Investigation of the skin microbiome: swabs vs. biopsies.

BACKGROUND: Human skin is populated by diverse bacteria and there is increasing evidence that resident bacteria play a key role initiating immune responses in cutaneous diseases such as atopic dermatitis, psoriasis and hidradenitis suppurativa. Bacteria are present at all layers of the skin but many studies have relied on swabs to profile the skin microbiota. OBJECTIVES: As the pathogenesis of many skin conditions is dermal, we wanted to compare the microbiota obtained in swabs (surface) and biopsies (dermis). METHODS: Using 16S rRNA gene sequencing we established the microbial profiles of skin swabs and skin biopsies in 16 patients. RESULTS: We found differences in both diversity and taxonomic composition of the microbiome obtained from swabs and biopsies of the same individual. Several taxa were found to be more abundant in the swabs, which displayed significantly higher community richness, but Clostridiales and Bacteroidetes were significantly enriched in the biopsies. Most published research on cutaneous microbiota has been based on skin swabs, which represent the surface of the skin. CONCLUSIONS: Our study demonstrated a clear difference between the microbiome observed from skin swabs and skin biopsies. These findings may be highly relevant in disorders such as psoriasis where pathogenesis arises in the dermis. What's already known about this topic? 16S RNA gene sequencing has facilitated study of the skin microbiome. Several studies have sequenced the microbiome sampled by skin swabs. What does this study add? The microbiome data obtained using swabs and biopsies were different. Diseases that are predominantly dermal should be studied using both swabs and biopsies.

An investigation into the nutritional composition and cost of gluten-free versus regular food products in the UK.

BACKGROUND: The gluten-free (GF) food market has expanded considerably, although there is limited comparative evidence for the nutritional quality and cost of GF food products. The present study aims to compare the nutrient composition and cost of GF and gluten-containing (regular) foods across 10 food categories in the UK. METHODS: Nutritional information and the cost of GF foods available in the UK (n = 679) and comparable regular foods (n = 1045) were systematically collected from manufacturer and supermarket websites. Foods were classified using UK front-of-pack labelling for content of fat, saturated fat, sugar and salt and nutrient content, and cost per 100 g were identified and compared between GF and regular foods. RESULTS: Overall, more GF foods were classified as containing high and medium fat, saturated fat, sugar and salt than regular foods, although this was not universally consistent. More GF bread and flour products contained high fat and sugar, whereas fewer GF crackers contained high fat and sugar compared to regular foods. High salt content was found more frequently in GF than regular products. On average, GF products were 159% more expensive than regular (£0.44/100 g versus £1.14/100 g). GF items were also more likely to be lower in fibre and protein content than regular foods. CONCLUSIONS: Differences exist in the nutritional composition of GF and regular food. GF food is unlikely to offer healthier alternatives to regular foods, except for those who require a GF diet for medically diagnosed conditions, and it is associated with higher costs.

Assessing the toxic effects of DMSO on cord blood to determine exposure time limits and the optimum concentration for cryopreservation.

BACKGROUND AND OBJECTIVES: Advantages of using cord blood (CB) over other sources of haematopoietic progenitor cells, such as bone marrow, include the ability to cryopreserve and bank the samples until requested for a transplant. Cryopreservation requires the addition of a cryoprotectant to prevent the formation of intracellular ice during freezing. Dimethyl sulphoxide (DMSO) is commonly used at a concentration of 10% (v/v); however, there is evidence to suggest this chemical is toxic to cells as well as to patients after infusion. MATERIALS AND METHODS: The toxic effects of DMSO were assessed through cell viability and in vitro functional assays in fresh and post-thaw CB samples before determining the maximum exposure time and optimal concentration for cryopreservation. RESULTS: A dose-dependent toxicity of DMSO was observed in fresh samples with 40% removing all viable and functional haematopoietic progenitor cells (HPC). In fresh and post-thaw analysis, minimal toxic effect was observed when cryopreservation was delayed for up to 1 h after 10% DMSO addition. After thawing, DMSO washout was superior to dilution or unmanipulated when maintained for long periods (advantage observed 1 h after thawing). Finally, the optimum concentration for cryopreserving CB was found to be 7.5 to 10% with detrimental effects observed outside of this range. CONCLUSION: These results support the use of 7.5-10% as the optimal DMSO concentration and the maximum exposure time should be limited to <1 h prior to freezing and 30 min post-thaw.

Is chronic plaque psoriasis triggered by microbiota in the skin?

There is a known association between psoriasis and Crohn disease (CD). Patients with CD are five times more likely to develop psoriasis, and, conversely, patients with psoriasis are more likely to develop CD. Many gastroenterologists now accept that CD results from a breakdown of immune tolerance to the microbiota of the intestine in genetically susceptible individuals. The microbiota of the skin have recently been investigated in psoriasis. Firmicutes was the most common phylum, and Streptococcus the most common genus identified. Beta-haemolytic streptococci have been implicated in both guttate and chronic plaque psoriasis. Furthermore, the innate immune system has been shown to be activated in psoriasis, and many of the genes associated with the disease are concerned with the signalling pathways of the innate immune system, notably interleukin-23 and nuclear factor κB. Patients with psoriasis also have an increased incidence of periodontitis, a disease thought to be due to an abnormal response to normal oral commensals. Based on the similarities between CD and psoriasis, we propose that psoriasis is due to a breakdown of immune tolerance to the microbiota of the skin. In support of this hypothesis we provide evidence for microbiota in the skin, activation of the innate immune system, and genetic abnormalities involving the innate immune system.

Evidence that a locus for familial psoriasis maps to chromosome 4q.

Psoriasis is an inflammatory skin disease that affects 2% of the population. It is characterised by red, scaly skin patches which are usually found on the scalp, elbows and knees, and may be associated with severe arthropathy. The lesions are caused by abnormal keratinocyte proliferation, and infiltration of inflammatory cells into the dermis and epidermis. The usual age of onset of psoriasis is between 15 and 30 years, although it can present at any age. Psoriasis is recognised to have a large genetic component. Twin studies show the concordance in monozygotic twins to be between 65-70%, compared to between 15-20% in dizygotic twins. Family studies estimate the risk to first degree relatives at between 8-23%. However, there are also several environmental factors, including streptococcal infection and stress, that affect the onset and presentation of the disease. The mode of inheritance of psoriasis is unclear. We conducted a genome-wide scan to search for psoriasis susceptibility loci in a single large multiplex family. Parametric linkage analysis indicated that a susceptibility locus for familial psoriasis was located on chromosome 4q. Investigation of this locus in five further multiplex families using both parametric and non-parametric methods gave significant localisation to chromosome 4q. The maximum total pairwise lod score obtained was 3.03 with the microsatellite marker D4S1535 at theta = 0.08. Non-parametric multipoint analysis with GENEHUNTER- demonstrated significant excess allele sharing, with a P value of 0.0026, at the same locus.

Concepts in psoriasis: psoriasis and the extracellular matrix.

Post-streptococcal guttate psoriasis only sometimes progresses to the persistent plaque psoriasis. We have previously proposed that psoriasis is driven by multiple extradomain A+ fibronectin (EDA+ FN) 'feedback loops'. The psoriasis-specific loop, the 'keratinocyte loop', depends upon expression by keratinocytes of α5ß1 integrin. In normal skin, α5ß1 expression is minimal or absent in resting epithelial cells, where basal cells are anchored to the laminin component of the basement membrane (via integrins α6ß4 and α3ß1). However, when the laminin layer is disrupted, due to wounding for instance, α5ß1 is then strongly expressed by basal cells as a means for anchoring. At uninvolved skin sites in patients with psoriasis the laminin layer within the basement membrane is disrupted, with a consequential increase in the expression by basal keratinocytes of EDA+ FN and α5ß1. We are postulating that in guttate psoriasis streptococcal lytic enzymes, and in particular streptokinase, may - via plasminogen activation - initiate a disruption of the laminin layer in the basement membrane. It is also possible that the expression by proliferating keratinocytes of plasminogen-activating factor might result in additional laminin digestion. The suggestion is that progression of guttate psoriasis to full-blown plaque psoriasis, or to its spontaneous resolution, is governed largely by the triggering of humoral immune responses. Thus, if streptococcal lytic enzymes are neutralized by host antibodies in advance of irreversible and lasting damage to the basement membrane of the dermal papillae, then guttate psoriasis will resolve spontaneously. In contrast, if permanent damage is induced prior to effective neutralization of the relevant bacterial toxins, then keratinocytes will become chronically destabilized. The consequence will be to liberate keratinocytes to proliferate in response to various stimuli, and to continue to elaborate plasminogen activation enzymes; the combined effect being, in concert with immunological priming, to precipitate and sustain plaque psoriasis.

Psoriasis and extra domain A fibronectin loops.

We have previously postulated that as well as T-helper (Th) 1 and Th17 cells, the transforming growth factor (TGF)-beta/fibronectin (FN)/alpha5beta1 pathway is central to psoriasis pathogenesis. EDA+ FN refers to an alternatively spliced isoform of FN with an additional domain known as extra domain A. EDA+ FN has two important properties pertinent to psoriasis lesions: it stimulates keratinocyte hyperproliferation, and, through stimulation of Toll-like receptor (TLR) 4, stimulates production of proinflammatory cytokines. EDA+ FN production induced by TGF-beta stimulation can be maintained in psoriasis lesions via two main feedback loops. Firstly, EDA+ FN stimulates proliferation of keratinocytes, which, in an autocrine fashion, will release more EDA+ FN. Secondly, EDA+ FN stimulates TLR4 expressed by antigen-presenting cells resulting in the production of proinflammatory cytokines such as tumour necrosis factor-alpha, interleukin (IL)-1, IL-6 and IL-12. The resultant promotion of cutaneous inflammation results in the recruitment of Th1 cells, which also produce EDA+ FN. We propose that these 'FN loops' contribute to the maintenance and progression of psoriatic lesions. Finally, although the association between psoriasis and heart/thrombotic disease remains unclear one plausible link may be the promotion of atherosclerosis and thrombotic heart disease by EDA+ FN.

Bile leak from the duct of Luschka.

A 64-year-old diabetic man underwent an open cholecystectomy for acute necrotizing cholecystitis. Post-operatively he developed a biloma which was drained percutaneously. A bile leak was suspected and he underwent an ERCP. Initial cholangiography was normal, but upon continued injection of contrast agent, a bile leak originating from a branch of the right hepatic duct or duct of Luschka became evident. A sphincterotomy was performed and a plastic stent was placed into the common bile duct. The leak resolved and the plastic stent was removed 6 weeks later.

Psoriasis and streptococci: the natural selection of psoriasis revisited.

We have previously postulated that surviving invasive streptococcal infections may have been a factor in psoriasis becoming a common skin disease in some parts of the world. Many of the candidate genes linked to psoriasis are associated with the acquired or innate immune system, which are also important in host defence to invasive streptococcal infections. High rates of positive streptococcal throat swabs among patients with chronic plaque psoriasis suggest that they are efficient at internalizing/carrying beta-haemolytic streptococci. Internalization of streptococci in the throat is dependent upon the transforming growth factor (TGF)-beta/fibronectin/alpha 5 beta 1 integrin pathway. The immune cell Th17 and its related cytokine network are important in mucosal defence, being very effective against extracellular microbes but having little effect on intracellular organisms. The TGF-beta/fibronectin/alpha 5 beta 1 integrin pathway and the Th17 cell network also appear to be operative in psoriasis, animal models of both TGF-beta and alpha 5 beta 1 cutaneous overexpression being associated with characteristic psoriasis lesions. We postulate that some of the genotypic/phenotypic changes in different immunological pathways in psoriasis, including the acquired T-cell response, the innate immune response, the TGF-beta/fibronectin/alpha 5 beta 1 integrin pathway and the Th17 cell system, confer protection against mortality during epidemics of invasive streptococcal infections, heightened efficiency in internalizing and allowing carriage of streptococci as well as predisposition to the development of psoriasis.

Double-balloon enteroscopy-assisted virtual chromoendoscopy for small-bowel disorders: a case series.

The Fujinon intelligent color enhancement (FICE) system is a new, virtual chromoendoscopy technique that enhances mucosal visibility. The aim of this study was to assess the utility of double-balloon enteroscopy (DBE) with FICE technology (EPX-4400 processor, Japan) for the characterization of various small-bowel diseases. Overall, a total of 574 endoscopic pictures were obtained and analyzed. FICE was found to be a helpful method for the evaluation of adenomatous small-bowel polyps and angiodysplasias. Its use for the characterization of celiac and Crohn's disease appears to be limited. Overall, FICE may become a useful method that aids in characterization and provides new insights to small-bowel pathologies.

A retrospective analysis of emergency double-balloon enteroscopy for small-bowel bleeding.

Although the role of emergency esophagogastroduodenoscopy (EGD) and colonoscopy for upper and lower gastrointestinal bleeding (GIB) is well defined, there are no data on the concept of emergency double-balloon enteroscopy (DBE) for small-bowel bleeding. The aim of this study was to retrospectively evaluate the concept of emergency DBE in overt obscure GIB and assess its impact on patient management. A total of 17 emergency DBEs for overt obscure GIB were carried out in ten patients (six women, four men; mean age 68 years, range 35 - 83). The following diagnoses were made: actively bleeding Dieulafoy lesions of the small bowel, n = 2; bleeding tumors, n = 4 (carcinoids n = 2, adenocarcinoma n = 1, lipoma n = 1); bleeding angiodysplasias and/or large arteriovenous malformation (AVM), n = 2; multiple ulcers, n = 1; and no diagnosis, n = 1. Endoscopic therapies included argon plasma coagulation (n = 6), injection of epinephrine (n = 3), and use of fibrin glue (n = 1). It appears that emergency DBE is technically feasible, facilitates both diagnosis and therapy and enables management of patients with massive overt obscure GIB. This study is a first step in establishing the concept of emergency DBE for patients with suspected small-bowel bleeding.

A possible role for vaccination in the treatment of psoriasis?

Psoriasis is a multifactorial immune skin disease whose etiology involves a strong genetic component, involving several genes encoding proteins involved in epidermal differentiation and immune, inflammatory and pathogen responses, in combination with microbial environmental factors. Although various microorganisms appear to provoke or aggravate the disease, including Staphylococcus aureus, Malassezia and Candida albicans, the association between S. pyogenes throat infections and guttate psoriasis is supported by the strongest clinical evidence. Furthermore, the identification of peptidoglycan-specific T cells in psoriatic skin lesions has led to the proposal that cell wall peptidoglycan may mediate the link between streptococcal infection in the tonsils and the subsequent induction of skin lesions. These findings suggest that psoriasis may be a possible candidate for therapeutic streptococcal vaccination. Current treatments for psoriasis have several limitations including toxicity and an increased risk of infection and malignancy. In contrast, vaccination could potentially induce long-term tolerance without the side effects caused by global immunosuppression. Future research will need to address the identity of the triggering microbial antigen(s); such knowledge could open the way for vaccination as a therapeutic tool for psoriasis.

HLA class II alpha chain gene polymorphisms in patients with insulin-dependent diabetes mellitus, dermatitis herpetiformis, and celiac disease.

We have investigated DNA polymorphism of the class II alpha chain genes in HLA typed patients with insulin dependent diabetes mellitus (IDDM; n = 79), celiac disease (CD; n = 46), dermatitis herpetiformis (DH; n = 53), and controls (n = 86). Preferential allelic associations of HLA genes and gene products have thus been constructed for susceptibility to these diseases. DR alpha and DQ alpha gene polymorphisms indicated heterogeneity of HLA DR3, DRw6, and DR7, and HLA DR2 and DRw6, respectively. In DR7 positive CD patients a 3.8-kilobase (kb) DR alpha fragment, which correlated with DQw3, was found in only 11% of patients compared with 45% of corresponding controls (P less than 0.05). An increased frequency of a DX alpha genotype UU in all three diseases was found (IDDM 59%, DH 45%, CD 48%, compared to 21% in controls, P less than 0.001), which is not explained solely by the increased frequencies of DR3-DX alpha U. We therefore conclude part of the genetic susceptibility for these three conditions is encoded by genes within the DQ-DX subregion.

Characterization of yellow plaques found in the small bowel during double-balloon enteroscopy.

BACKGROUND AND STUDY AIMS: The aim of this study was to characterize yellow (or whitish) plaques of the small bowel that were found during double-balloon enteroscopy (DBE) performed for small-bowel evaluation. PATIENTS AND METHODS: Patients who were being evaluated for small-bowel pathology at our institution (for a variety of indications) were included in the study. In 16 patients, DBE revealed yellow or whitish submucosal plaques, defined as small, raised, submucosal lesions that were well circumscribed and covered by normal-appearing small-bowel mucosa. Biopsy tissue obtained during the procedures was stained with hematoxylin and eosin and with periodic acid-Schiff stain, and was subjected to immunochemical testing using endothelial markers (anti-CD31 and anti-CD34). RESULTS: These 16 patients were identified out of a total of 150 DBE procedures performed in 120 patients (eight men, eight women; mean age 62, range 33 - 78). The lesions were mostly single (range 1 to > 5 lesions), ranging in size from 2 mm to 15 mm, and were slightly raised (from 1 mm to 2 mm). In four cases the plaques could not be biopsied because the patient had a coagulation disorder or because the DBE was being performed to investigate severe acute bleeding. In the other 12 patients, a characteristic white-yellow liquid exudated from the biopsy site in 80 % of lesions, and these 12 patients were shown to have lymphangiectasias. No association with an infiltrative disorder could be detected. CONCLUSIONS: Yellow and white submucosal plaques are found in up to 13 % of patients undergoing DBE. They are most likely to be lymphangiectasias and are a normal anatomical variant. They do not require further work-up.

What are we injecting with our drugs?

In preparation for a case, an anaesthetist opened a 20 ml glass vial of propofol and aspirated the propofol into a syringe via a blunt drawing-up needle. Increased resistance was felt with aspiration. On inspection, a shard of glass was found at the tip of the drawing-up needle. The shard was presumed to be from the propofol ampoule, and to have fallen into the solution upon snapping open its glass tip. This illustrative case raises the issue of contamination of drugs by particles introduced during the drawing-up process. It also highlights the possibility that during the drawing-up process, intravenous drugs may become contaminated not just with particles, but with microorganisms on the surface of the particles. In this article, we discuss relevant recent research of the implications of this type of drug contamination. We draw attention to the need for meticulous care in drawing up and administering intravenous drugs during anaesthesia, particularly propofol.