Mechanisms of fluoroquinolone resistance in Pseudomonas aeruginosa isolates from canine infections

Chronic otitis externa in dogs is often associated with Pseudomonas aeruginosa infection. Fluoroquinolones are often used for treating such infections. Fluoroquinolone resistance mechanisms were characterized in 10 strains of P. aeruginosa isolated from chronic canine otitis externa. Nine out of ten strains harbored a mutation in the gyrA gene and presented an overexpression of efflux pump(s). There was a good correlation between the lipophilicity of the fluoroquinolone being tested and the effect of the efflux pump inhibitor in the final MIC. Therefore, both mechanisms, mutation in the gyrA gene and increased efflux pump(s), seem to play an important role in the acquisition of fluoroquinolone resistance in veterinary clinical isolates of P. aeruginosa. Levels of resistance to fluoroquinolones suggest that they could not be a good choice for systemic therapy of Pseudomonas otitis.     

Antimicrobial effect of combinations of EDTA-Tris and amikacin or neomycin on the microorganisms associated with otitis externa in dogs

Combinations of EDTA-Tris and two aminoglycoside antibiotics (amikacin and neomycin) were tested for synergistic activities against the microorganisms associated with otitis externa in dogs and for the solutions' stability over time. Synergistic activity was observed when EDTA-Tris plus amikacin and EDTA-Tris plus neomycin were tested againstStaphylococcus intermedius, Proteus mirabilis, Pseudomonas aeruginosa, andEscherichia coli, but not againstCandida albicans. Stability studies over a 3-month period indicated that the test solutions were stable at room temperature and that their antimicrobial activity was maintained.Abbreviations CFU colony-forming units - EDTA (ethylenedinitrilo)tetraacetic acid, disodium salt - FIC fractional inhibitory concentration - MBC minimal bactericidal concentration - MIC minimal inhibitory concentration - Tris tris(hydroxymethyl)aminomethane     

Uptake of antibodies in gram-negative bacteria exposed to EDTA-Tris

Ethylene diamine tetra-acetic acid disodium salt (3.22 mM) and Tris (hydroxymethyl aminomethane) (50 mM) was found to cause potentiation of antibiotic actions and/or increased uptake of antibiotics when assessed by bacterial inhibition kinetic studies and uptake of radiolabeled antibiotics. Solutions of EDTA-Tris potentiate the effect of tetracycline, dihydrostreptomycin and penicillin when reacted with Escherichia coli. A similar effect was observed for tetracycline when incubated with Proteus vulgaris and penicillin, chloramphenicol, dihydrostreptomycin and tetracycline when incubated with Pseudomonas aeruginosa. These effects were noted by decreasing bacterial counts in the bacterial inhibition kinetic studies and increased uptake of radiolabeled antibiotics.     

Potentiating effect of EDTA-Tris on the activity of antibiotics against resistant bacteria associated with otitis, dermatitis and cystitis

Possible synergistic effects of the combination of EDTA-tromethamine (EDTA-Tris) and three antimicrobial agents (cephaloridine, kanendomycin and enrofloxacin) against resistant Gram-positive and Gram-negative bacteria are reported. Bacteria were isolated from eight cases of chronic otitis externa, five cases of chronic dermatitis and four cases of recurrent cystitis in dogs which had previously been treated with one of the three antibiotics without success. Animals exposed to EDTA-tromethamine plus the antibiotic recovered completely within 10 days, and were controlled clinically and bacteriologically for 180 days. Local irrigation with EDTA-tromethamine solution was well tolerated and no side effects were recorded.     

Development of resistant bacteria isolated from dogs with otitis externa or urinary tract infections after exposure to enrofloxacin in vitro

Minimum inhibitory concentrations for enrofloxacin were determined for 63 bacterial isolates from dogs with otitis externa or urinary tract infections. Development of resistant mutants was determined after exposing the isolates to enrofloxacin in vitro for up to five serial passages. Results indicated that Pseudomonas aeruginosa and Enterococcus spp isolates exposed to enrofloxacin developed resistance rapidly, whereas Klebsiella, Proteus, and Streptococcus spp were less likely to develop resistance. Despite the presence of enrofloxacin pressure, no resistant bacteria developed in the Escherichia coli and staphylococcal isolates. In many isolates, susceptibility patterns changed from susceptible to intermediate.     

Antibacterial susceptibility patterns of Pseudomonas strains isolated from chronic canine otitis externa

Development of antibiotic resistance in bacteria is a problem of great concern. It is important to establish the convenience of antimicrobial susceptibility tests in animal infections. The aim of this study was to test the susceptibility to antibiotics of Pseudomonas strains isolated from chronic canine otitis externa. We tested 23 strains of Pseudomonas: 19 Ps. aeruginosa, three Ps. fluorescens and one Pseudomonas spp. The most effective antibiotics were tobramycin (100% susceptible), marbofloxacin (91.3%) and ceftazidime (91.3%). Ticarcillin and gentamicin, commonly used for the treatment of otitis externa also showed good results (susceptibility of strains was 86 and 65.2% respectively). Lower susceptibility was found using enrofloxacin (52.1%) probably due to its indiscriminate use. We emphasize the need for a rational policy of antibiotic prescribing in order to prevent the selection of resistant strains.     

The role of Pityrosporum pachydermatis in otitis externa of dogs: evaluation of a treatment with miconazole.

The bacterial and mycotic flora were assessed in 158 ears of dogs with otitis externa and in 101 ears of healthy control dogs. Pityrosporum pachydermatis occurred in 57 per cent of ears with otitis externa and in 17 per cent of clinically healthy ears. Staphylococci and Pseudomonas aeruginosa were the predominant bacteria in otitic ears, micrococci and Bacillus spp were the most frequent isolates from clinically healthy ears. P pachydermatis, Ps aeruginosa and Candida tropicalis occurred in monoculture in a significant number of mainly chronic cases of otitis externa. A combination preparation, containing miconazole, polymyzin B and prednisolone, was highly effective in controlling the clinical signs of otitis externa and eliminating flora from the affected ears. The data presented suggest that yeasts, and especially P pachydermatis, may be significant pathogens in otitis externa and that antimycotic treatment is an essential part of the treatment of otitis externa in dogs.     

Antimicrobial susceptibility of Pseudomonas aeruginosa isolates from dogs with otitis externa

Pseudomonas aeruginosa is a common cause of otitis externa in dogs, and treatment of these infections is becoming problematic because of the increasing number of multiresistant strains. The aim of the present study was to compare the in vitro activities of cefepime, ceftazidime, enrofloxacin, ciprofloxacin, gentamicin and ticarcillin/clavulanic acid against 104 strains of P aeruginosa isolated from dogs with otitis externa. Antimicrobial susceptibility and minimum inhibitory concentrations, in μg/ml, were evaluated by the E test (bioMérieux). The most active compound was ceftazidime, with 100 per cent efficiency. The majority of tested strains were susceptible to ticarcillin/clavulanic acid (89.4 per cent), followed by ciprofloxacin (88.5 per cent) and cefepime (60.6 per cent). The highest resistance was observed to enrofloxacin (51.9 per cent) and gentamicin (43.3 per cent). Large numbers of strains were intermediately susceptible to antibiotics registered for use in veterinary medicine in Croatia--enrofloxacin (47.1 per cent) and gentamicin (41.3 per cent).     

Minimum inhibitory concentrations of some antimicrobial drugs against bacteria causing uterine infections in cattle

The minimum inhibitory concentrations (MICs) of oxytetracycline, cephapirin, cephapirin/mecillinam, cefquinome, ceftiofur and enrofloxacin, candidate antibiotics for the principal bacteria associated with uterine infections: Escherichia coli, Arcanobacterium pyogenes and the anaerobic bacteria Fusobacterium necrophorum and Prevotella melaninogenicus, were determined by the agar dilution method. The bacteria were isolated from animals with clinical metritis and/or endometritis. For E coli, cefquinome and enrofloxacin had the lowest MIC90 and MIC50 values (< 0.06 microg/ml), and oxytetracycline and cephapirin had the highest values. For A pyogenes, oxytetracycline had the highest MIC50 value (16 microg/ml), but all the cephalosporins had values below 0.06 microg/ml. For the anaerobic bacteria, enrofloxacin and oxytetracycline had the highest MIC50 values but all the cephalosporins had values of 0.06 microg/ml or below.     

Update on antimicrobial susceptibilities of bacterial isolates from canine and feline otitis externa

The in vitro susceptibility of a total of 1819 bacterial isolates from canine and 103 isolates from feline otitis externa cases to 13 antimicrobial drugs over a 5-year period was evaluated. Among topically used drugs, 90% of isolates were susceptible to gentamicin. Susceptibility of Pseudomonas aeruginosa to gentamicin was 85%, and to polymyxin B 100%. For isolates other than the Pseudomonas sp., susceptibility was highest to amoxicillin-clavulanic acid.     

Purulent rhinitis and otitis caused by Pseudomonas aeruginosa in sheep showered with contaminated 'shower wash'

Three crossbred lowland ewes developed a severe purulent rhinitis and another three ewes developed a severe purulent otitis externa/media after being showered with a wash that had been used 24 to 48 hours before on a separate group of Cheviot ewes with lesions of dermatitis. Pseudomonas aeruginosa was isolated in pure growth from the aural and nasal abscesses and also from the dermatitis lesions. Extended antibiotic susceptibility testing and the random amplification of polymorphic DNA indicated that a single clonal type was associated with the rhinitis and otitis and with the dermatitis, providing strong evidence of an epidemiological link between the lesions of dermatitis and the aural and nasal abscesses through the use of the contaminated 'shower wash'.     

Evaluation of a method using Proteus mirabilis and Pseudomonas aeruginosa to experimentally induce dual infection of the urinary bladder in dogs

OBJECTIVE: To evaluate a method using Proteus mirabilis and Pseudomonas aeruginosa to experimentally induce dual infection of the urinary bladder in dogs. ANIMALS: 6 healthy mixed-breed dogs. PROCEDURE: Dogs were anesthetized, and cystitis was induced by infusing a solution of salicylic acid in ethanol into the bladder, followed by an inoculum containing field isolates of P. mirabilis and P. aeruginosa. Dogs were examined daily for 21 days after induction of cystitis. On day 21, dogs were euthanatized, and urinary bladder, renal pelvis, and prostate specimens were submitted for bacterial culture. RESULTS: After induction of cystitis, all dogs had evidence of thickening of the bladder wall, dysuria, tenesmus, and hematuria. Urinalysis revealed proteinuria, hematuria, and pyuria. All urine samples obtained on day 21 yielded growth of P. mirabilis, but P. aeruginosa was not cultured from any of these samples. Proteus mirabilis was isolated from bladder, renal pelvis, or prostate specimens from 4 dogs; P. aeruginosa was not isolated from any of the tissue specimens. CONCLUSION: Results suggest that the method used in the present study fails to induce dual infection of the urinary bladder with P. mirabilis and P. aeruginosa. The inability to establish a persistent dual infection with this method may have been a result of insufficient pathogenicity of the Pseudomonas isolate or an inadequacy of the experimental design.     

Bacteriology and mycology of otitis externa in dogs]

The bacterial and fungal flora of 1118 ears of dogs with otitis externa and 100 ears of healthy control dogs were studied in order to isolate the causative agents. The yeast Malassezia pachydermatis (56%) was by far the most common organism in otitic dogs followed by the bacteria Staphylococcus intermedius (23%), Pseudomonas aeruginosa (12%), Proteus spp. (6%) and Streptococcus canis (5%). A statistical analysis of observed results showed that the incidence of these organisms is significant in otitic dogs. Many strains of S.intermedius, P.aeruginosa and Proteus spp. are resistant to antimicrobial agents commonly used to treat otitis externa. Therefore an antimicrobial susceptibility testing was performed using "Cobas Bact" for these bacterias. Furthermore, 80 strains of M.pachydermatis were submitted to identification-kits (API 20 CAUX, API STAPH, Cobas Micro). The observed results showed that an identification with these tests was not possible.     

Variability of laboratory identification and antibiotic susceptibility reporting of Pseudomonas spp. isolates from dogs with chronic otitis externa

The purpose of this study was to evaluate interlaboratory variation in isolation and antibiotic susceptibility pattern of Pseudomonas spp. as reported to veterinarians for cases of canine chronic bacterial otitis externa. Twenty-six dogs with unilateral or bilateral bacterial otitis externa from multiple referral practices were included in this prospective study. Triplicate samples collected simultaneously from the same location in the external ear canal were randomly submitted to three laboratories for culture and susceptibility testing. Pseudomonas spp. were isolated from 18 of 34 (53%) ears. All three laboratories agreed on the presence of Pseudomonas spp. in 15 (83.3%) ears sampled. However, two laboratories agreed on two (11.1%) occasions, and on one occasion (5.5%) Pseudomonas spp. were identified in only one laboratory. Minimum inhibitory concentration (MIC) susceptibilities to 11 antibiotics were compared between laboratories B and C. Using laboratory-defined susceptibility of sensitive (S), intermediate (I) and resistant (R), none of the 16 Pseudomonas spp. with MIC data reported had identical patterns of antibiotic susceptibility. Agreement in susceptibility to individual antibiotics was observed in 13 of 16 (81%) occasions for amikacin and gentamicin, 10 of 16 (63%) occasions for ticarcillin, and nine of 16 (56%) for enrofloxacin. These results indicate that Pseudomonas spp. were identified by all three laboratories chosen for this study in 83% of the time. Moreover, antibiotic susceptibility patterns and MIC values reported to veterinarians may not agree between laboratories. Veterinarians should interpret bacterial culture and susceptibility results with multiple caveats including variability between laboratories.     

The microbial environment of the ear canal in health and disease

Otitis externa is an important disease of dogs and, to a lesser degree, cats. The yeast Malassezia canis is the most common organism demonstrated in ear infections. Coagulase-positive staphylococci are the most common bacteria isolated, often occurring with M. canis. Pseudomonas aeruginosa and Proteus are gram-negative bacteria that are frequently encountered. Other bacteria, such as beta-hemolytic streptococci, enterococci, Escherichia coli, and Corynebacterium are also important. Malassezia canis, coagulase-positive staphylococci, and Pseudomonas aeruginosa commonly occur as monoinfections, whereas other bacteria are typically associated with mixed infections. Smears may provide rapid diagnosis on which to base empiric treatment. Cultures should be considered in recurrent or refractory cases, especially those involving gram-negative bacteria. In practices that are not equipped to perform culture and susceptibility tests, it is advisable to seek the advice of a microbiology laboratory.     

Identification and antimicrobial susceptibility patterns of bacteria causing otitis externa in dogs

Bacterial agents are considered important pathogens causing external otitis in dogs. It is essential to carry out bacterial culture and antimicrobial susceptibility test in the case of otitis externa, particularly for chronic or recurring cases. Sterile swab samples were obtained from terminal part of vertical ear canals of 74 dogs with otitis externa for cytology, bacterial culture and antimicrobial susceptibility test. Cytologic smears were stained using Gram and Giemsa staining methods. Aerobic bacterial culture performed on blood agar and MacConkey agar. Among total number of 92 isolated bacteria, 68 were Staphylococcus intermedius. Other isolated bacteria included: Pseudomonas aeruginosa, Proteus mirabilis, Escherichia coli, Pasteurella canis, and six other species of coagulase-negative Staphylococcus. Antimicrobial susceptibility test were performed for all isolated bacteria using 14 antibiotics. Based on the results of this study, all isolated Staphylococcus spp. were sensitive to amikacin, enrofloxacin, and rifampin, and had low resistance to gentamicin, cephalothin and ceftriaxone. More than half of gram-positive isolates were resistant to penicillin and ampicillin. Generally, all isolated gram-negative bacteria, were sensitive to amikacin and enrofloxacin, and had low resistance to ceftriaxone and gentamicin. They were highly resistant to penicillin, eythromycin, and cephalothin. Regarding the results of this study, in cases of uncomplicated otitis externa, it is possible to select antimicrobial drugs merely based on cytology, but it is recommended to perform bacterial culture and antimicrobial susceptibility test. However, in complicated or refractory cases, antimicrobials should be selected based on bacterial culture and antimicrobial susceptibility test.     

Antibacterial effect of bovine lactoferrin against udder pathogens

The antibacterial effect of lactoferrin (Lf) was tested on isolates of Escherichia coli (E. coli), Staphylococcus aureus (S. aureus), and coagulase-negative staphylococci (CNS) as well as on Pseudomonas aeruginosa (P. aeruginosa) and Klebsiella pneumoniae (K. pneumoniae), originally isolated from bovine mastitis. Concentrations of Lf used were 0.67 mg/ml, 1.67 mg/ml, and 2.67 mg/ml. Growth of udder pathogens was monitored by turbidometry either in broth culture or in whey prepared from normal milk. We focused on 3 different growth variables: lag time, slope, and maximum absorbance of bacterial growth curves. Growth inhibition was seen in the broth but hardly at all in whey. The isolates of E. coli and CNS did not grow sufficiently well in whey to draw any conclusions. The most effective inhibitory activity of Lf was seen against E. coli and P. aeruginosa. All 5 E. coil isolates had similar growth patterns. Inhibition of growth by Lf was concentration-dependent. The concentration of 0.67 mg/ml in broth and whey was generally too low for a significant inhibitory effect.     

Effects of beta-thujaplicin on anti-Malassezia pachydermatis remedy for canine otitis externa

The antifungal activity of beta-thujaplicin was evaluated against 51 Malassezia pachydermatis strains isolated from canine ear canals with or without otitis externa. For comparison, sensitivity tests were performed on M. pachydermatis isolates for nystatin, ketoconazole, and terbinafine HCl, all clinically available antifungal agents. The minimal inhibition concentrations over 50% of the tested isolates (MIC50) were 3.13 microg/ml for beta-thujaplicin and nystatin, 0.016 microg/ml for ketoconazole, and 1.56 microg/ml for terbinafine HCl. The antifungal effect for M. pachydermatis of beta-thujaplicin compared favorably with commercial antifungal agents. None of the 51 M. pachydermatis isolates showed resistance against any of the tested antibiotics investigated in this study. Ten representative isolates of M. pachydermatis were subcultured for 30 generations at concentrations close to the MIC levels of beta-thujaplicin, nystatin, ketoconazole, and terbinafine HCl, and examined to determine whether they had acquired resistance to each drug. As a result, M. pachydermatis was found to achieve resistance more easily for ketoconazole and terbinafine HCl than for beta-thujaplicin or nystatin. The MIC50 of beta-thujaplicin did not change during the course of subculture, and it is thought that the potential development of a resistant strain is low, even with continuous infusion for otitis externa therapy. beta-Thujaplicin is an inexpensive and safe treatment with anti-inflammatory and deodorant effects that can be recommended as an effective remedy for canine otitis externa.     

EDTA-Tris对不同消毒药物消毒效能的影响

当EDTA-Tris与不同消毒药物联合使用时,两者之间可能表现出增强、协同或拮抗作用。试验表明,当EDTA-Tris与新洁尔灭或甲醛联合应用时,其对试验菌的杀菌效能均表现出协同或增强作用,而与戊二醛、聚维酮碘或十二烷基硫酸钠联合应用时,其对不同菌株的杀菌作用则表现不一,与聚维酮碘联合应用时,对大肠杆菌的抑杀效能表现为拮抗作用,与十二烷基硫酸钠联合应用时,对链球菌的抑杀效能则表现为拮抗作用,而与戊二醛联合应用时,则对链球菌和大肠杆菌的抑杀效能均表现出拮抗作用。     

苦豆子提取物对多重耐药大肠杆菌的体外抑菌作用

为探究苦豆子提取物对多重耐药大肠杆菌的体外抑菌作用,本研究以从蛋鸡粪便样本中分离的1株多重耐药大肠杆菌为研究对象,测定了苦豆子提取物的最小抑菌浓度(MIC)以及与抗菌药物联用的抑菌效果,并测定了对多重耐药大肠杆菌生长曲线、生物被膜以及运动能力的影响。结果表明,苦豆子提取物MIC为125 mg/mL,与头孢噻呋联用呈现协同作用;与对照组相比,1/2MIC和MIC的苦豆子提取物可明显抑制菌株的生长;不同浓度苦豆子提取物对菌株生物被膜具有一定程度的抑制和清除作用,1/2MIC和MIC组可极显著降低菌株的运动能力(P<0.01)。表明苦豆子提取物对多重耐药大肠杆菌具有良好的体外抑菌作用。