Antimicrobial resistance in Escherichia coli isolates from Japanese raccoon dogs (Nyctereutes viverrinus) in Kanagawa Prefecture, Japan: Emergence of extended-spectrum cephalosporin-resistant human-related clones.

Introduction. Wild animals are one of the putative reservoirs of antimicrobial-resistant bacteria, but the significance of raccoon dogs remains to be investigated.Hypothesis. Raccoon dogs can be a reservoir of antimicrobial-resistant bacteria.Aim. This study aimed to explore the prevalence of antimicrobial resistance, mainly extended-spectrum cephalosporins resistance, in Escherichia coli isolates from faeces of 80 Japanese raccoon dogs in Kanagawa Prefecture, Japan.Methodology. All of the 80 faecal samples were streaked onto deoxycholate-hydrogen sulfate-lactose (DHL) and cefotaxime (CTX)-supplemented DHL (DHL-CTX) agars. Susceptibilities to ten antimicrobials were determined using the agar dilution method. Additionally, extended-spectrum ß-lactamases (ESBLs) and AmpC-type ß-lactamases (ABLs) were identified in addition to sequence types (STs), in ESC-resistant isolates by a polymerase chain reaction and sequencing.Results. Out of all the samples, 75 (93.8 %) and 20 (25.0 %) E. coli isolates were isolated by DHL and DHL-CTX agars, respectively. Significantly higher resistance rates to most of the drugs were found in DHL-CTX-derived isolates than DHL-derived isolates (P<0.01). Genetic analysis identified CTX-M-14 (n=6), CTX-M-2 (n=2), CTX-M-1 (n=1) and CTX-M-55 (n=1) as ESBLs, and CMY-2 (n=8) and DHA-1 (n=1) as ABLs in 20 DHL-CTX-derived isolates. Most of the detected STs were related to Japanese humans (i.e. ST10, ST58, ST69, ST131, ST357, ST648 and ST4038). Notably, this is the first report on ST69, ST131, ST155 and ST648, which are well-known international high-risk clones in Japanese raccoon dogs.Conclusion. Our findings underscore the need to understand the significance of raccoon dogs as an antimicrobial-resistant bacteria reservoir using one health approach.

Antimicrobial resistance and multilocus sequence types of Stenotrophomonas maltophilia isolated from dogs and cats in Japan.

As the representative multidrug-resistant pathogen, Stenotrophomonas maltophilia has multiple intrinsic and acquired resistances, including carbapenem resistance. In companion animals, the antimicrobial susceptibility and sequence types (STs) of S. maltophilia are not well understood due to its limited isolation rate. We investigated the antimicrobial susceptibilities and multilocus sequence types (MLSTs) of 38 S. maltophilia strains isolated from dogs and cats in Japan. Prevalence of resistance was detected for imipenem (100 %), aztreonam (94.7 %), piperacillin (65.8 %), trimethoprim-sulfamethoxazole (65.8 %), and ceftazidime (60.5 %). Rates of resistances to chloramphenicol, minocycline, and levofloxacin were low (2.6-5.3 %). MLST analysis revealed that all 38 strains were assigned to 34 STs, including 11 previously reported STs and 23 newly identified STs. Phylogenetic analysis of MLSTs enabled categorization of 13 isolates (34.2 %) into genogroup 6, which is a major genogroup of human isolates. Multinational surveillance would be needed to clarify the significance of antimicrobial-resistant S. maltophilia isolates from companion animals.

Sacran, a sulfated polysaccharide, suppresses the absorption of lipids and modulates the intestinal flora in non-alcoholic steatohepatitis model rats.

AIMS: The objective of this study was to investigate the effects of administering sacran, a sulfated polysaccharide, on liver biology, gut microbiota, oxidative stress, and inflammation on stroke-prone spontaneously hypertensive (SHRSP5/Dmcr) rats that develop fibrotic steatohepatitis with histological similarities to that of non-alcoholic steatohepatitis (NASH). MAIN METHODS: Four groups of 8-week-old SHRSP5/Dmcr rats were fed a high fat-cholesterol (HFC) diet for 4 and 8 weeks and administered either sacran (80 mg/kg/day) or a non-treatment, respectively. Liver function was evaluated by biochemical and histopathological analyses. Hepatic inflammatory markers were measured using mRNA expression. Fecal microbial profiles were determined via 16S rRNA sequencing. A triglyceride (TG) absorption test was administered to the 8-week-old Sprague-Dawley (SD) rats. KEY FINDING: Sacran administration was observed to decrease the extent of oxidative stress and hepatic biochemical parameters in serum and hepatic injury with the levels of transforming growth factor-beta (TGF-ß1) and tumor necrosis factor-alpha (TNF-α), being increased compared to those of the non-treatment group. At the genus level, sacran administration caused a significant decrease in the harmful Prevotella genus, and a significant increase in the useful Blautia genus was observed. Sacran administration also decreased the serum TG increase that was induced by administering corn oil to the SD rats. SIGNIFICANCE: We conclude that sacran administration has the potential to reduce the absorption of lipids into blood and to improve several gut microbiotas, in the gastrointestinal tract, thereby inhibiting the subsequent development of oxidative stress and hepatic markers in the systematic circulation on NASH.

Urinary Pharmacokinetic and Pharmacodynamic Profiles of Fosfomycin against Extended-Spectrum ß-Lactamase-Producing Escherichia coli with Canine Ex Vivo Modeling: A Pilot Study.

Fosfomycin is a candidate drug for extended-spectrum ß-lactamase (ESBL)-producing bacteria, but its efficacy is yet to be investigated in dogs. This study investigated the urinary pharmacokinetic/pharmacodynamic (PK/PD) profile of fosfomycin orally administered at 80 mg/kg to six healthy dogs to assess its efficacy for canine urinary tract infections (UTIs) caused by ESBL-producing bacteria. Four strains of ESBL-producing Escherichia coli (ESBL-EC) characterized by fosfomycin minimum inhibitory concentrations (MICs) of 0.5, 1, 2, and 32 µg/mL were used. Urine samples for the measurement of urinary drug concentrations and urinary bactericidal titers (UBTs) were obtained after drug administration. The urinary concentrations (µg/mL, mean ± SE) were 1348.2 ± 163.5, 1191.6 ± 260.2, and 661.1 ± 190.4 at 0-4, 4-8, and 8-12 h, respectively, after drug administration. The mean urinary area under the curve during the test period (AUC0-12) of fosfomycin was estimated to be 12,803.8 µg·h/mL. The UBTs for all tested strains fluctuated closely with urine concentration during the test period (r = 0.944-1.000), and the area under the UBT-versus-time curve correlated with the urinary AUC/MIC of each strain (r = 0.991). According to the optimal urinary PK/PD target value, fosfomycin at 80 mg/kg twice daily may be suitable for the treatment of canine UTIs caused by ESBL-EC presenting MIC ≤ 128 µg/mL.

Characterization of Antimicrobial Resistance in Serratia spp. and Citrobacter spp. Isolates from Companion Animals in Japan: Nosocomial Dissemination of Extended-Spectrum Cephalosporin-Resistant Citrobacter freundii.

In many countries including Japan, the status of emerging antimicrobial resistance among Serratia spp. and Citrobacter spp. in companion animals remains unknown because these genera are rarely isolated from animals. In this study, 30 Serratia spp. and 23 Citrobacter spp. isolates from companion animals underwent susceptibility testing for 10 antimicrobials. Phenotypic and genetic approaches were used to identify the mechanisms of extended-spectrum cephalosporins (ESC). Subsequently, ESC-resistant Citrobacter spp. strains underwent multilocus sequence typing and pulsed-field gel electrophoresis (PFGE). A significantly higher rate (34.8%) of ESC resistance was observed in Citrobacter spp. isolates than in Serratia spp. isolates (0%). ESC resistance was detected in five C. freundii strains, two C. portucalensis strains, and one C. koseri strain. All of the ESC-resistant Citrobacter spp. strains harbored CMY-type and/or DHA-type AmpC ß-lactamases. Three C. freundii strains harbored the CTX-M-3-type extended-spectrum ß-lactamases. Notably, the three blaCTX-3-producing and two blaCMY-117-bearing C. freundii strains (obtained from different patients in one hospital) had the same sequence type (ST156 and ST18, respectively) and similar PFGE profiles. We believe that ESC-resistant Citrobacter spp. are important nosocomial pathogens in veterinary medicine. Therefore, infection control in animal hospitals is essential to prevent dissemination of these resistant pathogens.

Assessment of urinary pharmacokinetic and pharmacodynamic profiles of faropenem against extended-spectrum ß-lactamase-producing Escherichia coli with canine ex vivo modelling: a pilot study.

This study was carried out to investigate the urinary pharmacokinetics and pharmacodynamics of faropenem administered orally at 5 mg kg-1 in six healthy dogs to assess the efficacy of the drug for canine urinary tract infections (UTIs) with extended-spectrum ß-lactamase (ESBL)-producing bacteria. Six strains of ESBL-producing Escherichia coli (ESBL-EC) with the following faropenem minimum inhibitory concentrations (MICs) were used: 1 µg ml-1 (n=2), 2 µg ml-1 (n=2), 4 µg ml-1 (n=1) and 16 µg ml-1 (n=1). Urine samples were obtained every 4 h for the first 12 h after administration to measure urinary drug concentration and urinary bactericidal titres (UBTs). Both the urine concentration of faropenem and the UBTs for all tested strains peaked at 0-4 h after administration, and decreased markedly at 8-12 h. The mean urinary concentration of faropenem at 8-12 h (23±5.2 µg ml-1) exceeded the MIC of 1 µg ml-1 by fourfold, which is required to inhibit the growth of 90  % of ESBL-EC. These findings indicate that faropenem administered twice daily at a dose of 5 mg kg-1 is acceptable for the treatment of most dogs with ESBL-EC-related UTIs.

Species distribution, virulence factors, and antimicrobial resistance of Acinetobacter spp. isolates from dogs and cats: a preliminary study.

We investigated the prevalence of virulence factors and antimicrobial resistance among 67 Acinetobacter spp. isolates, consisting of 21 Acinetobacter baumannii and 46 non-baumannii Acinetobacter from companion animals. The PCR analysis showed that the most prevalent virulence gene was afa/draBC (29.9%), followed by papC (22.4%) and cvaC (20.9%). Antimicrobial susceptibility testing revealed that resistance to gentamicin (14.9%) and ciprofloxacin (11.9%) was relatively prevalent. Five gentamicin- and/or ciprofloxacin-resistant A. baumannii strains were assigned to ST25, ST149, ST164, ST203, and ST1198. All ciprofloxacin-resistant isolates harbored point mutations in gyrA and/or parC. This is the first preliminary monitoring of animal-origin Acinetobacter spp. in Japan.

In vitro efficacy of 16 antimicrobial drugs against a large collection of ß-lactamase-producing isolates of extraintestinal pathogenic Escherichia coli from dogs and cats.

PURPOSE: The aim of this study was to assess the in vitro efficacy of candidate antimicrobials against extended-spectrum ß-lactamase (ESBL)-producing isolates of extraintestinal pathogenic Escherichia coli (ExPEC) from companion animals. METHODOLOGY: A total of 90 ESBL-producing ExPEC isolates from dogs and cats were tested for susceptibility to 16 antimicrobials with the agar dilution method. We also identified the ESBLs and AmpC ß-lactamases of these isolates with PCR and DNA sequencing.Results/Key findings. All isolates were susceptible to meropenem, tebipenem and amikacin (AMK), and various proportions were susceptible to latamoxef (LMX, 97.8 %), fosfomycin (FOM, 97.8 %), faropenem (FPM, 96.7 %), nitrofurantoin (NFT, 96.7 %), flomoxef (FMX, 93.3 %), piperacillin/tazobactam (PTZ, 92.2 %), cefmetazole (CMZ, 91.1 %), chloramphenicol (80.0 %), trimethoprim/sulfamethoxazole (64.4 %), amoxicillin/clavulanic acid (63.3 %), ceftibuten (60.0 %), tetracycline (52.2 %) and enrofloxacin (10.0 %). A genetic analysis showed that 83 of the 90 (92.2 %) isolates were positive for CTX-M-type genes: CTX-M-14 (n=26), CTX-M-27 (n=20), CTX-M-55 (n=17), CTX-M-15 (n=12), CTX-M-2 (n=5), CTX-M-24 (n=2), CTX-M-104 (n=2) and CTX-M-3 (n=1). Eight isolates also expressed AmpC ß-lactamase phenotypes. CONCLUSION: This study demonstrates that the susceptibility rates to PTZ, CMZ, LMX, AMK, FOM, FPM, NFT and FMX were similar to those to carbapenems (>90 %), implying that these drugs are available alternatives to carbapenems for the treatment of companion animals infected with ExPEC-producing CTX-M-type ESBLs. Further in vivo studies of the effective use of these antimicrobials are required.

Assessment of urinary pharmacokinetics and pharmacodynamics of orbifloxacin in healthy dogs with ex vivo modelling.

PURPOSE: The aim of this study was to investigate the urinary pharmacokinetics (PK) of orbifloxacin (OBFX) administered at 5 mg kg-1 in six healthy dogs. A further aim was to use an ex vivo model to evaluate the urinary PK and pharmacodynamics (PD) of OBFX to determine its urinary bactericidal titre (UBT), which represents the maximal dilution of urine allowing bactericidal activity. METHODOLOGY: Fourteen urinary tract infection (UTI) pathogenic strains of five bacterial species (Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Proteus mirabilis and Staphylococcuspseudintermedius) were used. Urine samples were obtained every 4 h for the first 24 h after OBFX administration, for measurement of urine drug concentration and UBT.Results/Key findings. The urine OBFX concentration peaked at 0-4, 4-8 or 4-8 h after administration, with a maximum concentration of 383±171 µg ml-1. Overall, the fluctuation in median UBT closely correlated with that of the mean urine OBFX concentration. In addition, the median areas under the UBT-time curves (AUBTs) were significantly inversely correlated with the MICs for OBFX in the tested strains (P<0.01). Notably, median UBTs and AUBTs were extremely low (0-0.5 and 2-5, respectively) in OBFX-resistant E. coli strains with MIC ≥8 µg ml-1. CONCLUSION: The fluctuation of UBTs closely correlated with that of urine concentration, and UBT values depended on the susceptibility of the bacterial strains to OBFX. We believe that ex vivo modelling to determine UBTs is useful to evaluate the urinary PK/PD of antimicrobials indicated for UTIs in dogs.

Phenotypic and molecular characterization of antimicrobial resistance in Enterobacter spp. isolates from companion animals in Japan.

The emergence of antimicrobial resistance among Enterobacter spp., including resistance to extended-spectrum cephalosporins (ESC), is of great concern in both human and veterinary medicine. In this study, we investigated the prevalence of antimicrobial resistance among 60 isolates of Enterobacter spp., including E. cloacae (n = 44), E. aerogenes (n = 10), and E. asburiae (n = 6), from clinical specimens of dogs and cats from 15 prefectures in Japan. Furthermore, we characterized the resistance mechanisms harbored by these isolates, including extended-spectrum ß-lactamases (ESBLs) and plasmid-mediated quinolone resistance (PMQR); and assessed the genetic relatedness of ESC-resistant Enterobacter spp. strains by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). Antimicrobial susceptibility testing demonstrated the resistance rates to ampicillin (93.3%), amoxicillin-clavulanic acid (93.3%), cefmetazole (93.3%), chloramphenicol (46.7%), ciprofloxacin (43.3%), tetracycline (40.0%), ceftazidime (33.3%), cefotaxime (33.3%), trimethoprim/sulfamethoxazole (28.3%), gentamicin (23.3%), and meropenem (0%). Phenotypic testing detected ESBLs in 16 of 18 ESC-resistant E. cloacae isolates but not in the other species. The most frequent ESBL was CTX-M-15 (n = 8), followed by SHV-12 (n = 7), and CTX-M-3 (n = 1). As for AmpC ß-lactamases, CMY-2 (n = 2) and DHA-1 (n = 2) were identified in ESC-resistant E. cloacae strains with or without ESBLs. All of the ESC-resistant E. cloacae strains also harbored one or two PMQRs, including qnrB (n = 15), aac(6')-Ib-cr (n = 8), and qnrS (n = 2). Based on MLST and PFGE analysis, E. cloacae clones of ST591-SHV-12, ST171-CTX-M-15, and ST121-CTX-M-15 were detected in one or several hospitals. These results suggested intra- and inter-hospital dissemination of E. cloacae clones co-harboring ESBLs and PMQRs among companion animals. This is the first report on the large-scale monitoring of antimicrobial-resistant isolates of Enterobacter spp. from companion animals in Japan.

Determination of minimum biofilm eradication concentrations of orbifloxacin for canine bacterial uropathogens over different treatment periods.

Biofilm formation can cause refractory urinary tract infections (UTIs) in dogs; however, minimum biofilm eradication concentrations (MBECs) of veterinary drugs against canine uropathogens remain to be investigated. In this study, the MBECs of orbifloxacin (OBFX), trimethoprim-sulfamethoxazole (TMS) and amoxicillin/clavulanate (ACV) over different time periods for treatment of canine uropathogenic Escherichia coli (n = 10) were determined. The MBECs of OBFX for other bacterial uropathogens, including Staphylococcus pseudintermedius (n = 5), Pseudomonas aeruginosa (n = 5), Klebsiella pneumoniae (n = 5) and Proteus mirabilis (n = 5) were also determined. Minimum inhibitory concentrations (MICs) were identified for all strains by broth microdilution, and MBECs were determined at 24, 72, and 168 hr using the Calgary biofilm method. The 24 hr MBECs of OBFX, TMS and ACV for the E. coli strains were significantly higher than the MICs (P < 0.05), and the 72 and 168 hr MBECs were significantly lower than those at 24 hr (P < 0.05). In addition, the 24 hr OBFX MBECs for the four other uropathogens were significantly higher than the corresponding MICs (P < 0.05). The 72 and/or 168 hr OBFX MBECs for S. pseudintermedius, K. pneumoniae and P. mirabilis were significantly lower than the 24 hr concentrations (P < 0.05), whereas for P. aeruginosa, no significant difference was found between any of the MBECs (P > 0.05). These data indicate that the administration period and uropathogenic bacterial species are important factors affecting the efficacy of OBFX treatment of biofilm-related UTIs in dogs.

Phenotypic and Molecular Characterization of Antimicrobial Resistance in Klebsiella spp. Isolates from Companion Animals in Japan: Clonal Dissemination of Multidrug-Resistant Extended-Spectrum ß-Lactamase-Producing Klebsiella pneumoniae.

The emergence of antimicrobial resistance in Klebsiella spp., including resistance to extended-spectrum cephalosporins (ESC) and fluoroquinolones, is of great concern in both human and veterinary medicine. In this study, we investigated the prevalence of antimicrobial resistance in a total of 103 Klebsiella spp. isolates, consisting of Klebsiella pneumoniae complex (KP, n = 89) and K. oxytoca (KO, n = 14) from clinical specimens of dogs and cats in Japan. Furthermore, we characterized the resistance mechanisms, including extended-spectrum ß-lactamase (ESBL), plasmid-mediated AmpC ß-lactamase (PABL), and plasmid-mediated quinolone resistance (PMQR); and assessed genetic relatedness of ESC-resistant Klebsiella spp. strains by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE). Antimicrobial susceptibility testing demonstrated that resistance rates to ampicillin, cephalothin, enrofloxacin, ciprofloxacin, trimethoprim/sulfamethoxazole, cefotaxime, gentamicin, tetracycline, chloramphenicol, amoxicillin-clavulanic acid, and cefmetazole were 98.1, 37.9, 37.9, 35.9, 35.0, 34.0, 31.1, 30.1, 28.2, 14.6, and 6.8%, respectively. Phenotypic testing detected ESBLs and/or AmpC ß-lactamases in 31 of 89 (34.8%) KP isolates, but not in KO isolates. Resistances to 5 of the 12 antimicrobials tested, as well as the three PMQRs [qnrB, qnrS, and aac(6')-Ib-cr], were detected significantly more frequently in ESBL-producing KP, than in non-ESBL-producing KP and KO. The most frequent ESBL was CTX-M-15 (n = 13), followed by CTX-M-14 (n = 7), CTX-M-55 (n = 6), SHV-2 (n = 5), CTX-M-2 (n = 2), and CTX-M-3 (n = 2). Based on the rpoB phylogeny, all ESBL-producing strains were identified as K. pneumoniae, except for one CTX-M-14-producing strain, which was identified as K. quasipneumoniae. All of AmpC ß-lactamase positive isolates (n = 6) harbored DHA-1, one of the PABLs. Based on MLST and PFGE analysis, ST15 KP clones producing CTX-M-2, CTX-M-15, CTX-M-55, and/or SHV-2, as well as KP clones of ST1844-CTX-M-55, ST655-CTX-M-14, and ST307-CTX-M-15, were detected in one or several hospitals. Surprisingly, specific clones were detected in different patients at an interval of many months. These results suggest that multidrug-resistant ESBL-producing KP were clonally disseminated among companion animals via not only direct but also indirect transmission. This is the first report on large-scale monitoring of antimicrobial-resistant Klebsiella spp. isolates from companion animals in Japan.

Serological surveillance for antibodies against Erysipelothrix species in wild boar and deer in Japan.

We investigated the seroprevalence of antibodies against Erysipelothrix in wild animals in Japan. Serum samples were collected from 48 wild boar, 26 Yezo deer and 26 Japanese deer in Japan. Growth agglutination (GA) test was performed to estimate antibody titers. As a result, positive results were obtained from 32 (66.7%), 1 (3.6%) and 6 (23.1%) samples from wild boar, Yezo deer and Japanese deer, respectively. Our findings suggest that wild animals may be an important reservoir of Erysipelothrix.

Effects of oral orbifloxacin on fecal coliforms in healthy cats: a pilot study.

The study objective was to determine the effect of oral orbifloxacin (ORB) on antimicrobial susceptibility and composition of fecal coliforms in cats. Nine cats were randomized to two groups administered a daily oral dose of 2.5 and 5.0 mg ORB/kg for 7 days and a control group (three cats per group). Coliforms were isolated from stool samples and were tested for susceptibilities to ORB and 5 other drugs. ORB concentration in feces was measured using high-performance liquid chromatography (HPLC). The coliforms were undetectable after 2 days of ORB administration, and their number increased in most cats after termination of the administration. Furthermore, only isolates of Escherichia coli were detected in all cats before administration, and those of Citrobacter freundii were detected after termination of the administration. E. coli isolates exhibited high ORB susceptibility [Minimum inhibitory concentration (MIC), ≤0.125 µg/ml] or relatively low susceptibility (MIC, 1-2 µg/ml) with a single gyrA mutation. C. freundii isolates largely exhibited intermediate ORB susceptibility (MIC, 4 µg/ml), in addition to resistance to ampicillin and cefazolin, and harbored qnrB, but not a gyrA mutation. HPLC revealed that the peaks of mean concentration were 61.3 and 141.0 µg/g in groups receiving 2.5 and 5.0 mg/kg, respectively. Our findings suggest that oral ORB may alter the total counts and composition of fecal coliform, but is unlikely to yield highly fluoroquinolone-resistant mutants of E. coli and C. freundii in cats, possibly because of the high drug concentration in feces.

First case of Propionibacterium acnes urinary tract infection in a dog.

BACKGROUND: Propionibacterium acnes has been rarely isolated as a commensal from dogs, but there is little evidence of pathogenicity. Urinary tract infections are common in dogs and are typically caused by various commensal bacteria. Here we present the first case report of a urinary tract infection caused by P. acnes. CASE PRESENTATION: A 6-year-old female Japanese Shiba Inu was hospitalized for polyuria, polydipsia, and severe hematuria. At admission, blood tests revealed leukocytosis, slight anemia, decreased albumin, and slightly elevated blood urea nitrogen. Computerized tomography showed gas accumulation on the inner side of the bladder wall. Urinalysis revealed proteinuria and bilirubinuria without glycosuria. The urine sediment contained large numbers of erythrocytes and leukocytes. Additionally, rod-shaped bacteria were detected by Diff-Quik staining. Enrofloxacin and metronidazole were administered empirically; however, the renal function declined sharply and the patient died 2 days later. Bacteriological examination revealed that the causative agent was Propionibacterium acnes, which was identified as sequence type 53 via multilocus sequence typing. This isolate showed high susceptibility to ampicillin, amoxicillin/clavulanic acid, cefoxitin, imipenem, clindamycin, tetracycline, chloramphenicol, and enrofloxacin, but was resistant to metronidazole. CONCLUSION: To the best of our knowledge, this is the first case report of a dog with urinary tract infection caused by P. acnes.

Phenotypic and molecular characterization of antimicrobial resistance in Proteus mirabilis isolates from dogs.

Large-scale monitoring of resistance to 14 antimicrobial agents was performed using 103 Proteus mirabilis strains isolated from dogs in Japan. Resistant strains were analysed to identify their resistance mechanisms. Rates of resistance to chloramphenicol, streptomycin, enrofloxacin, trimethoprim/sulfamethoxazole, kanamycin, ampicillin, ciprofloxacin, cephalothin, gentamicin, cefoxitin and cefotaxime were 20.4, 15.5, 12.6, 10.7, 9.7, 8.7, 5.8, 2.9, 2.9, 1.9 and 1.9%, respectively. No resistance to ceftazidime, aztreonam or imipenem was found. Class 1 and 2 integrases were detected in 2.9 and 11.7% of isolates, respectively. Class 1 integrons contained aadB or aadB-catB-like-blaOXA10-aadA1, whereas those of class 2 contained sat-aadA1, dhfr1-sat-aadA1 or none of the anticipated resistance genes. Of five distinct plasmid-mediated quinolone-resistance (PMQR) genes, only qnrD gene was detected in 1.9% of isolates. Quinolone-resistance determining regions (QRDRs) of gyrA and parC from 13 enrofloxacin-intermediate and -resistant isolates were sequenced. Seven strains had double mutations and three had single mutations. Three of nine ampicillin-resistant isolates harboured AmpC-type ß-lactamases (i.e. blaCMY-2, blaCMY-4 and blaDHA-1). These results suggest that canine Proteus mirabilis deserves continued surveillance as an important reservoir of antimicrobial resistance determinants. This is the first report, to our knowledge, describing integrons, PMQRs and QRDR mutations in Proteus mirabilis isolates from companion animals.

Identification of the cause of severe skin infection by Fournier transform infrared spectroscopy: a case of Fournier's gangrene caused by fish bone.

Fournier's gangrene (FG) is an infrequent but highly lethal infection. Here we report a 74-year-old man who presented with genital swelling and severe malaise. Based on the physical and imaging examination results, the diagnosis of FG was confirmed. Intraoperative findings showed dirty necrosis of soft tissue, and a splinter-shaped foreign body was found in the perirectal region. The foreign body was thought to be the cause of the condition, and it was analyzed using Fourier transform infrared spectroscopy. We found that the foreign body was a mixture of calcium phosphate and protein, suggesting that the splinter was a bone. Moreover, during the medical interview, the patient mentioned about intake of fish around the time of onset of symptoms. Therefore, to confirm the results of the analysis, DNA was extracted from the foreign body, and genomic PCR with subsequent sequence analysis was performed. The DNA sequence was identical to that of Oncorhynchus kisutch, a salmon that is a very popular food in Japan. On the basis of these findings, we concluded that FG in this case was caused by the penetration into the rectum of an accidentally ingested fish bone. Although some cases of intra-abdominal abscess due to accidental ingestion of fish bone have been reported, FG caused by fish bone is extremely rare.