Dataset of assembly and annotation of the mitogenomes of Triatoma dimidiata and Triatoma huehuetenanguensis captured from Yucatán, México.

Triatoma dimidiata is a species complex, and its members are responsible for the transmission of Trypanosoma cruzi, the causative agent of Chagas disease. We present the assembly and annotation of the mitogenome of the Triatoma dimidiata (Latreille, 1811) and Triatoma huehuetenanguensis Lima-Cordón & Justi, 2019. The mitochondrial genomes were successfully sequenced using the Illumina Nextseq 500 platform, 2×75 cycles, and 5 million reads per sample. Contigs were assembled and annotated using the reference genomes of T. dimidiata and T. huehuetenanguensis available in Genbank (NC_002609 and NC_050325.1, respectively). The mitogenomes of T. dimidiata have lengths of 17,008 bp, while those of T. huehuetenanguensis are 15,910 bp and 15,909 bp. The genome comprises 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes, and a control region. The mitogenomes will be valuable to scholars and students focused on integrative taxonomy, phylogeography, and evolutionary studies of the Triatoma dimidiata complex and the transmission of Chagas diseases.

Activation of murine macrophages by membrane proteins from Tritrichomonas foetus grown on iron- and calcium-rich conditions.

Tritrichomonas foetus is a protozoan parasite that causes a venereal disease in cattle limiting reproduction by abortions and sterility. The immune response against this parasite is poorly understood. Since the iron and calcium ions are important regulators of the microenvironment of the urogenital tract in cattle, we decided to evaluate the role of these divalent cations on the antigenicity of membrane proteins of T. foetus on macrophage activation as one of the first inflammatory responses towards this pathogen. Colorimetric methods and ELISA were used to detect the nitric oxide and oxygen peroxide production and expression of cytokines in culture supernatant from macrophage incubated with membrane proteins from T. foetus cultured in iron- and calcium-rich conditions. qRT-PCR assays were used to evaluate the transcript expression of genes involved in the inflammatory response on the macrophages. The membrane proteins used for in vitro stimulation caused the up-regulation of the iNOS and NOX-2 genes as well as the generation of NO and H2 O2 in murine macrophages on a dependent way of the metal concentrations. Additionally, after stimulation, macrophages showed a considerable rise in pro-inflammatory cytokines and a downregulation of anti-inflammatory cytokines, as well as up-regulation in the transcription of the TLR4 and MyD88 genes. These data suggest that membrane proteins of T. foetus induced by iron and calcium can activate an inflammatory specific macrophage response via TLR4/MyD88 signalling pathway.

Exposure of Tritrichomonas foetus to sublethal doses of metronidazole induces a specific proinflammatory response in murine macrophages.

Tritrichomonas foetus is a flagellated parasite that primarily infects the reproductive tissues of livestock, causing bovine trichomoniasis. The cytoplasmic membrane of T. foetus contains various compounds that contribute to adherence, colonization, and pathogenicity. Metronidazole (MTZ) is the main treatment for trichomoniasis, but the emergence of drug-resistant strains is a concern due to improper use and dosing. T. foetus infection induces inflammation, and macrophages are key players in the immune response. However, our understanding of the host's immune response to T. foetus is limited, and the specific mechanisms underlying these responses are not well understood. This study aimed to investigate the impact of T. foetus surface proteins from trophozoites cultured under different sublethal MTZ conditions (MTZ-treated T. foetus MPs) on macrophage activation. By analyzing cytokine levels and gene expression in murine macrophages, we demonstrated that MTZ-treated T. foetus MPs induce a specific proinflammatory response. MTZ-treated T. foetus MPs-exposed macrophages exhibited a higher NO and H2 O2 production and overexpression of iNOS and NOX-2 genes in comparison to untreated T. foetus. Additionally, MTZ-treated T. foetus MPs triggered a significant induction of the proinflammatory cytokines IL-1ß, IL-6, TNF-α, and IFN-γ, as well as the overexpression of the TLR4, MyD88, and NF-κB genes on murine macrophages. The study aimed to unravel the immunological response and potential proinflammatory pathways involved in T. foetus infection and MTZ stress. Understanding the immune responses and mechanisms through which T. foetus surface proteins activate macrophages can contribute to the development of new therapeutic strategies for controlling bovine trichomoniasis.

Mites associated with budgerigars Melopsittacus undulatus (Psittaciformes: Psittacidae) and the first report of Ornithonyssus bursa (Mesostigmata: Macronyssidae) in Mexico.

Background: Hematophagous mites affect numerous bird species, causing severe injuries to the budgerigars. Some species can cause dermatitis in humans. Aims: The purpose was to morphologically identify the mites related to budgerigars (Melopsittacus undulatus) and their nests in Yucatan, Mexico. Methods: In May 2022, a private budgerigar hatchery was visited and mites were collected from the bodies of the birds and their nests. The morphological traits of the mites were confirmed by scanning electron microscopy. Results: Four of 30 birds showed severe clinical signs of mite infestation. The Budgerigars revealed lesions in the cere, nostrils, eyelids, beak, and paws. The bird's skin showed signs of dryness and beige coloring. The birds with severe damage also presented anorexia and had deformed paws and beaks. The parasitosis was caused by the "burrowing mites," Knemidocoptes pilae. The burrowing mites and the Grallacheles bakeri were recovered and identified from paw scabs. To eliminate mites, a topical application of Ivermectin was administered to the necks of the birds. The dose was a single, which has a residuality of 21 days. Two drops (0.115 mg/ml) of ivermectin were applied to each bird. A gradual reduction in crusted lesions due to mite mortality was noted. The "tropical fowl mite" Ornithonyssus bursa was identified in the nests, which represents the first record in Mexico. Conclusions: Three species of mites were discovered in a single budgerigar hatchery. This emphasizes the importance of deworming birds and keeping a clean environment in their cages to reduce the potential for parasitic mite infestation.

Impacto indirecto de COVID-19 en las tasas de incidencia de enfermedades transmitidas por vector en México / Indirect impact of COVID-19 on the incidence rates of vector-borne diseases in Mexico

Resumen Introducción: La distribución espacial y temporal de la infección por SARS-CoV-2 sobrepasa las áreas endémicas de enfermedades transmitidas por vector (ETV), cuya vigilancia en México ha cambiado sustancialmente a partir del primer caso confirmado de COVID-19. Objetivos: Estimar y comparar las tasas de incidencia de las ETV antes y después de la introducción del SARS-CoV-2 en México. Métodos: Estudio retrospectivo de casos de ETV de 2014 a 2021. Las tasas de incidencia de cada ETV en el periodo previo (2014-2019) y posterior (2020-2021) a la introducción del SARS-CoV-2 en México fueron calculadas y comparadas. Resultados: Antes de la introducción del SARS-CoV-2, las tasas de incidencia de las ETV fueron altas y posterior a la introducción del coronavirus hubo un descenso en los índices epidemiológicos; sin embargo, solo se identificó diferencia estadística significativa en la tasa de incidencia de la malaria (p ≤ 0.05) y otras rickettsias (p ≤ 0.05). Conclusiones: Algunas medidas para reducir los casos de COVID-19, como el distanciamiento social, el confinamiento domiciliario, la reducción en el aforo en el transporte público y el trabajo en casa, probablemente contribuyeron a disminuir temporalmente el número de casos de las ETV; sin embargo, puede haber rebrote de las ETV en el futuro cercano.

Abstract Introduction: SARS-CoV-2 infection spatial and temporal distribution overlaps with endemic areas of vector-borne diseases (VBD), whose surveillance in Mexico has substantially changed since the first COVID-19 confirmed case. Objectives: To estimate and compare the incidence rates of VBDs before and after the introduction of SARS-CoV-2 in Mexico. Methods: Retrospective study of VBD cases from 2014 to 2021. The incidence rates of each VBD in the period before (2014-2019) and after (2020-2021) the introduction of SARS-CoV-2 in Mexico were calculated and compared. Results: Before the introduction of SARS-CoV-2, the incidence rates of VBDs were high and after the introduction of coronavirus there was a decrease in epidemiological indices; however, there was only statistically significant difference in the incidence rate of malaria (p ≤ 0.05) and other rickettsiae (p ≤ 0.05). Conclusions: Some measures to reduce COVID-19 cases, such as social distancing, home confinement, reductions in public transport and working at home (home office), probably temporarily decreased the number of VBD cases; however, there may be a resurgence of VBDs in the near future.

Indirect impact of COVID-19 on the incidence rates of vector-borne diseases in Mexico.

INTRODUCTION: SARS-CoV-2 infection spatial and temporal distribution overlaps with endemic areas of vector-borne diseases (VBD), whose surveillance in Mexico has substantially changed since the first COVID-19 confirmed case. OBJECTIVES: To estimate and compare the incidence rates of VBDs before and after the introduction of SARS-CoV-2 in Mexico. METHODS: Retrospective study of VBD cases from 2014 to 2021. The incidence rates of each VBD in the period before (2014-2019) and after (2020-2021) the introduction of SARS-CoV-2 in Mexico were calculated and compared. RESULTS: Before the introduction of SARS-CoV-2, the incidence rates of VBDs were high and after the introduction of coronavirus there was a decrease in epidemiological indices; however, there was only statistically significant difference in the incidence rate of malaria (p ≤ 0.05) and other rickettsiae (p ≤ 0.05). CONCLUSIONS: Some measures to reduce COVID-19 cases, such as social distancing, home confinement, reductions in public transport and working at home (home office), probably temporarily decreased the number of VBD cases; however, there may be a resurgence of VBDs in the near future.

INTRODUCCIÓN: La distribución espacial y temporal de la infección por SARS-CoV-2 sobrepasa las áreas endémicas de enfermedades transmitidas por vector (ETV), cuya vigilancia en México ha cambiado sustancialmente a partir del primer caso confirmado de COVID-19. OBJETIVOS: Estimar y comparar las tasas de incidencia de las ETV antes y después de la introducción del SARS-CoV-2 en México. MÉTODOS: Estudio retrospectivo de casos de ETV de 2014 a 2021. Las tasas de incidencia de cada ETV en el periodo previo (2014-2019) y posterior (2020-2021) a la introducción del SARS-CoV-2 en México fueron calculadas y comparadas. RESULTADOS: Antes de la introducción del SARS-CoV-2, las tasas de incidencia de las ETV fueron altas y posterior a la introducción del coronavirus hubo un descenso en los índices epidemiológicos; sin embargo, solo se identificó diferencia estadística significativa en la tasa de incidencia de la malaria (p ≤ 0.05) y otras rickettsias (p ≤ 0.05). CONCLUSIONES: Algunas medidas para reducir los casos de COVID-19, como el distanciamiento social, el confinamiento domiciliario, la reducción en el aforo en el transporte público y el trabajo en casa, probablemente contribuyeron a disminuir temporalmente el número de casos de las ETV; sin embargo, puede haber rebrote de las ETV en el futuro cercano.

Four Species of Under-reported Parasitic Arthropods in Mexico and their Potential Role as Vectors of Pathogens.

Numerous arthropod taxa are important in human and veterinary medicine. The salivary secretions and feces of arthropods can cause allergic reactions in host vertebrates or harbor pathogens. Also, bites can be a risk factor for secondary infections. Documenting the diversity of arthropods of medical and veterinary importance remains an important aspect of disease control and prevention. We provide new records of ectoparasitic arthropods from Mexico that are of potential medical or veterinary relevance. Scanning electron microscopy along with amplification and sequencing of a fragment of the mitochondrial gene (16S rRNA) was used to confirm some species identities. We report the cat louse Felicola subrostratus from cats and the chewing louse Heterodoxus spiniger from dogs, which are common ectoparasites but largely not reported in Mexico. The chigger Eutrombicula alfreddugesi is common on wild lizards (Squamata). For the first time, E. alfreddugesi is reported on Hemidactylus frenatus (common house gecko). This reptile has a close relationship with humans and its chiggers can cause dermatitis (i.e., trombiculiasis) or transmit pathogens. In addition, the common bed bug Cimex lectularius is reported for the first time in the state of Yucatan, an atypical area for its natural distribution. Phylogenetic analysis revealed that Cimex lectularius from Yucatan is closely related to genetic sequences of Cimex lectularius from China. Knowing the regional distribution of arthropods allows the design and implementation of prevention strategies for those that have potential roles as reservoirs or vectors.

Levels of Myeloperoxidase and Metalloproteinase-9 in Gingival Crevicular Fluid from Diabetic Subjects with and without Stage 2, Grade B Periodontitis.

OBJECTIVE: The present study aimed to compare levels of matrix metalloproteinase-9 (MMP-9) and myeloperoxidase (MPO) in gingival crevicular fluid (GCF) from subjects with controlled and noncontrolled Type 2 Diabetes Mellitus (T2D), with and without stage 2 grade B periodontitis (POD2B) versus healthy (H) subjects. METHODS: The levels of both enzymes, from 80 GCF samples collected with PerioPaper strips, were analyzed by a Multiplex/Luminex assay. Five groups were formed, all current patients at the Institutional Dentistry Service, and distributed as follows: two groups of diabetics (one controlled and one poorly controlled); two groups with the previous conditions and diagnosed with POD2B; and one H group. RESULTS: The highest concentration of MMP-9 corresponded to the H group, while the lowest corresponded to the T2D controlled group. Regarding MPO levels, the highest levels were associated with the T2D controlled with POD2B group and the lowest with the T2D controlled group. CONCLUSIONS: No apparent relationship between the elevation of MMP-9 and MPO levels was observed among subjects with T2D, with and without POD2B, compared to H subjects.

Toxoplasma gondii in Captive Wild Felids of Mexico: Its Frequency and Capability to Eliminate Oocysts.

There is little information about Toxoplasma gondii in wild felids, even when these species have been associated with cases of toxoplasmosis in humans. In this study, samples of serum and whole blood were collected from 42 felids from 10 different species, in 4 Mexican zoos. Stool samples from 36 animals were also collected, corresponding to 82% of the felids included in the study. Stool samples were used for the search of oocysts by light field microscopy and PCR. Serum samples were analyzed by indirect immunoglobulin G (IgG) enzyme-linked immunosorbent assay (ELISA) and indirect fluorescent antibody test (IFAT). DNA samples were purified from whole blood and stool for the amplification of a fragment of the SAG1 gene of T. gondii by a nested PCR (nPCR). The seroprevalence of IgG anti-T. gondii-specific antibodies by means of the ELISA was 100% (42/42) and 52.4% (22/42) by IFAT. The titers obtained varied from 1:80 to 1:2560. DNA of T. gondii was detected in 9.5% (4/42) of the blood samples by using nPCR. No oocysts were observed in the stool samples analyzed by light field microscopy. However, the DNA of the parasite was identified in 14.3% (5/35) of the stool samples evaluated. These results indicate a high prevalence of T. gondii in the studied populations of wild felids in captivity, with evidence of parasitemia and elimination of few oocysts even in adult hosts.

Frequency of Trypanosoma cruzi Infection in Synanthropic and Wild Rodents Captured in a Rural Community in Southeast of Mexico.

The protozoan parasite Trypanosoma cruzi is the causative agent of the Chagas disease, which is endemic in southeastern Mexico and is transmitted by the vector Triatoma dimidiata (triatomide). T. cruzi infect a great variety of domestic and wild mammals; rodents are considered one of the most important reservoirs of the parasite in the transmission cycles of T. cruzi. The objective of this study was to determine the frequency of T. cruzi infection and to determine the parasitic load in synanthropic and wild rodents from the rural community of southern Mexico. A total of 41 blood samples and 68 heart tissue samples were collected from various species of synanthropic (n= 48 in 2 species) and wild rodents (n= 35 in 5 species). DNA was extracted from samples to detect the presence of T. cruzi through quantitative PCR (qPCR). T. cruzi DNA was detected in the 9.75% of the blood samples of the synanthropic species (4/41) (14.28%) for Rattus rattus samples and 25% for Ototylomys phyllotis samples, with an average of parasitic load of 4.80 ± 1.17 parasites/µL. In the case of heart tissue samples, 10.29% were positive for T. cruzi (7/68) (8.7% for Rattus rattus, 40% for Peromyscus yucatanicus, and 42.8% for Ototylomys phyllotis) with an average parasite load of 3.15 ± 1.98 eq-parasites/mg. The active and chronic infection of T. cruzi in synanthropic or wild rodents of the rural community of southern Mexico evidences the natural infection in these reservoirs which contribute to maintaining the agent in the wild and domestic environments and can represent a risk of infection for the human population when the vector is present.

Quantitative and histological assessment of maternal-fetal transmission of Trypanosoma cruzi in guinea pigs: An experimental model of congenital Chagas disease.

OBJECTIVE: We evaluated the effect of Trypanosoma cruzi infection on fertility, gestation outcome, and maternal-fetal transmission in guinea pigs (Cavia porcellus). METHODS: Animals were infected with T. cruzi H4 strain (TcI lineage) before gestation (IBG) or during gestation (IDG). Tissue and sera samples of dams and fetuses were obtained near parturition. RESULTS: All IBG and IDG dams were seropositive by two tests, and exhibited blood parasite load of 1.62±2.2 and 50.1±62 parasites/µl, respectively, by quantitative PCR. Histological evaluation showed muscle fiber degeneration and cellular necrosis in all infected dams. Parasite nests were not detected in infected dams by histology. However, qPCR analysis detected parasites-eq/g heart tissue of 153±104.7 and 169.3±129.4 in IBG and IDG dams, respectively. All fetuses of infected dams were positive for anti-parasite IgG antibodies and tissue parasites by qPCR, but presented a low level of tissue inflammatory infiltrate. Fetuses of IDG (vs. IBG) dams exhibited higher degree of muscle fiber degeneration and cellular necrosis in the heart and skeletal tissues. The placental tissue exhibited no inflammatory lesions and amastigote nests, yet parasites-eq/g of 381.2±34.3 and 79.2±84.9 were detected in IDG and IBG placentas, respectively. Fetal development was compromised, and evidenced by a decline in weight, crow-rump length, and abdominal width in both groups. CONCLUSIONS: T. cruzi TcI has a high capacity of congenital transmission even when it was inoculated at a very low dose before or during gestation. Tissue lesions, parasite load, and fetal under development provide evidence for high virulence of the parasite during pregnancy. Despite finding of high parasite burden by qPCR, placentas were protected from cellular damage. Our studies offer an experimental model to study the efficacy of vaccines and drugs against congenital transmission of T. cruzi. These results also call for T. cruzi screening in pregnant women and adequate follow up of the newborns in endemic areas.

Serological survey of Leptospira interrogans, Toxoplasma gondii and Trypanosoma cruzi in free roaming domestic dogs and cats from a marginated rural area of Yucatan Mexico.

To evaluate the serological status for Trypanosoma cruzi, Toxoplasma gondii and Leptospira interrogans antibodies in free roaming dogs and cats from a marginated rural community in Yucatan Mexico, 100 households were visited and animals sampled. From the 106 samples, 93 were from dogs and 13 were from cats. Frequency of positive results for T. gondii, T. cruzi and Leptospira spp was 97.8%, 9.7% and 45.2% for dogs and 92.3%, 0.0% and 15.2% for cats, respectively. No associations with age, sex and body condition was found for T. gondii and Leptospira spp neither for the place where pets sleep, fumigation or presence of triatomes in the household in the case of T. cruzi. For leptospirosis the most common serovars found were Canicola, Autralis and Bratislava in dogs and cats with titres of 100 or 200 with exception of one dog with a titre of 400. The high frequency of seropositive dogs suggests a high circulation of the agents in the population of free roaming owned dogs and cats probably due to the lack of control of the reservoirs and vectors involved. Domestic animals in those rural communities can be sentinels to assess the risk of human exposure in the rural communities.

Comparative 2-D electrophoresis of salivary proteins in Triatoma dimidiata and Rhodnius prolixus (Hemiptera: Reduviidae) and major cross-reactive antigens

An immune response to triatomine's saliva is an immunological marker of exposure to triatomine bites. However, considerable variability in salivary protein profiles did exist among species. In the present work, we compare salivary proteins from Mexican Triatoma dimidiata and Rhodnius prolixus using 2-D electrophoresis. A clear differential saliva profile was found to exist between these two triatomine species. Fewer protein spots were detected in R. prolixus than in T. dimidiata. More than half of the proteins had an isoelectric point between 5 and 7 and a molecular weight between 10 and 30 kDa in T. dimidiata. Mice exposed to T. dimidiata saliva mount an immune response to three major cross-reacting antigens in R. prolixius saliva with weights of 10 kDa and 55 kDa. Our findings may alert for the presence of cross-reacting antigens between triatomine species in regions where two or more species are overlapping in the same geographical area.

Presence of Toxoplasma gondii in Drinking Water from an Endemic Region in Southern Mexico.

Toxoplasmosis can be acquired through the ingestion of contaminated drinking water with oocysts of Toxoplasma gondii, highly resistant to the routinely disinfection processes; based on chlorination commonly used in the water supply industry. The aim of this study was to determine the presence of T. gondii DNA in samples of public drinking water from an endemic region of southern Mexico. In total 74 samples of water (5 L each) were collected from the three well fields (I, II, and III) and 71 independent wells, distributing public drinking water to the city of Merida Yucatan, after passing through the chlorination process. Water samples were filtered and concentrated by a sucrose solution, then DNA was extracted and evaluated through a nested-PCR (nPCR) specific for T. gondii. Positive samples were detected in 5.4% (4/74) of the water samples. This is the first report of the presence of T. gondii DNA in public drinking water from a large city in southern Mexico, where their consumption without any postpurification treatment could pose a risk for acquiring the infection in the urban population.

Pre-exposure to faeces or saliva of Triatoma dimidiata decreases parasitemia in mice challenged with Trypanosoma cruzi: a description of the inflammatory reaction at the inoculation site

Under natural conditions, Trypanosoma cruzi infection is transmitted to mammals when faeces contaminated with metacyclic trypomastigotes gain access through skin lesions, mucosa or bite wounds. Natural infection of bugs with T. cruzi can vary greatly from less than 1% up to 70%, depending on triatomine species: in the case of Triatoma dimidiata, the percentage of infection is around 30%. In this work uses biological fluids (saliva and faeces) from Triatoma dimidiata to inoculate experimental animals once or multiple times, before inoculation with faeces contaminated with metacyclic trypomastigotes discrete type unit Ia (TcI). The site of infection was analyzed for histological changes based on hematoxile-eosine technique and toluide blue stain for mast cells. Inoculation with saliva led to the recruitment of eosinophils and mononuclear cells at the inoculation site, whereas inoculation with faeces led to the recruitment of neutrophils. Mice inoculated multiple times exhibited a strong inflammatory reaction from the first hour. Mono- or multi-exposure to T. dimidiata fluids before inoculation with metacyclic trypomastigotes helped to control the level of parasitemia. Previous contact with saliva or faeces of T. dimidiata reduces parasitemia in T. cruzi I -infected mice.

Presence of Toxoplasma gondii in Pork Intended for Human Consumption in Tropical Southern Mexico.

BACKGROUND: Toxoplasmosis is caused by the protozoon Toxoplasma gondii, which is one of the most widespread parasites that infect animals and humans worldwide. One of the main routes of infection for humans is through the consumption of infected meat containing bradyzoites in tissue cysts. Pork is one of the foremost meat types associated with outbreaks of acute toxoplasmosis in humans. MATERIALS AND METHODS: Sixty blood samples were collected from finished pigs at slaughter and their sera was evaluated by an indirect-IgG ELISA. Matched muscle samples were obtained from the tongue and loin. Whole blood and tissue samples were evaluated to search for T. gondii DNA using a nested-polymerase chain reaction. RESULTS: Seroprevalence of T. gondii was 96.6% (58/60) of sampled pigs. Meanwhile, T. gondii DNA was present in 23.21% of tongue tissue samples (13/56), 7% of loin tissues (4/57), and 0% in blood samples (0/44), respectively. Two pigs were serologically indeterminate. CONCLUSION: This is the first report of the presence of T. gondii DNA in tissue samples obtained from finalized pigs. Results from the present study suggest a high exposure to T. gondii in pigs intended for human consumption from the tropical region of Mexico. Thus, the consumption of some undercooked pork meat meals typical from the southern region of Mexico could represent a significant risk for acquiring infection for the human population.

American trypanosomiasis and associated risk factors in owned dogs from the major city of Yucatan, Mexico.

BACKGROUND: The American trypanosomiasis is a zoonosis caused by the protozoa Trypanosoma cruzi (T. cruzi). The disease is widely distributed throughout the American continent, affecting a wide range of hosts, including dogs. It is present in the canine population in the State of Yucatan, Mexico. However, no significant studies in owned dogs have been performed in the metropolitan area of Merida. A transversal study was conducted in 370 owned dogs from Merida, Yucatan, Mexico. METHODS: A cross-sectional study including 370 dogs was performed in a major city of Yucatan, Mexico, to detect IgG antibodies against T. cruzi. A commercial ELISA test kit was used and a chi-square test used to evaluate associated risk factors; odds ratio (OR) and 95 % confidence interval (CI) were also estimated. RESULTS: The indirect ELISA and western blot (WB) tests were used to detect specific immunoglobulin G antibodies against T. cruzi in serum samples. A prevalence of 12.2 % was found; age and area of residence were statistically associated with seropositivity in dogs (p <0.05). CONCLUSIONS: Results from the present study suggests the presence and abundance of the vector in urban conditions where a high number of seropositive cases of T. cruzi cases were found.

Multiplex Real-Time PCR Assay Using TaqMan Probes for the Identification of Trypanosoma cruzi DTUs in Biological and Clinical Samples.

BACKGROUND: Trypanosoma cruzi has been classified into six Discrete Typing Units (DTUs), designated as TcI-TcVI. In order to effectively use this standardized nomenclature, a reproducible genotyping strategy is imperative. Several typing schemes have been developed with variable levels of complexity, selectivity and analytical sensitivity. Most of them can be only applied to cultured stocks. In this context, we aimed to develop a multiplex Real-Time PCR method to identify the six T. cruzi DTUs using TaqMan probes (MTq-PCR). METHODS/PRINCIPAL FINDINGS: The MTq-PCR has been evaluated in 39 cultured stocks and 307 biological samples from vectors, reservoirs and patients from different geographical regions and transmission cycles in comparison with a multi-locus conventional PCR algorithm. The MTq-PCR was inclusive for laboratory stocks and natural isolates and sensitive for direct typing of different biological samples from vectors, reservoirs and patients with acute, congenital infection or Chagas reactivation. The first round SL-IR MTq-PCR detected 1 fg DNA/reaction tube of TcI, TcII and TcIII and 1 pg DNA/reaction tube of TcIV, TcV and TcVI reference strains. The MTq-PCR was able to characterize DTUs in 83% of triatomine and 96% of reservoir samples that had been typed by conventional PCR methods. Regarding clinical samples, 100% of those derived from acute infected patients, 62.5% from congenitally infected children and 50% from patients with clinical reactivation could be genotyped. Sensitivity for direct typing of blood samples from chronic Chagas disease patients (32.8% from asymptomatic and 22.2% from symptomatic patients) and mixed infections was lower than that of the conventional PCR algorithm. CONCLUSIONS/SIGNIFICANCE: Typing is resolved after a single or a second round of Real-Time PCR, depending on the DTU. This format reduces carryover contamination and is amenable to quantification, automation and kit production.