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Biomedical subjects

K E Johansson

Publications and source records attributed to K E Johansson.

At least 19 recordsLinked to original sources
PubMed2005

Survival of clostridium perfringens during simulated transport and stability of some plasmid-borne toxin genes under aerobic conditions.

Clostridium perfringens is a pathogen of great concern in veterinary medicine, because it causes enteric diseases and different types of toxaemias in domesticated animals. It is important that bacteria in tissue samples, which have been collected in the field, survive and for the classification of C. perfringens into the correct toxin group, it is crucial that plasmid-borne genes are not lost during transportation or in the diagnostic laboratory. The objectives of this study were to investigate the survival of C. perfringens in a simulated transport of field samples and to determine the stability of the plasmid-borne toxin genes cpb1 and etx after storage at room temperature and at 4 degrees C. Stability of the plasmid-borne genes cpb1 and etx of C. perfringens CCUG 2035, and cpb2 from C. perfringens CIP 106526, JF 2255 and 6 field isolates in aerobic atmosphere was also studied. Survival of C. perfringens was similar in all experiments. The cpbl and etx genes were detected in all isolates from samples stored either at room temperature or at 4 degrees C for 24-44 h. Repeated aerobic treatment of C. perfringens CCUG 2035 and CIP 106526 did not result in the loss of the plasmid-borne genes cpb1, cpb2 or etx. Plasmid-borne genes in C. perfringens were found to be more stable than generally reported. Therefore, C. perfringens toxinotyping by PCR can be performed reliably, as the risk of plasmid loss seems to be a minor problem.

Clostridium perfringens↗
PubMed2002

Prospective evaluation of laparoscopic and open 360 degree fundoplication in mild and severe gastro-oesophageal reflux disease.

OBJECTIVE: To investigate the relationship between five-year control of reflux and early postoperative oesophageal function after total fundoplication done either laparoscopically or through a laparotomy in severe and mild reflux disease. DESIGN: Prospective open study. SETTING: University hospital, Sweden. PATIENTS: In the group with severe disease 9 patients had a laparotomy and 7 laparoscopy. The corresponding figures for the group with mild disease were 21 and 34 respectively. RESULTS: The increase in lower oesophageal sphincter pressure 6 months after operation in patients with recurrent disease was significantly less than that for patients with good reflux control (p < 0.01). In patients who had laparotomy, including 30% (9/30) with severe reflux disease, good long-term reflux control was found in 93% (27/29). In patients operated on laparoscopically including 17% (7/41) with severe reflux disease good long-term reflux control was found in 90% (35/39). CONCLUSION: The mechanism of recurrence differed between patients with severe disease who had a laparotomy and patients with mild disease operated on laparoscopically. Early postoperative manometry was prognostic for recurrence. Long-term reflux control seems to be similar after laparotomy and laparoscopy. Further randomised studies are needed.

Diagnostic Techniques, Digestive System↗
PubMed2001

Detection of Mycobacterium avium subsp. paratuberculosis in tissue samples by single, fluorescent and nested PCR based on the IS900 gene.

The aim of this study was to determine if fluorescent PCR could be used instead of nested PCR, for the detection of Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) in clinical specimens, to improve the sensitivity without increasing the risk for cross-contamination. We investigated and compared the sensitivity of single PCR, fluorescent PCR and nested PCR for the detection of IS900, an insertion sequence specific for M. paratuberculosis. A previously described extraction method for clinical specimens, based on xylene, was evaluated regarding its suitability for routine diagnostic work. The sensitivity of each PCR system was assessed by analysing a serial dilution of M. paratuberculosis DNA. To improve the reliability of the PCR and to facilitate the interpretation of the PCR results, a positive internal control molecule ("mimic") was developed and used for single and fluorescent PCR. In nested PCR, an existing mimic was used. The efficiency of recovering DNA of M. paratuberculosis from clinical specimens by the extraction method and detection of the organism by PCR was studied by analysing spiked ileum mucosa specimens. The final evaluation was performed on seventeen ileum mucosa specimens, previously found positive for M. paratuberculosis by bacterial culture. Twelve of the samples were positive by fluorescent PCR and nested PCR, and 10 samples were positive by single PCR. The use of mimics showed inhibition in specimens harbouring few M. paratuberculosis organisms, illustrating the effect of inhibitory substances in combination with small amounts of M. paratuberculosis DNA. We conclude that the extraction method was not adequate to recover small amounts of M. paratuberculosis and that inhibitory substances were still present in the processed specimens, but that the method is useful for identifying positive samples. Fluorescent PCR was a suitable alternative to both single PCR and nested PCR for the detection of M. paratuberculosis.

Animals↗
PubMed2001

Genetic diversity and evolution of Mycoplasma capricolum subsp. capripneumoniae strains from eastern Africa assessed by 16S rDNA sequence analysis.

Mycoplasma capricolum subsp. capripneumoniae (M. capripneumoniae), the causal agent of contagious caprine pleuropneumonia (CCPP), is a member of the so-called Mycoplasma mycoides cluster. These mycoplasmas have two rRNA operons in which intraspecific variations have been demonstrated. The sequences of the 16S rRNA genes of both operons from 13 field strains of M. capripneumoniae from three neighbouring African countries (Kenya, Ethiopia, and Tanzania) were determined. Four new and unique polymorphism patterns reflecting the intraspecific variations were found. Two of these patterns included length differences between the rrnA and rrnB operons. The length difference in one of the patterns was caused by a two-nucleotide insert (TG) in the rrnB operon and the length difference in the other pattern was due to a three-nucleotide deletion, also in the rrnB operon. Another pattern was characterised by a polymorphic position caused by a mutation that is known to cause streptomycin resistance in other bacterial species. The strain with this pattern was also found to be resistant to streptomycin. Streptomycin resistant clones were selected from four M. capripneumoniae strains to further investigate the correlation of this mutation to streptomycin resistance. Mutations in the 16S rRNA genes had occurred in two of these strains. The fourth pattern included a new polymorphism in position 1059. The results show that polymorphisms in M. capripneumoniae strains can be used as epidemiological markers for CCPP in smaller geographical areas and to study the molecular evolution of this species.

Animals↗
PubMed2001

Re-evaluation of the classical Mycoplasma lipophilum cluster (Weisburg et al. 1989) and description of two new clusters in the hominis group based on 16S rDNA sequences.

The Mycoplasma lipophilum cluster (Weisburg et al. 1989) in the hominis group of the mollicutes is re-evaluated in this work to update the phylogenetic framework for classification of species within the genus Mycoplasma. Therefore, sequences of the 16S rRNA gene were determined from previously described species, and 11 were found to be closely related to the M. lipophilum cluster. A selection of members of the other hitherto defined clusters of the hominis group was included for phylogenetic analysis, revealing that the classical M. lipophilum cluster could be re-organized into two clusters, namely the M. lipophilum cluster and the Mycoplasma bovis cluster. The former was found to contain two species, while the latter contained 20 species. The two clusters were closely related, sharing an ancestral branch with the Mycoplasma synoviae cluster. Furthermore, the M. bovis cluster could be divided into subclusters. Interestingly, two species, Mycoplasma equigenitalium and Mycoplasma elephantis, formed a distinct and early branch of the M. lipophilum, M. bovis and M. synoviae clusters. This entity was termed the M. equigenitalium cluster. The clusters and subclusters could be verified by using neighbour-joining and maximum-likelihood analyses on a variety of data sets, bootstrap calculations, secondary structure analysis and signature nucleotides. Therefore, the new 16S rDNA data presented in this work were used to re-evaluate the M. lipophilum cluster, leading to the definition of two additional clusters. At present, the mollicutes belonging to the hominis group can be classified into ten evolutionary lineages.

Bacterial Typing Techniques↗
PubMed2001

Proposal to transfer some members of the genera Haemobartonella and Eperythrozoon to the genus Mycoplasma with descriptions of 'Candidatus Mycoplasma haemofelis', 'Candidatus Mycoplasma haemomuris', 'Candidatus Mycoplasma haemosuis' and 'Candidatus Mycoplasma wenyonii'.

Cell-wall-less uncultivated parasitic bacteria that attach to the surface of host erythrocytes currently are classified in the order Rickettsiales, family Anaplasmataceae, in the genera Haemobartonella and Eperythrozoon. Recently 16S rRNA gene sequences have been determined for four of these species: Haemobartonella felis and Haemobartonella muris and Eperythrozoon suis and Eperythrozoon wenyonii. Phylogenetic analysis of these sequence data shows that these haemotrophic bacteria are closely related to species in the genus Mycoplasma (class Mollicutes). These haemotrophic bacteria form a new phylogenetic cluster within the so-called pneumoniae group of Mycoplasma and share properties with one another as well as with other members of the pneumoniae group. These studies clearly indicate that the classification of these taxa should be changed to reflect their phylogenetic affiliation and the following is proposed: (i) that Haemobartonella felis and Haemobartonella muris should be transferred to the genus Mycoplasma as 'Candidatus Mycoplasma haemofelis' and 'Candidatus Mycoplasma haemomuris' and (ii) that Eperythrozoon suis and Eperythrozoon wenyonii should be transferred to the genus Mycoplasma as 'Candidatus Mycoplasma haemosuis' and 'Candidatus Mycoplasma wenyonii'. The former Haemobartonella and Eperythrozoon species described here represent a new group of parasitic mycoplasmas that possess a pathogenic capacity previously unrecognized among the mollicutes. These haemotrophic mycoplasmas have been given the trivial name haemoplasmas. These results call into question the affiliation of the remaining officially named species of Haemobartonella and Eperythrozoon which should be considered species of uncertain affiliation pending the resolution of their phylogenetic status.

Anaplasmataceae↗
PubMed2001

Phylogeny of the seal mycoplasmas Mycoplasma phocae corrig., Mycoplasma phocicerebrale corrig. and Mycoplasma phocirhinis corrig. based on sequence analysis of 16S rDNA.

The nucleotide sequences of the 16S rRNA genes from the type strains of three seal mycoplasmas, Mycoplasma phocicerebrale, Mycoplasma phocae and Mycoplasma phocirhinis (formerly Mycoplasma phocacerebrale, Mycoplasma phocidae and Mycoplasma phocarhinis, respectively), were determined by direct DNA cycle sequencing. Polymorphisms were found in all three 16S rRNA gene sequences, showing the existence of two different rRNA operons. In M. phocae, a length difference was found between the operons, caused by an insertion or a deletion of an adenosine in one of the operons. The sequence information was used to construct phylogenetic trees. All three species were found to belong to the hominis group, but to different clusters. M. phocicerebrale and M. phocae were found to be members of the Mycoplasma hominis cluster, within which M. phocicerebrale grouped in the Mycoplasma alkalescens subcluster. M. phocirhinis was found to be a member of the Mycoplasma bovigenitalium subcluster of the Mycoplasma bovis cluster. The 16S rRNA gene sequences of all hitherto validly described species within the M. hominis and M. bovis clusters have now been determined.

Animals↗
PubMed2000

Effects of anti-reflux surgery on chronic cough and asthma in patients with gastro-oesophageal reflux disease.

This two-group prospective study evaluated the effect of anti-reflux surgery (fundoplication) on 24 patients with severe gastro-oesophageal reflux disease (GORD) and concomitant asthma (n=13) or chronic cough (n=11). Twenty-four hour oesophageal pH monitoring and lung function tests (FEV1, FVC) were done before and within 1 year after anti-reflux surgery. A diary was kept by the patient during the 4-week period prior to surgery and during 4-week periods 6 and 12 months postoperatively, with daily monitoring of peak expiratory flow rate, respiratory and reflux symptoms and medication. In non-asthmatic patients, coughing was reduced by 47% and 80% during the day and night, respectively, 12 months after surgery (P < 0.01). Concomitant hoarseness and expectoration were also significantly reduced (P<0.05). No effect on lung function was seen. In patients with asthma, small, non-significant reductions in asthma symptom scores and consumption of rescue medication were seen. Twenty-two patients were completely free from their GORD symptoms after surgery. In conclusion, anti-reflux surgery in patients with GORD had a more favourable effect on concomitant cough than concomitant asthma.

Adult↗
PubMed2000

Updated phylogenetic description of the Mycoplasma hominis cluster (Weisburg et al. 1989) based on 16S rDNA sequences.

The fastidious nature of the mollicutes (mycoplasmas), their lack of a classic bacterial cell wall, and their very small genome, make phylogenetic placements of new species in this enlarging group of prokaryotes an important and valuable aid in their classification. In this report we have determined the phylogeny of the Mycoplasma hominis cluster of the hominis group. The 16S rDNA sequences from several previously described Mycoplasma species were determined and ten species were found to belong to the M. hominis cluster. With almost complete sequences available, the phylogenetic analysis revealed that the M. hominis cluster currently comprises 19 species, forming a distinct clade as judged from branch lengths, bootstrap percentage values, nucleotide signature analysis, and structural elements in the 16S rRNA molecule. The 16S rRNA gene sequences of species in the M. hominis cluster were found to be > or = 94% similar and the range within which similarities can be used in the classification of new species is discussed. Members of the M. hominis cluster all share a major biochemical property of M. hominis, in that they hydrolyse arginine and are incapable of fermenting glucose. This consistency in phenotypic pattern has not been found in any of the other phylogenetic clusters of the hominis group. Two species, the non-cultivable agent of Grey Lung disease in rodents (tentatively named 'Candidatus Mycoplasma ravipulmonis') and the avian species Mycoplasma gypis strain B1/T1T, were regarded as close relatives to the M. hominis cluster, but are clearly separated from the species of this cluster. Both species formed early branches of the M. hominis cluster and should be regarded as individual lines containing one species.

DNA, Bacterial↗
PubMed1999

Mycoplasma cavipharyngis and Mycoplasma fastidiosum, the closest relatives to Eperythrozoon spp. and Haemobartonella spp.

The 16S rRNA gene sequences of Mycoplasma cavipharyngis and Mycoplasma fastidiosum have been determined. Phylogenetic analysis showed that these species formed a new cluster within the so-called pneumoniae group of the mollicutes (class Mollicutes). This cluster will be referred to as the M. fastidiosum cluster. Interestingly, the M. fastidiosum cluster formed a sister lineage to the haemotrophic bacteria. Eperythrozoon spp. and Haemobartonella spp. The two latter genera, formerly classified as rickettsias, formed a stable phylogenetic entity in the tree as judged from branch lengths, bootstrap values and sequence signatures. Thus, the members of the M. fastidiosum cluster are the closest known relatives to the haemotrophic bacteria. Our data strongly support that the haemotrophic bacteria should be reclassified to reflect their actual phylogenetic affiliation.

Anaplasmataceae↗
PubMed1999

Genetic basis of macrolide and lincosamide resistance in Brachyspira (Serpulina) hyodysenteriae.

Macrolide antibiotic resistance is widespread among Brachyspira hyodysenteriae (formerly Serpulina hyodysenteriae) isolates. The genetic basis of macrolide and lincosamide resistance in B. hyodysenteriae was elucidated. Resistance to tylosin, erythromycin and clindamycin in B. hyodysenteriae was associated with an A-->T transversion mutation in the nucleotide position homologous with position 2058 of the Escherichia coli 23S rRNA gene. The nucleotide sequences of the peptidyl transferase region of the 23S rDNA from seven macrolide and lincosamide resistant and seven susceptible strains of Brachyspira spp. were determined. None of the susceptible strains were mutated whereas all the resistant strains had a mutation in position 2058. Susceptible strains became resistant in vitro after subculturing on agar containing 4 micrograms ml-1 of tylosin. Sequencing of these strains revealed an A-->G transition mutation in position 2058.

Anti-Bacterial Agents↗
PubMed1999

Single PCR and nested PCR with a mimic molecule for detection of Mycobacterium avium subsp. paratuberculosis.

Mycobacterium avium subsp. paratuberculosis is the causative agent of Johne's disease in ruminants. The current methods for detection of M. avium subsp. paratuberculosis are slow and insensitive. We report the use of a polymerase chain reaction (PCR) based on IS900 to confirm growth of M. avium subsp. paratuberculosis in primary bacterial cultures from bovine tissue and fecal samples. The use of PCR on single colonies reduced the time for analysis by 2 months compared with conventional methods. We also report the development of a nested PCR based on IS900 and the development of a positive internal control molecule, a so-called mimic. The system was tested with spiked tissue samples, and the sensitivity was estimated to 10 CFU per sample. Seventeen tissue samples, previously found M. avium subsp. paratuberculosis positive by microbiological culture, were analyzed by nested PCR and the efficiency of the PCR was checked by co-amplification of the mimic. Absence of the mimic amplicon indicated inhibition of the amplification. Ten of the samples were positive and five were negative, as judged from the presence or absence of the IS900 PCR product. Two negative samples could not be judged because of inhibition revealed by mimic molecules. It was concluded that the nested PCR, together with the mimic, could be a useful tool in screening tissue materials.

Animals↗
PubMed1999

Detection and identification of Mycoplasma conjunctivae in infectious keratoconjunctivitis by PCR based on the 16S rRNA gene.

A specific PCR assay based on unique sequences of the rrs genes (16S rRNA) of Mycoplasma conjunctivae was developed for direct detection and identification of this pathogen from clinical material. DNA from eye swabs was amplified after a simple lysis step by either a single PCR with the M. conjunctivae specific primer pair McoR1 and McoF1, or by a nested PCR with the Mycoplasma genus specific primer pair MOLIGEN1-L and 16UNI-R in the first step and McoR1 and McoF1 in the second step. The specificity of the primer pair McoR1 and McoF1 was verified with purified DNA from the type strain, from 17 field isolates of M. conjunctivae and from several Mollicutes which are phylogenetically related to M. conjunctivae or which can be isolated from the same host animals. This method identified mycoplasma isolates from goat, sheep, ibex and chamois originating from different countries as M. conjunctivae. No cross amplifications with other mycoplasmas which are related to M. conjunctivae were observed. Eye swab samples containing known numbers of M. conjunctivae cells were analysed after direct lysis of the material. The detection level was estimated to be 20 cells per swab when the nested PCR procedure was used and 2 x 10(5) by the single PCR method. In an experimental infection model of sheep, the nested PCR method for detection of M. conjunctivae gave results which were comparable to mycoplasmal culture. These are the implications for diagnostic purposes: M. conjunctivae isolates can be identified by the one-step PCR method, whereas for detection and identification of M. conjunctivae in clinical material the two-step method should be used (higher sensitivity).

Animals↗
PubMed1999

Diagnosis of contagious bovine pleuropneumonia by PCR-laser- induced fluorescence and PCR-restriction endonuclease analysis based on the 16S rRNA genes of Mycoplasma mycoides subsp. mycoides SC.

As contagious bovine pleuropneumonia (CBPP) is spreading fast in many African countries, there is an increasing demand for rapid and sensitive diagnostic methods that can be used to confirm the initial diagnosis based on clinical symptoms or pathological findings. Two PCR-based diagnostic systems for identification of the infectious agent, Mycoplasma mycoides subsp. mycoides SC (M. mycoides SC), in various samples are presented. Both systems involve group-specific amplification of the two 16S rRNA genes from mycoplasmas of the M. mycoides cluster. The laser-induced fluorescence assay is based on a unique sequence length difference between the two 16S rRNA genes in M. mycoides SC. This region was amplified by PCR, and the products were separated by polyacrylamide gel electrophoresis in a DNA sequencer. The resulting electropherogram showed two peaks for strains of M. mycoides SC and one peak for all other members of the M. mycoides cluster. The second system was based on restriction endonuclease analysis and agarose gel electrophoresis. Restriction of amplicons from a region containing a polymorphism, which is found in M. mycoides SC only, resulted in an extra band on the agarose gel because an AluI site is lacking in the rrnA operon. Specimens from cows with postmortem signs of CBPP were analyzed with the two PCR systems. M. mycoides SC was clearly identified in pleural fluid and lung tissue, and the methods were found to be robust and rapid. The results were in agreement with those obtained by conventional diagnostic techniques.

Animals↗
PubMed1998

Susceptibility of alpine ibex to conjunctivitis caused by inoculation of a sheep-strain of Mycoplasma conjunctivae.

We evaluated the susceptibility of alpine ibex (Capra ibex ibex) to mycoplasmal conjunctivitis induced by a strain of Mycoplasma conjunctivae isolated from domestic sheep by inoculation of three alpine ibexes with 1.2 x 10(6) colony forming units of M. conjunctivae in the conjunctival sac of both eyes. One more ibex was exposed to the infection by contact. Experimental animals were free of M. conjunctivae and ocular Chlamydia infection before inoculation. Conjunctivitis and serous to mucous lachrymation became apparent in all four ibexes. Clinical signs began within 2 days in inoculated animals and 22 days after the beginning of the experiment in the contact ibex. M. conjunctivae was demonstrated up to the 63th day post-inoculation by cultural and PCR-methods. After 63 days, histopathologic examination revealed nearly normal ocular tissues, and M. conjunctivae could be detected from two eyes only. No other infectious agents which might cause conjunctivitis or keratitis, including Chlamydia psittaci and Branhamella ovis, were involved. Our investigation indicates that sheep-strains of M. conjunctivae can induce conjunctivitis in alpine ibex, thus showing pathogenicity of this organism for Caprinae species other than domestic sheep and goats.

Animals↗