Bacterial Culture Collection Research Articles

Gut microbiota carcinogen metabolism causes distal tissue tumours

Exposure to environmental pollutants and human microbiome composition are important predisposition factors for tumour development1,2. Similar to drug molecules, pollutants are typically metabolized in the body, which can change their carcinogenic potential and affect tissue distribution through altered toxicokinetics3. Although recent studies demonstrated that human-associated microorganisms can chemically convert a wide range of xenobiotics and influence the profile and tissue exposure of resulting metabolites4,5, the effect of microbial biotransformation on chemical-induced tumour development remains unclear. Here we show that the depletion of the gut microbiota affects the toxicokinetics of nitrosamines, which markedly reduces the development and severity of nitrosamine-induced urinary bladder cancer in mice6,7. We causally linked this carcinogen biotransformation to specific gut bacterial isolates in vitro and in vivo using individualized bacterial culture collections and gnotobiotic mouse models, respectively. We tested gut communities from different human donors to demonstrate that microbial carcinogen metabolism varies between individuals and we showed that this metabolic activity applies to structurally related nitrosamine carcinogens. Altogether, these results indicate that gut microbiota carcinogen metabolism may be a contributing factor for chemical-induced carcinogenesis, which could open avenues to target the microbiome for improved predisposition risk assessment and prevention of cancer.

Alginate- and Chitosan-Modified Gelatin Hydrogel Microbeads for Delivery of E. coli Phages.

Bacterial infections are among the most significant health problems/concerns worldwide. A very critical concern is the rapidly increasing number of antibiotic-resistant bacteria, which requires much more effective countermeasures. As nature's antibacterial entities, bacteriophages shortly ("phages") are very important alternatives to antibiotics, having many superior features compared with antibiotics. The development of phage-carrying controlled-release formulations is still challenging due to the need to protect their activities in preparation, storage, and use, as well as the need to create more user-friendly forms by considering their application area/site/conditions. Here, we prepared gelatin hydrogel microbeads by a two-step process. Sodium alginate was included for modification within the initial recipes, and these composite microbeads were further coated with chitosan. Their swelling ratio, average diameters, and Zeta potentials were determined, and degradations in HCl were demonstrated. The target bacteria Escherichia coli (E.coli) and its specific phage (T4) were obtained from bacterial culture collections and propagated. Phages were loaded within the microbeads with a simple method. The phage release characteristics were investigated comparatively and were demonstrated here. High release rates were observed from the gelatin microbeads. It was possible to reduce the phage release rate using sodium alginate in the recipe and chitosan coating. Using these gelatin-based microbeads as phage carrier matrices-especially in lyophilized forms-significantly improved the phage stability even at room temperature. It was concluded that phage release from gelatin hydrogel microbeads could be further controlled by alginate and chitosan modifications and that user-friendly lyophilized phage formulations with a much longer shelf life could be produced.

Eight complete genome sequences of bacteria isolated from laboratory stock of giant kelp gametophytes.

We report whole-genome sequences of eight bacteria isolated from laboratory-kept Macrocystis pyrifera gametophytes. The bacterial culture collection is maintained in cryostorage and will be utilized in future applications as inoculants. The genomes were assembled using Oxford Nanopore Technology long-read sequencing.

Bacterial clade-specific analysis identifies distinct epithelial responses in inflammatory bowel disease

Abnormal immune responses to the resident gut microbiome can drive inflammatory bowel disease (IBD). Here, we combine high-resolution, culture-based shotgun metagenomic sequencing and analysis with matched host transcriptomics across three intestinal sites (terminal ileum, cecum, rectum) from pediatric IBD (PIBD) patients (n= 58) and matched controls (n= 42) to investigate this relationship. Combining our site-specific approach with bacterial culturing, we establish a cohort-specific bacterial culture collection, comprising 6,620 isolates (170 distinct species, 32 putative novel), cultured from 286 mucosal biopsies. Phylogeny-based, clade-specific metagenomic analysis identifies key, functionally distinct Enterococcus clades associated with either IBD or health. Strain-specific invitro validation demonstrates differences in cell cytotoxicity and inflammatory signaling in intestinal epithelial cells, consistent with the colonic mucosa-specific response measured in patients with IBD. This demonstrates the importance of strain-specific phenotypes and consideration of anatomical sites in exploring the dysregulated host-bacterial interactions in IBD.

Hemolysis and hypersensitive tests ease culture collection management of antagonistic bacteria

A biosafety assessment is a crucial first step in the management of microbial culture collection to screen and determine unexpected potential plant and human pathogenic bacteria. It is common to collect and store as many fresh culture collections from natural resources before being further evaluated for antagonist bacteria. As a result, a bulk of isolates must be preserved which required more effort and budget. Safety evaluations based on the hemolysis and hypersensitive reactions offer simple tests to ease culture collection management of antagonist bacteria. The study aimed to evaluate the safety of bacterial culture collection for their hemolysis and hypersensitive reactions. Ninety-five isolates of rhizosphere and endophytic bacterial isolates from the culture collections of the Department of Plant Protection-IPB University, Indonesian Industrial and Beverages Crops Research Institute, and Indonesian Spice and Medicinal Crops Research Institute, were evaluated their safety using the hemolysis and hypersensitive tests. The hemolysis test was conducted using blood agar media, from which isolates with a negative (?) reaction were then tested for the hypersensitivity reaction on tobacco leaves. Bacterial isolates passed from both hemolysis and hypersensitivity tests were then preserved by the lyophilization method for long-term storage of culture collection. Based on the hemolysis test, 68 out of 95 bacterial isolates (71.57%) were found to be positive (? or ?) reactions. The hypersensitive test showed that 22 of 27 negative hemolysis isolates did not trigger hypersensitivity reactions in tobacco leaves, therefore, they were preserved by lyophilization. The study indicated that a high number of bacterial isolates in the present collection, 68 positive hemolysis, and 5 hypersensitive, need to be re-evaluated due to their safety concerns. The present study highlights the importance of biosafety tests performed in an early stage before the to permanent collection of antagonist isolates.

Characterization of a Bacterial Culture Collection from Terrestrial Subsurface Habitats in Colombia

Characterization of a Bacterial Culture Collection from Terrestrial Subsurface Habitats in Colombia

Comparative study of the rhizosphere microbiome of Coffea arabica grown in different countries reveals a small set of prevalent and keystone taxa

Coffee (Coffea arabica) is a globally important crop and its cultivation demands the use of agrochemicals and pesticides. Recently, plant-associated microorganisms have emerged as an unexploited resource to enhance crop performance and sustainability. In particular, the rhizosphere microbiome has been identified as a hotspot for plant health, by providing tolerance to biotic and abiotic stresses as well as regulating phenological traits. However, its composition varies widely across crops and environmental conditions. Hence, the knowledge of the “common core” taxa, which are a subset of the microbiome that is persistently associated, across different environmental gradients and conditions, to a host species, represent valuable targets for the development of microbiome-based management approaches. This study aimed to characterize and determine the rhizosphere-inhabiting microbial groups (common core and most prevalent taxa), associated with the Coffea arabica plants in different geographical locations. To assess the structure of the coffee rhizosphere microbiome, we performed the 16 S rRNA amplicon analysis of samples collected across six locations spanning four countries and grown under different climate conditions, soil type, agroecosystem management, plant vegetative stage and age. Our findings allowed us to delineate and identify a robust set of six bacterial taxa, namely Sphingobium, Rhizobium group, Acidibacter, Sphingomonas, Burkholderia group and Amycolatopsis, which are likely to play an important role for the growth and health of coffee plants. Using network analysis, we observed that some of the six “common core” taxa were highly interconnected with the resident community, being also keystone taxa of the coffee rhizosphere microbiome. Furthermore, we performed a high-throughput targeted isolation resulting into 162 rhizosphere bacterial strains, some of them related to the most prevalent taxa. These isolates represent a working bacterial culture collection to be used in the future for the development of microbiome-based solutions for a more sustainable coffee farming.

Multiplexed mobilization and expression of biosynthetic gene clusters

Bacterial genomes contain large reservoirs of biosynthetic gene clusters (BGCs) that are predicted to encode unexplored natural products. Heterologous expression of previously unstudied BGCs should facilitate the discovery of additional therapeutically relevant bioactive molecules from bacterial culture collections, but the large-scale manipulation of BGCs remains cumbersome. Here, we describe a method to parallelize the identification, mobilization and heterologous expression of BGCs. Our solution simultaneously captures large numbers of BGCs by cloning the genomes of a strain collection in a large-insert library and uses the CONKAT-seq (co-occurrence network analysis of targeted sequences) sequencing pipeline to efficiently localize clones carrying intact BGCs which represent candidates for heterologous expression. Our discovery of several natural products, including an antibiotic that is active against multi-drug resistant Staphylococcus aureus, demonstrates the potential of leveraging economies of scale with this approach to systematically interrogate cryptic BGCs contained in strain collections.

Nanocarriers based on bacterial membrane materials for cancer vaccine delivery.

Here we present a protocol for the construction and use of two types of nanocarrier based on bacterial membrane materials for cancer vaccine delivery. Cancer vaccines induce tumor regression through triggering the specific T-cell responses against tumor neoantigens, a process that can be enhanced by nanocarrier delivery. Inspired by the body's natural immune defenses against bacterial invasion, we have developed two different types of nanocarrier based on bacterial membrane materials, which employ genetically engineered outer-membrane vesicles (OMVs), or hybrid membrane vesicles containing bacterial cytoplasmic membrane, respectively. The OMV-based nanocarriers can rapidly display different tumor antigens through the surface modified Plug-and-Display system, suitable for customized cancer vaccines when the tumor neoantigens can be identified. The hybrid membrane-based nanocarriers are prepared through fusion of the bacterial cytoplasmic membrane and the primary tumor cell membrane from surgically removed tumor tissues, possessing unique advantages as personalized cancer vaccines when the neoantigens are not readily available. Compared with chemically synthesized nanocarriers such as liposomes and polymer without intrinsic adjuvant properties, owing to the large amounts of pathogen-associated molecular patterns, the two nanocarriers can activate the antigen-presenting cells while delivering multiple antigens, thus inducing effective antigen presentation and robust adaptive immune activation. Excluding bacterial culture and tumor tissue collection, the preparation of OMV- and hybrid membrane-based nanocarriers takes ~8 h and 10 h for tumor vaccine construction, respectively. We also detail how to use these nanocarriers to create cancer nanovaccines and evaluate their immunostimulatory and antitumor effects.

Building More Resilient Culture Collections: A Call for Increased Deposits of Plant-Associated Bacteria.

Biological collections preserve our past, while helping protect our future and increase future knowledge. Plant bacterial culture collections are our security for domestic and global biosecurity. This feature article will provide an introduction to the global position of plant bacterial collections. The role of collections in monitoring plant pathogenic bacteria will be explored through the presentation of five cases studies. These case studies demonstrate why culture collections were imperative for the outcome in each situation. We discuss what we believe should be the best practices to improve microbial preservation and accessioning rates, and why plant bacterial culture collections must increase deposits to be prepared for future emerging pathogens. This is not only the case for global culture collections, but on a much bigger scale, our future scientific successes, our biosecurity decisions and responses, and our knowledge are contingent upon preserving our valuable bacterial strains. It is hoped that once you read this article, you will see the need to deposit your strains in registered public collections and make a concerted effort to build better bacterial culture collections with us.

Identification of the contaminating psychrotrophic bacteria in refrigerated bulked raw milk and the assessment of their deteriorating potential

This work aimed to characterize, identify, and determine the deteriorating potential of the contaminating psychrotrophic bacteria in refrigerated raw milk. Samples were submitted to serial dilutions and plated in specific culture media to form a bacterial culture collection. The isolates were characterized for their morphology and biochemical characteristics. The deteriorating potential of the isolates was determined according to the proteolytic, lipolytic and lecithinase activities at 4.0 ºC, 6.5 ºC, and 25.0 ºC. The results obtained for deterioration potential were assessed by the multivariate statistical method and by the principal components analysis (PCA). A total of 159 isolates were characterized, and of these, 46 strongly proteolytic Gram-negative isolates were selected for identification using the API 20 NE kit. The predominant bacteria were Gram-negative and oxidase and catalase positive, with a predominance of bacteria of the genus Pseudomonas. Using PCA, it was shown that the bacteria with the greatest deterioration potential were lecithinase producers, and that, in the autumn, proteolytic bacteria predominated at 4.0 ºC. Of the 46 isolates identified, more than 80% belonged to the species Pseudomonas fluorescens. Thus, attention should be given to the importance of implementing microbial contamination prevention measures in the bulking process, since, even under refrigeration, psychrotrophic bacteria multiply and produce enzymes that deteriorate lipids and proteins, with consequent quality losses of the milk and its derivatives, yield losses in the production of dairy products, and economic losses.

A Bacterial Genome and Culture Collection of Gut Microbial in Weanling Piglet.

ABSTRACTThe microbiota hosted in the pig gastrointestinal tract are important to health of this biomedical model. However, the individual species and functional repertoires that make up the pig gut microbiome remain largely undefined. Here we comprehensively investigated the genomes and functions of the piglet gut microbiome using culture-based and metagenomics approaches. A collection included 266 cultured genomes and 482 metagenome-assembled genomes (MAGs) that were clustered to 428 species across 10 phyla was established. Among these clustered species, 333 genomes represent potential new species. Less matches between cultured genomes and MAGs revealed a substantial bias for the acquisition of reference genomes by the two strategies. Glycoside hydrolases was the dominant category of carbohydrate-active enzymes. Four-hundred forty-five secondary metabolite biosynthetic genes were predicted from 292 genomes with bacteriocin being the most. Pan genome analysis of Limosilactobacillus reuteri uncover the biosynthesis of reuterin was strain-specific and the production was experimentally determined. This study provides a comprehensive view of the microbiome composition and the function landscape of the gut of weanling piglets and a valuable bacterial resource for further experimentations.IMPORTANCE The microorganism communities resided in mammalian gastrointestinal tract impacted the health and disease of the host. Our study complements metagenomic analysis with culture-based approach to establish a bacteria and genome collection and comprehensively investigate the microbiome composition and function of the gut of weanling piglets. We provide a valuable resource for further study of gut microbiota of weanling piglet and development of probiotics for prevention of disease.

Investigation of the Role of Vitamin C In Enhancing the Activity of Antibacterial Agents and Biofilm Formation

Resistance to antibiotics is rapidly spreading across the globe, posing a new health-care challenge for all countries. Bacterial biofilms are three-dimensional formations made up of cells encased in a matrix of polymeric, making cells resistant to the drugs and the immune system. As a result, new tactics for inhibiting the production of the EPS matrix may lead to more efficient use of already available antibiotics. The mechanism of vitamins C effect on boosting the effectiveness of several anti-bacterial drugs was investigated in this study, the target isolates were obtained from University of Mosul/ Biology department/ bacterial culture collections and evaluated qualitatively and quantitatively. The isolates involved: Staphylococcus aureus, Escherichia coli ,Klebsiella sp., Serratia marcescens and Pseudomonas aeroginosa. Antibiotic sensitivity tests and biofilm producing assay results revealed that the majority of the isolates were resistant to a range of antibiotics and had a large capacity for biofilm formation when grown on a cover glass surface. Vitamin C is an antioxidant, a scavenger of active metabolites. The Minimum Inhibitory Concentration (MIC) of Vitamin C against selected isolates had been determined, and all further experiments used concentrations below the MIC. Our results showed that Vit.C pre-treatment enhance the bactericidal effect of antibiotic and increases bacterial susceptibility to antibiotics. Using light microscopy, experiments of sub inhibitory doses of Vitamin C revealed good suppression of selected isolates biofilm development on the cover glass surface. Vitamin C can be utilized as an antibiotic adjuvant in combination with antibiotic and has effective biofilm inhibition, caused by multidrug-resistant bacteria, according to evidence.

Host preference and invasiveness of commensal bacteria in the Lotus and Arabidopsis root microbiota

Roots of different plant species are colonized by bacterial communities, that are distinct even when hosts share the same habitat. It remains unclear to what extent the host actively selects these communities and whether commensals are adapted to a specific plant species. To address this question, we assembled a sequence-indexed bacterial culture collection from roots and nodules of Lotus japonicus that contains representatives of most species previously identified using metagenomics. We analysed taxonomically paired synthetic communities from L. japonicus and Arabidopsis thaliana in a multi-species gnotobiotic system and detected signatures of host preference among commensal bacteria in a community context, but not in mono-associations. Sequential inoculation experiments revealed priority effects during root microbiota assembly, where established communities are resilient to invasion by latecomers, and that host preference of commensal bacteria confers a competitive advantage in their cognate host. Our findings show that host preference in commensal bacteria from diverse taxonomic groups is associated with their invasiveness into standing root-associated communities.

Screening of Human Gut Bacterial Culture Collection Identifies Species That Biotransform Quercetin into Metabolites with Anticancer Properties.

We previously demonstrated that flavonoid metabolites inhibit cancer cell proliferation through both CDK-dependent and -independent mechanisms. The existing evidence suggests that gut microbiota is capable of flavonoid biotransformation to generate bioactive metabolites including 2,4,6-trihydroxybenzoic acid (2,4,6-THBA), 3,4-dihydroxybenzoic acid (3,4-DHBA), 3,4,5-trihyroxybenzoic acid (3,4,5-THBA) and 3,4-dihydroxyphenylacetic acid (DOPAC). In this study, we screened 94 human gut bacterial species for their ability to biotransform flavonoid quercetin into different metabolites. We demonstrated that five of these species were able to degrade quercetin including Bacillus glycinifermentans, Flavonifractor plautii, Bacteroides eggerthii, Olsenella scatoligenes and Eubacterium eligens. Additional studies showed that B. glycinifermentans could generate 2,4,6-THBA and 3,4-DHBA from quercetin while F. plautii generates DOPAC. In addition to the differences in the metabolites produced, we also observed that the kinetics of quercetin degradation was different between B. glycinifermentans and F. plautii, suggesting that the pathways of degradation are likely different between these strains. Similar to the antiproliferative effects of 2,4,6-THBA and 3,4-DHBA demonstrated previously, DOPAC also inhibited colony formation ex vivo in the HCT-116 colon cancer cell line. Consistent with this, the bacterial culture supernatant of F. plautii also inhibited colony formation in this cell line. Thus, as F. plautii and B. glycinifermentans generate metabolites possessing antiproliferative activity, we suggest that these strains have the potential to be developed into probiotics to improve human gut health.

Antibacterial effect of polyphenols enriched drumstick plant leaves (Moringa oleifera) extract: A research study

Microorganisms are raising resistance against available antibiotics due to usage of antibiotics in a wrong way. There is lot of demand for finding /searching new natural antimicrobial agents as they are inexpensive, easily available, most of them are edible and mainly non-toxic even at the rate of milligram quantity. The aim was to study the efficiency of the anti-bacterial effect of Polyphenols enriched extract of Drum stick plant leaves (Moringa oleifera) against staphylococcus aureus. The materials involved in this study include Drum stick plant leaves, micro-organism staphylococcus aureus in the bacterial type culture collection, agar, and blood-agar plates. At 10% concentration, of Polyphenols enriched extract of Drum stick leaves had zero anti-bacterial activity, while between 20 to 25% concentrations revealed high activity against the bacteria. Thus, increased in the anti-bacterial activity was promising as the concentration augmented from 20 to 25%. The results acquired from this study points that polyphenol enriched extract of Drum stick leaves (Moringa oleifera) had antibacterial property against Staphylococcus aureus when obtained to a necessary concentration.

In Silico Comparison Shows that the Pan-Genome of a Dairy-Related Bacterial Culture Collection Covers Most Reactions Annotated to Human Microbiomes.

The diversity of the human microbiome is positively associated with human health. However, this diversity is endangered by Westernized dietary patterns that are characterized by a decreased nutrient variety. Diversity might potentially be improved by promoting dietary patterns rich in microbial strains. Various collections of bacterial cultures resulting from a century of dairy research are readily available worldwide, and could be exploited to contribute towards this end. We have conducted a functional in silico analysis of the metagenome of 24 strains, each representing one of the species in a bacterial culture collection composed of 626 sequenced strains, and compared the pathways potentially covered by this metagenome to the intestinal metagenome of four healthy, although overweight, humans. Remarkably, the pan-genome of the 24 strains covers 89% of the human gut microbiome’s annotated enzymatic reactions. Furthermore, the dairy microbial collection covers biological pathways, such as methylglyoxal degradation, sulfate reduction, γ-aminobutyric (GABA) acid degradation and salicylate degradation, which are differently covered among the four subjects and are involved in a range of cardiometabolic, intestinal, and neurological disorders. We conclude that microbial culture collections derived from dairy research have the genomic potential to complement and restore functional redundancy in human microbiomes.

Composition and structure of the culturable gut bacterial communities in Anopheles albimanus from Colombia.

The understanding of factors affecting the gut bacterial communities in malaria vectors is essential for the design of vector control interventions, such as those based on a paratransgenic approach. One of the requirements of this method is the availability of bacteria from the mosquito susceptible to culture. Thus, the aim of this study was to evaluate the composition and structure of the culturable gut bacterial communities in field mosquitoes Anopheles albimanus from Colombia, in addition to generate a bacterial collection to further analyze microbial functional activity. Gut bacteria were isolated from An. albimanus larvae and adult mosquitoes collected in localities of the Atlantic and Pacific Coasts. The bacterial isolates were grouped in 28 morphospecies that corresponded to three phyla, three classes, nine families and 14 genera. The larvae guts from San Antero (Atlantic Coast) and Buenaventura (Pacific Coast) shared the genera Bacillus and Lysinibacillus and in adults, Bacillus and Bacillus cereus Group were registered in both localities. Gut bacterial richness was higher in adults from the Pacific with respect to the Atlantic Coast, while larval richness was similar in samples of both coasts. The Shannon index indicated uniformity in morphospecies abundances in both localities. Finally, the characterization of morphospecies from the gut of Anopheles albimanus mosquitoes from Colombia by culture-dependent methods complemented with 16S rRNA gene sequencing allowed the definition, at a finer resolution, of the composition and structure of these microbial communities. In addition, the obtained bacterial culture collection will allow further evaluation of the microorganisms for their potential as biocontrol agents.

Screening the cultivable cave microbial mats for the production of antimicrobial compounds and antibiotic resistance

The current work extends the phenotypic characterization of a bacterial culture collection obtained from white, yellow, grey or pink microbial cave wall colonies that are common in the caves of Slovenian Karst. We have determined antibiotic resistance to 22 natural and synthetic antibiotics in 69 isolates from the microbial mats. Thirty-eight isolates (52%) were resistant to 1-5 antibiotics; another 27 isolates (37%) were resistant to 6-10 antibiotics; and 7 isolates (0.1%) were resistant to 11-17 antibiotics. We screened for production of antimicrobial compounds by growing cave isolates on five different media and overlaying individual cultures with ten Gram-positive and Gram negative multidrug resistant isolates. Out of 78 isolates tested, 15.3% showed antimicrobial activity against Escherichia coli DH5, 15.3% against extended-spectrum β-lactamase producing Escherichia coli, 3.8% against Salmonella enterica serovar Typhimurium TL747, 9% against Klebsiella pneumoniae ATCC BAA-1706, 9% against carbapenem-resistant Klebsiella pneumoniae ATCC BAA-1705, 7.7% against Bacillus cereus, 20.5% against Bacillus subtilis, 9% against Listeria monocytogenes, 19.2% against methicillin-resistant Staphylococcus aureus, and 20.5% against methicillin-resistant Staphylococcus pseudointermedius. Results showed the potential of cave microbes to suppress the growth of multi-resistant pathogens, and, a relatively high resistance to antibiotics.

The present & the past: a review of newly accessioned bacterial strains into the UK’s National Collection of Type Cultures

The National Collection of Type Cultures (NCTC) is a bacterial culture collection which is taxonomically and biologically diverse. NCTC holds approximately 6000 different strains from over 900 different species; among them strains originally isolated in the 19th century, strains for use as controls as stipulated EUCAST and ISO guidelines, type strains on which the description of bacterial species are based and other strains from a variety of backgrounds. The remit of NCTC is to provide authentic bacterial cultures of medical and veterinary interest to the scientific community, to support and enhance the reproducibility of scientific research and to improve global public health. To fulfil this remit, remain scientifically relevant and to preserve the legacy of contemporary medical bacteriology for future scientists, NCTC accessions strains of clinical significance: such as recently circulating and outbreak strains, diagnostic escape mutants and strains with novel antimicrobial resistance profiles. In 2018, 166 bacterial strains were accessioned into the NCTC and made available to the scientific community. These include NCTC 14052: a reference strain for emergent hyper-virulent K. pneumoniae, 4 type strains of newly described bacterial species, 82 strains accessioned from the Murray Collection of pre-antibiotic era Enterobacteriaceae and 8 strains with antimicrobial resistance mechanisms previously unrepresented in the collection, including NCTC 14208: a N. gonorrhoeae isolated from an instance of combined ceftriaxone and azithromycin treatment failure. Through literature review we have highlighted their value to the scientific community, both in their own right and in the context of bacterial strains already held by the NCTC.