Sylvia Páz Díaz-Camacho

Solution-phase parallel synthesis of substituted chalcones and their antiparasitary activity against Giardia lamblia

A library of 25-membered chalcones was prepared by parallel synthesis. Substituted acetophenones and benzaldehydes were condensed using the Claisen-Schmidt base-catalyzed aldol condensation. Several chalcones showed in vitro antiparasitic activity against Giardia lamblia. The highest activity observed for the IC(50) values were 12.72, 15.05 and 15.31 microg/mL, respectively; these are potential leads for the development of antigiardial compounds.

A novel sigma-like glutathione transferase of Taenia solium metacestode☆

GSTs are a group of multifunctional enzymes, whose major functions involve catalysis of conjugation of glutathione thiolate anion with a multitude of bi-substrates or transportation of a range of hydrophobic ligands. Helminth GSTs are intimately involved in the scavenging of endogenously/exogenously-derived toxic compounds and xenobiotics. In this study, we identified a novel GST gene of Taenia solium metacestodes (TsMs), which is a causative agent of neurocysticercosis. The 804 bp-long cDNA encoded a 639 bp open reading frame (212 amino acid polypeptide), which exhibited the structural motif and domain organisation characteristic of GST. It formed a strong clade with trematode and insect sigmaGSTs. We designated this cDNA as TsM sigma-like GST (TsMsigmaGST). Native TsMsigmaGST identified through gel filtration combined with compatible immunoproteomics consisted of four isoforms at approximately 25 kDa with different pIs between 8.2 and 8.7. TsMsigmaGST showed an enzyme activity as a homodimer and was specifically expressed in the scolex cytosol. The recombinant TsMsigmaGST expressed in Escherichia coli showed sigma-like activity with 1-chloro-2,4-dinitrobenzene (CDNB). The Vmax and Km for CDNB and glutathione (GSH) were 1.08 and 0.78 micromol/min/mg, and 0.16 and 0.17 mM, respectively. Its optimal activity was observed at pH 8.0 and at 40 degrees C. The enzyme activity was potently inhibited by bromosulfophthalein, and to a lesser extent by rose bengal and triphenyltin chloride. Albendazole and praziquantel non-competitively inhibited both G- and H-sites of the enzyme. To our knowledge this is the first description of the sigma-class GST in cestode parasites. The enzyme might be involved in scavenging of intracellularly generated xenobiotics during homeostatic processes and anthelminthic metabolisms. Revelation of biochemical and biological properties of TsMsigmaGST might allow us to understand pathobiological events inherent to this long-standing parasitic disease, and thus to target therapeutic intervention.

Microwave-assisted synthesis of hydropyridines and study of the DPPH-scavenging activity

A series of hydropyridines (HPs) were synthesized using dimedones, ethyl acetoacetate, ammonium acetate and appropriate aldehydes under solvent-free conditions and microwave-irradiation through Hantzsch reaction. All the synthesized HPs were tested in vitro for the 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging activity and structure–activity relationship was discussed; only 12 out of the 36 HPs showed antioxidant activity. Polyhydroquinolines (PHQs) were the most active compounds among the studied HPs, they were characterized by the lack of an ester-like structure substituent at carbon 5.

Serodiagnostic reliability of single-step enriched low-molecular weight proteins of Taenia solium metacestode of American and Asian isolates

The low-molecular weight proteins (LMWPs) of Taenia solium metacestode (TsM) constituted pertinent serodiagnostic antigens for cysticercosis. We established a novel single-step purification of the LMWPs from TsM cyst fluid (CF). When the CF was precipitated with trichloroacetic acid/acetone mixture at the final concentrations of 5 and 50%, most LMWPs (ranging 7-38kDa) remained in the supernatant fraction. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot analyses revealed that the LMWPs consisted mainly with the previously reported subunit proteins of the 120- and 150-kDa antigen complexes. Serum samples from neurocysticercosis (NC) and other helminthic infections, as well as those from healthy controls were tested by both immunoblotting and micro-ELISA. In 135 cases of active stage NC patients, 132 cases (97.7%) showed positive reactions. Serum samples from other helminthic diseases (n=125) and healthy controls (n=100) exhibited no positive reactions except for cystic echinococcosis, of which 12% (3/25 cases) exhibited low levels of cross-reactivity. The LMWPs from different geographical regions (Korea and Mexico) showed diagnostic sensitivity and specificity of 97.7% and 98.7% against active stage NC. Our single-step separation method for the LMWPs provided excellent performance with easy applicability and high reproducibility, which has a great benefit for preparation of potent antigen in endemic areas.

Structural and Binding Properties of Two Paralogous Fatty Acid Binding Proteins of Taenia solium Metacestode

Background Fatty acid (FA) binding proteins (FABPs) of helminths are implicated in acquisition and utilization of host-derived hydrophobic substances, as well as in signaling and cellular interactions. We previously demonstrated that secretory hydrophobic ligand binding proteins (HLBPs) of Taenia solium metacestode (TsM), a causative agent of neurocysticercosis (NC), shuttle FAs in the surrounding host tissues and inwardly transport the FAs across the parasite syncytial membrane. However, the protein molecules responsible for the intracellular trafficking and assimilation of FAs have remained elusive. Methodology/Principal Findings We isolated two novel TsMFABP genes (TsMFABP1 and TsMFABP2), which encoded 133- and 136-amino acid polypeptides with predicted molecular masses of 14.3 and 14.8 kDa, respectively. They shared 45% sequence identity with each other and 15–95% with other related-members. Homology modeling demonstrated a characteristic β-barrel composed of 10 anti-parallel β-strands and two α-helices. TsMFABP2 harbored two additional loops between β-strands two and three, and β-strands six and seven, respectively. TsMFABP1 was secreted into cyst fluid and surrounding environments, whereas TsMFABP2 was intracellularly confined. Partially purified native proteins migrated to 15 kDa with different isoelectric points of 9.2 (TsMFABP1) and 8.4 (TsMFABP2). Both native and recombinant proteins bound to 11-([5-dimethylaminonaphthalene-1-sulfonyl]amino)undecannoic acid, dansyl-DL-α-amino-caprylic acid, cis-parinaric acid and retinol, which were competitively inhibited by oleic acid. TsMFABP1 exhibited high affinity toward FA analogs. TsMFABPs showed weak binding activity to retinol, but TsMFABP2 showed relatively high affinity. Isolation of two distinct genes from an individual genome strongly suggested their paralogous nature. Abundant expression of TsMFABP1 and TsMFABP2 in the canal region of worm matched well with the histological distributions of lipids and retinol. Conclusions/Significance The divergent biochemical properties, physiological roles and cellular distributions of the TsMFABPs might be one of the critical mechanisms compensating for inadequate de novo FA synthesis. These proteins might exert harmonized or independent roles on lipid assimilation and intracellular signaling. The specialized distribution of retinol in the canal region further implies that cells in this region might differentiate into diverse cell types during metamorphosis into an adult worm. Identification of bioactive systems pertinent to parasitic homeostasis may provide a valuable target for function-related drug design.

Discovery of an endemic area of Gnathostoma turgidum infection among opossums, Didelphis virginiana, in Mexico.

Abstract Gnathostomosis, caused by Gnathostoma binucleatum, is a serious public health issue in Mexico. Although 2 other Gnathostoma spp., G. turgidum and G. lamothei, have been found in wild animals, their natural life cycle or their relation to human disease remains unclear. While we were conducting an epidemiological survey on Gnathostoma spp. in Sinaloa State, Mexico, we found an endemic area for G. turgidum in common opossums, Didelphis virginiana, located in Tecualilla, Sinaloa. The species identification was carried out by morphological and molecular biological methods. This is the first record of an endemic area for G. turgidum infection in opossums, D. virginiana, in the Americas.

Identification of estuarine fish Dormitator latifrons as an intermediate host and Eleotris picta as a paratenic host for Gnathostoma binucleatum in Sinaloa, Mexico.

Gnathostomosis is a typical fish-borne zoonotic parasitosis and is currently a serious public health issue in Mexico. Among several Gnathostoma species present in wild animals in Mexico, Gnathostoma binucleatum is the only proven species responsible for human diseases, and the advanced third stage larvae (AL3) of G. binucleatum have been found in over 20 species of fish in this country. In Sinaloa State, two fish species, Dormitator latifrons and Eleotris picta, were heavily contaminated with G. binucleatum AL3. When we analyzed the relationship between the size of the fish and the density of infection with G. binucleatum AL3, the distribution patterns of AL3 were markedly different between these two fish species. Apparent size-dependent accumulation was observed in E. picta but not in D. latifrons, suggesting that E. picta is a paratenic host whereas D. latifrons is a second intermediate host.

Synthesis of 1H-benzoxazine-2,4-diones from heterocyclic anhydrides: evaluation of antioxidant and antimicrobial activities

A facile one-step synthesis of 1H-benzoxazine-2,4-diones from heterocyclic anhydrides and TMSA was described. This paper determines their antimicrobial activity against nine human bacterial pathogens by the broth microdilution method; antioxidant activity by DPPH• inactivation and a ferric-reducing power assay; and toxicity by a brine shrimp, Artemia salina, assay. The 1H-benzoxazine-2,4-dione yields were in the range of 57 to 98%. The novel compound 1H-pyrazino[2,3-][1,3]oxazine-2,4-dione 4c showed the highest antioxidant capacity (DPPH 35.4% and FRAP 0.063 µmol TEs/µmol).

Is Gnathostoma turgidum an Annual Parasite of Opossums? Drastic Seasonal Changes of Infection in Didelphis virginiana in Mexico

Abstract Gnathostoma turgidum is a nematode that parasitizes the stomach of opossums, Didelphis virginiana. Despite its wide distribution in the Americas, its natural life cycle is poorly understood. Recently, we found an endemic area for G. turgidum infection in Sinaloa, Mexico (Díaz-Camacho et al., 2009). Based on sporadic surveys for several years, the prevalence was apparently high in summer and extremely low in winter. To confirm that this is really a seasonal variance, we conducted a longitudinal survey on G. turgidum infection in opossums from November 2007 to November 2008. The results showed amazing seasonal changes in the prevalence, with synchronized migration and maturation of worms in opossums. Between February and March, many juvenile worms, with occasional AL3, were found in the liver, but no worms were found in the stomach. Mature adult worms began to appear in the stomach around April and rapidly increased in number toward July, when all worms resided in the stomach. Then, the worms disappeared almost completely by November. These results suggest that G. turgidum is an annual parasite of the opossum, D. virginiana, in Mexico.

Giardia duodenalis genotypes among schoolchildren and their families and pets in urban and rural areas of Sinaloa, Mexico

INTRODUCTION Giardiasis is a human health concern worldwide, especially among schoolchildren. Giardia duodenalis genotypes A and B are infective to humans, but their zoonotic potential remains controversial. In Mexico, the most prevalent genotype is A, but B was also detected in southeastern Mexico. In Sinaloa state, northwestern Mexico, giardiasis is highly prevalent, but Giardia genotypes have been poorly studied. METHODOLOGY This study aimed to investigate the distribution and clinical-epidemiological correlation of G. duodenalis genotypes in schoolchildren and their families and pets in urban and rural areas of Sinaloa state, Mexico. RESULTS Among 395 schoolchildren (274 urban, 121 rural), 76 (49 urban, 27 rural) were infected with G. duodenalis. In total, 22 families (15 urban, 7 rural) of infected schoolchildren, consisting of 60 family members (41 urban, 19 rural) and 21 pet dogs (15 urban, 6 rural) were examined; 10 family members (5 urban, 5 rural) and 5 pet dogs (3 urban, 2 rural) of 10 families (6 urban, 4 rural) were infected. After PCR-RFLP analyses of vsp417 and gdh genes, genotype prevalence among infected urban schoolchildren was 79.5% AI, 12.8% AII, and 7.7% mixed AI+B. However, only AI genotype was found in family members and pets. In the rural area, only the AI genotype was detected. Genotypes were not correlated with clinical manifestations. CONCLUSIONS This paper shows the presence of B genotype in northwestern Mexico for the first time. Detection of AI genotype in dogs suggested the possible role of dogs as the reservoir for human giardiasis in Sinaloa, Mexico.