Jan Ursing

Clinical strains of Acinetobacter classified by DNA-DNA hybridization.

A collection of Acinetobacter strains consisting of 168 consecutive clinical strains and 30 type and reference strains was studied by DNA‐DNA hybridization and a few phenotypic tests. The field strains could be allotted to 13 DNA groups. By means of reference strains ten of these could be identified with groups described by Bouvet & Grimont (1986), while three groups were new; they were given the numbers 13‐15. The type strain of A. radioresistens‐ recently described by Nishimura et al. (1988) ‐ was shown to be a member of DNA group 12, which comprised 31 clinical isolates. Of the 19 strains of A. junii, eight showed hemolytic activity on sheep and human blood agar and an additional four strains on human blood agar only. Strains of this species have previously been regarded as non‐hemolytic. Reciprocal DNA pairing data for the reference strains of the DNA groups were treated by UPGMA clustering. The reference strains for A. calcoaceticus, A. baumannii and for DNA groups 3 and 13 formed a cluster with about 70% relatedness within the cluster. Other DNA groups joined at levels below 60%.

Yersinia frederiksenii: A new species of enterobacteriaceae composed of rhamnose-positive strains (formerly called atypicalyersinia enterocolitica orYersinia enterocolitica-Like)

Yersinia frederiksenii sp. nov. is defined biochemically and genetically.Y. frederiksenii stains belong to three separate DNA relatedness groups, each of which is separable fromY. enterocolitica, Y. intermedia, Y. kristensenii, Y. pseudotuberculosis, andYersinia biotypes X1 and X2. The threeY. frederiksenii DNA relatedness groups, 6175, 2581-77, and 867, were represected by 10, 3, and 1 strain, respectively. All three groups were phenotypically similar. Pending additional study, it was decided to retain them all inY. frederiksenii. The positive rhamnose reaction separatesY. frederiksenii fromY. enterocolitica, Y. kristensenii, andYersinia biotype X1. A positive sucrose reaction distinguishesY. frederiksenii from the rhamnose-positive, sucrose-negativeYersinia biotype X2. Negative reactions for melibiose, raffinose, and α-methyl-d-glucoside distinguishY. frederiksenii fromY. intermedia. A negative melibiose reaction and positive reactions for ornithine decarboxylase, indole, sucrose, sorbose, sorbitol, inositol, and Voges-Proskauer separateY. frederiksenii fromY. pseudotuberculosis. Strain 6175 (=CIP 80-29) is proposed as the type strain forY. frederiksenii.

Genotypic and Phenotypic Diversity of Pseudomonas stutzeri

Summary A total of 49 strains of Pseudomonas stutzeri including reference strains from culture collections were phenotypically and genotypically studied. They were isolated from clinical sources, marine sediments, waste-water treatment plants and soil. DNA-DNA hybridization using the thermal melting point of the hybrids as parameter established seven genomic groups, four of them containing less than three strains. The G+C content ranged from 60.9–64,9 mol%. Phenotypical investigation using 102 characters demonstrated a considerable heterogeneity within the genomic groups. Because of the lack of useful differential tests there is still no sound basis for division of P. stutzeri at the species level. It is proposed that the term ‘genomovar’ be used to denote genomic groups of a nomenspecies.

ThermotolerantCampylobacter with no or weak catalase activity isolated from dogs

ThermotolerantCampylobacter strains isolated from dog feces were characterized by phenotypical tests, DNA base composition, and DNA-DNA-hybridization. Out of 98 strains, 63 were catalase negative or weakly reacting (CNW); they were found in diarrheic as well as in healthy dogs. The CNW strains were all nalidixic-acid sensitive, hippurate negative, and grew at 42°C but not at 25°C. Seven strains were further investigated. They were sensitive to 2,3,5-triphenyl-tetrazolium chloride, reduced nitrate, and produced H2S in the lead acetate test but not in triple-sugar-iron agar. The mol% G+C for five CNW isolates ranged from 35.2–35.8, which is higher than reported for any thermotolerantCampylobacter species. The strains also formed a well-delimitated DNA homology group with 80% or more intragroup relatedness and about 40% related toC. coli andC. jejuni.

Numerical Analysis of Cell Envelope Protein Profiles of Acinetobacter Strains Classified by DNA-DNA Hybridization

Summary Cell emvelope protein profiles, obtained by sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE), of Acinetobacter strains were compared with DNA hybridization data of the strains. The collection of strains comprised 98 field strains mostly from clinical sources in Sweden and the Netherlands and 22 strains from culture collections. Thirteen DNA groups were represented. Protein profiles were densitometrically recorded and subjected to numerical analysis. When the complete profile was used for analysis, the heavily stained bands outweighed the less heavily stained bands and the clustering did not correlate well with DNA groups. When the analysis was restricted to the upper part of the profile, consisting of minor bands of relatively high molecular weight, most strains belonging to the same DNA group clustered together. Thus, this part of the profile may help to identify strains at the DNA group level, while the complete profile can be used to differentiate strains within DNA groups.

A quantitative bacterial dot method for DNA-DNA hybridization and its correlation to the hydroxyapatite method

A filter hybridization method employing bacterial samples and [125I]labeled chromosomal DNA as a probe was used for DNA-DNA hybridization. It was found that the hybrids had a thermal melting temperature very similar to that of duplexes formed by purified filterbound DNA. The difference in thermal denaturation midpoint between homologous and heterologous duplexes was determined for a number of strains ofAcinetobacter spp. andEnterobacter agglomerans. A comparison with the corresponding data obtained by the hydroxyapatite method showed good correlation between the two methods. The use of bacterial samples in filter hybridization omits the time-consuming DNA preparation procedure necessary for traditional DNA-DNA hybridization procedures. A simplified, two-step elution procedure is suggested for processing large numbers of strains.

Characterization of Human Invasive Isolates of Listeria monocytogenes in Sweden 1986-2007

Since 1986, 68% of the Listeria monocytogenes isolates from human cases of invasive listeriosis in Sweden are available for retrospective studies. The aim of the present study was to characterize 601 human invasive isolates of L. monocytogenes in Sweden from 1986 to 2007 by using serotyping and pulsed-field gel electrophoresis. Since 1996, serovar 4b was permanently reduced to the second or third most common serovar in human cases in Sweden. During the latter period, 2000-2007, only 13% belonged to serovar 4b and 71% to 1/2a. The dendrogram, based on pulsovars, reveals two clusters with different serovars. Cluster 1 exhibits serovars 4b and 1/2b, whereas cluster 2 consists of serovar 1/2a. Serovar 1/2a seems to be more heterogeneous than serovar 4b.

Description of Campylobacter upsaliensis sp. nov. Previously Known as the CNW Group

Summary A new species, with the name Campylobacter upsaliensis , is proposed for catalase negative or weakly reacting campylobacters, which are thermotolerant, hippurate negative, nitrate positive, nitrite negative, unable to produce H 2 S in triple sugar iron agar and iron metabisulphite medium, intolerant to 1.5% NaCl and susceptible to nalidixic acid. G+C content of DNA is 35–36 mol%. The description is based on 37 human and canine C. upsaliensis strains identified by DNA-DNA hybridization (quantitative bacterial dot method). Campylobacter upsaliensis has been found in the feces of humans, dogs and cats with or without, signs of enteritis. It has also been found in blood from diseased humans with different clinical pictures including enteritis. The type strain is designated C231 T (= NCTC 11541 T ) and the G+C content of its DNA is 35 mol%.

Characterization of Listeria strains isolated from soft cheese.

Three soft cheeses were exposed to quantitative analysis for listeria and found to contain a large number of listeria. Thirty-five of the listeria strains isolated from the three cheeses were characterized by use of biochemical tests, serotyping, phagetyping and DNA restriction enzyme analysis. Seven isolates were identified as Listeria innocua and 28 as Listeria monocytogenes. Two to four different clones of L. monocytogenes could be identified from each cheese. In contrast, only one clone could be detected among the L. innocua isolates. From an epidemiological point of view the findings of different clones of L. monocytogenes in the same cheese emphasize the need for typing several listeria isolates from one and the same food sample. It is concluded that the best overview of the population of the listeria strains is obtained after direct plating of the sample followed by enumeration, isolation and extensive typing.

Genomic groups and biochemical profiles of clinical isolates of Enterobacter cloacae

A collection of 123 clinical strains presumptively identified as Enterobacter cloacae and 12 type and reference strains of Enterobacter spp. were genotypically investigated by a quantitative bacterial dot method for DNA‐DNA hybridization, giving an estimate of Δm (difference in thermal denaturation midpoint between homologous and heterologous duplexes). The API 20E system was used for phenotypic characterization. Using discontinuities in the values of Δm as criterion, five genomic groups of E. cloacae could be demonstrated, eleven ungrouped strains representing at least one additional group. Nine API profiles were found, the ideal phenotype of E. cloacae (i.e. the phenotype showing the most common reaction for the species in all tests studied) being the most frequently found. The type strain of E. cloacae and the reference strain CDC 1347–71 represented rather small genomic groups of three and eight strains respectively, most of them inositol positive. A majority of 98 isolates formed one single genomic group, biochemically dominated by the ideal phenotype. The genomic groups could not be differentiated phenotypically. At present there seems to be no reason for an attempt to split E. cloacae into two or more species. The type strain of E. dissolvens showed itself to be closely related to the type strain of E. cloacae (Δm 2.3°C), indicating that the two species may be regarded as subjective synonyms.