Structural Diversity among Edwardsiellaceae Core Oligosaccharides.

The Edwardsiella genus presents five different pathogenic species: Edwardsiella tarda, E. anguillarum, E. piscicida, E. hoshinae and E. ictaluri. These species cause infections mainly in fish, but they can also infect reptiles, birds or humans. Lipopolysaccharide (endotoxin) plays an important role in the pathogenesis of these bacteria. For the first time, the chemical structure and genomics of the lipopolysaccharide (LPS) core oligosaccharides of E. piscicida, E. anguillarum, E. hoshinae and E. ictaluri were studied. The complete gene assignments for all core biosynthesis gene functions were acquired. The structure of core oligosaccharides was investigated by ¹H and 13C nuclear magnetic resonance (NMR) spectroscopy. The structures of E. piscicida and E. anguillarum core oligosaccharides show the presence of →3,4)-L-glycero-α-D-manno-Hepp, two terminal ß-D-Glcp, →2,3,7)-L-glycero-α-D-manno-Hepp, →7)-L-glycero-α-D-manno-Hepp, terminal α-D-GlcpN, two →4)-α-D-GalpA, → 3)-α-D-GlcpNAc, terminal ß-D-Galp and →5-substituted Kdo. E. hoshinare core oligosaccharide shows only one terminal ß-D-Glcp, and instead of terminal ß-D-Galp a terminal α-D-GlcpNAc. E. ictaluri core oligosaccharide shows only one terminal ß-D-Glcp, one →4)-α-D-GalpA and do not have terminal α-D-GlcpN (see complementary figure).

Characteristics of Environmental Klebsiella pneumoniae and Klebsiella oxytoca Bacteriophages and Their Therapeutic Applications.

In recent years, multidrug-resistant (MDR) strains of Klebsiella pneumoniae have spread globally, being responsible for the occurrence and severity of nosocomial infections. The NDM-1-kp, VIM-1 carbapenemase-producing isolates as well as extended-spectrum beta lactamase-producing (ESBL) isolates along with Klebsiella oxytoca strains have become emerging pathogens. Due to the growing problem of antibiotic resistance, bacteriophage therapy may be a potential alternative to combat such multidrug-resistant Klebsiella strains. Here, we present the results of a long-term study on the isolation and biology of bacteriophages active against K. pneumoniae, as well as K. oxytoca strains. We evaluated biological properties, morphology, host specificity, lytic spectrum and sensitivity of these phages to chemical agents along with their life cycle parameters such as adsorption, latent period, and burst size. Phages designated by us, vB_KpnM-52N (Kpn52N) and VB_KpnM-53N (Kpn53N), demonstrated relatively broad lytic spectra among tested Klebsiella strains, high burst size, adsorption rates and stability, which makes them promising candidates for therapeutic purposes. We also examined selected Klebsiella phages from our historical collection. Notably, one phage isolated nearly 60 years ago was successfully used in purulent cerebrospinal meningitis in a new-born and has maintained lytic activity to this day. Genomic sequences of selected phages were determined and analyzed. The phages of the sequenced genomes belong to the Slopekvirus and Jiaodavirus genus, a group of phages related to T4 at the family level. They share several features of T4 making them suitable for antibacterial therapies: the obligatorily lytic lifestyle, a lack of homologs of known virulence or antibiotic resistance genes, and a battery of enzymes degrading host DNA at infection.

The Mutation in wbaP cps Gene Cluster Selected by Phage-Borne Depolymerase Abolishes Capsule Production and Diminishes the Virulence of Klebsiella pneumoniae.

Klebsiella pneumoniae is considered one of the most critical multidrug-resistant pathogens and urgently requires new therapeutic strategies. Capsular polysaccharides (CPS), lipopolysaccharides (LPS), and exopolysaccharides (EPS) are the major virulence factors protecting K. pneumoniae against the immune response and thus may be targeted by phage-based therapeutics such as polysaccharides-degrading enzymes. Since the emergence of resistance to antibacterials is generally considered undesirable, in this study, the genetic and phenotypic characteristics of resistance to the phage-borne CPS-degrading depolymerase and its effect on K. pneumoniae virulence were investigated. The K63 serotype targeting depolymerase (KP36gp50) derived from Klebsiella siphovirus KP36 was used as the selective agent during the treatment of K. pneumoniae 486 biofilm. Genome-driven examination combined with the surface polysaccharide structural analysis of resistant mutant showed the point mutation and frameshift in the wbaP gene located within the cps gene cluster, resulting in the loss of the capsule. The sharp decline in the yield of CPS was accompanied by the production of a larger amount of smooth LPS. The modification of the surface polysaccharide layers did not affect bacterial fitness nor the insensitivity to serum complement; however, it made bacteria more prone to phagocytosis combined with the higher adherence and internalization to human lung epithelial cells. In that context, it was showed that the emerging resistance to the antivirulence agent (phage-borne capsule depolymerase) results in beneficial consequences, i.e., the sensitization to the innate immune response.

Identification of the Primary Structure of Selenium-Containing Polysaccharides Selectively Inhibiting T-Cell Proliferation.

We previously described the biosynthesis, isolation, and immunosuppressive activity of the selenium-containing polysaccharide fraction isolated from the mycelial culture of Lentinula edodes. Structural studies have shown that the fraction was a protein-containing mixture of high molar mass polysaccharides α- and ß-glucans. However, which of the components of the complex fraction is responsible for the immunosuppressive activity non-typical for polysaccharides of fungal origin has not been explained. In the current study, we defined four-polysaccharide components of the Se-containing polysaccharide fraction determined their primary structure and examined the effect on T- and B-cell proliferation. The isolated Se-polysaccharides, α-1,4-glucan (Mw 2.25 × 106 g/mol), unbranched ß-1,6-d-glucan, unbranched ß-1,3-d-glucan and ß-1,3-branched ß-1,6-d-glucan (Mw 1.10 × 105 g/mol), are not typical as components of the cell wall of L. edodes. All are biologically active, but the inhibitory effect of the isolated polysaccharides on lymphocyte proliferation was weaker, though more selective than that of the crude fraction.

Structural analysis of Edwardsiella tarda PCM 1155 O-polysaccharide revealed the presence of unique ß-L-RhapNAc3NAc derivative.

The chemical structure of the lipopolysaccharide O-polysaccharide repeating unit of Edwardsiella tarda strain PCM 1155 was studied for the first time. The complete structure of repeating unit was investigated by chemical methods, 1H and 13C nuclear magnetic resonance (NMR) spectroscopy, and matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The rarely occurring monosaccharide, 2,3-diacetamido-2,3,6-trideoxy-l-mannose (L-RhapNAc3NAc) was identified. The following structure was established.

Isolation and Characterization of Phages Active against Paenibacillus larvae Causing American Foulbrood in Honeybees in Poland.

The aim of this study was the isolation and characterization, including the phage effect on honeybees in laboratory conditions, of phages active against Paenibacillus larvae, the causative agent of American Foulbrood-a highly infective and easily spreading disease occurring in honeybee larva, and subsequently the development of a preparation to prevent and treat this dangerous disease. From the tested material (over 2500 samples) 35 Paenibacillus spp. strains were obtained and used to search for phages. Five phages specific to Paenibacillus were isolated and characterized (ultrastructure, morphology, biological properties, storage stability, and genome sequence). The characteristics were performed to obtain knowledge of their lytic potential and compose the final phage cocktail with high antibacterial potential and intended use of future field application. Preliminary safety studies have also been carried out on healthy bees, which suggest that the phage preparation administered is harmless.

The Impact of Insertion Sequences on O-Serotype Phenotype and Its O-Locus-Based Prediction in Klebsiella pneumoniae O2 and O1.

Klebsiella pneumoniae is a nosocomial pathogen, pointed out by the World Helth Organisation (WHO) as "critical" regarding the highly limited options of treatment. Lipopolysaccharide (LPS, O-antigen) and capsular polysaccharide (K-antigen) are its virulence factors and surface antigens, determining O- and K-serotypes and encoded by O- or K-loci. They are promising targets for antibody-based therapies (vaccines and passive immunization) as an alternative to antibiotics. To make such immunotherapy effective, knowledge about O/K-antigen structures, drift, and distribution among clinical isolates is needed. At present, the structural analysis of O-antigens is efficiently supported by bioinformatics. O- and K-loci-based genotyping by polymerase chain reaction (PCR) or whole genome sequencing WGS has been proposed as a diagnostic tool, including the Kaptive tool available in the public domain. We analyzed discrepancies for O2 serotyping between Kaptive-based predictions (O2 variant 2 serotype) and the actual phenotype (O2 variant 1) for two K. pneumoniae clinical isolates. Identified length discrepancies from the reference O-locus resulted from insertion sequences (ISs) within rfb regions of the O-loci. In silico analysis of 8130 O1 and O2 genomes available in public databases indicated a broader distribution of ISs in rfbs that may influence the actual O-antigen structure. Our results show that current high-throughput genotyping algorithms need to be further refined to consider the effects of ISs on the LPS O-serotype.

Lipopolysaccharide-Linked Enterobacterial Common Antigen (ECALPS) Occurs in Rough Strains of Escherichia coli R1, R2, and R4.

Enterobacterial common antigen (ECA) is a conserved surface antigen characteristic for Enterobacteriaceae. It is consisting of trisaccharide repeating unit, →3)-α-d-Fucp4NAc-(1→4)-ß-d-ManpNAcA-(1→4)-α-d-GlcpNAc-(1→, where prevailing forms include ECA linked to phosphatidylglycerol (ECAPG) and cyclic ECA (ECACYC). Lipopolysaccharide (LPS)-associated form (ECALPS) has been proved to date only for rough Shigella sonnei phase II. Depending on the structure organization, ECA constitutes surface antigen (ECAPG and ECALPS) or maintains the outer membrane permeability barrier (ECACYC). The existence of LPS was hypothesized in the 1960-80s on the basis of serological observations. Only a few Escherichia coli strains (i.e., R1, R2, R3, R4, and K-12) have led to the generation of anti-ECA antibodies upon immunization, excluding ECAPG as an immunogen and conjecturing ECALPS as the only immunogenic form. Here, we presented a structural survey of ECALPS in E. coli R1, R2, R3, and R4 to correlate previous serological observations with the presence of ECALPS. The low yields of ECALPS were identified in the R1, R2, and R4 strains, where ECA occupied outer core residues of LPS that used to be substituted by O-specific polysaccharide in the case of smooth LPS. Previously published observations and hypotheses regarding the immunogenicity and biosynthesis of ECALPS were discussed and correlated with presented herein structural data.

De­O­acylated lipooligosaccharide of E. coli B reduces the number of metastatic foci via downregulation of myeloid cell activity.

Lipopolysaccharides are the main surface antigens and virulence factors of gram­negative bacteria. Removal of four ester­bound fatty acid residues from hexaacyl lipid A of Escherichia coli lipooligosaccharide (LOS) resulted in the de­O­acylated derivative E. coli LOS­OH (LOS­OH). This procedure caused a significant reduction in the toxicity of this compound compared to the native molecule. We investigated the effect of such a structural LOS modification on its biological activity using in vitro assays with monocytic cells of the RAW264.7 line, dendritic cells of the JAWS II line, bone marrow­derived dendritic cells (BM­DCs), and spleen cells. Furthermore, in in vivo experiments with a melanoma B16 metastasis model, the anti­metastatic activity of the compounds and spleen cell reactivity mediated by them representing a systemic response were analyzed. The results revealed that LOS­OH demonstrated weaker ability than LOS to stimulate and polarize an immune response both in vitro and in vivo. It induced lower cytokine production by cells of myeloid lines. Multiple applications of LOS­OH into mice injected intravenously with B16 cells significantly (P<0.05; P<0.01) reduced the number of metastatic foci in the lungs, presumably via silencing of myeloid cell reactivity as well as the inability to stimulate lymphoid cells both directly and indirectly. These findings suggest that LOS­OH maintained in the body of metastasis­bearing mice appears to modulate or downregulate the innate response, leading to the inability of blood myeloid cells to support the migration of melanoma cells to lung tissue.

The New Structure of Core Oligosaccharide Presented by Proteus penneri 40A and 41 Lipopolysaccharides.

The new type of core oligosaccharide in Proteus penneri 40A and 41 lipopolysaccharides has been investigated by ¹H and 13C NMR spectroscopy, electrospray ionization mass spectrometry and chemical methods. Core oligosaccharides of both strains were chosen for structural analysis based on the reactivity of LPSs with serum against P. penneri 40A core oligosaccharide-diphtheria toxoid conjugate. Structural analyses revealed that P. penneri 40A and 41 LPSs possess an identical core oligosaccharide.

Discovery of monoclonal antibodies cross-reactive to novel subserotypes of K. pneumoniae O3.

Klebsiella pneumoniae is responsible for nosocomial infections causing significant morbidity and mortality. Treatment of newly emerging multi-drug resistant strains is hampered due to severely limited antibiotic choices. Passive immunization targeting LPS O-antigens has been proposed as an alternative therapeutic option, given the limited variability of Klebsiella O-antigens. Here we report that the O3 serogroup, previously considered to have uniform O-antigen built of mannan, represents three different subtypes differing in the number of mannose residues within the O-antigen repeating units. Genetic analysis of the genes encoding mannose polymerization revealed differences that underline the observed structural alterations. The O3 variants represent antigenically different types based on the different reactivity pattern of murine monoclonal antibodies raised against a K. pneumoniae O3 strain. Typing of a collection of K. pneumoniae O3 clinical isolates showed that strains expressing the novel O3b antigen, the tri-mannose form, were more prevalent than those having the penta-mannose form, traditionally called O3, while the tetra-mannose variant, termed here O3a, seems to be rare. A monoclonal antibody cross-reacting with all three O3 sub-serogroups was also selected and shown to bind to the surface of various K. pneumoniae strains expressing different O3 subtypes and capsular antigens.

The Complete Structure of the Core Oligosaccharide from Edwardsiella tarda EIB 202 Lipopolysaccharide.

The chemical structure and genomics of the lipopolysaccharide (LPS) core oligosaccharide of pathogenic Edwardsiella tarda strain EIB 202 were studied for the first time. The complete gene assignment for all LPS core biosynthesis gene functions was acquired. The complete structure of core oligosaccharide was investigated by ¹H and 13C nuclear magnetic resonance (NMR) spectroscopy, electrospray ionization mass spectrometry MSn, and matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry. The following structure of the undecasaccharide was established: The heterogeneous appearance of the core oligosaccharide structure was due to the partial lack of ß-d-Galp and the replacement of α-d-GlcpNAcGly by α-d-GlcpNGly. The glycine location was identified by mass spectrometry.

Identification of d-Galactan-III As Part of the Lipopolysaccharide of Klebsiella pneumoniae Serotype O1.

Klebsiella pneumoniae is a Gram-negative, ubiquitous bacterium capable of causing severe nosocomial infections in individuals with impaired immune system. Emerging multi-drug resistant strains of this species and particularly carbapenem-resistant strains pose an urgent threat to public health. The lipopolysaccharide (LPS) O-antigen is the main surface antigen. It contributes to the virulence of this species and determines the O-serotype of K. pneumoniae isolates. Among the nine main O-serotypes of K. pneumoniae, O1-and O2-type pathogens are causative agents of over 50% of all infections. Serotype O1, the most common O-serotype, expresses complex LPS consisting of d-galactan-I (a polymer built of → 3)-ß-d-Galf-(1 → 3)-α-d-Galp-(1 → repeating units) capped by d-galactan-II (built of [ → 3)-α-d-Galp-(1 → 3)-ß-d-Galp-(1 →] repeating units). Galactan-I is present as the sole polymer in O2 serotype. Recently, in case of serotype O2, conversion of galactan-I to galactan-III (→ 3)-ß-d-Galf-(1 → 3)-[α-d-Galp-(1 → 4)]-α-d-Galp-(1 →) was reported. Substitution of → 3)-α-d-Galp by a branching terminal α-d-Galp was dependent on the presence of the gmlABC operon and had a major impact on the antigenicity of the galactan polymer. Genetic analysis indicated that 40% of the O1 clinical isolates also carry the gmlABC locus; therefore we aimed to characterize the corresponding phenotype of LPS O-antigens. The presence of galactan-III among O1 strains was proven using galactan-III-specific monoclonal antibodies and confirmed by structural analyses performed using sugar and methylation analysis as well as classical and high-resolution magic angle spinning NMR spectroscopy. By using an isogenic mutant pair, we demonstrated that galactan-III expression was dependent on the presence of glycosyltransferases encoded by gmlABC, as was shown previously for the O2 serotype. Furthermore, the galactan-II structures in O1gml+ strains remained unaffected corroborating no functional interactions between the biosynthesis of galactan-III and galactan-II polymers.

Means to Facilitate the Overcoming of Gastric Juice Barrier by a Therapeutic Staphylococcal Bacteriophage A5/80.

In this article we compare the efficacy of different pharmacological agents (ranitidine, and omeprazole) to support phage transit from stomach to distal portions of the gastrointestinal tract in rats. We show that a temporal modification of environment in the animal stomach may protect Twort-like therapeutic antistaphylococcal phage A5/80 (from bacteriophage collection of the Hirszfeld Institute of Immunology and Experimental Therapy PAS in Wroclaw, Poland) from the inactivation by gastric juice effectively enough to enable a significant fraction of orally administered A5/80 to pass to the intestine. Interestingly, we found that yogurt may be a relatively strong in enhancing phage transit. Given the immunomodulating activities of phages our data may suggest that phages and yogurt can act synergistically in mediating their probiotic activities and enhancing the effectiveness of oral phage therapy. We also demonstrate that orally applied phages of similar size, morphology, and sensitivity to acidic environment may differ in their translocation into the bloodstream. This was evident in mice in which a therapeutic staphylococcal phage A5/80 reached the blood upon oral administration combined with antacid agent whilst T4 phage was not detected even when applied in 103 times higher dose. Our findings also suggest that phage penetration from digestive tract to the blood may be species-specific.

Structure-Activity Relationship of Plesiomonas shigelloides Lipid A to the Production of TNF-α, IL-1ß, and IL-6 by Human and Murine Macrophages.

Plesiomonas shigelloides is a Gram-negative bacterium that is associated with diarrheal disease in humans. Lipopolysaccharide (LPS) is the main surface antigen and virulence factor of this bacterium. The lipid A (LA) moiety of LPS is the main region recognized by target cells of immune system. Here, we evaluated the biological activities of P. shigelloides LA for their abilities to induce the productions of proinflammatory cytokines (TNF-α, IL-1ß, and IL-6) by human and murine macrophages [THP-1 macrophages and immortalized murine bone marrow-derived macrophages (iBMDM)]. Four native P. shigelloides LA preparations differing in their phosphoethanolamine (PEtn) substitution, length, number, and saturation of fatty acids were compared with Escherichia coli O55 LA. The bisphosphorylated, hexaacylated, and asymmetric forms of the P. shigelloides and E. coli LA molecules had similar activities in human and murine macrophages, indicating that shortening of the acyl chains in P. shigelloides LA had no effect on its in vitro activities. The PEtn decoration also had no impact on the interaction with the toll-like receptor 4/MD-2 receptor complex. The heptaacylated form of P. shigelloides LA decorated with 16:0 exhibited strong effect on proinflammatory activity, significantly decreasing the levels of all tested cytokines in both murine and human macrophages. Our results revealed that despite the presence of shorter acyl chains and an unsaturated acyl residue (16:1), the bisphosphorylated, hexaacylated, and asymmetric forms of P. shigelloides LA represent highly immunostimulatory structures.

The O-antigen of Plesiomonas shigelloides serotype O36 containing pseudaminic acid.

The structure of the repeating unit of O-antigen of Plesiomonas shigelloides serotype O36 has been investigated by 1H and 13C NMR spectroscopy, matrix-assisted laser-desorption/ionization time-of-flight mass spectrometry and chemical methods. The new structure of trisaccharide has been established: [Formula: see text] These trisaccharide O-antigen units substitute the core undecasaccharide at C-4 of the ß-D-GlcpNAc residue. The core oligosaccharide and lipid A are identical with these of the serotype O17 (PCM 2231) (Maciejewska, A., Lukasiewicz, J., Kaszowska, M., Jachymek, W., Man-Kupisinska, A.; Lugowski, C. Mar. Drugs.2013, 11 (2), 440-454; Lukasiewicz, J., Dzieciatkowska, M., Niedziela, T., Jachymek, W., Augustyniuk, A., Kenne, L., Lugowski, C. Biochemistry, 2006, 45, 10434-10447).

Oral Administration of Polymyxin B Modulates the Activity of Lipooligosaccharide E. coli B against Lung Metastases in Murine Tumor Models.

INTRODUCTION: Polymyxin B (PmB) belongs to the group of cyclic peptide antibiotics, which neutralize the activity of LPS by binding to lipid A. The aim of this study was to analyze the effect of PmB on the biological activity of lipooligosaccharide (LOS E. coli B,rough form of LPS) in vitro and in experimental metastasis models. RESULTS: Cultures of murine macrophage J774A.1 cells and murine bone marrow-derived dendritic cells (BM-DC) stimulated in vitro with LOS and supplemented with PmB demonstrated a decrease in inflammatory cytokine production (IL-6, IL-10, TNF-α) and down-regulation of CD40, CD80, CD86 and MHC class II molecule expression. Additionally, PmB suspended in drinking water was given to the C57BL/6 mice seven or five days prior to the intravenous injection of B16 or LLC cells and intraperitoneal application of LOS. This strategy of PmB administration was continued throughout the duration of the experiments (29 or 21 days). In B16 model, statistically significant decrease in the number of metastases in mice treated with PmB and LOS (p<0.01) was found on the 14th day of the experiments, whereas the most intensive changes in surface-antigen expression and ex vivo production of IL-6, IL-1ß and TNF-α by peritoneal cells were observed 7 days earlier. By contrast, antigen expression and ex vivo production of IL-6, IL-10, IFN-γ by splenocytes remained relatively high and stable. Statistically significant decrease in LLC metastases number was observed after the application of LOS (p<0.01) and in the group of mice preconditioned by PmB and subsequently treated with LOS (LOS + PmB, p<0.01). CONCLUSIONS: In conclusion, prolonged in vivo application of PmB was not able to neutralize the LOS-induced immune cell activity but its presence in the organism of treated mice was important in modulation of the LOS-mediated response against the development of metastases.

Both clades of the epidemic KPC-producing Klebsiella pneumoniae clone ST258 share a modified galactan O-antigen type.

Klebsiella pneumoniae ST258 is a globally disseminated, extremely drug resistant, nosocomial clone with limited treatment options. We show that the vast majority of ST258 isolates express modified d-galactan-I lipopolysaccharide O-antigen, termed hereinafter as D-galactan-III. The genetic determinant required for galactan-III synthesis was identified as a distinct operon adjacent to the rfb (wb) locus encoding D-galactan-I synthesis. The three genes within the operon encode predicted glycosyltransferases. Testing an isogenic transformant pair revealed that expression of D-galactan-III, in comparison to D-galactan-I, conferred improved survival in the presence of human serum. Eighty-three percent of the more than 200 ST258 draft genome sequences currently available carries the corresponding operon and hence these isolates are predicted to express galactan-III antigens. A D-galactan-III specific monoclonal antibody (mAb) was shown to bind to extracted LPS from a panel of ST258 isolates. The same mAb confirmed accessibility of galactan-III in surface staining of ST258 irrespective of the distinct capsular antigens expressed by both clades described previously. Based on these data, the galactan-III antigen may represent an attractive target for active and passive immunization approaches against K. pneumoniae ST258.

Core oligosaccharide of Escherichia coli B-the structure required for bacteriophage T4 recognition.

The structure of Escherichia coli B strain PCM 1935 core oligosaccharide has been investigated by (1)H and (13)C NMR spectroscopy, MALDI-TOF MS and ESI MS(n). It was concluded that the core oligosaccharide is a pentasaccharide with the following structure: ESI MS/MS analysis revealed that the glycine (a minor component) is linked to the →3,7)-l-α-d-Hepp-(1→ residue.

NMR study of the O-specific polysaccharide and the core oligosaccharide from the lipopolysaccharide produced by Plesiomonas shigelloides O24:H8 (strain CNCTC 92/89).

The structures of the O-specific polysacccharide and core oligosaccharide of the lipopolysaccharide from Plesiomonas shigelloides O24:H8, strain CNCTC 92/89, have been investigated by NMR spectroscopy and ESI mass spectrometry. The O-specific polysaccharide was found to be composed of a tetrasaccharide repeating unit consisting of [→3)-α-FucpNAc-(1→3)-α-GalpNAcA-(1→3)-α-QuipNAc-(1→] and of α-RhapNAc (1→4) linked to the GalpNAcA residue. An identical structure has been reported for the capsular polysaccharide of the clinical isolate of Vibrio vulnificus strain BO62316 [1]. The core oligosaccharide was composed of a decasaccharide which structure is identical with these in P. shigelloides serotype O54 [2] and serotype O37 [3].