As part of an ongoing investigation of filamentous fungi for anticancer leads, an active culture was identified from the Mycosynthetix library (MSX 70741, of the order Hypocreales, Ascomycota). The fungal extract exhibited cytotoxic activity against the H460 (human non-small cell lung carcinoma) cell line, and bioactivity-directed fractionation yielded peptaibols 1–12 and harzianums A (13) and B (14). Structure elucidation of 1–12 was facilitated by high-resolution MS/MS obtained on a Thermo LTQ Orbitrap XL using Higher-Energy Collisional Dissociation (HCD) and by high field NMR (950 MHz). The absolute configuration was determined by Marfey’s analysis of the individual amino acids; the time required for such analysis was decreased via the development of a 10 min UPLC method. The isolated peptaibols (1–12), along with three other peptaibols isolated and elucidated from a different fungus (MSX 57715) of the same Order (15–17), were examined for activity in a suite of biological assays, including those for cytotoxic, antibacterial, and anthelmintic activities.

We have been collaborating since 1992 in studies on southern sea otters (Enhydra lutris nereis) as part of a program to define factors, which may be responsible for limiting the growth of the southern sea otter population. We previously demonstrated Toxoplasma gondii in sea otters. We postulated that cat feces containing oocysts could be entering the marine environment through storm run-off or through municipal sewage since cat feces are often disposed down toilets by cat owners. The present study examined the sporulation of T. gondii oocysts in seawater and the survival of sporulated oocysts in seawater. Unsporulated oocysts were placed in 15 ppt artificial seawater, 32 ppt artificial seawater or 2% sulfuric acid (positive control) at 24 C in an incubator. Samples were examined daily for 3 days and development monitored by counting 100 oocysts from each sample. From 75 to 80% of the oocysts were sporulated by 3 days post-inoculation under all treatment conditions. Groups of 2 mice were fed 10,000 oocysts each from each of the 3 treatment groups. All inoculated mice developed toxoplasmosis indicating that oocysts were capable of sporulating in seawater. Survival of sporulated oocysts was examined by placing sporulated T. gondii oocysts in 15 ppt seawater at room temperature 22-24 C (RT) or in a refrigerator kept at 4 C. Mice fed oocysts that had been stored at 4C or RT for 6 months became infected. These results indicate that T. gondii oocysts can sporulate and remain viable in seawater for several months.

Toxoplasma gondii is a well-recognized cause of disease in congenitally infected and immunocompromised individuals. Histone deacetylases (HDAC) comprise a family of enzymes that participate in the regulation of chromatin structure, gene expression, and cell signaling in eukaryotes. Toxoplasma gondii expresses a HDAC Class I enzyme homologous to human hdac3. Previous work showed that the histone deacetylase inhibitors (HDI) apicidin and valproic acid inhibit T. gondii infections in vitro. The present study compares the activity of hydroxamic-acid histone deacetylase inhibitors against the RH strain of T. gondii growing in HS68 human foreskin fibroblast cells. Nanomolar concentrations of suberoylanilide hydroxamic acid (SAHA), suberic bishydroxamic acid (SBHA), scriptaid, and trichostatin A (TSA) inhibited T. gondii tachyzoite proliferation. Scriptaid was the most potent hydroxamic acid inhibitor (IC50 = 39 nM). In comparison, the carboxylate histone deacetylase inhibitors sodium valproate, sodium butyrate, and 4-phenylbutyrate were less potent (IC50 range 1-5 mM). All of the inhibitors tested, except SBHA, completely protected the HS68 monolayers from T. gondii at concentrations 3-6 times greater than their respective IC50. In contrast, nicotinamide, an inhibitor of NADI-dependent Class III HDAC, had minimal activity against T. gondii in our in vitro assays. We conclude that the hydroxamic acid class of histone deacetylase inhibitors exhibit potent anti-T. gondii activity in vitro.

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The synthesis of prostaglandins F2α and E2 by guinea‐pig uterine homogenates was inhibited by p‐bromophenacyl bromide (PBPAB), an inhibitor of phospholipase A2. Metabolism of prostaglandin F2α by uterine homogenates was undetectable; this was not affected by PBPAB. There was no significant difference between the amounts of arachidonic acid released from uterine homogenates on days 7 and 15 of the oestrous cycle. Small amounts of dihomo‐γ‐linolenic acid were detected in the homogenates. The release of arachidonic acid from uterine homogenates was greatly inhibited by PBPAB. Addition of exogenous arachidonic acid to uterine homogenates did not overcome the inhibition of uterine prostaglandin F2α synthesis produced by PBPAB. It is concluded that PBPAB inhibits both the release of arachidonic acid from the guinea‐pig uterus and its subsequent conversion into prostaglandins.

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We examined the effects of 5 microg/ml ponazuril treatment on developing tachyzoites of Neospora caninum and merozoites of Sarcocystis neurona to better determine the mode of action of this anticoccidial drug. Both parasites develop asexually by endogenesis. Neospora caninum was selected for study because it develops by endodyogeny, which results in two tachyzoites being produced internally, and S. neurona was selected because it develops by endopolygeny which results in many merozoites being produced internally. Ponazuril inhibited development of N. caninum after approximately 48 h post-exposure. Treated tachyzoites of N. caninum developed vacuoles and underwent degeneration. Ponazuril also inhibited development of merozoites of S. neurona. Treated merozoites and maturing schizonts of S. neurona developed vacuoles and underwent degeneration. The ability of S. neurona schizonts to undergo cytokinesis was inhibited. Our results are discussed in relation to previous ultrastructural research on endogenesis of tachyzoites of Toxoplasma gondii undergoing endodyogeny which indicated that ponazuril induced multinucleate stage formation and inhibited cytokinesis. Ponazuril is believed to act on the apicoplast and our study demonstrates that this agent may express its inhibitory effects in different phenotypic manners on different apicomplexan parasites. The enzyme/enzyme systems that are the inhibitory target of ponazuril may be different in these apicomplexans, or the results of inhibition may affect different pathways downstream of its initial site of action in these parasites.

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