Veterinarians contacted to identify cats diagnosed with osteoarthritis (OA) provided information on signalment, method of diagnosis, treatment and concurrent disease. Owners of 50 cats were interviewed to collect information on specific OA signs observed in the home, relating to mobility, self-maintenance, social and exploratory behavior, and activity and habits at diagnosis and after treatment. Mean age at diagnosis was 12 y; concurrent diseases were common (44%). Owner-reported abnormalities led to OA diagnosis in most cases; either as the primary finding (30%), or combined with abnormal physical examination or radiographic findings (64%). Owners frequently reported changes in mobility, particularly gait, jumping, and use of stairs. Oral or injectable disease-modifying osteoarthritis drugs were the most common treatments (71%). Feline OA diagnosis and therapeutic monitoring appear to rely heavily on owner-perceived signs; physical examination abnormalities may not be detected. Questioning of owners revealed various observable signs potentially useful in OA detection and monitoring. Ultrasound-guided percutaneous antegrade pyelography with computed tomography for the diagnosis of spontaneous partial ureteral rupture in a dog Swan Specchi, Giuseppe Lacava, Marc-André d’Anjou, Eric Zini, Edoardo Auriemma Abstract — A 10-year-old spayed female dalmatian dog developed acute vomiting and abdominal pain. A 10-year-old spayed female dalmatian dog developed acute vomiting and abdominal pain. Ultrasound examination of the abdomen showed right hydronephrosis and proximal ureter dilation with mild retroperitoneal free fluid. Computed tomography (CT) of the abdomen confirmed the ultrasonographic findings and revealed, additionally, a right ureteral stone. Spontaneus rupture of the right ureter was confirmed with CT post ultrasound-guided percutaneous antegrade pyelography. Pyelo-ureteral rupture and the presence of a ureteral stone were confirmed at surgery. An unusual case of urinary incontinence in an intersex West Highland white terrier Neil A. Connery, Tim Spotswood Abstract — A 5-year-old neutered female West Highland white terrier dog was presented with a history of A 5-year-old neutered female West Highland white terrier dog was presented with a history of congenital urinary incontinence that had become refractory to medical management. Complex urogenital anomalies including urethrovestibular and vestibuloperineal fistulae with low vulvar position along with a penoclitoris were present. Vaginectomy with perineal urethral reconstruction resolved the incontinence. Putative peanut allergy-induced urticaria in a dog Min-Hee Kang, Hee-Myung Park A 9-year-old, spayed male schnauzer dog was presented with vomiting, diarrhea, generalized erythema, pruritic urticaria and conjunctival hyperemia after ingestion of peanut. The history, clinical signs, and histopathology of the lesions were compatible with a hypersensitivity reaction. The clinical signs resolved rapidly after treatment with prednisolone and antihistamine. This is the first report of urticaria caused by peanut ingestion in a dog. Necrotizing fasciitis caused by methicillin-resistant Staphylococcus pseudintermedius at a previously irradiated site in a dog Monique N. Mayer, Joseph E. Rubin Abstract — A great Dane dog was presented with a small, superficial wound on the left tarsus that rapidly A great Dane dog was presented with a small, superficial wound on the left tarsus that rapidly progressed to a large necrotic area. The dog had undergone radiation therapy in the left tarsal region 33 months previously. Necrotizing fasciitis was diagnosed on histopathological examination, and bacterial culture revealed methicillin-resistant Staphylococcus pseudintermedius.

To determine the prevalence of plasmid-mediated quinolone resistance (PMQR) determinants and investigate mutations in gyrase and topoisomerase genes that may contribute to increased fluoroquinolone resistance in canine and feline Escherichia coli isolates in the USA that displayed reduced susceptibility to extended-spectrum cephalosporins. This study was undertaken because previous epidemiological studies identified a potential correlation between extended-spectrum cephalosporins and fluoroquinolone resistance.Isolates (n54) with reduced susceptibility to ceftazidime or cefotaxime were screened by PCR for the presence of PMQR determinants and gyrase and topoisomerase genes were sequenced. Isolates were further characterized by conjugation and phylogenetic analyses.PMQR determinants aac(6)-Ib-cr, qnrS and qepA were identified in 30, 23 and 5 isolates, respectively. Multiple mutations were identified in the quinolone resistance-determining region, including the novel substitutions of Glu-84Ala and Leu-88Gln in ParC and Arg-432Ser and Glu-460Val in ParE. The isolate that exhibited the highest level of enrofloxacin resistance (MIC 256 mg/L) had a double mutation in gyrA (Ser-83Leu and Asp-87Asn) and a triple mutation in parC (Ser-80Ile, Glu-84Gly and a novel mutation, Leu-88Gln). The presence of PMQR genes increased the ciprofloxacin MIC values 4-fold to 8-fold in transconjugants relative to the recipient strain. Approximately 39 of the isolates belonged to phylogenetic group D and 30 to group B2, which typically contain an increased number of virulence determinants compared with other groups.Novel mutations in topoisomerase genes and PMQR determinants aac(6)-Ib-cr, qnrS and qepA genes were detected among extended-spectrum -lactamase-producing E. coli in the USA.

The rapid spread of extended-spectrum β-lactamases (ESBLs) and quinolone resistance in Escherichia coli in companion animals (1-2) has increased concern among veterinarians9 to find an alternative therapy to treat clinical infections caused by these resistant organisms.…

In the last decade, antimicrobial resistance has been widespread in several bacterial species. This increase in resistance could be associated with an increase in the use of different antimicrobials to treat infections caused by pathogenic bacteria. While resistance to antimicrobials is often attributed to known mechanisms, other mechanisms are still under investigation for many bacterial species. Detection of antimicrobial resistance often involves conventional agar, broth or disk diffusion assays. However, these methods can be cumbersome and time consuming compared to molecular methods. Consequently, several polymerase chain reaction (PCR) techniques have been developed to expedite the detection of antimicrobial resistance in bacterial pathogens. PCR-based technologies are rapid, sensitive and specific for detecting antimicrobial resistance. Application of such technologies in diagnostic laboratories can provide insight into emerging mechanisms of antimicrobial resistance in veterinary pathogens. In this chapter, we describe molecular mechanisms of drug resistance in microbial pathogens and the potential advantages and disadvantages of PCR-based methods.

Percent resistance and minimum inhibitory concentrations (MIC) were described for canine (n = 301) and feline (n = 75) pathogenic Escherichia coli (E. coli) isolates solicited during May 2005 to Sep 2005 from the Clinical Pharmacology Laboratory at Auburn University (n = 165) or commercial diagnostic laboratories ([CDL]; n = 211) from four regions in the USA. Drugs tested were amoxicillin (AMX), amoxicillin trihydrate/clavulanate potassium (AMXC), cefpodoxime (CFP), doxycycline (DXY), enrofloxacin (ENR), gentamicin (GM) and trimethoprim-sulfamethoxazole (TMS). Urinary isolates were most common (n = 174). Percent resistance was greatest for isolates from the respiratory tract, urine, and skin compared with the ear. Resistance was also greatest for samples sent from the south and central states compared with the western states (P ≤ 0.001). Percent resistance by drug was AMX (46 ± 2.6%) > AMXC (37 ± 2.5%) > CFP (21.8 ± 2%) = DXY (22 ± 2.1%) = ENR (20 ± 2.1%) = TMS (19 ± 2%) > GM (12 ± 1.7%). There was a significant difference in resistance between the different antibiotic drugs (P ≤ 0.001). Population MIC distributions were bimodal, and MICs were highest in samples from the southern states (P ≤ 0.001). E. coli resistance may limit its empirical treatment. For susceptible isolates, AMX and AMXC may be least effective and TMS most effective.

Since the early 1990s, there has been a substantial increase in the emergence and dissemination of antimicrobial resistance in foodborne and zoonotic pathogens, such as Campylobacter, Salmonella and Escherichia coli causing human infections in several developed countries. This increase has been attributed to the potential use of antimicrobials for prophylactic treatment of food animals and humans. While these drugs were successful in the treatment of most diseases in animals and humans, they have contributed to the emergence of drug-resistant bacteria. Several studies have addressed the public health significance of antimicrobial resistance in foodborne pathogens. Attention has been particularly focused on the pros and cons of short term and long term use of antimicrobial agents in food of animals for growth promotion and disease prevention. The public health risk of emergence, spread and transmission of drug-resistant foodborne pathogens in the farm-to-the-fork continuum warrants appropriate actions by both the scientific community and the regulatory agencies in advocating restrictions on the approval and use of new and existing drugs. In this chapter, we have described the prevalence, specific mechanisms (based on the class of drugs) and dissemination of antimicrobial resistance in Campylobacter, Salmonella and E. coli. These three pathogens were selected because they constitute the bulk of bacterial illnesses worldwide. The review will highlight the importance of addressing the issue of conventional and emerging drug-resistant foodborne bacteria in foods, animals and humans so that steps can be taken to minimize their spread in the environment.

ABSTRACT Resistance to extended-spectrum cephalosporins (ESC) among members of the family Enterobacteriaceae occurs worldwide; however, little is known about ESC resistance in Escherichia coli strains from companion animals. Clinical isolates of E. coli were collected from veterinary diagnostic laboratories throughout the United States from 2008 to 2009. E. coli isolates (n = 54) with reduced susceptibility to ceftazidime or cefotaxime (MIC ≥ 16 μg/ml) and extended-spectrum-β-lactamase (ESBL) phenotypes were analyzed. PCR and sequencing were used to detect mutations in ESBL-encoding genes and the regulatory region of the chromosomal gene ampC. Conjugation experiments and plasmid identification were conducted to examine the transferability of resistance to ESCs. All isolates carried the blaCTX-M-1-group β-lactamase genes in addition to one or more of the following β-lactamase genes: blaTEM, blaSHV-3, blaCMY-2, blaCTX-M-14-like, and blaOXA-1. Different blaTEM sequence variants were detected in some isolates (n = 40). Three isolates harbored a blaTEM-181 gene with a novel mutation resulting in an Ala184Val substitution. Approximately 78% of the isolates had mutations in promoter/attenuator regions of the chromosomal gene ampC, one of which was a novel insertion of adenine between bases −28 and −29. Plasmids ranging in size from 11 to 233 kbp were detected in the isolates, with a common plasmid size of 93 kbp identified in 60% of isolates. Plasmid-mediated transfer of β-lactamase genes increased the MICs (≥16-fold) of ESCs for transconjugants. Replicon typing among isolates revealed the predominance of IncI and IncFIA plasmids, followed by IncFIB plasmids. This study shows the emergence of conjugative plasmid-borne ESBLs among E. coli strains from companion animals in the United States, which may compromise the effective therapeutic use of ESCs in veterinary medicine.

OBJECTIVE To investigate the contribution of gyrA mutation and efflux pumps to fluoroquinolone resistance and multidrug resistance among Escherichia coli isolates from dogs and cats. SAMPLE POPULATION 536 clinical isolates of E coli. PROCEDURES Minimum inhibitory concentrations (MICs) were determined for enrofloxacin and 6 other drug classes by use of broth microdilution techniques. Real-time PCR assay was used to determine the mutation in gyrA; Phe-Arg-β-naphthylamide, an efflux pump inhibitor, was used to examine the contribution of efflux pump overexpression. RESULTS The MIC for fluoroquinolones increased in a stepwise fashion and was lowest in the absence of mutations, higher with a single point mutation, and highest with 2 point mutations. Level of resistance in the latter category was high (8 times the breakpoint), but this was associated with expression of the AcrAB efflux pump. Inhibition of the efflux pump resulted in a reduction in the MIC to less than the susceptible breakpoint for isolates with an MIC ≤ 4 mg/L, regardless of the presence of a mutation. The greatest magnitude in MIC decrease (MIC was decreased by a factor of > 67 fold) was for isolates with a single mutation but the greatest absolute decrease in MIC (124 mg/L) was for isolates with 2 mutations. Inhibition of the AcrAB efflux pump in isolates characterized by multidrug resistance decreased the MIC of drugs structurally unrelated to fluoroquinolone. CONCLUSIONS AND CLINICAL RELEVANCE Fluoroquinolone resistance in E coli appeared to be a stepwise phenomenon, with MIC increasing as the number of point mutations in gyrA increased, but high-level resistance and multidrug resistance associated with fluoroquinolone resistance reflected overexpression of the AcrAB efflux pump.

Integrons are mobile genetic elements that incorporate an open reading frame or gene cassettes. They have an important role in the acquisition and dissemination of antimicrobial resistance genes. Yet the occurrence of integrons carrying antimicrobial resistance genes in bacterial pathogens of pet animals is seldom addressed. The purpose of this study was to describe the incidence of class 1 and 2 integrons in clinical isolates of Escherichia coli (n = 82) from cats and dogs provided by diagnostic laboratories in five States of the USA. An association between resistance genes in the integrons and the isolates’ phenotypes was found. Integrons were detected using PCR and then further characterized by restriction fragment-length polymorphism analysis and amplicon sequencing. Class 1 integrons were detected in 27% of the isolates, while only 2% (n = 2) of the isolates were positive for the presence of class 2 integrons. Seventy-two percent (n = 59) of the isolates did not carry integrons. Eleven gene cassettes were found either alone or in combination with other gene cassettes, which encoded resistance to aminoglycosides (aadA1, aadA2, aadA5, aacA4, and aadB), trimethoprim (dfrA1, dhfrA17, and dfrA12), chloramphenicol (catB3 and cmlA6), and streptothricin (sat1), respectively. All integron-positive isolates were characterized by resistance to least two drug classes and 35% produced extended-spectrum β-lactamases. The association of integrons carried on plasmids and antimicrobial resistance was confirmed by curing experiments for three isolates. Resistance was resolved once large plasmids (size range 97–169 kb) carrying the class 1 integron were lost. Therefore, integrons appear to have an essential role in facilitating the dissemination of the resistance genes and contributing to the creation of multi-drug resistant phenotypes.

Fluoroquinolones are used to treat infections caused by Escherichia coli in canine and feline veterinary patients, particularly those infecting the urinary tract. The gyrA gene is a primary target causing fluoroquinolone resistance in gram negative coliforms, with mutations in codons 83 and 87 generally associated with high-level of resistance E. coli clinical isolates. We have developed a fluorescence resonance energy transfer (FRET) quantitative PCR to identify enrofloxacin-resistance in clinical E. coli isolates that carry mutations in codons 83 and 87 of gyrA. This real-time quantitative PCR assay is rapid, economical, and sensitive compared with cultured antimicrobial susceptibility testing. The assay identified as few as four genome copies per reaction from culture and 19 genome copies in urine. For the 70 isolates tested, the sensitivity was 87.5% (95% CI=75-95.3%) (n=42/48), specificity was 100% (95% CI=87.3-100%) (n=22/22), whereas accuracy was 91.4% (95% CI=82.3-97%) (n=64/70). Furthermore, we were able to accurately differentiate between the wild type and mutants E. coli directly from infected canine urine samples (n=5) within 2 h. These results were confirmed by sequence alignments of the PCR products and comparison with the susceptibility testing. The FRET-PCR assay appears to have promising clinical application as an early diagnostic tool for rapid and sensitive detection and differentiation of the level of fluoroquinolone resistance among clinical E. coli isolates that may facilitate design of the dosing regimen.

Background: Antimicrobial resistance is emerging among canine and feline clinical E. coli isolates. Objectives: To generate baseline data on the prevalence and molecular basis of antimicrobial resistance in the canine and feline clinical Escherichia coli isolates. Animals: E. coli pathogens isolated from dogs and cats. Methods: A total of 377 E. coli isolates were collected from dogs or cats with spontaneous infection between May and September 2005. Isolates were phenotypically characterized based on the susceptibility to seven antimicrobial agents by E-test® according to CLSI guidelines. Selected isolates from each phenotype were genotyped using pulsed-field gel electrophoresis (PFGE). Results: Of the 377 isolates, 43 phenotypes were generated. Almost 44% (n= 85/194) of the resistance isolates expressed single drug resistance, with resistance to beta-lactams predominating (83%; n=71/85). Only two isolates expressed single drug resistance to

ABSTRACT The survival of Campylobacter jejuni and Campylobacter coli at pH 7.0, 6.0, 5.0 and 4.0 for up to 24 h, and the induction of an acid adaptation response in 12 C. jejuni and 10 C. coli strains in early exponential or late stationary phases in tryptic soy broth (TSB) and Brucella broth were investigated. C. coli strains were more sensitive than C. jejuni in media adjusted to pH 5.0 or 4.0 with hydrochloric acid. Five log10 cfu/mL of C. coli were inhibited at 12 h in pH 5.0, but only 2.5 log10 cfu/mL of C. jejuni cells were inhibited under the same conditions. No viable cells were detected at 4 h in pH 4.0 from an inoculum of 6 log10 cfu/mL of C. coli. Late stationary phase cells of C. jejuni exposed to pH 5.0 for 4 h in TSB exhibited a significant (P ≤ 0.05) acid tolerance response when compared to nonexposed cells. Late stationary phase cells of C. coli exposed to pH 5.0 for 3 h in TSB and late stationary and early exponential phase cells exposed to pH 5.0 for 3 h in Brucella broth showed a significant (P ≤ 0.05) acid tolerance response when compared to nonexposed cells. The acid tolerance responses observed for C. jejuni and C. coli conferred adapted cells no more than 1 log10 cfu/mL survival advantage over nonadapted cells. Brucella broth appeared to be more protective than TSB for the survival of C. jejuni and C. coli at pH 4.0.