Gallbladder microbiota in healthy dogs and dogs with mucocele formation.

To date studies have not investigated the culture-independent microbiome of bile from dogs, a species where aseptic collection of bile under ultrasound guidance is somewhat routine. Despite frequent collection of bile for culture-based diagnosis of bacterial cholecystitis, it is unknown whether bile from healthy dogs harbors uncultivable bacteria or a core microbiota. The answer to this question is critical to understanding the pathogenesis of biliary infection and as a baseline to exploration of other biliary diseases in dogs where uncultivable bacteria could play a pathogenic role. A pressing example of such a disease would be gallbladder mucocele formation in dogs. This prevalent and deadly condition is characterized by excessive secretion of abnormal mucus by the gallbladder epithelium that can eventually lead to rupture of the gallbladder or obstruction of bile flow. The cause of mucocele formation is unknown as is whether uncultivable, and therefore unrecognized, bacteria play any systematic role in pathogenesis. In this study we applied next-generation 16S rRNA gene sequencing to identify the culture-negative bacterial community of gallbladder bile from healthy dogs and gallbladder mucus from dogs with mucocele formation. Integral to our study was the use of 2 separate DNA isolations on each sample using different extraction methods and sequencing of negative control samples enabling recognition and curation of contaminating sequences. Microbiota findings were validated by simultaneous culture-based identification, cytological examination of bile, and fluorescence in-situ hybridization (FISH) performed on gallbladder mucosa. Using culture-dependent, cytological, FISH, and 16S rRNA sequencing approaches, results of our study do not support existence of a core microbiome in the bile of healthy dogs or gallbladder mucus from dogs with mucocele formation. Our findings further document how contaminating sequences can significantly contribute to the results of sequencing analysis when performed on samples with low bacterial biomass.

Detection of Escherichia coli and Enterococcus spp. in dogs with polymicrobial urinary tract infections: A 5-year retrospective study.

BACKGROUND: Urinary tract infections (UTI) caused by Escherichia coli and Enterococcus spp., which are frequently coisolated in polymicrobial UTI, cause morbidity among dogs and warrant antimicrobial therapy. OBJECTIVES: To evaluate clinical features of dogs with polymicrobial E. coli and Enterococcal UTI. ANIMALS: Forty-four client-owned dogs with polymicrobial bacteriuria and groups of 100 client-owned dogs with E. coli and Enterococcal monomicrobial bacteriuria. METHODS: Retrospective cohort study of medical records of dogs at a university teaching hospital from 2014 to 2019. Prevalence of recurrent UTI and isolate antimicrobial resistance were determined. Clinical outcomes of dogs with recurrent UTI from groups including cost and hospital visits were compared. RESULTS: Recurrent UTI was more prevalent (P = .05) in dogs with polymicrobial bacteriuria (57%, 95% confidence interval [95% CI]: 42%-70%) compared to the Enterococcal monomicrobial group (40%, 95% CI: 31%-50%). Escherichia coli from polymicrobial bacteriuria were more frequently resistant to doxycycline (P < .01, 43%, 95% CI: 29%-58%) and gentamicin (P = .03, 17%, 95% CI: 9%-31%) compared to E. coli from monomicrobial bacteriuria (17% and 5%, 95% CI: 11%-26% and 2%-11% for doxycycline and gentamicin, respectively). Dogs with recurrent UTI from the polymicrobial UTI group had significantly (P = .05) more hospital visits (mean = 6 visits, 95% CI: 1.7-9.8) compared to recurrent monomicrobial UTI dogs (mean = 4 and 3 visits, 95% CI: 1.0 to 4.4 and -0.7 to 7.7 for E. coli and Enterococcal monomicrobial UTI, respectively). CONCLUSIONS AND CLINICAL IMPORTANCE: Escherichia coli and Enterococcus spp. polymicrobial UTI had more frequent adverse clinical outcomes for dogs.

Comparison of the Intestinal Pharmacokinetics of Two Different Florfenicol Dosing Regimens and Its Impact on the Prevalence and Phenotypic Resistance of E. coli and Enterococcus over Time.

In order to mitigate the food animal sector's role in the growing threat of antimicrobial resistance (AMR), the World Health Organization (WHO) suggests the use of lower tier antimicrobials, such as florfenicol. Florfenicol has two dosing schemes used to treat primarily bovine respiratory disease. In this study, the objective was to characterize the plasma and gastrointestinal pharmacokinetics of each dosing regimen and assess the effect of these dosing regimens on the prevalence of resistant indicator bacteria over time. Twelve steers underwent abdominal surgery to facilitate the placement of ultrafiltration probes within the lumen of the ileum and colon, as well as placement of an interstitial probe. Following surgery, cattle were dosed with either 20 mg/kg IM every 48 h of florfenicol given twice (n = 6) or a single, subcutaneous dose (40 mg/kg, n = 6). Plasma, interstitial fluid, gastrointestinal ultrafiltrate, and feces were collected. Pharmacokinetic analysis demonstrated high penetration of florfenicol within the gastrointestinal tract for both the high and low dose group (300%, 97%, respectively). There was no significant difference noted between dosing groups in proportion or persistence of phenotypically resistant bacterial isolates; however, the percent of resistant isolates was high throughout the study period. The recommendation for the use of a lower tier antimicrobial, such as florfenicol, may allow for the persistence of co-resistance for antibiotics of high regulatory concern.

Sterility and concentration of liposomal bupivacaine single-use vial when used in a multiple-dose manner.

OBJECTIVE: To evaluate the sterility of bupivacaine liposome injectable suspension (Nocita®) used in a multiple-dose fashion for 5 days. STUDY DESIGN: Triplicate liposomal bupivacaine vials were stored under two conditions, (1) room temperature (24°C) and (2) refrigerated temperature (5°C). A 3-mL aliquot was withdrawn from each vial daily. Samples were inoculated in tryptic soy broth in triplicate and then incubated for 24 hours at 37°C and subcultured every 48 hours onto blood agar and Sabouraud dextrose agar, respectively. Separate 1.5-mL aliquots of liposomal bupivacaine were centrifuged at 3500 g to separate liposome-encapsulated bupivacaine from the solution. Concentration of unencapsulated bupivacaine was analyzed via high-pressure liquid chromatography. Data were analyzed by using mixed effects procedure with multiple comparisons. SAMPLE POPULATION: Ten 20-mL vials of bupivacaine liposome injectable suspension stored under two conditions, (1) room temperature (24°C) and (2) refrigerated temperature (5°C). RESULTS: Five days of repeated withdrawal from the single-use vials yielded no bacterial growth. One control vial, which was opened and punctured once on the last day of the experiment, yielded fungal growth of an Aspergillus spp, likely an environmental contaminant. The concentration of free bupivacaine did not significantly differ until the fifth day of sampling. CONCLUSION: When aseptic technique was used, liposomal bupivacaine remained sterile for 5 days. Concentrations of free bupivacaine were unchanged from baseline for 4 days in both refrigerated and room temperature conditions. CLINICAL SIGNIFICANCE: Single-use liposomal bupivacaine vials can be used extralabel in a multiple-dose fashion for up to 4 days when stored either refrigerated or room temperature when sterile technique is used.

Efficacy of vaporized hydrogen peroxide for repeated sterilization of a single-use single-incision laparoscopic surgery port.

OBJECTIVE: To determine the ability of vaporized hydrogen peroxide (VHP) to sterilize a single-use single-incision laparoscopic surgery port and its associated components after repeated simulated uses. STUDY DESIGN: Prospective in vitro experimental study. SAMPLE POPULATION: Six single-use single-incision laparoscopic surgery ports with associated cannulas and insufflation tubing. METHODS: Ports, cannulas, and tubing were subjected to 10 cycles of simulated use, bacterial inoculation with Staphylococcus pseudintermedius and Escherichia coli, decontamination and sterilization, and testing via culture based on their treatment group designation. Bacteriological scores were compared among the negative control, positive control, and 4 treated ports and components. RESULTS: There was no difference in bacteriological scores between treated ports, cannulas, and insufflation tubing and the negative control port and components. Bacteriological scores of ports and components undergoing 6-10 cycles were not significantly different from scores of ports and components undergoing 5 or fewer sterilization cycles. No difference was found in detection of bacteria from treated ports by biopsy of the foam versus sampling via wash. CONCLUSION: This study suggests that a single-use single-incision laparoscopic port and its associated components can be effectively sterilized after multiple simulated uses by using VHP. CLINICAL SIGNIFICANCE: Reuse of a single-use single-incision laparoscopic port is a safe and effective method of cost reduction in veterinary patients.

Listeria monocytogenes septicemia in an immunocompromised dog.

An 11-year-old, male castrated, Boston Terrier was presented to the North Carolina State University College of Veterinary Medicine Small Animal Emergency Service with a 2-day history of progressive ataxia, left-sided head tilt, and anorexia. The dog had previously been diagnosed with chronic lymphoid leukemia and suspected immune-mediated destruction of his bone marrow precursor cells, possibly due to therapy with immunosuppressive dosages of prednisone and azathioprine. During the physical examination, abnormal findings included an increased body temperature and horizontal nystagmus. Diagnostic investigations included a computed tomography (CT) scan, which confirmed bilateral otitis media, and a blood culture, which was positive for Listeria monocytogenes serotype 4b (epidemic clone 1). Upon treatment with ampicillin/sulbactam, enrofloxacin, and minocycline, the dog became normothermic and the neurologic signs improved. L monocytogenes serotype 4b (epidemic clone 1) has been associated with outbreaks of human listeriosis originating from food contamination. Although rare case reports of Listeria spp. infection in dogs exist, an actual infection with the epidemic clone 1 strain has never before been reported in a dog. It should be included in the differential diagnoses in immunocompromised dogs with clinical signs of septicemia.

Implantation of an ultrafiltration device in the ileum and spiral colon of steers to continuously collect intestinal fluid.

Collection of fluid from the lumen of the gastrointestinal tract is commonly necessary for research projects, but presents challenges including intestinal motility and potential for leakage of intestinal contents. In this study, ultrafiltration collection devices were surgically implanted in the ileum and spiral colon of 12 steers for repeated collection of intestinal fluid over 48 hours. There were no significant complications associated with surgery or during the post-operative period, nor were there any significant pathologic changes found at necropsy 3 or 4 days post-surgery. Over 48 hours, we obtained 88% of the desired 212 samples. Only two devices failed to routinely collect samples. Use of ultrafiltration probes is a novel, consistent and humane method to repeatedly sample the gastrointestinal contents.