LongDat: an R package for covariate-sensitive longitudinal analysis of high-dimensional data

We introduce LongDat, an R package that analyzes longitudinal multivariable (cohort) data while simultaneously accounting for a potentially large number of covariates. The primary use case is to differentiate direct from indirect effects of an intervention (or treatment) and to identify covariates (potential mechanistic intermediates) in longitudinal data. LongDat focuses on analyzing longitudinal microbiome data, but its usage can be expanded to other data types, such as binary, categorical, and continuous data. We tested and compared LongDat with other tools (i.e., MaAsLin2, ANCOM, lgpr, and ZIBR) on both simulated and real data. We showed that LongDat outperformed these tools in accuracy, runtime, and memory cost, especially when there were multiple covariates. The results indicate that the LongDat R package is a computationally efficient and low-memory-cost tool for longitudinal data with multiple covariates and facilitates robust biomarker searches in high-dimensional datasets

DNA sonication inverse PCR for genome scale analysis of uncharacterized flanking sequences

1. There are few available tools to comprehensively and economically identify uncharacterized flanking regions that are not extremely labour intensive and which exploit the advantages of emerging long‐read sequencing platforms. 2. We describe SIP; a sonication‐based inverse PCR high‐throughput sequencing strategy to investigate uncharacterized flanking region sequences, including those flanking mobile DNA. SIP combines unbiased fragmentation by sonication and target enrichment by coupling outward facing PCR priming with long‐read sequencing technologies. 3. We demonstrate the effectiveness of SIP by determining retroviral integrations which are high copy and challenging to characterize. We further describe SIP's workflow, examine retroviral (proviral) enrichment and characterize viral structural variants identified. When SIP was coupled with long‐read sequencing using the PacBio RS II platform, proviral integration was extensively characterized at high sequence depth per integration. By interrogating the sequence data, we were also able to test several intrinsic factors including SIP's propensity to form chimeric sequences and adapter ligation efficiencies. 4. SIP is an adaption of a traditional molecular biology technique that can be used to characterize any unknown genomic flanking sequence or to extend any sequence for which only minimal sequence information is available. SIP can be applied broadly to study complex biological systems such as mobile genetic elements with high throughput

The effectiveness of two silicone dressings for sacral and heel pressure ulcer prevention compared with no dressings in high‐risk intensive care unit patients: a randomized controlled parallel‐group trial

Background There is a high incidence of pressure ulcers in high-risk settings such as intensive care. There is emerging evidence that the application of dressings to pressure ulcer predilection areas (sacrum and heels) improves prevention strategies. Objectives To determine whether preventive dressings, applied to the sacrum and heels of high-risk patients in intensive care units, in addition to standard prevention, reduces the incidence of pressure ulcers. Methods Between June 2015 and July 2018, a randomized, controlled, two-arm, superiority pragmatic study was performed with a concealed 1 : 1 allocation to the intervention and control group. Patients assigned to the intervention group had dressings applied to the sacrum and heels. Results In total, 7575 patients were screened for eligibility and 475 patients were included and allocated to both groups. Finally, 212 patients in the intervention group and 210 in the control group were analysed. The mean age was 63 center dot 5 years and the majority of patients were male (65 center dot 4%). The cumulative pressure ulcer incidence category II and above was 2 center dot 8% in the intervention, and 10 center dot 5% in the control group (P = 0 center dot 001). Compared with the control group, the relative risk in the intervention group was 0 center dot 26 [95% confidence interval (CI) 0 center dot 11-0 center dot 62] and the absolute risk reduction was 0 center dot 08 (95% CI 0 center dot 03-0 center dot 13). Conclusions The results indicate that the application of dressings, in addition to standard prevention, in high-risk intensive care unit patients is effective in preventing pressure ulcers at the heels and sacrum. What's already known about this topic? Pressure ulcers are severe soft tissue injuries and wounds, which occur worldwide in all healthcare settings. Despite preventive interventions, pressure ulcers still develop. There is emerging evidence that dressings help to prevent pressure ulcers. What does this study add? The incidence of pressure ulcers in intensive care units among high-risk patients remains high. The application of dressings to the sacrum and heels, in addition to standard preventive measures, reduces the relative and absolute risks for the development of pressure ulcers. The application of preventive dressings at the heels and sacrum seems to be feasible in intensive care settings

The gut microbiome in Myalgic Encephalomyelitis (ME)/Chronic Fatigue Syndrome (CFS)

Myalgic encephalomyelitis (ME) or Chronic Fatigue Syndrome (CFS) is a neglected, debilitating multi-systemic disease without diagnostic marker or therapy. Despite evidence for neurological, immunological, infectious, muscular and endocrine pathophysiological abnormalities, the etiology and a clear pathophysiology remains unclear. The gut microbiome gained much attention in the last decade with manifold implications in health and disease. Here we review the current state of knowledge on the interplay between ME/CFS and the microbiome, to identify potential diagnostic or interventional approaches, and propose areas where further research is needed. We iteratively selected and elaborated on key theories about a correlation between microbiome state and ME/CFS pathology, developing further hypotheses. Based on the literature we hypothesize that antibiotic use throughout life favours an intestinal microbiota composition which might be a risk factor for ME/CFS. Main proposed pathomechanisms include gut dysbiosis, altered gut-brain axis activity, increased gut permeability with concomitant bacterial translocation and reduced levels of short-chain-fatty acids, D-lactic acidosis, an abnormal tryptophan metabolism and low activity of the kynurenine pathway. We review options for microbiome manipulation in ME/CFS patients including probiotic and dietary interventions as well as fecal microbiota transplantations. Beyond increasing gut permeability and bacterial translocation, specific dysbiosis may modify fermentation products, affecting peripheral mitochondria. Considering the gut-brain axis we strongly suspect that the microbiome may contribute to neurocognitive impairments of ME/CFS patients. Further larger studies are needed, above all to clarify whether D-lactic acidosis and early-life antibiotic use may be part of ME/CFS etiology and what role changes in the tryptophan metabolism might play. An association between the gut microbiome and the disease ME/CFS is plausible. As causality remains unclear, we recommend longitudinal studies. Activity levels, bedridden hours and disease progression should be compared to antibiotic exposure, drug intakes and alterations in the composition of the microbiota. The therapeutic potential of fecal microbiota transfer and of targeted dietary interventions should be systematically evaluated

The effect of probiotic and synbiotic supplementation on appetite-regulating hormones and desire to eat: a systematic review and meta-analysis of clinical trials

UNLABELLED: Recent studies have demonstrated the effect of probiotics, prebiotics, and synbiotics on adiponectin and leptin levels; however, those findings remain contested. The present study aimed to explore the impact of probiotics/synbiotics on appetite-regulating hormones and the desire to eat. METHODS: A systematic review was conducted by searching the Medline (PubMed) and Scopus databases from inception to December 2021, using relevant keywords and MeSH terms, and appropriate randomized controlled trials (RCTs) were extracted. The standardized mean differences (SMD) and 95% confidence intervals (95%CIs) were calculated as part of the meta-analysis using a random-effect model to determine the mean effect sizes. Analysis of Galbraith plots and the Cochrane Chi-squared test were conducted to examine heterogeneity. RESULTS: Meta-analysis of data from a total of 26 RCTs (n = 1536) showed a significant decrease in serum/plasma leptin concentration following probiotic/synbiotic supplementation (SMD: -0.38, 95%CI= -0.638, -0.124); P-value= 0.004; I(2)= 69.4%; P heterogeneity <0.001). The leptin level decrease from probiotic/synbiotic supplementation was higher in patients with NAFLD than those with overweight/obesity or type 2 diabetes mellitus/ metabolic syndrome/ prediabetes. Probiotic/synbiotic supplementation was associated with a trending increase in adiponectin levels, stronger in patients with type 2 diabetes mellitus, metabolic syndrome, and prediabetes (SMD: 0.25, 95%CI= 0.04, 0.46) µg/mL; P-value=0.021; I(2)=16.8%; P heterogeneity= 0.30). Additionally, supplementation with probiotic/synbiotic was linked to a slight increase in desire to eat (SMD: 0.34, 95%CI= 0.03, 0.66) P-value = 0.030; I(2)=39.4%; P heterogeneity= 0.16). CONCLUSION: Our meta-analysis indicates a favorable impact of probiotic/synbiotic supplementation on regulating leptin and adiponectin secretion

Guts within guts: the microbiome of the intestinal helminth parasite Ascaris suum is derived but distinct from its host

BACKGROUND: Intestinal helminths are extremely prevalent among humans and animals. In particular, intestinal roundworms affect more than 1 billion people around the globe and are a major issue in animal husbandry. These pathogens live in intimate contact with the host gut microbiota and harbor bacteria within their own intestines. Knowledge of the bacterial host microbiome at the site of infection is limited, and data on the parasite microbiome is, to the best of our knowledge, non-existent. RESULTS: The intestinal microbiome of the natural parasite and zoonotic macropathogen, Ascaris suum was analyzed in contrast to the diversity and composition of the infected host gut. 16S sequencing of the parasite intestine and host intestinal compartments showed that the parasite gut has a significantly less diverse microbiome than its host, and the host gut exhibits a reduced microbiome diversity at the site of parasite infection in the jejunum. While the host's microbiome composition at the site of infection significantly determines the microbiome composition of its parasite, microbial signatures differentiate the nematodes from their hosts as the Ascaris intestine supports the growth of microbes that are otherwise under-represented in the host gut. CONCLUSION: Our data clearly indicate that a nematode infection reduces the microbiome diversity of the host gut, and that the nematode gut represents a selective bacterial niche harboring bacteria that are derived but distinct from the host gut

The gut microbiota is associated with the small intestinal paracellular permeability and the development of the immune system in healthy children during the first two years of life

BACKGROUND: The intestinal barrier plays an important role in the defense against infections, and nutritional, endocrine, and immune functions. The gut microbiota playing an important role in development of the gastrointestinal tract can impact intestinal permeability and immunity during early life, but data concerning this problem are scarce. METHODS: We analyzed the microbiota in fecal samples (101 samples in total) collected longitudinally over 24 months from 21 newborns to investigate whether the markers of small intestinal paracellular permeability (zonulin) and immune system development (calprotectin) are linked to the gut microbiota. The results were validated using data from an independent cohort that included the calprotectin and gut microbiota in children during the first year of life. RESULTS: Zonulin levels tended to increase for up to 6 months after childbirth and stabilize thereafter remaining at a high level while calprotectin concentration was high after childbirth and began to decline from 6 months of life. The gut microbiota composition and the related metabolic potentials changed during the first 2 years of life and were correlated with zonulin and calprotectin levels. Faecal calprotectin correlated inversely with alpha diversity (Shannon index, r = − 0.30, FDR P (Q) = 0.039). It also correlated with seven taxa; i.a. negatively with Ruminococcaceae (r = − 0.34, Q = 0.046), and Clostridiales (r = − 0.34, Q = 0.048) and positively with Staphylococcus (r = 0.38, Q = 0.023) and Staphylococcaceae (r = 0.35, Q = 0.04), whereas zonulin correlated with 19 taxa; i.a. with Bacillales (r = − 0.52, Q = 0.0004), Clostridiales (r = 0.48, Q = 0.001) and the Ruminococcus (torques group) (r = 0.40, Q = 0.026). When time intervals were considered only changes in abundance of the Ruminococcus (torques group) were associated with changes in calprotectin (β = 2.94, SE = 0.8, Q = 0.015). The dynamics of stool calprotectin was negatively associated with changes in two MetaCyc pathways: pyruvate fermentation to butanoate (β = − 4.54, SE = 1.08, Q = 0.028) and Clostridium acetobutylicum fermentation (β = − 4.48, SE = 1.16, Q = 0.026). CONCLUSIONS: The small intestinal paracellular permeability, immune system-related markers and gut microbiota change dynamically during the first 2 years of life. The Ruminococcus (torques group) seems to be especially involved in controlling paracellular permeability. Staphylococcus, Staphylococcaceae, Ruminococcaceae, and Clostridiales, may be potential biomarkers of the immune system. Despite observed correlations their clear causation and health consequences were not proven. Mechanistic studies are required

Dynamic changes in microbiome composition following Mare's milk intake for prevention of collateral antibiotic effect

INTRODUCTION: Probiotics and prebiotics are widely used for recovery of the human gut microbiome after antibiotic treatment. High antibiotic usage is especially common in children with developing microbiome. We hypothesized that dry Mare's milk, which is rich in biologically active substances without containing live bacteria, could be used as a prebiotic in promoting microbial diversity following antibiotic treatment in children. The present pilot study aims to determine the impacts of dry Mare's milk on the diversity of gut bacterial communities when administered during antibiotic treatment and throughout the subsequent recovery phase. METHODS: Six children aged 4 to 5 years and diagnosed with bilateral bronchopneumonia were prescribed cephalosporin antibiotics. During the 60 days of the study, three children consumed dry Mare's milk whereas the other three did not. Fecal samples were collected daily during antibiotic therapy and every 5 days after antibiotic therapy. Total DNA was isolated and taxonomic composition of gut microbiota was analyzed by 16S rRNA amplicon sequencing. To assess the immune status of the gut, stool samples were analyzed by bead-based multiplex assays. RESULTS: Mare's milk treatment seems to prevent the bloom of Mollicutes, while preventing the loss of Coriobacteriales. Immunological analysis of the stool reveals an effect of Mare’s milk on local immune parameters under the present conditions