47 ± 0.42 5.54 (4.12-7.45) 6.23 pgaC (R)   0 ± 1.05 1(0.48-1.07)   apxIVA (T)

RTX toxin protein -3.01 ± 1.12 8.06 (3.69-17.61) 6.5 apxIVA (R)   0 ± 0.60 1 (0.65-1.52)   relA (T) GTP pyrophosphokinase -0.95 ± 0.42 2.0 (1.44-2.56) 6.30 relA (R)   0 ± 0.59 1(0.66-1.51)   lamB (T)2 Maltoporin 1.03 ± 0.39 0.49 Go6983 manufacturer (0.37-0.64) na3 lamB (R)   0 ± 0.23 1 (0.85-1.17)   1Fold change is the fold increase or decrease in the level of find more expression of a gene in the malT mutant (target sample, abbreviated as T) relative to the level of expression of the gene in the wild type (calibrator or reference sample, abbreviated as R) in BALF except for the lamB gene2 whose expression was compared in BHI to examine the effect of the malT knockout mutation on the expression of the lamB gene. 3 Not applicable. Values in the parentheses represent the range in the fold change.

Discussion Expression of maltose-regulon genes by BALF-exposed A. pleuropneumoniae CM5 After exposure of A. pleuropneumoniae CM5 to BALF for 30 minutes, a gene that appeared to be lamB homologue was shown to be up-regulated by the organism in RT-PCR DD experiments (Figure 1). We selected 30 min for incubation of the organism in BALF, as the medium conditions should remain fairly constant during this time as might be seen in the animal during early infection when there is constant replenishment of alveolar fluid. As shown in real-time PCR studies, the genes encoding intrinsic membrane transport system proteins (MalF and PAK5 MalG), maltodextrin phosphorylase (MalP), amylomaltase (MalQ), AZD8931 cell line ATP-binding cassette of the maltodextrin transporter (MalK) of the maltose regulon were also up-regulated in BALF, although some at very low levels (Table 1). Comparison of gene expression in BALF- and BHI-incubated cells by DNA

microarrays [15] showed that malF and malG were up-regulated in BALF. However, no differential expression was seen in malT, malK, malP or malQ genes. This disparate finding could be because only small quantities of these proteins are required for function, and small changes in gene expression are difficult to detect. For further study, we focused on the lamB and malT genes of the maltose regulon as LamB is a cell surface protein that lies at the host-pathogen interface and MalT is a transcriptional regulator that might control the expression of genes other than those involved in the maltose and maltodextrin transport and metabolism. malT and lamB are the components of a functional maltose regulon in A. pleuropneumoniae CM5 All of the strains of A. pleuropneumoniae sequenced so far possess homologs of the maltose regulon genes malEFG, malK-lamB-malM, malT and malPQ. As demonstrated by microarray-based comparative genomic profiling, these genes are present in the reference strains of all 15 serovars of A. pleuropneumoniae [16].