Babesia bovis: Effects of cysteine protease inhibitors on in vitro growth

Okubo, Ken; Yokoyama, Naoki; Yadav, Govind; Alhassan, Andy; Igarashi, Ikuo

doi:10.1016/j.exppara.2007.04.009

Cited by 36 publications

(27 citation statements)

References 27 publications

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“…6,8,37 Conversely, effective doses of pepstatin A and mefloquine for bovine Babesia parasites were higher than those of other drugs tested in previous studies. 10,29,[38][39][40][41] For equine Babesia parasites, the IC 50 values of pepstatin A and mefloquine were similar to those reported in previous studies 6,7,16,42 but significantly lower than the IC 50 values reported for equine Babesia. Moreover, the combination of pepstatin A and mefloquine produced antagonistic effects on in vitro-cultured parasites.…”

Section: Discussionsupporting

confidence: 60%

“…[3][4][5] Although some anti-babesial agents have been used to control the disease, continuous searches for the development of new drugs against Babesia are caused by toxic side effects, repeated relapse of parasite infections, and the possibility of emerging drug-resistant parasites. 6 Several novel antibabesial drugs, such as triclosan, 6 artesunate, pyrimethamine, pamaquine, 7 heparin, 8 imidazole derivatives, staurosporine, 9 and cysteine protease inhibitors, 10 have been successfully studied by using in vitro and in vivo models. However, these drugs have not been evaluated for field application.…”

Section: Introductionmentioning

confidence: 99%

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Inhibitory Effects of Pepstatin A and Mefloquine on the Growth of Babesia Parasites

Munkhjargal¹,

AbouLaila²,

Terkawi³

et al. 2012

The American Society of Tropical Medicine and Hygiene

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Abstract. We evaluated the inhibitory effects of pepstatin A and mefloquine on the in vitro and in vivo growths of Babesia parasites. The in vitro growth of Babesia bovis, B. bigemina, B. caballi, and B. equi was significantly inhibited (P 0.05) by micromolar concentrations of pepstatin A (50% inhibitory concentrations = 38.5, 36.5, 17.6, and 18.1 μM, respectively) and mefloquine (50% inhibitory concentrations = 59.7, 56.7, 20.7, and 4 μM, respectively). Furthermore, both reagents either alone at a concentration of 5 mg/kg or in combinations (2.5/2.5 and 5/5 mg/kg) for 10 days significantly inhibited the in vivo growth of B. microti in mice. Mefloquine treatment was highly effective and the combination treatments were less effective than other treatments. Therefore, mefloquine may antagonize the actions of pepstatin A against babesiosis and aspartic proteases may play an important role in the asexual growth cycle of Babesia parasites.

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Section: Discussionsupporting

confidence: 60%

Section: Introductionmentioning

confidence: 99%

Inhibitory Effects of Pepstatin A and Mefloquine on the Growth of Babesia Parasites

Munkhjargal¹,

AbouLaila²,

Terkawi³

et al. 2012

The American Society of Tropical Medicine and Hygiene

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“…In general, inhibition of tick and parasite proteases is of interest as both the tick and the parasite genomes encode for several cysteine proteases important for blood digestion (Sojka et al, 2008) and host invasion (Florin-Christensen and Schnittger, 2009), respectivelly. Addition of various cysteine protease inhibitors into the B. bovis culture resulted in parasite growth inhibition (Okubo et al, 2007). The cysteine proteases inhibitor called cystatin-2 (Hlcyst2) from H. longicornis (Zhou et al, 2006) was overexpressed in midgut and hemocytes after Babesia infection.…”

Section: Tick Interactions With Transmitted Pathogensmentioning

confidence: 99%

Interaction of the tick immune system with transmitted pathogens

Hajdušek¹,

Šíma²,

Ayllón³

et al. 2013

Front. Cell. Infect. Microbiol.

209

199

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“…However, evidence of their importance for the survival of these parasites was obtained by the observation that specific inhibitors of these enzymes impaired merozoite growth in vitro [24]. As in P. falciparum , cysteine peptidases may be involved in the erythrocyte egress of B. bovis merozoites, a prerequisite for the maintenance of the asexual propagation of the parasite, and/or nutrition of the trophozoite and merozoite parasite stages that reside within the erythrocyte through degradation of hemoglobin.…”

Section: Introductionmentioning

confidence: 99%

Bovipain-2, the falcipain-2 ortholog, is expressed in intraerythrocytic stages of the tick-transmitted hemoparasite Babesia bovis

et al. 2010

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BackgroundCysteine proteases have been shown to be highly relevant for Apicomplexan parasites. In the case of Babesia bovis, a tick-transmitted hemoparasite of cattle, inhibitors of these enzymes were shown to hamper intraerythrocytic replication of the parasite, underscoring their importance for survival.ResultsFour papain-like cysteine proteases were found to be encoded by the B. bovis genome using the MEROPS database. One of them, the ortholog of Plasmodium falciparum falcipain-2, here named bovipain-2, was further characterized. Bovipain-2 is encoded in B. bovis chromosome 4 by an ORF of 1.3 kb, has a predicted molecular weight of 42 kDa, and is hydrophilic with the exception of a transmembrane region. It has orthologs in several other apicomplexans, and its predicted amino acid sequence shows a high degree of conservation among several B. bovis isolates from North and South America. Synteny studies demonstrated that the bovipain-2 gene has expanded in the genomes of two related piroplasmids, Theileria parva and T. annulata, into families of 6 and 7 clustered genes respectively. The bovipain-2 gene is transcribed in in vitro cultured intra-erythrocyte forms of a virulent and an attenuated B. bovis strain from Argentina, and has no introns, as shown by RT-PCR followed by sequencing. Antibodies against a recombinant form of bovipain-2 recognized two parasite protein bands of 34 and 26 kDa, which coincide with the predicted sizes of the pro-peptidase and mature peptidase, respectively. Immunofluorescence studies showed an intracellular localization of bovipain-2 in the middle-rear region of in vitro cultured merozoites, as well as diffused in the cytoplasm of infected erythrocytes. Anti-bovipain-2 antibodies also reacted with B. bigemina-infected erythrocytes giving a similar pattern, which suggests cross-reactivity among these species. Antibodies in sera of two out of six B. bovis-experimentally infected bovines tested, reacted specifically with recombinant bovipain-2 in immunoblots, thus demonstrating expression and immunogenicity during bovine-infecting stages.ConclusionsOverall, we present the characterization of bovipain-2 and demonstrate its in vitro and in vivo expression in virulent and attenuated strains. Given the involvement of apicomplexan cysteine proteases in essential parasite functions, bovipain-2 constitutes a new vaccine candidate and potential drug target for bovine babesiosis.

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Babesia bovis: Effects of cysteine protease inhibitors on in vitro growth

Cited by 36 publications

References 27 publications

Inhibitory Effects of Pepstatin A and Mefloquine on the Growth of Babesia Parasites

Inhibitory Effects of Pepstatin A and Mefloquine on the Growth of Babesia Parasites

Interaction of the tick immune system with transmitted pathogens

Bovipain-2, the falcipain-2 ortholog, is expressed in intraerythrocytic stages of the tick-transmitted hemoparasite Babesia bovis

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