Characterization and Pathogenicity of Vibrio cholerae and Vibrio vulnificus from Marine environments

The genus Vibrioof the family Vibrionaceae are Gram negative, oxidasepositive, rod- or curved- rodshaped facultative anaerobes, widespread in marine and estuarine environments. Vibrio species are opportunistic human pathogens responsible for diarrhoeal disease, gastroenteritis, septicaemia and wound infections and are also pathogens of aquatic organisms, causing infections to crustaceans, bivalves and fishes. In the present study, marine environmental samples like seafood and water and sediment samples from aquafarms and mangroves were screened for the presence of Vibrio species. Of the134 isolates obtained from the various samples, 45 were segregated to the genus Vibrio on the basis of phenotypic characterization.like Gram staining, oxidase test, MoF test and salinity tolerance. Partial 16S rDNA sequence analysis was utilized for species level identification of the isolates and the strains were identified as V. cholerae(N=21), V. vulnificus(N=18), V. parahaemolyticus(N=3), V. alginolyticus (N=2) and V. azureus (N=1). The genetic relatedness and variations among the 45 Vibrio isolates were elucidated based on 16S rDNA sequences. Phenotypic characterization of the isolates was based on their response to 12 biochemical tests namely Voges-Proskauers’s (VP test), arginine dihydrolase , tolerance to 3% NaCl test, ONPG test that detects β-galactosidase activity, and tests for utilization of citrate, ornithine, mannitol, arabinose, sucrose, glucose, salicin and cellobiose. The isolates exhibited diverse biochemical patterns, some specific for the species and others indicative of their environmental source.Antibiogram for the isolates was determined subsequent to testing their susceptibility to 12 antibiotics by the disc diffusion method. Varying degrees of resistance to gentamycin (2.22%), ampicillin(62.22%), nalidixic acid (4.44%), vancomycin (86.66), cefixime (17.77%), rifampicin (20%), tetracycline (42.22%) and chloramphenicol (2.22%) was exhibited. All the isolates were susceptible to streptomycin, co-trimoxazole, trimethoprim and azithromycin. Isolates from all the three marine environments exhibited multiple antibiotic resistance, with high MAR index value. The molecular typing methods such as ERIC PCR and BOX PCR revealed intraspecies relatedness and genetic heterogeneity within the environmental isolatesof V. cholerae and V. vulnificus. The 21 strains of V. choleraewere serogroupedas non O1/ non O139 by screening for the presence O1rfb and O139 rfb marker genes by PCR. The virulence/virulence associated genes namely ctxA, ctxB, ace, VPI, hlyA, ompU, rtxA, toxR, zot, nagst, tcpA, nin and nanwere screened in V. cholerae and V. vulnificusstrains.The V. vulnificusstrains were also screened for three species specific genes viz., cps, vvhand viu. In V. cholerae strains, the virulence associated genes like VPI, hlyA, rtxA, ompU and toxR were confirmed by PCR. All the isolates, except for strain BTOS6, harbored at least one or a combination of the tested genes and V. choleraestrain BTPR5 isolated from prawn hosted the highest number of virulence associated genes. Among the V. vulnificusstrains, only 3 virulence genes, VPI, toxR and cps, were confirmed out of the 16 tested and only 7 of the isolates had these genes in one or more combinations. Strain BTPS6 from aquafarm and strain BTVE4 from mangrove samples yielded positive amplification for the three genes. The toxRgene from 9 strains of V. choleraeand 3 strains of V. vulnificus were cloned and sequenced for phylogenetic analysis based on nucleotide and the amino acid sequences. Multiple sequence alignment of the nucleotide sequences and amino acid sequences of the environmental strains of V. choleraerevealed that the toxRgene in the environmental strains are 100% homologous to themselves and to the V. choleraetoxR gene sequence available in the Genbank database. The 3 strains of V. vulnificus displayed high nucleotide and amino acid sequence similarity among themselves and to the sequences of V. cholerae and V. harveyi obtained from the GenBank database, but exhibited only 72% homology to the sequences of its close relative V. vulnificus. Structure prediction of the ToxR protein of Vibrio cholerae strain BTMA5 was by PHYRE2 software. The deduced amino acid sequence showed maximum resemblance with the structure of DNA-binding domain of response regulator2 from Escherichia coli k-12 Template based homology modelling in PHYRE2 successfully modelled the predicted protein and its secondary structure based on protein data bank (PDB) template c3zq7A. The pathogenicity studies were performed using the nematode Caenorhabditiselegansas a model system. The assessment of pathogenicity of environmental strain of V. choleraewas conducted with E. coli strain OP50 as the food source in control plates, environmental V. cholerae strain BTOS6, negative for all tested virulence genes, to check for the suitability of Vibrio sp. as a food source for the nematode;V. cholerae Co 366 ElTor, a clinical pathogenic strain and V. cholerae strain BTPR5 from seafood (Prawn) and positive for the tested virulence genes like VPI, hlyA, ompU,rtxA and toxR. It was found that V. cholerae strain BTOS6 could serve as a food source in place of E. coli strain OP50 but behavioral aberrations like sluggish movement and lawn avoidance and morphological abnormalities like pharyngeal and intestinal distensions and bagging were exhibited by the worms fed on V. cholerae Co 366 ElTor strain and environmental BTPR5 indicating their pathogenicity to the nematode. Assessment of pathogenicity of the environmental strains of V. vulnificus was performed with V. vulnificus strain BTPS6 which tested positive for 3 virulence genes, namely, cps, toxRand VPI, and V. vulnificus strain BTMM7 that did not possess any of the tested virulence genes. A reduction was observed in the life span of worms fed on environmental strain of V. vulnificusBTMM7 rather than on the ordinary laboratory food source, E. coli OP50. Behavioral abnormalities like sluggish movement, lawn avoidance and bagging were also observed in the worms fed with strain BTPS6, but the pharynx and the intestine were intact. The presence of multi drug resistant environmental Vibrio strainsthat constitute a major reservoir of diverse virulence genes are to be dealt with caution as they play a decisive role in pathogenicity and horizontal gene transfer in the marine environments.

Iron Uptake and Homeostasis in Microorganisms

Preface. Iron is considered to be a minor element employed, in a variety of forms, by nearly all living organisms. In some cases, it is utilised in large quantities, for instance for the formation of magnetosomes within magnetotactic bacteria or during use of iron as a respiratory donor or acceptor by iron oxidising or reducing bacteria. However, in most cases the role of iron is restricted to its use as a cofactor or prosthetic group assisting the biological activity of many different types of protein. The key metabolic processes that are dependent on iron as a cofactor are numerous; they include respiration, light harvesting, nitrogen fixation, the Krebs cycle, redox stress resistance, amino acid synthesis and oxygen transport. Indeed, it is clear that Life in its current form would be impossible in the absence of iron. One of the main reasons for the reliance of Life upon this metal is the ability of iron to exist in multiple redox states, in particular the relatively stable ferrous (Fe2+) and ferric (Fe3+) forms. The availability of these stable oxidation states allows iron to engage in redox reactions over a wide range of midpoint potentials, depending on the coordination environment, making it an extremely adaptable mediator of electron exchange processes. Iron is also one of the most common elements within the Earth’s crust (5% abundance) and thus is considered to have been readily available when Life evolved on our early, anaerobic planet. However, as oxygen accumulated (the ‘Great oxidation event’) within the atmosphere some 2.4 billion years ago, and as the oceans became less acidic, the iron within primordial oceans was converted from its soluble reduced form to its weakly-soluble oxidised ferric form, which precipitated (~1.8 billion years ago) to form the ‘banded iron formations’ (BIFs) observed today in Precambrian sedimentary rocks around the world. These BIFs provide a geological record marking a transition point away from the ancient anaerobic world towards modern aerobic Earth. They also indicate a period over which the bio-availability of iron shifted from abundance to limitation, a condition that extends to the modern day. Thus, it is considered likely that the vast majority of extant organisms face the common problem of securing sufficient iron from their environment – a problem that Life on Earth has had to cope with for some 2 billion years. This struggle for iron is exemplified by the competition for this metal amongst co-habiting microorganisms who resort to stealing (pirating) each others iron supplies! The reliance of micro-organisms upon iron can be disadvantageous to them, and to our innate immune system it represents a chink in the microbial armour, offering an opportunity that can be exploited to ward off pathogenic invaders. In order to infect body tissues and cause disease, pathogens must secure all their iron from the host. To fight such infections, the host specifically withdraws available iron through the action of various iron depleting processes (e.g. the release of lactoferrin and lipocalin-2) – this represents an important strategy in our defence against disease. However, pathogens are frequently able to deploy iron acquisition systems that target host iron sources such as transferrin, lactoferrin and hemoproteins, and thus counteract the iron-withdrawal approaches of the host. Inactivation of such host-targeting iron-uptake systems often attenuates the pathogenicity of the invading microbe, illustrating the importance of ‘the battle for iron’ in the infection process. The role of iron sequestration systems in facilitating microbial infections has been a major driving force in research aimed at unravelling the complexities of microbial iron transport processes. But also, the intricacy of such systems offers a challenge that stimulates the curiosity. One such challenge is to understand how balanced levels of free iron within the cytosol are achieved in a way that avoids toxicity whilst providing sufficient levels for metabolic purposes – this is a requirement that all organisms have to meet. Although the systems involved in achieving this balance can be highly variable amongst different microorganisms, the overall strategy is common. On a coarse level, the homeostatic control of cellular iron is maintained through strict control of the uptake, storage and utilisation of available iron, and is co-ordinated by integrated iron-regulatory networks. However, much yet remains to be discovered concerning the fine details of these different iron regulatory processes. As already indicated, perhaps the most difficult task in maintaining iron homeostasis is simply the procurement of sufficient iron from external sources. The importance of this problem is demonstrated by the plethora of distinct iron transporters often found within a single bacterium, each targeting different forms (complex or redox state) of iron or a different environmental condition. Thus, microbes devote considerable cellular resource to securing iron from their surroundings, reflecting how successful acquisition of iron can be crucial in the competition for survival. The aim of this book is provide the reader with an overview of iron transport processes within a range of microorganisms and to provide an indication of how microbial iron levels are controlled. This aim is promoted through the inclusion of expert reviews on several well studied examples that illustrate the current state of play concerning our comprehension of how iron is translocated into the bacterial (or fungal) cell and how iron homeostasis is controlled within microbes. The first two chapters (1-2) consider the general properties of microbial iron-chelating compounds (known as ‘siderophores’), and the mechanisms used by bacteria to acquire haem and utilise it as an iron source. The following twelve chapters (3-14) focus on specific types of microorganism that are of key interest, covering both an array of pathogens for humans, animals and plants (e.g. species of Bordetella, Shigella, , Erwinia, Vibrio, Aeromonas, Francisella, Campylobacter and Staphylococci, and EHEC) as well as a number of prominent non-pathogens (e.g. the rhizobia, E. coli K-12, Bacteroides spp., cyanobacteria, Bacillus spp. and yeasts). The chapters relay the common themes in microbial iron uptake approaches (e.g. the use of siderophores, TonB-dependent transporters, and ABC transport systems), but also highlight many distinctions (such as use of different types iron regulator and the impact of the presence/absence of a cell wall) in the strategies employed. We hope that those both within and outside the field will find this book useful, stimulating and interesting. We intend that it will provide a source for reference that will assist relevant researchers and provide an entry point for those initiating their studies within this subject. Finally, it is important that we acknowledge and thank wholeheartedly the many contributors who have provided the 14 excellent chapters from which this book is composed. Without their considerable efforts, this book, and the understanding that it relays, would not have been possible. Simon C Andrews and Pierre Cornelis

Differential induction of apoptosis in human colonic carcinoma cells (Caco-2) by Atopobium, and commensal, probiotic and enteropathogenic bacteria: mediation by the mitochondrial pathway

The induction of apoptosis in mammalian cells by bacteria is well reported. This process may assist infection by pathogens whereas for non-pathogens apoptosis induction within carcinoma cells protects against colon cancer. Here, apoptosis induction by a major new gut bacterium, Atopobium minutum, was compared with induction by commensal (Escherichia coli K-12 strains), probiotic (Lactobacillus rhamnosus, Bifidobacterium latis) and pathogenic (E. coli: EPEC and VTEC) gut bacteria within the colon cancer cell line, Caco-2. The results show a major apoptotic effect for the pathogens, mild effects for the probiotic strains and A. minutum, but no effect for commensal E. coli. The mild apoptotic effects observed are consistent with the beneficial roles of probotics in protection against colon cancer and suggest, for the first time, that A. minutum possesses similar advantageous, anti-cancerous activity. Although bacterial infection increased Caco-2 membrane FAS levels, caspase-8 was not activated indicating that apoptosis is FAS independent. Instead, in all cases, apoptosis was induced through the mitochondrial pathway as indicated by BAX translocation, cytorchrome c release, and caspase-9 and -3 cleavage. This suggests that an intracellular stimulus initiates the observed apoptosis responses.

Exposure of Escherichia coli and Salmonella enterica serovar Typhimurium to triclosan induces a species-specific response, including drug detoxification

Objectives: The use of triclosan within various environments has been linked to the development of multiple drug resistance (MDR) through the increased expression of efflux pumps such as AcrAB-ToIC. In this work, we investigate the effect of triclosan exposure in order to ascertain the response of two species to the presence of this widely used biocide. Methods: The transcriptomes of Salmonella enterica serovar Typhimurium SL1344 and Escherichia coli K-12 MG1655 after exposure to the MIC of triclosan (0.12 mg/L) were determined in microarray experiments. Phenotypic validation of the transcriptomic data included RT-PCR, ability to form a biofilm and motility assays. Results: Despite important differences in the triclosan-dependent transcriptomes of the two species, increased expression of efflux pump component genes was seen in both. Increased expression of soxS was observed in Salmonella Typhimurium, however, within E. coli, decreased expression was seen. Expression of fabBAGI in Salmonella Typhimurium was decreased, whereas in E. coli expression of fabABFH was increased. Increased expression of ompR and genes within this regulon (e.g. ompC, csgD and ssrA) was seen in the transcriptome of Salmonella Typhimurium. An unexpected response of E. coli was the differential expression of genes within operons involved in iron homeostasis; these included fhu, fep and ent. Conclusions: These data indicate that whilst a core response to triclosan exposure exists, the differential transcriptome of each species was different. This suggests that E. coli K-12 should not be considered the paradigm for the Enterobacteriaceae when exploring the effects of antimicrobial agents.

Genetic diversity among Escherichia coli O157 : H7 isolates and identification of genes linked to human infections

An Escherichia coli oligonucleotide microarray based on three sequenced genomes was validated for comparative genomic microarray hybridization and used to study the diversity of E. coli O157 isolates from human infections and food and animal sources. Among 26 test strains, 24 (including both Shiga toxin [Stx]-positive and -negative strains) were found to be related to the two sequenced E. coli O157:117 strains, EDL933 and Sakai. However, these strains showed much greater genetic diversity than those reported previously, and most of them could not be categorized as either lineage I or H. Some genes were found more often in isolates from human than from nonhuman sources; e.g., ECs1202 and ECs2976, associated with stx2AB and stx1AB, were in all isolates from human sources but in only 40% of those from nonhuman sources. Some (but not all) lineage I-specific or -dominant genes were also more frequently associated with isolates from human. The results suggested that it might be more effective to concentrate our efforts on finding markers that are directly related to infection rather than those specific to certain lineages. In addition, two Stx-negative O157 cattle isolates (one confirmed to be 117) were significantly different from other Stx-positive and -negative E. coli O157:117 strains and were more similar to MG1655 in their gene content. This work demonstrates that not all E. coli O157:117 strains belong to the same clonal group, and those that were similar to E. coli K-12 might be less virulent.

Comparative genomic indexing reveals the phylogenomics of Escherichia coli pathogens

The Escherichia coli O26 serogroup includes important food-borne pathogens associated with human and animal diarrheal disease. Current typing methods have revealed great genetic heterogeneity within the O26 group; the data are often inconsistent and focus only on verotoxin (VT)-positive O26 isolates. To improve current understanding of diversity within this serogroup, the genomic relatedness of VT-positive and -negative O26 strains was assessed by comparative genomic indexing. Our results clearly demonstrate that irrespective of virulence characteristics and pathotype designation, the O26 strains show greater genomic similarity to each other than to any other strain included in this study. Our data suggest that enteropathogenic and VT-expressing E. coli O26 strains represent the same clonal lineage and that W-expressing E. coli O26 strains have gained additional virulence characteristics. Using this approach, we established the core genes which are central to the E. coli species and identified regions of variation from the E. coli K-12 chromosomal backbone.

Identification of core and variable components of the Salmonella enterica subspecies I genome by microarray

We have performed microarray hybridization studies on 40 clinical isolates from 12 common serovars within Salmonella enterica subspecies I to identify the conserved chromosomal gene pool. We were able to separate the core invariant portion of the genome by a novel mathematical approach using a decision tree based on genes ranked by increasing variance. All genes within the core component were confirmed using available sequence and microarray information for S. enterica subspecies I strains. The majority of genes within the core component had conserved homologues in Escherichia coli K-12 strain MG1655. However, many genes present in the conserved set which were absent or highly divergent in K-12 had close homologues in pathogenic bacteria such as Shigella flexneri and Pseudomonas aeruginosa. Genes within previously established virulence determinants such as SPI1 to SPI5 were conserved. In addition several genes within SPI6, all of SPI9, and three fimbrial operons (fim, bcf, and stb) were conserved within all S. enterica strains included in this study. Although many phage and insertion sequence elements were missing from the core component, approximately half the pseudogenes present in S. enterica serovar Typhi were conserved. Furthermore, approximately half the genes conserved in the core set encoded hypothetical proteins. Separation of the core and variant gene sets within S. enterica subspecies I has offered fundamental biological insight into the genetic basis of phenotypic similarity and diversity across S. enterica subspecies I and shown how the core genome of these pathogens differs from the closely related E. coli K-12 laboratory strain.

Magnetic structures of quaternary intermetallic borocarbides RCo(2)B(2)C (R = Dy, Ho, Er)

The magnetic structures of the title compounds have been studied by neutron diffraction. In contrast to the isomorphous RNi(2)B(2)C compounds, wherein a variety of exotic incommensurate modulated structures has been observed, the magnetic structure of ErCo(2)B(2)C is found to be a collinear antiferromagnet with k = (12, 0, 12) while those of HoCo(2)B(2)C and DyCo(2)B(2)C are observed to be simple ferromagnets. For all studied compounds, the moments are found to be confined within the basal plane and their magnitudes are comparable to the values obtained from the low-temperature isothermal magnetization measurements. The absence of modulated magnetic structures in the RCo(2)B(2)C series (for ErCo(2)B(2)C, verified down to 50 mK) is attributed to the quenching of the Fermi surface nesting features.

Search for squeezed-pair correlations at RHIC

Squeezed correlations of particle-antiparticle pairs, also called back-to-back correlations (BBC), are predicted to appear if the hadron masses are modified in the hot and dense hadronic medium formed in high energy nucleus nucleus collisions. Although well established theoretically, the squeezed-particle correlations have not yet been searched for experimentally in high energy hadronic or heavy ion collisions, clearly requiring optimized forms to experimentally search for this effect. Within a non-relativistic treatment developed earlier we show that one promising way to search for the BBC signal is to look into the squeezed correlation function of pairs of phi's at RHIC energies, plotted in terms of the average momentum of the pair, K(12) = 1/2 (k(1) + k(2)). This variable's modulus, 2 vertical bar K(12)vertical bar, is the non-relativistic limit of the variable Q(bbc), introduced herewith. Some squeezing effects on the HBT correlation function are also discussed.

Squeezed correlations among particle-antiparticle pairs

The hadronic correlation among particle-antiparticle pairs was highlighted in the late 1990's, culminating with the demonstration that it should exist if the masses of the hadrons were modified in the hot and dense medium formed in high energy heavy ion collisions. They were called Back-to-Back Correlations (BBC) of particle-antiparticle pairs, also known as squeezed correlations. However, even though they are well-established theoretically, such hadronic correlations have not yet been experimentally discovered. Expecting to compel the experimentalists to search for this effect, we suggest here a clear way to look for the BBC signal, by constructing the squeezed correlation function of phi phi and K(+)K(-) pairs at RHIC energies, plotted in terms of the average momentum of the pair, K(12)=1/2(k(1) + k(2)), inspired by procedures adopted in Hanbury-Brown & Twiss (HBT) correlations.

SQUEEZED PARTICLE-ANTIPARTICLE CORRELATIONS

A novel type of correlation involving particle-antiparticle pairs was found out in the 1990's. Currently known as squeezed or Back-to-Back Correlations (BBC), they should be present if the hadronic masses are modified in the hot and dense medium formed in high energy heavy ion collisions. Although well-established theoretically, such hadronic correlations have not yet been observed experimentally. In this phenomenological study we suggest a promising way to search for the BBC signal, by looking into the squeezed correlation function of phi phi and K(+)K(-) pairs at RHIC energies, as function of the pair average momentum, K(12) = (k(1) + k(2))/2. The effects of in-medium mass-shift on the identical particle correlations (Hanbury-Brown and Twiss effect) are also discussed.

A Chloro-Bridged Linear Chain Imine-Copper(II) Complex and Its Application as an Enzyme-Free Amperometric Biosensor for Hydrogen Peroxide

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Nutrição e produtividade do crambe em razão da adubação de semeadura na safrinha

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Paleomagnetismo de rochas vulvânicas do Nordeste do Brasil e a época da abertura do Oceano Atlântico Sul

Na primeira parte deste trabalho foram desenvolvidos estudos de magnetismo de rochas e paleomagnetismo em amostras de rochas vulcânicas do Nordeste brasileiro. As idades das amostras compreende os períodos Jurássico e Cretáceo. Com este objetivo foram amostradas quatro áreas tendo sido estudado um total de 496 amostras em 55 sítios. Para a coleta foi utilizada uma perfuradora portátil que extrai amostras de 2.5 cm de diâmetro. A orientação das amostras foi feita por meio de uma bússola magnética e de um clinômetro. Os espécimes foram submetidos a desmagnetizações por campo magnético alternado e em alguns poucos casos foi empregada a desmagnetização térmica. Atribuindo-se peso unitário a cada sítio foi determinada a direção média da magnetização remanescente característica de cada uma das áreas estudadas. As rochas vulcânicas do período Jurássico, localizadas na borda oeste da Bacia do Maranhão (Porto Franco-Estreito), apresentaram uma direção media em que D= 3.9°, I= -17.9° com α₉₅= 9.3°, k= 17.9, N= 15 e todos os sítios apresentaram polaridade normal. Para esta área foi determinado o polo paleomagnético de coordenadas 85.3°N, 82.5°E (A₉₅= 6.9º) que se localiza próximo a outros polos paleomagnéticos conhecidos para esse período. As rochas da borda leste da Bacia do Maranhão (Teresina-Picos-Floriano) de idade cretácica inferior apresentaram uma direção média de magnetização remanescente característica tal que D= 174.7°, I= +6.0º com α₉₅= 2.8º, k= 122, N= 21 e todos os sítios apresentaram polaridade reversa. O polo paleomagnético associado a elas apresentou por coordenadas 83.6°N, 261.0°E (A₉₅=1.9°) e mostrou concordância com outros polos sul americanos de mesma idade. No Rio Grande do Norte foi estudado um enxame de diques toleíticos também de idade cretácica inferior, cuja direção média da magnetização remanescente característica encontrada foi D= 186.6º, I= +20.6º com α₉₅= 14.0° e k= 12.9, N= 10. Os sítios desta área apresentaram magnetizações com polaridades normal a reversa. O polo paleomagnético obtido se localiza em 80.6°N e 94.8°E com A₉₅= 9.5°. O estudo das rochas vulcânicas da província magnética do Cabo de Santo Agostinho indicou para a região um valor de D= 0.4º, I= -20.6º com α₉₅= 4.8° e k= 114, N= 9 para a magnetização remanescente característica. Todos os sítios apresentaram polaridade normal e o polo paleomagnético determinado apresentou as seguintes coordenadas: 87.6ºN, 135ºE com A₉₅= 4.5º. Foi discutida a eliminação da variação secular das direções obtidas, de forma que cada polo apresentado nesta dissertação é verdadeiramente um polo paleomagnético. A análise dos minerais magnéticos portadores da remanência, efetuada por curvas termomagnéticas ou por difração de Raio-X, indicou na maior parte das ocorrências, a presença de titanomagnetita pobre em titânio. A presença de maguemita e algumas vezes hematita, na maior parte das vezes resultado de intemperismo, não anulou a magnetização termoremanente associada à época de formação da rocha, que foi determinada após a aplicação de técnicas de desmagnetização aos espécimes. Pelas curvas termomagnéticas obteve-se, para a maioria das amostras, uma temperatura de Curie entre 500 e 600ºC. Os casos mais freqüentes indicaram a ocorrência de titanomagnetita exsolvida, em que foram observadas a presença de uma fase próxima à magnetita e outra fase rica em titânio, próxima à ilmenita, resultado de oxidação de alta temperatura. A segunda parte do trabalho diz respeito à determinação da época de abertura do oceano Atlântico Sul por meio de dados paleomagnéticos. Entretanto ao invés de se utilizar o procedimento comumente encontrado na literatura, e que se baseia nas curvas de deriva polar aparente de cada continente, foi aplicado um teste estatístico que avalia a probabilidade de determinada posição relativa entre os continentes ser válida ou não, para determinado período em estudo. Assim foi aplicado um teste F a polos paleomagnéticos da África e da América do Sul, dos períodos Triássico, Jurássico, Cretáceo Inferior e Cretáceo Médio-Superior, tendo sido estudadas situações que reconstituem a posição pré-deriva dos continentes e configurações que simulem um afastamento entre eles. Os resultados dos testes estatísticos indicaram, dentro de uma probabilidade de erro de menos de 5%, que a configuração pré-deriva de Martin et al (1981) é compatível com os dados paleomagnéticos do Triássico, mas apresenta uma diferença significativa para os paleopolos de Jurássico, Cretáceo Inferior, Cretáceo Médio-Superior. Outras reconstruções pré-deriva testadas apresentaram o mesmo resultado. A comparação entre os polos paleomagnéticos da América do Sul e da África, segundo uma reconstrução que admite uma pequena abertura entre os continentes, como a proposta por Sclater et al (1977) para 110 m.a. atrás, indicou que os dados do Triássico não são compatíveis com este afastamento. Por outro lado os paleopolos do Jurássico e do Cretáceo Inferior, embora mais antigos que a data sugerida pela reconstrução, são consistentes com esta separação dentro de uma probabilidade de erro de menos de 5%. Os dados do Cretáceo Médio-Superior se mostraram consistentes com a reconstrução sugerida para 80 m.a. atrás por Francheteau (1973) e que propõe uma separação maior entre os continentes. Com base na premissa de deslocamentos de blocos continentais rígidos a análise dos resultados obtidos indicou que América do Sul e África estavam unidas por suas margens continentais opostas no período Triássico e que uma pequena separação entre estes continentes, provavelmente devida a uma rutura inicial, ocorreu no Jurássico e se manteve, então, aproximadamente estacionária até o início do Cretáceo Inferior. Esta conclusão difere da maior parte dos trabalhos que discutem a abertura do oceano Atlântico Sul. Os dados do Cretáceo Médio-Superior são compatíveis com um afastamento rápido e significativo entre os continentes naquele período.

A educação física no ensino médio em escolas particulares: as perspectivas de alunos, diretores, coordenadores e professores de educação física

Although physical education is regulated by law and recognized as a mandatory component of basic education class, which includes high school, there are many reports that these classes do not actually happen. Such evidence shows the situation experienced by the author of this study, which brought this research to the attention. Therefore, the objective was to investigate and understand the perspective of students, physical education teachers, principals and coordinators forward their ideas to this reality, about the high school physical education in private schools. The research made use of qualitative approach, guided by an exploratory study. The technique was undertaken to collect the questionnaire with open and closed questions, involving 263 students enrolled in high school of four schools in two cities in the state of Sao Paulo and also the director, coordinator and professor of physical education of each one of these institutions, resulting in 12 education professionals. The categories of analysis were defined as: 1. Regarding physical education in high school, 2. The occurrence of physical rducation in high school, 3. Content and dynamic strategies developed in class 4. Student participation in class. The results show that despite, although some schools have physical education classes, there are still many students who do not take them. This reveals a double challenge to the area, which is: although it is essential that such a curriculum component is included in the grades (respecting the regional law legal system) that is not enough students that actually participate in it. As teachers, coordinators and directors recognize the importance of effective physical education, there are numerous of factors that deserve consideration to better understand this reality. One relates to the content of sportivization, another contrast to the period of occurrence of classes in relation to other curriculum components. Additionally, some others are relat...