Saturday, October 10, 2009

What's new for 'Trypanosomatids' in PubMed

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Search kinetoplastids OR kinetoplastid OR Kinetoplastida OR "trypanosoma brucei" OR leishmania OR brucei OR leishmaniasis OR "African trypanosomiasis"
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PubMed Results
Items 1 -10 of 13

1: Am J Trop Med Hyg. 2009 Oct;81(4):665-74.

Development of an Alamar Blue viability assay in 384-well format for high throughput whole cell screening of Trypanosoma brucei brucei bloodstream form strain 427.

Sykes ML, Avery VM.

Eskitis Institute for Cell and Molecular Therapies, Griffith University, Nathan, Queensland, Australia. m.sykes@griffith.edu.au

There is an urgent need for new compounds for the drug development pipeline for treatment of patients with African sleeping sickness. One approach for identifying such compounds is by high throughput screening (HTS) of compound collections. For time and cost considerations, there is a need for the development of an assay that uses at least 384-well formats. To our knowledge, there are currently no viability assays for whole cell screening of trypanosomes in the 384-well plate format. We have developed and optimized an Alamar Blue viability assay in a 384-well format for Trypanosoma brucei brucei bloodstream form strain 427 (BS427). The assay had a Z' > 0.5 and tolerated a final dimethyl-sulfoxide concentration of 0.42%. Drug sensitivity was compared with those reported from previously developed 96-well methods and was found to be comparable. The sensitivity and cost benefit of the Alamar Blue assay make it an excellent candidate for HTS application.

PMID: 19815884 [PubMed - in process]

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2: Am J Trop Med Hyg. 2009 Oct;81(4):578-82.

Sensitive, specific, and rapid detection of Leishmania donovani DNA by loop-mediated isothermal amplification.

Takagi H, Itoh M, Islam MZ, Razzaque A, Ekram AR, Hashighuchi Y, Noiri E, Kimura E.

Department of Parasitology, Aichi Medical University School of Medicine, Nagakute, Aichi, Japan.

We have applied a loop-mediated isothermal amplification (LAMP) technique to detect Leishmania donovani DNA. The LAMP technique detected 1 fg of L. donovani DNA, which was 10-fold more sensitive than a conventional polymerase chain reaction (PCR). All nested PCR-positive blood samples from visceral leishmaniasis patients were positive with the LAMP technique, and DNA samples from L. infantum, L. major, L. mexicana, L. tropica, L. braziliensis, Plasmodium falciparum, and healthy humans were negative with the LAMP technique. The advantages of the LAMP method are its shorter reaction time, a lack of requirement of sophisticated equipment, and visual judgment of positivity based on the turbidity of reaction mixture. Our LAMP technique can be a better alternative to a conventional PCR, especially under field conditions.

PMID: 19815869 [PubMed - in process]

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3: Am J Trop Med Hyg. 2009 Oct;81(4):572-7.

Vector incrimination of sand flies in the most important visceral leishmaniasis focus in Iran.

Oshaghi MA, Ravasan NM, Javadian EA, Mohebali M, Hajjaran H, Zare Z, Mohtarami F, Rassi Y.

Department of Medical Entomology and Vector Control, School of Public Health and Institute of Health Research, Tehran University of Medical Sciences, Tehran, Iran. oshaghima@yahoo.com

The prevalence, host preference, and rate of Leishmania spp. infection of sand fly species are important parameters for incrimination of parasite vectors. We applied polymerase chain reaction (PCR)-based and enzyme-linked immunosorbent assay (ELISA) methods to detect Leishmania spp. parasites and blood meals within individual sand flies in the most important visceral leishmaniasis (VL) focus in northwestern Iran. Leishmania spp. minicircles (kinetoplast DNA) were found in 14 (0.9%) of 1,569 female specimens. Sequence analysis of 650 basepairs of an internal transcribed spacer ribosomal DNA gene identified L. infantum/L. donovani in 12 specimens and L. adleri-like parasites in 2 specimens. Nine (64.3%) of 14 of the Leishmania spp.-positive sand flies were Phlebotomus perfeliewi transcaucasicus. Blood meal identification of host DNA within sand flies by PCR-based and ELISA methods showed that 30% and 28%, respectively, were positive for human blood. Results of this study showed that P. perfeliewi transcaucasicus is the most prevalent, infected, and anthropophagic sand fly and plays a major role in VL transmission in the region studied.

PMID: 19815868 [PubMed - in process]

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4: Am J Trop Med Hyg. 2009 Oct;81(4):565-71.

Molecular epidemiology of American tegumentary leishmaniasis in Panama.

Miranda A, Carrasco R, Paz H, Pascale JM, Samudio F, Saldaña A, Santamaría G, Mendoza Y, Calzada JE.

Instituto Conmemorativo Gorgas de Estudios de la Salud, Panama City, Panama.

American tegumentary leishmaniasis is an increasing public health problem in Panama. This study describes the clinical characteristics and the molecular epidemiology of leishmaniasis in Panama over a 5-year period (2004-2008). Additionally, we applied a polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP)-based assay to identify Leishmania species in clinical isolates, skin scrapings, and sandflies specimens. Whereas 60.3% of cases were detected with conventional parasitologic techniques (smear or in vitro culture), the PCR detected 72% positive patients. Our clinical-epidemiologic data corroborate the high incidence of L. (Viannia) panamensis and provide evidence of peridomestic and/or domestic transmission. Mucosal involvement was observed in 4.2% of the patients. The overall natural infection rate with Leishmania in 103 pools of sandflies was 0.46%. Lutzomyia gomezi and Lutzomya panamensis were the prevalent species incriminated as vectors at the capture sites in central Panama. This study contributes to a better knowledge of the current epidemiology of tegumentary leishmaniasis in Panama.

PMID: 19815867 [PubMed - in process]

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5: Am J Trop Med Hyg. 2009 Oct;81(4):559-64.

Tegumentary leishmaniasis as the cause of immune reconstitution inflammatory syndrome in a patient co-infected with human immunodeficiency virus and Leishmania guyanensis.

Chrusciak-Talhari A, Ribeiro-Rodrigues R, Talhari C, Silva RM Jr, Ferreira LC, Botileiro SF, Santos LO, Dietze R, Talhari S.

Fundação de Medicina Tropical do Amazonas, Manaus, Amazonas, Brazil.

We report a case of immune reconstitution inflammatory syndrome (IRIS) in a 32-year-old man infected with human immunodeficiency virus and Leishmania guyanensis. Three months after initiation of highly active anti-retroviral therapy (HAART), the patient had disseminated cutaneous leishmaniasis and started anti-leishmanial therapy. The patient's leishmaniasis manifestations during HAART ranged form an anergic response (46 CD4+ T cells/microL) to a disseminated cutaneous leishmaniasis (112 CD4+ T cells/microL). Eight weeks later (168 CD4+ T cells/microL, skin biopsy specimens showed inflammatory infiltrates with no detectable amastigotes. The patient then became comatose. Prednisone therapy (60 mg/day) was initiated with a significant improvement within 48 hours. Three months later (CD4+ T cell count = 184 cell/microL), localized, classic, cutaneous leishmaniasis developed in the patient and anti-leishmanial treatment was re-introduced. On that occasion, frequency of T regulatory cells was 1.82% of all CD4+ cells. Our data suggest a pivotal role for CD4+ T cells in the onset of IRIS and lesion ulceration and their association with a low frequency of T regulatory cells.

PMID: 19815866 [PubMed - in process]

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6: Am J Trop Med Hyg. 2009 Oct;81(4):555-8.

Co-Infection of Leishmania (Viannia) braziliensis and HIV: report of a case of mucosal leishmaniasis in Cochabamba, Bolivia.

Torrico F, Parrado R, Castro R, Marquez CJ, Torrico MC, Solano M, Reithinger R, García AL.

Facultad de Medicina, Universidad Mayor de San Simón, Cochabamba, Bolivia.

We describe the first case of Leishmania/HIV co-infection reported in Bolivia. Initially hospitalized with a diagnosis of pneumonia and bronchitis, the patient had numerous cutaneous and mucosal lesions caused by Leishmania (Viannia) braziliensis. The patient was also diagnosed as severely immunocompromised because of HIV infection.

PMID: 19815865 [PubMed - in process]

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7: Proc Natl Acad Sci U S A. 2009 Oct 6. [Epub ahead of print]

Distinct and overlapping roles for two Dicer-like proteins in the RNA interference pathways of the ancient eukaryote Trypanosoma brucei.

Patrick KL, Shi H, Kolev NG, Ersfeld K, Tschudi C, Ullu E.

Departments of Epidemiology and Public Health, Internal Medicine, and Cell Biology, Yale University Medical School, 295 Congress Avenue, New Haven, CT 06536-0812.

Trypanosoma brucei is one of the most ancient eukaryotes where RNA interference (RNAi) is operational and is the only single-cell pathogen where RNAi has been extensively studied and used as a tool for functional analyses. Here, we report that the T. brucei RNAi pathway, although relying on a single Argonaute protein (AGO1), is initiated by the activities of two distinct Dicer-like enzymes. Both TbDCL1, a mostly cytoplasmic protein, and the previously undescribed nuclear enzyme TbDCL2 contribute to the biogenesis of siRNAs from retroposons. However, TbDCL2 has a predominant role in generating siRNAs from chromosomal internal repeat transcripts that accumulate at the nucleolus in RNAi-deficient cells and in initiating the endogenous RNAi response against retroposons and repeats alike. Moreover, siRNAs generated by both TbDCL1 and TbDCL2 carry a 5'-monophosphate and a blocked 3' terminus, suggesting that 3' end modification is an ancient trait of siRNAs. We thus propose a model whereby TbDCL2 fuels the T. brucei nuclear RNAi pathway and TbDCL1 patrols the cytoplasm, posttranscriptionally silencing potentially harmful nucleic acid parasites that may access the cytoplasm. Nevertheless, we also provide evidence for cross-talk between the two Dicer-like enzymes, because TbDCL2 is implicated in the generation of 35- to 65-nucleotide intermediate transcripts that appear to be substrates for TbDCL1. Our finding that dcl2KO cells are more sensitive to RNAi triggers than wild-type cells has significant implications for reverse genetic analyses in this important human pathogen.

PMID: 19815526 [PubMed - as supplied by publisher]

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8: Protein Expr Purif. 2009 Oct 5. [Epub ahead of print]

Overproduction, purification and characterisation of Tbj1, a novel Type III Hsp40 from Trypanosoma brucei, the African Sleeping Sickness parasite.

Louw CA, Ludewig MH, Blatch GL.

Biomedical Biotechnology Research Unit, Department of Biochemistry, Microbiology and Biotechnology, Rhodes University, Grahamstown 6140, South Africa.

The heat shock protein 40 (Hsp40) family of proteins act as co-chaperones of the heat shock protein 70 (Hsp70) chaperone family, and together they play a vital role in the maintenance of cellular homeostasis. The Type III class of Hsp40s are diverse in terms of both sequence identity and function and have not been extensively characterised. The Trypanosoma brucei parasite is the causative agent of Human African Trypanosomiasis, and possesses an unusually large Hsp40 complement, consisting mostly of Type III Hsp40s. A novel T. brucei Type III Hsp40, Tbj1, was heterologously expressed, purified, and found to exist as a compact monomer in solution. Using polyclonal antibodies to the full-length recombinant protein, Tbj1 was found by Western analysis to be expressed in the T. brucei bloodstream form. Tbj1 was found to be able to assist two different Hsp70 proteins in the suppression of protein aggregation in vitro, despite being unable to stimulate their ATPase activity. This indicated that while Tbj1 did not possess independent chaperone activity, it potentially functioned as a novel co-chaperone of Hsp70 in T. brucei.

PMID: 19815073 [PubMed - as supplied by publisher]

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9: Mol Biochem Parasitol. 2009 Oct 5. [Epub ahead of print]

Validationof a New Method for Immobilising Kinetoplastid Parasites for Live Cell Imaging.

Price HP, Maclean L, Marrison J, O'Toole PJ, Smith DF.

Centre for Immunology and Infection, Department of Biology/Hull York Medical School, University of York, Heslington, York YO10 5YW, UK.

The kinetoplastid parasites are responsible for three of the ten most neglected tropical diseases as classified by the WHO. Recent advances in molecular and cellular analyses have allowed rapid progress in our understanding of the biology of these lethal pathogens. In this study we validate a new method for immobilising Trypanosoma brucei and Leishmania major parasites while maintaining a high level of viability. This allows reproducible live cell imaging of these highly motile organisms, thus enabling a full complement of advanced microscopic techniques to be utilised to better understand these pathogenic species.

PMID: 19815033 [PubMed - as supplied by publisher]

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10: Acta Otorrinolaringol Esp. 2009 Jul-Aug;60(4):298-300. Epub 2009 Jun 24.

[Mucocutaneous Leishmaniasis: An imported illness with ENT repercussions.]

[Article in Spanish]

González M, Benito F, García L, Iglesias A.

Servicio de ORL y PCF, Hospital Clínico Universitario de Salamanca, España.

We report a case of mucocutaneous Leishmaniasis, an uncommon illness in our area. Leishmaniasis covers a group of diseases caused by protozoa of the genus Leishmania with several pathogenic species transmitted by Phlebotomus mosquitoes. Leishmania braziliensis is endemic in parts of South America and is responsible for the mucocutaneous Leishmaniasis reported here. The initial lesion is cutaneous and appears as an ulcer on arms or legs, leaving a scar. In some untreated cases, a late phase may affect the ENT area with hard to treat chronic destructive and mutilating lesions.

PMID: 19814979 [PubMed - in process]

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