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M Achtman

Publications and source records attributed to M Achtman.

At least 37 records · Page 2Linked to original sources
PubMed1999

The relative contributions of recombination and mutation to the divergence of clones of Neisseria meningitidis.

Multilocus sequence typing (MLST) is a recently developed nucleotide sequence-based method for the definitive assignment of isolates within bacterial populations to specific clones. MLST uses the same principles as multilocus enzyme electrophoresis and provides data that can be used to investigate aspects of the population genetics and evolution of bacterial species. We used an MLST data set consisting of the sequences of approximately 450-bp fragments from seven housekeeping loci from a large strain collection of Neisseria meningitidis to estimate the relative impact of recombination compared with point mutation in the diversification of N. meningitidis clonal complexes. 126 meningococcal isolates were assigned to 10 clonal complexes, 9 of which contained minor clonal variants. The allelic variation within each complex was classified as a recombinational exchange or a putative point mutation through a comparison of the sequences of each variant allele with that of the allele typically found in the clonal complex. The nine clonal complexes contained a total of 23 allelic variants, and analysis of the sequences of these variant alleles revealed that a single nucleotide site in a meningococcal housekeeping gene is at least 80-fold more likely to change as a result of recombination than as a result of mutation. This value is estimated to be 10-50-fold for Escherichia coli and approximately 50-fold for Streptococcus pneumoniae.

Alleles↗
PubMed1998

Free recombination within Helicobacter pylori.

Sequences of three gene fragments (flaA, flaB, and vacA) from Helicobacter pylori strains isolated from patients in Germany, Canada, and South Africa were analyzed for diversity and for linkage equilibrium by using the Homoplasy Test and compatibility matrices. Horizontal genetic exchange in H. pylori is so frequent that different loci and polymorphisms within each locus are all at linkage equilibrium. These results indicate that H. pylori is panmictic. Comparisons with sequences from Escherichia coli, Neisseria meningitidis, and Drosophila melanogaster showed that recombination in H. pylori was much more frequent than in other species. In contrast, when multiple family members infected with H. pylori were investigated, some strains were indistinguishable at all three loci. Thus, H. pylori is clonal over short time periods after natural transmission.

Alleles↗
PubMed1998

Multilocus sequence typing: a portable approach to the identification of clones within populations of pathogenic microorganisms.

Traditional and molecular typing schemes for the characterization of pathogenic microorganisms are poorly portable because they index variation that is difficult to compare among laboratories. To overcome these problems, we propose multilocus sequence typing (MLST), which exploits the unambiguous nature and electronic portability of nucleotide sequence data for the characterization of microorganisms. To evaluate MLST, we determined the sequences of approximately 470-bp fragments from 11 housekeeping genes in a reference set of 107 isolates of Neisseria meningitidis from invasive disease and healthy carriers. For each locus, alleles were assigned arbitrary numbers and dendrograms were constructed from the pairwise differences in multilocus allelic profiles by cluster analysis. The strain associations obtained were consistent with clonal groupings previously determined by multilocus enzyme electrophoresis. A subset of six gene fragments was chosen that retained the resolution and congruence achieved by using all 11 loci. Most isolates from hyper-virulent lineages of serogroups A, B, and C meningococci were identical for all loci or differed from the majority type at only a single locus. MLST using six loci therefore reliably identified the major meningococcal lineages associated with invasive disease. MLST can be applied to almost all bacterial species and other haploid organisms, including those that are difficult to cultivate. The overwhelming advantage of MLST over other molecular typing methods is that sequence data are truly portable between laboratories, permitting one expanding global database per species to be placed on a World-Wide Web site, thus enabling exchange of molecular typing data for global epidemiology via the Internet.

Bacterial Typing Techniques↗
PubMed1998

Microevolution during epidemic spread of Neisseria meningitidis.

Similar to many other naturally transformable bacteria, Neisseria meningitidis has yielded many examples where horizontal genetic exchange has resulted in genetic variation of individual strains. Epidemic strains are purified of genetic variants due to bottlenecks during the spread from country to country, resulting in clonal descent. Occasionally, clonal replacement also occurs during epidemic spread. These processes occur rapidly in serogroup A meningococci; such that after their descent from a common ancestor, clonally related bacteria have diversified at numerous loci within the last decades.

Disease Outbreaks↗
PubMed1998

Meningitis caused by a serogroup W135 clone of the ET-37 complex of Neisseria meningitidis in West Africa.

Meningococci belonging to serogroup W135 caused several cases of meningococcal meningitis in The Gambia in 1995 and were isolated during a serogroup A epidemic in Mali in 1994. The eight isolates tested belonged to the same clone of the ET-37 complex and differed in several bands from the pulsed-field gel electrophoresis restriction pattern of serogroup C meningococci of the ET-37 complex isolated in Mali. Three of 6 patients infected in The Gambia died, indicating that this W135 clone is virulent. Vaccines that protect only against infections with meningococci belonging to serogroups A and C are usually used to control outbreaks in Africa, although vaccines containing the W135 polysaccharide are available. The findings of this study indicate that outbreaks of meningococcal meningitis in Africa can be associated with serogroup W135 infections and that serogrouping is essential before vaccination campaigns are started.

Adolescent↗
PubMed1998

Sequence diversity, predicted two-dimensional protein structure, and epitope mapping of neisserial Opa proteins.

The sequence diversity of 45 Opa outer membrane proteins from Neisseria meningitidis, Neisseria gonorrhoeae, Neisseria sicca, and Neisseria flava indicates that horizontal genetic exchange of opa alleles has been rare between these species. A two-dimensional structural model containing four surface-exposed loops was constructed based on rules derived from porin crystal structure and on conservation of sequence homology within transmembrane beta-strands. The minimal continuous epitopes recognized by 23 monoclonal antibodies were mapped to loops 2 and 3. Some of these epitopes are localized on the bacterial cell surface, in support of the model.

Amino Acid Sequence↗
PubMed1998

Randomly amplified polymorphic DNA genotyping of serogroup A meningococci yields results similar to those obtained by multilocus enzyme electrophoresis and reveals new genotypes.

Randomly amplified polymorphic DNA (RAPD) genotyping was applied to one representative strain of each of the 84 electrophoretic types (ETs) of Neisseria meningitidis serogroup A previously defined by multilocus enzyme electrophoresis (MEE) (J.-F. Wang et al., Infect. Immun. 60:5267-5282, 1992). Twenty-seven additional isolates comprising six ETs were also tested. MEE and RAPD genotyping yielded similar dendrograms at the subgroup level. Similar results were obtained by both methods for 18 serogroup A meningococci isolated in The Netherlands between 1989 and 1993. Ten of these isolates defined a new subgroup, designated subgroup IX. One isolate belonged to the ET-5 complex, normally associated with serogroup B strains (D. A. Caugant et al., Proc. Natl. Acad. Sci. USA 83:4927-4931, 1986). By RAPD genotyping, meningococci can be linked to previously characterized genotypes by using a computerized database, and dendrograms based on cluster analyses can easily be generated. RAPD analysis offers advantages over MEE since intermediate numbers of isolates of serogroup A meningococci can quickly be assigned to known subgroups and new subgroups can be defined.

Bacterial Typing Techniques↗
PubMed1998

Necessity of molecular techniques to distinguish between Neisseria meningitidis strains isolated from patients with meningococcal disease and from their healthy contacts.

Serogroup C strains of Neisseria meningitidis were isolated from a Germany patient with severe meningococcal disease after a trip to the Czech Republic. These strains (case isolates) were characterized by classical and molecular techniques, as were other strains (carrier isolates) isolated from healthy contacts. Five of 10 carrier isolates had switched off the expression of capsular polysaccharide, as demonstrated by a serogroup-specific PCR. The two case isolates were indistinguishable by multilocus sequence typing and belonged to the ET-37 complex. The carrier isolates belonged to four different sequence types, all unrelated to that of the case strains. Pulsed-field gel electrophoresis showed that the case isolates differed from reference ET-37 complex strains from the Czech Republic and Canada as well as from all the carrier isolates. The isolate from the patient's nasopharynx was indistinguishable from the blood isolate except for a 40,000-bp chromosomal deletion that had occurred during systemic spread.

Adolescent↗
PubMed1997

Microevolution and epidemic spread of serogroup A Neisseria meningitidis--a review.

An extensive and representative strain collection of serogroup A Neisseria meningitidis was established. These bacteria were obtained from different endemic and epidemic/pandemic sources and include strains from diseased patients and healthy carriers. The genetic relationships of the bacteria were defined by multi-locus enzyme electrophoresis and sequence polymorphisms of genetically variable antigens have been analyzed in closely-related groupings. The results are interpreted as reflecting a balance of recombination events, which disrupt clonal relationships, and sequential bottlenecks, which purify the bacterial population of genetic variants during epidemic spread.

Alleles↗
PubMed1997

Persistence of antibodies to meningococcal IgA1 protease versus decay of antibodies to group A polysaccharide and Opc protein.

Sera were taken over a 5 year period from Gambian children vaccinated in 1983, when aged 1-4 years, with A + C meningococcal capsular polysaccharide, ELISA tests were devised to determine the concentrations of immunoglobulin A, G and M reacting with A polysaccharide and of IgG reacting with Opc protein, IgA1 protease and an internal 104 mer peptide derived from IgA1 protease. Vaccination resulted in a brief rise of antibodies to A polysaccharide followed by decline to pre-immunization levels. IgM levels were very high even before vaccination. Antibodies to Opc protein stimulated by natural exposure also declined over the 5 year period. In contrast, antibodies stimulated by natural exposure to IgA1 protease or to the internal peptide remained constant or increased (final geometric mean level of 47 micrograms IgG ml-1). We speculate that healthy carriage of Neisseria meningitidis or Haemophilus influenzae is responsible for this increase in IgG concentration.

Amino Acid Sequence↗
PubMed1997

Two-dimensional structure of the Opc invasin from Neisseria meningitidis.

A two-dimensional structural model was devised for the Opc outer membrane protein invasin which contains 10 transmembrane strands and five surface-exposed loops. One continuous epitope recognized by three monoclonal antibodies was localized to the tip of loop 2 by synthetic peptides and site-directed mutagenesis while a second, discontinuous epitope recognized by a fourth antibody was localized to loops 4 and 5 by insertion mutagenesis. These monoclonal antibodies are bactericidal and inhibit adhesion and invasion. Most of the T-cell epitopes defined by Wiertz et al. (1996) were localized to the transmembrane strands. Oligonucleotides encoding a foreign epitope (nabla) from Semliki Forest virus were inserted into BglII restriction sites created by site-directed mutagenesis. The nabla epitopes inserted in all five predicted loops were recognized on the cell surface of live Escherichia coli bacteria by a monoclonal antibody and are exposed while nabla epitopes in the N-terminus or three predicted turns were not. The results thus confirm important predictions of the model and define five permissive sites within surface-exposed loops which can be used to insert foreign epitopes.

Adhesins, Bacterial↗
PubMed1997

Clonal descent and microevolution of Neisseria meningitidis during 30 years of epidemic spread.

Serogroup A meningococci of subgroups III, IV-1 and IV-2 are probably descended from a common ancestor that existed in the nineteenth century. The 10.5kb sequences spanning five distinct chromosomal loci, encoding cell-surface antigens, a secreted protease or housekeeping genes and intergenic regions, were almost identical in strains of those subgroups isolated in 1966, 1966 and 1917 respectively. During the subsequent two to three decades, all of these loci varied as a result of mutation, translocation or import of DNA from unrelated neisseriae. Thus, microevolution occurs frequently in naturally transformable bacteria. Many variants were isolated only once or within a single geographical location and disappeared thereafter. Other variants achieved genetic fixation within months or a few years. The speed with which sequence variation is either eliminated or fixed may reflect sequential bottlenecks associated with epidemic spread and contrasts with the results of phylogenetic analyses from bacteria that do not cause epidemics.

Alleles↗
PubMed1996

Allelic polymorphism and site-specific recombination in the opc locus of Neisseria meningitidis.

The opc gene is widespread in epidemic and endemic Neisseria meningitidis, but most strains of certain epidemic clones (ET-37 complex, Cluster A4) and a few random endemic isolates lack an opc gene. Four percent of the 1148 bp that contain opc plus the surrounding intergenic region was polymorphic (18 alleles), and many of the alleles contained a 230 bp insertion at a fixed location in the intergenic region. The presence or absence of the insertion reflects site-specific recombination. The alleles are stably inherited within clonal groupings for up to at least 50 years, with rare cases of horizontal genetic exchange. Most statistical methods indicated significant intragenic recombination events within this dataset.

Alleles↗
PubMed1996

T-cell responses to outer membrane proteins of Neisseria meningitidis: comparative study of the Opa, Opc, and PorA proteins.

Former studies have shown that the class 5 outer membranes proteins (Opa and Opc proteins) of Neisseria meningitidis are at least as immunogenic as meningococcal porin proteins. High antibody titers to class 5 proteins have been observed in sera obtained during convalescence after meningococcal infection. A strong increase in anti-class 5 antibodies has also been observed in vaccinees who received a meningococcal outer membrane vesicle preparation. The enhanced B-cell response to class 5 proteins may be due to the presence of immunodominant helper T-cell epitopes in these proteins. In order to investigate this hypothesis, we tested purified Opa, Opc, and class 1 proteins for recognition by human T cells. a hierarchy of T-cell immunogenicity was observed among the outer membrane proteins, the Opa protein being more immunogenic than the other proteins. In most cases, the proliferative responses elicited by Opc were higher than the responses observed for the class 1 protein. The epitopes recognized by the immune T cells were identified by using overlapping synthetic peptides spanning the protein sequences of OpaB, Opa5d, and Opc.

Adult↗
PubMed1996

The porA alleles are identical in subgroup III serogroup A Neisseria meningitidis strains isolated in China in the 1960s and 1980s.

The porA genes from serogroup A, subgroup III strains isolated in the People's Republic of China in 1966 and in 1984 and 1985 were amplified by PCR at an annealing temperature of 75 degrees C. The DNA sequences (5 strains) and the restriction patterns generated by MspI (14 strains) were identical, unlike the results reported by Peixuan et al. (Z. Peixuan, H. Xujing, and X. Li, J. Clin. Microbiol. 33:458-462, 1995). Furthermore, PCR products which were amplified at an annealing temperature of 60 degrees C, as described by Peixuan et al., were heterogeneous in our study.

Alleles↗
PubMed1996

[Molecular epidemiology of meningococcal meningitis in Mali: isolation of a new class 1 protein variant (P1.y)].

The study deals with 570 strains of Neisseriaceae isolated between 1989 and 1994 in Mali: 396 of the strains were isolated from samples of cerebrospinal fluid and 174 from the throat. Serogroup C accounted for 55% of all strains. Antigenic structure was determined by ELISA, SDS-PAGE and transfer to nitrocellulose membrane for immunoblotting with monoclonal antibodies produced at the Max Planck Institute for Molecular Genetics. For serogroup A, the class 1 protein types found were P1.7 for strains isolated prior to 1994 and P1.9 for strains isolated in 1994. P1.7 is specific to clone IV-1 and P1.9 to clone III-1, which was responsible for the 1994 epidemic. All strains of serogroup C isolated from fluid CSF and most strains isolated from the throat exhibit a new type of class 1 protein which the authors have designated P1.y. P1.y is characteristic of Malian strains of serogroup C; it is rare or absent in strains from other countries (Burkina Faso, Ghana, Italy, USA). The nucleotide sequence of the gene expressing P1.y and the corresponding amino acid sequence were determined at the National Institute for Biological Standards and Control, England.

Bacterial Proteins↗