The effects of bacterial leaching on metal partitioning in sewage sludge

The partitioning of Mn, Al, Zn, Cu and Ti ions in municipal sewage sludge was investigated before and after bioleaching processes effectuated by Acidithiobacillus ferrooxidans and Acidithiobacillus thiooxidans. Oxidation reduction potential increase and pH decrease were obtained as a result of bacterial activity. A less pronounced and constant decrease was obtained with A. ferrooxidans, whereas A. thiooxidans presented a lag phase before a steep pH decrease. Metal solubilization was accomplished in experimental systems supplemented with energy source, Fe2+ for A. ferrooxidans and S-0 for A. thiooxidans. Solubilization efficiency differed for each metal except for Al, and was relatively similar for either organism. Metal partitioning was conducted using five-step sequential extraction procedure before and after the bioleaching. The results indicated that Zn and Mn ions were mostly associated with the organic fraction, whereas Cu, Al and Ti ions with the sulphide/ residue fraction. The bioleaching process caused prompt solubilization of metals mostly associated with the more labile fractions (exchangeable, adsorbed and organically bound metals), whereas those associated to the less labile ones (EDTA and sulphide/residue fractions) were exchanged towards more labile fractions.

Effect of anaerobic digestion and initial pH on metal bioleaching from sewage sludge

The effects of anaerobic digestion and initial pH on the bioleaching of metals from sewage sludge were investigated in shake flask experiments. A strain of Acidithiobacillus thiooxidans was employed in the assays using secondary and anaerobic sludges, which resulted in similar solubilization yields of the metals chromium, copper, lead, nickel, and zinc for both the sludges investigated. The effect of initial pH (7.0 and 4.0) on metal bioleaching was assayed by using the anaerobic sludge inoculated with indigenous sulfur-oxidizing thiobacilli. Although the time required to reach the end of the experiment (final pH close to 1.0) was shortened at initial pH of 4.0, final metal solubilization was not significantly different for both initial pH values, resulting in higher solubilization yields for copper, nickel, and zinc (higher than 80%). Chromium and lead presented solubilization yields close to 50%. The results obtained in this work showed that the metal bioleaching process can be applied to sewage sludge regardless of the type of sludge and without the requirement of pH adjustment.

Morphological and molecular characterization of Colletotrichum spp. from citrus orchards affected by postbloom fruit drop in Brazil

Brazilian isolates of Colletotrichum spp. from citrus orchards affected by postbloom fruit drop were examined for colony colour, mycelial growth, benomyl-resistance, pathogenicity, and genetic variability by random amplified polymorphic DNA (RAPD) analysis. All isolates were obtained from flowers and persistent calyxes from different citrus hosts from São Paulo, Brazil. DNA polymorphisms detected after amplification with random 10-mer primers were used to classify the isolates into two groups. Group I isolates grew rapidly on potato-dextrose agar (PDA) and were sensitive to benomyl, and group II isolates grew slowly on PDA and were benomyl-resistant. Colletotrichum acutatum was analyzed by RAPD and had high genetic similarity with group II isolates of Colletotrichum from citrus. Probably, the group I is C, gloeosporioides and group II is C. acutatum.

Physiological and genomic variations in rice cells recovered from direct immersion and storage in liquid nitrogen

The use of cryoprotectants and slow cooling rates are routine procedures for the cryopreservation of plant cell lines. However, our results with rice (Oryza sativa L,, ev. Taipei 309) show that calli can be cryopreserved by direct immersion and stored in liquid nitrogen without any cryoprotection, the efficiency of recovery using this method, as well as a conventional method was generally increased with a previous abscisic acid (ABA) treatment. Following cryopreservation, calli demonstrated some differences with respect to unfrozen calli of the same lines, Thus, resistance to freezing stress (- 20 degrees C for 2 h) increased significantly in all lines tested, irrespective of their pre-incubation with ABA, Calli that had been directly stored in liquid nitrogen also demonstrated a higher competence for genetic transformation than their unfrozen counterparts, as indicated by the transient gene expression levels obtained after particle bombardment, These differences might lead to further biotechnological applications, A genetic analysis of amplified DNA polymorphisms was performed with three independent lines that had been subjected to four combinations of ABA treatment and direct immersion in liquid nitrogen, At the loci screened with the randomly amplified polymorphic DNA (RAPD) markers tested, the genetic variations among lines and among calli of the same line appear to bd more related to tissue-culture-induced somaclonal variation than to cryoselection.

Use of primers derived from a new sequence of the bovine Y chromosome for sexing Bos taurus and Bos indicus embryos

A bovine male-specific marker was identified in our laboratory through random amplified polymorphic DNA (RAPD) analysis. This fragment of 3216 bp was cloned, sequenced and mapped by fluorescent in situ hybridization (FISH) on the taurine Yq. Primers derived from this sequence were initially screened by polymerase chain reaction (PCR) for their ability to detect Y-specific segments in zebu and taurine genomic DNA. Two of these primers amplified a 655 bp Y-specific sequence present in taurine and zebu male genomic DNA. These primers were then used for detecting the 655 bp male sequence in DNA from 173 zebu and 30 taurine embryos, which had been previously sexed using primers for the sequence BC 1.2. The results revealed an accuracy of 100%. (C) 2002 Elsevier B.V. All rights reserved.

An evaluation of the genetic diversity of Xylella fastidiosa isolated from diseased citrus and coffee in São Paulo, Brazil

Strains of Xylella fastidiosa, isolated from sweet orange trees (Citrus sinensis) and coffee trees (Coffea arabica) with symptoms of citrus variegated chlorosis and Requeima do Cafe, respectively, were indistinguish able based on repetitive extragenic palindromic polymerase chain reaction (PCR) and enterobacterial repetitive intergenic consensus PCR assays. These strains were also indistinguishable with a previously described PCR assay that distinguished the citrus strains from all other strains of Xylella fastidiosa. Because we were not able to document any genomic diversity in our collection of Xylella fastidiosa strains isolated from diseased citrus, the observed gradient of increasing disease severity from southern to northern regions of São Paulo State is unlikely due to the presence of significantly different strains of the pathogen in the different regions. When comparisons were made to reference strains of Xylella fastidiosa isolated from other hosts using these methods, four groups were consistently identified consistent with the hosts and regions from which the strains originated: citrus and coffee, grapevine and almond, mulberry, and elm, plum, and oak. Independent results from random amplified polymorphic DNA (RAPD) PCR assays were also consistent with these results; however, two of the primers tested in RAPD-PCR were able to distinguish the coffee and citrus strains. Sequence comparisons of a PCR product amplified from all strains of Xylella fastidiosa confirmed the presence of a CfoI polymorphism that can be used to distinguish the citrus strains from all others. The ability to distinguish Xylella fastidiosa strains from citrus and coffee with a PCR-based assay will be useful in epidemiological and etiological studies of this pathogen.

Tolerance to copper and zinc of Acidithiobacillus thiooxidans isolated from sewage sludge

The effect of copper and zinc ions on sulphur oxidation by Acidithiobacillus thiooxidans, strain SFR01, isolated from anaerobic sewage sludge was assessed, resulting in tolerance levels up to 20 and 200 mmol l(-1) for copper and zinc, respectively. The tolerance levels obtained were higher than the concentration of copper and zinc usually found in the collected sewage sludge. The tolerance levels obtained indicate no constraints for sludge bioleaching of those metals due to their toxicities to the indigenous A. thiooxidans.

An evaluation into the potential of biological processing for the removal of metals from sewage sludges

The use of sewage sludge in agricultural land as a means of sludge disposal and recycling has been shown to be economical and suitable because of the presence of nutrients such as nitrogen and phosphorus. However, municipal sludges often contain high quantities of toxic metals and other compounds that must be removed for its safe use in agricultural soils. The biological leaching of metals from sewage sludges has been shown to be a promising technique for metal detoxifying in such complex matrix. The process efficiency is dependent on several physico-chemical parameters, such as total solids concentration, metal forms, pH-ORP, and temperature. Scale-up of the process has not yet been defined and is still pursuing the correct operational design. Current research involving the bioleaching of metals from sewage sludge and its application to land, which affects soil physical properties, are presented and discussed.

Genetic variation within and among species of genus Arachis, section Rhizomatosae

The genus Arachis is endemic to South America and comprises 80 species, 69 of which have already been described and eleven not yet published. The genus includes the cultivated peanut ( A. hypogaea) and several forage species, the most important ones being A. glabrata and A. pintoi. Accessions of section Rhizomatosae, including three tetraploid species 2n = 4x = 40 (A. glabrata, A. pseudovillosa and A. nitida nom. nud.) and one diploid species 2n = 2x = 20 (A. burkartii), were evaluated using RAPD markers to assay genetic variability within and among species. The ten random primers used yielded a total of 113 polymorphic bands. The data were scored as the presence or absence of each band in each sample. A distance matrix and dendrogram were obtained using Link's coefficient and the neighbor-joining method. Most accessions analyzed grouped into two major clusters: the first comprised most accessions of A. glabrata and accessions of A. nitida, and the second cluster comprised accessions of A. burkartii. Arachis pseudovillosa and a few accessions of A. glabrata and A. nitida were placed between these major clusters. The diploid and tetraploid species were grouped quite separately, suggesting that the tetraploids did not originate from the diploid species analyzed.

Solubilization profiles of metal ions from bioleaching of sewage sludge as a function of pH

Two patterns of solubilization of metal ions resulting from bioleaching of sewage sludge by sulphur-oxidizing Thiobacillus spp. were established as a function of pH. Chromium and copper ions required a pH of 2-3 to initiate their solubilization, whereas nickel and zinc ions had their solubilization initiated at pH 6-6.5. The patterns obtained were independent of the sludge solids concentrations investigated (10, 17, 25, 32.5 and 40 g l(-1)).

Identification of a molecular marker linked to apomixis in Brachiaria humidicola (Poaceae)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Relationships among Pratylenchus jaehni and P. coffeae populations from Brazil.

Wilcken, S.R.S., E.S. Mori, M. Bacci, L.C.C.B. Ferraz, C.M.G. Oliveira & M.M. Inomoto. 2008. Relationships among Pratylenchus jaehni and P. coffeae populations from Brazil. The relationships among Pratylenchus jaehni (C) and six amphimitic Pratylenchus populations from Brazil (three from banana, PcB1, PcB2, and PcB3; one from Aglaonema sp., M2; one from coffee, K5; and one from citrus, C) were studied through morphological and molecular analysis, and pathogenicity test. The analysis of morphometric characteristics of PcB1, M2, C2 and K5, compared with P. jaehni and P. coffeae (K6) data obtained from literature, delineated three groups: P. coffeae (K6), PcB1 and M2; P. jaehni (C1) and C2; and K5. RAPD analysis of PcB1, PcB2, PcB3, M2, C2, K5 and P.jaehni (C1) demonstrated that these populations form three distinct genetic groups: PcB1, PcB2, PcB3 and M2; P.jaehni and C2; and K5. In pathogenicity test, K5 reproduced well on coffee and caused extensive root necrosis, but C2 did not. Population C2 reproduced well on Rangpur lime, which was previously rated as poor host to K5. Therefore, the results demonstrated that C2 is P. jaehni, K5 is an undescribed species of Pratylenchus, and PcB1, PcB2 and PcB3 are P. coffeae. The taxonomic status of M2 remains inconclusive.

Genetic variation between several species of sections Extranervosae, Caulorrhizae, Heteranthae, and Triseminatae (genus Arachis) estimated by DNA polymorphism

Genetic variation within and among accessions of the genus Arachis representing sections Extranervosae, Caulorrhizae, Heteranthae, and Triseminatae was evaluated using RFLP and RAPD markers. RAPD markers revealed a higher level of genetic diversity than did RFLP markers, both within and among the species evaluated. Phenograms based on various band-matching algorithms revealed three major clusters of similarity among the sections evaluated. The first group included the species from section Extranervosae, the second group consisted of sections Triseminatae, Caulorrhizae, and Heteranthae, and the third group consisted of one accession of Arachis hypogaea, which had been included as a representative of section Arachis. The phenograms obtained from the RAPD and RFLP data were similar but not identical. Arachis pietrarellii, assayed only by RAPD, showed a high degree of genetic similarity with Arachis villosulicarpa. This observation supported the hypothesis that these two species are closely related. It was also shown that accession V 7786, previously considered to be Arachis sp. aff. pietrarellii, and assayed using both RFLPs and RAPDs, was possibly a new species from section Extranervosae, but very distinct from A. pietrarellii.

Evaluation of microsatellite markers in cultivars of sweet orange (Citrus sinensis [L.] Osbeck)

Sweet orange is considered a very important species in the citrus world market and presents wide morphological variability. However, its characterization at the molecular level by random amplified polymorphic DNA (RAPD) and isozyme markers is not appropriate. Microsatellite or simple sequence repeats (SSRs) have been suggested as ideal for studies in cultures of vegetative propagation and as value markers for mapping in several species. However, information on microsatellite polymorphism in citrus species is scarce. In this work, microsatellite markers (AG-repeats) were developed from an enrichment library of genomic DNA of sweet orange cv. Pera (Citrus sinensis [L.] Osbeck), and 31 cultivars of sweet orange were evaluated. Evaluation of 18 microsatellite primers did not permit differentiation of the varieties studied. New microsatellite primers are being evaluated with the aim of detecting polymorphisms among the cultivars and closely related species to be used in genetic mapping programs.

Characterization of Rhizoctonia solani associated with soybean in Brazil

Rhizoctonia solani causes pre- and post-emergence damping-off, root and hypocotyl rot and foliar blight in soybean. Foliar blight has resulted in yield losses of 31-60% in north and northeast Brazil. The aim of this study was to characterize isolates of R. solani associated with soybean in Brazil. Among 73 Rhizoctonia isolates examined, six were binucleate and 67 were multinucleate. The multinucleate isolates were characterized according to hyphal anastomosis reaction, mycelial growth rate, thiamine requirement, sclerotia production, and RAPD molecular markers. Four isolates that caused hypocotyl rot belonged to AG-4 and using RAPD analysis they grouped together with the HGI subgroup. Another isolate that caused root and hypocotyl rots was thiamine auxotrophic, grew at 35 °C, and belonged to AG-2-2 IIIB. All 62 isolates that caused foliar blight belonged to AG-1 IA. RAPD analysis of R. solani AG-1 IA soybean isolates showed high genetic similarity to a tester strain of AG-1 IA, confirming their classification. The teleomorph of R. solani, Thanatephorus cucumeris was produced in vitro by one AG-1 IA isolate from soybean. The AG-4 and AG-2-2 IIIB isolates caused damping-off and root and hypocotyl rots of soybean seedlings cv. 'FT-Cristalina', under greenhouse conditions. The AG-2-2 IIIB isolate caused large lesions on the cortex tissue, that was distinct from the symptoms caused by AG-4 isolates. The AG-1 IA isolates caused foliar blight in adult soybean plants cv. 'Xingu' under the greenhouse and also in a detached-leaf assay.