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1.
Maria Rebelo Claudia Sousa Howard M. Shapiro Maria M. Mota Martin P. Grobusch Thomas H?nscheid 《PloS one》2013,8(4)
Resistance of Plasmodium falciparum to almost all antimalarial drugs, including the first-line treatment with artemisinins, has been described, representing an obvious threat to malaria control. In vitro antimalarial sensitivity testing is crucial to detect and monitor drug resistance. Current assays have been successfully used to detect drug effects on parasites. However, they have some limitations, such as the use of radioactive or expensive reagents or long incubation times. Here we describe a novel assay to detect antimalarial drug effects, based on flow cytometric detection of hemozoin (Hz), which is rapid and does not require any additional reagents. Hz is an optimal parasite maturation indicator since its amount increases as the parasite matures. Due to its physical property of birefringence, Hz depolarizes light, hence it can be detected using optical methods such as flow cytometry. A common flow cytometer was adapted to detect light depolarization caused by Hz. Synchronized in vitro cultures of P. falciparum were incubated for 48 hours with several antimalarial drugs. Analysis of depolarizing events, corresponding to parasitized red blood cells containing Hz, allowed the detection of parasite maturation. Moreover, chloroquine resistance and the inhibitory effect of all antimalarial drugs tested, except for pyrimethamine, could be determined as early as 18 to 24 hours of incubation. At 24 hours incubation, 50% inhibitory concentrations (IC50) were comparable to previously reported values. These results indicate that the reagent-free, real-time Hz detection assay could become a novel assay for the detection of drug effects on Plasmodium falciparum. 相似文献
2.
Maturity Onset Diabetes of the Young (MODY) is a heterogeneous group of genetic diseases characterized by a primary defect in insulin secretion and hyperglycemia, non-ketotic disease, monogenic autosomal dominant mode of inheritance, age at onset less than 25 years, and lack of auto-antibodies. It accounts for 2–5% of all cases of non-type 1 diabetes. MODY subtype 2 is caused by mutations in the glucokinase (GCK) gene. In this study, we sequenced the GCK gene of two volunteers with clinical diagnosis for MODY2 and we were able to identify four mutations including one for a premature stop codon (c.76C>T). Based on these results, we have developed a specific PCR-RFLP assay to detect this mutation and tested 122 related volunteers from the same family. This mutation in the GCK gene was detected in 21 additional subjects who also had the clinical features of this genetic disease. In conclusion, we identified new GCK gene mutations in a Brazilian family of Italian descendance, with one due to a premature stop codon located in the second exon of the gene. We also developed a specific assay that is fast, cheap and reliable to detect this mutation. Finally, we built a molecular ancestry model based on our results for the migration of individuals carrying this genetic mutation from Northern Italy to Brazil. 相似文献
3.
The influence of several polymeric additives on specific glucose uptake rate of flocs of a S. cerevisiae strain — S. cerevisiae NRRLY 265 was studied. A special continuous membrane microreactor was used to measure glucose uptake on the presence of calcium and of the tested additives — two cationic polymers — bis(polyoxyethylene-bis(amine)) 20,000 and BPA 1,000 and one anionic polymer — Magna Floc LT25.An increase on glucose uptake rate was always observed when comparing with calcium bound flocs. For bis(polyoxyethylene-bis(amine)) 20,000 the increase was only 19% but for BPA 1,000 a value of more than 50% was observed. For Magna Floc LT25 a two fold increase was measured.The determination of floc size and porosity in the presence of the additives indicated that, on the basis of these parameters, it was not possible to explain the observed glucose uptake rates. The floc porosites in additive bound flocs were similar and 10% larger than for calcium bound flocs and glucose uptake rate was larger for the largest flocs — Magna Floc LT25 bound flocs were the largest followed by BPA 1,000, bis(polyoxyethylene-bis(amine)) 20,000 and calcium bound flocs. These values disagree with what should be expected in diffusion controlled processes.The calculation of intercellular floc distance indicated that polymeric additives act on the reduction of diffusional limitations by increasing the available flux area for glucose inside the flocs. By analysing different kinds of packings, it was also observed that the packing arrangement for yeast cells in flocs is close to the cubic packing. The simulation of this arrangement for the obtained floc sizes confirmed that the 10% increase in floc porosity is sufficient to explain the increase in the available flux area. 相似文献
4.
M. Aluja J. Guillen P. Liedo M. Cabrera E. Rios G. de la Rosa H. Celedonio D. Mota 《BioControl》1990,35(1):39-48
A total of 1,302 parasitoids representing 8 species and 4 families were recovered from 9,818 fruit fly host fruits sampled.
The most common parasitoid species wasDiachasmimorpha longicaudata (Ashmead). Average percent parasitism ranged between 0.44 and 29.23%. Parasitoid emergence data indicate thatAnastrepha ludens (Loew),A. obliqua (Sein),A. serpentina (Wiedeman),A. striata (Schiner) andToxotrypana curvicauda (Gerstaecker) were subject to parasitism. We provide information on the population fluctuation ofAnastrepha ludens, A. obliqua, A. serpentina, A. distincta (Greene),A. striata, A. fraterculus (Wiedeman),A. chiclayae (Greene),A. montei (Costa Lima),A. leptozona (Hendel) andA. tripunctata (Wulp).Anastrepha ludens andA. obliqua were the most common species, representing 95.3% of all fruit fly species caught in McPhail traps.
相似文献
5.
Dr M. T. S. Peraçoli M. T. Rezkallah-Iwasso N. G. S. Mota M. R. Montenegro 《Mycopathologia》1993,121(3):149-156
The effect of dialysable leukocyte extracts (DLE) obtained from hamsters immunized withParacoccidioides brasiliensis (immune DLE) and from non-immunized hamsters (non-immune DLE) was studied in hamsters inoculated withP. brasiliensis by the intratesticular route. Treatment with immune or non-immune DLE was started during the third week of infection and was repeated at 7, 11, 15 and 19 weeks. A group of untreated infected animals was used as control. Animals were submitted to the delayed hypersensitivity skin test toP. brasiliensis antigen (PbAg) in vivo and assayed in vitro by the macrophage migration inhibition test in the presence of Phytohemagglutinin (PHA) and PbAg and by immunodiffusion for specific antibody. The animals were sacrificed at 4, 8, 12, 16 and 20 weeks. The morphology and extension of the lesions were studied at the inoculation site, and in lymph nodes, lungs, liver, spleen and kidneys. In contrast to the controls, animals treated with both DLEs maintained a positive cell-mediated immune response throughout the experiment and developed less extensive infection with a significantly lower number of fungi in the lesions. The results suggest that immune and non-immune DLE preparations modified the evolution of experimental paracoccidioidomycosis with equal efficiency. This similarity may be explained by the immunoregulatory activities of both extracts. 相似文献
6.
Interaction between nonflocculent and flocculent cells of Saccharomyces cerevisiae was studied. Adhesion experiments were done using three types of nonflocculent cells and a flocculent one. Two types of nonflocculent cells were obtained from the flocculent strain by changing environmental growth conditions. The integration of nonflocculent cells in the flocs was observed by two different methods: measurement of the sedimentation capacity of mixtures and microscopic observation of stained nonflocculent cells blended with flocculent cells. It was possible to verify that cell-cell interaction corresponds to a true stable binding and not to a simple entrapment inside the floc matrix. 相似文献
7.
J Fuentes Mota J Fernández-Bola?os Guzmán J M García Fernández W Moreda C Ortiz M A Pradera I Robina C Welsh 《Carbohydrate research》1992,232(1):47-57
The preparation of 2,3,4-tri-O-benzyl- (3), 2,3,4-tri-O-acetyl- (4), and 2,3,4-tri-O-benzoyl-N-(2,2-diethoxycarbonylvinyl)-6-O-trityl-beta- D-glucopyranosylamine (5) is described. The reaction of 3-5 with 2,3,4,6-tetra-O-acetyl-alpha-D-glucopyranosyl bromide yields 2,3,4-tri-O-benzyl- (9), 2,3,4-tri-O-acetyl- (10), and 2,3,4-tri-O-benzoyl-N-(2,2-diethoxycarbonylvinyl)-6-O-(2,3,4,6-tet ra-O- acetyl-beta-D-glucopyranosyl)-beta-D-glucopyranosylamine (11), respectively. 2,3,4-Tri-O-benzyl- (6), 2,3,4-tri-O-acetyl- (7), and 2,3,4-tri-O- benzoyl-N-(2,2-diethoxycarbonylvinyl)-beta-D-glucopyranosylamine (8) are also described. 相似文献
8.
Male hamsters (105) received intratesticular injection of suspension of a live yeast phase culture ofParacoccidioides brasiliensis and were sacrificed weekly during 20 weeks. Humoral immunity was studied by the agar-gel immunodiffusion (ID) and indirect immunofluorescence (IF) tests. Cell-mediated immunity was determined by the macrophage migration inhibition test in the presence of phytohemagglutinin (PHA) andParacoccidioides brasiliensis soluble antigen (PbAg). The morphology of the lesions was studied in the inoculation site, lymph nodes, lung, liver, spleen and kidneys.Disseminated paracoccidioidomycosis was observed in 100% of the animals after the first week. The lesions were initially made up of fungi surrounded by polymorphonuclear neutrophils and macrophages. Up to the 10th week the majority of the lesions appeared as compact confluent ephitelioid granulomas containing rare large fungi, some showing signs of degeneration. At this time, the specific antibody titers and the cellular immune response to PHA and PbAg were highest.From the 11th week on the granulomas became less compact, edematous with the epithelioid cells loosely arranged. This change was accompanied by an increase in the number of fungi showing reproductive activity and was associated with renal amyloidosis and progressive decline of cellular immune response both to PHA and PbAg. Contrariwise the titers of circulating antibodies were maintained.In the present model, disseminated paracoccidioidomycosis of the hamster was associated with depression of cellular immunity, change in the pattern of the granuloma, intense fungi proliferation and amyloidosis. 相似文献
9.
Sophie Mallez Chantal Castagnone Margarida Espada Paulo Vieira Jonathan D. Eisenback Manuel Mota Thomas Guillemaud Philippe Castagnone-Sereno 《PloS one》2013,8(3)
The pinewood nematode, Bursaphelenchus xylophilus, native to North America, is the causative agent of pine wilt disease and among the most important invasive forest pests in the East-Asian countries, such as Japan and China. Since 1999, it has been found in Europe in the Iberian Peninsula, where it also causes significant damage. In a previous study, 94 pairs of microsatellite primers have been identified in silico in the pinewood nematode genome. In the present study, specific PCR amplifications and polymorphism tests to validate these loci were performed and 17 microsatellite loci that were suitable for routine analysis of B. xylophilus genetic diversity were selected. The polymorphism of these markers was evaluated on nematodes from four field origins and one laboratory collection strain, all originate from the native area. The number of alleles and the expected heterozygosity varied between 2 and 11 and between 0.039 and 0.777, respectively. First insights into the population genetic structure of B. xylophilus were obtained using clustering and multivariate methods on the genotypes obtained from the field samples. The results showed that the pinewood nematode genetic diversity is spatially structured at the scale of the pine tree and probably at larger scales. The role of dispersal by the insect vector versus human activities in shaping this structure is discussed. 相似文献
10.
Marlene Lopes Carla Oliveira Lucília Domingues Manuel Mota Isabel Belo 《Biotechnology progress》2014,30(5):1040-1047
Pichia pastoris is a widely used host for the production of heterologous proteins. In this case, high cell densities are needed and oxygen is a major limiting factor. The increased air pressure could be used to improve the oxygen solubility in the medium and to reach the high oxygen demand of methanol metabolism. In this study, two P. pastoris strains producing two different recombinant proteins, one intracellular (β‐galactosidase) and other extracellular (frutalin), were used to investigate the effect of increased air pressure on yeast growth in glycerol and heterologous protein production, using the methanol AOX1‐inducible system. Experiments were carried out in a stainless steel bioreactor under total air pressure of 1 bar and 5 bar. The use of an air pressure raise of up to 5 bar proved to be applicable for P. pastoris cultivation. Moreover, no effects on the kinetic growth parameters and methanol utilization (Mut) phenotype of strains were found, while an increase in recombinant β‐galactosidase‐specific activity (ninefold) and recombinant frutalin production was observed. Furthermore, the air pressure raise led to a reduction in the secreted protease specific activity. This work shows for the first time that the application of an air pressure of 5 bar may be used as a strategy to decrease protease secretion and improve recombinant protein production in P. pastoris. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 30:1040–1047, 2014 相似文献