The biological role of the enigmatic C3larvinAB toxin of the honey bee pathogenic bacterium Paenibacillus larvae

Abstract

Summary Paenibacillus larvae is the causative agent of the notifiable epizootic American foulbrood, a fatal bacterial disease of honey bee larvae. The species P. larvae has been classified into four differentially virulent and prevalent genotypes (ERIC I‐IV), which also differ in their virulence factor equipment. Recently, a novel P. larvae toxin, the C3‐like C3larvin, has been described. Genome analysis now revealed that the C3larvin gene is actually a part of a toxin locus encompassing two genes encoding a binary AB toxin with the A subunit being C3larvin (C3larvinA) and a putative B subunit (C3larvinB) encoded by the second gene. Sequence and structural analyses demonstrated that C3larvinB is a homologue of the Bacillus anthracis protective antigen (PA), the B subunit of anthrax toxin. The C3larvinAB toxin locus was interrupted by point mutations in all analysed P. larvae ERIC I and ERIC II strains. Only one P. larvae ERIC III/IV strain harboured an uninterrupted toxin locus comprising full‐length genes for C3larvinA and B. Exposure bioassays did not substantiate a role as virulence factor for C3larvinAB in P. larvae ERIC I/II. However, the PA homologue C3larvinB had an influence on the virulence of the unique P. larvae strain expressing the functional C3larvinAB locus.

Characterization of C3larvinA, a novel RhoA-targeting ADP-ribosyltransferase toxin produced by the honey bee pathogen,Paenibacillus larvae

“…Consistent with this observation, P. larvae ERIC I and ERIC II gene inactivation mutants lacking C3larvin trunc expression did not cause larval mortality compared with WT strains when used for experimental infection [39]. These data suggested that despite its enzymatic activity in biochemical assays [38], C3larvin trunc does not influence the virulence of P. larvae [39]. Further in silico analyses then revealed that in P. larvae ERIC I and ERIC II, the C3larvin trunc gene is part of a binary AB toxin locus which had been annotated as non-functional due to several disruptions of the open-reading frames coding for the A-and B-subunits [34].…”

“…However, C3larvin trunc was shown to lack N-terminal sequences responsible for cell-entry activity, and indeed, the toxin was unable to invade mouse macrophages [38]. Consistent with this observation, P. larvae ERIC I and ERIC II gene inactivation mutants lacking C3larvin trunc expression did not cause larval mortality compared with WT strains when used for experimental infection [39]. These data suggested that despite its enzymatic activity in biochemical assays [38], C3larvin trunc does not influence the virulence of P. larvae [39].…”

“…In this context, a full-length C3larvinAB locus comprising non-interrupted genes for the A-(C3larvinA) and the B-subunits (C3larvinB) was found in a singular P. larvae ERIC III/IV strain. Remarkably, C3larvinA contains the N-terminal sequences [39] not present in the originally described, inactive C3larvin trunc [38].…”

“…The 248-residue (28.6 kDa) C3larvinA protein produced by P. larvae strain ERIC III/IV strain, 11-8051 [39] is a single-domain mART toxin that possesses a 26-residue leader sequence. The mature protein is 222 residues (25.7 kDa).…”

“…C3larvinA has the hallmark catalytic Q-X-E signature of C3-like mART toxins ( Figure 1A) [39]. The ten C3-like exotoxins known so far are shown in the sequence identity matrix ( Figure 1B) and share a minimum of 27% sequence identity, and the most similar pair includes C3larvin trunc and C3larvinA at 99% identity ( Figure 1A,B).…”