Botulinum neurotoxins are diverse proteins. They are currently represented by at least seven serotypes and more than 40 subtypes. New clostridial strains that produce novel neurotoxin variants are being identified with increasing frequency, which presents challenges when organizing the nomenclature surrounding these neurotoxins. Worldwide, researchers are faced with the possibility that toxins having identical sequences may be given different designations or novel toxins having unique sequences may be given the same designations on publication. In order to minimize these problems, an ad hoc committee consisting of over 20 researchers in the field of botulinum neurotoxin research was convened to discuss the clarification of the issues involved in botulinum neurotoxin nomenclature. This publication presents a historical overview of the issues and provides guidelines for botulinum neurotoxin subtype nomenclature in the future.

ABSTRACTClostridium botulinumtypes C and D, as well as their mosaic variants C-D and D-C, are associated with avian and mammalian botulism. This study reports on the development of low-density macroarrays based on the GeneDisc cycler platform (Pall-GeneDisc Technologies) applied to the simultaneous detection of theC. botulinumsubtypes C, C-D, D, and D-C. The limit of detection of the PCR assays was 38 fg of total DNA, corresponding to 15 genome copies. Artificially contaminated samples of cecum showed a limit of detection below 50 spores/g. The tests were performed with a large variety of bacterial strains, includingC. botulinumtypes C (n= 12), C-D (n= 29), D (n= 5), and D-C (n= 10), other botulinum neurotoxin (BoNT)-producingClostridiumstrains (n= 20), non-BoNT-producing clostridia (n= 20), and other bacterial species (n= 23), and showed a high specificity. These PCR assays were compared to previously published real-time PCRs for the detection ofC. botulinumin 292 samples collected from cases of botulism events in four European regions. The majority of the samples originated from wild birds (n= 108), poultry (n= 60), and bovines (n= 56). Among the 292 samples, 144 were positive for either thebont/C-D or thebont/D-C gene by using the GeneDisc arrays. The reliability of the results tallied to 97.94%. Interestingly, only BoNT mosaics, types C-D and D-C, were found in naturally contaminated samples whatever their animal origin and their geographical location. Further investigations should now be performed in order to check that mosaic types dominate in Europe and that acquisition of mosaic types helps in survival or adaptation to particular niche.

A two year study (2008-2009) was carried out to monitor the Usutu virus (USUV) circulation in Italy. Sentinel horses and chickens, wild birds and mosquitoes were sampled and tested for the presence of USUV and USUV antibodies within the WND National Surveillance plan. Seroconversion evidenced in sentinel animals proved that in these two years the virus has circulated in Tuscany, Emilia Romagna, Veneto and Friuli Venezia Giulia regions. In Veneto USUV caused a severe blackbird die-off disease involving at least a thousand birds. Eleven viral strains were detected in organs of 9 blackbirds (52.9%) and two magpies (0.5%) originating from Veneto and Emilia Romagna regions. USUV was also detected in a pool of Culex pipiens caught in Tuscany. According to the alignment of the NS5 partial sequences, no differences between the Italian USUV strains isolated from Veneto, Friuli and Emilia Romagna regions were observed. The Italian North Eastern strain sequences were identical to those of the strain detected in the brain of a human patient and shared a high similarity with the isolates from Vienna and Budapest. Conversely, there were few differences between the Italian strains which circulated in the North Eastern regions and the USUV strain detected in a pool of C. pipiens caught in Tuscany. A high degree of similarity at both nucleotide and amino acid level was also found when the full genome sequence of the Italian North Eastern isolate was compared with that of the strains circulating in Europe. The North Eastern Italian strain sequence exhibited 97% identity to the South African reference strain SAAR-1776. The deduced amino acid sequences of the Italian strain differed by 10 and 11 amino-acids from the Budapest and Vienna strains, respectively, and by 28 from the SAAR-1776 strain. According to this study two strains of USUVs are likely to have circulated in Italy between 2008 and 2009. They have developed strategies of adaptation and evolution to spread into new areas and to become established.

The genome of Weissella oryzae SG25T was recently sequenced and a botulinum neurotoxin (BoNT) like gene was identified by bioinformatics methods. The typical three-domains organization of BoNTs with a N-terminal metalloprotease domain, a translocation and a cell binding domains could be identified. The BoNT family of neurotoxins is rapidly growing, but this was the first indication of the possible expression of a BoNT toxin outside the Clostridium genus. We performed molecular modeling and dynamics simulations showing that the 50 kDa N-terminal domain folds very similarly to the metalloprotease domain of BoNT/B, whilst the binding part is different. However, neither the recombinant metalloprotease nor the binding domains showed cross-reactivity with the standard antisera that define the seven serotypes of BoNTs. We found that the purified Weissella metalloprotease cleaves VAMP at a single site untouched by the other VAMP-specific BoNTs. This site is a unique Trp-Trp peptide bond located within the juxtamembrane segment of VAMP which is essential for neurotransmitter release. Therefore, the present study identifies the first non-Clostridial BoNT-like metalloprotease that cleaves VAMP at a novel and relevant site and we propose to label it BoNT/Wo.

No abstract

No abstract

No abstract

No abstract