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.

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.

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.

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.

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.

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.

Evolução anual da qualidade do leite cru refrigerado processado em uma indústria de Minas Gerais / Annual evolution of raw milk quality processed in a dairy industry of Minas Gerais state, Brazil

Analisou-se a evolução anual da qualidade de leite cru refrigerado pelo levantamento e análise estatística do banco de dados das análises individuais de leite dos tanques refrigeradores, computadas mensalmente entre abril de 2002 e dezembro de 2008 em uma indústria de Minas Gerais. O volume de leite em conformidade com a IN51 para contagem bacteriana total para o ano de 2008 aumentou de 74,3% em 2002 para 85,8% em 2008. Considerando-se a IN51 para o ano de 2011, esses valores seriam de 30,5% e 46,6%, respectivamente. O volume de leite em conformidade com a IN51 para contagem de células somáticas reduziu-se de 93,5% em 2002 para 90,1% em 2008. Para o ano de 2011, esses valores seriam de 64,9 e 60,5%, respectivamente. Em relação à composição sólida do leite, o volume de leite em conformidade com a IN51 para os teores de gordura, de proteína e de extrato seco desengordurado foi de 95,2%, 98,2% e 89,6%, respectivamente. As variações sazonais influenciaram a qualidade do leite cru refrigerado de tanques de expansão. A implantação de um sistema de pagamento por qualidade do leite ao produtor foi eficaz na melhoria dos índices de qualidade da matéria-prima.

The present study aimed to analyze the annual evolution of cooled raw milk quality, through analytical analysis of the data base of individual bulk milk tanks analysis, monthly computed from April/2002 to December/2008 at a dairy industry in Minas Gerais. The mean volume of milk in compliance with IN51 for total bacterial count for year 2008 increased from 74.3% in 2002 to 85.8% in 2008. Considering IN51 for year 2011, these values would be 30.5% and 46.6%, respectively. The mean volume of milk in compliance with IN51 for somatic cells count reduced from 93.5% in 2002 to 90.1% in 2008. Considering the legal standard for the year of 2011, these values would be 64.9 and 60.5%, respectively. In relation to the solid composition of milk, the mean volume of milk in compliance with IN51 for fat, protein and solids nonfat were 95.2%, 98.2% and 89.6%, respectively. The season variations affected the quality of the bulk tank cooled raw milk from bulk tanks. The implantation of a payment system for the quality of milk to the producer revealed to be efficient in its objective to improve the quality indexes of raw milk.

Seropositivity for and intestinal colonization with Entamoeba histolytica and entamoeba dispar in individuals in northeastern Brazil.

In a slum community in northeastern Brazil 20% of a sample population was colonized with Entamoeba histolytica or Entamoeba dispar and 10.6% was colonized with E. histolytica alone. No correlation between seropositivity for anti-Ga1NAc lectin antibody and colonization was found. These results suggest that colonization does not necessarily produce immunity to reinfection.

Seroepidemiology of Entamoeba histolytica in a slum in northeastern Brazil.

Infection with the human pathogenic parasite Entamoeba histolytica has not been well-characterized in northeastern Brazil. In this study, the prevalence of E. histolytica infection in a slum in northeastern Brazil was assayed using an enzyme-linked immunosorbent assay (ELISA) for antibodies against the galactose/N-acetyl-D-galactosamine (Gal/GalNAc)-inhibitable adherence lectin of E. histolytica. Sera from a total of 335 individuals were examined for anti-Gal/GalNAc lectin antibodies. The overall seropositivity was 24.7%; 29.4% of females and 19.4% of males were positive. Among different age groups there was a peak of 40% positivity in the 6-14-year-old age group. There was also familial clustering of seropositivity. To examine colonization, stool samples from 155 people were examined microscopically for the presence of the parasite. Fourteen of 155 stools (9.0%) were identified as containing E. histolytica or nonpathogenic E. dispar. These 14 positive stools were analyzed with an ELISA that detects Gal/GalNAc lectin antigen and can distinguish between E. histolytica and E. dispar. Four stools (29%) were positive for E. histolytica and the remaining 10 were identified as E. dispar-positive. Although the overall colonization rate by microscopy was only 9%, with a third identified as E. histolytica, up to 40% of older children develop serologic evidence of having experienced pathogenic E. histolytica infection. The results of this study demonstrate that this community in northeastern Brazil is highly endemic for E. histolytica with infection rates similar to other developing nations.

A middle path: slavery and natural increase in nineteenth-century Minas Gerais.

"This article challenges the conventional notion that all regions of Brazil...were entirely dependent upon the international slave trade for the maintenance or growth of their slave populations.... The arguments will be based on data relating to the nineteenth-century slave population of the province of Minas Gerais.... Despite the size of its slave contingent, Minas Gerais was at best weakly linked to the export sector.... In light of data from the 1830s it will be argued that this economic transformation away from the export sector had a profound impact on slave demography.... New evidence shows that the Mineiro slave population achieved a positive rate of natural increase within a single generation of the termination of the Atlantic slave trade."