Parasite Burden Research Articles

Assessment of health problems of sheep and goats based on ante-mortem and post-mortem inspection at Addis Ababa Abattoir, Ethiopia.

Ethiopia has a rapidly growing small ruminant sector, which faces low productivity due to husbandry practices and poor health condition of the animals. A study was conducted in Ethiopia's largest municipal abattoir with the objective to assess the health problems of sheep and goats presented for slaughter using standard ante-mortem and post-mortem methodology. A cross-sectional study using systematic random sampling was conducted on 384 sheep and 384 goats from January to July 2014. Soiled skin (69.1%), poor body condition (24.3%), and nostril discharge (19.5%) were common among both species at ante-mortem examination. Gross lesions were frequent in livers (39.7%) and lungs (37.2%), while pneumonia (18.1%) and adhesions (13.8%) were frequent in the lungs of sheep and goats, indicating stress-related illness. Parasitic lesions, especially fasciolosis (19.3%) and hydatid cysts (8.1%) were significantly more common in sheep livers (p 0.05). The direct financial loss from lesions in both species was 1,077,015 ETB or 53,851 USD per year, most of which was estimated to occur from carcass bruising. The findings indicate that reducing parasite burden and preventing carcass bruising through improved handling could significantly increase the profitability of the small ruminant meat sector in Ethiopia.

Infection dynamics following experimental challenge of pigs orally dosed with different stages of two archetypal genotypes of Toxoplasma gondii

Toxoplasma gondii is a food-borne zoonotic parasite widespread in a variety of hosts, including humans. With a majority of infections in Europe estimated to be meat-borne, pork, as one of the most consumed meats worldwide, represents a potential risk for consumers. Therefore, we aimed to investigate the progress of T. gondii infection and tissue tropism in experimentally infected pigs, using different T. gondii isolates and infectious stages, i.e. tissue cysts or oocysts. Twenty-four pigs were allocated to treatment in four groups of six, with each group inoculated orally with an estimated low dose of either 400 oocysts or 10 tissue cysts of two European T. gondii isolates, a type II and a type III isolate. The majority of pigs seroconverted two weeks post-inoculation. Pigs infected with the type III isolate had significantly higher levels of anti-T. gondii antibodies compared to those infected with the type II isolate. Histopathological exams revealed reactive hyperplasia of the lymphatic tissue of all pigs. Additionally, a selected set of nine tissues was collected during necropsy at 50 dpi from each of the remaining 22 pigs for T. gondii DNA detection by quantitative real-time PCR. A positive result was obtained in 29.8 % (59/139) of tested tissues. The brain was identified as the most frequently positive tissue in 63.6 % (14/22) of the animals. In contrast, liver samples tested negative in all animals. The highest mean parasite load, calculated by interpolating the average Cq values on the standard curve made of ten-fold serial dilutions of the genomic DNA, corresponding to 100 to 104 tachyzoites/µL, was observed in shoulder musculature with an estimated concentration of 84.4 [0.0–442.5] parasites per gram of tissue. The study highlights the variability in clinical signs and tissue distribution of T. gondii in pigs based on the combination of parasite stages and strains, with type III isolates, particularly oocysts, causing a stronger antibody response and higher tissue parasite burden. These findings suggest the need for further investigation of type III isolates to better understand their potential risks to humans.

Protective Efficacy of the Epitope-Conjugated Antigen N-Tc52/TSkb20 in Mitigating Trypanosoma cruzi Infection through CD8+ T-Cells and IFNγ Responses.

Chagas disease, caused by the protozoan Trypanosoma cruzi, remains a major public health challenge affecting millions in Latin America and worldwide. Although significant progress has been made in vector control, no vaccine exists to prevent infection or mitigate disease pathogenesis. We developed a rationally designed chimeric protein vaccine, N-Tc52/TSkb20, incorporating immunodominant epitopes from two T. cruzi antigens, the amino-terminal portion of Tc52 and the TSkb20 epitope derived from trans-sialidase. The objectives of this study were to construct and characterize the antigen and evaluate its protective potential in an immunoprophylactic murine model of T. cruzi infection. The N-Tc52/TSkb20 protein was recombinantly expressed in E. coli and its identity was confirmed using mass spectrometry and Western blotting. Immunization with the chimeric protein significantly controlled parasitemia and reduced the heart, colon, and skeletal muscle parasite burdens compared to non-vaccinated mice. Protection was superior to vaccination with the individual parental antigen components. Mechanistically, the vaccine induced potent CD8+ T-cell and IFNγ responses against the incorporated epitopes and a protective IgG antibody profile. A relatively low IL-10 response favored early parasite control. These results validate the promising multi-epitope approach and support the continued development of this type of rational vaccine design strategy against Chagas disease.

Gyrodactylus in the spotlight: how exposure to light impacts disease and the feeding behavior of the freshwater tropical guppy (Poecilia reticulata).

Artificial light at night (ALAN) negatively impacts organisms in many ways, from their feeding behaviors to their response and ability to deal with disease. Our knowledge of ALAN is focused on hosts, but we must also consider their parasites, which constitute half of all described animal species. Here, we assessed the impact of light exposure on a model host-parasite system (Poecilia reticulata and the ectoparasitic monogenean Gyrodactylus turnbulli). First, parasite-free fish were exposed to 12:12 h light:dark (control) or 24:0 h light:dark (ALAN) for 21 days followed by experimental infection. Second, naturally acquired G. turnbulli infections were monitored for 28 days during exposure of their hosts to a specified light regime (6:18 h, 12:12 h, or 24:0 h light:dark). Experimentally infected fish exposed to constant light had, on average, a greater maximum parasite burden than controls, but no other measured parasite metrics were impacted. Host feeding behavior was also significantly affected: fish under ALAN fed faster and took more bites than controls, whilst fish exposed to reduced light fed slower. Thus, ALAN can impact parasite burdens, even in the short term, and altering light conditions will impact fish feeding behavior. Such responses could initiate disease outbreaks or perturb food-webs with wider ecological impacts.

ACV with/without IVM: a new talk on intestinal CDX2 and muscular CD34 and Cyclin D1 during Trichinella spiralis infection.

The current study assessed the efficacy of Acyclovir (ACV) and Ivermectin (IVM) as monotherapies and combined treatments for intestinal and muscular stages of Trichinella spiralis infection. One-hundred Swiss albino mice received orally 250 ± 50 infectious larvae and were divided into infected-untreated (Group-1), IVM-treated (Group-2), ACV-treated (Group-3), combined IVM+ACV (Group-4), and healthy controls (Group-5). Each group was subdivided into subgroup-A-enteric phase (10 mice, sacrificed day-7 p.i.) and subgroup-B-muscular phase (10 mice, sacrificed day-35 p.i.). Survival rate and body weight were recorded. Parasite burden and intestinal histopathology were assessed. In addition, immunohistochemical expression of epithelial CDX2 in the intestinal phase and CyclinD1 as well as CD34 in the muscular phase were evaluated. Compared, IVM and ACV monotherapies showed insignificant differences in the amelioration of enteric histopathology, except for lymphocytic counts. In the muscle phase, monotherapies showed variable disruptions in the encapsulated larvae. Compared with monotherapies, the combined treatment performed relatively better improvement of intestinal inflammation and reduction in the enteric and muscular parasite burden. CDX2 and CyclinD1 positively correlated with intestinal inflammation and parasite burden, while CD34 showed a negative correlation. CDX2 positively correlated with CyclinD1. CD34 negatively correlated with CDX2 and CyclinD1. IVM +ACV significantly ameliorated CDX2, CyclinD1, and CD34 expressions compared with monotherapies. Conclusion. T. spiralis infection-associated inflammation induced CDX2 and CyclinD1 expressions, whereas CD34 was reduced. The molecular tumorigenic effect of the nematode remains questionable. Nevertheless, IVM +ACV appeared to be a promising anthelminthic anti-inflammatory combination that, in parallel, rectified CDX2, CyclinD1, and CD34 expressions.

Virus-like particles expressing microneme-associated antigen of Plasmodium berghei confer better protection than those expressing apical membrane antigen 1.

Malaria is a global disease affecting a large portion of the world's population. Although vaccines have recently become available, their efficacies are suboptimal. We generated virus-like particles (VLPs) that expressed either apical membrane antigen 1 (AMA1) or microneme-associated antigen (MIC) of Plasmodium berghei and compared their efficacy in BALB/c mice. We found that immune sera acquired from AMA1 VLP- or MIC VLP-immunized mice specifically interacted with the antigen of choice and the whole P. berghei lysate antigen, indicating that the antibodies were highly parasite-specific. Both VLP vaccines significantly enhanced germinal center B cell frequencies in the inguinal lymph nodes of mice compared with the control, but only the mice that received MIC VLPs showed significantly enhanced CD4+ T cell responses in the blood following P. berghei challenge infection. AMA1 and MIC VLPs significantly suppressed TNF-α and interleukin-10 production but had a negligible effect on interferon-γ. Both VLPs prevented excessive parasitemia buildup in immunized mice, although parasite burden reduction induced by MIC VLPs was slightly more effective than that induced by AMA1. Both VLPs were equally effective at preventing body weight loss. Our findings demonstrated that the MIC VLP was an effective inducer of protection against murine experimental malaria and should be the focus of further development.

Monocytes, particularly nonclassical ones, lose their opsonic and nonopsonic phagocytosis capacity during pediatric cerebral malaria.

Innate immunity is crucial to reducing parasite burden and contributing to survival in severe malaria. Monocytes are key actors in the innate response and, like macrophages, are plastic cells whose function and phenotype are regulated by the signals from the microenvironment. In the context of cerebral malaria (CM), monocyte response constitutes an important issue to understand. We previously demonstrated that decreased percentages of nonclassical monocytes were associated with death outcomes in CM children. In the current study, we postulated that monocyte phagocytosis function is impacted by the severity of malaria infection. To study this hypothesis, we compared the opsonic and nonopsonic phagocytosis capacity of circulant monocytes from Beninese children with uncomplicated malaria (UM) and CM. For the CM group, samples were obtained at inclusion (D0) and 3 and 30 days after treatment (D3, D30). The phagocytosis capacity of monocytes and their subsets was characterized by flow cytometry and transcriptional profiling by studying genes known for their functional implication in infected-red blood cell (iRBC) elimination or immune escape. Our results confirm our hypothesis and highlight the higher capacity of nonclassical monocytes to phagocyte iRBC. We also confirm that a low number of nonclassical monocytes is associated with CM outcome when compared to UM, suggesting a mobilization of this subpopulation to the cerebral inflammatory site. Finally, our results suggest the implication of the inhibitory receptors LILRB1, LILRB2, and Tim3 in phagocytosis control. Taken together, these data provide a better understanding of the interplay between monocytes and malaria infection in the pathogenicity of CM.

Amphotericin B resistance in Leishmania amazonensis: In vitro and in vivo characterization of a Brazilian clinical isolate.

In Brazil, Leishmania amazonensis is the etiological agent of cutaneous and diffuse cutaneous leishmaniasis. The state of Maranhão in the Northeast of Brazil is prevalent for these clinical forms of the disease and also has high rates of HIV infection. Here, we characterized the drug susceptibility of a L. amazonensis clinical isolate from a 46-year-old man with diffuse cutaneous leishmaniasis coinfected with HIV from this endemic area. This patient underwent several therapeutic regimens with meglumine antimoniate, liposomal amphotericin B, and pentamidine, without success. In vitro susceptibility assays against promastigotes and intracellular amastigotes demonstrated that this isolate had low susceptibility to amphotericin B, when compared with the reference strain of this species that is considered susceptible to antileishmanial drugs. Additionally, we investigated whether the low in vitro susceptibility would affect the in vivo response to amphotericin B treatment. The drug was effective in reducing the lesion size and parasite burden in mice infected with the reference strain, whereas those infected with the clinical isolate and a resistant line (generated experimentally by stepwise selection) were refractory to amphotericin B treatment. To evaluate whether the isolate was intrinsically resistant to amphotericin B in animals, infected mice were treated with other drugs that had not been used in the treatment of the patient (miltefosine, paromomycin, and a combination of both). Our findings demonstrated that all drug schemes were able to reduce lesion size and parasite burden in animals infected with the clinical isolate, confirming the amphotericin B-resistance phenotype. These findings indicate that the treatment failure observed in the patient may be associated with amphotericin B resistance, and demonstrate the potential emergence of amphotericin B-resistant L. amazonensis isolates in an area of Brazil endemic for cutaneous leishmaniasis.

MAPK/ERK activation in macrophages promotes Leishmania internalization and pathogenesis

The obligate intracellular parasite Leishmania binds several receptors to trigger uptake by phagocytic cells, ultimately resulting in visceral or cutaneous leishmaniasis. A series of signaling pathways in host cells, which are critical for establishment and persistence of infection, are activated during Leishmania internalization. Thus, preventing Leishmania uptake by phagocytes could be a novel therapeutic strategy for leishmaniasis. However, the host cellular machinery mediating promastigote and amastigote uptake is not well understood. Here, using small molecule inhibitors of Mitogen-activated protein/Extracellular signal regulated kinases (MAPK/ERK), we demonstrate that ERK1/2 mediates Leishmania amazonensis uptake and (to a lesser extent) phagocytosis of beads by macrophages. We find that inhibiting host MEK1/2 or ERK1/2 leads to inefficient amastigote uptake. Moreover, using inhibitors and primary macrophages lacking spleen tyrosine kinase (SYK) or Abl family kinases, we show that SYK and Abl family kinases mediate Raf, MEK, and ERK1/2 activity and are necessary for uptake. Finally, we demonstrate that trametinib, a MEK1/2 inhibitor used to treat cancer, reduces disease severity and parasite burden in Leishmania-infected mice, even if it is started after lesions develop. Our results show that maximal Leishmania infection requires MAPK/ERK and highlight potential for MAPK/ERK-mediated signaling pathways to be novel therapeutic targets for leishmaniasis.

Landscape drives zoonotic malaria prevalence in non-human primates.

Zoonotic disease dynamics in wildlife hosts are rarely quantified at macroecological scales due to the lack of systematic surveys. Non-human primates (NHPs) host Plasmodium knowlesi, a zoonotic malaria of public health concern and the main barrier to malaria elimination in Southeast Asia. Understanding of regional P. knowlesi infection dynamics in wildlife is limited. Here, we systematically assemble reports of NHP P. knowlesi and investigate geographic determinants of prevalence in reservoir species. Meta-analysis of 6322 NHPs from 148 sites reveals that prevalence is heterogeneous across Southeast Asia, with low overall prevalence and high estimates for Malaysian Borneo. We find that regions exhibiting higher prevalence in NHPs overlap with human infection hotspots. In wildlife and humans, parasite transmission is linked to land conversion and fragmentation. By assembling remote sensing data and fitting statistical models to prevalence at multiple spatial scales, we identify novel relationships between P. knowlesi in NHPs and forest fragmentation. This suggests that higher prevalence may be contingent on habitat complexity, which would begin to explain observed geographic variation in parasite burden. These findings address critical gaps in understanding regional P. knowlesi epidemiology and indicate that prevalence in simian reservoirs may be a key spatial driver of human spillover risk.

Landscape drives zoonotic malaria prevalence in non-human primates

Zoonotic disease dynamics in wildlife hosts are rarely quantified at macroecological scales due to the lack of systematic surveys. Non-human primates (NHPs) host Plasmodium knowlesi, a zoonotic malaria of public health concern and the main barrier to malaria elimination in Southeast Asia. Understanding of regional P. knowlesi infection dynamics in wildlife is limited. Here, we systematically assemble reports of NHP P. knowlesi and investigate geographic determinants of prevalence in reservoir species. Meta-analysis of 6322 NHPs from 148 sites reveals that prevalence is heterogeneous across Southeast Asia, with low overall prevalence and high estimates for Malaysian Borneo. We find that regions exhibiting higher prevalence in NHPs overlap with human infection hotspots. In wildlife and humans, parasite transmission is linked to land conversion and fragmentation. By assembling remote sensing data and fitting statistical models to prevalence at multiple spatial scales, we identify novel relationships between P. knowlesi in NHPs and forest fragmentation. This suggests that higher prevalence may be contingent on habitat complexity, which would begin to explain observed geographic variation in parasite burden. These findings address critical gaps in understanding regional P. knowlesi epidemiology and indicate that prevalence in simian reservoirs may be a key spatial driver of human spillover risk.

Risk-Based Gastrointestinal Parasite Control in a Tropical Zoological Institute

The surveillance and treatment of parasites are important features of preventative healthcare plans in zoological institutes. The parasite control strategies employed in temperate regions often involve prophylactic anthelmintic treatments during seasons where the burden of gastrointestinal parasites in fecal testing is high. These strategies are, however, not applicable in the tropics, where temperatures remain high throughout the year, allowing continuous parasitic development. A risk-based parasite management strategy was adopted by a tropical zoological institute. For parasite surveillance, routine fecal direct smears and magnesium sulfate flotations were performed to determine parasitic prevalence. The frequency of fecal checks for the year was determined by the frequency at which clinically relevant parasitism (fecal tests that resulted in the animal being treated) was detected during routine fecal checks in the previous year. A yearly anthelmintic drug-class rotation schedule was also implemented. The total number of fecal tests performed per year and the number of animals with clinically significant parasitic disease decreased by 30.0% (637/2126) and 46.9% (207/451), respectively, over the four-year period of the study. Anthelmintic class rotation also improved the efficacy of fenbendazole in treating Strongyloides spp. infecting the group of orangutans. This parasite control strategy is a targeted approach to managing preventative healthcare, reducing the work required to perform routine surveillance tests whilst maintaining the health of the collection of animals.

Preclinical evaluation of combined therapy with amiodarone and low-dose benznidazole in a mouse model of chronic Trypanosoma cruzi infection

Chagasic chronic cardiomyopathy (CCC) is the primary clinical manifestation of Chagas disease (CD), caused by Trypanosoma cruzi. Current therapeutic options for CD are limited to benznidazole (Bz) and nifurtimox. Amiodarone (AMD) has emerged as most effective drug for treating the arrhythmic form of CCC. To address the effects of Bz and AMD we used a preclinical model of CCC. Female C57BL/6 mice were infected with T. cruzi and subjected to oral treatment for 30 consecutive days, either as monotherapy or in combination. AMD in monotherapy decreased the prolonged QTc interval, the incidence of atrioventricular conduction disorders and cardiac hypertrophy. However, AMD monotherapy did not impact parasitemia, parasite load, TNF concentration and production of reactive oxygen species (ROS) in cardiac tissue. Alike Bz therapy, the combination of Bz and AMD (Bz/AMD), improved cardiac electric abnormalities detected T. cruzi-infected mice such as decrease in heart rates, enlargement of PR and QTc intervals and increased incidence of atrioventricular block and sinus arrhythmia. Further, Bz/AMD therapy ameliorated the ventricular function and reduced parasite burden in the cardiac tissue and parasitemia to a degree comparable to Bz monotherapy. Importantly, Bz/AMD treatment efficiently reduced TNF concentration in the cardiac tissue and plasma and had beneficial effects on immunological abnormalities. Moreover, in the cardiac tissue Bz/AMD therapy reduced fibronectin and collagen deposition, mitochondrial damage and production of ROS, and improved sarcomeric and gap junction integrity. Our study underlines the potential of the Bz/AMD therapy, as we have shown that combination increased efficacy in the treatment of CCC.

Leishmania exploits host cAMP/EPAC/calcineurin signaling to induce an IL-33–mediated anti-inflammatory environment for the establishment of infection

Host anti-inflammatory responses are critical for the progression of visceral leishmaniasis and the pleomorphic cytokine IL-33 was found to be upregulated in infection. The underlying mechanism is not yet known. Here, we documented that IL-33 induction is a consequence of elevated cAMP-mediated EPAC/calcineurin-dependent signaling and is essential for the sustenance of infection. L. donovani-infected RAW and bone marrow-derived macrophages showed significant up-regulation of IL-33 and its neutralization by anti-IL-33 antibody resulted in decreased parasite survival and increased inflammatory responses. Infection-induced elevated cAMP was involved in IL-33 production and of its two downstream effectors PKA and EPAC, only the latter was responsible for elevated IL-33 levels. EPAC initiated Rap-dependent phospholipase C activation, which triggered the release of intracellular calcium followed by calcium/calmodulin complex formation. Screening of calmodulin-dependent enzymes affirmed the involvement of the phosphatase calcineurin in cAMP/EPAC/calcium/calmodulin signaling-induced IL-33 production and parasite survival. Activated calcineurin thereby ensured nuclear localization of the transcription factors NFATc1 and HIF-1α for IL-33 transcription and we further ascertained their role in IL-33 transcription using a ChIP assay. Administering specific inhibitors of NFATc1 and HIF-1α in a BALB/c mouse model of visceral leishmaniasis led to decreased liver and spleen parasite burden with concomitant decrease in IL-33 levels. Splenocyte supernatants of inhibitor-treated infected mice further documented an increase in TNF-α and IL-12 with simultaneous decrease of IL-10, thereby indicating an overall disease-escalating effect of IL-33. Thus, this study demonstrates that cAMP/EPAC/calcineurin signaling is crucial for the activation of IL-33 and in effect creates anti-inflammatory responses, essential for infection.

In Vivo Bioluminescence Imaging Reveals Differences in Leishmania infantum Parasite Killing Kinetics by Antileishmanial Reference Drugs.

The bioluminescent Leishmania infantum BALB/c mouse model was used to evaluate the parasiticidal drug action kinetics of the reference drugs miltefosine, paromomycin, sodium stibogluconate, and liposomal amphotericin B. Infected mice were treated for 5 days starting from 7 days post-infection, and parasite burdens were monitored over time via bioluminescence imaging (BLI). Using nonlinear regression analyses of the BLI signal, the parasite elimination half-life (t1/2) in the liver, bone marrow, and whole body was determined and compared for the different treatment regimens. Significant differences in parasiticidal kinetics were recorded. A single intravenous dose of 0.5 mg/kg liposomal amphotericin B was the fastest acting with a t1/2 of less than 1 day. Intraperitoneal injection of paromomycin at 320 mg/kg for 5 days proved to be the slowest with a t1/2 of about 5 days in the liver and 16 days in the bone marrow. To conclude, evaluation of the cidal kinetics of the different antileishmanial reference drugs revealed striking differences in their parasite elimination half-lives. This BLI approach also enables an in-depth pharmacodynamic comparison between novel drug leads and may constitute an essential tool for the design of potential drug combinations.

Analysis of intestinal epithelial cell responses to Cryptosporidium highlights the temporal effects of IFN-γ on parasite restriction.

The production of IFN-γ is crucial for control of multiple enteric infections, but its impact on intestinal epithelial cells (IEC) is not well understood. Cryptosporidium parasites exclusively infect epithelial cells and the ability of interferons to activate the transcription factor STAT1 in IEC is required for parasite clearance. Here, the use of single cell RNA sequencing to profile IEC during infection revealed an increased proportion of mid-villus enterocytes during infection and induction of IFN-γ-dependent gene signatures that was comparable between uninfected and infected cells. These analyses were complemented by in vivo studies, which demonstrated that IEC expression of the IFN-γ receptor was required for parasite control. Unexpectedly, treatment of Ifng-/- mice with IFN-γ showed the IEC response to this cytokine correlates with a delayed reduction in parasite burden but did not affect parasite development. These data sets provide insight into the impact of IFN-γ on IEC and suggest a model in which IFN-γ signalling to uninfected enterocytes is important for control of Cryptosporidium.

Parasitological examination results of zoo animals in Germany between 2012 and 2022

Parasitic infections in zoo animals are a critical concern for both animal health and management. The aim of this study was to assess the occurrence of endo- and ectoparasites among zoo animals in Germany. A retrospective analysis of the submitted samples of a diverse range of zoo animals (5768) from a ten-year period (2012–2022) was conducted. Overall, 31.1% of those samples tested positive for at least one parasite. In the examined samples, helminths (28.4%) were found more often than protozoans (10.3%) or ectoparasites (0.8%). Among the various animal groups the following parasites were found most commonly: Artiodactyla: Coccidia (34.6%), Strongylida (23.4%); Perissodactyla: Strongylida (19.3%), Ascaridida (12.0%); Carnivora: Ascaridida (16.6%), Coccidia (8.1%); Rodentia: Oxyurida (18.2%), Coccidia (10.5%); Marsupialia: Coccidia (9.4%), Oxyurida (5.9%); Primates: Trichuris spp. (9.7%), Oxyurida (2.2%); Aves: Capillaria (7.8%), Ascaridida (7.6%); Reptilia, Amphibia, Insecta: Oxyurida (18.7%); Pisces: Ciliates (6.2%). Furthermore, potentially zoonotic parasites were identified, including Toxoplasma gondii (0.1%), Cryptosporidium sp. (0.1%). By examining the occurrence of specific parasites, these findings demonstrate the importance of parasites in the context of zoo animal health. They also highlight the need for effective strategies to control parasite burden to improve the overall welfare of zoo animals.

Natural resistance to meglumine antimoniate is associated with treatment failure in cutaneous leishmaniasis caused by Leishmania (Viannia) panamensis.

The multifactorial basis of therapeutic response can obscure the relation between antimicrobial drug susceptibility and clinical outcome. To discern the relationship between parasite susceptibility to meglumine antimoniate (SbV) and therapeutic outcome of cutaneous leishmaniasis, risk factors for treatment failure were considered in evaluating this relationship in ninety-one cutaneous leishmaniasis patients and corresponding clinical strains of Leishmania (Viannia) panamensis. Parasite susceptibility to 32 μg SbV/mL (plasma Cmax) was evaluated in primary human macrophages, PBMCs, and U937 macrophages. Early parasitological response to treatment was determined in lesions of a subgroup of patients, and pathogenicity of Sb-resistant and sensitive clinical strains was compared in BALB/c mice. Parasite survival in cell models and patient lesions was determined by qRT-PCR of Leishmania 7SLRNA transcript. Parasite loads in BALB/c mice were quantified by limiting dilution analysis. The disparate Sb-susceptibility of parasite subpopulations distinguished by isoenzyme profiles (zymodemes) was manifest in all cell models. Notably, Sb-resistance defined by parasite survival, was most effectively discerned in U937 macrophages compared with primary human host cells, significantly higher among strains from patients who failed treatment than cured and, significantly associated with treatment failure. Each unit increase in transformed survival rate corresponded to a 10.6-fold rise in the odds of treatment failure. Furthermore, treatment failure was significantly associated with naturally Sb-resistant zymodeme 2.3 strains, which also produced larger lesions and parasite burdens in BALB/c mice than Sb-sensitive zymodeme 2.2 strains. The confounding effect of host risk factors for treatment failure in discerning this association was evidenced in comparing strains from patients with and without the defined risk factors for treatment failure. These results establish the association of natural resistance to meglumine antimoniate with treatment failure, the importance of host risk factors in evaluating drug susceptibility and treatment outcome, and the clinical and epidemiological relevance of natural Sb-resistance in L. (V.) panamensis subpopulations.

CCR5 promotes the migration of pathological CD8+ T cells to the leishmanial lesions.

Cytolytic CD8+ T cells mediate immunopathology in cutaneous leishmaniasis without controlling parasites. Here, we identify factors involved in CD8+ T cell migration to the lesion that could be targeted to ameliorate disease severity. CCR5 was the most highly expressed chemokine receptor in patient lesions, and the high expression of CCL3 and CCL4, CCR5 ligands, was associated with delayed healing of lesions. To test the requirement for CCR5, Leishmania-infected Rag1-/- mice were reconstituted with CCR5-/- CD8+ T cells. We found that these mice developed smaller lesions accompanied by a reduction in CD8+ T cell numbers compared to controls. We confirmed these findings by showing that the inhibition of CCR5 with maraviroc, a selective inhibitor of CCR5, reduced lesion development without affecting the parasite burden. Together, these results reveal that CD8+ T cells migrate to leishmanial lesions in a CCR5-dependent manner and that blocking CCR5 prevents CD8+ T cell-mediated pathology.

Characterization of intestinal mononuclear phagocyte subsets in young ruminants at homeostasis and during Cryptosporidium parvum infection.

Cryptosporidiosis is a poorly controlled zoonosis caused by an intestinal parasite, Cryptosporidium parvum, with a high prevalence in livestock (cattle, sheep, and goats). Young animals are particularly susceptible to this infection due to the immaturity of their intestinal immune system. In a neonatal mouse model, we previously demonstrated the importance of the innate immunity and particularly of type 1 conventional dendritic cells (cDC1) among mononuclear phagocytes (MPs) in controlling the acute phase of C. parvum infection. These immune populations are well described in mice and humans, but their fine characterization in the intestine of young ruminants remained to be further explored. Immune cells of the small intestinal Peyer's patches and of the distal jejunum were isolated from naive lambs and calves at different ages. This was followed by their fine characterization by flow cytometry and transcriptomic analyses (q-RT-PCR and single cell RNAseq (lamb cells)). Newborn animals were infected with C. parvum, clinical signs and parasite burden were quantified, and isolated MP cells were characterized by flow cytometry in comparison with age matched control animals. Here, we identified one population of macrophages and three subsets of cDC (cDC1, cDC2, and a minor cDC subset with migratory properties) in the intestine of lamb and calf by phenotypic and targeted gene expression analyses. Unsupervised single-cell transcriptomic analysis confirmed the identification of these four intestinal MP subpopulations in lamb, while highlighting a deeper diversity of cell subsets among monocytic and dendritic cells. We demonstrated a weak proportion of cDC1 in the intestine of highly susceptible newborn lambs together with an increase of these cells within the first days of life and in response to the infection. Considering cDC1 importance for efficient parasite control in the mouse model, one may speculate that the cDC1/cDC2 ratio plays also a key role for the efficient control of C. parvum in young ruminants. In this study, we established the first fine characterization of intestinal MP subsets in young lambs and calves providing new insights for comparative immunology of the intestinal MP system across species and for future investigations on host-Cryptosporidium interactions in target species.