Ecology and identification of environmental fungi and metabolic processes involved in the biodeterioration of Brazilian soapstone historical monuments.

This study aimed to evaluate the action of organic acids produced by the fungal population associated with the biodeterioration process of the Twelve Prophets of Aleijadinho, a set of soapstone sculptures in Congonhas, state of Minas Gerais, Brazil. For this, samples of fungi were obtained from the surface of each of the 12 outdoor stone sculptures that comprise the set of Prophets. The identification of the colonizing filamentous fungi was performed by classical microbiology and molecular methods. Some species of filamentous fungi-dependent cultivation were detected, and the presence of species Aspergillus versicolor, Curvularia lunata, Epicoccum nigrum, Penicillium citrinum and Pseudocercospora norchiensis indicated a connection with the excretion of organic acids. The acids produced by each of these fungal species were analysed quantitatively by chromatographic methods, revealing potential biodeterioration by the action of acidic metabolites excreted in the stone. SIGNIFICANCE AND IMPACT OF THE STUDY: Minas Gerais, Brazil, is vulnerable to the activities of mineral extraction industries, posing an imminent risk to United Nations Educational, Scientific and Cultural Organization (UNESCO) recognized cities, e.g. Congonhas. Many of these municipalities hold many soapstone religious sculptures and historical monuments. Consequently, soapstone is susceptible to filamentous fungi attack causing irreversible biodeterioration. Despite the concern related to nondestructive sampling of 18th century sculptures, in this study, we have discussed the factors that lead to biodeterioration of soapstone due to organic acid excretion by the fungi that damage the stone, thereby providing an insight in conserving and preserving the soapstone monuments.

REVIEW-ARTICLE Intermediate alleles of Huntington's disease HTT gene in different populations worldwide: a systematic review.

Huntington's disease (HD) is an autosomal dominant progressive neurodegenerative disorder caused by a dynamic mutation due to the expansion of CAG repeats in the HTT gene (4p16.3). The considered normal alleles have less than 27 CAG repeats. Intermediate alleles (IAs) show 27 to 35 CAG repeats and expanded alleles have more than 35 repeats. The IAs apparently have shown a normal phenotype. However, there are some reported associations between individuals that bear an IA and clinical HD signs, such as behavioral disturbs. The association of IAs with the presence of clinical signs gives clinical relevance to these patients. We emphasized the importance of determining the frequency of IA alleles in the general population as well as in HD families. Therefore, the aim of this study was to conduct a systematic review, in order to investigate the frequency of IAs in the overall chromosomes of different ethnic groups and of families with HD history worldwide as well as the frequency of individuals who bear the intermediate alleles. We searched indexed articles from the following electronic databases: U.S. National Library of Medicine and the National Institutes of Health (PubMed), Pubmed Central (PMC) and Virtual Health Library (VHL). Therefore, 488 articles were obtained and, of these, 33 had been published in more than one database. We accepted the article of only one database and ended up with 455 articles for this review. The frequency of IAs within the chromosomes of the general population ranged from 0.45 to 8.7% and of individuals with family history of HD ranged from 0.05 to 5.1%. The higher frequency of IAs in the general population (8.7%) was found in one Brazilian cohort.

Technical note: A facility for respiration measurements in cattle.

A respiration system consisting of 4 climate-controlled chambers and 1 set of flowmeters and analyzers was constructed and validated. Each chamber had volume of 21.10m(3) (3.68×2.56×2.24m) and was made from steel with double-glazed windows on either side enabling visual contact between animals. The chambers are independently climate-controlled and can maintain temperature and relative humidity in a range from 5 to 45°C and 30 to 80%, respectively. A flow generator and mass flowmeter continuously pull air from each chamber and a slight negative pressure inside the chamber is ensured. Air from all chambers and ambient air share a common gas analysis and data acquisition system for monitoring O2, CO2, and CH4 concentrations over the measurement period, with the cycle time set to 20min. Analyzers are regularly calibrated and the chambers have mean recoveries of 99.0 and 98.0% for CO2 and CH4, respectively. The chambers are equipped with infrared cameras and electronic feed and water bins for intake measurements, as well as sensors for monitoring animal position and heart rate. Data acquisition and analysis software is used to calculate the rate of consumption of O2 and production of CO2 and CH4. The dynamic respiration measurements are integrated with feed intake data and other sensors. The daily gas exchanges are estimated by integration to determine methane emission and heat production. We conducted a trial with 12 lactating 3/4 Holstein × 1/4 Gyr crossbred dairy cows (6 multiparous and 6 primiparous) under 2 feeding regimens (ad libitum or restricted) to validate the system. Two 22-h respiration measurements were obtained from each cow. Restricted-fed cows showed lower values for milk yield, methane emission, and heat production compared with ad libitum-fed animals. We found no difference between groups for CH4 produced per kilogram of dry matter intake. Repeatability for CH4 emission and heat production was high (0.97 and 0.92, respectively). The respiration system described herein is a useful tool for measuring the dynamic and accumulated data of heat production, methane emission, and feed intake.

Notification of Huntington's disease as primary cause of death in Brazil from 1984 to 2008.

The aim of this article was to conduct a retrospective observational study on reported deaths due to Huntington's disease (HD) in Brazil in the past 25 years (from 1984 to 2008). Data were obtained from the Brazilian Mortality Information System (SIM/DATASUS), the official system of Brazilian Mortality Database. The data obtained included information regarding the gender of the deceased and the number of death notifications, which we stratified by demographic regions and states. HD mortality per 100,000 was calculated and plotted in a graph. Linear regression was calculated using ordinary least square technique. We observed that the mortality due to HD recorded by SIM/DATASUS from 1984 to 2008 had increased at much higher rates than the population in the same period. Also, some Brazilian regions still show very low rates of HD mortality compared to the national average of deaths due to HD. These findings suggest that HD mortality has been underestimated. Ignorance about the disease as well as the fact that death from HD can occur as a consequence of heart disease, pneumonia, or suicide can strongly contribute to the misguided notification of HD as the cause of death in the official reports.

Mini-Review: Monosomy 1p36 syndrome: reviewing the correlation between deletion sizes and phenotypes.

The major clinical features of monosomy 1p36 deletion are developmental delay and hypotonia associated with short stature and craniofacial dysmorphisms. The objective of this study was to review the cases of 1p36 deletion that was reported between 1999 and 2014, in order to identify a possible correlation between the size of the 1p36-deleted segment and the clinical phenotype of the disease. Scientific articles published in the (National Center for Biotechnology Information; NCBI http://www.ncbi.nlm.nih.gov/pubmed) and Scientific Electronic Library Online (www.scielo.com.br) databases were searched using key word combinations, such as "1p36 deletion", "monosomy 1p36 deletion", and "1p36 deletion syndrome". Articles in English or Spanish reporting the correlation between deletion sizes and the respective clinical phenotypes were retrieved, while letters, reviews, guidelines, and studies with mouse models were excluded. Among the 746 retrieved articles, only 17 (12 case reports and 5 series of cases), comprising 29 patients (9 males and 20 females, aged 0 months (neonate) to 22 years) bearing the 1p36 deletions and whose clinical phenotypes were described, met the inclusion criteria. The genotype-phenotype correlation in monosomy 1p36 is a challenge because of the variability in the size of the deleted segment, as well as in the clinical manifestations of similar size deletions. Therefore, the severity of the clinical features was not always associated with the deletion size, possibly because of the other influences, such as stochastic factors, epigenetic events, or reduced penetration of the deleted genes.

Prader-Willi-like phenotypes: a systematic review of their chromosomal abnormalities.

Prader-Willi syndrome (PWS) is caused by the lack of expression of genes located on paternal chromosome 15q11-q13. This lack of gene expression may be due to a deletion in this chromosomal segment, to maternal uniparental disomy of chromosome 15, or to a defect in the imprinting center on 15q11-q13. PWS is characterized by hypotonia during the neonatal stage and in childhood, accompanied by a delay in neuropsychomotor development. Overeating, obesity, and mental deficiency arise later on. The syndrome has a clinical overlap with other diseases, which makes it difficult to accurately diagnose. The purpose of this article is to review the Prader-Willi-like phenotype in the scientific literature from 2000 to 2013, i.e., to review the cases of PWS caused by chromosomal abnormalities different from those found on chromosome 15. A search was carried out using the "National Center for Biotechnology Information" (www.pubmed.com) and "Scientific Electronic Library Online (www.scielo.br) databases and combinations of key words such as "Prader-Willi-like phenotype" and "Prader-Willi syndrome phenotype". Editorials, letters, reviews, and guidelines were excluded. Articles chosen contained descriptions of patients diagnosed with the PWS phenotype but who were negative for alterations on 15q11-q13. Our search found 643 articles about PWS, but only 14 of these matched with the Prader-Willi-like phenotype and with the selected years of publication (2000-2013). If two or more articles reported the same chromosomal alterations for Prader-Willi-like phenotype, the most recent was chosen. Twelve articles of 14 were case reports and 2 reported series of cases.

Phenotyping of Passiflora edulis, P. setacea, and their hybrids by a multivariate approach.

Morphological characterization is the most accessible and used method to quantify the genetic diversity of the available germplasm. The multivariate statistical method is highly important for this purpose. This study aimed to characterize parents and hybrids of Passiflora according to morphoagronomic descriptors and estimate the genetic divergence between them based on the joint analysis of qualitative and quantitative variables using the Ward-modified location model (MLM) procedure. One hundred and thirty-eight individuals were assessed (10 P. edulis, 10 P. setacea, and 118 interspecific hybrids) using 23 quantitative and 12 qualitative descriptors. The values for the quantitative descriptors were measured and subjected to multivariate statistics using the Ward-MLM strategy. Large genetic variability was detected by the morphoagronomic data in the 138 genotypes that were evaluated, and the hybrids presented higher variability than the parents. Pseudo-F and pseudo-t2 criteria showed that the optimal number of groups was three. Group I was composed of 118 hybrid genotypes; group II was composed of the 10 P. setacea genotypes, and group III was composed of the 10 P. edulis genotypes. The longest distance was found between groups II and III (474.96). The shortest distance was detected between groups I and II (198.78), which indicates that the segregating population is genetically closer to P. setacea than to P. edulis. The Ward-MLM procedure is a useful tool to detect genetic diversity and group accessions using both qualitative and quantitative variables.

A systematic review of the intergenerational aspects and the diverse genetic profiles of Huntington's disease.

Huntington's disease (HD) is a rare progressive and fatal neurogenetic degenerative disease, characterized by movement and personality disorders and by progressive dementia. Its prevalence varies by ethnic origin and different genetic profiles predisposing individuals to HD in each population. The prevalence of HD is 5-10 per 100,000 individuals in Caucasian populations of North America and Western Europe. It is an autosomal dominant disease associated with the expansion of CAG-type repetitive DNA sequences in the HTT gene. This gene, located on the short arm of chromosome 4, encodes the protein huntingtin. In this study, we reviewed 17 articles about HD that report data from 2400 affected individuals from various countries around the world, including Venezuela, China, Croatia, Turkey, Germany, Italy, Brazil, Spain, Taiwan, India, the Netherlands, Russia, and the USA, with a focus on genetic profiles and intergenerational expansions or contractions of expanded alleles responsible for causing HD. We discuss the genetic characteristics of HD in different populations and any atypical cases reported in these studies.

Biocontrol potential of actinobacteria against Pantoea ananatis, the causal agent of maize white spot disease.

Pantoea ananatis is the causal agent of maize white spot, a foliar disease responsible for significant maize yield reduction worldwide, especially in Brazil. In general, the maize foliar diseases control involves the adoption of resistant genotypes and pesticides application. However, the use of agrochemicals can significantly cause increase production costs, damage to human health and negative environmental impacts. In this sense, the use of biological control agents has been considered among the most promising eco-friendly technologies for sustainable agriculture. Actinobacteria, particularly of Streptomyces genus, has been widely recognized as agroindustrially important microorganism due to its potential in producing diverse range of secondary metabolites, including antibiotics and enzymes. Thus, the aim of this work is to characterize and to evaluate the potential of soil actinobacteria for P. ananatis control. We observed that 59 actinobacteria strains (85%) exhibited proteolytic or chitinolytic activity. Only the strains Streptomyces pseudovenezuelae ACSL 470, that also exhibited high proteolytic activity, S. novaecaesareae ACSL 432 and S. laculatispora ACP 35 demonstrated high or moderate antagonist activity in vitro against P. ananatis. Temporal analysis of metabolites produced by these strains growth in different liquid media indicated greater antibacterial activity at 72 h. In this condition, chromatographic and mass spectrometry analysis revealed that S. pseudovenezuelae ACSL 470 strain produced neomycin, an aminoglycoside antibiotic that displayed high bactericidal activity in vitro against P. ananatis. This is the first report of actinobacteria acting as potential microbial antagonists for P. ananatis control. Further studies are needed to determine the control efficacy of maize white spot disease by Streptomyces strains or their metabolites in greenhouse and field conditions.

Low humidity environmental challenge causes barrier disruption and cornification of the mouse corneal epithelium via a c-jun N-terminal kinase 2 (JNK2) pathway.

Patients with tear dysfunction often experience increased irritation symptoms when subjected to drafty and/or low humidity environmental conditions. The purpose of this study was to investigate the effects of low humidity stress (LHS) on corneal barrier function and expression of cornified envelope (CE) precursor proteins in the epithelium of C57BL/6 and c-jun N-terminal kinase 2 (JNK2) knockout (KO) mice. LHS was induced in both strains by exposure to an air draft for 15 (LHS15D) or 30 days (LHS30D) at a relative humidity <30%RH. Nonstressed (NS) mice were used as controls. Oregon-green-dextran uptake was used to measure corneal barrier function. Levels of small proline-rich protein (SPRR)-2, involucrin, occludin, and MMP-9 were evaluated by immunofluorescent staining in cornea sections. Wholemount corneas immunostained for occludin were used to measure mean apical cell area. Gelatinase activity was evaluated by in situ zymography. Expression of MMP, CE and inflammatory cytokine genes was evaluated by qPCR. C57BL/6 mice exposed to LHS15D showed corneal barrier dysfunction, decreased apical corneal epithelial cell area, higher MMP-9 expression and gelatinase activity and increased involucrin and SPRR-2 immunoreactivity in the corneal epithelium compared to NS mice. JNK2KO mice were resistant to LHS-induced corneal barrier disruption. MMP-3,-9,-13, IL-1α, IL-1ß, involucrin and SPRR-2a RNA transcripts were significantly increased in C57BL/6 mice at LHS15D, while no change was noted in JNK2KO mice. LHS is capable of altering corneal barrier function, promoting pathologic alteration of the TJ complex and stimulating production of CE proteins by the corneal epithelium. Activation of the JNK2 signaling pathway contributes to corneal epithelial barrier disruption in LHS.

Effectiveness of a palliative care outpatient programme in improving cancer-related symptoms among ambulatory Brazilian patients.

The focuses of palliative care are to provide symptom relief and improve quality of life through an interdisciplinary approach. Previous studies conducted to evaluate the effectiveness of palliative care in reducing symptom distress among outpatients confirmed the importance of the palliative care approach. To our knowledge, there is no published information from Brazil regarding the impact of a palliative care outpatient programme in reducing symptom distress. Symptom scores from 232 patients were measured using Edmonton Symptom Assessment System scales in two consecutive consults. Changes in symptoms at follow-up visit were analysed using Wilcoxon signed-rank paired test. The symptom subtraction indices (SSI) (follow-up scores minus baseline scores) were calculated and then analysed with Spearman's correlation. Edmonton Symptom Assessment System median scores at follow-up visits were statistically significant reduced in all symptoms evaluated. All the SSI positively correlated with well-being-SSI. Other important SSI correlations were: fatigue-SSI and anxiety-SSI, and fatigue-SSI and dyspnoea-SSI. Our palliative care outpatient programme was able to provide a significant improvement in the symptoms evaluated. The well-being-SSI was positively correlated with all the SSI, verifying that the control of symptoms in palliative care is essential for the patient well-being. Adequate/inadequate control of specifically symptoms seems to indirectly improve/worsen other symptoms.

Microorganisms from corn stigma with biocontrol potential of Fusarium verticillioides.

The mycotoxigenic fungus Fusarium verticillioides is the primary maize pathogen and causes the maize stalk and ear rot diseases with significant economic losses. Furthermore, the excessive use of fungicides to control F. verticillioides constitutes threats to the environment and human health. Thus, sustainable alternatives such as biological control are needed to minimize the hazards associated with the current method. Although much is known about the vulnerability of the maize silks as a gateway for several fungal pathogens invading the developing grains, studies on the chemical properties of silk extracts and their resident microbiota are scarce. This study isolated and characterized bacteria and fungi that colonize the maize stigma to assess new potential biocontrol agents. The samples were collected from maize fields in the Brazilian localities of Sete Lagoas-MG, Sidrolândia-MS, Sertaneja-PR, and Goiânia-GO. One hundred sixty-seven microorganisms were isolated, 46% endophytic and 54% epiphytic. First, the antagonist activity was evaluated by the agar disc diffusion method performed in triplicate, and 83% of the isolates showed antagonist activity against F. verticillioides. Then, the 42 most efficient isolates were identified based on the partial sequencing of the bacterial 16S rRNA gene and fungi ITS region. The bacteria belong to the genera Bacillus (57.1%), Burkholderia (23.8%), Achromobacter (7.1%), Pseudomonas (2.4%), and Serratia (2.4%), while the fungi are Penicillium (2.4%), Candida (2.4), and Aspergillus (2.4%). The results showed that microorganisms from maize stigma might represent new promising agents for F. verticillioides control.

Thermo-resistant enzyme-producing microorganisms isolated from composting.

Organo-mineral fertilizers supplemented with biological additives are an alternative to chemical fertilizers. In this study, thermoresistant microorganisms from composting mass were isolated by two-step procedures. First, samples taken at different time points and temperatures (33 days at 52 ºC, 60 days at 63 ºC, and over 365 days at 26 ºC) were pre-incubated at 80 oC for 30 minutes. Second, the microbial selection by in vitro culture-based methods and heat shock at 60 oC and 100 oC for 2h and 4h. Forty-one isolates were able to grow at 60 °C for 4h; twenty-seven at 100 °C for 2h, and two at 100 °C for 4h. The molecular identification by partial sequencing of the 16S ribosomal gene using universal primers revealed that thirty-five isolates were from eight Bacillus species, one Brevibacillus borstelensis, three Streptomyces thermogriseus, and two fungi (Thermomyces lanuginosus and T. dupontii). Data from amylase, phytase, and cellulase activity assays and the enzymatic index (EI) showed that 38 of 41 thermo-resistant isolates produce at least one enzyme. For amylase activity, the highest EI value was observed in Bacillus licheniformis (isolate 21C2, EI= 4.11), followed by Brevibacillus borstelensis (isolate 6C2, EI= 3.66), Bacillus cereus (isolate 18C2, EI= 3.52), and Bacillus paralicheniformis (isolate 20C2, EI= 3.34). For phytase, the highest EI values were observed for Bacillus cereus (isolate 18C2, EI= 2.30) and Bacillus licheniformis (isolate 3C1, EI= 2.15). Concerning cellulose production, B. altitudinis (isolate 6C1) was the most efficient (EI= 6.40), followed by three Bacillus subtilis (isolates 9C1, 16C2, and 19C2) with EI values of 5.66, 5.84, and 5.88, respectively, and one B. pumilus (isolate 27C2, EI= 5.78). The selected microorganisms are potentially useful as a biological additive in organo-mineral fertilizers and other biotechnological processes.

Diesel exhaust particles override natural injury-limiting pathways in the lung.

Induction of effective inflammation in the lung in response to environmental and microbial stimuli is dependent on cooperative signaling between leukocytes and lung tissue cells. We explored how these inflammatory networks are modulated by diesel exhaust particles (DEP) using cocultures of human monocytes with epithelial cells. Cocultures, or monoculture controls, were treated with DEP in the presence or absence of LPS or flagellin. Production of cytokines was explored by Western blotting and ELISA; cell signaling was analyzed by Western blotting. Here, we show that responses of epithelial cells to DEP are amplified by the presence of monocytes. DEP amplified the responses of cellular cocultures to very low doses of TLR agonists. In addition, in the presence of DEP, the responses induced by LPS or flagellin were less amenable to antagonism by the physiological IL-1 antagonist, IL-1ra. This was paralleled by the uncoupling of IL-1 production and release from monocytes, potentially attributable to an ability of DEP to sequester or degrade extracellular ATP. These data describe a model of inflammation where DEP amplifies responses to low concentrations of microbial agonists and alters the nature of the inflammatory milieu induced by TLR agonists.

Absence of transforming growth factor-beta type II receptor is associated with poorer prognosis in HER2-negative breast tumours.

BACKGROUND: The clinical relevance of transforming growth factor-beta (TGF-beta)-signalling pathway in breast carcinomas (BCs) remained elusive. This study aimed to evaluate the prognostic value of TGF-beta1 and transforming growth factor-beta type II receptor (TGF-betaRII) expression levels in tumour cells and their association with the established biomarkers in BC. PATIENTS AND METHODS: In 324 BC from patients with long-term follow-up, the TGF-beta1 and TGF-betaRII transcript and protein expression levels were assessed. RESULTS: TGF-beta1 and TGF-betaRII down-expression was significantly associated with BC. Negative TGF-beta1 and TGF-betaRII protein status was associated with the development of distant metastasis (P = 0.003 and P = 0.029, respectively). In multivariate analysis, TGF-beta1-positive tumours were associated with increased disease-free survival (DFS) [hazard ratio (HR) = 0.489, P = 0.003]. TGF-betaRII positivity was an independent prognostic factor for DFS (HR = 0.439, P = 0.001) and overall survival (OS) (HR = 0.409, P = 0.003) in human epidermal growth factor receptor-2 (HER2)-negative patients. Absence of TGF-beta1 and TGF-betaRII proteins in breast tumour cells was significantly associated with metastasis development. CONCLUSIONS: To the best of our knowledge, this is the first report indicating the relevance of HER2 status in discriminating TGF-betaRII as a prognostic marker for DFS and OS in human BC. These data indicate that TGF-betaRII protein analysis in tumour cells could be introduced in clinical practice as additional prognostic biomarker in HER2-negative BC.

Microsatellite instability in Lactuca sativa chronically exposed to cadmium.

Cadmium (Cd) is a cytotoxic, mutagenic and carcinogenic metal. Mutagenesis is indicative of genetic instability and can be assayed by use of microsatellite markers (simple sequence repeats, SSRs). These are tandem-repeated tracts of DNA composed of units that are 1-6 base pairs (bp) long, spread throughout the genome and highly polymorphic. SSRs can be used in the detection of genomic DNA damage and/or mutational events (e.g. deletions, insertions, point mutations). In order to study chronic exposure to cadmium, Lactuca sativa L. seeds were germinated in distilled water and grown on modified Hoagland's medium, both supplemented with 0, 10 and 100microM Cd(NO(3))(2). After 28 days of exposure, the plants were harvested to assess shoot and root length and accumulation of cadmium. DNA was extracted from young and expanded leaves and roots in order to analyse microsatellite instability (MSI). Mutagenic effects of cadmium were evaluated on nine microsatellite loci. No MSI was found in leaves, but a 2-bp deletion in one lettuce root SSR was detected among the SSRs that were analysed. Thus, SSR analyses may provide a complementary tool in the assessment of different genotoxic effects of compounds on plants.

Thermo-resistant enzyme-producing microorganisms isolated from composting

Abstract Organo-mineral fertilizers supplemented with biological additives are an alternative to chemical fertilizers. In this study, thermoresistant microorganisms from composting mass were isolated by two-step procedures. First, samples taken at different time points and temperatures (33 days at 52 ºC, 60 days at 63 ºC, and over 365 days at 26 ºC) were pre-incubated at 80 oC for 30 minutes. Second, the microbial selection by in vitro culture-based methods and heat shock at 60 oC and 100 oC for 2h and 4h. Forty-one isolates were able to grow at 60 °C for 4h; twenty-seven at 100 °C for 2h, and two at 100 °C for 4h. The molecular identification by partial sequencing of the 16S ribosomal gene using universal primers revealed that thirty-five isolates were from eight Bacillus species, one Brevibacillus borstelensis, three Streptomyces thermogriseus, and two fungi (Thermomyces lanuginosus and T. dupontii). Data from amylase, phytase, and cellulase activity assays and the enzymatic index (EI) showed that 38 of 41 thermo-resistant isolates produce at least one enzyme. For amylase activity, the highest EI value was observed in Bacillus licheniformis (isolate 21C2, EI= 4.11), followed by Brevibacillus borstelensis (isolate 6C2, EI= 3.66), Bacillus cereus (isolate 18C2, EI= 3.52), and Bacillus paralicheniformis (isolate 20C2, EI= 3.34). For phytase, the highest EI values were observed for Bacillus cereus (isolate 18C2, EI= 2.30) and Bacillus licheniformis (isolate 3C1, EI= 2.15). Concerning cellulose production, B. altitudinis (isolate 6C1) was the most efficient (EI= 6.40), followed by three Bacillus subtilis (isolates 9C1, 16C2, and 19C2) with EI values of 5.66, 5.84, and 5.88, respectively, and one B. pumilus (isolate 27C2, EI= 5.78). The selected microorganisms are potentially useful as a biological additive in organo-mineral fertilizers and other biotechnological processes.

Resumo Os fertilizantes organo-minerais suplementados com aditivos biológicos são uma alternativa aos adubos químicos. Neste estudo, microrganismos termoresistentes foram isolados de compostagem por procedimentos de duas etapas. Inicialmente, as amostras tomadas em diferentes períodos e temperaturas (33 dias a 52 ºC, 60 dias a 63 ºC e mais de 365 dias a 26 ºC) foram pré-incubadas a 80 oC por 30 minutos. Posteriormente, a seleção microbiana foi conduzida por métodos baseados em cultura in vitro e choque térmico a 60 oC e 100 oC por 2h e 4h. Quarenta e um isolados foram capazes de crescer a 60 °C por 4h; vinte e sete a 100 °C por 2h e dois a 100 °C por 4h. A identificação molecular por sequenciamento parcial do gene ribossomal 16S usando primers universais revelou que trinta e cinco isolados eram de oito espécies de Bacillus, um Brevibacillus borstelensis, três Streptomyces thermogriseus e dois fungos (Thermomyces lanuginosus e T. dupontii). Os dados dos ensaios de atividade de amilase, fitase e celulase e o índice enzimático (IE) mostraram que 38 dos 41 isolados termorresistentes produziram pelo menos uma enzima. Para a atividade da amilase, o maior valor de IE foi observado em Bacillus licheniformis (isolado 21C2, IE = 4,11), seguido por Brevibacillus borstelensis (isolado 6C2, IE = 3,66), Bacillus cereus (isolado 18C2, IE = 3,52) e Bacillus paralicheniformis (isolado 20C2, IE = 3,34). Para a fitase, os maiores valores de IE foram observados para B. cereus (isolado 18C2, IE = 2,30) e B. licheniformis (isolado 3C1, IE = 2,15). Em relação à produção de celulose, B. altitudinis (isolado 6C1) foi o mais eficiente (IE = 6,40), seguido por três Bacillus subtilis (isolados 9C1, 16C2 e 19C2) com valores de IE de 5,66, 5,84 e 5,88, respectivamente, e um B. pumilus (isolado 27C2, IE = 5,78). Pode-se inferir que os microrganismos selecionados são potencialmente úteis como aditivos biológicos em fertilizantes organo-minerais e outros processos biotecnológicos.

Thermo-resistant enzyme-producing microorganisms isolated from composting / Microrganismos produtores de enzimas termo-resistentes isolados de compostagem

Organo-mineral fertilizers supplemented with biological additives are an alternative to chemical fertilizers. In this study, thermoresistant microorganisms from composting mass were isolated by two-step procedures. First, samples taken at different time points and temperatures (33 days at 52 ºC, 60 days at 63 ºC, and over 365 days at 26 ºC) were pre-incubated at 80 oC for 30 minutes. Second, the microbial selection by in vitro culture-based methods and heat shock at 60 oC and 100 oC for 2h and 4h. Forty-one isolates were able to grow at 60 °C for 4h; twenty-seven at 100 °C for 2h, and two at 100 °C for 4h. The molecular identification by partial sequencing of the 16S ribosomal gene using universal primers revealed that thirty-five isolates were from eight Bacillus species, one Brevibacillus borstelensis, three Streptomyces thermogriseus, and two fungi (Thermomyces lanuginosus and T. dupontii). Data from amylase, phytase, and cellulase activity assays and the enzymatic index (EI) showed that 38 of 41 thermo-resistant isolates produce at least one enzyme. For amylase activity, the highest EI value was observed in Bacillus licheniformis (isolate 21C2, EI= 4.11), followed by Brevibacillus borstelensis (isolate 6C2, EI= 3.66), Bacillus cereus (isolate 18C2, EI= 3.52), and Bacillus paralicheniformis (isolate 20C2, EI= 3.34). For phytase, the highest EI values were observed for Bacillus cereus (isolate 18C2, EI= 2.30) and Bacillus licheniformis (isolate 3C1, EI= 2.15). Concerning cellulose production, B. altitudinis (isolate 6C1) was the most efficient (EI= 6.40), followed by three Bacillus subtilis (isolates 9C1, 16C2, and 19C2) with EI values of 5.66, 5.84, and 5.88, respectively, and one B. pumilus (isolate 27C2, EI= 5.78). The selected microorganisms are potentially useful as a biological additive in organo-mineral fertilizers and other biotechnological processes.

Os fertilizantes organo-minerais suplementados com aditivos biológicos são uma alternativa aos adubos químicos. Neste estudo, microrganismos termoresistentes foram isolados de compostagem por procedimentos de duas etapas. Inicialmente, as amostras tomadas em diferentes períodos e temperaturas (33 dias a 52 ºC, 60 dias a 63 ºC e mais de 365 dias a 26 ºC) foram pré-incubadas a 80 oC por 30 minutos. Posteriormente, a seleção microbiana foi conduzida por métodos baseados em cultura in vitro e choque térmico a 60 oC e 100 oC por 2h e 4h. Quarenta e um isolados foram capazes de crescer a 60 °C por 4h; vinte e sete a 100 °C por 2h e dois a 100 °C por 4h. A identificação molecular por sequenciamento parcial do gene ribossomal 16S usando primers universais revelou que trinta e cinco isolados eram de oito espécies de Bacillus, um Brevibacillus borstelensis, três Streptomyces thermogriseus e dois fungos (Thermomyces lanuginosus e T. dupontii). Os dados dos ensaios de atividade de amilase, fitase e celulase e o índice enzimático (IE) mostraram que 38 dos 41 isolados termorresistentes produziram pelo menos uma enzima. Para a atividade da amilase, o maior valor de IE foi observado em Bacillus licheniformis (isolado 21C2, IE = 4,11), seguido por Brevibacillus borstelensis (isolado 6C2, IE = 3,66), Bacillus cereus (isolado 18C2, IE = 3,52) e Bacillus paralicheniformis (isolado 20C2, IE = 3,34). Para a fitase, os maiores valores de IE foram observados para B. cereus (isolado 18C2, IE = 2,30) e B. licheniformis (isolado 3C1, IE = 2,15). Em relação à produção de celulose, B. altitudinis (isolado 6C1) foi o mais eficiente (IE = 6,40), seguido por três Bacillus subtilis (isolados 9C1, 16C2 e 19C2) com valores de IE de 5,66, 5,84 e 5,88, respectivamente, e um B. pumilus (isolado 27C2, IE = 5,78). Pode-se inferir que os microrganismos selecionados são potencialmente úteis como aditivos biológicos em fertilizantes organo-minerais e outros processos biotecnológicos.

Age-related spontaneous lacrimal keratoconjunctivitis is accompanied by dysfunctional T regulatory cells.

In both humans and animal models, the development of Sjögren syndrome (SS) and non-SS keratoconjunctivitis sicca (KCS) increases with age. Here, we investigated the ocular surface and lacrimal gland (LG) phenotype of NOD.B10.H2b mice at 7-14, 45-50, and 96-100 weeks. Aged mice develop increased corneal permeability, CD4+ T-cell infiltration, and conjunctival goblet cell loss. Aged mice have LG atrophy with increased lymphocyte infiltration and inflammatory cytokine levels. An increase in the frequency of CD4+Foxp3+ T regulatory cells (Tregs) was observed with age in the cervical lymph node (CLN), spleen, and LG. These CD4+CD25+ cells lose suppressive ability, while maintaining expression of Foxp3 (forkhead box P3) and producing interleukin-17 (IL-17) and interferon-γ (IFN-γ). An increase of Foxp3+IL-17+ or Foxp3+IFN-γ+ cells was observed in the LG and LG-draining CLN. In adoptive transfer experiments, recipients of either purified Tregs or purified T effector cells from aged donors developed lacrimal keratoconjunctivitis, whereas recipients of young Tregs or young T effector cells failed to develop disease. Overall, these results suggest inflammatory cytokine-producing CD4+Foxp3+ cells participate in the pathogenesis of age-related ocular surface disease.