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The composition of the gut microbiota plays an important role in maintaining the balance between health and disease. However, there is considerably less information on the composition of the gut microbiota of non-Western communities. This study was designed to investigate the evolution in the gut microbiota in a cohort of Nigerian infants within the first year of life. Faecal samples were obtained monthly from 28 infants from birth for one year. The infants had been born by a mix of natural birth and caesarean section and were either breast-fed or mixed fed. Sequencing of the V1-V2 region of the 16S rRNA gene was used to characterise the microbiota. Short chain fatty acids and lactate present in each faecal sample were identified by gas chromatography. Microbial differences were observed between the vaginal and caesarean section delivered infants in samples collected within 7 days of life, although these differences were not observed in later samples. Exclusively breastfed infants had predominance of Ruminococcus gnavus , Collinsella , and Sutterella species. Different Bifidobacterium species dominated breast-fed compared to mixed fed infants. Clostridium , Enterococcus , Roseburia , and Coprococcus species were observed once the infants commenced weaning. Butyrate was first detected when weaning started between months 4–6 in the majority of the infants while total short chain fatty acid concentrations increased, and acetate and lactate remained high following the introduction of solid foods. The observed taxonomic differences in the gut microbiota between Nigerian infants, as well as butyrate production during weaning, were strongly influenced by diet, and not by birthing method. Introduction of local/solid foods encouraged the colonisation and evolution of specific marker organisms associated with carbohydrate metabolism.

Background Characterisation of the bacterial composition of the gut microbiota is increasingly carried out with a view to establish the role of different bacterial species in causation or prevention of disease. It is thus essential that the methods used to determine the microbial composition are robust. Here, several widely used molecular techniques were compared to establish the optimal methods to assess the bacterial composition in faecal samples from babies, before weaning. Results The bacterial community profile detected in the faeces of infants is highly dependent on the methodology used. Bifidobacteria were the most abundant bacteria detected at 6 weeks in faeces from two initially breast-fed babies using fluorescent in situ hybridisation (FISH), in agreement with data from previous culture-based studies. Using the 16S rRNA gene sequencing approach, however, we found that the detection of bifidobacteria in particular crucially depended on the optimisation of the DNA extraction method, and the choice of primers used to amplify the V1–V3 regions of 16S rRNA genes prior to subsequent sequence analysis. Bifidobacteria were only well represented among amplified 16S rRNA gene sequences when mechanical disruption (bead-beating) procedures for DNA extraction were employed together with optimised “universal” PCR primers. These primers incorporate degenerate bases at positions where mismatches to bifidobacteria and other bacterial taxa occur. The use of a DNA extraction kit with no bead-beating step resulted in a complete absence of bifidobacteria in the sequence data, even when using the optimised primers. Conclusions This work emphasises the importance of sample processing methodology to downstream sequencing results and illustrates the value of employing multiple approaches for determining microbiota composition.

The gut microbiota has recently been proposed as a novel component in the regulation of host homeostasis and immunity. We have assessed for the first time the role of the gut microbiota in a mouse model of leukemia (transplantation of BaF3 cells containing ectopic expression of Bcr-Abl), characterized at the final stage by a loss of fat mass, muscle atrophy, anorexia and inflammation. The gut microbial 16S rDNA analysis, using PCR-Denaturating Gradient Gel Electrophoresis and quantitative PCR, reveals a dysbiosis and a selective modulation of Lactobacillus spp. (decrease of L. reuteri and L. johnsonii/gasseri in favor of L. murinus/animalis) in the BaF3 mice compared to the controls. The restoration of Lactobacillus species by oral supplementation with L. reuteri 100-23 and L. gasseri 311476 reduced the expression of atrophy markers (Atrogin-1, MuRF1, LC3, Cathepsin L) in the gastrocnemius and in the tibialis, a phenomenon correlated with a decrease of inflammatory cytokines (interleukin-6, monocyte chemoattractant protein-1, interleukin-4, granulocyte colony-stimulating factor, quantified by multiplex immuno-assay). These positive effects are strain- and/or species-specific since L. acidophilus NCFM supplementation does not impact on muscle atrophy markers and systemic inflammation. Altogether, these results suggest that the gut microbiota could constitute a novel therapeutic target in the management of leukemia-associated inflammation and related disorders in the muscle.

Background Faecal samples are frequently used to characterise the gut microbiota in health and disease, yet there is considerable debate about how representative faecal bacterial profiles are of the overall gut community. A particular concern is whether bacterial populations associated with the gut mucosa are properly represented in faecal samples, since these communities are considered critical in the aetiology of gastrointestinal diseases. In this study we compared the profiles of the faecal and mucosal microbiota from ten healthy volunteers using bacterial culturing (culturomics) and next-generation sequencing targeting the 16S ribosomal nucleic acid (rRNA) gene. Paired fresh rectal biopsies and faecal samples were processed under stringent anaerobic conditions to maintain the viability of the bacteria. Four different sample types were analysed: faecal (F), faecal homogenised (FHg), biopsy tissue (B) and biopsy wash (BW) samples.  Results There were no significant statistical differences in either bacterial richness or diversity between biopsy washes (BW) and faecal (F) or faecal homogenised (FHg) samples. Principal coordinates analysis of a Bray–Curtis distance matrix generated from sequence variant tables did not show distinct clustering between these samples (PERMANOVA; p  = 0.972) but showed strong clustering of samples from individual donors. However, the rectal biopsy tissue (B) samples had a significantly altered bacterial signature with greater abundance of Proteobacteria and Acidobacteria compared to faecal (F) and faecal homogenised (FHg) samples. A total of 528 bacteria encompassing 92 distinct bacterial species were isolated and cultured from a subset of six volunteer samples (biopsy washes and faeces). This included isolation of 22 novel bacterial species. There was significant similarity between the bacterial species grown in anaerobic culture and those identified by 16S rRNA gene sequencing (Spearman correlation; rho = 0.548, p  = 0.001). Conclusion This study showed that the bacterial profiles of paired faecal and rectal biopsy wash samples were very similar, validating the use of faecal samples as a convenient surrogate for rectal biopsy-associated microbiota. Anaerobic bacterial culture results showed similar taxonomic patterns to the amplicon sequence analysis disproving the dogma that culture-based methods do not reflect findings of molecular assessments of gut bacterial composition. FGviAy1dgRvRfr2cUx9EBx Video abstract

Background/Objectives: Alzheimer’s disease (AD) is the most common form of dementia, characterized by an irreversible decline in cognitive function. The pathogenesis of several neurodegenerative disorders has been linked to changes in the gut microbiota, transmitted through the gut-brain axis. Methods: We set out to establish by case-control study methodology whether there were any differences in the composition and/or function of the gut microbiota between older resident adults in care homes with or without an AD diagnosis via analysis of the microbial composition from fecal samples. Results: The microbial composition, determined by 16S rRNA gene profiling, indicated that AD sufferers had significantly increased proportions of Escherichia/Shigella and Clostridium_sensu_stricto_1, and significantly decreased proportions of Bacteroides, Faecalibacterium, Blautia, and Roseburia species. The increase in potentially pro-inflammatory bacteria was consistent with slightly higher concentrations of calprotectin, a biomarker of gut inflammation. Fecal concentrations of most microbial metabolites measured were similar across groups, although participants with AD had significantly increased proportions of the branched-chain fatty acid, iso-butyrate, and lower overall concentrations of total short chain fatty acids. Conclusions: Participants with Alzheimer’s disease have several key differences within their gut microbiota profile, in contrast to care home residents without Alzheimer’s disease. The altered microbiome included both compositional and functional changes linked to poorer health and gut inflammation.

The human gut harbours a wide range of bacterial communities that play key roles in supplying nutrients and energy to the host through anaerobic fermentation of dietary components and host secretions. This fermentative process involves different functional groups of microorganisms linked in a trophic chain. Although the diversity of the intestinal microbiota has been studied extensively using molecular techniques, the functional aspects of this biodiversity remain mostly unexplored. The aim of the present work was to enumerate the principal metabolic groups of microorganisms involved in the fermentative process in the gut of healthy humans. These functional groups of microorganisms were quantified by a cultural approach, while the taxonomic composition of the microbiota was assessed by in situ hybridization on the same faecal samples. The functional groups of microorganisms that predominated in the gut were the polysaccharide-degrading populations involved in the breakdown of the most readily available exogenous and endogenous substrates and the predominant butyrate-producing species. Most of the functional groups of microorganisms studied appeared to be present at rather similar levels in all healthy volunteers, suggesting that optimal numbers of these various bacterial groups are crucial for efficient gut fermentation, as well as for host nutrition and health. Significant interindividual differences were, however, confirmed with respect to the numbers of methanogenic archaea, filter paper-degrading and acetogenic bacteria and the products formed by lactate-utilizing bacteria.

A patient presented with fever, generalised rash, confusion, orofacial movements and myoclonus after receiving the first dose of mRNA-1273 vaccine from Moderna. MRI was unremarkable while cerebrospinal fluid showed leucocytosis with lymphocyte predominance and hyperproteinorrachia. The skin evidenced red, non-scaly, oedematous papules coalescing into plaques with scattered non-follicular pustules. Skin biopsy was consistent with a neutrophilic dermatosis. The patient fulfilled the criteria for Sweet syndrome. A thorough evaluation ruled out alternative infectious, autoimmune or malignant aetiologies, and all manifestations resolved with glucocorticoids. While we cannot prove causality, there was a temporal correlation between the vaccination and the clinical findings.

Testing 21 different fecal DNA extraction protocols in multiple laboratories results in a standardized protocol with the potential to improve comparability across human gut microbiome studies. Technical variation in metagenomic analysis must be minimized to confidently assess the contributions of microbiota to human health. Here we tested 21 representative DNA extraction protocols on the same fecal samples and quantified differences in observed microbial community composition. We compared them with differences due to library preparation and sample storage, which we contrasted with observed biological variation within the same specimen or within an individual over time. We found that DNA extraction had the largest effect on the outcome of metagenomic analysis. To rank DNA extraction protocols, we considered resulting DNA quantity and quality, and we ascertained biases in estimates of community diversity and the ratio between Gram-positive and Gram-negative bacteria. We recommend a standardized DNA extraction method for human fecal samples, for which transferability across labs was established and which was further benchmarked using a mock community of known composition. Its adoption will improve comparability of human gut microbiome studies and facilitate meta-analyses.

Based on responses to controlled diagnostic blocks of cervical facet joints, the prevalence of cervical facet joint pain in chronic neck pain has been shown to range from 54% to 67%, with false-positive results of 27% to 63% with a single diagnostic block. Other confounding factors claimed to influence the diagnostic validity of cervical facet joint blocks include administration of anxiolytics and narcotics prior to or during the procedure. To evaluate the effect of midazolam and fentanyl on the validity of diagnosis of cervical facet joint pain. Randomized, prospective, double-blind, placebo-controlled evaluation. The study was undertaken in an interventional pain management practice. The design consisted of a placebo group receiving sodium chloride solution and two experimental groups receiving either midazolam or fentanyl. The patients included in the study were treated in the past and were presenting for repeat treatment after a significant period of symptom relief. Outcomes were assessed at baseline and after the administration of 1 of the 3 solutions (Group I, sodium chloride solution; Group II, midazolam; or Group III, fentanyl). Outcome measures included numeric pain scale, proportion of pain relief, and ability to perform prior painful movements. Pain relief of > or = 80% was noted in 5% of the patients in Group I, 8% in Group II, and 8% in Group III. However, > or = 50% relief was noted in 8% of the patients in Group I, 13% in Group II, and 27% in Group III. Overall, 8% of the patients in Group I, 13% in Group II, and 27% in Group III were able to perform movements which were painful prior to injection. The administration of sedation with midazolam or fentanyl is a confounding factor in the diagnosis of cervical facet joint pain in patients with chronic neck pain. However, if > or = 80% pain relief with ability to perform prior painful movements is used as the standard for evaluating the effect of controlled local anesthetic blocks, the diagnostic validity of cervical facet joint nerve blocks may be preserved.

Zygapophysial or facet joint pain in patients suffering with chronic spinal pain without disc herniation or radiculopathy may be diagnosed with certainty by the use of controlled diagnostic blocks. But, in patients suffering with either lumbar or cervical facet joint pain, even this diagnostic approach may be confounded by false-positives when using a single diagnostic block. It may also be confounded by the administration of anxiolytics and narcotics prior to, or during, the controlled diagnostic facet joint blocks. The effect of sedation on the validity and potential differential results in patients suffering with combined cervical and lumbar facet joint pain has not been evaluated. To assess the effects of midazolam and fentanyl on the diagnostic validity of facet joint blocks in patients suffering with both cervical and lumbar facet joint pain. Randomized, double-blind, placebo-controlled study. The design consisted of a placebo group receiving a sodium chloride solution and two experimental groups receiving either midazolam or fentanyl. Patients included in the study had been diagnosed with facet joint pain using controlled comparative local anesthetic blocks of the medial branches and L5 dorsal rami. They had been treated with lumbar and cervical facet joint nerve blocks and experienced good pain relief; and were presenting for repeat treatment after a period of symptom relief. The study was performed in an interventional pain management practice in the United States; a total of 60 patients participated with 20 patients randomly allocated into each group. Outcome measures included numeric pain scores, proportion of pain relief, and ability to perform prior painful movements. Outcomes were assessed at baseline and after the administration of 1 of the 3 solutions (Group I, sodium chloride solution; Group II, midazolam; or Group III, fentanyl). Overall, 50% of the patients were relaxed or sedated in the placebo group, while 100% of the patients in the midazolam and fentanyl groups were relaxed or sedated. As many as 10% of the patients reported significant relief (>= 80%) with the ability to perform prior painful movements. Perioperative administration of sodium chloride, midazolam, or fentanyl can confound results in the diagnosis of combined cervical and lumbar facet joint pain. False-positive results with placebo or sedation may be seen in a small proportion of patients.