Polymerase chain reaction and DNA probe hybridization to assess the efficacy of diminazene treatment in Trypanosoma brucei-infected cattle

Four of eight Ankole longhorn cattle experimentally infected with Trypanoso ma brucei were treated with 7 mg/kg diminazene aceturate (Berenil, Hoechst AG, Germany) at day 71 postinfection. The trypanocidal activity was monitor ed using polymerase chain reaction (PCR) and DNA probe hybridization. When extracted parasite DNA (without host DNA) was used, as little as 1 fg per r eaction, which is equivalent to about 1-10% of the DNA in a single trypanos ome, produced a specific product that was visible as a 177-bp band in an ag arose gel. In infected cattle, specific PCR products could be amplified at as early as 1 day postinfection. PCR signals remained positive during infec tion, except in one sample, although aparasitemic phases occurred. In cases where treatment resulted in a significant clinical improvement, PCR signal s disappeared at 3-4 days after the administration of the drug. By contrast , in cattle that showed clinical signs of CNS involvement after treatment, although aparasitemic. and died before the termination of the experiment. s pecific products could be amplified on several occasions following treatmen t. The PCR signals generated after treatment could be further enhanced by s ubsequent slot-blot hybridization with a T. brucei-specific DNA probe. We c onclude that PCR coupled with DNA probe hybridization provides a highly sen sitive tool for the assessment of therapeutic efficiency and disease progre ssion in trypanosome infections, especially in chronic infections when the level of parasitemia is low or when trypanosomes are sequestered at cryptic sites.