Alexander Kimler
1962 Journal of Bacteriology  
TPNH Specific formed activity rnjmoles C. burnetii 94 0.109 NYS + heated C. burnetii 3 0.0063 * Isocitrate, 8.5 mM; MnCI2, 0.11 mM; TPN+, 0.54 mM; tris buffer, pH 7.5, 0.05 M; disrupted C. burnetii (LD50 10-4-8]), 0.72 mg N/ml; NYS + heated C. burnetii, 0.38 mg N/ml; final vol, 1.45 ml, incubated 15 min, 25 C. J. Biol. Chem. 193:405, 1951), employing a known reference sample of 6PG. The intermediate formation of 6-phospho-6-gluconolactone was shown by trapping the lactone in hydroxyla-mine
more » ... and Lipmann, J. Biol. Chem. 194:417, 1951). In two experiments 0.10 and 0.20 ,umoles of lactone were formed from 7 ,umoles of G6P. Disrupted C. burnetii preparations also oxidized isocitrate in the presence of TPN ( Table 2 ). The reaction was TPN-specific; diphosphopyridine nucleotide was not reduced when it replaced TPN in the incubation mixture. TPN reduction was linear, with A OD = 0.04/min. Rickettsial ability to oxidize G6P identifies an important energy source for the organisms. The oxidation of isocitrate is consistent with the demonstration of citrate synthesis by C. burnetii and with reports of rickettsial oxidation of other tricarboxylic acid cycle intermediates. In several recent studies on staphylococci, a number of personal observations were made which may prove useful in the clinical laboratory. In the performance of the coagulase test, fresh rabbit plasma gave higher positive results than dehydrated plasmas. The substitution of human plasma should be considered with caution, for Streitfeld, Sallman, and Shoelson (Nature 184:1665(Nature 184: , 1959 found that pooled gamma globulin inhibited coagulase production by staphylococci. Any new plasma, fresh or reconstituted, should be checked with known controls. One of the controls should be a weak coagulase producer or a strain which takes at least 24 hr to give a positive test. Approximately 80% of Staphylococcus aureus strains isolated from patients reduced triphenyltetrazolium chloride in 30 to 40 min. S. epidermidis took about twice as long. The medium used was tryptose agar (Difco), pH 7.2. After autoclaving, it was cooled to 45 to 50 C and the following added: Seitz-filtered 10% D-mannitol, final concentration 1 %, and a sterile (autoclaved) solution of triphenyltetrazolium chloride 1:350, final solution having a concentration of 1:35,000. S. aureus grew well on nutrient agar with a pH of 8.5, whereas other staphylococci were either completely inhibited or showed little growth. In correlating production of coagulase with fermentation of mannitol, a number of discrepancies were apparent. A small percentage of staphylococci, approximately 1%, were coagulase positive and did not ferment mannitol. About 8% were coagulase negative and fermented mannitol. When the coagulase test was used as the standard, most of the media currently available for the isolation and identification of staphylococci showed more "false positives" (coagulase negative) than "false negatives" (coagulase positive). A small percentage of coagulase-negative staphylococci, about 0.5 to 1.0%, gave positive results when tested with all of the following media: (i) mannitol broth; (ii) deoxyribonucleic acid agar (DiSalvo, Med. Technicians Bull., U.S.
doi:10.1128/jb.83.1.207-208.1962 fatcat:7rksilsx5rcr3gzhmy65ev3ora