and classification of new types of defence systems. Many new defence systems have recently been reported but there is a lack of accessible tools available to identify homologs of these systems in different genomes. Here, we report the Prokaryotic Antiviral Defence LOCator (PADLOC), a flexible and scalable open-source tool for defence system identification. With PADLOC, defence system genes are identified using HMM-based homologue searches, followed by validation of system completeness using gene presence/absence and synteny criteria specified by customisable system classifications. We show that PADLOC identifies defence systems with high accuracy and sensitivity. Our modular approach to organising the HMMs and system classifications allows additional defence systems to be easily integrated into the PADLOC database. To demonstrate application of PADLOC to biological questions, we used PADLOC to identify six new subtypes of known defence systems and a putative novel defence system comprised of a helicase, methylase and ATPase. PADLOC is available as a standalone package (https://github.com/padlocbio/padloc) and as a webserver (https://padloc.otago.ac.nz)., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Military vehicle underbody blast (UBB) is the cause of many serious injuries in theatre today; however, the effects of these chaotic events on the human body are not well understood. The purpose of this research was to replicate UBB loading conditions on the human pelvis and investigate the resulting response in a controlled laboratory setting. In addition to better understanding the response of the human pelvis to high rate vertical loading, this test series also aimed to identify high rate injury thresholds. Twenty-seven post mortem human surrogate (PMHS) component pelvis tests were completed using the University of Virginia's (UVa) simulated blast rig under a range of loading conditions and postures. Of those tests, 17 were in the anteriorly-tilted posture and used to construct the human injury probability curve. Average seat pan (rigid) accelerations for this subset of tests ranged from 300 to 2400 g over 2 to 3 ms of positive phase duration. Post-test computed tomography (CT) scans and necropsies were performed to determine injuries and revealed a frequent occurrence of anterior and posterior injuries, resulting in unstable pelvis ring fractures. The resulting Human Injury Probability Curve (HIPC) yielded mean forces of 5529, 8516, and 12431 N as measured by mass compensated seat platen loadcells applied through the rigid seat to the bilateral ischium are associated with a 10, 25, and 50% risk for unstable pelvic ring sacrum fractures in an anteriorly-tilted pelvis (28° from vertical), respectively., (© 2020. Biomedical Engineering Society.)

Wastewater treatment plants (WWTPs) are useful environments for investigating the occurrence, diversity, and evolution of plasmids encoding clinically relevant antibiotic resistance genes (ARGs). Our objective was to isolate and sequence plasmids encoding meropenem resistance from bacterial hosts within Canadian WWTPs. We used two enrichment culture approaches for primary plasmid isolation, followed by screening for antibiotic resistance, conjugative mobility, and stability in enteric bacteria. Isolated plasmids were sequenced using Illumina MiSeq and Sanger sequencing methods. Bioinformatics analyses resolved a multi-resistance IncF /MOB F12 plasmid, pFEMG (209 357 bp), harbouring resistance genes to β-lactam ( bla CMY-42 , bla TEM-1β , and bla NDM-5 ), macrolide ( mphA - mrx-mphR ), tetracycline ( tetR - tetB - tetC-tetD ), trimethoprim ( dfrA12 ), aminoglycoside ( aadA2 ), and sulfonamide ( sul1 ) antibiotic classes. We also isolated an IncI1 /MOB P12 plasmid pPIMR (172 280 bp) carrying similar β-lactamase and a small multi-drug efflux resistance gene cluster ( bla CMY-42 -blc-sugE ) to pFEMG. The co-occurrence of different ARGs within a single 24 552 bp cluster in pFEMG - interspersed with transposons, insertion sequence elements, and a class 1 integron - may be of significant interest to human and veterinary medicine. Additionally, the presence of conjugative and plasmid maintenance genes in the studied plasmids corresponded to observed high conjugative transfer frequencies and stable maintenance. Extensive investigation is required to further understand the fitness trade-offs of plasmids with different types of conjugative transfer and maintenance modules.

Members of the Mesorhizobium genus are soil bacteria that often form nitrogen-fixing symbioses with legumes. Most characterised Mesorhizobium spp. genomes are ~8 Mb in size and harbour extensive pangenomes including large integrative and conjugative elements (ICEs) carrying genes required for symbiosis (ICESyms). Here, we document and compare the conjugative mobilome of 41 complete Mesorhizobium genomes. We delineated 56 ICEs and 24 integrative and mobilizable elements (IMEs) collectively occupying 16 distinct integration sites, along with 24 plasmids. We also demonstrated horizontal transfer of the largest (853,775 bp) documented ICE, the tripartite ICE M spSym AA22 . The conjugation systems of all identified ICEs and several plasmids were related to those of the paradigm ICESym ICE Ml Sym R7A , with each carrying conserved genes for conjugative pilus formation ( trb ), excision ( rdfS ), DNA transfer ( rlxS ) and regulation ( fseA ). ICESyms have likely evolved from a common ancestor, despite occupying a variety of distinct integration sites and specifying symbiosis with diverse legumes. We found extensive evidence for recombination between ICEs and particularly ICESyms, which all uniquely lack the conjugation entry-exclusion factor gene trbK . Frequent duplication, replacement and pseudogenization of genes for quorum-sensing-mediated activation and antiactivation of ICE transfer suggests ICE transfer regulation is constantly evolving. Pangenome-wide association analysis of the ICE identified genes potentially involved in symbiosis, rhizosphere colonisation and/or adaptation to distinct legume hosts. In summary, the Mesorhizobium genus has accumulated a large and dynamic pangenome that evolves through ongoing horizontal gene transfer of large conjugative elements related to ICE Ml Sym R7A .

Biobeds are agriculture-based bioremediation tools used to safely contain and microbially degrade on-farm pesticide waste and rinsate, thereby reducing the negative environmental impacts associated with pesticide use. While these engineered ecosystems demonstrate efficient pesticide removal, the microbiomes in these environments remain largely understudied both taxonomically and functionally. This study used metagenomic and metatranscriptomic techniques to characterize the microbial community in a two-cell Canadian biobed system before and after a field season of pesticide application. These culture-independent approaches identified an enrichment of xenobiotic-degrading bacteria, such as Afipia, Sphingopyxis and Pseudomonas, and enrichment and transcription of xenobiotic-degrading genes, such as peroxidases, oxygenases, and hydroxylases, among others; we were able to directly link the transcription of these genes to Pseudomonas, Oligotropha, Mesorhizobium, Rhodopseudomonas, and Stenotrophomonas taxa., (© 2021 Society for Applied Microbiology and John Wiley & Sons Ltd.)

Introduction: X-linked Dystonia-Parkinsonism (XDP) is a progressive, disabling disease characterized by the devastating impairment of bulbar function, including speech and swallowing. Despite these detrimental impacts, bulbar impairments in this population are not well characterized., Objectives: To identify impairments in the bulbar system measured by oromotor performance in individuals with XDP relative to healthy controls. Secondarily, to detect diagnostic bulbar markers that are sensitive and specific to the initial years of XDP., Methods: This case-control study included 25 healthy controls and 30 participants with XDP, divided into two subgroups based on the median of their disease length. Multiple clinical and instrumental oromotor tasks and measures were used to evaluate bulbar motor function., Results: Differences were found between both the subgroups with XDP and healthy controls on almost all measures, including maximum performance tasks such as tongue strength, alternating motion rate (AMR), and sequential motion rate (SMR) (p < 0.05). Differences were found between the XDP subgroups and the control group for the percentage of pause time during the speech, a rating of speech severity, and a swallowing task (ps < 0.05). Scores on self-reported questionnaires, tongue strength, the number of repetitions produced during an AMR, percent pause, and speech severity demonstrated good sensitivity and specificity to differentiate the initial years of XDP onset from healthy controls., Conclusions: Our findings revealed impairments across bulbar functions in participants within the first 7 years of the XDP onset. Highly sensitive and specific bulbar impairment measures were detected in instrumental and self-reported measures that are fundamental for monitoring disease., (Copyright © 2021. Published by Elsevier Ltd.)

Introduction/aims: Dysphagia worsens mortality and quality of life for persons diagnosed with amyotrophic lateral sclerosis (ALS), yet our understanding of its incidence and timing remains limited. In this study we sought to estimate dysphagia incidence and dysphagia-free survival over time., Methods: Using data from the Pooled Resource Open-Access ALS Clinical Trials Database, we compared characteristics of persons with and without dysphagia upon study entry. To account for competing mortality risk, we used Kaplan-Meier curves to estimate the cumulative incidence of dysphagia and the median number of days until the development of dysphagia or death in those without dysphagia at study entry., Results: Patients with dysphagia upon study entry were more likely to have bulbar onset and had faster rates of functional decline and shorter diagnostic delays. The cumulative incidence of new-onset dysphagia was 44% at 1 year and 64% at 2 years after trial enrollment for those with spinal onset, and 85% and 92% for those with bulbar onset. The median duration of dysphagia-free survival after trial enrollment was 11.5 months for those with spinal onset and 3.2 months for those with bulbar onset., Discussion: Our findings underscore the high risk for dysphagia development and support the need for early dysphagia referral and evaluation to minimize the risk of serious dysphagia-related complications., (© 2021 Wiley Periodicals LLC.)

Background: The impact of tongue dysfunction on deglutition in persons diagnosed with amyotrophic lateral sclerosis (ALS) is not well understood. This information is needed to improve our understanding of the mechanisms of swallowing impairment, for identifying risk factors of dysphagia, and for establishing impairment-specific treatments aimed at slowing the loss of swallow function., Objectives: The goals of this study were to determine the relation between biomechanical measures of oral tongue movements using electromagnetic articulography (EMA) and measures of swallow physiology, swallow safety and efficiency, and self-reported swallowing function., Methods: Participants were diagnosed with ALS by a neurologist following the El Escorial Criteria from the World Federation of Neurology. Twelve participants underwent (1) EMA to derive biomechanical measures of the tongue, (2) videofluoroscopic evaluation to measure swallow physiology, safety, and efficiency, and (3) maximal tongue strength testing using the Iowa Oral Pressure Instrument (IOPI). Participants completed self-reported functional assessments. Spearman's rank correlations assessed for associations between lingual biomechanics and swallowing physiology, swallow safety and efficiency, and self-reported bulbar function., Results: Results demonstrated strong associations between biomechanical and swallowing physiology, swallow safety, and self-reported measures. Notably, swallowing safety during thin liquid intake was associated with tongue speed (r = - 0.7, p < 0.05) and range of motion (r = - 0.71, p < 0.05), and swallowing safety during puree intake was associated with tongue strength (r = - 0.69, p < 0.05)., Conclusions: Our findings underscore the importance of tongue movements on swallowing physiology and safety, help improve our understanding of mechanisms of swallowing impairment, and highlight a potential clinical tool to index bulbar impairment.

Mesorhizobium phage vB&#95;MloS&#95;Cp1R7A-A1 was isolated from soil planted with chickpea in Saskatchewan. It is dissimilar in sequence and morphology to previously described rhizobiophages. It is a B3 morphotype virus with a distinct prolate capsid and belongs to the tailed phage family Siphoviridae . Its genome has a GC content of 60.3% and 238 predicted genes. Putative functions were predicted for 57 genes, which include 27 tRNA genes with anticodons corresponding to 18 amino acids. This represents the highest number of tRNA genes reported yet in a rhizobiophage. The gene arrangement shows a partially modular organization. Most of the structural genes are found in one module, whereas tRNA genes are in another. Genes for replication, recombination, and nucleotide metabolism form the third module. The arrangement of the replication module resembles the replication module of Enterobacteria phage T5, raising the possibility that it uses a recombination-based replication mechanism, but there is also a suggestion that a T7-like replication mechanism could be used. Phage termini appear to be long direct repeats of just over 12 kb in length. Phylogenetic analysis revealed that Cp1R7A-A1 is more closely related to PhiCbK-like Caulobacter phages and other B3 morphotype phages than to other rhizobiophages sequenced thus far.

Bacteria currently included in Rhizobium leguminosarum are too diverse to be considered a single species, so we can refer to this as a species complex (the Rlc). We have found 429 publicly available genome sequences that fall within the Rlc and these show that the Rlc is a distinct entity, well separated from other species in the genus. Its sister taxon is R. anhuiense . We constructed a phylogeny based on concatenated sequences of 120 universal (core) genes, and calculated pairwise average nucleotide identity (ANI) between all genomes. From these analyses, we concluded that the Rlc includes 18 distinct genospecies, plus 7 unique strains that are not placed in these genospecies. Each genospecies is separated by a distinct gap in ANI values, usually at approximately 96% ANI, implying that it is a 'natural' unit. Five of the genospecies include the type strains of named species: R. laguerreae, R. sophorae, R. ruizarguesonis , "R. indicum" and R. leguminosarum itself. The 16S ribosomal RNA sequence is remarkably diverse within the Rlc, but does not distinguish the genospecies. Partial sequences of housekeeping genes, which have frequently been used to characterize isolate collections, can mostly be assigned unambiguously to a genospecies, but alleles within a genospecies do not always form a clade, so single genes are not a reliable guide to the true phylogeny of the strains. We conclude that access to a large number of genome sequences is a powerful tool for characterizing the diversity of bacteria, and that taxonomic conclusions should be based on all available genome sequences, not just those of type strains.

Despite signs of facial nerve recovery within a few months following face transplantation, speech deficits persist for years. Behavioral speech modifications (e.g., slower-than-normal speaking rate and increased loudness) have shown promising potential to enhance speech intelligibility in populations with dysarthric speech. However, such evidence-based practice approach is lacking in clinical management of speech in individuals with facial transplantation. Because facial transplantation involves complex craniofacial reconstruction and facial nerve coaptation, it is unknown to what extent individuals with face transplant are capable of adapting their motor system to task-specific articulatory demands. The purpose of this study was to identify the underlying articulatory mechanisms employed by individuals with face transplantation in response to speech modification cues at early and late stages of neuromotor recovery. In addition, we aimed to identify speech modifications that conferred improved speech clarity. Participants were seven individuals who underwent full or partial facial vascularized composite allografts that included lips and muscles of facial animation and were in early (~2 months) or late (~42 months) stages of recovery. Participants produced repetitions of the sentence "Buy Bobby a puppy" in normal, fast, loud, and slow speech modifications. Articulatory movement traces were recorded using a 3D optical motion capture system. Kinematic measures of average speed (mm/s) and range of movement (mm 3 ) were extracted from the lower lip (± jaw) marker. Two speech language pathologists rated speech clarity for each speaker using a visual analog scale (VAS) approach. Results demonstrated that facial motor capacity increased from early to late stages of recovery. While individuals in the early group exhibited restricted capabilities to adjust their motor system based on the articulatory demands of each speech modification, individuals in the late group demonstrated faster speed and larger-than-normal range of movement for loud speech, and slower speed and larger-than-normal range of movement for slow speech. In addition, subjects in both groups showed overreliance on jaw rather than lip articulatory function across all speech modifications, perhaps as a compensatory strategy to optimize articulatory stability and maximize speech function. Finally, improved speech clarity was associated with loud speech in both stages of recovery., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Eshghi, Perry, Richburg, Ventresca, Pomahac and Green.)

Rhizobium leguminosarum symbiovar trifolii strains TA1 and CC275e are nitrogen-fixing microsymbionts of Trifolium spp. and have been used as commercial inoculant strains for clovers in pastoral agriculture in Australia and New Zealand. Here we present the complete genome sequences of both strains, resolving their multipartite genome structures and allowing for future studies using genomic approaches.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Mesorhizobium is a genus of soil bacteria, some isolates of which form an endosymbiotic relationship with diverse legumes of the Loteae tribe. The symbiotic genes of these mesorhizobia are generally carried on integrative and conjugative elements termed symbiosis islands (ICESyms). Mesorhizobium strains that nodulate Lotus spp. have been divided into host-range groupings. Group I (GI) strains nodulate L. corniculatus and L. japonicus ecotype Gifu, while group II (GII) strains have a broader host range, which includes L. pedunculatus . To identify the basis of this extended host range, and better understand Mesorhizobium and ICESym genomics, the genomes of eight Mesorhizobium strains were completed using hybrid long- and short-read assembly. Bioinformatic comparison with previously sequenced mesorhizobia genomes indicated host range was not predicted by Mesorhizobium genospecies but rather by the evolutionary relationship between ICESym symbiotic regions. Three radiating lineages of Loteae ICESyms were identified on this basis, which correlate with Lotus spp. host-range grouping and have lineage-specific nod gene complements. Pangenomic analysis of the completed GI and GII ICESyms identified 155 core genes (on average 30.1 % of a given ICESym). Individual GI or GII ICESyms carried diverse accessory genes with an average of 34.6 % of genes unique to a given ICESym. Identification and comparative analysis of NodD symbiotic regulatory motifs - nod boxes - identified 21 branches across the NodD regulons. Four of these branches were associated with seven genes unique to the five GII ICESyms. The nod boxes preceding the host-range gene nodZ in GI and GII ICESyms were disparate, suggesting regulation of nodZ may differ between GI and GII ICESyms. The broad host-range determinant(s) of GII ICESyms that confer nodulation of L. pedunculatus are likely present amongst the 53 GII-unique genes identified.

Establishment of the symbiotic relationship that develops between rhizobia and their legume hosts is contingent upon an interkingdom signal exchange. In response to host legume flavonoids, NodD proteins from compatible rhizobia activate expression of nodulation genes that produce lipochitin oligosaccharide signaling molecules known as Nod factors. Root nodule formation commences upon legume recognition of compatible Nod factor. Rhizobium leguminosarum was previously considered to contain one copy of nodD ; here, we show that some strains of the Trifolium (clover) microsymbiont R. leguminosarum bv. trifolii contain a second copy designated nodD2. nodD2 genes were present in 8 out of 13 strains of R. leguminosarum bv. trifolii, but were absent from the genomes of 16 R. leguminosarum bv. viciae strains. Analysis of single and double nodD1 and nodD2 mutants in R. leguminosarum bv. trifolii strain TA1 revealed that NodD2 was functional and enhanced nodule colonization competitiveness. However, NodD1 showed significantly greater capacity to induce nod gene expression and infection thread formation. Clover species are either annual or perennial and this phenological distinction is rarely crossed by individual R. leguminosarum bv. trifolii microsymbionts for effective symbiosis. Of 13 strains with genome sequences available, 7 of the 8 effective microsymbionts of perennial hosts contained nodD2 , whereas the 3 microsymbionts of annual hosts did not. We hypothesize that NodD2 inducer recognition differs from NodD1, and NodD2 functions to enhance competition and effective symbiosis, which may discriminate in favor of perennial hosts. IMPORTANCE Establishment of the rhizobium-legume symbiosis requires a highly specific and complex signal exchange between both participants. Rhizobia perceive legume flavonoid compounds through LysR-type NodD regulators. Often, rhizobia encode multiple copies of nodD , which is one determinant of host specificity. In some species of rhizobia, the presence of multiple copies of NodD extends their symbiotic host-range. Here, we identified and characterized a second copy of nodD present in some strains of the clover microsymbiont Rhizobium leguminosarum bv. trifolii. The second nodD gene contributed to the competitive ability of the strain on white clover, an important forage legume. A screen for strains containing nodD2 could be utilized as one criterion to select strains with enhanced competitive ability for use as inoculants for pasture production., (Copyright © 2020 American Society for Microbiology.)

Background: Pantoea is a genus within the Enterobacterales whose members encompass free-living and host-associated lifestyles. Despite our growing understanding of the role of mobile genetic elements in the biology, ecology, and evolution of this bacterial group, few Pantoea bacteriophages have been identified and characterized. Materials and Methods: A bacteriophage that could infect Pantoea agglomerans was isolated from barnyard soil. We used electron microscopy and complete genome sequencing to identify the viral family, and evaluated its host range across 10 different Pantoea species groups using both bacterial lawn and phage lawn assays. The latter assays were carried out using a scalable microplate assay to increase throughput and enable spectrophotometric quantitation. We also performed a phylogenetic analysis to determine the closest relatives of our phage. Results: Phage vB&#95;PagP-SK1 belongs to the genus Teseptimavirus of the Podoviridae family in the order Caudovirales. The 39,938 bp genome has a modular structure with early, middle, and late genes, along with the characteristic direct terminal repeats of 172 bp. Genome composition and synteny were similar to that of the Erwinia amylovora phage, vB&#95;EamP-L1, with the exception of a few loci that are most similar to genes of phage infecting other members of the Enterobacteriaceae. A total of 94 Pantoea strains were surveyed and vB&#95;PagP-SK1 was found to infect 15 Pantoea strains across three species, predominantly P. agglomerans , along with one Erwinia billingiae strain. Conclusions: vB&#95;PagP-SK1 belongs to the Teseptimavirus genus and has a host range that spans multiple species groups, and is most closely related to the E. amylovora phage, vB&#95;EamP-L1. The presence of xenologous genes in its genome indicates that the genome is a mosaic of multiple Teseptimavirus phages that infect members of the Enterobacteriaceae ., Competing Interests: No competing financial interests exist., (Copyright 2020, Mary Ann Liebert, Inc., publishers.)

Cultivation of dedicated soil plots called 'landfarms' is an effective technology for bioremediation of hydrocarbon waste generated by various industrial practices. To understand the influence of soil conditions on landfarm microbial communities, analysis of bacterial and fungal community structure using next-generation sequencing at different sections and depths was performed across a hydrocarbon-waste landfarm in Regina, Saskatchewan, Canada. While a core set of hydrocarbon-associated bacterial and fungal taxa are present throughout the landfarm, unique bacterial and fungal operational taxonomic units are differentially abundant at sections within the landfarm, which correlate with differences in soil physiochemical properties and management practices. Increased frequency of waste application resulted in strong positive correlations between bacterial community assemblages and elevated amounts of oil, grease and F3 - F4 hydrocarbon fractions. In areas of standing water and lower application of hydrocarbon, microbial community structure correlated with soil pH, trace nutrients and metals. Overall, diversity and structure of bacterial communities remain relatively stable across the landfarm, while in contrast, fungal community structure appears more responsive to soil oxygen conditions. Results are consistent with the hypothesis that years of bioremediation activity have shaped microbial communities; however, several management practices can be undertaken to increase efficiency of remediation, including the removal of standing water and soil tilling across the landfarm., (© 2019 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.)

Early identification of bulbar involvement in persons with ALS is critical for improving diagnosis and prognosis; however, efficacious diagnostic markers have not yet been identified. The purpose of this study was to determine whether kinematic changes of the tongue and jaw during swallowing, measured using 3D electromagnetic articulography (EMA), predate clinically identifiable symptoms of speech and swallowing impairment in persons diagnosed with ALS. Data were collected from 16 adults diagnosed with ALS and 18 neurotypical controls. Groups were aged matched. Eligible participants with ALS were tolerating an unrestricted diet (FOIS = 7), produced intelligible speech (> 97%), and had a speaking rate greater than 150 words per minute. Participants completed a 3-mL water swallow task, during which EMA recorded kinematic measures of the anterior and posterior regions of tongue including lingual speed, range of motion, duration, coordination, and efficiency. Jaw speed and range of motion were also recorded. Persons diagnosed with ALS demonstrated reduced posterior lingual range of motion (11.40 mm ± 4.01 vs. 16.07 mm ± 5.27), slower posterior lingual speeds (83.67 mm/s ± 47.96 vs. 141.35 mm/s ± 66.54), increased lingual movement duration (13.46 s ± 6.75 vs. 9.21 s ± 3.28), and reduced lingual coordination (0.04 s ± 0.11 vs. 17 s ± 0.19) during the 3-oz water swallow task compared to controls. Persons diagnosed with ALS demonstrated increased range of motion (9.86 mm ± 5.38 vs. 6 mm ± 3.78) and increased jaw speed (68.62 mm/s ± 50.13 vs. 34.72 mm/s ± 17.75) during swallowing compared to controls. The current findings suggest that changes in lingual and jaw motor performance during a simple water swallow task are present in persons with ALS who are pre-symptomatic of clinically detectable bulbar impairment.

Aims: This study evaluated the variability of bacterial and fungal communities within unique pesticide remediation biobeds., Methods and Results: Four biobeds receiving different applied pesticide rinsates, were sampled throughout an operational season to determine pesticide removal efficacy and microbial communities. Biomixture samples collected from different biobed depths, were subjected to Illumina sequencing of the 16S rRNA (bacteria) and ITS2 (fungi) genes. Pesticide removal rates for all biobeds averaged 99%, with microbial community analysis revealing biobeds shared 60-70% of the most abundant bacterial and fungal orders, respectively. Though biobed depth did not greatly impact microbial community profile or diversity, bacterial and fungal taxa profiles between biobeds notably diverge at levels of genera and OTU. Biobed bacterial communities exhibited greater diversity than fungal communities between and within all biobeds., Conclusions: Biobeds receiving variable pesticide rinsates share a 'core' microbial community, exhibiting greater bacterial diversity relative to fungal diversity. Pesticide exposure increased bacterial diversity throughout the biobeds, while fungal diversity was variable, meriting further understanding of fungicide application to biobed fungal community stability., Significance and Impact of the Study: Biobeds achieve high treatment efficacy of unique pesticide rinsates, regardless of differentiation of specific genera in response to specific compounds; supporting biobeds as a robust engineered system for pesticide rinsates bioremediation., (© 2018 The Society for Applied Microbiology.)

Deinococcus sp. strain UR1, a resilient bacterium isolated from the surface of a stainless steel sign located on the University of Regina campus in Saskatchewan, Canada, was sequenced to 56-fold coverage to produce 73 contigs with a consensus length of 4,472,838 bp and a G+C content of 69.37%., (Copyright © 2018 Suchan et al.)

Herein, we report the genome sequence of Delftia acidovorans strain RAY209, a plant growth-promoting rhizobacterium that is used in commercial inoculants for canola and soybean. The genome of RAY209 has a consensus of 6,528,879 bp and an estimated 5,721 coding sequences., (Copyright © 2017 Perry et al.)

Purpose: Facial transplantation is a relatively new option for individuals with severe facial disfigurements. Clinical case studies on existing patients indicate many instances of persistent facial motor impairment for facial expression, speech, and swallowing. These preliminary findings motivate additional research on the impact of lip-strengthening exercises following facial transplantation., Method: In this study, we assessed the efficacy of an 8-week, biofeedback-driven, lip closure-strengthening exercise program in a single patient 1-year status post-full facial transplantation. Exercise was at 60% of peak strength. Outcome measures included instrumental measures of lip strength and mobility, clinical measures of speech, and patient-reported outcomes in feeding and facial expression., Results: Results revealed improvements in labial strength, speed of lip movement, and range of motion during speech. A 3-point improvement in sentence speech intelligibility was also observed following strength-training exercise. The patient reported improvements in her ability to drink from a straw and communicate via facial expression., Conclusion: These preliminary findings motivate additional research on the efficacy of lip-strengthening exercises following facial transplantation.

Bedaquiline (BDQ), a diarylquinoline antibiotic that targets ATP synthase, is effective for the treatment of Mycobacterium tuberculosis infections that no longer respond to conventional drugs. While investigating the off-label use of BDQ as salvage therapy, seven of 13 patients with Mycobacterium intracellulare lung disease had an initial microbiological response and then relapsed. Whole-genome comparison of pretreatment and relapse isolates of M. intracellulare uncovered mutations in a previously uncharacterized locus, mmpT5 Preliminary analysis suggested similarities between mmpT5 and the mmpR5 locus, which is associated with low-level BDQ resistance in M. tuberculosis Both genes encode transcriptional regulators and are adjacent to orthologs of the mmpS5-mmpL5 drug efflux operon. However, MmpT5 belongs to the TetR superfamily, whereas MmpR5 is a MarR family protein. Targeted sequencing uncovered nonsynonymous mmpT5 mutations in isolates from all seven relapse cases, including two pretreatment isolates. In contrast, only two relapse patient isolates had nonsynonymous changes in ATP synthase subunit c (atpE), the primary target of BDQ. Susceptibility testing indicated that mmpT5 mutations are associated with modest 2- to 8-fold increases in MICs for BDQ and clofazimine, whereas one atpE mutant exhibited a 50-fold increase in MIC for BDQ. Bedaquiline shows potential for the treatment of M. intracellulare lung disease, but optimization of treatment regimens is required to prevent the emergence of mmpT5 variants and microbiological relapse., (Copyright © 2017 American Society for Microbiology.)

The genome of Salmonella enterica subspecies enterica serovar Enteritidis phage type 8 strain EN1660, isolated from an outbreak in Thunder Bay, Canada, was sequenced to 46-fold coverage using an Illumina MiSeq with 300-bp paired-end sequencing chemistry to produce 28 contigs with an N 50 value of 490,721 bp., (Copyright © 2017 Perry et al.)

Insertion sequencing (INSeq) analysis of Rhizobium leguminosarum bv. viciae 3841 (Rlv3841) grown on glucose or succinate at both 21% and 1% O 2 was used to understand how O 2 concentration alters metabolism. Two transcriptional regulators were required for growth on glucose (pRL120207 [eryD] and RL0547 [phoB]), five were required on succinate (pRL100388, RL1641, RL1642, RL3427, and RL4524 [ecfL]), and three were required on 1% O 2 (pRL110072, RL0545 [phoU], and RL4042). A novel toxin-antitoxin system was identified that could be important for generation of new plasmidless rhizobial strains. Rlv3841 appears to use the methylglyoxal pathway alongside the Entner-Doudoroff (ED) pathway and tricarboxylic acid (TCA) cycle for optimal growth on glucose. Surprisingly, the ED pathway was required for growth on succinate, suggesting that sugars made by gluconeogenesis must undergo recycling. Altered amino acid metabolism was specifically needed for growth on glucose, including RL2082 (gatB) and pRL120419 (opaA, encoding omega-amino acid:pyruvate transaminase). Growth on succinate specifically required enzymes of nucleobase synthesis, including ribose-phosphate pyrophosphokinase (RL3468 [prs]) and a cytosine deaminase (pRL90208 [codA]). Succinate growth was particularly dependent on cell surface factors, including the PrsD-PrsE type I secretion system and UDP-galactose production. Only RL2393 (glnB, encoding nitrogen regulatory protein PII) was specifically essential for growth on succinate at 1% O 2 , conditions similar to those experienced by N 2 -fixing bacteroids. Glutamate synthesis is constitutively activated in glnB mutants, suggesting that consumption of 2-ketoglutarate may increase flux through the TCA cycle, leading to excess reductant that cannot be reoxidized at 1% O 2 and cell death., Importance: Rhizobium leguminosarum, a soil bacterium that forms N 2 -fixing symbioses with several agriculturally important leguminous plants (including pea, vetch, and lentil), has been widely utilized as a model to study Rhizobium-legume symbioses. Insertion sequencing (INSeq) has been used to identify factors needed for its growth on different carbon sources and O 2 levels. Identification of these factors is fundamental to a better understanding of the cell physiology and core metabolism of this bacterium, which adapts to a variety of different carbon sources and O 2 tensions during growth in soil and N 2 fixation in symbiosis with legumes., (Copyright © 2016 American Society for Microbiology.)

Background: Facial self-inflicted gunshot wounds (SIGSWs) cause a devastating midfacial defect and pose a challenging problem to the reconstructive surgeon. Face transplantation (FT) has the potential for near-normal restoration in otherwise non-reconstructible defects. Two out of 7 FT recipients at Brigham and Women's Hospital (BWH) sustained SIGSWs. In this study, we illustrate the role of FT in the management of SIGSWs through an aesthetic, functional, and psychosocial examination of outcomes., Methods: We performed a retrospective analysis of individuals with SIGSWs who were screened at BWH between 2008 and 2015. We then collected data of the injuries, modes of conventional reconstruction (CR), and deficits. For the FT recipients, we critically reviewed the psychosocial screening process and post-transplantation aesthetic, functional, and psychosocial outcomes., Results: A total of six individuals post-SIGSWs were screened for FT. All of them had undergone CR, with five receiving loco-regional flaps and free tissue transfers, and one undergoing serial debridement and primary soft-tissue repair. Following CR, all suffered from residual functional and aesthetic deficits. Two underwent partial FT and one is currently undergoing FT screening. We describe the pre-transplant psychosocial screening process and the aesthetic, functional, and psychosocial outcomes of the SIGSW FT recipients., Conclusions: We examined the facial SIGSW injury, outcomes of CR, and the mechanism of FT to offer a potential solution to the shortcomings of CR. More importantly, we highlight the critical nature of the psychosocial component of the multidisciplinary evaluation given the history of mental illness and suicidal behavior in this subset of patients., (Copyright © 2016 British Association of Plastic, Reconstructive and Aesthetic Surgeons. Published by Elsevier Ltd. All rights reserved.)

The free-living legume symbiont Rhizobium leguminosarum is of significant economic value because of its ability to provide fixed nitrogen to globally important leguminous food crops, such as peas and lentils. Discovery based research into the genetics and physiology of R. leguminosarum provides the foundational knowledge necessary for understanding the bacterium's complex lifestyle, necessary for augmenting its use in an agricultural setting. Transposon insertion sequencing (INSeq) facilitates high-throughput forward genetic screening at a genomic scale to identify individual genes required for growth in a specific environment. In this study we applied INSeq to screen the genome of R. leguminosarum bv. viciae strain 3841 (RLV3841) for genes required for growth on minimal mannitol containing medium. Results from this study were contrasted with a prior INSeq experiment screened on peptide rich media to identify a common set of functional genes necessary for basic physiology. Contrasting the two growth conditions indicated that approximately 10% of the chromosome was required for growth, under both growth conditions. Specific genes that were essential to singular growth conditions were also identified. Data from INSeq screening on mannitol as a sole carbon source were used to reconstruct a metabolic map summarizing growth impaired phenotypes observed in the Embden-Meyerhof-Parnas pathway, Entner-Doudoroff pathway, pentose phosphate pathway, and tricarboxylic acid cycle. This revealed the presence of mannitol dependent and independent metabolic pathways required for growth, along with identifying metabolic steps with isozymes or possible carbon flux by-passes. Additionally, genes were identified on plasmids pRL11 and pRL12 that are likely to encode functional activities important to the central physiology of RLV3841.

Bacteriophages may play an important role in regulating population size and diversity of the root nodule symbiont Rhizobium leguminosarum, as well as participating in horizontal gene transfer. Although phages that infect this species have been isolated in the past, our knowledge of their molecular biology, and especially of genome composition, is extremely limited, and this lack of information impacts on the ability to assess phage population dynamics and limits potential agricultural applications of rhizobiophages. To help address this deficit in available sequence and biological information, the complete genome sequence of the Myoviridae temperate phage PPF1 that infects R. leguminosarum biovar viciae strain F1 was determined. The genome is 54,506 bp in length with an average G+C content of 61.9 %. The genome contains 94 putative open reading frames (ORFs) and 74.5 % of these predicted ORFs share homology at the protein level with previously reported sequences in the database. However, putative functions could only be assigned to 25.5 % (24 ORFs) of the predicted genes. PPF1 was capable of efficiently lysogenizing its rhizobial host R. leguminosarum F1. The site-specific recombination system of the phage targets an integration site that lies within a putative tRNA-Pro (CGG) gene in R. leguminosarum F1. Upon integration, the phage is capable of restoring the disrupted tRNA gene, owing to the 50 bp homologous sequence (att core region) it shares with its rhizobial host genome. Phage PPF1 is the first temperate phage infecting members of the genus Rhizobium for which a complete genome sequence, as well as other biological data such as the integration site, is available.

Copy number variations associate with different developmental phenotypes and represent a major cause of congenital anomalies of the kidney and urinary tract (CAKUT). Because rare pathogenic copy number variations are often large and contain multiple genes, identification of the underlying genetic drivers has proven to be difficult. Here we studied the role of rare copy number variations in 80 patients from the KIMONO study cohort for which pathogenic mutations in three genes commonly implicated in CAKUT were excluded. In total, 13 known or novel genomic imbalances in 11 of 80 patients were absent or extremely rare in 23,362 population controls. To identify the most likely genetic drivers for the CAKUT phenotype underlying these rare copy number variations, we used a systematic in silico approach based on frequency in a large data set of controls, annotation with publicly available databases for developmental diseases, tolerance and haploinsufficiency scores, and gene expression profile in the developing kidney and urinary tract. Five novel candidate genes for CAKUT were identified that showed specific expression in the human and mouse developing urinary tract. Among these genes, DLG1 and KIF12 are likely novel susceptibility genes for CAKUT in humans. Thus, there is a significant role of genomic imbalance in the determination of kidney developmental phenotypes. Additionally, we defined a systematic strategy to identify genetic drivers underlying rare copy number variations.

Rheinheimera sp. KL1 was isolated from an algal bloom in Katepwa Lake, Saskatchewan, Canada. The isolate shares genetic and physiological similarities with Rheinheimera tangshanensis. The genome is estimated to be 4,295,060 bp in length with a GC content of 46.37%. Sequence analysis suggests the strain carries a previously uncharacterized prophage., (Copyright © 2015 O’Connor et al.)

Here, we report the first draft genome sequence of Salmonella enterica subsp. enterica serovar Livingstone. This S. Livingstone strain CKY-S4 displayed biofilm formation and cellulose production and could persist on lettuce. This genome may help the study of mechanisms by which enteric pathogens colonize food crops., (Copyright © 2015 Tambalo et al.)

Importance: The relatively high and possibly rising incidence of mouth squamous cell carcinoma in nonsmokers, especially women, without obvious cause has been noted by previous authors. Is chronic dental trauma and irritation a carcinogen, and what is its importance compared with human papillomavirus (HPV) oropharyngeal cancer in nonsmokers?, Objective: To determine whether oral cavity cancers occurred more commonly at sites of dental trauma and how the position of these cancers varied between nonsmokers lacking major identified carcinogens and smokers. If these cancers occurred more frequently at sites of chronic trauma, especially in nonsmokers, it would suggest chronic dental trauma as a possible carcinogen., Design, Setting, and Participants: A retrospective analysis of 881 patients with oral cavity or oropharyngeal cancers seen through a tertiary referral hospital between 2001 and 2011 was performed., Main Outcomes and Measures: Patient medical records were analyzed to determine the location of the tumor within the oral cavity and oropharynx and how it relates to patient demographics, smoking and alcohol histories, and comorbidities. Dental histories were also sought, including use of dentures., Results: Nonsmokers comprised 87 of 390 patients with mouth cancer (22%) and 48 of 334 patients with oropharyngeal cancer (14%). Female nonsmoking patients included 53 with oral cancer (61%) but only 12 with oropharyngeal squamous cell carcinoma (25%). Oral cancers occurred on the lateral tongue, a potential site of chronic dental trauma, in 57 nonsmokers (66%) compared with 107 smokers/ex-smokers (33%) (P < .001). Gingival and floor of mouth lesions occurred in older patients, possibly from chronic denture rubbing. Twenty-six patients had dental abnormalities recorded in close proximity to where their tumor developed., Conclusions and Relevance: Oral cavity cancers occur predominantly at sites of potential dental and denture trauma, especially in nonsmokers without other risk factors. Recognizing teeth irritation as a potential carcinogen would have an impact on prevention and treatment strategies.

Background: The Rhizobiaceae family of Gram-negative bacteria often engage in symbiosis with plants of economic importance. Historically, genetic studies to identify the function of individual genes, and characterize the biology of these bacteria have relied on the use of classical transposon mutagenesis. To increase the rate of scientific discovery in the Rhizobiaceae there is a need to adapt high-throughput genetic screens like insertion sequencing for use in this family of bacteria. Here we describe a Rhizobiaceae compatible MmeI-adapted mariner transposon that can be used with insertion sequencing for high-throughput genetic screening., Results: The newly constructed mariner transposon pSAM&#95;Rl mutagenized R. leguminosarum, S. meliloti, and A. tumefaciens at a high frequency. In R. leguminosarum, mutant pools were generated that saturated 88% of potential mariner insertions sites in the genome. Analysis of the R. leguminosarum transposon insertion sequencing data with a previously described hidden Markov model-based method resulted in assignment of the contribution of all annotated genes in the R. leguminosarum 3841 genome for growth on a complex medium. Good concordance was observed between genes observed to be required for growth on the complex medium, and previous studies., Conclusions: The newly described Rhizobiaceaee compatible mariner transposon insertion sequencing vector pSAM&#95;Rl has been shown to mutagenize at a high frequency and to be an effective tool for use in high-throughput genetic screening. The construction and validation of this transposon insertion sequencing tool for use in the Rhizobiziaceae will provide an opportunity for researchers in the Rhizobiaceae community to use high-throughput genetic screening, allowing for significant increase in the rate of genetic discovery, particularly given the recent release of genome sequences from many Rhizobiaceae strains.

G(M1)-gangliosidosis is a rare progressive neurodegenerative disorder due to an autosomal recessively inherited deficiency of lysosomal β-galactosidase. We have identified seven American black bears (Ursus americanus) found in the Northeast United States suffering from G(M1)-gangliosidosis. This report describes the clinical features, brain MRI, and morphologic, biochemical and molecular genetic findings in the affected bears. Brain lipids were compared with those in the brain of a G(M1)-mouse. The bears presented at ages 10-14 months in poor clinical condition, lethargic, tremulous and ataxic. They continued to decline and were humanely euthanized. The T(2)-weighted MR images of the brain of one bear disclosed white matter hyperintensity. Morphological studies of the brain from five of the bears revealed enlarged neurons with foamy cytoplasm containing granules. Axonal spheroids were present in white matter. Electron microscopic examination revealed lamellated membrane structures within neurons. Cytoplasmic vacuoles were found in the liver, kidneys and chondrocytes and foamy macrophages within the lungs. Acid β-galactosidase activity in cultured skin fibroblasts was only 1-2% of control values. In the brain, ganglioside-bound sialic acid was increased more than 2-fold with G(M1)-ganglioside predominating. G(A1) content was also increased whereas cerebrosides and sulfatides were markedly decreased. The distribution of gangliosides was similar to that in the G(M1)-mouse brain, but the loss of myelin lipids was greater in the brain of the affected bear than in the brain of the G(M1) mouse. Isolated full-length cDNA of the black bear GLB1 gene revealed 86% homology to its human counterpart in nucleotide sequence and 82% in amino acid sequence. GLB1 cDNA from liver tissue of an affected bear contained a homozygous recessive T(1042) to C transition inducing a Tyr348 to His mutation (Y348H) within a highly conserved region of the GLB1 gene. The coincidence of several black bears with G(M1)-gangliosidosis in the same geographic area suggests increased frequency of a founder mutation in this animal population., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Background: Congenital abnormalities of the kidney and the urinary tract are the most common cause of pediatric kidney failure. These disorders are highly heterogeneous, and the etiologic factors are poorly understood., Methods: We performed genomewide linkage analysis and whole-exome sequencing in a family with an autosomal dominant form of congenital abnormalities of the kidney or urinary tract (seven affected family members). We also performed a sequence analysis in 311 unrelated patients, as well as histologic and functional studies., Results: Linkage analysis identified five regions of the genome that were shared among all affected family members. Exome sequencing identified a single, rare, deleterious variant within these linkage intervals, a heterozygous splice-site mutation in the dual serine-threonine and tyrosine protein kinase gene (DSTYK). This variant, which resulted in aberrant splicing of messenger RNA, was present in all affected family members. Additional, independent DSTYK mutations, including nonsense and splice-site mutations, were detected in 7 of 311 unrelated patients. DSTYK is highly expressed in the maturing epithelia of all major organs, localizing to cell membranes. Knockdown in zebrafish resulted in developmental defects in multiple organs, which suggested loss of fibroblast growth factor (FGF) signaling. Consistent with this finding is the observation that DSTYK colocalizes with FGF receptors in the ureteric bud and metanephric mesenchyme. DSTYK knockdown in human embryonic kidney cells inhibited FGF-stimulated phosphorylation of extracellular-signal-regulated kinase (ERK), the principal signal downstream of receptor tyrosine kinases., Conclusions: We detected independent DSTYK mutations in 2.3% of patients with congenital abnormalities of the kidney or urinary tract, a finding that suggests that DSTYK is a major determinant of human urinary tract development, downstream of FGF signaling. (Funded by the National Institutes of Health and others.).

We examined the burden of large, rare, copy-number variants (CNVs) in 192 individuals with renal hypodysplasia (RHD) and replicated findings in 330 RHD cases from two independent cohorts. CNV distribution was significantly skewed toward larger gene-disrupting events in RHD cases compared to 4,733 ethnicity-matched controls (p = 4.8 × 10(-11)). This excess was attributable to known and novel (i.e., not present in any database or in the literature) genomic disorders. All together, 55/522 (10.5%) RHD cases harbored 34 distinct known genomic disorders, which were detected in only 0.2% of 13,839 population controls (p = 1.2 × 10(-58)). Another 32 (6.1%) RHD cases harbored large gene-disrupting CNVs that were absent from or extremely rare in the 13,839 population controls, identifying 38 potential novel or rare genomic disorders for this trait. Deletions at the HNF1B locus and the DiGeorge/velocardiofacial locus were most frequent. However, the majority of disorders were detected in a single individual. Genomic disorders were detected in 22.5% of individuals with multiple malformations and 14.5% of individuals with isolated urinary-tract defects; 14 individuals harbored two or more diagnostic or rare CNVs. Strikingly, the majority of the known CNV disorders detected in the RHD cohort have previous associations with developmental delay or neuropsychiatric diseases. Up to 16.6% of individuals with kidney malformations had a molecular diagnosis attributable to a copy-number disorder, suggesting kidney malformations as a sentinel manifestation of pathogenic genomic imbalances. A search for pathogenic CNVs should be considered in this population for the diagnosis of their specific genomic disorders and for the evaluation of the potential for developmental delay., (Copyright © 2012 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

To identify gene loci associated with steroid-resistant nephrotic syndrome (SRNS), we utilized homozygosity mapping and exome sequencing in a consanguineous pedigree with three affected siblings. High-density genotyping identified three segments of homozygosity spanning 33.6 Mb on chromosomes 5, 10, and 15 containing 296 candidate genes. Exome sequencing identified two homozygous missense variants within the chromosome 15 segment; an A159P substitution in myosin 1E (MYO1E), encoding a podocyte cytoskeletal protein; and an E181K substitution in nei endonuclease VIII-like 1 (NEIL1), encoding a base-excision DNA repair enzyme. Both variants disrupt highly conserved protein sequences and were absent in public databases, 247 healthy controls, and 286 patients with nephrotic syndrome. The MYO1E A159P variant is noteworthy, as it is expected to impair ligand binding and actin interaction in the MYO1E motor domain. The predicted loss of function is consistent with the previous demonstration that Myo1e inactivation produces nephrotic syndrome in mice. Screening 71 additional patients with SRNS, however, did not identify independent NEIL1 or MYO1E mutations, suggesting larger sequencing efforts are needed to uncover which mutation is responsible for the phenotype. Our findings demonstrate the utility of exome sequencing for rapidly identifying candidate genes for human SRNS.

To serve African-American families effectively, marriage and family therapists need to develop a level of cultural competence. This content analysis of the relevant treatment literature was conducted to examine the most common expert recommendations for family therapy with African Americans. Fifteen specific guidelines were generated, including orient the family to therapy, do not assume familiarity, address issue of racism, intervene multi-systemically, do home visits, use problem-solving focus, involve religious leader, incorporate the father, and acknowledge strengths. Conceptual and empirical support for each guideline is discussed, and conclusions are made regarding culturally competent therapy with African-American families.

As the Hispanic population of the United States continues to grow, so will the need for therapists who have been trained to work with Hispanic families. This content analysis of the available treatment literature generated several specific guidelines that can be used in training and evaluating culturally competent therapists. Guidelines included: Use family therapy, act as advocate for the family, assess immigration experience, assess acculturation, respect father, interview family subsystems separately, do not force changes, provide concrete suggestions, and warmly engage the family. Empirical and conceptual support for each guideline is discussed and several conclusions are made regarding culturally competent therapy with Hispanic families.

We investigated the pulmonary vascular effects of E4021, a potent inhibitor of cGMP-specific phosphodiesterase, in control late-gestation fetal lambs, and in newborn lambs with persistent pulmonary hypertension (PPHN) after prenatal ligation of the ductus arteriosus. E4021 alone significantly relaxed fifth-generation pulmonary arteries isolated from control fetal lambs, an effect completely blocked after inhibition of nitric oxide synthase (NOS). In contrast, E4021 did not relax pulmonary arteries isolated from hypertensive lambs. Pretreatment with E4021 (10(-7) M) significantly enhanced relaxations to the NO donor S-nitrosyl-acetyl-penicilamine (SNAP) in arteries from both control and hypertensive lambs. In control, fully instrumented fetal lambs, infusions of E4021 (31 microgram/min) selectively dilated the pulmonary circulation, an effect again blocked after inhibition of NO synthase. Further studies were performed in newborn lambs with PPHN to study the vascular effects of E4021 alone, and in combination with inhaled NO. E4021 alone (1 to 100 microgram/kg/min) decreased pulmonary artery pressure (Ppa) in a dose-dependent fashion, and had minimal effect on systemic pressure. At the highest dose (100 microgram/kg/min), the dilation was selective for the pulmonary circulation. In subsequent protocols, E4021 (10 microgram/kg/min) significantly decreased Ppa and pulmonary vascular resistance (PVR), but these pulmonary vascular effects were not enhanced after NO inhalation at 0.5 or 5 ppm. We speculate that the lack of enhancement was due to the dramatic effects of E4021 alone. Potent, specific phosphodiesterase inhibitors such as E4021 may prove to be useful in the treatment of PPHN.