Bacteria isolated from Aedes aegypti with potential vector control applications.

Highly anthropophilic and adapted to urban environments, Aedes aegypti mosquitoes are the main vectors of arboviruses that cause human diseases such as dengue, zika, and chikungunya fever, especially in countries with tropical and subtropical climates. Microorganisms with mosquitocidal and larvicidal activities have been suggested as environmentally safe alternatives to chemical or mechanical mosquito control methods. Here, we analyzed cultivable bacteria isolated from all stages of the mosquito life cycle for their larvicidal activity against Ae. aegypti. A total of 424 bacterial strains isolated from eggs, larvae, pupae, or adult Ae. aegypti were analyzed for the pathogenic potential of their crude cultures against larvae of this same mosquito species. Nine strains displayed larvicidal activity comparable to the strain AM65-52, reisolated from commercial BTi-based product VectoBac® WG. 16S rRNA gene sequencing revealed that the set of larvicidal strains contains two representatives of the genus Bacillus, five Enterobacter, and two Stenotrophomonas. This study demonstrates that some bacteria isolated from Ae. aegypti are pathogenic for the mosquito from which they were isolated. The data are promising for developing novel bioinsecticides for the control of these medically important mosquitoes.

Antigen-Specific Antibody Signature Is Associated with COVID-19 Outcome.

Numerous studies have focused on inflammation-related markers to understand COVID-19. In this study, we performed a comparative analysis of spike (S) and nucleocapsid (N) protein-specific IgA, total IgG and IgG subclass response in COVID-19 patients and compared this to their disease outcome. We observed that the SARS-CoV-2 infection elicits a robust IgA and IgG response against the N-terminal (N1) and C-terminal (N3) region of the N protein, whereas we failed to detect IgA antibodies and observed a weak IgG response against the disordered linker region (N2) in COVID-19 patients. N and S protein-specific IgG1, IgG2 and IgG3 response was significantly elevated in hospitalized patients with severe disease compared to outpatients with non-severe disease. IgA and total IgG antibody reactivity gradually increased after the first week of symptoms. Magnitude of RBD-ACE2 blocking antibodies identified in a competitive assay and neutralizing antibodies detected by PRNT assay correlated with disease severity. Generally, the IgA and total IgG response between the discharged and deceased COVID-19 patients was similar. However, significant differences in the ratio of IgG subclass antibodies were observed between discharged and deceased patients, especially towards the disordered linker region of the N protein. Overall, SARS-CoV-2 infection is linked to an elevated blood antibody response in severe patients compared to non-severe patients. Monitoring of antigen-specific serological response could be an important tool to accompany disease progression and improve outcomes.

Using an Aluminum Hydroxide-Chitosan Matrix Increased the Vaccine Potential and Immune Response of Mice against Multi-Drug-Resistant Acinetobacter baumannii.

Acinetobacter baumannii is a Gram-negative, immobile, aerobic nosocomial opportunistic coccobacillus that causes pneumonia, septicemia, and urinary tract infections in immunosuppressed patients. There are no commercially available alternative antimicrobials, and multi-drug resistance is an urgent concern that requires emergency measures and new therapeutic strategies. This study evaluated a multi-drug-resistant A. baumannii whole-cell vaccine, inactivated and adsorbed on an aluminum hydroxide-chitosan (mAhC) matrix, in an A. baumannii sepsis model in immunosuppressed mice by cyclophosphamide (CY). CY-treated mice were divided into immunized, non-immunized, and adjuvant-inoculated groups. Three vaccine doses were given at 0D, 14D, and 28D, followed by a lethal dose of 4.0 × 108 CFU/mL of A. baumannii. Immunized CY-treated mice underwent a significant humoral response, with the highest IgG levels and a higher survival rate (85%); this differed from the non-immunized CY-treated mice, none of whom survived (p < 0.001), and from the adjuvant group, with 45% survival (p < 0.05). Histological data revealed the evident expansion of white spleen pulp from immunized CY-treated mice, whereas, in non-immunized and adjuvanted CY-treated mice, there was more significant organ tissue damage. Our results confirmed the proof-of-concept of the immune response and vaccine protection in a sepsis model in CY-treated mice, contributing to the advancement of new alternatives for protection against A. baumannii infections.

An Electrochemical Immunosensor Based on Carboxylated Graphene/SPCE for IgG-SARS-CoV-2 Nucleocapsid Determination.

The COVID-19 pandemic has emphasized the importance and urgent need for rapid and accurate diagnostic tests for detecting and screening this infection. Our proposal was to develop a biosensor based on an ELISA immunoassay for monitoring antibodies against SARS-CoV-2 in human serum samples. The nucleocapsid protein (N protein) from SARS-CoV-2 was employed as a specific receptor for the detection of SARS-CoV-2 nucleocapsid immunoglobulin G. N protein was immobilized on the surface of a screen-printed carbon electrode (SPCE) modified with carboxylated graphene (CG). The percentage of IgG-SARS-CoV-2 nucleocapsid present was quantified using a secondary antibody labeled with horseradish peroxidase (HRP) (anti-IgG-HRP) catalyzed using 3,3',5,5'-tetramethylbenzidine (TMB) mediator by chronoamperometry. A linear response was obtained in the range of 1:1000-1:200 v/v in phosphate buffer solution (PBS), and the detection limit calculated was 1:4947 v/v. The chronoamperometric method showed electrical signals directly proportional to antibody concentrations due to antigen-antibody (Ag-Ab) specific and stable binding reaction.

Bacillus subtilis spores as delivery system for nasal Plasmodium falciparum circumsporozoite surface protein immunization in a murine model.

Malaria remains a widespread public health problem in tropical and subtropical regions around the world, and there is still no vaccine available for full protection. In recent years, it has been observed that spores of Bacillus subtillis can act as a vaccine carrier and adjuvant, promoting an elevated humoral response after co-administration with antigens either coupled or integrated to their surface. In our study, B. subtillis spores from the KO7 strain were used to couple the recombinant CSP protein of P. falciparum (rPfCSP), and the nasal humoral-induced immune response in Balb/C mice was evaluated. Our results demonstrate that the spores coupled to rPfCSP increase the immunogenicity of the antigen, which induces high levels of serum IgG, and with balanced Th1/Th2 immune response, being detected antibodies in serum samples for 250 days. Therefore, the use of B. subtilis spores appears to be promising for use as an adjuvant in a vaccine formulation.

Yeasts with Fermentative Potential Associated with Fruits of Camu-Camu (Myrciaria dubia, Kunth) from North of Brazilian Amazon.

Considering the high biotechnological potential of yeasts associated to edible fruits, a screening for these microorganisms, capable of alcoholic fermentation, was performed in ripe fruits of camu-camu (Myrciaria dubia, Kunth). The fruits were collected from north of Brazilian Amazon, in the floodplain of the Cauamé River. Yeasts were isolated, and fermentation capability was evaluated using Durham tubes. Quantitative assays were performed to calculate ethanol yield (g g-1), specific growth rate (h-1), and ethanol productivity (g L-1·h-1). Taxonomic identification was performed by ribosomal gene nucleotide sequence analysis by alignment using BLASTN. A total of fifteen yeast colonies were isolated, and three of them presented promising ability to ferment glucose to ethanol. These isolates were identified as Candida orthopsilosis, Pichia kudriavzevii, and Meyerozyma caribbica. When cultured in broth containing 180 g·L-1 of glucose, M. caribbica CC003 reached 91.7 percent of the maximum theoretical ethanol concentration (84.4 g·L-1), presenting an ethanol yield and productivity of 0.4688 g·g-1 and 0.781 g·L-1·h-1, respectively. These results indicate a promising potential of this isolate for bioprocess applications. This paper is a rare report of C. orthopsilosis with endophytic habit because most of the references indicate it as a human pathogen. Besides this, M. caribbica is a promising fermenter for alcoholic beverages due to its osmotolerance and high ethanol yield. This is the first paper reporting endophytic yeasts associated with fruits of Myrciaria dubia.

Enzymatic Hydrolysis of Lignocellulosic Biomass Using an Optimized Enzymatic Cocktail Prepared from Secretomes of Filamentous Fungi Isolated from Amazonian Biodiversity.

The use of lignocellulosic biomass (LCB) has emerged as one of the main strategies for generating renewable biofuels. For the efficient use of such feedstock, pre-treatments are essential. The hydrolysis of cellulose - major component of LCB - demands enzymatic cocktails with improved efficiency to generate fermentable sugars. In this scenario, lignocellulolytic fungi have enormous potential for the development of efficient enzyme platforms. In this study, two enzymatic cocktails were developed for hydrolysis of two lignocellulosic biomasses: industrial cellulose pulp and cassava peel. The solid biomass ratio in relation to the protein content of the enzyme cocktail was performed by experimental design. The optimized cocktail for the hydrolysis of cellulose pulp (AMZ 1) was composed, in protein base, by 43% of Aspergillus sp. LMI03 enzyme extract and 57% of T. reesei QM9414, while the optimal enzyme cocktail for cassava peel hydrolysis (AMZ 2) was composed by 50% of Aspergillus sp. LMI03 enzyme extract, 25% of the extract of P. citrinum LMI01 and 25% of T. reesei. The ratio between solids and protein loading for AMZ 1 cocktail performance was 52 g/L solids and 30 mg protein/g solids, resulting in a hydrolytic efficiency of 93%. For the AMZ 2 cocktail, the hydrolytic efficiency was 78% for an optimized ratio of 78 g/L solids and 19 mg protein/g solids. These results indicate that cocktails formulated with enzymatic extracts of P. citrinum LMI01, Aspergillus sp. LMI03, and T. reesei QM9414 are excellent alternatives for efficient hydrolysis of plant biomass and for other processes that depend on biocatalysis.

Physical Mapping of the Anopheles (Nyssorhynchus) darlingi Genomic Scaffolds.

The genome assembly of Anopheles darlingi consists of 2221 scaffolds (N50 = 115,072 bp) and has a size spanning 136.94 Mbp. This assembly represents one of the smallest genomes among Anopheles species. Anopheles darlingi genomic DNA fragments of ~37 Kb were cloned, end-sequenced, and used as probes for fluorescence in situ hybridization (FISH) with salivary gland polytene chromosomes. In total, we mapped nine DNA probes to scaffolds and autosomal arms. Comparative analysis of the An. darlingi scaffolds with homologous sequences of the Anopheles albimanus and Anopheles gambiae genomes identified chromosomal rearrangements among these species. Our results confirmed that physical mapping is a useful tool for anchoring genome assemblies to mosquito chromosomes.

Mutational data and population profiling of 23 Y-STRs in three Brazilian populations.

Y-chromosomal STRs are important markers in forensic genetics, due to some peculiar characteristics. The absence of recombination makes them a useful tool to infer kinship in complex cases involving distant paternal relatives, or to infer paternal bio-geographic ancestry. The presence of a single copy, being transmitted from father to son, allow tracing mutational events in Y-STRs without ambiguity. For the statistical interpretation of forensic evidences based on Y-STR profiles, it is necessary to have estimates on both mutation rates and haplotype frequencies. In this work, 407 father-son duos from São Paulo and Rio de Janeiro states and 204 unrelated individuals from Manaus were analyzed. Haplotype frequencies and mutation rates for the Y-STRs from the PowerPlex Y23 commercial kit were estimated. Thirty-six mutations were observed in 15 of the 22 Y-STRs analyzed, for an average mutation rate of 3.84 × 10-3 (95 % CI 2.69 × 10-3 to 5.32 × 10-3). All mutations in GAAA repeats occurred in alleles with 13 or more uninterrupted units. Mutations in GATA repeats were observed in alleles with 9-17 uninterrupted units. An analysis carried out in different father's age groups showed an increase of 2.48 times the mutation rate in the age group of 40-50 years, when compared to the 20-30 age group, in agreement with the described for autosomal STRs. A high haplotype diversity was found in the three Brazilian populations. Pairwise genetic distance analysis (FST) showed no significant differences between the three populations in this study, which were also close to populations with strong European influence. The highest distances among the Brazilian populations were with São Gabriel da Cachoeira, which has a high Native American ancestry.

Frequency of gene ace i polymorphism i-d in athletes of different sports / Frequência do polimorfismo i-d do gene eca i em atletas de diferentes esportes / Frecuencia del polimorfismo i-d del gen eca i en atletas de diferentes deportes

ABSTRACT Introduction: The angiotensin-converting enzyme I-D (ACE) polymorphism gene is one of the most widely investigated genetic variations in sports science. Apparently, allele I is related to endurance sports, while allele D is related to power-strength activities. Nevertheless, studies have presented controversial results when it comes as to its occurrence in a variety of sports. Objective: This study aims to evaluate the frequency of gene ACE polymorphism I-D in professional athletes of collective or individual sports. Methods: Five mL blood were collected from 189 subjects divided into two groups: athletes (AG, n=127, wrestling, taekwondo, soccer, futsal and handball) and non-athletes (NAG, n=62). The athletes group was subdivided by group modalities, into: collective and individual. Both groups were further subdivided into male and female. Thus, we have the groups FAC= collective female, FAI= individual female, MAC= collective male, and MAI= individual male. The statistical analysis was carried out by frequency test, and the Hardy- Weinberg equilibrium by the x² test. Results: The results for the AG group indicated the following frequencies: DD=7%, ID=44% and II=49%. Allele frequency: D=29% and I=71%. For the NAG, the results were: DD=6.5%, ID=45.2% and II=48%. Allele frequency: D=29% and I=71%. The AG genotypic and allele frequencies did not differ statistically from those of the NAG (p= 0.982 and p= 0.984, respectively). However, we noticed that the genotypes II and ID frequencies were significantly higher than those of the DD. Conclusion: It can be concluded that the genotypic and allelic I-D frequencies of the ACE gene do not seem to influence performance in either group or individual sports. ACTN3 genotype frequencies did not vary significantly between male and female control subjects, and overall, there was no significant deviation from Hardy-Weinberg (H-W) equilibrium. Level of evidence I; Diagnostic studies-Investigating diagnostic test.

RESUMO Introdução: O polimorfismo I-D do gene da enzima conversora da angiotensina (ECA) é uma das variações genéticas mais amplamente investigadas na ciência do esporte. Aparentemente, o alelo I está relacionado aos esportes de resistência e o alelo D às atividades de força. Entretanto, os estudos têm apresentado resultados controversos quanto a sua ocorrência em diversos esportes. Objetivo: O presente estudo pretende avaliar a frequência do polimorfismo I-D do gene da ECA em atletas profissionais de esportes coletivos ou individuais. Métodos: Cinco mL de sangue foram coletados de 189 indivíduos divididos em dois grupos: atletas (GA, n=127, praticantes de luta livre, taekwondo, futebol, futsal ou handebol) e não atletas (GNA, n=62). O grupo de atletas foi subdividido de acordo com a modalidade: coletiva e individual. Ambos os grupos foram também subdivididos em masculino e feminino. Portanto, temos os grupos FAC = feminino coletivo, FAI = feminino individual, MAC = masculino coletivo, MAI = masculino individual. A análise estatística foi realizada através do teste de frequência e o equilíbrio de Hardy-Weinberg pelo teste x². Resultados: Os resultados para o GA indicaram as seguintes frequências: DD=7%, ID=44% e II=49%. Frequência alélica: D=29% e I=71%. Para o GNA, os resultados foram: DD=6,5%, ID=45,2% e II=48%. Frequência alélica: D=29% e I=71%. As frequências genotípicas e alélicas do GA não se diferiram estatisticamente daquelas do GNA (p= 0,982 e p= 0,984, respectivamente). Entretanto, notamos que as frequências dos genótipos II e ID se apresentaram significativamente maiores do que aquelas do DD. Conclusão: Pode-se concluir que as frequências I-D genotípicas e alélicas do gene da ECA não pareceram influenciar o desempenho tanto nos esportes individuais como coletivos. As frequências do genótipo ACTN3 não variaram significativamente entre os indivíduos de controle de ambos os sexos, e, no geral, não houve um desvio significativo do equilíbrio de Hardy-Weinberg (H-W). Nível de evidência I; Estudos diagnósticos-Investigação de um exame para diagnóstico.

RESUMEN Introducción: El polimorfismo I-D del gen de la enzima convertidora de la angiotensina (ECA) es una de las variaciones genéticas más ampliamente investigadas en la ciencia del deporte. Aparentemente, el alelo I está relacionado a los deportes de resistencia y el alelo D a las actividades de fuerza. Entretanto, los estudios han presentado resultados controvertidos cuanto a su ocurrencia en diversos deportes. Objetivo: El presente estudio pretende evaluar la frecuencia del polimorfismo I-D del gen de la ECA en atletas profesionales de deportes colectivos o individuales. Métodos: Cinco mL de sangre fueron recolectados de 189 individuos divididos en dos grupos: atletas (GA, n=127 practicantes de lucha libre, taekwondo, fútbol, futsal y hándbol) y no atletas (GNA, n=62). El grupo de atletas fue subdividido de acuerdo con la modalidad: colectiva e individual. Ambos grupos fueron también subdivididos en masculino y femenino: Por lo tanto, tenemos los grupos FAC= colectivo femenino, FAI= femenino individual, MAC= masculino colectivo, MAI= masculino individual. El análisis estadístico fue realizado a través del test de frecuencia, y el equilibrio de Hardy-Weinberg por el test x². Resultados: Los resultados de GA indicaran las siguientes frecuencias: DD=7%, ID=44% y II=49%. Frecuencia alélica: D=29% e I=71%. Para el GNA, los resultados fueron: DD=6,5%, ID=45,2% y II=48%. Frecuencia alélica: D=29% e I=71%. Las frecuencias genotípicas y alélicas del GA no difirieron estadísticamente de las del GNA (p=0,982 y p=0,984, respectivamente). Entretanto, notamos que las frecuencias de los genotipos II e ID se presentaron significativamente mayores que aquellas del DD. Conclusión: Se puede concluir que las frecuencias I-D genotípicas y alélicas del gen de la ECA no parecieron influenciar el desempeño tanto en los deportes individuales como colectivos. Las frecuencias del genotipo ACTN3 no variaron significativamente entre los individuos de control de ambos sexos y, en general, no hubo desvío significativo del equilibrio de Hardy-Weinberg (H-W). Nivel de evidencia I; Estudios diagnósticos-Investigación de un examen para diagnóstico.

Draft Genome Sequence of the Novel Enterobacter cloacae Strain amazonensis, a Highly Heavy Metal-Resistant Bacterium from a Contaminated Stream in Amazonas, Brazil.

Here, we report the draft genome of the Enterobacter cloacae strain amazonensis, a bacterium highly resistant to mercury that was isolated from a metal- and sewage-contaminated stream in Amazonas, Brazil. The exploration of the 5.0-Mb genome revealed 104 genes encoding resistance to toxic compounds and heavy metals, highlighting the potential biotechnological applications of this strain.

Draft Genome Sequence of Burkholderia gladioli Coa14, a Bacterium with Petroleum Bioremediation Potential Isolated from Coari Lake, Amazonas, Brazil.

Burkholderia gladioli Coa14 is a bacterium isolated from water collected from Coari Lake (Amazonas, Brazil) that shows a capacity for survival in a medium containing only oil as a carbon source. Here, we report its draft genome sequence, highlighting some genes involved with petroleum derivative degradation.

Assessment of petroleum biodegradation for Bacillus toyonensis by the using redox indicator 2,6 dichlorophenol indophenol / Avaliação da biodegradação do petróleo por Bacillus toyonensis, usando o indicador redox 2,6 diclorofenol indofenol

Petroleum degrading microorganisms have been isolated from different environments with the purpose of being used in bioremediation processes in areas impacted by petroleum spills. The objective of this study was to evaluate the ability of Bacillus toyonensis AM07 strain to metabolize petroleum compounds. The strain was isolated from the effluent dike of the Urucu Petroleum Province, Coari - Amazonas, Brazil. The degrading activity of B. toyonensis was evaluated by the colorimetric method using the redox indicator 2,6-dichlorophenol indophenol (DCPIP). Thus, the microorganism was inoculated into minimal medium with DCPIP, and with petroleum as the sole carbon source. The degradation potential of the microorganism was found by changing the DCPIP staining and absorbance readings 600nm. The results obtained demonstrated that the bacterial strain was able to degrade petroleum by altering the color of the medium from blue to colorless and by reducing the concentration of the indicator in the absorbance readings. B. toyonensis AM07 strain has shown good performance in the petroleum degradation assays and may be used in the future in remediation technologies for hydrocarbon impacted environments.

Microrganismos degradadores de petróleo têm sido isolados de diferentes ambientes com a finalidade de serem utilizados em processos de biorremediação de áreas impactadas com derrames de petróleo. O objetivo deste estudo foi avaliar a capacidade da linhagem de Bacillus toyonensis AM07, isolada do dique de efluente da Província Petrolífera de Urucu, Coari - Amazonas, Brasil, em metabolizar compostos do petróleo. A atividade degradadora do B. toyonensis foi avaliada pelo método colorimétrico, utilizando indicador redox 2,6-diclorofenol indofenol (DCPIP). Assim, o microrganismo foi inoculado em meio mínimo com DCPIP e petróleo como única fonte de carbono. O potencial de degradação do microrganismo foi constatado mediante a mudança de coloração DCPIP e leituras de absorbância 600nm. Os resultados obtidos demonstraram que a cepa bacteriana foi capaz de degradar petróleo, alterando a coloração do meio de azul para incolor e reduzindo a concentração do indicador nas leituras de absorbâncias. A cepa de B. toyonensis AM07 mostrou bom desempenho nos ensaios de degradação do petróleo, podendo ser utilizada, no futuro, em tecnologias de remediação de ambientes impactados por hidrocarbonetos.

Production of thermostable ß-glucosidase and CMCase by Penicillium sp: LMI01 isolated from the Amazon region

Background: Cellulolytic enzymes of microbial origin have great industrial importance because of their wide application in various industrial sectors. Fungi are considered the most efficient producers of these enzymes. Bioprospecting survey to identify fungal sources of biomass-hydrolyzing enzymes from a high-diversity environment is an important approach to discover interesting strains for bioprocess uses. In this study, we evaluated the production of endoglucanase (CMCase) and ß-glucosidase, enzymes from the lignocellulolytic complex, produced by a native fungus. Penicillium sp. LMI01 was isolated from decaying plant material in the Amazon region, and its performance was compared with that of the standard isolate Trichoderma reesei QM9414 under submerged fermentation conditions. Results: The effectiveness of LMI01 was similar to that of QM9414 in volumetric enzyme activity (U/mL); however, the specific enzyme activity (U/mg) of the former was higher, corresponding to 24.170 U/mg of CMCase and 1.345 U/mg of ß-glucosidase. The enzymes produced by LMI01 had the following physicochemical properties: CMCase activity was optimal at pH 4.2 and the ß-glucosidase activity was optimal at pH 6.0. Both CMCase and ß-glucosidase had an optimum temperature at 60°C and were thermostable between 50 and 60°C. The electrophoretic profile of the proteins secreted by LMI01 indicated that this isolate produced at least two enzymes with CMCase activity, with approximate molecular masses of 50 and 35 kDa, and ß-glucosidases with molecular masses between 70 and 100 kDa. Conclusions: The effectiveness and characteristics of these enzymes indicate that LMI01 can be an alternative for the hydrolysis of lignocellulosic materials and should be tested in commercial formulations.

Petroleum biodegrading and co-resistance to antibiotics by Serratia marcescens strain isolated in Coari, Amazonas / Biodegradação do petróleo e corresistência a antibióticos por Serratia marcescens isolada em Coari, Amazonas

Serratia marcescens is a Gram-negative bacillus, anaerobic facultative belonging to the family Enterobacteriaceae. S. marcescens strains are able to grow in the presence of different xenobiotic compounds, among them, petroleum and heavy metals. Xenobiotic resistant strains develop concomitant resistance to multiple antibiotics, referred to as co-resistance. The AMS212 strain was submitted to the microplate qualitative DCPIP - redox 2,6 dichlorophenol indophenol method. The quantitative test was carried out in Erlenmeyer flasks, followed by the change of color with the absorbance readings, trough the colorimetric method. The antibiotic resistance profile was evaluated by the Kirby -Bauer method. In the qualitative assay, the AMS212 strain altered the color of the DCPIP, which changed from blue to colorless, confirming that petroleum biodegradation occurred. In the quantitative test, the readings were decreasing, confirming that the concentration of DCPIP decreased as a function of the incubation time. The susceptibility test revealed that the AMS212 strain presented multiresistance to four different antibiotics. S. marcescens presented high performance in the biodegradation of petroleum, opening possibility to use it in projects involving the remediation of impacted areas. The expression of the antibiotic co-resistance phenotype confirms that the AMS212 strain is able to withstand different environmental aggressions.

Serratia marcescens é um bacilo Gram-negativo, anaeróbio facultativo, pertencente à família Enterobacteriaceae. Linhagens de S. marcescens são capazes de crescer na presença de diferentes compostos xenobióticos, dentre eles, petróleo e metais pesados. Linhagens resistentes a xenobióticos desenvolvem concomitante resistência a múltiplos antibióticos, denominada corresistência. A linhagem AMS212 foi submetida ao método colorimétrico com indicador DCPIP - redox 2,6 diclorofenol indofenol, qualitativo, em microplacas. O teste quantitativo foi realizado em frascos Erlenmeyer, acompanhando-se a mudança de coloração, com as leituras das absorbâncias. Avaliou-se o perfil de resistência a antibióticos pelo método de Kirby-Bauer. No ensaio qualitativo, a linhagem AMS212 alterou a cor do DCPIP, que passou de azul para incolor, confirmando que ocorreu biodegradação do petróleo. No teste quantitativo, as leituras foram decrescentes, confirmando que a concentração do DCPIP diminuiu em função do tempo de incubação. O teste de susceptibilidade revelou que a linhagem AMS212 apresenta multirresistência a quatro antibióticos diferentes. S. marcescens apresentou alto desempenho na biodegradação do petróleo, abrindo possibilidade de utilizá-la em projetos envolvendo a remediação de áreas impactadas. A expressão do fenótipo de corresistência a antibióticos confirma que a linhagem AMS212 é capaz de resistir a diferentes agressões ambientais.

Isolation and Characterization of Yeasts Able to Assimilate Sugarcane Bagasse Hemicellulosic Hydrolysate and Produce Xylitol Associated with Veturius transversus (Passalidae, Coleoptera, and Insecta).

Yeasts are an important component of insect gut microbial content, playing roles such as degradation of polymers and toxic compounds, biological control, and hormone, vitamin, and digestive enzyme production. The xylophagous beetle gut is a hyperdiverse habitat and a potential source of new species with industrial abilities such as enzyme production, pentose fermentation, and biodetoxification. In this work, samples of Veturius transversus (Passalidae, Coleoptera, and Insecta) were collected from the Central Amazon Rainforest. Their guts were dissected and a total of 20 microbial colonies were isolated using sugarcane bagasse hemicellulosic hydrolysate. They were identified as having 10 distinct biochemical profiles, and genetic analysis allowed identification as three clades in the genera Candida, Williopsis, and Geotrichum. All colonies were able to assimilate D-xylose and 18 were able to produce xylitol, especially a strain of Geotrichum, with a maximum yield of 0.502 g·g-1. These results agree with a previous prediction that the microbial community associated with xylophagous insects is a promising source of species of biotechnological interest.

Xylitol production and furfural consumption by a wild type Geotrichum sp

Background: Xylitol is a five carbons polyol with promising medical applications. It can be obtained from chemical D-xylose reduction or by microbial fermentation of Sugarcane Bagasse Hemicellulosic Hydrolysate. For this last process, some microbial inhibitors, as furfural, constitute severe bottleneck. In this case, the use of strains able to produce xylitol simultaneously to furfural neutralization is an interesting alternative. A wild-type strain of Geotrichum sp. was detected with this ability, and its performance in xylitol production and furfural consumption was evaluated. Furthermore, were analyzed its degradation products. Results: Geotrichum sp. produced xylitol from D-xylose fermentation with a yield of 0.44 g-g-1. Furfural was fully consumed in fermentation assay and when provided in the medium until concentration of 6 g-L-1. The furfural degradation product is not an identified molecule, presenting a molecular weight of 161 g-mol-1, an uncommon feature for the microbial metabolism of this product. Conclusion: This strain presents most remarkable potential in performing furfural consumption simultaneous to xylitol production. Subsequent efforts must be employed to establish bioprocess to simultaneous detoxification and xylitol production by Geotrichum sp.

Endophytic cultivable bacterial community obtained from the Paullinia cupana seed in Amazonas and Bahia regions and its antagonistic effects against Colletotrichum gloeosporioides.

Guarana (Paullinia cupana var. sorbilis) is a plant from the Amazonas region with socio-economic importance. However, guarana production has been increasingly affected by unfavorable conditions resulting from anthracnose, caused by the Colletotrichum fungal genus, which primarily affects mainly the Amazonas region. The aim of the present study was to isolate bacterial endophytes from the seeds of guarana plants obtained from Amazonas region and the Northeast state of Bahia, a region where this disease is not a problem for guarana plantations. The number of bacterial Colony Forming Units (CFU/g seeds) was 2.4 × 10(4) from the Bahia and 2.9 × 10(4) from the Amazonas region. One hundred and two isolated bacteria were evaluated in vitro against the phytopathogenic strain Colletotrichum gloeosporioides L1. These isolates were also analyzed for the enzymatic production of amylase, cellulase, protease, pectinase, lipase and esterase. Approximately 15% of isolates, showing high antagonistic activity, and the production of at least one enzyme were identified through the partial sequencing of 16S rDNA. The genus Bacillus was the most frequently observed, followed by Paenibacillus, Ochrobactrum, Microbacterium and Stenotrophomonas. Proteolytic activity was observed in 24 isolates followed by amylolytic, pectinolytic and cellulolytic activities. No esterase and lipase production was detected. Most of the isolates, showing antagonistic effects against C. gloeosporioides and high enzymatic activities, were isolated from the anthracnose-affected region. A biocontrol method using the endophytes from guarana seeds could be applied in the future, as these bacteria are vertically transferred to guarana seedlings.

Molecular characterization of the gene feminizer in the stingless bee Melipona interrupta (Hymenoptera: Apidae) reveals association to sex and caste development.

In highly eusocial insects, development of reproductive traits are regulated not only by sex determination pathway, but it also depends on caste fate. The molecular basis of both mechanisms in stingless bees and possible interaction with each other is still obscure. Here, we investigate sex determination in Melipona interrupta, focusing on characterization and expression analysis of the feminizer gene (Mi-fem), and its association to a major component of caste determination, the juvenile hormone (JH). We present evidence that Mi-fem mRNA is sex-specifically spliced in which only the female splice variant encodes the full length protein, following the same principle known for other bee species. We quantified Mi-fem expression among developmental stages, sexes and castes. Mi-fem expression varies considerably throughout development, with higher expression levels in embryos. Also, fem levels in pupae and newly emerged adults were significantly higher in queens than workers and males. Finally, we ectopically applied JH in cocoon spinning larvae, which correspond to the time window where queen/worker phenotypes diverge. We observed a significantly increase in Mi-fem expression compared to control groups. Since up to 100% of females turn into queens when treated with JH (while control groups are composed mainly of workers), we propose that fem might act to regulate queens' development. Our findings provide support for the conserved regulatory function of fem in Melipona bees and demonstrate a significant correlation between key elements of sex and caste determination pathways, opening the avenue to further investigate the molecular basis of these complex traits.

Endophytic bacterial diversity in the phyllosphere of Amazon Paullinia cupana associated with asymptomatic and symptomatic anthracnose.

Endophytes colonize an ecological niche similar to that of phytopathogens, which make them candidate for disease suppression. Anthracnose is a disease caused by Colletotrichum spp., a phytopathogen that can infect guarana (Paullinia cupana), an important commercial crop in the Brazilian Amazon. We investigated the diversity of endophytic bacteria inhabiting the phyllosphere of asymptomatic and symptomatic anthracnose guarana plants. The PCR-denaturation gradient gel electrophoresis (PCR-DGGE) fingerprints revealed differences in the structure of the evaluated communities. Detailed analysis of endophytic bacteria composition using culture-dependent and 16S rRNA clone libraries revealed the presence of Firmicutes, Proteobacteria, Actinobacteria, Bacteroidetes, and Acidobacteria phyla. Firmicutes comprised the majority of isolates in asymptomatic plants (2.40E(-4)). However, cloning and sequencing of 16S rRNA revealed differences at the genus level for Neisseria (1.4E(-4)), Haemophilus (2.1E(-3)) and Arsenophonus (3.6E(-5)) in asymptomatic plants, Aquicella (3.5E(-3)) in symptomatic anthracnose plants, and Pseudomonas (1.1E(-3)), which was mainly identified in asymptomatic plants. In cross-comparisons of the endophytic bacterial communities as a whole, symptomatic anthracnose plants contained higher diversity, as reflected in the Shannon-Weaver and Simpson indices estimation (P < 0.05). Similarly, comparisons using LIBSHUFF and heatmap analysis for the relative abundance of operational taxonomic units (OTUs) showed differences between endophytic bacterial communities. These data are in agreement with the NMSD and ANOSIM analysis of DGGE profiles. Our results suggest that anthracnose can restructure endophytic bacterial communities by selecting certain strains in the phyllosphere of P. cupana. The understanding of these interactions is important for the development of strategies of biocontrol for Colletotrichum.