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The objective of this study was to identify genetic single nucleotide polymorphisms (SNPs) in the coding region of the KRTAP11.1 gene from the Suri alpaca population from Puno, Peru. Genomic DNA (n = 119) was used to amplify the 636 bp fragment of the coding region from the KRTAP11.1 gene. The polymerase chain reaction products of the KRTAP11.1 gene were sequenced by both strands and polymorphism type SNPs were identified. All SNPs (n = 3) were in Hardy-Weinberg equilibrium and had a high level of genotypic linkage disequilibrium with the presence of two haplotypes. All genetic polymorphisms generate non-synonymous amino acids changes: SNP 110 T>G (Ser>Ala), SNP 363 T>G (Phe>Cys) and SNP 375 A>C (Tyr>Ser). The results indicated the presence of moderate genetic diversity in the coding region of the KRTAP11.1 gene from the Suri alpaca population.

Objetivo. Producir anticuerpos recombinantes de cadena única de alpaca que se unan con alta afinidad y especificidad al antígeno excretado-secretado (ES) de Fasciola hepatica para el desarrollo de tecnologías nuevas de diagnóstico de fascioliasis humana y animal. Materiales y métodos. Se ha construido una genoteca de cADNde los dominios variables de anticuerpos de cadena única pesada, conocidos como VHH, a partir de células mononucleares de sangre periférica de una alpaca inmunizada con el antígeno ES de F. hepatica. La genoteca fue tamizada con el antígeno ES por despliegue diferencial de fagos (phage display), seleccionando diez VHH que se unen específicamente a ES. El VHH anti ES fue clonado en un vector de expresión, la proteína recombinante (VHH-ES1) de 15,3 kDa fue producida por fermentación en E. coli y purificada a homogeneidad por cromatografía de afinidad. La unión del VHH-ES1 al antígeno ES fue evaluada por ELISA usando VHH-ES1 como anticuerpo de captura, antisuero policlonal anti-ES de conejo y conjugado anti IgG de conejo con peróxidasa de rábano. Resultados. Se ha identificado y producido un VHH-ES1 recombinante que se une al antígeno ES (VHH-ES1) que correspondía a un anticuerpo de la subclase IgG2 de bisagra larga. La unión del anticuerpo VHH-ES1 al antígeno muestra linealidad respecto a la concentración de ES en el rango de 50-5000 ng/mL y el valor límite de detección del antígeno está en el rango de 30-170 ng/mL de ES (R²=0,99). Conclusión. El VHH-ES1 se une con afinidad y especificidad al antígeno ES de F. hepatica y es un anticuerpo promisorio a evaluar para el desarrollo de nuevas tecnologías de diagnóstico de fascioliasis. Objectives. To produce recombinant single-chain antibodies from alpaca that will bind to the excreted-secreted (ES) Fasciola hepatica antigen with high affinity and specificity, so as to develop new diagnostic technologies of human and animal fascioliasis. Materials and Methods. A gene bank of DNA of the variable dominions of heavy single-chain antibodies (VHH) has been created, based on mononuclear cells of peripheral blood of an alpaca immunized with the ES antigen of F. hepatica. The gene bank was screened with the ES antigen by differential phage display, selecting ten VHH that bind specifically to ES. The anti-ES VHH was cloned in an expression vector, the recombinant protein (VHH-ES1) of 15.3 kDa was produced by fermentation in E. coli and purified to homogeneity by affinity chromatography. The binding of VHH-ES1 to the ES antigen was evaluated by ELISA using VHH-ES1 as capture antibody, policlonal anti-ES serum of rabbit and conjugated rabbit anti IgG with radish peroxidase. Results. A VHH that binds to the ES antigen (VHH-ES1) has been identified through differential phage display and produced by fermentation in E. coli; this corresponds to an antibody of the long-hinge IgG2 subclass. The binding of the VHH-ES1 antibody to the antigen shows linearity with respect to the concentration of ES in the 50-5,000 ng/mL range and the limit of detection value of the antigen is in the 30-170 ng/mL range of ES (R2=0.99). Conclusions. The VHH-ES1 binds with affinity and specificity to the ES antigen of F. hepatica and is a promissory antibody to be assessed for the development of new fascioliasis diagnostic technologies.

Power converter circuits with grid interaction are gaining increasing attention especially with emerging renewable energy technologies. Passive filters are inevitable for power electronic devices with grid coupling. In this context LCL filter topologies are a recognized cost and space reducing alternative to simple single coil solutions for grid current harmonic attenuation. Since this type of filter represents a third order plant, a simple PI controller would cause instabilities once applied to control the grid current of the undamped filter. A common approach is the addition of a so called active damping to the PI-controller. Documented solutions refer to the feedback of specific state variables and/or implementation of additional filters. The corresponding parameter determination turns out to be generally quite complicated as it often deals with several variables. This article provides practicing engineers with a simple and effective recipe to determine just one active damping parameter for PI current controllers applied to three phase power converters with grid connection via LCL filters. A simple and generally valid formula is derived via the Hurwitz stability criterion that determines a safe operating area of this single parameter.

To produce recombinant single-chain antibodies from alpaca that will bind to the excreted-secreted (ES) Fasciola hepatica antigen with high affinity and specificity, so as to develop new diagnostic technologies of human and animal fascioliasis.A gene bank of DNA of the variable dominions of heavy single-chain antibodies (VHH) has been created, based on mononuclear cells of peripheral blood of an alpaca immunized with the ES antigen of F. hepatica. The gene bank was screened with the ES antigen by differential phage display, selecting ten VHH that bind specifically to ES. The anti-ES VHH was cloned in an expression vector, the recombinant protein (VHH-ES1) of 15.3 kDa was produced by fermentation in E. coli and purified to homogeneity by affinity chromatography. The binding of VHH-ES1 to the ES antigen was evaluated by ELISA using VHH-ES1 as capture antibody, policlonal anti-ES serum of rabbit and conjugated rabbit anti IgG with radish peroxidase.A VHH that binds to the ES antigen (VHH-ES1) has been identified through differential phage display and produced by fermentation in E. coli; this corresponds to an antibody of the long-hinge IgG2 subclass. The binding of the VHH-ES1 antibody to the antigen shows linearity with respect to the concentration of ES in the 50-5,000 ng/mL range and the limit of detection value of the antigen is in the 30-170 ng/mL range of ES (R2=0.99).The VHH-ES1 binds with affinity and specificity to the ES antigen of F. hepatica and is a promissory antibody to be assessed for the development of new fascioliasis diagnostic technologies.Producir anticuerpos recombinantes de cadena única de alpaca que se unan con alta afinidad y especificidad al antígeno excretado-secretado (ES) de Fasciola hepatica para el desarrollo de tecnologías nuevas de diagnóstico de fascioliasis humana y animal.Se ha construido una genoteca de cADNde los dominios variables de anticuerpos de cadena única pesada, conocidos como VHH, a partir de células mononucleares de sangre periférica de una alpaca inmunizada con el antígeno ES de F. hepatica. La genoteca fue tamizada con el antígeno ES por despliegue diferencial de fagos (phage display), seleccionando diez VHH que se unen específicamente a ES. El VHH anti ES fue clonado en un vector de expresión, la proteína recombinante (VHH-ES1) de 15,3 kDa fue producida por fermentación en E. coli y purificada a homogeneidad por cromatografía de afinidad. La unión del VHH-ES1 al antígeno ES fue evaluada por ELISA usando VHH-ES1 como anticuerpo de captura, antisuero policlonal anti-ES de conejo y conjugado anti IgG de conejo con peróxidasa de rábano.Se ha identificado y producido un VHH-ES1 recombinante que se une al antígeno ES (VHH-ES1) que correspondía a un anticuerpo de la subclase IgG2 de bisagra larga. La unión del anticuerpo VHH-ES1 al antígeno muestra linealidad respecto a la concentración de ES en el rango de 50-5000 ng/mL y el valor límite de detección del antígeno está en el rango de 30-170 ng/mL de ES (R2=0,99).El VHH-ES1 se une con afinidad y especificidad al antígeno ES de F. hepatica y es un anticuerpo promisorio a evaluar para el desarrollo de nuevas tecnologías de diagnóstico de fascioliasis.

International audience; Introduction: The etiology of several hepatocellular carcinoma (HCC) cases remains largely unknown. Although Fasciola hepatica has been associated with liver fibrosis in Latin America, it has not yet been associated with HCC. This study aimed to determine the existence of specific IgG antibodies against F. hepatica in the serum samples of HCC patients. Methods: In total, 13 serum samples from 13 HCC patients were screened using Fas2-ELISA. Results: Fas2-ELISA demonstrated negative results in all HCC patients included in this study. Conclusions: The pre-existence of F. hepatica infection in HCC patients needs to be further investigated in epidemiological and experimental studies.

Background Fascioliasis is an infectious disease caused by parasites Fasciola hepatica and F. gigantica. Humans are infected by the consumption of vegetables and water contaminated with the infective form of the parasite. Materials and methods In this study, an IgM-ELISA with the cysteine proteinase Fas2 antigen was evaluated with sera from 76 patients infected with F. hepatica, 24 patients with other parasite infections and 84 healthy volunteers. Results IgM-ELISA resulted in 43% positives in F. hepatica patients with positive serology to Fas2-ELISA, but no positives resulted from testing healthy volunteers and individuals infected with other parasites. The IgM-ELISA diagnostic parameters showed a sensitivity of 43.4% (95% CI 0.321-0.553), specificity of 100% (95% CI 0.957-1), and no cross-reactivity with other parasitic infection. Interference by rheumatoid factor in the IgM immunoassay was controlled by treating sera with rheumatoid factor absorbent before testing. Conclusions Fas2 antigen is detected by circulating IgM in patients infected with F. hepatica and IgM-ELISA using Fas2 appears as a specific immunoassay to detect the acute phase of the acute phase of F. hepatica infection in humans.

Genetic tagging is the unique identification of individuals by their DNA profile. This technique is well established in mammals, but it has not yet been widely adopted for birds. Extraction methods for minute amounts of DNA even enable the use of genetic tagging from non-invasive samples, like hair, scat, or feather. In this study, we evaluate the potential for non-invasive genetic tagging by using molted feathers of two sympatric macaw species in the Peruvian Amazon. Correct species identification is critical when relying on feathers for genetic analysis, so we describe multilocus methods for species identification. We evaluate the quality of naturally shed macaw feathers in tropical environmental conditions and present new primers for molecular sexing on the feather samples. We successfully validated 11 microsatellite markers for use in genetic tagging studies on large macaws and confirmed that DNA from blood and feather samples yields equivalent population genetic patterns. The techniques described here can be implemented for other birds with higher conservation concern.

Fascioliasis is a zoonotic parasitic disease caused by Fasciola hepatica and Fasciola gigantica. Of both species, F. hepatica is the only one described in the Americas, mainly transmitted by lymnaeid snail vectors of the Galba/. Fossaria group. Human fascioliasis endemic areas are mainly located in high altitude areas of Andean countries. Given the necessity to characterize F. hepatica populations involved, the phenotypic features of fasciolid adults infecting sheep present in human fascioliasis endemic areas were analysed in the Cajamarca Valley and Mantaro Valley (valley transmission patterns) and the northern Bolivian Altiplano (altiplanic transmission pattern). A computer image analysis system (CIAS) was applied on the basis of standardized measurements. The aforementioned highland populations were compared to standard lowland natural and experimental populations of European origin. Liver fluke size was studied by multivariate analyses. Two phenotypic patterns could be distinguished in F. hepatica adult size: the valley pattern (Cajamarca and Mantaro, Peru) and the altiplanic pattern (northern Altiplano, Bolivia). Results showed that the Andean valley population and European standard populations presented a phenotypic homogeneity. The Altiplano population showed a large size range with a pronouncedly lower minimum size indicating that uterus gravidity is reached at a smaller size than in valley populations. The results of this study demonstrate that there is no apparent relationship between the shape of fasciolid adults with regard to altitudinal difference or geographical origin and that allometry-free shape appears as a more stable trait than size in fasciolid species. Results are analysed in terms of intensity/crowding effect aspects and permanent/seasonal transmission characteristics.

This research was supported by funds from the Rufford Small Grant Foundation, Loro Parque Foundation, and The Australian National University.

Racial variation, twin studies, segregation analyses, linkage and association studies all suggest that genetic factors play an important role in predisposition to tuberculosis. Many previous studies have been performed with pulmonary TB patients, as the most prevalent form of clinical TB (nearly 95%), and very few of them have considered extrapulmonary TB. The present study evaluates the effects of variation in eight candidate genes (LTA, TNF, IL1B, IL1RN, IL10, TGFB1, TIRAP and P2X7) with pulmonary, pleural, miliary and other extrapulmonary forms of TB in a Peruvian population from the North of Lima. 626 TB cases and 513 healthy controls were enrolled in this study. LTA(+368) and IL10(-592) were associated with different clinical forms of TB (P

The prevalence of Fasciola hepatica infection, in endemic countries, in patients with established cirrhosis is unknown. We hypothesized that, in endemic countries, the presence of fascioliasis may be detected in a serum pool of cirrhotic patients. Forty-four previously stored serum samples of patients with established liver cirrhosis, in the Hospital Nacional Cayetano Heredia in Lima, Peru, were collected from 1998 to 2003 and assessed for hepatitis B, C and fascioliasis antibodies (Fas2 ELISA). Hepatitis B surface antigen (HBsAg) was positive in 8.8% (n = 34), hepatitis B core antibody (anti-HBc) in 32.5% (n = 34), hepatitis C antibodies (anti-HCV) in 9.1% (n = 33), and 9.1% (n = 44) were Fas2 ELISA positive. This disease is an example of an emerging tropical infection which can be present in chronic liver diseases, requiring greater clinician awareness especially in endemic rural areas. Further clinical studies are warranted.

We conducted a case-control study to clarify the risk factors for human fascioliasis in the Northern Peruvian Altiplano. Cases (n=61) were individuals who were diagnosed as having chronic fascioliasis by faecal and/or serologic (Fas2-ELISA) studies. Controls (n=61) had negative serologic and coprological results for Fasciola hepatica. We identified an association between fascioliasis and four variables (40 analysed): the habit of drinking alfalfa juice (OR=4.5; 95% CI 1.8-11.1; P

High prevalence rates of human fascioliasis have been described in several regions of Peru. We surveyed 20 families in an endemic area of Peru in order to determine the proportion of infection with F. hepatica in relatives of diagnosed subjects and in order to identify associated risk factors. The study included feces and blood samples of 93 subjects. Ages ranged from one to 53 (mean = 18.6; SD = 14.2). The overall prevalence of fascioliasis by fecal examinations was 33.3% (n = 83) and by serology, 51.9% (n = 86). The prevalence in age group I (< 19 years old) by coprological and serological tests was 61.4% and 75.9%, respectively; in group II (>; 19 years old) 15.4% and 37.5%. The main associated risk factor with fascioliasis was eating salads (OR = 3.29, CI = 1.2 - 9.0, p = 0.02). In conclusion, human fascioliasis is highly prevalent in the relatives of index cases and the most significant risk factor of acquiring fascioliasis in the family is eating salads in endemic areas. Altas tasas de fasciolosis humana han sido descritas en varias regiones del Perú. Estudiamos 20 familias en una área endémica del Perú para determinar la proporción de infección con F. hepatica en los familiares de los sujetos diagnosticados y para identificar factores de riesgo asociados. El estudio incluyó un total de 93 sujetos, quienes contribuyeron con muestras de heces y sangre. Las edades comprendieron desde 1 a 53 años (media = 18.6; DS = 14.2). La prevalencia general de fasciolosis por exámenes de heces fue 33.3% (n = 83) y por serología, 51.9% (n = 86). La prevalencia en el grupo de edad I (< 19 años de edad) por pruebas coprológicas y serológicas fueron 61.4% y 75.9%, respectivamente; en el grupo II (>; 19 años de edad) 15.4% y 37.5%. El principal factor de riesgo asociado con fasciolosis fue comer ensaladas (OR = 3.29, IC = 1.2-9.0, p = 0.02). En conclusión, la fasciolosis humana es altamente prevalente en familiares de los casos índices y el factor de riesgo más significante para adquirirla en la familia es comer ensaladas en las áreas endémicas.

Circulating antibody against Fasciola hepatica antigens was determined by enzyme-linked immunosorbent assay (ELISA) and immunoelectrophoresis in alpacas naturally exposed to F. hepatica. Serological assay parameters were established by using sera from eight infected animals and seven controls with no record of this parasitic infection. Excretory–secretory (ES-) products, Fas1- and Fas2-ELISA were used to survey 307 alpacas from a F. hepatica endemic area in the Peruvian Andes. Seroprevalence of F. hepatica infection varied from 56.7, 64.8 and 66.8% measured by Fas1-, Fas2- and ES-ELISA, respectively. The sensitivity for ES-ELISA was 95%, corresponding Fas1- and Fas2-ELISA sensitivity values were 90 and 95%. In this population, 7% of animals were positive for F. hepatica eggs in faeces, other parasites detected were Trichuris sp. (40%), Nematodirus sp. (34.6%), Lamanema sp. (12.8%) and Eimeria sp. (11.8%). The results show that F. hepatica infected animals elicit circulating antibodies against ES, Fas1 and Fas2. Fas2-ELISA may be proposed as a sensitive assay for the immunodiagnosis of fasciolosis in alpacas.

Espinoza, Jose R., Giancarlo Alvarez, Fabiola León-Velarde, Hugo F. Ju Preciado, Jose-Luis Macarlupu, Maria Rivera-Ch, Jorge Rodriguez, Judith Favier, Anne-Paule Gimenez-Roqueplo, and Jean-Paul Richalet. Vascular endothelial growth factor-A is associated with chronic mountain sickness in Andean population. High Alt Med Biol. 15:146–154, 2014.—A study of chronic mountain sickness (CMS) with a candidate gene—vascular endothelial growth factor A (VEGFA)—was carried out in a Peruvian population living at high altitude in Cerro de Pasco (4380 m). The study was performed by genotyping of 11 tag SNPs encompassing 2.2 kb of region of VEGFA gene in patients with a diagnosis of CMS (n=131; 49.1±12.7 years old) and unrelated healthy controls (n=84; 47.2±13.4 years old). The VEGFA tag SNP rs3025033 was found associated with CMS (p0.36, p

A 58-year-old man presented with an asymptomatic 5cm left anterior cervical tumor that had been present for over a year. Histological analysis revealed chronic inflammation and granuloma with giant cells surrounding Fasciola hepatica eggs. Fas2 antigen was detected in the tissue by immunohistochemistry. Serology for F. hepatica was positive, but stool examinations were negative and eosinophil count was normal. A month later, the tumor recurred, and an adult parasite was found during excision. The patient received triclabendazole, and after 6 months serology was negative. Erratic localization of F. hepatica reveals a pathology involving chronic inflammation caused by a sexually mature parasite, although according to theory only immature parasites are located in ectopic lesions.

Fasciola hepatica adult worm cysteine proteinases were active-site, affinity radio-labeled with benzyloxicarbonyl-L-tyrosine-L-alanine diazomethylketone (Z-Tyr125I-Ala-CHN2). Sera from patients with fascioliasis and from rabbits experimentally infected with F. hepatica immunoprecipitated the radiolabeled parasite cysteine proteinases in immunoelectrophoresis assays. Two purified antigens were identified as part of the complex mosaic of antigens immunoprecipitated by the sera of infected patients. These antigens (Fas1 and Fas2) have been shown to be an important part of the Fharc2 precipitin band used for serologic diagnosis in humans and cattle. They showed cysteine proteinase activity with different proteolytic specificities and partial identity in double immunodiffusion assays. The results obtained in this work show that the Fas1 and Fas2 antigens are sensitive and specific antigens for diagnosis of this serious helminthic disease in humans and other susceptible hosts.

Genetic diversity within and between tandemly arrayed copies of the Gp63 gene occurs in laboratory isolates of Leishmania spp., but the extent to which this represents natural genetic diversity has not been assessed. Here, the Gp63 locus is examined in 58 fresh isolates of L. peruviana , and clones derived from them, collected throughout the Peruvian Andes. Extensive polymorphism is observed, both in size of Gp63 containing chromosomes, and for restriction-fragment-length polymorphisms (RFLPs) at the Gp63 locus. All clones within an isolate are identical, including those with two distinct Gp63-hybridising chromosomal-sized pulsed-field gel electrophoresis (PFGE) bands, consistent with diploidy but with size differences in homologous chromosomes. For RFLP analysis, three enzymes were selected to cut within the coding region ( Pst I), in the intergenic region ( Sal I) and outside ( Eco RI) the Gp63 gene cluster. Pst I gave identical banding patterns across all isolates/clones. For Eco RI and Sal I, all clones within an isolate were identical, but isolates were polymorphic for fragments at 13 (2–30 kb) and 8 (2.6–8.8 kb) different molecular mass locations generating 19 and 16 distinct RFLP patterns or genotypes for each enzyme, respectively. Eco RI restriction patterns, analysed by PFGE, were consistent with the presence of two clusters of Gp63 genes on each homologous chromosome, one contained within Eco RI fragments large enough to carry from 3 to 10 copies of the Gp63 gene, the second on fragments which could carry 1 or 2 copies of the gene. Sal I patterns indicated variable restriction sites within clusters, but not within every intergenic region. A hierarchical analysis of variance of allele frequencies, expressed in terms of Wright's F -statistic, indicated significant barriers to gene flow at all levels, valleys within regions (north/south), villages within valleys, and individuals within villages. This extreme polymorphism at the Gp63 locus of L. peruviana demonstrates the great potential for generation of genetic diversity in parasite populations.

This paper presents the first evaluation of the molecular epidemiology of Mycobacterium tuberculosis in Peru. We characterised 323 isolates using spoligotyping and mycobacterial interspersed repetitive units variable number tandem repeats (MIRU-VNTR) typing. We aimed to determine the levels of genetic diversity and genetic differentiation among and within Peruvian isolates and the epidemiological factors which may be driving patterns of population structure and evolution of M. tuberculosis in Peru. Our results compared to the fourth international spoligotyping database (SpolDB4) and MIRU-VNTRplus, show that the main M. tuberculosis families present are Latin American-Mediterranean, Haarlem, T, and Beijing. Bayesian clustering recovered 15 groups in the Peruvian M. tuberculosis isolates, among which two were composed mainly of orphans, implying the presence of native “Peruvian” strains not previously reported. Variable levels of association with drug resistance were observed, with Beijing genotypes not showing any association with multidrug resistance, while in other groups MIRU-VNTR loci 2, 23, 31, and 40 were found to be associated with the multidrug-resistant tuberculosis (MDR-TB) phenotype, suggesting that a linkage disequibrium between these MIRU and drug resistance loci may be present. Genetic differentiation was present among drug resistant and sensitive strains. Ethambutol appeared to be the main driver of differentiation, suggesting that strong selection pressure could have been exerted by drug treatment in Peru over recent years.

Background Emergence of human fascioliasis prompted a worldwide control initiative including a pilot study in a few countries. Two hyperendemic areas were chosen: Huacullani, Northern Altiplano, Bolivia, representing the Altiplanic transmission pattern with high prevalences and intensities; Cajamarca valley, Peru, representing the valley pattern with high prevalences but low intensities. Coprological sample collection, transport and study procedures were analyzed to improve individual diagnosis and subsequent treatments and surveillance activities. Therefore, a coproantigen-detection technique (MM3-COPRO ELISA) was evaluated, using classical techniques for egg detection for comparison. Methodology and Findings A total of 436 and 362 stool samples from schoolchildren of Huacullani and Cajamarca, respectively, were used. Positive samples from Huacullani were 24.77% using the MM3-COPRO technique, and 21.56% using Kato-Katz. Positive samples from Cajamarca were 11.05% using MM3-COPRO, and 5.24% using rapid sedimentation and Kato-Katz. In Huacullani, using Kato-Katz as gold standard, sensitivity and specificity were 94.68% and 98.48%, respectively, and using Kato-Katz and COPRO-ELISA test together, they were 95.68% and 100%. In Cajamarca, using rapid sedimentation and Kato-Katz together, results were 94.73% and 93.58%, and using rapid sedimentation, Kato-Katz and copro-ELISA together, they were 97.56% and 100%, respectively. There was no correlation between coproantigen detection by optical density (OD) and infection intensity by eggs per gram of feces (epg) in Cajamarca low burden cases (, Author Summary A coproantigen-detection technique (MM3-COPRO ELISA) was evaluated in 436 and 362 schoolchildren of Huacullani, Bolivia, and Cajamarca valley, Peru, respectively. Classical techniques for egg detection were used for comparison. In Huacullani, using Kato-Katz as gold standard, sensitivity and specificity were 94.68% and 98.48%, respectively, and using Kato-Katz and COPRO-ELISA test together, they were 95.68% and 100%, respectively. In Cajamarca, using rapid sedimentation and Kato-Katz together, these results were 94.73% and 93.58%, respectively, and using rapid sedimentation, Kato-Katz and copro-ELISA together, results were 97.56% and 100%, respectively. There was no correlation between coproantigen detection by optical density (OD) and infection intensity by eggs per gram of feces (epg) in Cajamarca low burden cases (

The aim of this study was to assess and apply a microsatellite multiplex system for parentage determination in alpacas. An approach for parentage testing based on 10 microsatellites was evaluated in a population of 329 unrelated alpacas from different geographical zones in Peru. All microsatellite markers, which amplified in two multiplex reactions, were highly polymorphic with a mean of 14.5 alleles per locus (six to 28 alleles per locus) and an average expected heterozygosity (H(E)) of 0.8185 (range of 0.698-0.946). The total parentage exclusion probability was 0.999456 for excluding a candidate parent from parentage of an arbitrary offspring, given only the genotype of the offspring, and 0.999991 for excluding a candidate parent from parentage of an arbitrary offspring, given the genotype of the offspring and the other parent. In a case test of parentage assignment, the microsatellite panel assigned 38 (from 45 cases) offspring parentage to 10 sires with LOD scores ranging from 2.19 x 10(+13) to 1.34 x 10(+15) and Delta values ranging from 2.80 x 10(+12) to 1.34 x 10(+15) with an estimated pedigree error rate of 15.5%. The performance of this multiplex panel of markers suggests that it will be useful in parentage testing of alpacas.

The performance of Fas2-ELISA for the diagnosis of Fasciola hepatica infection in children living in areas of high endemicity for fascioliasis in the Peruvian Andes is analyzed. Fas2-ELISA is based on the detection of circulating IgG antibodies elicited in infected individuals against a F. hepatica antigen termed Fas2. The study was conducted in three Andean localities, Huertas-Julcan in Junin, Asillo in Puno, and Cajamarca, with a total population of 634 children in an age range 1 to 16 years old. Child fascioliasis prevalence was 21.1% in Huertas-Julcan, 25.4% in Asillo, and 24% in Cajamarca, estimated by coprological inspection. The seroprevalence of F. hepatica infection, determined by Fas2-ELISA, was 27.8% in Huertas-Julcan, 44.6% in Asillo, and 29.1% in Cajamarca. The overall sensitivity of Fas2-ELISA was 92.4%, the specificity 83.6%, and the negative predictive value 97.2%. No association between OD(450) Fas2-ELISA and infection intensity measured by egg counting was observed. Results show that Fas2-ELISA is a highly sensitive immunodiagnostic test for the detection of F. hepatica infection in children living in human fascioliasis endemic areas.

Fasciola hepaticahas recently emerged as a major pathogen of humans from reports on areas of endemicity and hyper-endemicity for fascioliasis. This situation is aggravated by the lack of standard assays for the screen diagnosis ofF. hepaticainfection in humans living in endemic areas. Our laboratory has developed an enzyme-linked immunosorbent assay (Fas2-ELISA) based on the capture of IgG antibody by a purified protein Fas2, which is an adult fluke cysteine proteinase. Fas2-ELISA exhibited 95% sensitivity and 100% specificity in 38 individuals infected withF. hepaticadiagnosed by finding eggs in stools and 46 serum samples from healthy volunteers. No cross-reaction was observed with 54 serum samples from patients with ten different parasitic infections including the trematodesParagonimus westermaniandSchistosoma mansoni. The high antigenicity of Fas2 is suggested by the fact that antibodies to Fas2 rise rapidly by 1–2 weeks of infection and rise until patency at 8 weeks of infection in experimentally infected alpacas. Field screening for human fascioliasis using Fas2-ELISA and coprology in three endemic locations of the Peruvian Andes resulted in 95.5% sensitivity, 86.6% specificity in a population of 664 children in an age range of 1 to 16 years old. These results provide evidence of the clinical potential of Fas2-ELISA to diagnose fascioliasis in humans exposed to liver fluke infection in endemic areas for this parasite. Fas2-ELISA is currently developed as a standard assay for both field screening for fascioliasis in people living in endemic areas and detecting occasionallyF. hepaticainfected patients in clinical laboratories.

Polymorphism in SLC11A1 has been implicated in host susceptibility to tuberculosis. We have studied associations between INT4, D543N, and 3′UTR polymorphisms of SLC11A1 and different clinical forms of TB. Analysis used 507 patients with pulmonary TB, 123 with extra pulmonary TB and 513 controls. INT4 and D543N showed allelic association with pulmonary TB ( P = 0.02 and 0.03 respectively). INT4–D543N–3′UTR haplotypes showed an association with pulmonary TB ( P = 0.03). No association of SLC11A1 with miliary TB was observed, and a possible association of D543N to the pleural form ( P = 0.08) was suggested. These results support association between SLC11A1 and TB, particularly to the common pulmonary form.

The immunodiagnosis of human infections with Fasciola hepatica using purified parasite cysteine proteinases as antigens is reported in this paper. IgG enzyme-linked immunosorbent assays (ELISAs) with 2 cysteine proteinases of 26 kDa (Fas1) and 25 kDa (Fas2), obtained from the regurgitated material of adult worms, were evaluated with serum samples from 38 patients infected with F. hepatica, from 54 patients with other parasitic infections and from 46 healthy volunteers. The diagnostic sensitivities for detection of F. hepatica infection at a serum dilution of 1 in 500 were 89% and 95% with Fas1 and Fas2 respectively. The specificity of the ELISAs was 98% for Fas1 and 100% for Fas2. IgG ELISA with Fas2 is a highly sensitive and specific assay for the immunodiagnosis of human fascioliasis.

Publisher Summary Genome analysis is a global approach centered on a large-scale synthesis of genetic and physical mapping information and has emerged as one of the most effective tools for molecular genetics. Protozoan parasites have several common genomic features: (1) asexual or clonal growth is a normal part of the life cycle, (2) the haploid genome sizes are about the size of the smallest human chromosome, (3) the chromosomes can generally be resolved by pulsed-field gel electrophoresis (PFGE), (4) chromosome size polymorphisms among natural isolates are relatively frequent, and (5) the genomes appear to have relatively little dispersed-repetitive DNA. The trypanosomatids that include Trypanosoma and Leishmania species share some unusual properties: (1) all messenger RNAs are trans-spliced with a 5' spliced leader or miniexon 39-nucleotide sequence and (2) many genes are polycistronically transcribed and may exist as tandem arrays. Some of these characteristics can be used to tailor methods specifically for the analysis of protozoan parasite genomes. This chapter describes the way the ability to separate whole chromosomes by gel electrophoresis simplifies the assignation of genes to chromosomes, the way the large number of telomere ends may be used to generate chromosome tags, and the way the spliced leader sequence aids in the construction of a directionally cloned, full-length complementary DNA library.

We have evaluated the possible mechanisms of liver fibrosis caused by Fasciola hepatica in an animal model and in culture using immortalized human stellate cells. Liver biopsies of F. hepatica-infected rats were performed at wk 8 and 16. Serum-starved LX-2 cells, a human stellate cell line, were exposed to increasing concentrations of Fas2 antigen. The expression of key fibrosis-related genes was evaluated by qRT-PCR. There was a significant correlation between fibrogenic gene expression and both intensity and duration of infection. LX-2 cells exposed to Fas2 showed progressively increased expression of mRNAs for Collagen I, alpha-smooth muscle-actin, platelet-derived growth factor beta receptor, and tissue inhibitor of metalloproteinase II; inhibition of Fas2 cysteine proteinase activity by E-64 abrogated these increases, suggesting that the protease activity of Fas2 is involved in fibrogenic stimulation. In summary, F. hepatica infection is associated with up-regulation of mRNAs associated with hepatic fibrogenesis in vivo and in activated hepatic stellate cells.

Fasciola hepatica es el agente etiológico de la fasciolosis en el Perú. La fasciolosis es un problema de salud pública por la alta prevalencia de la infección humana, especialmente niños y un problema veterinario de importancia por las altas tasas de infección del ganado en la mayoría de regiones del País. La fasciolosis es endémica en la sierra y la costa, y esporádica en la región amazónica. La infección humana se reporta en 18 regiones y la animal en 21 de las 24 regiones del Perú. La transmisión humana ocurre en poblaciones andinas rurales dedicadas a la agricultura, pero hay un creciente número de casos en ciudades. Diferentes situaciones epidemiológicas de la infección humana se presentan: i) Regiones con casos no autóctonos, donde la infección es por consumo de vegetales contaminados traídos de zonas endémicas; ii) Regiones con poblados hipo- y mesoendémicos, transmisión local y prevalencia < 10%; y iii) Regiones con poblados hiperendémicos, transmisión local y prevalencia ≥ 10%. El parásito infecta a vacunos, ovinos, equinos, caprinos, porcinos, camélidos, cobayos y conejos. La pérdida ganadera anual por la fasciolosis es no menor de US$ 50 millones, estimada por la prevalencia de la infección y los decomisos de hígados de vacunos en mataderos. Es difícil estimar el impacto económico de la fasciolosis humana por su estatus de enfermedad desatendida, lo cierto es que la fasciolosis es endémica, y en algunos casos hiperendémica, en las regiones más pobres de nuestro país, donde la situación debe ser reconocida como una emergencia de salud pública. Fasciola hepatica is the causative agent of fasciolosis in Peru; the disease is an important public health problem by the high prevalence of the human infection affecting mainly children and a major veterinary problem by the high rates of infected livestock. The human disease is endemic in the Sierra and the Coast but sporadic in the Amazonia, and reported in 18 Departments, while the animal infection in 21 of 24 Departments of Peru. Transmission occurs in Andean rural populations engaged in agriculture, but recently an increasing number of people became infected in the cities. The epidemiological situation in Peru includes i) Departments with non-autochtonous cases, where infection occurs by consumption of contaminated vegetables brought from endemic areas or infection is acquired by visit to endemic areas; ii) Departments with hypoendemic and mesoendemic villages, where transmission occurs by ingest of contaminated vegetables and prevalence ≤10%; and iii) Departments with hyper-endemic villages with human prevalence >10 %, with an intense transmission by consumption of contaminated vegetables. The disease affects bovine, sheep, goat, swine, equine, South American camelids, rabbits and guinea pigs. The negative impact of fasciolosis in the livestock economy is not lesser than US$ 50 million per year, estimation based on the prevalence and the number of condemned livers in the abattoirs. It is difficult to estimate the economic impact of this infection in the human health due to its status of neglected disease, but fasciolosis is hyper-endemic in the poorest Andean areas of Peru where the situation has to be recognized as a public health emergency.