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  • 1. Ahrén, C M
    et al.
    Gothefors, Leif
    Umeå University, Faculty of Medicine, Department of Clinical Sciences, Paediatrics.
    Stoll, B J
    Salek, M A
    Svennerholm, A M
    Comparison of methods for detection of colonization factor antigens on enterotoxigenic Escherichia coli.1986In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 23, no 3, p. 586-91Article in journal (Refereed)
    Abstract [en]

    Fecal Escherichia coli isolates from 196 patients with watery diarrhea and 68 healthy individuals (controls) were analyzed in Bangladesh immediately after isolation for the presence of colonization factor antigen (CFA) I or II (CFA/I or CFA/II, respectively) by a mannose-resistant hemagglutination (MRHA) test with six species of erythrocytes and by a slide agglutination test with absorbed CFA/I or CFA/II antisera. The presence of CFAs was confirmed by immunodiffusion analyses done in Sweden. By these methods, it was found that 49 of 69 enterotoxin-producing E. coli strains isolated from patients carried CFA/I or CFA/II, whereas none of the nonenterotoxigenic E. coli isolates or the three toxin-positive strains isolated from healthy individuals carried these adhesins. All E. coli strains retained their MRHA ability after transportation to Sweden followed by one subculture and after storage at -70 degrees C (but not at room temperature) for 1 to 2 years without further subculturing. After 5 to 10 subcultures of the fresh isolates, however, 70% of the initially CFA/I- and 80% of the initially CFA/II-carrying strains analyzed did not hemagglutinate. The efficacy of different methods for detecting CFAs on the fresh isolates was compared with that of immunodiffusion. The sensitivity of MRHA with human blood group A erythrocytes for the detection of CFA/I was high (97%), but the specificity was only 69%. The sensitivity of MRHA with bovine erythrocytes for the detection of CFA/II in Bangladesh was very low but increased considerably when chicken erythrocytes were also used. Whereas both false-positive and false-negative reactions were obtained when absorbed CFA antisera were used for agglutination, antisera against purified CFAs were equally effective as immunodiffusion in identifying CFA/I and CFA/II-carrying strains.

  • 2.
    Alexeyev, Oleg A
    et al.
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Marklund, Ingrid
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Shannon, Beverley
    Golovleva, Irina
    Olsson, Jan
    Umeå University, Faculty of Medicine, Department of Medical Biosciences, Pathology.
    Andersson, Charlotte
    Eriksson, Irene
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Cohen, Ronald
    Elgh, Fredrik
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Direct visualization of Propionibacterium acnes in prostate tissue by multicolor fluorescent in situ hybridization assay.2007In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 45, no 11, p. 3721-8Article in journal (Refereed)
    Abstract [en]

    Prostate tissues from patients with prostate cancer and benign prostatic hyperplasia (BPH) frequently contain histological inflammation, and a proportion of these patients show evidence of Propionibacterium acnes infection in the prostate gland. We developed a multicolor fluorescent in situ hybridization (FISH) assay targeting P. acnes 23S rRNA along with a 14-kb region of the P. acnes genome. This assay was used to analyze prostate tissues from patients with prostate cancer and BPH. P. acnes infection of the prostate gland was demonstrated in prostatic tissue in 5 of 10 randomly selected prostate cancer patients. FISH analysis and confocal laser microscopy imaging revealed intracellular localization and stromal biofilm-like aggregates as common forms of P. acnes infection in prostate tissues from both prostate cancer and BPH patients. A sequential analysis of prostate tissue from individual patients suggested that P. acnes can persist for up to 6 years in the prostate gland. These results indicate that P. acnes can establish a persistent infection in the prostate gland. Further study is needed to clarify the link between this bacterium and prostatic inflammation which may contribute to the development of BPH and prostate cancer.

  • 3.
    Allard, A
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Albinsson, B
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Wadell, G
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Rapid typing of human adenoviruses by a general PCR combined with restriction endonuclease analysis.2001In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 39, no 2, p. 498-505Article in journal (Refereed)
    Abstract [en]

    We have developed a system for rapid typing of adenoviruses (Ads) based on a combination of PCR and restriction endonuclease (RE) digestion (PCR-RE digestion). Degenerated consensus primers were designed, allowing amplification of DNA from all 51 human Ad prototype strains and altogether 44 different genome variants of Ad serotypes 1, 3, 4, 5, 7, 11, 19, 40, and 41. The 301-bp amplimer of 22 prototype strains representing all six subgenera and the genome variant was selected as a target for sequencing to look for subgenus and genome type variabilities. The sequences obtained were used to facilitate the selection of specific REs for discrimination purposes in a diagnostic assay by following the concept of cleavage or noncleavage of the 301-bp amplimer. On the basis of these results, a flowchart was constructed, allowing identification of subgenus B:2 and D serotypes and almost complete distinction of subgenus A, B:1, C, E, and F serotypes. Application of the PCR-RE digestion system to clinical samples allowed typing of 34 of 40 clinical samples positive for Ad. The genome type determined by this method was identical to that obtained by traditional RE typing of full-length Ad DNA. The remaining six samples were positive only after a nested PCR. Therefore, to reduce the risk of false-negative results, samples scored negative by the PCR-RE digestion system should be evaluated by the described nested PCR. Used in combination, the PCR-RE digestion method and the nested PCR provide a reliable and sensitive system that can easily be applied to all kinds of clinical samples when rapid identification of adenoviruses is needed.

  • 4.
    Aspevall, O
    et al.
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Molecular and Clinical Medicine.
    Osterman, Björn
    Dittmer, R
    Stén, L
    Lindbäck, E
    Forsum, Urban
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Molecular and Clinical Medicine. Östergötlands Läns Landsting, Centre for Laboratory Medicine, Department of Clinical Microbiology.
    Performance of four chromogenic urine culture media after one or two days of incubation compared with reference media.2002In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 40, p. 1500-1503Article in journal (Refereed)
    Abstract [en]

    Four chromogenic urine culture media were compared to culture on blood agar, MacConkey agar, and CLED (cysteine-, lactose-, and electrolyte-deficient) agar for detection of uropathogens in 1,200 urine specimens. After 2 nights of incubation, 96% of all isolates were recovered on blood agar, 96% were recovered on CLED agar, 92% were recovered on CPS ID2, 96% were recovered on CHROMagar Orientation from BBL, 95% were recovered on CHROMagar Orientation from The CHROMagar Company, and 95% were recovered on Chromogenic UTI Medium.

  • 5.
    Bergqvist, Cecilia
    et al.
    Stockholm University, Faculty of Science, Department of Neurochemistry. Public Health Agency of Sweden, Sweden.
    Holmström, Petra
    Lindegren, Gunnel
    Lagerqvist, Nina
    Leijon, Mikael
    Falk, Kerstin I.
    Multiplex Nucleic Acid Suspension Bead Arrays for Detection and Subtyping of Filoviruses2015In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 53, no 4, p. 1368-1370Article in journal (Refereed)
    Abstract [en]

    Here we describe multiplex suspension bead array systems that allow fast and reliable detection of reverse transcriptase (RT) PCR amplified filovirus genomes and also enable subtyping of Ebola virus species and Marburg virus strains. These systems have an analytical sensitivity equivalent to that of RT-PCR.

  • 6. Bom, Reinier J. M.
    et al.
    Christerson, Linus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    van der Loeff, Maarten F. Schim
    Coutinho, Roel A.
    Herrmann, Björn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Bruisten, Sylvia M.
    Evaluation of High-Resolution Typing Methods for Chlamydia trachomatis in Samples from Heterosexual Couples2011In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 49, no 8, p. 2844-2853Article in journal (Refereed)
    Abstract [en]

    We aimed to compare conventional ompA typing of Chlamydia trachomatis with multilocus sequence typing (MLST) and multilocus variable-number tandem-repeat (VNTR) analysis (MLVA). Previously used MLST and MLVA systems were compared to modified versions that used shorter target regions and nested PCR. Heterosexual couples were selected from among persons with urogenital C. trachomatis infections visiting the sexually transmitted infection outpatient clinic in Amsterdam, The Netherlands. We identified 30 couples with a total of 65 degrees C. trachomatis-positive samples on which MLST and MLVA for eight target regions were performed. All regions were successfully sequenced in 52 samples, resulting in a complete profile for 18 couples and 12 individuals. Nine ompA genovars from D to K, with two variants of genovar G, were found. The numbers of sequence type and MLVA type profiles were 20 for MLST and 21 for MLVA, and a combination of MLST and MLVA yielded 28 profiles, with discriminatory indexes (D) ranging from 0.95 to 0.99. Partners in 17 couples shared identical profiles, while partners in 1 couple had completely different profiles. Three persons had infections at multiple anatomical locations, and within each of these three individuals, all profiles were identical. The discriminatory capacity of all MLST and MLVA methods is much higher than that of ompA genotyping (D = 0.78). No genotype variation was found within the samples of the same person or from heterosexual couples with a putative single transmission. This shows that the chlamydial genome in clinical specimens has an appropriate polymorphism to enable epidemiological cluster analysis using MLST and MLVA.

  • 7.
    Boman, Jens
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Gaydos, Charlotte
    Department of Medicine, The Johns Hopkins University, Baltimore, Maryland.
    Juto, Per
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Wadell, Göran
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Quinn, Thomas C
    Department of Medicine, The Johns Hopkins University, Baltimore, Maryland.
    Failure to detect Chlamydia trachomatis in cell culture by using a monoclonal antibody directed against the major outer membrane protein1997In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 35, no 10, p. 2679-2680Article in journal (Refereed)
    Abstract [en]

    Two commercially available monoclonal antibodies for cell culture confirmation of Chlamydia trachomatis were compared in two prospective studies and one large retrospective study. In total, more than 33,000 genital specimens were cultured in parallel and stained with both antibodies, one of which was directed against the major outer membrane protein (MOMP) and one uf which was directed against the lipopolysaccharide (LPS). We found the anti-LPS-based assay to be more sensitive and as specific as the anti-MOMP-based assay for C. trachomatis cell culture confirmation of genital specimens.

  • 8. Broekhuijsen, Martien
    et al.
    Larsson, Pär
    Umeå University, Faculty of Medicine, Clinical Microbiology. Umeå University, Faculty of Medicine, Clinical Microbiology, Clinical Bacteriology.
    Byström, Mona
    Eriksson, Ulla
    Larsson, Eva
    Prior, Richard G.
    Sjöstedt, Anders
    Umeå University, Faculty of Medicine, Clinical Microbiology. Umeå University, Faculty of Medicine, Clinical Microbiology, Clinical Bacteriology.
    Titball, Richard W.
    Forsman, Mats
    FOI, Umeå.
    Genome-wide DNA microarray analysis of Francisella tularensis strains demonstrates extensive genetic conservation within the species but identifies regions that are unique to the highly virulent F. tularensis subsp. tularensis.2003In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 41, no 7, p. 2924-2931Article in journal (Refereed)
  • 9.
    Broman, T.
    et al.
    Department of Molecular Biology, Umeå University, Umeå, Sweden.
    Palmgren, H.
    Department of Molecular Biology, Umeå University, Umeå, Sweden; Department of Infectious Diseases, Umeå University, Umeå, Sweden.
    Bergström, S.
    Department of Molecular Biology, Umeå University, Umeå, Sweden.
    Sellin, M.
    Department of Clinical Microbiology, Umeå University, Umeå, Sweden .
    Waldenström, J.
    Department of Animal Ecology, Lund University, Lund, Sweden.
    Danielsson-Tham, Marie-Louise
    Department of Food Hygiene, Swed. Univ. of Agricultural Sciences, Uppsala, Sweden .
    Olsen, B.
    Department of Infectious Diseases, Umeå University, Umeå, Sweden; Res. Inst. Zoonotic Ecol./Epidemiol., Färjestaden, Sweden .
    Campylobacter jejuni in black-headed gulls (Larus ridibundus): prevalence, genotypes, and influence on C. jejuni epidemiology2002In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 40, no 12, p. 4594-4602Article in journal (Refereed)
    Abstract [en]

    Campylobacteriosis is a zoonotic disease in which birds have been suggested to play an important role as a reservoir. We investigated the prevalence of Campylobacter jejuni subsp. jejuni in black-headed gulls (Larus ridibundus) in southern Sweden with the aim of examining the nature of C. jejuni infection in this bird species. Birds were sampled in four sampling series each year during 1999 (n = 419) and 2000 (n = 365). Longitudinally sampled C. jejuni isolates from individual gulls were subjected to macrorestriction profiling (MRP) by pulsed-field gel electrophoresis to investigate the genotypical stability during the natural course of infection. Furthermore, a subset (n = 76) of black-headed gull isolates was compared to isolates from broiler chickens (n = 38) and humans (n = 56) originating from the same geographic area. We found a pronounced seasonal variation in C. jejuni carriage, with the highest rates found in late autumn. MRP similarities were higher between isolates of human and broiler chicken origin, than between those of wild bird origin and either of the other two hosts. However, identical MRPs were found in two gull isolates and one human isolate after digestion with two restriction enzymes, strongly indicating that they may have been colonized by the same clone of C. jejuni. The MRPs most prevalent in gull isolates did not occur among isolates from humans and broiler chickens, suggesting the existence of a subpopulation of C. jejuni adapted to species-specific colonization or environmental survival.

  • 10. Bucardo, F
    et al.
    Karlsson, B
    Nordgren, J
    Paniagua, M
    Gonzalez, A
    Amador, JJ
    Espinoza, F
    Svensson, Lennart
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Molecular and Clinical Medicine, Molecular Virology.
    Mutated G4P[8] rotavirus associated with a nationwide outbreak of gastroenteritis in Nicaragua in 20052007In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 45, no 3, p. 990-997Article in journal (Refereed)
    Abstract [en]

    During February and March 2005, one of the largest national recorded outbreaks of severe acute gastroenteritis occurred in Nicaragua, affecting ≥64,000 individuals and causing ≥56 deaths, predominantly in children under 5 years of age. Through a nationwide laboratory-based study, stool samples were collected and investigated for rotavirus. Of 108 stool samples examined, 72 (67%) were positive for rotavirus. While 69% (50/72) of the positive samples were found in children less than 2 years of age, 50% (6/12) of the adult samples were positive. A mutated G4P[8] strain was the most commonly recognized strain (85%), followed by mixed G strains (8%) and G9P[8] (7%) strains. Phylogenetic analysis of the VP7 gene revealed that the G4 strains belonged to the emerging lineage Ic and was distantly related to the ST3 and VA70 G4 strains. Secondary structure predictions of the VP7 G4 protein revealed an insert of an asparagine residue in position 76, which, combined with additional mutations, surprisingly modified two downstream β-sheets at amino acid positions 80 to 85 and 115 to 119. The 2005 G4P[8] strain compared to a G4P[8] strain from 2002 had a substitution of an asparagine residue for threonine (Asn→Thr) at position 96 within antigenic region A, thus eliminating a potential glycosylation site. The mutated G4 virus was introduced in Nicaragua after 2002 and probably emerged from Brazil, Argentina, or Uruguay. Copyright © 2007, American Society for Microbiology. All Rights Reserved.

  • 11.
    Bucardo, Filemon
    et al.
    Department of Microbiology, University of León, UNAN-León, Nicaragua.
    Nordgren, Johan
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Carlsson, Beatrice
    Linköping University, Department of Clinical and Experimental Medicine, Molecular Virology . Linköping University, Faculty of Health Sciences.
    Paniagua, Margarita
    Microbiology, Tumor and Cell Biology Centre, Karolinska Institutet, S-171 77 Stockholm, Sweden.
    Lindgren, Per-Eric
    Linköping University, Department of Clinical and Experimental Medicine, Medical Microbiology . Linköping University, Faculty of Health Sciences.
    Espinoza, Felix
    Department of Microbiology, University of León, UNAN-León, Nicaragua.
    Svensson, Lennart
    Linköping University, Department of Clinical and Experimental Medicine, Molecular Virology . Linköping University, Faculty of Health Sciences.
    Pediatric norovirus diarrhea in Nicaragua2008In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 46, no 8, p. 2573-2580Article in journal (Refereed)
    Abstract [en]

    Information about norovirus (NoV) infections in Central America is limited. Through a passive community and hospital pediatric diarrhea surveillance program, a total of 542 stool samples were collected between March 2005 and February 2006 in León, Nicaragua. NoV was detected in 12% (65/542) of the children; of these, 11% (45/409) were in the community and 15% (20/133) were in the hospital, with most strains (88%) belonging to genogroup II. NoV infections were age and gender associated, with children of <2 years of age (P < 0.05) and girls (P < 0.05) being most affected. Breast-feeding did not reduce the number of NoV infections. An important proportion (57%) of NoV-infected children were coinfected with diarrheagenic Escherichia coli. A significant proportion (18/31) of NoV-positive children with dehydration required intravenous rehydration. Nucleotide sequence analysis (38/65) of the N-terminal and shell region in the capsid gene revealed that at least six genotypes (GI.4, GII.2, GII.4, GII.7, GII.17, and a potentially novel cluster termed "GII.18-Nica") circulated during the study period, with GII.4 virus being predominant (26/38). The majority (20/26) of those GII.4 strains shared high nucleotide homology (99%) with the globally emerging Hunter strain. The mean viral load was approximately 15-fold higher in children infected with GII.4 virus than in those infected with other G.II viruses, with the highest viral load observed for the group of children infected with GII.4 and requiring intravenous rehydration. This study, the first of its type from a Central American country, suggests that NoV is an important etiological agent of acute diarrhea among children of <2 years of age in Nicaragua.

  • 12.
    Christerson, Linus
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Microbiology and Infectious Medicine, Clinical Bacteriology.
    Bom, Reinier J. M.
    Bruisten, Sylvia M.
    Yass, Resha
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Microbiology and Infectious Medicine, Clinical Bacteriology.
    Hardick, Justin
    Bratt, Goran
    Gaydos, Charlotte A.
    Morre, Servaas A.
    Herrmann, Bjorn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Microbiology and Infectious Medicine, Clinical Bacteriology.
    Chlamydia trachomatis Strains Show Specific Clustering for Men Who Have Sex with Men Compared to Heterosexual Populations in Sweden, the Netherlands, and the United States2012In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 50, no 11, p. 3548-3555Article in journal (Refereed)
    Abstract [en]

    High-resolution genotyping of Chlamydia trachomatis improves the characterization of strains infecting different patient groups and sexual networks. In this study, multilocus sequence typing (MLST) and ompA sequence determination were used for an analysis of C. trachomatis strains from 203 men who have sex with men (MSM) from Sweden, the Netherlands, and the United States. The results obtained were compared with data from 153 heterosexual women from Sweden and the Netherlands. The overlap in MLST/ompA profiles between MSM from Sweden and the Netherlands was 68%, while the overlap between heterosexual populations from these countries was only 18%. The distribution of genotypes in MSM from the United States was less similar to that in MSM from the European countries, with 45% and 46% overlaps for MSM in Sweden and the Netherlands, respectively. Minimum-spanning-tree analysis of MLST/ompA sequence types identified two large clusters that contained almost exclusively samples from MSM and comprised 74% of all MSM samples. Three other clusters were predominated by samples from women but also contained MSM specimens. Of 19 detected variants of the MLST target CT144, three variants were highly associated with MSM. Our study supports the hypotheses of both tissue tropism as well as epidemiological network structures as explanations for the linkage between specific genetic variants and sexual orientation.

  • 13.
    Christerson, Linus
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Ruettger, Anke
    Friedrich-Loeffler-Institut (Federal Research Institute for Animal Health), Institute of Molecular Pathogenesis, Jena, Germany.
    Gravningen, Kirsten
    Department of Microbiology and Infection Control, University Hospital of North Norway, Tromsø, Norway.
    Ehricht, Ralf
    Alere Technologies GmbH, Jena, Germany.
    Sachse, Konrad
    Friedrich-Loeffler-Institut (Federal Research Institute for Animal Health), Institute of Molecular Pathogenesis, Jena, Germany.
    Herrmann, Björn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    High-Resolution Genotyping of Chlamydia trachomatis by Use of a Novel Multilocus Typing DNA Microarray2011In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 49, no 8, p. 2838-2843Article in journal (Refereed)
    Abstract [en]

    Typing of Chlamydia trachomatis is important to understandingits epidemiology. Currently used methods such as DNA sequencingof the ompA gene and multilocus sequence typing (MLST) eitheroffer limited epidemiological resolution or are laborious andexpensive, or both. DNA microarray technology using the ArrayStripformat is an affordable alternative for genotyping. In thisstudy, we developed a new multilocus typing (MLT) DNA microarray,based on the target regions of a high-resolution MLST systemas well as software for easy analysis. Validation of the arraywas done by typing 80 previously MLST-typed clinical specimensfrom unselected adolescents in school. The MLT array showed100% specificity and provided 2.4-times-higher resolution thanompA sequencing, separating the commonly predominating ompAE/Bour genotype into 7 MLT array genotypes. The MLT array reproducedepidemiological findings revealed by the MLST system and showedsufficient sensitivity to work with clinical specimens. Comparedto MLST analysis, the expenses needed for testing a sample withthe MLT array are considerably lower. Moreover, testing canbe completed within 1 working day rather than 3 or 4 days, withdata analysis not requiring highly specialized personnel. Thepresent MLT array represents a powerful alternative in C. trachomatisgenotyping.

  • 14. Claesson, B A
    et al.
    Lagergård, T
    Trollfors, B
    Gothefors, Leif
    Umeå University, Faculty of Medicine, Department of Clinical Sciences, Paediatrics.
    Jodal, U
    Serum antibody response to capsular polysaccharide, outer membrane, and lipooligosaccharide in children with invasive Haemophilus influenzae type b infections.1987In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 25, no 12, p. 2339-43Article in journal (Refereed)
    Abstract [en]

    Serum antibodies against capsular polysaccharide (CPS), outer membrane (OM), and lipooligosaccharide (LOS) from Haemophilus influenzae type b were measured by enzyme-linked immunosorbent assay in acute- and convalescent-phase sera from 21 children between 3 months and 4 years of age with invasive H. influenzae type b infections. As expected, the levels of anti-CPS antibodies in the acute-phase serum samples were low or not detectable, as were the levels of antibodies against LOS. In contrast, all children had detectable antibodies against the OM in the acute-phase serum sample, indicating that they are of little or no importance for protection. An antibody response to CPS was noted in 13 of the 21 patients, mainly in the older children. An antibody response to the OM was seen in 16 patients, with no evident relation to age. The antibody response to the OM preparation, which consisted of proteins and LOS, was probably directed mainly against the OM proteins, since only six children showed a response, usually of low magnitude, of antibodies to LOS.

  • 15.
    Demczuk, Walter H.B.
    et al.
    National Microbiology Laboratory, Winnipeg, Canada.
    Sidhu, S.
    National Microbiology Laboratory, Winnipeg, Canada.
    Unemo, Magnus
    WHO Collaborating Centre for Gonorrhoea and Other STIs, Örebro University Hospital, Örebro, Sweden; School of Medical Sciences, Örebro University, Örebro, Sweden.
    Whiley, David M.
    Centre for Clinical Research, The University of Queensland, Brisbane, Australia.
    Allen, Vanessa G.
    Public Health Ontario Laboratories, Toronto , Canada.
    Dillon, Jeremiah R.
    Department of Microbiology and Immunology, University of Saskatchewan, Saskatoon, Canada.
    Cole, Michelle J.
    Public Health England, London, United Kingdom.
    Seah, Christine
    Public Health Ontario Laboratories, Toronto, Canada.
    Trembizki, Ella
    Centre for Clinical Research, The University of Queensland, Brisbane, Australia.
    Trees, David L.
    Centers for Disease Control and Prevention, Atlanta GA, United States.
    Kersh, Ellen N.
    Centers for Disease Control and Prevention, Atlanta GA, United States.
    Abrams, A. Jeanine
    Centers for Disease Control and Prevention, Atlanta GA, United States.
    de Vries, Henry J.C.
    STI Outpatient Clinic, Department of Infectious Diseases, Public Health Service Amsterdam, Amsterdam, the Netherlands; Department of Dermatology, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands; Center for Infection and Immunity Amsterdam, Academic Medical Center, University of Amsterdam, Amsterdam, the Netherlands.
    van Dam, Alje P.
    Public Health Laboratory, Public Health Service Amsterdam, Amsterdam, the Netherlands; Department of Medical Microbiology, OLVG General Hospital, Amsterdam, the Netherlands; .
    Medina, I.
    National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg MB, Canada.
    Bharat, Amrita
    National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg MB, Canada.
    Mulvey, Michael Richard
    National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg MB, Canada.
    Van Domselaar, Gary
    National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg MB, Canada.
    Martin, Irene E.
    National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg MB, Canada.
    Neisseria gonorrhoeae Sequence Typing for Antimicrobial Resistance: a Novel Antimicrobial Resistance Multilocus Typing Scheme for Tracking Global Dissemination of N. gonorrhoeae Strains2017In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 55, no 5, p. 1454-1468Article in journal (Refereed)
    Abstract [en]

    A curated Web-based user-friendly sequence typing tool based on antimicrobial resistance determinants in Neisseria gonorrhoeae was developed and is publicly accessible (https://ngstar.canada.ca). The N. gonorrhoeae Sequence Typing for Antimicrobial Resistance (NG-STAR) molecular typing scheme uses the DNA sequences of 7 genes (penA, mtrR, porB, ponA, gyrA, parC, and 23S rRNA) associated with resistance to β-lactam antimicrobials, macrolides, or fluoroquinolones. NG-STAR uses the entire penA sequence, combining the historical nomenclature for penA types I to XXXVIII with novel nucleotide sequence designations; the full mtrR sequence and a portion of its promoter region; portions of ponA, porB, gyrA, and parC; and 23S rRNA sequences. NG-STAR grouped 768 isolates into 139 sequence types (STs) (n = 660) consisting of 29 clonal complexes (CCs) having a maximum of a single-locus variation, and 76 NG-STAR STs (n = 109) were identified as unrelated singletons. NG-STAR had a high Simpson's diversity index value of 96.5% (95% confidence interval [CI] = 0.959 to 0.969). The most common STs were NG-STAR ST-90 (n = 100; 13.0%), ST-42 and ST-91 (n = 45; 5.9%), ST-64 (n = 44; 5.72%), and ST-139 (n = 42; 5.5%). Decreased susceptibility to azithromycin was associated with NG-STAR ST-58, ST-61, ST-64, ST-79, ST-91, and ST-139 (n = 156; 92.3%); decreased susceptibility to cephalosporins was associated with NG-STAR ST-90, ST-91, and ST-97 (n = 162; 94.2%); and ciprofloxacin resistance was associated with NG-STAR ST-26, ST-90, ST-91, ST-97, ST-150, and ST-158 (n = 196; 98.0%). All isolates of NG-STAR ST-42, ST-43, ST-63, ST-81, and ST-160 (n = 106) were susceptible to all four antimicrobials. The standardization of nomenclature associated with antimicrobial resistance determinants through an internationally available database will facilitate the monitoring of the global dissemination of antimicrobial-resistant N. gonorrhoeae strains.

  • 16.
    Demczuk, Walter
    et al.
    Bacteriology and Enteric Diseases Program, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, Canada.
    Martin, Irene
    Bacteriology and Enteric Diseases Program, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, Canada.
    Peterson, Shelley
    Bacteriology and Enteric Diseases Program, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, Canada.
    Bharat, Amrita
    Bacteriology and Enteric Diseases Program, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, Canada.
    Van Domselaar, Gary
    Science Technology Cores and Services Division, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, Manitoba, Canada; Department of Medical Microbiology, University of Manitoba, Winnipeg, Canada.
    Graham, Morag
    Science Technology Cores and Services Division, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, Manitoba, Canada; Department of Medical Microbiology, University of Manitoba, Winnipeg, Canada.
    Lefebvre, Brigitte
    Laboratoire de Santé Publique du Québec, Ste-Anne-de-Bellevue QC, Canada.
    Allen, Vanessa
    Public Health Ontario Laboratories, Toronto ON, Canada.
    Hoang, Linda
    British Columbia Centres for Disease Control Public Health Microbiology & Reference Laboratory, Vancouver BC, Canada.
    Tyrrell, Greg
    Provincial Laboratory for Public Health, Edmonton, Canada.
    Horsman, Greg
    Saskatchewan Disease Control Laboratory, Regina, Canada.
    Wylie, John
    Cadham Provincial Laboratory, Winnipeg, Canada.
    Haldane, David
    Queen Elizabeth II Health Sciences Centre, Halifax, Canada.
    Archibald, Chris
    Centre for Communicable Diseases and Infection Control, Public Health Agency of Canada, Ottawa, Canada.
    Wong, Tom
    First Nations and Inuit Health Branch, Health Canada, Ottawa, Canada.
    Unemo, Magnus
    Örebro University, School of Health Sciences. Department of Laboratory Medicine, Microbiology, Faculty of Medicine and Health, Örebro University, Örebro, Sweden.
    Mulvey, Michael R
    Bacteriology and Enteric Diseases Program, National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, Canada.
    Genomic epidemiology and molecular resistance mechanisms of azithromycin resistant Neisseria gonorrhoeae in Canada from 1997 to 20142016In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 54, no 5, p. 1304-1313Article in journal (Refereed)
    Abstract [en]

    The emergence of Neisseria gonorrhoeae with decreased susceptibility to cephalosporins and azithromycin resistance (AZM-R) represent a public health threat of untreatable gonorrhoea infections. Genomic epidemiology through whole genome sequencing was used to describe the emergence, dissemination, and spread of AZM-R strains. The genomes of 213 AZM-R and 23 AZM-susceptible N. gonorrhoeae isolates collected in Canada from 1989 to 2014 were sequenced. Core single nucleotide polymorphism (SNP) phylogenomic analysis resolved 246 isolates into 13 lineages. High-level AZM-R (minimum inhibitory concentration ≥256 μg/ml) was found in 5 phylogenetically diverse isolates, all of which possessed the A2059G mutation (Escherichia coli numbering) in all four 23S rRNA alleles. One high-level AZM-R isolate collected in 2009 concurrently had decreased susceptibility to ceftriaxone (MIC=0.125 μg/ml). An increase in the number of 23S rRNA alleles with the C2611T mutations (E. coli numbering) conferred low to moderate AZM-R (2 to 4 and 8 to 32 μg/mL, respectively). Low level AZM-R was also associated with mtrR promoter mutations including -35A deletion and the presence of N. meningitidis-like sequences. Geographic and temporal phylogenetic clustering indicate emergent AZM-R strains arise independently and can then rapidly expand clonally in a region through local sexual networks.

  • 17.
    Donà, Valentina
    et al.
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
    Kasraian, Sara
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
    Lupo, Agnese
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
    Guilarte, Yuvia N.
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
    Hauser, Christoph
    Department of Infectious Diseases, Bern University Hospital, University of Bern, Bern, Switzerland.
    Furrer, Hansjakob
    Department of Infectious Diseases, Bern University Hospital, University of Bern, Bern, Switzerland.
    Unemo, Magnus
    Örebro University, School of Health Sciences.
    Low, Nicola
    Department of Infectious Diseases, Bern University Hospital, University of Bern, Bern, Switzerland.
    Endimiani, Andrea
    Institute for Infectious Diseases, University of Bern, Bern, Switzerlanda.
    Multiplex real-time PCR assay with high-resolution melting analysis for characterization of antimicrobial resistance in neisseria gonorrhoeae2016In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 54, no 8, p. 2074-2081Article in journal (Refereed)
    Abstract [en]

    Resistance to antibiotics used against Neisseria gonorrhoeae infections is a major public health concern. Antimicrobial resistance (AMR) testing relies on time-consuming culture-based methods. Development of rapid molecular tests for detecting AMR determinants could provide valuable tools for surveillance, epidemiological studies and to inform individual case management. We developed a fast (<1.5 hrs) SYBR-green based real-time PCR method with high resolution melting (HRM) analysis. One triplex and three duplex reactions included two sequences for N. gonorrhoeae identification and seven determinants of resistance to extended-spectrum cephalosporins (ESCs), azithromycin, ciprofloxacin, and spectinomycin. The method was validated by testing 39 previously fully-characterized N. gonorrhoeae strains, 19 commensal Neisseria spp., and an additional panel of 193 gonococcal isolates. Results were compared with culture-based AMR determination. The assay correctly identified N. gonorrhoeae and the presence or absence of the seven AMR determinants. There was some cross-reactivity with non-gonococcal Neisseria species and the detection limit was 10(3)-10(4) gDNA copies/reaction. Overall, the platform accurately detected resistance to ciprofloxacin (sensitivity and specificity, 100%), ceftriaxone (sensitivity 100%, specificity 90%), cefixime (sensitivity 92%, specificity 94%), azithromycin and spectinomycin (both sensitivity and specificity, 100%). In conclusion, our methodology accurately detects mutations generating resistance to antibiotics used to treat gonorrhea. Low assay sensitivity prevents direct diagnostic testing of clinical specimens but this method can be used to screen collections of gonococcal isolates for AMR more quickly than with current culture-based AMR testing.

  • 18.
    Donà, Valentina
    et al.
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
    Smid, Joost H.
    Institute of Social and Preventive Medicine, University of Bern, Bern, Switzerland.
    Kasraian, Sara
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
    Egli-Gany, Dianne
    Institute of Social and Preventive Medicine, University of Bern, Bern, Switzerland.
    Dost, Ferah
    Ambulatorium Kanonengasse, Zurich, Switzerland.
    Imeri, Fatime
    Laborgemeinschaft 1, Zurich, Switzerland.
    Unemo, Magnus
    Örebro University, School of Medical Sciences. Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, Örebro University Hospital, Örebro, Sweden.
    Low, Nicola
    Institute of Social and Preventive Medicine, University of Bern, Bern, Switzerland.
    Endimiani, Andrea
    Institute for Infectious Diseases, University of Bern, Bern, Switzerland.
    Mismatch Amplification Mutation Assay (MAMA)-Based Real-Time PCR for Rapid Detection of Neisseria gonorrhoeae and Antimicrobial Resistance Determinants in Clinical Specimens2018In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 56, no 9, article id e00365-18Article in journal (Refereed)
    Abstract [en]

    Molecular methods are often used for Neisseria gonorrhoeae (NG) detection, but complete definition of antimicrobial resistance (AMR) patterns still requires phenotypic tests. We developed an assay that both identifies NG and detects AMR determinants in clinical specimens.We designed a mismatch amplification mutation assay (MAMA)-based SYBR Green real-time PCR targeting: one NG-specific region (opa); mosaic penA alleles (Asp345 deletion, Gly545Ser) associated with decreased susceptibility to cephalosporins; alterations conferring resistance to ciprofloxacin (GyrA: Ser91Phe), azithromycin (23S rRNA: A2059G and C2611T) and spectinomycin (16S rRNA: C1192T). We applied the real-time PCR to 489 clinical specimens, of which 94 had paired culture isolates, and evaluated its performance by comparison with commercial diagnostic molecular and phenotypic tests.Our assay exhibited a sensitivity/specificity of 93%/100%, 96%/85%, 90%/91%, 100%/100% and 100%/90% for the detection of NG directly from urethral, rectal, pharyngeal, cervical and vaginal samples, respectively. The MAMA strategy allowed the detection of AMR mutations by comparing cycle threshold values with the reference opa reaction. The method accurately predicted the phenotype to four antibiotic classes when compared with the MIC values obtained from 94 paired cultures (sensitivity/specificity for cephalosporins, azithromycin, ciprofloxacin and spectinomycin resistance: 100%/95%, 100%/100%, 100%/100% and not applicable (NA)/100%, respectively, in genital specimens; NA/72%, NA/98%, 100%/97%, and NA/96%, respectively, in extra-genital specimens). False-positive results, particularly for the penA Asp345del reaction were observed predominantly in pharyngeal specimens.Our real-time PCR assay is a promising rapid method to identify NG and predict AMR directly in genital specimens, but further optimization for extra-genital specimens is needed.

  • 19. Dzieciatkowska, Monika
    et al.
    Liu, Xin
    Heikema, Astrid P.
    Houliston, R. Scott
    van Belkum, Alex
    Schweda, Elke K. H.
    Södertörn University, School of Life Sciences.
    Gilbert, Michel
    Richards, James C.
    Li, Jianjun
    Rapid method for sensitive screening of oligosaccharide epitopes in the lipooligosaccharide from Campylobacter jejuni strains isolated from Guillain-Barre syndrome and Miller Fisher syndrome patients2008In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 46, no 10, p. 3429-3436Article in journal (Refereed)
    Abstract [en]

    Campylobacter jejuni lipooligosaccharide (LOS) can trigger Guillain-Barre syndrome (GBS) due to its similarity to human gangliosides. Rapid and accurate structural elucidation of the LOS glycan of a strain isolated from a GBS patient could help physicians determine the spectrum of anti-ganglioside antibodies likely to be found and therefore provide valuable assistance in establishing an appropriate course of treatment. The ability of implemented mass spectrometry-based approaches in a clinical setting has been limited by the laborious and time-consuming nature of the protocols, typically 3 to 4 days, used to prepare LOS. In order to improve the analytical throughput, microwave-assisted enzymatic digestion was investigated. In this study, the bacterial cells were suspended in 50 mu l of 20 mM ammonium acetate buffer containing DNase and RNase and treated by direct microwave irradiation for 3 min. Then, proteinase K was added and the samples were again microwaved. The intact LOS samples were analyzed using electrophoresis-assisted open-tubular liquid chromatography-mass spectrometry. The reliability of the rapid, high-throughput technique was demonstrated through analysis of LOS glycans from 73 C. jejuni strains. The structure was elucidated using material from a single colony. The total time for sample preparation and MS analysis is less than 60 min.

  • 20. Ejrnaes, Karen
    et al.
    Sandvang, Dorthe
    Lundgren, Bettina
    Ferry, Sven
    Umeå University, Faculty of Medicine, Clinical Microbiology. Umeå University, Faculty of Medicine, Clinical Microbiology, Clinical Bacteriology.
    Holm, Stig
    Göteborgs Universitet.
    Monsen, Tor
    Umeå University, Faculty of Medicine, Clinical Microbiology. Umeå University, Faculty of Medicine, Clinical Microbiology, Clinical Bacteriology.
    Lundholm, Rolf
    Umeå University, Faculty of Medicine, Clinical Microbiology. Umeå University, Faculty of Medicine, Clinical Microbiology, Clinical Bacteriology.
    Frimodt-Moller, Niels
    Pulsed-field gel electrophoresis typing of Escherichia coli strains from samples collected before and after pivmecillinam or placebo treatment of uncomplicated community-acquired urinary tract infection in women.2006In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 44, no 5, p. 1776-81Article in journal (Refereed)
    Abstract [en]

    The primary infecting Escherichia coli strains from 156 women with community-acquired uncomplicated urinary tract infection (UTI) randomized to pivmecillinam or placebo and the E. coli strains causing UTI at two follow-up visits were typed using pulsed-field gel electrophoresis (PFGE). In the pivmecillinam treatment group PFGE showed that among patients having a negative urine culture at the first follow-up 77% (46/60) had a relapse with the primary infecting E. coli strain and 23% (14/60) had reinfection with a new E. coli strain at the second follow-up. Among patients having E. coli at the first follow-up PFGE showed that 80% (32/40) had persistence with the primary infecting E. coli strain, 15% (6/40) had reinfection with a new E. coli strain, and 5% (2/40) had different E. coli strains at the two follow-up visits (one had reinfection followed by relapse, and the other had persistence followed by reinfection). In the placebo group the majority had E. coli at the first follow-up. PFGE showed that among these patients 96% (50/52) had persistence with the primary infecting E. coli strain and 4% (2/50) had different E. coli strains at the two follow-up visits (both had persistence followed by reinfection). The finding that the majority of UTIs at follow-up are caused by the primary infecting E. coli strain supports the theory of a vaginal and rectal reservoir but could also support the recent discovery that E. coli strains are able to persist in the bladder epithelium despite appropriate antibiotic treatment, constituting a reservoir for recurrent UTI.

  • 21.
    Elgh, Fredrik
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Lundkvist, A
    Alexeyev, O A
    Stenlund, H
    Avsic-Zupanc, T
    Hjelle, B
    Lee, H W
    Smith, K J
    Vainionpää, R
    Wiger, D
    Wadell, G
    Juto, P
    Serological diagnosis of hantavirus infections by an enzyme-linked immunosorbent assay based on detection of immunoglobulin G and M responses to recombinant nucleocapsid proteins of five viral serotypes.1997In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 35, no 5, p. 1122-30Article in journal (Refereed)
    Abstract [en]

    Worldwide, hantaviruses cause more than 100,000 human infections annually. Rapid and accurate methods are important both in monitoring acute infections and for epidemiological studies. We and others have shown that the amino termini of hantavirus nucleocapsid proteins (Ns) are sensitive tools for the detection of specific antibodies in hantavirus disease. Accordingly, we expressed truncated Ns (amino acids 1 to 117) in Escherichia coli from the five hantaviruses known to be pathogenic to man; Hantaan (HTN), Seoul (SEO), Dobrava (DOB), Sin Nombre (SN), and Puumala (PUU) viruses. In order to obtain pure antigens for use in an enzyme-linked immunosorbent assay (ELISA), the recombinant proteins were purified by polyhistidine-metal chelate affinity chromatography. Polyclonal animal antisera and a panel of serum specimens from hantavirus-infected individuals from Scandinavia, Slovenia, Russia, Korea, China, and the United States were used to evaluate the usefulness of the method. With both human and animal sera, it was possible to designate the antibody response into two groups: those with HTN, SEO, and DOB virus reactivity on the one hand and those with SN and PUU virus reactivity on the other. In sera from Scandinavia, European Russia, and the United States, the antibody response was directed mainly to the PUU and SN virus group. The sera from Asia reacted almost exclusively with the HTN, SEO, and DOB types of viruses. This was true for both the immunoglobulin M (IgM) and IgG antibody responses, indicating that this type of discrimination can be done during the acute phase of hantavirus infections. Both the HTN, SEO, and DOB virus and the PUU and SN virus types of antibody response patterns were found in patients from the Balkan region (Solvenia).

  • 22.
    Ericsson, Henrik
    et al.
    Department of Food Hygiene, Faculty of Veterinary Medicine, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Eklöw, Annelie
    Department of Food Hygiene, Faculty of Veterinary Medicine, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Danielsson-Tham, Marie-Louise
    Department of Food Hygiene, Faculty of Veterinary Medicine, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Loncarevic, Semir
    Department of Food Hygiene, Faculty of Veterinary Medicine, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Mentzing, L.-O.
    Communicable Disease Center for Värmland County, Karlstad Central Hospital, Karlstad, Sweden.
    Persson, I.
    Communicable Disease Center for Värmland County, Karlstad Central Hospital, Karlstad, Sweden.
    Unnerstad, Helle
    Department of Food Hygiene, Faculty of Veterinary Medicine, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    Tham, Wilhelm
    Department of Food Hygiene, Faculty of Veterinary Medicine, Swedish University of Agricultural Sciences, Uppsala, Sweden.
    An outbreak of listeriosis suspected to have been caused by rainbow trout1997In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 35, no 11, p. 2904-2907Article in journal (Refereed)
    Abstract [en]

    An outbreak of listeriosis in Sweden, consisting of nine cases, was investigated by means of molecular typing of strains from patients and strains isolated from suspected foodstuffs, together with interviews of the patients. Listeria monocytogenes was isolated from six of the patients, and all isolates were of the same clonal type. This clonal type was also isolated from a 'gravad' rainbow trout, made by producer Y, found in the refrigerator of one of the patients. Unopened packages obtained from producer y were also found to contain the same clonal type of L. monocytogenes. Based on the interview results and the bacteriological typing, we suspect that at least six of the nine cases were caused by gravad or cold-smoked rainbow trout made by producer Y. To our knowledge, this is the first rainbow trout-borne outbreak of listeriosis ever reported.

  • 23.
    Eriksson, Lorraine
    et al.
    Örebro University, School of Medical Sciences. Department of Laboratory Medicine, Faculty of Health and Medical Sciences, Örebro University, Örebro, Sweden.
    Hedberg, Sara Thulin
    Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Jacobsson, Susanne
    Örebro University, School of Medical Sciences. Örebro University, School of Health Sciences. Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Fredlund, Hans
    Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Mölling, Paula
    Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden.
    Stenmark, Bianca
    Örebro University, School of Medical Sciences. Department of Laboratory Medicine, Faculty of Health and Medical Sciences, Örebro University, Örebro, Sweden.
    Whole-Genome Sequencing of Emerging Invasive Neisseria meningitidis Serogroup W in Sweden2018In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 56, no 4, article id e01409-17Article in journal (Refereed)
    Abstract [en]

    Invasive disease caused by Neisseria meningitidis serogroup W (MenW) has historically had a low incidence in Sweden, with an average incidence of 0.03 case/100,000 population from 1995 to 2014. In recent years, a significant increase in the incidence of MenW has been noted in Sweden, to an average incidence of 0.15 case/100,000 population in 2015 to 2016. In 2017 (1 January to 30 June), 33% of invasive meningococcal disease cases (7/21 cases) were caused by MenW. In the present study, all invasive MenW isolates from Sweden collected in 1995 to June 2017 (n = 86) were subjected to whole-genome sequencing to determine the population structure and to compare isolates from Sweden with historical and international cases. The increase of MenW in Sweden was determined to be due to isolates belonging to the South American sublineage of MenW clonal complex 11, namely, the novel U.K. 2013 lineage. This lineage was introduced in Sweden in 2013 and has since been the dominant lineage of MenW.

  • 24.
    Evander, Magnus
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Eriksson, Irene
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Pettersson, Lisa
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Juto, Per
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Ahlm, Clas
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Infectious Diseases.
    Olsson, Gert E
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Bucht, Göran
    Umeå University, Faculty of Science and Technology, Department of Molecular Biology (Faculty of Science and Technology).
    Allard, Annika
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Puumala hantavirus viremia diagnosed by real-time reverse transcriptase PCR using samples from patients with hemorrhagic fever and renal syndrome.2007In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 45, no 8, p. 2491-7Article in journal (Refereed)
    Abstract [en]

    Puumala virus (PUUV) is the endemic hantavirus in northern Sweden and causes nephropathia epidemica (NE), a milder form of hemorrhagic fever with renal syndrome. There is a need for fast and reliable diagnostics to differentiate the disease from other infections. By aligning virus RNA sequences isolated from 11 different bank voles and one human patient, we designed a real-time reverse transcriptase (RT) PCR method for detection of PUUV RNA. The real-time RT-PCR assay showed linearity from 20 to 2 x 10(6) virus copies with a correlation coefficient above 0.98 to 0.99 for all experiments. The detection threshold for PUUV cDNA was two copies per reaction. A two-step qualitative RT-PCR to detect PUUV RNA showed 100% concordance with the real-time RT-PCR assay. PUUV RNA viremia was detected in 33 of 34 PUUV immunoglobulin M (IgM)-positive patients with typical clinical NE disease from the region of endemicity. One PUUV IgM-negative sample had PUUV RNA, and 4 days later, the patient was IgM positive. Of samples with indeterminate IgM, 43% were PUUV RNA positive. The kinetics of antibody titers and PUUV viremia were studied, and five of six NE patients displayed a decrease in PUUV viremia a few days after disease outbreak coupled with an increase in PUUV IgM and IgG. In one patient with continuously high PUUV RNA levels but low IgM and no IgG response, the infection was lethal. These findings demonstrated that real-time RT-PCR is a useful method for diagnosis of PUUV viremia and for detecting PUUV RNA at early time points, before the appearance of IgM antibodies.

  • 25. Foucault, C
    et al.
    La Scola, B.
    Lindroos, Hillevi
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Evolution, Genomics and Systematics, Molecular Evolution.
    Andersson, Siv
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Evolution, Genomics and Systematics, Molecular Evolution.
    Raoult, D.
    Multispacer typing technique for sequence-based typing of Bartonella quintana2005In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 43, no 1, p. 41-48Article in journal (Refereed)
    Abstract [en]

    Bartonella quintana is a worldwide fastidious bacterium of the Alphaproteobacteria responsible for bacillary angiomatosis, trench fever, chronic lymphadenopathy, and culture-negative endocarditis. The recent genome sequencing of a B. quintana isolate allowed us to propose a genome-wide sequence-based typing method. To ensure sequence discrimination based on highly polymorphic areas, we amplified and sequenced 34 spacers in a large collection of B. quintana isolates. Six of these exhibited polymorphisms and allowed the characterization of 4 genotypes. However, the strain variants suggested by the noncoding sequences did not correlate with the results of pulsed-field gel electrophoresis (PFGE), which suggested a higher degree of variability. Modification of the PFGE profile of one isolate after nine subcultures confirmed that rearrangement frequencies are high in this species, making PFGE unreliable for epidemiological purposes. The low extent of sequence heterogeneity in the species suggests a recent emergence of this bacterium as a human pathogen. Direct typing of natural samples allowed the identification of a fifth genotype in the DNA extracted from a human body louse collected in Burundi. We have named the typing technique herein described multispacer typing.

  • 26. Garnier, Fabien
    et al.
    Janapatla, Rajendra Prasad
    Charpentier, Emmanuelle
    Umeå University, Faculty of Medicine, Molecular Infection Medicine Sweden (MIMS).
    Masson, Geoffrey
    Grélaud, Carole
    Stach, Jean François
    Denis, François
    Ploy, Marie-Cécile
    Insertion sequence 1515 in the ply gene of a type 1 clinical isolate of Streptococcus pneumoniae abolishes pneurnolysin expression2007In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 45, no 7, p. 2296-2297Article in journal (Refereed)
    Abstract [en]

    Abstract: A serotype 1 Streptococcus pneumoniae strain isolated by blood culture from a woman with pneumonia was found to harbor insertion sequence (IS) 1515 in the pneumolysin gene, abolishing pneumolysin expression. To our knowledge, this is the first report of an IS in the pneumolysin gene of S. pneumoniae.

  • 27. Gisselsson-Solén, Marie
    et al.
    Bylander, Anita
    Wilhelmsson, Christina
    Hermansson, Ann
    Melhus, Åsa
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    The Binax NOW test as a tool for diagnosis of severe acute otitis media and associated complications2007In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 45, no 9, p. 3003-3007Article in journal (Refereed)
    Abstract [en]

    The diagnosis of acute otitis media (AOM) is often difficult, depending heavily on the experience and skills of the examiner. However, it is important to identify episodes of AOM that involve the risk of complications and to treat these episodes appropriately. The present study was performed in order to evaluate the use of a rapid antigen assay for Streptococcus pneumoniae, the Binax NOW test, as a diagnostic tool in patients with severe AOM and associated complications. The study included 70 patients with 74 episodes of AOM, 18 of them with complications. Cultures, Binax NOW tests, and a PCR assay were performed on nasopharyngeal secretions, middle ear fluid, and in some cases mastoid bone, cerebrospinal fluid, and urine. According to culture and PCR of the middle ear fluid, 30 (41%) of the episodes were caused by S. pneumoniae. The Binax NOW test was positive in 24 of these episodes (80%). It identified pneumococcal AOM independent of antibiotic treatment, and it was easily adapted to bone tissue. The test yielded sensitivity, specificity, and positive and negative predictive values for middle ear specimens of 85%, 100%, 100%, and 89%, respectively. The corresponding positive and negative values for predicting the bacterial etiology with nasopharyngeal secretions were 51% and 75%. This study showed that the Binax NOW test is a useful diagnostic tool for patients with severe AOM with or without complications.

  • 28.
    Golparian, Daniel
    et al.
    WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Faculty of Medicine and Health, Örebro University Hospital, Örebro, Sweden.
    Boräng, Stina
    Department of Clinical Microbiology, Karolinska University Hospital Huddinge, Stockholm, Sweden .
    Sundqvist, Martin
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Unemo, Magnus
    Örebro University, School of Health and Medical Sciences, Örebro University, Sweden. WHO Collaborating Centre for Gonorrhoea and other Sexually Transmitted Infections, Swedish Reference Laboratory for Pathogenic Neisseria, Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Evaluation of the New BD Max GC Real-Time PCR Assay, Analytically and Clinically as a Supplementary Test for the BD ProbeTec GC Qx Amplified DNA Assay, for Molecular Detection of Neisseria gonorrhoeae2015In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 53, no 12, p. 3935-3937Article in journal (Refereed)
    Abstract [en]

    The new BD Max GC real-time PCR assay showed high clinical and analytical sensitivity and specificity. It can be an effective and accurate supplementary test for the BD ProbeTec GC Qx amplified DNA assay, which had suboptimal specificity, and might also be used for initial detection of Neisseria gonorrhoeae.

  • 29.
    Gullsby, Karolina
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Centre for Research and Development, Gävleborg. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infection medicine.
    Olsen, Björn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Bondeson, Kåre
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infection medicine.
    Molecular typing of Mycoplasma pneumoniae strains in Sweden, 1996–2017, and the emergence of a new P1 cytadhesin gene, Variant 2e2019In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 57, no 6, article id e00049-19Article in journal (Refereed)
    Abstract [en]

    Mycoplasma pneumoniae causes respiratory infections, such as community-acquired pneumonia (CAP), with epidemics recurring every 3 to 7 years. In 2010 and 2011, many countries experienced an extraordinary epidemic peak. The cause of these recurring epidemics is not understood, but decreasing herd immunity and shifts in the strains' antigenic properties have been suggested as contributing factors. M. pneumoniae PCR-positive samples were collected between 1996 and 2017 from four neighboring counties inhabited by 12% of Sweden's population. A total of 578 isolates were characterized directly from 624 clinical samples using P1 typing by sequencing and multilocus variable number tandem repeat analysis (MLVA). A fluorescence resonance energy transfer (FRET)-PCR approach was also used to detect mutations associated with macrolide resistance in the 23S rRNA gene. Through P1 typing, the strains were classified into type 1 and type 2, as well as variants 2a, 2b, 2c, and a new variant found in nine of the strains, denoted variant 2e. Twelve MLVA types were distinguished, and 3-5-6-2 (42.4%), 4-5-7-2 (37.4%), and 3-6-6-2 (14.9%) predominated. Several P1 and MLVA types cocirculated each year, but type 2/variant 2 strains and MLVA types 3-5-6-2 and 4-5-7-2 predominated during the epidemic period comprising the peak of 2010 and 2011. In 2016 and 2017, type 1 became more common, and MLVA type 4-5-7-2 predominated. We also found that 0.2% (1/578) of the strains carried a macrolide resistance-associated mutation, indicating a very low prevalence of macrolide resistance in this region of Sweden.

  • 30.
    Gullsby, Karolina
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Centre for Research and Development, Gävleborg.
    Storm, Martin
    Bondeson, Kåre
    Simultaneous detection of Chlamydophila pneumonia and Mycoplasma pneumoniae by idé of molecular bravons in a duplex real-time PCR2008In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660XArticle in journal (Refereed)
  • 31. Gullsby, Karolina
    et al.
    Storm, Martin
    Bondeson, Kåre
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Virology.
    Simultaneous detection of Chlamydophila pneumoniae and Mycoplasma pneumoniae by use of molecular beacons in a duplex real-time PCR2008In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 46, no 2, p. 727-31Article in journal (Refereed)
    Abstract [en]

    A real-time PCR was designed for detection of Chlamydophila pneumoniae and Mycoplasma pneumoniae such that each pathogen could be detected in a single tube and differentiated using molecular beacons marked with different fluorochromes. This duplex PCR, targeting the P1 adhesion gene for M. pneumoniae and the ompA gene for C. pneumoniae, was compared with two conventional PCR assays targeting the 16S rRNA gene and the ompA gene. A total of 120 clinical throat and nasopharyngeal swab samples were tested. DNA extraction was performed using an alkali denaturation/neutralization method, and real-time amplification, detection, and data analysis were performed using a Rotor-Gene 2000 real-time rotary analyzer (Corbett Life Science, Sydney, Australia). Using conventional PCR as a reference in an analysis of 120 samples, 13 of 14 samples positive for C. pneumoniae were detected by the novel real-time PCR. In an analysis of M. pneumoniae, 22 samples were positive in the conventional PCR and the novel assay detected 24 positive samples. When using the conventional PCR as a reference, sensitivity and specificity were 93% and 100%, respectively, for C. pneumoniae and 100% and 98%, respectively, for M. pneumoniae. With an overall agreement of 98.8%, this suggests that performance of the new duplex real-time PCR is comparable to that of conventional PCR.

  • 32. Gyarmati, Péter
    et al.
    Conze, Tim
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology, Molecular tools.
    Zohari, Siamak
    LeBlanc, Neil
    Nilsson, Mats
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology, Molecular tools.
    Landegren, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology, Molecular tools.
    Banér, Johan
    Belák, Sándor
    Simultaneous genotyping of all hemagglutinin and neuraminidase subtypes of avian influenza viruses by use of padlock probes2008In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 46, no 5, p. 1747-1751Article in journal (Refereed)
    Abstract [en]

    A subtyping assay for both the hemagglutinin (HA) and neuraminidase (NA) surface antigens of the avian influenza virus (AIV) has been developed. The method uses padlock probe chemistry combined with a microarray output for detection. The outstanding feature of this assay is its capability to designate both the HA and the NA of an AIV sample from a single reaction mixture. A panel of 77 influenza virus strains was tested representing the entire assortment of the two antigens. One hundred percent (77/77) of the samples tested were identified as AIV, and 97% (75/77) were subtyped correctly in accordance with previous examinations performed by classical diagnostic methods. Testing of heterologous pathogens verified the specificity of the assay. This assay is a convenient and practical tool for the study of AIVs, providing important HA and NA data more rapidly than conventional methods.

  • 33.
    Gylfe, Åsa
    et al.
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Olsen, Björn
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Infectious Diseases.
    Strasevicius, Darius
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Marti Ras, Nuria
    Weihe, Pál
    Noppa, Laila
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology.
    Östberg, Yngve
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Baranton, Guy
    Bergström, Sven
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Isolation of Lyme disease Borrelia from puffins (Fratercula arctica) and seabird ticks (Ixodes uriae) on the Faeroe Islands1999In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 37, no 4, p. 890-896Article in journal (Refereed)
    Abstract [en]

    This is the first report on the isolation of Lyme disease Borrelia from seabirds on the Faeroe Islands and the characteristics of its enzootic cycle. The major components of the Borrelia cycle include the puffin (Fratercula arctica) as the reservoir and Ixodes uriae as the vector. The importance of this cycle and its impact on the spread of human Lyme borreliosis have not yet been established. Borrelia spirochetes isolated from 2 of 102 sampled puffins were compared to the borreliae previously obtained from seabird ticks, I. uriae. The rrf-rrl intergenic spacer and the rrs and the ospC genes were sequenced and a series of phylogenetic trees were constructed. Sequence data and restriction fragment length polymorphism analysis grouped the strains together with Borrelia garinii. In a seroepidemiological survey performed with residents involved in puffin hunting on the Faeroe Islands, 3 of 81 serum samples were found to be positive by two commonly used clinical tests: a flagellin-based enzyme-linked immunosorbent assay (ELISA) and Western blotting. These three positive serum samples also had high optical density values in a whole-cell ELISA. The finding of seropositive Faeroe Islanders who are regularly exposed to I. uriae indicate that there may be a transfer of B. garinii by this tick species to humans.

  • 34.
    Henning, Claes
    et al.
    Sahlgrenska University Södra Älvsborgs Hospital.
    Aygül, Nilsu
    Karolinska Institutet.
    Dinnétz, Patrik
    Södertörn University, School of Natural Sciences, Technology and Environmental Studies, Environmental Science.
    Wallgren, Karin
    Karolinska University Hospital.
    Özenci, Volkan
    Karolinska Institutet / Karolinska University Hospital.
    Detailed analysis of the characteristics of sample volume in blood culture bottles2019In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 57, no 8, article id e00268-19Article in journal (Refereed)
    Abstract [en]

    Blood volume is the most important variable, for detection of microorganisms in blood cultures (BC). Most standards recommend 40-60 ml blood, collected in several BC bottles filled up to 10 ml. We measured blood volume in individual BC bottles, and analysed the association of hospital, bottle type, day of the week, daily sampling time, and age and gender of the patient, with sampling volume and BC result. The variation in blood volume per BC bottle was analysed in a mixed linear model using hospital, bottle type, weekday, sampling time, age and gender as fixed factors, and patient ID and episode as random factors to control for repetitive sampling of individual patients. Only 18 % of all bottles were filled with the recommended 8-10 ml, and 47 % were filled with less than 8 ml. The mean (±SE) volume was larger in positive 9.09 (±0.15) compared to negative bottles 8.47 (±0.07) (p<0.001). Blood volume was larger in BacT/ALERT-FA Plus than in -FN Plus BC bottles (p<0.001). There was significantly lower volumes collected during the night (p<0.001). The volume of blood collected decreased significantly with increasing patient age (p<0.001). Larger volumes were collected from males compared to females, 8.78 (±0.06) vs. 8.36 (±0.06) ml (mean ± SE) respectively (p<0.001). The odds of detecting a positive patient increases with 13 % for each additional ml blood drawn. Our results show that we need to work actively with development of blood sampling routines to overcome age and gender effects, and to optimize blood sampling volumes.

  • 35. Hepojoki, Satu
    et al.
    Rusanen, Juuso
    Hepojoki, Jussi
    Nurmi, Visa
    Vaheri, Antti
    Lundkvist, Åke
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Hedman, Klaus
    Vapalahti, Olli
    Competitive Homogeneous Immunoassay for Rapid Serodiagnosis of Hantavirus Disease2015In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 53, no 7, p. 2292-2297Article in journal (Refereed)
    Abstract [en]

    In this study, we describe a competitive homogeneous immunoassay that makes use of Forster resonance energy transfer (FRET) in rapid detection of pathogen-specific antibodies. The assay principle is based on competition between a monoclonal antibody (MAb) and serum antibodies to a given antigen. In the assay, named competitive FRET immunoassay (CFRET-IA), the FRET signal is induced if MAb carrying a donor label binds to an acceptor-labeled antigen. Specific antibodies in serum compete for antigen binding, resulting in reduced FRET signal. The proof-of-principle for the assay was obtained using donor-labeled Puumala virus nucleocapsid protein (PUUV-N) and acceptor-labeled anti-PUUV-N MAb. The assay was evaluated by analyzing 329 clinical samples comprising 101 from individuals with acute PUUV infection, 42 from individuals with past infection, and 186 from individuals with PUUV-seronegative sera, and the results were compared to those of reference tests. The rapid serodiagnostic test we introduced herein performed with 100% sensitivity and 99% specificity for diagnosing acute hantavirus disease.

  • 36. Herrera, Phabiola M
    et al.
    Mendez, Melissa
    Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
    Velapatiño, Billie
    Santivañez, Livia
    Balqui, Jacqueline
    Finger, S Alison
    Sherman, Jonathan
    Zimic, Mirko
    Cabrera, Lilia
    Watanabe, Jose
    Rodríguez, Carlos
    Gilman, Robert H
    Berg, Douglas E
    DNA-level diversity and relatedness of Helicobacter pylori strains in shantytown families in Peru and transmission in a developing-country setting.2008In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 46, no 12, p. 3912-3918Article in journal (Refereed)
    Abstract [en]

    The efficiency of transmission of a pathogen within families compared with that between unrelated persons can affect both the strategies needed to control or eradicate infection and how the pathogen evolves. In industrialized countries, most cases of transmission of the gastric pathogen Helicobacter pylori seems to be from mother to child. An alternative model, potentially applicable among the very poor in developing countries, where infection is more common and the sanitary infrastructure is often deficient, invokes frequent transmission among unrelated persons, often via environmental sources. In the present study, we compared the genotypes of H. pylori from members of shantytown households in Peru to better understand the transmission of H. pylori in developing-country settings. H. pylori cultures and/or DNAs were obtained with informed consent by the string test (a minimally invasive alternative to endoscopy) from at least one child and one parent from each of 62 families. The random amplified polymorphic DNA fingerprints of 57 of 81 (70%) child-mother strain pairs did not match, nor did the diagnostic gene sequences (>1% DNA sequence difference), independent of the child's age (range, 1 to 39 years). Most strains from siblings or other paired family members were also unrelated. These results suggest that H. pylori infections are often community acquired in the society studied. Transmission between unrelated persons should facilitate the formation of novel recombinant genotypes by interstrain DNA transfer and selection for genotypes that are well suited for individual hosts. It also implies that the effective prevention of H. pylori infection and associated gastroduodenal disease will require anti-H. pylori measures to be applied communitywide.

  • 37. Herrmann, Bjorn
    et al.
    Isaksson, Jenny
    Ryberg, Martin
    Tångrot, Jeanette
    Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine).
    Saleh, Isam
    Versteeg, Bart
    Gravningen, Kirsten
    Bruisten, Sylvia
    Global Multilocus Sequence Type Analysis of Chlamydia trachomatis Strains from 16 Countries2015In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 53, no 7, p. 2172-2179Article in journal (Refereed)
    Abstract [en]

    The Uppsala University Chlamydia trachomatis multilocus sequence type (MLST) database (http://mlstdb.bmc.uu.se) is based on five target regions (non-housekeeping genes) and the ompA gene. Each target has various numbers of alleles-hctB, 89; CT058, 51; CT144, 30; CT172, 38; and pbpB, 35-derived from 13 studies. Our aims were to perform an overall analysis of all C. trachomatis MLST sequence types (STs) in the database, examine STs with global spread, and evaluate the phylogenetic capability by using the five targets. A total of 415 STs were recognized from 2,089 specimens. The addition of 49 ompA gene variants created 459 profiles. ST variation and their geographical distribution were characterized using eBURST and minimum spanning tree analyses. There were 609 samples from men having sex with men (MSM), with 4 predominating STs detected in this group, comprising 63% of MSM cases. Four other STs predominated among 1,383 heterosexual cases comprising, 31% of this group. The diversity index in ocular trachoma cases was significantly lower than in sexually transmitted chlamydia infections. Predominating STs were identified in 12 available C. trachomatis whole genomes which were compared to 22 C. trachomatis full genomes without predominating STs. No specific gene in the 12 genomes with predominating STs could be linked to successful spread of certain STs. Phylogenetic analysis showed that MLST targets provide a tree similar to trees based on whole-genome analysis. The presented MLST scheme identified C. trachomatis strains with global spread. It provides a tool for epidemiological investigations and is useful for phylogenetic analyses.

  • 38.
    Herrmann, Björn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Microbiology and Infectious Medicine, Clinical Bacteriology.
    Isaksson, Jenny
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Microbiology and Infectious Medicine, Clinical Bacteriology.
    Ryberg, Martin
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Organismal Biology, Systematic Biology.
    Tangrot, Jeanette
    Saleh, Isam
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Microbiology and Infectious Medicine, Clinical Bacteriology.
    Versteeg, Bart
    Gravningen, Kirsten
    Bruisten, Sylvia
    Global Multilocus Sequence Type Analysis of Chlamydia trachomatis Strains from 16 Countries2015In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 53, no 7, p. 2172-2179Article in journal (Refereed)
    Abstract [en]

    The Uppsala University Chlamydia trachomatis multilocus sequence type (MLST) database (http://mlstdb.bmc.uu.se) is based on five target regions (non-housekeeping genes) and the ompA gene. Each target has various numbers of alleles-hctB, 89; CT058, 51; CT144, 30; CT172, 38; and pbpB, 35-derived from 13 studies. Our aims were to perform an overall analysis of all C. trachomatis MLST sequence types (STs) in the database, examine STs with global spread, and evaluate the phylogenetic capability by using the five targets. A total of 415 STs were recognized from 2,089 specimens. The addition of 49 ompA gene variants created 459 profiles. ST variation and their geographical distribution were characterized using eBURST and minimum spanning tree analyses. There were 609 samples from men having sex with men (MSM), with 4 predominating STs detected in this group, comprising 63% of MSM cases. Four other STs predominated among 1,383 heterosexual cases comprising, 31% of this group. The diversity index in ocular trachoma cases was significantly lower than in sexually transmitted chlamydia infections. Predominating STs were identified in 12 available C. trachomatis whole genomes which were compared to 22 C. trachomatis full genomes without predominating STs. No specific gene in the 12 genomes with predominating STs could be linked to successful spread of certain STs. Phylogenetic analysis showed that MLST targets provide a tree similar to trees based on whole-genome analysis. The presented MLST scheme identified C. trachomatis strains with global spread. It provides a tool for epidemiological investigations and is useful for phylogenetic analyses.

  • 39.
    Herrmann, Björn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Larsson, Viviana Cavaglia
    Klinisk virologi.
    Rubin, Carl-Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Sund, Fredrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Eriksson, Britt-Marie
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Arvidson, Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Women's and Children's Health, Pediatrics.
    Yun, Zhibing
    Bondeson, Kåre
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Virology.
    Blomberg, Jonas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Virology.
    Comparison of a duplex quantitative real-time PCR assay and the COBAS Amplicor CMV Monitor test for detection of cytomegalovirus2004In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 42, no 5, p. 1909-14Article in journal (Refereed)
    Abstract [en]

    A duplex quantitative real-time PCR (qPCR) assay was designed to detect both the polymerase gene (pol) and the glycoprotein gene (gB) of cytomegalovirus (CMV). The detection limit of the qPCR was determined to be 1 to 3 copies/reaction and the linear measure interval was 10(3) to 10(8) copies/ml. The qPCR system was compared to the COBAS Amplicor CMV Monitor test (COBAS) by an analysis of 138 plasma samples. Both systems detected CMV in 71 cases and had negative results for 33 samples. In addition, 34 samples were positive by qPCR and negative by the COBAS assay, but in no case was the COBAS result positive and the qPCR result negative. Thus, qPCR detected 48% more positive cases than the COBAS method. For samples with > or = 10(5) copies/ml by qPCR, a saturation effect was seen in the COBAS assay and quantification required dilution. Copy numbers for pol and gB by qPCR generally agreed. However, the reproducibility of qPCR assays and the need for an international standard are discussed. Discrepant copy numbers for pol and gB by qPCR were found for samples from two patients, and sequence analysis revealed that the corresponding CMV strains were mismatched at four nucleotide positions compared with the gB fragment primer sequences. In conclusion, a duplex qPCR assay in a real-time format facilitates quantitative measurements and minimizes the risk of false-negative results.

  • 40.
    Herrmann, Björn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Winqvist, Ola
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Internal Medicine.
    Mattsson, Jens G
    Kirsebom, Leif A
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology, Microbiology.
    Differentiation of Chlamydia spp by sequence determination and restriction endonuclease cleavage of RNase P RNA genes1996In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 34, no 8, p. 1897-1902Article in journal (Refereed)
    Abstract [en]

    The amplification of DNA from Chlamydia trachomatis by PCR with degenerated primers yielded a 345-bp fragment of the putative RNase P RNA gene. From the deduced DNA sequence of this gene in C. trachomatis, a modified primer pair was designed. The primer pair was subsequently used to obtain the corresponding gene products from Chlamydia pneumoniae and Chlamydia psittaci. Sequence comparisons revealed similarities of 76.6% between C. trachomatis and C. pneumoniae, 79.5% between C. trachomatis and C. psittaci, and 84.7% between C. pneumoniae and C. psittaci. Furthermore, the three species were differentiated by fragment length polymorphism analysis after restriction enzyme cleavage of the PCR products. Sequence variations among 14 serotypes of C. trachomatis were confined to one purine base substitution in the putative RNase P RNA gene of lymphogranuloma venereum strains L1 to L3. Complete sequence similarity was found for nine strains of C. pneumoniae of different geographic origins. Taken together, our results indicate a possibility of the general application of this method in clinical bacteriology. Analysis of the secondary structures of the putative RNase P RNA genes from the different Chlamydia species suggested that a novel structural element in the domain of RNase P RNA is involved in base pairing with the 3'-terminal CCA motif of a tRNA precursor. This structure has not previously been found among RNase P RNAs of members of the division Bacteria.

  • 41.
    Heymans, Raymond
    et al.
    Public Health Laboratory, Cluster of Infectious Diseases, Health Service of Amsterdam, Amsterdam, Netherlands.
    Golparian, Daniel
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sverige; Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Bruisten, Sylvia M.
    Public Health Laboratory, Cluster of Infectious Diseases, Health Service of Amsterdam, Amsterdam, Netherlands; Department of Experimental Virology, University of Amsterdam, Amsterdam, Netherlands.
    Schouls, Leo M.
    Laboratory for Infectious Diseases and Perinatal Screening, National Institute for Public Health and the Environment (RIVM), Bilthoven, Netherlands.
    Unemo, Magnus
    Örebro University Hospital. WHO Collaborating Centre for Gonorrhoea and other STIs, National Reference Laboratory for Pathogenic Neisseria, Örebro University Hospital, Örebro, Sverige; Department of Laboratory Medicine, Microbiology, Örebro University Hospital, Örebro, Sweden.
    Evaluation of Neisseria gonorrhoeae Multiple-Locus Variable-Number Tandem-Repeat Analysis, N. gonorrhoeae Multiantigen Sequence Typing, and Full-Length porB Gene Sequence Analysis for Molecular Epidemiological Typing2012In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 50, no 1, p. 180-183Article in journal (Refereed)
    Abstract [en]

    The performance characteristics of Neisseria gonorrhoeae multilocus variable-number tandem-repeat analysis were evaluated, by comparison with N. gonorrhoeae multiantigen sequence typing and full-length porB sequence typing. Assessment of the relatedness of intra- and interpatient isolates showed that all three genotyping techniques display a high resolution and typeability.

  • 42. Hjelle, B
    et al.
    Jenison, S
    Torrez-Martinez, N
    Herring, B
    Quan, S
    Polito, A
    Pichuantes, S
    Yamada, T
    Morris, C
    Elgh, Fredrik
    Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Virology.
    Lee, H W
    Artsob, H
    Dinello, R
    Rapid and specific detection of Sin Nombre virus antibodies in patients with hantavirus pulmonary syndrome by a strip immunoblot assay suitable for field diagnosis.1997In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 35, no 3, p. 600-8Article in journal (Refereed)
    Abstract [en]

    To develop a rapid antibody test for Sin Nombre hantavirus (SNV) infection for diagnosis of hantavirus pulmonary syndrome (HPS) in field settings where advanced instrumentation is not available, a strip immunoblot assay bearing four immobilized antigens for SNV and a recombinant nucleocapsid protein antigen of Seoul hantavirus (SEOV) was prepared. The SNV antigens included a full-length recombinant-expressed nucleocapsid (N) protein (rN), a recombinant-expressed G1 protein (residues 35 to 117), and synthetic peptides derived from N (residues 17 to 59) and G1 (residues 55 to 88). On the basis of the observed reactivities of hantavirus-infected patient and control sera, we determined that a positive assay requires reactivity with SNV or SEOV rN antigen and at least one other antigen. Isolated reactivity to either viral rN antigen is indeterminate, and any pattern of reactivity that does not include reactivity to an rN antigen is considered indeterminate but is unlikely to represent hantavirus infection. Fifty-eight of 59 samples from patients with acute SNV-associated HPS were positive according to these criteria, and one was initially indeterminate. Four of four samples from patients with HPS due to other hantaviruses were positive, as were most samples from patients with SEOV and Puumala virus infections. Of 192 control serum samples, 2 (1%) were positive and 2 were indeterminate. Acute SNV infection was distinguishable from remote SNV infection or infection with hantaviruses other than SNV by the presence of G1 peptide antigen reactivities in the former. The strip immunoblot assay shows promise for the detection of SNV antibodies early in the course of HPS.

  • 43.
    Hjorth, SV
    et al.
    Statens Serum Institut, DK-2300 Copenhagen .
    Björnelius, Eva
    Karolinska Institutet.
    Lidbrink, Peter
    Karolinska Institutet.
    Falk, Lars
    Örebro University Hospital.
    Dohn, B
    Statens Serum Institut, DK-2300 Copenhagen .
    Berthelsen, L
    Statens Serum Institut, DK-2300 Copenhagen .
    Ma, L
    Louisiana State University Health Sciences Center.
    Martin, DH
    Louisiana State University Health Sciences Center.
    Jensen, Jörgen Skov
    Statens Serum Institut, DK-2300 Copenhagen .
    Sequence-based typing of Mycoplasma genitalium reveals sexual transmission2006In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 44, no 6, p. 2078-2083Article in journal (Refereed)
    Abstract [en]

    Mycoplasma genitalium causes male nonchlamydial, nongonococcal urethritis and is associated with cervicitis and pelvic inflammatory disease in women. Epidemiological studies indicate that M. genitalium is sexually transmitted, and the aim of the present study was to further substantiate this by means of a DNA typing system. A typing assay based on a diagnostic mgpB gene PCR was developed, evaluated, and applied directly to urogenital specimens. The assay had a low limit of detection and hence a high typeability. Sequences of isolates from 52 unrelated patients were divided into 29 different sequence types, giving a discriminatory index of 0.95. Two to six M. genitalium-positive specimens were collected from each of 44 patients over a median interval of 56 days (range, 11 to 1,395). Forty had the same sequence type in consecutive specimens. Specimens collected from two men were repeatedly positive at intervals of 472 and 1,395 days, respectively, but the sequence types had changed. A new strain was introduced in one sexual dyad, and the sequence types changed subsequently. Seventy-nine M. genitalium-positive specimens from 19 couples were investigated, and all partners initially had concordant sequence types, but one couple had discordant types at one time point before a newly introduced strain took over. The present typing system is simple and reproducible and has an excellent discriminatory capacity which might prove useful in studies of sexual networks and for evaluation of treatment failures. In the laboratory, this system may document the uniqueness of newly isolated M. genitalium strains.

  • 44. Hogberg, Liselotte
    et al.
    Geli, Patricia
    Stockholm University, Faculty of Science, Department of Mathematics.
    Ringberg, Hakan
    Melander, Eva
    Lipsitch, Marc
    Ekdahl, Karl
    Age- and serogroup-related differences in observed durations of nasopharyngeal carriage of penicillin-resistant pneumococci2007In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 45, no 3, p. 948-952Article in journal (Refereed)
    Abstract [en]

    Using data from an ongoing Swedish intervention project, the observed durations of nasopharyngeal carriage of penicillin-nonsusceptible Streptococcus pneumoniae (PNSP) (MIC of penicillin G of >= 0.5 mu g/ml) stratified by both pneumococcal serogroup and age of the carrier were compared. The means and 95% confidence intervals (CIs) were estimated by fitting a gamma distribution to the observed duration of carriage for each age and serogroup stratum. The mean observed duration of carriage for all cases was 37 days (95% CI, 35 to 38 days). Children below the age of 5 years carried PNSP for significantly longer periods (43 days; 95% CI, 41 to 45 days) compared with older individuals (25 days; 95% CI, 24 to 27 days). There were also differences within the group of cases below the age of 5 years, as the duration of carriage became significantly shorter for each increasing age step: < 1, 1 to 2, and 3 to 4 years. In addition, patients < 5 years of age carried serogroups 9 and 14 for significantly shorter periods than groups 6 and 23. Serogroup 9 was also carried for significantly shorter periods than group 19. For patients aged 5 years or older, no significant difference in carriage duration for different ages or serogroups could be noted. As young children have the longest duration of PNSP carriage, interventions aiming to reduce the prevalence in this group are of great importance. The results highlight the importance of taking both serogroup and age of the carriers into account when studying the dynamics of pneumococcal transmission in young children.

  • 45.
    Holmberg, Martin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    McGill, Svena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Ehrenborg, Christian
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Wesslen, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Hjelm, Eva
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Darelid, J
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Blad, Lars
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Engstrand, L
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Regnery, Russell
    Friman, Göran
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Evaluation of human seroreactivity to Bartonella species in Sweden1999In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 37, no 5, p. 1381-1384Article in journal (Refereed)
    Abstract [en]

    Among the species that compose the expanding genus Bartonella, thus far only B. henselae and B. quintana have reportedly been isolated from humans in Europe. To evaluate the prevalence of Bartonella infection in Sweden,we conducted a retrospective serological examination of 126 human serum samples. These samples were analyzed for antibodies to B. henselae, B. quintana, and B. elizabethae, Serum samples from 100 blood donors, who spanned the ages of 20 to 60 and had no apparent clinical signs of illness, were also studied as a control group. An immunoglobulin G indirect fluorescence antibody assay revealed 4 and 8.3% Bartonella positivity rates for the blood donor and patient group, respectively, when a cutoff titer of greater than or equal to 64 was chosen. Among the blood donors, four were seropositive to B, elizabethae; one of these also had concordant positive titer to B. henselae, In the patient group, 14 serum samples were positive against Bartonella spp, These serum specimens represented nine patients. In three of these seropositive patients, paired serum samples displayed a fourfold increase in antibody titer to at least one of the three antigens, These three patients are discussed. In this report we also present a case study of a 60-year-old Swedish male with fatal myocarditis, Postmortem serological analysis revealed a high titer against B. elizabethae, PCR and nucleotide sequencing of the myocardial tissue from this patient, and of Liver tissue from one of the other three patients, showed sequences similar to B. quintana, The age, geographical origin, animal contacts, and serological response pattern to the different Bartonella antigens differed among the four patients. This study substantiates the presence of Bartonella spp, in Sweden, documents the seroreactivity to three Bartonella antigens in Swedish patients, and reports the first two cases of B. quintana-like infections in Sweden.

  • 46. Innings, Åsa
    et al.
    Ullberg, Måns
    Johansson, Anders
    Linköping University, Faculty of Health Sciences. Linköping University, Department of Molecular and Clinical Medicine, Clinical Microbiology. Östergötlands Läns Landsting, Centre for Laboratory Medicine, Department of Clinical Microbiology.
    Rubin, Carl Johan
    Noreus, Niklas
    Isaksson, Magnus
    Herrman, Björn
    Multiplex real-time PCR targeting the RNase P RNA gene for detection and identification of Candida species in blood2007In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 45, no 3, p. 874-880Article in journal (Refereed)
    Abstract [en]

    We have developed a single-tube multiplex real-time PCR method for the detection of the eight most common Candida species causing septicemia: Candida albicans, C. dubliniensis, C. famata, C. glabrata, C. guilliermondii, C. krusei, C. parapsilosis, and C. tropicalis. The method developed targets the RNase P RNA gene RPR1. Sequences of this geiie were determined for seven of the Candida species and showed surprisiRgly large sequence variation. C. glabrata was found to have a gene that was five times longer gene than those of the other species, and the nucleotide sequence similarity between C. krusei and C. albicans was as low as 55%. The multiplex PCR contained three probes that enabled the specific detection of C. albicans, C. glabrata, and C. krusei and a fourth probe that allowed the general detection of the remaining species. The method was able to detect 1 to 10 genome copies when the detection limit was tested repeatedly for the four species C. albicans, C. glabrata, C. krusei, and C. guilliermondii. No significant difference in the detection limit was seen when the multiplex format was compared with single-species PCR, i.e., two primers and one probe. The method detected eight clinically relevant Candida species and did not react with other tested non-Candida species or human DNA. The assay was applied to 20 blood samples from nine patients and showed a sensitivity similar to that of culture. Copyright © 2007, American Society for Microbiology. All Rights Reserved.

  • 47.
    Innings, Åsa
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Ullberg, Måns
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Johansson, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Rubin, Carl Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Noreus, Niklas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Isaksson, Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Herrmann, Björn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Multiplex real-time PCR targeting the RNase P RNA gene for detection and identification of Candida species in blood2007In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 45, no 3, p. 874-880Article in journal (Refereed)
    Abstract [en]

    We have developed a single-tube multiplex real-time PCR method for the detection of the eight most common Candida species causing septicemia: Candida albicans, C. dubliniensis, C. famata, C. glabrata, C. guilliermondii, C. krusei, C. parapsilosis, and C. tropicalis. The method developed targets the RNase P RNA gene RPR1. Sequences of this geiie were determined for seven of the Candida species and showed surprisiRgly large sequence variation. C. glabrata was found to have a gene that was five times longer gene than those of the other species, and the nucleotide sequence similarity between C. krusei and C. albicans was as low as 55%. The multiplex PCR contained three probes that enabled the specific detection of C. albicans, C. glabrata, and C. krusei and a fourth probe that allowed the general detection of the remaining species. The method was able to detect 1 to 10 genome copies when the detection limit was tested repeatedly for the four species C. albicans, C. glabrata, C. krusei, and C. guilliermondii. No significant difference in the detection limit was seen when the multiplex format was compared with single-species PCR, i.e., two primers and one probe. The method detected eight clinically relevant Candida species and did not react with other tested non-Candida species or human DNA. The assay was applied to 20 blood samples from nine patients and showed a sensitivity similar to that of culture.

  • 48.
    Jakobsson, Tell
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Obstetrics and gynecology. Linköping University, Faculty of Health Sciences.
    Forsum, Urban
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Microbiology. Linköping University, Faculty of Health Sciences.
    Lactobacillus iners: a marker of changes in the vaginal flora?2007In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 45, no 9, p. 3145-Article in journal (Other academic)
  • 49.
    Jakobsson, Tell
    et al.
    Linköping University, Department of Clinical and Experimental Medicine, Obstetrics and gynecology. Linköping University, Faculty of Health Sciences.
    Forsum, Urban
    Linköping University, Department of Clinical and Experimental Medicine, Clinical Microbiology. Linköping University, Faculty of Health Sciences. Östergötlands Läns Landsting, Centre for Laboratory Medicine, Department of Clinical Microbiology.
    The predominant Human vaginal Lactobacillus flora during IVF treatment2008In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 7, no 14, p. 1-9Article in journal (Refereed)
    Abstract [en]

    Background: Signature matching of nucleotide sequences in the V1 and V3 regions 16S rRNA genes using pyrosequencing technology is a powerful tool for typing vaginal Lactobacilli to the species level and has been used for investigating the vaginal microbial niche.

    Methods: This study has characterized the normal cultivable vaginal Lactobacillus flora at varying estradiol levels in plasma; the study comprised 17 patients undergoing ovarian stimulation for In Vitro Fertilization (IVF) treatment. The vaginal status of each participant was initially assessed as normal according to Amsel and Nugent criteria.

    Results: L. crispatus, L. gasseri and/or L. jensenii were present in 10 of the patients throughout the study period, and little variation among these three species was encountered in individual patients. The flora of three women was dominated by L. delbrüeckii, L. rhamnosus or L. vaginalis. One woman exhibited a dominance of L. iners. The flora of the remaining three women were initially dominated by L. rhamnosus or L. reuteri, but as their estrogen levels rose, their flora composition altered, to become dominated by one of the three species most common in a normal, healthy vagina.

    Conclusion: Signature matching of nucleotide sequences in the V1 and V3 regions of 16S rRNA genes is a discriminative tool for the study of vaginal Lactobacilli and can be used to track the Lactobacillus flora under a variety of physiological conditions.

  • 50.
    Johansson, Anders
    Umeå University, Faculty of Medicine, Clinical Microbiology. Umeå University, Faculty of Medicine, Clinical Microbiology, Infectious Diseases.
    Comparative analysis of PCR versus culture for diagnosis of ulceroglandular tularemia2000In: Journal of Clinical Microbiology, ISSN 0095-1137, E-ISSN 1098-660X, Vol. 38, no 1, p. 22-26Article in journal (Refereed)
    Abstract [en]

    PCR and culture were comparatively evaluated for their abilities to demonstrate Francisella tularensis in wound specimens from tularemia patients during an outbreak in Sweden in 1998. For transport of the specimens used for PCR, a buffer solution containing a nuclease inhibitor was used, and for transport of the specimens used for culture, a commercial transport system was selected after experimental comparison of various systems. Of 40 patients with culture- and/or serology-verified ulceroglandular tularemia, PCR detected F. tularensis DNA in 30 (75%) patients, whereas culture detected bacterial growth in 25 (62%) patients. Compared to data from a previous study, the present inclusion of a nuclease inhibitor in the transport medium did not improve the sensitivity of the PCR, whereas the sensitivity of the culture procedure was significantly increased by selection of the system used for transport. Among eight patients with clinically suspected tularemia but with negative serology and culture, specimens from four patients showed detectable DNA. In three of these patients the diagnosis was verified by the demonstration of an F. tularensis-specific T-cell response in vitro. In conclusion, PCR was more sensitive than culture for demonstration of F. tularensis in wound specimens. Besides, we showed that tularemia may proceed without development of serum antibodies, and in these patients, PCR may be of special importance for verification of the diagnosis.

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