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Lobsters are ubiquitous, economically important decapod crustaceans with apparently conflicting evolutionary relationships. Here, we describe the chromosomal location of the major (45S rDNA) and minor (5S rDNA) ribosomal gene families in four species of Astacidea and five of Achelata, using two-color FISH. The major ribosomal family is located in 4–16 sites per diploid chromosome set, with Nephrops norvegicus (Nephropidae) showing the highest number described so far in Decapoda. The 5S rDNA is located in two sites in eight species; only in the crayfish Procambarus clarkii the 5S FISH signals were detected in four sites together with additional weaker signals. Furthermore, in N. norvegicus the minor ribosomal genes are syntenic with one major ribosomal cluster. Moreover, we located by two-color FISH the pentanucleotide (TTAGG)n telomeric repeat in the Nephropidae studied, showing the occurrence of a colocalization with 45S ribosomal sequences in Homarus gammarus. The comparison of chromosomal locations of repetitive sequences in Mediterranean, Atlantic, and South African lobster species as well as in marine and freshwater ones provides information on chromosomal evolution and cytotaxonomy of Decapoda. [ABSTRACT FROM AUTHOR]

Probiotic bacteria, capable of conferring benefits to the host, can present challenges in design, development, scale-up, manufacturing, commercialization, and life cycle management. Strain identification is one of the main quality parameters; nevertheless, this task can be challenging since established methodologies can lack resolution at the strain level for some microorganisms and\or are labor-intensive and time-consuming. Fourier transform infrared spectroscopy (FTIRS) has been largely used for the investigation of pathogenic species in the clinical field, whereas only recently has been proposed for the identification of probiotic strains. Within the probiotic industrial production, bacterial strains can be subjected to stressful conditions that may affect genomic and phenotypic characteristics; therefore, real-time monitoring of all the sequential growth steps is requested. Considering the fast, low-cost, and high-throughput features, FTIRS is an innovative and functional technology for typing probiotic strains from bench-top experiments to large-scale industrial production, allowing the monitoring of stability and identity of probiotic strains. In this study, the discriminatory power of FTIRS was assessed for four Lactiplantibacillus plantarum probiotic strains grown under different conditions, including temperatures (30 and 37°C) and medium (broth and agar), after consecutive sub-culturing steps. A comparison between the generated spectra with pulsed-field gel electrophoresis (PFGE) profiles was also performed. FTIRS was not only able to distinguish the strains of L. plantarum under different growth conditions but also to prove the phenotypic stability of L. plantarum type strain LP-CT after six growing steps. Regardless of the growth conditions, FTIRS spectra related to LP-CT constituted a unique hierarchical cluster, separated from the other L. plantarum strains. These results were confirmed by a PFGE analysis. In addition, based on FTIRS data, broth cultures demonstrated a higher reproducibility and discriminatory power with respect to agar ones. These results support the introduction of FTIRS in the probiotic industry, allowing for the step-by-step monitoring of massive microbial production while also guaranteeing the stability and purity of the probiotic strain. The proposed novel approach can constitute an impressive improvement in the probiotic manufacturing process., Competing Interests: MP, FD, and TG were employees of Probiotical Research S.r.L, Novara (Italy). MC was employee of Bruker Daltonics, Bremen (Germany). The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Deidda, Cordovana, Bozzi Cionci, Graziano, Di Gioia and Pane.)

Autism spectrum disorders (ASDs) represent a diagnostic challenge with a still partially uncertain etiology, in which genetic and environmental factors have now been assessed. Among the hypotheses underlying the involvement of biological and environmental factors, the gut-brain axis is of particular interest in autism spectrum disorders. Several studies have highlighted the related incidence of particular gastrointestinal symptoms (GISs) in children suffering from ASDs. Probiotics have shown success in treating several gastrointestinal dysbiotic disorders; therefore, it is plausible to investigate whether they can alleviate behavioral symptoms as well. On these bases, a randomized double-blind crossover study with a placebo was conducted, evaluating the effects of a mixture of probiotics in a group of 61 subjects aged between 24 months and 16 years old with a diagnosis of ASD. Behavioral evaluation was performed through the administration of a questionnaire including a Parenting Stress Index (PSI) test and the Vineland Adaptive Behavior Scale (VABS). The Psycho-Educational Profile and the Autism Spectrum Rating Scale (ASRS) were also evaluated. Microbial composition analyses of fecal samples of the two groups was also performed. The study showed significant improvements in GISs, communication skills, maladaptive behaviors, and perceived parental stress level after the administration of probiotics. Microbiome alpha diversity was comparable between treatment arms and no significant differences were found, although beta diversity results were significantly different in the treatment group between T0 and T1 time points. Streptococcus thermophilus , Bifidobacterium longum , Limosilactobacillus fermentum , and Ligilactobacillus salivarius species were identified as some of the most discriminant taxa positively associated with T1 samples. This preliminary study corroborates the relationship between intestinal microbiota and ASD recently described in the literature.

Aim: Acinetobacter baumannii is a pathogen of serious concern, often exhibiting multiple antibiotic resistance, frequently associated with hospital outbreaks in intensive care units. A prompt detection and tracking of these isolates is crucial. Reference methods for typing (pulsed-field gel electrophoresis, whole-genome sequencing) are accurate, but expensive and time-consuming, therefore limited to retrospective analysis. Materials & methods: In this study, the application of the FTIR-based IR Biotyper ® (IRBT) to track and monitor in real-time the spread of a multidrug-resistant A. baumannii  outbreak was investigated. The index case and the multidrug-resistant  A. baumannii isolates collected in the following 3 weeks were investigated. Results: IR Biotyper ® clustering results were fully confirmed by pulsed-field gel electrophoresis results. Conclusions: IR Biotyper represent a promising tool for real-time hospital hygiene, enabling a prompt and reliable typing.

Fourier transform infrared (FTIR) spectroscopy, a technology traditionally used in chemistry to determine the molecular composition of a wide range of sample types, has gained growing interest in microbial typing. It is based on the different vibrational modes of the covalent bonds between atoms of a given sample, as bacterial cells, induced by the absorption of infrared radiation. This technique has been largely used for the study of pathogenic species, especially in the clinical field, and has been proposed also for the typing at different subspecies levels. The high throughput, speed, low cost, and simplicity make FTIR spectroscopy an attractive technique also for industrial applications, in particular, for probiotics. The aim of this study was to compare FTIR spectroscopy with established genotyping methods, pulsed-field gel electrophoresis (PFGE), whole-genome sequencing (WGS), and multilocus sequence typing (MLST), in order to highlight the FTIR spectroscopy potential discriminatory power at strain level. Our study focused on bifidobacteria, an important group of intestinal commensals generally recognized as probiotics. For their properties in promoting and maintaining health, bifidobacteria are largely marketed by the pharmaceutical, food, and dairy industries. Strains belonging to Bifidobacterium longum subsp. longum and Bifidobacterium animalis subsp. lactis were taken into consideration together with some additional type strains. For B. longum subsp. longum , it was possible to discriminate the strains with all the methods used. Although two isolates were shown to be strictly phylogenetically related, constituting a unique cluster, based on PFGE, WGS, and MLST, no clustering was observed with FTIR. For B. animalis subsp. lactis group, PFGE, WGS, and MLST were non-discriminatory, and only one strain was easily distinguished. On the other hand, FTIR discriminated all the isolates one by one, and no clustering was observed. According to these results, FTIR analysis is not only equivalent to PFGE, WGS, and MLST, but also for some strains, in particular, for B. animalis subsp. lactis group, more informative, being able to differentiate strains not discernible with the other two methods based on phenotypic variations likely deriving from certain genetic changes. Fourier transform infrared spectroscopy has highlighted the possibility of using the cell surface as a kind of barcode making tracing strains possible, representing an important aspect in probiotic applications. Furthermore, this work constitutes the first investigation on bifidobacterial strain typing using FTIR spectroscopy., Competing Interests: MP and FD are employees of Probiotical Research S.r.L., Novara, Italy. MC is employee of Bruker Daltonik, Bremen, Germany. FF and IC are employees of Microbion S.r.L., San Giovanni Lupatoto, Verona, Italy. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Deidda, Bozzi Cionci, Cordovana, Campedelli, Fracchetti, Di Gioia, Ambretti and Pane.)

Background: Defective alleles within the PRF1 gene, encoding the pore-forming protein perforin, in combination with environmental factors, cause familial type 2 hemophagocytic lymphohistiocytosis (FHL2), a rare, severe autosomal recessive childhood disorder characterized by massive release of cytokines-cytokine storm., Objective: The aim of this study was to determine the function of hypomorph PRF1:p.A91V g.72360387 G > A on multiple sclerosis (MS) and type 1 diabetes (T1D)., Methods: We cross-compare the association data for PRF1:p.A91V mutation derived from GWAS on adult MS and pediatric T1D in Sardinians. The novel association with T1D was replicated in metanalysis in 12,584 cases and 17,692 controls from Sardinia, the United Kingdom, and Scotland. To dissect this mutation function, we searched through the coincident association immunophenotypes in additional set of general population Sardinians., Results: We report that PRF1:p.A91V , is associated with increase of lymphocyte levels, especially within the cytotoxic memory T-cells, at general population level with reduced interleukin 7 receptor expression on these cells. The minor allele increased risk of MS, in 2903 cases and 2880 controls from Sardinia p  = 2.06 × 10 -4 , odds ratio OR = 1.29, replicating a previous finding, whereas it protects from T1D p  = 1.04 × 10 -5 , OR = 0.82., Conclusion: Our results indicate opposing contributions of the cytotoxic T-cell compartment to MS and T1D pathogenesis.

The Muraenidae is one of the largest and most complex anguilliform families. Despite their abundance and important ecological roles, morays are little studied, especially cytogenetically, and both their phylogenetic relationships and the taxonomy of their genera are controversial. With the aim of extending the karyology of this fish group, the chromosomal mapping of the 5S ribosomal gene family was performed on seven species belonging to the genera Muraena and Gymnothorax from both the Atlantic and Pacific oceans. Fluorescence in situ hybridisation (FISH) experiments were realized using species-specific 5S rDNA probes; in addition, two-colour FISH was performed to investigate the possible association with the 45S ribosomal gene family. Multiple 5S rDNA clusters, located either in species-specific or in possibly homoeologous chromosomes, were found. Either a syntenic or different chromosomal location of the two ribosomal genes was detected. Our results revealed variability in the number and location of 5S rDNA clusters and confirmed a substantial conservation of the number and location of the 45S rDNA.

We report on the influence of ~22 million variants on 731 immune cell traits in a cohort of 3,757 Sardinians. We detected 122 significant (P < 1.28 × 10 -11 ) independent association signals for 459 cell traits at 70 loci (53 of them novel) identifying several molecules and mechanisms involved in cell regulation. Furthermore, 53 signals at 36 loci overlapped with previously reported disease-associated signals, predominantly for autoimmune disorders, highlighting intermediate phenotypes in pathogenesis. Collectively, our findings illustrate complex genetic regulation of immune cells with highly selective effects on autoimmune disease risk at the cell-subtype level. These results identify drug-targetable pathways informing the design of more specific treatments for autoimmune diseases.

Exclusive breastfeeding is currently recommended until at least 6 months of postnatal age, due to maternal breast milk (BM) unique composition and beneficial properties. In fact, BM modifies itself according to gestational age (GA) at birth, adapting its composition to neonatal requests during lactation. Multiple births represent about 3% of the whole pregnancies; such neonates result more vulnerable than full-term newborns, due to lower GA and birth weight (BW) and the higher incidence of perinatal complications. Although an adequate nutrition is fundamental for twins and other multiples, studies on this topic are lacking. We collected and analyzed BM from mothers of 19 twins and 5 triplets showing GA < 33 weeks and BW < 1500 g, comparing it to a control group of 28 preterm singletons. As a result, at GA ≤ 28 weeks, we observed that protein content is higher in BM for multiples (1.53 vs. 1.29 g per 100 ml), lactose concentration is greater in BM for singletons (6.72 vs. 6.34 g per 100 ml) and GA results the most relevant factor influencing BM protein composition. BM for multiples results higher in proteins and lower in lactose, if compared with singleton's samples; this could promote and sustain growth and organ development in this vulnerable category. BM from multiples shows a trophic and immunologic role, since these neonates often show lower GA and BW instead of singletons. These findings could help in optimizing nutritional strategies and improving BM individualized fortification.

Background: Genomewide association studies of autoimmune diseases have mapped hundreds of susceptibility regions in the genome. However, only for a few association signals has the causal gene been identified, and for even fewer have the causal variant and underlying mechanism been defined. Coincident associations of DNA variants affecting both the risk of autoimmune disease and quantitative immune variables provide an informative route to explore disease mechanisms and drug-targetable pathways., Methods: Using case-control samples from Sardinia, Italy, we performed a genomewide association study in multiple sclerosis followed by TNFSF13B locus-specific association testing in systemic lupus erythematosus (SLE). Extensive phenotyping of quantitative immune variables, sequence-based fine mapping, cross-population and cross-phenotype analyses, and gene-expression studies were used to identify the causal variant and elucidate its mechanism of action. Signatures of positive selection were also investigated., Results: A variant in TNFSF13B, encoding the cytokine and drug target B-cell activating factor (BAFF), was associated with multiple sclerosis as well as SLE. The disease-risk allele was also associated with up-regulated humoral immunity through increased levels of soluble BAFF, B lymphocytes, and immunoglobulins. The causal variant was identified: an insertion-deletion variant, GCTGT→A (in which A is the risk allele), yielded a shorter transcript that escaped microRNA inhibition and increased production of soluble BAFF, which in turn up-regulated humoral immunity. Population genetic signatures indicated that this autoimmunity variant has been evolutionarily advantageous, most likely by augmenting resistance to malaria., Conclusions: A TNFSF13B variant was associated with multiple sclerosis and SLE, and its effects were clarified at the population, cellular, and molecular levels. (Funded by the Italian Foundation for Multiple Sclerosis and others.).

Goals: To investigate the possible use of Lactobacillus strains in the prophylaxis and/or adjuvant therapy of acute vulvovaginal candidiasis and other vaginal infections sustained by Candida yeasts., Background: The incidence of Candida infections has substantially increased in recent years. Treatment of vaginal infections with lactobacilli has a long tradition, starting with Döderlein's description of the vaginal microbiota., Materials and Methods: We assessed the activity of serially diluted fluconazole and miconazole (from 3 ng/mL to 1 mg/mL) against Candida strains. Serial dilutions of the azoles were prepared in Sabouraud Dextrose Broth in the presence of Candida strains. Broths were incubated under aerobic condition at 30°C, and the optical density was measured at 560 nm. Minimum inhibitory concentration was defined as the lowest concentration of the antibiotic that completely inhibited visible growth., Results: An evident resistance to the azoles used was recorded for all species of Candida, with the exception of Candida parapsilosis. For this species, a minimum inhibitory concentration ≤1 mg/mL was obtained, thus confirming the slight sensitivity to fluconazole and miconazole.All Lactobacillus strains tested, namely LF5, LF09, LF10, and LF11, have the ability to significantly inhibit the growth of the five species of Candida of at least 4 logarithms. Furthermore, the best result obtained with miconazole on C. parapsilosis is still 2 logarithms lower., Conclusions: The use of beneficial bacteria, especially lactobacilli, could be regarded as a good alternative for the prevention and treatment of Candida infections.

Goals: The aim of the study was to unequivocally demonstrate the nontransmissibility of the genes mediating the resistance of the strain Bifidobacterium longum W11 (LMG P-21586) to rifaximin., Background: Most antibiotic treatments can induce unfavorable side effects such as antibiotic-associated diarrhea, which is largely attributable to the disruption of the intestinal microbiota. The parallel intake of probiotic bacteria might reduce these events, even if with generally very poor results. In this regard, the use of antibiotic-resistant beneficial bacteria could represent a worthy strategy., Study: Rifaximin was tested in parallel with rifampicin, rifapentine, and rifabutin, all rifamycin derivates, using 5 different concentrations. Susceptibility tests were performed by the disc diffusion method of Kirby-Bauer, and inhibition zones were measured after incubation at 37°C. B. longum BL03 was used as comparison. The B. longum W11 genome was sequenced on Illumina MiSeq with a 250 PE reads module. After mapping the reads with the reference bacterial genome, the alignment data were processed using FreeBayes software., Results: B. longum BL03 was inhibited by all antibiotics even at the lowest concentration. In contrast, the W11 strain was inhibited by rifampicin, rifabutin, and rifaximin only at the highest concentration (512 μg/mL). The genomic analysis showed a mutation into the chromosomal DNA. No transposable elements were found, and the genetic locus was not flanked by close mobile genetic elements., Conclusions: B. longum W11 could be used in combined therapy with rifaximin, thus opening new focused frontiers in the probiotic era while preserving the necessary safety of use for consumers.

Goals: To investigate the modulation of human cytokines by Bifidobacterium longum strains isolated from Centenarians. In particular, we measured the production of interleukin (IL)-12p70, interferon-γ, IL-17A, and IL-4 from human peripheral blood mononuclear cells after stimulation with live bacteria., Background: Probiotics may inhibit pathogens and modulate the immune system, bringing a beneficial effect on human health. Among the probiotic strains, bifidobacteria play a key role in the maturation of the host's immune system. At present, only a few comparative data are available on the effects of bifidobacteria associations on cytokine production., Study: Peripheral blood mononuclear cells were isolated, cultured, and stimulated (ratio 1:1) with B. longum DLBL07, B. longum DLBL08, B. longum DLBL09, B. longum DLBL10, or B. longum DLBL11, either alone or in association. Cytokine production was determined by an enzyme-linked immunosorbent assay., Results: Both the B. longum DLBL mixture and the individual B. longum DLBL strains induced similar levels of IL-4, interferon-γ, and IL-17A. Under all conditions tested, no IL-12p70 release was detected., Conclusions: The fact that B. longum strains were obtained from Centenarians suggests a perfect homeostasis between this specific species and the host. Moreover all the B. longum strains from Centenarians used in our study share some biological similarities.

Goals: This study was undertaken to demonstrate the ability of Lactobacillus fermentum LF5 (DSM 32277) to inhibit in vitro different Candida species and Gardnerella vaginalis to weigh its potential effectiveness even in mixed vaginal infections., Background: A wide female population is suffering from various vulvovaginal infections. These diseases are often associated with a decrease in the concentration of Lactobacilli in the vagina. Mixed vaginal infections represent >20% of women with vulvovaginal infection., Study: LF5 strain was cocultured in De Man, Rogosa and Sharpe with Candida according to a 1:100 ratio in favor of the yeast. Each culture was sampled after 24 hours of incubation for the selective enumeration of the yeasts performed on yeast extract glucose chloramphenicol agar medium.The growth of Gardnerella alone (positive control) and in the presence of different concentrations of neutralized supernatants of L. fermentum LF5 ranging from 5% to 20% was quantified by means of optical density at 600 nm (OD600)., Results: L. fermentum LF5 demonstrated the ability to inhibit significantly the growth of the 5 species of Candida by at least 4 logarithms.Furthermore, L. fermentum LF5 showed a significant activity after both 24 and 48 hours (46% and 82% with 20% of neutralized supernatant, respectively). A significant dose-dependent growth inhibition was recorded in particular after 48 hours of incubation, even achieving a 80% inhibition of G. vaginalis growth., Conclusions: The biotherapeutic LF5 could be the only documented strain effective in mixed forms. For this purpose, a human clinical trial is in progress.

Goals: To determine the in vitro antimicrobial activity of selected Lactobacillus strains isolated from the feces of healthy humans against Klebsiella pneumoniae., Background: Klebsiella is ubiquitous in nature and may colonize the skin, the pharynx, or the gastrointestinal tract of humans. Despite the widespread use of antibiotic molecules with a broad spectrum in hospitalized patients, an increased overall load of klebsiellae as well as the subsequent development of multidrug-resistant strains able to synthesize extended-spectrum beta-lactamase have been registered. These strains are particularly virulent, express capsular-type K55, and have a considerable ability to propagate., Study: The 4 strains Lactobacillus paracasei LPC01 (CNCM I-1390), Lactobacillus rhamnosus LR04 (DSM 16605), Bifidobacterium longum B2274 (DSM 24707), and Lactobacillus delbrueckii subsp. delbrueckii LDD01 (DSM 22106) were tested. The analysis was performed using both a disc-diffusion assay and the broth-dilution procedure, also including an evaluation of the supernatants obtained from a fresh broth culture of each bacterium., Results: L. delbrueckii subsp. delbrueckii LDD01 demonstrated the best inhibitory results among all the tested strains. The antibacterial activity of the supernatant was retained even after treatment with α-amylase and neutralization with NaOH 1N, thus suggesting the protein structure of the inhibitory molecule. In contrast, it was completely lost after treatment with proteinase K., Conclusions: Overall results suggest that the inhibitory effect of L. delbrueckii subsp. delbrueckii LDD01 should be attributed to the production of a bacteriocin. This strain may be prospectively useful for strengthening probiotic formulations and possibly counteract infections by K. pneumoniae in humans.

Background: The use of breast milk presents numerous early and long-term advantages for ELBW preterms. However, breast milk without fortification does not cover the high nutritional needs of such patients. The aim of our study was to assess the effect of individualized fortification of breast milk on the growth of ELBWs hospitalized in a neonatal intensive care unit (NICU)., Methods: Retrospective single-center observational study., Results: We assessed the growth of 41 consecutive ELBWs (21 females, 20 males) with gestational ages between 23 and 30 weeks (mean GA 26.31±1.8) fed with breast milk in an individualized way. The rate of growth as the mean weight increase with breast milk fortification was 16.04±3.13 g/kg/day, more than the growth of the fetus in the uterus (~15 g/kg/day). This result was confirmed also among the ELBWs of lower GA. However, only 24.4% of all the ELBWs at the time of discharge from the NICU presented an appropriate weight for their gestational age. No cases of Necrotizing Enterocolitis (NEC) were observed., Conclusions: Despite high growth velocity, the ELBWs failed to remain in the same percentiles of birth and, at discharge, only 27.7% had a weight of >10 centiles. Further studies are needed to improve growth during early critical phases of development., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Background: Leaky gut, or intestinal permeability, is the phenomenon of the gut wall exhibiting increased absorbency. It is pretty well recognised that an altered or damaged bowel lining or gut wall may result from unbalanced diet, parasites, infection, or medications and that this allows substances such as toxins, microbes, undigested food, or waste to leak through. As a natural consequence, this prompts the body to initiate an immune reaction leading to potentially severe health conditions. Different strategies may be used to improve, at least temporarily, the physiological intestinal barrier. The use of specific beneficial microorganisms, such as lactobacilli and bifidobacteria, has been suggested as an innovative tool to counteract an improper level of intestinal permeability. The association of bacteria with specific gelling agents, such as gums, may represent an improvement since these molecules are able to form hydrophilic gels that distribute uniformly over the inner intestinal surface. This pilot study was undertaken to evaluate intestinal permeability in subjects treated with a gelling complex, an association of tara gum and the microorganism Streptococcus thermophilus ST10 (DSM 25246), which has a well-demonstrated in vitro ability to synthesise and secrete exopolysaccharides (EPSs)., Methods: Twenty-five healthy subjects were enrolled in this human intervention, double-blind, placebo-controlled, pilot trial (age between 21 and 57 y, mean 37.7±11.2). Subjects were then randomised into 2 groups: group A (13 subjects) was given an active formulation containing 250 mg of tara gum and 1 billion viable cells of S. thermophilus ST10, whereas group B (12 subjects) was given a placebo formulation. All the subjects participating in the study were directed to take 1 dose per day for 30 consecutive days. The presence and concentration of exopolysaccharides (EPSs) in the faeces was determined at time 0 (d0), after 30 days of treatment (d30), and at the end of the 2-week follow-up period (d45). The monosaccharide composition of EPSs was used to quantify the possible contribution of tara gum to the amount of polysaccharides detected in the faecal material. Intestinal permeability was evaluated at the same time by means of the lactitol/mannitol ratio (small intestine permeability) and sucralose concentration (colonic permeability) in urine specimens sampled after specified times. A statistical comparison was made between the concentration of EPSs, the lactulose/mannitol ratio, and the amount of excreted sucralose in the 2 groups at d0, d30, and d45., Results: In the active group, supplementation with S. thermophilus ST10 and tara gum was able to significantly increase the faecal EPSs concentration compared with placebo (from 0.169 mg/g to 0.633 mg/g after 30 d, P<0.001). An interesting decrease in intestinal permeability, both of the small bowel and in the colon, was also recorded. The L/M ratio diminished from 0.021 in the active group to 0.014 and 0.015 after 30 and 45 days, respectively (P=0.045 and P=0.033 compared with placebo). The sucralose concentration decreased from 35.8 mg to 27.9 mg and 29.1 mg (P=0.038 and P=0.026 compared with placebo) at the end of the supplementation period and after the follow-up, respectively. No significant differences were recorded in the placebo after 30 days or at the end of the follow-up., Conclusions: The association of the EPSs produced by S. thermophilus ST10 and tara gum seems capable of significantly improving the intestinal functional barrier in healthy subjects. A wider study in subjects presenting impaired gut permeability would be useful in the future to confirm the positive results from this pilot trial. In any case, our findings are consistent with the parallel increase in exopolysaccharide concentration in the faecal material, thus suggesting the effective ability of the strain used to secrete EPSs in the gut lumen. An innovative approach of this type may be useful in helping to restore the physiological barrier by means of a merely natural and mechanical action.

The complex network of specialized cells and molecules in the immune system has evolved to defend against pathogens, but inadvertent immune system attacks on "self" result in autoimmune disease. Both genetic regulation of immune cell levels and their relationships with autoimmunity are largely undetermined. Here, we report genetic contributions to quantitative levels of 95 cell types encompassing 272 immune traits, in a cohort of 1,629 individuals from four clustered Sardinian villages. We first estimated trait heritability, showing that it can be substantial, accounting for up to 87% of the variance (mean 41%). Next, by assessing ∼8.2 million variants that we identified and confirmed in an extended set of 2,870 individuals, 23 independent variants at 13 loci associated with at least one trait. Notably, variants at three loci (HLA, IL2RA, and SH2B3/ATXN2) overlap with known autoimmune disease associations. These results connect specific cellular phenotypes to specific genetic variants, helping to explicate their involvement in disease., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Genetic variation within the male-specific portion of the Y chromosome (MSY) can clarify the origins of contemporary populations, but previous studies were hampered by partial genetic information. Population sequencing of 1204 Sardinian males identified 11,763 MSY single-nucleotide polymorphisms, 6751 of which have not previously been observed. We constructed a MSY phylogenetic tree containing all main haplogroups found in Europe, along with many Sardinian-specific lineage clusters within each haplogroup. The tree was calibrated with archaeological data from the initial expansion of the Sardinian population ~7700 years ago. The ages of nodes highlight different genetic strata in Sardinia and reveal the presumptive timing of coalescence with other human populations. We calculate a putative age for coalescence of ~180,000 to 200,000 years ago, which is consistent with previous mitochondrial DNA-based estimates.

The evolutionary pathway of Palinuridae (Crustacea, Decapoda) is still controversial, uncertain and unexplored, expecially from a karyological point of view. Here we describe the South African spiny lobster Jasus lalandii karyotype: n and 2n values, heterochromatin distribution, nucleolar organizer region (NOR) location and telomeric repeat structure and location. To compare the genomic and chromosomal organization in Palinuridae we located NORs in Panulirus regius, Palinurus gilchristi and Palinurus mauritanicus: all species showed multiple NORs. In J. lalandii NORs were located on three chromosome pairs, with interindividual polymorphism. In P. regius and in the two Palinurus species NORs were located on two chromosome pairs. In the two last species 45S ribosomal gene loci were also found on B chromosomes. In addition, the nature and location of telomeric repeats were investigated by FISH in J. lalandii, P. gilchristi, P. mauritanicus Palinurus elephas, and P. regius (Palinuridae, Achelata), and in Scyllarus arctus (Scyllaridae, Achelata): all these Achelata species showed the (TTAGG)n pentameric repeats. Furthermore, in J. lalandii these repeats occurred in all the telomeres and in some interstitial chromosomal sites, associated with NORs.

Background: Lactobacilli and bifidobacteria are often associated with health-promoting effects. These live microorganisms, defined as probiotics, are commonly consumed as part of fermented foods, such as yoghurt and fermented milks, or as dietary supplements. Escherichia coli is a gram-negative, rod-shaped bacterium commonly found in the lower intestine of warm-blooded organisms. As a part of the normal gut microbiota, this microorganism colonizes the gastrointestinal tract of animals and humans within a few hours after birth. All E. coli strains can produce a wide variety of biogenic amines responsible for potentially harmful systemic intoxications. Enterohemorrhagic E. coli serotype O157:H7 is a pathotype of diarrhoeagenic strains with a large virulence plasmid pO157 able to produce 1 or more Shiga toxins., Methods: The overall aim of this study was to determine the inhibitory effects of different strains of probiotics on E. coli serotypes, including E. coli O157:H7 (CQ9485). In particular, the antagonistic activity of 4 Bifidobacterium strains (Probiotical SpA, Italy) and 16 lactic acid bacteria, more specifically 14 Lactobacillus spp. and 2 Streptococcus spp., was assessed against selected E. coli biotypes (ATCC 8739, ATCC 10536, ATCC 35218, and ATCC 25922). The diarrhoeagenic serotype O157:H7 was also tested., Results: The experimental data collected demonstrated an in vitro significant inhibitory effect of 6 Lactobacillus strains, namely L. rhamnosus LR04, L. rhamnosus LR06, L. plantarum LP01, L. plantarum LP02, L. pentosus LPS01, and L. delbrueckii subsp. delbrueckii LDD01, and 2 Bifidobacterium strains, B. breve BR03 and B. breve B632. The inhibiting extent was slightly different among these strains, with L. delbrueckii subsp. delbrueckii LDD01 showing the highest activity on E. coli O157:H7., Conclusions: Most of the probiotics studied are able to antagonize the growth of the 5 strains of E. coli tested, including the O157:H7 biotype, well known for their characteristic to produce a wide variety of biogenic amines considered responsible for dangerous systemic intoxications.

Identifying the genes that influence levels of pro-inflammatory molecules can help to elucidate the mechanisms underlying this process. We first conducted a two-stage genome-wide association scan (GWAS) for the key inflammatory biomarkers Interleukin-6 (IL-6), the general measure of inflammation erythrocyte sedimentation rate (ESR), monocyte chemotactic protein-1 (MCP-1), and high-sensitivity C-reactive protein (hsCRP) in a large cohort of individuals from the founder population of Sardinia. By analysing 731,213 autosomal or X chromosome SNPs and an additional ∼1.9 million imputed variants in 4,694 individuals, we identified several SNPs associated with the selected quantitative trait loci (QTLs) and replicated all the top signals in an independent sample of 1,392 individuals from the same population. Next, to increase power to detect and resolve associations, we further genotyped the whole cohort (6,145 individuals) for 293,875 variants included on the ImmunoChip and MetaboChip custom arrays. Overall, our combined approach led to the identification of 9 genome-wide significant novel independent signals-5 of which were identified only with the custom arrays-and provided confirmatory evidence for an additional 7. Novel signals include: for IL-6, in the ABO gene (rs657152, p = 2.13×10(-29)); for ESR, at the HBB (rs4910472, p = 2.31×10(-11)) and UCN119B/SPPL3 (rs11829037, p = 8.91×10(-10)) loci; for MCP-1, near its receptor CCR2 (rs17141006, p = 7.53×10(-13)) and in CADM3 (rs3026968, p = 7.63×10(-13)); for hsCRP, within the CRP gene (rs3093077, p = 5.73×10(-21)), near DARC (rs3845624, p = 1.43×10(-10)), UNC119B/SPPL3 (rs11829037, p = 1.50×10(-14)), and ICOSLG/AIRE (rs113459440, p = 1.54×10(-08)) loci. Confirmatory evidence was found for IL-6 in the IL-6R gene (rs4129267); for ESR at CR1 (rs12567990) and TMEM57 (rs10903129); for MCP-1 at DARC (rs12075); and for hsCRP at CRP (rs1205), HNF1A (rs225918), and APOC-I (rs4420638). Our results improve the current knowledge of genetic variants underlying inflammation and provide novel clues for the understanding of the molecular mechanisms regulating this complex process., Competing Interests: The authors have declared that no competing interests exist.

A genome-wide association scan of approximately 6.6 million genotyped or imputed variants in 882 Sardinian individuals with multiple sclerosis (cases) and 872 controls suggested association of CBLB gene variants with disease, which was confirmed in 1,775 cases and 2,005 controls (rs9657904, overall P = 1.60 x 10(-10), OR = 1.40). CBLB encodes a negative regulator of adaptive immune responses, and mice lacking the ortholog are prone to experimental autoimmune encephalomyelitis, the animal model of multiple sclerosis.

Background and Objectives: Hereditary hemochromatosis (HH) is an autosomal recessive disorder of iron metabolism. The HFE gene implicated in this disorder has been identified on chromosome 6 (6p21.3). The most prevalent mutation in HH patients changes the 282 cysteine residue to tyrosine (C282Y). The role of a second mutation which changes the 63 histidine to aspartic acid (H63D) in iron overload has been controversial. The aim of this study was to evaluate the effect of the H63D mutation on the ferritin levels of beta-thalassemia carriers., Design and Methods: beta-thalassemia carriers have a tendency to increase iron absorption because of mild anemia and slightly increased erythropoiesis. Differences in ferritin levels between homozygotes for H63D and wild type may indicate a modulator effect of the HFE mutation on iron absorption. We studied 152 healthy males, heterozygous for beta-thalassemia. Serum ferritin was measured by chemiluminescence. H63D genotypes were determined by digestion of polymerase chain reaction (PCR) products with MboI restriction enzyme., Results: Forty-five subjects were H63D heterozygotes and four subjects were H63D homozygotes. Ferritin levels were (mean +/- SD): 250 +/- 138 microg/L in homozygotes for the wild type H/H; 295 +/- 186 microg/L in H/D heterozygotes; and 389 +/- 75 microg/L in homozygotes for the mutation D/D. The difference in ferritin values between H/H and D/D is statistically significant (p=0.022)., Interpretation and Conclusions: beta-thalassemia carriers who are homozygotes for the H63D mutation have higher ferritin levels than beta-thalassemia carriers with the H/H genotype, suggesting that the H63D mutation may have a modulating effect on iron absorption.