Safety profile and effects on the peripheral immune response of fecal microbiota transplantation in clinically healthy dogs.

BACKGROUND: Fecal microbiota transplantation (FMT) is increasingly used for gastrointestinal and extra-gastrointestinal diseases in veterinary medicine. However, its effects on immune responses and possible adverse events have not been systematically investigated. HYPOTHESIS/OBJECTIVES: Determine the short-term safety profile and changes in the peripheral immune system after a single FMT administration in healthy dogs. ANIMALS: Ten client-owned, clinically healthy dogs as FMT recipients, and 2 client-owned clinically healthy dogs as FMT donors. METHODS: Prospective non-randomized clinical trial. A single rectal enema of 5 g/kg was given to clinically healthy canine recipients. During the 28 days after FMT administration, owners self-reported adverse events and fecal scores. On Days 0 (baseline), 1, 4, 10, and 28 after FMT, fecal and blood samples were collected. The canine fecal dysbiosis index (DI) was calculated using qPCR. RESULTS: No significant changes were found in the following variables: CBC, serum biochemistry, C-reactive protein, serum cytokines (interleukins [IL]-2, -6, -8, tumor necrosis factor [TNF]-α), peripheral leukocytes (B cells, T cells, cluster of differentiation [CD]4+ T cells, CD8+ T cells, T regulatory cells), and the canine DI. Mild vomiting (n = 3), diarrhea (n = 4), decreased activity (n = 2), and inappetence (n = 1) were reported, and resolved without intervention. CONCLUSIONS AND CLINICAL IMPORTANCE: Fecal microbiota transplantation did not significantly alter the evaluated variables and recipients experienced minimal adverse events associated with FMT administration. Fecal microbiota transplantation was not associated with serious adverse events, changes in peripheral immunologic variables, or the canine DI in the short-term.

Unbiased serum metabolomic analysis in cats with naturally occurring chronic enteropathies before and after medical intervention.

Chronic enteropathies (CE) are common disorders in cats and the differentiation between the two main underlying diseases, inflammatory bowel disease (IBD) and low-grade intestinal T-cell lymphoma (LGITL), can be challenging. Characterization of the serum metabolome could provide further information on alterations of disease-associated metabolic pathways and may identify diagnostic or therapeutic targets. Unbiased metabolomics analysis of serum from 28 cats with CE (14 cats with IBD, 14 cats with LGITL) and 14 healthy controls identified 1,007 named metabolites, of which 129 were significantly different in cats with CE compared to healthy controls at baseline. Random Forest analysis revealed a predictive accuracy of 90% for differentiating controls from cats with chronic enteropathy. Metabolic pathways found to be significantly altered included phospholipids, amino acids, thiamine, and tryptophan metabolism. Several metabolites were found to be significantly different between cats with IBD versus LGITL, including several sphingolipids, phosphatidylcholine 40:7, uridine, pinitol, 3,4-dihydroxybenzoic acid, and glucuronic acid. However, random forest analysis revealed a poor group predictive accuracy of 60% for the differentiation of IBD from LGITL. Of 129 compounds found to be significantly different between healthy cats and cats with CE at baseline, 58 remained different following treatment.

A Comparison of the Oral Microbiota in Healthy Dogs and Dogs with Oral Tumors.

The aim of this study was to further describe the oral microbiota of healthy dogs by DNA shotgun sequencing and compare those to dogs with oral tumors. Oral swabs (representative of all niches of the oral cavity) were collected from healthy dogs (n = 24) and from dogs with different oral tumors (n = 7). DNA was extracted from the swabs and shotgun metagenomic sequencing was performed. Only minor differences in microbiota composition were observed between the two groups. At the phylum level, the Bacteroidota, Proteobacteria, Actinobacteriota, Desulfobacterota and Firmicutes were most abundant in both groups. Observed Operational Taxonomic Units-OTUs (species richness) was significantly higher in the healthy patients, but there was no significant difference in the Shannon diversity index between the groups. No significant difference was found in beta diversity between the groups. The core oral microbiota consisted of 67 bacterial species that were identified in all 24 healthy dogs. Our study provides further insight into the composition of the oral microbiota of healthy dogs and in dogs with oral tumors.

Fecal Concentrations of Long-Chain Fatty Acids, Sterols, and Unconjugated Bile Acids in Cats with Chronic Enteropathy.

Chronic enteropathy (CE) in cats encompasses food-responsive enteropathy, chronic inflammatory enteropathy (or inflammatory bowel disease), and low-grade intestinal T-cell lymphoma. While alterations in the gut metabolome have been extensively studied in humans and dogs with gastrointestinal disorders, little is known about the specific metabolic profile of cats with CE. As lipids take part in energy storage, inflammation, and cellular structure, investigating the lipid profile in cats with CE is crucial. This study aimed to measure fecal concentrations of various fatty acids, sterols, and bile acids. Fecal samples from 56 cats with CE and 77 healthy control cats were analyzed using gas chromatography-mass spectrometry, targeting 12 fatty acids, 10 sterols, and 5 unconjugated bile acids. Fecal concentrations of nine targeted fatty acids and animal-derived sterols were significantly increased in cats with CE. However, fecal concentrations of plant-derived sterols were significantly decreased in cats with CE. Additionally, an increased percentage of primary bile acids was observed in a subset of cats with CE. These findings suggest the presence of lipid maldigestion, malabsorption, and inflammation in the gastrointestinal tract of cats with CE. Understanding the lipid alterations in cats with CE can provide insights into the disease mechanisms and potential future therapeutic strategies.

Analytical validation, sample stability, and clinical evaluation of a new high-sensitivity cardiac troponin I immunoassay for use in dogs, with comparison to a previous ultrasensitive assay.

Cardiac troponin I (cTnI) is considered the gold standard biomarker for myocardial injury and shows a high degree of homology between humans and dogs. The ADVIA Centaur XP High-Sensitivity Troponin I (AC-cTnI-HS) assay has been validated for use in humans but not dogs. The study objectives were to analytically validate the AC-cTnI-HS assay in dogs, to assess correlation between the AC-cTnI-HS and a previous ADVIA Centaur TnI-Ultra (AC-cTnI-U) assay, to assess cTnI sample storage stability, and to clinically evaluate the AC-cTnI-HS assay in healthy dogs and dogs with cardiac disease. Canine serum samples were used for analytical validation. Intra- and inter-assay variability, dilutional parallelism, and spiking recovery were assessed. Samples from 196 client-owned dogs were evaluated (healthy dogs (n = 39) or dogs with congenital heart disease (n = 54), myxomatous mitral valve disease (n = 68), dilated cardiomyopathy (n = 15), or myocarditis (n = 20)). Inter- and intra-assay coefficient of variation (%CV) was between 2.8-41.4% and 3.8-30.2%, respectively, with pools with concentrations >20 pg/mL all having %CVs <10%. The observed to expected ratios for dilutional parallelism and spiking recovery experiments ranged between 92.3 and 266.7.0% and 84.3 and 108%, respectively. A strong correlation between the AC-cTnI-HS and AC-cTnI-U assays was observed (Spearman's ρ = 0.927), though a proportional bias existed, with AC-cTnI-HS assay concentrations being proportionally lower than AC-cTnI-U assay concentrations. Serum samples stored at -80°C had stable cTnI measurements for up to 2.7 years and after a single freeze-thaw cycle. Healthy dogs and dogs with congenital heart disease had significantly lower cTnI concentrations than dogs in the other three groups. The AC-cTnI-HS assay precisely, reproducibly, and accurately measures cTnI concentrations in dog serum with cTnI concentrations >20 pg/mL.

Fecal S100A12 concentrations in cats with chronic enteropathies.

OBJECTIVES: The aim of this study was to compare fecal S100A12 concentrations in cats diagnosed with chronic enteropathy (CE) with healthy control cats. METHODS: This was a prospective, cross-sectional study. Forty-nine cats that had gastrointestinal signs for >3 weeks and a complete diagnostic work-up, including bloodwork, abdominal ultrasound and upper and/or lower gastrointestinal endoscopic biopsies, were enrolled into the CE group. Nineteen cats from the CE group were diagnosed with inflammatory bowel disease (IBD) or chronic inflammatory enteropathy (CIE) and 30 with alimentary lymphoma (LSA), based on histopathology results and additional testing with immunohistochemistry or molecular clonality testing with PCR if indicated. Nineteen apparently healthy control cats were included in the study. One fecal sample was collected from each cat and S100A12 concentrations were quantified by an analytically validated in-house ELISA. RESULTS: Fecal S100A12 concentrations differed between cats with LSA (median 110 ng/g; interquartile range [IQR] 18-548) and control cats (median 4 ng/g; IQR 2-25 [P <0.001]) and between cats with IBD (median 34 ng/g; IQR 15-973) and control cats (P <0.003). S100A12 concentrations in CE cats (median 94 ng/g; IQR 16-548) were statistically significantly higher compared with control cats (P <0.001). The area under the receiver operating characteristic curve (AUROC) to separate healthy cats from CE cats was 0.81 (95% confidence interval [CI] 0.70-0.92) and was statistically significant (P <0.001). The AUROC to separate cats with IBD from cats with LSA was 0.51 (95% CI 0.34-0.68) and was not statistically significant (P = 0.9). CONCLUSIONS AND RELEVANCE: Fecal S100A12 concentrations at the time of diagnostic investigation were higher in cats with CIE and LSA than in healthy controls but did not differ between cats with LSA and those with CIE/IBD. This study is an initial step toward evaluating a novel non-invasive marker of feline CIE. Further studies are needed to determine the diagnostic utility of fecal S100A12 concentrations in cats with CE, including comparing cats with IBD/CIE and LSA, and to compare them with cats with extra-gastrointestinal disease.

Fecal Microbiota, Bile Acids, Sterols, and Fatty Acids in Dogs with Chronic Enteropathy Fed a Home-Cooked Diet Supplemented with Coconut Oil.

Medium-chain fatty acids (MCFAs) are considered to be interesting energy sources for dogs affected by chronic enteropathies (CE). This study analyzed the clinical scores, fecal microbiota, and metabolomes of 18 CE dogs fed a home-cooked diet (HCD) supplemented with virgin coconut oil (VCO), a source of MCFA, at 10% of metabolizable energy (HCD + VCO). The dogs were clinically evaluated with the Canine Chronic Enteropathy Activity Index (CCECAI) before and at the end of study. Fecal samples were collected at baseline, after 7 days of HCD, and after 30 days of HCD + VCO, for fecal score (FS) assessment, microbial analysis, and determination of bile acids (BA), sterols, and fatty acids (FA). The dogs responded positively to diet change, as shown by the CCECAI improvement (p = 0.001); HCD reduced fecal fat excretion and HCD + VCO improved FS (p < 0.001), even though an increase in fecal moisture occurred due to HCD (p = 0.001). HCD modified fecal FA (C6:0: +79%, C14:0: +74%, C20:0: +43%, C22:0: +58%, C24:0: +47%, C18:3n-3: +106%, C20:4n-6: +56%, and monounsaturated FA (MUFA): -23%, p < 0.05) and sterol profile (coprostanol: -27%, sitostanol: -86%, p < 0.01). VCO increased (p < 0.05) fecal total saturated FA (SFA: +28%, C14:0: +142%, C16:0 +21%, C22:0 +33%) and selected MCFAs (+162%; C10:0 +183%, C12:0 +600%), while reducing (p < 0.05) total MUFA (-29%), polyunsaturated FA (-26%), campesterol (-56%) and phyto-/zoosterols ratio (0.93:1 vs. 0.36:1). The median dysbiosis index was <0 and, together with fecal BA, was not significantly affected by HCD nor by VCO. The HCD diet increased total fecal bacteria (p = 0.005) and the abundance of Fusobacterium spp. (p = 0.028). This study confirmed that clinical signs, and to a lesser extent fecal microbiota and metabolome, are positively influenced by HCD in CE dogs. Moreover, it has been shown that fecal proportions of MCFA increased when MCFAs were supplemented in those dogs. The present results emphasize the need for future studies to better understand the intestinal absorptive mechanism of MCFA in dogs.

Chronic Inflammatory Enteropathy and Low-Grade Intestinal T-Cell Lymphoma Are Associated with Altered Microbial Tryptophan Catabolism in Cats.

Chronic inflammatory enteropathy (CIE) and low-grade intestinal T-cell lymphoma (LGITL) are common chronic enteropathies (CE) in cats. Enteric microbiota dysbiosis is implicated in the pathogenesis of CE; however, the mechanisms of host-microbiome interactions are poorly understood in cats. Microbial indole catabolites of tryptophan (MICT) are gut bacterial catabolites of tryptophan that are hypothesized to regulate intestinal inflammation and mucosal barrier function. MICTs are decreased in the sera of humans with inflammatory bowel disease and previous studies identified altered tryptophan metabolism in cats with CE. We sought to determine whether MICTs were decreased in cats with CE using archived serum samples from cats with CIE (n = 44) or LGITL (n = 31) and healthy controls (n = 26). Quantitative LC-MS/MS was used to measure serum concentrations of tryptophan, its endogenous catabolites (kynurenine, kynurenate, serotonin) and MICTs (indolepyruvate, indolealdehyde, indoleacrylate, indoleacetamide, indoleacetate, indolelactate, indolepropionate, tryptamine). Serum concentrations of tryptophan, indolepropionate, indoleacrylate, indolealdehyde, indolepyruvate, indolelactate were significantly decreased in the CIE and LGITL groups compared to those in healthy controls. Indolelactate concentrations were significantly lower in cats with LGITL compared to CIE (p = 0.006). Significant correlations were detected among serum MICTs and cobalamin, folate, fPLI, and fTLI. Our findings suggest that MICTs are promising biomarkers to investigate the role of gut bacteria in the pathobiology of chronic enteropathies in cats.

Analytical Validation of an Assay for Concurrent Measurement of Amino Acids in Dog Serum and Comparison of Amino Acid Concentrations between Whole Blood, Plasma, and Serum from Dogs.

Amino acids play an important role in metabolism. Comprehensive analytical validation of an assay for the concurrent measurement of a large number of amino acids in dogs is lacking, which precludes its usefulness in a clinical setting. Amino acids are often measured in plasma or whole blood. However, serum is commonly used for gastrointestinal diagnostic testing in dogs and is therefore convenient to use. This study aimed to analytically validate an assay for the concurrent measurement of amino acids in dog serum and to evaluate differences in amino acid concentrations in whole blood, plasma, and serum in dogs. Analytical validation of the assay (Biochrom 30+ Amino Acid Analyzer) was performed on fresh or banked serum samples from dogs. Whole blood, plasma, and serum from 36 healthy dogs were analyzed, and concentrations of the three sample types were compared. The assay was demonstrated to be precise, reproducible, accurate, linear, and stable for the measurement of the majority of compounds detected in dog serum. Cystine, glutamic acid, and ethanolamine were shown to be unstable at conditions commonly encountered in clinical settings. Significant differences in concentrations were identified between whole blood, plasma, and serum for 33 of 42 compounds. Amino acid profiles in serum and plasma were more similar to each other than to those in whole blood. While some amino acids are present in similar concentrations in whole blood, plasma, and serum, others are highly dependent on the type of biofluid, and measurements warrant strict adherence to sample type-based reference intervals.

Whole blood and plasma taurine reference intervals in adult Cavalier King Charles Spaniels and correlations between taurine concentration, diet and mitral valve disease.

OBJECTIVE: To determine breed-specific reference intervals for whole blood (WB) and plasma taurine concentrations in adult, overtly healthy Cavalier King Charles Spaniels (CKCSs) and determine whether taurine concentrations differ across preclinical myxomatous mitral valve disease (MMVD) stages or between CKCSs eating diets that meet World Small Animal Veterinary Association (WSAVA) nutritional guidelines versus other diets. ANIMALS: 200 privately owned CKCSs. PROCEDURES: Clinically healthy adult CKCSs were recruited prospectively. Diet and supplement history was collected. Dogs were staged by echocardiography using MMVD consensus guidelines. Taurine concentrations were measured in deproteinized lithium heparin WB and plasma samples with the postcolumn ninhydrin derivatization method on a dedicated amino acid analyzer. RESULTS: There were 12 stage A (6%), 150 stage B1 (75%), and 38 stage B2 (19%) CKCSs. Seventy-eight dogs (39%) were reported by their owners to be eating diets meeting WSAVA nutritional guidelines; 116 (58%) were not. Taurine concentrations in plasma (P = .444) and WB (P = .073) were not significantly different across MMVD stages or between CKCSs eating diets meeting WSAVA nutritional guidelines versus other diets (P = .345 and P = .527, respectively). Reference intervals for WB taurine (152 to 373 µM) and plasma taurine (51 to 217 µM) concentrations in CKCSs were generated. CLINICAL RELEVANCE: In CKCSs, taurine concentrations do not differ significantly based on preclinical MMVD stage, nor do they differ significantly based on consumption of a diet that does or does not meet WSAVA nutritional guidelines.

Detection of invasive Escherichia coli in dogs with granulomatous colitis using immunohistochemistry.

Granulomatous colitis in dogs can be associated with infection of the colonic mucosa by invasive strains of Escherichia coli. To date, fluorescence in situ hybridization (FISH) is the gold-standard method to assess intramucosal and intracellular bacterial invasion. However, FISH requires expensive fluorescence microscopy equipment and is therefore not widely available. We investigated the use of immunohistochemistry (IHC) as an alternative method to detect invasive E. coli in dogs with granulomatous colitis. Archived paraffin-embedded blocks were selected from 26 dogs with colitis, in which FISH had been performed by an outside laboratory. Using a polyclonal antibody, IHC for E. coli was performed on sections cut from the same blocks, and the presence of invasive E. coli was recorded. All 11 specimens in which FISH had detected E. coli were also positive on IHC, with strong immunolabeling in the cytoplasm of macrophages and extracellularly in the lamina propria; all 15 specimens that were negative for invasive bacteria on FISH were also negative on IHC. We found that IHC is a sensitive technique for the detection of invasive E. coli in dogs with granulomatous colitis.

Dysbiosis index to evaluate the fecal microbiota in healthy cats and cats with chronic enteropathies.

OBJECTIVES: Previous studies have identified various bacterial taxa that are altered in cats with chronic enteropathies (CE) vs healthy cats. Therefore, the aim of this study was to develop a targeted quantitative molecular method to evaluate the fecal microbiota of cats. METHODS: Fecal samples from 80 client-owned healthy cats and 68 cats with CE were retrospectively evaluated. A panel of quantitative PCR (qPCR) assays was used to measure the fecal abundance of total bacteria and seven bacterial taxa: Bacteroides, Bifidobacterium, Clostridium hiranonis, Escherichia coli, Faecalibacterium, Streptococcus and Turicibacter. The nearest centroid classifier algorithm was used to calculate a dysbiosis index (DI) based on these qPCR abundances. RESULTS: The abundances of total bacteria, Bacteroides, Bifidobacterium, C hiranonis, Faecalibacterium and Turicibacter were significantly decreased, while those of E coli and Streptococcus were significantly increased in cats with CE (P <0.027 for all). The DI in cats with CE was significantly higher compared with healthy cats (P <0.001). When the cut-off value of the DI was set at 0, it provided 77% (95% confidence interval [CI] 66-85) sensitivity and 96% (95% CI 89-99) specificity to differentiate the microbiota of cats with CE from those of healthy cats. Fifty-two of 68 cats with CE had a DI >0. CONCLUSIONS AND RELEVANCE: A qPCR-based DI for assessing the fecal microbiota of cats was established. The results showed that a large proportion of cats with CE had an altered fecal microbiota as evidenced by an increased DI. Prospective studies are warranted to evaluate the utility of this assay for clinical assessment of feline CE.

Serum pancreatic lipase immunoreactivity in sick dogs after chronic administration of supraphysiologic doses of glucocorticoids.

BACKGROUND: Glucocorticoids (GC) are commonly used for a long term to treat a multitude of immune-mediated, inflammatory, and neoplastic diseases in dogs. Conflicting results of published studies on the effects of exogenous and endogenous GCs on serum canine pancreatic lipase immunoreactivity (cPLI) raise the question of whether cPLI concentrations can be reliably interpreted in patients receiving GCs. OBJECTIVE: We sought to determine the effect of long-term GC administration at supraphysiologic doses on serum cPLI concentrations in sick dogs. METHODS: Serum samples were collected from 35 client-owned dogs. Dogs were administered prednisone at a dose of ≥0.5 mg/kg per day for ≥3 weeks. Serum cPLI was measured prior to the initiation and after ≥3 weeks of GC therapy. RESULTS: There was a significant increase in serum cPLI between baseline (median 101 µg/L; range 30-1997 µg/L) and following the administration of ≥0.5 mg/kg/day of prednisone (median 173 µg/L; range 30-2000 µg/L) in dogs (P = 0.025). However, the median change was small (31 µg/L). There was no suspicion of pancreatitis in any of the dogs. Diagnostic interpretation changed in 6/35 dogs, with no apparent dose-response relationship. CONCLUSIONS: There was a statistically significant difference from baseline in serum cPLI measurements in sick dogs receiving long-term prednisone. Although the change was small and often clinically insignificant, it could pose a clinical interpretation dilemma in some dogs. It is unknown whether these observations are coincidental due to subclinical pancreatitis or caused by another effect of GCs on pancreatic acinar cells.

Analysis of the gut microbiome in dogs and cats.

The gut microbiome is an important immune and metabolic organ. Intestinal bacteria produce various metabolites that influence the health of the intestine and other organ systems, including kidney, brain, and heart. Changes in the microbiome in diseased states are termed dysbiosis. The concept of dysbiosis is constantly evolving and includes changes in microbiome diversity and/or structure and functional changes (eg, altered production of bacterial metabolites). Molecular tools are now the standard for microbiome analysis. Sequencing of microbial genes provides information about the bacteria present and their functional potential but lacks standardization and analytical validation of methods and consistency in the reporting of results. This makes it difficult to compare results across studies or for individual clinical patients. The Dysbiosis Index (DI) is a validated quantitative PCR assay for canine fecal samples that measures the abundance of seven important bacterial taxa and summarizes the results as one single number. Reference intervals are established for dogs, and the DI can be used to assess the microbiome in clinical patients over time and in response to therapy (eg, fecal microbiota transplantation). In situ hybridization or immunohistochemistry allows the identification of mucosa-adherent and intracellular bacteria in animals with intestinal disease, especially granulomatous colitis. Future directions include the measurement of bacterial metabolites in feces or serum as markers for the appropriate function of the microbiome. This article summarizes different approaches to the analysis of gut microbiota and how they might be applicable to research studies and clinical practice in dogs and cats.

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.

Effects of dietary macronutrient profile on apparent total tract macronutrient digestibility and fecal microbiota, fermentative metabolites, and bile acids of female dogs after spay surgery.

Obesity and estrogen reduction are known to affect the gut microbiota and gut microbial-derived metabolites in some species, but limited information is available in dogs. The aim of this study was to determine the effects of dietary macronutrient profile on apparent total tract macronutrient digestibility, fecal microbiota, and fecal metabolites of adult female dogs after spay surgery. Twenty-eight adult intact female beagles (age: 3.02 ± 0.71 yr, BW: 10.28 ± 0.77 kg; BCS: 4.98 ± 0.57) were used. After a 5-wk baseline phase (week 0), 24 dogs were spayed and randomly allotted to one of three experimental diets (n = 8 per group): 1) control (CO) containing moderate protein and fiber (COSP), 2) high-protein, high-fiber (HPHF), or 3) high-protein, high-fiber plus omega-3 and medium-chain fatty acids (HPHFO). Four dogs were sham-operated and fed CO (COSH). All dogs were fed to maintain BW for 12 wk after spay and then allowed to consume twice that amount for 12 wk. Fecal samples were collected at weeks 0, 12, and 24 for digestibility, microbiota, and metabolite analysis. All data were analyzed using repeated measures and linear mixed models procedure of SAS 9.4, with results reported as a change from baseline. Apparent organic matter and energy digestibilities had greater decreases in HPHF and HPHFO than COSH and COSP. Increases in fecal acetate, total short-chain fatty acids, and secondary bile acids were greater and decreases in primary bile acids were greater in HPHF and HPHFO. Principal coordinates analysis of weighted UniFrac distances revealed that HPHF and HPHFO clustered together and separated from COSH and COSP at weeks 12 and 24, with relative abundances of Faecalibacterium, Romboutsia, and Fusobacterium increasing to a greater extent and Catenibacterium, Bifidobacterium, Prevotella 9, Eubacterium, and Megamonas decreasing to a greater extent in HPHF or HPHFO. Our results suggest that high-protein, high-fiber diets alter nutrient and energy digestibilities, fecal metabolite concentrations, and fecal gut microbiota, but spay surgery had minor effects. Future research is needed to investigate how food intake, nutrient profile, and changes in hormone production influence gut microbiota and metabolites of dogs individually and how this knowledge may be used to manage spayed pets.

Untargeted fecal metabolome analysis in obese dogs after weight loss achieved by feeding a high-fiber-high-protein diet.

INTRODUCTION: In humans and companion animals, obesity is accompanied by metabolic derangements. Studies have revealed differences in the composition of the fecal microbiome between obese dogs and those with an ideal body weight. OBJECTIVES: We have previously reported that the fecal microbiome in obese dogs changes after controlled weight reduction, induced by feeding a diet high in fiber and protein. Despite these findings, it is unclear if taxonomic differences infer differences at the functional level between obese dogs and those with an ideal body weight. METHODOLOGY: Untargeted fecal metabolome analysis was performed on dogs with obesity before and after weight loss achieved by feeding a high-fiber-high-protein diet. RESULTS: Fecal metabolome analysis revealed a total of 13 compounds that changed in concentration in obese dogs after weight loss. Of these compounds, metabolites associated with bacterial metabolism decreased after weight loss including purine, L-(-)-methionine, coumestrol, and the alkaloids 1-methylxanthine and trigonelline. Conversely, the polyphenols (-)-epicatechin and matairesinol and the quinoline derivatives 1,5-isoquinolinediol and 2-hydroxiquinoline increased after weight loss. CONCLUSION: These results suggest differences in intestinal microbiome at the functional level after weight loss, but further studies are needed to determine the role of these compounds in the etiology of obesity and weight loss.

Metabolomics Analysis Reveals Deranged Energy Metabolism and Amino Acid Metabolic Reprogramming in Dogs With Myxomatous Mitral Valve Disease.

Background Myxomatous mitral valve disease (MMVD), a naturally occurring heart disease, affects 10% to 15% of the canine population. Canine MMVD shares many similarities with human MMVD. Untargeted metabolomics was performed to identify changes in metabolic pathways and biomarkers with potential clinical utilities. Methods and Results Serum samples from 27 healthy, 22 stage B1, 18 stage B2 preclinical MMVD dogs, and 17 MMVD dogs with a history of congestive heart failure (CHF) were analyzed. Linear regression analysis identified 173 known metabolites whose concentrations were different among the 4 groups (adjusted P<0.05), of which 40% belonged to amino acid super pathways, while 30% were lipids. More than 50% of significant metabolites were correlated with left atrial diameter but not left ventricular dimension. Acylcarnitines, tricarboxylic acid cycle intermediates, and creatine accumulated in proportion to MMVD severity. α-Ketobutyrate and ketone bodies were increased as MMVD advanced. Nicotinamide, a key substrate of the main nicotinamide adenine dinucleotide (NAD+) salvage pathway, was decreased, while quinolinate of the de novo NAD+ biosynthesis was increased in CHF dogs versus healthy dogs. 3-Methylhistidine, marker for myofibrillar protein degradation, was higher in CHF dogs than non-CHF dogs. Trimethylamine N-oxide (TMAO) and TMAO-producing precursors, including carnitine, phosphatidylcholine, betaine, and trimethyllysine, were increased in CHF dogs versus non-CHF dogs. Elevated levels of uremic toxins, including guanidino compounds, TMAO, and urea, were observed in CHF dogs. Pathway analysis highlighted the importance of bioenergetics and amino acid metabolism in canine MMVD. Conclusions Our study revealed altered energy metabolism, amino acid metabolic programming, and reduced renal function in the development of MMVD and CHF. Complex interplays along the heart-kidney-gut axis were implicated.

Untargeted metabolomic analysis in cats with naturally occurring inflammatory bowel disease and alimentary small cell lymphoma.

Feline chronic enteropathy (CE) is a common gastrointestinal disorder in cats and mainly comprises inflammatory bowel disease (IBD) and small cell lymphoma (SCL). Differentiation between IBD and SCL can be diagnostically challenging. We characterized the fecal metabolome of 14 healthy cats and 22 cats with naturally occurring CE (11 cats with IBD and 11 cats with SCL). Principal component analysis and heat map analysis showed distinct clustering between cats with CE and healthy controls. Random forest classification revealed good group prediction for healthy cats and cats with CE, with an overall out-of-bag error rate of 16.7%. Univariate analysis indicated that levels of 84 compounds in cats with CE differed from those in healthy cats. Polyunsaturated fatty acids held discriminatory power in differentiating IBD from SCL. Metabolomic profiles of cats with CE resembled those in people with CE with significant alterations of metabolites related to tryptophan, arachidonic acid, and glutathione pathways.