Prevalence of netB-positive Clostridium perfringens in Italian poultry flocks by environmental sampling.

Clostridium perfringens type G is one of the pathogens involved in enteric diseases in poultry. NetB, a pore-forming toxin, is considered the main virulence factor responsible for necrotic enteritis during C. perfringens infection. We carried out a field study involving 14 farms to evaluate the occurrence of netB-positive C. perfringens and the impact of infection in Italian poultry flocks. Environmental samples (n = 117) and 50 carcasses were screened by microbiologic and molecular methods. Microbiologic investigations yielded 82 C. perfringens isolates. DNA was extracted from all samples and screened for α-toxin and NetB encoding genes by real-time PCR. The C. perfringens α-toxin gene was detected in 151 of 167 extracts (90.4%), and 31 of 151 (20.5%) were netB gene positive also. Sixteen isolates from a turkey flock with mild enteric disorders were also netB positive, demonstrating their occurrence not only in broiler but also in turkey flocks. A pulsed-field gel electrophoresis protocol was optimized to evaluate the diversity among isolates and revealed high genetic heterogeneity. The complete NetB toxin-coding gene of 2 C. perfringens isolates from turkey and broiler flocks were analyzed and showed very high relatedness with analogous sequences worldwide.

Cryopreservation of turkey spermatozoa without permeant cryoprotectants.

In avian species, cryopreservation of semen is necessary for developing sperm cryobanks. It is very difficult, however to cryopreserve turkey sperm and have sperm be viable after thawing. Glycerol, the commonly used sperm cryoprotectant in many species, is toxic to sperm of avian species. The aim of this study was to evaluate whether the non-permeating dextran was effective for the cryopreservation and maintenance of turkey spermatozoa viability after thawing, avoiding the use of permeating cryoprotectants. Turkey sperm were diluted with a medium supplemented with 11% glycerol or dextran with a 1,000 molecular weight (MW), dextran with a 10,000 MW, or dextran with a 20,000 MW each at a 2%, 5%, or 10% concentration. Sperm kinetic characteristics, membrane and acrosome integrity (AI), and the capacity of spermatozoa to interact with the autologous perivitelline layer were evaluated after equilibration and cryopreservation. Results indicate that with use of glycerol and the 1,000 MW dextran there was lesser sperm viability after both equilibration and cryopreservation, compared with use of the 10,000 or 20,000 MW dextran compounds. There was a greater cryoprotective effect with the 10,000 and 20,000 MW dextran compounds at the 10% concentration with spermatozoa maintaining a greater functionality and capacity to interact with the autologous perivitelline layer. In conclusion, the results of this study indicate turkey spermatozoa could be effectively cryopreserved in extender without the use of glycerol as a penetrating cryoprotectant but with the use of the 10,000 or 20,000 MW dextran compounds at a 10% extender concentration.