Molecular subtyping of poultry-associated type A Clostridium perfringens isolates by repetitive-element PCR.
Clostridium perfringens strains (type A) isolated from an integrated poultry operation were subtyped using repetitive-element PCR with Dt primers. Isolates were obtained from fecal, egg shell, fluff, and carcass rinse samples as part of a previously reported temporally linked epidemiological survey. A total of 48 isolates of C. perfringens were obtained from different stages of the broiler chicken production chain from two separate breeder farms that supplied a single hatchery that in turn provided chicks to a single grow-out farm whose flocks were processed at a single plant. All 48 isolates were typeable (100% typeability) by repetitive-element PCR with Dt primers. This subtyping method was highly reproducible and discriminatory. By repetitive-element PCR with Dt primers, isolates were classified into four major branches with 12 subgroups or clades. The Simpson's index of discrimination was calculated to be 0.96 for groupings of >95% correlation. Toxin gene profiles of the isolates indicated that all of the isolates were C. perfringens alpha-toxin gene positive and 46 of 48 isolates were beta2-toxin gene positive. All strains were negative for beta- and epsilon-toxin genes. Repetitive sequence-based PCR was found to be a technically practical and reproducible means of subtyping C. perfringens libraries from specific epidemiological or production environment settings.
Siragusa
G R
GR
Agricultural Research Service, United States Department of Agriculture, Russell Research Center, Athens, Georgia 30604-5677, USA. siragusa@saa.ars.usda.gov
Danyluk
M D
MD
Hiett
K L
KL
Wise
M G
MG
Craven
S E
SE
D000818:Animals; D001427:Bacterial Toxins / Q000235:genetics; D015373:Bacterial Typing Techniques / Q000379:methods; D002135:Calcium-Binding Proteins / Q000235:genetics; D002645:Chickens / Q000382:microbiology*; D003016:Clostridium perfringens / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D005516:Food Microbiology; D005798:Genes, Bacterial; D016133:Polymerase Chain Reaction / Q000379:methods*; D012091:Repetitive Sequences, Nucleic Acid; D010738:Type C Phospholipases / Q000235:genetics
D001427:Bacterial Toxins; D002135:Calcium-Binding Proteins; D004269:DNA, Bacterial; C109896:cpb2 protein, Clostridium perfringens; D010738:Type C Phospholipases; C040649:alpha toxin, Clostridium perfringens