scholarly journals High-resolution melting analysis identifies reservoir hosts of zoonotic Leishmania parasites in Tunisia

2022 ◽  
Vol 15 (1) ◽  
Author(s):  
Moufida Derghal ◽  
Abir Tebai ◽  
Ghofrane Balti ◽  
Hajer Souguir-Omrani ◽  
Jomaa Chemkhi ◽  
...  
Keyword(s):  
High Resolution ◽  
Leishmania Major ◽  
High Resolution Melting ◽  
Reservoir Host ◽  
Sand Fly ◽  
Canine Leishmaniasis ◽  
Multiple Infections ◽  
Epidemiological Surveys ◽  
Reservoir Hosts

Abstract Background Leishmaniasis is endemic in Tunisia and presents with different clinical forms, caused by the species Leishmania infantum, Leishmania major, and Leishmania tropica. The life cycle of Leishmania is complex and involves several phlebotomine sand fly vectors and mammalian reservoir hosts. The aim of this work is the development and evaluation of a high-resolution melting PCR (PCR-HRM) tool to detect and identify Leishmania parasites in wild and domestic hosts, constituting confirmed (dogs and Meriones rodents) or potential (hedgehogs) reservoirs in Tunisia. Methods Using in vitro-cultured Leishmania isolates, PCR-HRM reactions were developed targeting the 7SL RNA and HSP70 genes. Animals were captured or sampled in El Kef Governorate, North West Tunisia. DNA was extracted from the liver, spleen, kidney, and heart from hedgehogs (Atelerix algirus) (n = 3) and rodents (Meriones shawi) (n = 7) and from whole blood of dogs (n = 12) that did not present any symptoms of canine leishmaniasis. In total, 52 DNA samples were processed by PCR-HRM using both pairs of primers. Results The results showed melting curves enabling discrimination of the three Leishmania species present in Tunisia, and were further confirmed by Sanger sequencing. Application of PCR-HRM assays on reservoir host samples showed that overall among the examined samples, 45 were positive, while seven were negative, with no Leishmania infection. Meriones shawi were found infected with L. major, while dogs were infected with L. infantum. However, co-infections with L. major/L. infantum species were detected in four Meriones specimens and in all tested hedgehogs. In addition, multiple infections with the three Leishmania species were found in one hedgehog specimen. Sequence analyses of PCR-HRM products corroborated the Leishmania species found in analyzed samples. Conclusions The results of PCR-HRM assays applied to field specimens further support the possibility of hedgehogs as reservoir hosts of Leishmania. In addition, we showed their usefulness in the diagnosis of canine leishmaniasis, specifically in asymptomatic dogs, which will ensure a better evaluation of infection extent, thus improving elaboration of control programs. This PCR-HRM method is a robust and reliable tool for molecular detection and identification of Leishmania and can be easily implemented in epidemiological surveys in endemic regions. Graphical Abstract

scholarly journals Polymorphisms in Plasmodium falciparum dihydropteroate synthetase and dihydrofolate reductase genes in Nigerian children with uncomplicated malaria using high-resolution melting technique

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Adeyemi T. Kayode ◽  
Fehintola V. Ajogbasile ◽  
Kazeem Akano ◽  
Jessica N. Uwanibe ◽  
Paul E. Oluniyi ◽  
...  
Keyword(s):  
High Resolution ◽  
Dihydrofolate Reductase ◽  
Uncomplicated Malaria ◽  
High Resolution Melting ◽  
Intermittent Preventive Treatment ◽  
Preventive Treatment ◽  
Triple Mutant ◽  
Seasonal Malaria Chemoprevention

AbstractIn 2005, the Nigerian Federal Ministry of Health revised the treatment policy for uncomplicated malaria with the introduction of artemisinin-based combination therapies (ACTs). This policy change discouraged the use of Sulphadoxine-pyrimethamine (SP) as the second-line treatment of uncomplicated falciparum malaria. However, SP is used as an intermittent preventive treatment of malaria in pregnancy (IPTp) and seasonal malaria chemoprevention (SMC) in children aged 3–59 months. There have been increasing reports of SP resistance especially in the non-pregnant population in Nigeria, thus, the need to continually monitor the efficacy of SP as IPTp and SMC by estimating polymorphisms in dihydropteroate synthetase (dhps) and dihydrofolate reductase (dhfr) genes associated with SP resistance. The high resolution-melting (HRM) assay was used to investigate polymorphisms in codons 51, 59, 108 and 164 of the dhfr gene and codons 437, 540, 581 and 613 of the dhps gene. DNA was extracted from 271 dried bloodspot filter paper samples obtained from children (< 5 years old) with uncomplicated malaria. The dhfr triple mutant I51R59N108, dhps double mutant G437G581 and quadruple dhfr I51R59N108 + dhps G437 mutant haplotypes were observed in 80.8%, 13.7% and 52.8% parasites, respectively. Although the quintuple dhfr I51R59N108 + dhps G437E540 and sextuple dhfr I51R59N108 + dhps G437E540G581 mutant haplotypes linked with in-vivo and in-vitro SP resistance were not detected, constant surveillance of these haplotypes should be done in the country to detect any change in prevalence.

scholarly journals Some Epidemiological Aspects of Cutaneous Leishmaniasis in a New Focus, Central Iran

2015 ◽  
Vol 2015 ◽  
pp. 1-5 ◽  
Author(s):  
M. R. Yaghoobi-Ershadi ◽  
N. Marvi-Moghadam ◽  
R. Jafari ◽  
A. A. Akhavan ◽  
H. Solimani ◽  
...  
Keyword(s):  
Cutaneous Leishmaniasis ◽  
Primary Schools ◽  
Leishmania Major ◽  
Reservoir Host ◽  
Human Infection ◽  
Native People ◽  
Sand Fly ◽  
Animal Reservoir ◽  
Rural Districts ◽  
Tatera Indica

Following the epidemic of cutaneous leishmaniasis in Khatam County, Yazd Province, this study was carried out to determine vector, and animal reservoir host(s) and investigate the human infection during 2005-2006. Four rural districts where the disease had higher prevalence were selected. Sticky paper traps were used to collect sand flies during April to November, biweekly. Meanwhile rodents were captured using Sherman traps from August to November. Households and primary schools were visited and examined for human infection in February 2006. The parasite was detected by RAPD-PCR method. The rate of ulcers and scars among the inhabitants was 4.8% and 9.8%, respectively. Three rodent species were captured during the study:Meriones libycus, Rhombomys opimus, andTatera indica. Six sand fly species were also collected and identified; among themPhlebotomus papatasihad the highest frequency.Leishmania majorwas detected as the agent of the disease in the area. It was detected fromR. opimusand native people.

scholarly journals Transcriptional and genomic parallels between the monoxenous parasiteHerpetomonas muscarumandLeishmania

2019 ◽  
Author(s):  
Megan A. Sloan ◽  
Karen Brooks ◽  
Thomas D. Otto ◽  
Mandy J. Sanders ◽  
James A. Cotton ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Leishmania Major ◽  
Neglected Tropical Diseases ◽  
Fruit Fly ◽  
Sand Fly ◽  
Suitable Model ◽  
Insect Host ◽  
Causative Agents ◽  
Trypanosomatid Parasites

AbstractTrypanosomatid parasites are causative agents of important human and animal diseases such as sleeping sickness and leishmaniasis. Most trypanosomatids are transmitted to their mammalian hosts by insects, often belonging to Diptera (or true flies). These are called dixenous trypanosomatids since they infect two different hosts, in contrast to those that infect just insects (monoxenous). However, it is still unclear whether dixenous and monoxenous trypanosomatids interact similarly with their insect host, as fly-monoxenous trypanosomatid interaction systems are rarely reported and under-studied – despite being common in nature. Here we present the genome of monoxenous trypanosomatidHerpetomonas muscarumand discuss its transcriptome duringin vitroculture and during infection of its natural insect hostDrosophila melanogaster. TheH. muscarumgenome is broadly syntenic with that of human parasiteLeishmania major. We also found strong similarities between theH. muscarumtranscriptome during fruit fly infection, and those ofLeishmaniaduring sand fly infections. Overall this suggestsDrosophila-Herpetomonasis a suitable model for less accessible insect-trypanosomatid host-parasite systems such as sandfly-Leishmania.Author SummaryTrypanosomes andLeishmaniaare parasites that cause serious Neglected Tropical Diseases (NTDs) in the world’s poorest people. Both of these are dixenous trypanosomatids, transmitted to humans and other mammals by biting flies. They are called dixenous as they can establish infections in two different types of hosts – insect vectors and mammals. In contrast, monoxenous trypanosomatids usually only infect insects. Despite establishment in the insect’s midgut being key to transmission of NTDs, events during early establishment inside the insect are still unclear in both dixenous and monoxenous parasites. Here, we study the interaction between a model insect – the fruit flyDrosophila melanogaster– and its natural monoxenous trypanosomatid parasiteHerpetomonas muscarum. We show that both the genome of this parasite, and gene regulation at early stages of infection have strong parallels withLeishmania. This work has begun to identify evolutionarily conserved aspects of the process by which trypanosomatids establish in insects, thus potentially highlighting key checkpoints necessary for transmission of dixenous parasites. In turn, this might inform new strategies to control trypanosomatid NTDs.

scholarly journals Evaluation of a New Topical Treatment for the Control of Cutaneous Leishmaniasis

2020 ◽  
Vol 8 (11) ◽  
pp. 1803
Author(s):  
Berenice Martínez-Salazar ◽  
Vanessa Carregaro Pereira ◽  
Yazmin Hauyon-La-Torre ◽  
Ali Khamesipour ◽  
Fabienne Tacchini-Cottier
Keyword(s):  
Cutaneous Leishmaniasis ◽  
Topical Application ◽  
Topical Treatment ◽  
Leishmania Major ◽  
Sand Fly ◽  
Control Group ◽  
Type 2 Immune Response ◽  
Metacyclic Promastigotes ◽  
Control Lesion

Leishmania major (L. major) causes cutaneous leishmaniasis in the Old World. The infection mostly induces a localized lesion restricted to the sand fly bite. The costs and the side effects of current treatments render imperative the development of new therapies that are affordable and easy to administrate. Topical treatment would be the ideal option for the treatment of cutaneous leishmaniasis. MF29 is a 3-haloacetamidobenzoate that was shown in vitro to inhibit tubulin assembly in Leishmania. Here, we tested a topical cream formulated with MF29. BALB/c mice were infected in the ear dermis with L. major metacyclic promastigotes and once the lesion appeared, mice were treated with different concentrations of MF29 and compared to the control group treated with the cream used as the vehicle. We observed that topical application of MF29 reduced the progression of the infection while control groups developed an unhealing lesion that became necrotic. The treatment decreased the type 2 immune response. Comparison with SinaAmphoLeish, another topical treatment, revealed that MF29 treatment once a day was sufficient to control lesion development, while application SinaAmphoLeish needed applications twice daily. Collectively, our data suggest that MF-29 topical application could be a promising topical treatment for cutaneous leishmaniasis.

scholarly journals Molecular detection of Leishmania donovani, Leishmania major, and Trypanosoma species in Sergentomyia squamipleuris sand flies from a visceral leishmaniasis focus in Merti sub-County, eastern Kenya

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Barrack O. Owino ◽  
Jackline Milkah Mwangi ◽  
Steve Kiplagat ◽  
Hannah Njiriku Mwangi ◽  
Johnstone M. Ingonga ◽  
...  
Keyword(s):  
Visceral Leishmaniasis ◽  
Mus Musculus ◽  
Blood Meal ◽  
Leishmania Donovani ◽  
Leishmania Major ◽  
Sand Fly ◽  
Health Concern ◽  
Sand Flies ◽  
Reservoir Hosts ◽  
Blood Meals

Abstract Background Visceral leishmaniasis (VL) and zoonotic cutaneous leishmaniasis (ZCL) are of public health concern in Merti sub-County, Kenya, but epidemiological data on transmission, vector abundance, distribution, and reservoir hosts remain limited. To better understand the disease and inform control measures to reduce transmission, we investigated the abundance and distribution of sand fly species responsible for Leishmania transmission in the sub-County and their blood-meal hosts. Methods We conducted an entomological survey in five villages with reported cases of VL in Merti sub-County, Kenya, using CDC miniature light traps and castor oil sticky papers. Sand flies were dissected and identified to the species level using standard taxonomic keys and PCR analysis of the cytochrome c oxidase subunit 1 (cox1) gene. Leishmania parasites were detected and identified by PCR and sequencing of internal transcribed spacer 1 (ITS1) genes. Blood-meal sources of engorged females were identified by high-resolution melting analysis of vertebrate cytochrome b (cyt-b) gene PCR products. Results We sampled 526 sand flies consisting of 8 species, Phlebotomus orientalis (1.52%; n = 8), and 7 Sergentomyia spp. Sergentomyia squamipleuris was the most abundant sand fly species (78.71%; n = 414) followed by Sergentomyia clydei (10.46%; n = 55). Leishmania major, Leishmania donovani, and Trypanosoma DNA were detected in S. squamipleuris specimens. Humans were the main sources of sand fly blood meals. However, we also detected mixed blood meals; one S. squamipleuris specimen had fed on both human and mouse (Mus musculus) blood, while two Ph. orientalis specimens fed on human, hyrax (Procavia capensis), and mouse (Mus musculus) blood. Conclusions Our findings implicate the potential involvement of S. squamipleuris in the transmission of Leishmania and question the dogma that human leishmaniases in the Old World are exclusively transmitted by sand flies of the Phlebotomus genus. The presence of Trypanosoma spp. may indicate mechanical transmission, whose efficiency should be investigated. Host preference analysis revealed the possibility of zoonotic transmission of leishmaniasis and other pathogens in the sub-County. Leishmania major and L. donovani are known to cause ZCL and VL, respectively. However, the reservoir status of the parasites is not uniform. Further studies are needed to determine the reservoir hosts of Leishmania spp. in the area.

scholarly journals A novel experimental system reveals immunoregulatory responses as mediators of persistent orthohantavirus infections in a rodent reservoir host

2019 ◽  
Author(s):  
Tomas Strandin ◽  
Teemu Smura ◽  
Paula Ahola ◽  
Kirsi Aaltonen ◽  
Tarja Sironen ◽  
...  
Keyword(s):  
Immune Responses ◽  
Virus Replication ◽  
Bank Vole ◽  
Experimental System ◽  
Reservoir Host ◽  
Myodes Glareolus ◽  
Bank Voles ◽  
Human Infections ◽  
Reservoir Hosts

AbstractOrthohantaviruses are globally emerging zoonotic pathogens. Human infections are characterized by an overt immune response that is efficient at counteracting virus replication but can also cause severe tissue damage. In contrast, orthohantavirus infections in rodent reservoir hosts are persistent and asymptomatic. The mechanisms facilitating asymptomatic virus persistence in reservoir hosts are not well understood but could help to guide therapeutic strategies for human infections. Here we report on a study using in vivo and in vitro experiments to investigate immune responses associated with persistent Puumala orthohantavirus (PUUV) infections in the bank vole (Myodes glareolus), its reservoir host. We examined adaptive cellular and humoral responses by quantifying changes in T-cell related gene expression in the spleen and immunoglobulin (Ig) responses in blood, respectively. Since existing Vero E6-cell adapted hantavirus isolates have been demonstrated to have lost their wild-type infection characteristics, infections were conducted with a novel PUUV strain isolated on a bank vole cell line. Whole virus genome sequencing revealed that only minor sequence changes occurred during the isolation process, and critically, experimental infections of bank voles with the new isolate resembled natural infections. In vitro infection of bank vole splenocytes with the novel isolate demonstrated that PUUV promotes immunoregulatory responses by inducing interleukin-10, a cytokine strongly associated with chronic viral infections. A delayed virus-specific humoral response occurred in experimentally infected bank voles, which is likely to allow for initial virus replication and the establishment of persistent infections. These results suggest that host immunoregulation facilitates persistent orthohantavirus infections in reservoir hosts.ImportanceOrthohantaviruses are a group of global pathogens that regularly spillover from rodent reservoirs into humans and can cause severe disease. Conversely, infections in reservoir hosts do not cause obvious adverse effects. The mechanisms responsible for persistent asymptomatic reservoir infections are unknown, and progress has been hindered by the absence of an adequate experimental system. Knowledge on these mechanisms could help provide strategies to treat human infections. We developed and validated an experimental system based on an orthohantavirus isolated in cells of its vole reservoir host. Using animal and cell culture experiments in the reservoir host system, we demonstrated that infection suppresses immunity in the vole reservoir via specific mechanisms, likely allowing the virus to take hold and preventing immune responses that can cause self-damage.

scholarly journals Zoonotic cutaneous leishmaniasis and Leishmania infection among Meriones shawi population in Setif Province, Algeria

2021 ◽  
Vol 22 (7) ◽  
Author(s):  
Amina Fellahi ◽  
NACER DJIRAR ◽  
ABDELKADER CHERIEF ◽  
ABDELKRIM BOUDRISSA ◽  
NAOUEL EDDAIKRA
Keyword(s):  
Cutaneous Leishmaniasis ◽  
Molecular Detection ◽  
Leishmania Major ◽  
Positive Association ◽  
Reservoir Host ◽  
Negative Association ◽  
Leishmania Infection ◽  
Zoonotic Cutaneous Leishmaniasis ◽  
Almost All ◽  
Reservoir Hosts

Abstract. Fellahi A, Djirar N, Cherief A, Boudrissa A, Eddaikra N. 2021. Zoonotic cutaneous leishmaniasis and Leishmania infection among Meriones shawi population in Setif Province, Algeria. Biodiversitas 22: 2547-2554. Zoonotic cutaneous leishmaniasis (ZCL) is the most endemic disease in Algeria. Leishmania major is the causative agent, and Gerbils Meriones shawi and Psammomys obesus are the main reservoir hosts. The Province of Setif in Algeria has become a very active focus for this zoonosis. Our study was carried out from January 2017 until January 2019 in ten representative stations of the study area, with an aim to identify the association between zoonosis and reservoir host species. Eighty-six (86) specimens of M. shawi and three (3) specimens of P. obesus were captured and determined. Superficial lesions were subjected to Giemsa stained smears. In addition, Livers and spleens were tested for Leishmania DNA using ITS1 PCR. The results showed the presence of M. shawi in almost all communities of Setif Province and the highest population with 50 specimens (58.13%) was in the center and south. The microscopic and molecular detection of Leishmania showed high infestation in most specimens of Meriones (32/86) with a rate of 37.20 %, however, no infestation in Psammomys has been detected. The PCA results showed a negative association between rainfall and ZCL cases; however, positive association was found between temperature and the human CL cases. Also, positive association was demonstrated between ZCL in south and center of Setif Province and Meriones captured. Our finding, confirm that M. shawi is the principal reservoir host (76.78%) of the ZCL in Setif Province, since P. obesus was nearly absent (2.67%).

High resolution mapping of nucleic acid containing complexes in vitro and in situ

1996 ◽  
Vol 54 ◽  
pp. 48-49
Author(s):  
D. P. Bazett-Jones ◽  
M. J. Hendzel
Keyword(s):  
Nucleic Acid ◽  
High Resolution ◽  
Transcription Initiation ◽  
Molecular Mechanisms ◽  
Heavy Atom ◽  
Regulation Of Transcription ◽  
High Exposure ◽  
Resolution Mapping ◽  
Tata Sequence

Structural analysis of combinations of nucleosomes and transcription factors on promoter and enhancer elements is necessary in order to understand the molecular mechanisms responsible for the regulation of transcription initiation. Such complexes are often not amenable to study by high resolution crystallographic techniques. We have been applying electron spectroscopic imaging (ESI) to specific problems in molecular biology related to transcription regulation. There are several advantages that this technique offers in studies of nucleoprotein complexes. First, an intermediate level of spatial resolution can be achieved because heavy atom contrast agents are not necessary. Second, mass and stoichiometric relationships of protein and nucleic acid can be estimated by phosphorus detection, an element in much higher proportions in nucleic acid than protein. Third, wrapping or bending of the DNA by the protein constituents can be observed by phosphorus mapping of the complexes. Even when ESI is used with high exposure of electrons to the specimen, important macromolecular information may be provided. For example, an image of the TATA binding protein (TBP) bound to DNA is shown in the Figure (top panel). It can be seen that the protein distorts the DNA away from itself and much of its mass sits off the DNA helix axis. Moreover, phosphorus and mass estimates demonstrate whether one or two TBP molecules interact with this particular promoter TATA sequence.

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