RESUMO
Probiotics are live microorganisms that, when administered in adequate quantities, provide health benefits to the host. In this study, phenotypic and genotypic methods were used to evaluate the probiotic properties of Bacillus altitudinis 1.4. The isolate was sensitive to all antimicrobials tested and presented a positive result in the hemolysis test. B. altitudinis 1.4 spores were more resistant than vegetative cells, when evaluated in simulation of cell viability in the gastrointestinal tract, as well as adhesion to the intestinal mucosa. The isolate was capable of self-aggregation and coaggregation with pathogens such as Escherichia coli ATCC 25922 and Salmonella Enteritidis ATCC 13076. Genomic analysis revealed the presence of genes with probiotic characteristics. From this study it was possible to evaluate the gene expression of pro-inflammatory and anti-inflammatory cytokines for different treatments. Viable vegetative cells of B. altitudinis 1.4 increased the transcription of pro-inflammatory factors, in addition to also increasing the transcription of IL-10, indicating a tendency to stimulate a pro-inflammatory profile. Given the results presented, B. altitudinis 1.4 showed potential to be applied in the incorporation of this microorganism into animal feed, since the spores could tolerate the feed handling and pelletization processes.
Assuntos
Bacillus , Genoma Bacteriano , Probióticos , Probióticos/farmacologia , Bacillus/genética , Fatores Imunológicos/farmacologia , Citocinas/metabolismo , Citocinas/genética , Escherichia coli/genética , Esporos Bacterianos/genética , Aderência Bacteriana , Salmonella enteritidis/genética , Ração Animal/microbiologia , Antibacterianos/farmacologia , AnimaisRESUMO
Gelatin, a versatile protein derived from collagen, is widely used in the food, pharmaceutical and medical sectors. However, bacterial contamination by spore-forming bacteria during gelatin processing represents a significant concern for product safety and quality. In this study, an investigation was carried out to explore the heat and chemical resistance, as well as the identification and characterization of spore-forming bacteria isolated from gelatin processing. The methodologies involved chemical resistance tests with drastic pH in microplates and thermal resistance tests in capillary tubes of various isolates obtained at different processing stages. In addition, phenotypic and genotypic analyses were carried out to characterize the most resistant isolates of spore-forming bacteria. The findings of this study revealed the presence of several species, including Bacillus cereus, Bacillus licheniformis, Bacillus sonorensis, Bacillus subtilis, Geobacillus stearothermophilus, and Clostridium sporogenes, with some isolates exhibiting remarkable chemical and heat resistances. In addition, a significant proportion of the most resistant isolates showed gelatinase activity (n = 19/21; 90.5 %) and the presence of heat resistance (n = 5/21; 23.8 %), and virulence genes (n = 11/21; 52.4 %). The results of this study suggest that interventions should be done in quality control practices and that process parameter adjustments and effective contamination reduction strategies should be implemented through gelatin processing.
Assuntos
Gelatina , Temperatura Alta , RNA Ribossômico 16S , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Esporos Bacterianos , Esporos Bacterianos/genética , RNA Ribossômico 16S/genética , Virulência/genética , Microbiologia de Alimentos , Bacillus/genética , Bacillus/isolamento & purificaçãoRESUMO
Streptomyces are soil bacteria with complex life cycle. During sporulation Streptomyces linear chromosomes become highly compacted so that the genetic material fits within limited spore volume. The key players in this process are nucleoid-associated proteins (NAPs). Among them, HU (heat unstable) proteins are the most abundant NAPs in the cell and the most conserved in bacteria. HupS, one of the two HU homologues encoded by the Streptomyces genome, is the best-studied spore-associated NAP. In contrast to other HU homologues, HupS contains a long, C-terminal domain that is extremely rich in lysine repeats (LR domain) similar to eukaryotic histone H2B and mycobacterial HupB protein. Here, we have investigated, whether lysine residues in HupS are posttranslationally modified by reversible lysine acetylation. We have confirmed that Streptomyces venezuelae HupS is acetylated in vivo. We showed that HupS binding to DNA in vitro is controlled by the acetylation. Moreover, we identified that CobB1, one of two Sir2 homologues in Streptomyces, controls HupS acetylation levels in vivo. We demonstrate that the elimination of CobB1 increases HupS mobility, reduces chromosome compaction in spores, and affects spores maturation. Thus, our studies indicate that HupS acetylation affects its function by diminishing DNA binding and disturbing chromosome organization.
Assuntos
Proteínas de Bactérias , Esporos Bacterianos , Streptomyces , Streptomyces/genética , Streptomyces/metabolismo , Esporos Bacterianos/genética , Esporos Bacterianos/metabolismo , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Acetilação , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , DNA Bacteriano/metabolismo , DNA Bacteriano/genética , Ligação Proteica , Lisina/metabolismoRESUMO
Alicyclobacillus spp. is the cause of great concern for the food industry due to their spores' resistance (thermal and chemical) and the spoilage potential of some species. Despite this, not all Alicyclobacillus strains can spoil fruit juices. Thus, this study aimed to identify Alicyclobacillus spp. strains isolated from fruit-based products produced in Argentina, Brazil, and Italy by DNA sequencing. All Alicyclobacillus isolates were tested for guaiacol production by the peroxidase method. Positive strains for guaiacol production were individually inoculated at concentration of 103 CFU/mL in 10 mL of orange (pH 3.90) and apple (pH 3.50) juices adjusted to 11°Brix, following incubation at 45 °C for at least 5 days to induce the production of the following spoilage compounds: Guaiacol, 2,6-dichlorophenol (2,6-DCP) and 2,6-dibromophenol (2,6-DBP). The techniques of micro-solid phase extraction by headspace (HS-SPME) and gas-chromatography with mass spectrometry (GC-MS) were used to identify and quantify the spoilage compounds. All GC-MS data was analyzed by principal component analysis (PCA). The effects of different thermal shock conditions on the recovery of Alicyclobacillus spores inoculated in orange and apple juice (11°Brix) were also tested. A total of 484 strains were isolated from 48 brands, and the species A. acidocaldarius and A. acidoterrestris were the most found among all samples analyzed. In some samples from Argentina, the species A. vulcanalis and A. mali were also identified. The incidence of these two main species of Alicyclobacillus in this study was mainly in products from pear (n = 108; 22.3 %), peach (n = 99; 20.5 %), apple (n = 86; 17.8 %), and tomato (n = 63; 13 %). The results indicated that from the total isolates from Argentina (n = 414), Brazil (n = 54) and Italy (n = 16) were able to produce guaiacol: 107 (25.8 %), 33 (61.1 %) and 13 (81.2 %) isolates from each country, respectively. The PCA score plot indicated that the Argentina and Brazil isolates correlate with higher production of guaiacol and 2,6-DCP/2,6-DBP, respectively. Heatmaps of cell survival after heat shock demonstrated that strains with different levels of guaiacol production present different resistances according to spoilage ability. None of the Alicyclobacillus isolates survived heat shocks at 120 °C for 3 min. This work provides insights into the incidence, spoilage potential, and thermal shock resistance of Alicyclobacillus strains isolated from fruit-based products.
Assuntos
Alicyclobacillus , Sucos de Frutas e Vegetais , Frutas , Cromatografia Gasosa-Espectrometria de Massas , Guaiacol , Esporos Bacterianos , Alicyclobacillus/isolamento & purificação , Alicyclobacillus/genética , Alicyclobacillus/classificação , Alicyclobacillus/crescimento & desenvolvimento , Sucos de Frutas e Vegetais/microbiologia , Guaiacol/análogos & derivados , Guaiacol/metabolismo , Guaiacol/farmacologia , Frutas/microbiologia , Esporos Bacterianos/crescimento & desenvolvimento , Esporos Bacterianos/isolamento & purificação , Microbiologia de Alimentos , Contaminação de Alimentos/análise , Brasil , Microextração em Fase Sólida , Argentina , Malus/microbiologia , Itália , Temperatura Alta , Citrus sinensis/microbiologiaRESUMO
The production of whole-liquid eggs is of significant economic and nutritional importance. This study aimed to assess the phenotypic and genotypic diversity of mesophilic aerobic spore-forming bacteria (nâ¯=â¯200) isolated from pasteurized whole liquid egg and liquid egg yolk. The majority of the isolates were identified as belonging to the genera Bacillus (86â¯%), followed by Brevibacillus (10â¯%) and Lysinibacillus (4â¯%). For the phenotypic characterization, isolates were subjected to various heat shocks, with the most significant reductions observed at 80⯰C/30â¯min and 90⯰C/10â¯min for isolates recovered from raw materials. On the other hand, the decrease was similar for isolates recovered from raw material and final product at 100⯰C/5 min and 110⯰C/5 min. Genotypic genes related to heat resistance (cdnL, spoVAD, dacB, clpC, dnaK, and yitF/Tn1546) were examined for genotypic characterization. The dnaK gene showed a positive correlation with the highest thermal condition tested (110⯰C/5 min), while 100⯰C/5 min had the highest number of positively correlated genes (clpC, cdnL, yitF/Tn1546, and spoVAD). Whole Genome Sequencing of four strains revealed genes related to sporulation, structure formation, initiation and regulation, stress response, and DNA repair in vegetative cells. The findings of this study indicate that these mesophilic aerobic spore-forming bacteria may adopt several strategies to persist through the process and reach the final product. As the inactivation of these microorganisms during egg processing is challenging, preventing raw materials contamination and their establishment in processing premises must be reinforced.
Assuntos
Bacillus , Esporos Bacterianos , Esporos Bacterianos/genética , Bactérias , Cognição , Gema de OvoRESUMO
Sporeforming bacteria are a concern in some food raw materials, such as cocoa powder. Samples (n = 618) were collected on two farms and at several stages during cocoa powder manufacture in three commercial processing lines to determine the impact of each stage on bacterial spore populations. Mesophilic aerobic, mesophilic anaerobic, thermophilic aerobic, and Bacillus cereus spore populations were enumerated in all the samples. Genetic diversity in B. cereus strains (n = 110) isolated from the samples was examined by M13 sequence-based PCR typing, partial sequencing of the panC gene, and the presence/absence of ces and cspA genes. The counts of different groups of sporeforming bacteria varied amongst farms and processing lines. For example, the counts of mesophilic aerobic spore-forming (MAS) populations of cocoa bean fermentation were lower than 1 log spore/g in Farm 1 but higher than 4 log spore/g in Farm 2. B. cereus isolated from cocoa powder was also recovered from cocoa beans, nibs, and samples after roasting, refining, and pressing, which indicated that B. cereus spores persist throughout cocoa processing. Phylogenetic group IV was the most frequent (73%), along with processing. Strains from phylogenetic group III (14 %) did not show the ces gene's presence.
Assuntos
Bacillus cereus , Chocolate , Bacillus cereus/genética , Filogenia , Anaerobiose , Esporos Bacterianos/genética , Microbiologia de Alimentos , Contagem de Colônia MicrobianaRESUMO
SUMMARYThe metabolic conditions that prevail during bacterial growth have evolved with the faithful operation of repair systems that recognize and eliminate DNA lesions caused by intracellular and exogenous agents. This idea is supported by the low rate of spontaneous mutations (10-9) that occur in replicating cells, maintaining genome integrity. In contrast, when growth and/or replication cease, bacteria frequently process DNA lesions in an error-prone manner. DNA repairs provide cells with the tools needed for maintaining homeostasis during stressful conditions and depend on the developmental context in which repair events occur. Thus, different physiological scenarios can be anticipated. In nutritionally stressed bacteria, different components of the base excision repair pathway may process damaged DNA in an error-prone approach, promoting genetic variability. Interestingly, suppressing the mismatch repair machinery and activating specific DNA glycosylases promote stationary-phase mutations. Current evidence also suggests that in resting cells, coupling repair processes to actively transcribed genes may promote multiple genetic transactions that are advantageous for stressed cells. DNA repair during sporulation is of interest as a model to understand how transcriptional processes influence the formation of mutations in conditions where replication is halted. Current reports indicate that transcriptional coupling repair-dependent and -independent processes operate in differentiating cells to process spontaneous and induced DNA damage and that error-prone synthesis of DNA is involved in these events. These and other noncanonical ways of DNA repair that contribute to mutagenesis, survival, and evolution are reviewed in this manuscript.
Assuntos
Bacillus subtilis , Reparo do DNA , Mutagênese , Reparo do DNA/genética , Bacillus subtilis/genética , Bacillus subtilis/fisiologia , Estresse Fisiológico/genética , Dano ao DNA , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Replicação do DNA , DNA Bacteriano/genética , Esporos Bacterianos/genética , Esporos Bacterianos/crescimento & desenvolvimentoRESUMO
Two growth modes have been described for the filamentous Streptomyces bacteria. Their classic developmental life cycle culminates in the formation of dormant spores, where movement to new environments is mediated through spore dispersal. In contrast, exploratory growth proceeds as a rapidly expanding vegetative mycelium that leads to extensive surface colonization and is associated with the release of volatile compounds that promote alkalinization (and reduced iron bioavailability) of its surrounding environment. Here, we report that exploratory growth in Streptomyces venezuelae can proceed in tandem with classic sporulating development in response to specific nutritional cues. Sporulating exploration is not accompanied by a rise in environmental pH but has the same iron acquisition requirements as conventional exploration. We found that mutants that were defective in their ability to sporulate were unaffected in exploration, but mutants undergoing precocious sporulation were compromised in their exploratory growth and this appeared to be mediated through premature activation of the developmental regulator WhiI. Cell envelope integrity was also found to be critical for exploration, as mutations in the cell envelope stress-responsive extracytoplasmic function sigma factor SigE led to a failure to explore robustly under all exploration-promoting conditions. Finally, in expanding the known exploration-promoting conditions, we discovered that the model species Streptomyces lividans exhibited exploration capabilities, supporting the proposal that exploration is conserved across diverse streptomycetes. IMPORTANCE: Streptomyces bacteria have evolved diverse developmental and metabolic strategies to thrive in dynamic environmental niches. Here, we report the amalgamation of previously disparate developmental pathways, showing that colony expansion via exploration can proceed in tandem with colony sporulation. This developmental integration extends beyond phenotype to include shared genetic elements, with sporulation-specific repressors being required for successful exploration. Comparing this new exploration mode with previously identified strategies has revealed key differences (e.g., no need for environmental alkalinization), and simultaneously allowed us to define unifying requirements for Streptomyces exploration. The "reproductive exploration" phenomenon reported here represents a unique bet-hedging strategy, with the Streptomyces colony engaging in an aggressive colonization strategy while transporting a protected genetic repository.
Assuntos
Streptomyces , Animais , Streptomyces/metabolismo , Fatores de Transcrição/metabolismo , Ferro/metabolismo , Estágios do Ciclo de Vida , Esporos Bacterianos , Proteínas de Bactérias/metabolismoRESUMO
Clostridium botulinum is a foodborne pathogen responsible for severe neuroparalytic disease associated with the ingestion of pre-formed toxin in food, with processed meats and canned foods being the most affected. Control of this pathogen in meat products is carried out using the preservative sodium nitrite (NaNO2), which in food, under certain conditions, such as thermal processing and storage, can form carcinogenic compounds. Therefore, the objective was to use nanoemulsified essential oils (EOs) as natural antimicrobial agents, with the aim of reducing the dose of NaNO2 applied in mortadella. The antimicrobial activity of nanoemulsions prepared with mixtures of EOs of garlic, clove, pink pepper, and black pepper was evaluated on endospores and vegetative cells of C. botulinum and Clostridium sporogenes (surrogate model) inoculated in mortadella prepared with 50 parts per million NaNO2. The effects on the technological (pH, water activity, and color) and sensory characteristics of the product were also evaluated. The combinations of EOs and their nanoemulsions showed sporicidal effects on the endospores of both tested microorganisms, with no counts observed from the 10th day of analysis. Furthermore, bacteriostatic effects on the studied microorganisms were observed. Regarding the technological and sensorial characteristics of the product, the addition of the combined EOs had a negative impact on the color of the mortadella and on the flavor/aroma. Despite the strong commercial appeal of adding natural preservatives to foods, the effects on flavor and color must be considered. Given the importance of controlling C. botulinum in this type of product, as well as the reduction in the amount of NaNO2 used, this combination of EOs represents a promising antimicrobial alternative to this preservative, encouraging further research in this direction.
Assuntos
Clostridium botulinum , Clostridium , Produtos da Carne , Óleos Voláteis , Óleos Voláteis/farmacologia , Clostridium botulinum/efeitos dos fármacos , Produtos da Carne/microbiologia , Clostridium/efeitos dos fármacos , Microbiologia de Alimentos , Nitrito de Sódio/farmacologia , Emulsões , Humanos , Conservação de Alimentos/métodos , Esporos Bacterianos/efeitos dos fármacos , Conservantes de Alimentos/farmacologia , Paladar , Antibacterianos/farmacologiaRESUMO
Among the milk contaminating microorganisms, those which are able to form heat-resistant spores are concerning, especially for dairy companies that use ultra-high temperature (UHT) technology. These spores, throughout storage, can germinate and produce hydrolytic enzymes that compromise the quality of the final product. This study evaluated 184 UHT milk samples from different batches collected from seven Brazilian dairy companies with a possible microbial contamination problem. The bacteria were isolated, phenotypically characterized, clustered by REP-PCR, and identified through 16S rDNA sequencing. The presence of Bacillus sporothermodurans was verified using biochemical tests (Gram staining, catalase and oxidase test, glucose fermentation, esculin hydrolysis, nitrate reduction, and urease test). According to these tests, none of the isolates presented typical characteristics of B. sporothermodurans. In sequence, the isolates, that presented rod-shapes, were submitted to molecular analyses in order to determine the microbial biodiversity existing among them. The isolates obtained were grouped into 16 clusters, four of which were composed of only one individual. A phylogenetic tree was constructed using the sequences obtained from the 16S rDNA sequencing and some reference strains of species close to those found using BLAST search in the NCBI nucleotide database. Through this tree, it was possible to verify the division of the isolates into two large groups, the Bacillus subtilis and the Bacillus cereus groups. Furthermore, most isolates are phylogenetically closely related, which makes it even more difficult to identify them at the species level. In conclusion, it was possible to assess, in general, the groups of sporulated contaminants in Brazilian UHT milk produced in the regions evaluated. In addition, it was also possible to determine the biodiversity of spore-forming bacteria found in UHT milk samples, thus opening up a range of possible research topics regarding the effects of the presence of these microorganisms on milk quality.