Structural and Genetic Basis for the Serological Differentiation of Pasteurella multocida Heddleston Serotypes 2 and 5

Abstract

Pasteurella multocida is classified into 16 serotypes according to the Heddleston typing scheme. As part of a comprehensive study to define the structural and genetic basis of this scheme, we have determined the structure of the lipopolysaccharide (LPS) produced by P. multocida strains M1404 (B:2) and P1702 (E:5), the type strains for serotypes 2 and 5, respectively. The only difference between the LPS structures made by these two strains was the absence of a phosphoethanolamine (PEtn) moiety at the 3 position of the second heptose (Hep II) in M1404. Analysis of the lpt-3 gene, required for the addition of this PEtn residue, revealed that the gene was intact in P1702 but contained a nonsense mutation in M1404. Expression of an intact copy of lpt-3 in M1404 resulted in the attachment of a PEtn residue to the 3 position of the Hep II residue, generating an LPS structure identical to that produced by P1702. We identified and characterized each of the glycosyltransferase genes required for assembly of the serotype 2 and 5 LPS outer core. Monoclonal antibodies raised against serotype 2 LPS recognized the serotype 2/5-specific outer core LPS structure, but recognition of this structure was inhibited by the PEtn residue on Hep II. These data indicate that the serological classification of strains into Heddleston serotypes 2 and 5 is dependent on the presence or absence of PEtn on Hep II.

Development of a Rapid Multiplex PCR Assay To Genotype Pasteurella multocida Strains by Use of the Lipopolysaccharide Outer Core Biosynthesis Locus

“…We have shown previously that the 16 unique LPS outer core structures produced by the P. multocida Heddleston type strains are PM993 H8 2002 Duck PM995 H3 2002 Chicken PM1075 H16 2004 Chicken PM1098 H15 2004 Unknown PM1099 H10 2004 Unknown PM1103 H10 2004 Unknown PM1113 NT 2004 Avian PM1120 NT 2005 Chicken PM1124 H1, H4, H12 2005 Unknown PM1128 H10 2005 Bovine PM1132 H1, H3, H4, H10, H14 2005 Pig PM1153 H1, H3, H7 2005 Avian PM1165 H1 2006 Duck PM1193 H3 2006 Duck PM1205 H1 2007 Emu PM1258 NT 2010 Chicken PM1268 NT 2010 Chicken PM1300 H4 2009 Turkey PM1304 H1 2009 Chicken PM1315 H1 2009 Chicken PM1316 H4 2009 Unknown PM1317 H3 2009 Unknown PM1320 H10, H13, H14 2010 Chicken PM1369 H1 2010 Chicken PM1396 H1, H3 2010 Unknown generated from only eight distinct genetic loci, which we have named L1 to L8 ( Fig. 1) (9)(10)(11)(12)(13)(14)(15)20)<...>…”

“…The sugar composition of the LPS from selected strains was determined by mass spectrometry as previously described (22). The predicted LPS structures produced by the P. multocida isolates were determined using MS compositional analysis and comparison with the known compositions and structures of the 16 Heddleston serovar type strains (9)(10)(11)(12)(13)(14)(15)(16).…”

“…We have recently carried out a comprehensive analysis of the LPS structures expressed by the 16 Heddleston type strains and identified the genes required for LPS assembly in each strain (9)(10)(11)(12)(13)(14)(15)(16). These combined analyses showed that the LPS produced by all strains consisted of a highly conserved inner core and a variable outer core and revealed that each of the 16 Heddleston type strains expresses structurally distinct LPS.…”

Pasteurella multocida Heddleston Serovar 3 and 4 Strains Share a Common Lipopolysaccharide Biosynthesis Locus but Display both Inter- and Intrastrain Lipopolysaccharide Heterogeneity

“…The LPS structures produced by P. multocida type strains representing the Heddleston serovars 1, 2, 5, 8, 9, 13, and 14 have been determined, as well as the structure of the LPS expressed by the genome-sequenced serovar 3 strain Pm70 (11)(12)(13)(14)(15)(16)(17). Some P. multocida strains can produce two LPS inner core glycoforms simultaneously, termed glycoform A and glycoform B.…”

Phenotypic and Genotypic Characterisation of Pasteurella Multocida Strains Isolated from Pigs in Poland

“…For capsular serotyping, Carter (6) used indirect haemagglutination test. Heddleston (22) used the gel diffusion precipitation test and distinguished 16 serotypes. Townsend et al (36) developed PCR for capsular typing of Pm strains.…”