Bacterial Communities in the Midgut of Ponerine Ants (Hymenoptera: Formicidae: Ponerinae)

Symbiotic microorganisms are directly related to the ecological success of host insects, influencing many aspects of their biology. The present study is the first to investigate the microbiota associated with ants of the subfamily Ponerinae and aims to identify the bacterial midgut communities of Dinoponera lucida, Pachycondyla curvinodis, Pachycondyla striata, Odontomachus brunneus and Odontomachus bauri. After dissecting the midguts of these ants, DNA was extracted, and the bacterial 16S rRNA gene was amplified via PCR using the universal primers pair 27F/1492R. The obtained PCR products were cloned and sequenced using an ABI 3500 automated sequencer. The sequences were grouped into operational taxonomic units (OTUs) based on a 97% similarity criterion using MOTHUR. The greatest species richness was observed in O. bauri, with 15 OTUs, followed by D. lucida with five OTUs, O. brunneus, with four OTUs, and P. curvinodis and P. striata, both with three OTUs. There were representatives of the phyla Actinobacteria, Proteobacteria, Tenericutes and Firmicutes, including the genera Bartonella, Mesoplasma, Mesorhizobium, Spiroplasma, Wolbachia and Serratia in the guts of the studied Ponerine ants. The low microbial diversity observed given the predatory trophic habits of the species studied suggests that there is selection for these microorganisms, predominantly preserving symbionts with functional roles that are able to colonize this environment. It is also valid to infer that the identified bacteria are predominant in the gut and exhibit mutualistic functions that are important for immunity, reproduction and nutrition; moreover, a subset may be parasites that have considerable impacts on the studied ants

Fungal communities in pressmud composting harbour beneficial and detrimental fungi for human welfare

Pressmud is a substrate derived from sugarcane juice filtrate, and around 26-40  kg of this residue are produced per ton of sugarcane. It is mainly used as fertilizer in crops, and its application in the field is often made without any prior treatment, but, in this research, it was studied for the risk this practice poses for human health. This research was stimulated by previous results indicating the presence of opportunistic pathogens in residues used in various composting systems and the extensive use of fresh pressmud in agriculture. Here, It was assessed the fungal diversity present in both fresh and composting pressmud using 454 pyrosequencing. In addition, heat-tolerant fungi were isolated and surveyed for their enzymatic repertoire of biomass-degrading enzymes (cellulase, xylanase, laccase and polygalacturonase). A wide range of opportunistic pathogens was found among the most abundant taxa in the fresh pressmud, such as Lomentospora prolificans (43.13 %), Trichosporon sp. (10.07 %), Candida tropicalis (7.91 %), and Hormographiella aspergillata (8.19 %). This indicates that fresh pressmud might be a putative source of human pathogenic fungi, presenting a potential threat to human health if applied as fertilizer without any treatment. With regard to the heat-tolerant fungi found in this substrate, all the 110 isolates screened were able to produce at least one of the tested enzymes. The pressmud composting process not only effectively reduces the load of pathogenic fungi, but also creates an interesting environment for fungi able to produce thermostable hydrolytic and oxidative enzymes with biotechnological applications

Comparison in the Trichoderma longibrachiatum xyloglucanase production using tamarind (Tamarindus indica) and jatobá (Hymenaea courbaril) seeds: factorial design and immobilization on ionic supports

in the control of the stretching and expansion of the plant cell wall. There are five types of enzymes known to be capable of cleaving the linear chain of xyloglucan, the most famous of them being the xyloglucanase (XEG). The immobilization can be used to solve problems related to stability, besides the economic benefits brought by the possibility of repeated use and recovery, decreasing the costs of production. Therefore, this study aims the optimization of the production of a xyloglucanase from Trichoderma longibrachiatum, with the aid of factorial design, using tamarind (Tamarindus indica) and jatobá (Hymenaea courbaril) seeds as carbon source; and the immobilization of the enzyme on ionic supports, such as MANAE (monoamino-N-aminoethyl), DEAE (diethylaminoethyl)-cellulose, CM (carboxymethyl)-cellulose and PEI (polyethyleneimine). High concentrations of carbon source in the culture medium, especially tamarind seeds, were the most favorable conditions for the greater activity of the xyloglucanase from T. longibrachiatum. The scaling up from Erlenmeyer flasks to the bioreactor was an essential strategy to increase the content of secreted enzyme. Regarding the biochemical characterization of the crude extract, the optimal temperature was 50-55 °C and the optimal pH 5.0. Regarding the stabilities to pH and to temperature, the enzyme was not stable for prolonged periods, which was crucial for the performing of immobilization on ionic resins (CM-cellulose, DEAE-cellulose, MANAE, and PEI), being the first time described in literature the immobilization of a xyloglucanase on these supports.We thank the Fundação de Amparo à Pesquisa do estado de São Paulo (process 2018/07522-6; 2014/50884-5), and Conselho Nacional de Dsenvolvimento Científico (process 301963/2017-7; 465319/2014-9).info:eu-repo/semantics/publishedVersio

Fungal communities in pressmud composting harbour beneficial and detrimental fungi for human welfare

Pressmud is a substrate derived from sugarcane juice filtrate, and around 26-40  kg of this residue are produced per ton of sugarcane. It is mainly used as fertilizer in crops, and its application in the field is often made without any prior treatment, but, in this research, it was studied for the risk this practice poses for human health. This research was stimulated by previous results indicating the presence of opportunistic pathogens in residues used in various composting systems and the extensive use of fresh pressmud in agriculture. Here, It was assessed the fungal diversity present in both fresh and composting pressmud using 454 pyrosequencing. In addition, heat-tolerant fungi were isolated and surveyed for their enzymatic repertoire of biomass-degrading enzymes (cellulase, xylanase, laccase and polygalacturonase). A wide range of opportunistic pathogens was found among the most abundant taxa in the fresh pressmud, such as Lomentospora prolificans (43.13 %), Trichosporon sp. (10.07 %), Candida tropicalis (7.91 %), and Hormographiella aspergillata (8.19 %). This indicates that fresh pressmud might be a putative source of human pathogenic fungi, presenting a potential threat to human health if applied as fertilizer without any treatment. With regard to the heat-tolerant fungi found in this substrate, all the 110 isolates screened were able to produce at least one of the tested enzymes. The pressmud composting process not only effectively reduces the load of pathogenic fungi, but also creates an interesting environment for fungi able to produce thermostable hydrolytic and oxidative enzymes with biotechnological applications

Genome mining for peptidases in heat-tolerant and mesophilic fungi and putative adaptations for thermostability

Abstract Background Peptidases (EC 3.4) consist of a large group of hydrolytic enzymes that catalyze the hydrolysis of proteins accounting for approximately 65% of the total worldwide enzyme production. Peptidases from thermophilic fungi have adaptations to high temperature that makes them adequate for biotechnological application. In the present study, we profiled the genomes of heat-tolerant fungi and phylogenetically related mesophilic species for genes encoding for peptidases and their putative adaptations for thermostability. Results We generated an extensive catalogue of these enzymes ranging from 241 to 820 peptidase genes in the genomes of 23 fungi. Thermophilic species presented the smallest number of peptidases encoding genes in relation to mesophilic species, and the peptidases families with a greater number of genes were the most affected. We observed differences in peptidases in thermophilic species in comparison to mesophilic counterparts, at (i) the genome level: a great reduction in the number of peptidases encoding genes that harbored a higher number of copies; (ii) in the primary protein structure: shifts in proportion of single or groups of amino acids; and (iii) in the three-dimensional structure: reduction in the number of internal cavities. Similar results were reported for extremely thermophilic proteins, but here we show for the first time that several changes also occurred on the moderate thermophilic enzymes of fungi. In regards to the amino acids composition, peptidases from thermophilic species in relation to the mesophilic ones, contained a larger proportion of Ala, Glu, Gly, Pro, Arg and Val residues and a lower number of Cys, His, Ile, Lys, Met, Asn, Gln, Ser, Thr and Trp residues (P < 0.05). Moreover, we observed an increase in the proportion of hydrophobic and charged amino acids and a decrease in polar amino acids. Conclusions Although thermophilic fungi present less genes encoding for peptidases, these have adaptations that could play a role in thermal resistance from genome to protein structure level

Percepção do consumidor sobre a higiene na comercialização de carnes em feira livre da cidade de Garanhuns – PE

Este trabalho teve como objetivo avaliar a percepção de consumidores em relação à higiene na comercialização de produtos cárneos na feira livre do bairro Heliópolis, na cidade de Garanhuns, Pernambuco. Para tanto, foi realizada uma pesquisa de caráter quantitativo, quando foram aplicados 50 questionários aos consumidores da feira. Os dados obtidos foram tabulados e analisados por meio da estatística descritiva. Os resultados revelaram que os consumidores estavam cientes que a forma de comercialização na feira livre era precária, com deficiências no aspecto higiênicossanitário, desde a obtenção da matéria-prima, acondicionamento, manipulação e transporte dos alimentos. Metade dos pesquisados (50%) apresentaram uma escolaridade elevada. Contudo, isto não influenciou no hábito em frequentar a feira, uma vez que, independentemente da escolaridade, todos (100%) concordaram com a continuidade desta forma de comercialização dos produtos cárneos. Apesar de relatarem disposição em pagar um valor mais elevado por um produto de melhor qualidade, a influência cultural é responsável pelo hábito em frequentar a feira semanalmente e adquirir tais produtos. Esta forma de comercialização representa riscos à saúde do consumidor, o que constitui um problema de saúde pública, que deve sofrer intervenção por parte dos órgãos fiscalizadores. A população deve ser alertada sobre tais riscos, enquanto os comerciantes, capacitados sobre os diversos aspectos relacionados a manipulação segura dos alimentos comercializados

Heterologous production and biochemical characterization of a new highly glucose tolerant GH1 β-glucosidase from Anoxybacillus thermarum

The enzymatic lignocellulosic biomass conversion into value-added products requires the use of enzyme-rich cocktails, including β-glucosidases that hydrolyze cellobiose and cellooligosaccharides to glucose. During hydrolysis occurs accumulation of monomers causing inhibition of some enzymes; thus, glucose/xylose tolerant β-glucosidases could overcome this drawback. The search of new tolerant enzymes showing additional properties,such as high activity, wide-pH range, and thermal stability is very relevant to improve the bioprocess. We describe a novel β-glucosidase GH1 from the thermophilic Anoxybacillus thermarum (BgAt), which stood out by the robustness combination of great glucose/xylose tolerance, thermal stability, and high Vmax. The recombinant his-tagged-BgAt was overexpressed in Escherichia coli, was purified in one step, showed a high glucose/xylose tolerance, and activity stimulation (presence of 0.4M glucose/1.0M xylose). The optimal activity was at 65 °C - pH 7.0. BgAt presented an extraordinary temperature stability (48 h – 50 °C), and pH stability (5.5–8.0). The novel enzyme showed outstanding Vmax values compared to other β-glucosidases. Using p-nitrophenyl-β-D-glucopyranoside as substrate the values were Vmax (7614 U/mg), and KM (0.360 mM). These values suffer a displacement in Vmax to 14,026 U/mg (glucose), 14,886 U/mg (xylose), and KM 0.877mM (glucose), and 1.410mM (xylose).This work was supported by Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP, process no 2018/07522-6) and a scholarship to Tassio B. Oliveira (grant 2017/09000-4). MLTMP is a Research Fellow of Conselho de Desenvolvimento Científico e Tecnológico (CNPq, process 301963/2017-7). The project also received grants from National Institute of Science and Technology of Bioethanol,INCT, CNPq 465319/2014-9/FAPESP nº 2014/50884-5). PZA was a fellow from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) funding code 001 and Programa de Doutorado Sanduiche no Exterior, PDSE no 88881.135684/2016-01.Peer reviewe

Comparison of <i>Trichoderma longibrachiatum</i> Xyloglucanase Production Using Tamarind (<i>Tamarindus indica</i>) and Jatoba (<i>Hymenaea courbaril</i>) Seeds: Factorial Design and Immobilization on Ionic Supports

Xyloglucan (XG) is the predominant hemicellulose in the primary cell wall of superior plants. It has a fundamental role in controlling the stretching and expansion of the plant cell wall. There are five types of enzymes known to cleave the linear chain of xyloglucan, and the most well-known is xyloglucanase (XEG). The immobilization process can be used to solve problems related to stability, besides the economic benefits brought by the possibility of its repeated use and recovery. Therefore, this study aims at the optimization of the xyloglucanase production of Trichoderma longibrachiatum using a central composite rotatable design (CCRD) with tamarind and jatoba seeds as carbon sources, as well as XEG immobilization on ionic supports, such as MANAE (monoamine-N-aminoethyl), DEAE (diethylaminoethyl)-cellulose, CM (carboxymethyl)-cellulose, and PEI (polyethyleneimine). High concentrations of carbon sources (1.705%), at a temperature of 30 °C and under agitation for 72 h, were the most favorable conditions for the XEG activity from T. longibrachiatum with respect to both carbon sources. However, the tamarind seeds showed 23.5% higher activity compared to the jatoba seeds. Therefore, this carbon source was chosen to continue the experiments. The scaling up from Erlenmeyer flasks to the bioreactor increased the XEG activity 1.27-fold (1.040 ± 0.088 U/mL). Regarding the biochemical characterization of the crude extract, the optimal temperature range was 50–55 °C, and the optimal pH was 5.0. Regarding the stabilities with respect to pH and temperature, XEG was not stable for prolonged periods, which was crucial to immobilizing it on ionic resins. XEG showed the best immobilization efficiency on CM-cellulose and DEAE-cellulose, with activities of 1.16 and 0.89 U/g of the derivative (enzyme plus support), respectively. This study describes, for the first time in the literature, the immobilization of a fungal xyloglucanase using these supports

Saccharification of different sugarcane bagasse varieties by enzymatic cocktails produced by Mycothermus thermophilus and Trichoderma reesei RP698 cultures in agro-industrial residues

Lignocellulosic biomasses are promising sources for the obtaining of clean energy through their bioconversion into ethanol. Their saccharification involves a multi-enzymatic system. Here, we evaluated the hydrolysis of a mixture of sugarcane bagasse varieties (SCB), Energy cane fiber (EC), and sugarcane bagasse from the SP80-3280 variety, all in natura, using fungal enzymatic extracts obtained from Mycothermus thermophilus and Trichoderma reesei RP698 cultures supplemented with various agro-industrial residues. The enzymatic extracts from both fungi, when grown in a corn cob and corn straw, led to the highest sugarcane hydrolysis. For M. thermophilus, the reducing sugars released (mg/mL) were 1.21 ± 0.12, 1.25 ± 0.14, and 0.98 ± 0.05 for SCB, EC and SP80-3280, respectively; for T. reesei, the reducing sugars were 0.84 ± 0.08, 0.89 ± 0.06 and 0.68 ± 0.03 for SCB, SP80-3280, and EC, respectively. The cocktail obtained from the co-cultivation of these fungi in corn straw at 35 °C showed the best hydrolysis results, the release of sugars (mg/mL) was 1.31 ± 0.06 (SCB), 2.18 ± 0.08 (EC) and 1.67 ± 0.09 (SP80-3280). Scanning electron microscopy and thermogravimetric analysis confirmed changes in the fiber structures after enzymatic hydrolysis. Thus, these fungi were shown to be promising for an enzymatic cocktail design and sugarcane biomass saccharification.This study was financed by the Coordenaçao de Aperfeiçoamento de Pessoal de Nível Superior - Brazil (CAPES) - Finance Code 001; Fundaçao de Amparo ~ a Pesquisa do Estado de Sao Paulo ~ (FAPESP, Grant 2014/50884-5, 2017/09000e4, 10/52322e3; 2018/07522e6), Portuguese Foundation for Science and Technology (FCT) e POCI-01-0145-FEDER-032206) and Conselho de Desenvolvimento Científico e Tecnologico (CNPq, Grant 465319/2014 e9). MLTMP is a Research Productivity fellow of CNPq (process 301963/2017e7).info:eu-repo/semantics/publishedVersio