A small proportion of Zebu genetic background in crossbred calves may not be enough to improve resistance against natural bovine Babesia spp. infections.

The main objective of cattle breeders in tropical and subtropical regions is to acquire animals with taurine-productive traits adapted to the broad weather range of these regions. However, one of the main challenges on using taurine genetics in these areas is the high susceptibility of these animals to tick-borne diseases. Consequently, the present study evaluated from 10 November 2021-19 April 2022, the over 13 assessments, the Babesia bovis and Babesia bigemina DNA loads and the IgG anti-B. bovis and anti-B. bigemina levels in Angus (n = 17, 100% Taurine) and Ultrablack (n = 14, ∼82% taurine and 18% Zebu) calves. Data were analyzed using a multivariate mixed model with repeated measures of the same animal including the fixed effects of evaluation, genetic group, sex, Babesia spp., and their interactions. The repeatability values were estimated from the (co)variances matrix and expressed for each species. The correlations between the DNA loads (CNlog) and IgG titers (S/P) values for the two species were also estimated using the same model. Regarding the specific IgG antibody titers for both Babesia spp., no significant differences were observed between the two genetic groups. However, for B. bovis and B. bigemina DNA loads, Ultrablack calves presented significantly higher values than Angus calves. Under the conditions evaluated in this study, our findings suggest that the low percentage of Zebu genetic in the Ultrablack breed was insufficient to improve resistance against babesiosis. Further studies must demonstrate if the low percentages of Zebu genetics in Taurine breeds can modify the susceptibility to babesiosis infections.

Molecular monitoring of treatment efficacy in human visceral leishmaniasis.

BACKGROUND: Focused efforts of the visceral leishmaniasis elimination program have led to a drastic decline in cases, and the present challenge is disease monitoring, which this study aimed to assess. METHODS: A Leishmania kinetoplastid-targeted qPCR quantified parasite load at disease presentation, and following treatment completion (n=49); an additional 80 cases were monitored after completion of treatment. RESULTS: The parasite load at disease presentation was 13 461.00 (2560.00-37764.00)/µg gDNA, which upon completion of treatment reduced in 47 of 49 cases to 1(1-1)/µg gDNA, p<0.0001. In 80 cases that presented >2 months post-treatment, their parasite burden similarly decreased to 1(1-1)/µg gDNA except in 6 of 80 cases, which were qPCR positive. CONCLUSION: In 129 cases of visceral leishmaniasis, qPCR by quantification of parasite burden proved effective for monitoring treatment.

Preparing Collared Peccary (Pecari tajacu Linnaeus, 1758) for Reintroduction into the Wild: A Screening for Parasites and Hemopathogens of a Captive Population.

The reintroduction of captive animals to the wild helps restore endangered species, but it risks pathogen transmission, harming wild populations. Such transmission can impact the genetic diversity and long-term viability of these populations. This study assessed parasite diversity and load in captive Pecari tajacu, a species native to the Americas and culturally significant to Brazilian indigenous culture, prior to reintroduction. Samples from 24 peccaries were analyzed for ectoparasites, hemopathogens, and stool parasites with direct and molecular analysis. Findings showed that various parasites were present. Two peccaries (8.3%) were infested by the adult tick Amblyomma sculptum. Six (25.0%) tested positive for Trypanosoma evansi, four (16.7%) for hemobacteria of the family Anaplasmataceae, twelve (50.0%) for hemotropic Mycoplasma, and seven (29.2%) for Leishmania braziliensis. Stool samples indicated multiple parasites, with sixteen (66.7%) peccaries infected by Strongylida order parasites, Spiruridae in three (12.5%), and Ascaris suum in one (4.2%) animal. Cysts of Balantidium sp. were found in twenty (83.3%), Entamoeba polecki in five (20.8%), and Iodamoeba bütschlii in two (8.3%) peccaries. To our current knowledge, this is the first global report of Leishmania braziliensis, Iodamoeba bütschlii, and Entamoeba polecki in P. tajacu, irrespective of the environment, including both captivity and wild conditions. Some of these parasites are common in domestic animals, and others are zoonotic, indicating potential interspecies pathogen transmission.

Ivermectin decreases parasite load, testosterone, and potentially antler length in a group of captive red deer males (Cervus elaphus).

Secondary sex traits (SSTs) can favour males in intra-sexual competition, allowing females to reliably assess their quality. They can also be connected to other aspects of fitness, such as resistance to parasites and pathogens, as parasites have negative effects on the development of SSTs. Antlers are one of the most recognizable examples of SSTs whose development is regulated by testosterone and reflects the actual condition of the bearer. Elevated testosterone can exaggerate the size of SSTs while impairing the function of the immune system ("The Immunocompetence Handicap Hypothesis") posing a trade-off between antler development and immune function. In this study, we experimentally manipulated the parasite load in captive red deer (Cervus elaphus) males with Ivermectin during antler development for two consecutive years. Expecting an inverse proportionality between parasite load and antler size, we hypothesized the treated deer to have larger antlers than the untreated ones. Our results showed that, following the immunocompetence handicap hypothesis, parasite load was positively associated with testosterone levels. However, the application of Ivermectin suppressed the parasite load of the treated animals but did not lead to the development of larger antlers. Instead, it significantly suppressed the concentration of testosterone in the treated animals, whilst the animals that had higher testosterone also had the highest parasite load. Our findings show that Ivermectin can potentially decrease the levels of testosterone and, consequently, antler size. These findings have important implications for the management of captive populations, especially in contexts where the development of large trophies is desired.

MicroRNAs regulating macrophages infected with Leishmania L. (V.) Braziliensis isolated from different clinical forms of American tegumentary leishmaniasis.

Background: Leishmaniasis is an infectious disease caused by protozoa of the genus Leishmania. There are still no vaccines, and therapeutic options are limited, indicating the constant need to understand the fine mechanisms of its pathophysiology. An approach that has been explored in leishmaniasis is the participation of microRNAs (miRNAs), a class of small non-coding RNAs that act, in most cases, to repress gene expression. miRNAs play a role in the complex and plastic interaction between the host and pathogens, either as part of the host's immune response to neutralize infection or as a molecular strategy employed by the pathogen to modulate host pathways to its own benefit. Methods: Monocyte-derived macrophages from healthy subjects were infected with isolates of three clinical forms of L. braziliensis: cutaneous (CL), mucosal (ML), and disseminated (DL) leishmaniasis. We compared the expression of miRNAs that take part in the TLR/NFkB pathways. Correlations with parasite load as well as immune parameters were analyzed. Results: miRNAs -103a-3p, -21-3p, 125a-3p -155-5p, -146a-5p, -132- 5p, and -147a were differentially expressed in the metastatic ML and DL forms, and there was a direct correlation between miRNAs -103a-3p, -21-3p, -155-5p, -146a-5p, -132-5p, and -9-3p and parasite load with ML and DL isolates. We also found a correlation between the expression of miR-21-3p and miR-146a-5p with the antiapoptotic gene BCL2 and the increase of viable cells, whereas miR-147a was indirectly correlated with CXCL-9 levels. Conclusion: The expression of miRNAs is strongly correlated with the parasite load and the inflammatory response, suggesting the participation of these molecules in the pathogenesis of the different clinical forms of L. braziliensis.

Epidemiology of schistosomiasis in the town of Manjo, littoral - Region,Cameroon.

Background: Schistosomiasis is endemic in Cameroon and continues to cause serious public health problems, especially among populations in rural areas. This study aimed at determining the prevalence and risk factors of urinary and intestinal schistosomiasis in Manjo. Method: A cross-sectional study was conducted in the city of Manjo in 2020. Stool and urine samples were collected from 400 participants. These stool and urine samples were examined by the Kato Katz, and centrifugation methods respectively. Results: The results obtained showed an overall prevalence of 6.25%, with 5% and 1.25% for S. mansoni and S. haematobium respectively. A significant difference (p < 0.05) was revealed among occupations, age groups, neighborhood, water usage, educational level, knowledge of the disease meanwhile no significant difference was observed between gender and occupation according to prevalence. The most infected ages were] 50-; + [and]20-35] with 13.36% and 11.86% respectively. S. haematobium revealed a low infection intensity while S. mansoni showed moderate infection intensity. The mean parasite load for S. haematobium was 6 ± 3.225 Eggs/10 ml in females and 7 ± 4.243 Eggs/10 ml for males; while the mean parasitic load in S. mansoni was 180 ± 142.441 Epg in females and 146.67 ± 82.286 Epg in males. Conclusion: Manjo can be classified as a low endemic area with a prevalence rate of 6.25% and species observed were S. haematobium and S. mansoni. Also, risk factors where observed including the use of water from the river for domestic purposes. Therefore, the intensification of health education campaigns among the population would delay the development of this disease in the locality.

Validation of the NAT Chagas IVD Kit for the Detection and Quantification of Trypanosoma cruzi in Blood Samples of Patients with Chagas Disease.

In the absence of validated biomarkers to control the cure of Chagas disease, PCR-based diagnosis is being used as the main tool for an early indication of therapeutic failure. However, since it is considered a technique of complex reproducibility, mainly due to difficulties in establishing accurate controls to guarantee the quality of the reaction, the use of PCR for Chagas disease diagnosis is restricted to specialized centers. In an effort to disseminate the molecular diagnosis of Chagas disease and its applications, new diagnostic kits based on qPCR have been made available in the market in recent years. Here, we show the results of the validation of the NAT Chagas kit (Nucleic Acid Test for Chagas Disease) for the detection and quantification of T. cruzi in blood samples of patients suspected of Chagas disease infection. The kit, composed of a TaqMan duplex reaction targeting the T. cruzi satellite nuclear DNA and an exogenous internal amplification control, presented a reportable range from 104 to 0.5 parasite equivalents/mL and a limit of detection (LOD) of 0.16 parasite equivalents/mL of blood. In addition, the NAT Chagas kit detected T. cruzi belonging to all six discrete typing units (DTUs-TcI to TcVI), similarly to the in-house real-time PCR performed with commercial reagents, which has been selected as the best performance assay in the international consensus for the validation of qPCR for Chagas disease. In the clinical validation presented here, the kit showed 100% sensitivity and 100% specificity when compared to the consensus in-house real-time PCR assay. Thus, the NAT Chagas kit, which is produced entirely in Brazil under the international standards of good manufacturing practices (GMP), appears as an excellent alternative to enable the molecular diagnosis of Chagas disease in public and private diagnostic centers, as well as to facilitate the monitoring of patients under etiological treatment participating in clinical trials.

Molecular Diagnosis of Leishmaniasis in Spain: Development and Validation of Ready-To-Use Gel-Form Nested and Real-Time PCRs To Detect Leishmania spp.

Leishmaniasis is an endemic parasitic disease in at least 98 countries. In Spain, it is considered a zoonosis caused by Leishmania infantum, with an annual incidence of 0.62 cases/100,000 inhabitants. The predominant clinical manifestations are the cutaneous (CL) and visceral forms (VL), and the diagnosis is performed by parasitological, serological, and molecular tests. At the WHO Collaborating Center for Leishmaniasis (WHOCCLeish), routine diagnostic tests are based on a nested PCR (Ln-PCR), culture, and serological tests. To simplify our PCR protocol, we aimed to develop and validate a ready-to-use nested gel-form PCR (LeishGelPCR) and a duplex real-time PCR (qPCR) that allowed simultaneous detection of Leishmania and mammalian DNA as an internal control (Leish-qPCR). Clinical validation was performed in 200 samples from the WHOCCLeish collection; 92 and 85 out of 94 and 87 samples were positive by LeishGelPCR and Leish-qPCR, respectively, showing a sensitivity of 98% in both approaches. The specificity was 100% for LeishGelPCR and 98% for Leish-qPCR. The limits of detection of both protocols were similar (0.5 and 0.2 parasites/reaction). Parasite loads in VL and CL forms were similar, although high loads were observed when invasive samples were tested. In conclusion, LeishGelPCR and Leish-qPCR showed excellent performance in the diagnosis of leishmaniasis. These new forms of 18S rRNA gene PCR are equivalent to Ln-PCR and can be introduced in the algorithm for CL and VL diagnosis. IMPORTANCE Although the gold standard for diagnosis of leishmaniasis is the microscopic observation of amastigotes, molecular techniques are becoming a cost-efficient alternative. Currently, PCR is a routine resource that is used in many reference microbiology laboratories. In this article, we have described two ways to improve the reproducibility and usability of the molecular detection of Leishmania spp. These new approaches could be introduced even in middle- and low-resource laboratories; one is a ready-to-use gel-form system of a nested PCR and the other is a real-time PCR. We show why molecular diagnosis is the best methodology to confirm a clinical suspicion of leishmaniasis with higher sensitivity than traditional methods, thus facilitating early diagnosis and timely treatment of human leishmaniasis.

Establishment of a murine model of congenital toxoplasmosis and validation of a qPCR assay to assess the parasite load in maternal and fetal tissues.

Toxoplasma gondii is the causative agent of toxoplasmosis, a disease that affects warm-blooded animals and one third of the human population worldwide. Pregnant women who have never been exposed to the parasite constitute an important risk group, as infection during pregnancy often leads to congenital toxoplasmosis, the most severe form of the disease. Current therapy for toxoplasmosis is the same as it was 50 years ago and has little or no effect when vertical transmission occurs. Therefore, it is urgent to develop new strategies to prevent mother-to-fetus transmission. The implementation of experimental animal models of congenital toxoplasmosis that reproduces the transmission rates and clinical signs in humans opens an avenue of possibilities to interfere in the progression of the disease. In addition, knowing the parasite load in maternal and fetal tissues after infection, which may be related to organ abnormalities and disease outcome, is another important step in designing a promising intervention strategy. Therefore, we implemented here a murine model of congenital toxoplasmosis with outbred Swiss Webster mice infected intravenously with tachyzoites of the ME49 strain of T. gondii that mimics the frequency of transmission of the parasite, as well as important clinical signs of human congenital toxoplasmosis, such as macrocephaly, in addition to providing a highly sensitive quantitative real-time PCR assay to assess parasite load in mouse tissues. As the disease is not restricted to humans, also affecting several domestic animals, including companion animals and livestock, they can also benefit from the model presented in this study.

Parasitological, Molecular, and Histopathological Investigation of the Potential Activity of Propolis and Wheat Germ Oil against Acute Toxoplasmosis in Mice.

Toxoplasmosis is one of the most common parasitic zoonoses that affects all vertebrates. The drugs most commonly used against toxoplasmosis have many side effects, making the development of new antiparasitic drugs a big challenge. The present study evaluated the therapeutic effectiveness of novel herbal treatments, including propolis and wheat germ oil (WGO), against acute toxoplasmosis. A total of 50 albino mice were divided into five groups: group 1 (G1) (non-infected and non-treated); group 2 (G2) (infected without treatment); group 3 (G3) (treated with propolis); group 4 (G4) (treated with WGO); group 5 (G5) (treated with a combination of propolis and WGO). The effects of the herbal substances on different organs, mainly liver, spleen, and lungs, were investigated using parasitological, molecular, and histopathological examinations. The results of parasitological examination demonstrated statistically significant (p < 0.05) differences in the parasitic load between treated groups (G3, G4, and G5) compared to the control positive group (G2). These differences were represented by a significant reduction in the parasite load in stained tissue smears from the liver obtained from the animals treated with propolis (G3) compared to the parasite load in the positive control group. Similarly, animals (G4) treated with WGO exhibited a significant reduction in the parasite load versus the positive control group, while the lowest parasite load was found in G5, treated with propolis and WGO. Quantification of the parasite burden through molecular methods (PCR) revealed similar findings represented by reduction in the parasite burden in all treated groups with WGO and propolis as compared to the control group. Importantly, these previous parasitological and molecular findings were accompanied by a marked improvement in the histopathological picture of the liver, spleen, and lungs. In conclusion, propolis and WGO showed a good combination of therapeutic efficacy against acute toxoplasmosis.

The Use of an Adjuvant System Improves Innate and Adaptive Immune Response When Associated with a Leishmania (Viannia) braziliensis Antigen in a Vaccine Candidate against L. (Leishmania) infantum Infection.

BACKGROUND: The adjuvants' optimal dose and the administration route can directly influence the epitope recognition patterns and profiles of innate response. We aimed to establish the effect and the optimal dose of adjuvant systems for proposing a vaccine candidate to be employed with Leishmania (Viannia) braziliensis. METHODS: We evaluated the adjuvants saponin (SAP), monophosphoryl lipid A (MPL) and resiquimod (R-848) isolated and combined as adjuvant systems in a lower dose corresponding to 25%, 33%, and 50% of each adjuvant total dose. Male outbred BALB/c mice were divided into 13 groups, SAP, MPL, and R-848 isolated, and the adjuvant systems SAP plus MPL (SM), SAP plus R-848 (SR), and MPL plus R-848 (MR). RESULTS: SM50 increased levels of all chemokines analyzed and TNF production, while it presented an increased inflammatory cell infiltrate in the skin with macrophage recruitment. Thus, we proposed a vaccine candidate employing L. (V.) braziliensis antigen associated with the SM adjuvant system against experimental L. (Leishmania) infantum challenge. We observed a significant increase in the frequency of cells expressing the central and effector memory CD4+ T cells phenotype in immunized mice with the LBSM50. In the liver, there was a decreased parasite load when mice received LBSM50. CONCLUSIONS: When combined with L. (V.) braziliensis antigen, SM50 increases TNF and IFN-γ, which generates central and effector memory CD4+ T cells. Therefore, using an adjuvant system can promote an effective innate immune response with the potential to compose future vaccines.

Development of quantitative PCR and digital PCR for the quantification of Leishmania infantum in dogs.

Leishmaniasis is a zoonotic disease with worldwide distribution. In the Americas, the causative agent of the visceral form is the protozoa Leishmania (Leishmania) infantum. Transmission to the host or vertebrate reservoir occurs through the bite of infected arthropod females like Lutzomyia longipalpis. The epidemiological connection between the infection in dogs and humans generate constant studies about the relationship between the parasite and the canine host, including the development of methods and tests for the detection and quantification ofLeishmania (L.) infantum. Both conventional PCR (cPCR) and quantitative PCR (qPCR) can be used in the diagnosis of the parasite. Dropet Digital PCR (ddPCR) is another useful tool. Knowing the parasite load and its relationship with the clinical signs of naturally infected dogs is useful in research development and for establishing treatments that reduce the transmission of the disease. In this study, thirty-nine clinical samples of spleen from dogs naturaly infected by L. infantum were collected after necropsy. Two molecular tools were used to quantify the parasite load (qPCR and ddPCR) and there was 100% agreement in the results of the them. The tools developed in this work are important for the detection of L. infantum in dogs and humans. Droplet Digital PCR does not require a standard curve and is easy to standardize. In such manner, this new tool can generate more in-depth information in the broad debate about parasitic loads and the pathogenesis of leishmaniasis.

Helminths associated with terrestrial slugs in Swedish agricultural fields.

Slugs are important agricultural pests causing yearly yield losses. However, parasitizing helminths potentially could affect the size of the slug population. Here, a survey of terrestrial slug-parasitic helminths (nematodes and trematodes) was conducted for the first time in Sweden. In total, 268 terrestrial slugs were collected from 27 agricultural field edges in three seasons over 2020 and 2021 and dissected for presence of helminth parasites. Slugs belonging to the genus Arion were molecularly identified by mitochondrial DNA cytochrome c oxidase subunit I (COI) while parasites were identified using ribosomal RNA (18S). Overall, 13% of the collected slugs had helminth parasites and the likelihood of a slug being parasitized was highest in autumn. Slugs identified as Arion vulgaris were more likely to be parasitized than native slug species. The prevalence of nematodes and trematodes were similar; the dominant species found were Alloionema appendiculatum and Brachylaima thompsoni, respectively. This is the first record of the presence of these two species in Sweden.

Trypanosoma cruzi Survey in Poached Pichis (Zaedyus pichiy; Mammalia, Cingulata) from Mendoza, Argentina.

The transmission cycles of Trypanosoma cruzi, the causative agent of Chagas disease, include a wide variety of mammals and hematophagous triatomine insects. Infection with this blood parasite has been confirmed in many armadillo species; however, information on infection in Zaedyus pichiy, a small armadillo that inhabits areas endemic to Chagas disease, is scarce. Our objective was to determine the infection frequency and parasite load of T. cruzi in 49 wild Z. pichiy confiscated dead from poachers in Mendoza, Argentina, 2010-2017. We detected T. cruzi DNA in 32 of 49 armadillos (65%) using real-time PCR, confirming infection with T. cruzi in a high proportion of confiscated pichis. No differences were found related to sex, age, or ecoregion origin of the assessed pichis. Parasite loads ranged between <0.1 and 8.88 parasite equivalents/microgram cardiac tissue. Additional studies on the infection status of Z. pichiy are needed to determine their role in the maintenance of the sylvatic transmission cycle and the potential zoonotic risk from hunted pichis.

Large scale systemic control short-circuits pathogen transmission by interrupting the sand rat (Psammomys obesus)-to-sand fly (Phlebotomus papatasi) Leishmania major transmission cycle.

Systemic control uses the vertebrate hosts of zoonotic pathogens as "Trojan horses," killing blood-feeding female vectors and short-circuiting host-to-vector pathogen transmission. Previous studies focused only on the effect of systemic control on vector abundance at small spatial scales. None were conducted at a spatial scale relevant for vector control and none on the effect of systemic control on pathogen transmission rates. We tested the application of systemic control, using Fipronil-impregnated rodent baits, in reducing Leishmania major (Kinetoplastida: Trypanosomatidae; Yakimoff & Schokhor, 1914) infection levels within the vector, Phlebotomus papatasi (Diptera: Psychodidae; Scopoli, 1786) population, at the town-scale. We provided Fipronil-impregnated food-baits to all Psammomys obesus (Mammalia:Muridae; Cretzschmar, 1828), the main L. major reservoir, burrows along the southern perimeter of the town of Yeruham, Israel, and compared sand fly abundance and infection levels with a non-treated control area. We found a significant and substantial treatment effect on L. major infection levels in the female sand fly population. Sand fly abundance was not affected. Our results demonstrate, for the first time, the potential of systemic control in reducing pathogen transmission rates at a large, epidemiologically relevant, spatial scale.

Toxoplasma gondii SAG2, SAG3 and GRA6 alleles and single nucleotide polymorphism in congenital infections with known parasite load and clinical outcome

ABSTRACT Amniotic fluid DNA samples were genotyped by multilocus-nested-PCR-RFLP, but only three of 11 markers amplified 113 of 122 (92.6%) samples, resulting in 12 untyped and 101 partial non-archetypal genotypes. The 101 typed samples were subdivided into four groups: G1 with 73 samples (5'and 3' SAG2 allele I + SAG3 allele III + GRA6 allele III), 53 had parasite load ≤ 102 parasites/mL (43 asymptomatic, 10 mild infections), 17 had load > 102 and ≤ 103 (one mild, 13 moderate and three severe), and three had load > 103 parasites/mL (three severe); G2 with 22 samples (5'and 3' SAG2 allele I + SAG3 allele III), all parasite load levels ≤ 102 parasites/mL (18 asymptomatic and four mild); G3 with five samples (5' and 3' SAG2 allele I + SAG3 allele II), parasite load ≤ 102 parasites/mL (three asymptomatic and two mild); G4 with one sample (5' and 3' SAG2 allele II + SAG3 allele II + GRA6 allele I), a parasite load < 102 parasites/mL in an asymptomatic infant. After DNA sequencing, restriction sites confirmed SAG2, SAG3 and GRA6 alleles in 98.7%, 100% and 100% of the cases, respectively, while single nucleotide polymorphisms confirmed 90% of 5'-SAG2 allele I; 98.7% of 3'-SAG2 allele I; 98% of SAG-3 allele III, but only 40% of GRA6 allele III results. For the moment, partial non-archetypal genotypes of parasites did not show any relationship with either parasite load in amniotic fluid samples or clinical outcome of infants at the age of 12 months.

Trypanosoma cruzi Parasite Burdens of Several Triatomine Species in Colombia.

Trypanosoma cruzi, the causal agent of Chagas disease, is mainly transmitted by insects of the Triatominae subfamily. In Colombia, there are 26 triatomine species, and 16 of them are naturally infected with the parasite. The parasite loads of naturally infected vectors can be significant in targeting specific species that can affect the epidemiology of the disease. Studying their ecology and behavior is vital to understand their role in T. cruzi transmission dynamics. We evaluated the parasite loads of 182 field-collected triatomines corresponding to 10 species in 13 departments across Colombia. We standardized a methodology to quantify T. cruzi DNA in these insects. We obtained a LOD (limit of detection) of 3.05 p-eq/mL. The 82% of triatomines we evaluated were positive for T. cruzi infection, with loads ranging from hundreds to millions of equivalent parasites per milliliter. Panstrongylus geniculatus, Rhodnius prolixus, and Triatoma dimidiata were the species with the highest loads of T. cruzi; however, other species whose role as vectors is still unknown were also found with high loads of parasites. Our results suggest the relevance of secondary species for T. cruzi transmission in Colombia. We hope our data can help improve entomological surveillance and vector control programs in the country and the region.

Host-Parasite Relationship-Nematode Communities in Populations of Small Mammals.

Nematode burdens and variation in morphological characteristics were assessed in eighty-eight animals from three host species (Apodemus sylvaticus, Apodemus flavicollis, and Myodes glareolus) from eight localities in Serbia. In total, 15 species of nematodes were identified, and the overall mean parasite species richness (IndPSR) was 1.61 per animal (1.98 in A. flavicollis, 1.43 in M. glareolus, and 0.83 in A. sylvaticus). Furthermore, the studied host species significantly differed in individual parasite load (IndPL) and in the following morphological characters: spleen mass, body condition index (BCI), and body mass. We aimed to analyze the relationship between the burden of intestinal nematodes, on one hand, and the body conditions of the host and its capability to develop immune defends on the other. Spleen mass was considered as a measure of immune response. In all host species, larger animals with a better condition (higher BCI) were infected with more parasites species (IndPSR), while parasite load was not related to BCI. Only in A. flavicollis were males significantly larger, but females of the same sizes were infected with more parasite species. This female-biased parasitism is contrary to the theoretical expectation that males should be more parasitized, being larger, more active, with a wider home range. Although the spleen size was significantly correlated with body condition and body mass, IndPSR was not related to spleen mass in any studied species, but in M. galareolus, we found that a smaller spleen was related to higher infection intensity (IndPL).

Parasitemia Levels in Trypanosoma cruzi Infection in Spain, an Area Where the Disease Is Not Endemic: Trends by Different Molecular Approaches.

Trypanosoma cruzi infection has expanded globally through human migration. In Spain, the mother-to-child route is the mode of transmission contributing to autochthonous Chagas disease (CD); however, most people acquired the infection in their country of origin and were diagnosed in the chronic phase (imported chronic CD). In this context, we assessed the quantitative potential of the Loopamp Trypanosoma cruzi detection kit (Sat-TcLAMP) based on satellite DNA (Sat-DNA) to determine parasitemia levels compared to those detected by real-time quantitative PCRs (qPCRs) targeting Sat-DNA (Sat-qPCR) and kinetoplast DNA minicircles (kDNA-qPCR). This study included 173 specimens from 39 autochthonous congenital and 116 imported chronic CD cases diagnosed in Spain. kDNA-qPCR showed higher sensitivity than Sat-qPCR and Sat-TcLAMP. According to all quantitative approaches, parasitemia levels were significantly higher in congenital infection than in chronic CD (1 × 10-1 to 5 × 105 versus >1 × 10-1 to 6 × 103 parasite equivalents/mL, respectively [P < 0.001]). Sat-TcLAMP, Sat-qPCR, and kDNA-qPCR results were equivalent at high levels of parasitemia (P = 0.381). Discrepancies were significant for low levels of parasitemia and older individuals. Differences between Sat-TcLAMP and Sat-qPCR were not qualitatively significant, but estimations of parasitemia using Sat-TcLAMP were closer to those by kDNA-qPCR. Parasitemia changes were assessed in 6 individual cases in follow-up, in which trends showed similar patterns by all quantitative approaches. At high levels of parasitemia, Sat-TcLAMP, Sat-qPCR, and kDNA-qPCR worked similarly, but significant differences were found for the low levels characteristic of late chronic CD. A suitable harmonization strategy needs to be developed for low-level parasitemia detection using Sat-DNA- and kDNA-based tests. IMPORTANCE Currently, molecular equipment has been introduced into many health care centers, even in low-income countries. PCR, qPCR, and loop-mediated isothermal amplification (LAMP) are becoming more accessible for the diagnosis of neglected infectious diseases. Chagas disease (CD) is spreading worldwide, and in countries where the disease is not endemic, such as Spain, the parasite Trypanosoma cruzi is transmitted from mother to child (congenital CD). Here, we explore why LAMP, aimed at detecting T. cruzi parasite DNA, is a reliable option for the diagnosis of congenital CD and the early detection of reactivation in chronic infection. When the parasite load is high, LAMP is equivalent to any qPCR. In addition, the estimations of T. cruzi parasitemia in patients living in Spain, a country where the disease is not endemic, resemble natural evolution in areas of endemicity. If molecular tests are introduced into the diagnostic algorithm for congenital infection, early diagnosis and timely treatment would be accomplished, so the interruption of vertical transmission can be an achievable goal.