Anionic Sites, Fucose Residues and Class I Human Leukocyte Antigen Fate During Interaction of Toxoplasma gondii with Endothelial Cells

Toxoplasma gondii invades and proliferates in human umbilical vein endothelial cells where it resides in a parasitophorous vacuole. In order to analyze which components of the endothelial cell plasma membrane are internalized and become part of the parasitophorous vacuole membrane, the culture of endothelial cells was labeled with cationized ferritin or UEA I lectin or anti Class I human leukocytte antigen (HLA) before or after infection with T. gondii. The results showed no cationized ferritin and UEA I lectin in any parasitophorous vacuole membrane, however, the Class I HLA molecule labeling was observed in some endocytic vacuoles containing parasite until 1 h of interaction with T. gondii. After 24 h parasite-host cell interaction, the labeling was absent on the vacuolar membrane, but presents only in small vesicles near parasitophorous vacuole. These results suggest the anionic site and fucose residues are excluded at the time of parasitophorous vacuole formation while Class I HLA molecules are present only on a minority of Toxoplasma-containig vacuoles.

Molluscicidal activity of Physalis angulata L. extracts and fractions on Biomphalaria tenagophila (d'Orbigny, 1835) under laboratory conditions

The main objective of this research is to evaluate the molluscicide activity of Physalis angulata L. Biomphalaria tenagophila specimens under laboratory conditions. Extracts and fractions were supplied by the Laboratório de Química de Produtos Naturais, Farmanguinhos-Fiocruz. Experiments were performed according to the methodology described by the World Health Organization for molluscicide tests using the concentrations from 0.1 to 500 mg/l of the extracts, fractions and of a pool of physalins modified steroids present in this species. The results show that ethyl acetate and acetone extracts from the whole plant, the ethanolic extracts of the roots and the physalins pool from stems and leaves were active. Only the whole plant extracts were available in sufficient quantity for the determination of LD50 and LD90 values.

Emergence of dhfrXVb and blaCARB-4 gene cassettes in class 1 integrons from clinical Pseudomonas aeruginosa isolated in Amazon region

Integrons play a role in horizontal acquisition and expression of genes, as well as gene reservoir, contributing for the resistance phenotype, particularly relevant to bacteria of clinical importance. We aimed to determine the composition and the organization of the class 1 integron variable region present in Pseudomonas aeruginosa clinical isolates from Brazil. Strains carrying class 1 integrons were resistant to the majority of antibiotics tested, except to imipenem and ceftazidime. Sequence analysis of the integron variable region revealed the presence of the blaCARB-4 gene into two distinct cassette arrays: aacA4-dhfrXVb-blaCARB-4 and aadB-aacA4-blaCARB-4 . dhfrXVb gene cassette, which is rare in Brazil and in P. aeruginosa species, was found in one isolate. PFGE analysis showed the spread of blaCARB-4 among P. aeruginosa clones. The occurrence of blaCARB-4 and dhfrXVb in Brazil may contribute for developing resistance to clinically important antibiotics, and shows a diversified scenarium of these elements occurring in Amazon clinical settings, where no study about integron dinamycs was performed to date.

blaCTX-M-2 and blaCTX-M-28 extended-spectrum β-lactamase genes and class 1 integrons in clinical isolates of Klebsiella pneumoniae from Brazil

Twenty-eight Klebsiella pneumoniae clinical isolates that exhibited an extended-spectrum cephalosporin-resistance profile from a city in the Northeast of Brazil were analysed by PCR and DNA sequencing in order to determine the occurrence of blaCTX-M genes and class 1 integrons. We determined the occurrence of the blaCTX-M-2 gene in six K. pneumoniae isolates and describe the first detection of the blaCTX-M-28 gene in South America. Seven isolates carried class 1 integrons. Partial sequencing analysis of the 5'-3'CS variable region in the class 1 integrons of three isolates revealed the presence of aadA1, blaOXA-2 and dfr22 gene cassettes.

Occurrence and composition of class 1 and class 2 integrons in clinical and environmental O1 and non-O1/non-O139 Vibrio cholerae strains from the Brazilian Amazon

This study identified and characterised class 1 and 2 integrons in clinical and environmental Vibrio cholerae O1 and non-O1/non-O139 strains isolated from the Brazilian Amazon. The aadA2 and aadA7 gene cassettes were found in class 1 integrons in two genotypes of environmental V. cholerae non-O1/non-O139. Empty integrons were found in strains from the Brazilian cholera epidemic. A class 2 integron was detected in one strain from the V. cholerae Amazonia lineage harbouring sat1 and aadA1 genes. All isolates were resistant to aminoglycosides, indicating aadA functionality. These findings suggest that environmental bacteria act as cassette reservoirs that favour the emergence of resistant pathogens.

Association analysis of human leukocyte antigen class II (DRB1) alleles with leprosy in individuals from São Luís, state of Maranhão, Brazil

Epidemiological studies have demonstrated that the variability of the clinical response to infection caused by Mycobacterium leprae is associated with host genetic factors. The present study investigated the frequency of human leukocyte antigen (HLA) class II (DRB1) alleles in patients with leprosy from São Luís, Maranhão, Brazil. A case-control study was performed in 85 individuals with leprosy and 85 healthy subjects. All samples were analysed via polymerase chain reaction-sequence specific oligonucleotide probes. The HLA-DRB1*16 allele showed a higher frequency in the group with leprosy [(9.41% vs. 4.12%) odds ratio (OR) = 2.41 95% confidence interval (CI) (0.96-6.08) p = 0.05], whereas the HLA-DRB1*11 allele was less frequent in the group with leprosy [(6.47% vs. 11.76%) OR = 0.51 95% CI (0.23-1.12) p = 0.09]. The frequency of HLA-DRB1* alleles between the control group and leprosy patient subgroups presenting different forms of the disease showed that the HLA-DRB1*16 (16.13% vs. 8.24%, OR = 4.10, CI = 1.27-13.27, p = 0.010) and HLA-DRB1*14 (5% vs. 3.53%, OR = 4.63, CI = 1.00-21.08, p = 0.032) alleles were significantly more frequent in patients with different clinical subtypes of leprosy. The sample size was a limitation in this study. Nevertheless, the results demonstrated the existence of a genetic susceptibility associated with the clinical forms of leprosy. The low frequency of the HLA-DRB1*11 allele should be further studied to investigate the possible protective effect of this allele.

In vitro and in vivo antimalarial activity and cytotoxicity of extracts, fractions and a substance isolated from the Amazonian plant Tachia grandiflora (Gentianaceae)

Tachia sp. are used as antimalarials in the Amazon Region and in vivo antimalarial activity of a Tachia sp. has been previously reported. Tachia grandiflora Maguire and Weaver is an Amazonian antimalarial plant and herein its cytotoxicity and antimalarial activity were investigated. Spectral analysis of the tetraoxygenated xanthone decussatin and the iridoid aglyone amplexine isolated, respectively, from the chloroform fractions of root methanol and leaf ethanol extracts was performed. In vitro inhibition of the growth of Plasmodium falciparum Welch was evaluated using optical microscopy on blood smears. Crude extracts of leaves and roots were inactive in vitro. However, chloroform fractions of the root and leaf extracts [half-maximal inhibitory concentration (IC50) = 10.5 and 35.8 µg/mL, respectively] and amplexine (IC50= 7.1 µg/mL) were active in vitro. Extracts and fractions were not toxic to type MRC-5 human fibroblasts (IC50> 50 µg/mL). Water extracts of the roots of T. grandiflora administered by mouth were the most active extracts in the Peters 4-day suppression test in Plasmodium berghei-infected mice. At 500 mg/kg/day, these extracts exhibited 45-59% inhibition five to seven days after infection. T. grandiflora infusions, fractions and isolated substance have potential as antimalarials.

Nitrogen fractions in the microbial biomass in soils of southern Brazil

The reaction of nitrogen compounds with ninhydrin can be used as an indicator of cytoplasmic materials released from microbial cells killed by fumigation. Total-N, ninhydrin-reactive-N (NR-N), ammonium-N (A-N), and α-amino-N in the microbial biomass of soils from the State of Rio Grande do Sul, Brazil, were determined, in 1996, in 0.5 mol L-1 K2SO4 extracts of fumigated and non-fumigated soils. Total-N varied from 20.3 to 104.4 mg kg-1 and the ninhydrin-reactive-N corresponded, in average, to 27% of this. The ninhydrin-reactive-N was made up of 67% ammonium-N and 33% aminoacids with the amino group at the α-carbon position. It was concluded that colorimetric analysis of NR-N and A-N may be used as a direct measure of microbial N in soil. This simple and rapid procedure is adequate for routine analyses.

Relationships between soil class and nutritional status of coffee plantations

Farm planning requires an assessment of the soil class. Research suggest that the Diagnosis and Recommendation Integrated System (DRIS) has the capacity to evaluate the nutritional status of coffee plantations, regardless of environmental conditions. Additionally, the use of DRIS could reduce the costs for farm planning. This study evaluated the relationship between the soil class and nutritional status of coffee plants (Coffea canephora Pierre) using the Critical Level (CL) and DRIS methods, based on two multivariate statistical methods (discriminant and multidimensional scaling analyses). During three consecutive years, yield and foliar concentration of nutrients (N, P, K, Ca, Mg, S, B, Zn, Mn, Fe and Cu) were obtained from coffee plantations cultivated in Espírito Santo state. Discriminant analysis showed that the soil class was an important factor determining the nutritional status of the coffee plants. The grouping separation by the CL method was not as effective as the DRIS one. The bidimensional analysis of Euclidean distances did not show the same relationship between plant nutritional status and soil class. Multidimensional scaling analysis by the CL method indicated that 93.3 % of the crops grouped into one cluster, whereas the DRIS method split the fields more evenly into three clusters. The DRIS method thus proved to be more consistent than the CL method for grouping coffee plantations by soil class.

Soil fertility, organic carbon and fractions of the organic matter at different distances from eucalyptus stumps

Knowledge on variations in vertical, horizontal and temporal characteristics of the soil chemical properties under eucalyptus stumps left in the soil is of fundamental importance for the management of subsequent crops. The objective of this work was to evaluate the effect of eucalyptus stumps (ES) left after cutting on the spatial variability of chemical characteristics in a dystrophic Yellow Argisol in the eastern coastal plain region of Brazil. For this purpose, ES left for 31 and 54 months were selected in two experimental areas with similar characteristics, to assess the decomposition effects of the stumps on soil chemical attributes. Soil samples were collected directly around these ES, and at distances of 30, 60, 90, 120 and 150 cm away from them, in the layers 0-10, 10-20 and 20-40 cm along the row of ES, which is in-between the rows of eucalyptus trees of a new plantation, grown at a spacing of 3 x 3 m. The soil was sampled in five replications in plots of 900 m² each and the samples analyzed for pH, available P and K (Mehlich-1), exchangeable Al, Ca and Mg, total organic carbon (TOC) and C content in humic substances (HS) and in the free light fraction. The pH values and P, K, Ca2+, Mg2+ and Al3+ contents varied between the soil layers with increasing distance from the 31 and 54-monthold stumps. The highest pH, P, K, Ca2+ and Mg2+ values and the lowest Al3+ content were found in the surface soil layer. The TOC of the various fractions of soil organic matter decreased with increasing distance from the 31 and 54-month-old ES in the 0-10 and 10-20 cm layers, indicating that the root (and stump) cycling and rhizodeposition contribute to maintain soil organic matter. The C contents of the free light fraction, of the HS and TOC fractions were higher in the topsoil layer under the ES left for 31 months due to the higher clay levels of this layer, than in those found under the 54-month-old stumps. However, highest C levels of the different fractions of soil organic matter in the topsoil layer reflect the deposition and maintenance of forest residues on the soil surface, mainly after forest harvest.

Soil phosphorus fractions in sandy soils amended with cattle manure for long periods

Phosphorus fractions were determined in soil samples from areas fertilized or not with farmyard cattle manure (FYM) and in samples of FYM used in the semi-arid region of Paraiba state, Brazil. Soil samples were taken from the 0-20; 20-40 and 40-60 cm layers of 18 cultivated areas, which, according to interviews with farmers, had been treated with 12 to 20 t ha-1 FYM annually, for the past 2 to 40 years. Soil samples were also collected from four unfertilized pasture areas as controls. Phosphorus in the soil samples was sequentially extracted with water (Pw), resin (Pres), NaHCO3 (Pi bic and Po bic), NaOH (Pi hid and Po hid), H2SO4 (Pacid) and, finally, by digestion with H2SO4/H2O2 (Presd). Nine FYM samples were extracted with water, resin, Mehlich-1, H2SO4, NaOH or digestion with H2SO4/H2O2, not sequentially, and the extracts analyzed for P. The sampled areas had homogeneous, sandy and P-deficient soils; increases in total soil P (Pt) above the mean value of the control areas (up to 274 mg kg-1 in the 0-20 cm layer of the most P-enriched samples) were therefore attributed to FYM applications, which was the only external P input in the region. Regression analysis was used to study the relationship between soil P fractions and Pt. The Pacid fraction, related to Ca-P forms, showed the greatest increases (p < 0.01) as a result of FYM applications, rising from 8.4 mg kg-1 in a non-fertilized sample to 43.8 mg kg-1 in the sample with the highest Pt content. The sum of Pw, Pres and Pi bic, considered as labile P, showed comparable increases with Pacid, while Pi hid showed the smallest increase due to FYM applications. Organic P forms also increased, more so the fraction Po hid, considered less labile, than the more labile one, Po bic. The residual P fraction was practically half of Pt, independently of the Pt value. Increases in labile P, Pacid and organic P were justified by the high average concentration of Pw (36 %), Pacid (34 %), and Po hid (30 %) in the FYM. Significant changes in the proportion of P forms among soil layers indicated the downward movement of P in organic forms.

The effect of plantation silviculture on soil organic matter and particle-size fractions in Amazonia

Eucalyptus grandis and other clonal plantations cover about 3.5 million ha in Brazil. The impacts of intensively-managed short-rotation forestry on soil aggregate structure and Carbon (C) dynamics are largely undocumented in tropical ecosystems. Long-term sustainability of these systems is probably in part linked to maintenance of soil organic matter and good soil structure and aggregation, especially in areas with low-fertility soils. This study investigated soil aggregate dynamics on a clay soil and a sandy soil, each with a Eucalyptus plantation and an adjacent primary forest. Silvicultural management did not reduce total C stocks, and did not change soil bulk density. Aggregates of the managed soils did not decrease in mass as hypothesized, which indicates that soil cultivation in 6 year cycles did not cause large decreases in soil aggregation in either soil texture. Silt, clay, and C of the sandy plantation soil shifted to greater aggregate protection, which may represent a decrease in C availability. The organic matter in the clay plantation soil increased in the fractions considered less protected while this shift from C to structural forms considered more protected was not observed.

Physical properties and particle-size fractions of soil organic matter in crop-livestock integration

Crop-livestock integration represents an interesting alternative of soil management, especially in regions where the maintenance of cover crops in no-tillage systems is difficult. The objective of this study was to evaluate soil physical and chemical properties, based on the hypothesis that a well-managed crop-livestock integration system improves the soil quality and stabilizes the system. The experiment was set up in a completely randomized design, with five replications. The treatments were arranged in a 6 x 4 factorial design, to assess five crop rotation systems in crop-livestock integration, and native forest as reference of soil undisturbed by agriculture, in four layers (0.0-0.05; 0.05-0.10; 0.10-0.15 and 0.15-0.20 m). The crop rotation systems in crop-livestock integration promoted changes in soil physical and chemical properties and the effects of the different systems were mainly detected in the surface layer. The crops in integrated crop-livestock systems allowed the maintenance of soil carbon at levels equal to those of the native forest, proving the efficiency of these systems in terms of soil conservation. The systems influenced the environmental stability positively; the soil quality indicator mineral-associated organic matter was best related to aggregate stability.

Microbiological properties and oxidizable organic carbon fractions of an oxisol under coffee with split phosphorus applications and irrigation regimes

Phosphorus fertilization and irrigation increase coffee production, but little is known about the effect of these practices on soil organic matter and soil microbiota in the Cerrado. The objective of this study was to evaluate the microbiological and oxidizable organic carbon fractions of a dystrophic Red Latossol under coffee and split phosphorus (P) applications and different irrigation regimes. The experiment was arranged in a randomized block design in a 3 x 2 factorial design with three split P applications (P1: 300 kg ha-1 P2O5, recommended for the crop year, of which two thirds were applied in September and the third part in December; P2: 600 kg ha-1 P2O5, applied at planting and then every two years, and P3: 1,800 kg ha-1 P2O5, the requirement for six years, applied at once at planting), two irrigation regimes (rainfed and year-round irrigation), with three replications. The layers 0-5 and 5-10 cm were sampled to determine microbial biomass carbon (MBC), basal respiration (BR), enzyme activity of acid phosphatase, the oxidizable organic carbon fractions (F1, F2, F3, and F4), and total organic carbon (TOC). The irrigation regimes increased the levels of MBC, microbial activity and acid phosphatase, TOC and oxidizable fractions of soil organic matter under coffee. In general, the form of dividing P had little influence on the soil microbial properties and OC. Only P3 under irrigation increased the levels of MBC and acid phosphatase activity.