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S Löfdahl

Publications and source records attributed to S Löfdahl.

17 recordsLinked to original sources
PubMed1992

Comparison of the amino acid sequences of nine different serotypes of hepatitis B surface antigen and genomic classification of the corresponding hepatitis B virus strains.

The surface (S) genes of 12 hepatitis B viruses (HBVs) encoding nine different serotypes of hepatitis B surface antigen (HBsAg) were amplified by the polymerase chain reaction and sequenced. These represented the eight strains of HBV, P1 to P8, defined at an international workshop on HBsAg subtypes in Paris in 1975, and the adrq- subtype. The S genes from additional HBV strains, one ayw4, one adw4 and one ayw1, of sub-Saharan African origin, were also sequenced. The relationship of these 12 new S gene sequences to those of the 20 published previously was investigated by constructing a phylogenetic tree, which confirmed a previous classification into four groups, designated A to D, based on 18 complete HBV genomes. When relating our sequenced S genes to these genomic groups, ayw1 of African origin and P6 (adw2) were both allocated to group A, the reference P1 (ayw1 of Vietnamese origin) was allocated to group B, P5 (ayr), P8 (adr) and adrq- were all related to group C, and P2 (ayw2) and P3 (ayw3) could both be allocated to group D. Interestingly, the S genes of w4 serotype viruses, i.e. P4 (ayw4) and P7 (adw4q-), differed by 4% or more from both previous groups and from each other, suggesting their classification into two new groups, for which the designations E and F are proposed. Genomes specifying ayw were also found in groups A and B; previously sequenced genomes specifying the ayw subtype have all been confined to group D. There were indications that the epitope for subdeterminants of w resided at amino acid positions 125 to 127. Thus, at positions 125 and 127, ayw1, ayw2 and adw2 had T and P residues, respectively, whereas M and T residues were at the corresponding positions of ayw3. Both ayw4 and adw4 had L at residue 127, and all strains expressing r, apart from P5, had an I instead of a T residue at position 126.

Amino Acid Sequence↗
PubMed1992

Monoclonal antibodies specific for elongation factor Tu and complete nucleotide sequence of the tuf gene in Mycobacterium tuberculosis.

Monoclonal antibodies against mycobacterial antigens were produced by immunizing LOU/C rats with live Mycobacterium bovis BCG. The antibodies were characterized by an enzyme-linked immunosorbent assay and by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by Western blotting (immunoblotting). One antibody, MAMB 2, reactive with a 47-kDa protein was used to screen a lambda gt11 M. tuberculosis gene library (R. A. Young, B. R. Bloom, C. M. Grosskinsky, J. Ivanji, D. Thomas, and R. W. Davis, Proc. Natl. Acad. Sci. USA 82:2583-2587, 1985). Three recombinant phages reactive with MAMB 2 in plaque lysates were isolated, and part of the insert was sequenced. The mycobacterial inserts were all expressed as proteins fused with beta-galactosidase when the phages were induced as lysogens in Escherichia coli. The entire M. tuberculosis tuf gene was obtained by screening the lambda gt11 library with a DNA probe specific for the primary clones. A phage isolated from this screening was able to express the native protein in E. coli when introduced as a lysogen. A comparison of the entire gene sequence and the deduced protein sequence with the EMBL DNA and Swiss-Prot protein data libraries revealed strong homologies with elongation factors of bacteria, yeast mitochondria, and a plant chloroplast.

Amino Acid Sequence↗
PubMed1990

DNA hybridization for diagnosis of pertussis.

The aim of the present study was to evaluate a mixed phase DNA hybridization assay for detection of Bordetella pertussis and Bordetella parapertussis in nasopharyngeal aspirates from patients with suspected pertussis. Among 179 consecutive patients with own or parental suspicion of pertussis, the diagnosis was confirmed in 103 patients by serology and in 52/103 (50%) cases also by culture. The remaining 76 patients served as nonpertussis controls. Direct hybridization was positive in 38% samples with serology as reference method, a non-significant difference to the 50% sensitivity for culture. Preculture of samples on membranes for 24, 48 and 72 h gave a significantly higher sensitivity only with 72 h preculture, 69% vs 50% (P = 0.007). The 72h preculture gave, however, also a significant decrease of specificity, 87% vs 100% for routine culture (P = 0.001) and is not a more rapid diagnostic method. The result shows that rapid diagnosis by DNA hybridization can be achieved in a large proportion of pertussis cases. The presence of smaller numbers of bacteria in samples only positive after preculture indicate that DNA hybridization could be a highly sensitive diagnostic method with further development of more rapid amplification systems.

Bordetella↗
PubMed1990

A DNA hybridization test for detection of Bordetella in nasopharyngeal specimens.

A cloned Bam H1 fragment of genomic Bordetella pertussis DNA which recognized a frequently repeated sequence in the genome of B. pertussis was used as a probe in a DNA hybridization assay for the detection of Bordetella. Extensive studies on cross-reactivity were carried out in standardized strains and in cultured swab specimens from patients without suspected pertussis. Hybridizations of cultured clinical specimens from 142 patients with suspected pertussis were in complete agreement with the standard identification methods. The recovery rate of B. pertussis from nasopharyngeal swabs was less than 50%. Therefore the possibility to detect low numbers of B. pertussis in solution (nasopharyngeal aspirates) was investigated. The detection limit of direct hybridization by dot-blot technique was 5 x 10(3)-10(4) B. pertussis. Culturing bacteria on membranes placed on agar plates prior to hybridization showed that the detection limit could be lowered to 10(4), 10(2), and 10(1) cfu after 1, 2 and 3 days' culture, respectively. DNA hybridization under these conditions was found to be sufficiently sensitive and specific for further evaluation in clinical specimens for diagnosis of pertussis.

Bacteriological Techniques↗
PubMed1990

Drug-resistant coagulase-negative skin staphylococci. Evaluation of four marker systems and epidemiology in an orthopaedic ward.

Drug-resistant coagulase-negative staphylococci (DRCNS) in orthopaedic patients and ward staff were studied. A significant increase in the DRCNS carriage rate was observed among the 16 patients studied after 14 days of hospitalization with levels approaching that of the staff. Patients receiving dicloxacillin prophylaxis (n = 9) were more likely to be colonized with methicillin-resistant CNS, while patients receiving no antibiotics (n = 7) became to a larger extent colonized with multiple DRCNS. The combined data from species determination, biochemical, plasmid, and antibiogram typing revealed a considerable diversity among DRCNS; 64 types were distinguished among 112 DRCNS isolates selected for study after exclusion of apparently duplicate isolates. Plasmid plus antibiogram typing yielded almost as many types (61); whereas species determination plus antibiogram distinguished only 33 types. Although a novel computerized 96-reaction biotyping method alone enabled differentiation of 17 biotypes, most DRCNS isolates belonged to one of three major biotypes limiting the usefulness of this method. Ten of the 64 (16%) DRCNS types identified comprised 50 of the 112 (45%) isolates. These were isolated from staff and from patients on day 14, suggesting a nosocomial origin.

Adult↗
PubMed1989

Identification and nucleotide sequence of the delta-lysin gene, hld, adjacent to the accessory gene regulator (agr) of Staphylococcus aureus.

A Tn551 insertional mutation in the accessory gene regulator (agr) locus of the Staphylococcus aureus chromosome resulted in the decreased production of at least seven extracellular toxins and enzymes and a simultaneous increase in the production of protein A and coagulase (Recsei et al. 1986). Adjacent to this locus we have now identified another gene, hld, transcribed into a 0.5 kb RNA which codes for the staphylococcal delta-lysin. The expression of hld was totally repressed in a strain carrying the agr insertional mutation. Hybridization with strand-specific probes and primer extension analysis revealed that hld and agr are transcribed in opposite directions, starting 188 nucleotides apart. The hld gene is mainly expressed during the post-exponential growth phase and is totally repressed during early exponential growth. Determination of hld mRNA half-life in different growth phases indicated that this regulation is at the level of transcription.

Amino Acid Sequence↗
PubMed1989

Identification of a DNA fragment in the genome of Bordetella pertussis carrying repeated DNA sequences also present in other Bordetella species.

A DNA fragment (3.8 kbp) which hybridized to repeated sequences in the genome of Bordetella species has been cloned from Bordetella pertussis chromosomal DNA. Eleven subclones were constructed from this fragment. They exhibited distinct inter-species hybridization patterns in genomic blots of each of the Bordetella used in the study. One subclone revealed intraspecies variability among B. pertussis strains and another did not hybridize to B. parapertussis. The 3.8 kbp DNA fragment possesses a middle sequence surrounded by repeated sequences organized in an opposite symmetrical orientation. The external inverted repeats of it hybridized to a 680 bp DNA sequence which is located about 800 bp upstream of the pertussis toxin operon. The novel structural organization of the 3.8 kbp fragment suggests the possibility that this DNA segment is an IS-like element which may have an important function in the expression of virulence determinants in Bordetella bacteria.

Blotting, Southern↗
PubMed1988

Cloning of a chromosomal locus (exp) which regulates the expression of several exoprotein genes in Staphylococcus aureus.

Insertion of the erythromycin resistance transposon Tn551 into a single site of the Staphylococcus aureus chromosome resulted in decreased production of alpha-toxin, serine and metallo-proteinases and several other extracellular proteins and a simultaneous increase in the production of protein A. The site of insertion, designated exp, was separate from the structural gene for alpha-toxin and protein A. Hybridization analysis showed that the effect of the insertional mutation on the expression of the alpha-toxin and protein A was at the level of transcription. The chromosomal DNA flanking the transposon and the corresponding DNA of the wild-type strain was cloned in Escherichia coli. Northern blot hybridization experiments revealed that the exp locus codes for a major RNA of approximately 3.5 kb. This RNA was not found in the insertional mutant nor in a spontaneous exp mutant. A map of the exp locus constructed by Northern blot and restriction enzyme analysis showed that the insertional mutation was located in the middle of the coding sequence of the 3.5 kb RNA. The insertional mutant was reverted to wild type by inserting a recombinant plasmid containing most of the coding sequence of the 3.5 kb RNA.

Bacterial Proteins↗
PubMed1988

Epidemiological typing of Corynebacterium diphtheriae isolated in Sweden 1984-1986.

From September 1984 to December 1986 121 cases of culture-positive diphtheria were reported to the National Bacteriological Laboratory in Stockholm. Toxigenic Corynebacterium diphtheriae nongravis was isolated from all but one of 33 patients with disease and from 69 healthy carriers. 63/65 toxigenic isolates, available for epidemiological typing, had the same phage type, 20, and the same restriction enzyme pattern, RE2. This included strains isolated both from patients inside and outside of the traditional risk groups of people abusing alcohol and drugs. Non-toxigenic strains gave different phage types and RE patterns.

Bacteriophage Typing↗
PubMed1987

Expression and secretion of pertussis toxin subunit S1 in Bacillus subtilis.

Pertussis toxin (PT) is an important virulence determinant of Bordetella pertussis and one of the major protective antigens against whooping cough. The genes coding for PT have recently been cloned, but attempts to express them in Escherichia coli have been unsuccessful. We therefore explored the possibility of expressing these genes in Bacilius subtilis for which efficient vectors are available. The lack of endotoxin in the Gram-positive Bacillus might be an additional advantage for the production of a vaccine component. A DNA fragment coding for S1, one of the subunits of pertussis toxin, was inserted into an alpha-amylase secretion vector and the recombinant plasmid was introduced into B. subtilis. This resulted in high expression of S1, most of which was secreted and therefore found in the culture supernatant. This supernatant had ADP-ribosylating activity similar to that of PT. Western blot with antiserum to B. pertussis holotoxin showed several proteins ranging in size from 28 kDa to 20 kDa reacting in specific manner. About 10% of the protein recognized by the antiserum was of the size expected for native-size S1. The total amount of S1 proteins (full size and truncated) in the culture supernatant was about 100 mg/l. S1 protein made in B. subtilis was partially purified using chromatography with P-cellulose and Blue Sepharose. This preparation was used to immunize rabbits; the immune serum thus obtained recognized subunit S1 of native pertussis toxin.

Amino Acid Sequence↗
PubMed1978

A vector for recombinant DNA in Staphylococcus aureus.

Staphylococcal plasmids pS194 and pSC194 which confer streptomycin and streptomycin-chloramphenicol resistance respectively have been used as vectors for construction of recombinant DNA, since they each carry one single recipient site for endonuclease EcoRI. Hybrid DNA does not express streptomycin resistance, a marker which is present in both vectors, presumably because the marker gene is cleaved by EcoRI. A chloramphenicol marker present in pSC194 was used for positive hybrid selection. Hybrid plasmids generated by joining pSC194 with one or more of the four EcoRI fragments of the large (18.1-10(6) daltons) staphylococcal plasmid pI258 were constructed and permitted us to develop a physical map for pI258.

Chloramphenicol↗
PubMed1978

Biological characteristics of a type I restriction-modification system in Staphylococcus aureus.

Two restriction-modification systems, S1 and S2, are present in Staphylococcus aureus RN450 (S. Iordanescu and M. Surdeanu, J. Gen. Microbiol., 96:277-281, 1976). System S2 affects phage multiplication after both infection and transfection. Unmodified plasmid and chromosomal DNAs are also not expressed following transduction and transformation into a restrictive host. Restricted phages are, however, capable of conferring phage-mediated competence, although the state of competence does not affect the restriction-modification system. The restricting activity of system S2 is inactivated by heat treatment of the cells. An enzymatic activity that restricts unmodified phage DNA in the presence of ATP, Mg2+, and S-adenosylmethionine was recovered from cell-free extracts of a strain RN450 derivative.

DNA Restriction Enzymes↗
PubMed1978

Characterization of small plasmids from Staphylococcus aureus.

Small molecular weight plasmids from Staphylococcus aureus were characterized with respect to size, restriction enzyme cleavage pattern and transforming capacity. The plasmids pS194 and pC194 which encode streptomycin and chloramphenicol resistance respectively contained 3.0 and 2.0 megadaltons of DNA as determined by zonal rate centrifugation and electron-microscopy. Both plasmids transformed S. aureus wigh high efficiency. Plasmid pC194 contained only one cleavage site for endonuclease HindIII and pS194 contained single cleavage sites for HindIII and EcoRI. A natural recombinant between these two plasmids, pSC194, shared the high transforming capacity of the parental plasmids and contained one EcoRI site and two HindIII sites. pSC194 DNA also transformed B. subtilis with high efficiency. The recombinant plasmid pSC194 may be used as an EcoRI vector for construction and propagation of hybrid DNA in S. aureus as shown in the following paper (Löfdahl et al., 1978).

Bacillus subtilis↗
PubMed1975

Transformation reveals a chromosomal locus of the gene(s) for methicillin resistance in Staphylococcus aureus.

The localization of the gene(s) mediating methicillin (mecr) in Staphylococcus aureus was determined by transformation with deoxyribonucleic acid (DNA) from a natural mecr strain (DU 4916) and transformation obtained with DNA from this strain. Streptomycin resistance genes (strr) and novobiocin resistance genes (novr) were used concurrently as representatives for chromosomal genes; penicillinase (PI254) and tetracycline plasmids were used as examples of medium- and small-size extrachromosomal genes, respectively. Superinfection of the lysogenic recipients with the competence-inducing phage phi11 or 83A enhanced transformation for all markers. Phenotypic expression of cadmium (cadr), tetracycline (tetr), or methicillin resistance (mecr) did not appear to require a host recombination system since a recA1 mutant could serve as the recipient provided it was superinfected with a competence-inducing phage. There was, furthermore, no requirement for preexisting plasmids for phenotypic expression. Ultraviolet irradiation of transforming DNA enhanced at low doses the transformation frequency for chromosomal genes strr and novr but not for mecr, cadr, or tetr. The gene(s) for mecr was transformed with chromosomal DNA after sodium dodecyl sulfate-sodium chloride extraction and after neutral sucrose gradient centrifugation of bulk DNA from wild-type strain DU 4916 and the transformats. No cavalently closed circular DNA or open circular DNA carrying the methicillin resistance gene(s) could be detected in the wild type or the transformants either by ethidium bromide-cesium chloride gradient centrifugation or by zonal rate centrifugation of cells directly lysed on top of the gradients. The mecr gene(s) is thus probably of chromosomal nature but possibly under recombinational control of phage genes, since transfer of mecr is independent of the recA1 gene(s) but can be accomplished in this strain after superinfection with a competence-inducing phage. Ultraviolet light inactivation of transforming DNA shows first-order kinetics for mecr transformability similar to that observed for both transfecting and plasmid DNA.

Chromosome Mapping↗
PubMedDate not supplied

Evaluation of an improved DNA probe for diagnosis of pertussis.

A Bordetella pertussis specific subclone, pRZ61, of a Bordetella genus-specific clone, pB23, was evaluated on nasopharyngeal aspirates of 179 patients with suspected pertussis. Hybridization was performed directly after spotting or after 1-3 days of preculture of the nylon membranes on solid culture medium. A direct comparison of the two probes was obtained by reprobing with the subclone the same membranes that had been hybridized with the parent probe. pRZ61 detected 50% of the serologically defined cases of pertussis, that is, had the same sensitivity as standard culture. Specificity as compared with serology was close to 100%. The increasing sensitivity and the corresponding decreasing specificity after preculture noted for pB23 was not seen with the subclone. The study showed that the improved probe represents a rapid diagnostic method in pertussis.

Bordetella pertussis↗