Development of a highly tolerant bacterial consortium for asphaltene biodegradation in soils.

Asphaltenes are the most polar and heavy fraction of petroleum, and their complex structure and toxicity make them resistant to biodegradation. The ability to tolerate high asphaltene concentrations is crucial to reducing the toxicity-related inhibition of microbial growth and improving their capacity for adaptation, survival, and biodegradation in soils highly contaminated with asphaltenes. This study developed a highly tolerant consortium for efficient asphaltene biodegradation in soils from 22 bacterial isolates obtained from heavy-crude oil-contaminated soils. Isolates corresponded to the Rhodococcus, Bacillus, Stutzerimonas, Cellulosimicrobium, Pseudomonas, and Paenibacillus genera, among others, and used pure asphaltenes and heavy crude oil as the only carbon sources. Surface plate assays were used to evaluate the tolerance of individual isolates to asphaltenes, and the results showed variations in the extension and inhibition rates with maximum tolerance levels at 60,000 mg asphaltenes l-1. Inhibition assays were used to select non-antagonistic bacterial isolates among those showing the highest tolerance levels to asphaltenes. A consortium made up of the five most tolerant and non-antagonistic bacterial isolates was able to degrade up to 83 wt.% out of 10,000 mg asphaltenes kg-1 in the soil after 52 days. Due to its biological compatibility, high asphaltene tolerance, and ability to utilise it as a source of energy, the degrading consortium developed in this work has shown a high potential for soil bioremediation and is a promising candidate for the treatment of aged soil areas contaminated with heavy and extra-heavy crude oil. This would be the first research to assess and consider extreme bacterial tolerance and microbial antagonism between individual degrading microbes, leading to the development of an improved consortium capable of efficiently degrading high amounts of asphaltenes in soil.

Effect of Essential Oil from Lippia origanoides on the Transcriptional Expression of Genes Related to Quorum Sensing, Biofilm Formation, and Virulence of Escherichia coli and Staphylococcus aureus.

Microbial infections resistant to conventional antibiotics constitute one of the most important causes of mortality in the world. In some bacterial species, such as Escherichia coli and Staphylococcus aureus pathogens, biofilm formation can favor their antimicrobial resistance. These biofilm-forming bacteria produce a compact and protective matrix, allowing their adherence and colonization to different surfaces, and contributing to resistance, recurrence, and chronicity of the infections. Therefore, different therapeutic alternatives have been investigated to interrupt both cellular communication routes and biofilm formation. Among these, essential oils (EO) from Lippia origanoides thymol-carvacrol II chemotype (LOTC II) plants have demonstrated biological activity against different biofilm-forming pathogenic bacteria. In this work, we determined the effect of LOTC II EO on the expression of genes associated with quorum sensing (QS) communication, biofilm formation, and virulence of E. coli ATCC 25922 and S. aureus ATCC 29213. This EO was found to have high efficacy against biofilm formation, decreasing-by negative regulation-the expression of genes involved in motility (fimH), adherence and cellular aggregation (csgD), and exopolysaccharide production (pgaC) in E. coli. In addition, this effect was also determined in S. aureus where the L. origanoides EO diminished the expression of genes involved in QS communication (agrA), production of exopolysaccharides by PIA/PNG (icaA), synthesis of alpha hemolysin (hla), transcriptional regulators of the production of extracellular toxins (RNA III), QS and biofilm formation transcriptional regulators (sarA) and global regulators of biofilm formation (rbf and aur). Positive regulation was observed on the expression of genes encoding inhibitors of biofilm formation (e.g., sdiA and ariR). These findings suggest that LOTCII EO can affect biological pathways associated with QS communication, biofilm formation, and virulence of E. coli and S. aureus at subinhibitory concentrations and could be a promising candidate as a natural antibacterial alternative to conventional antibiotics.

Effect of Essential Oils on Growth Inhibition, Biofilm Formation and Membrane Integrity of Escherichia coli and Staphylococcus aureus.

Biofilm as a cellular conformation confers survival properties to microbial populations and favors microbial resistance. Here, we investigated the antimicrobial, antibiofilm, antimotility, antihemolytic activity, and the interaction with synthetic membranes of 15 essential oils (EOs) on E. coli ATCC 25922 and S. aureus ATCC 29213. Antimicrobial activity of EOs was determined through microdilution method; development of the biofilm was assessed using the crystal violet assay and SEM microscopy. Results indicate that Lippia origanoides thymol-carvacrol II chemotype (LTC II) and Thymus vulgaris (TV) exhibited a significant antibacterial activity, with MIC values of 0.45 and 0.75 mg/mL, respectively. The percentage of biofilm formation inhibition was greater than 70% at subinhibitory concentrations (MIC50) for LTC II EO. The results demonstrate that these two oils had significantly reduced the hemolytic effect of S. aureus by 54% and 32%, respectively, and the mobility capacity by swimming in E. coli with percentages of decrease of 55% and 47%, respectively. The results show that LTC II and TV EOs can interact with the hydrophobic core of lipid bilayers and alter the physicochemical properties of membranes. The findings suggest that LTC II and TV oils may potentially be used to aid in the treatment of S. aureus and E. coli infections.

Genome-Resolved Meta-Analysis of the Microbiome in Oil Reservoirs Worldwide.

Microorganisms inhabiting subsurface petroleum reservoirs are key players in biochemical transformations. The interactions of microbial communities in these environments are highly complex and still poorly understood. This work aimed to assess publicly available metagenomes from oil reservoirs and implement a robust pipeline of genome-resolved metagenomics to decipher metabolic and taxonomic profiles of petroleum reservoirs worldwide. Analysis of 301.2 Gb of metagenomic information derived from heavily flooded petroleum reservoirs in China and Alaska to non-flooded petroleum reservoirs in Brazil enabled us to reconstruct 148 metagenome-assembled genomes (MAGs) of high and medium quality. At the phylum level, 74% of MAGs belonged to bacteria and 26% to archaea. The profiles of these MAGs were related to the physicochemical parameters and recovery management applied. The analysis of the potential functional core in the reservoirs showed that the microbiota was specialized for each site, with 31.7% of the total KEGG orthologies annotated as functions (1690 genes) common to all oil fields, while 18% of the functions were site-specific, i.e., present only in one of the oil fields. The oil reservoirs with a lower level of intervention were the most similar to the potential functional core, while the oil fields with a long history of water injection had greater variation in functional profile. These results show how key microorganisms and their functions respond to the distinct physicochemical parameters and interventions of the oil field operations such as water injection and expand the knowledge of biogeochemical transformations in these ecosystems.

Cambios en el perfil protéico de E. coli O157:H7 frente al tratamiento con Ib-M1 e IONP@Ib-M1 / Changes in the protein profile of E. coli O157: H7 against treatment with Ib-M1 and IONP@Ib-M1 / Identificação de alterações no perfil protetor de E. coli O157: H7 contra o tratamento com Ib-M1 e IONP@Ib-M1

Resumen Escherichia coli 0157:H7 es una bacteria patógena reconocida por su capacidad de resistencia a diversos antibióticos; razón por la cual, se generan complicaciones en el tratamiento de infecciones producidas por esta bacteria. El péptido Ib-M1 y el bioconjugado I0NP@Ib-M1 han surgido como una nueva alternativa antimicrobiana contra E. coli 0157:H7. El mecanismo de acción de Ib-Mi e I0NP@Ib-M1 contra esta bacteria aún es desconocido; por lo tanto, el objetivo de esta investigación fue identificar el cambio en el perfil de proteínas de E. coli 0157:H7 luego del tratamiento con Ib-M1 e I0NP@ Ib-M1 como primer paso para determinar su mecanismo de acción. Para esto, se llevó a cabo la obtención de proteínas, posteriormente se realizó una electroforesis bidimensional para finalmente realizar la determinación de la variabilidad de los perfiles proteicos. Una vez obtenidos estos perfiles, se llevó a cabo un análisis de varianza (AN0VA). Se identificaron 72 proteínas expresadas diferencialmente, las cuales pueden relacionarse con el efecto sobre el crecimiento de la bacteria en presencia de Ib-M1 e I0NP@Ib-M. Estas proteínas se encuentran involucradas en procesos de transferencia de grupos acilo (proteína Yhbs), translocación de lipoproteínas (proteína LolA) y transporte de aminoácidos (proteína GpmA), entre otros.

Abstract Escherichia coli 0157: H7 is a pathogenic bacterium which is recognized for the ability to resist multiple antibiotics; accordingly, complications occur in the treatment of infections caused by this bacterium. The Ib-M1 peptide and the I0NP @ Ib-M1 bioconjugate have emerged as a new antimicrobial alternatives against E. coli 0157: H7. The mechanism of action of Ib-M1 and I0NP @ Ib-M1 against this bacterium is still unknown; therefore, the goal of this research was to identify the change in the proteins profile of E. coli 0157: H7 after treatment with Ib-M1 and I0NP @ Ib-M1 as a first step to determine its mechanism of action. For this, the proteins were obtained first, and then a two-dimensional electrophoresis was performed to finally determine the variability of the protein profiles. 0nce the protein profiles were obtained, an analysis of variance (AN0VA) was carried out. 72 differentially expressed proteins were identified, which can be connected to the effect on the bacterium's growth in the presence of Ib-M1 and I0NP @ Ib-M. These proteins are involved in acyl groups transfer processes (Yhbs protein), lipoprotein translocation (LolA protein) and amino acid transport (GpmA protein), among others.

Resumo Escherichia coli O157: H7 é uma bactéria patogênica reconhecida por sua capacidade de resistir a vários antibióticos; razão pela qual, complicações são geradas no tratamento de infecções produzidas por essa bactéria. O peptídeo Ib-M1 livre e imobilizado em nanopartículas magnéticas de óxido de ferro (IONP @ Ib-M1) surgiu como uma nova alternativa antimicrobiana contra E. coli O157: H7 e isolados clínicos desta bactéria. O mecanismo de ação de Ib-M1 e IONP @ Ib-M1 contra E. coli O157: H7 ainda é desconhecido; Portanto, o objetivo desta pesquisa foi identificar a alteração no perfil proteico de E. coli O157: H7 após o tratamento com Ib-M1 e IONP @ Ib-M1 como um primeiro passo para determinar seu mecanismo de ação. Para isso, foi realizada a obtenção das proteínas, posteriormente foi realizada uma eletroforese bidimensional para finalmente determinar a variabilidade dos perfis protéicos. Uma vez obtidos os perfis de proteínas, foi realizada uma análise de variância (ANOVA). Os resultados mostram a identificação de proteínas expressas diferencialmente e que estão envolvidas em processos de transferência de grupos acila (proteína Yhbs), translocação de lipoproteínas (proteína LolA) e transporte de aminoácidos (proteína GpmA), entre outros.

Construction of PAH-degrading mixed microbial consortia by induced selection in soil.

Bioremediation of polycyclic aromatic hydrocarbons (PAHs)-contaminated soils through the biostimulation and bioaugmentation processes can be a strategy for the clean-up of oil spills and environmental accidents. In this work, an induced microbial selection method using PAH-polluted soils was successfully used to construct two microbial consortia exhibiting high degradation levels of low and high molecular weight PAHs. Six fungal and seven bacterial native strains were used to construct mixed consortia with the ability to tolerate high amounts of phenanthrene (Phe), pyrene (Pyr) and benzo(a)pyrene (BaP) and utilize these compounds as a sole carbon source. In addition, we used two engineered PAH-degrading fungal strains producing heterologous ligninolytic enzymes. After a previous selection using microbial antagonism tests, the selection was performed in microcosm systems and monitored using PCR-DGGE, CO2 evolution and PAH quantitation. The resulting consortia (i.e., C1 and C2) were able to degrade up to 92% of Phe, 64% of Pyr and 65% of BaP out of 1000 mg kg-1 of a mixture of Phe, Pyr and BaP (1:1:1) after a two-week incubation. The results indicate that constructed microbial consortia have high potential for soil bioremediation by bioaugmentation and biostimulation and may be effective for the treatment of sites polluted with PAHs due to their elevated tolerance to aromatic compounds, their capacity to utilize them as energy source.

Comparative metagenomic analysis of PAH degradation in soil by a mixed microbial consortium.

In this study, we used a taxonomic and functional metagenomic approach to analyze some of the effects (e.g. displacement, permanence, disappearance) produced between native microbiota and a previously constructed Polycyclic Aromatic Hydrocarbon (PAH)-degrading microbial consortium during the bioremediation process of a soil polluted with PAHs. Bioaugmentation with a fungal-bacterial consortium and biostimulation of native microbiota using corn stover as texturizer produced appreciable changes in the microbial diversity of polluted soils, shifting native microbial communities in favor of degrading specific populations. Functional metagenomics showed changes in gene abundance suggesting a bias towards aromatic hydrocarbon and intermediary degradation pathways, which greatly favored PAH mineralization. In contrast, pathways favoring the formation of toxic intermediates such as cytochrome P450-mediated reactions were found to be significantly reduced in bioaugmented soils. PAH biodegradation in soil using the microbial consortium was faster and reached higher degradation values (84% after 30 d) as a result of an increased co-metabolic degradation when compared with other mixed microbial consortia. The main differences between inoculated and non-inoculated soils were observed in aromatic ring-hydroxylating dioxygenases, laccase, protocatechuate, salicylate and benzoate-degrading enzyme genes. Based on our results, we propose that several concurrent metabolic pathways are taking place in soils during PAH degradation.

Biodegradation of polycyclic aromatic hydrocarbons by Trichoderma species: a mini review.

Fungi belonging to Trichoderma genus are ascomycetes found in soils worldwide. Trichoderma has been studied in relation to diverse biotechnological applications and are known as successful colonizers of their common habitats. Members of this genus have been well described as effective biocontrol organisms through the production of secondary metabolites with potential applications as new antibiotics. Even though members of Trichoderma are commonly used for the commercial production of lytic enzymes, as a biological control agent, and also in the food industry, their use in xenobiotic biodegradation is limited. Trichoderma stands out as a genus with a great range of substrate utilization, a high production of antimicrobial compounds, and its ability for environmental opportunism. In this review, we focused on the recent advances in the research of Trichoderma species as potent and efficient aromatic hydrocarbon-degrading organisms, as well as aimed to provide insight into its potential role in the bioremediation of soils contaminated with heavy hydrocarbons. Several Trichoderma species are associated with the ability to metabolize a variety of both high and low molecular weight polycyclic aromatic hydrocarbons (PAHs) such as naphthalene, phenanthrene, chrysene, pyrene, and benzo[a]pyrene. PAH-degrading species include Trichoderma hamatum, Trichoderma harzianum, Trichoderma reesei, Trichoderma koningii, Trichoderma viride, Trichoderma virens, and Trichoderma asperellum using alternate enzyme systems commonly seen in other organisms, such as multicooper laccases, peroxidases, and ring-cleavage dioxygenases. Within these species, T. asperellum stands out as a versatile organism with remarkable degrading abilities, high tolerance, and a remarkable potential to be used as a remediation agent in polluted soils.

Morphological changes and growth of filamentous fungi in the presence of high concentrations of PAHs.

In this study, we evaluated the effect of low and high molecular weight polycyclic aromatic hydrocarbons (PAHs), i.e., Phenanthrene, Pyrene and Benzo[a]pyrene, on the radial growth and morphology of the PAH-degrading fungal strains Aspergillus nomius H7 and Trichoderma asperellum H15. The presence of PAHs in solid medium produced significant detrimental effects on the radial growth of A. nomius H7 at 4,000 and 6,000 mg L(-1) and changes in mycelium pigmentation, abundance and sporulation ability at 1,000-6,000 mg L(-1). In contrast, the radial growth of T. asperellum H15 was not affected at any of the doses tested, although sporulation was observed only up to 4,000 mg L(-1) and as with the H7 strain, some visible changes in sporulation patterns and mycelium pigmentation were observed. Our results suggest that fungal strains exposed to high doses of PAHs significantly vary in their growth rates and sporulation characteristics in response to the physiological and defense mechanisms that affect both pigment production and conidiation processes. This finding is relevant for obtaining a better understanding of fungal adaptation in PAH-polluted environments and for developing and implementing adequate strategies for the remediation of contaminated soils.

Morphological changes and growth of filamentous fungi in the presence of high concentrations of PAHs

In this study, we evaluated the effect of low and high molecular weight polycyclic aromatic hydrocarbons (PAHs), i.e., Phenanthrene, Pyrene and Benzo[a]pyrene, on the radial growth and morphology of the PAH-degrading fungal strains Aspergillus nomius H7 and Trichoderma asperellum H15. The presence of PAHs in solid medium produced significant detrimental effects on the radial growth of A. nomius H7 at 4,000 and 6,000 mg L⁻¹ and changes in mycelium pigmentation, abundance and sporulation ability at 1,000–6,000 mg L⁻¹. In contrast, the radial growth of T. asperellum H15 was not affected at any of the doses tested, although sporulation was observed only up to 4,000 mg L⁻¹ and as with the H7 strain, some visible changes in sporulation patterns and mycelium pigmentation were observed. Our results suggest that fungal strains exposed to high doses of PAHs significantly vary in their growth rates and sporulation characteristics in response to the physiological and defense mechanisms that affect both pigment production and conidiation processes. This finding is relevant for obtaining a better understanding of fungal adaptation in PAH-polluted environments and for developing and implementing adequate strategies for the remediation of contaminated soils.

Degradation of polycyclic aromatic hydrocarbons in soil by a tolerant strain of Trichoderma asperellum.

Trichoderma asperellum H15, a previously isolated strain characterized by its high tolerance to low (LMW) and high molecular weight (HMW) PAHs, was tested for its ability to degrade 3-5 ring PAHs (phenanthrene, pyrene, and benzo[a]pyrene) in soil microcosms along with a biostimulation treatment with sugarcane bagasse. T. asperellum H15 rapidly adapted to PAH-contaminated soils, producing more CO2 than uncontaminated microcosms and achieving up to 78 % of phenanthrene degradation in soils contaminated with 1,000 mg Kg(-1) after 14 days. In soils contaminated with 1,000 mg Kg(-1) of a three-PAH mixture, strain H15 was shown to degrade 74 % phenanthrene, 63 % pyrene, and 81 % of benzo[a]pyrene. Fungal catechol 1,2 dioxygenase, laccase, and peroxidase enzyme activities were found to be involved in the degradation of PAHs by T. asperellum. The results demonstrated the potential of T. asperellum H15 to be used in a bioremediation process. This is the first report describing the involvement of T. asperellum in LMW and HMW-PAH degradation in soils. These findings, along with the ability to remove large amounts of PAHs in soil found in the present work provide enough evidence to consider T. asperellum as a promising and efficient PAH-degrading microorganism.

Direct analysis of genetic variability in Trypanosoma cruzi populations from tissues of Colombian chagasic patients.

The clinical symptoms of Chagas disease are highly variable and are correlated with geographical distribution and parasite genetic group. Trypanosoma cruzi group I is associated with chagasic cardiomyopathy in Colombia and other countries in northern South America. However, in southern South America, T cruzi group II predominates and is associated with cardiomyopathy and digestive forms of the disease. The aim of this work was to determine the correlation between the genetic profiles of T cruzi groups circulating in the biological cycle and those present in tissues from patients with Chagas disease. We genotyped T cruzi in 10 heart tissue samples from patients with cardiomyopathy from a highly endemic area of Colombia. The genotyping was performed using nuclear and mitochondrial genes and low-stringency single-specific primer polymerase chain reaction. As expected, the predominant genetic group was T cruzi group I; however, we also detected T cruzi group II. Microsatellite analyses suggested a predominance of monoclonal populations, and sequence alignments showed similarities with Colombian strains. In addition, kinetoplast DNA signatures obtained by low-stringency single-specific primer polymerase chain reaction allowed us to group strains into the 2 genetic groups. Thus, we conclude that both T cruzi genetic groups are producing severe cases of Chagas disease in Colombia. We did not observe any correlation between low-stringency single-specific primer polymerase chain reaction profiles, histopathologic findings, clinical forms, and severity of Chagas disease.

Mixed infection of Trypanosoma cruzi I and II in a Colombian cardiomyopathic patient.

The Trypanosoma cruzi taxon is composed of 2 major lineages, T cruzi I and T cruzi II. The clinical symptoms of Chagas disease are highly variable, and their geographic distribution is correlated with the distribution of the parasite lineages. In Colombia and northern South America, T cruzi I lineage is associated with chagasic cardiomyopathy. Alternatively, in the countries south cone of South America, there is a predominance of T cruzi II, which is associated with cardiomyopathy and digestive diseases. We report for the first time a mixed infection consisting of both T cruzi I and T cruzi II detected in the esophagus and in the heart, respectively, of a cardiomyopathic patient from an endemic area in Santander, Colombia. This finding has epidemiological relevance related to the association of T cruzi II with the clinical manifestations of Chagas disease and its frequency in Colombia and countries in northern South America.

Evidence of Trypanosoma cruzi II infection in Colombian chagasic patients.

Trypanosoma cruzi is genetically classified into at least two major lineages named T. cruzi I (also named Tc I) and T. cruzi II (also named Tc IIb). T. cruzi II is associated with Chagas' disease in the southern cone of South America, while T. cruzi I is the only one so far identified in chagasic patients of Central America and in the northern part of South America. Herein we identified T. cruzi IIb directly in 9.9% of blood of chronic chagasic patients of Colombia. This finding establishes that in this region, the two T. cruzi lineages are associated with the pathology of Chagas' disease and have implications in the morbidity and epidemiology of the disease.

Polymorphisms of toll-like receptor 2 and 4 genes in Chagas disease.

The aim of this study was to test the possible implication of toll-like receptor 2 (TLR2) and TLR4 gene polymorphisms in determining the susceptibility to Chagas' disease. Genotypes were determined by polymerase chain reaction-restriction fragment length polymorphism in 475 individuals from Colombia, 143 seropositive with chagasic cardiomyopathy, 132 seropositive asymptomatic and 200 seronegative. The TLR2 arginine to glutamine substitution at residue 753(Arg753Gln) polymorphism was absent in the groups analyzed. The TLR4 Asp299Gly and Thr399Ile polymorphisms are in linkage disequilibrium and we observed a very low frequency of these polymorphisms in our study population (2.6% and 1.8% respectively). The overall TLR2 and TLR4 alleles and genotype distribution in seronegative and seropositive were not significantly different. We compared the frequencies between asymptomatic patients and those with chagasic cardiomyopathy and we did not observe any significant differences in the distribution of alleles or genotypes. In summary, this study corroborates the low frequency of TLR2 and TLR4 polymorphisms observed in other populations and suggest that these do not play an important role in Chagas' disease. The validation of these findings in independent cohorts is needed to firmly establish a role for TLR2 and TLR4 variants in Chagas' disease.

Polymorphisms of toll-like receptor 2 and 4 genes in Chagas disease

The aim of this study was to test the possible implication of toll-like receptor 2 (TLR2) and TLR4 gene polymorphisms in determining the susceptibility to Chagas' disease. Genotypes were determined by polymerase chain reaction-restriction fragment length polymorphism in 475 individuals from Colombia, 143 seropositive with chagasic cardiomyopathy, 132 seropositive asymptomatic and 200 seronegative. The TLR2 arginine to glutamine substitution at residue 753(Arg753Gln) polymorphism was absent in the groups analyzed. The TLR4 Asp299Gly and Thr399Ile polymorphisms are in linkage disequilibrium and we observed a very low frequency of these polymorphisms in our study population (2.6 percent and 1.8 percent respectively). The overall TLR2 and TLR4 alleles and genotype distribution in seronegative and seropositive were not significantly different. We compared the frequencies between asymptomatic patients and those with chagasic cardiomyopathy and we did not observe any significant differences in the distribution of alleles or genotypes. In summary, this study corroborates the low frequency of TLR2 and TLR4 polymorphisms observed in other populations and suggest that these do not play an important role in Chagas' disease. The validation of these findings in independent cohorts is needed to firmly establish a role for TLR2 and TLR4 variants in Chagas' disease.

[Seroprevalence of Chagas disease and associated risk factors in a population of Morroa, Sucre]. / Seroprevalencia de la enfermedad de Chagas y factores de riesgo asociados en una población de Morroa, Sucre.

INTRODUCTION: Chagas disease is a major public health problem in Latin America. In Colombia, a large area has the ecoepidemiological conditions which favor the active transmition of this infection. OBJECTIVE: This study was undertaken in a population from the municipality of Morroa, Sucre Province, to evaluate risk factors and to determine the seroprevalence to Chagas disease. MATERIALS AND METHODS: A questionnaire was given to a sample population of 122 people classed as rural (n=76) or urban area (n=46). A serological screening was undertaken by Elisa test, with confirmation of seropositives with IHA (Chagatest) and parasitological confirmation by polymerase chain reaction (PCR). RESULTS: Four people were positive by Elisa test (3.3%); however, they were negative by IHA and PCR. One of the four positives by Elisa was positive by indirect immuno flourescence (IFAT) as well. The sample showed a low presence of seropositives against Trypanosoma cruzi. However, the presence of parasite could not be confirmed by the PCR test. The main risk factors were houses thatched with palm roofs, clay floors, wood walls, and presence of domestic animal reservoirs. CONCLUSIONS: The study population presented risk factors for the establishment of active transmission. The presence of triatomines must be verified in this area and establishment of control measures are necessary for preventiving the resurgence of the Chagas disease in Morroa.

Polymorphism in the 3' UTR of the IL12B gene is associated with Chagas' disease cardiomyopathy.

The aim of this study was to investigate the possible influence of a 3' untranslated region (3' UTR) polymorphism of the IL12B gene in susceptibility to Trypanosoma cruzi infection or in the development to cardiomyopathy in Chagas' disease (CD). We determined the IL12B 3' UTR genotypes in a sample of 200 seronegative individuals and 260 serologically positive patients (130 with Chagasic cardiomyopathy and 130 asymptomatic). All individuals are from a Colombian region where T. cruzi infection is endemic. Genotyping was performed by the PCR-restriction fragment length polymorphism (RFLP) method. The overall distribution of the IL12B 3' UTR alleles and genotypes in seronegative compared with seropositive individuals was not statistically significant. Interestingly, we found that the IL12B 3' UTR CC genotype was significantly increased among cardiomyopathic patients when compared to asymptomatic individuals (16% versus 5%; P=0.005; P(c)=0.015; OR=3.39; 95% CI 1.3-9.15). In addition, we observed that the IL12B 3' UTR C allele was present at significantly higher frequency in cardiomyopathic (33% versus 22%; P=0.008; P(c)=0.016; OR=1.69; 95% CI 1.12-2.55) as compared to asymptomatic. Our results suggest that IL12B 3' UTR gene polymorphisms may influence the susceptibility to develop Chagasic cardiomyopathy.

Interleukin-1 gene cluster polymorphism in chagas disease in a Colombian case-control study.

The aim of this study was to assess the possible association between the IL1A, IL1B and IL1RN gene polymorphisms and Chagas disease. Our study population consisted of 130 serologically positive cardiomyopathic patients and 130 seropositive and asymptomatic individuals from a Colombian population where Trypanosoma cruzi infection is endemic. Genotyping of the IL1A (-889C/T, +4845G/T), IL1B (-511C/T, -31T/C, +3954T/C, +5810G/A) and IL1RN (+8006T/C, +8061C/T, +11100T/C) polymorphisms was performed by polymerase chain reaction-restriction fragment length polymorphism and polymerase chain reaction sequence-specific primer methods. Statistically significant differences in the distribution of the IL1B +5810 genotypes were observed comparing cardiomyopathic patients and asymptomatic individuals (p = 0.036). Frequency of the GG genotype was higher in the cardiomyopathic patient group than in the asymptomatic group (13% versus 5%, p = 0.03, odds ratio [OR] = 2.64, 95% confidence interval [CI] = 0.99-7.33). Differences in the distribution of the allele frequencies were also observed, being the +5810G allele overrepresented in patients with cardiomyopathy (37% versus 27%, p = 0.014, OR = 1.59, 95% CI = 1.08-2.36). Examination of markers in the IL1A (-889 and +4845), IL1B (-511, -31, and +3954) and IL1RN (+11100) genes revealed that the overall distribution of alleles and genotypes in patients with chagasic cardiomyopathy and asymptomatic were not significantly different. Our results show that in Colombian population the IL1B+5810G allele was associated with an increased risk chagasic cardiomyopathy. In addition, we demonstrated that homozygosity for the IL1B +5810G risk allele increased significantly the susceptibility to cardiomyopathy. This implies that the effect of IL1B gene on chagasic cardiomyopathy predisposition is dose dependent. We found that the haplotype CT of IL1B -31 and +3954 polymorphisms showed higher association with risk to chagasic cardiomyopathy (p(c) = 0.008, OR = 12.53) and the extended haplotype (CCTCATT) was significantly more frequent in asymptomatic than in cardiomyopathic patients (p = 0.0014, p(c) = 0.011, OR = 0.17). Therefore this study suggests that IL1 gene cluster polymorphisms may play a relevant role in the susceptibility to development of chagasic chronic cardiomyopathy.