Effects of a Veterinary Gastrointestinal Low-Fat Diet on Fecal Characteristics, Metabolites, and Microbiota Concentrations of Adult Dogs Treated with Metronidazole.

Antibiotics are known to cause loose stools, disrupt the fecal microbiota, and alter fecal bile acid (BA) profiles of dogs. Recovery may be aided by diet, but little research has been conducted. The objective of this study was to determine how a veterinary low-fat diet affected the fecal characteristics, metabolites, BA, and microbiota of dogs receiving antibiotics. Twenty-four healthy adult dogs [7.38 ± 1.95 yr; 7.67 ± 0.76 kg body weight (BW)] were used in an 8-wk completely randomized design study. During a 2-wk baseline, all dogs were fed a leading grocery brand dry kibble diet (GBD). Over the next 2 wk, dogs were fed GBD and received metronidazole orally (20 mg/kg BW twice daily). At wk 4, dogs were randomly allotted to one of two treatments [GBD or Blue Buffalo Natural Veterinary Diet GI Gastrointestinal Support Low-Fat (BB)] and fed for 4 wk. Fecal scores were recorded daily and fresh fecal samples were collected at wk 2, 4, 5, 6, 7, and 8 for measurement of pH, dry matter content, and metabolite and BA concentrations. Fecal microbiota populations were analyzed by 16S rRNA gene amplicon sequencing and qPCR-based dysbiosis index (DI). All data were analyzed as repeated measures using the Mixed Models procedure of SAS 9.4, testing for effects of treatment, time, and treatment*time and significance set at P<0.05. Metronidazole increased (P<0.0001) fecal scores (looser stools), reduced fecal short-chain fatty acid, branched-chain fatty acid, phenol, and indole concentrations, increased primary BA concentrations, and decreased secondary BA concentrations. Metronidazole also reduced fecal bacterial alpha diversity, altered the abundance of 58 bacterial genera, and increased DI. During antibiotic recovery, change in fecal pH, dry matter percentage, and metabolite and immunoglobulin A concentrations were altered (P<0.05) by diet. Fecal BA concentrations recovered quickly for all dogs. Change in lithocholic acid was affected (P<0.0001) by diet, but other BA were not. Recovery of over 25 bacterial genera was impacted by diet (P<0.05). While many bacterial taxa returned to baseline levels after 4 wk, others did not fully recover. DI and bacterial alpha diversity measures recovered quickly for all dogs, but were not impacted by diet. In conclusion, metronidazole drastically altered the fecal microbiota and metabolites of dogs. While most variables returned to baseline by wk 8, diet may be used to aid in recovery.

Effects of a veterinary gastrointestinal diet on fecal characteristics, metabolites, and microbiota concentrations of adult cats treated with metronidazole.

Antibiotics are used to treat gastrointestinal diseases or infections but are known to negatively affect stool quality and gut microbiota in cats and dogs. Therefore, identifying dietary strategies that may aid in antibiotic recovery is of interest. The objective of this study was to determine how a veterinary gastrointestinal diet affected the fecal characteristics, microbiota, and metabolite and bile acid (BA) concentrations of cats recovering from metronidazole administration. Twenty-four healthy adult cats were used in an 8-wk completely randomized design study. During a 2-wk baseline, all cats consumed a leading grocery brand diet (GBD). Over the next 2 wk, cats consumed GBD and received metronidazole (20 mg/kg body weight twice daily). At week 4, cats were randomly allotted to one of 2 treatments [GBD; BLUE Natural Veterinary Diet GI Gastrointestinal Support (BB)] and fed for 4 wk. Fecal scores were recorded daily and fresh fecal samples were collected at weeks 2, 4, 5, 6, 7, and 8 for measurement of pH, dry matter (DM) %, metabolites, and microbiota. Microbiota was analyzed by 16S rRNA gene sequencing and qPCR, which was used to calculate dysbiosis index. Data were analyzed as repeated measures using the Mixed Models procedure of SAS 9.4, testing for effects of diet, time and diet*time. Metronidazole had dramatic effects on all outcomes, including increased fecal scores (looser stools), reduced fecal pH and DM%, reduced fecal short-chain fatty acid, branched-chain fatty acid, ammonia, phenol, and indole concentrations, and altered fecal BA concentrations (increased primary BA; reduced secondary BA). Metronidazole reduced fecal bacterial alpha diversity, increased dysbiosis index, and altered the relative abundance of 78 bacterial genera. Fecal outcomes partially recovered over the next 4 wk, with some being impacted by diet. Fecal acetate concentrations were higher after metronidazole in cats fed BB. Dysbiosis index and alpha diversity measures slowly recovered over 4 wk, without diet differences. Recovery of 16 bacterial genera was impacted by diet. Fecal BA profiles demonstrated a prolonged impairment of primary to secondary BA conversion, with cholic acid being lower after metronidazole in cats fed BB. In conclusion, our data demonstrate that metronidazole is a powerful antibiotic that has long-lasting effects on the fecal microbiota and metabolites of cats. Outcome variables slowly recovered over time, but a gastrointestinal diet may aid in recovery.

The objective of this study was to determine how a veterinary gastrointestinal diet impacts the fecal characteristics, metabolites, and microbiota concentrations of adult cats treated with metronidazole. All cats were fed a leading grocery brand diet (GBD) during a 2-wk baseline, dosed orally with metronidazole (20 mg/kg BW twice daily) for 2 wk, then randomly allotted to one of 2 treatments [GBD; BLUE Natural Veterinary Diet GI Gastrointestinal Support (BB)] and fed for 4 wk. Fecal scores were recorded daily and fresh fecal samples were collected at weeks 2, 4, 5, 6, 7, and 8 to assess fecal characteristics, microbiota populations, and metabolite and bile acid (BA) concentrations. Metronidazole increased fecal scores (looser stools), altered relative abundances of 78 bacterial genera and BA profiles (increased primary BA; reduced secondary BA), and increased dysbiosis index. Fecal outcomes varied during recovery. Recovery of 16 bacterial genera was impacted by diet. Dysbiosis index and bacterial alpha diversity were not affected by diet. Our data demonstrate that metronidazole is a powerful antibiotic and may have long-lasting effects on the gut microbiota and metabolites of cats. Feeding a gastrointestinal diet may aid in the recovery of several markers of microbial function (e.g., metabolites).

Effects of a milk oligosaccharide biosimilar on fecal characteristics, microbiota, and bile acid, calprotectin, and immunoglobulin concentrations of healthy adult dogs treated with metronidazole.

In recent dog and cat experiments, a novel milk oligosaccharide biosimilar (GNU100) positively modulated fecal microbiota and metabolite profiles, suggesting benefits to gastrointestinal health. The objective of this study was to investigate the effects of GNU100 on the fecal characteristics, microbiota, and bile acid (BA) concentrations of healthy adult dogs treated with antibiotics. Twelve healthy adult female dogs (mean age: 3.74 ± 2.4 yr) were used in an 8-wk crossover design study (dogs underwent both treatments). All dogs were fed a control diet during a 2-wk baseline, then randomly allotted to 1 of 2 treatments (diet only or diet + 1% GNU100) for another 6 wk. From weeks 2 to 4, dogs were orally administered metronidazole (20 mg/kg BW) twice daily. Fecal scores were recorded daily and fresh fecal samples were collected at weeks 2, 4, 5, 6, and 8 for measurement of pH, dry matter, microbiota populations, and BA, immunoglobulin A, and calprotectin concentrations. On weeks 0, 4, and 8, blood samples were collected for serum chemistry and hematology analysis. All data were analyzed as repeated measures using the Mixed Models procedure of SAS version 9.4, with significance considered P < 0.05. Metronidazole increased (P < 0.0001) fecal scores (looser stools) and modified (P < 0.05) fecal microbiota and BA profiles. Using qPCR, metronidazole reduced fecal Blautia, Fusobacterium, Turicibacter, Clostridium hiranonis, and Faecalibacterium abundances, and increased fecal Streptococcus and Escherichia coli abundances. DNA sequencing analysis demonstrated that metronidazole reduced microbial alpha diversity and influenced the relative abundance of 20 bacterial genera and families. Metronidazole also increased primary BA and reduced secondary BA concentrations. Most antibiotic-induced changes returned to baseline by week 8. Fecal scores were more stable (P = 0.01) in GNU100-fed dogs than controls after antibiotic administration. GNU100 also influenced fecal microbiota and BA profiles, reducing (P < 0.05) the influence of metronidazole on microbial alpha diversity and returning some fecal microbiota and secondary BA to baseline levels at a quicker (P < 0.05) rate than controls. In conclusion, our results suggest that GNU100 supplementation provides benefits to dogs treated with antibiotics, providing more stable fecal scores, maintaining microbial diversity, and allowing for quicker recovery of microbiota and secondary BA profiles which play an essential role in gut health.

Our objective was to test the effects of a novel milk oligosaccharide biosimilar (GNU100) on the fecal characteristics, microbiota, and bile acid (BA) concentrations of healthy adult dogs treated with antibiotics. Dogs were fed a control diet during a 2-wk baseline, then randomly allotted to 1 of 2 treatments (diet only or diet + 1% GNU100) for another 6 wk. From weeks 2 to 4, dogs were given an oral antibiotic. Fecal scores were recorded and fresh fecal samples were collected over time to assess fecal characteristics, microbiota populations, and BA concentrations. The antibiotic was shown to increase fecal scores (looser stools) and modify fecal microbiota populations (altered diversity and ~20 bacterial genera and families) and BA profiles (increased primary and reduced secondary BA). Most antibiotic-induced changes returned to baseline by week 8. In dogs fed GNU100, fecal scores were more stable and changes to microbial diversity were lower than controls after antibiotic administration. Fecal microbiota and secondary BA of GNU100-fed dogs also returned to baseline levels at a quicker rate than controls. These results suggest that GNU100 provides benefits to dogs given antibiotics, providing more stable fecal scores, maintaining microbial diversity, and allowing for quicker recovery of microbiota and BA profiles.

Correlation between Targeted qPCR Assays and Untargeted DNA Shotgun Metagenomic Sequencing for Assessing the Fecal Microbiota in Dogs.

DNA shotgun sequencing is an untargeted approach for identifying changes in relative abundances, while qPCR allows reproducible quantification of specific bacteria. The canine dysbiosis index (DI) assesses the canine fecal microbiota by using a mathematical algorithm based on qPCR results. We evaluated the correlation between qPCR and shotgun sequencing using fecal samples from 296 dogs with different clinical phenotypes. While significant correlations were found between qPCR and sequencing, certain taxa were only detectable by qPCR and not by sequencing. Based on sequencing, less than 2% of bacterial species (17/1190) were consistently present in all healthy dogs (n = 76). Dogs with an abnormal DI had lower alpha-diversity compared to dogs with normal DI. Increases in the DI correctly predicted the gradual shifts in microbiota observed by sequencing: minor changes (R = 0.19, DI < 0 with any targeted taxa outside the reference interval, RI), mild-moderate changes (R = 0.24, 0 < DI < 2), and significant dysbiosis (R = 0.54, 0.73, and 0.91 for DI > 2, DI > 5, and DI > 8, respectively), compared to dogs with a normal DI (DI < 0, all targets within the RI), as higher R-values indicated larger dissimilarities. In conclusion, the qPCR-based DI is an effective indicator of overall microbiota shifts observed by shotgun sequencing in dogs.

Weight loss and high-protein, high-fiber diet consumption impact blood metabolite profiles, body composition, voluntary physical activity, fecal microbiota, and fecal metabolites of adult dogs.

Canine obesity is associated with reduced lifespan and metabolic dysfunction, but can be managed by dietary intervention. This study aimed to determine the effects of restricted feeding of a high-protein, high-fiber (HPHF) diet and weight loss on body composition, physical activity, blood metabolites, and fecal microbiota and metabolites of overweight dogs. Twelve spayed female dogs (age: 5.5 ± 1.1 yr; body weight [BW]: 14.8 ± 2.0 kg, body condition score [BCS]: 7.9 ± 0.8) were fed a HPHF diet during a 4-wk baseline phase to maintain BW. After baseline (week 0), dogs were first fed 80% of baseline intake and then adjusted to target 1.5% weekly weight loss for 24 wk. Body composition using dual-energy x-ray absorptiometry and blood samples (weeks 0, 6, 12, 18, and 24), voluntary physical activity (weeks 0, 7, 15, and 23), and fresh fecal samples for microbiota and metabolite analysis (weeks 0, 4, 8, 12, 16, 20, and 24) were measured over time. Microbiota data were analyzed using QIIME 2. All data were analyzed statistically over time using SAS 9.4. After 24 wk, dogs lost 31.2% of initial BW and had 1.43 ± 0.73% weight loss per week. BCS decreased (P < 0.0001) by 2.7 units, fat mass decreased (P < 0.0001) by 3.1 kg, and fat percentage decreased (P < 0.0001) by 11.7% with weight loss. Many serum metabolites and hormones were altered, with triglycerides, leptin, insulin, C-reactive protein, and interleukin-6 decreasing (P < 0.05) with weight loss. Relative abundances of fecal Bifidobacterium, Coriobacteriaceae UCG-002, undefined Muribaculaceae, Allobaculum, Eubacterium, Lachnospira, Negativivibacillus, Ruminococcus gauvreauii group, uncultured Erysipelotrichaceae, and Parasutterella increased (P < 0.05), whereas Prevotellaceae Ga6A1 group, Catenibacterium, Erysipelatoclostridium, Fusobacterium, Holdemanella, Lachnoclostridium, Lactobacillus, Megamonas, Peptoclostridium, Ruminococcus gnavus group, and Streptococcus decreased (P < 0.01) with weight loss. Despite the number of significant changes, a state of dysbiosis was not observed in overweight dogs. Fecal ammonia and secondary bile acids decreased, whereas fecal valerate increased with weight loss. Several correlations between gut microbial taxa and biological parameters were observed. Our results suggest that restricted feeding of a HPHF diet and weight loss promotes fat mass loss, minimizes lean mass loss, reduces inflammatory marker and triglyceride concentrations, and modulates fecal microbiota phylogeny and activity in overweight dogs.

Canine obesity is associated with reduced lifespan and metabolic dysfunction, but dietary intervention may aid in its management. This study aimed to determine the effects of restricted feeding of a high-protein, high-fiber (HPHF) diet and weight loss on body composition, physical activity, blood metabolites, and fecal bacteria and metabolites of overweight dogs. Twelve overweight dogs were fed a HPHF diet during a 4-wk baseline to maintain body weight and then fed to lose weight for 24 wk. Body composition, blood samples, voluntary physical activity, and fresh fecal samples were measured over time. After 24 wk, dogs lost over 30% of their initial body weight and had 1.4% weight loss per week. As expected, serum triglycerides, leptin, insulin, C-reactive protein, and interleukin-6 decreased with weight loss. The relative abundances of 4 bacterial phyla and over 30 bacterial genera were altered with weight loss. Fecal ammonia and secondary bile acid concentrations decreased, whereas fecal valerate concentrations increased with weight loss. Several correlations between fecal bacteria and physiological parameters were identified. Our results suggest that a HPHF diet and weight loss promote fat mass loss, reduce inflammatory marker and triglyceride concentrations, and modulate fecal bacterial populations and activity in overweight dogs.

Dietary supplementation with fiber, "biotics," and spray-dried plasma affects apparent total tract macronutrient digestibility and the fecal characteristics, fecal microbiota, and immune function of adult dogs.

A variety of functional ingredients, including fibers, prebiotics, probiotics, and postbiotics may be added to pet foods to support gastrointestinal and immune health. While many of these ingredients have been tested individually, commercial foods often include blends that also require testing. This study was conducted to evaluate the effects of diets containing blends of fibers, "biotics," and/or spray-dried plasma on apparent total tract digestibility (ATTD), stool quality, fecal microbiota and metabolites, and immune health outcomes of adult dogs. A total of 12 healthy adult intact English pointer dogs (6 M, 6 F; age = 6.4 ± 2.0 yr; BW = 25.8 ± 2.6 kg) were used in a replicated 3 × 3 Latin square design to test diets formulated to: 1) contain a low concentration of fermentative substances (control diet, CT); 2) be enriched with a fiber-prebiotic-probiotic blend (FPPB); and 3) be enriched with a fiber-prebiotic-probiotic blend + immune-modulating ingredients (iFFPB). In each 28-d period, 22 d of diet adaptation was followed by a 5-d fecal collection phase and 1 d for blood sample collection. All data were analyzed using SAS 9.4, with significance being P < 0.05 and trends being P < 0.10. FPPB and iFPPB diets led to shifts in numerous outcome measures. Dry matter (DM), organic matter, fat, fiber, and energy ATTD were lower (P < 0.01), fecal scores were lower (P < 0.01; firmer stools), and fecal DM% was higher (P < 0.0001) in dogs fed FPPB or iFPPB than those fed CT. Serum triglycerides and cholesterol were lower (P < 0.01) in dogs fed FPPB or iFPPB than those fed CT. Fecal protein catabolites (isobutyrate, isovalerate, indole, and ammonia) and butyrate were lower (P < 0.05), while fecal immunoglobulin A (IgA) was higher (P < 0.01) in dogs fed FPPB and iFPPB than those fed CT. Fecal microbiota populations were affected by diet, with alpha-diversity being lower (P < 0.05) in dogs fed iFPPB and the relative abundance of 20 bacterial genera being altered in dogs fed FPPB or iFPPB compared with CT. The circulating helper T cell:cytotoxic T cell ratio was higher (P < 0.05) in dogs fed iFPPB than those fed CT. Circulating B cells were lower (P < 0.05) in dogs fed FPPB than those fed iFPPB, and lower (P < 0.05) in dogs fed iFPPB than those fed CT. Our results demonstrate that feeding a fiber-prebiotic-probiotic blend may provide many benefits to canine health, including improved stool quality, beneficial shifts to fecal microbiota and metabolite profiles, reduced blood lipids, and increased fecal IgA.

A variety of functional ingredients­those that provide benefits beyond their nutritional value­may be added to pet foods to support gastrointestinal and immune health. While many of these ingredients have been tested individually, commercial foods often include blends that also require testing. This study was conducted to evaluate the effects of diets containing blends of dietary fibers and other functional ingredients on nutrient digestibility and the stool characteristics and immune health outcomes of adult dogs consuming them. Treatments included a control diet containing low amounts of dietary fiber, a diet containing a fiber­prebiotic­probiotic blend, and a diet containing the fiber­prebiotic­probiotic blend as well as immune-modulating ingredients. The test diets were shown to shift many outcome measures. First, they were shown to reduce nutrient digestibility and decrease fecal scores (more firm stool). Second, test diets reduced blood lipids and beneficially altered fecal metabolite concentrations. Third, test diets increased fecal immunoglobulin A concentrations, suggesting enhanced gut immunity. Lastly, the test diets shifted fecal bacterial populations. Our results demonstrate that feeding a fiber­prebiotic­probiotic blend may provide many benefits to canine health, including improved stool quality, beneficial shifts to fecal bacteria and metabolite profiles, reduced blood lipids, and enhanced gut immunity.