An initial genomic blueprint of the healthy human oesophageal microbiome.

Background: The oesophageal microbiome is thought to contribute to the pathogenesis of oesophageal cancer. However, investigations using culture and molecular barcodes have provided only a low-resolution view of this important microbial community. We therefore explored the potential of culturomics and metagenomic binning to generate a catalogue of reference genomes from the healthy human oesophageal microbiome, alongside a comparison set from saliva. Results: Twenty-two distinct colonial morphotypes from healthy oesophageal samples were genome-sequenced. These fell into twelve species clusters, eleven of which represented previously defined species. Two isolates belonged to a novel species, which we have named Rothia gullae. We performed metagenomic binning of reads generated from UK samples from this study alongside reads generated from Australian samples in a recent study. Metagenomic binning generated 136 medium or high-quality metagenome-assembled genomes (MAGs). MAGs were assigned to 56 species clusters, eight representing novel Candidatus species, which we have named Ca. Granulicatella gullae, Ca. Streptococcus gullae, Ca. Nanosynbacter quadramensis, Ca. Nanosynbacter gullae, Ca. Nanosynbacter colneyensis, Ca. Nanosynbacter norwichensis, Ca. Nanosynococcus oralis and Ca. Haemophilus gullae. Five of these novel species belong to the recently described phylum Patescibacteria . Although members of the Patescibacteria are known to inhabit the oral cavity, this is the first report of their presence in the oesophagus. Eighteen of the metagenomic species were, until recently, identified only by hard-to-remember alphanumeric placeholder designations. Here we illustrate the utility of a set of recently published arbitrary Latinate species names in providing user-friendly taxonomic labels for microbiome analyses.Our non-redundant species catalogue contained 63 species derived from cultured isolates or MAGs. Mapping revealed that these species account for around half of the sequences in the oesophageal and saliva metagenomes. Although no species was present in all oesophageal samples, 60 species occurred in at least one oesophageal metagenome from either study, with 50 identified in both cohorts. Conclusions: Recovery of genomes and discovery of new species represents an important step forward in our understanding of the oesophageal microbiome. The genes and genomes that we have released into the public domain will provide a base line for future comparative, mechanistic and intervention studies.

Development and validation of ester impregnated pH strips for locating nasogastric feeding tubes in the stomach-a multicentre prospective diagnostic performance study.

BACKGROUND: NG (nasogastric) tubes are used worldwide as a means to provide enteral nutrition. Testing the pH of tube aspirates prior to feeding is commonly used to verify tube location before feeding or medication. A pH at or lower than 5.5 was taken as evidence for stomach intubation. However, the existing standard pH strips lack sensitivity, especially in patients receiving feeding and antacids medication. We developed and validated a first-generation ester-impregnated pH strip test to improve the accuracy towards gastric placements in adult population receiving routine NG-tube feeding. The sensitivity was improved by its augmentation with the action of human gastric lipase (HGL), an enzyme specific to the stomach. METHODS: We carried out a multi-centred, prospective, two-gate diagnostic accuracy study on patients who require routine NG-tube feeding in 10 NHS hospitals comparing the sensitivity of the novel pH strip to the standard pH test, using either chest X-rays or, in its absence, clinical observation of the absence of adverse events as the reference standard. We also tested the novel pH strips in lung aspirates from patients undergoing oesophageal cancer surgeries using visual inspection as the reference standard. We simulated health economics using a decision analytic model and carried out adoption studies to understand its route to commercialisation. The primary end point is the sensitivity of novel and standard pH tests at the recommended pH cut-off of 5.5. RESULTS: A total of 6400 ester-impregnated pH strips were prepared based on an ISO13485 quality management system. A total of 376 gastric samples were collected from adult patients in 10 NHS hospitals who were receiving routine NG-tube feeding. The sensitivities of the standard and novel pH tests were respectively 49.2% (95% CI 44.1­54.3%) and 70.2% (95% CI 65.6­74.8%) under pH cut-off of 5.5 and the novel test has a lung specificity of 89.5% (95% CI 79.6%, 99.4%). Our simulation showed that using the novel test can potentially save 132 unnecessary chest X-rays per check per every 1000 eligible patients, or direct savings of £4034 to the NHS. CONCLUSIONS: The novel pH test correctly identified significantly more patients with tubes located inside the stomach compared to the standard pH test used widely by the NHS. TRIAL REGISTRATION: http://www.isrctn.com/ISRCTN11170249 , Registered 21 June 2017-retrospectively registered.

Endogenous aldehyde accumulation generates genotoxicity and exhaled biomarkers in esophageal adenocarcinoma.

Volatile aldehydes are enriched in esophageal adenocarcinoma (EAC) patients' breath and could improve early diagnosis, however the mechanisms of their production are unknown. Here, we show that weak aldehyde detoxification characterizes EAC, which is sufficient to cause endogenous aldehyde accumulation in vitro. Two aldehyde groups are significantly enriched in EAC biopsies and adjacent tissue: (i) short-chain alkanals, and (ii) medium-chain alkanals, including decanal. The short-chain alkanals form DNA-adducts, which demonstrates genotoxicity and confirms inadequate detoxification. Metformin, a putative aldehyde scavenger, reduces this toxicity. Tissue and breath concentrations of the medium-chain alkanal decanal are correlated, and increased decanal is linked to reduced ALDH3A2 expression, TP53 deletion, and adverse clinical features. Thus, we present a model for increased exhaled aldehydes based on endogenous accumulation from reduced detoxification, which also causes therapeutically actionable genotoxicity. These results support EAC early diagnosis trials using exhaled aldehyde analysis.

Mass-Spectrometry Analysis of Mixed-Breath, Isolated-Bronchial-Breath, and Gastric-Endoluminal-Air Volatile Fatty Acids in Esophagogastric Cancer.

A noninvasive breath test has the potential to improve survival from esophagogastric cancer by facilitating earlier detection. This study aimed to investigate the production of target volatile fatty acids (VFAs) in esophagogastric cancer through analysis of the ex vivo headspace above underivatized tissues and in vivo analysis within defined anatomical compartments, including analysis of mixed breath, isolated bronchial breath, and gastric-endoluminal air. VFAs were measured by PTR-ToF-MS and GC-MS. Levels of VFAs (acetic, butyric, pentanoic, and hexanoic acids) and acetone were elevated in ex vivo experiments in the headspace above esophagogastric cancer compared with the levels in samples from control subjects with morphologically normal and benign conditions of the upper gastrointestinal tract. In 25 patients with esophagogastric cancer and 20 control subjects, receiver-operating-characteristic analysis for the cancer-specific VFAs butyric acid ( P < 0.001) and pentatonic acid ( P = 0.005) within in vivo gastric-endoluminal air gave an area under the curve of 0.80 (95% confidence interval of 0.65 to 0.93, P = 0.01). Compared with mixed- and bronchial-breath samples, all examined VFAs were found in highest concentrations within esophagogastric-endoluminal air. In addition, VFAs were higher in all samples derived from cancer patients compared with in the controls. Equivalence of VFA levels within the mixed and bronchial breath of cancer patients suggests that their origin within breath is principally derived from the lungs and, by inference, from the systemic circulation as opposed to direct passage from the upper gastrointestinal tract. These findings highlight the potential to utilize VFAs for endoluminal-gas biopsies and noninvasive mixed-exhaled-breath testing for esophagogastric-cancer detection.