A total of 175 clinical and non-clinical isolates of Klebsiella were subjected to numerical analysis of electrophoretic patterns of their soluble proteins. Of these strains, 160 clustered into five major groups, ten clustered in groups of two or three strains, and five strains remained ungrouped. Selected strains of the major groups were analyzed by DNA hybridization competition experiments. Almost all strains of the species K. pneumoniae, K. ozaenae and K. rhinoscleromatis occurred in three different electrophoretic groups. The high degree of hybridization obtained between DNA from these strains confirmed their genotypic relatedness and supported the classification proposed by Ørskov. Another group was designated as K. mobilis as its strains shared reassociation values of 88 to 100% with the type strain of K. mobilis. The majority of the remaining strains were clustered in three electrophoretic groups which correlated well with DNA hybridization and phenotypic studies. These were identified as K. oxytoca, K. planticola and K. terrigena.

We determined the taxonomic position of 17 strains of bacteria that were previously classified as “Klebsiella-like” strains, corresponding to group K of Gavini et al. [Ann. Microbiol. (Paris) 128B:45–59, 1977]. These strains were isolated from sewage, surface waters, drinking waters, and unpolluted soils. Deoxyribonucleic acid (DNA)-DNA hybridizations and a numerical analysis of electrophoretic protein patterns were used in this study. At least 82% of the group K strains investigated formed a tight protein electrophoretic cluster. The DNA homology values for all of the selected group K strains were higher than 73% and indicated the genetic homogeneity of this group. Two strains, CUETM 77-177 and CUETM 78-134, which were classified as group L strains by Gavini et al., were identified as true members of group K by the above-mentioned methods. The protein gel electrophoretic technique permitted distinction of group K strains from other species of the genus Klebsiella. DNA-DNA hybridization experiments revealed relatedness values of 40 to 71% between the reference strain of group K and the species Klebsiella mobilis, Klebsiella pneumoniae, Klebsiella oxytoca, and Klebsiella terrigena. Phenotypic characteristics, protein electrophoretic patterns, and the results of DNA-DNA hybridizations supported the individuality of group K, and we propose the name Klebsiella trevisanii sp. nov. for the strains of this group. These strains were positive in the Voges-Proskauer, urease, and β-galactosidase tests, grew at 4 and 41°, utilized histamine as a carbon source, did not ferment melezitose, and did not use m-hydroxybenzoate; indole was produced by 42% of the isolates. Strain CIP 81-36 (= CUETM 78-120) was designated the type strain of this species.

We propose to maintain the genera Buttiauxella and Kluyvera, which are phenotypically similar, as separate genera in the Enterobacteriaceae. This separation is supported by the following findings: (i) strains of Kluyvera ascorbata and Kluyvera cryocrescens were related to Buttiauxella agrestis ATCC 33320T (T = type strain) at levels of 32 to 36% and 30 to 31%, respectively, as determined by deoxyribonucleic acid relatedness (nitrocellulose filter method at 52.8°C); (ii) the guanine-plus-cytosine ratios of Buttiauxella and Kluyvera deoxyribonucleic acids 48 to 50 mol% and 55 to 57 mol%, respectively; and (iii) production of indole and lysine decarboxylase and fermentation of sucrose in 2 days could differentiate Kluyvera from Buttiauxella.

Deoxyribonucleic acid (DNA)-DNA hybridization and numerical analysis of electrophoretic protein patterns were carried out to determine the taxonomic position of 37 “Klebsiella pneumoniae-like” strains from nonclinical origin (soil and water) and previously studied by numerical taxonomy (group L of Gavini et al., Ann. Microbiol. [Inst. Pasteur] 128B:45-59, 1977). The DNA interrelatedness for 20 selected strains of group L was at least 87%, except for two strains (63 and 69%). Protein electrophoretograms indicated likewise that at least 85% of the L strains constitute a genetically homogeneous group. The latter technique enabled the distinction of strains of group L from those of K. pneumoniae, K. oxytoca, and an unnamed Klebsiella cluster (group K) related to K. pneumoniae and K. oxytoca. On the basis of their phenotypic characters, protein electrophoretic patterns, and DNA-DNA relatedness, we propose for these strains the name Klebsiella terrigena sp. nov., with strain CIP 80-07 (= CUETM 77-176) as the type strain. The DNA of this strain was found to be 49 to 64% homologous to K. oxytoca DNA, 48 to 63% to K. pneumoniae DNA, and 51% to K. ozaenae DNA.

In this communication we report on the taxonomic position of strains previously alloted to group F by numerical phenotypic analysis (Gavini et al., 1976b) according to DNA-DNA hybridization studies. This group comprising 28 strains isolated from drinking water and soil is related to the genus Citrobacter on the basis of the IMViC tests. It differs from this taxon with respect to the numerical analysis of its phenotype and also in its G + C mol % of DNA (Ferragut et al., 1978a).

Twenty strains of Enterobacteriaceae isolated from water are assigned to a new species in the genus Serratia on the basis of phenetic (numerical analysis) and genetic (deoxyribonucleic acid [DNA]-DNA hybridization) evidence. They are closely related to each other (75 to 91% DNA-DNA relatedness) and show 50% DNA relatedness with species of Serratia but only 20 to 30% relatedness with other members of the family Enterobacteriaceae. The strains are acetoin, gelatinase, and deoxyribonuclease negative, but lysine, ornithine decarboxylase, and Tween-esterase positive. They are metabolically very active, fermenting all the sugars and alcohols tested except melezitose, inulin and sorbose. The name Serratia fonticola is proposed for these strains. Strain 11 (= ATCC 29844) is the type strain of the species.

This study is a measurement of the moles percent guanine plus cytosine of 34 strains of bacteria belonging or related to the genus Enterobacter. All of the strains included in the sampling have been previously studied by numerical taxonomy, which has shown four new classes: H1, H2, H3, and d. The average guanine-plus-cytosine contents for the defined species and of the new classes are: Enterobacter cloacae (10 strains), 54.5 mol% (standard deviation, 1.32); Hafnia alvei (5 strains), 48.1 mol% (standard deviation, 1.0); Enterobacter agglomerans (syn. Erwinia herbicola) (1 strain), 52.4 mol% (standard deviation, 0.41); Serratia liquefaciens (1 strain), 50.9 mol% (standard deviation, 1.22); Enterobacter aerogenes (1 strain), 53.5 mol% (standard deviation, 0.29); class H1 (5 strains), 52.2 mol% (standard deviation, 1.3); class H2 (5 strains), 53.4 mol% (standard deviation, 1.9); class H3 (5 strains), 54.2 mol% (standard deviation, 0.1); class d (1 strain), 52.6 mol% (standard deviation, 1.45). The importance of the guanine-plus-cytosine contents for discriminating defined species and new classes is discussed.

The deoxyribonculeic acid (DNA) of 106 strains of Enterobacteria was analysed for the guanine + cytosine (GC) content. These strains, whose origin and principal characters are described in the text, belong to the genera Citrobacter (C. freundii H2S-) and Levinea (L. malonatica and L. amalonatica). Four other groups or classes named C.D.E. and F. could not be classified on the base of the usual phenotypic criteria. DNA from the strains of Levinea has a GC% of 50.3 to 53.3, while DNA from the strains of C. freundii H2S- has a GC% of 48.6 to 51.7. The representative values from the new classes are C, 50.9%; D, 54%; E, 52.7%; F, 49.5%. For the latter a genomic heterogeneity was shown, expressing itself as two subpopulations whose average GC% are 51.7 and 48.6 respectively. Statistical analysis of the averages give a significant individuality to these new classes.

Melting point curves of DNA are used to calculate average GC content. Four methods of Tm determination are described and are used in the case of all DNA samples studied. The results presented confirme that thermal denaturation curves are asymetrical, and this limits the use of "normal probability paper" and "regression lines" methods. Best reproductibility is obtained by a study of the function (see article) followed by a linear interpolation of the median. This approach yields at the same time interesting information about bacterial DNA. The graphic determination of Tm appears to be the method for choice for routine analysis.

This work studies the classification, by numerical procedure, of 122 strains belonging or related to the genus Klebsiella. Four classes can be defined: J (= K. pneumoniae: 29 strains for the most isolated from human faeces), K (related to K. pneumoniae and to K. oxytoca: 24 strains isolated from water), L (related to K. pneumoniae although, tetrathionate-reductase+, m-hydrobenzoate+: 37 strains isolated from water or soil), M (= K. oxytoca: 22 strains for the most isolated from water). The importance of the origins of the strains is emphasized.

This study is a measurement of the moles percent guanine plus cytosine of 34 strains of bacteria belonging or related to the genus Enterobacter. All of the strains included in the sampling have been previously studied by numerical taxonomy, which has shown four new classes: HI, HZ, HB, and d. The average guanine-plus-cytosine contents for the defined species and of the new classes are: Enterobacter cloacae (10 strains), 54.5 mol% (standard deviation, 1.32); Hafnia aluei (5 strains), 48.1 mol% (standard deviation, 1.0); Enterobacter agglomerans (syn. Erwinia herbicola) (1 strain), 52.4 mol% (standard deviation, 0.41); Serratia liquefaciens (1 strain), 50.9 mol% (standard deviation, 1.22); Enterobacter aerogenes (1 strain), 53.5 mol% (standard deviation, 0.29); class H1 (5 strains), 52.2 mol% (standard deviation, 1.3); class Hz (5 strains), 53.4 mol% (standard deviation, 1.9); class H3 (5 strains), 54.2 mol% (standard deviation, 0.1); class d (1 strain), 52.6 mol% (standard deviation, 1.45). The importance of the guanine-plus- cytosine contents for discriminating defined species and new classes is discussed.