[Basic study of Beutin's washed sheep blood agar plate used for selective screening of verocytotoxin-producing/enterohemorrhagic Escherichia coli (VTEC/EHEC)].

Enterohemolysin production of Beutin's washed sheep blood agar plate has been used as an epidemiological marker for selective screening of Verocytotoxin-producing/Enterohemorrhagic Escherichia coli (VTEC/EHEC). In this study, we examined about the component of Beutin's washed sheep blood agar for further improvement of hemolysin production media. The following items were studied: the numbers of washings of sheep blood with phosphate buffered saline (PBS) (pH 7.2), concentration of sheep blood, kind and concentration of divalent metal ion (Ca2+, Mg2+) and basal medium. Twenty-seven strains of VT-producing E. coli of 7 different O-serotypes, 74 strains of VT-nonproducing E. coli of 24 different O-serotypes and one strain of O157 coded Escherichia hermanii were used for this basic study. In comparison of washing times of sheep blood with PBS, 5 times washing was better than 3 times, the original. In sheep blood concentration, supplement with 4% sheep blood was best for hemolysis observation. In experiment of addition of 2 divalent metal ions, Ca2+ and Mg2+, supplement with Ca2+ was more suitable than Mg2+ for hemolysis, and the supplement with 10 mM CaCl2, the original, was the best concentration. On the basal medium used in Beutin's sheep washed blood agar, 4 kinds of media were compared. In addition to Soybean-Casein Digest (SCD) agar, the original, Nutrient agar, Heart Infusion (HI) agar and Brain Heart Infusion (BHI) agar were examined, HI agar was the best blood agar base in the four media. As the above experimental results, the composition of the better Beutin's washed sheep blood agar may be summarize as follows: Heart Infusion agar 'Eiken' used for blood agar base, supplemented with 10 mM CaCl2 and 4% defibrinated sheep blood (Japan biosupp. Center) washed five times in phosphate-buffered saline, pH 7.2.

[Comparison of O-serotype distribution of Escherichia coli isolated from faecal specimens of patients with sporadic diarrhea and healthy persons and regional difference in Japan].

To investigate the isolation frequency of O-serotype of Escherichia coli, a total of 1,563 faecal specimens obtained from patients with sporadic diarrhea in Ishikawa between July 1997 and June 1998, were examined. As a result of O-serotyping of isolated strains using commercially E. coli antisera (43 different types), 247 strains of 29 different O-serotypes were isolated. Isolation rate was 15.8%. Most predominant O-serotype was O1 (128 strains, 52%), followed by O18 (26 strains, 11%), O6 (17 strains, 7%), O111 (16 strains, 6%), and these 4 different O-serotypes took up three quarters of the isolated E. coli. Between August 1996 and May 1997, E. coli isolation from faecal samples of 51,893 healthy persons and O-serotyping of isolated strains using commercial antisera to 6 predominant O-serotypes (O-26, 111, 114, 128, 157 and O1) of VTEC/EHEC were carried out. Among 6 O-serotypes, the most predominant O-serotype was O1 (93% of isolates), followed by O26, 111, 128 (6%) and O114, 157 (1%). These isolation frequencies in patients were 80%, 18%, 2%, respectively, have resembled each other in healthy persons in many points. In a similar way, of these distributions of O-serotype of strains hemolysed on Beutin's blood agar plates, we compared patients with healthy persons. Fifty-six strains (3.6% of the total) of E. coli of different O-serotypes were isolated from 1,563 patients and 57 strains (2.8% of the total) belonging to 11 serotypes from 2,036 healthy persons. As a result of O-serotype frequency, both groups resembled each other. O18 and O6, the most predominant O-serotypes, occupied 64% of the isolated strains in patients and 74% in healthy persons. Next in patients, O1, 26 were 7%--level, O28 ac, 152, 157 were 4%--level, respectively, and in healthy persons. O1 was 5%--level, O28 ac, 55, 146, 152 were 4%--level respectively. In the comparison of O-serotype frequency of E. coli isolated from sporadic diarrhea in other 5 areas (Kanto district, Tokyo, Oita, Aichi and Ishikawa), O1, 6, 8, 18, 25, 26, 55, 86a, 111, 125, 126, 127a, 128, 146, 148, 157 and 166 (17 types) have covered a wide area. On the other hand, O29, 44, 78, 112ac, 115, 136, 143, 152, 168 and 169 (10 types) have a tendency to distribute in local areas, we believe that there are regional differences even in the same Japanese territory.

[Selective screening of enterohemorrhagic Escherichia coli depending on hemolysis on Beutin's washed sheep blood agar plates].

Recently, Beutin et al have studied hemolysin production in a large number of serologically diverse verotoxin (VT) producing Escherichia coli (VTEC) strains and found a close association between hemolysin production and VT production. This study was to examine whether hemolysin production on sheep blood agar can be used as an epidemiological marker for microbiological screening of enterohemorrhagic Escherichia coli (EHEC). In comparison of hemolysis on washed blood agar plates prepared in our laboratory and commercially available blood agar, all VT producing strains exhibited a typical hemolytic phenotype on the former, on the other hand, VT non-producers were less hemolytic on the former than the latter. On the washed blood agar, 11 VT1 producing strains belonging to 4 different serotypes, 1 strain of VT2 producer, 5 VT1&2 producers belonging to 2 different serotypes, all the 17 strains of VTEC were positive for hemolysis. The 85 VT non-producing strains belonging to 18 different serotypes isolated from faecal specimens from patients with diarrhea were tested, 10 strains (12%) belonging to 4 different serotypes (O6, 18, 26, UT: untypable) produced hemolysin. Two-hundred four (10%) of the 2,036 strains isolated from faecal specimens from 507 healthy persons exhibited typical hemolysis. As a result of identification of these 204 hemolytic strains, 148 strains of E. coli 13 strains of Citrobacter and 10 strains of other enterobacteria, and 13 strains of Pseudomonas, 12 strains of Aeromonas, 5 strains of Proteus and 3 strains of Vibrio were detected. 148 hemolytic strains of E. coli were carried out O-serotyping with commercially available pathogenic E. coli antisera, 57 strains (39%) were classified into 11 different serotypes, but 91 strains were not typable. Most frequent serotype was O18 (24 strains), next O6 (18 strains), and other 8 different serotypes (O25, 26, 28ac, 29, 55, 146, 152, 159) were less than 2 strain respectively.

[The working adaptation in visual display terminal (VDT) workers].

The working adaptation in VDT workers was conducted by investigating from two different standpoints: one was to indicate an improvement of working conditions and environments around the VDT workers, evaluating the fundamental data of their visual physiological functions, and the other was to investigate the relation between their performances and production. The results were as followed: 1. Eye strain induced by VDT work was able to take an objective view as accommodative and convergent depression including the tear productive function, followed by making good working conditions. 2. The measurement of eye movement was able to analyze the response time and the response process for the informations indicating on the display, and the concept of recovery time for deteriorated accommodative function was useful to improve the display design. 3. It was suggested that the measurement of visual evoked potential was more simple method to evaluate the workers' attention. 4. The relation between performance and psychological stress and the time course analysis of performance were studied, so that it was given suggestions as to assign suitably the work management and the working design to each VDT worker.