Publications by authors named "Maggie Williams"

55 Publications

Heterozygous ANKRD17 loss-of-function variants cause a syndrome with intellectual disability, speech delay, and dysmorphism.

Am J Hum Genet 2021 Apr 21. Epub 2021 Apr 21.

Institute for Human Genetics, University of California, San Francisco, San Francisco, CA 94143, USA.

ANKRD17 is an ankyrin repeat-containing protein thought to play a role in cell cycle progression, whose ortholog in Drosophila functions in the Hippo pathway as a co-factor of Yorkie. Here, we delineate a neurodevelopmental disorder caused by de novo heterozygous ANKRD17 variants. The mutational spectrum of this cohort of 34 individuals from 32 families is highly suggestive of haploinsufficiency as the underlying mechanism of disease, with 21 truncating or essential splice site variants, 9 missense variants, 1 in-frame insertion-deletion, and 1 microdeletion (1.16 Mb). Consequently, our data indicate that loss of ANKRD17 is likely the main cause of phenotypes previously associated with large multi-gene chromosomal aberrations of the 4q13.3 region. Protein modeling suggests that most of the missense variants disrupt the stability of the ankyrin repeats through alteration of core structural residues. The major phenotypic characteristic of our cohort is a variable degree of developmental delay/intellectual disability, particularly affecting speech, while additional features include growth failure, feeding difficulties, non-specific MRI abnormalities, epilepsy and/or abnormal EEG, predisposition to recurrent infections (mostly bacterial), ophthalmological abnormalities, gait/balance disturbance, and joint hypermobility. Moreover, many individuals shared similar dysmorphic facial features. Analysis of single-cell RNA-seq data from the developing human telencephalon indicated ANKRD17 expression at multiple stages of neurogenesis, adding further evidence to the assertion that damaging ANKRD17 variants cause a neurodevelopmental disorder.
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http://dx.doi.org/10.1016/j.ajhg.2021.04.007DOI Listing
April 2021

Extended-Spectrum β-Lactamase and Carbapenemase Genes are Substantially and Sequentially Reduced during Conveyance and Treatment of Urban Sewage.

Environ Sci Technol 2021 05 22;55(9):5939-5949. Epub 2021 Apr 22.

Department of Environmental Engineering, Technical University of Denmark, Lyngby 2800, DK.

Urban wastewater systems (UWSs) are a main receptacle of excreted antibiotic resistance genes (ARGs) and their host microorganisms. However, we lack integrated and quantitative observations of the occurrence of ARGs in the UWS to characterize the sources and identify processes that contribute to their fate. We sampled the UWSs from three medium-size cities in Denmark, Spain, and the United Kingdom and quantified 70 clinically important extended-spectrum β-lactamase and carbapenemase genes along with the mobile genetic elements and microbial communities. Results from all three countries showed that sewage-especially from hospitals-carried substantial loads of ARGs (10-10 copies per person equivalent), but these loads progressively declined along sewers and through sewage treatment plants, resulting in minimal emissions (10-10 copies per person equivalent). Removal was primarily during sewage conveyance (65 ± 36%) rather than within sewage treatment (34 ± 23%). The extended-spectrum β-lactamase and carbapenemase genes were clustered in groups based on their persistence in the UWS compartments. The less-persistent groups were associated to putative host taxa (especially Enterobacteriaceae and Moraxellaceae), while the more persistent groups appeared horizontally transferred and correlated significantly with total cell numbers and mobile genetic elements. This documentation of a substantial ARG reduction during sewage conveyance provides opportunities for antibiotic resistance management and a caution for sewage-based antibiotic resistance surveillance.
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http://dx.doi.org/10.1021/acs.est.0c08548DOI Listing
May 2021

Consensus statement on standards and guidelines for the molecular diagnostics of Alport syndrome: refining the ACMG criteria.

Eur J Hum Genet 2021 Apr 15. Epub 2021 Apr 15.

Medical Genetics Unit, Department of Clinical and Experimental Biomedical Sciences "Mario Serio", University of Florence, Florence, Italy.

The recent Chandos House meeting of the Alport Variant Collaborative extended the indications for screening for pathogenic variants in the COL4A5, COL4A3 and COL4A4 genes beyond the classical Alport phenotype (haematuria, renal failure; family history of haematuria or renal failure) to include persistent proteinuria, steroid-resistant nephrotic syndrome, focal and segmental glomerulosclerosis (FSGS), familial IgA glomerulonephritis and end-stage kidney failure without an obvious cause. The meeting refined the ACMG criteria for variant assessment for the Alport genes (COL4A3-5). It identified 'mutational hotspots' (PM1) in the collagen IV α5, α3 and α4 chains including position 1 Glycine residues in the Gly-X-Y repeats in the intermediate collagenous domains; and Cysteine residues in the carboxy non-collagenous domain (PP3). It considered that 'well-established' functional assays (PS3, BS3) were still mainly research tools but sequencing and minigene assays were commonly used to confirm splicing variants. It was not possible to define the Minor Allele Frequency (MAF) threshold above which variants were considered Benign (BA1, BS1), because of the different modes of inheritances of Alport syndrome, and the occurrence of hypomorphic variants (often Glycine adjacent to a non-collagenous interruption) and local founder effects. Heterozygous COL4A3 and COL4A4 variants were common 'incidental' findings also present in normal reference databases. The recognition and interpretation of hypomorphic variants in the COL4A3-COL4A5 genes remains a challenge.
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http://dx.doi.org/10.1038/s41431-021-00858-1DOI Listing
April 2021

Syndromic disorders caused by gain-of-function variants in KCNH1, KCNK4, and KCNN3-a subgroup of K channelopathies.

Eur J Hum Genet 2021 Feb 16. Epub 2021 Feb 16.

Institute of Human Genetics, University Medical Center Hamburg-Eppendorf, Hamburg, Germany.

Decreased or increased activity of potassium channels caused by loss-of-function and gain-of-function (GOF) variants in the corresponding genes, respectively, underlies a broad spectrum of human disorders affecting the central nervous system, heart, kidney, and other organs. While the association of epilepsy and intellectual disability (ID) with variants affecting function in genes encoding potassium channels is well known, GOF missense variants in K channel encoding genes in individuals with syndromic developmental disorders have only recently been recognized. These syndromic phenotypes include Zimmermann-Laband and Temple-Baraitser syndromes, caused by dominant variants in KCNH1, FHEIG syndrome due to dominant variants in KCNK4, and the clinical picture associated with dominant variants in KCNN3. Here we review the presentation of these individuals, including five newly reported with variants in KCNH1 and three additional individuals with KCNN3 variants, all variants likely affecting function. There is notable overlap in the phenotypic findings of these syndromes associated with dominant KCNN3, KCNH1, and KCNK4 variants, sharing developmental delay and/or ID, coarse facial features, gingival enlargement, distal digital hypoplasia, and hypertrichosis. We suggest to combine the phenotypes and define a new subgroup of potassium channelopathies caused by increased K conductance, referred to as syndromic neurodevelopmental K channelopathies due to dominant variants in KCNH1, KCNK4, or KCNN3.
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http://dx.doi.org/10.1038/s41431-021-00818-9DOI Listing
February 2021

AIFM1-associated X-linked spondylometaphyseal dysplasia with cerebral hypomyelination.

Am J Med Genet A 2021 04 13;185(4):1228-1235. Epub 2021 Jan 13.

Department of Clinical Genetics, University Hospitals Bristol and Weston NHS Foundation Trust, Bristol, United Kingdom.

Spondylometaphyseal dysplasia with cerebral hypomyelination (SMD-H) is a very rare but distinctive phenotype, unusually combining spondylometaphyseal dysplasia with hypomyelinating leukodystrophy. Recently, SMD-H has been associated with variants confined to a specific intra-genic locus involving Exon 7, suggesting that AIFM1 plays an important role in bone development and metabolism as well as cerebral myelination. Here we describe two further affected boys, one with a novel intronic variant associated with skipping of Exon 7 of AIFM1 and the other a synonymous variant within Exon 7 of AIFM1. We describe their clinical course and radiological and genetic findings, providing further insight into the natural history of this condition.
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http://dx.doi.org/10.1002/ajmg.a.62072DOI Listing
April 2021

Response to First Course of Intensified Immunosuppression in Genetically Stratified Steroid Resistant Nephrotic Syndrome.

Clin J Am Soc Nephrol 2020 07 21;15(7):983-994. Epub 2020 Apr 21.

Bristol Renal, Translational Health Sciences, Bristol Medical School, University of Bristol, Bristol, United Kingdom

Background And Objectives: Intensified immunosuppression in steroid-resistant nephrotic syndrome is broadly applied, with disparate outcomes. This review of patients from the United Kingdom National Study of Nephrotic Syndrome cohort aimed to improve disease stratification by determining, in comprehensively genetically screened patients with steroid-resistant nephrotic syndrome, if there is an association between response to initial intensified immunosuppression and disease progression and/or post-transplant recurrence.

Design, Setting, Participants, & Measurements: Pediatric patients with steroid-resistant nephrotic syndrome were recruited the UK National Registry of Rare Kidney Diseases. All patients were whole-genome sequenced, whole-exome sequenced, or steroid-resistant nephrotic syndrome gene-panel sequenced. Complete response or partial response within 6 months of starting intensified immunosuppression was ascertained using laboratory data. Response to intensified immunosuppression and outcomes were analyzed according to genetic testing results, pattern of steroid resistance, and first biopsy findings.

Results: Of 271 patients, 178 (92 males, median onset age 4.7 years) received intensified immunosuppression with response available. A total of 4% of patients with monogenic disease showed complete response, compared with 25% of genetic-testing-negative patients (=0.02). None of the former recurred post-transplantation. In genetic-testing-negative patients, 97% with complete response to first intensified immunosuppression did not progress, whereas 44% of nonresponders developed kidney failure with 73% recurrence post-transplant. Secondary steroid resistance had a higher complete response rate than primary/presumed resistance (43% versus 23%; =0.001). The highest complete response rate in secondary steroid resistance was to rituximab (64%). Biopsy results showed no correlation with intensified immunosuppression response or outcome.

Conclusions: Patients with monogenic steroid-resistant nephrotic syndrome had a poor therapeutic response and no post-transplant recurrence. In genetic-testing-negative patients, there was an association between response to first intensified immunosuppression and long-term outcome. Patients with complete response rarely progressed to kidney failure, whereas nonresponders had poor kidney survival and a high post-transplant recurrence rate. Patients with secondary steroid resistance were more likely to respond, particularly to rituximab.
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http://dx.doi.org/10.2215/CJN.13371019DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7341765PMC
July 2020

Employees' knowledge and practices on occupational exposure to tuberculosis at specialised tuberculosis hospitals in South Africa.

Curationis 2020 Apr 1;43(1):e1-e8. Epub 2020 Apr 1.

Department of Nursing Science, Nelson Mandela University, Port Elizabeth.

Background: To prevent the spread of infection of tuberculosis (TB), sufficient knowledge and safe practices regarding occupational exposure are crucial for all employees working in TB hospitals.

Objectives: To explore and describe the knowledge and practices of employees working in three specialised TB hospitals in Nelson Mandela Bay, Eastern Cape, regarding occupational exposure to TB.

Methods: A quantitative, descriptive and contextual study was conducted using convenience sampling to have 181 employees at the three hospitals elected to complete the self-administered questionnaire, which was distributed in December 2016. Three scores on a scale of 0-10 were calculated per participant: knowledge, personal practice and institutional practice. Descriptive and inferential statistics were utilised.

Results: Approximately, one-third (34%) of the participants were between the ages of 36 and 45 years. Most of the participants (63%) attended high school and less than one-third (28%) had a tertiary qualification. The majority of participants (62%) had not received any clinical training. Participants displayed high scores ( 6) for knowledge (75%; mean = 6.65), personal practice (68%; mean = 6.12) and institutional practice (51%; mean = 6.15). The correlation between knowledge and personal practice was found to be non-significant (r = 0.033). An analysis of variance revealed that Knowledge is significantly related to age and education level.

Conclusion: Employees' knowledge regarding occupational TB exposure was generally high, but they were not necessarily practicing what they knew. Further research is required regarding appropriate managerial interventions to ensure that employees' practices improve, which should reduce the risk of occupational TB exposure.
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http://dx.doi.org/10.4102/curationis.v43i1.2039DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7203248PMC
April 2020

Association of circulating adipokine concentrations with indices of adiposity and sex in healthy, adult client owned cats.

BMC Vet Res 2019 Sep 18;15(1):332. Epub 2019 Sep 18.

Department of Production Animal Health, Faculty of Veterinary Medicine, University of Calgary, 3330 Hospital Dr. NW, Calgary, AB, T2N 4N1, Canada.

Background: Both diabetes mellitus (DM) and obesity are common in cats. The adipokines leptin, adiponectin, resistin and omentin are thought to have important roles in human obesity and glucose homeostasis; however, their functions in the pathophysiology of feline diabetes mellitus and obesity are poorly understood. We determined whether sexual dimorphism exists for circulating concentrations of these adipokines, whether they are associated with adiposity, and whether they correlate with basic indices of insulin sensitivity in cats. Healthy, client-owned male and female cats that were either ideal weight or obese were recruited into the study. Fasting blood glucose, fructosamine, cholesterol, triglycerides, insulin and plasma concentrations of adipokines were evaluated.

Results: Obese cats had greater serum concentrations of glucose and triglycerides than ideal weight cats, but fructosamine and cholesterol concentrations did not differ between groups. Body weight and body mass index were greater in male than female cats, but circulating metabolite cocentrations were similar between sexes of both the ideal weight and obese groups. Plasma concentrations of insulin and leptin were greater in obese than ideal weight cats, with reciprocal reduction in adiponectin concentrations in obese cats; there were no sex differences in these hormones. Interestingly, plasma omentin concentrations were greater in male than female cats but with no differences between obese and ideal weight states.

Conclusion: Together our findings suggest that rather than gender, body weight and adiposity are more important determinants of circulating concentrations of the adipokines leptin and adiponectin. On the contrary, the adipokine omentin is not affected by body weight or adiposity but instead exhibits sexual dimorphism in cats.
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http://dx.doi.org/10.1186/s12917-019-2080-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6749635PMC
September 2019

Classic galactosaemia in the Greek Cypriot population: An epidemiological and molecular study.

Ann Hum Genet 2019 09 17;83(5):291-298. Epub 2019 Apr 17.

Department of Biochemical Genetics, The Cyprus Institute of Neurology and Genetics, Nicosia, Cyprus.

Classic galactosaemia is an inherited metabolic disorder of galactose metabolism caused by deficiency of the enzyme galactose-1-phosphate uridyltransferase (GALT) resulting from mutations in the GALT gene. The objectives of the present study were the determination of the carrier frequency of classic galactosaemia in the Greek Cypriot population and the molecular characterization of the disease alleles. We performed an epidemiological study involving 528 Greek Cypriots originating from all parts of Cyprus. Carriers were identified by measuring GALT activity in red blood cells and were subsequently subjected to mutation analysis. A total of five mutations were identified in patients and carriers of classic galactosaemia: a large deletion of 8.5 kb previously reported by us (55% of alleles), the known mutations p.Lys285Asn (30%), p.Pro185Ser (5%), and c.820+13A>G (5%), and a novel mutation c.378-12G>A (5%). Interestingly, the most common mutation in European populations, p.Gln188Arg, was not identified in this Cypriot cohort. The carrier frequency for classic galactosaemia among Greek Cypriots was estimated to be 1:88, predicting a homozygote incidence of 1:31,000 births. The Duarte 1 and Duarte 2 variants were found to be present at a frequency of 5.5% and 2.5%, respectively.
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http://dx.doi.org/10.1111/ahg.12318DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6766971PMC
September 2019

Direct or DNA Extraction-Free Amplification and Quantification of Foodborne Pathogens.

Methods Mol Biol 2019 ;1918:21-33

Department of Civil and Environmental Engineering, Michigan State University, East Lansing, MI, USA.

The use of direct nucleic acid amplification of pathogens from food matrices has the potential to reduce time to results over DNA extraction-based approaches as well as traditional culture-based approaches. Here we describe protocols for assay design and experiments for direct amplification of foodborne pathogens in food sample matrices using loop-mediated isothermal amplification (LAMP) and polymerase chain reaction (PCR). The examples provided include the detection Escherichia coli in milk samples and Salmonella in pork meat samples. This protocol includes relevant reagents and methods including obtaining target sequences, assay design, sample processing, and amplification. These methods, though used for specific example matrices, could be applied to many other foodborne pathogens and sample types.
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http://dx.doi.org/10.1007/978-1-4939-9000-9_2DOI Listing
June 2019

Toward interprofessional service-learning and social accountability in health: One South African University's process-oriented-participatory journey.

J Interprof Care 2019 May-Jun;33(3):291-294. Epub 2018 Oct 15.

b Department of Family and Preventive Medicine , University of Arkansas for Medical Sciences , Little Rock , USA.

The Nelson Mandela University in Port Elizabeth, South Africa, seeks to transform its health professions curricula in order to achieve equity in health outcomes. Integral to this are interprofessional education service-learning initiatives attendant to socially accountable objectives. We describe one such initiative, the Zanempilo Mobile Health Education Platform (MHEP), which engages interprofessional healthcare students and faculty members in delivering health services to underserved communities. The Zanempilo MHEP consists of a converted 13-ton truck as a mobile clinic from where student-run services are provided. We illustrate the intentional process by which we, an interprofessional health science working group, created socially accountable learning goals appropriate to the above platform. We developed, employed, and refined a process-oriented-participatory approach rooted in theories of social constructivism and social network development that included the following phases: orientation, analysis, synthesis, production, and dissemination. Out of this approach emerged several socially accountable learning goals for students and faculty members working on the Zanempilo MHEP. These goals incorporated five educational domains, namely knowledge, attitudes, skills, intentions, and relationships. We anticipate using these goals to identify future curricular objectives and competencies.
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http://dx.doi.org/10.1080/13561820.2018.1532399DOI Listing
December 2019

Clinical utility of the polygenic LDL-C SNP score in familial hypercholesterolemia.

Atherosclerosis 2018 10;277:457-463

Institute of Cardiovascular Science, University College London, 5 University St, London, WC1E 6JF, United Kingdom. Electronic address:

Mutations in any of three genes (LDLR, APOB and PCSK9) are known to cause autosomal dominant FH, but a mutation can be found in only ∼40% of patients with a clinical diagnosis of FH. In the remainder, a polygenic aetiology may be the cause of the phenotype, due to the co-inheritance of common LDL-C raising variants. In 2013, we reported the development of a 12-SNP LDL-C "SNP-Score" based on common variants identified as LDL-C raising from genome wide association consortium studies, and have confirmed the validity of this score in samples of no-mutation FH adults and children from more than six countries with European-Caucasian populations. In more than 80% of those with a clinical diagnosis of FH but with no detectable mutation in LDLR/APOB/PCSK9, the polygenic explanation is the most likely for their hypercholesterolaemia. Those with a low score (in the bottom two deciles) may have a mutation in a novel gene, and further research including whole exome or whole genome sequencing is warranted. Only in families where the index case has a monogenic cause should cascade testing be carried out, using DNA tests for an unambiguous identification of affected relatives. The clinical utility of the polygenic explanation is that it supports a more conservative (less aggressive) treatment care pathway for those with no mutation. The ability to distinguish those with a clinical diagnosis of FH who have a monogenic or a polygenic cause of their hypercholesterolaemia is a paradigm example of the use of genomic information to inform Precision Medicine using lipid lowering agents with different efficacy and costs.
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http://dx.doi.org/10.1016/j.atherosclerosis.2018.06.006DOI Listing
October 2018

COMPARISON OF THE EFFICACY AND SAFETY OF MEDETOMIDINE-KETAMINE VERSUS MEDETOMIDINE-AZAPERONE-ALFAXALONE COMBINATION IN FREE-RANGING ROCKY MOUNTAIN BIGHORN SHEEP ( OVIS CANADENSIS).

J Zoo Wildl Med 2018 Sep;49(3):662-670

Chemical immobilization is a key aspect of wildlife management. To minimize dose-dependent adverse effects, immobilization protocols often include two or more synergistic agents, which allows for reductions in individual drug dosages. Free-ranging bighorn sheep ( Ovis canadensis) in Canada ( n = 74) were remotely injected with a combination of medetomidine (0.16 ± 0.04 mg/kg) and ketamine (4.0 ± 1.4 mg/kg) (MK), or combination of medetomidine (0.14 ± 0.06 mg/kg), azaperone (0.21 ± 0.11 mg/kg), and alfaxalone (0.45 ± 0.21 mg/kg) (MAA). Once recumbency was achieved, arterial blood samples were collected and immediately analyzed for blood gas and acid-base status. Rectal temperature, heart rate, and respiratory rate were recorded upon recumbency and throughout anesthesia at 5-15 min intervals. At conclusion of the procedures, medetomidine was reversed by intramuscular atipamezole at five times the medetomidine dose. Induction times (mean ± standard deviation) of animals that became immobilized with one dart (8.7 ± 3.2 min, 7.3 ± 3.9 min) and recovery times of all animals (3.4 ± 1.5 min, 3.9 ± 1.6 min) were not significantly different between MK and MAA groups, respectively. Both MK and MAA groups experienced severe hypoxemia (PaO 42 ± 9 mmHg, 40 ± 10 mmHg, respectively). PaCO was significantly higher ( P = 0.0248) in the MK group (median 54 mmHg) than the MAA group (median 48 mmHg) with a trend towards lower pH (7.40 vs 7.42, respectively, P = 0.07). Initially, MK animals had higher heart rates than MAA animals (median 49 vs 40 beats/min), which decreased over time. In bighorn sheep, both MK and MAA produced reliable, reversible immobilization with smooth inductions and recoveries. However, less respiratory depression was seen with MAA than MK.
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http://dx.doi.org/10.1638/2017-0161.1DOI Listing
September 2018

Antimicrobial Resistance in the Environment.

Water Environ Res 2018 Oct;90(10):865-884

Department of Civil and Environmental Engineering, Michigan State University, East Lansing, MI, USA.

This review su mmarizes selected publications from 2017 highlighting the occurrence of antimicrobial resistance (AMR) genes in the environment with emphasis on the aquatic environment. The review also covers different treatment technologies being developed for AMR genes as an environmental contaminant. The progress made in the area of AMR gene databases and tools is also reviewed. Besides a brief introduction, the content is categorized into three main sections: i) Occurrence of AMR in the Environment, ii) Treatment technologies for AMR, and iii) AMR databases and tools.
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http://dx.doi.org/10.2175/106143018X15289915807056DOI Listing
October 2018

Primer set 2.0 for highly parallel qPCR array targeting antibiotic resistance genes and mobile genetic elements.

FEMS Microbiol Ecol 2018 09;94(9)

Department of Civil and Environmental Engineering, Michigan State University, East Lansing, Michigan 48824, USA.

The high-throughput antibiotic resistance gene (ARG) qPCR array, initially published in 2012, is increasingly used to quantify resistance and mobile determinants in environmental matrices. Continued utility of the array; however, necessitates improvements such as removing or redesigning questionable primer sets, updating targeted genes and coverage of available sequences. Towards this goal, a new primer design tool (EcoFunPrimer) was used to aid in identification of conserved regions of diverse genes. The total number of assays used for diverse genes was reduced from 91 old primer sets to 52 new primer sets, with only a 10% loss in sequence coverage. While the old and new array both contain 384 primer sets, a reduction in old primer sets permitted 147 additional ARGs and mobile genetic elements to be targeted. Results of validating the updated array with a mock community of strains resulted in over 98% of tested instances incurring true positive/negative calls. Common queries related to sensitivity, quantification and conventional data analysis (e.g. Ct cutoff value, and estimated genomic copies without standard curves) were also explored. A combined list of new and previously used primer sets is provided with a recommended set based on redesign of primer sets and results of validation.
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http://dx.doi.org/10.1093/femsec/fiy130DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7250373PMC
September 2018

Evaluation of Nucleic Acid Isothermal Amplification Methods for Human Clinical Microbial Infection Detection.

Front Microbiol 2017 12;8:2211. Epub 2017 Dec 12.

Department of Osteopathic Medical Specialties, Section of Emergency Medicine, College of Osteopathic Medicine, Michigan State University, East Lansing, MI, United States.

Battling infection is a major healthcare objective. Untreated infections can rapidly evolve toward the condition of sepsis in which the body begins to fail and resuscitation becomes critical and tenuous. Identification of infection followed by rapid antimicrobial treatment are primary goals of medical care, but precise identification of offending organisms by current methods is slow and broad spectrum empirical therapy is employed to cover most potential pathogens. Current methods for identification of bacterial pathogens in a clinical setting typically require days of time, or a 4- to 8-h growth phase followed by DNA extraction, purification and PCR-based amplification. We demonstrate rapid (70-120 min) genetic diagnostics methods utilizing loop-mediated isothermal amplification (LAMP) to test for 15 common infection pathogen targets, called the Infection Diagnosis Panel (In-Dx). The method utilizes filtration to rapidly concentrate bacteria in sample matrices with lower bacterial loads and direct LAMP amplification without DNA purification from clinical blood, urine, wound, sputum and stool samples. The In-Dx panel was tested using two methods of detection: (1) real-time thermocycler fluorescent detection of LAMP amplification and (2) visual discrimination of color change in the presence of Eriochrome Black T (EBT) dye following amplification. In total, 239 duplicate samples were collected (31 blood, 122 urine, 73 mucocutaneous wound/swab, 11 sputum and two stool) from 229 prospectively enrolled hospital patients with suspected clinical infection and analyzed both at the hospital and by In-Dx. Sensitivity (Se) of the In-Dx panel targets pathogens from urine samples by In-Dx was 91.1% and specificity (Sp) was 97.3%, with a positive predictive value (PPV) of 53.7% and a negative predictive value (NPV) of 99.7% as compared to clinical microbial detection methods. Sensitivity of detection of the In-Dx panel from mucocutaneous swab samples was 65.5% with a Sp of 99.3%, and a PPV of 84% and NPV of 98% as compared to clinical microbial detection methods. Results indicate the LAMP-based In-Dx panel allows rapid and precise diagnosis of clinical infections by targeted pathogens across multiple culture types for point-of-care utilization.
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http://dx.doi.org/10.3389/fmicb.2017.02211DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5732957PMC
December 2017

Expanding the clinical spectrum of recessive truncating mutations of to a Bohring-Opitz-like phenotype.

J Med Genet 2017 12 26;54(12):830-835. Epub 2017 Oct 26.

Inserm UMR 1231 GAD Team, Genetics of Developmental Anomalies, Université de Bourgogne-Franche Comté, Dijon, France.

Background: Bohring-Opitz syndrome (BOS) is a rare genetic disorder characterised by a recognisable craniofacial appearance and a typical 'BOS' posture. BOS is caused by sporadic mutations of. However, several typical patients with BOS have no molecular diagnosis, suggesting clinical and genetic heterogeneity.

Objectives: To expand the phenotypical spectrum of autosomal recessive variants of , reported as causing Crisponi syndrome/cold-induced sweating syndrome type 1 (CS/CISS1)-like syndrome.

Methods: We performed whole-exome sequencing in two families with a suspected recessive mode of inheritance. We used the Matchmaker Exchange initiative to identify additional patients.

Results: Here, we report six patients with microcephaly, facial dysmorphism, including exophthalmos, nevus flammeus of the glabella and joint contractures with a suspected BOS posture in five out of six patients. We identified autosomal recessive truncating mutations in the gene. encodes a BTB-kelch protein implicated in the cell cycle and in protein degradation by the ubiquitin-proteasome pathway. Recently, biallelic mutations in the gene were reported in four families and associated with CS/CISS1, characterised by clinical features overlapping with our patients.

Conclusion: We have expanded the clinical spectrum of autosomal recessive variants by describing a syndrome with features overlapping CS/CISS1 and BOS.
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http://dx.doi.org/10.1136/jmedgenet-2017-104748DOI Listing
December 2017

Chronic pain has a strong impact on quality of life in facioscapulohumeral muscular dystrophy.

Muscle Nerve 2018 03 7;57(3):380-387. Epub 2017 Nov 7.

MRC Centre for Neuromuscular Diseases, Institute of Genetic Medicine, Newcastle University, Central Parkway Newcastle upon Tyne NE1 3 BZ, United Kingdom.

Introduction: Earlier small case series and clinical observations reported on chronic pain playing an important role in facioscapulohumeral dystrophy (FSHD). The aim of this study was to determine the characteristics and impact of pain on quality of life (QoL) in patients with FSHD.

Methods: We analyzed patient reported outcome measures collected through the U.K. FSHD Patient Registry.

Results: Of 398 patients, 88.6% reported pain at the time of study. The most frequent locations were shoulders and lower back. A total of 203 participants reported chronic pain, 30.4% of them as severe. The overall disease impact on QoL was significantly higher in patients with early onset and long disease duration. Chronic pain had a negative impact on all Individualised Neuromuscular Quality of Life Questionnaire domains and overall disease score.

Discussion: Our study shows that pain in FSHD type 1 (FSHD1) is frequent and strongly impacts on QoL, similar to other chronic, painful disorders. Management of pain should be considered when treating FSHD1 patients. Muscle Nerve 57: 380-387, 2018.
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http://dx.doi.org/10.1002/mus.25991DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5836962PMC
March 2018

Isothermal amplification of environmental DNA (eDNA) for direct field-based monitoring and laboratory confirmation of Dreissena sp.

PLoS One 2017 16;12(10):e0186462. Epub 2017 Oct 16.

Department of Civil and Environmental Engineering, Michigan State University, East Lansing, Michigan, United States of America.

Loop-mediated isothermal amplification (LAMP) of aquatic invasive species environmental DNA (AIS eDNA) was used for rapid, sensitive, and specific detection of Dreissena sp. relevant to the Great Lakes (USA) basin. The method was validated for two uses including i) direct amplification of eDNA using a hand filtration system and ii) confirmation of the results after DNA extraction using a conventional thermal cycler run at isothermal temperatures. Direct amplification eliminated the need for DNA extraction and purification and allowed detection of target invasive species in grab or concentrated surface water samples, containing both free DNA as well as larger cells and particulates, such as veligers, eggs, or seeds. The direct amplification method validation was conducted using Dreissena polymorpha and Dreissena bugensis and uses up to 1 L grab water samples for high target abundance (e.g., greater than 10 veligers (larval mussels) per L for Dreissena sp.) or 20 L samples concentrated through 35 μm nylon screens for low target abundance, at less than 10 veligers per liter water. Surface water concentrate samples were collected over a period of three years, mostly from inland lakes in Michigan with the help of a network of volunteers. Field samples collected from 318 surface water locations included i) filtered concentrate for direct amplification validation and ii) 1 L grab water sample for eDNA extraction and confirmation. Though the extraction-based protocol was more sensitive (resulting in more positive detections than direct amplification), direct amplification could be used for rapid screening, allowing for quicker action times. For samples collected between May and August, results of eDNA direct amplification were consistent with known presence/absence of selected invasive species. A cross-platform smartphone application was also developed to disseminate the analyzed results to volunteers. Field tests of the direct amplification protocol using a portable device (Gene-Z) showed the method could be used in the field to obtain results within one hr (from sample to result). Overall, the direct amplification has the potential to simplify the eDNA-based monitoring of multiple aquatic invasive species. Additional studies are warranted to establish quantitative correlation between eDNA copy number, veliger, biomass or organismal abundance in the field.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0186462PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5643059PMC
November 2017

MicroRNAs-Based Inter-Domain Communication between the Host and Members of the Gut Microbiome.

Front Microbiol 2017 27;8:1896. Epub 2017 Sep 27.

Department of Civil and Environmental Engineering, Michigan State University, East Lansing, MI, United States.

The gut microbiome is an important modulator of host gene expression, impacting important functions such as the innate immune response. Recent evidence suggests that the inter-domain communication between the gut microbiome and host may in part occur via microRNAs (small, non-coding RNA molecules) which are often differentially expressed in the presence of bacteria and can even be released and taken up by bacteria. The role of microRNAs in microbiome-host communication in intestinal diseases is not fully understood, particularly in diseases impacted by exposure to environmental toxicants. Here, we review the present knowledge in the areas of microbiome and microRNA expression-based communication, microbiome and intestinal disease relationships, and microRNA expression responses to intestinal diseases. We also examine potential links between host microRNA-microbiota communication and exposure to environmental toxicants by reviewing connections between (i) toxicants and microRNA expression, (ii) toxicants and gut diseases, and (iii) toxicants and the gut microbiome. Future multidisciplinary research in this area is needed to uncover these interactions with the potential to impact how gut-microbiome associated diseases [e.g., inflammatory bowel disease (IBD) and many others] are managed.
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http://dx.doi.org/10.3389/fmicb.2017.01896DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5624305PMC
September 2017

Antimicrobial Resistance in the Environment.

Water Environ Res 2017 Oct;89(10):921-941

This review summarizes selected publications of 2016 with emphasis on occurrence and treatment of antibiotic resistance genes and bacteria in the aquatic environment and wastewater and drinking water treatment plants. The review is conducted with emphasis on fate, modeling, risk assessment and data analysis methodologies for characterizing abundance. After providing a brief introduction, the review is divided into the following four sections: i) Occurrence of AMR in the Environment, ii) Treatment Technologies for AMR, iii) Modeling of Fate, Risk, and Environmental Impact of AMR, and iv) ARG Databases and Pipelines.
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http://dx.doi.org/10.2175/106143017X15023776270179DOI Listing
October 2017

Modulatory Influence of Segmented Filamentous Bacteria on Transcriptomic Response of Gnotobiotic Mice Exposed to TCDD.

Front Microbiol 2017 7;8:1708. Epub 2017 Sep 7.

Department of Civil and Environmental Engineering, East LansingMI, United States.

Environmental toxicants such as 2,3,7,8-tetrachlorodibenzo--dioxin (TCDD), an aryl hydrocarbon receptor (AhR), are known to induce host toxicity and structural shifts in the gut microbiota. Key bacterial populations with similar or opposing functional responses to AhR ligand exposure may potentially help regulate expression of genes associated with immune dysfunction. To examine this question and the mechanisms for AhR ligand-induced bacterial shifts, C57BL/6 gnotobiotic mice were colonized with and without segmented filamentous bacteria (SFB) - an immune activator. Mice were also colonized with polysaccharide A producing - an immune suppressor to serve as a commensal background. Following colonization, mice were administered TCDD (30 μg/kg) every 4 days for 28 days by oral gavage. Quantified with the nCounter mouse immunology panel, opposing responses in ileal gene expression (e.g., genes associated with T-cell differentiation via the class II major histocompatibility complex) as a result of TCDD dosing and SFB colonization were observed. Genes that responded to TCDD in the presence of SFB did not show a significant response in the absence of SFB, and vice versa. Regulatory T-cells examined in the mesenteric lymph-nodes, spleen, and blood were also less impacted by TCDD in mice colonized with SFB. TCDD-induced shifts in abundance of SFB and compared with previous studies in mice with a traditional gut microbiome. With regard to the mouse model colonized with individual populations, results indicate that TCDD-induced host response was significantly modulated by the presence of SFB in the gut microbiome, providing insight into therapeutic potential between AhR ligands and key commensals.
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http://dx.doi.org/10.3389/fmicb.2017.01708DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5594080PMC
September 2017

A splice-site variant in ANKRD11 associated with classical KBG syndrome.

Am J Med Genet A 2017 Oct 17;173(10):2844-2846. Epub 2017 Aug 17.

University Hospitals Bristol NHS Trust, Bristol, England.

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http://dx.doi.org/10.1002/ajmg.a.38397DOI Listing
October 2017

TCDD administered on activated carbon eliminates bioavailability and subsequent shifts to a key murine gut commensal.

Appl Microbiol Biotechnol 2017 Oct 15;101(19):7409-7415. Epub 2017 Aug 15.

Department of Civil and Environmental Engineering, East Lansing, MI, 48824, USA.

Activated carbon (AC) is an increasingly attractive remediation alternative for the sequestration of dioxins at contaminated sites globally. However, the potential for AC to reduce the bioavailability of dioxins in mammals and the residing gut microbiota has received less attention. This question was partially answered in a recent study examining 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced hallmark toxic responses in mice administered with TCDD sequestered by AC or freely available in corn oil by oral gavage. Results from that study support the use of AC to significantly reduce the bioavailability of TCDD to the host. Herein, we examined the bioavailability of TCDD sequestered to AC on a key murine gut commensal and the influence of AC on the community structure of the gut microbiota. The analysis included qPCR to quantify the expression of segmented filamentous bacteria (SFB) in the mouse ileum, which has responded to TCDD-induced host toxicity in previous studies and community structure via sequencing the 16S ribosomal RNA (rRNA) gene. The expression of SFB 16S rRNA gene and functional genes significantly increased with TCDD administered with corn oil vehicle. Such a response was absent when TCDD was sequestered by AC. In addition, AC appeared to have a minimal influence on murine gut community structure and diversity, affecting only the relative abundance of Lactobacillaceae and two other groups. Results of this study further support the remedial use of AC for eliminating bioavailability of TCDD to host and subsequent influence on the gut microbiome.
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http://dx.doi.org/10.1007/s00253-017-8460-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5909190PMC
October 2017

Clinical genetic testing using a custom-designed steroid-resistant nephrotic syndrome gene panel: analysis and recommendations.

J Med Genet 2017 12 5;54(12):795-804. Epub 2017 Aug 5.

Bristol Renal, School of Clinical Sciences, University of Bristol, Bristol, UK.

Background: There are many single-gene causes of steroid-resistant nephrotic syndrome (SRNS) and the list continues to grow rapidly. Prompt comprehensive diagnostic testing is key to realising the clinical benefits of a genetic diagnosis. This report describes a bespoke-designed, targeted next-generation sequencing (NGS) diagnostic gene panel assay to detect variants in 37 genes including the ability to identify copy number variants (CNVs).

Methods: This study reports results of 302 patients referred for SRNS diagnostic gene panel analysis. Phenotype and clinical impact data were collected using a standard proforma. Candidate variants detected by NGS were confirmed by Sanger sequencing/Multiplex Ligation-dependent Probe Amplification with subsequent family segregation analysis where possible.

Results: Clinical presentation was nephrotic syndrome in 267 patients and suspected Alport syndrome (AS) in 35. NGS panel testing determined a likely genetic cause of disease in 44/220 (20.0%) paediatric and 10/47 (21.3%) adult nephrotic cases, and 17/35 (48.6%) of haematuria/AS patients. Of 71 patients with genetic disease, 32 had novel pathogenic variants without a previous disease association including two with deletions of one or more exons of or .

Conclusion: Gene panel testing provides a genetic diagnosis in a significant number of patients presenting with SRNS or suspected AS. It should be undertaken at an early stage of the care pathway and include the ability to detect CNVs as an emerging mechanism for genes associated with this condition. Use of clinical genetic testing after diagnosis of SRNS has the potential to stratify patients and assist decision-making regarding management.
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http://dx.doi.org/10.1136/jmedgenet-2017-104811DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5740557PMC
December 2017

Modeling Hybridization Kinetics of Gene Probes in a DNA Biochip Using FEMLAB.

Microarrays (Basel) 2017 May 29;6(2). Epub 2017 May 29.

Department of Civil and Environmental Engineering, Michigan State University, East Lansing, MI 48823,USA.

Microfluidic DNA biochips capable of detecting specific DNA sequences are useful in medical diagnostics, drug discovery, food safety monitoring and agriculture. They are used as miniaturized platforms for analysis of nucleic acids-based biomarkers. Binding kinetics between immobilized single stranded DNA on the surface and its complementary strand present in the sample are of interest. To achieve optimal sensitivity with minimum sample size and rapid hybridization, ability to predict the kinetics of hybridization based on the thermodynamic characteristics of the probe is crucial. In this study, a computer aided numerical model for the design and optimization of a flow-through biochip was developed using a finite element technique packaged software tool (FEMLAB; package included in COMSOL Multiphysics) to simulate the transport of DNA through a microfluidic chamber to the reaction surface. The model accounts for fluid flow, convection and diffusion in the channel and on the reaction surface. Concentration, association rate constant, dissociation rate constant, recirculation flow rate, and temperature were key parameters affecting the rate of hybridization. The model predicted the kinetic profile and signal intensities of eighteen 20-mer probes targeting vancomycin resistance genes (VRGs). Predicted signal intensities and hybridization kinetics strongly correlated with experimental data in the biochip (R² = 0.8131).
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http://dx.doi.org/10.3390/microarrays6020009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5487956PMC
May 2017

Respiratory involvement in ambulant and non-ambulant patients with facioscapulohumeral muscular dystrophy.

J Neurol 2017 Jun 26;264(6):1271-1280. Epub 2017 May 26.

John Walton Muscular Dystrophy Research Centre, MRC Centre for Neuromuscular Diseases, Institute of Genetic Medicine, Newcastle University, Central Parkway, Newcastle upon Tyne, NE1 3BZ, UK.

Understand the occurrence and predictors of respiratory impairment in FSHD. Data from 100 FSHD patients was collected regarding demographics, genetics, respiratory status and pulmonary function tests, clinical manifestations and Clinical Severity Scale (CSS) scores. Patients were assigned to two severity groups using CSS: mild (scores <3.5) and moderate/severely affected (scores ≥3.5). Forced Vital Capacity (FVC) was classified as severely impaired if less than 50% of the predicted. Statistical analysis was performed using IBM SPSS Statistics 23, tests were two-tailed and the level of significance set at 5%. Spirometry was available for 94 patients; 41.5% had abnormal results with a restrictive pattern in 38.3% patients. There was a correlation between FVC; CSS score and D4Z4 fragment length with a higher probability of severe respiratory involvement in the early onset group, moderate/severe disease and D4Z4 fragments <18 kb. Patients with severe respiratory involvement showed a high prevalence of sleep-disordered breathing. FVC decline over time was indicative of three progression groups. Respiratory involvement for both ambulant and non-ambulant patients with FSHD is more frequent and severe than previously suggested. Sleep-disordered breathing is frequent and negatively influences the respiratory status. Annual screening of the respiratory status with spirometry and clinical assessment is thus warranted in FSHD patients, even while ambulant.
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http://dx.doi.org/10.1007/s00415-017-8525-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5486574PMC
June 2017

Quantification of microRNAs directly from body fluids using a base-stacking isothermal amplification method in a point-of-care device.

Biomed Microdevices 2017 Sep;19(3):45

Department of Civil and Environmental Engineering, Michigan State University, East Lansing, MI, 48824, USA.

MicroRNAs have been proposed to be a class of biomarkers of disease as expression levels are significantly altered in various tissues and body fluids when compared to healthy controls. As such, the detection and quantification of microRNAs is imperative. While many methods have been established for quantification of microRNAs, they typically rely on time consuming handling such as RNA extraction, purification, or ligation. Here we describe a novel method for quantification of microRNAs using direct amplification in body fluids without upstream sample preparation. Tested with a point-of-care device (termed Gene-Z), the presence of microRNA promotes base-stacking hybridization, and subsequent amplification between two universal strands. The base-stacking approach, which was achieved in <60 min, provided a sensitivity of 1.4 fmol per reaction. Tested in various percentages of whole blood, plasma, and faeces, precision (coefficient of variation = 2.6%) was maintained and comparable to amplification in pristine samples. Overall, the developed method represents a significant step towards rapid, one-step detection of microRNAs.
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http://dx.doi.org/10.1007/s10544-017-0191-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5896010PMC
September 2017

On-filter direct amplification of Legionella pneumophila for rapid assessment of its abundance and viability.

Water Res 2017 09 13;121:162-170. Epub 2017 May 13.

Department of Civil and Environmental Engineering, Michigan State University, East Lansing, MI, USA; Center for Microbial Ecology, Michigan State University, East Lansing, MI, USA. Electronic address:

Guidelines and regulations to control Legionella pneumophila in cooling water systems of large buildings are evolving due to the increasing number of outbreaks. Rapid, on-site, simple, and sensitive quantification methods that are also able to assess viability may be extremely useful in monitoring and control. Culture-based methods for measuring L. pneumophila may take 4-10 days and qPCR-based methods are also slow, requiring at least a day from sample to result, albeit mainly due to the need for sample transport to a centralized laboratory. This study reports a rapid isothermal amplification method for L. pneumophila concentration and detection with live/dead differentiation under field conditions. Using an on-filter direct amplification (i.e., amplification of cells without DNA extraction and purification) approach with propidium monoazide (PMA), and a real time isothermal amplification platform (Gene-Z), L. pneumophila could be detected in 1-2 h at ∼1 cfu/100 ml of tap water. Signature sequences from 16S rRNA and cadA genes were used as genetic markers for L. pneumophila and loop-mediated isothermal amplification (LAMP) primers were designed using Primer Explorer V4. Result were also compared with direct amplification of cells spiked into distilled, tap, and cooling water samples as well as extracted DNA by qPCR. This method may be useful to managers of cooling water systems in large buildings for rapid detection of L. pneumophila. The overall approach of on-site sample concentration, on-filter amplification, and live/dead differentiation may be extended to other organisms where analytical sensitivity and speed are equally important.
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http://dx.doi.org/10.1016/j.watres.2017.05.028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5525548PMC
September 2017