Downloaded from www.microbiologyresearch.org by IP: 54.146.78.28 On: Sat, 03 Dec 2016 07:56:00 Journal of General Microbiology (1986), 132, 1891-1 894. Printed in Great Britain 1891 Enzyme-linked Immunosorbent Assay (ELISA) as a Means of Taxonomic Analysis of Streptomyces and Related Organisms By R. KIRBY* AND E. P. RYBICKI Department of Microbiology, University of Cape Town, Rondebosch 7700, South Africa (Received 2 December 1985 ; revised 19 February 1986) Fourteen Streptomyces strains from various numerical taxonomic classes and representatives of three other genera of actinomycetes were studied using an indirect enzyme-linked immunosorbent assay (IND-ELISA) to determine their serological relationships. The IND- ELISA results agreed with those from previous numerical taxonomic analyses and Ouchterlony double-diffusion studies. The IND-ELISA method is quicker, more quantitative and less subjective than Ouchterlony assays and thus should be useful in Streptomyces taxonomy. The results indicated that Frankia sp- CpI 1 was related to Streptomyces. INTRODUCTION Three major approaches have been used to study the taxonomy of Streptomyces: morphology and pigment production, numerical taxonomy and nucleic acid hybridization. Identification of Streptomyces has generally relied on morphology and pigment production. This is subjective in many cases, especially where there is variation in the growth media used and the point in the growth cycle that examination takes place. Numerical taxonomy is more reliable as it uses a wide variety of criteria, but much work is needed to create a comprehensive data base. Nucleic acid hybridization (DNA-RNA or DNA-DNA) requires the use of radiochemicals, the careful preparation of nucleic acid and a high degree of control of hybridization conditions (Mordarski et al., 1981). In a few cases, serology has been used to study the taxonomy of Streptomyces and related species with some success (Ridell, 1981; Ridell & Williams, 1983). Serological techniques allow direct study of the proteins produced by the various Streptomyces and therefore should allow quantification of the divergence of these proteins between species. In previous studies using serological techniques, Ouchterlony double-diffusion assays have been used to measure the major protein similarities of Streptomyces species (Ridell & Williams, 1983). This approach has three disadvantages. Firstly, only the major soluble antigens can be studied and any minor components are ignored. Secondly, the assay is semiquantitative as only the number of major antigens detected is counted and not their degree of cross-reaction. Finally, the identification of precipitin lines and spurs is subjective and difficult to reproduce in a reaction involving a very complex antigen mixture such as that from total soluble bacterial cell contents. Moreover, the Ouchterlony technique is far less sensitive than indirect enzyme-linked immunosorbent assay (IND-ELISA), which has the added advantage of being usable for membrane bound or insoluble antigens as well as for soluble ones. IND-ELISA involves the coating of the plastic wells of an ELISA plate with the antigen to be tested, the binding of a known antibody to its specific antigen in the coated well and then detection of the binding by an enzyme-linked second antibody and a specific substrate reaction. The result is a direct quantitative analysis of the amount of antibody bound to the coated antigen. In this study we investigate the potential of IND-ELISA as a taxonomic tool. ’ Abbreviation : IND-ELISA, Indirect enzyme-linked immunosorbent assay. 0001-3128 0 1986 SGM