Sterile inflammation via TRPM8 RNA-dependent TLR3-NF-kB/IRF3 activation promotes antitumor immunity in prostate cancer.

Inflammation is a common condition of prostate tissue, whose impact on carcinogenesis is highly debated. Microbial colonization is a well-documented cause of a small percentage of prostatitis cases, but it remains unclear what underlies the majority of sterile inflammation reported. Here, androgen- independent fluctuations of PSA expression in prostate cells have lead us to identify a prominent function of the Transient Receptor Potential Cation Channel Subfamily M Member 8 (TRPM8) gene in sterile inflammation. Prostate cells secret TRPM8 RNA into extracellular vesicles (EVs), which primes TLR3/NF-kB-mediated inflammatory signaling after EV endocytosis by epithelial cancer cells. Furthermore, prostate cancer xenografts expressing a translation-defective form of TRPM8 RNA contain less collagen type I in the extracellular matrix, significantly more infiltrating NK cells, and larger necrotic areas as compared to control xenografts. These findings imply sustained, androgen-independent expression of TRPM8 constitutes as a promoter of anticancer innate immunity, which may constitute a clinically relevant condition affecting prostate cancer prognosis.

Adaptation Response in Sheep: Ewes in Different Cortisol Clusters Reveal Changes in the Expression of Salivary miRNAs.

Farm procedures have an impact on animal welfare by activating the hypothalamic-pituitary-adrenal axis that induces a wide array of physiological responses. This adaptive system guarantees that the animal copes with environmental variations and it induces metabolic and molecular changes that can be quantified. MicroRNAs (miRNAs) play a key role in the regulation of homeostasis and emerging evidence has identified circulating miRNAs as promising biomarkers of stress-related disorders in animals. Based on a clustering analysis of salivary cortisol trends and levels, 20 ewes were classified into two different clusters. The introduction of a ram in the flock was identified as a common farm practice and reference time point to collect saliva samples. Sixteen miRNAs related to the adaptation response were selected. Among them, miR-16b, miR-21, miR-24, miR-26a, miR-27a, miR-99a, and miR-223 were amplified in saliva samples. Cluster 1 was characterized by a lower expression of miR-16b and miR-21 compared with Cluster 2 (p < 0.05). This study identified for the first time several miRNAs expressed in sheep saliva, pointing out significant differences in the expression patterns between the cortisol clusters. In addition, the trend analyses of these miRNAs resulted in clusters (p = 0.017), suggesting the possible cooperation of miR-16b and -21 in the integrated stress responses, as already demonstrated in other species as well. Other research to define the role of these miRNAs is needed, but the evaluation of the salivary miRNAs could support the selection of ewes for different profiles of response to sources of stressors common in the farm scenario.

Productive Performances of Slow-Growing Chicken Breeds and Their Crosses with a Commercial Strain in Conventional and Free-Range Farming Systems.

Local chicken breeds play a vital role in promoting sustainability by preserving genetic diversity, enhancing resilience, and supporting local economies. These breeds are adapted to local climates and conditions, requiring fewer external resources and inputs for their maintenance. By conserving and utilizing local chicken breeds, sustainable farming practices can be incentivized, maintaining ecosystem balance and ensuring food security for future generations. The present study aimed at evaluating the growth performance and slaughter traits of two local Italian chicken breeds (Bionda Piemontese and Robusta Maculata) and their crosses with a medium-growth genotype (Sasso chicken®) reared in conventional and free-range farming systems. The conventional system used a high-energy high-protein diet in a closed barn with controlled temperature, humidity, and lighting, and a stocking density of 33 kg/m2. The free-range system used a low-input diet (low-energy low-protein diet composed of local and GMO-free feed ingredients), uncontrolled environmental conditions, and a stocking density of 21 kg/m2 in a barn with free access to an outdoor area. The birds were slaughtered at 84 days of age in both systems. The crossbred chickens showed the best results for growth performance in both farming systems compared to local breeds. Within genotype, the final live weight of chickens was similar in the two farming systems. In conclusion, slow-growth crossbreeds should be used in alternative farming systems, demonstrating better performance than pure local breeds.

The role of tumor microenvironment in drug resistance: emerging technologies to unravel breast cancer heterogeneity.

Breast cancer is a highly heterogeneous disease, at both inter- and intra-tumor levels, and this heterogeneity is a crucial determinant of malignant progression and response to treatments. In addition to genetic diversity and plasticity of cancer cells, the tumor microenvironment contributes to tumor heterogeneity shaping the physical and biological surroundings of the tumor. The activity of certain types of immune, endothelial or mesenchymal cells in the microenvironment can change the effectiveness of cancer therapies via a plethora of different mechanisms. Therefore, deciphering the interactions between the distinct cell types, their spatial organization and their specific contribution to tumor growth and drug sensitivity is still a major challenge. Dissecting intra-tumor heterogeneity is currently an urgent need to better define breast cancer biology and to develop therapeutic strategies targeting the microenvironment as helpful tools for combined and personalized treatment. In this review, we analyze the mechanisms by which the tumor microenvironment affects the characteristics of tumor heterogeneity that ultimately result in drug resistance, and we outline state of the art preclinical models and emerging technologies that will be instrumental in unraveling the impact of the tumor microenvironment on resistance to therapies.

Challenging the Hypothesis of in Utero Microbiota Acquisition in Healthy Canine and Feline Pregnancies at Term: Preliminary Data.

At present, there are no data on the presence of bacteria in healthy canine and feline pregnancies at term. Here, we investigated the uterine microbiome in bitches (n = 5) and queens (n = 3) undergoing elective cesarean section in two facilities. Samples included swabs from the endometrium, amniotic fluid, and meconium, and environmental swabs of the surgical tray as controls. Culture and 16S rRNA gene sequencing were used to investigate the presence of bacteria. Culture was positive for 34.3% of samples (uterus n = 3, amniotic fluid n = 2, meconium n = 4, controls n = 0), mostly with low growth of common contaminant bacteria. With sequencing techniques, the bacterial abundance was significantly lower than in environmental controls (p < 0.05). Sequencing results showed a species-specific pattern, and significant differences between canine and feline bacterial populations were found at order, family, and genus level. No differences were found in alpha and beta diversities between feto-maternal tissues and controls (p > 0.05). Dominant phyla were Bacteroidetes, Firmicutes, and Proteobacteria in different proportions based on tissue and species. Culture and sequencing results suggest that the bacterial biomass is very low in healthy canine and feline pregnancies at term, that bacteria likely originate from contamination from the dam's skin, and that the presence of viable bacteria could not be confirmed most of the time.

Potential Impact of Environmental Pollution by Human Antivirals on Avian Influenza Virus Evolution.

Antiviral (AV) drugs are the main line of defense against pandemic influenza. However, different administration policies are applied in countries with different stocks of AV drugs. These policies lead to different occurrences of drug metabolites in the aquatic environment, altering animal behavior with evolutionary consequences on viruses. The aim of this study was to investigate the potential impact of environmental pollution by human antivirals, such as oseltamivir carboxylate (OC), on the evolutionary rate of avian influenza. We used NA, HA, NP, and MP viral segments from two groups of neighboring countries sharing migratory routes of wild birds and characterized by different AV stockpiles. BEAST analyses were performed using the uncorrelated lognormal clock evolutionary model and the Bayesian skyline tree prior model. The ratios between the rate of evolution of the NA gene and the HA, NP, and MP segments were considered. The two groups of countries were compared by analyzing the differences in the ratio distributions. Our analyses highlighted a possible different behavior in the evolution of H5N1 2.3 clade viral strains when OC environmental pollution is present. In conclusion, the widespread consumption of antivirals and their presence in wastewater could influence the selective pressure on viruses.

Dog Owners' Attitude toward Veterinary Antibiotic Use and Antibiotic Resistance with a Focus on Canine Diarrhea Management.

An ad hoc questionnaire was designed in order to investigate AMR knowledge amongst Italian dog owners, owner expectations concerning pharmacological treatment of canine AD, and client attitudes towards and compliance with alternative strategies to antimicrobial administration. A total of 250 questionnaires were returned. Most of respondents were female, aged 36-70 and workers. More than a half of participants owned one dog with mixed breed, with Labrador retriever, golden retriever, dachshund, and border collie being the most represented breeds. On average, each dog was treated with an oral antibiotic 1.044 times per year. Intestinal diseases were among the main reasons (19%) for antibiotic prescription. Oral antibiotic courses without veterinary consultation (21%) and anticipated termination of the therapy (17.1%) were less common than reported elsewhere. The majority of respondents knew the meaning of AMR with a significant inverse association between the level of education and the tendency to administer antimicrobials without consulting a clinician (p = 0.004). Most of the owners expected a rapid recovery of clinical signs after a first episode of AD and accepted natural dietary supplementation for treating the condition. Ninety-five percent of the respondents believed that public funding should be spent to study AMR. Even though an acceptable degree of AMR awareness emerged, we feel that further efforts should be made to increase public AMR knowledge and to stimulate proactive measures to fight the phenomenon. On the other hand, the development of guidelines for the treatment of uncomplicated canine AD would help clinicians to rationalize antimicrobial use.

Predicting outcome in dogs with diffuse large B-cell lymphoma with a novel immune landscape signature.

Canine diffuse large B-cell lymphoma (cDLBCL) is characterized by high mortality and clinical heterogeneity. Although chemo-immunotherapy improves outcome, treatment response remains mainly unpredictable. To identify a set of immune-related genes aberrantly regulated and impacting the prognosis, we explored the immune landscape of cDLBCL by NanoString. The immune gene expression profile of 48 fully clinically characterized cDLBCLs treated with chemo-immunotherapy was analyzed with the NanoString nCounter Canine IO Panel using RNA extracted from tumor tissue paraffin blocks. A Cox proportional-hazards model was used to design a prognostic gene signature. The Cox model identified a 6-gene signature (IL2RB, BCL6, TXK, C2, CDKN2B, ITK) strongly associated with lymphoma-specific survival, from which a risk score was calculated. Dogs were assigned to high-risk or low-risk groups according to the median score. Thirty-nine genes were differentially expressed between the 2 groups. Gene set analysis highlighted an upregulation of genes involved in complement activation, cytotoxicity, and antigen processing in low-risk dogs compared with high-risk dogs, whereas genes associated with cell cycle were downregulated in dogs with a lower risk. In line with these results, cell type profiling suggested the abundance of natural killer and CD8+ cells in low-risk dogs compared with high-risk dogs. Furthermore, the prognostic power of the risk score was validated in an independent cohort of cDLBCL. In conclusion, the 6-gene-derived risk score represents a robust biomarker in predicting the prognosis in cDLBCL. Moreover, our results suggest that enhanced tumor antigen recognition and cytotoxic activity are crucial in achieving a more effective response to chemo-immunotherapy.

Antibiotic resistant Escherichia coli in wild birds hospitalised in a wildlife rescue centre.

The aim of this work was to evaluate the consequence of a hospitalisation period on antimicrobial resistance in Escherichia coli isolated from wild bird species admitted in the wildlife rescue centre of the Department of Veterinary Sciences (Turin University, Italy). Samples were collected from 121 raptors and 51 synanthropic animals, at the time of arrival as well as 5 and 10 days afterwards for a total of 372 faecal samples, and the susceptibility of E. coli strains was tested to a panel of seven antibacterials. Of the total, 109 animals (63.37 %) presented at least one sample positive for E. coli, 36 strains (39.6 %) were multi-drug resistant (MDR) and 12 (13.2 %) were extended spectrum beta-lactamase (ESBL)-producing E. coli. During the first 10 days of hospitalisation E. coli strains increased the number of resistances towards each antimicrobial principle, the number of ESBL E. coli and the therapy with fluoroquinolones developed resistance towards ceftriaxone, marbofloxacin, sulfamethoxazole-trimethoprim and tetracycline. Our results suggest that wild birds act as reservoirs of MDR bacteria, being potential sources for their spreading in the environment and to other species.

Feasibility of 16S rRNA sequencing for cerebrospinal fluid microbiome analysis in cattle with neurological disorders: a pilot study.

Bacterial infection of the central nervous system (CNS) in cattle requires prompt and adequate antimicrobial treatment. The current gold standard for antemortem etiological diagnosis is cerebrospinal fluid (CSF) culture, which often yields false negative results. CSF has long been considered a sterile district in healthy patients, but this notion has been recently challenged. For this pilot study, we used 16S rRNA gene sequencing to investigate the microbial composition of CSF of cattle presenting with CNS disorders and to compare it between subjects with CNS infections and with CNS disorders of other nature. The study sample was 10 animals: 4 presenting with CNS infectious-inflammatory diseases and 6 with other CNS disorders, based on definitive diagnosis. Since the initial round of a standard 16S rRNA PCR did not yield sufficient genetic material for sequencing in any of the samples, the protocol was modified to increase its sensitivity. Bacterial genetic material was identified in 6 animals and 2 groups were formed: an infectious inflammatory (n = 3) and a noninfectious inflammatory group (n = 3). The most frequently expressed bacterial families were Pseudomonadaceae (44.61%), Moraxellaceae (19.54%), Mycobacteriaceae (11.80%); the genera were Pseudomonas (45.42%), Acinetobacter (19.91%), Mycobacterium (12.01%). There were no detectable differences in the CSF microbial composition of the samples from the two groups. Sequencing of bacterial DNA present in the CSF was possible only after increasing PCR sensitivity. The results of 16S rRNA sequencing showed the presence of a microbial community in the CSF in cattle with neurological disorders. Further studies, in which CSF samples from healthy animals and samples from the environment are included as controls, are needed.

Epidural anesthesia in dogs undergoing hindlimb orthopedic surgery: effects of two injection sites.

This prospective clinical trial evaluated the effects of epidural anesthesia (EA) placed at the lumbosacral compared to the L5-L6 junction in dogs undergoing hindlimb orthopedic surgery. In all, 98 dogs were randomly assigned to receive injection at either L7-S1 (LS group) or L5-L6 (LL group) at the same local anesthetic regimen (1 mg/kg bupivacaine 0.5% and 0.1 mg/kg morphine 1%). Fentanyl (1 µg/kg) was the intraoperative rescue analgesia (iRA) administered if mean arterial pressure increased by 30% above pre-stimulation value. Procedural failure, iRA, hypotension, motor block resolution, and postoperative side effects were recorded. There were 7/47 (15%) epidural procedural failures in the LS group and 8/51 (16%) (P=1.00) in the LL group; iRA was administered in 21/40 (52%) LS group dogs and in 13/43 (30%) LL group dogs, respectively (P=0.047). The incidence of hypotension was 10/40 (25%) and 16/43 (37%) in the LS group and the LL group, respectively (P=0.25). Proprioceptive residual deficit at 8 hr after EA was recorded in 3/26 (12%) in group LS dogs and in 13/26 (50%) group LL dogs, respectively (P=0.01). The proprioceptive residual deficit at 24 hr in one dog (LL group) resolved within 36 hr. No episodes of postoperative urinary retention, pruritus or neurological damage were recorded. The L5-L6 EA decreased significantly iRA but delays the proprioceptive recovery time. Further studies are needed to determine whether a lower bupivacaine dose reduces the duration of the residual block retaining the same incidence of iRA.

Circulating skeletal muscle related microRNAs profile in Piedmontese cattle during different age.

Piedmontese cattle is known for double-muscle phenotype. MicroRNAs (miRNAs) play important role as regulators in skeletal muscle physiological processes, and we hypothesize that plasma miRNAs expression profiles could be affected by skeletal muscle growth status related to age. Plasma samples of cattle were collected during four different ages from first week of life until the time of commercial end of the fattening period before slaughter. Small-RNA sequencing data analysis revealed the presence of 40% of muscle-related miRNAs among the top 25 highly expressed miRNAs and, 19 miRNAs showed differential expression too. Using qRT-PCR, we validated in a larger bovine population, miRNAs involved in skeletal muscle physiology pathways. Comparing new-born with the other age groups, miR-10b, miR-126-5p, miR-143 and miR-146b were significantly up-regulated, whereas miR-21-5p, miR-221, miR-223 and miR-30b-5p were significantly down-regulated. High expression levels of miR-23a in all the groups were found. Myostatin, a negative regulator of skeletal muscle hypertrophy, was predicted as the target gene for miR-23a and miR-126-5p and we demonstrated their direct binding. Correlation analysis revealed association between miRNAs expression profiles and animals' weights along the age. Circulating miRNAs could be promising for future studies on their biomarker potentialities to beef cattle selection.

The RNA-Binding Protein ESRP1 Modulates the Expression of RAC1b in Colorectal Cancer Cells.

RNA binding proteins are well recognized as critical regulators of tumorigenic processes through their capacity to modulate RNA biogenesis, including alternative splicing, RNA stability and mRNA translation. The RNA binding protein Epithelial Splicing Regulatory Protein 1 (ESRP1) can act as a tumor suppressor or promoter in a cell type- and disease context-dependent manner. We have previously shown that elevated expression of ESRP1 in colorectal cancer cells can drive tumor progression. To gain further insights into the pro-tumorigenic mechanism of action of ESRP1, we performed cDNA microarray analysis on two colorectal cells lines modulated for ESRP1 expression. Intriguingly, RAC1b was highly expressed, both at mRNA and protein levels, in ESRP1-overexpressing cells, while the opposite trend was observed in ESRP1-silenced CRC cells. Moreover, RAC1 and RAC1b mRNA co-immunoprecipitate with ESRP1 protein. Silencing of RAC1b expression significantly reduced the number of soft agar colonies formed by ESRP1-overexpressing cells, suggesting that ESRP1 acted, at least partially, through RAC1b in its tumor-promoting activities in CRC cells. Thus, our data provide molecular cues on targetable candidates in CRC cases with high ESRP1 expression.

In vivo and in vitro Digestibility of an Extruded Complete Dog Food Containing Black Soldier Fly (Hermetia illucens) Larvae Meal as Protein Source.

Growing attention is being directed toward insects as a novel and sustainable source of protein for pet food. The aim of the study was to evaluate nutrient digestibility of a diet containing black soldier fly larvae as its main protein source. Moreover, the purpose of the study was to compare the traditional in vivo total collection method with the in vivo marker method and in vitro digestibility method. Two isonitrogenous and isoenergetic dry diets containing either venison meal (CTRL diet) or black soldier fly larvae meal (BSF diet) as their primary sources of proteins were fed to six adult dogs, according to a Latin square design. The digestibility of nutrients was determined using both in vivo ("total collection" and "internal marker" approaches) and in vitro methods. The two diets showed similar nutrient digestibility values for dry matter, organic matter, ether extract, ash, and phosphorus. However, a statistical trend (p = 0.066) was observed indicating greater protein digestibility in the BSF diet compared with the CTRL diet. Calcium digestibility was higher in the BSF diet compared with the CTRL diet (p = 0.018). On the contrary, fiber digestibility was lower in the insect-based diet compared with the venison diet (p < 0.001). There was no difference between total collection and internal marker methods in the assessment of in vivo digestibility for any of the nutrients considered. The in vitro digestibility values for dry matter, organic matter, and crude protein, as well as the estimated in vivo digestibility of organic matter and crude protein by the means of the predictive equation, were aligned with the in vivo results, although in vitro estimations were consistently higher compared with those obtained by in vivo analysis. Digestibility analysis of a dog food containing insect meal as the sole source of protein (36.5% inclusion) showed promising results in terms of it presenting similar values as a meat-based diet, indicating its suitability as a sustainable protein source for pet food. Moreover, the study showed that both the in vivo marker method and the in vitro method could be possible alternatives to the traditional total collection method in digestibility trials.

Pseudogenes as Competitive Endogenous RNAs: Target Prediction and Validation.

Pseudogenes may regulate expression of their parental genes as well as other protein-coding genes through various mechanisms. One such mechanism is the ability to act as competitive endogenous RNA (ceRNA) and participate in microRNA-mediated cross-regulation. Here, we outline how to predict the targets of pseudogene ceRNAs bioinformatically and how to validate them experimentally.

Evaluation of prognostic impact of pre-treatment neutrophil to lymphocyte and lymphocyte to monocyte ratios in dogs with oral malignant melanoma treated with surgery and adjuvant CSPG4-antigen electrovaccination: an explorative study.

The role of systemic inflammation in cancer's progression has been widely investigated, especially in melanoma in humans. Pre-treatment leukocyte counts and ratios play a recognized prognostic role in several types of malignancies, but no information is available regarding canine oral malignant melanoma (COMM). The purpose of this explorative retrospective study was to investigate the prognostic impact of pre-treatment neutrophil to lymphocyte (NLR) and lymphocyte to monocyte (LMR) ratios in dogs with oral malignant melanoma that underwent surgical resection and immunotherapy with adjuvant CSPG4-antigen electrovaccination. Thirty-nine dogs with histologically confirmed oral melanoma and with available pre-treatment haematological analyses, performed at maximum 60 days before the first treatment, were retrospectively enrolled. Statistical analysis was performed to explore possible correlations among NLR and LMR with age, clinical stage, tumour pigmentation, tumour size, nuclear atypia, mitotic index, Ki67, CSPG4 expression, ulceration, bone invasion and excision margins status. The impact of NLR and LMR on overall survival time (OST) was explored among various ratio cut off and across different time points with Kaplan-Meier method. No significant relationship was identified between leukocytes ratios and histological parameters, CSPG4 expression, excision margin status, age, tumour size and clinical stage. NLR and LMR did not display a prognostic impact on the survival time of the entire population. Pre-treatment leukocyte ratios may not represent a useful prognostic factor in dogs with oral melanoma, especially in absence of distant metastatic disease.

Competing endogenous RNAs and cancer: How coding and non-coding molecules cross-talk can impinge on disease.

Cancers are characterized by several dramatic biological changes. Among the many post-transcriptional regulatory mechanisms, microRNAs are known as fine-tune regulators for their transcript silencing ability. Competing endogenous RNAs (ceRNAs) are transcripts that share microRNA binding elements and can compete for them, thus regulating each other indirectly. ceRNA networks interconnect the regulatory control of different transcript classes of the coding and non-coding space and co-operate with other cellular and molecular regulatory mechanisms. Altered ceRNA networks are involved in tumor formation and progression as well as in chemoresistance, in invasion and in the onset of metastases. The analysis of changes in the balance between ceRNA transcripts could offer hints to identify novel pathways for diagnosis, prognosis and therapies in precision medicine interventions. Moreover, the possibility to query highly specific tumor databases, such as TCGA, and to combine clinical data, transcript expression and sequence information is allowing to develop specific predictive tools for precision medicine.

The p140Cap adaptor protein as a molecular hub to block cancer aggressiveness.

The p140Cap adaptor protein is a scaffold molecule encoded by the SRCIN1 gene, which is physiologically expressed in several epithelial tissues and in the neurons. However, p140Cap is also strongly expressed in a significant subset of cancers including breast cancer and neuroblastoma. Notably, cancer patients with high p140Cap expression in their primary tumors have a lower probability of developing a distant event and ERBB2-positive breast cancer sufferers show better survival. In neuroblastoma patients, SRCIN1 mRNA levels represent an independent risk factor, which is inversely correlated to disease aggressiveness. Consistent with clinical data, SRCIN1 gain or loss of function mouse models demonstrated that p140Cap may affect tumor growth and metastasis formation by controlling the signaling pathways involved in tumorigenesis and metastatic features. This study reviews data showing the relevance of SRCIN1/p140Cap in cancer patients, the impact of SRCIN1 status on p140Cap expression, the specific mechanisms through which p140Cap can limit cancer progression, the molecular functions regulated by p140Cap, along with the p140Cap interactome, to unveil its key role for patient stratification in clinics.

Analysis of Early Assessable Risk Factors for Poor Outcome in Dogs With Cluster Seizures and Status Epilepticus.

Status epileptics (SE) and cluster seizures (CS) are serious neurological emergencies associated with poor outcome in epileptic patients. Data on risk factors associated with outcome in epileptic patients affected by CS and SE have not been studied extensively to date. In the present retrospective study potential risk factors predictive of negative outcome in a population of dogs affected by CS and SE were analyzed. Ninety-three dogs were included in the study: 21/93 patients (23%) presented with SE and 72/93 (77%) with CS. Based on multivariate statistical analysis, factors statistically associated with a poor outcome were the occurrence of the first epileptic seizure outside the defined idiopathic interval (6 months-6 years), a condition of hyperthermia at presentation and the absence of previous antiepileptic drugs (AEDs) in case of previous history of seizures. The results of the present study implement data on risk factors associated with poor outcome in dogs affected by CS or SE and can aid in the creation of an ad hoc scoring system, similar to systems currently applied in human medicine upon hospital admission to benchmark performances and establish protocols for triage and therapeutic management.

Risk factors for type 1 diabetes, including environmental, behavioural and gut microbial factors: a case-control study.

Type 1 diabetes (T1D) is a common autoimmune disease that is characterized by insufficient insulin production. The onset of T1D is the result of gene-environment interactions. Sociodemographic and behavioural factors may contribute to T1D, and the gut microbiota is proposed to be a driving factor of T1D. An integrated preventive strategy for T1D is not available at present. This case-control study attempted to estimate the exposure linked to T1D to identify significant risk factors for healthy children. Forty children with T1D and 56 healthy controls were included in this study. Anthropometric, socio-economic, nutritional, behavioural, and clinical data were collected. Faecal bacteria were investigated by molecular methods. The findings showed, in multivariable model, that the risk factors for T1D include higher Firmicutes levels (OR 7.30; IC 2.26-23.54) and higher carbohydrate intake (OR 1.03; IC 1.01-1.05), whereas having a greater amount of Bifidobacterium in the gut (OR 0.13; IC 0.05 - 0.34) was a protective factor for T1D. These findings may facilitate the development of preventive strategies for T1D, such as performing genetic screening, characterizing the gut microbiota, and managing nutritional and social factors.