First Draft Genome for a Burkholderia mallei Isolate Originating from a Glanderous Mule from Brazil.

Burkholderia mallei is the etiological agent of glanders. Here, we present the draft genome sequence of Burkholderia mallei strain 16-2438_BM#8 that was isolated from a mule found dead in Pernambuco, northeast Brazil. It is the first available genomic sequence from a strain isolated on the American continent.

Occurrence of C. botulinum in healthy cattle and their environment following poultry botulism outbreaks in mixed farms.

Ten cattle farms located in an area with a recent history of poultry botulism outbreaks were investigated to evaluate the occurrence of toxigenic C. botulinum in healthy cattle. Environmental samples in the 10 cattle farms and bovine fecal contents in farms with a confirmed environmental contamination were collected. Detection of C. botulinum toxin genes C, D, C/D, D/C and E was performed using real-time PCR. 4.9% (7/143) of the environmental samples collected in the 10 investigated cattle farms were positive for C. botulinum type C/D. Theses samples (boot-swabs in stalls and on pasture and water of a stream) were collected in 3 different farms. One cow dung sample and 3 out of 64 fecal contents samples collected in a single farm were also positive for C. botulinum type C/D. This study demonstrates that cattle are probably indirectly contaminated via poultry botulism in the area and that they can be intermittent carrier of C. botulinum type C/D after poultry botulism outbreaks in mixed farms.

Investigation of Clostridium botulinum in commercial poultry farms in France between 2011 and 2013.

Between 2011 and 2013, 17 poultry botulism outbreaks were investigated in France. All cases were associated with Clostridium botulinum type C-D. Presence of C. botulinum was studied in seven areas: poultry house, changing room, ventilation system, surroundings, animal reservoirs, water, and feed. Swabs, litter, soil, darkling beetles, rodents and wild bird droppings, feed and water samples were collected. The presence of C. botulinum type C-D in the environment of affected flocks was detected in 39.5% of the 185 samples analysed by real-time polymerase chain reaction. C. botulinum type C-D was reported in each area. Four areas were more frequently contaminated, being found positive in more than one-half of farms: darkling beetles (9/11), poultry house (14/17), water (13/16) and surroundings (11/16). After cleaning and disinfection, the ventilation system and/or the soil (in the houses and the surroundings) returned positive results in four out of eight poultry farms. Consequently, darkling beetles, the drinking water, the ventilation system and the soil in the surroundings and the houses were identified as the main critical contaminated areas to consider in poultry farms to prevent recurrence of botulism outbreaks.

An innovative molecular detection tool for tracking and tracing Clostridium botulinum types A, B, E, F and other botulinum neurotoxin producing Clostridia based on the GeneDisc cycler.

Rapid and specific detection of botulinum neurotoxin (BoNT) producing Clostridia is a priority for public health authorities, in case of both natural and intentional botulism outbreaks. This study reports on the evaluation of a detection system based on the GeneDisc Cycler designed for simultaneously testing the bont/A, bont/B, bont/E and bont/F genes encoding for the botulinum neurotoxins types A, B, E and F. BoNT-producing Clostridia (n = 102) and non-BoNT-producing bacteria (n = 52) isolated from clinical, food and environmental samples were tested using this macro-array and results were compared to the reference lethality test on mice. The bont genes were correctly detected in all C. botulinum type A, B, E and F strains available, as well as in toxigenic C. baratii type F and toxigenic C. butyricum type E. No cross reactivity was observed with non human-toxigenic bacteria, C. botulinum types C, D and G. The identification of the bont genotype using the macro-array was correlated to toxino-typing of the BoNTs as determined by the mouse bioassay. An "evaluation trial" of the GeneDisc array performed blind in four European laboratories with 77 BoNT-producing Clostridia as well as 10 food and clinical samples showed that the developed macro-array is specific and reliable for identifying BoNT/A-, BoNT/B-, BoNT/E- and BoNT/F-producing clostridial strains and for screening naturally contaminated food and fecal samples. The test is robust, has a low detection limit (c.a. 5 to 50 genome copies in the PCR reaction microwell) and is promising for monitoring BoNT-producing Clostridia in different kinds of samples including food and clinical samples.

Molecular characterization of IRF3 and IRF7 in rainbow trout, Oncorhynchus mykiss: functional analysis and transcriptional modulation.

Interferon regulatory factors (IRF) 3 and 7 in mammals are known to be crucial in regulating the type I interferon (IFN) response to viral infection as part of transcriptional complexes binding to IRF-binding elements (IRF-Es) and interferon stimulatory response elements (ISREs) within IFN and interferon-stimulated genes (ISGs). Here we report the sequencing and characterization of full-length cDNA homologues of rainbow trout (rt)IRF7 and, for the first time in fish, IRF3. RtIRF3 consists of 2127 bp with a 159 bp 5'-UTR-containing two upstream AUGs and a 573 bp 3'-UTR. RtIRF7 was found to be 2055 bp, with a 102 bp 5'-UTR and a 705 bp 3'-UTR. The open reading frames (ORFs) translate into 464 amino acid and 415 amino acid proteins, respectively, each possessing a putative DNA-binding domain (DBD) containing a tryptophan cluster, which is characteristic of all IRF family members. The presence of putative IRF association domain (IAD)s, serine-rich C terminal domains (poorly conserved in trout IRF3), and phylogenetic analysis places the two genes in the IRF3 subfamily. Both genes were found to be upregulated by poly I:C, type I recombinant rainbow trout (r) IFN (second isoform, type I rIFN), type II rIFN (rIFNgamma), LPS, and rIL-1beta in the trout macrophage cell line, RTS-11. Poly I:C and type I rIFN also induced IRF3 and IRF7 expression in a trout fibroblast cell line (RTG-2). Transient transfection of RTG-2 cells with each IRF fused to GFP revealed a predominant cytoplasmic distribution found most intensely around the nucleus and, to a lesser extent, within cell nuclei. Transient transfection of rtIRF3 in the Mx-1-luciferase reporter cell line, RTG-P1, revealed a modest increase in luciferase activity relative to the vehicle control, which was lost in cells over-expressing a DBD-truncated form of rtIRF3. Both full-length and DBD-truncated forms of rtIRF7 increased reporter activity relative to the control, although to a non-significant extent. Electromobility shift assays (EMSAs) did not reveal a specific interaction between each IRF and the ISRE element found in the Mx-1 promoter, although the Mx-1 ISRE bound specifically to endogenous transcriptional complexes. These data support the premise that rtIRF3 and rtIRF7 are important molecules in the regulation of antiviral responses in fish, with the impact of rIFNgamma on rtIRF3/7 expression implying a role for these IRFs in immune processes other than type I IFN-driven antiviral responses.