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(The FASEB Journal. 1999;13:331-337.)
© 1999 FASEB

RESEARCH COMMUNICATION

Invasive isolates of Neisseria meningitidis possess enhanced immunoglobulin A1 protease activity compared to colonizing strains

SRDJAN VITOVSKI, ROBERT C. READ and JON R. SAYERS 1

Division of Molecular and Genetic Medicine, The University of Sheffield, Royal Hallamshire Hospital, Sheffield, S10 2JF, United Kingdom


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae possess the ability to cleave human IgA1 antibodies, and all successfully colonize and occasionally invade the human upper respiratory tract. N. meningitidis invades the bloodstream after a period of nasopharyngeal colonization. We directly compared levels of IgA1 protease activity in strains (n=52) derived from the cerebrospinal fluid or blood of patients with meningococcal disease with strains of N. meningitidis obtained from asymptomatic carriers (n=25). IgA1 protease activity was determined by a sensitive semiquantitative ELISA assay. Levels of IgA1 protease activity were significantly higher (P<0.0001) in strains associated with invasive meningococcal disease (98% with detectable activity, mean = 580 mU) than with those obtained from asymptomatic carriers (76% with detectable activity, mean = 280 mU). Despite marked variation in enzyme activity, almost all strains (96%) possessed the gene for IgA1 protease. Given the panmictic population structure of the bacterial isolates investigated, these data, obtained from two groups infected with N. meningitidis, but with markedly different clinical outcomes, provide the first quantitative evidence that IgA1 protease activity is a virulence determinant that contributes to the pathogenic phenotype, and suggest IgA1 protease as a potential target for prophylaxis.—Vitovski, S., Read, R. C., Sayers, J. R. Invasive isolates of Neisseria meningitidis possess enhanced immunoglobulin A1 protease activity compared to colonizing strains.


Key Words: human • bacterial infections • virulence • meningitis • carrier state


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
IMMUNOGLOBULIN A1 (IgA1)2 is the most abundant antibody associated with human mucosal surfaces, including the upper respiratory tract (1 , 2 ). IgA1 contributes to immune exclusion by binding the surface of colonizing bacteria, resulting in agglutination, mucin adherence, and steric hindrance of adhesin–epithelial receptor interactions (3 , 4 ), all of which contribute to mucociliary clearance.

Bacterial IgA1 proteases (IgA-specific postproline endopeptidases) are extracellular enzymes able to cleave IgA1 within the hinge region, thus separating the antigen binding fragment (Fab) and constant regions (Fc) (5 , 6 ). The consequences of this cleavage are twofold. First, agglutination and mechanical clearance are impeded; second, the released Fab fragment is still able to bind cognate antigen and is thus capable of masking epitopes from subsequent recognition by other intact immunoglobulins (7) .

Bacteria that produce IgA1 proteases include the three most common causative agents of bacterial meningitis: Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae 8-10) . These organisms comprise some of the flora present in the normal human nasopharynx. Colonization of the upper respiratory tract may lead to asymptomatic carriage or subsequent invasion of the host, causing serious disease (11) . Since IgA1 is the predominant isotype of the upper respiratory tract (12) , it has been suggested that possession of IgA1 protease is a virulence determinant of disease in which colonization of this site is a component. Other mucosal pathogens such as N. gonorrhoeae, Streptococcus sanguis, and Prevotella spp. also produce potent IgA1 proteases 8-10) .

Several other lines of indirect evidence for a role in pathogenesis have been reported: nonpathogenic species of Neisseria and Haemophilus lack IgA1 protease activity (13 , 14 ); the products of IgA1 cleavage have been found in the cerebrospinal fluid of patients with bacterial meningitis, in vaginal washings from gonorrhea cases, and in other secretions from individuals infected with known IgA1 protease-producing bacteria (15 , 16 ). However, more direct evidence for the role of bacterial IgA1 proteases in microbial pathogenesis is lacking. Animal model experiments are precluded due to specificity of the enzyme for human IgA1.

In the 25 years since their discovery (17) , most studies of bacterial IgA1 proteases have concentrated on their distribution among different micro-organisms (18) or on biochemical and genetic characteristics 19-22) . To date, there have been no reported attempts to quantify the IgA1 protease activity of individual strains of N. meningitidis apart from very recently reported work describing the activities of four strains (23) . Specifically, the relationship between IgA1 protease activity in N. meningitidis isolated from carriers and clinical cases has not been reported.

The aim of this work was to compare IgA1 protease activity in a number of heterogeneous (i.e., serologically diverse) isolates of N. meningitidis obtained from clinical cases of meningococcal disease with those cultured from the throats of asymptomatic carriers.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Bacterial strains
Invasive strains (52 isolates) from cerebrospinal fluid (CSF) or blood cultures were obtained from patients of all ages with meningococcal disease admitted to hospitals in North Trent over the 2 year period of 1995–1996. Carriage strains (25 isolates) were obtained from throat swabs of asymptomatic patients of all ages attending general pediatric or medical units of the Central Sheffield University Hospitals from North Trent communities over the 6 month period from July to December of 1996. All isolates resembling meningococci by oxidase, Gram reaction and microscopic morphology were stored in glycerol broth at -70°C until further investigation. Confirmatory identification, grouping, typing, and subtyping with monoclonal antibodies were conducted at the Meningococcal Reference Laboratory, Manchester, U.K. Three strains—NMB, K454, and MC58 24-26) —have been subjects of previous studies. The characteristics of the strains used in this study are presented in Table 1 and Table 2 . All isolates were grown on chocolate agar plates at 37°C under 5% CO2. Broth cultures were grown in 10 ml of brain–heart infusion medium containing 10% horse serum.


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  		Table 1. Characteristics of invasive N. meningitidisisolatesa


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  		Table 2. Characteristics of N. meningitidis strains obtained from asymptomatic carriersa

DNA isolation and PCR amplification
To identify the IgA1 mature protease domain in each strain studied, the oligonucleotides (forward, 5'-CGTGACTTTGCAGAAAACAA-3' and reverse, 5'-AAATAACGGAGAGCCGCTATCGCC-3') were used as primers for polymerase chain reaction (PCR) amplification, and were derived from the published sequence of the N. meningitidis HF13 (accession number X82474) IgA1 protease gene (27) . The primers produce a product corresponding to positions 553-1261 bp of the published sequence. Crude genomic DNA was prepared by transferring a loopful of bacteria from a single plate, resuspension in 0.5 ml of 10 mM Tris-HCl, pH 8 buffer, and boiling for 10 min. Amplification was performed with 1 unit of AmpliTaqDNA polymerase (Perkin Elmer, Norwalk, Conn.) and buffer supplied by the manufacturer supplemented with 200 µM of each dNTP, 200 ng of each primer and 5 µl of crude genomic DNA in total volume of 50 µl. Thirty cycles of amplification consisting of 30 s denaturation at 94°C, 90 s annealing at 55°C, and 120 s extension at 74°C were performed in a programmable DNA thermal cycler (Perkin-Elmer Cetus Corp). Negative controls were performed with PCR mixtures lacking either bacterial DNA or primers. PCR products were analyzed by electrophoresis of 10 µl of the amplification mixture on a 1% agarose gel and detected by staining with ethidium bromide. The appearance of the 724 bp band was scored as a positive amplification. Amplification of Neisseria 16S ribosomal RNA genes was performed as a control for DNA concentration and quality for each strain prior to IgA1 protease gene detection by using previously published primer sequences (28) . The sequences of two representative PCR products were determined using the dye termination method with an ABI Taq FS sequencing kit (Applied Biosystems, Cheshire, U.K.) analyzed on an ABI 373 A Stretch automated sequencing machine (Applied Biosystems). DNA sequence analysis was performed using Mac Vector and AssemblyLIGN (International Biotechnologies, Cambridge, U.K.) software packages.

IgA1 protease assay
IgA1 protease activities were measured using a modification of an enzyme-linked immunoassay (ELISA) that has already been described (29) . A loopful of bacteria from chocolate agar plates was inoculated into 10 ml of liquid medium. Bacterial cultures were grown until late log phase and absorbance at 550 nm of each culture was measured. A new 10 ml culture was started using 0.1–0.5 ml (depending on the growth rate) of the primary culture as an inoculum. This culture was allowed to reach mid-log phase (between 0.4 and 0.5 A550). At that point, 4 ml of each bacterial culture was filtered through a 0.22 µm disposable filter unit (Gelman) and the filtrate was used immediately in the ELISA as follows: IgA1 protease substrate (purified IgA1, 3.2 µg/ml (Calbiochem, San Diego, Calif.), or purified human colostrum IgA, 3.2 µg/ml (Sigma, St. Louis), were bound to the surface of a polystyrene microtitration plate (Immulon 2, Dynex) through their Fab using rabbit anti-human {lambda} light chain antibody (Dako, Glostrup, Denmark). Immobilized IgA substrates were exposed to filtered culture supernatants (150 µl) for 1 h. Incubation of such bound substrates with bacterial culture supernatants resulted in release and loss of the Fc{alpha} region upon rinsing, whereas the Fab fragment is retained after washing. Loss of Fc{alpha} was detected indirectly through a reduced binding of peroxidase-conjugated rabbit anti-human Fc{alpha} antibody (Dako) as assayed with the chromogenic substrate o-phenylenediamine dihydrochloride (Sigma). The difference in signal intensity between the undigested substrate molecules in control wells and the wells containing bacterial supernatant was recorded as {Delta} A490nm value for each bacterial strain. Relative activity was calculated by dividing the {Delta} A490nm value by the absorbance of the same culture (measured at 550 nm). One unit of enzyme activity was defined as the amount of enzyme able to effect a change in optical density of one absorbance unit (at 490 nm) in 1 h at 37°C. Each bacterial strain was assayed in triplicate for both IgA1 and S-IgA.

Statistical analysis
Statistical analysis was performed by an unpaired Student's t test of the differences in the mean activities displayed by each group of bacteria on each substrate assayed. Strains with no detectable IgA1 protease activity were included in all statistical calculations. The null hypothesis assumed there was no difference in the activity levels produced by the two groups of bacteria (invasive and carriage).


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Serotyping of Neisseria meningitidis strains
Strain nomenclature, group, type and subtypes, IgA1 protease activities (serum IgA1 and secretory IgA), and presence of the IgA1 protease gene for both invasive (Table 1) and colonizing strains (Table 2) are summarized. The clinical isolates, though diverse, were predominantly serogroups B (54%) and C (39%). As expected, the carriage strains were more heterogeneous, comprising mostly nontypeable isolates (56%).

Detection of the iga gene by PCR
The diagnostic PCR test confirmed the presence of an IgA1 protease in 98% of the clinical and 92% of carriage strains used in this study. Failure to detect a PCR product could be due to the absence of a suitable complimentary nucleotide sequence (caused by loss of the gene or a mutation abolishing the priming reaction) or, alternatively, to inadequate preparation of bacterial DNA samples. We excluded the latter by demonstrating that all isolates gave a positive PCR signal with primers designed to amplify a region of the highly conserved 16S Neisseria ribosomal RNA genes (data not shown).

Sequence analysis
The sequences of two representative PCR products obtained from strains MC58 and SVG69 were determined and found to be identical to the published sequences of N. meningitidis iga genes (data not shown; accession number AF012203).

IgA1 protease activity
A summary of the IgA1 protease activity assays is presented graphically in Fig. 1 . The levels of IgA1 protease activity associated with individual strains are also presented (Tables 1 and 2) . Examination of these data revealed a wide variation in detectable levels of enzymatic activity, e.g., strain SVG21 produced up to 24-fold more activity than SVG27. All but one of the invasive isolates (SVG44) displayed detectable IgA1 protease activity (mean activity = 580 mU, standard error 30 mU for IgA1 and 500 mU, standard error 30 mU for S-IgA) whereas the carriage isolates mostly displayed lower activity (mean activity 280 mU, standard error 50 mU for IgA1 and 250 mU, standard error 40 mU for S-IgA). This result is statistically significant at the P < 0.0001 level. Six of the 25 carriage isolates displayed no detectable activity in this assay system. All protease-negative supernatants were incubated with substrate for extended periods (up to 12 h), but still failed to show detectable activity.



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  		Figure 1. Graphical representation comparing the levels of IgA1 protease activity associated with invasive (I) and carriage (C) strains of N. meningitidis assayed on both IgA1 (open bars) and secretory IgA (S-IgA, shaded bars) substrates as described in Materials and Methods. Error bars represent the standard error of each mean.

We investigated the reliability of the ELISA assay used by analyzing three representative isolates as follows: five individual colonies of each of three axenic isolates were assayed in triplicate for protease production as described above. The means and ranges of activity determined for each isolate were as follows: strain 1 mean = 530 mU, range 500–560 mU; strain 2 mean = 756, range 740–770 mU; and strain 3 mean = 576 mU, range = 560–600 mU. The standard deviations for these means were less than 5%. The influence of subculturing on IgA1 protease production was investigated. Five individual colonies from one axenic culture agar plate were grown in liquid culture and assayed for IgA1 protease activity as described (day 1 cultures). A small sample of each was then subcultured in fresh media and further subcultured to produce five day 2 cultures. Day 3 cultures were prepared similarly with inocula from day 2 samples. Comparison of the IgA1 protease activities produced by these samples showed that there was no significant variation in the activity produced by the day 1, day 2, and day 3 cultures. Indeed, for all cultures tested in this manner the range of activities recorded was 710–780 mU, mean = 740.


   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
That clinical isolates of N. meningitidis produce higher levels of IgA1 protease activity than carriage isolates could be explained either on the basis that the protease can contribute significantly to virulence or is merely a marker of a linked virulence factor gene. The latter is unlikely since N. meningitidis is known to have a panmictic population structure (except in the case of clonal epidemic isolates) resulting in the random association of alleles (30) . The isolates examined in this study were diverse, as shown by serological characterization. This eliminates the possibility that elevated levels of IgA1 protease activity are merely associated with some other virulence factor. Thus, our data suggest that IgA1 protease is an important virulence determinant of N. meningitidis in that relatively high levels of IgA protease activity were observed in strains that had translocated from the nasopharynx to the bloodstream. Until now, possession of IgA1 protease by colonizing bacteria that use mucosal niches has been an interesting but apparently inconsequential property.

We used two approaches to investigate bacterial IgA1 proteases in these isolates: determination of the presence of the IgA1 protease gene by PCR amplification and semiquantitative measurement of enzyme activity by an ELISA. With regard to IgA1 protease gene detection, all strains tested positive apart from one invasive strain (SVG22) and two carriage strains (SVG23 and SVG26). Although strains SVG22 and SVG23 did not possess an amplifiable IgA1 protease gene sequence when using our primers, they were positive for enzyme activity in the ELISA. The lack of a positive signal for the iga gene in these cases can be explained by chromosomal mutation of nucleotides complementary to the 3' end of the oligonucleotide primers used in this study. Other workers have examined Neisseria cohorts for the presence of an iga gene but have not correlated this with quantified enzyme activity (27 , 31 ).

We applied an ELISA to determine levels of IgA1 protease activity in two large groups of N. meningitidis derived from individuals with polarized disease phenotypes. Previous studies of large cohorts of IgA1 protease-producing bacteria have been qualitative, establishing the presence or absence of enzyme activity, or have determined cleavage types using immunoelectrophoresis or sodium dodecyl sulfate-polyacrylamide gel electrophoresis 32-34) . By using low concentrations of substrates and unconcentrated enzyme preparations, it was possible to detect variation in the levels of enzyme activity of individual strains in this study.

Quantitative determination of enzyme activity revealed several important facts: 1) the level of enzyme activity varies considerably among individual strains of N. meningitidis from undetectable to 950 mU; 2) there is a significant difference in the levels of enzyme activity between invasive and carriage strains (mean value 580 mU vs. 280 mU for IgA1 and 500 mU vs. 250 mU for S-IgA; Fig. 1 ); and 3) the number of strains showing no detectable activity under the assay conditions is much higher in the carriage group (24% vs. only 2% in the invasive group). Together, these findings strongly suggest that bacterial IgA1 proteases are important virulence factors, at least for N. meningitidis.

The ability to cleave S-IgA and IgA1 was comparable, though not identical, for each isolate. Some variation was to be expected when using IgA1 and S-IgA as substrates in the assay. The S-IgA contained a mixture of predominantly S-IgA1 and some S-IgA2, whereas a homogeneous preparation of purified monomeric IgA1 substrate was used. In addition, the observed differences may be a consequence of the presence of IgA1 protease neutralizing antibodies in the S-IgA batch used.

Almost all strains examined were shown to possess the iga gene, so why was there marked variation in enzyme activity? Observed variations in the level of enzyme activity could be explained by iga gene polymorphisms, which can arise naturally during the process of transformation of Neisseria strains and subsequent homologous recombination of DNA segments carrying the IgA1 protease gene. Neisseria is a naturally competent genus, able to incorporate exogenous DNA from related bacteria. For this to occur efficiently, a specific sequence, known as the DNA uptake signal, is required. The IgA1 protease gene is flanked by such sequences, and the polymorphic nature of the N. meningitidis gene iga has been reported (27) . Alternatively, variations in the enzyme activity observed could reflect differing levels of protease production, stability, catalytic efficiency, or even cleavage specificity (as we did not determine the exact site of cleavage in the IgA substrates). Comparison of serologically similar individual carriage and invasive strains also revealed that the latter tend to have elevated protease levels, e.g., in isolates SVG12 and SVG42, both characterized as C, 2a, NT (Tables 1 and 2) ; the former possessed approximately half the activity of the latter. The same patterns of production are seen in other pairs of serologically similar carriage and invasive isolates. For instance, consider SVG36 and SVG85 (B, 4, P1.14, 180 mU vs. 500 mU), SVG99 and SVG91 (C, 2b, P1.2 P1.5, 230 vs. 850 mU), and SVG64 and SVG5 (B, 1, P1.14, 640 vs. 910 mU): in all cases, the clinical isolate produced more protease than the carriage strains, which reflects analysis of the data as a whole.

How might IgA1 protease be involved in pathogenesis? Recent reports indicate that IgA1 protease may fulfill more than just the singular function of subverting the action of hostile antibodies. Two groups have recently demonstrated that N. gonorrhoeae IgA1 protease localizes to and is able to cleave the lysosome-associated membrane protein h-LAMP-1, a heavily glycosylated protein that forms a protective lining enclosing the terminal phagolysosome (35 , 36 ). Furthermore, it has been shown that an iga mutant of N. gonorrhoeae does not degrade LAMP1 and that growth of the mutant in epithelial cells is impaired. Thus, IgA1 protease could facilitate intracellular survival by interfering with phagolysosome maturation. However, there may be other functions of pathogenic importance. The recognition sites recognized by IgA protease include PP(S,T,A)P (37) , a motif present in numerous potentially relevant human proteins. These include macrophage colony-stimulating factor receptor (accession number P07333), the cytokine receptor common beta chain (P32917), interleukin 11 (P20809), interleukin 1 receptor-associated kinase (P51617), and the serum protein MSE55 (Q00587), as revealed by a search of the Swissprot database. In addition, a fragment of the iga gene product (the alpha peptide) has been shown to migrate to the epithelial cell nucleus, but the consequences of this remain obscure (38) . The pathologically relevant function of IgA1 protease is still unclear; a study of N. gonorrhoeae IgA1 protease suggested that it plays only a limited role in penetration of human mucosa in organ culture (39) . This assay was based on a comparison of the invasive potential of an iga mutant and its wild-type parent strain in a fallopian tube organ culture model. However, this report lacked data regarding the presence or absence of antibodies specific to the bacterial strain used. Obviously, the results presented here show that IgA1 protease contributes significantly to virulence but is not an absolute requirement for an invasive strain. As we have demonstrated that pathogenic N. meningitidis isolates possess elevated levels of IgA1 protease, it is possible that cleavage of S-IgA and interactions with subcellular components of the host epithelial cell (as yet unidentified) play a more important role in the development of disease states than was hitherto appreciated. Thus, IgA1 proteases could present a hitherto unrecognized target for the design of new antimicrobial agents or vaccines.


   ACKNOWLEDGMENTS
 
We thank the Special Trustees of the Former United Sheffield Hospitals, the National Meningitis Trust, and The Wellcome Trust for grant support. We acknowledge the assistance of Linda Goodwin, Dr. Trevor Winstanley (Department Medical Microbiology, University of Sheffield), Dr. P. Fenton (Doncaster Royal Infirmary), Dr. P. Norman (Sheffield Northern General Hospital), Dr. M. Osman (Barnsley District General Hospital), and Dr. A. Abbass (Rotherham District General Hospital) in collection of strains. We are indebted to Dr. E. Kaczmarski and other staff at the Meningococcal Reference Unit, Manchester PHL, for confirmation and typing of meningococcal strains.


   FOOTNOTES
 
1 Correspondence. E-mail: j.r.sayers{at}sheffield.ac.uk

2 Abbreviations: CSF, cerebrospinal fluid; Fab, antigen binding fragment; Fc, constant regions; IgA1, immunoglobulin A1; S-IgA, secretory immunoglobulin A; PCR, polymerase chain reaction; ELISA, enzyme-linked immunosorbent assay.

Received for publication July 8, 1998. Accepted for publication on November 24, 1998.
   REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

  1. Childers N. K., Bruce M. G., McGhee J. R.. Molecular mechanisms of immunoglobulin A defense. Annu. Rev. Microbiol. 1989;43:503-536.[Medline]
  2. Lamm M. E., Nedrud J. G., Kaetzel C. S., Mazanec M. B.. IgA and mucosal defense. APMIS 1995;103:241-246.[Medline]
  3. Brandtzaeg P.. Molecular and cellular aspects of the secretory immunoglobulin system. APMIS 1995;103:1-19.[Medline]
  4. Kraehenbuhl J. P., Neutra M. R.. Molecular and cellular basis of immune protection of mucosal surfaces. Physiol. Rev. 1992;72:853-879.[Free Full Text]
  5. Kilian M., Mestecky J., Russell M. W.. Defense mechanisms involving Fc-dependent functions of immunoglobulin A and their subversion by bacterial immunoglobulin A proteases. Microbiol. Rev. 1988;52:296-303.[Free Full Text]
  6. Mulks M. H., Shoberg R. J.. Bacterial immunoglobulin A1 proteases. Methods Enzymol 1994;235:543-554.[Medline]
  7. Mansa B., Kilian M.. Retained antigen-binding activity of Fab{alpha} fragments of human monoclonal immunoglobulin Al (IgAl) cleaved by IgAl protease. Infect. Immun. 1986;52:171-174.[Abstract/Free Full Text]
  8. Kornfeld S. J., Plaut A. G.. Secretory immunity and the bacterial IgA proteases. Rev. Infect. Dis. 1981;3:521-534.[Medline]
  9. Plaut A. G., Bachovchin W. W.. IgA-specific prolyl endopeptidasesserine type. Methods Enzymol 1994;244:137-151.[Medline]
  10. Kilian M., Reinholdt J., Lomholt H., Poulsen K., Frandsen E. V. G.. Biological significance of IgA1 proteases in bacterial colonization and pathogenesiscritical evaluation of experimental evidence. APMIS 1996;104:321-338.[Medline]
  11. Tunkel A. R., Scheld W. M.. Pathogenesis and pathophysiology of bacterial meningitis. Clin. Microbiol. Rev. 1993;6:118-139.[Abstract/Free Full Text]
  12. Delacroix D. L., Dive C., Rambaud J. C., Vaerman J. P.. IgA subclasses in various secretions and in serum. Immunology 1982;47:383-385.[Medline]
  13. Mulks M. H., Plaut A. G.. IgA protease production as a characteristic distinguishing pathogenic from harmless Neisseriaceae. New Engl. J. Med. 1978;299:973-976.[Abstract]
  14. Kilian M., Mestecky J., Schrohenloher J. E.. Pathogenic species of the genus Haemophilus and Streptococcus pneumoniae produce immunoglobulin A1 protease. Infect. Immun. 1979;26:143-149.[Abstract/Free Full Text]
  15. Kilian M., Reinholdt J., Mortensen S. B., Sorensen C. H.. Perturbation of mucosal immune defence mechanisms by bacterial IgA proteases. Bull. Eur. Phisiopathol. Resp. 1983;19:99-104.
  16. Ahl T., Reinholdt J.. Detection of immunoglobulin A1 protease-induced Fab{alpha} fragments on dental plaque bacteria. Infect. Immun. 1991;59:563-569.[Abstract/Free Full Text]
  17. Metha S., Plaut A. G., Calvanico N. J., Tomasi T. B.. Human immunoglobulin Aproduction of an Fc fragment by an enteric microbial proteolytic enzyme. J. Immunol. 1973;111:1274-1276.[Abstract/Free Full Text]
  18. Kilian M., Thomsen B., Petersen T. E., Bleeg H. S.. Occurrence and nature of bacterial IgA proteases. Ann. N.Y. Acad. Sci. 1983;409:612-624.[Medline]
  19. Koomey J. M., Gill R. E., Falkow S.. Genetic and biochemical analysis of gonococcal IgA1 protease cloning in Escherichia coli and construction of mutants of gonococci that fail to produce the activity. Proc. Natl. Acad. Sci. USA 1982;79:7881-7885.[Abstract/Free Full Text]
  20. Koomey J. M., Falkow S.. Nucleotide sequence homology between the immunoglobulin A1 protease genes of Neisseria gonorrhoeae, Neisseria meningitidis, and Hemophilus influenzae. Infect. Immun. 1984;43:101-107.[Abstract/Free Full Text]
  21. Halter R., Pohlner J., Meyer T. F.. IgA protease of Neisseria gonorrhoeaeisolation and characterization of the gene and its extracellular product. EMBO J 1984;3:1595-1601.[Medline]
  22. Pohlner J., Halter R., Beyreuther K., Meyer T. F.. Gene structure and extracellular secretion of Neisseria gonorrhoeae IgA protease. Nature (London) 1987;325:458-462.[Medline]
  23. Reinholdt J., Kilian M.. Comparative analysis of immunoglobulin A1 protease activity among bacteria representing different genera, species, and strains. Infect. Immun. 1997;65:4452-4459.[Abstract]
  24. Read R. C., Zimmerli S., Broaddus V. C., Sanan D. A., Stephens D. S., Ernst J. D.. The ({alpha}2-8)-linked polysialic acid capsule of group B Neisseria meningitidis modifies multiple steps during interaction with human macrophages. Infect. Immun. 1996;64:3210-3217.[Abstract]
  25. Read R. C., Fox A., Miller K., Gray T., Jones N., Borrows R., Jones D. M., Finch R. G.. Experimental infection of human nasal mucosal explants with Neisseria meningitidis. J. Med. Microbiol. 1995;42:353-361.[Abstract]
  26. Virji M., Makepeace K., Peak I. R. A., Ferguson D. J. P., Jennings M. P., Moxon E. R.. Opc-dependent and pilus-dependent interactions of meningococci with human endothelial cellsmolecular mechanisms and modulation by surface polysaccharides. Mol. Microbiol. 1995;18:741-754.[Medline]
  27. Lomholt H., Poulsen K., Kilian M.. Comparative characterization of the iga gene encoding IgA1 protease in Neisseria meningitidis, Neisseria gonorrhoeae and Haemophilus influenzae. Mol. Microbiol. 1995;15:495-506.[Medline]
  28. Greisen K., Loeffelholz M., Purohim A., Leong D.. PCR primers and probes for the 16S rRNA gene of most species of pathogenic bacteria, including bacteria found in cerebrospinal fluid. J. Clin. Microb. 1994;32:335-351.[Abstract/Free Full Text]
  29. Reinholdt J., Kilian M.. A sensitive enzyme linked immunosorbent assay for IgA protease activity. J. Immunol. Methods 1983;63:367-376.[Medline]
  30. Maynard Smith J., Smith N. H., O'Rourke M., Spratt B. G.. How clonal are bacteria?. Proc. Natl. Acad. Sci. USA 1993;90:4384-4388.[Abstract/Free Full Text]
  31. Lomholt H., Poulsen K., Caugant D. A., Kilian M.. Molecular polymorphism and epidemiology of Neisseria meningitidis immunoglobulin A1 proteases. Proc. Natl. Acad. Sci. USA 1992;89:2120-2124.[Abstract/Free Full Text]
  32. Plaut A. G., Gilbert J. V., Artenstain M. S., Capra J. D.. Neisseria gonorrhoeae and Neisseria meningitidisextracellular enzyme cleaves human immunoglobulin A. Science 1975;193:1103-1105.
  33. Mulks M. H., Plaut A. G., Feldman H. A., Frangione B.. IgA proteases of two distinct specificities are released by Neisseria meningitidis. J. Exp. Med. 1980;152:1442-1447.[Abstract/Free Full Text]
  34. Mulks M. H., Knapp J. S.. Immunoglobulin IgA1 protease types of Neisseria gonorrhoeae and their relationship to auxotype and serovar. Infect. Immun. 1992;55:931-936.
  35. Lin L., Ayala P., Larson J., Mulks M., Fukuda M., Carlsson S. R., Enns C., So M.. The Neisseria type 2 IgA1 protease cleaves LAMP1 and promotes survival of bacteria within epithelial cells. Mol. Microbiol. 1997;24:1083-1094.[Medline]
  36. Hauck C. R., Meyer T. F.. The lysosomal/phagosomal membrane protein h-lamp-1 is a target of the IgA1 protease of Neisseria gonorrhoeae. FEBS Lett 1997;405:86-90.[Medline]
  37. Klauser T., Pohlner J., Meyer T. F.. The secretion pathway of IgA protease-type proteins in Gram-negative bacteria. Bioessays 1993;15:799-805.[Medline]
  38. Pohlner J., Langenberg U., Wolk U., Beck S. C., Meyer T. F.. Uptake and nuclear transport of Neisseria Iga1 protease-associated alpha-proteins in human cells. Mol. Microbiol. 1995;17:1073-1083.[Medline]
  39. Cooper M. D., McGee Z. A., Mulks M. H., Koomey J. M., Hindman T. L.. Attachment to and invasion of human fallopian tube mucosa by an IgA1 protease deficient mutant of Neisseria gonorrhoeae and its wild-type parent. J. Infect. Dis. 1984;150:737-744.[Medline]



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Home page
 Infect. Immun.Home page
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Characterization of igaB, a Second Immunoglobulin A1 Protease Gene in Nontypeable Haemophilus influenzae.
Infect. Immun., October 1, 2006; 74(10): 5860 - 5870.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
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The Influences of Hinge Length and Composition on the Susceptibility of Human IgA to Cleavage by Diverse Bacterial IgA1 Proteases
J. Immunol., June 15, 2005; 174(12): 7792 - 7799.
[Abstract] [Full Text] [PDF]

Home page
 J Med MicrobiolHome page
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Neisseria meningitidis: an overview of the carriage state
J. Med. Microbiol., September 1, 2004; 53(9): 821 - 832.
[Abstract] [Full Text] [PDF]

Home page
 Infect. Immun.Home page
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Extracellular Enzymes with Immunomodulating Activities: Variations on a Theme in Streptococcus pyogenes
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[Full Text] [PDF]

Home page
 JAMAHome page
S. Vitovski, K. T. Dunkin, A. J. Howard, and J. R. Sayers
Nontypeable Haemophilus influenzae in Carriage and Disease: A Difference in IgA1 Protease Activity Levels
JAMA, April 3, 2002; 287(13): 1699 - 1705.
[Abstract] [Full Text] [PDF]

Home page
 Infect. Immun.Home page
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Neisserial Immunoglobulin A1 Protease Induces Specific T-Cell Responses in Humans
Infect. Immun., January 1, 2002; 70(1): 335 - 344.
[Abstract] [Full Text] [PDF]

Home page
 Infect. Immun.Home page
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Virulence Functions of Autotransporter Proteins
Infect. Immun., March 1, 2001; 69(3): 1231 - 1243.
[Full Text] [PDF]

Home page
 Clin. Microbiol. Rev.Home page
M. van Deuren, P. Brandtzaeg, and J. W. M. van der Meer
Update on Meningococcal Disease with Emphasis on Pathogenesis and Clinical Management
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[Abstract] [Full Text] [PDF]

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