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Trial 263 – ONGOING: Kluyveromyces marxianus fragilis B0399 for the health of the dog's gastrointestinal tract and beyond.

Background:

Gastrointestinal diseases in dogs are caused by infectious agents, intestinal parasites, diet, genetic predisposition, and environmental stress factors. In the absence of infectious agents and parasites, dysbiosis is observed in dogs (Scarsella et al., 2023), accounting for more than 50% of gastrointestinal disorders reported in this species. In these cases, the therapeutic strategy involves the administration of probiotics and prebiotics with the aim of alleviating symptoms and restoring the biodiversity of the intestinal microbial community.

Preliminary studies on certain probiotic yeasts from the genus Saccharomyces (Meineri et al., 2022) have shown that the use of probiotics in healthy dogs can positively affect the fecal microbiome, as well as the normal functioning of the digestive system and the overall well-being of the animal, directing it towards a balanced composition indicative of intestinal health, based on a large canine intestinal microbiome database (Scarsella et al., 2021). Another preliminary study on a small population of dogs (7 subjects) showed that Kluyveromyces marxianus fragilis B0399 supports intestinal health in terms of the dogs "fecal score" (link), similar to the effect repeatedly demonstrated in various tests and efficacy studies in other omnivorous monogastrics (pigs and humans) (Maccaferi et al., 2012; EFSA Journal, 2004), though its effect on the biodiversity and composition of the fecal microbiome has not yet been explored.

The ability to verify the composition of the fecal microbiome in dogs in relation to a dysbiosis condition represents an essential tool for characterizing the mode of action of the probiotic, proceeding with targeted administration, and evaluating its effect on the disorders progression and the stability of the microbiome over time.

To this end, the project plans to recruit dogs for veterinary studies, belonging to owners or caregivers, who are affected by non-infectious gastroenteritis or parasites, and administer the probiotic for at least 15/30 days or until the intestinal symptoms disappear (fecal consistency (dry matter); constipation and diarrhea; vomiting or regurgitation; increased and/or worsened flatulence/unpleasant odor; excessive drooling; reduced appetite, activity, and weight loss). The demographic data and characteristics of the subjects will be recorded at the time of enrollment, and clinical data and a fecal sample for DNA extraction and microbiome analysis will be collected at the beginning and end of the study period. Likewise, fecal samples will be collected from healthy dogs before and after the administration of the probiotic.

The microbiomes will then be compared with the available values in a database to verify the composition of microbial populations typical of intestinal health. The study results will allow the validation of the probiotics use in a relevant environment and will form the basis for further cohort studies on larger canine populations, stratified by age, breed, and types of diet.

Project Objectives:

Demonstration of the efficacy and determination of the "therapeutic" dosage of Kluyveromyces marxianus fragilis B0399 for supporting the treatment of gastrointestinal disorders and underlying intestinal dysbiosis in dogs.

Methodology:

• Enrollment of study subjects will be carried out in direct collaboration with veterinarians, canine nutritionists and caregivers; 1st Clinical visits & sample collection are performed (feces - collection and preservation) before starting the probiotic supplementation – time point “T0” of the study timeline.
• Study groups: Based on the anamnesis performed subjects will be assigned to either the "test" group, showing symptoms related to gastrointestinal disorders, or the group of healthy subjects serving as the reference, “control” group; Inclusion criterions for the test group: dogs exhibiting symptoms related to gastrointestinal disorders such as vomiting, diarrhea, flatulence, constipation, lack of appetite, dehydration, gastric rumbling, and also: fever, depression, and sudden inactivity. Absence of any gastrointestinal problems in the last month before the trial start is inclusion criterion for the “control” group
• Treatment: both groups will receive the active ingredient for 30 days
• Study parameters and quantification method:
  • Questionnaire: Owners of the dogs will be asked to keep a daily diary and fill up a specific questionnaire for the entire duration of administration, recording the symptoms related to the disorder that were noted at the start of the study. Statistical analysis of the frequency of symptoms present in the "test" group during the whole study.
  • Fecal samples will be collected on the beginning (“T0”), on the half (“T15”) and at the end (“T30”) of the trial period for the DNA extraction and total bacterial and yeast DNA sequencing. The sequencing results will be followed by bioinformatic analysis and microbial profiles of the fecal samples will be analyzed using the mathematical and statistical analysis developed and standardized by Scarsella et al., 2020.

This project is eligible for funding under the PR ERDF 2021-2027, Activity a1.3.1 – Non-refundable incentive tool for the creation of Proof of Concept (PoC), approved with DGR n. 1974/2023 (click to see the funding details)