Genomic and antigenic characterization of monomeric autotransporters of Haemophilus parasuis: an ongoing process of reductive evolution

Abstract

The genome of the highly pathogenic Haemophilus parasuis Nagasaki strain (serovar 5) was sequenced to 99 % completion. A genomic comparison with two other pathogenic serovar 5 H. parasuis strains identified six genes per genome (bmaA1-bmaA6) encoding b-barrel monomeric autotransporters, bmaA2 and bmaA3 being pseudogenes in at least one strain. The remaining encoded proteins were predicted to belong to the subtilisin (BmaA1 and BmaA4) and cysteine (BmaA5 and BmaA6) protease families. Allelic polymorphism was detected in other H. parasuis strains by comparative genomic hybridization using microarrays. Recombination events were observed, some of them leading to gene disruption in one of the three strains, although synteny around bmaA genes was conserved. These results suggest that bmaA genes are undergoing a process of reductive evolution. To evaluate their use as potential vaccine antigens, the products of the passenger domains of bmaA1, bmaA4, bmaA5 and bmaA6 were produced in Escherichia coli as recombinant proteins. They were detected by immunoblotting using sera of colostrum-deprived piglets recovering from a sublethal infection with H. parasuis (Nagasaki). The existence of specific antibodies after infection with H. parasuis also demonstrated in vivo expression. Using proteomics, only BmaA6 was detected in the in vitro-grown Nagasaki strain. Interestingly, the translocator domain was found in the outer membrane, while the passenger domain was located in supernatants. These results indicate that BmaA proteins could be considered as immunogen candidates to improve H. parasuis vaccines. However, their capacity to confer protective immunity needs to be studied further.

Development and application of an antibody detection ELISA for Haemophilus parasuis based on a monomeric autotransporter passenger domain

“…The antibodies induced by H. parasuis commercial killed vaccine and infection were both detectable via the rApd ELISA (Fig. 3), which is consistent with the findings that autotransporters of H. parasuis were able to be expressed in in vitro culture as well as in in vivo infection of pigs [15]. In addition, Apd was present in 15 serotype reference strains and five non-typeable strains tested; thus, the rApd ELISA exhibited the potential to be used for antibody detection to determine vaccine immunity and serological diagnosis of Glässer’s disease regardless of the serotype of strains.…”

“…Autotransporters consist of a cleavable N-terminal signal peptide, a functional passenger domain exposed to the outer membrane or released into the external environment, and a C-terminal region involved in the formation of a transmembrane pore [15]. In the present study, three autotransporter passenger domains of H. parasuis were determined to represent candidate antigens for H. parasuis indirect ELISA, with a monomeric a utotransporter p assenger d omain (Apd) being selected as the optimal antigen.…”

The passenger‐associated transport repeat promotes virulence factor secretion efficiency and delineates a distinct autotransporter subtype

“… The importance of the PATR within the AT family. A. Alignment of the PATR sites of IcsA and five other ATs : EprS ( Q9HY75 ), ShdA ( Q9XCJ4 ), subtilisin‐type serine protease PrtS ( P 09489) from S erratia marcescens (Shikata et al ., ), subtilisin‐type serine protease BmaA1 ( H6T4K9 ) of H aemophilus parasuis (Pina‐Pedrero et al ., ) and NalP ( Q8GKS5 ). Accessions are U ni P rot KB IDs .…”

“…An analysis of the usage of domains and motifs in all ATs within the UniProt Knowledgebase was also conducted by first identifying unique ATs using InterPro (Hunter et al, 2012) AT β-barrel identifiers IPR005546 and/or IPR006315 (7659 proteins). Figure 4B shows the different virulence A. Alignment of the PATR sites of IcsA and five other ATs: EprS (Q9HY75), ShdA (Q9XCJ4), subtilisin-type serine protease PrtS (P09489) from Serratia marcescens (Shikata et al, 1992), subtilisin-type serine protease BmaA1 (H6T4K9) of Haemophilus parasuis (Pina-Pedrero et al, 2012) and NalP (Q8GKS5). Accessions are UniProtKB IDs.…”

Genome comparison of three serovar 5 pathogenic strains of Haemophilus parasuis: insights into an evolving swine pathogen

“…Whole orthologous comparisons between H. parasuis and other micro-organisms within the Pasteurellaceae family showed that the swine pathogens H. parasuis and Actinobacillus pleuropneumoniae occupy close positions in this phylogenetic analysis, suggesting that the two species are probably derived from a recent common ancestor (Xu et al , 2011). In addition, a comparative analysis of monomeric autotransporters from three serovar 5 invasive strains (SH0165, 29755 and Nagasaki) suggested a reductive evolutionary state for these genes (Pina-Pedrero et al , 2012). More recently, a genome comparison of a H. parasuis serovar 12 strain (ZJ0906) with the serovar 5 strains SH0165 and 29755 showed that gene rearrangements are frequent in this species (Li et al , 2013).…”