Yasushi Kataoka

Leptospirosis is a zoonosis that affects humans and animals worldwide. Raccoons (Procyon lotor), adopted in urban environments, may act as potential reservoirs of Leptospira. We investigated the prevalence of pathogenic Leptospira in the kidney and urine samples of raccoons living in Tokyo, as well as anti-leptospiral antibodies in their serum, and aimed to examine the factors that expose raccoons to Leptospira. Polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) were used to detect leptospiral DNA and anti-leptospiral antibodies, respectively. Thirty-six of 156 raccoons (23.1%) were positive by PCR, and 16 of 165 raccoons (9.7%) were positive by ELISA. The prevalence and seroprevalence rates differed depending on the raccoon dispersal period. We used univariable logistic regression to estimate the environmental factors associated with pathogenic Leptospira and anti-leptospiral antibodies in raccoons. Significant differences were observed in the PCR results for the seasons (spring–summer) (p = 0.01), average monthly temperature (p < 0.01), and average monthly rainfall (p < 0.01). No significant difference was seen in the ELISA results, but raccoons in larger urban areas tended to have higher seroprevalence rates (p = 0.06). We identified a pattern of leptospiral spread in raccoon dispersal and environmental factors that expose raccoons to Leptospira.

The purpose of this retrospective study was to assess the efficacy of antimicrobial therapy for bovine acute Klebsiella pneumoniae mastitis. We evaluated data from cattle in Ehime, Japan, with naturally occurring acute mastitis due to K. pneumoniae (n=208) or Escherichia coli (n=201). Survival was significantly shorter in cattle with acute K. pneumoniae mastitis (median, 76 days) compared with the disease caused by E. coli (median 464 days). In 2004-2008, because both species were highly susceptible to cefazolin, cases of K. pneumoniae and E. coli mastitis were treated solely with cefazolin, yielding clinical cure rates of 52.8% for K. pneumoniae and 86.0% for E. coli. However, since 2009, the efficacy of treatment of K. pneumoniae mastitis with cefazolin alone has decreased. When cefazolin administered on the first disease day led to clinical improvement, treatment with cefazolin was continued. However, when cefazolin administered on the first disease day failed to yield clinical improvement, the antibiotic was switched to a fluoroquinolone on the second day, resulting in cure rates of 76.7% for K. pneumoniae and 80.0% for E. coli. These findings suggest that, when the first-line drug (e.g., cefazolin) is ineffective, promptly changing to a second-line drug (e.g., a fluoroquinolone) increases the cure rate for bovine K. pneumoniae mastitis.

In human medicine, fecal microbiota transplantation (FMT) is an effective treatment for recurrent Clostridioides difficile infection. It has also been tested as a treatment for multiple gastrointestinal diseases, including inflammatory bowel disease (IBD). However, only a few studies have focused on the changes in the microbiome following FMT for canine IBD. Here, we performed FMT in nine dogs with IBD using the fecal matter of healthy dogs and investigated the subsequent changes in the fecal microbiome and clinical signs. In three dogs, the fecal microbiome was examined by 16S rRNA sequencing. Fusobacteria were observed at a low proportion in dogs with IBD. However, the post-FMT microbiome became diverse and showed a significant increase in Fusobacteria proportion. Fusobacterium was detected in the nine dogs by quantitative polymerase chain reaction. The proportion of Fusobacterium in the post-FMT fecal microbiome was significantly increased (p<0.05). The changes in clinical signs (e.g., vomiting, diarrhea, and weight loss) were evaluated according to the canine inflammatory bowel disease activity index. The score of this index significantly decreased in all dogs (p<0.05) with improvements in clinical signs. These improvements were related to the changes in the proportion of microbes, particularly the increase in Fusobacterium. The dogs with IBD showed a lower proportion of Fusobacterium than healthy dogs. This suggests that a low proportion of Fusobacterium is a characteristic feature of canine IBD and that Fusobacterium is involved in this disease. The results of this study may help elucidate the pathogenesis of this disease and its association with Fusobacterium.

Cefapirin (CEP) and cefalonium (CNM) are first-generation cephalosporins widely used to treat bovine mastitis caused by Gram-positive bacteria including staphylococci. However, disks for susceptibility testing of those drugs in causative bacteria are not available. This study evaluated the efficacy of 10 µg and 30 µg pilot disks of CEP (CEP10 and CEP30) and CNM (CNM10 and CNM30) against 130 Staphylococcusaureus isolates from bovine mastitis. Scattergrams of minimum inhibitory concentrations (MICs) and zone diameters (ZDs) illustrated significant correlations between the MICs and ZDs of CEP10 (r = -0.912), CEP30 (r = -0.933), CNM10 (r = -0.847), and CNM30 (r = -0.807). The analysis by Normalized Resistance Interpretation indicated that the epidemiolocal cut-off value (ECV) of MIC for both cefapirin and cefalonium is ≤ 0.5 µg/mL, and the ECV of ZD for CEP10, CEP30, CNM10, and CNM30 were ≥ 22 mm, ≥ 25 mm, ≥ 22 mm, and ≥ 29 mm, respectively. We believe that both 10 μg and 30 μg CEP and CNM susceptibility disks will be helpful for guiding the appropriate use of these antibiotics for bovine mastitis. Further studies toward the establishment of clinical breakpoint of CEP and CNM would be needed for their routine use.

The effects of prescription diets on canine intestinal microbiota are unknown. In this study, we used next generation sequencing to investigate the impact of four commercially available prescription diet regimens on the fecal microbiome in six healthy dogs. The diet regimens used were as follows: weight-loss diet, low-fat diet, renal diet, and anallergenic diet. We found a significantly decreased proportion of phylum Actinobacteria with the weight-loss diet compared to the anallergenic diet. There were no significant differences in the proportion of phylum Bacteroidetes between the four diets. The proportion of phylum Firmicutes was significantly decreased with the weight-loss diet compared to the anallergenic diet. The proportion of phylum Fusobacteria was significantly increased with the weight-loss diet compared to the anallergenic diet. There were no significant differences in the proportion of phylum Proteobacteria after consumption of the four diets. We therefore demonstrated that commercial prescription diet influences the fecal microbiome in healthy dogs. These results might be useful when choosing a prescription diet for targeting a disease.

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 β-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 β-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.

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 f)-lactamase (ESBL), plasmid-mediated AmpC f)-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 3-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 [qnr-B, corS, 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 beta-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.

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 f)-lactamase (ESBL), plasmid-mediated AmpC f)-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 3-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 [qnr-B, corS, 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 beta-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.

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.

A total of 122 raptors bred as pet birds were sampled from 2007 to 2008 for the detection Salmonella and Escherichia coli. Cloacal swabs were collected from each bird and cultured. Salmonella spp. were not detected in any samples. A total of 98 E. coli strains were isolated from 122 fecal samples. A total of 18 of 98 E. coli isolates belonging to the following O serogroups were identified: O8 (9 strains), O1 (3 strains), O168 (3 strains), O86 (2 strains) and O128 (1 strain). In the order Accipitriformes, 37 (59.7%) of 62 strains were identified as verotoxin producing E. coli, and 1 strain (1.6%) was identified as heat-labile enterotoxin producing E. coli. On the other hand, in the order Strigiformes, 17 (47.2%) of 36 strains were identified as verotoxin producing E. coli, and 7 strains (19.4%) were identified as heat-labile enterotoxin producing E. coli. A comparison of resistance patterns of E. coli isolates from Accipitriformes and Strigiformes revealed that 33 (53.2%) of 62 isolates from Accipitriformes and 5 (13.9%) of 36 isolates from Strigiformes were antimicrobial sensitive (P<0.05).

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-bla(OXA10)-aadA1, whereas those of class 2 contained sat aadA 1, 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 (CIRDRs) 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 beta-lactamases (i.e. bla(CMY-2), bla(CMY-4) and bla(DHA-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.

In this study, we examined the antimicrobial susceptibility of the enterococci isolated from dogs and cats in Japan during 2011-2012. Fecal samples were collected from 84 dogs and 16 cats that underwent antibiotic treatment. Enterococci were detected in 70 of 84 dogs (83.3%) and 7 of 16 cats (43.8%). The most prevalent Enterococcus species was Enterococcus faecalis (64.9%); Enterococcus faecium and Enterococcus durans were also isolated from 14 of 77 (18.2%) and 5 of 77 (6.5%) of these animals, respectively. The most active resistance was observed for erythromycin (44.2%) and oxytetracycline (44.2%), and there was considerable resistance to lincomycin (41.6%), gentamicin (31.2%) and kanamycin (31.2%). Compared with the results of a similar study conducted in 2006 and 2007, enterococci susceptibility to enrofloxacin and ampicillin had significantly increased. Enterococcus gallinarum harboring vanC1 and Enterococcus casseliflavus harboring vanC2/3 were isolated from 4 of 77 enterococcal isolates. However, no enterococcal isolates were resistant to vancomycin. Multidrug resistance was found for as few as two and as many as nine antimicrobials regardless of the class. These results dem

In this study, we examined the antimicrobial susceptibility of the enterococci isolated from dogs and cats in Japan during 2011-2012. Fecal samples were collected from 84 dogs and 16 cats that underwent antibiotic treatment. Enterococci were detected in 70 of 84 dogs (83.3%) and 7 of 16 cats (43.8%). The most prevalent Enterococcus species was Enterococcus faecalis (64.9%); Enterococccus faecium and Enterococcus durans were also isolated from 14 of 77 (18.2%) and 5 of 77 (6.5%) of these animals, respectively. The most active resistance was observed for erythromycin (44.2%) and oxytetracycline (44.2%), and there was considerable resistance to lincomycin (41.6%), gentamicin (31.2%) and kanamycin (31.2%). Compared with the results of a similar study conducted in 2006 and 2007, enterococci susceptibility to enrofloxacin and ampicillin had significantly increased. Enterococcus gallinarum harboring van C] and Enterococcus casselifiavus harboring van C2/3 were isolated from 4 of 77 enterococcal isolates. However, no enterococcal isolates were resistant to vancomycin. Multidrug resistance was found for as few as two and as many as nine antimicrobials regardless of the class. These results demonstrate that dogs and cats treated with antibiotics are commonly colonized with antimicrobial-resistant enterococci.

The aim of this study was to show that a 39-kDa protein or OmpH of Pasteurella multocida strain P-1059 is essential for cross protection. Strain PBA322, a thinly capsulated strain of P. multocida strain P-1059, was used as a live vaccine in chickens. Strain PBA322 is a thinly capsulated strain in comparison with the parental strain P-1059. Chickens were vaccinated by single injection and then challenge-exposed with strains P-1059 or X-73 at two weeks post vaccination. Moreover, immune responses were also evaluated for both humoral and cellular immune response by ELISA and lymphocyte proliferation assay, respectively. The results showed that the live vaccine induced efficient immunity to protect chickens from challenge-exposure to the parent strain, but that the heterologous protection was poor. We concluded that the 39-kDa protein is essential for cross protection.

The purpose of the present study was to determine the prevalence of antibiotic-resistant enterococci in dogs and cats subjected to differing antibiotic pressures, and the prevalence of vancomycin resistance genes in isolates from these animals. Enterococci were isolated from fecal samples of 65 healthy dogs and 29 healthy cats brought to animal hospitals, from rectal swabs of 73 puppies and 15 kittens from five breeders and two pet shops, and from fecal samples of 20 dogs and 9 cats that were treated with antibiotics in Nippon Veterinary and Life Science University Animal Medical Center. The rates of resistance to ampicillin among isolates from the kitten puppy group and healthy dog cat group were 6.8 and 4.3%, respectively. In contrast, the rates of resistance to ampicillin in enterococci from the treatment group under antibiotic pressure were 37.5%. There was a significant difference between the antibiotic-treated group and the untreated group (P&lt;0.01). Similarly, in the treatment group, the rate of resistance to enrofloxacin was extremely high (75.0%). In comparison, in the healthy group and kitten puppy group, the rates of resistance to enrofloxacin were 23.4 and 12.1%, respectively. Among these groups, a significant difference was also observed in the apparent resistance rates (P&lt;0.01). Vancomycin-resistant enterococci (VRE) harboring vanA or vanB were not detected in any groups. Therefore, contamination of VRE in dogs and cats is still considered to be minimal in Japan.

Background: The mutant prevention concentration (MPC) is an important parameter to evaluate the likelihood of growth of fluoroquinolone-resistant mutants for antimicrobial-pathogen combinations. The MPCs of fluoroquinolones for different canine pathogens have not been compared. In this study, we compared for the first time orbifloxacin MPCs between susceptible strains of Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus pseudintermedius of canine origin. Methods: More than 10(10) CFU/ml of 10 strains of each bacterial species were inoculated onto Muller-Hinton agar supplemented with different concentrations of orbifloxacin from 1x to 64x minimum inhibitory concentration (MIC) and the MPCs were recorded. MICs of original strains and of mutants arising after exposure to sub-MPC concentrations (one per original strain) were determined in the presence or absence of efflux pump inhibitors (EPIs). The effects of quinolone resistance-determining region (QRDR) mutations were also examined. Results: MPCs were significantly higher for P. aeruginosa (16-128 mu g/ml) than for E. coli (0.5-32 mu g/ml). MPCs for S. pseudintermedius varied between the low-susceptible (16-128 mu g/ml) and the high-susceptible strains (4-16 mu g/ml) and were the most broadly distributed among the three species. Regarding resistance mechanisms, only one QRDR mutation in gyrA was found in all of the 10 mutants of E. coli and in 4 of the 10 mutants of P. aeruginosa, whereas mutations in both grlA and gyrA were found in 3 mutants and one mutation in grlA was found in 2 mutants among the 10 mutants of S. pseudintermedius. In the presence of an EPI, the MICs of P. aeruginosa mutants decreased markedly, those of E. coli mutants decreased moderately, and those of S. pseudintermedius mutants were unaffected. Conclusions: MPCs of orbifloxacin vary between bacterial species of canine pathogens, possibly due to the diversity of the main fluoroquinolone resistance mechanism among these species. Therefore, the type of bacterial species should be taken into consideration when using fluoroquinolone drugs such as orbifloxacin in canines.

Thirty-three cefazolin-resistant extraintestinal pathogenic Escherichia coli strains from companion animals were screened for blaCMY-1, blaCMY-2, blaSHV, blaTEM, and blaCTX-M genes. Extended-spectrum beta-lactamase-producing strains were further characterized by O serotyping and multilocus sequence typing. It was found that 20 and 17 isolates harbored TEM-1 and CMY-2 beta-lactamases, respectively, and 13 isolates harbored both beta-lactamases. One isolate harbored DHA-1 beta-lactamase. Eleven isolates were found to possess CTX-M beta-lactamases (CTX-M-27 [n= 6], CTX-M-14 [n= 3], CTX-M-15 [n= 1], and CTX-M-55 [n= 1]). Of 11 CTX-M-positive strains, four strains were O25b-ST131 clones harboring CTX-M-27, and the remaining seven strains belonged to O6-ST127, ONT-ST354, O159-ST539, O1-ST648, O8-ST1642, O25b-ST2042, and ONT-ST2178.

Orbifloxacin is a fluoroquinolone drug used widely in companion animal medicine. In this study, we firstly determined post-antibiotic effects (PAEs) and post-antibiotic sub-minimum inhibitory concentrations (MIC) effects (PA-SMEs) of orbifloxacin for two strains each of Escherichia coli and Pseudomonas aeruginosa from dogs, and these parameters were compared with those of enrofloxacin. At twice the MIC, the PAEs of orbifloxacin ranged from -0.28-0.93 h (mean, 0.29 h) for E. coli and -0.18-1.18 h (mean, 0.37 h) for P. aeruginosa. These parameters were not significantly different for E. coli and shorter for P. aeruginosa, compared to enrofloxacin (P &lt; 0.05). Continued exposure to 0.1, 0.2, and 0.3 the MIC of orbifloxacin resulted in average PA-SMEs of 0.55, 1.11, and 2.03 h, respectively, for E. coli, and 1.04, 1.40, and 2.47 h, respectively, for P. aeruginosa. These PA-SMEs, which had no significant differences with those of enrofloxacin, were significantly longer than the corresponding PAEs (P &lt; 0.05). These results suggest that the PA-SME of orbifloxacin for E. coli and P. aeruginosa can be meaningfully prolonged by increase of sub-MICs.

Objective-To assess the status of antimicrobial resistance (AMR), identify extraintestinal virulence factors (VFs) and phylogenetic origins, and analyze relationships among these traits in extraintestinal pathogenic Escherichia coli (ExPEC) isolates from companion animals. Sample-104 E coli isolates obtained from urine or genital swab samples collected between 2003 and 2010 from 85 dogs and 19 cats with urogenital infections in Japan. Procedures-Antimicrobial susceptibility of isolates was determined by use of the agar dilution method; a multiplex PCR assay was used for VF gene detection and phylogenetic group assessment. Genetic diversity was evaluated via randomly amplified polymorphic DNA analysis. Results-Of the 104 isolates, 45 (43.3%) were resistant to &gt; 2 antimicrobials. Phylogenetically, 64 (61.5%), 22 (21.2%), 13 (12.5%), and 5 (4.8%) isolates belonged to groups B2, D, B1, and A, respectively. Compared with other groups, group B2 isolates were less resistant to all tested antimicrobials and carried the pap, hly, and cnf genes with higher frequency and the aer gene with lower frequency. The aer gene was directly associated and the pap, sfa, hly, and cnf genes were inversely associated with AMR. Randomly amplified polymorphic DNA analysis revealed 3 major clusters, comprised mainly of group B1, B2, and D isolates; 2 subclusters of group B2 isolates had different VF and AMR status. Conclusions and Clinical Relevance-Prevalences of multidrug resistance and human-like phylogenetic origins among ExPEC isolates from companion animals in Japan were high. It is suggested that VFs, phylogenetic origins, and genetic diversity are significantly associated with AMR in ExPEC. (Am J Vet Res 2012;73:409-417)

In this study, fecal Escherichia coli isolates (n = 188) from 34 dog-owner pairs and 26 healthy control humans (2 isolates per individual) were tested for susceptibility to 6 antimicrobials and screened for virulence genes. Genetic diversity between canine and owner isolates was evaluated by pulsed-field gel electrophoresis (PFGE). Canine isolates exhibited significantly different rates of resistance to four and two antimicrobials, compared to control and owner isolates, respectively. Of the genes examined, the prevalence of sfa, hly, and cnf genes in canine isolates were higher than in control isolates, but not than in owner isolates. These results suggest that characteristics of owner isolates are somewhat similar to canine isolates, compared to isolates from non-dog owners. In addition, PFGE analysis revealed that transfer of E. coli between owners and their dogs had occurred within 3/34(8.8%) house-holds. Considering the effects of dog ownership on the population of E. coli isolates from owners, further epidemiological studies are required. (C) 2011 Elsevier Ltd. All rights reserved.

Seventy-three Pseudomonas aeruginosa isolates were collected from dogs and cats in Japan to investigate antimicrobial susceptibility and resistance mechanisms to anti-pseudomonal agents. Resistance rates against orbifloxacin, enrofloxacin, ciprofloxacin, cefotaxime, aztreonam and gentamicin were 34.2, 31.5, 20.5, 17.8, 12.3 and 4.1%, respectively. The degree of resistance to cefotaxime, orbifloxacin, and enrofloxacin was greatly affected by efflux pump inhibitors, indicating overexpression of efflux pump contributes to these resistances. Notably, orbifloxacin and enrofloxacin resistance was observed even in isolates without mutations in the target sites. This is the first report on cephalosporin- and fluoroquinolone-resistant isolates of P. aeruginosa from Japanese companion animals.

A mutant strain, PBA322, was constructed by electroporation of a phagemid containing the coding region of antisense RNA of the ompH gene, encoding 39 kDa capsular protein or OmpH, into the parental strain P-I 059 (serovar A:3) of Pasteurella multocida, and the pathogenicity was determined in mice and chickens. Grayish colonies of the mutant, indicating loss of capsule synthesis, were observed under a stereomicroseope using obliquely transmitted light, while iridescent colonies were observed for the parental strain. Moreover, strain PBA322 showed a low amount of OmpH compared with the parental strain on SDS-PAGE. Additionally, the capsule of strain PBA322 was thinner than that of the parental strain according to electron microscopy, correlating to the attenuation against chickens. In conclusion, strain PBA322, the mutant of P. multocida strain P-1059, was completely attenuated for chickens.

Campylobacter species were isolated from chicken meats in northeastern Thailand from December, 2007 to June, 2008. From 1,930 samples, Campylobacter spp. were obtained from 556 (28.8%). Campylobacter species were isolated from chicken livers at higher percentages compared to the other parts of chicken meats (p<0.001). Among 294 Campylobacter isolates, 187 (63.6%) were identified as C. jejuni, and 107 (36.4%) as C. coli. The results of serotyping by Penner's method showed that serotype L (22.7%) and serotype A (18.7%) were predominant. Antimicrobial susceptibility test revealed that 90.7, 37.3, 29.3 and 13.3% of C. jejuni were resistant to OFLX, DOXY, EM and CP, respectively. MIC50/MIC90 of OFLX, DOXY, EM and CP were 16/128, 4/256, 0.5/128 and 2/64 μg/ml, respectively. Precaution needs to be emphasized when attempt to use OFLX and DOXY for veterinary and human medicine due to the high percentage of resistance among C. jejuni isolated from chicken meat origin. These alarming figures should be notified to the general public.

Although the dog breeding industry is common in many countries, the presence of antimicrobial resistant bacteria among pups in kennels has been infrequently investigated. This study was conducted to better understand the epidemiology of antimicrobial-resistant Escherichia coli isolates from kennel pups not treated with antimicrobials. We investigated susceptibilities to 11 antimicrobials, and prevalence of extended-spectrum beta-lactamase (ESBL) in 86 faecal E. coli isolates from 43 pups in two kennels. Genetic relatedness among all isolates was assessed using pulsed-field gel electrophoresis (PFGE). Susceptibility tests revealed that 76% of the isolates were resistant to one or more of tested antimicrobials, with resistance to dihydrostreptomycin most frequently encountered (66.3%) followed by ampicillin (60.5%), trimethoprim-sulfamethoxazole (41.9%), oxytetracycline (26.7%), and chloramphenicol (26.7%). Multidrug resistance, defined as resistance against two or more classes of antimicrobials, was observed in 52 (60.5%) isolates. Three pups in one kennel harboured SHV-12 ESBL-producing isolates. A comparison between the two kennels showed that frequencies of resistance against seven antimicrobials and the variation in resistant phenotypes differed significantly. Analysis by PFGE revealed that clone sharing rates among pups of the same litters were not significantly different in both kennels (64.0% vs. 88.9%), whereas the rates among pups from different litters were significantly different between the two kennels (72.0% vs. 33.3%, P &lt; 0.05). The pups in the two kennels had antimicrobial-resistant E. coli clones, including multidrug-resistant and ESBL-producing clones. It is likely that resistant and susceptible bacteria can clonally spread among the same and/or different litters thus affecting the resistance prevalence.

Molecular epidemiology analyses of the 36 clinical isolates of Pasteurella multocida from various avian hosts in Japan between 1976 to 2007 including 5 reference strains from the U.S.A., Taiwan and Indonesia were performed by employing the single-enzyme amplified fragment length polymorphism (SE-AFLP) comparison with the classical Apal-based pulsed-field gel electrophoresis (PFGE). As the results. SE-AFLP gave 21 profiles while PFGE gave 20 profiles. The Simpson&apos;s index of diversity analysis indicated that SE-AFLP gave a high discrimination power than PFGE. This concluded that SE-AFLP is a higher discrimination power than PFGE to differentiate avian P. multocida isolates in Japan. In addition, the genetical profiles suggested that there is the evolution of somatic serotype 3 strain in the indigenous host of Japan.

For three years investigations from 1996 to 1998, we tried to isolate Escherichia coli O157:H7 from fecal samples collected from dogs and cats. In results, E. coli O157:H7 was isolated from 1 out of 614 samples (0.16%). This isolate produced Stx1 and Stx2 and was isolated from a dog kept by a human patient infected with E. coli O157:H7. Excluding in this case, dogs and cats as companion animals, therefore, may not give harbor to E. coli O157:H7.

Seven hundred thirty-nine animal strains and 662 livestock-farmer strains, consisting of Escherichia coli and enterococci, were examined for their pulsed-field gel electrophoresis (PFGE) and antimicrobial-resistance patterns. Two hundred fifty-eight and 203 PFGE patterns were found among 739 animal strains isolated from animals comprising broilers, pigs and cattle, and 662 human strains isolated from livestock farmers, respectively, from 27 farms in Japan. These results demonstrated that the PFGE patterns found among E. coli and enterococci strains from animals and livestock-farmers were heterogeneous and considerably diverse. The strains having both the identical PFGE pattern and the same drug-resistance pattern were defined as a single clone in this study. Seven types of E. coli and enterococci clones were shared among animals within the same farms and between the different farms housing the same animal species. The 25 strains (3.4%) of 739 E. coli and enterococci animal strains belonged to these seven types of clones. Only three types of E. coli clones were shared among animals between the different farms housing different animal species, but no identical E. faecalis or E. faecium clones were found between different animal species farms. The 15 strains (2.0%) of 739 E coli and enterococci animal strains belonged to these three types of clones. Additionally, the 11 strains (1.5%) of 739 E coli and enterococci strains isolated from animals were identical clones to strains isolated from livestock farmers of the same farm. These results suggest that the transmission of animal clones to livestock farmers or vice versa is less common.

A number of 689 Streptococcus suis isolates collected nationwide from diseased and healthy pigs from 1987 to 1996 were surveyed for antibiotic susceptibilities to 11 drugs. No isolates resistant to amoxicillin, chloramphenicol, and sulfamethoxazole/ trimethoprim were found. Isolates were highly susceptible to penicillins (penicillin G, ampicillin, and amoxicillin) except cloxacillin. They were not susceptible to tetracycline, streptomycin, and kanamaycin (MIC90 50 μg/ml, ≥100 μg/ml, and ≥100 μg/ml, respectively). Multiple-resistant isolates (≥3 antimicrobial agents) were found in 20.3% of all isolates tested.

Bactcrial isolation from slaughtered pigs with endocarditis was carried out from 1985 to 1994. A total of 495 (0.025%) out of 2,006,127 pigs were diagnosed as having endocarditis. Though bacteria were significantly isolated from 399 of the 495 pigs, bacteria could not be isolated in 96 pigs (19.4%). In 11 pigs, 2 bacterial species were isolated from heart lesion. Streptococcus suis was isolated from 127 cases (25.7%), Streptococcus dysgalactiae from 75 (15.2%), Erysipelothrix rhusiopathiae from 63 (12.7%), Actinomyces pyogenes from 39 (7.9%), Pasteurella multocida from 11 (2.2%), Staphylococcus aureus from 10 (2.0%), and Streptococcus porcinus from 8 (1.6%). Among the 99 isolates biochemically identified as S. suis, the major serotype was S. suis type 2 (35.4%). The remainder of the typable isolates were identified as serotypes 1/2 (2.0%) and 9 (1.0%). A total of 61 isolates (61.6%) were untypable.

Pasteurella multocida has been implicated in a variety of infections, including pulmonary, skin, bone, joint, cardiovascular, and central nervous system infections. Urinary tract infections caused by P. multocida, however, are rare. We studied a case of urinary tract infection caused by P. multocida in a 70-year-old woman with a 22-year history of non-insulin-dependent diabetes mellitus. Her urine was positive for gram-negative bacilli, which were identified as P. multocida. The same bacilli were isolated from the oral cavity of her daughter's pet dog. Both bacilli were found to be identical in terms of biological properties, drug susceptibility profile, serotype, and plasmid type. The clinical impact of the infection of this patient was minimal. Her hemoglobin A1C level had clearly improved one year after disappearance of the bacilli. The main reason for the improvement in the urinary findings was cessation of animal contact due to the death of the dog, with the antibiotic treatment only being temporarily effective. These findings suggest that P. multocida urinary tract infection is almost asymptomatic, even if such infection results in an increased hemoglobin A1C level. © 1997, THE JAPAN DIABETES SOCIETY. All rights reserved.

One hundred and one strains of enterotoxemic Escherichia coli (ETEEC) O-group 139 isolated from swine with edema disease were investigated for their adherence to HEp-2 cells in the presence of D-mannose. All strains adhered in large numbers to the cells (21 to 60 bacteria/cell). No correlation was found between the presence of F107 fimbria on the organisms and the adherence to the cells. Adhesion-inhibition tests showed that anti-K12 serum inhibited the adhesion ETEEC O-group 139 (an inhibition rate of 63 to 65%), but anti-F107 or anti-O139 sera did not. These results indicate that the capsular K12 antigen may be one of the pathogenic factors of ETEEC O-group 139.

In order to examine how the occurrences of drug-resistant bacterial strains in groups of pigs are influenced by the conditions for uses of antibiotics and by the differences in the methods for rearing and control, 328 strains in total of Escherichia coli derived from a SPF pig farm and a conventional pig farm were checked for sensitivity to 9 kinds of antibiotics and for conjugative R plasmids. The 9 antibiotics included TC, SM, KM, ABPC, CP, NA, CL, FRM, and SMX-TMP combination. The results were as described below.<br>1. Of the total 199 strains of Escherichia coli derived from a SPF pig farm, 131 strains (65.2%) were found to be resistant to one or more of the tested antibiotics, while 126 (97.7%) out of 129 strains of Escherichia coli derived from a conventional pig farm were found to be resistant.<br>2. In the comparison of the resistant strains, those derived from the SPF pig farm showed the largest share of sensitive strains of 67 strains (33.7%), of which those resistant to three agents and those resistant to two agents shared approximately. 20% each. In contrast, those resistant to multiple strains shared the majority of the strains derived from the conventional pig farm. More specifically, the strains resistant to 3 agents and those resistant to 2 agents had the largest share of 30.2% each. Those resistant to 4 agents shared 20.2%, while the share of the sensitive strains was as small as 3 strains (2.3%).<br>3. The ratio of detection by conjugation analysis resulted in 64.6% in the strains derived from the SPF pig farm, while that of the strains derived from the conventional pig farm amounted to 56.3%, showing hardly any difference between the SPF pig farm and the conventional pig farm in terms of resistance determinants on R plasmid.