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Adult rabbits were immunized with nine Achromobacter xylosoxidans strains by intravenous injection of Formalin-killed organisms. Antisera thus obtained were reciprocally titrated with the nine A. xylosoxidans strains, and seven sera were defined as serologically distinct. Three of nine antisera possessed one common antibody while also each having their own specific antibody. Ninety-five strains of A. xylosoxidans were examined for serotyping by a microtiter agglutination test with the nine antisera, and the strains were divided into seven serogroups. The distribution of the 95 strains among the serogroups was as follows: serogroup A, 15 strains; B, 17 strains; C, 43 strains; D, 2 strains; E, 4 strains; G, 8 strains; F, 2 strains. Four strains were not agglutinated with any of the antisera. Serogroups A, B, and F had the ability to reduce nitrate to nitrogen gas and to grow in heart infusion broth at 41 degrees C, although there were some exceptions. In contrast, most strains of serogroups C, D, and E could not produce nitrogen gas from nitrate, although they did produce nitrite. The strains of serogroup C could not grow at 41 degrees C, whereas those of serogroups D and E could. Thus, we concluded that serogroups A, B, and F referred to biotype IIIb described by Tatum, and serogroups C, D, and E referred to biotype IIIa. The serotyping of A. xylosoxidans may be useful for the analysis of nosocomial infections caused by these organisms.

Thirty-six Minitek (BBL Microbiology Systems, Cockeysville, Md.) tests were evaluated with 625 Kansai Medical University stock strains of 22 species and one group of nonfermentative gram-negative rods able to grow on ordinary peptone media. Among the 36 tests, 15 were selected because a clear-cut reaction was shown by all 625 Kansai Medical University strains. Of these 15 tests, 12 were further selected for routine use because they were regarded as useful for the identification of nonfermenters. The 12 tests were arranged into the following four groups: (i) lysine-arginine-ornithine, (ii) urea-ortho-nitrophenyl-beta-D-galactopyranoside-dextrose aerobic, (iii) maltose-xylose-starch, and (iv) esculin-nitrate reduction-indole. A new profile system for four digits, the Minitek Y-Y (Yabuuchi and Yamanaka) system, consisting of 64 numbers which represent each single species and 11 numbers which give two to four species, is herein proposed. The system was designed primarily for a less expensive identification of gram-negative rods already confirmed in a butt of either triple sugar iron or Kligler iron agar for their lack of ability to ferment dextrose. Among the 539 clinical isolates obtained from 3 hospitals, 511 strains identifiable by classical methods were also identified by the Minitek Y-Y system.

Two lactose-fermenting Salmonella typhi strains were isolated from bile and blood specimens of a typhoid fever patient who underwent a cholecystectomy due to cholelithiasis. One lactose-fermenting S. typhi strain was also isolated from a pus specimen which was obtained at the tip of the T-shaped tube withdrawn from the operative wound of the common bile duct of the patient. These three lactose-fermenting isolates: GIFU 11924 from bile, GIFU 11926 from pus, and GIFU 11927 from blood, were phenotypically identical to the type strain (GIFU 11801 = ATCC 19430 = NCTC 8385) of S. typhi, except that the three strains fermented lactose and failed to blacken the butt of Kligler iron agar or triple sugar iron agar medium. All three lactose-fermenting strains were resistant to chloramphenicol, ampicillin, sulfomethoxazole, trimethoprim, gentamicin, cephaloridine, and four other antimicrobial agents. The type strain was uniformly susceptible to these 10 drugs. The strain GIFU 11925, a lactose-negative dissociant from strain GIFU 11926, was also susceptible to these drugs, with the sole exception of chloramphenicol (minimal inhibitory concentration, 100 micrograms/ml).

Simple dot hybridization and fluorometric hybridization methods in microdilution wells were designed and established for rapid and routine genetic identification of viridans group streptococci. Reference DNA extracted from each strain of 24 reference Streptococcus species was fixed both on a nitrocellulose filter and in a microdilution well. A 1-ml portion of the bacterial suspension which matched the turbidity of McFarland no. 2 standard was prepared when a streptococcal strain was isolated. It was lysed with achromopeptidase, and the DNA was quickly labeled with photobiotin under a sunlamp for 15 min. Dot hybridization and fluorometric hybridization were then carried out between the labeled DNA of the unknown organism and 24 unlabeled reference DNAs. Hybridized fragments on a nitrocellulose filter were detected by using alkaline-phosphatase-conjugated streptavidin and analyzed with a color graphic analyzer. Hybridized fragments in microdilution wells were quantitatively detected by using an enzyme, streptavidin-conjugated beta-D-galactosidase, and a fluorogenic substrate, 4-methylumbelliferyl-beta-D-galactoside. Strains belonging to each genetically distinct species could be identified by this dot blot hybridization test. However, some clinical strains cross-hybridized with two or more reference species, and then they were difficult to differentiate by dot blot hybridization. In such a case, fluorometric identification provided reliable results because the fluorometric method was more quantitative than dot blot identification. By these methods, it was possible to determine species assignment within the viridans group.