Interestingly, the two analyzed strains of the mAb-subgroup Benidorm, 130b and Lens, cluster into
two distinct groups. This either indicates that the product of ORF 6 has probably no effect on the LPS structure of strains of the same monoclonal subgroup or that it has the same function despite low similarity. However, ORF 6 products might be involved in the establishment of a mAb-subgroup discriminating epitope. More precisely, only the mAb-subgroups LY2874455 in vivo Heysham and Knoxville react with mAb 3. This indicates a similar epitope which in turn could possibly be traced back to specific ORFs within the Sg1-specific region. However, strains of both mAb-subgroups were highly homologous regarding the whole LPS-biosynthesis with the exception of lag-1 which is present in Knoxville strains. (Figure 2B, Table 3). In addition, the
strain Camperdown 1, not reacting with mAb 3, carried a very similar LPS-biosynthesis locus as Heysham 1 and the Knoxville strains. However, it is the single ORF 6 in which Camperdown 1 clusters differently to Heysham 1. It can be assumed that the combination of ORF 6 to 9 which is exclusively found in Knoxville and Heysham strains leads to reactivity with mAb 3. Another ORF 6 as found in the genetically very similar strain Camperdown 1 could alter the LPS epitope and is thereby not recognized by mAb 3. Furthermore, the mAb 3 epitope was not influenced by O-acetylation of the legionaminic acid residue since the Knoxville strains were mAb 3/1+ and carried the lag-1 gene whereas the strain Heysham 1 is negative for both markers. Modification of legionaminic acid in transposon mutants Two additional FK506 price ORFs, ORF 8 and ORF 9, within in the highly variable region from ORF 6 to ORF 11 are most likely involved in O-antigen modification. The genetic nature of the
ORF 8 products displayed two different clusters which was comparable to the clustering of ORF 9. Both clusters share poor amino acid similarities of 31% (ORF 8) and 30.7% (ORF 9) (Table 3, Figure Morin Hydrate 2D). These differences in amino acid check details similarity were also reflected by the ORF orientation. Both ORFs were orientated into opposite directions in strains of the mAb-subgroups Knoxville, Camperdown and Heysham which form a separate cluster in both ORFs (Figure 1A). For the remaining mAb-subgroups (Philadelphia, Allentown, Benidorm, Bellingham and OLDA) the ORFs are oriented into identical directions. In silico analysis of these loci predicted a five-gene operon from ORF 8 to ORF 12 suggesting a coupled functional entity . These strains were also grouped into a single cluster. However, recent transcriptomic data obtained from strain Paris revealed a four-gene operon which lacks ORF 8 . For all strains regardless of the distance in the phylogenetic tree BLASTP predicted a methyltransferase function for ORF 8 [48, 52] and a siliac acid synthetase function (neuB family) for ORF 9 .