The extracellular matrix protein pattern in the canine neoplastic mammary gland.

Extracellular matrix (ECM) proteins in the mammary gland provide structure and regulate its development and homeostasis. Alterations in its structure can regulate and support pathogenesis, like breast tumors. Aiming to identify the health and tumoral canine mammary ECM scaffold protein profile by immunohistochemistry, the decellularization process was carried out to remove the cellular content. Additionally, it was verified the influence of health and tumoral ECM on the attachment of health and tumoral cells. The types I, III, IV, and V structural collagens were scarce in the mammary tumor, and ECM fibers were disorganized. Vimentin and CD44 were more common in mammary tumor stroma, suggesting a role in cell migration that results in tumor progression. Elastin, fibronectin, laminin, vitronectin, and osteopontin were similarly detected under healthy and tumor conditions, providing the attachment of normal cells in healthy ECM, while tumoral cells were able to attach in tumoral ECM. The protein pattern demonstrates ECM alteration in canine mammary tumorigenesis, presenting new knowledge on mammary tumor ECM microenvironment.

Reinstatement and phylogenetic allocation of the palm rust genus Cerradoa in the Pucciniaceae, and establishment of Pseudocerradoa, gen. nov.

The genus Cerradoa (type species Cerradoa palmaea) was established in 1978 by Hennen and Ono and named after the Brazilian Cerrado biome. The holotype collected in Planaltina, Federal District, Brazil, belonged to the first rust fungus reported on palms (Arecaceae). For decades, the status of Cerradoa as a distinct genus has been regarded as doubtful, representing a synonym of Edythea (Uropyxidaceae) starting with the second edition of the Illustrated Genera of Rust Fungi in 1983. Our molecular phylogenetic analyses, as well as our morphological investigations, allowed us to reject this synonymy, leading to the reinstatement of Cerradoa within the Pucciniaceae. Cerradoa, together with morphologically similar genera such as the newly established Pseudocerradoa with two species (Ps. paullula and Ps. rhaphidophorae) infecting araceous hosts, the fern rust Desmella, and also P. engleriana, could not be assigned to any of the seven identified major lineages within the Pucciniaceae. Edythea, instead of being maintained as a member of the Uropyxidaceae, was herein placed in Pucciniaceae, shown phylogenetically in close relationship to Cumminsiella mirabilissima, both infecting the Berberidaceae. Additionally, our extensive phylogenetic analyses add guidance for future taxonomic revisions in the highly polyphyletic genus Puccinia and other established taxa within the family Pucciniaceae.

A novel orchiectomy surgical procedure in donkeys ( Equus asinus africanus) with parascrotal access.

Donkeys are a public health concern in the Northeast region of Brazil, with thousands of stray animals. Orchiectomy is an important population control measure; however, the long postoperative period with daily treatment of open wounds in the scrotum makes it difficult to perform a large number of castrations in sheltering centers. We evaluate a novel surgical procedure for orchiectomy in donkeys using parascrotal access. Twelve donkeys were used, divided into two groups: I - submitted to orchiectomy through parascrotal surgical access (novel procedure), and II - submitted to orchiectomy through scrotal access (conventional). Postoperative evaluations consisted of a macroscopic evaluation of the surgical wound (bleeding and intensity of edema), hematological parameters, and peritoneal fluid, which occurred in both groups at the moments (M): M0 - before the surgical procedure. The others moments occurred after surgery: M12 (twelve hours); M24 (twenty-four hours); M48 (forty-eight hours); M72 (seventy-two hours); M8D (eight days); and M16D (sixteen days). The surgical techniques did not generate an important systemic inflammatory response to the point detected by the leukogram, fibrinogen dosage, and peritoneal fluid. The parascrotal technique required long surgery but promoted less bleeding, less edema, and faster healing. The techniques used did not promote sufficient systemic inflammation to alter the number of leukocytes and the fibrinogen concentration; however, evaluation of the peritoneal fluid proved to be important for evaluating inflammatory processes involving the scrotum and inguinal canal. We describe a novel surgical procedure for orchiectomy in Donkeys using a parascrotal access that promoted less risk of bleeding, shorter period of edema, and healing time, but required longer surgery time.

First report of Dematophora bunodes causing root rot of taro (Colocasia esculenta ) and leatherleaf fern (Rumohra adiantiformis) in Brazil.

Colocasia esculenta, taro (T), is a major staple food crop in the tropics, including Brazil. Rumohra adiantiformis, leatherhead fern (LF), is broadly cultivated for its ornamental fronds that are used as a component of flower arrangements. Soft root rot of T and LF, and accompanying rapid plant wilt and death, was observed in plantations in Espírito Santo (Brazil), at Venda Nova do Imigrante, in April 2014 (LF) and July 2015 (T). Great losses were observed. Firstly, a few individual scattered plants showed symptoms of disease in the plantations, then aggregates of plants and, after a few seasons, the majority of the plants in the field died before harvest, leading to the abandonment of the activity by farmers. A white mycelial matt was observed on the crown and roots ofying T and LF plants. Infected corms become necrotic and dark brown mycelial strands were observed internally in tissues. Diseased organs were carefully washed and surface sterilized in 10% sodium hypochlorite. Samples of tissue were removed from the boundary of necrotic tissues and placed on potato dextrose-agar (PDA) plates and incubated at 23±2 C in the dark. Homogeneous mycelial colonies were isolated from both T and LF and, upon observation of microscope mounts under an Olympus BX 53 light microscope, pear-shaped hyphal swellings at the septae (Castro et al. 2013) were observed. . A representative isolate from each host was deposited in the local culture collection as COAD 2911 (LF isolate) and COAD 2912 (T isolate). Additionally, DNA was extracted from each culture using the Wizard Genomic DNA Purification Kit (Promega) and the internal transcriptional spacer region was PCR amplified using the primers ITS5 and ITS1 (White et al. 1990). The amplicons were sequenced by MACROGEN (http://www.macrogen.com). Consensus sequences were deposited in GenBank: MW561595 (LF), MW561596 (T). Consensus regions were compared against other sequences available in Genbank. A BLASTn analysis resulted in LF and T sequences respectively 99% (526/531bp) and 98% (412/420 bp) identity with that of Dematophora bunodes (MN984619). Additionally, a phylogenetic analysis of a selected sequence alignment was performed on the CIPRES webportal (Miller et al., 2010) using MrBayes v.3.1.1 (Ronquist & Huelsenbeck, 2003). A phylogenetic tree was generated showing that the placement of LF and T isolates is in D. bunodes (Wittstein et al. 2020). Pathogenicity tests were performed for LF and T isolates against their original hosts. For inoculum, bags of twice-autoclaved parboiled rice were seeded separately with each isolate, which were allowed to colonize the rice for two weeks. Four healthy young LF and T plants were utilized. Two extra healthy plants grown in the same conditions, but not inoculated, served as controls. Thirty g of Dematophora-colonized rice was placed in direct contact with stems or roots of each LF or T plant. Plants were maintained in a dew chamber for 48 h after inoculation and then transferred to a greenhouse bench. All inoculated plants developed wilt and root rot and died after 15-20 days. Controls remained healthy. White mycelial colonies were formed over tissues of diseased LF and T and upon observation under the microscope, typical pear-shaped swellings were observed in slides prepared from newly obtained pure cultures from LF and T. Dematophora bunodes (formerly Rosellinia bunnodes) has a worldwide distribution and is well known as a polyphagous plant pathogen (Farr and Rossman, 2020) but has never been reported as a pathogen either of LF or T before in Brazil and worldwide. Its report on LF and T further expands an already large host-range and resolves the etiology of the disease on LF and T.

Utilização da lipoproteína de baixa densidade, em diferentes concentrações, em meio diluidor para criopreservação do sêmen ovino / Use of low density lipoprotein in different concentrations in diluent medium for cryopreservation of ovine semen

A utilização da gema de ovo dificulta a padronização de meios diluidores e apresenta riscos biológicos. Assim, este estudo avaliou diferentes concentrações da lipoproteína de baixa densidade (LDL), em substituição à gema de ovo, para a confecção de diluentes para criopreservação espermática em ovinos. Foram utilizados um diluidor controle (CTR= 20% de gema de ovo) e cinco tratamentos, substituindo-se a gema pelas diferentes proporções de LDL (T1=6%; T2=8%; T3=12%; T4=16%; T5=20%), todos à base de TRIS-glicerol. Para o estudo, utilizaram-se dois ejaculados, de seis reprodutores da raça Santa Inês. Sessenta dias após a criopreservação, as amostras foram descongeladas e avaliadas subjetivamente quanto à motilidade total (MT, %) e progressiva (MP, %), ao vigor (1-5) e à integridade funcional (choque hisposmótico com água destilada, %) e estrutural (corante supravital eosina, %) das membranas espermáticas. As avaliações de vigor e funcionalidade de membrana não diferiram (P>0,05) entre os grupos. Entretanto, os grupos T4 (P<0,01) e T5 (P<0,05) foram superiores ao CTR para os parâmetros MT, MP e integridade estrutural de membrana, o que confirma que as LDLs podem ser alternativas eficientes para substituição da gema de ovo em diluidores para criopreservação de sêmen ovino.(AU)

The use of egg yolk makes it difficult to standardize extenders and presents biological hazards. Thus, this study evaluated different concentrations of low-density lipoprotein (LDL) to replace yolk extenders for production of sperm for cryopreservation in ovine. A control extender was used (CTR= 20% yolk) and five treatments, replacing the yolk by different ratios of LDL (T1= 6%; T2= 8%, T3= 12%; T4= 16%; T5= 20%) all based on TRIS-glycerol. For the study, two ejaculates from six Santa Ines breeding were used. Sixty days after cryopreservation, the samples were thawed and evaluated for total motility (MT, %) and progressive motility (MP, %), vigor (1-5) and the functional integrity (hyposmotic shock with distilled water, %) and structural (supravital dye eosin, %) of the sperm membranes. The evaluations of strength and membrane functionality didn't differ (P> 0.05) between groups. However, T4 (P< 0.01) and T5 (P< 0.05) groups were superior to the CTR for the MT, MP, and membrane structural integrity parameters, which confirms that LDLs can be efficient alternatives for yolk replacement in extenders for cryopreservation of ovine semen.(AU)

Fungal Planet description sheets: 785-867.

Novel species of fungi described in this study include those from various countries as follows: Angola, Gnomoniopsis angolensis and Pseudopithomyces angolensis on unknown host plants. Australia, Dothiora corymbiae on Corymbia citriodora, Neoeucasphaeria eucalypti (incl. Neoeucasphaeria gen. nov.) on Eucalyptus sp., Fumagopsis stellae on Eucalyptus sp., Fusculina eucalyptorum (incl. Fusculinaceae fam. nov.) on Eucalyptus socialis, Harknessia corymbiicola on Corymbia maculata, Neocelosporium eucalypti (incl. Neocelosporium gen. nov., Neocelosporiaceae fam. nov. and Neocelosporiales ord. nov.) on Eucalyptus cyanophylla, Neophaeomoniella corymbiae on Corymbia citriodora, Neophaeomoniella eucalyptigena on Eucalyptus pilularis, Pseudoplagiostoma corymbiicola on Corymbia citriodora, Teratosphaeria gracilis on Eucalyptus gracilis, Zasmidium corymbiae on Corymbia citriodora. Brazil, Calonectria hemileiae on pustules of Hemileia vastatrix formed on leaves of Coffea arabica, Calvatia caatinguensis on soil, Cercospora solani-betacei on Solanum betaceum, Clathrus natalensis on soil, Diaporthe poincianellae on Poincianella pyramidalis, Geastrum piquiriunense on soil, Geosmithia carolliae on wing of Carollia perspicillata, Henningsia resupinata on wood, Penicillium guaibinense from soil, Periconia caespitosa from leaf litter, Pseudocercospora styracina on Styrax sp., Simplicillium filiforme as endophyte from Citrullus lanatus, Thozetella pindobacuensis on leaf litter, Xenosonderhenia coussapoae on Coussapoa floccosa. Canary Islands (Spain), Orbilia amarilla on Euphorbia canariensis. Cape Verde Islands, Xylodon jacobaeus on Eucalyptus camaldulensis. Chile, Colletotrichum arboricola on Fuchsia magellanica. Costa Rica, Lasiosphaeria miniovina on tree branch. Ecuador, Ganoderma chocoense on tree trunk. France, Neofitzroyomyces nerii (incl. Neofitzroyomyces gen. nov.) on Nerium oleander. Ghana, Castanediella tereticornis on Eucalyptus tereticornis, Falcocladium africanum on Eucalyptus brassiana, Rachicladosporium corymbiae on Corymbia citriodora. Hungary, Entoloma silvae-frondosae in Carpinus betulus-Pinus sylvestris mixed forest. Iran, Pseudopyricularia persiana on Cyperus sp. Italy, Inocybe roseascens on soil in mixed forest. Laos, Ophiocordyceps houaynhangensis on Coleoptera larva. Malaysia, Monilochaetes melastomae on Melastoma sp. Mexico, Absidia terrestris from soil. Netherlands, Acaulium pannemaniae, Conioscypha boutwelliae, Fusicolla septimanifiniscientiae, Gibellulopsis simonii, Lasionectria hilhorstii, Lectera nordwiniana, Leptodiscella rintelii, Parasarocladium debruynii and Sarocladium dejongiae (incl. Sarocladiaceae fam. nov.) from soil. New Zealand, Gnomoniopsis rosae on Rosa sp. and Neodevriesia metrosideri on Metrosideros sp. Puerto Rico, Neodevriesia coccolobae on Coccoloba uvifera, Neodevriesia tabebuiae and Alfaria tabebuiae on Tabebuia chrysantha. Russia, Amanita paludosa on bogged soil in mixed deciduous forest, Entoloma tiliae in forest of Tilia × europaea, Kwoniella endophytica on Pyrus communis. South Africa, Coniella diospyri on Diospyros mespiliformis, Neomelanconiella combreti (incl. Neomelanconiellaceae fam. nov. and Neomelanconiella gen. nov.) on Combretum sp., Polyphialoseptoria natalensis on unidentified plant host, Pseudorobillarda bolusanthi on Bolusanthus speciosus, Thelonectria pelargonii on Pelargonium sp. Spain, Vermiculariopsiella lauracearum and Anungitopsis lauri on Laurus novocanariensis, Geosmithia xerotolerans from a darkened wall of a house, Pseudopenidiella gallaica on leaf litter. Thailand, Corynespora thailandica on wood, Lareunionomyces loeiensis on leaf litter, Neocochlearomyces chromolaenae (incl. Neocochlearomyces gen. nov.) on Chromolaena odorata, Neomyrmecridium septatum (incl. Neomyrmecridium gen. nov.), Pararamichloridium caricicola on Carex sp., Xenodactylaria thailandica (incl. Xenodactylariaceae fam. nov. and Xenodactylaria gen. nov.), Neomyrmecridium asiaticum and Cymostachys thailandica from unidentified vine. USA, Carolinigaster bonitoi (incl. Carolinigaster gen. nov.) from soil, Penicillium fortuitum from house dust, Phaeotheca shathenatiana (incl. Phaeothecaceae fam. nov.) from twig and cone litter, Pythium wohlseniorum from stream water, Superstratomyces tardicrescens from human eye, Talaromyces iowaense from office air. Vietnam, Fistulinella olivaceoalba on soil. Morphological and culture characteristics along with DNA barcodes are provided.

Efeito da suplementação parenteral extra de cobre e zinco sobre a resposta imunológica de vacas Nelore / Effects of parenteral over-supplementation of cooper and zinc on immune response of Nellore cows

O objetivo deste estudo foi avaliar o efeito da suplementação mineral injetável extra de cobre (Cu) e zinco (Zn) sobre a resposta imunológica de vacas Nelore no período pré-parto. Foram avaliadas 60 vacas prenhes, as quais foram divididas em dois tratamentos, por meio da distribuição aleatória em delineamento inteiramente ao acaso. Aos 75 dias antes do parto, as vacas do tratamento testemunha (T) receberam soro fisiológico como placebo e os animais suplementados (S) receberam mineral injetável via subcutânea (75mg de cobre e 250mg de zinco, em dose única). Foram realizadas três coletas de sangue, duas antes da data prevista para o parto (75 e 10 dias) e uma 30 dias após o parto. Os teores de Cu, Zn, ceruloplasmina, imunoglobulinas G (IgG) e M (IgM) foram analisados durante as três coletas. A atividade fagocitária foi avaliada aos 30 dias pós-parto. Os dados foram examinados mediante análise de variância, com o uso do pacote estatístico do SAS, e os dados individuais da atividade fagocitária pelo PROC GLM. Os dados de Cu, Zn, IgG e IgM foram analisados como medidas repetidas no tempo de coleta por meio do PROC MIXED, com o nível de significância de 5%. Os teores de Cu, Zn, IgM, IgG, ceruloplasmina e a atividade fagocitária das vacas não sofreram influência dos tratamentos (P>0,05). O fornecimento de Cu e Zn injetável, nas doses utilizadas, aos 75 dias antes do parto para vacas Nelore, em dietas suficientes, não alterou os teores de Cu, Zn, ceruloplasmina e a resposta imunológica até 30 dias após o parto.(AU)

The aim of this study was to evaluate the effect of extra injectable mineral supplementation of copper (Cu) and zinc (Zn) on the immune response of Nellore cows in pre-partum period. Sixty pregnant cows were randomly distributed in a completely randomized design in two treatments. In the control treatment (T), cows received saline as placebo, and supplemented treatment (S) received mineral injection (75mg copper and 250mg of zinc, single dose) subcutaneously, 75 days prior to parturition. Blood was sampled three times, two before the expected date of parturition (75 and 10 days) and another at 30 days postpartum. Analyses were performed for Cu, Zn, ceruloplasmin, immunoglobulin G (IgG) and M (IgM) in the three periods and the phagocytic activity in the last period (30 days postpartum). The experimental data were subjected to analysis of variance using the statistical package SAS, being that the individual data phagocytic activity were analyzed by PROC GLM, and the Cu, Zn, IgG and IgM were analyzed as repeated measures in the time, using the PROC MIXED, with the significance level of 5%. The Cu, Zn, IgM, IgG, ceruloplasmin and the phagocytic activity of the cows were not affected by treatments (P>0.05). The supply of injectable Cu and Zn, at the doses used, 75 days before parturition to Nellore cows in sufficient diets, did not alter the serum contents of Cu, Zn, ceruloplasmin and the immune response up to 30 days after parturition.(AU)

Urbanization effects on the composition and structure of macrophytes communities in a lotic ecosystem of Pernambuco State, Brazil / Efeitos da urbanização na composição e estrutura de comunidades de macrófitas aquáticas em um ecossistema lótico do Estado de Pernambuco, Brasil

Abstract Population growth in urban areas changes freshwater ecosystems, and this can have consequences for macrophyte communities as can be seen in the municipalities that border the Capibaribe River, Pernambuco, Brazil. This study reports the effects of urbanization on the composition and structure of macrophyte communities in areas along that river. The following urbanized and non-urbanized sampling sites were chosen: Sites 1 and 2 (municipality of Santa Cruz do Capibaribe), Sites 3 and 4 (municipality of Toritama), and Sites 5 and 6 (metropolitan region of Recife). These sites were visited every two months from January to July 2013 to observe seasonal variation (wet and dry seasons). Thirty-one species were identified. Generally, the non-urbanized sites had a higher number of species. Multivariate analyses indicated significant overall differences between urbanized and non-urbanized areas (R = 0.044; p < 0.001) and between seasons (R = 0.018; p < 0.019). Owing to the large variation in physical, chemical, and biological characteristics between urbanized and non-urbanized areas, we found that urbanization significantly influenced the floristic composition and structure of macrophyte communities.

Resumo O crescimento populacional em áreas urbanas causa alterações em ecossistemas aquáticos continentais com consequência sobre as comunidades de macrófitas. Este fato vem ocorrendo nos municípios que margeiam o rio Capibaribe, Pernambuco, Brasil. Este trabalho analisa os efeitos da urbanização sobre a composição e estrutura das macrófitas em trechos do referido Rio. Levando em consideração áreas urbanizadas e não urbanizadas, foram escolhidos os seguintes Pontos de Coleta: Pontos 1 e 2 no Município de Santa Cruz do Capibaribe, Pontos 3 e 4 no Município de Toritama, Pontos 5 e 6 na Região Metropolitana do Recife. Estes pontos foram visitados bimestralmente (janeiro – julho/2013), para a observação da variação sazonal (estações seca e chuvosa). Foram identificadas 31 espécies. Geralmente, os pontos das áreas não urbanizadas apresentaram um número maior de espécies. As análises multivariadas indicaram diferenças globais significativas entre áreas urbanizadas e não urbanizadas (R = 0,044, p < 0,001) e também entre as estações (R = 0,018; p < 0,019). Devido à grande variação física, química e biológica entre as áreas urbanizadas e não urbanizadas, observou-se que o fator urbanização influenciou significativamente na composição florística e na estrutura das comunidades de macrófitas.

Neutrophil gelatinase-associated lipocalin is a biomarker of acute-on-chronic liver failure and prognosis in cirrhosis.

BACKGROUND & AIMS: Acute-on-chronic liver failure (ACLF) is a syndrome that occurs in cirrhosis characterized by organ failure(s) and high mortality rate. There are no biomarkers of ACLF. The LCN2 gene and its product, neutrophil gelatinase-associated lipocalin (NGAL), are upregulated in experimental models of liver injury and cultured hepatocytes as a result of injury by toxins or proinflammatory cytokines, particularly Interleukin-6. The aim of this study was to investigate whether NGAL could be a biomarker of ACLF and whether LCN2 gene may be upregulated in the liver in ACLF. METHODS: We analyzed urine and plasma NGAL levels in 716 patients hospitalized for complications of cirrhosis, 148 with ACLF. LCN2 expression was assessed in liver biopsies from 29 additional patients with decompensated cirrhosis with and without ACLF. RESULTS: Urine NGAL was markedly increased in ACLF vs. no ACLF patients (108(35-400) vs. 29(12-73)µg/g creatinine; p<0.001) and was an independent predictive factor of ACLF; the independent association persisted after adjustment for kidney function or exclusion of variables present in ACLF definition. Urine NGAL was also an independent predictive factor of 28day transplant-free mortality together with MELD score and leukocyte count (AUROC 0.88(0.83-0.92)). Urine NGAL improved significantly the accuracy of MELD in predicting prognosis. The LCN2 gene was markedly upregulated in the liver of patients with ACLF. Gene expression correlated directly with serum bilirubin and INR (r=0.79; p<0.001 and r=0.67; p<0.001), MELD (r=0.68; p<0.001) and Interleukin-6 (r=0.65; p<0.001). CONCLUSIONS: NGAL is a biomarker of ACLF and prognosis and correlates with liver failure and systemic inflammation. There is remarkable overexpression of LCN2 gene in the liver in ACLF syndrome. LAY SUMMARY: Urine NGAL is a biomarker of acute-on-chronic liver failure (ACLF). NGAL is a protein that may be expressed in several tissues in response to injury. The protein is filtered by the kidneys due to its small size and can be measured in the urine. Ariza, Graupera and colleagues found in a series of 716 patients with cirrhosis that urine NGAL was markedly increased in patients with ACLF and correlated with prognosis. Moreover, gene coding NGAL was markedly overexpressed in the liver tissue in ACLF.

Towards a phylogenetic reappraisal of Parmulariaceae and Asterinaceae (Dothideomycetes).

Members of the Asterinaceae and Parmulariaceae are obligate biotrophic fungi with a pantropical distribution that grow in direct association with living plant tissues and produce external ascomata and bitunicate asci. These fungi are poorly known, with limited information about their taxonomic position in the Dothideomycetes. Much of what is known is conjectural and based on observation of morphological characters. An assessment of the phylogenetic position of the Asterinaceae and Parmulariaceae is provided based on a phylogenetic analysis of the nrDNA operon (ITS) and the large subunit rDNA (LSU) sequence data obtained from fresh material of selected species collected in Brazil. Three key species were included and epitypified, namely Asterina melastomatis, which is the type species for the type genus of the Asterinaceae; Prillieuxina baccharidincola (Asterinaceae); and Parmularia styracis, which is the type species for the type genus of the Parmulariaceae. An LSU rDNA phylogenetic analysis was performed indicating the correct phylogenetic placement of the Asterinales within the Dothideomycetes. From this initial analysis it is clear that the Parmulariaceae as currently circumscribed is polyphyletic, and that the Asterinaceae and Parmulariaceae are related, which justifies the maintenance of the order Asterinales. Asterotexis cucurbitacearum is recognised as distinct from other Dothideomycetes and placed in the newly proposed family and order (Asterotexiaceae, Asterotexiales), while the higher order phylogeny of Inocyclus angularis remains unresolved. Additionally, Lembosia abaxialis is introduced as a novel species and the phylogenetic placement of the genera Batistinula and Prillieuxina is clarified.

First Report of Stigmina palmivora Causing Leaf Spots on Phoenix roebelenii in Brazil.

Phoenix roebelenii (Arecaceae), known as dwarf date (tamareira-anã in Brazil), is a palm native to Southeast Asia and widely cultivated worldwide because of its ornamental value and ease of adaptation to a broad range of climates and soil types (4). In June 2012, some individuals were observed in a private garden in the municipality of Viçosa (state of Minas Gerais, Brazil) bearing numerous necrotic lesions on its leaves. Representative samples were taken, dried in a plant press, and brought to the laboratory for examination. A fungus was regularly associated with the leaf spots. Fungal structures were mounted in lactophenol and slides were examined under a microscope (Olympus BX 51). Spores were taken from sporulating colonies with a sterile fine needle and plated on PDA for isolation. A pure culture was deposited in the culture collection of the Universidade Federal de Viçosa (accession COAD1338). A dried herbarium sample was deposited in the local herbarium (VIC39741). The fungus had the following morphology: conidiophores grouped on sporodochia, cylindrical, 12 to 29 × 5 to 6 µm, dark brown; conidiogenous cells, terminal, proliferating percurrently (annellidic), 8 to 20 × 5 to 6 µm, pale to dark brown; conidia obclavate to subcylindrical, straight, 58 to 147 × 5 to 6 µm, 6 to 16 septate, hila thickened and darkened with a thin-walled projecting papilla, dark brown, and verrucose. The morphology of the Brazilian collections agrees well with the description of Stigmina palmivora (2), a species known to cause leaf spots on P. roebelenii in the United States (Florida) and Japan (3). Pathogenicity was demonstrated through inoculation of leaves of healthy plants by placing 6 mm diameter cuture disks of COAD1338 on the leaf surface followed by incubation in a moist chamber for 48 h and then transferred to a greenhouse bench at 21 ± 3°C. Typical leaf spots were observed 15 days after inoculation. DNA was extracted from the isolate growing in pure culture and ITS and LSU sequences were generated and deposited in GenBank under the accession numbers KF656785 and KF656786, respectively. These were compared by BLASTn with other entries in GenBank, and the closest match for each region were Mycosphaerella colombiensis strain X215 and M. irregulariamosa strain CPC 1362 (EU514231, GU2114441) with 93% of nucleotide homology (over 100% query coverage) for ITS and 98% of nucleotide homology (over 100% query coverage) for LSU. There are no sequences for S. palmivora deposited in public databases for comparison, but for Stigmina platani, the type species in this genus, 86% and 96% nucleotide homology for ITS and LSU with S. palmivora were found. The genus Stigmina is regarded as being polyphyletic (1) and this is probably reflected by these low homology levels found in the BLASTn search. To our knowledge, this is the first report of Stigmina palmivora in Brazil. References: (1) P. W. Crous et al. Stud. Mycol. 75:37, 2012. (2) M. B. Ellis. Dematiaceous Hyphomycetes. Commonwealth Mycological Institute, Kew, UK, 1971. (3) D. F. Farr and A. Y. Rossman. Fungal Databases. Syst. Mycol. Microbiol. Lab. ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , 2013. (4) H. Lorenzi et al. Palmeira no Brasil: Exóticas e Nativas, 2nd ed. Editora Plantarum, Nova Odessa, Brazil, 2005.

Sizing up Septoria.

Septoria represents a genus of plant pathogenic fungi with a wide geographic distribution, commonly associated with leaf spots and stem cankers of a broad range of plant hosts. A major aim of this study was to resolve the phylogenetic generic limits of Septoria, Stagonospora, and other related genera such as Sphaerulina, Phaeosphaeria and Phaeoseptoria using sequences of the the partial 28S nuclear ribosomal RNA and RPB2 genes of a large set of isolates. Based on these results Septoria is shown to be a distinct genus in the Mycosphaerellaceae, which has mycosphaerella-like sexual morphs. Several septoria-like species are now accommodated in Sphaerulina, a genus previously linked to this complex. Phaeosphaeria (based on P. oryzae) is shown to be congeneric with Phaeoseptoria (based on P. papayae), which is reduced to synonymy under the former. Depazea nodorum (causal agent of nodorum blotch of cereals) and Septoria avenae (causal agent of avenae blotch of barley and rye) are placed in a new genus, Parastagonospora, which is shown to be distinct from Stagonospora (based on S. paludosa) and Phaeosphaeria. Partial nucleotide sequence data for five gene loci, ITS, LSU, EF-1α, RPB2 and Btub were generated for all of these isolates. A total of 47 clades or genera were resolved, leading to the introduction of 14 new genera, 36 new species, and 19 new combinations. TAXONOMIC NOVELTIES: New genera - Acicuseptoria Quaedvlieg, Verkley & Crous, Cylindroseptoria Quaedvlieg, Verkley & Crous, Kirstenboschia Quaedvlieg, Verkley & Crous, Neoseptoria Quaedvlieg, Verkley & Crous, Neostagonospora Quaedvlieg, Verkley & Crous, Parastagonospora Quaedvlieg, Verkley & Crous, Polyphialoseptoria Quaedvlieg, R.W. Barreto, Verkley & Crous, Ruptoseptoria Quaedvlieg, Verkley & Crous, Septorioides Quaedvlieg, Verkley & Crous, Setoseptoria Quaedvlieg, Verkley & Crous, Stromatoseptoria Quaedvlieg, Verkley & Crous, Vrystaatia Quaedvlieg, W.J. Swart, Verkley & Crous, Xenobotryosphaeria Quaedvlieg, Verkley & Crous, Xenoseptoria Quaedvlieg, H.D. Shin, Verkley & Crous. New species - Acicuseptoria rumicis Quaedvlieg, Verkley & Crous, Caryophylloseptoria pseudolychnidis Quaedvlieg, H.D. Shin, Verkley & Crous, Coniothyrium sidae Quaedvlieg, Verkley, R.W. Barreto & Crous, Corynespora leucadendri Quaedvlieg, Verkley & Crous, Cylindroseptoria ceratoniae Quaedvlieg, Verkley & Crous, Cylindroseptoria pistaciae Quaedvlieg, Verkley & Crous, Kirstenboschia diospyri Quaedvlieg, Verkley & Crous, Neoseptoria caricis Quaedvlieg, Verkley & Crous, Neostagonospora caricis Quaedvlieg, Verkley & Crous, Neostagonospora elegiae Quaedvlieg, Verkley & Crous, Paraphoma dioscoreae Quaedvlieg, H.D. Shin, Verkley & Crous, Parastagonospora caricis Quaedvlieg, Verkley & Crous, Parastagonospora poae Quaedvlieg, Verkley & Crous, Phlyctema vincetoxici Quaedvlieg, Verkley & Crous, Polyphialoseptoria tabebuiae-serratifoliae Quaedvlieg, Alfenas & Crous, Polyphialoseptoria terminaliae Quaedvlieg, R.W. Barreto, Verkley & Crous, Pseudoseptoria collariana Quaedvlieg, Verkley & Crous, Pseudoseptoria obscura Quaedvlieg, Verkley & Crous, Sclerostagonospora phragmiticola Quaedvlieg, Verkley & Crous, Septoria cretae Quaedvlieg, Verkley & Crous, Septoria glycinicola Quaedvlieg, H.D. Shin, Verkley & Crous, Septoria oenanthicola Quaedvlieg, H.D. Shin, Verkley & Crous, Septoria pseudonapelli Quaedvlieg, H.D. Shin, Verkley & Crous, Setophoma chromolaenae Quaedvlieg, Verkley, R.W. Barreto & Crous, Setoseptoria phragmitis Quaedvlieg, Verkley & Crous, Sphaerulina amelanchier Quaedvlieg, Verkley & Crous, Sphaerulina pseudovirgaureae Quaedvlieg, Verkley & Crous, Sphaerulina viciae Quaedvlieg, H.D. Shin, Verkley & Crous, Stagonospora duoseptata Quaedvlieg, Verkley & Crous, Stagonospora perfecta Quaedvlieg, Verkley & Crous, Stagonospora pseudocaricis Quaedvlieg, Verkley, Gardiennet & Crous, Stagonospora pseudovitensis Quaedvlieg, Verkley & Crous, Stagonospora uniseptata Quaedvlieg, Verkley & Crous, Vrystaatia aloeicola Quaedvlieg, Verkley, W.J. Swart & Crous, Xenobotryosphaeria calamagrostidis Quaedvlieg, Verkley & Crous, Xenoseptoria neosaccardoi Quaedvlieg, H.D. Shin, Verkley & Crous. New combinations - Parastagonospora avenae (A.B. Frank) Quaedvlieg, Verkley & Crous, Parastagonospora nodorum (Berk.) Quaedvlieg, Verkley & Crous, Phaeosphaeria papayae (Speg.) Quaedvlieg, Verkley & Crous, Pseudocercospora domingensis (Petr. & Cif.) Quaedvlieg, Verkley & Crous, Ruptoseptoria unedonis (Roberge ex Desm.) Quaedvlieg, Verkley & Crous, Septorioides pini-thunbergii (S. Kaneko) Quaedvlieg, Verkley & Crous, Sphaerulina abeliceae (Hiray.) Quaedvlieg, Verkley & Crous, Sphaerulina azaleae (Voglino) Quaedvlieg, Verkley & Crous, Sphaerulina berberidis (Niessl) Quaedvlieg, Verkley & Crous, Sphaerulina betulae (Pass.) Quaedvlieg, Verkley & Crous, Sphaerulina cercidis (Fr.) Quaedvlieg, Verkley & Crous, Sphaerulina menispermi (Thüm.) Quaedvlieg, Verkley & Crous, Sphaerulina musiva (Peck) Quaedvlieg, Verkley & Crous, Sphaerulina oxyacanthae (Kunze & J.C. Schmidt) Quaedvlieg, Verkley & Crous, Sphaerulina patriniae (Miura) Quaedvlieg, Verkley & Crous, Sphaerulina populicola (Peck) Quaedvlieg, Verkley & Crous, Sphaerulina quercicola (Desm.) Quaedvlieg, Verkley & Crous, Sphaerulina rhabdoclinis (Butin) Quaedvlieg, Verkley & Crous, Stromatoseptoria castaneicola (Desm.) Quaedvlieg, Verkley & Crous. Typifications: Epitypifications - Phaeosphaeria oryzae I. Miyake, Phaeoseptoria papayae Speg.; Neotypification - Hendersonia paludosa Sacc. & Speg.

Methyl jasmonate induces cell death and loss of hydrogenosomal membrane potential in Trichomonas vaginalis.

Trichomonas vaginalis is an important human parasite of the urogenital tract. Jasmonates are a group of small lipids that are produced in plants and function as stress hormones. Naturally occurring methyl jasmonate (MJ) has been used to treat several types of cancer cells and it is cytotoxic to protistan parasites. It has been suggested that mitochondria are the target organelles of jasmonates. Here, we tested this drug against T. vaginalis. Although metronidazole has been the drug of choice for trichomoniasis, side effects from this treatment are common, and nausea and dizziness have been reported in up to 12% of patients. In addition, there has been increased recognition of resistance to metronidazole. We demonstrate here using flow cytometry, JC-1 and scanning and transmission electron microscopy that MJ induced the cell death of T. vaginalis parasites. Our results are discussed with previous findings published by others.

Genotoxicity and morphological changes induced by the alkaloid monocrotaline, extracted from Crotalaria retusa, in a model of glial cells.

Plants of Crotalaria genus (Leguminosae) present large amounts of the pyrrolizidine alkaloid monocrotaline (MCT) and cause intoxication to animals and humans. Therefore, we investigated the MCT-induced cytotoxicity, morphological changes, and oxidative and genotoxic damages to glial cells, using the human glioblastoma cell line GL-15 as a model. The comet test showed that 24h exposure to 1-500microM MCT and 500microM dehydromonocrotaline (DHMC) caused significant increases in cell DNA damage index, which reached 42-64% and 53%, respectively. Cells exposed to 100-500microM MCT also featured a contracted cytoplasm presenting thin cellular processes and vimentin destabilisation. Conversely, exposure of GL-15 cells to low concentrations of MCT (1-10microM) clearly induced megalocytosis. Moreover, MCT also induced down regulation of MAPs, especially at the lower concentrations adopted (1-10microM). Apoptosis was also evidenced in cells treated with 100-500microM MCT, and a later cytotoxicity was only observed after 6 days of exposure to 500microM MCT. The data obtained provide support for heterogenic and multipotential effects of MCT on GL-15 cells, either interfering on cell growth and cytoskeletal protein expression, or inducing DNA damage and apoptosis and suggest that the response of glial cells to this alkaloid might be related to the neurological signs observed after Crotalaria intoxication.

Sclerotium rolfsii Causing Stem Rot of Impatiens walleriana in Brazil.

Impatiens walleriana, busy lizzy or balsam (local names in Brazil maria-sem-vergonha or beijo-de-frade), is an African member of the Balsaminaceae that has long ago been introduced and established in Brazil. It is now widely cultivated commercially as a potted plant and a popular garden plant (3). It also is a common weed along the coast and is particularly troublesome in some banana plantation areas. There are only two records of fungal pathogens attacking this plant in Brazil: Cercospora fukushiana (leaf spot) and Oidiopsis haplophylli (powdery mildew). In January 2009, a population of diseased plants of I. walleriana was found in a private garden in the city of Rio de Janeiro, Brazil. Plants had rotted and girdled stem bases, leading to a collapse of stems Necrotic areas were covered with fans of white mycelium as well as abundant spherical sclerotia. The fungus was isolated in pure culture by direct aseptic transfer of mycelial fragments and sclerotia to potato dextrose agar (PDA) plates. Colonies were white, cottony, often forming fans, primary hyphae 3.0 to 6.0 µm in diameter, and bearing clamp connections; sclerotia formed after 7 days, initially white becoming dark brown with age, and 0.8 to 1.85 mm in diameter. These are typical features of Sclerotium rolfsii. A specimen was deposited in the local herbarium (Herbarium VIC) under Accession No. VIC 30732. Koch's postulates were performed by inoculating three healthy potted I. walleriana plants (10 × 40 cm high) with approximately 100 freshly collected sclerotia placed in close vicinity with the stem bases. Noninoculated plants kept in a separate pot served as controls. Plants were incubated in a dew chamber for 48 h at 25 ± 2°C. All inoculated plants showed symptoms of stem rot 72 h after inoculation, whereas controls remained healthy. S. rolfsii is a highly polyphagous species that has been recorded to be causing rots (also known as Southern blights) in Brazil on numerous hosts but there are no records of it attacking any members of the Balsaminaceae in Brazil. The only other published records of S. rolfsii on Impatiens spp. are from the United States (Hawaii and Illinois) (2) and the Philippines (1). In South America, there is a single report from Argentina (4) where the disease is regarded as a major threat to the potted plant industry because I. walleriana is one of the most popular potted plants in that country. The potential for losses is also significant for Brazil. To our knowledge, this is the first report of S. rolfsii as a pathogen of I. walleriana in Brazil. Although very damaging to I. walleriana, it is unlikely that this fungus can be used as a natural enemy of this plant species in weed situations because of its wide host range. References: (1) T. O. Dizon and R. B. Pimentel. Philipp. Phytopathol. 29:101, 1993. (2) D. F. Farr and A. Y. Rossman. Fungal Databases, Systematic Mycology and Microbiology Laboratory. Online publication. ARS, USDA, 2009. (3) H. Lorenzi and H. M. Souza. Plantas Ornamentais no Brasil - Arbustivas, Herbáceas e Trepadeiras. Nova Odessa: Instituto Plantarum, 1995. (4). S. M. Wolcan and P. J. Grego. Australas. Plant Dis. Notes 4:54, 2009.

P2X7 modulatory web in Trypanosoma cruzi infection.

P2X7 is a member of the purinergic receptors family, with extracellular adenosine triphosphate (ATP) as the main agonist, promoting cations influx and membrane permeabilization that can lead to cell death. We previously proposed that extracellular ATP is involved in thymus atrophy induced by Trypanosoma cruzi infection through the induction of CD4+/CD8+ double-positive cell death and that P2X7 could be involved in this process. To further elucidate this possibility raised by in vitro assays, in this study, we used P2X7-/- mice and observed no difference in thymus atrophy or parasitemia when compared to C57Bl/6. We then decided to investigate other aspects of purinergic receptor interplay that could be better evidenced by the infection and observed that (1) thymocytes from infected and noninfected C57Bl/6 mice express P2X4 and P2X7 receptors (Western blotting), but ATP-induced membrane permeabilization only occurs in thymocytes from infected mice; (2) peritoneal macrophages from noninfected C57Bl/6 mice (P2X4+ and P2X7+) are permeabilized by ATP. Although macrophages from infected C57Bl/6 mice are P2X7- but P2X4+, they are resistant to ATP, either through permeabilization or Ca++ influx (fluorimetry); (3) using noninfected P2X7-/- mice, C57Bl/6 infected mice, and different agonistic stimuli, we observed interesting cross-talks among P2X and P2Y receptors (flow cytometry).

Anti-Trypanosoma cruzi activity of Pterodon pubescens seed oil: geranylgeraniol as the major bioactive component.

In the search for new therapeutic agents for Chagas' disease, we screened extracts obtained from the Brazilian plant Pterodon pubescens found commercially in the medicinal flora market. We investigated the potential trypanocidal effect of the oleaginous ethanolic extract of P. pubescens seeds and its fractions (PF1, PF1.1, PF1.2, and PF1.3) and of geranylgeraniol (GG-OH), the sole component of the hexane fraction (PF1.2). In experiments with bloodstream trypomastigotes of Trypanosoma cruzi, performed at 37 degrees C in culture medium, PF1.2 and GG-OH showed similar potency, while the oleaginous extract from P. pubescens seeds and the other fractions were about three times less active. GG-OH inhibited the proliferation of intracellular amastigotes, at concentrations which do not affect the mammalian host cell. Transmission electron microscopy and flow cytometry analysis indicate the mitochondrion, an organelle that plays a central role in apoptosis, of both epimastigotes and of trypomastigotes as the major target of GG-OH. On the other hand, the ultrastructural images of the endoplasmic reticulum profiles, myelin-like figures, and concentric membranous arrangements inside damaged mitochondrion are suggestive of an autophagic pathway leading to parasite death. Because the different forms of cell death share some morphological features such as mitochondrial collapse, further studies are needed to disclose the trypanocidal action of GG-OH.

Optimization of an improved analytical method for the determination of 1-nitropyrene in milligram diesel soot samples by high-performance liquid chromatography-mass spectrometry.

A method for determination of nitrated polycyclic aromatic hydrocarbons (NPAHs) in diesel soot by high-performance liquid chromatography-mass spectrometry with atmospheric pressure chemical ionization (APCI) and detection by ion-trap following ultrasonic extraction is described. The determination of 1-nitropyrene that it is the predominant NPAH in diesel soot was emphasized. Vaporization and drying temperatures of the APCI interface, electronic parameters of the MS detector and the analytical conditions in reversed-phase HPLC were optimized. The patterns of fragmentation of representative NPAHs were evaluated by single and multiple fragmentation steps and negative ionization led to the largest signals. The transition (247-->217) was employed for quantitative analysis of 1-nitropyrene. Calibration curves were linear between 1 and 15 microgL(-1) with correlation coefficients better than 0.999. Typical detection limit (DL) of 0.2 microgL(-1) was obtained. Samples of diesel soot and of the reference material (SRM-2975, NIST, USA) were extracted with methylene chloride. Recoveries were estimated by analysis of SRM 2975 and were between 82 and 105%. DL for 1-nitropyrene was better than 1.5 mg kg(-1), but the inclusion of an evaporation step in the sample processing procedure lowered the DL. The application of the method to diesel soot samples from bench motors showed levels

Experimental radiotherapy-induced enteritis: a probiotic interventional study.

OBJECTIVE: Acute radiation of the small intestine causes an immediate and potentially reversible effect on the sensitive regenerative epithelium of the intestinal mucosa while markedly altering the overall intestinal ecosystem. The aim of the present study was test a novel probiotic mixture formulation (Microflorana-F) in an experimental model of acute radiation enteritis with particular interest in endotoxinemia and bacterial translocation. MATERIALS AND METHODS: Male Wistar rats allocated to three groups were fed for 7 days with: (A) a standard balanced diet; (B) a standard diet with the addition of 1 mL t.i.d. of Microflorana-F and (C) the same probiotic but after heat inactivation. Under ketamine anesthesia, abdominal irradiation was performed at a single dose rate of 20 Gy. Sham-radiated healthy rats served as a control (D). Standard food and active/inactive probiotic supplementation schedule was maintained throughout the study period. When they were killed 14 days later a midline laparotomy and a medium sternotomy was carried out. The mesenteric lymph nodes, whole spleen and liver samples as well as blood, the portal vein and bile samples were cultured. Endotoxinemia was also measured. RESULTS: Early deaths (1 week) occurred mostly in rats fed standard food or inactivated probiotic. The endotoxin level significantly increased in irradiated rats fed standard food and inactivated probiotic while supplementation with the active form of the probiotic mixture significantly improved such parameters (P < 0.05). After radiation injury, mesenteric lymph nodes and portal blood were the samples most frequently yielding bacterial growth. Treatment with only the active form of probiotic significantly reduced the incidence of bacterial contamination in all samples. CONCLUSIONS: These data suggest that the manipulation of gut ecosystem by biologically effective probiotic preparations might be a worthwhile therapeutic and preventive tool in radiation-induced enteritis.