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1.
PLoS Pathog ; 16(12): e1009133, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33320907

RESUMO

The rapid and aggressive spread of artemisinin-resistant Plasmodium falciparum carrying the C580Y mutation in the kelch13 gene is a growing threat to malaria elimination in Southeast Asia, but there is no evidence of their spread to other regions. We conducted cross-sectional surveys in 2016 and 2017 at two clinics in Wewak, Papua New Guinea (PNG) where we identified three infections caused by C580Y mutants among 239 genotyped clinical samples. One of these mutants exhibited the highest survival rate (6.8%) among all parasites surveyed in ring-stage survival assays (RSA) for artemisinin. Analyses of kelch13 flanking regions, and comparisons of deep sequencing data from 389 clinical samples from PNG, Indonesian Papua and Western Cambodia, suggested an independent origin of the Wewak C580Y mutation, showing that the mutants possess several distinctive genetic features. Identity by descent (IBD) showed that multiple portions of the mutants' genomes share a common origin with parasites found in Indonesian Papua, comprising several mutations within genes previously associated with drug resistance, such as mdr1, ferredoxin, atg18 and pnp. These findings suggest that a P. falciparum lineage circulating on the island of New Guinea has gradually acquired a complex ensemble of variants, including kelch13 C580Y, which have affected the parasites' drug sensitivity. This worrying development reinforces the need for increased surveillance of the evolving parasite populations on the island, to contain the spread of resistance.


Assuntos
Anti-Infecciosos , Artemisininas , Resistência a Medicamentos/genética , Genes de Protozoários/genética , Plasmodium falciparum/genética , Anti-Infecciosos/uso terapêutico , Artemisininas/uso terapêutico , Estudos Transversais , Humanos , Malária Falciparum/tratamento farmacológico , Malária Falciparum/parasitologia , Mutação , Papua Nova Guiné
2.
PLoS One ; 15(10): e0236616, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33044964

RESUMO

Asexual blood stages of the malaria parasite are readily amenable to genetic modification via homologous recombination, allowing functional studies of parasite genes that are not essential in this part of the life cycle. However, conventional reverse genetics cannot be applied for the functional analysis of genes that are essential during asexual blood-stage replication. Various strategies have been developed for conditional mutagenesis of Plasmodium, including recombinase-based gene deletion, regulatable promoters, and mRNA or protein destabilization systems. Among these, the dimerisable Cre (DiCre) recombinase system has emerged as a powerful approach for conditional gene deletion in P. falciparum. In this system, the bacteriophage Cre is expressed in the form of two separate, enzymatically inactive polypeptides, each fused to a different rapamycin-binding protein. Rapamycin-induced heterodimerization of the two components restores recombinase activity. We have implemented the DiCre system in the rodent malaria parasite P. berghei, and show that rapamycin-induced excision of floxed DNA sequences can be achieved with very high efficiency in both mammalian and mosquito parasite stages. This tool can be used to investigate the function of essential genes not only in asexual blood stages, but also in other parts of the malaria parasite life cycle.


Assuntos
Deleção de Genes , Edição de Genes , Genes de Protozoários/genética , Integrases/metabolismo , Malária/parasitologia , Mutagênese , Plasmodium berghei/genética , Animais , Feminino , Integrases/química , Integrases/genética , Estágios do Ciclo de Vida , Malária/genética , Malária/metabolismo , Camundongos , Camundongos Endogâmicos C57BL
3.
Parasitol Res ; 119(10): 3327-3338, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32789533

RESUMO

The evolutionary history of Acanthamoeba has been substantially resolved by the 18S rDNA phylogeny which made it possible to delimit the main lines associated with some classical species. Some of them have proven to be polyphyletic, but the inappropriate use of treating under the same names unrelated strains persists. In this study, phylogenies based on the complete genes of nuclear and mitochondrial rDNA were compared, in order to verify the congruence of the different lines. Various groups can thus be identified, some of which associated with the type strains of given species. Recognizing them only by their species names would significantly reduce the current confusion, in addition to logically following basic taxonomic rules. In this manner, the well-known polyphyletic taxa A. castellanii and A. polyphaga, are restricted to the two lines specified by their type strains, while other widely used strains like Neff and Linc-AP1 that are often confused with the previous ones, can be assigned to their own lines. New species are potentially present in other groups and additional efforts are needed to delimit them.


Assuntos
Acanthamoeba/classificação , Filogenia , Acanthamoeba/genética , Animais , DNA Mitocondrial/genética , DNA Ribossômico/genética , Genes de Protozoários/genética , Genótipo
4.
Korean J Parasitol ; 58(3): 257-265, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32615739

RESUMO

The outbreak of human toxoplasmosis can be attributed to ingestion of food contaminated with Toxoplasma gondii. Toxoplasmosis recently increased in domestic and stray dogs and cats. It prompted studies on the zoonotic infectious diseases transmitted via these animals. Sero- and antigen prevalences of T. gondii in dogs and cats were surveyed using ELISA and PCR, and B1 gene phylogeny was analyzed in this study. Toxoplasmosis antibodies were measured on sera of 403 stray cats, 947 stray dogs, 909 domestic cats, and 2,412 domestic dogs collected at nationwide regions, Korea from 2017 to 2019. In addition, whole blood, feces, and tissue samples were also collected from stray cats (1,392), stray dogs (686), domestic cats (3,040), and domestic dogs (1,974), and T. gondii-specific B1 gene PCR was performed. Antibody prevalence of stray cats, stray dogs, domestic cats, and domestic dogs were 14.1%, 5.6%, 2.3%, and 0.04%, respectively. Antigen prevalence of these animals was 0.5%, 0.2%, 0.1%, and 0.4%, respectively. Stray cats revealed the highest infection rate of toxoplasmosis, followed by stray dogs, domestic cats, and domestic dogs. B1 gene positives were 5 of stray cats, and identified to high/moderate pathogenic Type I/III group. These findings enforce that preventive hygienic measure should be strengthened at One Health level in dogs and cats, domestic and stray, to minimize human toxoplasmosis infections.


Assuntos
Doenças do Gato/epidemiologia , Doenças do Gato/parasitologia , Gatos/parasitologia , Doenças do Cão/epidemiologia , Doenças do Cão/parasitologia , Cães/parasitologia , Genes de Protozoários/genética , Toxoplasma/genética , Toxoplasmose Animal/parasitologia , Animais , Filogenia , Reação em Cadeia da Polimerase , República da Coreia/epidemiologia , Estudos Soroepidemiológicos , Toxoplasmose/parasitologia , Toxoplasmose/prevenção & controle
5.
Arch Microbiol ; 202(10): 2689-2695, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32725599

RESUMO

Depression disorder is one of the most common psychological recognitions that characterized by sadness, low self-confidence, and disinterest in every activity. Considering evidence showing the effects of toxoplasmosis on the psychological disease, this study conducted to investigate the serological and molecular aspects of Toxoplasma gondii infection among patients with depression. In this study, after selecting the patients with depression and control groups under the supervision of a psychologist, the blood samples were collected and the serum samples and buffy coat were separated. The specific anti-Toxoplasma IgG antibodies in serum samples were evaluated using the commercial ELISA kit. Then the desired region of the Toxoplasma B1 gene was amplified using the specific primers. To confirm the specificity of primers to amplify the B1 gene of Toxoplasma, the extracted PCR product was sequenced. The overall prevalence of toxoplasmosis in patients with depression was 59.8 and 60.19% by ELISA and PCR, respectively. In the control group, the prevalence of Toxoplasma was 56.3 and 40.2% by serology and PCR. There was a significant correlation between the prevalence of toxoplasmosis and depression. Moreover, a significant difference was found between the variables of age, sex, kind of nutrition, level of education and toxoplasmosis among the two cases and control groups. The higher prevalence of Toxoplasma infection among patients with depression compared with the control group indicates the probable impact of this parasite on depression and exacerbates its symptoms, which requires special attention of specialist physicians and patient's relatives.


Assuntos
Anticorpos Antiprotozoários/sangue , Depressão/complicações , Depressão/parasitologia , Toxoplasma , Toxoplasmose/complicações , Toxoplasmose/epidemiologia , Anticorpos Antiprotozoários/genética , Estudos de Casos e Controles , Ensaio de Imunoadsorção Enzimática , Feminino , Genes de Protozoários/genética , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Masculino , Reação em Cadeia da Polimerase , Prevalência , Fatores de Risco , Toxoplasma/genética , Toxoplasma/imunologia , Toxoplasmose/imunologia
6.
Parasitol Res ; 119(7): 2359-2362, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32500368

RESUMO

Several Cryptosporidium species that infect reptiles, especially squamates, are well described, but there is limited data about Cryptosporidium species infecting crocodilians. In this study, we assess the occurrence of intestinal parasites using traditional microscopic examination and describe the prevalence and Cryptosporidium species in the captive-bred Chinese alligators (Alligator sinensis) in eastern China using molecular methods. The results of microscopic examination showed that no intestinal parasites were detected among the 491 fecal samples examined from the Chinese alligators. The overall prevalence for Cryptosporidium was 0.41% (2/491) by PCR detection using the SSU rRNA locus. Sequence and phylogenetic analysis of the SSU rRNA, COWP, and actin genes revealed the presence of Cryptosporidium testudinis, which has been isolated primarily from chelonians. This is the first detection of the specific DNA of C. testudinis in the feces of the Chinese alligator. This study expands our knowledge of the Cryptosporidium species involved in crocodiles, and more extensive studies are necessary to confirm the validity of C. testudinis in crocodiles.


Assuntos
Jacarés e Crocodilos/parasitologia , Criptosporidiose/parasitologia , Cryptosporidium/isolamento & purificação , Animais , China/epidemiologia , Criptosporidiose/epidemiologia , Cryptosporidium/classificação , Cryptosporidium/genética , DNA de Protozoário/genética , DNA Ribossômico/genética , Fezes/parasitologia , Genes de Protozoários/genética , Filogenia
7.
Parasitol Res ; 119(7): 2363-2367, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32500369

RESUMO

Rhipicephalus appendiculatus is the major tick vector of Theileria parva, an apicomplexan protozoan parasite that causes the most economically important and lethal disease of cattle in East and central Africa. The African cape buffalo (Syncerus caffer) is the major wildlife host of T. parva from southern Uganda and Kenya to southern Africa. We show herein that R. appendiculatus appears to be absent from the two largest national parks in northern Uganda. Syncerus caffer is common in both of these national parks, specifically Murchison falls (MFNP) and Kidepo Valley (KVNP). We re-confirmed the previously reported absence of T. parva in buffalo sampled in the two northern parks based on RLB data using a nested PCR based on the T. parva p104 gene. By contrast, T. parva-infected R. appendiculatus ticks and parasite-infected buffalo were present in Lake Mburo (LMNP) in South central Uganda. This suggests that the distribution of R. appendiculatus, which is predicted to include the higher rainfall regions of northern Uganda, may be limited by additional, as yet unknown factors.


Assuntos
Vetores Aracnídeos/parasitologia , Búfalos/parasitologia , Rhipicephalus/parasitologia , Theileria parva/fisiologia , Animais , Animais Selvagens/parasitologia , DNA de Protozoário/genética , Ecossistema , Genes de Protozoários/genética , Parques Recreativos , Theileria parva/genética , Theileriose/parasitologia , Theileriose/transmissão , Uganda/epidemiologia
8.
Parasitol Res ; 119(7): 2299-2307, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32476060

RESUMO

In the intermediate hosts, tachyzoites of T. gondii predominate in the acute stage while bradyzoites persist inside tissue cysts with the potential for reactivation. The two stages exhibit different metabolic and antigenic characters. The present study aimed to investigate temporal expression of Toxoplasma SAG1 and BAG1 genes in the brain tissue and the coincident parasitological and histopathological findings in mice models of toxoplasmosis. The study included group A: mice infected with RH strain and sacrificed 7 days post-infection (p.i.); group B: mice infected with RH strain and treated with sulfamethoxazole-trimethoprim (30 mg/kg/day and 150 mg/kg/day respectively) 24 h p.i. until sacrificed at days 5, 10, or 20 post-treatment; group C: mice infected with ME-49 strain and sacrificed at days 7, 27, 47, or 67 p.i; and group D: mice infected with ME-49 strain and received dexamethasone daily starting at day 68 p.i. and scarified at days 6 or 10 post-treatment. All mice were inspected daily for abnormal physical signs. Peritoneal exudate and brain homogenate were examined for detection of Toxoplasma stages. Brain sections were examined histopathologically. SAG1 and BAG1 gene expression was evaluated using reverse transcription real-time polymerase chain reaction and the ΔΔCt method. Results revealed that marked BAG1 upregulation is consistent with detection of Toxoplasma cysts and degenerative changes while predominance of tachyzoites and inflammatory infiltrate is compatible with SAG1 upregulation. The study sheds light on the potential for using stage-specific gene expression pattern as markers for evaluation of toxoplasmosis disease progression in clinical settings.


Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Estágios do Ciclo de Vida/genética , Toxoplasma/genética , Toxoplasmose Animal/patologia , Toxoplasmose Animal/parasitologia , Animais , Encéfalo/parasitologia , Encéfalo/patologia , Feminino , Genes de Protozoários/genética , Camundongos , Encistamento de Parasitas/genética , Toxoplasma/crescimento & desenvolvimento
9.
Vet Parasitol ; 281: 109115, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32361137

RESUMO

Giardia spp. and Cryptosporidium spp. are common gastrointestinal parasites with the potential for zoonotic transmission. This study aimed to (1) determine the genotypes occurring in dogs and coyotes occupying a similar urban area; (2) determine if these hosts were infected with potentially zoonotic genotypes; (3) provide baseline molecular data. In August and September 2012, 860 dog owners living in neighborhoods bordering six urban parks in Calgary, Alberta, Canada, provided faecal samples from their dogs. From March 2012 through July 2013, 193 coyote faeces were also collected from five of six of the same parks. Direct immunofluorescence microscopy (DFA) indicated that Giardia spp. and Cryptosporidium spp. infected a total of 64 (7.4%) and 21 (2.4%) dogs, as well as 15 (7.8%) and three (1.6%) coyotes, respectively. Semi-nested, polymerase chain reactions targeting the 16S small-subunit ribosomal ribonucleic acid (SSU rRNA) and 18S SSU rRNA genes of Giardia spp. and Cryptosporidium spp., respectively, were conducted on samples that screened positive by DFA, and products were sequenced and genotyped. Dogs were infected with Giardia intestinalis canid-associated assemblages C (n = 14), D (n = 13), and Cryptosporidium canis (n = 3). Similarly, G. intestinalis assemblages C (n = 1), D (n = 1) and C. canis (n = 1), were detected in coyotes, as well as G. intestinalis assemblage A (n = 1) and Cryptosporidium vole genotype (n = 1). Dogs and coyotes were predominantly infected with host-specific genotypes and few potentially zoonotic genotypes, suggesting that they may not represent a significant risk for zoonotic transmission of these parasites in urban areas where these hosts are sympatric.


Assuntos
Coiotes/parasitologia , Criptosporidiose/parasitologia , Cryptosporidium/genética , Doenças do Cão/parasitologia , Giardia/genética , Giardíase/parasitologia , Alberta/epidemiologia , Animais , Doenças do Cão/epidemiologia , Cães , Fezes/parasitologia , Genes de Protozoários/genética , Genes de RNAr/genética , Genótipo , Giardíase/epidemiologia , Parques Recreativos , Zoonoses/epidemiologia , Zoonoses/parasitologia
10.
Gene ; 743: 144624, 2020 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-32224274

RESUMO

The giant ciliate Stentor coeruleus (S. coeruleus) is a suitable model organism for studying morphogenesis and regeneration at the single-cell level. It contains a prominent structure on the anterior end of the cell, known as the oral apparatus (OA). OA can be induced to shed by urea treatment and then new OA regenerates via a series of defined morphological events and the cell resumes normal feeding activity. We identified OA constituents in S. coeruleus by mass spectrometry. A total of 882 OA-associated proteins were identified; the homologs of 181 of these are known OA constituents in other organisms. The expression pattern of OA-associated genes during regeneration was investigated using single-cell transcriptome sequencing. The expression of most OA-associated genes was high during regeneration, indicating their stable expression after OA shedding. We also identified OA-associated differentially expressed genes that may be involved in regulating OA reconstruction. In summary, this study gives preliminary insight into the molecular basis of OA in S. coeruleus.


Assuntos
Cilióforos/fisiologia , Genes de Protozoários/genética , Proteínas de Protozoários/metabolismo , Regeneração , Espectrometria de Massas , Proteômica , Proteínas de Protozoários/genética , Análise de Sequência de RNA , Análise de Célula Única
11.
Eur J Protistol ; 73: 125685, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32114251

RESUMO

The genus Cunea Kudryavtsev and Pawlowski, 2015 (Amoebozoa, Dactylopodida) was initially described from the oceanic benthos: C. profundata, from over 5 km depth in the Atlantic Ocean, and C. thuwala from the Red Sea benthos at ca. 60 m depth. Both species are identical to each other in morphology (including cell coat ultrastructure), but differ significantly in the gene sequence data, including barcoding loci of small subunit ribosomal RNA and cytochrome oxidase subunit 1 gene, as well as actin. This paper describes the third species of Cunea, C. russae n. sp. isolated from a brackish water habitat without a direct connection to the ocean, a small spring of brackish water (19‰) emerging from a 246 m deep hole in the earth. This species is morphologically identical to the previous two amoebae, but differs from them significantly in the gene sequence data and ecological preferences. In particular, this species has the broadest salinity tolerance range, being able to reproduce well already at 2.5‰. It is also capable of resisting cold temperatures, like C. profundata. The data obtained suggest that the genus Cunea may comprise a significant taxonomic diversity represented by morphologically identical, but quickly diverging species with significant ecological plasticity.


Assuntos
Amebozoários/classificação , Amebozoários/citologia , Amebozoários/genética , Biodiversidade , Temperatura Baixa , Genes de Protozoários/genética , Águas Salinas , Especificidade da Espécie
12.
Nat Commun ; 11(1): 1503, 2020 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-32198457

RESUMO

In the malaria parasite Plasmodium falciparum, the switch from asexual multiplication to sexual differentiation into gametocytes is essential for transmission to mosquitos. The transcription factor PfAP2-G is a key determinant of sexual commitment that orchestrates this crucial cell fate decision. Here we identify the direct targets of PfAP2-G and demonstrate that it dynamically binds hundreds of sites across the genome. We find that PfAP2-G is a transcriptional activator of early gametocyte genes, and identify differences in PfAP2-G occupancy between gametocytes derived via next-cycle and same-cycle conversion. Our data implicate PfAP2-G not only as a transcriptional activator of gametocyte genes, but also as a potential regulator of genes important for red blood cell invasion. We also find that regulation by PfAP2-G requires interaction with a second transcription factor, PfAP2-I. These results clarify the functional role of PfAP2-G during sexual commitment and early gametocytogenesis.


Assuntos
Malária/parasitologia , Plasmodium falciparum/crescimento & desenvolvimento , Plasmodium falciparum/genética , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Sistemas CRISPR-Cas , Eritrócitos/parasitologia , Regulação da Expressão Gênica , Genes de Protozoários/genética , Malária/transmissão , Malária Falciparum/parasitologia , Plasmodium falciparum/metabolismo , Proteínas de Protozoários/sangue , Fatores de Transcrição/metabolismo
13.
Sci Rep ; 10(1): 1583, 2020 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-32005846

RESUMO

Acanthamoeba spp. are predominant free-living amoebae of water and soil. They have been used as tools for the isolation and culture of microbes that resist after their phagocytosis, such as Legionella-like bacteria, and, more recently giant viruses for which differences in permissiveness have been reported. However, problems have been reported regarding their identification at the species level. The present work implemented specific PCR systems for the detection and identification of Acanthamoeba species through comparison of sequences and phylogenetic analyses. Thirty-three Acanthamoeba isolates were studied, including 20 reference strains and 13 isolates retrieved from water, soil or clinical samples. Previous delineation of a core genome encompassing 826 genes based on draft genome sequences from 14 Acanthamoeba species allowed designing PCR systems for one of these core genes that encodes an alanine-tRNA ligase. These primers allowed an efficient and specific screening to detect Acanthamoeba presence. In addition, they identified all 20 reference strains, while partial and complete sequences coding for 18S ribosomal RNA identified only 11 (55%). We found that four isolates may be considered as new Acanthamoeba species. Consistent with previous studies, we demonstrated that some Acanthamoeba isolates were incorrectly assigned to species using the 18S rDNA sequences. Our implemented tool may help determining which Acanthamoeba strains are the most efficient for the isolation of associated microorganisms.


Assuntos
Acanthamoeba/genética , Genes de Protozoários/genética , Acanthamoeba castellanii/genética , Alanina-tRNA Ligase/genética , Filogenia , Reação em Cadeia da Polimerase , RNA Ribossômico 18S/genética
14.
Parasitol Res ; 119(3): 1101-1108, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32006227

RESUMO

Limited data are available on infection rates and genetic identity of Enterocytozoon bieneusi and Giardia duodenalis in horses and donkeys. In this study, 865 fecal specimens were collected from donkeys (n = 540) and horses (n = 325) in three provinces and autonomous regions in northern China during 2015-2019. Enterocytozoon bieneusi was detected and genotyped by PCR and sequence analyses of the ribosomal internal transcribed spacer (ITS) and G. duodenalis was detected and genotyped by PCR and sequence analyses of the ß-giardin, glutamate dehydrogenase, and triosephosphate isomerase genes. The overall infection rates of E. bieneusi and G. duodenalis were 21.9% (118/540) and 11.5% (62/540) in donkeys, and 7.4% (24/325) and 2.8% (9/325) in horses, respectively. These differences in infection rates of E. bieneusi and G. duodenalis between donkeys and horses were significant (χ2 = 30.9, df = 1, P < 0.0001; χ2 = 20.4, df = 1, P < 0.0001, respectively). By age, the 28.9% infection rate of E. bieneusi in donkeys under 6 months was significantly higher than that in animals over 6 months (6.0%; χ2 = 35.2, df = 1, P < 0.0001). In contrast, donkeys of 6-12 months had higher infection rate (35.9%) of G. duodenalis than donkeys under 6 months (9.9%; χ2 = 22.1, df = 1, P < 0.0001) and over 12 months (8.7%; χ2 = 17.3, df = 1, P < 0.0001). In horses, animals of > 12 months had significantly higher infection rate (31.1%) of E. bieneusi than horses under 6 months (3.4%; χ2 = 29.4, df = 1, P < 0.0001) and 6-12 months (3.8%; χ2 = 26.1, df = 1, P < 0.0001). Twenty genotypes of E. bieneusi were detected, including six known ones and 14 new genotypes. Among them, nine genotypes in 45% E. bieneusi-positive specimens belonged to the zoonotic group 1. Similarly, three G. duodenalis assemblages were detected, including A (in 2 horses and 30 donkeys), B (in 6 horses and 29 donkeys), and E (in 1 horse); three donkeys had coinfections of assemblages A and B. The assemblage A isolates identified all belong to the sub-assemblage AI. These results indicate that unlike in other farm animals, there is a common occurrence of zoonotic E. bieneusi and G. duodenalis genotypes in horses and donkeys.


Assuntos
Enterocytozoon/fisiologia , Equidae/parasitologia , Giardia lamblia/fisiologia , Giardíase/veterinária , Cavalos/parasitologia , Microsporidiose/veterinária , Animais , Animais Domésticos/parasitologia , China/epidemiologia , Enterocytozoon/classificação , Enterocytozoon/genética , Fezes/parasitologia , Genes de Protozoários/genética , Genótipo , Giardia lamblia/classificação , Giardia lamblia/genética , Giardíase/epidemiologia , Giardíase/parasitologia , Especificidade de Hospedeiro , Microsporidiose/epidemiologia , Microsporidiose/parasitologia , Filogenia , Prevalência , Zoonoses/transmissão
15.
J Immunol ; 204(6): 1562-1570, 2020 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-31996457

RESUMO

In this study, we generated a tkl1 deletion mutant in the Toxoplasma gondii type 1 RH (RHΔtkl1) strain and tested the protective efficacies of vaccination using RHΔtkl1 tachyzoites against acute, chronic, and congenital T. gondii infections in Kunming mice. Mice vaccinated with RHΔtkl1 mounted a strong humoral and cellular response as shown by elevated levels of anti-T. gondii-specific IgG, IL-2, IL-12, IFN-γ, and IL-10. All RHΔtkl1-vaccinated mice survived a lethal challenge with 1 × 103 tachyzoites of type 1 RH or ToxoDB#9 (PYS or TgC7) strain as well as 100 cysts or oocysts of Prugniuad strain. All mock-vaccinated plus infected mice have died. Vaccination also protected against cyst- or oocyst-caused chronic infection, reduced vertical transmission caused by oocysts, increased litter size, and maintained body weight of pups born to dams challenged with 10 oocysts on day 5 of gestation. In contrast, all mock-vaccinated plus oocysts-infected dams had aborted, and no fetus has survived. Vaccinated dams remained healthy postinfection, and their brain cyst burden was significantly reduced compared with mock-vaccinated dams infected with oocysts. In vivo depletion of CD4+ T cells, CD8+ T cells, and B cells revealed that CD8+ T cells are involved in the protection of mice against T. gondii infection. Additionally, adoptive transfer of CD8+ T cells from RHΔtkl1-vaccinated mice significantly enhanced the survival of naive mice infected with the pathogenic strain. Together, these data reaffirm the importance of CD8+ T cell responses in future vaccine design for toxoplasmosis and present T. gondii tkl1 gene as a promising vaccine candidate.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Vacinas Protozoárias/administração & dosagem , Toxoplasma/imunologia , Toxoplasmose Animal/prevenção & controle , Toxoplasmose Congênita/prevenção & controle , Doença Aguda/terapia , Transferência Adotiva , Animais , Linfócitos T CD8-Positivos/transplante , Doença Crônica/prevenção & controle , Modelos Animais de Doenças , Feminino , Genes de Protozoários/genética , Genes de Protozoários/imunologia , Humanos , Transmissão Vertical de Doença Infecciosa/prevenção & controle , Gado/parasitologia , Masculino , Camundongos , Proteínas de Protozoários/genética , Proteínas de Protozoários/imunologia , Vacinas Protozoárias/genética , Vacinas Protozoárias/imunologia , Deleção de Sequência , Toxoplasma/genética , Toxoplasmose Animal/imunologia , Toxoplasmose Animal/parasitologia , Toxoplasmose Animal/transmissão , Toxoplasmose Congênita/imunologia , Toxoplasmose Congênita/parasitologia , Toxoplasmose Congênita/transmissão , Virulência/genética , Fatores de Virulência/genética , Fatores de Virulência/imunologia
16.
PLoS One ; 14(11): e0224743, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31751362

RESUMO

Human infections due to the monkey malaria parasite Plasmodium knowlesi are increasingly being reported from Malaysia. The parasite causes high parasitaemia, severe and fatal malaria in humans thus there is a need for urgent measures for its control. The MSP4 is a potential vaccine candidate, which is well studied in Plasmodium falciparum and Plasmodium vivax; however, no study has been conducted in the orthologous gene of P. knowlesi. In this study, we investigated the level of polymorphisms, haplotypes, natural selection and population structure of full-length pkmsp4 in 32 clinical samples from Malaysian Borneo along with 4 lab-adapted strains. We found low levels of polymorphism across the gene with exon I showing higher diversity than the exon II. The C- terminal epidermal growth factor (EGF) domains and GPI-anchored region within exon II were mostly conserved with only 2 non-synonymous substitutions. Although 21 amino acid haplotypes were found, the frequency of mutation at the majority of the polymorphic positions was low. We found evidence of negative selection at the exon II of the gene indicating existence of functional constraints. Phylogenetic haplotype network analysis identified shared haplotypes and indicated geographical clustering of samples originating from Peninsular Malaysia and Malaysian Borneo. High population differentiation values were observed within parasite populations originating from Malaysian Borneo (Kapit, Sarikei and Betong) and laboratory-adapted strains obtained from Peninsular Malaysia and Philippines indicating distinct population structure. This is the first study to genetically characterize the full-length msp4 gene from clinical isolates of P. knowlesi from Malaysia and thus would be very useful for future rational vaccine studies. Further studies with higher number of samples and functional characterization of the protein will be necessary.


Assuntos
Antígenos de Protozoários/genética , Variação Genética , Malária/parasitologia , Plasmodium knowlesi/genética , Proteínas de Protozoários/genética , Animais , Antígenos de Protozoários/isolamento & purificação , Bornéu , DNA de Protozoário/genética , DNA de Protozoário/isolamento & purificação , Genes de Protozoários/genética , Geografia , Humanos , Malásia , Merozoítos/genética , Merozoítos/isolamento & purificação , Filipinas , Filogenia , Plasmodium knowlesi/isolamento & purificação , Proteínas de Protozoários/isolamento & purificação , Seleção Genética , Análise de Sequência de DNA
17.
PLoS Comput Biol ; 15(9): e1007329, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31509524

RESUMO

Empirical evidence suggests that the malaria parasite Plasmodium falciparum employs a broad range of mechanisms to regulate gene transcription throughout the organism's complex life cycle. To better understand this regulatory machinery, we assembled a rich collection of genomic and epigenomic data sets, including information about transcription factor (TF) binding motifs, patterns of covalent histone modifications, nucleosome occupancy, GC content, and global 3D genome architecture. We used these data to train machine learning models to discriminate between high-expression and low-expression genes, focusing on three distinct stages of the red blood cell phase of the Plasmodium life cycle. Our results highlight the importance of histone modifications and 3D chromatin architecture in Plasmodium transcriptional regulation and suggest that AP2 transcription factors may play a limited regulatory role, perhaps operating in conjunction with epigenetic factors.


Assuntos
Biologia Computacional/métodos , Código das Histonas/genética , Modelos Estatísticos , Nucleossomos/genética , Plasmodium falciparum/genética , Eritrócitos/parasitologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/genética , Genes de Protozoários/genética , Humanos , Estágios do Ciclo de Vida/genética , Aprendizado de Máquina , Malária Falciparum , Modelos Biológicos , Plasmodium falciparum/citologia , Plasmodium falciparum/patogenicidade
18.
Nat Microbiol ; 4(12): 2226-2236, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31477896

RESUMO

The apicomplexan parasite Cryptosporidium is a leading global cause of severe diarrhoeal disease and an important contributor to early childhood mortality. Currently, there are no fully effective treatments or vaccines available. Parasite transmission occurs through ingestion of oocysts, through either direct contact or consumption of contaminated water or food. Oocysts are meiotic spores and the product of parasite sex. Cryptosporidium has a single-host life cycle in which both asexual and sexual processes occur in the intestine of infected hosts. Here, we genetically engineered strains of Cryptosporidium to make life cycle progression and parasite sex tractable. We derive reporter strains to follow parasite development in culture and in infected mice and define the genes that orchestrate sex and oocyst formation through mRNA sequencing of sorted cells. After 2 d, parasites in cell culture show pronounced sexualization, but productive fertilization does not occur and infection falters. By contrast, in infected mice, male gametes successfully fertilize female parasites, which leads to meiotic division and sporulation. To rigorously test for fertilization, we devised a two-component genetic-crossing assay using a reporter that is activated by Cre recombinase. Our findings suggest obligate developmental progression towards sex in Cryptosporidium, which has important implications for the treatment and prevention of the infection.


Assuntos
Criptosporidiose/parasitologia , Cryptosporidium parvum/crescimento & desenvolvimento , Cryptosporidium parvum/genética , Estágios do Ciclo de Vida/fisiologia , Desenvolvimento Sexual/fisiologia , Animais , Cryptosporidium parvum/citologia , Modelos Animais de Doenças , Feminino , Fertilização , Expressão Gênica , Genes de Protozoários/genética , Proteínas de Homeodomínio/genética , Interferon gama/genética , Masculino , Camundongos , Camundongos Knockout , Oocistos , Análise de Sequência de RNA
19.
Parasitol Res ; 118(10): 3043-3051, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31471745

RESUMO

Using a combination of morphological and molecular data, we describe a new apicomplexan parasite, Isospora svecica sp. n., from the white-spotted bluethroat, Luscinia svecica cyanecula, from the Czech Republic. Oocysts were found in its intestinal tract. Sporulation was exogenous and took 1-3 days. The oocysts were slightly ellipsoidal, of average size 26.17 × 20.33 µm, with a smooth bilayered wall. Micropyle, oocyst residuum, and polar granules were absent. Sporocysts were bottle-shaped, of an average size of 18.82 × 8.82 µm, with a thin, colourless wall. A conspicuous knob-like Stieda body was present. Substieda body was barely visible. Sporocyst residuum was present in the form of granules of various sizes. Sporozoites were banana-shaped and contained large anterior and small posterior refractile bodies. Partial DNA sequences of three genes were obtained from oocysts of Isospora svecica sp. n., being most closely related to other isosporans described from passerines. Little is known about the parasites of the avian family Muscicapidae, including coccidia, a highly prevalent parasitic protist group in all vertebrate classes. Only six species of the genus Isospora have so far been described in Muscicapidae, together with several "Isospora sp." that in fact most likely represent Isospora lacazei. The newly described Isospora svecica sp. n. differs morphologically from other coccidia reported from muscicapid birds, and represents the first coccidian species described from Luscinia svecica.


Assuntos
Isospora/classificação , Isosporíase/veterinária , Passeriformes/parasitologia , Animais , República Tcheca , Genes de Protozoários/genética , Intestinos/parasitologia , Isospora/citologia , Isospora/genética , Isospora/crescimento & desenvolvimento , Isosporíase/parasitologia , Oocistos/classificação , Oocistos/citologia , Oocistos/genética , Oocistos/crescimento & desenvolvimento , Esporozoítos/classificação , Esporozoítos/citologia , Esporozoítos/genética , Esporozoítos/crescimento & desenvolvimento
20.
Vet Parasitol ; 273: 32-35, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31442890

RESUMO

This study looked to assess the stability of Cryptosporidium parvum genotypes in calves between the final day of treatment with the antiprotozoal halofuginone lactate and seven days post-treatment. Paired faecal samples were collected on the final day of treatment and seven days later from 54 calves across seven farms in South-west England. The presence of Cryptosporidium species was detected using polymerase chain reaction targeting the 18 s rDNA. The presence and genotype of C. parvum was determined by PCR and amplicon sequencing targeting the gp60 locus. On farms where C. parvum was detected at both sampling times there was a distinct genotype shift. Detection of gp60 genotype IIaA15G2R1 decreased from 40% to 7% while IIaA17G1R1 increased from 0% to 41%, supplemented by IIaA16G3R1 in one sample. A shift in C. parvum genotypes present in calves within a one week sampling timeframe has not been described prior to this study, indicating that the timeframe is likely suitable for observing variation in C. parvum populations and interactions with antiprotozoal control strategies.


Assuntos
Doenças dos Bovinos/parasitologia , Criptosporidiose/parasitologia , Cryptosporidium parvum/genética , Animais , Antiprotozoários/uso terapêutico , Bovinos , Doenças dos Bovinos/tratamento farmacológico , Criptosporidiose/tratamento farmacológico , Inglaterra , Fezes/parasitologia , Genes de Protozoários/genética , Genótipo , RNA Ribossômico 18S/genética
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