RESUMEN
Human and yeast mitochondrial DNA polymerases (DNAPs), POLG and Mip1, are related by evolution to bacteriophage DNAPs. However, mitochondrial DNAPs contain unique amino and carboxyl-terminal extensions that physically interact. Here we describe that N-terminal deletions in Mip1 polymerases abolish polymerization and decrease exonucleolytic degradation, whereas moderate C-terminal deletions reduce polymerization. Similarly, to the N-terminal deletions, an extended C-terminal deletion of 298 amino acids is deficient in nucleotide addition and exonucleolytic degradation of double and single-stranded DNA. The latter observation suggests that the physical interaction between the amino and carboxyl-terminal regions of Mip1 may be related to the spread of pathogenic POLG mutant along its primary sequence.
Asunto(s)
ADN Polimerasa I/metabolismo , ADN de Hongos/biosíntesis , ADN Mitocondrial/biosíntesis , Proteínas Mitocondriales/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Dominio Catalítico , ADN Polimerasa I/genética , ADN Polimerasa gamma/genética , ADN Polimerasa gamma/metabolismo , ADN de Hongos/genética , ADN Mitocondrial/genética , Humanos , Proteínas Mitocondriales/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genéticaRESUMEN
The parasite responsible for autochthonous cutaneous leishmaniasis in Martinique island (French West Indies) was first isolated in 1995; its taxonomical position was established only in 2002, but it remained unnamed. In the present paper, the authors name this parasite Leishmania (Leishmania) martiniquensis Desbois, Pratlong & Dedet n. sp. and describe the type strain of this taxon, including its biological characteristics, biochemical and molecular identification, and pathogenicity. This parasite, clearly distinct from all other Euleishmania, and placed at the base of the Leishmania phylogenetic tree, is included in the subgenus Leishmania.
Asunto(s)
Enfermedades Endémicas , Leishmania/clasificación , Leishmaniasis Cutánea/parasitología , ADN Polimerasa I/genética , Humanos , Isoenzimas/genética , Leishmania/enzimología , Leishmania/genética , Leishmania/aislamiento & purificación , Leishmania/fisiología , Leishmaniasis Cutánea/epidemiología , Martinica/epidemiología , Proteínas Protozoarias/genética , ARN Polimerasa II/genética , ARN Protozoario/genética , ARN Ribosómico 18S/genética , Ribotipificación , Terminología como AsuntoRESUMEN
Venereal syphilis is a multi-stage, sexually transmitted disease caused by the spirochetal bacterium Treponema pallidum (Tp). Herein we describe a cohort of 57 patients (age 18-68 years) with secondary syphilis (SS) identified through a network of public sector primary health care providers in Cali, Colombia. To be eligible for participation, study subjects were required to have cutaneous lesions consistent with SS, a reactive Rapid Plasma Reagin test (RPR-titer > or = 1 : 4), and a confirmatory treponemal test (Fluorescent Treponemal Antibody Absorption test- FTA-ABS). Most subjects enrolled were women (64.9%), predominantly Afro-Colombian (38.6%) or mestizo (56.1%), and all were of low socio-economic status. Three (5.3%) subjects were newly diagnosed with HIV infection at study entry. The duration of signs and symptoms in most patients (53.6%) was less than 30 days; however, some patients reported being symptomatic for several months (range 5-240 days). The typical palmar and plantar exanthem of SS was the most common dermal manifestation (63%), followed by diffuse hypo- or hyperpigmented macules and papules on the trunk, abdomen and extremities. Three patients had patchy alopecia. Whole blood (WB) samples and punch biopsy material from a subset of SS patients were assayed for the presence of Tp DNA polymerase I gene (polA) target by real-time qualitative and quantitative PCR methods. Twelve (46%) of the 26 WB samples studied had quantifiable Tp DNA (ranging between 194.9 and 1954.2 Tp polA copies/ml blood) and seven (64%) were positive when WB DNA was extracted within 24 hours of collection. Tp DNA was also present in 8/12 (66%) skin biopsies available for testing. Strain typing analysis was attempted in all skin and WB samples with detectable Tp DNA. Using arp repeat size analysis and tpr RFLP patterns four different strain types were identified (14d, 16d, 13d and 22a). None of the WB samples had sufficient DNA for typing. The clinical and microbiologic observations presented herein, together with recent Cali syphilis seroprevalence data, provide additional evidence that venereal syphilis is highly endemic in this region of Colombia, thus underscoring the need for health care providers in the region to be acutely aware of the clinical manifestations of SS. This study also provides, for the first time, quantitative evidence that a significant proportion of untreated SS patients have substantial numbers of circulating spirochetes. How Tp is able to persist in the blood and skin of SS patients, despite the known presence of circulating treponemal opsonizing antibodies and the robust pro-inflammatory cellular immune responses characteristic of this stage of the disease, is not fully understood and requires further study.
Asunto(s)
Sífilis Cutánea/epidemiología , Sífilis Cutánea/patología , Treponema pallidum/aislamiento & purificación , Adolescente , Adulto , Anciano , Anticuerpos Antibacterianos/sangre , Proteínas Bacterianas/genética , Técnicas de Tipificación Bacteriana , Análisis por Conglomerados , Estudios de Cohortes , Colombia/epidemiología , Dermatoglifia del ADN , ADN Polimerasa I/genética , ADN Bacteriano/genética , ADN Bacteriano/aislamiento & purificación , Enfermedades Endémicas , Femenino , Genotipo , Humanos , Masculino , Persona de Mediana Edad , Epidemiología Molecular , Reacción en Cadena de la Polimerasa , Polimorfismo de Longitud del Fragmento de Restricción , Reaginas/sangre , Piel/microbiología , Piel/patología , Sífilis Cutánea/microbiología , Treponema pallidum/clasificación , Treponema pallidum/genética , Adulto JovenRESUMEN
Two cases of skin lesions similar to those caused by Leishmania parasites have been reported from Martinique. Parasites isolated from these lesions were unlike Leishmania reference strains by isoenzyme analysis and electron microscopy and were assumed to be monoxenous trypanosomatids which normally only infect invertebrates. Both strains have now been retyped by isoenzyme analysis and found to be identical to each other and distantly related to all other Leishmania species. The sequence of the 18S ribosomal RNA gene and partial sequences of the DNA polymerase alpha and RNA polymerase II largest subunit genes were obtained. These sequences indicated that the Martinique parasites clustered with L. enriettii and were basal to all other euleishmania. However, support for both the position basal to all euleishmania and the clustering with L. enriettii was low. The Martinique parasites may cluster with L. (Leishmania) or L. (Viannia) or form a novel clade within the euleishmania either with or without L. enriettii.
Asunto(s)
Leishmania/clasificación , Leishmaniasis Cutánea/parasitología , Animales , ADN Polimerasa I/química , ADN Polimerasa I/genética , ADN Protozoario/química , ADN Protozoario/genética , ADN Protozoario/aislamiento & purificación , ADN Ribosómico/química , ADN Ribosómico/genética , ADN Ribosómico/aislamiento & purificación , Electroforesis en Gel de Almidón , Humanos , Isoenzimas/metabolismo , Leishmania/enzimología , Leishmania/genética , Leishmaniasis Cutánea/patología , Martinica , Filogenia , Reacción en Cadena de la Polimerasa , ARN Polimerasa II/química , ARN Polimerasa II/genética , Alineación de Secuencia , Análisis de Secuencia de ADN , Homología de Secuencia de Ácido NucleicoRESUMEN
Thymine auxotrophy was shown to be associated with an increase in UV sensitivity both in Bacillus subtilis and in Escherichia coli. This UV sensitization became clearly evident in polA5 mutants of Bacillus subtilis: at UV doses of 16 J/m2, a reduction of more than 10-fold in the survivor population is observed in thymine requiring spontaneous mutants (polA5 thyA thyB) compared to the parental strains (polA5). Reversion of either thyA or thyB mutation led to a partial recovery in the UV resistance. This result suggests that DNA repair polymerization might be improved by the biosynthesis of thymidylate or some effect associated with such activity.