Goldstein Michael R, Kruth Stephen A, Bersenas Alexa M E, Holowaychuk Marie K, Weese J Scott
Department of Clinical Studies, Ontario Veterinary College, University of Guelph, Guelph, Ontario N1G 2W1.
Can J Vet Res. 2012 Jul;76(3):161-5.
Clostridium perfringens has been implicated as a cause of diarrhea in dogs. The objectives of this study were to compare 2 culture methods and to evaluate a multiplex polymerase chain reaction (PCR) assay to detect C. perfringens toxin genes alpha (α), beta (β ), beta 2 (β2), epsilon (ɛ), iota (ι), and C. perfringens enterotoxin (cpe) from canine isolates. Fecal samples were collected from clinically normal non-diarrheic (ND) dogs, (n = 105) and diarrheic dogs (DD, n = 54). Clostridium perfringens was isolated by directly inoculating stool onto 5% sheep blood agar (SBA) and enrichment in brain-heart infusion (BHI) broth, followed by inoculation onto SBA. Isolates were tested by multiplex PCR for the presence of α, β, β2, ɛ, ι, and cpe genes. C. perfringens was isolated from 84% of ND samples using direct culture and from 87.6% with enrichment (P = 0.79). In the DD group, corresponding isolation rates were 90.7% and 93.8% (P = 0.65). All isolates possessed the α toxin gene. Beta (β), β2, ɛ, ι, and cpe toxin genes were identified in 4.5%, 1.1%, 3.4%, 1.1%, and 14.8% of ND isolates, respectively. In the DD group, β and β2 were identified in 5%, ɛ and ι were not identified, and the cpe gene was identified in 16.9% of isolates. Enrichment with BHI broth did not significantly increase the yield of C. perfringens, but it did increase the time and cost of the procedure. C. perfringens toxin genes were present in equal proportions in both the ND and DD groups (P ≤ 0.15 to 0.6). Within the parameters of this study, culture of C. perfringens and PCR for toxin genes is of limited diagnostic usefulness due to its high prevalence in normal dogs and the lack of apparent difference in the distribution of toxin genes between normal and diarrheic dogs.
产气荚膜梭菌被认为是犬腹泻的病因之一。本研究的目的是比较两种培养方法，并评估一种多重聚合酶链反应（PCR）检测方法，以检测犬分离株中产气荚膜梭菌毒素基因α、β、β2、ε、ι和产气荚膜梭菌肠毒素（cpe）。从临床正常的非腹泻（ND）犬（n = 105）和腹泻犬（DD，n = 54）采集粪便样本。通过将粪便直接接种到5%绵羊血琼脂（SBA）上并在脑心浸液（BHI）肉汤中增菌，然后接种到SBA上，来分离产气荚膜梭菌。通过多重PCR检测分离株中α、β、β2、ε、ι和cpe基因的存在情况。使用直接培养法从84%的ND样本中分离出产气荚膜梭菌，增菌法的分离率为87.6%（P = 0.79）。在DD组中，相应的分离率分别为90.7%和93.8%（P = 0.65）。所有分离株均具有α毒素基因。在ND分离株中，分别有4.5%、1.1%、3.4%、1.1%和14.8%的分离株鉴定出β、β2、ε、ι和cpe毒素基因。在DD组中，5%的分离株鉴定出β，未鉴定出β2，16.9%的分离株鉴定出cpe基因，未鉴定出ε和ι。用BHI肉汤增菌并没有显著提高产气荚膜梭菌的分离率，但增加了操作时间和成本。产气荚膜梭菌毒素基因在ND组和DD组中的比例相同（P ≤ 0.15至0.6）。在本研究的参数范围内，产气荚膜梭菌培养和毒素基因PCR的诊断价值有限，因为其在正常犬中普遍存在，且正常犬和腹泻犬之间毒素基因分布缺乏明显差异。
