Primary Isolation Using Guinea Pigs and Embryonated Eggs 



ADDENDUM 



REAGENTS FOR GIMENEZ STAIN 



1. Carbol basic fuchsin stock solution: 



a. 10% basic fuchsin in 95% ethanol 100 ml 



b. 4%. aqueous phenol 250 ml 



(10 ml phenol in :50 ml distilled H, O) 



c. distilled H.O 650 ml 



Mix well; incubate at 37° C for 48 h before use. 



2. Stock buffers: 



a. 0.2MNaH2PO4 2.84 gm in 100 ml distilled water 



b. 0.2MNa2HPO4 2.76 gm in 100 ml distilled water 



3. Buffer solution: 



a. 0.2MNaH2PO4 3.5 ml 



b. 0.2MNa2HPO4 15.5 ml 



c. distilled water 19.0 ml 



4. Carbol basic fuchsin working solution: 



a. Carbol basic fuchsin stock ^ 4 ml 



b. 0.1 M sodium phosphate buffer solution, pH 7.45 1 ml 



c. Filter immediately and again before each use. Remains suitable for use for about 48 h. 



5. Malachite green oxalite: 



0.8% solution in distilled water 



REFERENCES 



1. Cherry, W.B., B. Pittnian. P.P. Hams, G.A. Hebert, B.M. Thomason, L. Thacker, and R.E. Weaver. 1978. 

 Detection of Legionnaires" disease bacteria by direct immunofluorescent staining. J. Clin. Microbiol. 8:329- 

 338. 



2. Feeley, J.C. R.J. Gibson, G.W. Gorman, N.C. Langford, J.K. Rasheed, D.C. Mackel. and W.B. Baine. CYE 

 agar: A primary isolation medium tor the Legionnaires' disease bacterium, (in press). 



3. Feeley, J.C, G.W. Gorman, R.E. Weaver, D.C. Mackel, and W.H. Smith. 1978. Priinary isolation media for 

 the Legionnaires' disease bacterium. J. Clin. Microbiol. 8:320-325. 



4. Kaufmann, A., et al. Pontiac fever: Isolation of the etiologic agent, its mode of transmission, and its 

 relationship to the Legionnaires' disease bacterium, (in preparation). 



5. Lewis, v., L. Thacker, C.C. Shepard. and J.E. McDade. 1978. In vivo susceptibihty of the Legionnaires' 

 disease bacterium in ten antimicrobial agents. Antimicrob. Agents Chemotherap. 13:78-80. 



6. McDade, J.E., and C.C. Shepard. 1979. Virulent to avirulent conversion in the Legionnaires' disease bacte- 

 rium and its effect on isolation techniques. J. Infect. Disease. (In press). 



7. McDade, J.E., C.C. Shepard, D.W. Eraser. T.R. Tsai, M.A. Redus, W.R. Dowdle, and the laboratory investiga- 

 tion team. 1977. Legionnaires' disease: Isolation of a bacterium and demonstration of its role in other 

 respiratory disease. N. Engl. J. Med. 297: 1 197-1203, 



8. McKinney, R.M., B.M. Thomason. P.P. Harris, L. Thacker, K.R. Lewallen. H.W. Wilkinson, G.A. Hebert, and 

 C.W. Moss. 1978. Recognition of a new serogroup of the Legionnaires' disease bacterium. J. Clin. Microbiol. 

 9:103-107. 



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