Toxoplasmosis, caused by Toxoplasma gondii (Tg), is one of the most prevalent zoonotic diseases worldwide. Currently, safe and efficient therapeutic options for this disease are still being developed, and are urgently needed. Agrimonia pilosa (A. pilosa) is a rosaceous plant, and widely distributed in Asia. The bioactive compounds in A. pilosa Ledeb include flavonoid, triterpenoids, and isocoumarin, and have been reported to show many pharmacological effects including anti-inflammatory, antioxidant, and antibacterial activities.The aim of this study was to determine Anti-protozoal efficacy of Agrimonia pilosa ethanol extract (APE) against Toxoplasma gondii (Tg). In this study, APE was examined and screened for anti-Tg activity. Seventy percent ethanol was used for herbal extraction. For anti-Tg activity screening, in vitro study with APE using tachyzoit of Tg RH strain infected HeLa cells were performed. Also, in vivo anti- Tg study using the mouse infection model was conducted. As the results, the selectivity of APE was 3.35, which was higher than Sulfadiazine selectivity (2.08). Also, we performed in vivo animal study to evaluate the anti-Tg activity of APE with mouse model. As the results, Anti-toxoplasma activity of the APE was as high as sulfadiazine. These results demonstrate that it can be successfully cured against Tg infection with APE. It could be a promising native herb treatment for the prevention of Tg infection.

인수공통 감염증의 하나인 톡소포자충의 검출을 위해서는 대부분은 ELISA 법이 사용 되고 있으나, 충체가 사멸 된 후에도 양성반응이 나타나는 등 사용에 제한이 있다. 반면 유전자 검출법은 현재 감염상태를 확인 할 수 있기 때문에 식중독 원인조사 등에 적합하다고 판단되어 이를 활용하여 본 연구를 진행하였다. 톡소포자충의 유전정보를 통해 529 repeat region의 염기서열을 얻고, 프라이머 및 TaqMan 프로브를 설계하여 real-time PCR을 이용한 검 출법을 개발하였다. 검출한계(lower limit of detection) 및 적정곡선을 확인한 결과 10 genomic DNA copy가 검출한 계로 확인되었고, 정량을 위한 곡선은 101~106 DNA copies 까지 0.999의 R2값을 나타내었다. 개발된 검출법의 증폭효 율을 비교하기 위해 B1 gene 타겟 프라이머 세트와 타입 별 검출한계를 비교한 결과, type 1, 2, 3 톡소포자충에서 같거나 더 나은 검출한계를 보였다. 또한 식품에서 주로 분리되는 식중독 세균 14종 및 원충 3종에 대해 특이도를 비교한 결과, 모두 음성으로 나타났다. 개발된 검출법을 식육검체에 적용하였을 때 type 1, 2, 3에서 모두 원활한 검출결과를 보여 증폭방해물질이 존재하지 않은 것으로 확인되었다. 본 연구를 통해 개발된 유전자검출법은 국내 유통 중인 식육에서 인수 공통감염 원충의 하나인 톡소포 자충의 감염 여부를 확인하는 사전적 모니터링의 방법으로 활용될 예정이다.

The in vitro culture of Toxoplasma gondii was evaluated using a JNUCK cell strain that was independently developed by our research team. The sensitivity of the JNUCK strain was compared with those of MDCK and vero cell strains, which were previously shown to be sensitive to T. gondii. Morphological observation and 3H-uracil absorption tests showed that JNUCK cell strain has the highest sensitivity, while the MDCK and vero cell strains showed lower sensitivities. The optimal culture conditions using the JNUCK cell strain were determined by inoculating T. gondii at multiplicity of infection (MOI) 1, MOI 5, and MOI 25 and the numbers of T. gondii were measured 24, 48, and 72 hours after inoculation. The optimal medium composition was determined by adding different concentrations of FBS to DMEM medium for inoculation at MOI 5, and then calculating the numbers of tachyzoites inside the cells and in the medium. The numbers of tachyzoites inside the cell were highest for 10% FBS, while the numbers of tachyzoites in the medium peaked after addition of 30% FBS. These studies confirmed that the optimal culture condition of T. gondii using the JNUCK cell strain was achieved by inoculation at MOI 5, using medium containing 20% FBS and culturing for 72 hours.

Toxoplasma gondii (T. gondii) causes a life-threatening opportunistic infection. Despite its clinical importance, very few therapeutic drugs against T. gondii are available. Furthermore, these therapeutic regimens are not always suitable for prolonged treatment due to adverse side effects as well as the potential of clinical failure by selecting drug-resistant parasite variants. Dictamnus dasycarpus is known to have many medicinal properties, including anti-inflammatory, anti-fever, and anti-rheumatic activities. In this study, 70% ethanol extract of Dictamnus dasycarpus showed anti-T. gondii effects. Ethanolic extracts of Dictamnus dasycarpus used to treat T. gondii were tested in vitro for their anti-T. gondii activity and cytotoxicity. The selectivity of Dictamnus dasycarpus extract was 7.52, which was higher than that of Sulfadiazine (2.08). We conducted an in vivo animal test to evaluate the anti-T. gondii activity of Dictamnus dasycarpus extract as compared with that of Sulfadiazine. In T. gondii-infected mice, the inhibition rate of Dictamnus dasycarpus extract was high, similar to that of Sulfadiazine. This indicates that Dictamnus dasycarpus extract may be a source of new anti-T. gondii compounds.