Search

Between 1992 and 2007, a total of 86 isolates of Salmonella enterica Weltevreden were obtained from clinical human samples (n = 41), 45 farm animals and their environment on 20 farms, including poultry (n = 25), beef cattle (n = 5), swine (n = 5), dairy cattle (n = 3), mice (n = 2), pony (n = 1), fly (n = 1) and feed samples (n = 3), in Okinawa Prefecture, Japan. Only seven isolates (8.1%) of the isolates were resistance to one or more antimicrobial agents tested; six streptomycin (7.0%), six oxytetracycline (7.0%), two ampicillin (2.3%), two kanamycin, (2.3%), two chloramphenicol (2.3%), two suffamethoxazole/trimethoprim (2.3%), whereas all isolates were susceptible to gentamicin, sefuroxime, colistin, nalidixic acid, fosfomycin and ofloxacin. Drug resistance patterns showed six patterns; ABPC-SM-KM-OTC-CP-ST, ABPC-SM-ST, SM-KM-OTC, SM-OTC-CP, SM-OTC, OTC. Two ampicillin-resistant isolates harbored the blaTEM genes, streptomycin-resistant isolates (four aadA, two strA), tetracycline-resistant isolates (two tetA, three tetB), chloramphenicol-resistant isolates (two catA1), respectively.

Streptococcus suis, a major global porcine pathogen, is an emerging zoonosis in Southeast Asia that triggered a 2005 outbreak in China. S. suis causes meningitis, sepsis, and endocarditis in both pigs and humans and involves significant mortality. We report the case of a previously healthy 50-year-old dairy farmer who developed S. suis type 2 endocarditis complicated by pulmonary embolism and spondylitis. He experienced a high fever, chills, fatigue, and worsening low back pain in the 6 weeks prior to admission. On physical examination, he had lumbar spine tenderness and weakness of the left leg. Blood culture identified penicillin-sensitive S. suis type 2. Echocardiography showed vegetation on the tricuspid valve, and magnetic resonance imaging (MRI) showed signs of spondylitis. The man reported sudden chest pain several days after admission, which computed tomography (CT) showed what was diagnosed as a septic pulmonary embolism. He was treated with penicillin G for 4 weeks and gentamicin for the first 2 weeks, followed by 2 weeks of oral amoxicillin, after which his symptoms gradually improved. The infection source was probably his dairy herd, since calves often bit his fingers while feeding and S. suis was found in their oral mucus. Over 400 cases of human S. suis infection have been reported globally, but this is, to our knowledge, the first known case of bovine transmission. All of Japan's 8 other cases involved occupational swine exposure, 5 of whom had injuries to their fingers. This emerging situation should be made known to all possibly involved in unprotected direct contact with swine and cattle, particularly when the skin could be compromised by cuts or abrasions.

The Japanese Veterinary Antimicrobial Resistance Monitoring System (JVARM) was formed in 1999 in response to international concerns about the impact of antimicrobial resistance on public health. The aim of the present investigation was to provide insight into the occurrence of antimicrobial resistance among bacterial strains isolated from cattle, swine and poultry on farms across Japan. In this paper, we summarize a nationwide investigation (1999-2003) on antimicrobial susceptibility of the targeted bacterial species for the JVARM, namely Salmonella, Campylobacter, Enterococcus and Escherichia coli, isolated from the feces of healthy domestic animals. Minimum inhibitory concentrations (MICs) of 10-18 antimicrobial agents including the major generic groups were determined using an agar dilution method according to the international NCCLS guideline in principle. The outline of present results is as follows: (1) A total of 307 Salmonella, 3,225 Escherichia coli, 2,822 Enterococcus and 1,122 Campylobacter were isolated from healthy domestic animals and examined for antimicrobial susceptibility. (2) The isolates of all targeted bacterial species represent high rates of antimicrobial resistance to both oxytetracycline (OTC) and dihydrostreptomycin (DSM). (3) Among the isolates of Campylobacter and E. coli, the frequency of isolation of resistance to fluoroquinolones ranged from 14 to 24% and 2 to 3%, respectively. (4) Resistance to fluoroquinolones was not observed among the isolates of Salmonella or Enterococcus. Continious investigation at the nation level of antimicrobial resistance among the isolates from healthy domestic animals remains a high priority for the JVARM program. Furthermore, appropriate and prudent use of antimicrobials for diseased animals should be considered essential.

As a part of studies on the source of infection of Vero toxin-producing Escherichia coli (VTEC), O157:H7 strains isolated from human infectious enteritis between 1986 and 1995 and O157:H7 strains isolated from feces of milk cows between 2001 and 2003 were subjected to drug sensitivity test with drugs widely used as therapeutic drugs for various infectious diseases in humans and animals, and the following results were obtained. 1) Drug sensitivity tests with 20 drugs were performed in 52 strains derived human from diarrhea and 100 strains derived from milk cows, and resistance was noted in 115 strains (75.7%): 36 of the 52 human diarrhea-derived strains (69.2%) and 79 of the 100 milk cow-derived strains (79.0%). 2) The human diarrhea-derived strains and milk cow-derived strains were compared with regard to MIC90 of each drug. The antibacterial activity of the drugs was generally higher against the human diarrhea-derived strains than against the milk cow-derived strains. 3) In the 115 strains exhibiting resistance, the most frequent pattern of drug resistance was single drug resistance noted in 80 strains (68.4%), and multidrug resistance was noted in 35 strains (30.4%) consisting of 17 strains with resistance to 3 drugs, 14 strains with resistance to 2 drugs, and 2 strains each with resistance to 4 drugs and 5 drugs. More strains were multidrug-resistant in the milk cow-derived strains.

We examined enterohemolysin (Ehly) production, and detected the hlyA gene and the eaeA gene for the intestinal mucosal adherence factor intimin in 131 strains of human-derived verotoxin-producing Escherichia coli (VTEC) and 140 strains of livestock (cattle and swine) -derived VTEC to evaluate their hazards to humans. The hlyA gene was confirmed in 98.5% of human-derived, in 50.5% of cattle-derived, and in 10.3% of swine-derived VTEC strains. Ehly-positive rates were 96.2-97.7%, 45.9-55.0%, and 10.3-20.7% in human-, cattle-, and swine-derived VTEC strains, respectively. Thus, the positive rates differed among strains of different species origins. However, all 12 cattle-derived O157VTEC strains had hlyA, and were Ehly-positive. Although 97.7% of human-derived strains and all cattle-derived O157VTEC strains had eaeA, only 8.1% of cattle derived strains of serotypes other than O157 and 3.4% of swine-derived strains had eaeA. In human- and cattle-derived strains, the presence of eaeA was associated with Ehly: all eaeA-carrying strains had hlyA, and almost all of them were Ehly-positive. Cattle-derived eaeA-carrying strains accounted for 29.5-35.3% of Ehly-positive strains, compared to 100% in human-derived strains. Only 3-4% of Ehly-negative strains had eaeA, and none of the non-hlyA-carrying strains had eaeA. These findings suggest that 2 factors, eaeA and Ehly, serve as useful indicators for the evaluation of hazard to humans, and that Ehly is a useful indicator because cattle-derived Ehly-positive strains may have eaeA.

To elucidate the source and route of VTEC infection, we performed pulse field gel electrophoresis (PFGE) an 50 isolates from human diarrhea typed as serotypes O157, O111, and O26, which were very frequently isolated from patients with VTEC infection between 1986 and 1997, and 32 isolates from dairy cattle, a total of 82 isolates. The isolates were genetically analyzed based on the electrophoresis patterns of DNA, and a phylogenetic tree was prepared. The isolates were classified based on similarity > or = 89. The following results of the molecular epidemiological investigation were obtained. 1) Based on the electrophoresis patterns of DNA obtained by PFGE, 34 of the 49 O157 isolates (69.4%) were divided into groups 1-9, 15 of the 18 O111 isolates (83.3%) were divided into groups 1-3, and 12 of the 15 O26 isolates (80%) were divided into groups 1-3. Of the grouped isolates, group 8 of O157, groups 2 and 3 of O111, and group 3 of O26 included isolates from human diarrhea and dairy cattle, but the other groups included isolates from only one of the two sources. 2) With regard to regional investigation, groups 6 and 9 of O157 included human diarrhea-derived isolates from Yokohama and Ehime, and group 8 included a human diarrhea-derived isolate from Yokohama and a dairy cattle-derived isolate from Tokushima. Group 3 of O111 included a human diarrhea-derived isolate from Ehime and a dairy cattle-derived isolate from Hokkaido. Group 3 of O26 included human diarrhea-derived isolate from Ehime and dairy cattle-derived isolate from Sagamihara and Hokkaido. Since the above findings showed that although the frequency was low, isolates from human diarrhea and dairy cattle were included in the same groups, it was demonstrated that dairy cattle are closely related to the human infectious disease of the intestinal tract as a source of infection. However, classification using the PFGE method is difficult due to diversity of the electrophoresis pattern of DNA. It is necessary to investigate the classification by a combination of the PFGE method with phage typing, ribotyping, and RAPD-PCR, and to investigate more numbers of patient-derived and animal-derived isolates.

To clarify the source of infection and route of transmission of Verocytotoxin-producing Escherichia coli (VTEC) in humans, we collected fresh feces from healthy dairy cattle reared in Hokkaido, Fukushima, Kanagawa and Okinawa prefectures between June 1996 and March 1997, and attempted to isolate VTEC. The results are described below. 1) VTEC was isolated from 68 (27.1%) of 251 fecal samples tested. VTEC was isolated from 14 (28.0%) of 50 in Hokkaido, 13 (26.0%) of 50 in Fukushima, 20 (39.2%) of 51 in Kanagawa and 21 (21.0%) of 100 in Okinawa. There were no difference in the prevalence among the prefectures. 2) Toxin type and serotype of 85 isolates were determined. Thirty-three isolaties (38.8%) were classified into VT1 toxin and VT2 toxin, respectively, and 19 isolates (22.4%) were classified as the strain that produces both VT1 and VT2 toxins. The toxin types of these isolates were divided by serotypes. The VT1-producing isolates were the most frequent among O111:H-. The VT2-producing isolates included O2:H12, O2:H29, O2:H-, O82:H8, O82:HUT, O153:H19, O153:H42 and O153:H-. Among the isolates producing both VT1 and VT2 toxins, O153:H19 was relatively frequent. Based on findings that many bacterial strains coinciding with toxin types and serotypes of human-derived VTEC isolated from dairy cattle, it was suggested that dairy cattle are closely related to VTEC infection in human as a source of infection.

A familial outbreak of Verotoxin-producing Escherichia coli (VTEC) infection occurred in July 1996 in AKITA prefecture. Four VTEC strains harboring VT-1 and eaeA genes were isolated from three patients and a calf, breeding farm for which was located as close as 4 meters from the house where the patients lived in. All of the 4 VTEC isolates were serotyped as O63:H2 using commercially available sera kits. However, a patient isolate, EC-281, was serotyped as 0103:H2 at the International Escherichia and Klebsiella Centre. Titration and absorption tests using rabbit antisera raised against EC-281 confirmed that the serogroup of the remaining 3-VTEC isolates was also O103. Epidemiological characteristics including plasmid profile, antibiotic susceptibility patterns and pulsed-field gel electrophoresis patterns of the 4 VTEC isolates were completely the same, indicating that these isolates originated from a common source. These findings in conjunction with the results of epidemiological survey conducted by the Health Center suggested that a possible infectious source for this outbreak is the calf. Our present results strengthen the significance of calf as an infectious source of VTEC infection.

As part of the investigation on the source and route of infection with Verocytotoxin producing Escherichia coli (VTEC) in human beings, isolation of VTEC was attempted using fresh feces collected from healthy livestock (cattle, swine and goat) raised in Sagamihra, Yokohama and Hiratsuka cities from October, 1991 to March, 1992. VTEC could be isolated from 1 (1.0%) of 105 swine, 2 (3.6%) of 55 cattle and 2 (15.4%) of 13 goats. VTEC was isolated for the first time from goats in Japan. The combinations of the serotype and toxin type of the isolated strains were O116:H21 (VT2) and O163:H19 (VT2) for those isolated from the cattle, OUT:H19 (VT2vp) for that from the swine and OUT:H21 (VT1) for those from the goats. Since VTEC isolated from the cattle and goats were found to produce VT of the same serotypes as human VTEC, domestic animals were considered to be involved as a source of VTEC infection in human beings.

The vero cytotoxin (VT) is responsible for hemorrhagic colitis and hemolytic uremic syndrome. Polymerase chain reaction (PCR) was used to detect VT-producing coliform bacteria from dairy cattle. It was found that 39 (33.3%) of the 117 fecal samples examined were recognized with VT genes in BGLB enrichment broth by the PCR method (named BGLB-PCR). Of the VT-positive samples, 31 samples (26.5%) were found to have VT-producing Escherichia coli. Frequencies of isolation in younger cattle (under 5 months) were 31.3-32.9%. On the other hand, the PCR method using the bacterial suspension of some colonies from DHL selective isolation medium (named DHL-PCR), was used for 105 samples. The DHL-PCR was validated according to the number of colonies tested for detecting VTEC. When using E. coli strains which have been stored after isolation by the conventional culture method, the VT-producing strains found were 7 (10.3%) of the 68 isolates tested. The 101 out of the 108 VTEC strains from cattle were classified into 14 O groups. 4 O serogroups (O26, O111, O145, O157) from 60% of VTEC positive cattle, were also the most common in humans with diarrhea. All E. coli O157:H7 isolates failed to ferment after 48 hrs and to hydrolyze 4-methyl-umbelliferyl-beta-D-glucuronide (MUG). These results suggests that cattle may play an important role in human VTEC infections. The BGL B-PCR technique is usefull in ecological studies for VT-producing pathogens.

Two hundred and sixty-six piglets with diarrhea (from 4 farms), 73 healthy pregnant pig (from 2 farms), 27 calves with diarrhea (from 9 farms) and 47 healthy milk cows (from 1 farm) were examined for Vero-cytotoxin-producing Escherichia coli (VTEC), and 52, 11, 15 and 67 strains of VTEC were isolated from 17 piglets, 11 pregnant pigs, 6 calves and 23 milk cows, respectively. All VTEC strains from the piglets produced only VT2vp, while the strains from the healthy pigs did not produce VT2vp, but did VT1 and/or VT2. Most VTEC strains from calves and cows produced VT2vhb and some produced VT2 and VT1. Serotyping of the isolated strains showed that many strains from the piglets belonged either O139:H1, O141:H4 or O141:HUT, but the strains from the pigs were either R-form or O-untypable. Many strains from the calves and cows were serotyped into O116 or O113, but there were several R-form and O-untypable. From these results, it is suggested that VTEC strains, especially from the pregnant pigs, calves with diarrhea and healthy milk cows, which produced the same type of Verotoxins to that produced by human isolates, may become sources of human infections.

During a three-year period (1987 & 1990), a comprehensive attempt was made to isolate verocytotoxin-producing Escherichia coli (VTEC) from 96 bovine, 89 porcine, 67 canine and 18 feline cases suffering from gastroenteritis. VTEC were isolated from 11 cows and 1 cat. Bead-ELISA and oligonucteotide probes were used to type the verotoxins (VT) and it was found that the VTEC strains from cows produced not only the currently recognized VT1, VT2 and VT2vh but also produced two new VT's tentatively designated as VTx and VTy. The strain from the cat produced VTy. Serotyping of the strains revealed that some animal strains belonged to similar serotypes as those isolated from human patients with hemorrhagic colitis and hemolytic uremic syndrome. Cattle, especially cows, and domestic pets apparently are reservoirs of VTEC and probable sources of infection in our country as has been previously documented in Canada and USA.