Transmission of Alzheimer's disease-associated microbiota dysbiosis and its impact on cognitive function: evidence from mice and patients.

Spouses of Alzheimer's disease (AD) patients are at a higher risk of developing incidental dementia. However, the causes and underlying mechanism of this clinical observation remain largely unknown. One possible explanation is linked to microbiota dysbiosis, a condition that has been associated with AD. However, it remains unclear whether gut microbiota dysbiosis can be transmitted from AD individuals to non-AD individuals and contribute to the development of AD pathogenesis and cognitive impairment. We, therefore, set out to perform both animal studies and clinical investigation by co-housing wild-type mice and AD transgenic mice, analyzing microbiota via 16S rRNA gene sequencing, measuring short-chain fatty acid amounts, and employing behavioral test, mass spectrometry, site-mutations and other methods. The present study revealed that co-housing between wild-type mice and AD transgenic mice or administrating feces of AD transgenic mice to wild-type mice resulted in AD-associated gut microbiota dysbiosis, Tau phosphorylation, and cognitive impairment in the wild-type mice. Gavage with Lactobacillus and Bifidobacterium restored these changes in the wild-type mice. The oral and gut microbiota of AD patient partners resembled that of AD patients but differed from healthy controls, indicating the transmission of microbiota. The underlying mechanism of these findings includes that the butyric acid-mediated acetylation of GSK3ß at lysine 15 regulated its phosphorylation at serine 9, consequently impacting Tau phosphorylation. Pending confirmative studies, these results provide insight into a potential link between the transmission of AD-associated microbiota dysbiosis and development of cognitive impairment, which underscore the need for further research in this area.

Effects of Lifetime Exposures to Environmental Contaminants on the Adult Gut Microbiome.

Emerging experimental evidence indicates that toxicant-induced alterations in gut microbiota composition and activity may affect host homeostasis. However, data from human studies are scarce; to our knowledge, no previous studies have quantified the association of lifetime exposure to environmental chemicals, across multiple time points, with the composition of the adult gut microbiome. Here we studied 124 individuals born in the Faroe Islands in 1986-1987 who were followed approximately every seven years from birth through age 28 years. Organochlorine compounds, including polychlorinated biphenyls (PCBs) and pesticides, perfluoroalkyl substances (PFAS), and mercury (Hg), were measured in cord blood and longitudinally in participants' blood. At age 28, the gut microbiome was assessed using shotgun metagenomic sequencing. Historical contaminant exposures had little direct effect on the adult gut microbiome, while a small number of fastidious anaerobes were weakly linked to recent PFAS/PFOS exposures at age 28. In this cohort, our findings suggest no lasting effects of early life exposures on adult gut microbial composition, but proximal exposures may contribute to gut microbiome alterations. The methods developed and used for this investigation may help in future identification of small but lasting impacts of environmental toxicant exposure on the gut microbiome.

Differences of the Nasal Microbiome and Mycobiome by Clinical Characteristics of COPD Patients.

Rationale: While studies suggest that the lung microbiome may influence risk of chronic obstructive pulmonary disease (COPD) exacerbations, little is known about the relationship between the nasal biome and clinical characteristics of COPD patients. Methods: We sampled the nasal lining fluid by nasosorption of both nares of 20 people with moderate-to-severe COPD. All 40 samples, plus 4 negative controls, underwent DNA extraction, and 16SV4 ribosomal RNA (rRNA) (bacterial) and ribosomal internal transcribed spacer 2 (ITS2) (fungal) sequencing. We measured the proportion of variance (R2) in beta diversity explained by clinical factors, including age, sex, body mass index (BMI), COPD treatment, disease severity (forced expiratory volume in 1 second [FEV1], symptom/exacerbation frequency), peripheral eosinophil level (≥150 versus <150 cells/µL) and season of sampling, with the PERMANOVA test on the Bray-Curtis dissimilarities, accounting for within-person correlation of samples. We assessed the relative abundance of microbial features in the nasal community and their associations with clinical characteristics using the Microbiome Multivariable Association with Linear Models (MaAsLin2) package. Results: The most abundant nasal fluid bacterial taxa were Corynebacterium, Staphylococcus, Streptococcus, Moraxella, and Dolosigranulum, and fungal taxa were Malassezia, Candida, Malasseziales, Cladosporium and Aspergillus. Bacterial microbiome composition was associated with short-acting muscarinic antagonist use (R2 11.8%, p=0.002), sex (R2 8.3%, p=0.044), nasal steroid use (R2 7.7%, p=0.064), and higher eosinophil level (R2 7.6%, p=0.084). Mycobiome composition was associated with higher eosinophil level (R2 14.4%, p=0.004) and low FEV1 (R2 7.5%, p=0.071). No specific bacterium or fungus differed significantly in relative abundance by clinical characteristics in the multivariate per-feature analysis. Conclusion: The taxonomical composition of the nasal biome is heterogeneous in COPD patients and may be explained in part by clinical characteristics.

Dietary lignans, plasma enterolactone levels, and metabolic risk in men: exploring the role of the gut microbiome.

BACKGROUND: The conversion of plant lignans to bioactive enterolignans in the gastrointestinal tract is mediated through microbial processing. The goal of this study was to examine the relationships between lignan intake, plasma enterolactone concentrations, gut microbiome composition, and metabolic risk in free-living male adults. RESULTS: In 303 men participating in the Men's Lifestyle Validation Study (MLVS), lignan intake was assessed using two sets of 7-day diet records, and gut microbiome was profiled through shotgun sequencing of up to 2 pairs of fecal samples (n = 911). A score was calculated to summarize the abundance of bacteria species that were significantly associated with plasma enterolactone levels. Of the 138 filtered species, plasma enterolactone levels were significantly associated with the relative abundances of 18 species at FDR < 0.05 level. Per SD increment of lignan intake was associated with 20.7 nM (SEM: 2.3 nM) higher enterolactone concentrations among participants with a higher species score, whereas the corresponding estimate was 4.0 nM (SEM: 1.7 nM) among participants with a lower species score (P for interaction < 0.001). A total of 12 plasma metabolites were also significantly associated with these enterolactone-predicting species. Of the association between lignan intake and metabolic risk, 19.8% (95%CI: 7.3%-43.6%) was explained by the species score alone, 54.5% (95%CI: 21.8%-83.7%) by both species score and enterolactone levels, and 79.8% (95%CI: 17.7%-98.6%) by further considering the 12 plasma metabolites. CONCLUSION: We identified multiple gut bacteria species that were enriched or depleted at higher plasma levels of enterolactone in men. These species jointly modified the associations of lignan intake with plasma enterolactone levels and explained the majority of association between lignan intake and metabolic risk along with enterolactone levels and certain plasma metabolites.

Analysis of the Associations Between the Human Fecal Microbiome and Bone Density, Structure, and Strength: The Osteoporotic Fractures in Men (MrOS) Cohort.

In preclinical models, the composition and function of the gut microbiota have been linked to bone growth and homeostasis, but there are few available data from studies of human populations. In a hypothesis-generating experiment in a large cohort of community-dwelling older men (n = 831; age range, 78-98 years), we explored the associations between fecal microbial profiles and bone density, microarchitecture, and strength measured with total hip dual-energy X-ray absorptiometry (DXA) and high-resolution peripheral quantitative computed tomography (HRpQCT) (distal radius, distal and diaphyseal tibia). Fecal samples were collected and the 16S rRNA gene V4 hypervariable region sequenced. Sequences were bioinformatically processed through the DADA2 pipeline and then taxonomically assigned using SILVA. Generalized linear models as implemented in microbiome multivariable association with linear models (MaAsLin 2) were used to test for associations between skeletal measures and specific microbial genera. The abundances of four bacterial genera were weakly associated with bone density, structure, or strength (false discovery rate [FDR] ≤ 0.05), and the measured directions of associations of genera were generally consistent across multiple bone measures, supporting a role for microbiota on skeletal homeostasis. However, the associated effect sizes were small (log2 fold change < ±0.35), limiting power to confidently identify these associations even with high resolution skeletal imaging phenotypes, and we assessed the resulting implications for the design of future cohort-based studies. As in analogous examples from genomewide association studies, we find that larger cohort sizes will likely be needed to confidently identify associations between the fecal microbiota and skeletal health relying on 16S sequencing. Our findings bolster the view that the gut microbiome is associated with clinically important measures of bone health, while also indicating the challenges in the design of cohort-based microbiome studies. © 2022 American Society for Bone and Mineral Research (ASBMR).

Interplay between diet and gut microbiome, and circulating concentrations of trimethylamine N-oxide: findings from a longitudinal cohort of US men.

OBJECTIVES: Gut-produced trimethylamine N-oxide (TMAO) is postulated as a possible link between red meat intake and poor cardiometabolic health. We investigated whether gut microbiome could modify associations of dietary precursors with TMAO concentrations and cardiometabolic risk markers among free-living individuals. DESIGN: We collected up to two pairs of faecal samples (n=925) and two blood samples (n=473), 6 months apart, from 307 healthy men in the Men's Lifestyle Validation Study. Diet was assessed repeatedly using food-frequency questionnaires and diet records. We profiled faecal metagenome and metatranscriptome using shotgun sequencing and identified microbial taxonomic and functional features. RESULTS: TMAO concentrations were associated with the overall microbial compositions (permutational analysis of variance (PERMANOVA) test p=0.001). Multivariable taxa-wide association analysis identified 10 bacterial species whose abundance was significantly associated with plasma TMAO concentrations (false discovery rate <0.05). Higher habitual intake of red meat and choline was significantly associated with higher TMAO concentrations among participants who were microbial TMAO-producers (p<0.05), as characterised based on four abundant TMAO-predicting species, but not among other participants (for red meat, P-interaction=0.003; for choline, P-interaction=0.03). Among abundant TMAO-predicting species, Alistipes shahii significantly strengthened the positive association between red meat intake and HbA1c levels (P-interaction=0.01). Secondary analyses revealed that some functional features, including choline trimethylamine-lyase activating enzymes, were associated with TMAO concentrations. CONCLUSION: We identified microbial taxa that were associated with TMAO concentrations and modified the associations of red meat intake with TMAO concentrations and cardiometabolic risk markers. Our data underscore the interplay between diet and gut microbiome in producing potentially bioactive metabolites that may modulate cardiometabolic health.

The oral microbiome in relation to pancreatic cancer risk in African Americans.

BACKGROUND: African Americans have the highest pancreatic cancer incidence of any racial/ethnic group in the United States. The oral microbiome was associated with pancreatic cancer risk in a recent study, but no such studies have been conducted in African Americans. Poor oral health, which can be a cause or effect of microbial populations, was associated with an increased risk of pancreatic cancer in a single study of African Americans. METHODS: We prospectively investigated the oral microbiome in relation to pancreatic cancer risk among 122 African-American pancreatic cancer cases and 354 controls. DNA was extracted from oral wash samples for metagenomic shotgun sequencing. Alpha and beta diversity of the microbial profiles were calculated. Multivariable conditional logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) for associations between microbes and pancreatic cancer risk. RESULTS: No associations were observed with alpha or beta diversity, and no individual microbial taxa were differentially abundant between cases and control, after accounting for multiple comparisons. Among never smokers, there were elevated ORs for known oral pathogens: Porphyromonas gingivalis (OR = 1.69, 95% CI: 0.80-3.56), Prevotella intermedia (OR = 1.40, 95% CI: 0.69-2.85), and Tannerella forsythia (OR = 1.36, 95% CI: 0.66-2.77). CONCLUSIONS: Previously reported associations between oral taxa and pancreatic cancer were not present in this African-American population overall.

Multivariable association discovery in population-scale meta-omics studies.

It is challenging to associate features such as human health outcomes, diet, environmental conditions, or other metadata to microbial community measurements, due in part to their quantitative properties. Microbiome multi-omics are typically noisy, sparse (zero-inflated), high-dimensional, extremely non-normal, and often in the form of count or compositional measurements. Here we introduce an optimized combination of novel and established methodology to assess multivariable association of microbial community features with complex metadata in population-scale observational studies. Our approach, MaAsLin 2 (Microbiome Multivariable Associations with Linear Models), uses generalized linear and mixed models to accommodate a wide variety of modern epidemiological studies, including cross-sectional and longitudinal designs, as well as a variety of data types (e.g., counts and relative abundances) with or without covariates and repeated measurements. To construct this method, we conducted a large-scale evaluation of a broad range of scenarios under which straightforward identification of meta-omics associations can be challenging. These simulation studies reveal that MaAsLin 2's linear model preserves statistical power in the presence of repeated measures and multiple covariates, while accounting for the nuances of meta-omics features and controlling false discovery. We also applied MaAsLin 2 to a microbial multi-omics dataset from the Integrative Human Microbiome (HMP2) project which, in addition to reproducing established results, revealed a unique, integrated landscape of inflammatory bowel diseases (IBD) across multiple time points and omics profiles.

The Sulfur Microbial Diet and Risk of Colorectal Cancer by Molecular Subtypes and Intratumoral Microbial Species in Adult Men.

INTRODUCTION: We recently described the sulfur microbial diet, a pattern of intake associated with increased gut sulfur-metabolizing bacteria and incidence of distal colorectal cancer (CRC). We assessed whether this risk differed by CRC molecular subtypes or presence of intratumoral microbes involved in CRC pathogenesis (Fusobacterium nucleatum and Bifidobacterium spp.). METHODS: We performed Cox proportional hazards modeling to examine the association between the sulfur microbial diet and incidence of overall and distal CRC by molecular and microbial subtype in the Health Professionals Follow-Up Study (1986-2012). RESULTS: We documented 1,264 incident CRC cases among 48,246 men, approximately 40% of whom had available tissue data. After accounting for multiple hypothesis testing, the relationship between the sulfur microbial diet and CRC incidence did not differ by subtype. However, there was a suggestion of an association by prostaglandin synthase 2 (PTGS2) status with a multivariable adjusted hazard ratio for highest vs lowest tertile of sulfur microbial diet scores of 1.31 (95% confidence interval: 0.99-1.74, Ptrend = 0.07, Pheterogeneity = 0.04) for PTGS2-high CRC. The association of the sulfur microbial diet with distal CRC seemed to differ by the presence of intratumoral Bifidobacterium spp. with an adjusted hazard ratio for highest vs lowest tertile of sulfur microbial diet scores of 1.65 (95% confidence interval: 1.14-2.39, Ptrend = 0.01, Pheterogeneity = 0.03) for Bifidobacterium-negative distal CRC. We observed no apparent heterogeneity by other tested molecular markers. DISCUSSION: Greater long-term adherence to the sulfur microbial diet could be associated with PTGS2-high and Bifidobacterium-negative distal CRC in men. Additional studies are needed to further characterize the role of gut microbial sulfur metabolism and CRC.

New Approaches to Profile the Microbiome for Treatment of Neurodegenerative Disease.

Progressive neurodegenerative diseases represent some of the largest growing treatment challenges for public health in modern society. These diseases mainly progress due to aging and are driven by microglial surveillance and activation in response to changes occurring in the aging brain. The lack of efficacious treatment options for Alzheimer's disease (AD), as the focus of this review, and other neurodegenerative disorders has encouraged new approaches to address neuroinflammation for potential treatments. Here we will focus on the increasing evidence that dysbiosis of the gut microbiome is characterized by inflammation that may carry over to the central nervous system and into the brain. Neuroinflammation is the common thread associated with neurodegenerative diseases, but it is yet unknown at what point and how innate immune function turns pathogenic for an individual. This review will address extensive efforts to identify constituents of the gut microbiome and their neuroactive metabolites as a peripheral path to treatment. This approach is still in its infancy in substantive clinical trials and requires thorough human studies to elucidate the metabolic microbiome profile to design appropriate treatment strategies for early stages of neurodegenerative disease. We view that in order to address neurodegenerative mechanisms of the gut, microbiome and metabolite profiles must be determined to pre-screen AD subjects prior to the design of specific, chronic titrations of gut microbiota with low-dose antibiotics. This represents an exciting treatment strategy designed to balance inflammatory microglial involvement in disease progression with an individual's manifestation of AD as influenced by a coercive inflammatory gut.

Plant-Based Diet Index and Metabolic Risk in Men: Exploring the Role of the Gut Microbiome.

BACKGROUND: Healthy plant-based diet index (hPDI) is associated with a lower risk of cardiometabolic conditions, but its association as well as interactions with microbiome have not been elucidated. OBJECTIVES: We aimed to investigate the interrelations between hPDI, gut microbiome, and cardiometabolic risk markers. METHODS: hPDI was derived from dietary assessments by a validated FFQ and was examined in relation to metagenomic profiles of 911 fecal samples collected from 303 men aged 71 ± 4 y with an average BMI (in kg/m2) of 25.2 ± 3.6 in the Men's Lifestyle Validation Study. Principal coordinate (PCo) analysis based on Bray-Curtis dissimilarity was conducted, and interactions between hPDI and PCo were examined by using a metabolic risk score composed of blood lipids, BMI, and glycated hemoglobin. RESULTS: After multivariable adjustment, hPDI was significantly associated with the relative abundance of 7 species and 9 pathways. In particular, higher hPDI was significantly associated with a higher relative abundance of Bacteroides cellulosilyticus and Eubacterium eligens, amino acid biosynthesis pathways (l-isoleucine biosynthesis I and III and l-valine biosynthesis), and the pathway of pyruvate fermentation to isobutanol. A favorable association between hPDI and the metabolic risk score was more pronounced among men with a higher PCo characterized by higher abundance of Bacteroides uniformis and lower abundance of Prevotella copri. At the individual species level, a similar interaction was also observed between hPDI and P. copri, as well as with Clostridium clostridioforme or Blautia hydrogenotrophica (all P-interaction < 0.01). CONCLUSION: A greater adherence to a healthy plant-based diet by older men was associated with a microbial profile characterized by a higher abundance of multiple species, including B. cellulosilyticus and E. eligens, as well as pathways in amino acid metabolism and pyruvate fermentation. In addition, inverse associations between healthy plant-based diet and human metabolic risk may partially depend on microbial compositions.

A framework for microbiome science in public health.

Human microbiome science has advanced rapidly and reached a scale at which basic biology, clinical translation and population health are increasingly integrated. It is thus now possible for public health researchers, practitioners and policymakers to take specific action leveraging current and future microbiome-based opportunities and best practices. Here we provide an outline of considerations for research, education, interpretation and scientific communication concerning the human microbiome and public health. This includes guidelines for population-scale microbiome study design; necessary physical platforms and analysis methods; integration into public health areas such as epidemiology, nutrition, chronic disease, and global and environmental health; entrepreneurship and technology transfer; and educational curricula. Particularly in the near future, there are both opportunities for the incorporation of microbiome-based technologies into public health practice, and a growing need for policymaking and regulation around related areas such as prebiotic and probiotic supplements, novel live-cell therapies and fecal microbiota transplants.

Overview of the Microbiome Among Nurses study (Micro-N) as an example of prospective characterization of the microbiome within cohort studies.

A lack of prospective studies has been a major barrier for assessing the role of the microbiome in human health and disease on a population-wide scale. To address this significant knowledge gap, we have launched a large-scale collection targeting fecal and oral microbiome specimens from 20,000 women within the Nurses' Health Study II cohort (the Microbiome Among Nurses study, or Micro-N). Leveraging the rich epidemiologic data that have been repeatedly collected from this cohort since 1989; the established biorepository of archived blood, urine, buccal cell, and tumor tissue specimens; the available genetic and biomarker data; the cohort's ongoing follow-up; and the BIOM-Mass microbiome research platform, Micro-N furnishes unparalleled resources for future prospective studies to interrogate the interplay between host, environmental factors, and the microbiome in human health. These prospectively collected materials will provide much-needed evidence to infer causality in microbiome-associated outcomes, paving the way toward development of microbiota-targeted modulators, preventives, diagnostics and therapeutics. Here, we describe a generalizable, scalable and cost-effective platform used for stool and oral microbiome specimen and metadata collection in the Micro-N study as an example of how prospective studies of the microbiome may be carried out.

A 28 Day Clinical Assessment of a Lactic Acid-containing Antimicrobial Intimate Gel Wash Formulation on Skin Tolerance and Impact on the Vulvar Microbiome.

While intimate feminine hygiene products are widely used as part of daily cleansing routines, little is known about how these products impact the vulvovaginal area and its microbiome stability. This 4 week clinical study assessed tolerance of a novel gel wash containing lactic acid (pH 4.2) for external daily use when used on the external genital area and its effects on skin moisturization, vulvar skin pH, and the vulvar microbiome. After a 7 day pre-study conditioning period, 36 healthy females in three balanced age groups (18-29, 30-44, and 45-55 years) used the gel wash to cleanse their external genital area (mons pubis and vulva) and entire body at least once per day for 28 days. Skin tolerance of the gel wash was assessed by the gynecologist. Effects of the gel wash on vulvar skin microbiota were studied by performing bacterial 16S rRNA and fungal internal transcribed spacer (ITS) microbial richness and diversity analysis. Based on gynecologic assessment after 28 days of use, the gel wash showed acceptable tolerance, with no signs of increased dryness, redness, edema, itching, stinging, or burning. Use of the gel wash was associated with a significant increase in both short-term (single application) and longer-term (daily use for 28 days) skin moisturization. There was no significant change in vulvar skin pH over time with daily product use, and the gel wash did not significantly affect the natural vulvar microbiome species richness or diversity for bacteria or fungi. Results showed that this gel wash is a mild, moisturizing cleanser that maintains the natural pH and microbial diversity of vulvar skin. To our knowledge, this was the first study to assess the effect of an antimicrobial feminine gel wash on the natural pH and vulvar microbiome habitat of the skin using bacterial 16S rRNA and fungal ITS genetic sequencing techniques.

The interleukin-33 receptor contributes to pulmonary responses to ozone in male mice: role of the microbiome.

BACKGROUND: Interleukin-33 is released in the airways following acute ozone exposure and has the ability to cause airway hyperresponsiveness, a defining feature of asthma. Ozone causes greater airway hyperresponsiveness in male than female mice. Moreover, sex differences in the gut microbiome account for sex differences in this response to ozone. The purpose of this study was to determine whether there were sex differences in the role of interleukin-33 in ozone-induced airway hyperresponsiveness and to examine the role of the microbiome in these events. METHODS: Wildtype mice and mice genetically deficient in ST2, the interleukin-33 receptor, were housed from weaning with either other mice of the same genotype and sex, or with mice of the same sex but opposite genotype. At 15 weeks of age, fecal pellets were harvested for 16S rRNA sequencing and the mice were then exposed to air or ozone. Airway responsiveness was measured and a bronchoalveolar lavage was performed 24 h after exposure. RESULTS: In same-housed mice, ozone-induced airway hyperresponsiveness was greater in male than female wildtype mice. ST2 deficiency reduced ozone-induced airway hyperresponsiveness in male but not female mice and abolished sex differences in the response to ozone. However, sex differences in the role of interleukin-33 were unrelated to type 2 cytokine release: ozone-induced increases in bronchoalveolar lavage interleukin-5 were greater in females than males and ST2 deficiency virtually abolished interleukin-5 in both sexes. Since gut microbiota contribute to sex differences in ozone-induced airway hyperresponsiveness, we examined the role of the microbiome in these ST2-dependent sex differences. To do so, we cohoused wildtype and ST2 deficient mice, a situation that allows for transfer of microbiota among cage-mates. Cohousing altered the gut microbial community structure, as indicated by 16S rRNA gene sequencing of fecal DNA and reversed the effect of ST2 deficiency on pulmonary responses to ozone in male mice. CONCLUSIONS: The data indicate that the interleukin-33 /ST2 pathway contributes to ozone-induced airway hyperresponsiveness in male mice and suggest that the role of interleukin-33 is mediated at the level of the gut microbiome.

Obligate anaerobes are abundant in human necrotizing soft tissue infection samples - a metagenomics analysis.

Necrotizing soft tissue infections (NSTIs) are associated with high morbidity and mortality and are increasing in incidence. Proper identification of the microbial causes of NSTIs is a crucial step in diagnosis and treatment, but the majority of data collected are culture based, which is biased against fastidious organisms, including obligate anaerobes. The goal of this study was to address this gap in knowledge by characterizing NSTI microbial communities through molecular diagnostics. We performed 16S rRNA sequencing on human NSTI samples and identified five genera most commonly found in NSTIs (Prevotella, Bacteroides, Peptoniphilus, Porphyromonas, and Enterococcus). We found that a >70% contribution of obligate anaerobes to the bacterial population distribution was associated with NSTI mortality, and that NSTI samples, from both survivors and non-survivors, had an increased relative abundance of gram negative bacteria compared to those of abscess patients. Based on our data, we conclude that obligate anaerobes are abundant in NSTIs and a high relative abundance of anaerobes is associated with a worse outcome. We recommend increasing anaerobe antibiotic coverage during the treatment of NSTIs even when anaerobes are not found by traditional clinical microbiology methods, and especially when there is a clinical suspicion for anaerobe involvement.

Traditional culture methods fail to detect principle pathogens in necrotising soft tissue infection: a case report.

OBJECTIVE: Necrotising soft tissue infections (NSTIs) progress rapidly and mortality remains high, ranging from 10% to 30%, representing a significant challenge for health professionals. Early accurate diagnosis is crucial because timely and aggressive surgical intervention remains the number one indicator for a better clinical outcome. Understanding the microbial background of NSTIs would aid early diagnosis. PRESENTATION: We present a case of NSTI, in a seemingly healthy adult male, originating from a tooth abscess. The NSTI progressed rapidly, and eventually covered the patient's chest and abdominal skin and underlying soft tissue. RESULTS: Traditional blood and tissue culture only found Group C Streptococcus where 16S sequencing detected abundant Prevotella spp., a more likely causal organisms of the NSTI. The use of antibiotics with the approriate anaerobe coverage, in combination with timely surgical intervention, contributed to the ultimate successful clinical outcome. Complete wound healing and successful graft was achieved within one month of diagnosis of the microbes present. CONCLUSION: While surgical intervention remains the most important consideration in treatment of NSTI, correct identifcation of the microbial flora could also contribute to successful treatment.

Efficacy of hyperbaric oxygen therapy in bacterial biofilm eradication.

OBJECTIVE: Chronic wounds typically require several concurrent therapies, such as debridement, pressure offloading, and systemic and/or topical antibiotics. The aim of this study was to examine the efficacy of hyperbaric oxygen therapy (HBOT) towards reducing or eliminating bacterial biofilms in vitro and in vivo. METHOD: Efficacy was determined using in vitro grown biofilms subjected directly to HBOT for 30, 60 and 90 minutes, followed by cell viability determination using propidium monoazide-polymerase chain reaction (PMA-PCR). The efficacy of HBOT in vivo was studied by searching our chronic patient wound database and comparing time-to-healing between patients who did and did not receive HBOT as part of their treatment. RESULTS: In vitro data showed small but significant decreases in cell viability at the 30- and 90-minute time points in the HBOT group. The in vivo data showed reductions in bacterial load for patients who underwent HBOT, and ~1 week shorter treatment durations. Additionally, in patients' chronic wounds there was a considerable emergence of anaerobic bacteria and fungi between intermittent HBOT treatments. CONCLUSION: The data demonstrate that HBOT does possess a certain degree of biofilm killing capability. Moreover, as an adjuvant to standard treatment, more favourable patient outcomes are achieved through a quicker time-to-healing which reduces the chance of complications. Furthermore, the data provided insights into biofilm adaptations to challenges presented by this treatment strategy which should be kept in mind when treating chronic wounds. Further studies will be necessary to evaluate the benefits and mechanisms of HBOT, not only for patients with chronic wounds but other chronic infections caused by bacterial biofilms.

Cadaver Thanatomicrobiome Signatures: The Ubiquitous Nature of Clostridium Species in Human Decomposition.

Human thanatomicrobiome studies have established that an abundant number of putrefactive bacteria within internal organs of decaying bodies are obligate anaerobes, Clostridium spp. These microorganisms have been implicated as etiological agents in potentially life-threatening infections; notwithstanding, the scale and trajectory of these microbes after death have not been elucidated. We performed phylogenetic surveys of thanatomicrobiome signatures of cadavers' internal organs to compare the microbial diversity between the 16S rRNA gene V4 hypervariable region and V3-4 conjoined regions from livers and spleens of 45 cadavers undergoing forensic microbiological studies. Phylogenetic analyses of 16S rRNA gene sequences revealed that the V4 region had a significantly higher mean Chao1 richness within the total microbiome data. Permutational multivariate analysis of variance statistical tests, based on unweighted UniFrac distances, demonstrated that taxa compositions were significantly different between V4 and V3-4 hypervariable regions (p < 0.001). Of note, we present the first study, using the largest cohort of criminal cases to date, that two hypervariable regions show discriminatory power for human postmortem microbial diversity. In conclusion, here we propose the impact of hypervariable region selection for the 16S rRNA gene in differentiating thanatomicrobiomic profiles to provide empirical data to explain a unique concept, the Postmortem Clostridium Effect.