A global metagenomic map of urban microbiomes and antimicrobial resistance.

We present a global atlas of 4,728 metagenomic samples from mass-transit systems in 60 cities over 3 years, representing the first systematic, worldwide catalog of the urban microbial ecosystem. This atlas provides an annotated, geospatial profile of microbial strains, functional characteristics, antimicrobial resistance (AMR) markers, and genetic elements, including 10,928 viruses, 1,302 bacteria, 2 archaea, and 838,532 CRISPR arrays not found in reference databases. We identified 4,246 known species of urban microorganisms and a consistent set of 31 species found in 97% of samples that were distinct from human commensal organisms. Profiles of AMR genes varied widely in type and density across cities. Cities showed distinct microbial taxonomic signatures that were driven by climate and geographic differences. These results constitute a high-resolution global metagenomic atlas that enables discovery of organisms and genes, highlights potential public health and forensic applications, and provides a culture-independent view of AMR burden in cities.

Relationships between diet and gut microbiome in an Italian and Dutch cohort: does the dietary protein to fiber ratio play a role?

PURPOSE: To investigate the relationships between the habitual diet, the protein to fiber ratio (P/F), and the gut microbiome in one Italian and one Dutch cohort of healthy subjects consuming an omnivore diet. METHODS: The Italian cohort included 19 males (M_IT, BMI 25.2 ± 0.72 kg/m2, age 25.4 ± 0.96 years) and 20 females (F_IT, BMI 23.9 ± 0.81 kg/m2, age 23.8 ± 0.54 years); the Dutch cohort included 30 females (F_NL, BMI: 23.9 ± 0.81 kg/m2, age: 23.8 ± 0.54 years). Individual diets were recorded through Food Frequency Questionnaires and analyzed to assess the nutrient composition. Gut microbiome was assessed in fecal samples. RESULTS: M_IT consumed higher levels of proteins than F_NL and F_IT, whereas dietary fiber intake did not differ among groups. Data showed that consumption of plant protein to animal protein (PP/AP) and PP to total proteins ratio can determine a differentiation of F_NL more than the absolute amount of dietary fiber. Conversely, the protein to fiber (P/F) and AP to total proteins better characterized M_IT. M_IT harbored the highest abundance of proteolytic microorganisms and the lowest microbial gene richness. Conversely, F_NL had more fiber-degrading microorganisms like Bacteroides thetaiotaomicron, Bacteroides xylanisolvens, Roseburia sp., Coprococcus eutactus and Parabacteroides along with the highest number of genes encoding carbohydrate-active enzymes and gene richness. It was predicted that by each unit decrease in the P/F a 3% increase in gene richness occurred. CONCLUSION: Study findings suggested that dietary P/F, rather than the absolute amount of dietary fiber, could contribute to the shaping of the microbiome towards a more proteolytic or fiber-degrading gut ecosystem. CLINICALTRIALS: gov Identifier NCT04205045-01-10-2018, retrospectively registered. Dutch Trial Register NTR7531-05-10-2018.

Aetiology, risk factors and microbiota composition in children with prolonged diarrhoea: A prospective case-controlled cohort study.

AIM: Prolonged diarrhoea (ProD) refers to acute-onset diarrhoea that persists for longer than 1 week. As the aetiology, risk factors and management are poorly defined, we prospectively enrolled children hospitalised in a high-income setting to assess these outcomes and investigate the potential role of gut microbiota. METHODS: All children aged 30 days to 14 years admitted for acute-onset diarrhoea lasting 7-14 days were included. Children consecutively admitted in the same period for acute diarrhoea (AD) served as controls. High-throughput sequencing of 16S rRNA gene amplicons was used to analyse stool samples from a subset of patients and healthy controls. RESULTS: Sixty-eight with ProD and 104 with AD were enrolled. Intestinal infections were the main aetiology of diarrhoea in both groups (ProD 92.9% vs. AD 97.8%). ProD children showed a higher prevalence of bacterial infections compared to AD (30.8% vs. 8.9%, p = 0.024). Neither age, host-related factors, nor microbiome alterations were specifically linked to ProD. However, ProD children had a more severe initial clinical presentation than AD. CONCLUSION: ProD is often the result of an unusually severe intestinal infection that runs a course longer than expected but generally resolves without further problems. No specific management or therapies should be undertaken in most cases.

Psychrotrophic Bacteria Equipped with Virulence and Colonization Traits Populate the Ice Cream Manufacturing Environment.

Several microbial taxa have been associated with food processing facilities, and they might resist by attaching on tools and equipment even after sanitation procedures, producing biofilms that adhere to the surfaces and might embed other microorganisms, including spoilers and pathogens. There is increasing evidence that these communities can be transferred to the final product. To explore the microbial contamination routes in a facility producing ice creams, we collected foods and environmental swabs from industrial surfaces of equipment and tools and performed taxonomic and functional analyses of the microbial DNA extracted from the environmental samples. Our results suggest that complex communities dominated by psychrotrophic bacteria (e.g., Pseudomonas and Acinetobacter spp.) inhabit the food processing environment, and we demonstrate that these communities might be transferred from the surfaces to the products. Functional analysis performed on environmental samples highlighted the presence of several genes linked to antimicrobial resistance and adherence on abiotic surfaces; such genes were more abundant on food contact (FC) than on other surfaces. Metagenome-assembled genomes (MAGs) of Pseudomonas stutzeri showed genes linked with biofilm formation and motility, which are surely linked to colonizing capabilities in the processing lines. The study highlights clear potential advantages of applying microbiome mapping in the food industry for source tracking of microbial contamination and for planning appropriate ad hoc sanitization strategies. IMPORTANCE Several microbial species might permanently establish in food processing facilities, thus contributing to food loss. In fact, food contact surfaces might transfer microorganisms to intermediates and products, potentially representing a hazard to human health. In this work, we provide evidence of the existence of complex microbial communities overcoming sanitation in an ice cream-producing facility. These communities harbored several genes that could potentially lead to attachment to surfaces and antimicrobial resistance. Also, prediction of routes of contamination showed that several potential spoilage taxa might end up in the final product. Importantly, in this work, we show that mapping the environmental microbiome is a high-resolution technique that might help food business operators ensure food quality and safety through detection of potentially hazardous microorganisms.

Host phenotype classification from human microbiome data is mainly driven by the presence of microbial taxa.

Machine learning-based classification approaches are widely used to predict host phenotypes from microbiome data. Classifiers are typically employed by considering operational taxonomic units or relative abundance profiles as input features. Such types of data are intrinsically sparse, which opens the opportunity to make predictions from the presence/absence rather than the relative abundance of microbial taxa. This also poses the question whether it is the presence rather than the abundance of particular taxa to be relevant for discrimination purposes, an aspect that has been so far overlooked in the literature. In this paper, we aim at filling this gap by performing a meta-analysis on 4,128 publicly available metagenomes associated with multiple case-control studies. At species-level taxonomic resolution, we show that it is the presence rather than the relative abundance of specific microbial taxa to be important when building classification models. Such findings are robust to the choice of the classifier and confirmed by statistical tests applied to identifying differentially abundant/present taxa. Results are further confirmed at coarser taxonomic resolutions and validated on 4,026 additional 16S rRNA samples coming from 30 public case-control studies.

Outlook on next-generation probiotics from the human gut.

Probiotics currently available on the market generally belong to a narrow range of microbial species. However, recent studies about the importance of the gut microbial commensals on human health highlighted that the gut microbiome is an unexplored reservoir of potentially beneficial microbes. For this reason, academic and industrial research is focused on identifying and testing novel microbial strains of gut origin for the development of next-generation probiotics. Although several of these are promising for the prevention and treatment of many chronic diseases, studies on human subjects are still scarce and approval from regulatory agencies is, therefore, rare. In addition, some issues need to be overcome before implementing their wide application on the market, such as the best methods for cultivation and storage of these oxygen-sensitive taxa. This review summarizes the most recent evidence related to NGPs and provides an outlook to the main issues that still limit their wide employment.

Impact of oral antiviral therapy against HCV on gut microbiota. A prospective study.

The intestinal microbiota plays a fundamental role in physiological homeostasis as well as in pathologic conditions. Hepatitis C virus is the leading cause of chronic liver diseases worldwide. The treatment of this infection has been revolutionized by the availability of direct-acting antiviral agents which guarantee a high rate (about 95%) of viral clearance. Few studies have assessed the change in the gut microbiota of patients treated with direct-acting antiviral agents against HCV, and many aspects still need to be clarified. The aim of the study was to evaluate the effects of antiviral therapy on gut microbiota. We enrolled patients with HCV-related chronic liver disease attending the Infectious Diseases Unit of the A.O.U. Federico II of Naples from January 2017 to March 2018 and treated with DAAs. For each patient, a fecal sample was collected and analyzed for the assessment of microbial diversity before the start of therapy and by SVR12 time. We excluded patients who had received antibiotics in the previous 6 months. Twelve patients were enrolled (6 male, 8 genotype 1 (1 subtype 1a), 4 genotype 2). Fibrosis scores were F0 in 1 patient, F2 in 1 patient, F3 in 4 patients and cirrhosis in the remaining 6 (all in Child-Pugh class A). All were treated with DAAs for 12 weeks (5 with Paritaprevir-Ombitasvir-Ritonavir-Dasabuvir, 3 with Sofosbuvir-Ledipasvir, 1 with Sofosbuvir-Ribavirin, 1 with Sofosbuvir-Daclatasvir, 1 with Sofosbuvir-Velpatasvir) and 100% achieved SVR12. In all patients, we observed a trend in reduction of potentially pathogenic microorganisms (i.e., Enterobacteriaceae). Furthermore, a trend of increase in α-diversity was observed in patients by SVR12 compared to baseline. This trend was markedly more evident in patients without liver cirrhosis than in those with cirrhosis. Our study shows that viral eradication obtained with DAA is associated with a trend in restoring the heterogeneity of α-diversity and in reducing the percentage of potentially pathogenic microbial species, although this benefit is less evident in patients with cirrhosis. Further studies with larger sample size are needed to confirm these data.

Stool microRNA profiles reflect different dietary and gut microbiome patterns in healthy individuals.

OBJECTIVES: MicroRNA (miRNA) profiles have been evaluated in several biospecimens in relation to common diseases for which diet may have a considerable impact. We aimed at characterising how specific diets are associated with the miRNome in stool of vegans, vegetarians and omnivores and how this is reflected in the gut microbial composition, as this is still poorly explored. DESIGN: We performed small RNA and shotgun metagenomic sequencing in faecal samples and dietary recording from 120 healthy volunteers, equally distributed for the different diets and matched for sex and age. RESULTS: We found 49 miRNAs differentially expressed among vegans, vegetarians and omnivores (adj. p <0.05) and confirmed trends of expression levels of such miRNAs in vegans and vegetarians compared with an independent cohort of 45 omnivores. Two miRNAs related to lipid metabolism, miR-636 and miR-4739, were inversely correlated to the non-omnivorous diet duration, independently of subject age. Seventeen miRNAs correlated (|rho|>0.22, adj. p <0.05) with the estimated intake of nutrients, particularly animal proteins, phosphorus and, interestingly, lipids. In omnivores, higher Prevotella and Roseburia and lower Bacteroides abundances than in vegans and vegetarians were observed. Lipid metabolism-related miR-425-3p and miR-638 expression levels were associated with increased abundances of microbial species, such as Roseburia sp. CAG 182 and Akkermansia muciniphila, specific of different diets. An integrated analysis identified 25 miRNAs, 25 taxa and 7 dietary nutrients that clearly discriminated (area under the receiver operating characteristic curve=0.89) the three diets. CONCLUSION: Stool miRNA profiles are associated with specific diets and support the role of lipids as a driver of epigenetic changes and host-microbial molecular interactions in the gut.

Therapeutic effects elicited by the probiotic Lacticaseibacillus rhamnosus GG in children with atopic dermatitis. The results of the ProPAD trial.

BACKGROUND: Atopic dermatitis (AD) is a chronic inflammatory skin disease affecting up to 20% of the pediatric population associated with alteration of skin and gut microbiome. Probiotics have been proposed for AD treatment. The ProPAD study aimed to investigate the therapeutic effects of the probiotic Lacticaseibacillus rhamnosus GG (LGG) in children with AD. METHODS: In total, 100 AD patients aged 6-36 months were enrolled in a randomized, double-blind, controlled trial to receive placebo (Group A) or LGG (1 x 1010 CFU/daily) (Group B) for 12 weeks. The primary outcome was the evaluation of the efficacy of LGG supplementation on AD severity comparing the Scoring Atopic Dermatitis (SCORAD) index at baseline (T0) and at 12-week (T12). A reduction of ≥8.7 points on the SCORAD index was considered as minimum clinically important difference (MCID). The secondary outcomes were the SCORAD index evaluation at 4-week (T16) after the end of LGG treatment, number of days without rescue medications, changes in Infant Dermatitis Quality Of Life questionnaire (IDQOL), gut microbiome structure and function, and skin microbiome structure. RESULTS: The rate of subjects achieving MCID at T12 and at T16 was higher in Group B (p < .05), and remained higher at T16 (p < .05)The number of days without rescue medications was higher in Group B. IDQOL improved at T12 in the Group B (p < .05). A beneficial modulation of gut and skin microbiome was observed only in Group B patients. CONCLUSIONS: The probiotic LGG could be useful as adjunctive therapy in pediatric AD. The beneficial effects on disease severity and quality of life paralleled with a beneficial modulation of gut and skin microbiome.

Mediterranean diet consumption affects the endocannabinoid system in overweight and obese subjects: possible links with gut microbiome, insulin resistance and inflammation.

PURPOSE: To investigate whether a Mediterranean diet (MD) affected the plasma concentrations of endocannabinoids (ECs), N-acylethanolamines (NAEs) and their specific ratios in subjects with lifestyle risk factors for metabolic diseases. To identify the relationship between circulating levels of these compounds and gut microbiome, insulin resistance and systemic inflammation. METHODS: A parallel 8-week randomised controlled trial was performed involving 82 overweight and obese subjects aged (mean ± SEM) 43 ± 1.4 years with a BMI of 31.1 ± 0.5 kg/m2, habitual Western diet (CT) and sedentary lifestyle. Subjects were randomised to consume an MD tailored to their habitual energy and macronutrient intake (n = 43) or to maintain their habitual diet (n = 39). Endocannabinoids and endocannabinoid-like molecules, metabolic and inflammatory markers and gut microbiome were monitored over the study period. RESULTS: The MD intervention lowered plasma arachidonoylethanolamide (AEA, p = 0.02), increased plasma oleoylethanolamide/palmitoylethanolamide (OEA/PEA, p = 0.009) and OEA/AEA (p = 0.006) and increased faecal Akkermansia muciniphila (p = 0.026) independent of body weight changes. OEA/PEA positively correlated with abundance of key microbial players in diet-gut-health interplay and MD adherence. Following an MD, individuals with low-plasma OEA/PEA at baseline decreased homeostatic model assessment of insulin resistance index (p = 0.01), while individuals with high-plasma OEA/PEA decreased serum high-sensitive C-reactive protein (p = 0.02). CONCLUSIONS: We demonstrated that a switch from a CT to an isocaloric MD affects the endocannabinoid system and increases A. muciniphila abundance in the gut independently of body weight changes. Endocannabinoid tone and microbiome functionality at baseline drives an individualised response to an MD in ameliorating insulin sensitivity and inflammation. Clinical Trial Registry number and website NCT03071718; www.clinicaltrials.gov.

Mediterranean diet intervention in overweight and obese subjects lowers plasma cholesterol and causes changes in the gut microbiome and metabolome independently of energy intake.

OBJECTIVES: This study aimed to explore the effects of an isocaloric Mediterranean diet (MD) intervention on metabolic health, gut microbiome and systemic metabolome in subjects with lifestyle risk factors for metabolic disease. DESIGN: Eighty-two healthy overweight and obese subjects with a habitually low intake of fruit and vegetables and a sedentary lifestyle participated in a parallel 8-week randomised controlled trial. Forty-three participants consumed an MD tailored to their habitual energy intakes (MedD), and 39 maintained their regular diets (ConD). Dietary adherence, metabolic parameters, gut microbiome and systemic metabolome were monitored over the study period. RESULTS: Increased MD adherence in the MedD group successfully reprogrammed subjects' intake of fibre and animal proteins. Compliance was confirmed by lowered levels of carnitine in plasma and urine. Significant reductions in plasma cholesterol (primary outcome) and faecal bile acids occurred in the MedD compared with the ConD group. Shotgun metagenomics showed gut microbiome changes that reflected individual MD adherence and increase in gene richness in participants who reduced systemic inflammation over the intervention. The MD intervention led to increased levels of the fibre-degrading Faecalibacterium prausnitzii and of genes for microbial carbohydrate degradation linked to butyrate metabolism. The dietary changes in the MedD group led to increased urinary urolithins, faecal bile acid degradation and insulin sensitivity that co-varied with specific microbial taxa. CONCLUSION: Switching subjects to an MD while maintaining their energy intake reduced their blood cholesterol and caused multiple changes in their microbiome and metabolome that are relevant in future strategies for the improvement of metabolic health.

A Mediterranean Diet Intervention Reduces the Levels of Salivary Periodontopathogenic Bacteria in Overweight and Obese Subjects.

The human oral cavity is a complex ecosystem, and the alterations in salivary microbial communities are associated with both oral and nonoral diseases. The Mediterranean diet (MD) is a healthy dietary pattern useful for both prevention and treatment of several diseases. To further explore the effects of the MD on human health, in this study, we investigated the changes in the salivary microbial communities in overweight/obese subjects after an individually tailored MD-based nutritional intervention. Healthy overweight and obese subjects were randomized between two intervention groups. The MD group (Med-D group) increased their MD adherence during 8 weeks of intervention while the control diet (control-D) group did not change their dietary habits. The salivary microbiota was assessed at baseline and after 4 and 8 weeks of intervention. Despite no observed changes in the overall salivary microbiota composition, we found a significant decrease in the relative abundances of species-level operational taxonomic units annotated as Porphyromonas gingivalis, Prevotella intermedia, and Treponema denticola in the Med-D group compared to that in the control-D group after 8 weeks of intervention, which are known to be associated with periodontal disease. Such variations were significantly linked to dietary variables such as MD adherence rates and intakes of animal versus vegetable proteins. In addition, increased levels of Streptococcus cristatus were observed in the Med-D group, which has been reported as an antagonistic taxon inhibiting P. gingivalis gene expression. Our findings suggest that an MD-based nutritional intervention may be implicated in reducing periodontal bacteria, and an MD may be a dietary strategy supportive of oral homeostasis.IMPORTANCE Changes in dietary behavior with increased adherence to a Mediterranean diet can determine a reduction of periodontopathogenic bacterial abundances in the saliva of overweight subjects with cardiometabolic risk due to an unhealthy lifestyle, without any change in individual energy intake, nutrient intake, and physical activity.

Attenuated Lactococcus lactis and Surface Bacteria as Tools for Conditioning the Microbiota and Driving the Ripening of Semisoft Caciotta Cheese.

This study aimed at establishing the effects of attenuated starters and surface bacteria on various features of caciotta cheese. The cheese undergoes a ripening period during which the house microbiota contaminates the surface. Conventional cheese (the control cheese [CC]) is made using only primary starters. Primary starters and attenuated (i.e., unable to grow and synthesize lactic acid) Lactococcus lactis (Lc. lactis) subsp. lactis were used to produce caciotta cheese without (ATT cheese) or with an inoculum of surface bacteria: (i) Leuconostoc lactis (Le. lactis) (LL cheese), (ii) Vibrio casei (VC cheese), (iii) Staphylococcus equorum (SE cheese), (iv) Brochothrix thermosphacta (BX cheese), and (v) a mixture of all four (MIX cheese). Attenuated Lc. lactis increased microbial diversity during cheese ripening. At the core, attenuated starter mainly increased indigenous lactococci and Lactobacillus delbrueckii group bacteria. At the surface, the main effect was on Macrococcus caseolyticus Autochthonous Le. lactis strains took advantage of the attenuated starter, becoming dominant. Adjunct Le. lactis positively affected Lactobacillus sakei group bacteria on the LL cheese surface. Adjunct V. casei, S. equorum, and B. thermosphacta did not become dominant. Surfaces of VC, SE, and BX cheeses mainly harbored Staphylococcus succinus Peptidase activities were higher in cheeses made with attenuated starter than in CC, which had the lowest concentration of free amino acids. Based on the enzymatic activities of adjunct Le. lactis, LL and MIX cheeses exhibited the highest glutamate dehydrogenase, cystathionine-γ-lyase, and esterase activities. As shown by multivariate statistical analyses, LL and MIX cheeses showed the highest similarity for microbiological and biochemical features. LL and MIX cheeses received the highest scores for overall sensory acceptability.IMPORTANCE This study provides in-depth knowledge of the effects of attenuated starters and surface bacterial strains on the microbiota and related metabolic activities during cheese ripening. The use of attenuated Lc. lactis strongly impacted the microbiota assembly of caciotta cheese. This led to improved biochemical and sensory features compared to conventional cheese. Among surface bacterial strains, Le. lactis played a key role in the metabolic activities involved in cheese ripening. This resulted in an improvement of the sensory quality of caciotta cheese. The use of attenuated lactic acid bacteria and the surface adjunct Le. lactis could be a useful biotechnology to improve the flavor formation of caciotta cheese.

One ring to rule them all: an ecosystem engineer fungus fosters plant and microbial diversity in a Mediterranean grassland.

Species coexistence in grasslands is regulated by several environmental factors and interactions with the soil microbial community. Here, the development of the Basidiomycetes fungus Agaricus arvensis, forming fairy rings, in a species-rich Mediterranean grassland, is described. Effects of the mycelial front on plants, fungi and bacteria were assessed by vegetation survey and next generation sequencing approaches. Our results showed a fungal-dependent shift in the community structure operated by a wave-like spread of fairy rings that decreased plant, fungal and bacterial diversity, indicating a detrimental effect of fairy rings on most species. The fairy rings induced successional processes in plants that enhanced the replacement of a community dominated by perennial plants with short-living and fast-growing plant species. In parallel, fungal and bacterial communities showed evident differences in species composition with several taxa associated within distinct sampling zone across the fairy rings. Notably, bacteria belonging to the Burkholderia genus and fungi of the genus Trichoderma increased in response to the advancing mycelium of A. arvensis. The profound changes in community composition and the overall increase in taxa diversity at ecosystemic scale suggest that fairy ring-forming fungi may act as ecosystem engineer species in Mediterranean grasslands.

Altered gut microbiota and endocannabinoid system tone in vitamin D deficiency-mediated chronic pain.

Recent evidence points to the gut microbiota as a regulator of brain and behavior, although it remains to be determined if gut bacteria play a role in chronic pain. The endocannabinoid system is implicated in inflammation and chronic pain processing at both the gut and central nervous system (CNS) levels. In the present study, we used low Vitamin D dietary intake in mice and evaluated possible changes in gut microbiota, pain processing and endocannabinoid system signaling. Vitamin D deficiency induced a lower microbial diversity characterized by an increase in Firmicutes and a decrease in Verrucomicrobia and Bacteroidetes. Concurrently, vitamin D deficient mice showed tactile allodynia associated with neuronal hyperexcitability and alterations of endocannabinoid system members (endogenous mediators and their receptors) at the spinal cord level. Changes in endocannabinoid (anandamide and 2-arachidonoylglycerol) levels were also observed in the duodenum and colon. Remarkably, the anti-inflammatory anandamide congener, palmitoylethanolamide, counteracted both the pain behaviour and spinal biochemical changes in vitamin D deficient mice, whilst increasing the levels of Akkermansia, Eubacterium and Enterobacteriaceae, as compared with vehicle-treated mice. Finally, induction of spared nerve injury in normal or vitamin D deficient mice was not accompanied by changes in gut microbiota composition. Our data suggest the existence of a link between Vitamin D deficiency - with related changes in gut bacterial composition - and altered nociception, possibly via molecular mechanisms involving the endocannabinoid and related mediator signaling systems.

Strain-Level Diversity Analysis of Pseudomonas fragi after In Situ Pangenome Reconstruction Shows Distinctive Spoilage-Associated Metabolic Traits Clearly Selected by Different Storage Conditions.

Microbial spoilage of raw meat causes huge economic losses every year. An understanding of the microbial ecology associated with the spoilage and its dynamics during the refrigerated storage of meat can help in preventing and delaying the spoilage-related activities. The raw meat microbiota is usually complex, but only a few members will develop during storage and cause spoilage upon the pressure from several external factors, such as temperature and oxygen availability. We characterized the metagenome of beef packed aerobically or under vacuum during refrigerated storage to explore how different packaging conditions may influence the microbial composition and potential spoilage-associated activities. Different population dynamics and spoilage-associated genomic repertoires occurred in beef stored aerobically or in vacuum packaging. Moreover, the pangenomes of Pseudomonas fragi strains were extracted from metagenomes. We demonstrated the presence of specific, storage-driven strain-level profiles of Pseudomonas fragi, characterized by different gene repertoires and thus potentially able to act differently during meat spoilage. The results provide new knowledge on strain-level microbial ecology associated with meat spoilage and may be of value for future strategies of spoilage prevention and food waste reduction.IMPORTANCE This work provides insights on the mechanisms involved in raw beef spoilage during refrigerated storage and on the selective pressure exerted by the packaging conditions. We highlighted the presence of different microbial metagenomes during the spoilage of beef packaged aerobically or under vacuum. The packaging condition was able to select specific Pseudomonas fragi strains with distinctive genomic repertoires. This study may help in deciphering the behavior of different biomes directly in situ in food and in understanding the specific contribution of different strains to food spoilage.

The Intestinal Microbiota of Hermetia illucens Larvae Is Affected by Diet and Shows a Diverse Composition in the Different Midgut Regions.

The larva of the black soldier fly (Hermetia illucens) has emerged as an efficient system for the bioconversion of organic waste. Although many research efforts are devoted to the optimization of rearing conditions to increase the yield of the bioconversion process, microbiological aspects related to this insect are still neglected. Here, we describe the microbiota of the midgut of H. illucens larvae, showing the effect of different diets and midgut regions in shaping microbial load and diversity. The bacterial communities residing in the three parts of the midgut, characterized by remarkable changes in luminal pH values, differed in terms of bacterial numbers and microbiota composition. The microbiota of the anterior part of the midgut showed the highest diversity, which gradually decreased along the midgut, whereas bacterial load had an opposite trend, being maximal in the posterior region. The results also showed that the influence of the microbial content of ingested food was limited to the anterior part of the midgut, and that the feeding activity of H. illucens larvae did not significantly affect the microbiota of the substrate. Moreover, a high protein content compared to other macronutrients in the feeding substrate seemed to favor midgut dysbiosis. The overall data indicate the importance of taking into account the presence of different midgut structural and functional domains, as well as the substrate microbiota, in any further study that aims at clarifying microbiological aspects concerning H. illucens larval midgut.IMPORTANCE The demand for food of animal origin is expected to increase by 2050. Since traditional protein sources for monogastric diets are failing to meet the increasing demand for additional feed production, there is an urgent need to find alternative protein sources. The larvae of Hermetia illucens emerge as efficient converters of low-quality biomass into nutritionally valuable proteins. Many studies have been performed to optimize H. illucens mass rearing on a number of organic substrates and to quantitatively and qualitatively maximize the biomass yield. On the contrary, although the insect microbiota can be fundamental for bioconversion processes and its characterization is mandatory also for safety aspects, this topic is largely overlooked. Here, we provide an in-depth study of the microbiota of H. illucens larval midgut, taking into account pivotal aspects, such as the midgut spatial and functional regionalization, as well as microbiota and nutrient composition of the feeding substrate.

Laboratory medicine: health evaluation in elite athletes.

The need to evaluate the health status of an athlete represents a crucial aim in preventive and protective sports science in order to identify the best diagnostic strategy to improve performance and reduce risks related to physical exercise. In the present review we aim to define the main biochemical and haematological markers that vary significantly during and after sports training to identify risk factors, at competitive and professional levels and to highlight the set up of a specific parameter's panel for elite athletes. Moreover, we also intend to consider additional biomarkers, still under investigation, which could further contribute to laboratory sports medicine and provide reliable data that can be used by athlete's competent staff in order to establish personal attitudes and prevent sports injuries.

Large-scale mapping of microbial diversity in artisanal Brazilian cheeses.

Brazilian artisanal cheeses are characterized by the use of raw milk and in some cases, natural starter cultures, known as "pingo", as well as following simple and traditional manufacturing technology. In this study, a large-scale screening of the microbial ecology of 11 different types of artisanal cheeses produced in five geographical areas of Brazil was performed. Besides, the specific origin-related microbial signatures were identified. Clear geography- and technology-based differences in the microbiota were observed. Lactic acid bacteria dominated in all cheeses although Enterobacteriaceae and Staphylococcus also occurred in North, Northeast and Central cheeses. Differences in the lactic acid bacteria patterns were also highlighted: Streptococcus, Leuconostoc, Lactococcus and Lactobacillus were differently combined in terms of relative abundance according to product type and region of production. This study provides a comprehensive, unprecedented microbiological mapping of Brazilian cheeses, highlighting the impact of geographical origin and mode of production on microbial diversity. The results obtained will help to plan an evaluation of microbial contamination sources that will need to be studied for the improvement of cheese quality and safety.

Midgut microbiota and host immunocompetence underlie Bacillus thuringiensis killing mechanism.

Bacillus thuringiensis is a widely used bacterial entomopathogen producing insecticidal toxins, some of which are expressed in insect-resistant transgenic crops. Surprisingly, the killing mechanism of B. thuringiensis remains controversial. In particular, the importance of the septicemia induced by the host midgut microbiota is still debated as a result of the lack of experimental evidence obtained without drastic manipulation of the midgut and its content. Here this key issue is addressed by RNAi-mediated silencing of an immune gene in a lepidopteran host Spodoptera littoralis, leaving the midgut microbiota unaltered. The resulting cellular immunosuppression was characterized by a reduced nodulation response, which was associated with a significant enhancement of host larvae mortality triggered by B. thuringiensis and a Cry toxin. This was determined by an uncontrolled proliferation of midgut bacteria, after entering the body cavity through toxin-induced epithelial lesions. Consequently, the hemolymphatic microbiota dramatically changed upon treatment with Cry1Ca toxin, showing a remarkable predominance of Serratia and Clostridium species, which switched from asymptomatic gut symbionts to hemocoelic pathogens. These experimental results demonstrate the important contribution of host enteric flora in B. thuringiensis-killing activity and provide a sound foundation for developing new insect control strategies aimed at enhancing the impact of biocontrol agents by reducing the immunocompetence of the host.