Publications by authors named "Andrew Collins"

410 Publications

Establishing Stable Binary Cultures of Symbiotic Saccharibacteria from the Oral Cavity.

J Vis Exp 2021 Apr 13(170). Epub 2021 Apr 13.

The Forsyth Institute; Harvard School of Dental Medicine;

Many bacterial species cannot be cultured in the laboratory using standard methods, posing a significant barrier to studying the majority of microbial diversity on earth. Novel approaches are required to culture these uncultured bacteria so that investigators can effectively study their physiology and lifestyle using the powerful tools available in the laboratory. The Candidate Phyla Radiation (CPR) is one of the largest groups of uncultivated bacteria, comprising ~15% of the living diversity on earth. The first isolate of this group was a member of the Saccharibacteria phylum, 'Nanosynbacter lyticus' strain TM7x. TM7x is an unusually small bacterium that lives as a symbiont in direct contact with a bacterial host, Schaalia odontolytica, strain XH001. Taking advantage of the unusually small cell size and its lifestyle as a symbiotic organism, we developed a protocol to rapidly culture Saccharibacteria from dental plaque. This protocol will show how to filter a suspension of dental plaque through a 0.2 µm filter, then concentrate the collected Saccharibacteria cells and infect a culture of host organisms. The resulting coculture can be passaged as any normal bacterial culture and infection is confirmed by PCR. The resulting binary culture can be maintained in the laboratory and used for future experiments. While contamination is a possibility, the binary culture can be purified by either further filtering and reinfection of host, or by plating the binary culture and screening for infected colonies. We hope this protocol can be expanded to other sample types and environments, leading to the cultivation of many more species in the CPR.
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http://dx.doi.org/10.3791/62484DOI Listing
April 2021

Collection and storage of human white blood cells for analysis of DNA damage and repair activity using the comet assay in molecular epidemiology studies.

Mutagenesis 2021 Mar 23. Epub 2021 Mar 23.

Department of Pharmacology and Toxicology, University of Navarra, C/Irunlarrea 1, 31008 P#38lona, Spain.

DNA damage and repair activity are often assessed in blood s#38les from humans in different types of molecular epidemiology studies. However, it is not always feasible to analyse the s#38les on the day of collection without any type of storage. For instance, certain studies use repeated s#38ling of cells from the same subject or s#38les from different subjects collected at different time-points, and it is desirable to analyse all these s#38les in the same comet assay experiment. In addition, flawless comet assay analyses on frozen s#38les opens up for the possibility of using this technique on biobank material. In this article we discuss the use of cryopreserved peripheral blood mononuclear cells (PBMCs), buffy coat (BC) and whole blood (WB) for analysis of DNA damage and repair using the comet assay. The published literature and the authors' experiences indicate that various types of blood s#38les can be cryopreserved with only minor effect on the basal level of DNA damage. There is evidence to suggest that WB and PBMCs can be cryopreserved for several years without much effect on the level of DNA damage. However, care should be taken when cryopreserving WB and BCs. It is possible to use either fresh or frozen s#38les of blood cells, but results from fresh and frozen cells should not be used in the same dataset. The article outlines detailed protocols for the cryopreservation of PBMCs, BCs and WB s#38les.
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http://dx.doi.org/10.1093/mutage/geab012DOI Listing
March 2021

Clinical and Descriptive Study of Orofacial Clefts in Colombia: 2069 Patients From Operation Smile Foundation.

Cleft Palate Craniofac J 2021 Mar 19:10556656211000551. Epub 2021 Mar 19.

Human Genetics Group, 27989Universidad de La Sabana, Chía, Cundinamarca, Colombia.

Objective: To describe the population of patients with cleft lip and/or palate (CL/P) in terms of cleft phenotypes, gender, age, ethnic group, family history, clinical presentation (syndromic vs nonsyndromic), some environmental and behavioral factors, and some clinical features.

Design: Descriptive retrospective study.

Setting: Patients attending the genetics counseling practice in Operation Smile Foundation, Bogotá, Colombia, for over 8 years.

Participants: No screening was conducted. All patients requiring clinical genetics assessment in Operation Smile Foundation were included in the study.

Results: Left cleft lip and palate (CLP) and nonsyndromic forms were the most frequent types of malformations in this population. Psychomotor retardation and heart disease were the most frequent comorbidities in these patients. A low proportion of mothers exposed to passive smoking during pregnancy was observed and low birth weight accounted for an important number of cases. Aarskog, velocardiofacial, and orofaciodigital syndromes were the most frequent syndromic forms of CLP in this population.

Conclusions: In this study, the most frequent type of CL/P was the nonsyndromic complete left CLP. Aarskog, velocardiofacial, and orofaciodigital syndromes were the most frequent syndromic forms of CL/P in this population.
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http://dx.doi.org/10.1177/10556656211000551DOI Listing
March 2021

A snapshot of optometry teaching in Australia and New Zealand in response to COVID-19.

Clin Exp Optom 2021 Mar 1:1-5. Epub 2021 Mar 1.

Discipline of Optometry, Faculty of Health, The University of Canberra, Canberra, Australia.

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http://dx.doi.org/10.1080/08164622.2021.1878859DOI Listing
March 2021

The micronucleus cytome assay - A fast tool for DNA damage screening in human conjunctival epithelial cells.

Ocul Surf 2021 Apr 4;20:195-198. Epub 2021 Mar 4.

Laboratory of the Biology and Pathology of the Eye, Institute of Biology and Medical Genetics, First Faculty of Medicine, Charles University and General University Hospital in Prague, Czech Republic.

Purpose: To assess whether the micronucleus cytome assay (MCyt) reliably detects DNA damage occurring in control and pathological superficial epithelial cells from human conjunctiva.

Methods: Impression cytology samples from the bulbar conjunctiva of 33 healthy controls, eight patients with conjunctival intraepithelial neoplasia (CIN) and eight with mucous membrane pemphigoid (MMP) were examined using the MCyt modified for the ocular surface.

Results: The mean number of micronuclei (MNi) in control samples was 0.94 MNi/1000 epithelial cells, with no significant difference between conjunctival quadrants and independent of sex and age. The MCyt assay applied to CIN-affected eyes showed a significantly higher frequency of MNi (18.63/1000 cells), apoptotic cells, nuclear enlargement, multinucleated cells, and keratolysis compared with the corresponding unaffected paired eyes and with the control value. Although the mean MNi frequency in MMP eyes was also higher (1.73 MNi/1000 cells), it did not prove to be statistically different from the control samples. On the other hand, the MMP-affected eyes revealed significantly elevated percentages of cells with snake-like chromatin, multinucleated cells, apoptotic cells, and nuclear buds compared with controls.

Conclusions: Micronucleus cytome assay was adapted as a rapid screening test for genomic instability on the ocular surface. We have determined reference levels for MNi and other nuclear alterations on healthy conjunctiva and demonstrated that particularly frequencies of MNi are significantly elevated in conjunctiva affected by CIN. We demonstrate that MNi are more specific than other nuclear abnormalities and thus can be used for screening of ocular surface neoplasia.
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http://dx.doi.org/10.1016/j.jtos.2021.02.011DOI Listing
April 2021

Application of Next Generation Quality/Statistical Process Control and Expert-Led Case Review to Increase the Consistency of Diagnostic Rates in Precancerous Colorectal Polyps.

Qual Manag Health Care 2021 Jan 5;Publish Ahead of Print. Epub 2021 Jan 5.

Departments of Pathology and Molecular Medicine (Drs Bonert, Dmetrichuk, Al-Haddad, and Naqvi), Medicine (Dr Collins), and Oncology (Dr Major), McMaster University, Hamilton, Ontario, Canada; St. Joseph's Healthcare Hamilton, Hamilton, Ontario, Canada (Drs Bonert and Naqvi); Farncombe Family Digestive Health Research Institute/Division of Gastroenterology, Department of Medicine, McMaster University, Hamilton, Ontario, Canada (Dr Xenodemetropoulos); and Juravinski Hospital/Hamilton Health Sciences Centre/Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada (Ms Al-Haddad and Dr Major).

Background: Prior work suggests high interrater variability in the pathologist diagnostic rate (PDR) of the precancerous polyp sessile serrated adenoma (SSA).

Objectives: To improve the diagnostic consistency in the pathological evaluation of colorectal polyp specimens with diagnostic rate awareness, using funnel plots (FPs)/control charts (CCs), and a focused group case review.

Methods: All colorectal polyp specimen (CRPS) reports September 2015 to August 2017 were analyzed at one institution. PDRs were extracted using a hierarchical free-text string matching algorithm and visualized using FPs, showing pathologist specimen volume versus PDR, and CCs, showing pathologist versus normed PDR. The FPs/CCs were centered on the group median diagnostic rate (GMDR). Pathologists were shown their baseline SSA diagnostic rate in relation to the practice, and in January 2017, there was a focused group case review/open discussion of approximately 40 sequential cases signed as SSA with a gastrointestinal pathology expert.

Results: Nine pathologists interpreted more than 250 CRPSs per year. FPs/CCs for the first and second years showed 6/4 and 3/1 P < .05/P < .001 pathologist outliers, respectively, in relation to the GMDR for SSA and 0/0 and 0/0 P < .05/P < .001 pathologist outliers, respectively, in relation to the GMDR for tubular adenoma (TA). An in silico kappa (ISK) for SSA improved from 0.52 to 0.62.

Conclusion: Diagnostic rate awareness facilitated by FPs/CCs coupled with focused expert-led reviews may help calibrate PDR. Variation in SSA PDRs still remains high in relation to TA. ISK represents an intuitive, useful metric and Next Generation Quality/Statistical Process Control a promising approach for objectively increasing diagnostic consistency.
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http://dx.doi.org/10.1097/QMH.0000000000000299DOI Listing
January 2021

Editorial: Nomenclature - Avoiding Babylonian Speech Confusion in Present Day Immunology.

Front Immunol 2020 14;11:621100. Epub 2020 Dec 14.

Immunology Unit, Department of Biomedical Sciences, Medical School of the University of Barcelona, Barcelona, Spain.

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http://dx.doi.org/10.3389/fimmu.2020.621100DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7767845PMC
December 2020

The enzyme-modified comet assay: Past, present and future.

Food Chem Toxicol 2021 Jan 18;147:111865. Epub 2020 Nov 18.

Universidad de Navarra, Department of Pharmacology and Toxicology, Faculty of Pharmacy and Nutrition, Irunlarrea 1, 310008, Pamplona, Spain; IdiSNA, Navarra Institute for Health Research, Pamplona, Spain. Electronic address:

The enzyme-modified comet assay was developed in order to detect DNA lesions other than those detected by the standard version (single and double strand breaks and alkali-labile sites). Various lesion-specific enzymes, from the DNA repair machinery of bacteria and humans, have been combined with the comet assay, allowing detection of different oxidized and alkylated bases as well as cyclobutane pyrimidine dimers, mis-incorporated uracil and apurinic/apyrimidinic sites. The enzyme-modified comet assay has been applied in different fields - human biomonitoring, environmental toxicology, and genotoxicity testing (both in vitro and in vivo) - as well as in basic research. Up to now, twelve enzymes have been employed; here we describe the enzymes and give examples of studies in which they have been applied. The bacterial formamidopyrimidine DNA glycosylase (Fpg) and endonuclease III (EndoIII) have been extensively used while others have been used only rarely. Adding further enzymes to the comet assay toolbox could potentially increase the variety of DNA lesions that can be detected. The enzyme-modified comet assay can play a crucial role in the elucidation of the mechanism of action of both direct and indirect genotoxins, thus increasing the value of the assay in the regulatory context.
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http://dx.doi.org/10.1016/j.fct.2020.111865DOI Listing
January 2021

Isolation and cultivation of candidate phyla radiation (TM7) bacteria in coculture with bacterial hosts.

J Oral Microbiol 2020 Sep 6;12(1):1814666. Epub 2020 Sep 6.

Department of Microbiology, The Forsyth Institute, Cambridge, MA, USA.

Background: The vast majority of bacteria on earth have not yet been cultivated. There are many bacterial phyla with no cultivated examples including most members of the Candidate Phylum Radiation with the exception of human oral isolates from the phylum Saccharibacteria.

Aims: The aims of this research were to develop reproducible methods and validate approaches for the cultivation of human oral Saccharibacteria and to identify the conceptual pitfalls that delayed isolation of these bacteria for 20 years after their discovery.

Methods: Oral samples were dispersed and passed through 0.2 µm membrane filters. The ultrasmall saccharibacterial cells in the filtrate were pelleted, inoculated into broth cultures of potential bacterial host cells and passaged into fresh medium every 2-3 days.

Results: Thirty-two isolates representing four species of Saccharibacteria were isolated in stable coculture with three species of host bacteria from the phylum . Complete genome sequences were obtained for 16 isolates.

Conclusions: Human oral Saccharibacteria are obligate bacterial parasites that can be stably passaged in coculture with specific species of host bacteria. Isolating these important members of the human oral microbiome, and many natural environments, requires abandoning many of Koch's concepts and methods and embracing novel microbiological approaches.
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http://dx.doi.org/10.1080/20002297.2020.1814666DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7651992PMC
September 2020

An optimized comet-based in vitro DNA repair assay to assess base and nucleotide excision repair activity.

Nat Protoc 2020 12 16;15(12):3844-3878. Epub 2020 Nov 16.

Department of Pharmacology & Toxicology, School for Nutrition and Translational Research in Metabolism (NUTRIM), Maastricht University, Maastricht, the Netherlands.

This optimized protocol (including links to instruction videos) describes a comet-based in vitro DNA repair assay that is relatively simple, versatile, and inexpensive, enabling the detection of base and nucleotide excision repair activity. Protein extracts from samples are incubated with agarose-embedded substrate nucleoids ('naked' supercoiled DNA) containing specifically induced DNA lesions (e.g., resulting from oxidation, UVC radiation or benzo[a]pyrene-diol epoxide treatment). DNA incisions produced during the incubation reaction are quantified as strand breaks after electrophoresis, reflecting the extract's incision activity. The method has been applied in cell culture model systems, human biomonitoring and clinical investigations, and animal studies, using isolated blood cells and various solid tissues. Once extracts and substrates are prepared, the assay can be completed within 2 d.
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http://dx.doi.org/10.1038/s41596-020-0401-xDOI Listing
December 2020

Pediatric intrathecal baclofen management during the COVID-19 pandemic in the US and Canada.

J Pediatr Rehabil Med 2020;13(3):379-384

Physical Medicine and Rehabilitation, University of Miami Miller School of Medicine, Miami, FL, USA.

The COVID-19 pandemic has been a challenge to healthcare systems around the world. Within pediatric rehabilitation medicine, management of intrathecal baclofen has been particularly challenging. This editorial reviews how programs in the US and Canada coped with the quickly changing healthcare environment and how we can learn from this pandemic to be prepared for future crises.
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http://dx.doi.org/10.3233/PRM-200755DOI Listing
December 2020

Minimum Information for Reporting on the Comet Assay (MIRCA): recommendations for describing comet assay procedures and results.

Nat Protoc 2020 12 26;15(12):3817-3826. Epub 2020 Oct 26.

Department of Nutrition, University of Oslo, Oslo, Norway.

The comet assay is a widely used test for the detection of DNA damage and repair activity. However, there are interlaboratory differences in reported levels of baseline and induced damage in the same experimental systems. These differences may be attributed to protocol differences, although it is difficult to identify the relevant conditions because detailed comet assay procedures are not always published. Here, we present a Consensus Statement for the Minimum Information for Reporting Comet Assay (MIRCA) providing recommendations for describing comet assay conditions and results. These recommendations differentiate between 'desirable' and 'essential' information: 'essential' information refers to the precise details that are necessary to assess the quality of the experimental work, whereas 'desirable' information relates to technical issues that might be encountered when repeating the experiments. Adherence to MIRCA recommendations should ensure that comet assay results can be easily interpreted and independently verified by other researchers.
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http://dx.doi.org/10.1038/s41596-020-0398-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7688437PMC
December 2020

The repeatability and reproducibility of four techniques for measuring horizontal heterophoria: Implications for clinical practice.

J Optom 2020 Aug 11. Epub 2020 Aug 11.

School of Optometry and Vision Science, The University of Auckland, New Zealand.

Purpose: Convergence insufficiency, the most common binocular vision anomaly, is characterised by a receded near point of convergence and an exophoria which is at least 4 prism dioptres (Δ) larger at near than at distance. However, the repeatability of standard heterophoria measures are poorly understood. This study assessed the ability of four common heterophoria tests to detect differences of 4Δ by evaluating the inter- and intra-examiner variability of the selected techniques.

Methods: Distance and near horizontal heterophorias of 20 visually-normal adults were measured with the alternating prism cover test, von Graefe prism dissociation, Howell Card and Maddox Rod by two examiners at two separate visits using standardised instructions and techniques. We investigated inter- and intra-examiner variability using repeatability and reproducibility indices, as well as Bland-Altman analysis with acceptable limits of agreement defined as ±2Δ.

Results: The Howell card test had the lowest intra-examiner variability at both distance and near, as well as the best 95% limits of agreement (±1.6Δ for distance and ±3.7Δ for near). Inter-examiner reproducibility results were similar, although at near the alternating prism cover test had better repeatability (1.1Δ, 95% confidence intervals -1.1Δ to 4.0Δ) than the Howell card (1.4Δ, 95% confidence intervals -1.9Δ to 5.9Δ).

Conclusion: The low repeatability of many standard clinical heterophoria tests limits the ability to reliably detect a 4Δ difference. The Howell Card provided the most repeatable and reproducible results indicating that this technique should be used to detect small changes in heterophoria magnitude and direction.
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http://dx.doi.org/10.1016/j.optom.2020.05.005DOI Listing
August 2020

Medically Necessary Orthopaedic Surgery During the COVID-19 Pandemic: Safe Surgical Practices and a Classification to Guide Treatment.

J Bone Joint Surg Am 2020 Jul;102(14):e76

Department of Orthopaedics, Rothman Orthopaedics-New York, New York, NY.

Background: Coronavirus disease 2019 (COVID-19) has rapidly evolved as a viral pandemic. Countries worldwide have been affected by the recent outbreak caused by the SARS (severe acute respiratory syndrome)-CoV-2 virus. As with prior viral pandemics, health-care workers are at increased risk. Orthopaedic surgical procedures are common in health-care systems, ranging from emergency to elective procedures. Many orthopaedic surgical procedures are life or limb-saving and cannot be postponed during the COVID-19 pandemic because of potential patient harm. Our goal is to analyze how orthopaedic surgeons can perform medically necessary procedures during the pandemic and to help guide decision-making perioperatively.

Methods: We performed a review of the existing literature regarding COVID-19 and prior viral outbreaks to help guide clinical practice in terms of how to safely perform medically necessary orthopaedic procedures during the pandemic for both asymptomatic patients and high-risk (e.g., COVID-19-positive) patients. We created a classification system based on COVID-19 positivity, patient health status, and COVID-19 prevalence to help guide perioperative decision-making.

Results: We advocate that only urgent and emergency surgical procedures be performed. By following recommendations from the American College of Surgeons, the Centers for Disease Control and Prevention, and the recent literature, safe orthopaedic surgery and perioperative care can be performed. Screening measures are needed for patients and perioperative teams. Surgeons and perioperative teams at risk for contracting COVID-19 should use appropriate personal protective equipment (PPE), including N95 respirators or powered air-purifying respirators (PAPRs), when risk of viral spread is high. When preparing for medically necessary orthopaedic procedures during the pandemic, our classification system will help to guide decision-making. A multidisciplinary care plan is needed to ensure patient safety with medically necessary orthopaedic procedures during the COVID-19 pandemic.

Conclusions: Orthopaedic surgery during the COVID-19 pandemic can be performed safely when medically necessary but should be rare for COVID-19-positive or high-risk patients. Appropriate screening, PPE use, and multidisciplinary care will allow for safe medically necessary orthopaedic surgery to continue during the COVID-19 pandemic.

Level Of Evidence: Prognostic Level V. See Instructions for Authors for a complete description of levels of evidence.
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http://dx.doi.org/10.2106/JBJS.20.00599DOI Listing
July 2020

Guidance for publishing comet assay results.

Mutat Res 2020 Jun - Jul;854-855:503146. Epub 2020 Jan 23.

Department of Nutrition, University of Oslo, Norway. Electronic address:

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http://dx.doi.org/10.1016/j.mrgentox.2020.503146DOI Listing
October 2020

A Retrospective Thematic Analysis of Patient, Family, and Staff Creative Art Tiles Developed on a Tertiary Palliative Care Unit.

Am J Hosp Palliat Care 2021 Mar 29;38(3):266-275. Epub 2020 Jun 29.

Division of Palliative Care Medicine, Department of Oncology, University of Alberta, Edmonton, Canada.

Objectives: Clinical use of the creative arts in palliative care is well established, yet there are few evaluation studies of these programs.

Methods: In this first phase of a 3-phase evaluation of a creative arts program entitled "Tile Tales," we conducted a retrospective thematic analysis of 85 painted tiles and accompanying stories that were publically displayed on a tertiary palliative care unit. Each story was independently coded, using content analysis. Themes were derived through consensus, using the constant comparative method.

Results: Tiles were created by staff (n = 36, 42%), family (n = 32, 38%), patients (n = 9, 11%), or patients and family (n = 8, 9%). Six major themes emerged from the artwork: "Spirituality," "Relationships," "Journey," "Story," "Symbolism," and "Paradox."

Significance Of Results: These results illustrate how the creative arts can support the expression of diverse palliative care experiences, for patients, their families and palliative care staff, when words alone may not suffice.
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http://dx.doi.org/10.1177/1049909120935833DOI Listing
March 2021

The role of pain rehabilitation in an adolescent sickle cell disease population: a case series.

Pediatr Hematol Oncol 2020 Oct 13;37(7):637-644. Epub 2020 May 13.

Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA.

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http://dx.doi.org/10.1080/08880018.2020.1762811DOI Listing
October 2020

Potassium bromate as positive assay control for the Fpg-modified comet assay.

Mutagenesis 2020 09;35(4):341-348

Department of Nutrition, University of Oslo, Sognsvannsveien 9, 0372, Oslo, Norway.

The comet assay is a popular assay in biomonitoring studies. DNA strand breaks (or unspecific DNA lesions) are measured using the standard comet assay. Oxidative stress-generated DNA lesions can be measured by employing DNA repair enzymes to recognise oxidatively damaged DNA. Unfortunately, there has been a tendency to fail to report results from assay controls (or maybe even not to employ assay controls). We believe this might have been due to uncertainty as to what really constitutes a positive control. It should go without saying that a biomonitoring study cannot have a positive control group as it is unethical to expose healthy humans to DNA damaging (and thus potentially carcinogenic) agents. However, it is possible to include assay controls in the analysis (here meant as a cryopreserved sample of cells i.e. included in each experiment as a reference sample). In the present report we tested potassium bromate (KBrO3) as a positive comet assay control for the formamidopyrimidine DNA glycosylase (Fpg)-modified comet assay. Ten laboratories used the same procedure for treatment of monocytic THP-1 cells with KBrO3 (0.5, 1.5 and 4.5 mM for 1 h at 37°C) and subsequent cryopreservation. Results from one laboratory were excluded in the statistical analysis because of technical issues in the Fpg-modified comet assay. All other laboratories found a concentration-response relationship in cryopreserved samples (regression coefficients from 0.80 to 0.98), although with different slopes ranging from 1.25 to 11.9 Fpg-sensitive sites (%DNA in tail) per 1 mM KBrO3. Our results demonstrate that KBrO3 is a suitable positive comet assay control.
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http://dx.doi.org/10.1093/mutage/geaa011DOI Listing
September 2020

The role of the enzyme-modified comet assay in in vivo studies.

Toxicol Lett 2020 Jul 2;327:58-68. Epub 2020 Apr 2.

Department of Pharmacology and Toxicology, University of Navarra, C/Irunlarrea 1, 31009, Pamplona, Spain. Electronic address:

The in vivo comet assay is an established genotoxicity test, with an OECD test guideline, but in its standard form it measures only DNA strand breaks. Including in the assay an additional step, in which the DNA is incubated with a lesion-specific enzyme, can provide important information about the nature of the DNA damage. Formamidopyrimidine DNA glycosylase, 8-oxoguanine DNA glycosylase or endonuclease III are commonly used in the in vitro genotoxicity test and in human biomonitoring to detect oxidised bases, but in vivo applications are rarer. A systematic literature search has identified a total of 60 papers that report such in vivo experiments, testing a variety of agents. In many cases, strand breaks were not seen, but significant levels of enzyme-sensitive sites were induced - indicating a mechanism of action involving oxidative stress. Compounds such as methyl methanesulfonate (MMS) or ethyl methanesulfonate (EMS) could be used as positive controls in both the standard and the enzyme-modified in vivo comet assays.
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http://dx.doi.org/10.1016/j.toxlet.2020.03.016DOI Listing
July 2020

Application of the comet assay in human biomonitoring: An hCOMET perspective.

Mutat Res 2020 Jan - Mar;783:108288. Epub 2019 Nov 9.

Department of Public Health, Section of Environmental Health, University of Copenhagen, DK-1014 Copenhagen K, Denmark.

The comet assay is a well-accepted biomonitoring tool to examine the effect of dietary, lifestyle, environmental and occupational exposure on levels of DNA damage in human cells. With such a wide range of determinants for DNA damage levels, it becomes challenging to deal with confounding and certain factors are inter-related (e.g. poor nutritional intake may correlate with smoking status). This review describes the effect of intrinsic (i.e. sex, age, tobacco smoking, occupational exposure and obesity) and extrinsic (season, environmental exposures, diet, physical activity and alcohol consumption) factors on the level of DNA damage measured by the standard or enzyme-modified comet assay. Although each factor influences at least one comet assay endpoint, the collective evidence does not indicate single factors have a large impact. Thus, controlling for confounding may be necessary in a biomonitoring study, but none of the factors is strong enough to be regarded a priori as a confounder. Controlling for confounding in the comet assay requires a case-by-case approach. Inter-laboratory variation in levels of DNA damage and to some extent also reproducibility in biomonitoring studies are issues that have haunted the users of the comet assay for years. Procedures to collect specimens, and their storage, are not standardized. Likewise, statistical issues related to both sample-size calculation (before sampling of specimens) and statistical analysis of the results vary between studies. This review gives guidance to statistical analysis of the typically complex exposure, co-variate, and effect relationships in human biomonitoring studies.
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http://dx.doi.org/10.1016/j.mrrev.2019.108288DOI Listing
July 2020

Essentiality-specific pathogenicity prioritization gene score to improve filtering of disease sequence data.

Brief Bioinform 2021 Mar;22(2):1782-1789

The causal genetic variants underlying more than 50% of single gene (monogenic) disorders are yet to be discovered. Many patients with conditions likely to have a monogenic basis do not receive a confirmed molecular diagnosis which has potential impacts on clinical management. We have developed a gene-specific score, essentiality-specific pathogenicity prioritization (ESPP), to guide the recognition of genes likely to underlie monogenic disease variation to assist in filtering of genome sequence data. When a patient genome is sequenced, there are frequently several plausibly pathogenic variants identified in different genes. Recognition of the single gene most likely to include pathogenic variation can guide the identification of a causal variant. The ESPP score integrates gene-level scores which are broadly related to gene essentiality. Previous work towards the recognition of monogenic disease genes proposed a model with increasing gene essentiality from 'non-essential' to 'essential' genes (for which pathogenic variation may be incompatible with survival) with genes liable to contain disease variation positioned between these two extremes. We demonstrate that the ESPP score is useful for recognizing genes with high potential for pathogenic disease-related variation. Genes classed as essential have particularly high scores, as do genes recently recognized as strong candidates for developmental disorders. Through the integration of individual gene-specific scores, which have different properties and assumptions, we demonstrate the utility of an essentiality-based gene score to improve sequence genome filtering.
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http://dx.doi.org/10.1093/bib/bbaa029DOI Listing
March 2021

A Proposed New Nomenclature for the Immunoglobulin Genes of .

Front Immunol 2019 18;10:2961. Epub 2019 Dec 18.

School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, NSW, Australia.

Mammalian immunoglobulin (IG) genes are found in complex loci that contain hundreds of highly similar pseudogenes, functional genes and repetitive elements, which has made their investigation particularly challenging. High-throughput sequencing has provided new avenues for the investigation of these loci, and has recently been applied to study the IG genes of important inbred mouse strains, revealing unexpected differences between their IG loci. This demonstrated that the structural differences are of such magnitude that they call into question the merits of the current mouse IG gene nomenclatures. Three nomenclatures for the mouse IG heavy chain locus () are presently in use, and they are all positional nomenclatures using the C57BL/6 genome reference sequence as their template. The continued use of these nomenclatures requires that genes of other inbred strains be confidently identified as allelic variants of C57BL/6 genes, but this is clearly impossible. The unusual breeding histories of inbred mouse strains mean that, regardless of the genetics of wild mice, no single ancestral origin for the IG loci exists for laboratory mice. Here we present a general discussion of the challenges this presents for any IG nomenclature. Furthermore, we describe principles that could be followed in the formulation of a solution to these challenges. Finally, we propose a non-positional nomenclature that accords with the guidelines of the International Mouse Nomenclature Committee, and outline strategies that can be adopted to meet the nomenclature challenges if three systems are to give way to a new one.
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http://dx.doi.org/10.3389/fimmu.2019.02961DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6930147PMC
November 2020

The Impact of Spatial Distribution of Pain on Long-Term Trajectories for Chronic Pain Outcomes After Intensive Interdisciplinary Pain Treatment.

Clin J Pain 2020 03;36(3):181-188

Departments of Pediatrics.

Objectives: Evidence for the effectiveness of intensive interdisciplinary pain treatment (IIPT) for pediatric chronic pain is growing; however, little research has considered factors that contribute to differences in patients' treatment response. The present study utilized multilevel modeling to examine trajectory of change over time in functional disability from clinic assessment to 6-month follow-up in pediatric patients participating in IIPT, considering spatial distribution of pain, coping efficacy, and pain intensity.

Materials And Methods: Participants included 110 pediatric patients (Mage=14.65, SD=2.37; localized pain, 27.3%; widespread pain, 72.7%) consecutively admitted into IIPT. Patients completed self-report measures of functional disability, pain intensity, and coping efficacy at pretreatment assessment, admission, weekly during IIPT, discharge, and several follow-ups.

Results: Analysis identified a model with 3 three-way interactions, including time, time squared, pain intensity, coping efficacy, and pain distribution, on functional disability. The spatial distribution of pain had the greatest impact on treatment trajectory in patients with widespread pain, high pain intensity, and poor coping efficacy; these patients demonstrated greater functional disability compared with patients with localized pain. In addition, patients with widespread pain and poor coping efficacy had the most functional disability across all levels of pain intensity. Patients with localized pain and poor coping efficacy demonstrated initial treatment gains, but evidenced an increase in functional disability at follow-up.

Discussion: Overall, spatial distribution of pain, coping efficacy, and pain intensity played an important role in patients' response to treatment. Better understanding of these variables could improve treatment response, particularly for the most severely disabled chronic pain patients.
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http://dx.doi.org/10.1097/AJP.0000000000000793DOI Listing
March 2020

Linkage disequilibrium maps for European and African populations constructed from whole genome sequence data.

Sci Data 2019 10 17;6(1):208. Epub 2019 Oct 17.

Human Genetics & Genomic Medicine, Faculty of Medicine, University of Southampton, Southampton, UK.

Quantification of linkage disequilibrium (LD) patterns in the human genome is essential for genome-wide association studies, selection signature mapping and studies of recombination. Whole genome sequence (WGS) data provides optimal source data for this quantification as it is free from biases introduced by the design of array genotyping platforms. The Malécot-Morton model of LD allows the creation of a cumulative map for each choromosome, analogous to an LD form of a linkage map. Here we report LD maps generated from WGS data for a large population of European ancestry, as well as populations of Baganda, Ethiopian and Zulu ancestry. We achieve high average genetic marker densities of 2.3-4.6/kb. These maps show good agreement with prior, low resolution maps and are consistent between populations. Files are provided in BED format to allow researchers to readily utilise this resource.
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http://dx.doi.org/10.1038/s41597-019-0227-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6797713PMC
October 2019

Complete Genome Sequence of Strain PM004, a Novel Cultured Member of the Human Oral Microbiome from the Candidate Phylum (TM7).

Microbiol Resour Announc 2019 Oct 17;8(42). Epub 2019 Oct 17.

The Forsyth Institute, Cambridge, Massachusetts, USA

Strain PM004 is a cultured representative of human microbial taxon 955, a bacterium from the phylum It is an obligate parasite with a genome of <0.9 Mb and can be grown in coculture with its host, The complete genome sequence is presented here.
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http://dx.doi.org/10.1128/MRA.01159-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6797546PMC
October 2019

Complete Genome Sequence of Strain AC001, a Novel Cultured Member of the Human Oral Microbiome from the Candidate Phylum (TM7).

Microbiol Resour Announc 2019 Oct 17;8(42). Epub 2019 Oct 17.

The Forsyth Institute, Cambridge, Massachusetts, USA

Strain AC001 is a cultured representative of human microbial taxon 488, a bacterium from the candidate phylum It is an obligate parasite with a genome of <0.9 Mb and grows in coculture with its host, The complete genome sequence is presented here.
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http://dx.doi.org/10.1128/MRA.01158-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6797545PMC
October 2019

VDJbase: an adaptive immune receptor genotype and haplotype database.

Nucleic Acids Res 2020 01;48(D1):D1051-D1056

Bioengineering, Faculty of Engineering, Bar-Ilan University, Ramat Gan 5290002, Israel.

VDJbase is a publicly available database that offers easy searching of data describing the complete sets of gene sequences (genotypes and haplotypes) inferred from adaptive immune receptor repertoire sequencing datasets. VDJbase is designed to act as a resource that will allow the scientific community to explore the genetic variability of the immunoglobulin (Ig) and T cell receptor (TR) gene loci. It can also assist in the investigation of Ig- and TR-related genetic predispositions to diseases. Our database includes web-based query and online tools to assist in visualization and analysis of the genotype and haplotype data. It enables users to detect those alleles and genes that are significantly over-represented in a particular population, in terms of genotype, haplotype and gene expression. The database website can be freely accessed at https://www.vdjbase.org/, and no login is required. The data and code use creative common licenses and are freely downloadable from https://bitbucket.org/account/user/yaarilab/projects/GPHP.
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http://dx.doi.org/10.1093/nar/gkz872DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6943044PMC
January 2020

Gene-dense autosomal chromosomes show evidence for increased selection.

Heredity (Edinb) 2019 12 1;123(6):774-783. Epub 2019 Oct 1.

Genetic Epidemiology and Bioinformatics, Faculty of Medicine, University of Southampton, Duthie Building (808), Tremona Road, Southampton, SO16 6YD, UK.

Purifying selection tends to reduce nucleotide and haplotype diversity leading to increased linkage disequilibrium. However, detection of evidence for selection is difficult as the signature is confounded by wide variation in the recombination rate which has a complex relationship with selection. The effective bottleneck time (the ratio of the linkage disequilibrium map to the genetic map in Morgans) controls for variability in the recombination rate. Reduced effective bottleneck times indicate stronger residual linkage disequilibrium, consistent with increased selection. Using whole genome sequence data from one European and three Sub-Saharan African human populations we find, in the African samples, strong correlations between high gene densities and reduced effective bottleneck time for autosomal chromosomes. This suggests that gene-dense autosomes have been subject to increased purifying selection reducing effective bottleneck times compared to gene-poor autosomes. Although previous studies have shown unusually strong linkage disequilibrium for the sex chromosomes variation within the autosomes has not been recognised. The strongest relationship is between effective bottleneck time and the density of essential genes, which are likely targets of greater selective pressure (p = 0.006, for the 22 autosomes). The magnitude of the reduction in chromosome-specific effective bottleneck times from the least to the most gene-dense autosomes is ~17-21% for Sub-Saharan African populations. The effect size is greater in Sub-Saharan African populations, compared to a European sample, consistent with increased efficiency of selection in populations with larger effective population sizes which have not been subject to intense population bottlenecks as experienced by populations of European ancestry. The findings highlight the value of deeper analyses of selection within Sub-Saharan African populations.
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http://dx.doi.org/10.1038/s41437-019-0272-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6834609PMC
December 2019

OGRDB: a reference database of inferred immune receptor genes.

Nucleic Acids Res 2020 01;48(D1):D964-D970

Institute of Structural and Molecular Biology, Birkbeck College, University of London, London WC1E 7HX, UK.

High-throughput sequencing of the adaptive immune receptor repertoire (AIRR-seq) is providing unprecedented insights into the immune response to disease and into the development of immune disorders. The accurate interpretation of AIRR-seq data depends on the existence of comprehensive germline gene reference sets. Current sets are known to be incomplete and unrepresentative of the degree of polymorphism and diversity in human and animal populations. A key issue is the complexity of the genomic regions in which they lie, which, because of the presence of multiple repeats, insertions and deletions, have not proved tractable with short-read whole genome sequencing. Recently, tools and methods for inferring such gene sequences from AIRR-seq datasets have become available, and a community approach has been developed for the expert review and publication of such inferences. Here, we present OGRDB, the Open Germline Receptor Database (https://ogrdb.airr-community.org), a public resource for the submission, review and publication of previously unknown receptor germline sequences together with supporting evidence.
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http://dx.doi.org/10.1093/nar/gkz822DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6943078PMC
January 2020

Introduction to hCOMET special issue, 'Comet assay in vitro'.

Mutat Res 2019 09 3;845:403071. Epub 2019 Jul 3.

Department of Nutrition, Institute for Basic Medical Sciences, University of Oslo, Oslo, Norway.

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http://dx.doi.org/10.1016/j.mrgentox.2019.07.001DOI Listing
September 2019