Analysis of the genetic differences between Neisseria meningitidis and Neisseria gonorrhoeae: two closely related bacteria expressing two different pathogenicities.

Abstract

We have investigated genetic differences between the closely related pathogenic Neisseria species, Neisseria meningitidis and Neisseria gonorrhoeae, as a novel approach to the elucidation of the genetic basis for their different pathogenicities. N. meningitidis is a major cause of cerebrospinal meningitis, whereas N. gonorrhoeae is the agent of gonorrhoea. The technique of representational difference analysis was adapted to the search for genes present in the meningococcus but absent from the gonococcus. The libraries achieved are comprehensive and specific in that they contain sequences corresponding to the presently identified meningococcus-specific genes (capsule, frp, rotamase, and opc) but lack genes more or less homologous between the two species, e.g., ppk and pilCI. Of 35 randomly chosen clones specific to N. meningitidis, DNA sequence analysis has confirmed that the large majority have no homology with published neisserial sequences. Mapping of the cloned DNA fragments onto the chromosome of N. meningitidis strain Z2491 has revealed a nonrandom distribution of meningococcus-specific sequences. Most of the genetic differences between the meningococcus and gonococcus appear to be clustered in three distinct regions, one of which (region 1) contains the capsulerelated genes. Region 3 was found only in strains of serogroup A, whereas region 2 is present in a variety of meningococci belonging to different serogroups. At a time when bacterial genomes are being sequenced, we believe that this technique is a powerful tool for a rapid and directed analysis of the genetic basis of inter-or intraspecific phenotypic variations.The study of bacterial pathogenicity has greatly benefited from tools such as transposon mutagenesis, which have made possible the identification of virulence genes. However, there exist many bacteria for which mutagenesis is inefficient. One way to study virulence in such organisms is to take advantage of naturally occurring differences in pathogenicity between variants of the same species or between closely related species. In such a case, the differences in DNA sequence, including genes responsible for the differential pathogenicities, may be isolated from the generally similar genetic background by a subtractive technique. We set out to define genes or loci that may be responsible for the pathogenesis of meningococcal meningitis by comparing the chromosome of the meningococcus [Neisseria meningitidis (Nm)] with that of the gonococcus [Neisseria gonorrhoeae (Ng)]. These two human pathogens are very closely related, but cause notably different diseases. While Ng is generally responsible for localized inflammation of the urogenital tract, the meningococcus is the cause of lifethreatening disease, meningitis, that follows penetration of the blood-brain barrier and colonization of the meningeal membranes by the bacteria.In contrast to the great differences in pathogenic potential of the gonococcus and the meningococcus, the organisms are closely related at the level of genetic organiza...

Subtractive hybridization reveals a high genetic diversity in the fish pathogen Photobacterium damselae subsp. piscicida: evidence of a SXT-like element

“…specific for highly virulent isolates. Some of these differentially occurring genes may determine strain-specific characteristics such as virulence factors (Tinsley & Nassif, 1996;Groisman & Ochman, 1997;Reckseidler et al, 2001), which can be used as a valuable tool to establish the nature and severity of disease.…”

“…Representational difference analysis (RDA) (Lisitsyn, 1995;Tinsley & Nassif, 1996), which is based on suppression subtractive hybridization (SSH) (Akopyants et al, 1998;Diatchenko et al, 1996;Gurskaya et al, 1996), is a powerful technique for analysing the differences between two complex genomes, and for identifying DNA sequences that are present in one strain (the tester), but absent in another strain (the driver). This technique has been used in different bacterial species to identify genomic differences between virulent and avirulent strains (Calia et al, 1998;Mahairas et al, 1996;Zhang et al, 2000).…”

Aspectos fenotípicos, genotípicos e de diagnóstico da bactéria A. pleuropneumoniae

“…Por esta metodologia, os fragmentos de DNA presentes especificamente em um dos genomas podem ser isolados e caracterizados (CLAUS et al, 2000). Esta metodologia vem sendo utilizada com sucesso para caracterização de muitos patógenos bacterianos, como Escherichia coli e Vibrio cholerae (TINSLEY & NASSIF, 1998;CLAUS et al, 2000). Portanto, a técnica de RDA poderia ser aplicada na determinação de diferenças existentes entre isolados do sorotipo 3 obtidos no Brasil com os de outros países, respondendo algumas dúvidas a respeito dos mecanismos de patogenicidade desta bactéria para os suínos.…”

Validation of the suppressive subtractive hybridization method in Mycoplasma agalactiae species by the comparison of a field strain with the type strain PG2

“…The oligonucleotides RJ48 and RN48 were designed by the concatenation of the R24 sequence with the J24 or the N24 sequence, respectively. The rationale of this was to give us the possibility to switch from SSH to RDA, if desired, just by using two extra 12-mer oligonucleotides, Reco12 and Rbam12, which have been described elsewhere [26], to obtain the set of adapters for RDA.…”

“…Oligonucleotide sequences are given in Table II. The oligonucleotides R24, J24, N24, J12 and N12 were used for the comparison of Neisseria meningitidis and Neisseria gonorrhoeae by a subtractive hybridization technique, the Representational Difference Analysis (RDA) [26]; both the RDA and SSH methods are based on PCR enrichment of the target (tester-specific) sequences, but differ in their protocols. The oligonucleotides RJ48 and RN48 were designed by the concatenation of the R24 sequence with the J24 or the N24 sequence, respectively.…”