Trypanosoma cruzi Antigenic Proteins Shared with Acute Lymphoblastic Leukemia and Neuroblastoma.

Background. Research studies indicate that immunization with protein extracts of Trypanosoma cruzi, the protozoan parasite that causes Chagas disease, prevents the appearance of tumors in 60% of mice injected with the murine lung carcinoma tumor line. The molecular basis of this process is unknown, although the presence of specific antigens in tumor cells and on the surface of T. cruzi suggests an antiparasitic immune response, with an effective cross-reaction against cancer cells, hence the importance to identify the antigens involved and determine their potential as target cells in anticancer therapy. Aim. This study aimed to determine the presence of antigenic proteins of T. cruzi shared with acute lymphoblastic leukemia and neuroblastoma cells. Material and methods. To achieve this, polyclonal antibodies against T. cruzi were developed in rabbits, and reactivity was determined with protein extracts of acute lymphoblastic leukemia cells and neuroblastoma. The immunodetection of five different strains of T. cruzi against anti-T. cruzi polyclonal antibodies was also performed. Conclusion. The study allows the knowledge of the immunological interactions between cancer and parasites to be expanded and, therefore, contributes to the design of more and better projects that improve the therapeutic strategies applied in oncology.

Proteomic analysis of Plasmodium berghei in the ring phase during in vivo antiparasitic treatment with kramecyne.

Malaria is a parasitic disease of global importance due to its high annual death toll. The treatment for this infection is difficult for the increase in the populations of parasites resistant to the existing medicines, the development of new antimalarials is urgent needed. Several products developed for the control of malaria from herbalist have had a profound impact, for example, quinine obtained from the bark of the cinchona tree and recently those derived from artemisinin, whose discovery was the reason for the awarding of the 2015 Nobel Prize. The aim of the present study was to evaluate a compound named kramecyne extracted of "chayotillo" (Krameria cystisoides) plant used by the antiparasitic effect against some blood and intestinal protozoa (Giardia duodenalis y Trypanosoma cruzi). In addition is using for the treatment of inflammatory diseases. Measuring parasitaemia at different times, it was observed that in mice treated with kramecyne, it reached only 14% of parasitaemia at 7 days with a dose of 15 mg/kg, using chloroquine as a control drug, because it has not been demonstrated that parasites that infect rodents have developed resistance against this drug. Our results showed that kramecyne decreases the expression of parasite proteins that participate in biological processes, such as invasion, cytoadherence, pathogenicity and energy metabolism. With these results, it is proposed that this compound has repercussions on the metabolism of the parasite and could be useful for use as an antimalarial.

Aleatory Trypanosoma cruzi Vertical Transmission in Chiapas, Mexico: A Case Report.

Congenital Chagas disease is considered a form of dispersion of Trypanosoma cruzi related to human migration from endemic, often rural to previously non-endemic urban areas. This fact increases the Chagas disease establishment risk inside of family members by vertical transmission pathway. Congenital Chagas disease cases in newborns could not identified by the health professional even in endemic regions. Here we present the first family cluster of Chagas disease cases from Chiapas: one of the most important endemic areas in South of Mexico, where vertical T. cruzi transmission incidence rate is ranged between 2% to 22% revealing an important public health problem. Two cases inside a family from Chiapas, México with positive antibodies against T. cruzi detected by ELISA are presented; one of them got the infection through vertical pathway. We think that congenital Chagas disease should not be ignored in a newborn born from an asymptomatic Chagas disease mother, who may transmit the parasite infection randomly.

Co-infection of Dengue, Zika and Chikungunya in a group of pregnant women from Tuxtla Gutiérrez, Chiapas: Preliminary data. 2019.

INTRODUCTION: Dengue, Zika and Chikungunya are RNA Arboviruses present in some areas of Mexico, mainly in the endemic state of Chiapas that is characterized by presence of the vector that transmit them and an ecology that favors high transmission. According to the national epidemiological surveillance system, Dengue has intensified since 2018 and outbreaks continue in various states while for Zika and Chikungunya a decrease in cases has been reported in recent years. The main objective of this study was to determine the incidence of Dengue, Zika and Chikungunya infections during pregnancy in the state of Chiapas. PRINCIPAL FINDINGS: The presence of previous and current infections and coinfections diagnosed by molecular (RT-PCR) and immunological (ELISA for IgG determination) techniques indicates a wide circulation of viruses in asymptomatic people, specifically in pregnant women showing that silent infections in dry season contributes to the preservation of viruses. CONCLUSIONS: From 136 studied samples, 27.7% tested positive for DENV, 8% for ZIKV and 24.1% for CHIKV by RTPCR and the values of IgG in sera show that 83.9% were positive for IgG antibodies against DENV, 65% against ZIKV and 59.1% against CHIKV. Results demonstrated presence of ZIKV and CHIKV, not detected by the epidemiological surveillance system, so the importance of establishing proactive epidemiological systems more strict, especially because these infections in pregnant women can cause severe health problems for newborn children.

Injection of mRNA isolated from trophozoites of Giardia intestinalis induces expression of three types of chloride currents in Xenopus laevis oocytes.

Giardia lamblia is one of the most important worldwide causes of intestinal infections, yet little is known about its cellular physiology, especially the diversity of ionic channels that this parasite expresses. In this work, we show that injection of mRNA isolated from trophozoites of Giardia, into Xenopus laevis oocytes, induces expression of three types of chloride currents (here referred to as ICl-G1, ICl-G2, and ICl-G3), which have different biophysical and pharmacological properties. ICl-G1 currents show inward rectification and voltage dependence are enhanced by hypotonicity, show a selectivity sequence of (I > Br > Cl > F), and are inhibited by NPPB, DIDS, SITS, 9AC, DPC, and Zinc. These findings suggest that ICl-G1 is the result of expression of chloride channels related to ClC2. ICl-G2 currents show outward rectification and are dependent of intracellular calcium, its selectivity sequence is (Cl > Br > I > F) and are inhibited by NPPB, DIDS, SITS, 9AC, DPC, niflumic acid, tannic acid, and benzbromarone. These findings suggest that they are produced by calcium dependent chloride channels (CaCC). The third type of currents (ICl-G3) appears only after a hypoosmotic challenge, and has similar properties to those described for ICl-swell, such as outward rectification, instant activation, and slow inactivation at large depolarizing voltages. They were blocked by NPPB, DIDS, 9AC, NIf, DCPIB, and tamoxifen. Our results indicate that Giardia intestinalis has at least three types of anion conductances.

Análisis proteómico de los productos de excreción-secreción de cuatro aislados de Trichinella spiralis obtenidos de hospederos accidentales / Proteomic analysis of the excretion-secretion products of four Trichinella spiralis isolates obtained from accidental hosts

Resumen: Introducción: Trichinella spiralis es un nemátodo tisular que se aloja en el músculo esquelético de humanos y otros mamíferos y causa una serie de alteraciones fisiológicas. Las proteínas de los productos de excreción-secreción de T. spiralis juegan un papel importante en la aparición y regulación de estas alteraciones. Sin embargo, aún no se conoce el efecto de estos productos en la infección e invasión del parásito al hospedero. Métodos: Mediante un análisis electroforético en una dimensión, Western blot y espectrometría de masas, se evaluaron las diferencias y similitudes entre proteínas antigénicas y de superficie de cuatro aislados de T. spiralis obtenidos de hospederos accidentales (perros) y la cepa de referencia aislada de cerdos (MSUS/MEX/91/CM). Resultados: Utilizando ontología de genes, se encontraron cinco proteínas exclusivas de los hospederos accidentales. Después del análisis, se encontró que estas proteínas forman parte de la matriz extracelular del parásito, cuentan con actividad catalítica y están implicadas en la adhesión a las células del hospedero. La actividad antigénica de las cuatro cepas aisladas de hospederos accidentales es idéntica a la reportada para T. spiralis, visualizándose el triplete antigénico característico de 43, 45 y 47 kDa. Conclusiones: Las proteínas exclusivas de los hospederos accidentales proveen información para entender el mecanismo de acción de este parásito para penetrar el músculo y evadir la respuesta inmune en el hospedero.

Abstract: Background: Trichinella spiralis is an intestinal and tissue nematode specific for mammalian skeletal muscle, causing a series of physiological alterations. T. spiralis excretory-secretion products play an important role in the appearance and regulation of these alterations. However, the effect of these products on the infection and invasion of the parasite to the host is unknown. Methods: Differences and similarities between antigenic proteins and surface proteins of four accidental hosts isolates (dogs) of T. spiralis and the reference strain isolated from pigs (MSUS/MEX/91/CM) were assessed by electrophoresis, western blot and mass spectrometry. Results: Using gene ontology, five proteins exclusive to the accidental hosts were analyzed. The results showed that these proteins are part of the extracellular matrix of the parasite, present catalytic activity, and bind to host cells. The antigenic activity the four strains showed the antigenic triplet characteristic of T. spiralis of 43, 45 and 47 kDa. Conclusions: Five proteins exclusive to dog isolates provided information to understand the mechanism of action of this parasite to penetrate the muscle and evade the immune response in the host.

[Proteomic analysis of the excretion-secretion products of four Trichinella spiralis isolates obtained from accidental hosts]. / Análisis proteómico de los productos de excreción-secreción de cuatro aislados de Trichinella spiralis obtenidos de hospederos accidentales.

BACKGROUND: Trichinella spiralis is an intestinal and tissue nematode specific for mammalian skeletal muscle, causing a series of physiological alterations. T. spiralis excretory-secretion products play an important role in the appearance and regulation of these alterations. However, the effect of these products on the infection and invasion of the parasite to the host is unknown. METHODS: Differences and similarities between antigenic proteins and surface proteins of four accidental hosts isolates (dogs) of T. spiralis and the reference strain isolated from pigs (MSUS/MEX/91/CM) were assessed by electrophoresis, western blot and mass spectrometry. RESULTS: Using gene ontology, five proteins exclusive to the accidental hosts were analyzed. The results showed that these proteins are part of the extracellular matrix of the parasite, present catalytic activity, and bind to host cells. The antigenic activity the four strains showed the antigenic triplet characteristic of T. spiralis of 43, 45 and 47 kDa. CONCLUSIONS: Five proteins exclusive to dog isolates provided information to understand the mechanism of action of this parasite to penetrate the muscle and evade the immune response in the host.

Antigiardial Effect of Kramecyne in Experimental Giardiasis.

A variety of drugs are used in giardiasis treatment with different levels of efficiency, presence of side effects, and even formation of resistant strains, so that it is important to search new only-one-dose treatments with high efficiency and less side effects. Kramecyne, an anti-inflammatory compound isolated from methanolic extract of Krameria cytisoides, does not present toxicity, even at doses of 5,000 mg/kg. The objective was to determine the antigiardial effect of kramecyne over Giardia intestinalis in vitro and in vivo and analyze the expression of genes ERK1, ERK2, and AK on kramecyne treated trophozoites by Real Time Polymerase Chain Reaction (RTPCR). The median lethal dose (LD50) was 40 µg/mL and no morphological changes were observed by staining with blue trypan and light microscopy; experimental gerbil infection was eliminated with 320 µg/Kg of weight. After treatment there were no differences between intestines from treated and untreated gerbils. Kramecyne did not have significant effect over ERK1 and AK, but there are differences in ERK2 expression (p = 0.04). Results show antigiardial activity of kramecyne; however the mode of action is still unclear and the evaluation of ultrastructural damage and expressed proteins is an alternative of study to understand the action mechanism.

[Maternal-fetal transmission of Trypanosoma cruzi, a health problem slightly studied in Mexico: case Chiapas]. / Transmisión materno-fetal de Trypanosoma cruzi, un problema de salud poco estudiado en México: caso Chiapas.

OBJECTIVE: To determine the Trypanosoma cruzi infection prevalence in 1125 pregnant women and the transmission frequency to their children from Tapachula and Palenque, Chiapas. MATERIALS AND METHODS: We determined the prevalence by serology tests and the transmission frequency by polymerase chain reaction (PCR) and T. cruzi reactivity capacity after 12 months. RESULTS: Total maternal infection prevalence were 23/1 125 (2.04%), 9/600 (1.5%) were from Tapachula and 14/525 (2.6%) from Palenque. The seropositive women were between 20 and 35 years old, 31.8% have Premature Rapture of Membrane and 9.1% have history of perinatal death. The total percentage of positive newborns by PCR was 9/23 (39.13%), out of those 2/9 (22.2%) are from Tapachula and 7/14 (50%) from Palenque. The Maternal Fetal transmission frequency was. 2/9 (22.2%) in Tapachula and 1/14 (7.14%) in Palenque, all positive infants were asynthomatic. CONCLUSION: The maternal-fetal transmission rate in Chiapas State is variable; the reason could be the maternal immunological status and T. cruzi strain.

Transmisión materno-fetal de Trypanosoma cruzi, un problema de salud poco estudiado en México: caso Chiapas / Maternal-fetal transmission of Trypanosoma cruzi, a health problem slightly studied in Mexico: case Chiapas

Resumen: Objetivo: Determinar la prevalencia de infección de Trypanosoma cruzi en 1 125 mujeres embarazadas y la frecuencia con que éstas la transmiten a sus hijos en Tapachula y Palenque, Chiapas. Material y métodos: Se determinó la prevalencia materna por serología, la frecuencia de transmisión por reacción en cadena de la polimerasa (PCR) y la capacidad reactiva a T. cruzi a los 12 meses de edad del niño. Resultados: La prevalencia materna total fue 23/1 125 (2.04%); 9/600 (1.5%) en Tapachula y 14/525 (2.6%) en Palenque. Las mujeres seropositivas tenían entre 20 y 35 años; 31.8% tuvo ruptura prematura de membrana (RPM) y 9.1% tuvo antecedentes de muerte perinatal. La frecuencia de transmisión materno-fetal en Tapachula fue de 2/9(22.2%) y en Palenque de 1/14(7.14%); todos los niños positivos eran asintomáticos. Conclusión: La tasa de transmisión materno-fetal en el estado de Chiapas es variable; la razón podría ser el estado inmunológico de la madre o la cepa de T. cruzi.

Abstract: Objective: To determine the Trypanosoma cruzi infection prevalence in 1125 pregnant women and the transmission frequency to their children from Tapachula and Palenque, Chiapas. Materials and methods: We determined the prevalence by serology tests and the transmission frequency by polymerase chain reaction (PCR) and T. cruzi reactivity capacity after 12 months. Results: Total maternal infection prevalence were 23/1 125 (2.04%), 9/600 (1.5%) were from Tapachula and 14/525 (2.6%) from Palenque. The seropositive women were between 20 and 35 years old, 31.8% have Premature Rapture of Membrane and 9.1% have history of perinatal death. The total percentage of positive newborns by PCR was 9/23 (39.13%), out of those 2/9 (22.2%) are from Tapachula and 7/14 (50%) from Palenque. The Maternal Fetal transmission frequency was. 2/9 (22.2%) in Tapachula and 1/14 (7.14%) in Palenque, all positive infants were asynthomatic. Conclusion: The maternal-fetal transmission rate in Chiapas State is variable; the reason could be the maternal immunological status and T. cruzi strain.

In vitro anti-Giardia lamblia activity of 2-aryl-3-hydroxymethyl imidazo[1,2-a]pyridines and -pyrimidines, individually and in combination with albendazole.

Giardiasis is a major diarrheal disease found throughout the world, the causative agent being the flagellate protozoan Giardia intestinalis. Infection is more common in children than in adults. The appearance of drug resistance has complicated the treatment of several parasitic diseases, including giardiasis. Thus, the aim of this investigation was to make an in vitro evaluation of the antigiardia response of synthetic derivatives 2-aryl-3-hydroxymethylimidazo[1,2-a]pyridines 1 and -pyrimidines 2 against trophozoites of Giardia lamblia WB, in comparison with the reference drug, albendazole. Additionally, the synergistic action of albendazole in combination with each of the most active 2-aryl-3-hydroxymethyl imidazo[1,2-a]pyridines and pyrimidines was also assessed. Based on the IC50 values obtained, the best anti-Giardia activity was provided by the 3-hydroxymethyl-4-fluorophenylimidazo[1,2-a]pyrimidine derivative 2c and the corresponding imidazo[1,2-a]pyrimidine with the p-tolyl substituent 2d, followed by 2a and 2b. These four compounds showed effectiveness at a concentration similar to that of albendazole. Regarding synergism, the IC50 of the combination of albendazole with 2a, 2b or 2c gave the best anti-Giardia action, showing greater efficacy than albendazole alone. Hence, G. lamblia WB showed high susceptibility to some 2-aryl-3-hydroxymethyl imidazo[1,2-a] pyrimidines, which acted synergistically when used in combination with albendazole.

The MAK16 gene of Entamoeba histolytica and its identification in isolates from patients.

To identify sequences of Entamoeba histolytica associated with the development of amebic liver abscess (ALA) in hamsters, subtractive hybridization of cDNA from E. histolytica HM-1:IMSS under 2 growth conditions was performed: 1) cultured in axenic medium and 2) isolated from experimental ALA in hamsters. For this procedure, 6 sequences were obtained. Of these sequences, the mak16 gene was selected for amplification in 29 cultures of E. histolytica isolated from the feces of 10 patients with intestinal symptoms and 19 asymptomatic patients. Only 5 of the 10 isolates obtained from symptomatic patients developed ALA and amplified the mak16 gene, whereas the 19 isolates from asymptomatic patients did not amplify the mak16 gene nor did they develop ALA. Based on the results of Fisher's exact test (P<0.001), an association was inferred between the presence of the mak16 gene of E. histolytica and the ability to develop ALA in hamsters and with the patient's symptoms (P=0.02). The amplification of the mak16 gene suggests that it is an important gene in E. histolytica because it was present in the isolates from hamsters that developed liver damage.

Identificación de Microsporidium spp en pacientes con diagnóstico de leucemia linfoblástica aguda / Identification of Microsporidium spp in patients with acute lymphoblastic leukemia

Introducción. El grupo Microsporidia incluye parásitos intracelulares obligados que afectan varios huéspedes. Los géneros que infectan humanos son Enterocytozoon y Encephalitozoon . Microsporidia es considerado un organismo oportunista en pacientes inmunocomprometidos e inmunocompetentes, que se ha convertido en un importante problema de salud pública. El objetivo del trabajo fue identificar Microsporidium spp en muestras fecales de pacientes con leucemia linfoblástica aguda, en diferente etapa de su tratamiento. Métodos. Se analizaron 77 muestras de niños con diagnóstico de leucemia linfoblástica aguda menores de 12 años, por análisis coproparasitoscópico de Faust. Las muestras se tiñeron con Ziehl Nielsen, tricrómica de Masson y calcoflúor. Finalmente se realizó la amplificación por PCR del gen ribosomal. Resultados. De los pacientes analizados, 16/77 (20.77%) resultaron positivos para Microsporidia, independientemente de que presentaran diarrea. El PCR fue más efectivo para la identificación que el análisis microscópico de las muestras teñidas. Conclusiones. Este trabajo enfatiza la importancia de la microsporidiosis como infección emergente en pacientes con leucemia linfoblástica aguda bajo tratamiento quimioterapéutico, incrementando las complicaciones clínicas adicionales a la leucemia.

Background. The phylum Microsporidia includes obligate intracellular parasites that affect several hosts. The most frequent genera to affect humans are Enterocytozoon and Encephalitozoon. Microsporidium is considered an opportunistic parasite in both immunocompromised and immunocompetent patients and have become an important health problem. The aim of this study was to identify Microsporidium spp in fecal samples of patients with acute lymphoblastic leukemia (ALL), with and without diarrhea, at different treatment stages. Methods. Seventy seven samples from children <12 years old with diagnosis of ALL were analyzed by the Faust coproparasitoscopic method, Ziehl-Nielsen, trichrome and calcofluor staining methods and polymerase chain reaction. Results. Results showed that 16/77 (20.77%) children presented Microsporidium and there was no relationship between microsporidial infection and diarrhea. Polymerase chain reaction was more effective than analysis by light microscopy of staining samples in the identification of the parasite. Conclusions. This work emphasizes the importance of microsporidiosis as an emerging infection in patients with ALL undergoing chemotherapy, increasing the additional clinical complications of leukemia.

Voltage-dependent potassium currents expressed in Xenopus laevis oocytes after injection of mRNA isolated from trophozoites of Giardia lamblia (strain Portland-1).

Despite its importance as a health problem issue, almost nothing is known about the membrane physiology of Giardia lamblia and practically there exist no information so far regarding the variety and properties of ion channels that this protozoan parasite possesses. To address this subject we resorted to an indirect method, consisting in the injection of mRNA and further characterization of ion currents in Xenopus oocytes. In this work, we show that oocytes injected with mRNA isolated from cultured trophozoites of G. lamblia, strain Portland-1 express novel potassium currents that appear over the second day after injection and show time- and voltage-dependent activation followed by a slow inactivation. They start activating at -90 mV, with V1/2 of -30 mV; its time constant of activation (at +60 mV) is 0.11 sec, whereas that of inactivation is 1.92 sec, V1/2 = -44.6 mV. Such K currents were effectively blocked by K channel blockers TEA and 4AP, as well as Ba(2+), quinine, quinidine, charybdotoxin, dendrotoxin-1, capsaicin, margatoxin, and diltiazem. These results suggest that such currents are the result of expression of Giardia's voltage-gated K channels heterologously expressed in Xenopus laevis oocytes.

Molecular similarities and differences between Trichinella spp., isolated from canine skeletal muscle in Zacatecas, Mexico.

Four different isolates of Trichinella spp. (Z1, Z2, Z3, and Z4) obtained from the skeletal muscle of street dogs in the state of Zacatecas, Mexico were serial passaged in Wistar rats; infective larvae from the skeletal muscle of the rats were collected and frozen in liquid nitrogen. After centrifugation, DNA was extracted and the 5SRNAr and IsRNAr genes were amplified. The isolates were identified by the size of the amplified products from the 5SRNAr and IsRNAr genes (750 and 290 bp, respectively). The amplicons obtained by PCR were sequenced, aligned, and compared to the reference strain Trichinella spiralis MSUS/MEX/91//EM isolated from pigs. Based on our results, we determined that the Trichinella isolates from canine (Z1-Z4) belonged to the T. spiralis species and had 83% identity with the reference strain. The phylogenetic tree constructed from the sequences showed differences between the isolates from pig and dog. These genetic differences may be related to the immune response of the host or the pathogenicity of the isolates. Therefore, these findings have important epidemiological and public health implications.

Changes in beta-giardin sequence of Giardia intestinalis sensitive and resistant to albendazole strains.

Giardia intestinalis can develop resistance to albendazole, although the molecular mechanism is not understood. The aim of this study was to investigate the differences and permanent mutation in the beta-giardin gene of G. intestinalis strains: sensitive, resistant, or recovered-resistance to albendazole. The beta-giardin gene was amplified by nested polymerase chain reaction. The IC(50) values varied from 0.29 to 0.38 microg/mL for strains sensitive to albendazole. For resistant strains, the IC(50) range was 1.31-2.12 microg/mL. Recovered-sensitivity albendazole strains' IC(50) values were 0.33-0.49 microg/mL, and for strains with recovered-resistance, the IC(50) was 1.42-2.74 microg/mL. beta-giardin amplicon (720 bp) was sequenced and analysis sequence revealed several amino acid mutations from resistant and recovered-sensitive strains of G. intestinalis. Most of the mutations were located in the ROD domain of beta-giardin with a change from the sequence "TIARERA" in sensitive strains instead "IDRPRE" in resistant strains. A comparative sequence analysis in resistant, recovered-sensitive, and resistant-recovered strains revealed permanent mutation. This is the first report of combinatorial serine-proline-arginine repeats in the ROD domain of beta-giardin, whereas such repeats have been reported previously in the HEAD domain of SF-assemblin proteins. This is the first time that the resistance to albendazole correlates with genetics but it is not necessarily caused by mutations in the beta-giardin gene of G. intestinalis.

Frequency of Giardia intestinalis assemblages isolated from dogs and humans in a community from Culiacan, Sinaloa, Mexico using beta-giardin restriction gene.

The assemblage of 37 Giardia intestinalis samples was determined, 19 obtained from puppy feces, 12 from stools of different human subjects under 3 years of age and 6 from axenic culture. The assemblages were classified according to the restriction pattern of beta-giardin gene with Hae III enzyme. Results showed that dog assemblages were grouped AI (52.6%) and AII (47.4%), while 41.7% human samples belonged to genotype AI and 58.3% to genotype AII. All axenic cultures belonged to assemblage AI; types AI and AII were both found in dog and human feces by Hae III restriction enzyme assay, suggesting a similarity between human and dog parasites. These results suggest that domestic animals infected with Giardia could produce cysts potentially infective for humans.

Classification of Giardia intestinalis isolates by multiple polymerase chain reaction (multiplex).

Agarose gel electrophoresis of gdh gene fragments, amplified by Multiplex, was used to classify the assemblage of 24 Giardia isolates obtained from axenic cultures, children's stools, and feces of puppies from different dog breeds. Isolates were compared with seven reference strains of Giardia intestinalis. The results showed that 22/24 isolates (91%) belonged to assemblage A and could be further subclassified as assemblage A1 (18/22, 81%) and assemblage A2 (4/22, 19%). One sample revealed a mixture of A1/A2 genotypes, and another was assemblage G, indicating mixed infections by different strains in the same host, and an association with the assemblage reported in animals. The procedure described is useful to determine the Giardia genotype that parasitizes each host to conduct epidemiological studies assessing the close association between human- and animal-infecting strains and to monitor the adaptability of animal strains to humans.

Protein profile analysis from advanced third-stage larvae (AdvL3) and adult worms of Gnathostoma binucleatum (Nematoda: Spirurida).

Proteins from crude extracts of advanced third-stage larvae and adult Gnathostoma binucleatum nematode worms showed protein profiles in SDS-PAGE analysis similar to Echinococcus granulosus, Trichinella spiralis, Dipylidium caninum, Ancylostoma caninum, Ascaris lumbricoides and Toxocara canis. The immunoblot analysis of the human serum infected or suspected to be infected with G. binucleatum using the total larvae extract recognized the 40, 60, 80 and 115 kDa proteins and using the total adult worm extract recognized only the 80 and 115 kDa proteins. However, the 115 kDa protein showed cross-reactions with A. caninum, A. lumbricoides, T. canis and D. caninum with human serum positive to gnathostomosis, while the 40 kDa protein was only recognized with the G. binucleatum total larvae extract. The results obtained suggest that the use of antigens from the advanced third-stage larvae of the parasite were best recognized for immunodiagnosis of gnathostomosis.

Entamoeba histolytica sequences and their relationship with experimental liver abscesses in hamsters.

The purpose of the present paper was to analyse the association between sequences of Entamoeba histolytica and their relationship with the development of hepatic abscesses in hamsters, using a complementary DNA library for E. histolytica. From the sequences obtained, we designed oligonucleotides for amplification by PCR. Trophozoites were isolated from faeces of 11 patients in whom cysts from E. histolytica were identified, and these trophozoites were then subjected to monoaxenic culture. Then 1 x 10(5) trophozoites were inoculated into hamster liver, with three hamsters used for every culture. Sequences were obtained for ubiquitin, lectin surface precursor, replication factor MCM3 and surface antigen. The associations between sequences and hepatic abscesses were: 11/11 for ubiquitin, 9/11 for lectin precursor, 4/11 for replication factor and 1/11 for surface antigen. These results suggest that ubiquitin could be an important protein involved in the mechanism of E. histolytica invasion.