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Arend H. J. Kolk

Other affiliations: Royal Tropical Institute
Bio: Arend H. J. Kolk is an academic researcher from University of Amsterdam. The author has contributed to research in topics: Mycobacterium tuberculosis & Sputum. The author has an hindex of 15, co-authored 24 publications receiving 4237 citations. Previous affiliations of Arend H. J. Kolk include Royal Tropical Institute.

Papers
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Journal ArticleDOI
TL;DR: A novel method based on strain-dependent hybridization patterns of in vitro-amplified DNA with multiple spacer oligonucleotides was found to differentiate M. bovis from M. tuberculosis, a distinction which is often difficult to make by traditional methods.
Abstract: Widespread use of DNA restriction fragment length polymorphism (RFLP) to differentiate strains of Mycobacterium tuberculosis to monitor the transmission of tuberculosis has been hampered by the need to culture this slow-growing organism and by the level of technical sophistication needed for RFLP typing. We have developed a simple method which allows simultaneous detection and typing of M. tuberculosis in clinical specimens and reduces the time between suspicion of the disease and typing from 1 or several months to 1 or 3 days. The method is based on polymorphism of the chromosomal DR locus, which contains a variable number of short direct repeats interspersed with nonrepetitive spacers. The method is referred to as spacer oligotyping or "spoligotyping" because it is based on strain-dependent hybridization patterns of in vitro-amplified DNA with multiple spacer oligonucleotides. Most of the clinical isolates tested showed unique hybridization patterns, whereas outbreak strains shared the same spoligotype. The types obtained from direct examination of clinical samples were identical to those obtained by using DNA from cultured M. tuberculosis. This novel preliminary study shows that the novel method may be a useful tool for rapid disclosure of linked outbreak cases in a community, in hospitals, or in other institutions and for monitoring of transmission of multidrug-resistant M. tuberculosis. Unexpectedly, spoligotyping was found to differentiate M. bovis from M. tuberculosis, a distinction which is often difficult to make by traditional methods.

2,845 citations

Journal ArticleDOI
TL;DR: The results of the study show that the implementation of an effective system for monitoring sensitivity and specificity is required before the PCR can be used reliably in the diagnosis of tuberculosis.
Abstract: PCR is, in principle, a simple and rapid test for use in the detection of Mycobacterium tuberculosis. However, virtually no data are available on the reliability and reproducibility of the method. In order to assess the validity of PCR for the detection of mycobacteria in clinical samples, seven laboratories participated in a blinded study of 200 sputum, saliva, and water samples containing either known numbers of Mycobacterium bovis BCG cells or no added organisms. Each laboratory used its own protocol for pretreatment, DNA extraction, and detection of the amplification product. Insertion sequence IS6110 was the target for DNA amplification. Several participating laboratories reported high levels of false-positive PCR results, with rates ranging from 3 to 20% and with one extreme value of 77%. The levels of sensitivity also ranged widely among the different participants. A positive PCR result was reported for 2 to 90% of the samples with 10(3) mycobacteria. Although most participants did include control tests to check the sensitivity and specificity of the PCR, the sequence of operations from sample pretreatment to purification of DNA from bacteria was not always monitored adequately. During these procedures cross-contaminating DNA was introduced and/or bacterial DNA was lost. The results of the study show that the implementation of an effective system for monitoring sensitivity and specificity is required before the PCR can be used reliably in the diagnosis of tuberculosis.

421 citations

Journal ArticleDOI
TL;DR: The DNA sequence of a previously cloned Mycobacterium bovis BCG gene encoding an immunogenic 64-kilodalton protein, MbaA, was reported, supporting the previously observed strong reactivity of human T-cell clones with this, for mycobacteria, common antigen.
Abstract: We report the DNA sequence of a previously cloned Mycobacterium bovis BCG gene encoding an immunogenic 64-kilodalton protein. This protein, MbaA, was purified from overproducing Escherichia coli K-12 cells, and the presence of antibodies to MbaA in human sera was determined by an enzyme-linked immunosorbent assay. In about 80% of serum samples from tuberculosis patients and in about 60% of samples from BCG-vaccinated individuals, significant levels of anti-MbaA antibodies were found. Surprisingly, in about 30% of the control serum samples obtained from children, anti-MbaA antibodies were also observed. Guinea pigs sensitized with M. bovis BCG or MbaA showed a delayed-type hypersensitivity reaction after challenge with purified MbaA, supporting the previously observed strong reactivity of human T-cell clones with this, for mycobacteria, common antigen.

255 citations

Journal ArticleDOI
TL;DR: During the screening of a Mycobacterium tuberculosis lambda gt-11 gene library with monoclonal antibodies, a recombinant clone was detected which contained a mycobacterial DNA insert that hybridized specifically with DNA of M. tuberculosis complex strains.
Abstract: During the screening of a Mycobacterium tuberculosis lambda gt-11 gene library with monoclonal antibodies, we detected a recombinant clone, lambda PH7311, which contained a mycobacterial DNA insert that hybridized specifically with DNA of M. tuberculosis complex strains. Part of this insert was sequenced and used for the development of an M. tuberculosis complex-specific polymerase chain reaction (PCR). Only strains belonging to species of the M. tuberculosis complex group contained an amplifiable fragment of 158 base pairs (bp). This fragment was absent in all strains tested belonging to 15 other mycobacterial species. After amplification by PCR and dot blot hybridization with a digoxigenin-labeled oligonucleotide, the limit of detection of purified genomic M. tuberculosis DNA amounted to a quantity corresponding to 20 bacterial cells. By this technique about 10(3) M. tuberculosis bacteria were detectable in sputum. Using PCR, we were also able to detect M. tuberculosis cells in clinical material such as pleural fluid, bronchial washings, and biopsies, and these results were comparable with those obtained by classical bacterial culture. Of 34 M. tuberculosis strains, 5 did not carry the amplifiable 158-bp fragment, which occurs usually as a single copy in the chromosome. Evidence is presented that the 158-bp fragment is located near a repeated sequence in the chromosome. We presume that strains which did not carry the 158-bp fragment have lost a chromosomal segment by a genetic rearrangement induced by the repetitive DNA element. Images

238 citations

Journal Article
TL;DR: An international workshop organized and sponsored by the Immunology of Tuberculosis (IMMTUB) component of the World Health Organization (W.H.O.) Vaccine Development Programme to characterize the specificity and reaction patterns of murine monoclonal antibodies raised against various mycobacteria was held in Geneva, 3 to 5 June 1985.
Abstract: An international workshop organized and sponsored by the Immunology of Tuberculosis (IMMTUB) component of the World Health Organization (W.H.O.) Vaccine Development Programme to characterize the specificity and reaction patterns of murine monoclonal antibodies (MAbs) raised against various mycobacteria was held in Geneva, 3 to 5 June 1985. A total of 31 MAbs (28 ascites and 3 culture supernatants) generated in nine different laboratories using several mycobacterial antigens for immunization (Mycobocterium tuberculosis, virulent and avirulent strains, M. bovis BCG, and M. leprae) were submitted in early 1985 to the IMMTUB MAb bank located in the W.H.O. Immunology Research and Training Center (W.H.O./I.R.T.C.) in the Department of Pathology, University of Geneva. The samples were coded, aliquoted, and distributed to 12 laboratories for independent analysis by a variety of methods. Eight laboratories tested the reactivity and specificity of the MAbs using a spectrum of antigens prepared from up to 23 species of mycobacteria. The assay methods included enzymelinked immunosorbent assay (ELISA), radioimmunoassay (RIA), indirect immunofluorescence assay (IFA) and several additional related techniques, e.g., dot blots, Western blots, sodium dodecyl sulfate-polyacrylamide gel electrophoresis immunoperoxidase technique (SGIP), gel immunoradioassay (GIRA), etc. Several, laboratories used the same MAbs as probes to screen mycobacterium-derived recombinant DNA libraries for the expression of mycobacterium-specific protein antigens. In addition to the MAbs, several laboratories submitted various mycobacterial antigen preparations for testing. These antigens were derived by three different approaches: (i) preparations eluted from MAb immunoadsorbant columns, (ii) an antigen preparation (nonpurified) obtained by recombinant DNA techniques and expressed in Escherichia coli, and (iii) a synthetic peptide prepared by conventional solid-state peptide synthesis methods. All'of the tests were performed blindly on coded samples by the participating laboratories, and the results were sent to the W.H.O/I.R.T.C. in Geneva for compilation. The code was broken during the workshop discussion, and the results were analyzed and compared with regard to the following criteria: (i) reactivity and specificity ofMAbs for mycobacterial antigens, (ii) determination of the molecular size(s) and protease susceptibility of the antigen molecules, (iii) ability of the MAbs to detect phage in an M. tuberculosis Agtll recombinant DNA library which expressed mycobacteriumspecific target antigens, and (iv) stimulation of murine or human mycobacterium-specific T-cell populations and clones to proliferate and secrete lymphokines in vitro. The findings of individual investigators were discussed in detail during the workshop, and the results agreed upon by the participants are summarized in Tables 1 and 2. Table 1 lists the MAbs distributed, the results obtained for mycobacterial specificities, and the molecular size(s) and structure(s) of the antigens recognized by the MAbs. The specificity patterns of the MAbs for different mycobacteria varied and for the purpose of this report were divided into four categories: M. tuberculosis specific; M. tuberculosis complex (i.e., reacting with M. tuberculosis, M. bovis BCG, and M. africanum); limited cross-reactivity (reacting with a restricted number of additional mycobacteria), and broad cross-reactivity (reacting with a majority of the strains tested). The results for ELISA and RIA were similar for the vast majority of MAbs tested; hence, they were grouped together (with one exception, IT 21). There was substantial variation in the apparent molecular sizes of the antigens recognized by the MAbs (ranging from 12 to 80 kilodaltons [kDa], with single to multiple bands). However, several MAbs reacted with molecules of similar (if not identical) size, and in many cases the reaction pattern was related to the m.ycobacterial specificity, as summarized below. (i) None of the MAbs was found to be specific for a given mycobacterial species or strain when the results of all of the tests were taken into account. (ii) Six of the MAbs tested showed a specificity restricted to M. tuberculosis complex. They could be divided into two groups according to their reactivity patterns with two different molecular sizes: three MAbs (IT 1, 4, and 20) reacted with a 14-kDa molecule and another three (IT 15, 21, and 23) reacted with a 38-kDa molecule. (iii) The limited cross-reactivity pattern was represented by a series of MAbs which recognized a wide spectrum of molecular sizes: 12-kDa (IT 3), 71-kDa (IT 11), 19-kDa (IT 10, 12, 16, and 19), and multiband 65-kDa (IT 13, 31, and 33) antigens. (iv) Of the broadly cross-reactive MAbs, two (IT 17 apd 29) reacted with a 23-kDa molecule, another two (IT 9 and 32) showed multiband reactivity (20 to 80 kDa), and several others could not be classified due to poor reactions in the assay systems used. The antigens recognized by the MAbs were identified as proteins based on their sensitivity to treatment with proteolytic enzymes such as subtilisin. In several cases, however, the actual nature of the antigen could' not be established. There was no apparent relationship between the antigens and the immunization schedules used for production of the MAbs on the one hand and the characteristics of the MAbs as judged by their specificity and reaction patterns (Table 1) on the other. The MAbs were used as probes by two laboratories to detect phage in an M. tuberculosis Agtll recombinant DNA library which expressed target antigens recognized by the MAbs. Initially, a pool of the 33 MAbs was used to screen approximately 106 recombinant phage, and 134 positive signals were detected. Twenty-eight additional positive signals were obtained when a screening was carried out using a pool consisting of antibodies IT 1, 4, 13, 20, 21, 23, and 27. The phage corresponding to the positive signals were plaque purified and tested for reactivity with the individual MAbs to identify which target antigen was expressed. Six main groups of reactivity patterns were observed (Table 2). In one group (reactivity patterns A, B, C, and D), 22 phage reacted with MAbs IT 1, 4, and 20, 2 phages reacted with IT 1 and 4, 1 phage reacted with IT 4 and 20, and 1 phage reacted with IT 20 only. These data suggest that MAbs 1, 4, and 20 react with the same target antigen but not with the same epitopes on that antigen. This interpretation is consistent with crosscompetition binding data obtained using radiolabeled MAbs. Similarly, the reactivity patterns of a second group of phage (35 phage in patterns E, F, G, H, and I) indicate that antibodies IT 13, 31, and 33 react with the same antigen but

163 citations


Cited by
More filters
Journal ArticleDOI
TL;DR: A novel method based on strain-dependent hybridization patterns of in vitro-amplified DNA with multiple spacer oligonucleotides was found to differentiate M. bovis from M. tuberculosis, a distinction which is often difficult to make by traditional methods.
Abstract: Widespread use of DNA restriction fragment length polymorphism (RFLP) to differentiate strains of Mycobacterium tuberculosis to monitor the transmission of tuberculosis has been hampered by the need to culture this slow-growing organism and by the level of technical sophistication needed for RFLP typing. We have developed a simple method which allows simultaneous detection and typing of M. tuberculosis in clinical specimens and reduces the time between suspicion of the disease and typing from 1 or several months to 1 or 3 days. The method is based on polymorphism of the chromosomal DR locus, which contains a variable number of short direct repeats interspersed with nonrepetitive spacers. The method is referred to as spacer oligotyping or "spoligotyping" because it is based on strain-dependent hybridization patterns of in vitro-amplified DNA with multiple spacer oligonucleotides. Most of the clinical isolates tested showed unique hybridization patterns, whereas outbreak strains shared the same spoligotype. The types obtained from direct examination of clinical samples were identical to those obtained by using DNA from cultured M. tuberculosis. This novel preliminary study shows that the novel method may be a useful tool for rapid disclosure of linked outbreak cases in a community, in hospitals, or in other institutions and for monitoring of transmission of multidrug-resistant M. tuberculosis. Unexpectedly, spoligotyping was found to differentiate M. bovis from M. tuberculosis, a distinction which is often difficult to make by traditional methods.

2,845 citations

Journal ArticleDOI
TL;DR: CRISPRFinder is described, a web service offering tools to detect CRISPRs including the shortest ones including one or two motifs, define DRs and extract spacers, and get the flanking sequences to determine the leader.
Abstract: Clustered regularly interspaced short palindromic repeats (CRISPRs) constitute a particular family of tandem repeats found in a wide range of prokaryotic genomes (half of eubacteria and almost all archaea). They consist of a succession of highly conserved regions (DR) varying in size from 23 to 47 bp, separated by similarly sized unique sequences (spacer) of usually viral origin. A CRISPR cluster is flanked on one side by an AT-rich sequence called the leader and assumed to be a transcriptional promoter. Recent studies suggest that this structure represents a putative RNA-interference-based immune system. Here we describe CRISPRFinder, a web service offering tools to (i) detect CRISPRs including the shortest ones (one or two motifs); (ii) define DRs and extract spacers; (iii) get the flanking sequences to determine the leader; (iv) blast spacers against Genbank database and (v) check if the DR is found elsewhere in prokaryotic sequenced genomes. CRISPRFinder is freely accessible at http://crispr.u-psud.fr/Server/CRISPRfinder.php.

1,689 citations

Journal ArticleDOI
TL;DR: A novel family of repetitive DNA sequences that is present among both domains of the prokaryotes but absent from eukaryotes or viruses is studied, characterized by direct repeats, varying in size from 21 to 37 bp, interspaced by similarly sized non‐repetitive sequences.
Abstract: Using in silico analysis we studied a novel family of repetitive DNA sequences that is present among both domains of the prokaryotes (Archaea and Bacteria), but absent from eukaryotes or viruses. This family is characterized by direct repeats, varying in size from 21 to 37 bp, interspaced by similarly sized non-repetitive sequences. To appreciate their characteri-stic structure, we will refer to this family as the clustered regularly interspaced short palindromic repeats (CRISPR). In most species with two or more CRISPR loci, these loci were flanked on one side by a common leader sequence of 300-500 b. The direct repeats and the leader sequences were conserved within a species, but dissimilar between species. The presence of multiple chromosomal CRISPR loci suggests that CRISPRs are mobile elements. Four CRISPR-associated (cas) genes were identified in CRISPR-containing prokaryotes that were absent from CRISPR-negative prokaryotes. The cas genes were invariably located adjacent to a CRISPR locus, indicating that the cas genes and CRISPR loci have a functional relationship. The cas3 gene showed motifs characteristic for helicases of the superfamily 2, and the cas4 gene showed motifs of the RecB family of exonucleases, suggesting that these genes are involved in DNA metabolism or gene expression. The spatial coherence of CRISPR and cas genes may stimulate new research on the genesis and biological role of these repeats and genes.

1,639 citations

Journal ArticleDOI
06 Jun 1991-Nature
TL;DR: Extrachromosomal and integrative expression vectors carrying the regulatory sequences for major BCG heat-shock proteins have been developed and can elicit long-lasting humoral and cellular immune responses to foreign antigens in mice.
Abstract: BCG, a live attenuated tubercle bacillus, is the most widely used vaccine in the world and is also a useful vaccine vehicle for delivering protective antigens of multiple pathogens. Extrachromosomal and integrative expression vectors carrying the regulatory sequences for major BCG heat-shock proteins have been developed to allow expression of foreign antigens in BCG. These recombinant BCG strains can elicit long-lasting humoral and cellular immune responses to foreign antigens in mice.

1,476 citations

Journal ArticleDOI
TL;DR: The authors suggest that the spacer elements are the traces of past invasions by extrachromosomal elements, and hypothesize that they provide the cell immunity against phage infection, and more generally foreign DNA expression, by coding an anti-sense RNA.
Abstract: Numerous prokaryote genomes contain structures known as clustered regularly interspaced short palindromic repeats (CRISPRs), composed of 25-50 bp repeats separated by unique sequence spacers of similar length. CRISPR structures are found in the vicinity of four genes named cas1 to cas4. In silico analysis revealed another cluster of three genes associated with CRISPR structures in many bacterial species, named here as cas1B, cas5 and cas6, and also revealed a certain number of spacers that have homology with extant genes, most frequently derived from phages, but also derived from other extrachromosomal elements. Sequence analysis of CRISPR structures from 24 strains of Streptococcus thermophilus and Streptococcus vestibularis confirmed the homology of spacers with extrachromosomal elements. Phage sensitivity of S. thermophilus strains appears to be correlated with the number of spacers in the CRISPR locus the strain carries. The authors suggest that the spacer elements are the traces of past invasions by extrachromosomal elements, and hypothesize that they provide the cell immunity against phage infection, and more generally foreign DNA expression, by coding an anti-sense RNA. The presence of gene fragments in CRISPR structures and the nuclease motifs in cas genes of both cluster types suggests that CRISPR formation involves a DNA degradation step.

1,344 citations