Native and heterologous production of bacteriocins from gram-positive microorganisms

In nature, microorganisms can present several mechanisms for setting intercommunication and defense. One of these mechanisms is related to the production of bacteriocins, which are peptides with antimicrobial activity. Bacteriocins can be found in Gram-positive and Gram-negative bacteria. Nevertheless, bacteriocins produced by Gram-positive bacteria are of particular interest due to the industrial use of several strains that belong to this group, especially lactic acid bacteria (LAB), which have the status of generally recognized as safe (GRAS) microorganisms. In this work, we will review recent tendencies in the field of invention and state of art related to bacteriocin production by Gram-positive microorganism. Hundred-eight patents related to Gram-positive bacteriocin producers have been disclosed since 1965, from which 57% are related bacteriocins derived from Lactococcus, Lactobacillus, Streptococcus, and Pediococcus strains. Surprisingly, patents regarding heterologous bacteriocins production were mainly presented just in the last decade. Although the major application of bacteriocins is concerned to food industry to control spoilage and foodborne bacteria, during the last years bacteriocin applications have been displacing to the diagnosis and treatment of cancer, and plant disease resistance and growth promotion.     

Streptacidiphilus jiangxiensis sp. nov., a novel actinomycete isolated from acidic rhizosphere soil in China

The taxonomic position of three acidophilic actinomycetes isolated from acidic rhizosphere soil was established using a polyphasic approach. The morphological and chemical properties of the isolates were found to be consistent with their assignment to the genus Streptacidiphilus. Almost complete 16S rRNA gene sequences determined for the strains were aligned with corresponding sequences of representatives of the genera Kitasatospora, Streptacidiphilus and Streptomyces and phylogenetic trees inferred using three tree-making algorithms. The organisms formed a distinct subclade within the Streptacidiphilus 16S rRNA gene tree. They also shared nearly identical phenotypic profiles and rep-PCR fingerprint patterns that readily distinguished them from representatives of the established species of Streptacidiphilus. It is evident from the genotypic and phenotypic data that the three isolates form a new species in the genus Streptacidiphilus. The name proposed for this new species is Streptacidiphilus jiangxiensis, the type strain is isolate 33214T (= AS 4.1857T = JCM 12277T).     

Function inferences from a molecular structural model of bacterial ParE toxin

Toxin-antitoxin (TA) systems contribute to plasmid stability by a mechanism that relies on the differential stabilities of the toxin and antitoxin proteins and leads to the killing of daughter bacteria that did not receive a plasmid copy at the cell division. ParE is the toxic component of a TA system that constitutes along with RelE an important class of bacterial toxin called RelE/ParE superfamily. For ParE toxin, no crystallographic structure is available so far and rare in vitro studies demonstrated that the target of toxin activity is E. coli DNA gyrase. Here, a 3D Model for E. coli ParE toxin by molecular homology modeling was built using MODELLER, a program for comparative modeling. The Model was energy minimized by CHARMM and validated using PROCHECK and VERIFY3D programs. Resulting Ramachandran plot analysis it was found that the portion residues failing into the most favored and allowed regions was 96.8%. Structural similarity search employing DALI server showed as the best matches RelE and YoeB families. The Model also showed similarities with other microbial ribonucleases but in a small score. A possible homologous deep cleft active site was identified in the Model using CASTp program. Additional studies to investigate the nuclease activity in members of ParE family as well as to confirm the inhibitory replication activity are needed. The predicted Model allows initial inferences about the unexplored 3D structure of the ParE toxin and may be further used in rational design of molecules for structure-function studies.     

Detection of methanotrophs with highly divergent pmoA genes from Arctic soils

Tundra soil samples from the Canadian Arctic community, Kuujjuaq, were analyzed for the presence of the soluble (sMMO) and particulate (pMMO) methane monooxygenase genes. Total genomic DNA extracted from these soils was used as template for PCR using sMMO- and pMMO-specific primers, mmoX1-mmoX2 and A189-A682, respectively. pMMO and sMMO genes were detected in the Kuujjuaq soil samples. Isolation of sMMO-possessing methanotrophic microorganisms from the three soils, as determined by the colony naphthalene oxidation assay, was carried out using direct plating (5 degrees C) and methane enrichment studies (5 degrees C and 25 degrees C). Direct plating did not yield sMMO-possessing methanotrophic bacteria, whereas methane enrichments yielded isolates possessing and expressing sMMO activity. Analysis of derived amino acid sequences of pmoA genes and partial 16S rRNA genes obtained by PCR, using DNA isolated directly from this environment and from isolates, revealed the presence of highly divergent PmoA/AmoA sequences and 16S rRNA sequences that cluster closely with but are distinct from the genes from the genera Methylosinus and Methylocystis.     

Development of a panel of genome-wide ancestry informative markers to study admixture throughout the Americas

Most individuals throughout the Americas are admixed descendants of Native American, European, and African ancestors. Complex historical factors have resulted in varying proportions of ancestral contributions between individuals within and among ethnic groups. We developed a panel of 446 ancestry informative markers (AIMs) optimized to estimate ancestral proportions in individuals and populations throughout Latin America. We used genome-wide data from 953 individuals from diverse African, European, and Native American populations to select AIMs optimized for each of the three main continental populations that form the basis of modern Latin American populations. We selected markers on the basis of locus-specific branch length to be informative, well distributed throughout the genome, capable of being genotyped on widely available commercial platforms, and applicable throughout the Americas by minimizing within-continent heterogeneity. We then validated the panel in samples from four admixed populations by comparing ancestry estimates based on the AIMs panel to estimates based on genome-wide association study (GWAS) data. The panel provided balanced discriminatory power among the three ancestral populations and accurate estimates of individual ancestry proportions (R2 > 0.9 for ancestral components with significant between-subject variance). Finally, we genotyped samples from 18 populations from Latin America using the AIMs panel and estimated variability in ancestry within and between these populations. This panel and its reference genotype information will be useful resources to explore population history of admixture in Latin America and to correct for the potential effects of population stratification in admixed samples in the region.     

Melanoma RBPome identification reveals PDIA6 as an unconventional RNA-binding protein involved in metastasis

RNA-binding proteins (RBPs) have been relatively overlooked in cancer research despite their contribution to virtually every cancer hallmark. Here, we use RNA interactome capture (RIC) to characterize the melanoma RBPome and uncover novel RBPs involved in melanoma progression. Comparison of RIC profiles of a non-tumoral versus a metastatic cell line revealed prevalent changes in RNA-binding capacities that were not associated with changes in RBP levels. Extensive functional validation of a selected group of 24 RBPs using five different in vitro assays unveiled unanticipated roles of RBPs in melanoma malignancy. As proof-of-principle we focused on PDIA6, an ER-lumen chaperone that displayed a novel RNA-binding activity. We show that PDIA6 is involved in metastatic progression, map its RNA-binding domain, and find that RNA binding is required for PDIA6 tumorigenic properties. These results exemplify how RIC technologies can be harnessed to uncover novel vulnerabilities of cancer cells.     

Activin receptor ALK4 promotes adipose tissue hyperplasia by suppressing differentiation of adipocyte precursors

Adipocyte hyperplasia and hypertrophy are the two main processes contributing to adipose tissue expansion, yet the mechanisms that regulate and balance their involvement in obesity are incompletely understood. Activin B/GDF-3 receptor ALK7 is expressed in mature adipocytes and promotes adipocyte hypertrophy upon nutrient overload by suppressing adrenergic signaling and lipolysis. In contrast, the role of ALK4, the canonical pan-activin receptor, in adipose tissue is unknown. Here, we report that, unlike ALK7, ALK4 is preferentially expressed in adipocyte precursors, where it suppresses differentiation, allowing proliferation and adipose tissue expansion. ALK4 expression in adipose tissue increases upon nutrient overload and positively correlates with fat depot mass and body weight, suggesting a role in adipose tissue hyperplasia during obesity. Mechanistically, ALK4 signaling suppresses expression of CEBPα and PPARγ, two master regulators of adipocyte differentiation. Conversely, ALK4 deletion enhances CEBPα/PPARγ expression and induces premature adipocyte differentiation, which can be rescued by CEBPα knockdown. These results clarify the function of ALK4 in adipose tissue and highlight the contrasting roles of the two activin receptors in the regulation of adipocyte hyperplasia and hypertrophy during obesity.     

A novel application of entropy analysis for assessing changes in movement variability during cumulative tackles in young elite rugby league players

The aim of this study was to identify between-position (forwards vs. backs) differences in movement variability in cumulative tackle events training during both attacking and defensive roles. Eleven elite adolescent male rugby league players volunteered to participate in this study (mean ± SD, age; 18.5 ± 0.5 years, height; 179.5 ± 5.0 cm, body mass; 88.3 ± 13.0 kg). Participants performed a drill encompassing four blocks of six tackling (i.e. tackling an opponent) and six tackled (i.e. being tackled by an opponent while carrying a ball) events (i.e. 48 total tackles) while wearing a micro-technological inertial measurement unit (WIMU, Realtrack Systems, Spain). The acceleration data were used to calculate sample entropy (SampEn) to analyse the movement variability during tackles performance. In tackling actions SampEn showed significant between-position differences in block 1 (p = 0.0001) and block 2 (p = 0.0003). Significant between-block differences were observed in backs (block 1 vs 3, p = 0,0021; and block 1 vs 4, p = 0,0001) but not in forwards. When being tackled, SampEn showed significant between-position differences in block 1 (p = 0.0007) and block 3 (p = 0.0118). Significant between-block differences were only observed for backs in block 1 vs 4 (p = 0,0025). Movement variability shows a progressive reduction with cumulative tackle events, especially in backs and when in the defensive role (tackling). Forwards present lower movement variability values in all blocks, particularly in the first block, both in the attacking and defensive role. Entropy measures can be used by practitioners as an alternative tool to analyse the temporal structure of variability of tackle actions and quantify the load of these actions according to playing position.     

Bacterial division ring stabilizing ZapA versus destabilizing SlmA modulate FtsZ switching between biomolecular condensates and polymers

Cytokinesis is a fundamental process for bacterial survival and proliferation, involving the formation of a ring by filaments of the GTPase FtsZ, spatio-temporally regulated through the coordinated action of several factors. The mechanisms of this regulation remain largely unsolved, but the inhibition of FtsZ polymerization by the nucleoid occlusion factor SlmA and filament stabilization by the widely conserved cross-linking protein ZapA are known to play key roles. It was recently described that FtsZ, SlmA and its target DNA sequences (SlmA-binding sequence (SBS)) form phase-separated biomolecular condensates, a type of structure associated with cellular compartmentalization and resistance to stress. Using biochemical reconstitution and orthogonal biophysical approaches, we show that FtsZ-SlmA-SBS condensates captured ZapA in crowding conditions and when encapsulated inside cell-like microfluidics microdroplets. We found that, through non-competitive binding, the nucleotide-dependent FtsZ condensate/polymer interconversion was regulated by the ZapA/SlmA ratio. This suggests a highly concentration-responsive tuning of the interconversion that favours FtsZ polymer stabilization by ZapA under conditions mimicking intracellular crowding. These results highlight the importance of biomolecular condensates as concentration hubs for bacterial division factors, which can provide clues to their role in cell function and bacterial survival of stress conditions, such as those generated by antibiotic treatment.     

Paracoccidioidomycosis in people living with HIV/AIDS: A historical retrospective cohort study in a national reference center for infectious diseases,Rio de Janeiro,Brazil

Paracoccidioidomycosis (PCM) is one of the main endemic systemic mycoses in Latin America, usually occurring in rural areas. When PCM occurs simultaneously with underlying immunosuppressive conditions, it can present as an opportunistic disease. Between 2000 and 2017, literature reported around 200 PCM cases in people living with HIV/AIDS (PLWHA). To address research gaps on this co-infection and to study its possible temporal changes in the last decade, we performed an active co-infection case search on the HIV/AIDS and PCM cohorts from a Brazilian reference center database from 1989 to 2019. We found 20 PLWHA among 684 PCM patients (2.92%), predominantly male (70.0%) and urban workers (80.0%). The median age of patients was higher in the 2010–2019 decade (p = 0.006). The occurrence of PCM in PLWHA was lower when compared with other fungal diseases. Although 50.0% of the patients had already been diagnosed with HIV infection and presented CD4+ T cell counts greater than 200/mm³ at the time of PCM diagnosis, the suspicion of immunosuppression in the context of atypical and more severe clinical forms of PCM revealed the diagnosis of HIV infection in 35.0% of the patients. Two (10.0%) patients had an evolution compatible with immune reconstitution inflammatory syndrome (IRIS) after starting antiretroviral therapy (ART).We highlight the importance of considering a PCM diagnosis in PLWHA to prevent a late-onset treatment and progression to severe manifestations and unfavorable outcomes. In addition, HIV investigation is recommended in PCM patients, especially those with atypical and more severe clinical presentations.