Six species and six additional genovars are combined within the so-called Enterobacter cloacae complex, with one of them being the species Enterobacter hormaechei. In a recent population genetic study, two genetic clusters were found in close phylogenetic proximity to the genetic cluster of E. hormaechei. In order to prove the hypothesis that these three genetic clusters belong to the same species, we performed cross-hybridization experiments in microplates with DNAs of representatives of each genetic cluster. The close phylogenetic relationship among the clusters was reflected by their relatively low ⌬T m values, ranging from 0.3 to 4.8, confirming the hypothesis that the clusters are parts of the same species. These clusters can be distinguished from the other species of the E. cloacae complex, which have ⌬T m values of 5.6 to 10.3. Forty-eight E. hormaechei strains from the different genetic clusters were phenotypically characterized with 129 biochemical tests. In this way, E. hormaechei could be differentiated from the other species of the E. cloacae complex because it tests negative in the 3-hydroxy-butyrate test. The three genetic clusters of E. hormaechei could also be differentiated from each other by using phenotypic tests. Hence, we propose three new subspecies of E. hormaechei corresponding to genetic clusters VI, VII, and VIII of the E. cloacae complex. E. hormaechei subsp. hormaechei comb. The genus Enterobacter was first described by Hormaeche and Edwards (9). Since the transfer of Enterobacter agglomerans to the genus Pantoea (5), 14 species are included in the genus (1), including Enterobacter aerogenes, which is considered a homotypic synonym of Klebsiella mobilis because it has the same type strain (13). Around Enterobacter cloacae, six genetically related and phenotypically similar species have been combined within the so-called E. cloacae complex (7), i.e., E. cloacae, Enterobacter asburiae, Enterobacter dissolvens, Enterobacter hormaechei, Enterobacter kobei, and Enterobacter nimipressuralis. In addition to these species, at least six genetic clusters are phylogenetically delineated within the complex (7).Two of the most prominent clusters (clusters VI and VIII) are closely related to the species E. hormaechei, together forming the so-called E. hormaechei metacluster. They displayed 98 to 99% sequence identity in an analysis of three housekeeping genes and showed nearly identical restriction patterns of their ampC genes, which chromosomally code for a Bush class 1 beta-lactamase (7).E. hormaechei was first described on the basis of 23 isolates sent to the Centers for Disease Control and Prevention (Atlanta, Ga.) for identification. At that time, they could not be assigned to a species, since they were negative in D-sorbitol and melibiose tests and did not fit the biochemical profile of any established Enterobacter species. Later, this preselected set of isolates turned out to be genetically closely related to each other. The species E. hormaechei was proposed to be lactose, D-sorbitol, raffi...

The phenotypes of 153 strains belonging or related to the genus Bijidobacterium were studied. These organisms included 38 collection strains and 115 wild strains (41 strains of human origin, 56 strains of animal origin, and 18 strains obtained from rivers or sewage). Our phenotypic analysis revealed seven main groups that were subdivided into 20 subgroups. Seven subgroups contained no type or collection strain. Among the human strains, the type strains of BiJidobacterium pseudocatenulatum and B . catenulatum fell into group I, which contained the type strains of B. adolescentis (subgroup Ib), B . dentium (subgroup Ic), and B. angulatum (ungrouped). The type strain of B. breve belonged to subgroup IIIal, and the type strains of B . infantis and B . longum fell into subgroup IIIbl. Group VII comprised only wild strains that were isolated from human infant feces. Among the animal strains, group I1 consisted mainly of bifidobacteria that were isolated from pig feces and contained the type strains of B . suis (subgroup IIb), B . thermophilum (subgroup IIf), B . choerinum, and B. boum (ungrouped). Wild strains belonging to group V were isolated from pig, calf, cow, and chicken feces; this included the type strains of B . animalis (subgroup Va), B . magnum (subgroup Vb), B . pseudolongum, and B . globosum (subgroup Vc). The strains of human origin (groups I, 111, and VII) were well separated from the animal strains (groups 11, IV, and V). It was not surprising that the wild strains isolated from surface water or sewage were distributed in the animal groups as well as the human groups. Thus, bifidobacteria can be considered to be successful indicators of human or animal fecal pollution when they are correctly classified. The acidification patterns were not adequate to differentiate Bijidobacterium species, as determined previously by Mitsuoka

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A study was performed to evaluate a new manual miniaturized system, API Coryne (API-bioMerieux, Inc., La Balme les Grottes, France), in which conventional biochemical methods were used to identify 240 isolates of coryneform and related bacteria. A total of 40% of the isolates were excluded from the study because they could not be identified by conventional methods. Identifications of the 240 isolates obtained with API Coryne showed a 97.6% concordance with conventional methods (79% after 24 h of incubation, 21% after 48 h of incubation): 158 (65.8%) isolates were identified with no further testing, and extra testing was required for 76 (31.8%) isolates. In three (1.2%) cases, the organisms did not correspond to any key in the code book and could not be identified by the computer service of the manufacturer. Only three (1.2%) strains were misidentified. The system was shown to be reliable and rapid when compared with standard identification methods.

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API Listeria is a new 10-test strip for 24-h biochemical identification ofListeria isolates. With this commercial system, 85% of 646 Listeria strains, including atypical isolates selected for this study, were recognized at the species and subspecies level without a complementary test. A new test differentiates Listeria monocytogenes from L. innocua on the basis of the absence of arylamidase from the former. With this system, 97.7% (252 of 258) of the L. monocytogenes strains tested were correctly identified and differentiated from 99.4% (175 of 176) of the L. innocua strains also tested. Gram-positive bacteria other than Listeria spp. gave quite different biochemical patterns. This system considerably reduced the time needed for conventional identification, since results were available within 18 to 24 h.

Summary. Pyogenic streptococci of Lancefield group C or group G from human or animal sources were examined with a view to increasing the number of diagnostic tests useful for their differentiation. Human strains of group G produced L-prolyl-L-arginine aminopeptidase but isolates of Streptococcus equisimilis (group C) did not. Tests for a-L-glutamate aminopeptidase together with fermentation of glycogen or sorbitol distinguished S. dysgalactiae from strains of S. equisimilis isolated from animals. It was confirmed that fermentation tests were helpful in the study of S. equi and S. zooepidemicus and that enzyme reactions helped distinguish between S. canis and the human strains of group G.

A numerical analysis based on phenotypic characteristics (89 enzymatic tests and 49 carbohydrate acidification tests), in which experimental strips from Biomerieux-API, La Balme les Grottes, France, were used, was performed to characterize 82 new isolates belonging or related to Bijidobacterium longum, Bifidobacteriurn infantis, and Bafuiobucteriurn breve. A total of 72 strains were isolated from child or adult feces, and the other strains were obtained from human vaginas and bronchi. In this study we also included 38 type and reference strains that were representative of all species of the genus Bifidobucterium and 6 strains belonging to the genus LactobuciUus, DNA-DNA relationships between B. longurn and B. infantis were determined by using 19 strains related to these species, as determined by the numerical analysis, The degree of DNA binding was determined by the S1 nuclease method. The phenotypic study revealed that there were six main clusters, which were subdivided into nine subclusters. Subcluster Va contained the type strains of B. longum and B. infantis. The DNA-DNA relatedness values of some of the new isolates were very similar to the DNA-DNA relatedness values of the type strain of B. longurn. On the basis of these data, it was difficult to isolate B. infantis strains and then to define B. infantis as a single species separated from B. longurn. Subclusters lVb to IVf comprised reference strains of B. breve. Cluster I11 and subcluster Ia were not identified.At the species, present time, the genus Bifidobacterium includes 29 10 of which are of human origin. Four of these species, Bijidobacterium bijidum, Bijidobacterium longum, Bifidobacterium infantis, and Bifidobacterium breve, have been studied with increasing interest because of their role in physiological relationships in human (especially infant and child) gastrointestinal tracts (2,3,29). Furthermore, today, the use of these bacteria in food microbiology (dairy products) makes accurate differentiation of these organisms necessary (19).The type species of the genus, B. bifidum, is well separated from the other species on the basis of its phenotypic characteristics, as well as its DNA-DNA hybridization values. Identification of the three other species studied in this work is much more uncertain; in particular, distinction between B. longum and B. infantis is difficult.On the basis of the results of phenotypic studies, Reuter (18) defined and subdivided B. longum into two biovars, biovars a and b, and Mitsuoka (17)