Pathogenicity of Escherichia coli from dogs with UTI in relation to urovirulence factors.

Six strains of Escherichia coli, isolated from urine of dogs with urinary tract infection (UTI), were examined to assess of urovirulence factors (UVFs) in the pathogenesis of UTI in an experimental pyelonephritis mouse model. From the results of ID50 and LD50, isolates having different UVFs in the same O serotypes varied in pathogenicity, and isolates having the same UVFs in different O serotypes had nearly the same pathogenicity. Histopathogenic examination revealed that the presence of pap, hly and cnfl contributed greatly to the development of upper UTI. It has also been suggested that hly and cnfl significantly related to the LD50 of the strain in the mouse model, confirming that UVFs are closely related to the pathogenicity of canine UTI.

Clonality analysis of various hematopoietic disorders in cats naturally infected with feline leukemia virus.

The clonality analysis of the bone marrow cells was carried out by detecting the integrated proviruses of feline leukemia virus (FeLV) to understand the pathogenesis of FeLV-associated hematopoietic disorders in cats. Bone marrow cells from 4 cases with acute myeloid leukemia (AML), 9 cases with myelodysplastic syndromes (MDS), 2 cases with pure red cell aplasia (PRCA) and 3 healthy carriers infected with FeLV were subjected to Southern blot analyses using an exogenous FeLV probe. Clonal hematopoiesis was found in all the cases with AML and in 6 of the 9 cases with MDS, but not in the cases with both PRCA and healthy carriers infected with FeLV. In the 2 cases with MDS, it was thought that the same clones of the hematopoietic cells might proliferate before and after the progression of the disease irrespective of the changes of the hematological diagnoses by cytological examination. This study indicates that MDS in cats is a disease manifestation as a result of clonal proliferation of hematopoietic cells and can be recognized as a pre-leukemic state of AML.

Serotypes and virulence factors of Escherichia coli strains isolated from dogs and cats.

E. coli strains isolated from urine of dogs and cats with urinary tract infections (UTI) and from feces of healthy one's were serotyped, and the serotypes were correlated with uropathogenic virulence factors. The most prevalent O-serotypes, O4 and O6, were isolated from dogs and cats with UTI. In contrast, O11 and O102 strains were the most frequently found from feces of healthy dogs and cats. Most of type O4 and O6 strains possessed such virulence factors as pil, pap, sfa, hly, and cnf1, while most type O11 and O102 strains pil only or pil and aer. All strains of type O75 possessed afaI and aer. K1 antigen was negative in all strains obtained from UTI.

A study of metabolites isolated from the urine samples of cats and dogs administered orbifloxacin.

Urine samples of cats and dogs collected for 24 hr after a subcutaneous injection of orbifloxacin (OBFX) were analyzed. The metabolites were examined using HPLC. In the dog urine, 87% of total was the parent compound and 13% glucuronide compound of OBFX and 96% was parent and 4% metabolite in the cat urine. The metabolite of cat urine was identified as N-hydroxy OBFX, determined by comparison of the extraction of urine with chloroform with the standard compound of N-hydroxy OBFX, using LC/APCIMS. N-hydroxy OBFX had a weaker antibacterial activity against fluoroquinolone sensitive bacteria than the parent compound.

Distribution of uropathogenic virulence factors among Escherichia coli strains isolated from dogs and cats.

A variety of virulence factors (VFs) such as type 1 fimbriae, pilus associated with pyelonephritis, S fimbriae, afimbrial adhesin, alpha-hemolysin, aerobactin and cytotoxic necrotizing factor 1 are associated with uropathogenic Escherichia coli. In this study, 80 uropathogenic E. coli strains in 50 dogs and 30 cats suffering from UTI. In addition, 60 E. coli strains were isolated from fecal samples from 30 each of healthy dogs and cats. The distribution of VFs of uropathogenic E. coli strains isolated from dogs and cats suffering from urinary tract infections (UTI) were examined by the colony hybridization test with seven DNA probes specific for VFs, and the results were compared with those obtained in the studies on strains from humans with UTI. In uropathogenic E. coli strains isolated from dogs and cats suffering from UTI, VFs were detected as frequently as in the strains isolated from humans with UTI. Although less frequently, genes encoding these VFs especially pap, sfa, hly, and cnf 1 genes were also associated with E. coli strains isolated from feces of healthy cats, in contrast to the distribution pattern of uropathogenic E. coli observed in humans. Furthermore, all VFs except pil were significantly more frequently detected in strains isolated from urine of animals with cystitis than in those isolated from feces of healthy humans. These results indicate that VFs of E. coli contribute to the pathogenesis of UTI in dogs and cats.

Immunologic protection against canine heartworm infection.

This study was conducted to evaluate protective efficiency of three different protocols for vaccination in canine heartworm infection. To evaluate the three protocols of immunization, dogs were separately immunized with living larvae; 1) immunization with gamma-attenuated infective larvae, 2) with 50 micrograms/kg ivermectin-abbreviation, and 3) with chemical abbreviation plus Freund's complete adjuvant (FCA). Each group was composed of two dogs. All dogs used for this study were subcutaneously challenged with 100 intact third-stage larvae (L3) various days after the last immunization, and the worms in the pulmonary arteries and the right ventricle of the heart were recovered 17 to 25 weeks post-infection. The numbers and the sexes of the worms were determined. A mean of 38 worms was burdened in the group immunized with irradiated L3, 36 worms in the chemically-abbreviated group, but 15.5 worms in the group with chemical abbreviation plus FCA. The percentages of the protection in the former two groups were nearly 50%, but 72.3% in the group with ivermectin plus FCA. The adjuvant enhanced the protective immunity against L3 challenge. Obvious eosinophilia was observed in both immunized and control dogs except for two dogs. There was no correlation between the suppression of eosinophilia and the protective immunity in the present study.

Characteristics of nucleotide sequences flanking the trans-spliced leader SL1 exon in Dirofilaria immitis, Brugia malayi, and Brugia pahangi.

Nucleotide sequences surrounding the trans-spliced leader SL1 exon in the 5S rRNA gene spacer regions of Dirofilaria immitis, Brugia malayi, and B. pahangi were determined after PCR amplification, aligned with the genus Onchocerca for comparison, and used for the prediction of secondary structures. The nucleotide sequence of this region in B. pahangi was first shown in the present study. Hypothetical secondary structures of the spacer region suggested that the SL1 transcript is capable to form a stable stem-loop structure which may render transposition of the SL1 sequence to mRNA molecules. A homologous sequence to Sm-binding site was assigned on a bulge loop. No significant difference was observed in adult worms of D. immitis irrespective of sex or location. No difference was apparent between the two species in genus Brugia.

Differentiation of vaccine virus from field isolates of feline panleukopenia virus by polymerase chain reaction and restriction fragment length polymorphism analysis.

In an attempt to distinguish feline panleukopenia virus (FPLV) live vaccine strains from FPLV field isolates in Japan, we compared restriction fragment length polymorphisms (RFLP) of polymerase chain reaction (PCR)-amplified fragments of live FPLV vaccine strains with those of FPLV Japanese field isolates. On the basis of nucleotide sequence differences between PLI-IV, a live vaccine strain, and FPV-483, a recent field isolate, two restriction enzymes, Dra I and Afa I, were selected for PCR-RFLP analysis of nucleotide (nt) differences at nt 3695 and 4508, respectively. Three live vaccine strains including the PLI-IV strain could be distinguished from the Japanese field isolates by their PCR-RFLP patterns by Afa I, but one live vaccine strain was indistinguishable from the Japanese isolates when Dra I and Afa I were used. The Japanese field isolates were divided into two groups by the profile of PCR-RFLP patterns generated by Dra I and Afa I, suggesting that PCR-RFLP analysis using several enzymes provides a good genetic estimate of strain differentiation. No isolate that shows a Dra I-negative/Afa I-negative pattern has emerged in Japan, indicating the possibility that the live vaccine viruses with a Dra I-negative/Afa I-negative pattern, such as the PLI-Iv strain, are candidates for use as live FPLV vaccine strain in Japan where they can be genetically distinguished from field strains.

Assessment of the significance of virulence factors of uropathogenic Escherichia coli in experimental urinary tract infection in mice.

Four Escherichia coli strains, isolated from cystitis patients, belonging to serotype 02:H- and possessing different combinations of urovirulence factors were examined in an experimental pyelonephritis mouse model to assess the relative importance of virulence factors in causation of urinary tract infections (UTI). The results suggest not only that the each virulence factor has a role in causation of UTI but also that the presence of P fimbriae and production of hemolysin significantly reduced the LD50 and ID50 of the strains in the mouse model. The results also demonstrate that the presence of additional virulence factors acts in an additive or synergetic fashion enhancing the cumulative impact of the strain.

Detection and partial characterization of Pasteurella multocida found in feline skin lesions.

Pasteurella multocida was isolated from 21 of 105 purulent skin lesions in household cats. The bacterium was in pure culture in nine specimens and predominant in six specimens. Its viable counts were 10(2) to 10(7) colony forming units/ml. Of 21 isolates of P. multocida, seventeen were considered to be capsular type A. The predominant capsular and somatic type was the serotype A:3,4. Inoculation of the filter-sterilized supernatants of the isolates induced an erythematous response in guinea pig skins. These findings suggest that P. multocida is a candidate as a pathogen of feline skin lesions and the erythema-inducing activity of the bacterium may participate in the formation of skin lesions in household cats.

Two genomic variations in the E1 region of canine adenovirus type 2 strains.

We amplified the E1 region of canine adenovirus type 2 genomes by the polymerase chain reaction (PCR) and analyzed the PCR products by using eight restriction endonucleases. Restriction patterns of the E1 region cleaved with HaeIII and RsaI revealed two genomic variations among the canine adenovirus type 2 strains. Although the clinical significance of two distinct genotypes among the canine adenovirus type 2 strains is currently unknown, these genomic variations are well conserved among different strains in each genotype and suggest that the Japanese field strains, with reference to the E1 region, are different from the non-Japanese strains examined.

[Detection by PCR of genomic markers in canine parvovirus and feline panleukopenia virus]. / Détection par PCR de marqueurs génomiques dans le parvovirus canin et le virus de la panleucopénie féline.

The NS1 and VP1/VP2 genes of canine parvovirus and feline panleukopenia virus were amplified by the polymerase chain reaction (PCR). The restriction cleavage patterns of the amplified DNA fragments were compared among these parvoviruses including vaccine strains. Differences of the restriction site in the NS1 portions were observed between the vaccine strain and the wild type strain as well as between canine parvoviruses and feline panleukopenia viruses. The restriction patterns of feline panleukopenia viruses were distinct from those of canine parvoviruses, and showed differences between the vaccine strain and its wild type strain. This PCR-based restriction cleavage can be used for ecological study of such viruses.

Activity of AT-2266 compared with those of norfloxacin, pipemidic acid, nalidixic acid, and gentamicin against various experimental infections in mice.

AT-2266 (1-ethyl-6-fluoro-1,4-dihydro-4-oxo-7-(1-piperazinyl)-1, 8-naphthyridine-3-carboxylic acid) showed marked activity in vivo when administered orally to mice bearing systemic, pulmonary, dermal, or urinary tract infections due to variety of organisms. The activity of AT-2266 was uniformly higher than those of norfloxacin, pipemidic acid, and nalidixic acid against all of the infections. The activity of AT-2266 administered orally was almost comparable to that of gentamicin administered subcutaneously against urinary tract infections due to gram-negative organisms but was generally lower against other infections. AT-2266 exhibited significant activity against infections due to gentamicin-resistant and nalidixic acid-resistant organisms.

In vitro antibacterial properties of AT-2266, a new pyridonecarboxylic acid.

AT-2266, 1-ethyl-6-fluoro-1,4-dihydro-4-oxo-7-(1-piperazinyl)-1,8-naphthyridine-3 -carboxylic acid, is a new pyridonecarboxylic acid derivative with broad and potent antibacterial activity. It inhibited some gram-positive bacteria, such as staphylococci and Bacillus subtilis, and most gram-negative bacteria, including Serratia marcescens, Pseudomonas aeruginosa, Haemophilus influenzae, and Campylobacter jejuni, at concentrations of 0.1 to 0.78 microgram/ml, and most gram-positive bacteria, glucose-nonfermenters, and Mycoplasma pneumoniae at concentrations of 1.56 to 12.5 micrograms/ml. Most of the clinical isolates tested were as susceptible to AT-2266 as were laboratory strains. The antibacterial potency of AT-2266 was higher than those of pipemidic acid and nalidixic acid and similar to that of norfloxacin. AT-2266 was not cross-resistant with antibiotics and inhibited most highly nalidixic acid-resistant bacteria at concentrations of 1.56 to 3.13 micrograms/ml. Its activity was barely affected by the addition of horse serum or sodium cholate but weakened by lowering the medium pH or increasing the inoculum size. AT-2266 was bactericidal at concentrations near its minimal inhibitory concentrations. Frequencies of mutants resistant to 10 micrograms of AT-2266 per ml were lower than 4.0 x 10(-9).