Jejuni Lipooligosaccharide Research Articles

Th1/Th17-mediated Immunity and Protection from Peripheral Neuropathy in Wildtype and IL10-/- BALB/c Mice Infected with a Guillain-Barré Syndrome-associated Campylobacter jejuni Strain.

Campylobacter jejuni is an important cause of bacterial gastroenteritis worldwide and is linked to Guillain-Barré syndrome (GBS), a debilitating postinfectious polyneuropathy. The immunopathogenesis of GBS involves the generation of antibodies that are cross reactive to C. jejuni lipooligosaccharide and structurally similar peripheral nerve gangliosides. Both the C. jejuni infecting strain and host factors contribute to GBS development. GBS pathogenesis is associated with Th2-mediated responses in patients. Moreover, induction of IgG1 antiganglioside antibodies in association with colonic Th2-mediated immune responses has been reported in C. jejuni-infected C57BL/6 IL10-/- mice at 4 to 6 wk after infection. We hypothesized that, due to their Th2 immunologic bias, BALB/c mice would develop autoantibodies and signs of peripheral neuropathy after infection with a GBS patient-derived strain of C. jejuni (strain 260.94). WT and IL10-/- BALB/c mice were orally inoculated with C. jejuni 260.94, phenotyped weekly for neurologic deficits, and euthanized after 5 wk. Immune responses were assessed as C. jejuni-specific and antiganglioside antibodies in plasma and cytokine production and histologic lesions in the proximal colon. Peripheral nerve lesions were assessed in dorsal root ganglia and their afferent nerve fibers by scoring immunohistochemically labeled macrophages through morphometry. C. jejuni 260.94 stably colonized both WT and IL10-/- mice and induced systemic Th1/Th17-mediated immune responses with significant increases in C. jejuni-specific IgG2a, IgG2b, and IgG3 plasma antibodies. However, C. jejuni 260.94 did not induce IgG1 antiganglioside antibodies, colitis, or neurologic deficits or peripheral nerve lesions in WT or IL10-/- mice. Both WT and IL10-/- BALB/c mice showed relative protection from development of Th2-mediated immunity and antiganglioside antibodies as compared with C57BL/6 IL10-/- mice. Therefore, BALB/c mice infected with C. jejuni 260.94 are not an effective disease model but provide the opportunity to study the role of immune mechanisms and host genetic background in the susceptibility to post infectious GBS.

Molecular and in silico typing of the lipooligosaccharide biosynthesis gene cluster in Campylobacter jejuni and Campylobacter coli.

The extensive genetic variation in the lipooligosaccharide (LOS) core biosynthesis gene cluster has led to the development of a classification system; with 8 classes (I-VIII) for Campylobacter coli (C. coli) LOS region and with 23 classes (A-W) or four groups (1–4) for Campylobacter jejuni (C. jejuni) LOS region. PCR based LOS locus type identification for C. jejuni clinical isolates from a UK hospital as well as in silico LOS locus analysis for C. jejuni and C. coli genome sequences from GenBank was carried out to determine the frequencies of various LOS genotypes in C. jejuni and C. coli. Analysis of LOS gene content in 60 clinical C. jejuni isolates and 703 C. jejuni genome sequences revealed that class B (Group 1) was the most abundant LOS class in C. jejuni. The hierarchy of C. jejuni LOS group prevalence (group 1 > group 2 > group 3 > group 4) as well as the hierarchy of the frequency of C. jejuni LOS classes present within the group 1 (B > C > A > R > M > V), group 2 (H/P > O > E > W), group 3 (F > K > S) and group 4 (G > L) was identified. In silico analysis of LOS gene content in 564 C. coli genome sequences showed class III as the most abundant LOS locus type in C. coli. In silico analysis of LOS gene content also identified three novel LOS types of C. jejuni and previously unknown LOS biosynthesis genes in C. coli LOS locus types I, II, III, V and VIII. This study provides C. jejuni and C. coli LOS loci class frequencies in a smaller collection of C. jejuni clinical isolates as well as within the larger, worldwide database of C. jejuni and C. coli.

Multi-Locus Sequence Typing and Lipooligosaccharide Class Analysis of Campylobacter jejuni HS:19 Isolated in Japan.

Campylobacter jejuni is a major foodborne pathogen that causes enteritis in humans, and is also known to be an antecedent infectious factor for Guillain-Barré syndrome (GBS). The onset of GBS after C. jejuni infection results from molecular mimicry between human neuronal gangliosides and C. jejuni lipooligosaccharides (LOS). C. jejuni HS:19 has been previously isolated from GBS cases more frequently than other serotypes in Japan. Therefore, in this study, we performed molecular analysis of 88 HS:19 isolates from GBS cases, sporadic diarrhea patients, and poultry meat samples, using multi-locus sequence typing and LOS class analysis. As a result, 87 of the 88 HS:19 isolates were typed as ST22 / CC22 and LOS class A1, while one was typed as ST1947 / CC22 and LOS class A1. Furthermore, the analysis of another 331 isolates from sporadic enteritis cases showed that only 34 (10.3%) were classified as LOS class A, including HS:19 (25 isolates), HS:2 (8 isolates), and HS:4c (1 isolate). In conclusion, C. jejuni HS:19 had high clonality, regardless of its origin, compared to other capsule types in Japan.

Murine Models for the Investigation of Colonization Resistance and Innate Immune Responses in Campylobacter Jejuni Infections.

Human infections with the food-borne pathogen Campylobacter jejuni are progressively increasing worldwide and constitute a significant socioeconomic burden to mankind. Intestinal campylobacteriosis in humans is characterized by bloody diarrhea, fever, abdominal pain, and severe malaise. Some individuals develop chronic post-infectious sequelae including neurological and autoimmune diseases such as reactive arthritis and Guillain-Barré syndrome. Studies unraveling the molecular mechanisms underlying campylobacteriosis and post-infectious sequelae have been hampered by the scarcity of appropriate experimental in vivo models. Particularly, conventional laboratory mice are protected from C. jejuni infection due to the physiological colonization resistance exerted by the murine gut microbiota composition. Additionally, as compared to humans, mice are up to 10,000 times more resistant to C. jejuni lipooligosaccharide (LOS) constituting a major pathogenicity factor responsible for the immunopathological host responses during campylobacteriosis. In this chapter, we summarize the recent progress that has been made in overcoming these fundamental obstacles in Campylobacter research in mice. Modification of the murine host-specific gut microbiota composition and sensitization of the mice to C. jejuni LOS by deletion of genes encoding interleukin-10 or a single IL-1 receptor-related molecule as well as by dietary zinc depletion have yielded reliable murine infection models resembling key features of human campylobacteriosis. These substantial improvements pave the way for a better understanding of the molecular mechanisms underlying pathogen-host interactions. The ongoing validation and standardization of these novel murine infection models will provide the basis for the development of innovative treatment and prevention strategies to combat human campylobacteriosis and collateral damages of C. jejuni infections.

An Updated Classification System and Review of the Lipooligosaccharide Biosynthesis Gene Locus in Campylobacter jejuni.

Lipooligosaccharide (LOS) is an integral component of the Campylobacter cell membrane with a structure of core oligosaccharides forming inner and outer core regions and a lipid A moiety. The gene content of the LOS core biosynthesis cluster exhibits extensive sequence variation, which leads to the production of variable cell surface LOS structures in Campylobacter. Some LOS outer core molecules in Campylobacter jejuni are molecular mimics of host structures (such as neuronal gangliosides) and are thought to trigger neuronal disorders (particularly Guillain–Barré syndrome and Miller Fisher syndrome) in humans. The extensive genetic variation in the LOS biosynthesis gene cluster, a majority of which occurs in the LOS outer core biosynthesis gene content present between lgtF and waaV, has led to the development of a classification system with 23 classes (A–W) and four groups (1–4) for the C. jejuni LOS region. This review presents an updated and simplified classification system for LOS typing alongside an overview of the frequency of C. jejuni LOS biosynthesis genotypes and structures in various C. jejuni populations.

Novel Campylobacter concisus lipooligosaccharide is a determinant of inflammatory potential and virulence

The pathogenicity of Campylobacter concisus, increasingly found in the human gastrointestinal (GI) tract, is unclear. Some studies indicate that its role in GI conditions has been underestimated, whereas others suggest that the organism has a commensal-like phenotype. For the enteropathogen C. jejuni, the lipooligosaccharide (LOS) is a main driver of virulence. We investigated the LOS structure of four C. concisus clinical isolates and correlated the inflammatory potential of each isolate with bacterial virulence. Mass spectrometric analyses of lipid A revealed a novel hexa-acylated diglucosamine moiety with two or three phosphoryl substituents. Molecular and fragment ion analysis indicated that the oligosaccharide portion of the LOS had only a single phosphate and lacked phosphoethanolamine and sialic acid substitution, which are hallmarks of the C. jejuni LOS. Consistent with our structural findings, C. concisus LOS and live bacteria induced less TNF-α secretion in human monocytes than did C. jejuni Furthermore, the C. concisus bacteria were less virulent than C. jejuni in a Galleria mellonella infection model. The correlation of the novel lipid A structure, decreased phosphorylation, and lack of sialylation along with reduced inflammatory potential and virulence support the significance of the LOS as a determinant in the relative pathogenicity of C. concisus.

Molecular Mimicry and Guillain-Barré Syndrome

One-thirds of patients develop Guillain-Barré syndrome subsequent to Campylobacter jejuni enteritis. Molecular mimicry exists between C. jejuni lipo-oligosaccharides and human peripheral nerve gangliosides GM1 and GD1a. IgG antibodies against GM1 or GD1a are produced in one out of 5,000 patients with C. jejuni enteritis. The autoantibodies bind to gangliosides at the nodes of Ranvier in the peripheral motor nerves and activate complement in situ. This is followed by the disappearance of the voltage-gated sodium channel clusters at the nodes and disruption of axo-glial junctions at the paranodes. This results in the development of motor nerve conduction failure and muscle weakness in the four limbs.

Immunoglobulin G Fc N-glycosylation in Guillain-Barré syndrome treated with intravenous immunoglobulin.

Immunoglobulin G Fc N-glycosylation in Guillain-Barré syndrome treated with intravenous immunoglobulin.

Functional characterization of exopolyphosphatase/guanosine pentaphosphate phosphohydrolase (PPX/GPPA) of Campylobacter jejuni

The inorganic polyphosphate (poly-P) is a key regulator of stress responses and virulence in many bacterial pathogens including Campylobacter jejuni. The role of exopolyphosphatases/guanosine pentaphosphate (pppGpp) phosphohydrolases (PPX/GPPA) in poly-P homeostasis and C. jejuni pathobiology remains unexplored. Here, we analyzed deletion mutants (∆ppx1, ∆ppx2) and the double knockout mutant (dkppx), all ∆ppx mutants exhibited increased capacity to accumulate poly-P; however only ∆ppx1 and dkppx mutants showed decreased accumulation of ppGpp, an alarmone molecule that regulates stringent response in bacteria, suggesting potential dual role for PPX1/GPPA. Nutrient survival defect of ∆ppx mutants was rescued by the supplementation of specific amino acids implying that survival defect may be associated with decreased ppGpp and/ or increased poly-P in ∆ppx mutants. The ppk1 and spoT were upregulated in both ∆ppx1 and ∆ppx2 suggesting a compensatory role for SpoT and Ppk1 in poly-P and ppGpp homeostasis. The lack of ppx genes resulted in defects in motility, biofilm formation, nutrient stress survival, invasion and intracellular survival indicating that maintaining a certain level of poly-P is critical for ppx genes in C. jejuni pathophysiology. Both ppx1 and ppx2 mutants were resistant to human complement-mediated killing; however, the dkppx mutant was sensitive. The serum susceptibility did not occur in the presence of MgCl2 and EGTA suggesting an involvement of the classical or lectin pathway of complement mediated killing. Interestingly, the chicken serum did not have any effect on the ∆ppx mutants’ survival. The observed serum susceptibility was not related to C. jejuni surface capsule and lipooligosaccharide structures. Our study underscores the importance of PPX/GPPA proteins in poly-P and ppGpp homeostasis, two critical molecules that modulate environmental stress responses and virulence in C. jejuni.

Guillain-Barré Syndrome and Campylobacter jejuni Infection: A Review

Guillain-Barré syndrome (GBS), a neurologic disease that produces ascending paralysis, affects people all over the world. Acute infectious illness precedes 50%-75% of the GBS cases. Although many infectious agents have been associated with GBS, the strongest documented association is with Campylobacter infection. The first line of evidence supporting Campylobacter infection as a trigger of GBS is anecdotal reports. The second line of evidence is serological surveys, which have demonstrated that sera from GBS patients contain anti Campylobacter jejuni antibodies, consistent with recent infection. Finally, culture studies have proven that a high proportion of GBS patients have C. jejuni in their stools at the time of onset of neurological symptoms. One of every 1058 Campylobacter infections results in GBS. Sialic acid containing lipooligosaccharides (LOS) biosynthesis gene locus are associated with GBS and the expression of ganglioside mimicking structures. GM1a was the most prevalent ganglioside mimic in GBS associated strains. Molecular mimicry between C. jejuni LOS and gangliosides in human peripheral nerves, and cross-reactive serum antibody precipitate the majority of GBS cases in Bangladesh, like worldwide. DOI: http://dx.doi.org/10.3329/dmcj.v2i1.17794 Delta Med Col J. Jan 2014; 2(1): 28-35

Sialylation of Campylobacter jejuni Endotoxin Promotes Dendritic Cell–Mediated B Cell Responses through CD14-Dependent Production of IFN-β and TNF-α

Campylobacter jejuni is the most common bacterial cause of human gastroenteritis and often precedes development of Guillain-Barré syndrome (GBS), a life-threatening paralytic disease. The incorporation of the carbohydrate sialic acid into C. jejuni lipooligosaccharides (LOS) is associated with increased severity of gastroenteritis and with induction of GBS; however, the underlying mechanisms remain completely unknown. In this study, we demonstrate that sialic acids in C. jejuni endotoxin enhance the rapid production of IFN-β and TNF-α by human dendritic cells (DCs). Using neutralizing Abs and receptors it was shown that these DC-derived cytokines promote the proliferation of human mucosal B cells in a T cell-independent manner. The production of both IFN-β and TNF-α by DCs in response to LOS requires CD14, and the amplified response of DCs to sialylated C. jejuni LOS is CD14 dependent. Together, these results indicate that sialylation of C. jejuni LOS increases DC activation and promotes subsequent B cell responses through CD14-driven production of IFN-β and TNF-α. This enhanced DC/B cell response may explain the increased pathogenicity of sialylated C. jejuni and may be key to the initiation of B cell-mediated autoimmunity in GBS.

Campylobacter jejuni Lipooligosaccharide Sialylation, Phosphorylation, and Amide/Ester Linkage Modifications Fine-tune Human Toll-like Receptor 4 Activation

Campylobacter jejuni is a leading cause of acute gastroenteritis. C. jejuni lipooligosaccharide (LOS) is a potent activator of Toll-like receptor (TLR) 4-mediated innate immunity. Structural variations of the LOS have been previously reported in the oligosaccharide (OS) moiety, the disaccharide lipid A (LA) backbone, and the phosphorylation of the LA. Here, we studied LOS structural variation between C. jejuni strains associated with different ecological sources and analyzed their ability to activate TLR4 function. MALDI-TOF MS was performed to characterize structural variation in both the OS and LA among 15 different C. jejuni isolates. Cytokine induction in THP-1 cells and primary monocytes was correlated with LOS structural variation in each strain. Additionally, structural variation was correlated with the source of each strain. OS sialylation, increasing abundance of LA d-glucosamine versus 2,3-diamino-2,3-dideoxy-d-glucose, and phosphorylation status all correlated with TLR4 activation as measured in THP-1 cells and monocytes. Importantly, LOS-induced inflammatory responses were similar to those elicited by live bacteria, highlighting the prominent contribution of the LOS component in driving host immunity. OS sialylation status but not LA structure showed significant association with strains clustering with livestock sources. Our study highlights how variations in three structural components of C. jejuni LOS alter TLR4 activation and consequent monocyte activation.

Guillain-Barré Syndrome-Related Campylobacter jejuni in Bangladesh: Ganglioside Mimicry and Cross-Reactive Antibodies

Background Campylobacter jejuni is the predominant antecedent infection in Guillain-Barré syndrome (GBS). Molecular mimicry and cross-reactive immune responses to C. jejuni lipo-oligosaccharides (LOS) precipitate the development of GBS, although this mechanism has not been established in patients from developing countries. We determined the carbohydrate mimicry between C. jejuni LOS and gangliosides, and the cross-reactive antibody response in patients with GBS in Bangladesh.MethodologySera from 97 GBS patients, and 120 neurological and family controls were tested for antibody reactivity against LOS from C. jejuni isolates from GBS patients in Bangladesh (BD-07, BD-39, BD-10, BD-67 and BD-94) by enzyme-linked immunosorbent assay (ELISA). Cross-reactivity to LOS was determined by ELISA. The LOS outer core structures of C. jejuni strains associated with GBS/MFS were determined by mass spectrometry.Principle FindingsIgG antibodies to LOS from C. jejuni BD-07, BD-39, BD-10, and BD-67 IgG antibodies were found in serum from 56%, 58%, 14% and 15% of GBS patients respectively, as compared to very low frequency (<3%) in controls (p<0.001). Monoclonal antibodies specific for GM1 and GD1a reacted strongly with LOS from the C. jejuni strains (BD-07 and BD-39). Mass spectrometry analysis confirmed the presence of GM1 and GD1a carbohydrate mimics in the LOS from C. jejuni BD-07 and BD-39. Both BD-10 and BD-67 express the same LOS outer core, which appears to be a novel structure displaying GA2 and GD3 mimicry. Up to 90–100% of serum reactivity to gangliosides in two patients (DK-07 and DK-39) was inhibited by 50 µg/ml of LOS from the autologous C. jejuni isolates. However, patient DK-07 developed an anti-GD1a immune response while patient DK-39 developed an anti-GM1 immune response.ConclusionCarbohydrate mimicry between C. jejuni LOS and gangliosides, and cross-reactive serum antibody precipitate the majority of GBS cases in Bangladesh.

Characterization of Unique Modification of Flagellar Rod Protein FlgG by Campylobacter jejuni Lipid A Phosphoethanolamine Transferase, Linking Bacterial Locomotion and Antimicrobial Peptide Resistance

Gram-negative bacteria assemble complex surface structures that interface with the surrounding environment and are involved in pathogenesis. Recent work in Campylobacter jejuni identified a gene encoding a novel phosphoethanolamine (pEtN) transferase Cj0256, renamed EptC, that serves a dual role in modifying the flagellar rod protein, FlgG, and the lipid A domain of C. jejuni lipooligosaccharide with a pEtN residue. In this work, we characterize the unique post-translational pEtN modification of FlgG using collision-induced and electron transfer dissociation mass spectrometry, as well as a genetic approach using site-directed mutagenesis to determine the site of modification. Specifically, we show that FlgG is modified with pEtN at a single site (Thr(75)) by EptC and demonstrate enzyme specificity by showing that EptC is unable to modify other amino acids (e.g. serine and tyrosine). Using Campylobacter strains expressing site-directed FlgG mutants, we also show that defects in motility arise directly from the loss of pEtN modification of FlgG. Interestingly, alignments of FlgG from most epsilon proteobacteria reveal a conserved site of modification. Characterization of EptC and its enzymatic targets expands on the increasingly important field of prokaryotic post-translational modification of bacterial surface structures and the unidentified role they may play in pathogenesis.

Glycoconjugates Play a Key Role in Campylobacter jejuni Infection: Interactions between Host and Pathogen

Glycan based interactions between host and pathogen are critical in many bacterial and viral diseases. Glycan interactions range from initial receptor based adherence to protecting the infective agent from the host’s immune response through molecular mimicry. Campylobacter jejuni is an ideal model for studying the role of glycans in host–pathogen interactions, as well as the role of bacterial surface glycoconjugates in infection. Using glycan array analysis, C. jejuni has been shown to interact with a wide range of host glycoconjugates. Mannose and sialic acid residues appear to play a role in initial interactions between host and pathogen following environmental exposure, whereas fucose and galactose based interactions are likely to be required for prolonged colonization. Other studies have highlighted potential decoy receptor type interactions between host’s intestinal mucins and C. jejuni, demonstrating the importance of host glycoproteins as defense against C. jejuni infection as well as the role for glycoconjugates found in human breast milk in protection of breast feeding infants from infection with C. jejuni. C. jejuni can produce N- and O-linked glycoproteins, capsular polysaccharide (CPS) and/or lipooligosaccharide (LOS) which results in C. jejuni presenting its own diverse sugar coated displays on the cell surface. Bacterial glycans play an important and versatile role in infection and disease. Of these, the best understood is the molecular mimicry of human gangliosides presented by C. jejuni’s LOS and its link to the onset of autoimmune neuropathies such as the Guillain Barrè syndrome (GBS). However, the role of glycoconjugates presented by C. jejuni extends beyond expression of sialylated ganglioside structures involved in initiation of GBS. Expression of surface glycans by C. jejuni may also relate to the ability of this organism to interact with the glycoproteins for initial host–pathogen interactions and continued infectivity.

Guillain‐Barré Syndrome Animal Model: The First Proof of Molecular Mimicry in Human Autoimmune Disorder

Molecular mimicry between self and microbial components has been proposed as the pathogenic mechanism of autoimmune diseases, and this hypothesis is proven in Guillain-Barré syndrome. Guillain-Barré syndrome, the most frequent cause of acute neuromuscular paralysis, sometimes occurs after Campylobacter jejuni enteritis. Gangliosides are predominantly cell-surface glycolipids highly expressed in nervous tissue, whilst lipo-oligosaccharides are major components of the Gram-negative bacterium C. jejuni outer membrane. IgG autoantibodies to GM1 ganglioside were found in the sera from patients with Guillain-Barré syndrome. Molecular mimicry was demonstrated between GM1 and lipo-oligosaccharide of C. jejuni isolated from the patients. Disease models by sensitization of rabbits with GM1 and C. jejuni lipo-oligosaccharide were established. Guillain-Barré syndrome provided the first verification that an autoimmune disease is triggered by molecular mimicry. Its disease models are helpful to further understand the molecular pathogenesis as well as to develop new treatments in Guillain-Barré syndrome.

TLR4-Mediated Sensing of Campylobacter jejuni by Dendritic Cells Is Determined by Sialylation

In Guillain-Barré syndrome (GBS), ganglioside mimicry of Campylobacter jejuni lipo-oligosaccharide (LOS) drives the production of cross-reactive Abs to peripheral nerve gangliosides. We determined whether sialic acid residues in C. jejuni LOS modulate dendritic cell (DC) activation and subsequent B cell proliferation as a possible mechanism for the aberrant humoral immune response in GBS. Highly purified sialylated LOS of C. jejuni isolates from three GBS patients induced human DC maturation and secretion of inflammatory cytokines that were inhibited by anti-TLR4 neutralizing Abs. The extent of TLR4 signaling and DC activation was greater with LOS of the wild type isolates than with nonsialylated LOS of the corresponding sialyltransferase gene knockout (cst-II mutant) strains, indicating that sialylation boosts the DC response to C. jejuni LOS. Supernatants of LOS-activated DCs induced B cell proliferation after cross-linking of surface Igs in the absence of T cells. Lower B cell proliferation indices were found with DC supernatants after DC stimulation with cst-II mutant or neuraminidase desialylated LOS. This study showed that sialylation of C. jejuni LOS enhances human DC activation and subsequent B cell proliferation, which may contribute to the development of cross-reactive anti-ganglioside Abs found in GBS patients following C. jejuni infection.

Characteristics of Lipo-Oligosaccharide Loci of Campylobacter jejuni Isolates Associated with Guillain-Barré Syndrome from Hebei, China

Ganglioside mimicry by C.jejuni lipo-oligosaccharides (LOS) could induce the production of autoantibodies against gangliosides and the development of Guillain-Barré syndrome (GBS). The LOS biosynthesis region exhibits significant variation with different strains. Using PCR amplifications of genes from published LOS loci and sequencing the LOS biosynthesis loci, the eight GBS-associated C. jejuni strains from HeBei could be classified into four classes. The expression of sialylated LOS structures (class A) or non-sialylated LOS structures(class F, H and P) in the C. jejuni LOS is considered to be two different factors for the induction of GBS.

Effects of SequentialCampylobacter jejuni81-176 Lipooligosaccharide Core Truncations on Biofilm Formation, Stress Survival, and Pathogenesis

Campylobacter jejuni is a highly prevalent human pathogen for which pathogenic and stress survival strategies remain relatively poorly understood. We previously found that a C. jejuni strain 81-176 mutant defective for key virulence and stress survival attributes was also hyper-biofilm and hyperreactive to the UV fluorescent dye calcofluor white (CFW). We hypothesized that screening for CFW hyperreactive mutants would identify additional genes required for C. jejuni pathogenesis properties. Surprisingly, two such mutants harbored lesions in lipooligosaccharide (LOS) genes (waaF and lgtF), indicating a complete loss of the LOS outer core region. We utilized this as an opportunity to explore the role of each LOS core-specific moiety in the pathogenesis and stress survival of this strain and thus also constructed DeltagalT and DeltacstII mutants with more minor LOS truncations. Interestingly, we found that mutants lacking the LOS outer core (DeltawaaF and DeltalgtF but not DeltagalT or DeltacstII mutants) exhibited enhanced biofilm formation. The presence of the complete outer core was also necessary for resistance to complement-mediated killing. In contrast, any LOS truncation, even that of the terminal sialic acid (DeltacstII), resulted in diminished resistance to polymyxin B. The cathelicidin LL-37 was found to be active against C. jejuni, with the LOS mutants exhibiting modest but tiled alterations in LL-37 sensitivity. The DeltawaaF mutant but not the other LOS mutant strains also exhibited a defect in intraepithelial cell survival, an aspect of C. jejuni pathogenesis that has only recently begun to be clarified. Finally, using a mouse competition model, we now provide the first direct evidence for the importance of the C. jejuni LOS in host colonization. Collectively, this study has uncovered novel roles for the C. jejuni LOS, highlights the dynamic nature of the C. jejuni cell envelope, and provides insight into the contribution of specific LOS core moieties to stress survival and pathogenesis.

BAFF aids generation of IgG anti-ganglioside antibodies in response to Campylobacter jejuni lipo-oligosaccharide

Ganglioside mimicry of Campylobacter jejuni lipo-oligosaccharide (LOS) can induce the production of IgG anti-ganglioside antibodies, but the generation mechanism has yet to be clarified. B-cell activating factor belonging to the TNF family (BAFF) helped murine B cells produce anti-ganglioside antibodies against C. jejuni LOS. In splenocyte culture, however, anti-ganglioside antibodies were produced in the presence of a soluble transmembrane activator and calcium-modulating and cyclophilin ligand interactor immunoadhesin (TACI-Ig), a receptor for BAFF. TACI-Ig adenoviral vectors failed to decrease production of anti-ganglioside antibodies in mice sensitized with C. jejuni LOS and did not alter IgG subclasses, evidence that BAFF aids but is not essential for the generation of IgG anti-ganglioside antibodies in response to C. jejuni LOS.