The laboratory identification of gram-positive anaerobic cocci

A collection of 256 clinical strains and 40 reference strains of gram-positive anaerobic cocci (GPAC) was studied, to characterise the recognised species more fully and to define groups of strains which might correspond to previously undescribed species. The methods used were: gas-liquid chromatography (GLC) for the detection of volatile fatty acids (VFAs); determination of the pre-formed enzyme profile with a commercially available kit, ATB 32A; microscopic appearance; colonial morphology; and antibiotic sensitivity tests. Strains were placed in one of five VFA groups according to their GLC profile; 96% of strains were further assigned to 12 groups by their enzyme profile. There was less than 99% agreement between the two methods. Of 111 clinical strains in the VFA-negative group, 110 gave one of three distinct enzyme profiles corresponding to Peptostreptococcus magnus, P. micros and P. heliotrinreducens. The assignment of strains to groups based on their microscopic appearance and colonial morphology agreed well with groupings according to enzyme profile. Identification of butyrate-producing GPAC was unsatisfactory because it relied heavily on the enzyme profile; testing for indole production was of limited discriminative value. Most strains of P. asaccharolyticus and P. indolicus were very similar in enzyme profile, microscopic appearance and colonial morphology, but a sub-group of P. asaccharolyticus could be distinguished. A further indole-positive group corresponding to Hare group III was also noted. Strains of P. prevotii and P. tetradius were very similar, but easily distinguished from other butyrate-producing GPAC. However, 45% of the butyrate-producing cocci could not be assigned to recognised species; most of these were assigned to one of two new groups, the ADH group and the bGAL group, by their enzyme profile, microscopic appearance and smell. Four strains that produced a terminal VFA peak of isovaleric acid formed a new group designated 'ivoricus'. Reliable features for the identification of P. anaerobius were GLC (all GPAC that produced isocaproic acid were identified as P. anaerobius), enzyme profile and sensitivity to SPS. Two clinical strains that produced caproci acid were identified as Hare group VIII; they were distinguished from Peptococcus niger by their enzyme profile and colonial morphology. A phenotypic classification based on GLC and enzyme profile is presented, with a method for the identification of most strains of GPAC within 48 h of primary isolation.