Use of Variable-Number Tandem Repeats To Examine Genetic Diversity of Neisseria meningitidis

Abstract

Repetitive DNA motifs with potential variable-number tandem repeats (VNTR) were identified in the genome of Neisseria meningitidis and used to develop a typing method. A total of 146 meningococcal isolates recovered from carriers and patients were studied. These included 82 of the 107 N. meningitidis isolates previously used in the development of multilocus sequence typing (MLST), 45 isolates recovered from different counties in Norway in connection with local outbreaks, and 19 serogroup W135 isolates of sequence type 11 (ST-11), which were recovered in several parts of the world. The latter group comprised isolates related to the Hajj outbreak of 2000 and isolates recovered from outbreaks in Burkina Faso in 2001 and 2002. All isolates had been characterized previously by MLST or multilocus enzyme electrophoresis (MLEE). VNTR analysis showed that meningococcal isolates with similar MLST or MLEE types recovered from epidemiologically linked cases in a defined geographical area often presented similar VNTR patterns while isolates of the same MLST or MLEE types without an obvious epidemiological link showed variable VNTR patterns. Thus, VNTR analysis may be used for fine typing of meningococcal isolates after MLST or MLEE typing. The method might be especially valuable for differentiating among ST-11 strains, as shown by the VNTR analyses of serogroup W135 ST-11 meningococcal isolates recovered since the mid-1990s.Neisseria meningitidis is a major cause of meningitis and septicemia worldwide (8) and still remains one of the leading infectious causes of death in childhood in many industrialized countries. However, the greatest burden of the disease occurs in Africa and Asia, where large epidemics periodically affect the populations.Epidemiological studies of N. meningitidis have allowed a better understanding of the nature of meningococcal dissemination and of the relation between endemic and epidemic disease. During the last 2 decades, the classical phenotyping methods for epidemiological studies of meningococci have been complemented by a number of molecular methods, such as multilocus enzyme electrophoresis (MLEE), pulsed-field gel electrophoresis, randomly amplified polymorphic DNA, repetitive element-based PCR, insertion sequence analysis, ribotyping, restriction fragment length polymorphism analysis of PCR products, sequencing of individual genes that may be related to virulence (8), PorA VR typing (28), 16S rRNA sequencing (29), and multilocus sequence typing (MLST) (21). PorA VR typing, which is used to predict the amino acid sequences of the PorA outer membrane protein, is derived from DNA sequence analysis (28). A website for comparison of porA sequences from different laboratories (http://neisseria.org/nm/typing/pora) is now in widespread use by the research community and ensures consistency in identifying and naming variants of this protein.The gene encoding 16S rRNA has been used successfully to monitor and identify epidemiologically related isolates of meningococci (29), especially those associated wit...

Variable-Number Tandem Repeat Analysis of Meningococcal Isolates Belonging to the Sequence Type 162 Complex

“…In these complexes, the same patterns were obtained for epidemiologically related isolates from a relatively confined area and during a limited period of time (15). Therefore, the VNTR diversity demonstrated in the ST-162 complex in Greece suggests that this clonal complex has circulated for some time in the country and that the diseaseassociated isolates do not represent an outbreak situation.…”

“…We have previously demonstrated that VNTR typing is capable of distinguishing between meningococcal isolates belonging to the ST-32 complex and those belonging to the ST-11 complex (15). In these complexes, the same patterns were obtained for epidemiologically related isolates from a relatively confined area and during a limited period of time (15).…”

“…With the aim of establishing a variable-number tandem repeat (VNTR) method for molecular typing of N. meningitidis, we previously analyzed a total of 146 meningococci isolated from carriage, endemic, and epidemic cases (15). We concluded that, like pulsed-field gel electrophoresis (PFGE), VNTR analysis has high discriminatory power and may differentiate isolates identified as similar by multilocus enzyme electrophoresis or MLST (2,6,12).…”

“…(15). In brief, four VNTR areas in the genome of N. meningitidis were analyzed for genetic polymorphism by using the primers listed in Table 1.…”

Comparison of Multiple-Locus Variable-Number Tandem Repeat Analysis, Pulsed-Field Gel Electrophoresis, and Phage Typing for Subtype Analysis of Salmonella enterica Serotype Enteritidis

“…MLVA has been used to subtype a variety of species of bacteria, some of which have been difficult to subtype by other methods. Bacteria that have been typed by MLVA include Salmonella serotypes Typhimurium (22,24,35) and Typhi (26), Yersinia pestis (19), Francisella tularensis (9,17), Mycobacterium tuberculosis (1), Haemophilus influenzae (41), Bacillus anthracis (18), Bordetella pertussis (40), Escherichia coli O157:H7 (23,31), Enterococcus faecalis (47), Neisseria meningitidis (51), and Pseudomonas aeruginosa (33). MLVA has proven to be a rapid method and may be easy to standardize between laboratories.…”