Pii: s0001-706x(99)00071-6

Pathogenesis of and immunity to melioidosis Department of Microbiology and Infectious Diseases, Uni6ersity of Calgary Health Sciences Center, Calgary, Alta, Canada T2N 4N1 Abstract
While Burkholderia pseudomallei, the causative agent of melioidosis, is becoming increasingly recognized as a significant cause of morbidity and mortality in regions to which it is endemic, no licensed vaccine preparationcurrently exists for immunization against the disease. Therefore, one of the primary goals of our research has beento identify and characterize antigens expressed by B. pseudomallei isolates for the intended purpose of developing avaccine construct that can be used to actively immunize specific high risk populations against the disease. By utilizinga combination of biochemical, immunological and molecular approaches, our studies now indicate that some of themost promising candidates for this task include flagellin proteins and the endotoxin derived O-polysaccharide (PS)antigens expressed by the organism. In this review, we have attempted to summarize the current status of B.
pseudomallei research while endeavoring to provide a rationale for our approach towards the development of amelioidosis vaccine. 2000 Elsevier Science B.V. All rights reserved.
Keywords: Burkholderia pseudomallei; Melioidosis; Pathogenesis; Virulence factors; Protective antigens; Conjugate vaccine 1. Introduction
particularly high in South-East Asia and northernAustralia (Chaowagul et al., 1989; Leelarasamee Burkholderia pseudomallei, the etiological agent and Bovornkitti, 1989; Dance, 1991). In north- of melioidosis, is a Gram-negative, facultative eastern Thailand alone, an estimated 20% of com- anaerobic, motile bacillus that is responsible for a munity acquired septicemia and approximately broad spectrum of illnesses observed in both hu- 40% of deaths due to the complications associated with bacterial sepsis can be attributed to B. pseu- Leelarasamee and Bovornkitti, 1989; Sanford, domallei (Chaowagul et al., 1989). Although the organism is not strictly confined to the equatorial demonstrated that B. pseudomallei is endemic to regions, the probability of acquiring melioidosis regions which typically border 20° north and outside of these geographic domains is exceed- south of the equator, the incidence of disease is ingly low (Howe et al., 1971). Since the microbe isnutritionally diverse and is capable of resisting avariety of environmental extremes, it is puzzling * Corresponding author. Tel.: + 1-403-220-2564; fax: + 1- as to why a more uniform global distribution of E-mail address: woods@acs.ucalgary.ca (D.E. Woods) B. pseudomallei is not more apparent (Smith et al., 0001-706X/00/$ - see front matter 2000 Elsevier Science B.V. All rights reserved.
PII: S 0 0 0 1 - 7 0 6 X ( 9 9 ) 0 0 0 7 1 - 6 P.J. Brett, D.E. Woods / Acta Tropica 74 (2000) 201 – 210 1987). The organism can be readily isolated from tables percolate the organism up through the un- environmental niches such as rice paddies, still or derlying soils to the surface thus enhancing their stagnant waters and moist soils which predomi- potential for exposure to humans and animals.
nate in the tropics, and it is believed that these This route of transmission tends to explain the habitats are the primary reservoirs from which prevalence of disease amongst rice farmers and susceptible hosts acquire infections (Ellison et al., their families who labor in the rice paddies with- 1969; Leelarasamee and Bovornkitti, 1989).
out the benefit of protective clothing (Chaowagul The manifestations of melioidosis are com- et al., 1989; Leelarasamee and Bovornkitti, 1989).
monly represented by acute, sub-acute and Another important route of infection appears to chronic illnesses, with the clinical indications of be the inhalation and aspiration of contaminated some forms of the disease often being mistaken fomites. During the Vietnam war, a dispropor- for malaria, plague, pneumonia and miliary tuber- tionate number of helicopter crewmen succumbed culosis (Howe et al., 1971; Smith et al., 1987; to B. pseudomallei infections as compared to other Leelarasamee and Bovornkitti, 1989). Resistance soldiers stationed in the same regions. To explain to a variety of antimicrobial agents including this phenomenon, it has been proposed that the helicopter rotors acted to disturb infectious dust cephalosporins and many of the aminoglycosides particles around landing zones and thus facilitated is characteristic of B. pseudomallei clinical isolates the pulmonary inoculation of the crewmen with B. pseudomallei (Howe et al., 1971; Sanford, Bovornkitti, 1989; Godfrey et al., 1991; Weinberg 1990). Alternatively, ingestion of the organism and Heller, 1997). With this in mind, accurate and human to human transmission, although to identification of the organism, evaluation of the much lesser extents, have been implicated as severity of the infection and antibiotic susceptibil- routes of inoculation (McCormick et al., 1975).
ity testing are of paramount importance in devis- To date, there have been no reports of transmis- ing an effective chemotherapeutic strategy. While sion of disease between animals and humans the newer therapies that utilize combinations of (Leelarasamee and Bovornkitti, 1989; Dance, ceftazidime – cotrimoxazole or amoxicillin – clavu- lanate for treatment of disease are proving benefi- A number of physiological abnormalities have cial, the mortality rates associated with the acute been correlated with the predisposition of certain septicemic and pulmonary forms of melioidosis populations to B. pseudomallei infections. In par- are still unacceptably high (Smith et al., 1987; ticular, during a 1 year study of patients admitted Leelarasamee and Bovornkitti, 1989; Kanai and to a hospital in north-eastern Thailand for treat- Kondo, 1994; Weinberg and Heller, 1997; Ho et ment of septicemic melioidosis, 32% demonstrated al., 1997). Typically, prolonged oral therapy is pre-existing diabetes mellitus (Chaowagul et al., also recommended to assure the full clinical reso- 1989). Similarly, it has been shown in a diabetic lution of infections while reducing the potential infant rat model of infection that such animals are far more sensitive to challenge with B. pseudoma-llei isolates than are the healthy, non-diabetic rats(Woods et al., 1993). The reasons for this in- 2. Aspects of pathogenesis
creased susceptibility, however, are still being in-vestigated. Other health related factors which Melioidosis is primarily acquired via the inocu- appear to increase the probability of acquiring lation of compromised surface tissues by soils and melioidosis include impaired cellular immunity, waters contaminated with B. pseudomallei; the leukemia/lymphomas, HIV infections, renal disor- highest incidence of disease occurring during the ders, and debilitating afflictions such as alco- monsoon and rainy seasons (Chaowagul et al., holism and parenteral drug abuse (Whitmore and 1989; Leelarasamee and Bovornkitti, 1989). It is believed that during these periods, rising water Leelarasamee and Bovornkitti, 1989; Tanphaichi- P.J. Brett, D.E. Woods / Acta Tropica 74 (2000) 201 – 210 tra, 1989). Although B. pseudomallei related ill- been shown to manifest in dogs, cats, rats, rabbits nesses are documented in apparently healthy indi- and numerous other species. Animals reported to viduals, the organism is probably still best be immune to this organism are fowl, cattle and described as an opportunistic pathogen.
water buffalo (Smith et al., 1987). The latter The manifestations of the various forms of example is rather interesting since these animals melioidosis can be loosely defined as either acute, are frequently used to pull prows in rice paddies sub-acute and chronic (Howe et al., 1971). While and are, therefore, expected to encounter the mi- the incubation periods of B. pseudomallei infec- crobe on a regular basis. Epizootic outbreaks tions are not well defined, a review of the litera- have been documented in various regions of the ture does suggest that they can range from as little world. Notable examples include the death of 24 as a few days to upwards of 26 years (Mays and dolphins in a Hong Kong aquarium in 1975, and Ricketts, 1975; Leelarasamee and Bovornkitti, an outbreak which appeared in the Caribbean in 1989). The acute form of the disease can be 1957 resulting in the death of numerous sheep, subdivided into two groups; the acute pulmonary pigs and goats. Accounts from northern Australia and the acute septicemic forms. Acute pulmonary also relate multiple outbreaks of melioidosis in symptoms appear rapidly and are characterized lamb flocks (Sutmoller et al., 1957; Ketterer and by high fever and pulmonary distress. This is followed by the appearance of visceral abscesses Thus, melioidosis should be considered in any and death within a few days if left untreated. The febrile patient with a history of residence in a septicemic type illness is also rapidly fatal and major endemic region. If Gram-negative bipolar displays a high mortality rate when similarly left staining bacilli are observed in sputum, the organ- untreated. Clinical indications of this form in- ism can be readily cultured and identified (San- clude malaise, meningitis, cellulitis, as well as ford, 1990). In acute cases, blood and urine cutaneous and subcutaneous lesions. In many in- cultures are frequently positive, whereas, if stances the acute manifestations of melioidosis are chronic or sub-acute forms of the disease are associated with an appalling mortality rate, even suspected, biopsy may be required. (Sanford, when vigorous chemotherapeutic intervention is 1990). Serological studies can be helpful for diag- implemented (Howe et al., 1971; Leelarasamee nosing active and recrudescent disease, and an and Bovornkitti, 1989; Dance, 1990; Sanford, immunoglobulin M (IgM) immunofluorescence test is often positive in recent infections (Dance, Sub-acute melioidosis is probably best charac- 1991). Also, indirect hemagglutination and com- terized as a prolonged febrile illness. While multi- plement fixation and tests are available, but re- ple abscess formation can be observed on the quire the testing of paired sera over several weeks viscera, brain abscesses are seldom found. During to confirm the presence of an active infection the latter stages of the disease, the organism can (Smith et al., 1987; Chaowagul et al., 1989).
be readily cultured from blood, pus, urine andother bodily tissues and secretions (Smith et al.,1987; Leelarasamee and Bovornkitti, 1989). In 3. Virulence factors and protective antigens
many instances, death occurs within a few weeksto months in the absence of clinical intervention.
Although B. pseudomallei isolates are capable The sub-clinical or chronic form of the disease is of expressing an impressive array of both secreted considered to be the most common presentation and cell-associated antigens, the role(s) of these of melioidosis. It generally remains undiagnosed, products in the pathogenesis of disease have to however, until activated by a traumatic event or date been relatively ill defined. One of the primary upon post-mortem examination of the tissues reasons for this has been due to the lack of suitable techniques for genetically manipulating B. pseudomallei causes melioidosis in animals as the organism. Due to the recent application of a well as humans. In endemic areas, disease has Tn5 transposon based mutagenesis system for use P.J. Brett, D.E. Woods / Acta Tropica 74 (2000) 201 – 210 in B. pseudomallei, we have begun to identify and cross-reactive with this antigen (Sexton et al., characterize genetic loci which encode a number of these putative virulence determinants and pro- Most recently, via transposon mutagenesis, we tective antigens (DeShazer et al., 1997, 1998).
have identified an 11.8 kb chromosomal locus in Therefore, in the following sections we have at- B. pseudomallei, that demonstrates a high degree tempted to summarize the results of our most of homology to operons which encode for the current studies, as well as those from other products of the main terminal branch of general groups, in order to provide an overview of the secretory pathway (GSP) (Pugsley, 1993). Further antigens expressed by this B. pseudomallei.
characterization of the open reading frames in thislocus have confirmed that their orientation and physical arrangement are virtually identical to thepul gene cluster of Klebsiella oxytoca (Pugsley, The ability to acquire iron from host sources is 1993). Not surprisingly, the phenotypic analysis of a prerequisite for the successful establishment and the individual transposon mutants has also confi- maintenance of most bacterial infections. Yang et rmed their inability to secrete antigens associated al. have demonstrated that 84/84 B. pseudomallei with protease, lipase and lecithinase into the ex- strains examined during their studies tested posi- tracellular milieu. Interestingly, while we screened tive for siderophore production using the chrome more than 30 000 mutants for the loss of one or azurol S (CAS) assay. A structural and chemical more of the enzymatic activities, no protease, analysis of the siderophore synthesized by B.
lipase or lecithinase structural genes were iden- pseudomallei U7 confirmed that the molecule was tified. In order to asses the significance of GSP approximately 1000 Da in size, water soluble with secreted products in the pathogenesis of me- a yellow – green fluorescence and that it belonged lioidosis, we compared the virulence of the secre- to the hydroxamate class (Yang et al., 1991).
tion mutants to the wild type strain in the Syrian Furthermore, studies have also demonstrated the hamster model of infection. The results of these siderophore was capable of scavenging iron from studies indicated that while the protease, lipase both lactoferrin and transferrin in vitro (Yang et and lecithinase may play a small role in the al., 1993). The name malleobactin has been pro- pathogenesis of acute melioidosis, mutants defi- posed for this compound (Yang et al., 1991).
cient in their ability to secrete these particular It has been previously shown that B. pseudoma- exoenzymes were not severely attenuated in their llei isolates are capable of secreting antigens that ability to cause a fulminating illness (Woods, un- demonstrate biological activities consistent with proteases, lecithinases, lipases and hemolysins In the mid 1950s, several studies demonstrated (Esselman and Liu, 1961; Ashdown and Koehler, that filter sterilized B. pseudomallei culture super- 1990; Sexton et al., 1994). However, while these natants were lethal for mice and hamsters when factors have been implicated as important factors administered parenterally (Nigg et al., 1955; in the pathogenesis of the disease, only the Heckly and Nigg, 1958; Heckly, 1964). These protease has been characterized to date. Studies results were consistent with the fulminating ill- conducted by Sexton et al. have confirmed the nesses observed in animals following inoculation presence of a 36 000 kDa antigen with associated with viable bacteria, and suggested that B. pseu- proteolytic activities in B. pseudomallei culture domallei strains might be capable of secreting a supernatants. In particular, a protease expressed lethal toxin. In studies conducted by Ismail et al.
by B. pseudomallei 319a was found to be a metal- (1987) mouse lethal, thermolabile toxin was re- loenzyme requiring iron for maximal protease ac- portedly purified to homogeneity and character- tivity and demonstrated optimally activity at pH ized as a 31 000 MW protein. Haase et al. (1997) 8.0 and 60°C (Sexton et al., 1994). Furthermore, have also described the presence of cytotoxic ac- monoclonal antibodies (MAb) raised against a tivity in culture filtrates. Their results, however, Pseudomonas aeruginosa alkaline protease were P.J. Brett, D.E. Woods / Acta Tropica 74 (2000) 201 – 210 and that the cytotoxic activity is this instance is Haemophilus influenzae and Streptococcus pneu- most likely due to the presence of a small peptide.
moniae. These extracellular moieties enable bacte- Recently it has been reported that a rhamnolipid purified from B. pseudomallei culture supernatants inhibiting complement activation and phagocytic demonstrates a cytotoxic effect against HL60 and mediated killing (Joiner, 1988). A review of the HeLa cell lines (Haubler et al., 1998). Since this literature also indicates that B. pseudomallei activity can be neutralized by albumin, however, strains are capable of synthesizing capsular anti- it is unlikely to be of consequence in the patho- gens (Smith et al., 1987; Leelarasamee and genesis of B. pseudomallei infections. Curiously, Bovornkitti, 1989), and that they may play an while it has also been our experience that filter important role in the pathogenesis of melioidosis.
sterilized B. pseudomallei culture supernatants are While in vitro studies have determined that encap- lethal for a variety of tissue culture cell lines, we sulated B. pseudomallei strains are as susceptible have been unable to reproduce these effects in to phagocytic uptake by polymorphonuclear animal models, even when using preparations leukocytes (PMN) as non-encapsulated variants, concentrated by lyophilization (Brett et al., 1997, evidence tends to suggest that the presence of exopolysaccharide confers upon them the abilityto resist the bactericidal effects of the phagolyso- 3.2. Cell-associated antigens somal environment (Smith et al., 1987; Pruk-sachartvuthi et al., 1990). This is a feature of B.
A number of previous studies have confirmed pseudomallei strains that may help to explain why that B. pseudomallei is capable of synthesizing an these organisms are capable of remaining latent in acid phosphatase. It now appears that the molecule is a glycoprotein with optimal substrate Recently, Steinmetz et al. isolated and purified activity around pH 7.2 (Kanai and Kondo, 1991, a high molecular weight capsular antigen ( \150 1994; Kondo et al., 1996). While it has been kDa) from B. pseudomallei NCTC 7431 and suc- reported by Kanai and Kondo (1994) and Kondo ceeded in raising a MAb against it (Steinmetz et et al. (1996) that the acid phosphatase can be al., 1995). Via ELISA based techniques, they were readily isolated from both whole cell and superna- able to demonstrate the reactivity of both mucoid tant fractions, we have found the enzyme to be and non-mucoid strains with the MAb, thus sug- predominantly cell-associated (unpublished data).
gesting that the capsular antigen is constitutively Kanai et al. (1996) have also proposed that the expressed by B. pseudomallei strains. Interestingly, cell-associated form of the enzyme is a high temperature appeared to have little effect on the affinity receptor for insulin and that this receptor/ synthesis of the exopolysaccharide since B. pseu- ligand interaction may be responsible for modu- domallei strains grown at both 15 and 37°C re- lating the enzymatic profiles of B. pseudomallei acted with the MAb. Furthermore, an assay isolates. Recently, while screening transposon mu- utilizing a variety of Pseudomonas and Burkholde- tants for their inability to hydrolyze the chro- ria spp. as controls was able to confirm the spe- cificity of the MAb for B. pseudomallei and B.
phosphate (X-P), we successfully identified a mu- mallet strains only (Steinmetz et al., 1995). More tant devoid of acid phosphatase activity (unpub- recently, Masoud et al. (1997) have been success- ful at elucidating the chemical and structural chromosomal DNA flanking the transposon inser- characteristics of a capsular polysaccharide iso- tion demonstrated the presence of an open read- lated from the virulent clinical isolate B. pseudo- ing frame whose translated product was highly mallei 304b. Their results demonstrated that the homologous to an acid phosphatase expressed by exopolysaccharide was a linear unbranched poly- Francisella tularensis (var. novicida).
mer of repeating tetrasaccharide units having the Capsular polysaccharides are a common feature (-3)-2-O-Ac-b-D-Galp-(1-4)-a-D-Galp- (1-3)-b-D-Galp-(1-5)-b-D-KDOp-(2-).
P.J. Brett, D.E. Woods / Acta Tropica 74 (2000) 201 – 210 Nimtz et al. (1997) have demonstrated that a is required for the synthesis of the Type II antigen structurally identical capsular antigen is expressed and conferring the serum resistance phenotype by B. pseudomallei NCTC 7431. Studies by both (DeShazer et al., 1998). Using the infant diabetic groups have also shown that patient sera reacted rat model, we have also confirmed that Type II strongly with the purified carbohydrate antigens PS is probably a significant determinant in the indicating that this carbohydrate polymer is most pathogenesis of melioidosis since the LD50 value likely expressed in vivo (Steinmetz et al., 1995; associated with a Type II PS mutant is approxi- mately 140 fold higher than that of the wild type Previous studies have confirmed that the lipo- strain (Woods et al., 1993; DeShazer et al., 1998).
polysaccharide (LPS) antigens expressed by B.
Flagella are commonly recognized as important pseudomallei strains are highly conserved through- virulence determinants expressed by bacterial out this species (Pitt et al., 1992). In fact, serolog- pathogens since the motility phenotype imparted ical evidence suggests that there may be only one by these organelles often correlates with the abil- serotype of B. pseudomallei (Bryan et al., 1994). In ity of an organism to cause disease (Penn and order to investigate this phenomenon, Perry et al.
Luke, 1992; Moens and Vanderleyden, 1996).
has characterized the LPS antigens isolated from Therefore, we have focused a great deal of atten- a number of B. pseudomallei strains (Perry et al., tion towards determining the relative importance 1995). Their results demonstrated that B. pseudo- of motility in the pathogenesis of melioidosis. In mallei strains coordinately express two distinct previous studies we have demonstrated that a somatic O-antigens (PS) on their cell surface. The significant degree of size and antigenic homogene- Type I antigen consists of a high-molecular weight ity exists amongst flagellins expressed by B. pseu- unbranched 1,3-linked homopolymer of 2-O- domallei isolates. Furthermore, we have also shown that flagellin specific antiserum is capable residues, while the Type II antigen is an un- of passively protecting diabetic infant rats against branched heteropolymer consisting of (-3)-b-D- a B. pseudomallei challenge (Brett et al., 1994).
glucopyranose-(1-3)-6-deoxy-a-L-talopyranosyl- Curiously, however, in recent studies we have (1-disaccharide repeats (L-6dTalp: found that there was no significant difference between the virulence capacities associated with a lated) (Knirel et al., 1992; Perry et al., 1995).
wild-type strain of B. pseudomallei and non-motile While the simultaneous expression of two or more mutants in either the diabetic infant rat or Syrian LPS moieties is not an uncommon feature associ- hamster models of infection (DeShazer et al., ated with Gram-negative bacteria, the degree to 1997). Thus, taken together, these results indicate which the two PS antigens are conserved amongst that while flagella and/or motility may not be B. pseudomallei strains is quite remarkable (Perry major virulence determinants in the pathogenesis of melioidosis, purified flagellin may still serve as It has been previously reported that B. pseudo- a protective immunogen against B. pseudomallei mallei strains are resistant to the bactericidal ef- fects of normal human serum (Ismail et al., 1988).
Recently, we have established that the presence ofType II PS is essential for conferring this resis- 4. Conjugate vaccines
tance phenotype. Via the application of the trans-poson mutagenesis system, we have identified a Previous studies have demonstrated that both number of mutants that demonstrate a marked polyclonal antiserum and MAb’s raised against B.
sensitivity, in comparison to the parental strain, pseudomallei flagellin proteins, LPS and a tetanus to the bactericidal effects of normal serum. Se- toxin-PS glycoconjugate are capable of passively quence analysis of the chromosomal DNA flank- immunizing diabetic infant rats against challenge ing these transposon insertions has enabled us to with B. pseudomallei (Brett et al., 1994; Bryan et identify a 17.5 kb region of the chromosome that al., 1994). While these initial results were encour- P.J. Brett, D.E. Woods / Acta Tropica 74 (2000) 201 – 210 aging, it had become evident to us, however, that only the PS portion of LPS and not the toxic an active vaccine preparation would be the most component of LPS, namely lipid A, conjugated to practical for immunizing high risk populations flagellin protein obviates the toxic side effects of against melioidosis. Therefore, with this in mind, LPS but takes advantage of the protective re- we synthesized a glycoconjugate molecule that sponse to the PS moiety. Furthermore, the coordi- incorporated both flagellin protein and PS anti- nate presentation of multiple protective antigens gens. Based upon the preliminary success of these has served to enhance the immunological reper- studies, we believe that this preparation is a rea- toire of the vaccine recipient while providing im- sonable vaccine candidate for actively immunizing munity against a number of B. pseudomallei against the disease (Brett and Woods, 1996).
It has been well documented that T cell-inde- Since the size of the saccharide components pendent type 2 (TI-2) antigens such as bacterial displayed by a conjugate molecule appears to polysaccharides are capable of eliciting protective influence the immunogenicity of the preparation, antibody responses in adults (Dintzis, 1992; Jen- a heterogeneous combination of PS antigens were nings, 1992), but act poorly as immunogens in the covalently linked to the flagellin protein carrier elderly, in immunocompromised populations and (Dintzis et al., 1983, 1985; Jennings, 1992). The in children less than 18 months of age (Gold et incorporation of larger PS fragments act to in- al., 1977; Robbins, 1978; Jennings, 1983). In adult crease the size of the conjugate molecules such populations, TI-2 antigens typically stimulate the that the cross-linking of B cell mIg’s is maxi- synthesis of antigen specific immunoglobulin M mized. At the same time, the larger PS molecules (IgM), but often fail to evoke augmented immune also tend to retain conformationally stabilized epitopes which appear to be critical in the design boosting with carbohydrate preparations. This of a successful vaccine (Jennings, 1992). The phenomenon is primarily due to the inability of smaller oligosaccharide fragments appear to be activated B lymphocytes to recruit CD4+ T cell critical for stimulating immunoglobulin responses (Th) involvement via major histocompatibility against terminal epitopes of the carbohydrate complex type II (MHC II) restricted events (Hodgkin and Basten, 1995; Mond et al., 1995).
In order to remedy this situation, a number of raised against the glycoconjugate conjugate reacts bacterial carbohydrates (such as capsular polysac- with both purified flagellin protein and both Type charides or somatic O-antigens) have been cova- I and II PS; a situation that is obviously critical if lently coupled to protein or peptide carrier the immune serum is to be effective against a B.
molecules in order to facilitate T cell-dependent pseudomallei challenge. This confirmed that native (TD) immune responses against the particular car- epitopes displayed by the precursor molecules bohydrate moieties (Cryz et al., 1986; Insel and were maintained during the synthesis of the conju- Anderson, 1986; Winter et al., 1988; Watson et gate molecule. Furthermore, passive immuniza- al., 1992; Fattom et al., 1993; Konadu et al., 1994; tion studies have demonstrated that the IgG Lett et al., 1994). The presence of T cell epitopes fraction purified from the immune serum was inherent to peptide or proteinacious carriers is capable of protecting diabetic infant rats from a believed to facilitate these events (Braley-Mullen, challenge with B. pseudomallei (Brett and Woods, 1996). Based upon the success of these prelimi- Therefore, by conjugating the PS moieties to nary studies, we are currently characterizing the the flagellin carrier, we have been able to elicit efficacy of this vaccine preparation in active im- augmented immune responses against the PS com- ponents of the vaccine while concomitantly evok- Since B. pseudomallei isolates have been shown ing desirable immunoglobulin (Ig) class switching to express both the Type I and II PS antigens (an events (Brett and Woods, 1996). More impor- exception being 824a which displays only the tantly, the use of an active vaccine containing Type I antigen) and the flagellin proteins appear P.J. Brett, D.E. Woods / Acta Tropica 74 (2000) 201 – 210 to be antigenically conserved, we expect the con- Dance, D.A., Wuthiekanun, V., White, N.J., Chaowagul, W., jugate molecule to afford protection against the 1988. Antibiotic resistance in Pseudomonas pseudomallei.
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DeShazer, D., Brett, P.J., Carlyon, R., Woods, D.E., 1997.
This work was supported by the Canadian Bac- Mutagenesis of Burkholderia pseudomallei with Tn5- terial Diseases Network of Centers of Excellence.
OT182: isolation of motility mutants and molecular char- P.J.B. is the recipient of an Alberta Heritage acterization of the flagellin structural gene. J. Bacteriol.
179, 2116 – 2125.
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