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Parasites & Vectors Aug 2014Tryparedoxin peroxidase (TXNPx) participates in defence against oxidative stress as an antioxidant by metabolizing hydrogen peroxide into water molecules. Reports...
BACKGROUND
Tryparedoxin peroxidase (TXNPx) participates in defence against oxidative stress as an antioxidant by metabolizing hydrogen peroxide into water molecules. Reports suggest that drug-resistant parasites may increase the levels of TXNPx and other enzymes, thereby protecting them against oxidative stress.
METHODS
In this study, the gene encoding cytosolic TXNPx (cTXNPx) was characterized in lines of Leishmania (Viannia) braziliensis and Leishmania (Leishmania) infantum that are susceptible and resistant to potassium antimony tartrate (Sb(III)). We investigated the levels of mRNA and genomic organization of the cTXNPx gene. In addition, we transfected the Leishmania lines with the cTXNPx gene and analysed the susceptibility of transfected parasites to Sb(III) and to hydrogen peroxide (H2O2).
RESULTS
Northern blot and real-time reverse transcriptase polymerase chain reaction analyses revealed that the level of TXNPx mRNA was approximately 2.5-fold higher in the Sb(III)-resistant L. braziliensis line than in the parental line. In contrast, no significant difference in cTXNPx mRNA levels between the L. infantum lines was observed. Southern blot analyses revealed that the cTXNPx gene is not amplified in the genome of the Sb(III)-resistant Leishmania lines analysed. Functional analysis of cTXNPx was performed to determine whether overexpression of the enzyme in L. braziliensis and L. infantum lines would change their susceptibility to Sb(III). Western blotting analysis showed that the level of cTXNPx was 2 to 4-fold higher in transfected clones compared to non-transfected cells. Antimony susceptibility test (EC50 assay) revealed that L. braziliensis lines overexpressing cTXNPx had a 2-fold increase in resistance to Sb(III) when compared to the untransfected parental line. In addition, these clones are more tolerant to exogenous H2O2 than the untransfected parental line. In contrast, no difference in Sb(III) susceptibility and a moderate index of resistance to H2O2 was observed in L. infantum clones overexpressing cTXNPx.
CONCLUSION
Our functional analysis revealed that cTXNPx is involved in the antimony-resistance phenotype in L. braziliensis.
Topics: Antimony; Antiprotozoal Agents; Drug Resistance; Gene Expression Regulation, Enzymologic; Genomics; Hydrogen Peroxide; Leishmania braziliensis; Leishmania infantum; Oxidative Stress; Peroxidases; Protozoan Proteins; RNA, Messenger
PubMed: 25174795
DOI: 10.1186/1756-3305-7-406 -
Revista Brasileira de Parasitologia... 2015Canine visceral leishmaniasis (CVL) is difficult to diagnosis, mainly due to the presence of asymptomatic animals, the diversity of clinical symptoms and the difficulty...
Canine visceral leishmaniasis (CVL) is difficult to diagnosis, mainly due to the presence of asymptomatic animals, the diversity of clinical symptoms and the difficulty in obtaining diagnostic evidence of high sensitivity and specificity. The purpose of this study was to diagnose CVL in urinary sediment of 70 dogs of different breeds, sexes and ages from the veterinary hospital of the Federal University of Piauí and Zoonosis Control Center of Teresina, Brazil. The serological tests were TR DPP® for CVL and enzyme-linked immunosorbent assay (ELISA) for CVL, parasitological exams of bone marrow and lymph nodes and urine sediment cultures. Leishmania was detected in the bone marrow and/or lymph node of 61.0% of the animals (43/70), and urine sediment culture was positive in 9.30% (4/43) of these animals. In the serological exams, 70.0% (49/70) were reactive using the DPP and 78.2% (55/70) were reactive using ELISA. The goal of this study was to diagnose the presence of L. (infantum) chagasi in a culture of urinary sediment.
Topics: Animals; Dog Diseases; Dogs; Female; Leishmania infantum; Leishmaniasis, Visceral; Male; Urine
PubMed: 25909260
DOI: 10.1590/S1984-29612014086 -
Parasites & Vectors Sep 2021Leishmania tarentolae is a protozoan isolated from geckoes (Tarentola annularis, Tarentola mauritanica), which is considered non-pathogenic and is transmitted by...
BACKGROUND
Leishmania tarentolae is a protozoan isolated from geckoes (Tarentola annularis, Tarentola mauritanica), which is considered non-pathogenic and is transmitted by herpetophilic Sergentomyia spp. sand flies. This species occurs in sympatry with Leishmania infantum in areas where canine leishmaniasis is endemic. In the present study, we investigated the circulation of L. tarentolae and L. infantum in sand flies, dogs and lizards in a dog shelter in southern Italy, where canine leishmaniasis by L. infantum is endemic.
METHODS
Sheltered dogs (n = 100) negative for Leishmania spp. (March 2020) were screened by immunofluorescence antibody test (IFAT) using promastigotes of both species at two time points (June 2020 and March 2021). Whole blood from dogs, tissues of Podarcis siculus lizards (n = 28) and sand flies (n = 2306) were also sampled and tested by a duplex real-time PCR (dqPCR). Host blood meal was assessed in sand flies by PCR.
RESULTS
Overall, 16 dogs became positive for L. infantum and/or L. tarentolae by IFAT at one or both sampling periods. One canine blood sample was positive for L. infantum, whilst two for L. tarentolae by dqPCR. At the cytology of lizard blood, Leishmania spp. amastigote-like forms were detected in erythrocytes. Twenty-two tissue samples, mostly lung (21.4%), scored molecularly positive for L. tarentolae, corresponding to 10 lizards (i.e., 35.7%). Of the female Sergentomyia minuta sampled (n = 1252), 158 scored positive for L. tarentolae, four for L. infantum, and one co-infected. Two Phlebotomus perniciosus (out of 29 females) were positive for L. tarentolae. Engorged S. minuta (n = 10) fed on humans, and one P. perniciosus, positive for L. tarentolae, on lagomorphs.
CONCLUSIONS
Dogs and lacertid lizards (Podarcis siculus) were herein found for the first time infected by L. tarentolae. The detection of both L. tarentolae and L. infantum in S. minuta and P. perniciosus suggests their sympatric circulation, with a potential overlap in vertebrate hosts. The interactions between L. tarentolae and L. infantum should be further investigated in both vectors and vertebrate hosts to understand the potential implications for the diagnosis and control of canine leishmaniasis in endemic areas.
Topics: Animals; Dogs; Endemic Diseases; Female; Leishmania; Leishmania infantum; Leishmaniasis; Lizards; Male; Psychodidae; Zoonoses
PubMed: 34493323
DOI: 10.1186/s13071-021-04973-2 -
Brazilian Journal of Medical and... Mar 2010Fifteen symptomatic and seven asymptomatic dogs infected naturally with Leishmania chagasi were examined in order to identify the presence of parasites and changes in...
Fifteen symptomatic and seven asymptomatic dogs infected naturally with Leishmania chagasi were examined in order to identify the presence of parasites and changes in heart and lung. Histopathological, cytological, and immunohistochemical analyses were performed on samples of heart and lung tissues. An inflammatory reaction characterized by inflammatory mononuclear, perivascular and intermuscular infiltrates was observed in both symptomatic and asymptomatic animals on histopathological analysis of the heart. In the lung, there was thickening of the alveolar septa due to congestion, edema, inflammatory infiltrate, and fibroblast proliferation. A focal reaction was observed although a diffuse reaction was present in both groups. On cytological examination, heart and lung imprints revealed amastigotes in two symptomatic animals and heart imprints were found in 1 asymptomatic dog. Immunoperoxidase staining showed amastigotes in the lung and heart of only 1 of 6 symptomatic animals examined. Within the ethical principles and limits of this research, it can be inferred that the study of heart and lung alterations in canine visceral leishmaniasis is increasingly important for understanding the problem related to humans. Dogs with visceral leishmaniasis were a good experimental model, since infection was caused by the same agent and the animals developed clinical, pathological and immunological alterations similar to those observed in humans.
Topics: Animals; Antibodies, Protozoan; Dog Diseases; Dogs; Enzyme-Linked Immunosorbent Assay; Female; Fluorescent Antibody Technique, Indirect; Immunohistochemistry; Leishmania infantum; Leishmaniasis, Visceral; Lung; Male; Myocardium
PubMed: 20401439
DOI: 10.1590/s0100-879x2009007500037 -
Veterinary Parasitology Nov 2009Visceral leishmaniasis (VL) is primarily transmitted by an invertebrate vector, but transmission in the absence of the vector has been reported. Vertical transmission of...
Visceral leishmaniasis (VL) is primarily transmitted by an invertebrate vector, but transmission in the absence of the vector has been reported. Vertical transmission of VL has been described in man and dogs. The aim of this study was to evaluate the distribution of Leishmania amastigotes in fetal organs and histopathologic changes associated with parasitism and to determinate the frequency of transplacental transmission and potential of vertical transmission by symptomatic and asymptomatic pregnant bitches. Symptomatic (n=4) and asymptomatic (n=4) pregnant bitches, serologically and parasitologically positive for Leishmania sp., carrying a total of 53 fetuses (26 from symptomatic and 27 from asymptomatic bitches) were selected at the Veterinary Hospital of the National University of Asuncion, Paraguay. Samples of placenta and fetal organs such as liver, spleen, lymph nodes, bone marrow, kidney and heart were histologically evaluated and processed for immunodetection of amastigotes and PCR. There were no lesions compatible with VL in fetal tissues in spite of the presence of amastigotes, particularly in lymphoreticular tissues. However, fetal hepatocytes had marked degenerative changes that were independent of the presence of amastigotes in liver. Twenty-six out of 53 placentas (13 symptomatic and 13 asymptomatic) and a total of 17 fetuses out of 53 (nine symptomatic and eight asymptomatic) were PCR positive. Together these findings indicate a high frequency of transplacental transmission and no differences in the potential of transmission when symptomatic were compared to asymptomatic pregnant bitches.
Topics: Animals; Dog Diseases; Dogs; Female; Fetus; Infectious Disease Transmission, Vertical; Leishmania infantum; Leishmaniasis, Visceral; Placenta; Pregnancy
PubMed: 19647368
DOI: 10.1016/j.vetpar.2009.07.013 -
Frontiers in Cellular and Infection... 2019Despite the increasing number of studies concerning insect immunity, immune responses in the presence of infection has not been widely investigated. The few available...
Despite the increasing number of studies concerning insect immunity, immune responses in the presence of infection has not been widely investigated. The few available studies analyzed the role of the Toll and IMD pathways involved in response against and microbial infections. Nevertheless, effector molecules responsible for controlling sand fly infections have not been identified. In the present study we investigated the role a signal transduction pathway, the Transforming Growth Factor-beta (TGF-β) pathway, on the interrelation between and . We identified an homolog belonging to the multifunctional cytokine TGF-β gene family (-β), which is closely related to the activin/inhibin subfamily and potentially involved in responses to infections. We investigated this gene expression through the insect development and in adult flies infected with . Our results showed that -β was expressed in all developmental stages and was upregulated at the third day post infection, when protein levels were also higher as compared to uninfected insects. At this point blood digestion is finished and parasites are in close contact with the insect gut. In addition, we investigated the role of LlTGF-β on infection by using either gene silencing by RNAi or pathway inactivation by addition of the TGF-β receptor inhibitor SB431542. The blockage of the LlTGF-β pathway increased significantly antimicrobial peptides expression and nitric oxide levels in the insect gut, as expected. Both methods led to a decreased infection. Our results show that inactivation of the TGF-β signal transduction pathway reduce survival, therefore suggesting that under natural conditions the parasite benefits from the insect LlTGF-β pathway, as already seen in infection of mosquitoes.
Topics: Animals; Gene Expression Profiling; Host-Pathogen Interactions; Immunity, Innate; Insect Vectors; Leishmania infantum; Psychodidae; Signal Transduction; Survival Analysis; Transforming Growth Factor beta
PubMed: 30972305
DOI: 10.3389/fcimb.2019.00071 -
TheScientificWorldJournal 2015Leishmaniasis is considered by the World Health Organization as one of the infectious parasitic diseases endemic of great relevance and a global public health problem....
Leishmaniasis is considered by the World Health Organization as one of the infectious parasitic diseases endemic of great relevance and a global public health problem. Pentavalent antimonials used for treatment of this disease are limited and new phytochemicals emerge as an alternative to existing treatments, due to the low toxicity and cost reduction. Usnic acid is uniquely found in lichens and is especially abundant in genera such as Alectoria, Cladonia, Evernia, Lecanora, Ramalina, and Usnea. Usnic acid has been shown to exhibit antiviral, antiprotozoal, antiproliferative, anti-inflammatory, and analgesic activity. The aim of this study was to evaluate the antileishmanial activity of usnic acid on Leishmania infantum chagasi promastigotes and the occurrence of drug-induced ultrastructural damage in the parasite. Usnic acid was effective against the promastigote forms (IC50=18.30±2.00 µg/mL). Structural and ultrastructural aspects of parasite were analyzed. Morphological alterations were observed as blebs in cell membrane and shapes given off, increasing the number of cytoplasmic vacuoles, and cellular and mitochondrial swelling, with loss of cell polarity. We concluded that the usnic acid presented antileishmanial activity against promastigote forms of Leishmania infantum chagasi and structural and ultrastructural analysis reinforces its cytotoxicity. Further, in vitro studies are warranted to further evaluate this potential.
Topics: Animals; Benzofurans; In Vitro Techniques; Leishmania infantum
PubMed: 25767824
DOI: 10.1155/2015/617401 -
Genes Dec 2019Pathogen fitness landscapes change when transmission cycles establish in non-native environments or spill over into new vectors and hosts. The introduction of in the... (Review)
Review
Pathogen fitness landscapes change when transmission cycles establish in non-native environments or spill over into new vectors and hosts. The introduction of in the Americas into the Neotropics during European colonization represents a unique case study to investigate the mechanisms of ecological adaptation of this important parasite. Defining the evolutionary trajectories that drive fitness in this new environment are of great public health importance as they will allow unique insight into pathways of host/pathogen co-evolution and their consequences for region-specific changes in disease manifestation. This review summarizes current knowledge on genetic and phenotypic diversity in the Americas and its possible role in the unique epidemiology of visceral leishmaniasis (VL) in the New World. We highlight the importance of appreciating adaptive molecular mechanisms in to understand the parasites' successful establishment on the continent.
Topics: Atlantic Ocean; Evolution, Molecular; Genetic Fitness; Humans; Leishmania infantum; Leishmaniasis, Visceral; Phenotype
PubMed: 31861501
DOI: 10.3390/genes11010004 -
PloS One Apr 2011The life stages of Leishmania spp. include the infectious promastigote and the replicative intracellular amastigote. Each stage is phagocytosed by macrophages during the...
The life stages of Leishmania spp. include the infectious promastigote and the replicative intracellular amastigote. Each stage is phagocytosed by macrophages during the parasite life cycle. We previously showed that caveolae, a subset of cholesterol-rich membrane lipid rafts, facilitate uptake and intracellular survival of virulent promastigotes by macrophages, at least in part, by delaying parasitophorous vacuole (PV)-lysosome fusion. We hypothesized that amastigotes and promastigotes would differ in their route of macrophage entry and mechanism of PV maturation. Indeed, transient disruption of macrophage lipid rafts decreased the entry of promastigotes, but not amastigotes, into macrophages (P<0.001). Promastigote-containing PVs were positive for caveolin-1, and co-localized transiently with EEA-1 and Rab5 at 5 minutes. Amastigote-generated PVs lacked caveolin-1 but retained Rab5 and EEA-1 for at least 30 minutes or 2 hours, respectively. Coinciding with their conversion into amastigotes, the number of promastigote PVs positive for LAMP-1 increased from 20% at 1 hour, to 46% by 24 hours, (P<0.001, Chi square). In contrast, more than 80% of amastigote-initiated PVs were LAMP-1+ at both 1 and 24 hours. Furthermore, lipid raft disruption increased LAMP-1 recruitment to promastigote, but not to amastigote-containing compartments. Overall, our data showed that promastigotes enter macrophages through cholesterol-rich domains like caveolae to delay fusion with lysosomes. In contrast, amastigotes enter through a non-caveolae pathway, and their PVs rapidly fuse with late endosomes but prolong their association with early endosome markers. These results suggest a model in which promastigotes and amastigotes use different mechanisms to enter macrophages, modulate the kinetics of phagosome maturation, and facilitate their intracellular survival.
Topics: Animals; Caveolin 1; Cholesterol; Cricetinae; Leishmania infantum; Life Cycle Stages; Macrophages; Male; Membrane Microdomains; Mice; Mice, Inbred BALB C; Phagocytosis; Phagosomes
PubMed: 21552562
DOI: 10.1371/journal.pone.0019000 -
Infection and Immunity Jan 2011The vector-borne protozoan Leishmania infantum chagasi causes minimal inflammation after inoculation into skin but disseminates to cause fatal visceral leishmaniasis. To...
The vector-borne protozoan Leishmania infantum chagasi causes minimal inflammation after inoculation into skin but disseminates to cause fatal visceral leishmaniasis. To define the inflammatory response at the parasite inoculation site, we introduced metacyclic L. infantum chagasi promastigotes intradermally into BALB/c mouse ears and studied inflammatory cells over 7 days. Ly6G(+) neutrophils rapidly infiltrated the dermis, peaking after 6 to 24 h. Macrophages and NK cells next infiltrated the dermis, and NK followed by B cells expanded in draining lymph nodes. Parasite-containing phagocytes were tracked with fluorescent mCherry-labeled L. infantum chagasi. Ly6G(+) neutrophils contained the greatest proportion of intracellular parasites 6 to 24 h after inoculation, whereas dermal macrophages harbored the majority of intracellular parasites after 2 to 7 days. These observations were validated microscopically. Low doses of antibody transiently depleted mice of neutrophils, leaving other cells intact. Combined results of in vivo imaging, flow cytometry, and quantitative PCR showed that neutrophil depletion slowed the clearance of extracellular (luciferase-positive) promastigotes during the first 24 h after inoculation yet decreased the numbers of leukocytes containing intracellular (mCherry-positive) parasites. From 3 days onward, total L. infantum chagasi-containing dermal leukocytes and total L. infantum chagasi parasites in draining lymph nodes were similar in both groups. Nonetheless, a second wave of L. infantum chagasi-containing neutrophils occurred 7 days after parasite inoculation into neutrophil-depleted mice, corresponding to the time of neutrophil recovery. Thus, neutrophils were recruited to the dermis even late after inoculation, and L. infantum chagasi trafficked through neutrophils in both neutrophil-depleted and control mice, albeit with different kinetics. Recruitment of neutrophils and transient parasite residence in neutrophils may play a role in nonulcerative forms of leishmaniasis.
Topics: Animals; Female; Leishmania infantum; Leishmaniasis, Visceral; Leukocytes; Lymph Nodes; Mice; Mice, Inbred BALB C; Neutrophils; Skin
PubMed: 20937764
DOI: 10.1128/IAI.00338-10