Basic principles concerning the collection, transport, and processing of clinical specimens for the detection of Streptococcaceae are given. Identification of beta-hemolytic streptococci (S. pyogenes, S. agalactiae, Lancefield group C and G streptococci, S. anginosus) and enterococci is based on the careful observation of colony morphology and hemolytic pattern on sheep blood agar plates; subsequent genus or species confirmation is achieved by rather simple biochemical or enzymatic tests and by detection of streptococcal cell wall carbohydrate antigens (Lancefield grouping). Rapid antigen tests for the detection of group A and B streptococci directly from pharyngeal and vaginal swabs, respectively, are highly specific, thus allowing an immediate antibiotic therapy in patients with a positive test result. The reported sensitivities of these nonculture tests are too low to exclude streptococcal colonization or infection, however. The elucidation of the genetics of some major virulence factors of group A and B streptococci has contributed to knowledge of their association with disease, and molecular techniques have supplemented the traditional (mostly culture and serologic) methods for an improved understanding of the epidemiology and pathogenesis of streptococcal infections. Recently employed examples include the M protein gene typing of group A streptococci by oligonucleotide probes and the use of PCR assays for the detection of the genes encoding for the pyrogenic exotoxins. Restriction enzyme endonuclease digestions of bacterial DNA in association with DNA fragment separation by conventional or PFGE have been applied successfully to several species of Streptococcaceae (e.g., S. pyogenes, S. agalactiae). Enterococci are important pathogens in the hospital setting, exhibiting high morbidity and mortality rates in bacteremic patients with severe underlying disease. Molecular typing methods have clearly confirmed their potential to be nosocomially transmitted. E. faecalis and E. faecium still account for the majority of human infections, but some of the newer enterococcal species (at present 19 species are recognized) have been encountered as well. The definitive species identification of enterococci requires the performance of an array of biochemical tests. The increasing antimicrobial resistance of enterococci, including high-level resistance to penicillins and aminoglycosides and occasionally also to glycopeptides, has hampered standard therapeutic regimens. All enterococci isolated from serious infections should be tested for high-level gentamicin and streptomycin resistance by one of several methods evaluated; beta-lactamase production (primarily found in E. faecalis) is reliably detected by the nitrocefin test.(ABSTRACT TRUNCATED AT 400 WORDS)