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1.
PLoS One ; 15(8): e0237187, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32833981

RESUMO

INTRODUCTION: Infection of equids with Trypanosoma brucei (T. brucei) ssp. is of socioeconomic importance across sub-Saharan Africa as the disease often progresses to cause fatal meningoencephalitis. Loop-mediated isothermal amplification (LAMP) has been developed as a cost-effective molecular diagnostic test and is potentially applicable for use in field-based laboratories. PART I: Threshold levels for T. brucei ssp. detection by LAMP were determined using whole equine blood specimens spiked with known concentrations of parasites. Results were compared to OIE antemortem gold standard of T. brucei-PCR (TBR-PCR). RESULTS I: Threshold for detection of T. brucei ssp. on extracted DNA from whole blood was 1 parasite/ml blood for LAMP and TBR-PCR, and there was excellent agreement (14/15) between tests at high (1 x 103/ml) concentrations of parasites. Detection threshold was 100 parasites/ml using LAMP on whole blood (LWB). Threshold for LWB improved to 10 parasites/ml with detergent included. Performance was excellent for LAMP at high (1 x 103/ml) concentrations of parasites (15/15, 100%) but was variable at lower concentrations. Agreement between tests was weak to moderate, with the highest for TBR-PCR and LAMP on DNA extracted from whole blood (Cohen's kappa 0.95, 95% CI 0.64-1.00). PART II: A prospective cross-sectional study of working equids meeting clinical criteria for trypanosomiasis was undertaken in The Gambia. LAMP was evaluated against subsequent TBR-PCR. RESULTS II: Whole blood samples from 321 equids in The Gambia were processed under field conditions. There was weak agreement between LWB and TBR-PCR (Cohen's kappa 0.34, 95% CI 0.19-0.49) but excellent agreement when testing CSF (100% agreement on 6 samples). CONCLUSIONS: Findings support that LAMP is comparable to PCR when used on CSF samples in the field, an important tool for clinical decision making. Results suggest repeatability is low in animals with low parasitaemia. Negative samples should be interpreted in the context of clinical presentation.


Assuntos
Doenças dos Cavalos/parasitologia , Cavalos/parasitologia , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificação de Ácido Nucleico/métodos , Reação em Cadeia da Polimerase/métodos , Trypanosoma brucei brucei/genética , Tripanossomíase Africana/diagnóstico , Tripanossomíase Africana/veterinária , Animais , Estudos Transversais , DNA de Protozoário/sangue , DNA de Protozoário/genética , Feminino , Gâmbia , Masculino , Técnicas de Diagnóstico Molecular/economia , Técnicas de Amplificação de Ácido Nucleico/economia , Reação em Cadeia da Polimerase/economia , Estudos Prospectivos , Sensibilidade e Especificidade , Tripanossomíase Africana/parasitologia
2.
Parasitol Res ; 119(10): 3541-3548, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32803333

RESUMO

The aim of this study was to evaluate, through qPCR, the prevalence of parasitemia in sick kennel dogs naturally infected by canine leishmaniasis. An evaluation of daily changes of the parasitic load in peripheral blood was also performed. A comprehensive clinical examination and the collection of several samples (blood, lymph node, skin, and conjunctiva) were performed in 140 dogs living in an endemic area. Among these, only the dogs with clinically evident leishmaniasis were enrolled (39/140; 27.9%). Twelve (30.8%) out of 39 showed parasitemia, with a low load (median: 4 Leishmania/ml) despite a high lymph node parasite load (median: 4000 Leishmania/ml) and high IFAT titers (≥ 1:640). Seven sick dogs were sampled every 4 h for 6 times during a 24-h period, in order to obtain light- and dark-span samples. Only one (14.3%) out of the seven serial sampled dogs showed Leishmania DNA in the peripheral blood in two samples (2/42; 4.8%). Surprisingly, Leishmania DNA was also detected in the peripheral blood of asymptomatic dogs, negative to both serology and PCR performed on samples other than blood (6/101; 5.9%). The present study confirms that in canine leishmaniasis parasitemia is uncommon and even transitory. Even if recommended, microscopic examination is confirmed as a low sensitivity method with a lower diagnostic utility in canine leishmaniasis than qPCR. Moreover, circulating Leishmania DNA can be found even in healthy dogs. This finding is important in clinical practice because in endemic areas it suggests a transfusion risk and a possible transmission to the vector.


Assuntos
Doenças do Cão/parasitologia , Leishmania infantum/isolamento & purificação , Leishmaniose Visceral/veterinária , Parasitemia/veterinária , Animais , DNA de Protozoário/sangue , DNA de Protozoário/genética , Doenças do Cão/epidemiologia , Cães , Leishmania infantum/genética , Leishmaniose Visceral/epidemiologia , Leishmaniose Visceral/parasitologia , Carga Parasitária/veterinária , Parasitemia/epidemiologia , Parasitemia/parasitologia , Prevalência , Reação em Cadeia da Polimerase em Tempo Real/veterinária
3.
Artigo em Inglês | MEDLINE | ID: mdl-32752015

RESUMO

Malaria has been for millennia one of the best known and most destructive diseases affecting humans. Its high impact has aroused great interest for the development of new effective and reliable diagnostic techniques. Recently it has been recently published that hairs from mammal hosts are able to capture, hold and finally remove foreign DNA sequences of Leishmania parasites. The aim of this study was to check if Plasmodium falciparum (P. falciparum) DNA remains stable in blood samples deposited in Whatman paper after suffering different transport and storage conditions, and to compare the sensitivity of these results with those offered by thick a smear and Rapid Diagnostic Test, and besides to examine whether P. falciparum DNA would be detected and quantified by Real time quantitative PCR (qPCR) from hairs of people with different types of malaria. P. falciparum Histidine Repeat Protein II (pHRP-II) antigen detection and P. falciparum DNA were detected in 18 of 19 dry blood samples adhered to Whatman paper (94.74%), besides, Plasmodium DNA was also detected in seven out of 19 hair samples analyzed (36.84%), remaining stable until analysis for several months under the exposure to different environmental conditions. Although the sensitivity of PCR for the diagnosis of malaria in hair samples is not as high as blood analysis, the study of Plasmodium DNA presence in blood and hair could constitute a complementary tool with numerous advantages in sample collection, transport and storage. We suggest that the method could be also applied to medical, forensic and paleo-parasitological diagnosis, not only for malaria but also for searching many other pathogens in hair samples.


Assuntos
DNA de Protozoário , Malária Falciparum , Malária , Plasmodium falciparum , Animais , DNA de Protozoário/sangue , Testes Diagnósticos de Rotina , Feminino , Cabelo/parasitologia , Humanos , Malária/diagnóstico , Malária Falciparum/diagnóstico , Malária Falciparum/genética , Masculino , Plasmodium falciparum/genética , Reação em Cadeia da Polimerase em Tempo Real , Sensibilidade e Especificidade , Manejo de Espécimes
4.
Ann Parasitol ; 66(2): 143-156, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32531102

RESUMO

Recently, Leishmania infantum has increasingly been detected in stray cats in endemic regions of the world. Cats have been considered playing a role in the epidemiology of visceral leishmaniosis, an endemic zoonosis in Iran. The studies concerning feline leishmaniosis (FeL) allow the hypothesis that cats can be considered as potential reservoirs. The investigations on Leishmania infection in cats are very few in Iran and therefore we aimed to assess the L. infantum infection in stray cats and its possible role in transmission of the disease to human by direct agglutination test (DAT), ELISA, nested-PCR and confirmation via sequencing and phylogenetic analysis in Fars province, Iran. Whole blood samples were obtained from 174 stray cats. Anti-Leishmania antibodies were detected in the sera using DAT and ELISA. DNA was extracted from the buffy coat of each subject and PCR amplified, targeting Leishmania kDNA gene. PCR results were confirmed by sequence analysis. Prevalence of clinical signs in positive cats was 19.0%. Anti-Leishmania antibodies with different titers were detected in 48 (27.59%) and leishmanial DNA in 36 (20.69%) of the cats. The sequencing of PCR-positive cats revealed the parasite as L. infantum. A high seroprevalence of L. infantum was revealed, with higher levels in males, adult cats, and those living in rural districts and southern zones. Despite the reservoir task of cats in nature is still ambiguous, the high serological and molecular detection of L. infantum in stray cats indicates that cats are regularly bitten by infected sand flies in Fars province, southern Iran, and may have a potential reservoir role in the maintenance of L. infantum in the endemic areas of zoonotic visceral leishmaniosis in Iran. Anyway, Leishmania infection must be appraised in the differential diagnosis of cutaneous or systemic clinical signs in cats.


Assuntos
Anticorpos Antiprotozoários/sangue , Leishmania infantum , Leishmaniose Visceral/veterinária , Animais , Gatos , DNA de Protozoário/sangue , Feminino , Humanos , Irã (Geográfico)/epidemiologia , Leishmaniose Visceral/sangue , Leishmaniose Visceral/epidemiologia , Leishmaniose Visceral/transmissão , Masculino , Filogenia , Reação em Cadeia da Polimerase , Prevalência , Estudos Soroepidemiológicos
5.
Arch Microbiol ; 202(7): 1881-1888, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32448961

RESUMO

Due to defects and drawbacks of most conventional diagnostic methods including serology for the diagnosis of toxoplasmosis as a dangerous opportunistic infection in immunocompromised individuals, the accurate, rapid, and sensitive detection of infection in such patients is essential. In this study, the TaqMan probe-based real-time PCR and, a relatively new nucleic acid amplification method, the loop-mediated isothermal amplification (LAMP) technique was compared based on the repetitive elements (RE) sequence to detect Toxoplasma gondii (T. gondii) DNA in blood samples of immunocompromised individuals. During this study, 119 blood samples from immunocompromised cancer patients with renal failure, undergoing dialysis were studied. After DNA extraction from blood samples using the salt extraction method, the molecular techniques of TaqMan probe-based real-time PCR and LAMP were used to investigate the contamination of the samples with T. gondii, based on the 529 bp (RE) sequence of T. gondii. The analytical sensitivity of LAMP and real-time PCR was evaluated by duplicating the five-step serial dilutions of T. gondii tachyzoites from 0.25 to 5×105 spiked tachyzoites per milliliter of the Toxoplasma seronegative blood sample. The extracted DNA from other parasites and human chromosomal DNA were used to determine the specificity of the molecular methods. The obtained results were analyzed using Kappa statistical test and SPSS22 software. Out of 119 studied samples, 7 (5.8%) and 5 (4.2%) samples were positive for Toxoplasma by TaqMan probe-based real-time PCR and LAMP, respectively. The limits of detection of TaqMan probe-based real-time PCR and RE-LAMP in negative serum samples were one and five tachyzoites (CT 38), respectively. Both real-time PCR and LAMP methods were 100% specific for Toxoplasma detection. Positive results were obtained only with T. gondii DNA, while other DNA samples were negative. The TaqMan probe-based real-time PCR based on the RE sequence showed higher sensitivity to T. gondii DNA detection in blood samples of cancer patients and serial dilutions of parasitic tachyzoites. The results show that TaqMan probe-based real-time PRC is a sensitive and specific method for the detection of toxoplasmosis in immunocompromised individuals, as well as the LAMP assay, which can be used as a suitable alternative diagnostic method for the detection of toxoplasmosis in such patients, without need the for any expensive equipment.


Assuntos
DNA de Protozoário/genética , Técnicas de Amplificação de Ácido Nucleico , Infecções Oportunistas/diagnóstico , Parasitologia/métodos , Reação em Cadeia da Polimerase em Tempo Real , Toxoplasmose/diagnóstico , Animais , DNA de Protozoário/sangue , Humanos , Hospedeiro Imunocomprometido , Infecções Oportunistas/parasitologia , Sensibilidade e Especificidade , Toxoplasma/genética , Toxoplasmose/parasitologia
6.
J Parasitol ; 106(2): 312-315, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32330280

RESUMO

The purpose of this study was to describe the prevalence and incidence of Neospora caninum infection in dogs that are in close contact with dairy cattle and to identify possible risk factors associated with the infection in this population. Twenty-four dogs located in 8 different dairy farms of Aguascalientes, Mexico, were evaluated for a 6-mo period. Once a month a sample of serum and a sample of peripheral blood was collected. The serum was used to detect antibodies against N. caninum by means of the indirect immunofluorescence technique, and the blood was used to detect parasite's DNA. The association between seroprevalence and possible risk factors was estimated using logistic regression. The prevalence of anti-N. caninum antibodies was 54% in the first month, 62% in the last month, and the incidence was 8.69%. One farm had no positive cases. Antibody titers ranged from 1:50 to 1:800. Parasite DNA was not detected in any of the samples. Only the age (>6 yr) of the dogs was identified as a risk factor for infection by N. caninum (P ≤ 0.05).


Assuntos
Coccidiose/veterinária , Doenças do Cão/parasitologia , Neospora , Fatores Etários , Animais , Anticorpos Antiprotozoários/sangue , Bovinos , Coccidiose/epidemiologia , Coccidiose/parasitologia , DNA de Protozoário/sangue , DNA de Protozoário/isolamento & purificação , Indústria de Laticínios , Doenças do Cão/epidemiologia , Cães , Feminino , Incidência , Masculino , México/epidemiologia , Neospora/genética , Neospora/imunologia , Prevalência , Fatores de Risco
7.
PLoS Negl Trop Dis ; 14(4): e0008148, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32282820

RESUMO

BACKGROUND: Echinococcosis is a life-threatening parasitic disease caused by Echinococcus spp. tapeworms with over one million people affected globally at any time. The Echinococcus spp. tapeworms in the human body release DNA to the circulatory system, which can be a biomarker for echinococcosis. Cell-free DNA (cfDNA) is widely used in medical research and has been applied in various clinical settings. As for echinococcosis, several PCR-based tests had been trialed to detect cell-free Echinococcus spp. DNA in plasma or serum, but the sensitivity was about 20% to 25%. Low sensitivity of PCR-based methods might be related to our limited understanding of the features of cell-free Echinococcus spp. DNA in plasma, including its concentration, fragment pattern and release source. In this study, we applied ultra-high-throughput sequencing to comprehensively investigate the characteristics of cell-free Echinococcus spp. DNA in plasma of echinococcosis patients. METHODOLOGY/PRINCIPAL FINDINGS: We collected plasma samples from 23 echinococcosis patients. Total plasma cfDNA was extracted and sequenced with a high-throughput sequencing platform. An average of 282 million read pairs were obtained for each plasma sample. Sequencing data were analyzed with bioinformatics workflow combined with Echinococcus spp. sequence database. After identification of cell-free Echinococcus spp. reads, we found that the cell-free Echinococcus spp. reads accounted for 1.8e-5 to 4.0e-9 of the total clean reads. Comparing fragment length distribution of cfDNA between Echinococcus spp. and humans showed that cell-free Echinococcus spp. DNA of cystic echinococcosis (CE) had a broad length range, while that of alveolar echinococcosis (AE) had an obvious peak at about 135 bp. We found that most of the cell-free Echinococcus spp. DNA reads were from the nuclear genome with an even distribution, which might indicate a random release pattern of cell-free Echinococcus spp. DNA. CONCLUSIONS/SIGNIFICANCE: With ultra-high-throughput sequencing technology, we analyzed the concentration, fragment length, release source, and other characteristics of cell-free Echinococcus spp. DNA in the plasma of echinococcosis patients. A better understanding of the characteristics of cell-free Echinococcus spp. DNA in plasma may facilitate their future application as a biomarker for diagnosis.


Assuntos
DNA de Protozoário/sangue , Equinococose/diagnóstico , Equinococose/parasitologia , Echinococcus/genética , Echinococcus/isolamento & purificação , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Adolescente , Adulto , Animais , Sequência de Bases , Biomarcadores , Criança , DNA de Protozoário/isolamento & purificação , Feminino , Genoma de Protozoário , Humanos , Masculino , Pessoa de Meia-Idade , Plasma , Reação em Cadeia da Polimerase/métodos , Sensibilidade e Especificidade , Análise de Sequência de DNA , Adulto Jovem
8.
PLoS Negl Trop Dis ; 14(4): e0008253, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32324738

RESUMO

BACKGROUND: In the Mediterranean basin, Leishmania infantum is the causative agent of visceral leishmaniasis (VL), a zoonosis in which the dog is the primary domestic reservoir, although wildlife may have a leading role in the sylvatic cycle of the disease in some areas. Infections without disease are very frequent. There is limited information regarding the role that VL patients and asymptomatic infected individuals could be playing in the transmission of L. infantum. Xenodiagnosis of leishmaniasis has been used in this descriptive study to explore the role of symptomatic and asymptomatic infected individuals as reservoirs in a recent focus of leishmaniasis in southwestern Madrid, Spain. METHODOLOGY AND MAIN FINDINGS: Asymptomatic blood donors (n = 24), immunocompetent patients who were untreated (n = 12) or treated (n = 11) for visceral leishmaniasis (VL), and immunocompromised patients with VL (n = 3) were enrolled in the study. Their infectivity to Phlebotomus perniciosus was studied by indirect xenodiagnosis on peripheral blood samples. Quantitative polymerase chain reaction of blood samples from immunocompetent patients untreated for VL and immunocompromised untreated, treated and under secondary prophylaxis for VL was performed. Antibodies against Leishmania were studied by indirect fluorescent antibody and rK39-immunochromatographic tests. A lymphoproliferative assay with a soluble Leishmania antigen was used to screen for leishmaniasis infection in the healthy population. Sixty-two xenodiagnostic tests were carried out and 5,080 sand flies were dissected. Positive xenodiagnosis was recorded in four patients, with different sand fly infection rates: 1 immunosuppressed HIV / L. infantum coinfected asymptomatic patient, 1 immunosuppressed patient with multiple myeloma and symptomatic active VL, and 2 immunocompetent patients with untreated active VL. All blood donors were negative for both xenodiagnosis and conventional PCR. CONCLUSIONS / SIGNIFICANCE: There is no consensus amongst authors on the definition of an 'asymptomatic case' nor on the tools for screening; we, therefore, have adopted one for the sake of clarity. Immunocompetent subjects, both infected asymptomatics and those treated for VL, are limited in number and appear to have no epidemiological relevance. The impact is limited for immunocompetent patients with untreated active VL, whilst immunosuppressed individuals undergoing immunosuppressive therapy and immunosuppressed individuals HIV / L. infantum coinfected were the most infectious towards sand flies. It is noteworthy that the HIV / L. infantum coinfected patient with asymptomatic leishmaniasis was easily infectious to sand flies for a long time, despite being under continuous prophylaxis for leishmaniasis. Accordingly, screening for latent Leishmania infection in HIV-infected patients is recommended in scenarios where transmission occurs. In addition, screening for VL in HIV-infected patients who have spent time in VL-endemic areas should also be implemented in non-endemic areas. More research is needed to better understand if some asymptomatic coinfected individuals contribute to transmission as 'super-spreaders'.


Assuntos
Reservatórios de Doenças , Transmissão de Doença Infecciosa , Leishmania infantum/isolamento & purificação , Leishmaniose Visceral/epidemiologia , Leishmaniose Visceral/transmissão , Adulto , Animais , Anticorpos Antiprotozoários/sangue , Doenças Assintomáticas/epidemiologia , DNA de Protozoário/sangue , Feminino , Humanos , Leucócitos Mononucleares/imunologia , Masculino , Pessoa de Meia-Idade , Psychodidae/parasitologia , Espanha/epidemiologia
9.
Parasit Vectors ; 13(1): 115, 2020 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-32192533

RESUMO

BACKGROUND: An outbreak of leishmaniosis was studied in cats and dogs housed together with no separation in an animal shelter in Israel. METHODS: The study included recording of clinical signs, serology for Leishmania infection by ELISA, PCR of blood for Leishmania DNA by ITS1 HRM and kDNA PCR, parasite quantification, and trapping of sand flies around the shelter. RESULTS: Thirty-seven % (22/60) of the dogs and 75% (50/67) of the cats were seropositive to L. infantum with a significantly higher seropositivity rate in the cat population (χ2 = 42.160, P < 0.0001). Twenty-five percent (15/60) of the dogs were positive for Leishmania by blood PCR, 12% by the Leishmania ITS1 HRM PCR and 22% by kDNA PCR. Of the cats, 16% (11/67) were positive by kDNA PCR and none by ITS1 HRM PCR. All the PCR-positive animals were infected by L. infantum verified by DNA sequencing and there was no significant difference between the PCR-positivity in the dog and cat populations. Altogether, 43% (26/60) of the dogs and 79% (53/67) of the cats were positive by serology or PCR for L. infantum. The average Leishmania parasite load in the blood of PCR-positive dogs (42,967 parasites/ml) was significantly higher than in PCR-positive cats (1259 parasites/ml) (t(12) = 2.33, P = 0.037). Dogs that were positive by the Leishmania ITS1 HRM PCR and kDNA PCR had significantly higher parasite loads than dogs positive only by the kDNA PCR (t(11) = - 3.186580, P < 0.009). No significant effect was found for FIV seropositivity on Leishmania infection in the cats (χ2 = 0.506, P = 0.777). A higher percentage of Leishmania-positive dogs showed clinical signs compatible with leishmaniosis compared to Leishmania-positive cats (100 vs 52.8%, χ2 =15.242, P < 0.0001). Phlebotomus perfiliewi, a proven vector of L. infantum, comprised 92% of trapped sand flies. CONCLUSIONS: Comparisons of populations of cats and dogs exposed to sand flies and L. infantum under the same conditions indicated that although a high rate of exposure was detected in cats as manifested by a significantly greater degree of seropositivity, dogs had significantly higher blood parasite loads, and were likely to be more infectious to sand flies than cats.


Assuntos
Doenças do Gato/parasitologia , Surtos de Doenças/veterinária , Doenças do Cão/parasitologia , Leishmaniose/veterinária , Animais , Anticorpos Antiprotozoários/sangue , Doenças do Gato/epidemiologia , Gatos , DNA de Protozoário/sangue , Doenças do Cão/epidemiologia , Cães , Feminino , Israel/epidemiologia , Leishmania infantum , Leishmaniose/epidemiologia , Masculino , Carga Parasitária , Psychodidae/parasitologia , Estudos Soroepidemiológicos
10.
Comp Immunol Microbiol Infect Dis ; 70: 101453, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32163745

RESUMO

This study was undertaken to assess the effects of T. equi infection on serum concentrations of some important cytokines including interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), and interleukin-1 beta (IL-1ß), IL-1α, IL-4, IL-6, IL-8, IL-10, IL-12α, IL-12ß, IL-18, as well as total, protein and lipid binding sialic acids (TSA, PBSA and LBSA). Furthermore, any probable relation among the parasitemia, cytokines and sialic acids (SAs) were calculated using Pearson correlation and simple linear regression. Almost 300 draft horses (Kurdish-breed) with age of 3-4 years old from north-west of Iran were examined and an infected group comprised of 28 mares, naturally infected with T. equi, was identified and divided into 3 subgroups according to their parasitemia rates (low <1 %, moderate 1-3 % and high 3-5 %). Twenty healthy horses were considered as a control. Characterization and differentiation of piroplasmosis were conducted using routine hematological procedures and specific PCR assay. The results revealed a significant increase (P < 0.05) in all of the cytokines and SAs in a parasitic burden-dependent fashion. Additionally, a strong and positive relation was detected among the parasitemia, cytokines and SAs. Conclusively, T. equi infection is associated with induction of severe inflammatory processes in horses and SA plays a pivotal role in pathophysiology of the disease as it is tightly correlated with the parasitemia rate.


Assuntos
Citocinas/sangue , Doenças dos Cavalos/imunologia , Inflamação , Parasitemia/imunologia , Ácidos Siálicos/sangue , Theileriose/imunologia , Animais , Citocinas/imunologia , DNA de Protozoário/sangue , Feminino , Doenças dos Cavalos/parasitologia , Cavalos , Theileria
11.
J Parasitol ; 106(2): 211-220, 2020 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-32164026

RESUMO

Biogeography is known to have shaped the diversity and evolutionary history of avian haemosporidian parasites across the Neotropics. However, a paucity of information exists for the temperate Neotropics and especially from nonpasserine hosts. To understand the effect of biogeography in the temperate Neotropics on haemosporidians of nonpasserine hosts we screened ducks (Anseriformes) from central Chile for the presence of these parasites. Forty-two individuals of 4 duck species (Anas flavirostris, Anas georgica, Mareca sibilatrix, Spatula cyanoptera cyanoptera) were collected and assessed for haemosporidian parasite infections by real-time polymerase chain reaction screening and subsequent sequencing of the mitochondrial cytochrome b gene. Haemoproteus (subgenus Haemoproteus) and Plasmodium were detected in 2 host species, A. georgica and S. c. cyanoptera, with no Leucocytozoon found. Overall haemosporidian prevalence was low (14.2%), with the prevalence of Plasmodium (11.9%) being substantially greater than that of Haemoproteus (4.8%). Six haemosporidian cytochrome b lineages were recovered, 2 Haemoproteus and 4 Plasmodium, with all 6 lineages identified for the first time. In phylogenetic reconstruction, the Chilean Plasmodium lineages were more closely related to South American lineages from passerine birds than to known lineages from anseriforms. The subgenus Haemoproteus known from nonpasseriformes has never been identified from any anseriform host; however, we recovered 2 lineages from this subgenus, one from each A. georgica and S. c. cyanoptera. Further work is needed to determine if this presents true parasitism in ducks or only a spillover infection. The results of phylogenetic reconstruction demonstrate a unique evolutionary history of these Chilean parasites, differing from what is known for this host group. The unique geography of Chile, with a large part of the country being relatively isolated by the Atacama Desert in the north and the Andes in the east and south, would present opportunities for parasite diversification. Further work is needed to investigate how strongly the biogeographical isolation has shaped the haemosporidian parasites of this area. Our results add to the growing body of evidence that nonpasserine hosts support unique lineages of haemosporidian parasites, while also demonstrating the role of biogeography in haemosporidian parasite diversity in the temperate Neotropics.


Assuntos
Doenças das Aves/epidemiologia , Doenças das Aves/parasitologia , Patos/parasitologia , Haemosporida/isolamento & purificação , Infecções Protozoárias em Animais/epidemiologia , Animais , Teorema de Bayes , Evolução Biológica , Distribuição de Qui-Quadrado , Chile/epidemiologia , DNA de Protozoário/análise , DNA de Protozoário/sangue , DNA de Protozoário/isolamento & purificação , Haemosporida/classificação , Haemosporida/genética , Funções Verossimilhança , Fígado/parasitologia , Filogenia , Filogeografia , Reação em Cadeia da Polimerase/veterinária , Infecções Protozoárias em Animais/parasitologia
12.
Malar J ; 19(1): 28, 2020 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-31948448

RESUMO

BACKGROUND: The sensitivity of rapid diagnostic tests (RDTs) for malaria is inadequate for detecting low-density, often asymptomatic infections, such as those that can occur when screening pregnant women for malaria. The performance of the Alere™ Ultra-sensitive Malaria Ag Plasmodium falciparum RDT (uRDT) was assessed retrospectively in pregnant women in Indonesia. METHODS: The diagnostic performance of the uRDT and the CareStart™ Malaria HRP2/pLDH VOM (Plasmodium vivax, Plasmodium ovale and Plasmodium malariae) Combo RDT (csRDT) were assessed using 270 stored red blood cell pellets and plasma samples from asymptomatic pregnant women. These included 112 P. falciparum negative and 158 P. falciparum positive samples detected by a composite test (qPCR, LAMP, nPCR) as reference standard. Diagnostic indicators: sensitivity, specificity, negative predictive value (NPV), positive predictive value (PPV), diagnostic odds ratio (DOR) and the level of agreement (kappa) were calculated for comparison. RESULTS: Compared with the reference test, the uRDT had a sensitivity of 19.6% (95% CI 13.9-26.8) and specificity of 98.2% (93.1-99.7%). The csRDT was 22.8% (16.7-30.3) sensitive and 95.5% (89.4-98.3) specific for P. falciparum infections. Performance of the uRDT was non-significantly different to the csRDT (p = 0.169). RDT outcome was stratified by qPCR cycling threshold (Ct), and performance of the RDTs was found to be comparable across parasite loads. CONCLUSION: The uRDT performed similarly to the currently used csRDTs in detecting P. falciparum infections in asymptomatic pregnant women. In these settings, molecular diagnostics are currently the most sensitive for malaria.


Assuntos
Programas de Triagem Diagnóstica/normas , Malária Falciparum/diagnóstico , Complicações Parasitárias na Gravidez/diagnóstico , Coinfecção/diagnóstico , DNA de Protozoário/análise , DNA de Protozoário/sangue , DNA de Protozoário/isolamento & purificação , Eritrócitos/parasitologia , Feminino , Humanos , Indonésia , Razão de Chances , Plasmodium/genética , Plasmodium/imunologia , Plasmodium/isolamento & purificação , Valor Preditivo dos Testes , Gravidez , Estudos Retrospectivos , Sensibilidade e Especificidade
13.
Malar J ; 19(1): 20, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31941493

RESUMO

BACKGROUND: Zambia has set itself the ambitious target of eliminating malaria by 2021. To continue tracking transmission to zero, new interventions, tools and approaches are required. METHODS: Urban reactive case detection (RCD) was performed in Lusaka city from 2011 to 2015 to better understand the location and drivers of malaria transmission. Briefly, index cases were followed to their home and all consenting individuals living in the index house and nine proximal houses were tested with a malaria rapid diagnostic test and treated if positive. A brief survey was performed and for certain responses, a dried blood spot sample collected for genetic analysis. Aggregate health facility data, individual RCD response data and genetic results were analysed spatially and against environmental correlates. RESULTS: Total number of malaria cases remained relatively constant, while the average age of incident cases and the proportion of incident cases reporting recent travel both increased. The estimated R0 in Lusaka was < 1 throughout the study period. RCD responses performed within 250 m of uninhabited/vacant land were associated with a higher probability of identifying additional infections. CONCLUSIONS: Evidence suggests that the majority of malaria infections are imported from outside Lusaka. However there remains some level of local transmission occurring on the periphery of urban settlements, namely in the wet season. Unfortunately, due to the higher-than-expected complexity of infections and the small number of samples tested, genetic analysis was unable to identify any meaningful trends in the data.


Assuntos
Malária Falciparum/epidemiologia , Malária Falciparum/transmissão , Adolescente , Adulto , Fatores Etários , Animais , Criança , Pré-Escolar , DNA de Protozoário/sangue , Feminino , Humanos , Incidência , Malária Falciparum/diagnóstico , Masculino , Plasmodium falciparum/genética , Plasmodium falciparum/isolamento & purificação , Análise de Regressão , População Rural , Estações do Ano , Análise Espacial , Viagem , Saúde da População Urbana , Adulto Jovem , Zâmbia/epidemiologia
14.
J Infect Dis ; 221(5): 786-795, 2020 02 18.
Artigo em Inglês | MEDLINE | ID: mdl-31630194

RESUMO

Accurate malaria diagnosis is foundational for control and elimination, and Haiti relies on histidine-rich protein 2 (HRP2)-based rapid diagnostic tests (RDTs) identifying Plasmodium falciparum in clinical and community settings. In 2017, 1 household and 2 easy-access group surveys tested all participants (N = 32 506) by conventional and high-sensitivity RDTs. A subset of blood samples (n = 1154) was laboratory tested for HRP2 by bead-based immunoassay and for P. falciparum 18S rDNA by photo-induced electron transfer polymerase chain reaction. Both RDT types detected low concentrations of HRP2 with sensitivity estimates between 2.6 ng/mL and 14.6 ng/mL. Compared to the predicate HRP2 laboratory assay, RDT sensitivity ranged from 86.3% to 96.0% between tests and settings, and specificity from 90.0% to 99.6%. In the household survey, the high-sensitivity RDT provided a significantly higher number of positive tests, but this represented a very small proportion (<0.2%) of all participants. These data show that a high-sensitivity RDT may have limited utility in a malaria elimination setting like Haiti.


Assuntos
Testes Diagnósticos de Rotina/métodos , Malária Falciparum/diagnóstico , Malária Falciparum/transmissão , Plasmodium falciparum/genética , Plasmodium falciparum/imunologia , Adolescente , Antígenos de Protozoários/sangue , Antígenos de Protozoários/imunologia , Criança , Pré-Escolar , DNA de Protozoário/sangue , DNA de Protozoário/genética , DNA Ribossômico/sangue , DNA Ribossômico/genética , Ensaio de Imunoadsorção Enzimática/métodos , Feminino , Haiti/epidemiologia , Humanos , Lactente , Malária Falciparum/epidemiologia , Malária Falciparum/parasitologia , Masculino , Reação em Cadeia da Polimerase/métodos , Proteínas de Protozoários/sangue , Proteínas de Protozoários/imunologia , Sensibilidade e Especificidade
15.
BMC Vet Res ; 15(1): 446, 2019 Dec 09.
Artigo em Inglês | MEDLINE | ID: mdl-31818287

RESUMO

BACKGROUND: Leishmaniosis, a disease caused by pathogenic Leishmania parasites, remains an unresolved health problem in the New World and the Old World. It is well known that lizards can be infected by a subgenus of Leishmania parasites, i.e. Sauroleishmania, which is non-pathogenic to humans. However, evidence suggests that lizards may also harbor pathogenic Leishmania species including the undetermined Leishmania sp., discovered in our previous work. Leishmania DNA in lizard blood can be detected by using molecular methods, such as the polymerase chain reaction (PCR). RESULTS: Three hundred and sixteen lizards, representing 13 species of four genera, were captured for blood samples collection in Northwest China. Two reliable molecular markers (cytochrome b and heat shock protein 70 genes) were used for detection in the lizard blood samples, to confirm a widespread presence of pathogenic Leishmania parasites and the distribution pattern of Leishmania spp. in lizards from Northwest China. The PCR data indicated positive detection rate for Leishmania in all the tested lizards with an overall prevalence of 57.91% (183/316). Apart from lizard parasites like Leishmania tarentolae and Leishmania sp., several pathogenic Leishmania including L. turanica, L. tropica and L. donovani complex were identified by using phylogenetic analysis. Co-existence of different haplotypes was observed in most Leishmania DNA-positive lizards with an overall rate of 77.6% (142/183). Even mixed infections with different Leishmania species appeared to occur in the lizards with an overall rate of 37.7% (69/183). CONCLUSIONS: Lizards can harbor pathogenic Leishmania spp. Co-existence of different haplotypes or even species of Leishmania indicates mixed infections in natural lizard host. Lizards may contribute to the spread of Leishmania parasites. The pathogenic Leishmania species detected in lizards from Northwest China may be of great eco-epidemiological importance.


Assuntos
Leishmania/classificação , Leishmaniose/epidemiologia , Lagartos/parasitologia , Animais , China/epidemiologia , DNA de Protozoário/sangue , Haplótipos , Leishmania/genética , Lagartos/sangue , Filogenia , Reação em Cadeia da Polimerase , Análise de Sequência de DNA
16.
Ann Agric Environ Med ; 26(4): 538-543, 2019 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-31885225

RESUMO

INTRODUCTION AND OBJECTIVE: It is easier and non-invasive to obtain faecal samples compared with blood samples. Molecular techniques may enable detection of parasites even in tiny amounts of blood-containing faeces. We aimed to compare the sensitivity of detection of three Babesia species and Hepatozoon canis in blood and faecal samples, including samples derived from naturally infected hosts. MATERIAL AND METHODS: Three groups were involved: 1) Nine BALB/c mice infected with Babesia microti sampled during acute (n=3), post-acute (n=3) and chronic phases of infection (n=3); 2) Eight dogs with symptoms of babesiosis; 3) Six red foxes infected with B. vulpes, one fox infected with B. canis, four foxes infected with H. canis. Genomic DNA was extracted from blood and faeces by use of commercial kits and amplified with genus-specific primers in one-step or nested PCR reactions. Selected PCR products were sequenced. RESULTS: No positive results for faecal samples were obtained from H. canis-positive foxes in contrast to Babesia spp. infections. Positive results from PCRs were obtained for all BALB/c mice (100%), five dogs (62.5%) and four of seven foxes (57.1%). Successful sequencing was obtained for six selected murine samples (B. microti), four canine samples (B. canis) and for one fox sample (B. vulpes). The success of B. microti detection in murine faecal samples from acute, post-acute and chronic phases was identical (100%). CONCLUSIONS: Detectability of Babesia spp. infections was lower in naturally infected dogs and foxes, compared to experimentally infected mice. Detection of DNA in faecal samples can be useful in the detection of Babesia infection in populations from which blood samples are hard to obtain, but due regard must be given to the possibility that prevalence of infection may be severely underestimated.


Assuntos
Babesia/isolamento & purificação , Babesiose/parasitologia , Sangue/parasitologia , Coccídios/isolamento & purificação , Coccidiose/parasitologia , DNA de Protozoário/genética , Fezes/parasitologia , Animais , Babesia/genética , Coccídios/genética , DNA de Protozoário/sangue , Cães/parasitologia , Raposas/parasitologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Reação em Cadeia da Polimerase
17.
Rev. bras. parasitol. vet ; 28(4): 685-691, Oct.-Dec. 2019. tab
Artigo em Inglês | LILACS | ID: biblio-1057966

RESUMO

Abstract Equine piroplasmosis, an economically important disease in horses, has so far not been reported in Pernambuco state, Brazil. This study aimed to evaluate the seroprevalence of anti-Babesia caballi and anti-Theileria equi antibodies based on the detection of these agents in equine blood and in ticks on horses in the municipality of Petrolina, Pernambuco, northeastern Brazil. Blood samples were drawn from 393 horses and sera were examined by ELISA. The presence of tick infestations was evaluated, and 101 ticks were subjected to DNA amplification for the detection of Babesia spp. by polymerase chain reaction (PCR). No parasites were detected in the blood smears. Anti-B. caballi and anti-T. equi antibodies were found in 27.2% (107/393) and 34.8% (137/393) horses, respectively. Infestation by Dermacentor nitens was detected in 4.3% (17/393) of the horses. There was no DNA amplification of the agents in ticks. The risk factors for the presence of anti-T. equi antibodies (P < 0.05) were: purebred (P < 0.001), animals older than 156 months (P = 0.014), and the presence of ticks (P = 0.001). No risk factors for B. caballi were identified. This study confirmed the circulation of agents of equine piroplasmosis in the municipality of Petrolina, state of Pernambuco, Brazil.


Resumo Piroplasmose equina é uma doença economicamente importante em equinos e não possui relatos no Estado de Pernambuco, Brasil. O objetivo deste estudo foi avaliar a soroprevalência de anticorpos anti-B. caballi e anti-T. equi pela detecção destes agentes no sangue e carrapatos de equinos no município de Petrolina, Pernambuco, Nordeste do Brasil. Amostras de sangue de 393 equinos foram coletadas e submetidas ao esfregaço sanguíneo e ELISA. A presença de infestação por carrapatos foi avaliada, e 71 carrapatos foram submetidos à Reação em Cadeia da Polimerase (PCR) para Babesia spp. Nenhum parasito foi detectado na análise de esfregaços de sangue. Anticorpos anti-B. caballi e anti-T. equi foram verificados em 27,2% (107/393) e 34,8% (137/393) dos equinos, respectivamente. A infestação por Dermacentor nitens foi verificada em 4,3% (17/393) dos equinos. Não houve amplificação do DNA dos agentes nos 71 carrapatos submetidos à PCR. Os fatores de risco para presença de anticorpos anti-T. equi (P < 0,05) foram: raça definida (P < 0,001), animais > de 156 meses (P = 0,014) e presença de carrapatos (P = 0,001). Nenhum fator de risco foi identificado para B. caballi. Esse estudo permitiu a confirmação da presença de agentes da piroplasmose equina no município de Petrolina, Pernambuco.


Assuntos
Animais , Masculino , Feminino , Babesiose/epidemiologia , Carrapatos/microbiologia , Doenças dos Cavalos/epidemiologia , Filogenia , Babesia/genética , Babesia/imunologia , Babesiose/diagnóstico , Brasil/epidemiologia , Ensaio de Imunoadsorção Enzimática/veterinária , Estudos Soroepidemiológicos , Reação em Cadeia da Polimerase/veterinária , Prevalência , Fatores de Risco , DNA de Protozoário/sangue , Doenças dos Cavalos/diagnóstico , Doenças dos Cavalos/parasitologia , Cavalos
18.
Rev Bras Parasitol Vet ; 28(4): 685-691, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31691736

RESUMO

Equine piroplasmosis, an economically important disease in horses, has so far not been reported in Pernambuco state, Brazil. This study aimed to evaluate the seroprevalence of anti-Babesia caballi and anti-Theileria equi antibodies based on the detection of these agents in equine blood and in ticks on horses in the municipality of Petrolina, Pernambuco, northeastern Brazil. Blood samples were drawn from 393 horses and sera were examined by ELISA. The presence of tick infestations was evaluated, and 101 ticks were subjected to DNA amplification for the detection of Babesia spp. by polymerase chain reaction (PCR). No parasites were detected in the blood smears. Anti-B. caballi and anti-T. equi antibodies were found in 27.2% (107/393) and 34.8% (137/393) horses, respectively. Infestation by Dermacentor nitens was detected in 4.3% (17/393) of the horses. There was no DNA amplification of the agents in ticks. The risk factors for the presence of anti-T. equi antibodies (P < 0.05) were: purebred (P < 0.001), animals older than 156 months (P = 0.014), and the presence of ticks (P = 0.001). No risk factors for B. caballi were identified. This study confirmed the circulation of agents of equine piroplasmosis in the municipality of Petrolina, state of Pernambuco, Brazil.


Assuntos
Babesiose/epidemiologia , Doenças dos Cavalos/epidemiologia , Carrapatos/microbiologia , Animais , Babesia/genética , Babesia/imunologia , Babesiose/diagnóstico , Brasil/epidemiologia , DNA de Protozoário/sangue , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Doenças dos Cavalos/diagnóstico , Doenças dos Cavalos/parasitologia , Cavalos , Masculino , Filogenia , Reação em Cadeia da Polimerase/veterinária , Prevalência , Fatores de Risco , Estudos Soroepidemiológicos
19.
Vet Parasitol ; 276: 108977, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31775104

RESUMO

The aims of this study were to monitor the change in Theileria orientalis Ikeda type infection intensity, haematocrit, milk production and reproduction on three New Zealand spring calving dairy herds, over the 2014-2015 milking season. Three spring calving dairy farms, A, B and C, from high risk (endemically stable), low risk (endemically unstable), and zero risk (disease-free) tick areas respectively were followed through the 2014-2015 milking season. On Farms, A and B, 100 cows were randomly selected at the first visit, and the same cows blood sampled every month thereafter, whilst on Farm C, the whole herd was blood sampled bimonthly (140 cows). Blood samples were tested for haematocrit, by centrifugation, and Ikeda infection intensity, using qPCR. Animals that were Ikeda type PCR positive at the first sampling were described as prevalence cases and cows that were negative at the first sampling and became PCR positive during the sampling period were described as incidence cases. Production and reproduction data were accessed through LIC MINDA® and milk production data was standardised to energy corrected milk (ECM). In addition, the effect of buparvaquone (BPQ) treatment on milk production was estimated on Farm B. The prevalence of infection at the first sampling was 100 % on Farm A, 57 % on Farm B and 26 % on Farm C. The incidence risk of infection over the sampling period on Farms B and C was 25 % and 2 % and the incident rate was 0.026 and 0.002 cases per cow-month respectively. The average infection intensity for prevalence cases on all farms was low throughout the milking season, <7000 Ikeda organisms/µL however, cases of anaemia still occurred. There was no direct effect of infection intensity on milk production or from being a prevalence case compared to an uninfected cow on milk production, across all farms. However, on Farm B there was a loss of 266 kg (95 % CI 82 ̶ 450) ECM (∼20 kg milk solids) for incidence cases and a loss of 458 kg (95 % CI 211 ̶ 710) of ECM for buparvaquone treated cows, compared to uninfected cows. No significant effect of Ikeda infection on reproduction could be shown for Farms B and C, reproductive data for Farm A was not available. The effect of T. orientalis Ikeda type infection on production and reproduction appears to be minimal once animals have passed through the acute phase of infection and reached the chronic, asymptomatic carrier phase of infection.


Assuntos
Lactação , Reprodução , Theileriose/fisiopatologia , Animais , Antiprotozoários/uso terapêutico , Bovinos , DNA de Protozoário/sangue , Indústria de Laticínios , Feminino , Dosagem de Genes , Hematócrito/veterinária , Incidência , Estudos Longitudinais , Naftoquinonas/uso terapêutico , Prevalência , Estudos Prospectivos , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Theileria/efeitos dos fármacos , Theileria/genética , Theileriose/tratamento farmacológico , Theileriose/epidemiologia , Theileriose/parasitologia
20.
Biomedica ; 39(Supl. 2): 144-156, 2019 08 01.
Artigo em Inglês, Espanhol | MEDLINE | ID: mdl-31529841

RESUMO

Introduction: Toxoplasma gondii infection manifests differently in humans according to their immunity ranging from asymptomatic profiles to severe disease. There are multiple transmission mechanisms including blood transfusions, but little is known about the frequency of T. gondii infection in Colombia's blood banks. Objective: To determine the prevalence of T. gondii infection in blood donors of a blood bank in the city of Cúcuta by serological and molecular diagnostic techniques. Materials and methods: We identified IgG and IgM antibodies against T. gondii by immunoassay in serum from 348 donors. The frequency of T. gondii DNA was determined by polymerase chain reaction (PCR) in whole blood from seropositive donors and relevant variables were analyzed based on the information obtained from surveys during blood donor selection. Results: Out of the 348 enrolled donors, 134 (38.5%) showed IgG antibodies against T. gondii; two of them (0.6%) had both IgG and IgM, and in two of them (1.5%), parasite DNA was detected in blood samples. A bivariate analysis indicated an association between seropositivity to T. gondii and being over 26 years of age (p=0.020). Conclusions: The prevalence of T. gondii infection found in the blood donors of this study suggests a significant exposure to the infectious agent that becomes relevant when parasitemia is detected.


Assuntos
Anticorpos Antiprotozoários/sangue , Bancos de Sangue , Doadores de Sangue , DNA de Protozoário/sangue , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Parasitemia/epidemiologia , Toxoplasma/isolamento & purificação , Toxoplasmose/epidemiologia , Adolescente , Adulto , Idoso , Doadores de Sangue/estatística & dados numéricos , Colômbia/epidemiologia , Estudos Transversais , Feminino , Hospitais Universitários , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase em Tempo Real , Estudos Soroepidemiológicos , Fatores Socioeconômicos , Toxoplasma/genética , Toxoplasma/imunologia , Adulto Jovem
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