Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving nucleic acid
Reexamination Certificate
1997-09-23
2002-06-11
Graser, Jennifer E. (Department: 1645)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving nucleic acid
C536S023700, C536S024300, C536S024320, C435S320100, C435S975000
Reexamination Certificate
active
06403306
ABSTRACT:
FIELD OF THE INVENTION
This invention relates generally to the nucleotide sequences of serogroup-specific capsular polysaccharides genes and their use in a method for typing of serogroups of pathogenic bacteria, in particular
Neisseria meningitidis,
and further, relates to capsule gene switching in recombinant strains and the detection thereof.
BACKGROUND OF THE INVENTION
Contagious outbreaks of epidemic diseases constitute public health emergencies requiring rapid treatment and chemoprophylaxis of contacts. Vaccination of the population at risk can be considered if disease cases continue to occur. However, asymptomatic carriage of pathogens in humans is common and some of the adult population may be immunized from previous outbreaks. The factors leading from acquisition of the organism to invasive disease point to a clonal origin of the outbreaks and to an enhanced virulence or altered antigenicity of a particular clone.
Neisseria meningitidis
is a leading worldwide cause of
meningitis
and rapidly fatal sepsis in otherwise health individuals [Apicella, M. A. (1995) in
Principles and Practice of Infectious Diseases,
eds. Mandell, G. L., Douglas, R. G., and Bennett, J. E., Churchill Livingstone, New York, pp. 1896-1909]. In excess of 350,000 cases of meningococcal disease were estimated to have occurred in 1995 [WHO Report (1996) WHO, Geneva, ISBN 92 4 1561823]. The problem of meningococcal disease is emphasized by the recurrence of major epidemics due to serogroups A, B, and C
N. meningitidis
over the last 20 years, such as: the devastating serogroup A outbreak in sub-Saharan Africa in 1996 [WHO (1996)
Meningitis in Africa. The constant challenge of epidemics.
WHO 21:15 March]; the recent dramatic increases in the incidence of serogroup B and C meningococcal disease in parts of North America [CDC (1995)
MMWR
44:121-134; Jackson, L. A. et al. (1995)
JAMA
273:390-394; Wahlen, C. M. et al. (1995)
JAMA
273:383-389]; and the emergence in Europe and elsewhere of meningococci with decreased susceptibility to antibiotics [Campos, J. et al. (1992)
J. Infect. Dis.
166:173-177].
Differences in capsular polysaccharide chemical structure determine the meningococcal serogroups [Liu, T. Y. et al. (1971)
J. Biol. Chem.
246:2849-58; Liu, T. Y. et al. (1971)
J. Biol. Chem.
246:4703-12]. Meningococci of serogroups B, C, Y, and W-135 express capsules composed entirely of polysialic acid or sialic acid linked to glucose or galactose [Liu, T. Y. et al. (1971)
J. Biol. Chem.
246:4703-12; Bhattacharjee, A. K. et al. (1976)
Can. J. Biochem.
54:1-8], while the capsule of group A
N. meningitidis
is composed of N-acetyl mannosamine-1-phosphate [Liu, T. Y. et al. (1971)
J. Biol. Chem.
246:2849-58]. The currently available capsular polysaccharide vaccines for serogroups A, C, Y, or W-135
N. meningitidis
are effective for control of meningococcal outbreaks in older children and adults. However, because of poor immunogenicity in young children and short-lived immunity [Zollinger, W. D. and Moran, E. (1991)
Trans. R. Soc. Trop. Med. Hyg.
85:37-43], these vaccines are not routinely used for long-term prevention of meningococcal disease. In the case of group B
N. meningitidis,
whose (&agr;2→8)-linked polysialic capsule is an immunotolerized self antigen, a reliable polysaccharide vaccine is not yet available. However, rapid progress is being made in development of polysaccharide-protein conjugate vaccines and it is hoped that following the example of newly licensed
Haemophilus influenzae
type b vaccines, widespread introduction of the polysaccharide conjugates will lead to elimination of disease.
In some epidemic settings, simultaneous or closely-linked meningococcal outbreaks have occurred in the same population due to different serogroups [Sacchi, C. T. et al. (1994)
J. Clin. Microbiol.
32:1783-1787; CDC (1995)
MMWR
44:121-134; Krizova, P. and Musilek, M. (1994)
Centr. Eur. J. Publ. Hlth
3:189-194]. Further, Caugant et al. (Caugant, D. A. et al. (1986)
Proc. Natl. Acad. Sci. USA
83:4927-4931; Caugant, D. A. et al. (1987)
J. Bacteriol.
169:2781-2792] and others have noted that meningococcal isolates of different serogroups may be members of the same enzyme type (ET)-5, ET-37 or ET-4 clonal complexes.
Since 1993, the number of cases of serogroup B meningococcal disease in Oregon and adjacent counties in Washington State has doubled, and the overall incidence has been five-fold higher than rates observed in other parts of the United States [CDC (1995)
MMWR
44:121-134]. This increase was due to the first appearance in the U.S. of serogroup B meningococcal strains closely related to the ET-5 complex. ET-5 complex strains have been responsible for major epidemics in Norway, Iceland, Cuba and South America over the last twenty years (Caugant, D. A. et al. (1986)
Proc. Natl. Acad. Sci. USA
83:4927-4931; Sierra, G. V. et al. (1991)
NIPH Annals
14:195-207; Sacchi, C. T. et al. (1992)
J. Clin. Microbiol.
30:1734-1738]. Since 1994, cases of serogroup C meningococcal disease due to ET-5 complex strains were also noted in Oregon and Washington State. There exists a recurring need to understand the genetic basis for meningococcal capsule expression and to analyze the serogroup B and C ET-5 meningococcal strains responsible for the outbreak in the Pacific Northwest.
SUMMARY OF THE INVENTION
It is an object of the present invention to provide strains of
N. meningitidis
of a particular serogroup transformed in vitro to express a capsule polysaccharide marker of a different meningococcal strain serogroup. In a particular embodiment are provided prototype serogroup C, Y and W-135 meningococcal strains transformed in vitro with DNA comprising the synD of the serogroup B strain NMB. According to the present invention, conversion from one sialic acid expressing capsule serogroup to another can be accomplished by homologous recombination of the sequences encoding the serogroup-specific capsule polymerase. Such recombinant
N. meningitidis
strains are provided according to the invention as genetically engineered in vitro recombinations.
Also provided by the present invention are
Neisseria meningitidis
mutant serogroup strains which express different non-isogeneic capsular polysaccharides due to homologous recombination of the sequences encoding the serogroup-specific capsule polymerase. Specifically exemplified herein is a mutant
N. meningitidis
strain 1070 (serogroup B, ET-301) in which genetic markers are isogeneic to serogroup B except for the capsule polysaccharide, which is a serogroup C marker. Such meningococcal isolates comprise a recombinant or switched capsule gene and, in a particular embodiment, a switching or recombination event occurred from a serogroup B to a serogroup C capsule biosynthetic gene. Such recombinant
N. meningitidis
strains are provided according to the invention as naturally-occurring in vivo recombinant isolates.
It is also an object of the invention to provide meningococcal serogroup-specific capsule genes encoding characteristic capsular polysaccharide virulence determinants. In specific embodiments of the invention are provided capsule biosynthetic gene preparations of prototype serogroups A, B, C, Y and W-135, each serogroup-specific gene encoding a biosynthetic enzyme for a specific and distinguishing capsular polysaccharide.
It is an additional object of the invention to provide cloned DNA molecules which can be used to introduce an additional non-isogeneic capsular polysaccharide virulence determinant into strains of
N. meningitidis.
In a particular embodiment, the cloned DNA fragment containing the stable Tn916 insertion in the synD of the serogroup B
N. meningitidis
strain NMB was used to introduce the gene for the serogroup B (&agr;2→8)-linked capsule polysialyltransferase into other meningococcal strains to produce novel immunotypes. More generally, a cloned DNA fragment containing a stable inse
Stephens David S.
Swartley John S.
Emory University
Graser Jennifer E.
Greenlee Winner and Sullivan PC
LandOfFree
Serogroup-specific nucleotide sequences in the molecular... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Serogroup-specific nucleotide sequences in the molecular..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Serogroup-specific nucleotide sequences in the molecular... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2913816