Stratification of alopecia areata reveals involvement of CD4 T cell populations and altered faecal microbiota.

Alopecia areata (AA) is an immune-mediated disease that causes non-scarring hair loss. Autoreactive CD8 T cells are key pathogenic effectors in the skin, and AA has been associated both with atopy and with perturbations in intestinal homeostasis. This study aimed to investigate mechanisms driving AA by characterizing the circulating immunophenotype and faecal microbiome, and by stratifying AA to understand how identified signatures associated with heterogeneous clinical features of the condition. Flow cytometric analyses identified alterations in circulating B cells and CD4 T cells, while 16S sequencing identified changes in alpha and beta diversity in the faecal microbiome in AA. The proportions of transitional and naïve B cells were found to be elevated in AA, particularly in AA samples from individuals with >50% hair loss and those with comorbid atopy, which is commonly associated with extensive hair loss. Although significant changes in circulating CD8 T cells were not observed, we found significant changes in CD4+ populations. In individuals with <50% hair loss higher frequencies of CCR6+CD4 ("Th17") and CCR6+CXCR3+CD4 ("Th1/17") T cells were found. While microbial species richness was not altered, AA was associated with reduced evenness and Shannon diversity of the intestinal microbiota, again particularly in those with <50% hair loss. We have identified novel immunological and microbial signatures in individuals with alopecia areata. Surprisingly, these are associated with lower levels of hair loss, and may therefore provide a rationale for improved targeting of molecular therapeutics.

Drivers of ecological assembly in the hindgut of Atlantic Cod fed a macroalgal supplemented diet.

It is difficult to disentangle the many variables (e.g. internal or external cues and random events) that shape the microbiota in the gastrointestinal tract of any living species. Ecological assembly processes applied to microbial communities can elucidate these drivers. In our study, farmed Atlantic cod (Gadus morhua) were fed a diet of 10% macroalgae supplement (Ulva rigida [ULVA] or Ascophyllum nodosum [ASCO] or a non-supplemented control diet [CTRL]) over 12 weeks. We determined the influence of ecological assembly processes using a suite of null-modelling tools. We observed dissimilarity in the abundance of common OTUs over time, which was driven by deterministic assembly. The CTRL samples showed selection as a critical assembly process. While dispersal limitation was a driver of the gut microbiome for fish fed the macroalgae supplemented diet at Week 12 (i.e., ASCO and ULVA). Fish from the ASCO grouping diverged into ASCO_N (normal) and ASCO_LG (lower growth), where ASCO_LG individuals found the diet unpalatable. The recruitment of new taxa overtime was altered in the ASCO_LG fish, with the gut microbiome showing phylogenetic underdispersion (nepotistic species recruitment). Finally, the gut microbiome (CTRL and ULVA) showed increasing robustness to taxonomic disturbance over time and lower functional redundancy. This study advances our understanding of the ecological assembly and succession in the hindgut of juvenile Atlantic cod across dietary treatments. Understanding the processes driving ecological assembly in the gut microbiome, in fish research specifically, could allow us to manipulate the microbiome for improved health or resilience to disease for improved aquaculture welfare and production.

Genome erosion and evidence for an intracellular niche - exploring the biology of mycoplasmas in Atlantic salmon.

Mycoplasmas are the smallest autonomously self-replicating life form on the planet. Members of this bacterial genus are known to parasitise a wide array of metazoans including vertebrates. Whilst much research has been significant targeted at parasitic mammalian mycoplasmas, very little is known about their role in other vertebrates. In the current study, we aim to explore the biology of mycoplasmas in Atlantic Salmon, a species of major significance for aquaculture, including cellular niche, genome size structure and gene content. Using fluorescent in-situ hybridisation (FISH), mycoplasmas were targeted in epithelial tissues across the digestive tract (stomach, pyloric caecum and midgut) from different development stages (eggs, parr, subadult) of farmed Atlantic salmon (Salmo salar), and we present evidence for an intracellular niche for some of the microbes visualised. Via shotgun metagenomic sequencing, a nearly complete, albeit small, genome (~0.57 MB) as assembled from a farmed Atlantic salmon subadult. Phylogenetic analysis of the recovered genome revealed taxonomic proximity to other salmon derived mycoplasmas, as well as to the human pathogen Mycoplasma penetrans (~1.36 Mb). We annotated coding sequences and identified riboflavin pathway encoding genes and sugar transporters, the former potentially consistent with micronutrient provisioning in salmonid development. Our study provides insights into mucosal adherence, the cellular niche and gene catalog of Mycoplasma in the gut ecosystem of the Atlantic salmon, suggesting a high dependency of this minimalist bacterium on its host. Further study is required to explore and functional role of Mycoplasma in the nutrition and development of its salmonid host.

Temporal changes in the gut microbiota in farmed Atlantic cod (Gadus morhua) outweigh the response to diet supplementation with macroalgae.

BACKGROUND: Aquaculture successfully meets global food demands for many fish species. However, aquaculture production of Atlantic cod (Gadus morhua) is just 2.5% of total market production. For cod farming to be a viable economic venture specific challenges on how to increase growth, health and farming productivity need to be addressed. Feed ingredients play a key role here. Macroalgae (seaweeds) have been suggested as a functional feed supplement with both health and economic benefits for terrestrial farmed animals and fish. The impact of such dietary supplements to cod gut integrity and microbiota, which contribute to overall fish robustness is unknown. The objective of this study was to supplement the diet of juvenile Atlantic cod with macroalgae and determine the impacts on fish condition and growth, gut morphology and hindgut microbiota composition (16S rRNA amplicon sequencing). Fish were fed one of three diets: control (no macroalgal inclusion), 10% inclusion of either egg wrack (Ascophyllum nodosum) or sea lettuce (Ulva rigida) macroalgae in a 12-week trial. RESULTS: The results demonstrated there was no significant difference in fish condition, gut morphology or hindgut microbiota between the U. rigida supplemented fish group and the control group at any time-point. This trend was not observed with the A. nodosum treatment. Fish within this group were further categorised as either 'Normal' or 'Lower Growth'. 'Lower Growth' individuals found the diet unpalatable resulting in reduced weight and condition factor combined with an altered gut morphology and microbiome relative to the other treatments. Excluding this group, our results show that the hindgut microbiota was largely driven by temporal pressures with the microbial communities becoming more similar over time irrespective of dietary treatment. The core microbiome at the final time-point consisted of the orders Vibrionales (Vibrio and Photobacterium), Bacteroidales (Bacteroidetes and Macellibacteroides) and Clostridiales (Lachnoclostridium). CONCLUSIONS: Our study indicates that U. rigida macroalgae can be supplemented at 10% inclusion levels in the diet of juvenile farmed Atlantic cod without any impact on fish condition or hindgut microbial community structure. We also conclude that 10% dietary inclusion of A. nodosum is not a suitable feed supplement in a farmed cod diet.

Neutral Processes Dominate Microbial Community Assembly in Atlantic Salmon, Salmo salar.

In recent years, a wealth of studies has examined the relationships between a host and its microbiome across diverse taxa. Many studies characterize the host microbiome without considering the ecological processes that underpin microbiome assembly. In this study, the intestinal microbiota of Atlantic salmon, Salmo salar, sampled from farmed and wild environments was first characterized using 16S rRNA gene MiSeq sequencing analysis. We used neutral community models to determine the balance of stochastic and deterministic processes that underpin microbial community assembly and transfer across life cycle stage and between gut compartments. Across gut compartments in farmed fish, neutral models suggest that most microbes are transient with no evidence of adaptation to their environment. In wild fish, we found declining taxonomic and functional microbial community richness as fish mature through different life cycle stages. Alongside neutral community models applied to wild fish, we suggest that declining richness demonstrates an increasing role for the host in filtering microbial communities that is correlated with age. We found a limited subset of gut microflora adapted to the farmed and wild host environment among which Mycoplasma spp. are prominent. Our study reveals the ecological drivers underpinning community assembly in both farmed and wild Atlantic salmon and underlines the importance of understanding the role of stochastic processes, such as random drift and small migration rates in microbial community assembly, before considering any functional role of the gut microbes encountered.IMPORTANCE A growing number of studies have examined variation in the microbiome to determine the role in modulating host health, physiology, and ecology. However, the ecology of host microbial colonization is not fully understood and rarely tested. The continued increase in production of farmed Atlantic salmon, coupled with increased farmed-wild salmon interactions, has accentuated the need to unravel the potential adaptive function of the microbiome and to distinguish resident from transient gut microbes. Between gut compartments in a farmed system, we found a majority of operational taxonomic units (OTUs) that fit the neutral model, with Mycoplasma species among the key exceptions. In wild fish, deterministic processes account for more OTU differences across life stages than those observed across gut compartments. Unlike previous studies, our results make detailed comparisons between fish from wild and farmed environments, while also providing insight into the ecological processes underpinning microbial community assembly in this ecologically and economically important species.

Alopecia areata is characterized by dysregulation in systemic type 17 and type 2 cytokines, which may contribute to disease-associated psychological morbidity.

BACKGROUND: Alopecia areata (AA) is a common autoimmune disease, causing patchy hair loss that can progress to involve the entire scalp (totalis) or body (universalis). CD8+ NKG2D+ T cells dominate hair follicle pathogenesis, but the specific mechanisms driving hair loss are not fully understood. OBJECTIVES: To provide a detailed insight into the systemic cytokine signature associated with AA, and to assess the association between cytokines and depression. METHODS: We conducted multiplex analysis of plasma cytokines from patients with AA, patients with psoriatic arthritis (PsA) and healthy controls. We used the Hospital Anxiety and Depression Scale (HADS) to assess the occurrence of depression and anxiety in our cohort. RESULTS: Our analysis identified a systemic inflammatory signature associated with AA, characterized by elevated levels of interleukin (IL)-17A, IL-17F, IL-21 and IL-23 indicative of a type 17 immune response. Circulating levels of the type 2 cytokines IL-33, IL-31 and IL-17E (IL-25) were also significantly increased in AA. In comparison with PsA, AA was associated with higher levels of IL-17F, IL-17E and IL-23. We hypothesized that circulating inflammatory cytokines may contribute to wider comorbidities associated with AA. Our assessment of psychiatric comorbidity in AA using HADS scores showed that 18% and 51% of people with AA experienced symptoms of depression and anxiety, respectively. Using linear regression modelling, we identified that levels of IL-22 and IL-17E are positively and significantly associated with depression. CONCLUSIONS: Our data highlight changes in both type 17 and type 2 cytokines among people with AA, suggesting that complex systemic cytokine profiles may contribute both to the pathogenesis of AA and to the associated depression. What's already known about this topic? NKG2D+ CD8+ T cells cause hair loss in alopecia areata (AA) but the immunological mechanisms underlying the disease are not fully understood. AA is associated with changes in levels of interleukin (IL)-6, tumour necrosis factor-α, IL-1ß and type 17 cytokines. Psychiatric comorbidity is common among people with AA. What does this study add? People with AA have increased plasma levels of the type 2 cytokines IL-33, IL-31 and IL-17E (IL-25), in addition to the type 17 cytokines IL-17A, IL-21, IL-23 and IL-17F. Levels of IL-17E and IL-22 positively predict depression score. What is the translational message? AA is associated with increased levels of multiple inflammatory cytokines, implicating both type 17- and type 2 immune pathways. Our data indicate that therapeutic strategies for treating AA may need to address the underlying type 17- and type 2 immune dysregulation, rather than focusing narrowly on the CD8+ T-cell response. An immunological mechanism might contribute directly to the depression observed in people with AA.

Cold adaptation and replicable microbial community development during long-term low-temperature anaerobic digestion treatment of synthetic sewage.

The development and activity of a cold-adapting microbial community was monitored during low-temperature anaerobic digestion (LtAD) treatment of wastewater. Two replicate hybrid anaerobic sludge bed-fixed-film reactors treated a synthetic sewage wastewater at 12°C, at organic loading rates of 0.25-1.0 kg chemical oxygen demand (COD) m-3 d-1, over 889 days. The inoculum was obtained from a full-scale anaerobic digestion reactor, which was operated at 37°C. Both LtAD reactors readily degraded the influent with COD removal efficiencies regularly exceeding 78% for both the total and soluble COD fractions. The biomass from both reactors was sampled temporally and tested for activity against hydrolytic and methanogenic substrates at 12°C and 37°C. Data indicated that significantly enhanced low-temperature hydrolytic and methanogenic activity developed in both systems. For example, the hydrolysis rate constant (k) at 12°C had increased 20-30-fold by comparison to the inoculum by day 500. Substrate affinity also increased for hydrolytic substrates at low temperature. Next generation sequencing demonstrated that a shift in a community structure occurred over the trial, involving a 1-log-fold change in 25 SEQS (OTU-free approach) from the inoculum. Microbial community structure changes and process performance were replicable in the LtAD reactors.

Three-dimensional ultrasound imaging.

This review is about the development of three-dimensional (3D) ultrasonic medical imaging, how it works, and where its future lies. It assumes knowledge of two-dimensional (2D) ultrasound, which is covered elsewhere in this issue. The three main ways in which 3D ultrasound may be acquired are described: the mechanically swept 3D probe, the 2D transducer array that can acquire intrinsically 3D data, and the freehand 3D ultrasound. This provides an appreciation of the constraints implicit in each of these approaches together with their strengths and weaknesses. Then some of the techniques that are used for processing the 3D data and the way this can lead to information of clinical value are discussed. A table is provided to show the range of clinical applications reported in the literature. Finally, the discussion relating to the technology and its clinical applications to explain why 3D ultrasound has been relatively slow to be adopted in routine clinics is drawn together and the issues that will govern its development in the future explored.