NCTC3000: a century of bacterial strain collecting leads to a rich genomic data resource.

The National Collection of Type Cultures (NCTC) was founded on 1 January 1920 in order to fulfil a recognized need for a centralized repository for bacterial and fungal strains within the UK. It is among the longest-established collections of its kind anywhere in the world and today holds approximately 6000 type and reference bacterial strains - many of medical, scientific and veterinary importance - available to academic, health, food and veterinary institutions worldwide. Recently, a collaboration between NCTC, Pacific Biosciences and the Wellcome Sanger Institute established the NCTC3000 project to long-read sequence and assemble the genomes of up to 3000 NCTC strains. Here, at the beginning of the collection's second century, we introduce the resulting NCTC3000 sequence read datasets, genome assemblies and annotations as a unique, historically and scientifically relevant resource for the benefit of the international bacterial research community.

A feasibility randomised controlled trial of a Fibromyalgia Self-management Programme for adults in a community setting with a nested qualitative study (FALCON).

BACKGROUND: Fibromyalgia is a condition associated with widespread musculoskeletal pain, fatigue and sleep problems. Fibromyalgia treatment guidelines recommend non-pharmacological interventions and the development of self-management skills. An example of a programme that fits these guidelines is the Fibromyalgia Self-management Programme (FSMP) which consists of one 2.5-hour weekly session over six successive weeks and includes education about fibromyalgia, goal setting, pacing, sleep hygiene and nutritional advice. The FSMP is currently provided in a secondary care hospital setting and co-delivered by a multidisciplinary team. Delivery in a primary care setting has the potential to improve the accessibility of the programme to people with fibromyalgia. Therefore, this feasibility study aimed to determine the practicality and acceptability of conducting a future definitive randomised controlled trial of the FSMP in a community setting. METHOD: An exploratory, parallel-arm, one-to-one, randomised controlled trial. Participants were recruited from general practices across South West England, and the FSMP was co-delivered by physiotherapists and occupational therapists across two community sites. To determine the outcome measures for a future definitive trial several were tested. The Revised Fibromyalgia Impact Questionnaire, Arthritis Self-Efficacy Scale-8, Chalder Fatigue Scale, Short form 36, 5-Level EQ-5D version and Jenkins Sleep Scale were collected at baseline, 6 weeks and 6 months. Semi-structured interviews were conducted with patient participants, occupational therapists and physiotherapists to explore the acceptability and feasibility of delivering the FSMP in a community setting. RESULTS: A total of 74 participants were randomised to the FSMP intervention (n = 38) or control arm (n = 36). Attrition from the trial was 42% (31/74) at 6 months. A large proportion of those randomised to the intervention arm (34%, 13/38) failed to attend any sessions with six of the 13 withdrawing before the intervention commenced. The proportion of missing values was small for each of the outcome measures. Three overarching themes were derived from the interview data; (1) barriers and facilitators to attending the FSMP; (2) FSMP content, delivery and supporting documentation; and (3) trial processes. CONCLUSION: It is feasible to recruit people with fibromyalgia from Primary Care to participate in a randomised controlled trial testing the FSMP in a community setting. However, improvement in trial attrition and engagement with the intervention is needed. TRIAL REGISTRATION: The trial is registered with ISRCTN registry and was assigned on 29/04/2019. The registration number is ISRCTN10824225.

Deploying an In Vitro Gut Model to Assay the Impact of the Mannan-Oligosaccharide Prebiotic Bio-Mos on the Atlantic Salmon (Salmo salar) Gut Microbiome.

Alpha mannose-oligosaccharide (MOS) prebiotics are widely deployed in animal agriculture as immunomodulators as well as to enhance growth and gut health. Their mode of action is thought to be mediated through their impact on host microbial communities and their associated metabolism. Bio-Mos is a commercially available prebiotic currently used in the agri-feed industry, but studies show contrasting results of its effect on fish performance and feed efficiency. Thus, detailed studies are needed to investigate the effect of MOS supplements on the fish microbiome to enhance our understanding of the link between MOS and gut health. To assess Bio-Mos for potential use as a prebiotic growth promoter in salmonid aquaculture, we have modified an established Atlantic salmon in vitro gut model, SalmoSim, to evaluate its impact on the host microbial communities. The microbial communities obtained from ceca compartments from four adult farmed salmon were inoculated in biological triplicate reactors in SalmoSim. Prebiotic treatment was supplemented for 20 days, followed by a 6-day washout period. Inclusion of Bio-Mos in the media resulted in a significant increase in formate (P = 0.001), propionate (P = 0.037) and 3-methyl butanoic acid (P = 0.024) levels, correlated with increased abundances of several, principally, anaerobic microbial genera (Fusobacterium, Agarivorans, Pseudoalteromonas). DNA metabarcoding with the 16S rDNA marker confirmed a significant shift in microbial community composition in response to Bio-Mos supplementation with observed increase in lactic acid producing Carnobacterium. In conjunction with previous in vivo studies linking enhanced volatile fatty acid production alongside MOS supplementation to host growth and performance, our data suggest that Bio-Mos may be of value in salmonid production. Furthermore, our data highlights the potential role of in vitro gut models to complementin vivo trials of microbiome modulators. IMPORTANCE In this paper we report the results of the impact of a prebiotic (alpha-MOS supplementation) on microbial communities, using an in vitro simulator of the gut microbial environment of the Atlantic salmon. Our data suggest that Bio-Mos may be of value in salmonid production as it enhances volatile fatty acid production by the microbiota from salmon pyloric ceca and correlates with a significant shift in microbial community composition with observed increase in lactic acid producing Carnobacterium. In conjunction with previous in vivo studies linking enhanced volatile fatty acid production alongside MOS supplementation to host growth and performance, our data suggest that Bio-Mos may be of value in salmonid production. Furthermore, our data highlights the potential role of in vitro gut models to augment in vivo trials of microbiome modulators.

Optimizing Cognitive Assessment Outcome Measures for Alzheimer's Disease by Matching Wordlist Memory Test Features to Scoring Methodology.

Cognitive assessment with wordlist memory tests is a cost-effective and non-invasive method of identifying cognitive changes due to Alzheimer's disease and measuring clinical outcomes. However, with a rising need for more precise and granular measures of cognitive changes, especially in earlier or preclinical stages of Alzheimer's disease, traditional scoring methods have failed to provide adequate accuracy and information. Well-validated and widely adopted wordlist memory tests vary in many ways, including list length, number of learning trials, order of word presentation across trials, and inclusion of semantic categories, and these differences meaningfully impact cognition. While many simple scoring methods fail to account for the information that these features provide, extensive effort has been made to develop scoring methodologies, including the use of latent models that enable capture of this information for preclinical differentiation and prediction of cognitive changes. In this perspective article, we discuss prominent wordlist memory tests in use, their features, how different scoring methods fail or successfully capture the information these features provide, and recommendations for emerging cognitive models that optimally account for wordlist memory test features. Matching the use of such scoring methods to wordlist memory tests with appropriate features is key to obtaining precise measurement of subtle cognitive changes.

Genome Sequencing of a Historic Staphylococcus aureus Collection Reveals New Enterotoxin Genes and Sheds Light on the Evolution and Genomic Organization of This Key Virulence Gene Family.

We take advantage of a historic collection of 133 Staphylococcus aureus strains accessioned between 1924 and 2016, whose genomes have been long-read sequenced as part of a major National Collection of Type Cultures (NCTC) initiative, to conduct a gene family-wide computational analysis of enterotoxin genes. We identify two novel staphylococcal enterotoxin (pseudo)genes (sel29p and sel30), the former of which has not been observed in any contemporary strain to date. We provide further information on five additional enterotoxin genes or gene variants that either have recently entered the literature or for which the nomenclature or description is currently unclear (selz, sel26, sel27, sel28, and ses-2p). An examination of over 11,000 RefSeq genomes in search of wider support for these seven (pseudo)genes led to the identification of an additional three novel enterotoxin gene family members (sel31, sel32, and sel33) plus two new variants (seh-2p and ses-3p). We cast light on the genomic distribution of the enterotoxin genes, further defining their arrangement in gene clusters. Finally, we show that cooccurrence of enterotoxin genes is prevalent, with individual NCTC strains possessing as many as 18 enterotoxin genes and pseudogenes, and that clonal complex membership rather than time of isolation is the key factor in determining enterotoxin load.IMPORTANCEStaphylococcus aureus strains pose a significant health risk to both human and animal populations. Key among this species' virulence factors is the staphylococcal enterotoxin gene family. Certain enterotoxin forms can induce a potentially life-threatening immune response, while others are implicated in less fatal though often severe conditions such as food poisoning. Genetic characterization of staphylococcal enterotoxin gene family members has steadily accumulated over recent decades, with over 20 genes now established in the literature. Despite the current wealth of knowledge on this important gene family, questions remain about the presence of additional enterotoxin genes and the genomic composition of family members. This study further expands knowledge of the staphylococcal enterotoxins while shedding light on their evolution over the last century.

A feasibility randomised controlled trial of a fibromyalgia self-management programme in a community setting with a nested qualitative study (FALCON): Study protocol.

BACKGROUND: Fibromyalgia (FM) is a complex long-term condition associated with chronic widespread pain, fatigue, sleep problems, memory and concentration difficulties and irritable bowel syndrome. Current guidelines for the treatment of FM recommend nonpharmacological interventions. The Fibromyalgia Self-Management Programme (FSMP) is a nonpharmacological, multidisciplinary exercise and education group intervention. It aims to provide education and teach core skills, enabling those affected by FM to self-manage. The FSMP is currently codelivered by a multidisciplinary team within a secondary care service. The aim of this feasibility randomised controlled trial (RCT) is to determine the practicality and acceptability of delivering the FSMP in a community setting, informing a future RCT of effectiveness. METHODS: The feasibility RCT aims to recruit 70 people with FM. Participants will be randomised to either a community FSMP or control arm. All participants will be asked to complete six patient-reported outcome measures and one health economics questionnaire on three occasions; baseline, 6 weeks (end of the intervention) and 6 months. Between 12 and 16 participants and four therapists delivering the FSMP will be invited to take part in a semi-structured interview to explore their experiences of the FSMP. Patient participants will be purposively selected based upon key characteristics. ANALYSIS: Quantitative data will be analysed descriptively to summarise recruitment and attendance, participant reported outcomes and health economic data. Semi-structured interviews will be transcribed, anonymised and inductively coded. The codes will be grouped into categories and theoretically thematically analysed, comparing the results to existing literature. TRIAL REGISTRATION: The trial is registered with ISRCTN registry and was assigned on 29th of April 2020. The registration number is ISRCTN10824225.

Alterations in Intestinal Microbiota of Children With Celiac Disease at the Time of Diagnosis and on a Gluten-free Diet.

BACKGROUND AND AIMS: It is not clear whether alterations in the intestinal microbiota of children with celiac disease (CD) cause the disease or are a result of disease and/or its treatment with a gluten-free diet (GFD). METHODS: We obtained 167 fecal samples from 141 children (20 with new-onset CD, 45 treated with a GFD, 57 healthy children, and 19 unaffected siblings of children with CD) in Glasgow, Scotland. Samples were analyzed by 16S ribosomal RNA sequencing, and diet-related metabolites were measured by gas chromatography. We obtained fecal samples from 13 children with new-onset CD after 6 and 12 months on a GFD. Relationships between microbiota with diet composition, gastrointestinal function, and biomarkers of GFD compliance were explored. RESULTS: Microbiota α diversity did not differ among groups. Microbial dysbiosis was not observed in children with new-onset CD. In contrast, 2.8% (Bray-Curtis dissimilarity index, P = .025) and 2.5% (UniFrac distances, P = .027) of the variation in microbiota composition could be explained by the GFD. Between 3% and 5% of all taxa differed among all group comparisons. Eleven distinctive operational taxonomic units composed a microbe signature specific to CD with high diagnostic probability. Most operational taxonomic units that differed between patients on a GFD with new-onset CD vs healthy children were associated with nutrient and food group intake (from 75% to 94%) and with biomarkers of gluten ingestion. Fecal levels of butyrate and ammonia decreased during the GFD. CONCLUSIONS: Although several alterations in the intestinal microbiota of children with established CD appear to be effects of a GFD, specific bacteria were found to be distinct biomarkers of CD. Studies are needed to determine whether these bacteria contribute to pathogenesis of CD.

Complete genome sequences of Streptococcus pyogenes type strain reveal 100%-match between PacBio-solo and Illumina-Oxford Nanopore hybrid assemblies.

We present the first complete, closed genome sequences of Streptococcus pyogenes strains NCTC 8198T and CCUG 4207T, the type strain of the type species of the genus Streptococcus and an important human pathogen that causes a wide range of infectious diseases. S. pyogenes NCTC 8198T and CCUG 4207T are derived from deposit of the same strain at two different culture collections. NCTC 8198T was sequenced, using a PacBio platform; the genome sequence was assembled de novo, using HGAP. CCUG 4207T was sequenced and a de novo hybrid assembly was generated, using SPAdes, combining Illumina and Oxford Nanopore sequence reads. Both strategies yielded closed genome sequences of 1,914,862 bp, identical in length and sequence identity. Combining short-read Illumina and long-read Oxford Nanopore sequence data circumvented the expected error rate of the nanopore sequencing technology, producing a genome sequence indistinguishable to the one determined with PacBio. Sequence analyses revealed five prophage regions, a CRISPR-Cas system, numerous virulence factors and no relevant antibiotic resistance genes. These two complete genome sequences of the type strain of S. pyogenes will effectively serve as valuable taxonomic and genomic references for infectious disease diagnostics, as well as references for future studies and applications within the genus Streptococcus.

Fibromyalgia Self-Management: Mapping the behaviour change techniques used in a practice-based programme.

BACKGROUND: Fibromyalgia (FM) is a complex long-term condition associated with pain, fatigue and concentration difficulties. There is limited robust evidence for the effectiveness of pharmacological treatments for FM, with current guidelines recommending nonpharmacological interventions. The clinically developed Fibromyalgia Self-Management Programme (FSMP) is a nonpharmacological, multidisciplinary education group intervention. The FSMP aims to provide condition-specific, patient-centred education and exercise advice, supporting the development of core self-management skills. This research aimed to map the FSMP to a recommended behaviour change taxonomy (BCT). METHODS: Non-participatory observations of the 4- and 6-week FSMP were conducted. Detailed notes on the content of the course, therapist delivery and any additional content not included in the manual were recorded. Subsequently, semistructured interviews were conducted with both therapists (n = 4) and patients (n = 9). Observation and a review of the FSMP manual data were deductively coded to the BCT. Interview data were added to the framework. RESULTS: The review of the FSMP manual and observations of the course showed that the programme coded onto 12 of the 16 BCT domains, encompassing 22 behaviour change techniques. Both patient and therapist interviews indicated that patients made positive changes, including increased activity levels, pacing, better quality sleep and improved communication with family members. Patients reported improvements to symptoms as a result of attending the course. CONCLUSIONS: The FSMP utilises a range of behaviour change techniques. Patients who attend the course feel supported to make changes to their behaviour, enabling them to manage their symptoms more effectively.

New ECCO model documents for Material Deposit and Transfer Agreements in compliance with the Nagoya Protocol.

The European Culture Collections' Organisation presents two new model documents for Material Deposit Agreement (MDA) and Material Transfer Agreement (MTA) designed to enable microbial culture collection leaders to draft appropriate agreement documents for, respectively, deposit and supply of materials from a public collection. These tools provide guidance to collections seeking to draft an MDA and MTA, and are available in open access to be used, modified, and shared. The MDA model consists of a set of core fields typically included in a 'deposit form' to collect relevant information to facilitate assessment of the status of the material under access and benefit sharing (ABS) legislation. It also includes a set of exemplary clauses to be included in 'terms and conditions of use' for culture collection management and third parties. The MTA model addresses key issues including intellectual property rights, quality, safety, security and traceability. Reference is made to other important tools such as best practices and code of conduct related to ABS issues. Besides public collections, the MDA and MTA model documents can also be useful for individual researchers and microbial laboratories that collect or receive microbial cultures, keep a working collection, and wish to share their material with others.

Can proteomics elucidate mechanisms of antimicrobial resistance in Neisseria gonorrhoeae that whole genome sequencing is unable to identify? An analysis of protein expression within the 2016 WHO N. gonorrhoeae reference strains.

OBJECTIVES: Antimicrobial resistance (AMR) in Neisseria gonorrhoeae is of increasing concern. This study established a quantitative, scalable proteomics method to examine the WHO panel of N. gonorrhoeae isolates with completed closed genomic sequences and well-defined phenotypical and genotypical AMR patterns, to gain a greater understanding of AMR in N. gonorrhoeae. METHODS: 14 WHO reference strains were propagated, pooled stable isotope labelled lysates were used as an internal standard (IS). Protein lysates were mixed with IS, digested with trypsin and fractionated before analysis by nano-LC/MS/MS, in triplicate. The susceptible strain WHO F was used as reference to which the proteomic profiles of other strains were compared. Hierarchical clustering and permutation adjusted t-tests were performed to find proteins with significant fold changes. RESULTS: Standardised, reproducible protein expression profiles in N. gonorrhoeae reference strains were produced. Strains that have previously been shown to be highly similar using genomics, displayed different proteomic profiles. Several proteins from efflux pumps to stress responses, such as oxidative stress, toxin/antitoxin systems, were found to be altered in AMR strains. LtgE was upregulated in strains which displayed chromosomally mediated resistance to penicillin. MacB (the ATP hydrolysis part of macrolide efflux pump MacA-B), was ~twofold upregulated in WHO V (MIC of azithromycin >256 mg/L) and maybe associated with azithromycin resistance. CONCLUSIONS: A robust method was developed to study protein expression in N. gonorrhoeae. The proteome profiles could differentiate genetically similar stains. This study identified complex mechanisms in N. gonorrhoeae which may be associated with AMR.

Whole-Genome Sequences of Five Strains of Kocuria rosea, NCTC2676, NCTC7514, NCTC7512, NCTC7528, and NCTC7511.

Kocuria rosea is a Gram-positive coccus found in the environment and within normal human skin microbiota, and more recently, it has been potentially implicated as an opportunistic pathogen. Here, we describe the genome sequences of five strains of K. rosea (NCTC2676, NCTC7514, NCTC7512, NCTC7528, and NCTC7511).

Complete Whole-Genome Sequences of Two Raoultella terrigena Strains, NCTC 13097 and NCTC 13098, Isolated from Human Cases.

Raoultella terrigena is a bacterial species associated with soil and aquatic environments; however, sporadic cases of opportunistic disease in humans have been reported. Here, we report the first two complete genome sequences from clinical strains isolated from human sources that have been deposited in the National Collection of Type Cultures (NCTC).

Complete Whole-Genome Sequence of Haemophilus haemolyticus NCTC 10839.

Haemophilus haemolyticus is a Gram-negative bacterium that is a commensal of the respiratory tract in humans. Here, we report the complete genome sequence available for Haemophilus haemolyticus strain NCTC 10839, which was originally isolated from the nasopharynx of a child.

The history, genome and biology of NCTC 30: a non-pandemic Vibrio cholerae isolate from World War One.

The sixth global cholera pandemic lasted from 1899 to 1923. However, despite widespread fear of the disease and of its negative effects on troop morale, very few soldiers in the British Expeditionary Forces contracted cholera between 1914 and 1918. Here, we have revived and sequenced the genome of NCTC 30, a 102-year-old Vibrio cholerae isolate, which we believe is the oldest publicly available live V. cholerae strain in existence. NCTC 30 was isolated in 1916 from a British soldier convalescent in Egypt. We found that this strain does not encode cholera toxin, thought to be necessary to cause cholera, and is not part of V. cholerae lineages responsible for the pandemic disease. We also show that NCTC 30, which predates the introduction of penicillin-based antibiotics, harbours a functional ß-lactamase antibiotic resistance gene. Our data corroborate and provide molecular explanations for previous phenotypic studies of NCTC 30 and provide a new high-quality genome sequence for historical, non-pandemic V. cholerae.

Draft Genome Sequences of 64 Type Strains of 50 Species and 25 Subspecies of the Genus Staphylococcus Rosenbach 1884.

Members of the genus Staphylococcus have been isolated from humans, animals, and the environment. Accurate identification with whole-genome sequencing requires access to data derived from type strains. We provide sequence data for type strains of 64 taxa in the genus that at the time of this writing have standing in the nomenclature.

Mycobacterial biomaterials and resources for researchers.

There are many resources available to mycobacterial researchers, including culture collections around the world that distribute biomaterials to the general scientific community, genomic and clinical databases, and powerful bioinformatics tools. However, many of these resources may be unknown to the research community. This review article aims to summarize and publicize many of these resources, thus strengthening the quality and reproducibility of mycobacterial research by providing the scientific community access to authenticated and quality-controlled biomaterials and a wealth of information, analytical tools and research opportunities.

The effect of metabolic stress on genome stability of a synthetic biology chassis Escherichia coli K12 strain.

BACKGROUND: Synthetic organism-based biotechnologies are increasingly being proposed for environmental applications, such as in situ sensing. Typically, the novel function of these organisms is delivered by compiling genetic fragments in the genome of a chassis organism. To behave predictably, these chassis are designed with reduced genomes that minimize biological complexity. However, in these proposed applications it is expected that even when contained within a device, organisms will be exposed to fluctuating, often stressful, conditions and it is not clear whether their genomes will retain stability. RESULTS: Here we employed a chemostat design which enabled us to maintained two strains of E. coli K12 under sustained starvation stress: first the reduced genome synthetic biology chassis MDS42 and then, the control parent strain MG1655. We estimated mutation rates and utilised them as indicators of an increase in genome instability. We show that within 24 h the spontaneous mutation rate had increased similarly in both strains, destabilizing the genomes. High rates were maintained for the duration of the experiment. Growth rates of a cohort of randomly sampled mutants from both strains were utilized as a proxy for emerging phenotypic, and by association genetic variation. Mutant growth rates were consistently less than rates in non-mutants, an indicator of reduced fitness and the presence of mildly deleterious mutations in both the strains. In addition, the effect of these mutations on the populations as a whole varied by strain. CONCLUSIONS: Overall, this study shows that genome reductions in the MDS42 did not stabilize the chassis under metabolic stress. Over time, this could compromise the effectiveness of synthetic organisms built on chassis in environmental applications.

Complete Whole-Genome Sequence of Salmonella enterica subsp. enterica Serovar Java NCTC5706.

Salmonellae are a significant cause of morbidity and mortality globally. Here, we report the first complete genome sequence for Salmonella enterica subsp. enterica serovar Java strain NCTC5706. This strain is of historical significance, having been isolated in the pre-antibiotic era and was deposited into the National Collection of Type Cultures in 1939.