Fecal bile acid dysmetabolism and reduced ursodeoxycholic acid correlate with novel microbial signatures in feline chronic kidney disease.

Background: Microbial-derived secondary bile acids (SBAs) are reabsorbed and sensed via host receptors modulating cellular inflammation and fibrosis. Feline chronic kidney disease (CKD) occurs with progressive renal inflammation and fibrosis, mirroring the disease pathophysiology of human CKD patients. Methods: Prospective cross-sectional study compared healthy cats (n = 6) with CKD (IRIS Stage 2 n = 17, Stage 3 or 4 n = 11). Single timepoint fecal samples from all cats underwent targeted bile acid metabolomics. 16S rRNA gene amplicon sequencing using DADA2 with SILVA taxonomy characterized the fecal microbiota. Results: CKD cats had significantly reduced fecal concentrations (median 12.8 ng/mg, Mann-Whitney p = 0.0127) of the SBA ursodeoxycholic acid (UDCA) compared to healthy cats (median 39.4 ng/mg). Bile acid dysmetabolism characterized by <50% SBAs was present in 8/28 CKD and 0/6 healthy cats. Beta diversity significantly differed between cats with <50% SBAs and > 50% SBAs (PERMANOVA p < 0.0001). Twenty-six amplicon sequence variants (ASVs) with >97% nucleotide identity to Peptacetobacter hiranonis were identified. P. hiranonis combined relative abundance was significantly reduced (median 2.1%) in CKD cats with <50% SBAs compared to CKD cats with >50% SBAs (median 13.9%, adjusted p = 0.0002) and healthy cats with >50% SBAs (median 15.5%, adjusted p = 0.0112). P. hiranonis combined relative abundance was significantly positively correlated with the SBAs deoxycholic acid (Spearman r = 0.5218, adjusted p = 0.0407) and lithocholic acid (Spearman r = 0.5615, adjusted p = 0.0156). Three Oscillospirales ASVs and a Roseburia ASV were also identified as significantly correlated with fecal SBAs. Clinical and translational importance: The gut-kidney axis mediated through microbial-derived SBAs appears relevant to the spontaneous animal CKD model of domestic cats. This includes reduced fecal concentrations of the microbial-derived SBA UDCA, known to regulate inflammation and fibrosis and be reno-protective. Microbes correlated with fecal SBAs include bai operon containing P. hiranonis, as well as members of Oscillospirales, which also harbor a functional bai operon. Ultimately, CKD cats represent a translational opportunity to study the role of SBAs in the gut-kidney axis, including the potential to identify novel microbial-directed therapeutics to mitigate CKD pathogenesis in veterinary patients and humans alike.

The Effect of Renaltec on Serum Uremic Toxins in Cats with Experimentally Induced Chronic Kidney Disease.

Serum uremic toxins markedly increase in cats with chronic kidney disease (CKD) and have deleterious consequences. Renaltec is an oral adsorbent that binds uremic toxin precursors in the gut. In this prospective cohort study utilizing 13 purpose-bred cats with remnant kidney model-induced CKD (12 IRIS Stage 2, 1 IRIS Stage 3) eating a standardized renal diet, we aimed to assess the effect of Renaltec administration on serum indoxyl sulfate (IDS) and p-cresol sulfate (pCS) concentrations. Cats were sequentially treated with standard of care for 56 days, 500 mg Renaltec orally once daily for 56 days, and then three months later, 500 mg Renaltec orally twice daily for 56 days. Serum IDS and pCS concentrations were measured 28 and 56 days after the administration of Renaltec. Blood pressure and kidney function were measured before and 56 days after the administration of Renaltec. Significant decreases in serum IDS and pCS concentrations were observed for both once- and twice-daily dosing, particularly during the first 28 days of administration. More cats with BID dosing had clinically significant reductions in serum IDS and pCS concentrations than with SID dosing. Renaltec can reduce the serum concentrations of deleterious gut-derived uremic toxins in cats with CKD.

Effect of repeated administration of a parenteral feline herpesvirus-1, calicivirus, and panleukopenia virus vaccine on select clinicopathologic, immunological, renal histologic, and immunohistochemical parameters in healthy adult cats.

OBJECTIVE: To assess whether hyperinoculation of cats with a feline herpesvirus-1, calicivirus, and panleukopenia virus (FVRCP) vaccine could be used as a model to study interstitial nephritis and to assess humoral and cell-mediated immune responses toward vaccinal α-enolase. ANIMALS: 6 healthy young adult purpose-bred research cats. PROCEDURES: Baseline renal cortical biopsies, whole blood, serum, and urine were collected prior to administration of a commercial FVRCP parenteral vaccine. Vaccine hyperinoculation was defined as a total of 8 vaccinations given at 2-week intervals over a 14-week period. Blood samples were collected immediately prior to each vaccination, and a second renal biopsy was performed 2 weeks after hyperinoculation (week 16). Renal histopathology, renal α-enolase immunohistochemistry, and assays to detect humoral and cell-mediated immune reactions against Crandell-Rees feline kidney (CRFK) cell lysates and α-enolase were performed. An α-enolase immunoreactivity score for renal tubules and glomeruli based on signal intensity was determined by a blinded pathologist. RESULTS: Hyperinoculation with the vaccine was not associated with clinicopathologic evidence of renal dysfunction, and interstitial nephritis was not recognized by light microscopy in the time studied. The mean serum absorbance values for antibodies against CRFK antigen and α-enolase were significantly (P < 0.001) higher at weeks 4, 8, and 16 versus week 0. Renal tubular and glomerular α-enolase immunoreactivity scores were higher at week 16 compared to baseline. CLINICAL RELEVANCE: Findings suggested that systemic immunological reactions occurred and renal tissues were affected by vaccine hyperinoculation; however, short-term FVRCP vaccine hyperinoculation cannot be used to study interstitial nephritis in cats.

Urine bacterial culture growth and association with urine sedimentation and clinical findings in cats with acute kidney injury.

OBJECTIVES: A urine culture is often pursued in cats with acute kidney injury (AKI) to screen for bacterial growth in the urine, but it can be cost prohibitive. The aim of the study was to determine the ability of a urinalysis and lower urinary tract signs (LUTS) to predict urine culture results in cats with AKI. METHODS: Ninety-seven cats with AKI were included in this study. This was a retrospective, observational study. Medical records from 2008 to 2018 were reviewed to identify cats with AKI that had a paired urinalysis and urine bacterial culture. The sensitivity, specificity, positive predictive value and negative predictive values of microscopic bacteriuria, pyuria, hematuria and the presence of LUTS for predicting urine culture results was calculated. RESULTS: Thirty-two percent of cats (n = 31) had a positive urine culture. Of these, 28 (90%) had bacteriuria, 21 (68%) had pyuria, 13 (42%) had hematuria and 10 (32%) had LUTS. Of the 42 cats without hematuria or pyuria, seven had a positive urine culture (17%). Bacteriuria had a high sensitivity (90%) and specificity (92%) for predicting urine culture bacterial growth. The absence of bacteriuria had a high negative predictive value for no bacterial growth (95%). The odds of a positive urine culture were increased with bacteriuria (odds ratio [OR] 114, 95% confidence interval [CI] 29-621; P <0.001), pyuria (OR 21, 95% CI 7-70; P <0.001) and LUTS (OR 5, 95% CI 1.7-16; P = 0.004). Hematuria was not associated with a positive culture (sensitivity 42%, specificity 52%). CONCLUSIONS AND RELEVANCE: Microscopic bacteriuria and pyuria on urine sediment evaluation and LUTS can be helpful for predicting bacterial culture results in cats with AKI and in settings where submitting a urine culture may not be financially feasible.

Serum and Fecal Amino Acid Profiles in Cats with Chronic Kidney Disease.

The purpose of the study was to quantify serum and fecal amino acids (AA) in cats with chronic kidney disease (CKD) and compare to healthy cats. Thirty-five cats with International Renal Interest Society Stage 1-4 CKD and 16 healthy mature adult and senior client-owned cats were included in this prospective cross-sectional study. Sera were analyzed for 25 AA concentrations using an ion exchange chromatography AA analyzer with post column ninhydrin derivatization. Voided fecal samples were analyzed for 22 AA concentrations using liquid chromatography with tandem mass spectrometry. CKD cats had lower serum concentrations of phenylalanine (mean difference ± standard error of the mean: 12.7 ± 4.3 µM; p = 0.03), threonine (29.6 ± 9.2 µM; p = 0.03), tryptophan (18.4 ± 5.4 µM; p = 0.005), serine (29.8 ± 12.6 µM; p = 0.03), and tyrosine (11.6 ± 3.8 µM; p = 0.01) and higher serum concentrations of aspartic acid (4.7 ± 2.0 µM; p = 0.01), ß-alanine (3.4 ± 1.2 µM; p = 0.01), citrulline (5.7 ± 1.6 µM; p = 0.01), and taurine (109.9 ± 29.6 µM; p = 0.01) when compared to healthy cats. Fecal AA concentrations did not differ between healthy cats and CKD cats. 3-Methylhistidine-to-creatinine did not differ between healthy cats with and without muscle loss. Cats with CKD IRIS Stages 1-4 have a deranged serum amino acid profile compared to healthy cats.

Survey of defecation habits in apparently healthy and chronic kidney disease cats.

OBJECTIVES: Changes in bowel movements (BMs) are an important clinical sign in many diseases, including chronic kidney disease (CKD), and the purpose of this study was to collect information on BMs and fecal scores in both apparently healthy and CKD cats. A secondary aim was to assess owner awareness of BM frequency. METHODS: Owners were asked to complete an initial online questionnaire about their cat's health and litter box habits (including predicted BM frequency and fecal scores) and were then asked to clean the box daily for 7 days and report results (observed frequency of BMs and fecal scores) daily. Differences in BM frequency and fecal scores between apparently normal and CKD cats were compared using the Mann-Whitney test, and predicted vs observed data were compared using the Wilcoxon signed rank test. Difference in percentage of cats defecating more or less than once daily were assessed with Fisher's exact test. RESULTS: Survey data from 124 owners of apparently healthy cats and 43 owners of CKD cats who submitted two or more days of daily observations (in addition to the initial questionnaire) were analyzed. Eighty-five percent of apparently healthy cats were observed to defecate one or more times per day and 15% defecated less than once per day. Fifty-eight percent of CKD cats defecated one or more times per day and 42% defecated less than once per day. A significantly higher percentage of CKD cats defecated less than once per day in comparison with apparently healthy cats (P <0.0001). Observed BM frequency was significantly less in CKD cats compared with healthy cats (P = 0.02). Observed fecal scores were not significantly different between healthy and CKD cats. CONCLUSIONS AND RELEVANCE: The observed BM frequency of cats with CKD was less than apparently healthy cats and represents a clinically important variation from normal.

Impact of specimen type on findings for bacterial composition within the intestinal tract of dogs and cats with and without chronic enteropathy.

OBJECTIVE: To compare bacterial diversity and community composition among fecal, rectal swab, and colonic mucosal biopsy specimens from dogs and cats with and without chronic enteropathy (CE). ANIMALS: 9 healthy dogs, 8 dogs with CE, 8 healthy cats, and 9 cats with CE. PROCEDURES: In a cross-sectional study design, fecal, rectal swab, and colonic mucosal biopsy specimens were obtained by colonoscopy from healthy dogs and dogs and cats with CE. Fecal and rectal swab specimens were collected from healthy cats. Genomic DNA was extracted, the 16S rRNA V4 gene region was amplified, and sequencing was performed by use of primers 515F to 806R on a paired-end platform. RESULTS: For healthy dogs and dogs and cats with CE, bacterial diversity based on the Chao1 estimate of total species richness was higher for colonic mucosal biopsy specimens than for fecal specimens. Analysis of similarities by use of the Bray-Curtis dissimilarity index revealed that the bacterial communities captured in rectal swab specimens were similar to those captured in fecal specimens for healthy dogs and dogs with CE and similar to those captured in colonic mucosal biopsy specimens for both dog groups and cats with CE. CONCLUSIONS AND CLINICAL RELEVANCE: Rectal swab and colonic biopsy specimens were successfully used to characterize the bacteriome of the intestinal tract in dogs and cats by 16S rRNA gene sequencing. Although the specimen types evaluated in this study were not interchangeable in results, rectal swab specimens were practical to collect from dogs and cats to study bacterial composition within the intestinal tract and may provide an alternative to colonic mucosal biopsy and fecal specimens.

Evaluation of platelet function in cats with and without kidney disease: a pilot study.

OBJECTIVES: The aims of this study were to determine if stable chronic kidney disease (CKD) cats and uremic crisis cats have altered platelet function, and to determine the prevalence of positive fecal occult blood in CKD cats. METHODS: Platelet function in normal cats, clinically stable International Renal Interest Society (IRIS) stage 2-4 CKD cats and CKD cats experiencing a uremic crisis were evaluated using impedance aggregometry. Area under the curve (AUC) at 6 mins was calculated for saline, adenosine diphosphate (AUCADP) and arachidonic acid (AUCASPI). The AUC in addition to hematocrit, platelet count and mean platelet volume (MPV) were compared between groups using the Kruskal-Wallis test followed by Dunn's post-hoc analysis. Guaiac fecal occult blood tests were performed on fecal samples and results were compared between groups using a χ2 for trend test. RESULTS: AUCADP (P = 0.04) and AUCASPI (P = 0.05) were significantly higher in uremic crisis cats compared with normal cats at 6 mins. Hematocrit was significantly higher in normal cats when compared with IRIS stage 3 and 4 (P = 0.002) and uremic crisis (P = 0.0008) cats, with no difference among groups for platelet count or MPV. The proportion of cats with positive fecal occult blood samples was significantly different between groups (P = 0.0017); 50% uremic crisis cats, 33% IRIS stage 3 and 4 cats, and 10% IRIS stage 2 cats were positive, while no normal cats were positive. The proportion of cats with platelet clumping was significantly different between groups (P = 0.03). CONCLUSIONS AND RELEVANCE: Platelet hyper-reactivity may be occurring in CKD cats experiencing a uremic crisis. The etiology of positive fecal occult blood samples in CKD cats is unclear and did not appear to be related to decreased platelet function as measured in this study and requires further investigation.

Evaluation of nutrient content and caloric density in commercially available foods formulated for senior cats.

BACKGROUND: Cat foods marketed for senior cats (≥7 years) are available to owners. The variability in the nutritional content of these foods is unknown. OBJECTIVES: To measure the caloric density and caloric distribution of crude protein, crude fiber, crude fat, phosphorus, calcium, magnesium, sodium, potassium, and vitamin D3 in commercially available foods for senior cats and to compare nutrient content with foods for adult cats. SAMPLES: Thirty-one senior and 59 adult commercial nontherapeutic cat food products. METHODS: Descriptive study. Crude protein, crude fiber, and crude fat were measured using Dumas nitrogen combustion, Ankom filter bag technique, and acid hydrolysis, respectively. Mineral concentrations were measured using inductively coupled argon plasma-optical emission spectroscopy. Vitamin D3 was determined by liquid chromatography with tandem mass spectrometry. Caloric density was calculated using modified Atwater values. RESULTS: The evaluated nutrient concentrations in all foods for senior cats met the values of the Association of American Feed Control Officials Cat Food Nutrient Profile for adult maintenance. Foods for senior cats had significantly higher crude fiber content when compared to foods for adult cats (P < .0001). No significant difference in crude protein, crude fat and mineral concentrations was found between foods for senior and adult cats. CONCLUSIONS AND CLINICAL IMPORTANCE: Foods marketed for senior cats are highly variable in their caloric density and nutrient content and, except for crude fiber, are similar to foods for adult cats. Veterinarians should avoid broad recommendations regarding commercially available foods for senior cats, and dietary recommendations should reflect the patient's individual needs.

Evaluation of phosphorus, calcium, and magnesium content in commercially available foods formulated for healthy cats.

BACKGROUND: High dietary phosphorus (P) and low calcium-to-phosphorus ratio (Ca:P) are associated with kidney damage in cats. There are no established guidelines for dietary P maximum for cats. OBJECTIVES: To quantify crude protein, P, Ca, and magnesium (Mg) concentrations in cat foods and compare among food formats (dry, canned, raw), primary protein ingredients, protein concentrations (low, moderate, high), grain-free versus grain-containing foods, foods intended for adult maintenance versus all life stages, and cost. SAMPLES: Eighty-two commercial nonprescription cat foods. METHODS: Descriptive study. Mineral concentrations were measured using inductively coupled argon plasma-optical emission spectroscopy. Crude protein was measured using the Dumas nitrogen combustion method. Mineral and crude protein concentrations were compared among food categories. RESULTS: Twenty-seven foods contained ≥3.6 g P/1000 kcal metabolizable energy (ME), of which 7 exceeded 4.8 g/1000 kcal ME. Thirteen foods had low Ca:P ratio (≤1.0). The low-protein diet group had no products ≥3.6 g P/1000 kcal ME, which was significantly different compared to the high-protein diet group (52% of products had ≥3.6 g P/1000 kcal ME; P = .01). No significant differences in P content and Ca:P ratio were found among other diet categories. Canned foods had significantly lower Mg compared to dry (P < .001) and raw (P = .007) foods. Declared minimum P and Ca were significantly lower than analyzed concentrations (P = .0005 and P = .003, respectively). CONCLUSIONS AND CLINICAL IMPORTANCE: The high number of foods with high P and low Ca suggest that pet food regulatory reform should be considered.

Assessment of compounded transdermal mirtazapine as an appetite stimulant in cats with chronic kidney disease.

OBJECTIVES: The aim of this study was to assess the appetite stimulation properties of compounded transdermal mirtazapine (CTM) in cats with chronic kidney disease (CKD). METHODS: Two sequential double-blind placebo-controlled crossover prospective studies were performed in client-owned cats with stable stage 2 or 3 CKD and a history of decreased appetite. In the first study nine CKD cats were randomized to receive 3.75 mg/0.1 ml CTM gel or placebo on the inner pinna every other day for 3 weeks, then, after a 4 day washout period, the cats were crossed over to the alternate 3 week treatment. In a second study, 10 CKD cats were randomized to receive 1.88 mg/0.1 ml CTM or placebo on the same schedule. Physical examination and serum biochemistry were performed before and after each treatment period, and owners kept daily logs of appetite, activity and eating behaviors. Mirtazapine concentrations in CTM gels and steady-state mirtazapine serum concentrations were measured using liquid chromatography/tandem mass spectrometry. RESULTS: Administration of both 3.75 mg and 1.88 mg CTM resulted in a statistically significant increase in weight (P = 0.002 for both), increase in appetite (P = 0.01 and P = 0.005, respectively), and increase in rate of food consumption (P = 0.03 and P = 0.008, respectively). No significant difference in activity or vocalization was seen at either dose; however, individual cats experienced excessive meowing. Median weight increase for the 3.75 mg arm was 0.22 kg (range 0.04-0.44 kg), while median weight increase for the 1.88 mg arm was 0.26 kg (range -0.25 to 0.5 kg). Improvement in body condition score was seen in 5/9 cats in the 3.75 mg arm (P = 0.04) and 6/10 cats in the 1.88 mg arm (P = 0.004). CONCLUSIONS AND RELEVANCE: CTM increased appetite and resulted in weight gain in CKD cats despite significant inconsistencies in compounding, and may benefit cats in countries where an approved product is not available.

The fecal microbiome and serum concentrations of indoxyl sulfate and p-cresol sulfate in cats with chronic kidney disease.

BACKGROUND: Intestinal dysbiosis has been documented in humans with chronic kidney disease (CKD) and is thought to contribute to production of the uremic toxins indoxyl sulfate (IS) and p-cresol sulfate (pCS). Characteristics of the fecal microbiome in cats with CKD and correlation to serum concentrations of uremic toxins are unknown. OBJECTIVES: To characterize the fecal microbiome and measure serum IS and pCS concentrations of cats with CKD in comparison to healthy older cats. ANIMALS: Thirty client-owned cats with CKD (International Renal Interest Society stages 2-4) and 11 older (≥8 years) healthy control cats. METHODS: Prospective, cross-sectional study. Fecal samples were analyzed by sequencing of 16S rRNA genes and Escherichia coli quantitative PCR (qPCR). Serum concentrations of IS and pCS measured using liquid chromatography tandem mass spectrometry. RESULTS: Cats with CKD had significantly decreased fecal bacterial diversity and richness. Escherichia coli qPCR showed no significant difference in bacteria count between control and CKD cats. Cats with stage 2 (P = .01) and stages 3 and 4 (P = .0006) CKD had significantly higher serum IS concentrations compared to control cats. No significant difference found between stage 2 and stages 3 and 4 CKD. The pCS concentrations were not significantly different between CKD cats and control cats. CONCLUSIONS AND CLINICAL IMPORTANCE: Decreased fecal microbiome diversity and richness is associated with CKD in cats. Indoxyl sulfate concentration is significantly increased with CKD, and cats with stage 2 CKD may suffer from a similar uremic toxin burden as do cats with later stage disease.

Effect of modified live or inactivated feline herpesvirus-1 parenteral vaccines on clinical and laboratory findings following viral challenge.

Objectives The objective was to investigate the effect of one dose of an inactivated feline herpesvirus-1 (FHV-1), feline calicivirus (FCV) and panleukopenia virus (FPV) vaccine (FVRCP) or one dose of a modified live (ML) FVRCP vaccine on clinical signs and shedding of FHV-1 in specific pathogen-free kittens after challenge with FHV-1 7 days after vaccination. Methods Twenty-four FHV-1 seronegative 5-month-old kittens were randomized into three groups of eight kittens. Group 1 kittens were maintained as unvaccinated controls, group 2 kittens were administered one dose of the inactivated FVRCP vaccine subcutaneously (SC) and group 3 kittens were administered one dose of the ML FVRCP vaccine SC. All 24 cats were administered FHV-1 by nasal and oropharyngeal inoculation 7 days later and were observed daily for clinical signs of illness for 21 days. Results In the 21 days after FHV-1 challenge, both groups of vaccinated cats were less likely to be clinically ill (indicated by lower cumulative clinical scores) than control cats ( P <0.001). There was no statistical difference in total clinical score between the two vaccinated groups ( P = 0.97). Although the total clinical score was similar between both vaccines, signs of respiratory disease were significantly fewer in the kittens vaccinated with the inactivated FVRCP vaccine compared with the ML FVRCP vaccine ( P = 0.005) during the period after inoculation when the majority of clinical disease was observed. Conclusions and relevance Parenteral administration of either the inactivated FVRCP vaccine or the ML FVRCP vaccine can decrease clinical signs of illness due to FHV-1 on a day 7 challenge when compared with controls. Use of either vaccine product is indicated in cats at risk of acute exposure to FHV-1.