Levels of Small Extracellular Vesicles Containing hERG-1 and Hsp47 as Potential Biomarkers for Cardiovascular Diseases.

The diagnosis of cardiovascular disease (CVD) is still limited. Therefore, this study demonstrates the presence of human ether-a-go-go-related gene 1 (hERG1) and heat shock protein 47 (Hsp47) on the surface of small extracellular vesicles (sEVs) in human peripheral blood and their association with CVD. In this research, 20 individuals with heart failure and 26 participants subjected to cardiac stress tests were enrolled. The associations between hERG1 and/or Hsp47 in sEVs and CVD were established using Western blot, flow cytometry, electron microscopy, ELISA, and nanoparticle tracking analysis. The results show that hERG1 and Hsp47 were present in sEV membranes, extravesicularly exposing the sequences 430AFLLKETEEGPPATE445 for hERG1 and 169ALQSINEWAAQTT- DGKLPEVTKDVERTD196 for Hsp47. In addition, upon exposure to hypoxia, rat primary cardiomyocytes released sEVs into the media, and human cardiomyocytes in culture also released sEVs containing hERG1 (EV-hERG1) and/or Hsp47 (EV-Hsp47). Moreover, the levels of sEVs increased in the blood when cardiac ischemia was induced during the stress test, as well as the concentrations of EV-hERG1 and EV-Hsp47. Additionally, the plasma levels of EV-hERG1 and EV-Hsp47 decreased in patients with decompensated heart failure (DHF). Our data provide the first evidence that hERG1 and Hsp47 are present in the membranes of sEVs derived from the human cardiomyocyte cell line, and also in those isolated from human peripheral blood. Total sEVs, EV-hERG1, and EV-Hsp47 may be explored as biomarkers for heart diseases such as heart failure and cardiac ischemia.

Metabolic syndrome and arthritis among Mexican American older adults: findings from a 23-year follow-up.

PURPOSE: To examine the sex differences in the relationship of metabolic syndrome (MetS) criteria with arthritis and symptomatic arthritis among Mexican American older adults aged ≥ 65 without self-reported arthritis at baseline over 23-years of follow-up. METHODS: Participants (N = 1447) were from the Hispanic Established Population for the Epidemiologic Study of the Elderly (1993/94-2016). Measures included MetS criteria, arthritis defined as self-reported physician-diagnosed arthritis, socio-demographics, morbidities, depressive symptoms, pain on weight-bearing, cognitive and physical function, handgrip strength, mobility, and activities of daily living (ADLs) limitations. Symptomatic arthritis was defined as self-reported arthritis and having ≥ 1 of the following: pain, mobility limitation, or limited ADLs. RESULTS: At baseline, the mean age was 72.6 years and 730 (50.5%) of our participants were females. Female participants with 2 and 3 MetS criteria had greater odds of arthritis [odds ratio (OR) = 1.77, 95% Confidence Interval (Cl) = 1.28-2.45 and OR = 2.68, 95% CI = 1.69-4.27, respectively) and symptomatic arthritis (OR = 1.74, 95% Cl = 1.24-2.44 and OR = 3.27, 95% CI = 2.04-5.26, respectively) after controlling for covariates. Male participants with 2 and 3 MetS criteria had greater odds of arthritis (OR = 1.65, 95% Cl = 1.14-2.39 and OR = 2.52, 95% CI = 1.51-4.19, respectively) and symptomatic arthritis (OR = 1.93, 95% Cl = 1.30-2.86 and OR = 2.98, 95% CI = 1.62-5.47, respectively) after controlling for covariates. Both females and males with pain on weight-bearing had greater odds of arthritis than those without pain. CONCLUSIONS: At 23-years of follow-up, Mexican American older adults with MetS have an increased risk of arthritis and symptomatic arthritis. Early MetS screening and management may reduce arthritis in this population at high risk of disability.

Improved detection and phylogenetic analysis of plant proteins containing LysM domains.

Plants perceive N-acetyl-d-glucosamine-containing oligosaccharides that play a role in the interaction with bacteria and fungi, through cell-surface receptors containing a tight bundle of three LysM domains in their extracellular region. However, the identification of LysM domains of receptor-like kinases (RLK)/receptor-like proteins (RLP) using sequence based methods has led to some ambiguity, as some proteins have been annotated with only one or two LysM domains. This missing annotation was likely produced by the failure of the LysM hidden Markov model (HMM) from the Pfam database to correctly identify some LysM domains in proteins of plant origin. In this work, we provide improved HMMs for LysM domain detection in plants, that were built from the structural alignment of manually curated LysM domain structures from the Protein Data Bank and AlphaFold Protein Structure Database. Furthermore, we evaluated different sets of ligand-specific HMMs that were able to correctly classify a limited set of fully characterised RLK/Ps by their ligand specificity. In contrast, the phylogenetic analysis of the extracellular region of RLK/Ps, or of their individual LysM domains, was unable to discriminate these proteins by their ligand specificity. The HMMs reported here will allow a more sensitive detection of plant proteins containing LysM domains and help improve their characterisation.

Diversity, Genomics and Symbiotic Characteristics of Sinorhizobia That Nodulate Desmanthus spp. in Northwest Argentina.

Desmanthus spp. are legumes with the ability to associate with diverse α-proteobacteria-a microsymbiont-in order to establish nitrogen-fixing root nodules. A previous investigation from our laboratory revealed that the main bacteria associated with Desmanthus paspalaceus in symbiosis in central Argentina (Province of Santa Fe) were quite diverse and belonged to the genera Rhizobium and Mesorhizobium. To achieve a more extensive view of the local microsymbionts associated with Desmanthus spp., we sampled three different sites in Jujuy and Salta, in northwest Argentina. Matrix-assisted Laser-Desorption-Ionization Time-of-Flight mass spectrometry (MALDI-TOF) typing, 16S-rDNA analysis, and genome sequencing demonstrated that the dominant root-nodule microsymbionts belonged to the genus Sinorhizobium, with some sequenced genomes related to Sinorhizobium mexicanum, Sinorhizobium chiapanecum, and Sinorhizobium psoraleae. An analysis of nodA and nodC markers indicated that, in some of the isolates, horizontal gene transfer appeared to be responsible for the lack of congruence between the phylogenies of the chromosome and of the symbiotic region. These results revealed diverse evolutionary strategies for reaching the current Desmanthus-microsymbiont diversity. What is remarkable beside their observed genetic diversity is that the tolerance profiles of these isolates to abiotic stresses (temperature, salt concentration, pH) were quite coincident with the separation of the sinorhizobia according to place of origin, suggesting possible ecoedaphic adaptations. This observation, together with the higher aerial dry-weight matter that some isolates generated in Desmanthus virgatus cv. Marc when compared to the biomass generated by the commercial strain Sinorhizobium terangae CB3126, distinguish the collected sinorhizobia as constituting valuable germplasm for evaluation in local fields to select for more efficient symbiotic pairs.

Identification of Ensifer meliloti genes required for survival during peat-based bioinoculant maturation by STM-seq.

Rhizobial inoculants are sold either as rhizobia within a liquid matrix; or as rhizobia adhered to granules composed of peat prill or finely ground peat moss. During the production of peat-based inoculants, a series of physiological changes occur that result in an increased capability of the rhizobia to survive on the seeds. The number of viable rhizobia on preinoculated seeds at the point of sale, however, is often a limiting factor, as is the inefficiency of the inoculant bacteria to compete with the local rhizobia for the host colonization. In the present work, we used STM-seq for the genome-wide screening of Ensifer meliloti mutants affected in the survival during the maturation of peat-based inoculant formulations. Through this approach, we were able to identify a set of mutants whose behavior suggests that persistence in peat inoculants involves a complex phenotype that is connected to diverse cellular activities, mainly related to satisfying the requirements of bacterial nutrition (e.g., carbon sources, ions) and to coping with specific stresses (e.g., oxidative, mutational). These results also provide a base knowledge that could be used to more completely understand the survival mechanisms used by rhizobia during the maturation of peat-based inoculants, as well as for the design and implementation of practical strategies to improve inoculant formulations.

The Ischemia and Reperfusion Injury Involves the Toll-Like Receptor-4 Participation Mainly in the Kidney Cortex.

BACKGROUND/AIMS: The renal inflammatory response and kidney regeneration in ischemia-reperfusion injury (IRI) are associated with Toll-like receptor 4 (TLR4). Here we study the role of TLR4 during IRI in the renal cortex and medulla separately, using wild-type (TLR4-WT) and Knockout (TLR4-KO) TLR4 mice. METHODS: We used 30 minutes of bilateral renal ischemia, followed by 48 hours of reperfusion in C57BL/6 mice. We measured the expression of elements associated with kidney injury, inflammation, macrophage polarization, mesenchymal transition, and proteostasis in the renal cortex and medulla by qRT-PCR and Western blot. In addition, we studied kidney morphology by H/E and PAS. RESULTS: Renal ischemia (30min) and reperfusion (48hrs) induced the mRNA and protein of TLR4 in the renal cortex. In addition, Serum Creatinine (SCr), blood urea nitrogen (BUN), Neutrophil gelatinase-associated lipocalin (NGAL), and acute tubular necrosis (ATN) were increased in TLR4-WT by IRI. Interestingly, the SCr and BUN had normal levels in TLR-KO during IRI. However, ATN and high levels of NGAL were present in the kidneys of TLR4-KO mice. The pro-inflammatory (IL-6 and TNF-α) and anti-inflammatory (Foxp3 and IL-10) markers increased by IRI only in the cortex of TLR4-WT but not in TLR4-KO mice. Furthermore, the M1 (CD38 and Frp2) and M2 (Arg-I, Erg-2, and c-Myc) macrophage markers increased by IRI only in the cortex of TLR4-WT. The TLR4-KO blunted the IRI-upregulation of M1 but not the M2 macrophage polarization. Vimentin increased in the renal cortex and medulla of TLR4-WT animals but not in the cortex of TLR4-KO mice. In addition, iNOS and clusterin were increased by IRI only in the cortex of TLR4-WT, and the absence of TLR4 inhibited only clusterin upregulation. Finally, Hsp27 and Hsp70 protein levels increased by IRI in the cortex and medulla of TLR4-WT and TRL4-KO lost the IRI-upregulation of Hsp70. In summary, TLR4 participates in renal ischemia and reperfusion through pro-inflammatory and anti-inflammatory responses inducing impaired kidney function (SCr and BUN). However, the IRI-upregulation of M2 macrophage markers (cortex), iNOS (cortex), IL-6 (medulla), vimentin (medulla), and Hsp27 (cortex and medulla) were independent of TLR4. CONCLUSION: The TLR4 inactivation during IRI prevented the loss of renal function due to the inactivation of inflammation response, avoiding M1 and preserving the M2 macrophage polarization in the renal cortex.

1400W Prevents Renal Injury in the Renal Cortex But Not in the Medulla in a Murine Model of Ischemia and Reperfusion Injury.

BACKGROUND/AIMS: Acute kidney injury (AKI) carries high morbidity and mortality, and the inducible nitric oxide synthase (iNOS) is a potential molecular target to prevent kidney dysfunction. In previous work, we reported that the pharmacological inhibitions of iNOS before ischemia/reperfusion (I/R) attenuate the I/R-induced AKI in mice. Here, we study the iNOS inhibitor 1400W [N-(3-(Aminomethyl)benzyl] acetamide, which has been described to be much more specific to iNOS inhibition than other compounds. METHODS: We used 30 minutes of bilateral renal ischemia, followed by 24 hours of reperfusion in Balb/c mice. 1400w (10 mg/kg i.p) was applied before I/R injury. We measured the expression of elements associated with kidney injury, inflammation, macrophage polarization, mesenchymal transition, and nephrogenic genes by qRT-PCR in the renal cortex and medulla. The Periodic Acid-Schiff (PAS) was used to study the kidney morphology. RESULTS: Remarkably, we found that 1400W affects the renal cortex and medulla in different ways. Thus, in the renal cortex, 1400W prevented the I/R-upregulation of 1. NGAL, Clusterin, and signs of morphological damage; 2. IL-6 and TNF-α; 3. TGF-ß; 4. M2(Arg1, Erg2, cMyc) and M1(CD38, Fpr2) macrophage polarization makers; and 5. Vimentin and FGF2 levels but not in the renal medulla. CONCLUSION: 1400W conferred protection in the kidney cortex compared to the kidney medulla. The present investigation provides relevant information to understand the opportunity to use 1400W as a therapeutic approach in AKI treatment.

Exploring the Expression of Pro-Inflammatory and Hypoxia-Related MicroRNA-20a, MicroRNA-30e, and MicroRNA-93 in Periodontitis and Gingival Mesenchymal Stem Cells under Hypoxia.

Hypoxia associated with inflammation are common hallmarks observed in several diseases, and it plays a major role in the expression of non-coding RNAs, including microRNAs (miRNAs). In addition, the miRNA target genes for hypoxia-inducible factor-1α (HIF-1α) and nuclear factor of activated T cells-5 (NFAT5) modulate the adaptation to hypoxia. The objective of the present study was to explore hypoxia-related miRNA target genes for HIF-1α and NFAT5, as well as miRNA-20a, miRNA-30e, and miRNA-93 expression in periodontitis versus healthy gingival tissues and gingival mesenchymal stem cells (GMSCs) cultured under hypoxic conditions. Thus, a case-control study was conducted, including healthy and periodontitis subjects. Clinical data and gingival tissue biopsies were collected to analyze the expression of miRNA-20a, miRNA-30e, miRNA-93, HIF-1α, and NFAT5 by qRT-PCR. Subsequently, GMSCs were isolated and cultured under hypoxic conditions (1% O2) to explore the expression of the HIF-1α, NFAT5, and miRNAs. The results showed a significant upregulation of miRNA-20a (p = 0.028), miRNA-30e (p = 0.035), and miRNA-93 (p = 0.026) in periodontitis tissues compared to healthy gingival biopsies. NFAT5 mRNA was downregulated in periodontitis tissues (p = 0.037), but HIF-1α was not affected (p = 0.60). Interestingly, hypoxic GMSCs upregulated the expression of miRNA-20a and HIF-1α, but they downregulated miRNA-93e. In addition, NFAT5 mRNA expression was not affected in hypoxic GMSCs. In conclusion, in periodontitis patients, the expression of miRNA-20a, miRNA-30e, and miRNA-93 increased, but a decreased expression of NFAT5 mRNA was detected. In addition, GMSCs under hypoxic conditions upregulate the HIF-1α and increase miRNA-20a (p = 0.049) expression. This study explores the role of inflammatory and hypoxia-related miRNAs and their target genes in periodontitis and GMSCs. It is crucial to determine the potential therapeutic target of these miRNAs and hypoxia during the periodontal immune-inflammatory response, which should be analyzed in greater depth in future studies.

ubiF is involved in acid stress tolerance and symbiotic competitiveness in Rhizobium favelukesii LPU83.

The acidity of soils significantly reduces the productivity of legumes mainly because of the detrimental effects of hydrogen ions on the legume plants, leading to the establishment of an inefficient symbiosis and poor biological nitrogen fixation. We recently reported the analysis of the fully sequenced genome of Rhizobium favelukesii LPU83, an alfalfa-nodulating rhizobium with a remarkable ability to grow, nodulate and compete in acidic conditions. To gain more insight into the genetic mechanisms leading to acid tolerance in R. favelukesii LPU83, we constructed a transposon mutant library and screened for mutants displaying a more acid-sensitive phenotype than the parental strain. We identified mutant Tn833 carrying a single-transposon insertion within LPU83_2531, an uncharacterized short ORF located immediately upstream from ubiF homolog. This gene encodes a protein with an enzymatic activity involved in the biosynthesis of ubiquinone. As the transposon was inserted near the 3' end of LPU83_2531 and these genes are cotranscribed as a part of the same operon, we hypothesized that the phenotype in Tn833 is most likely due to a polar effect on ubiF transcription.We found that a mutant in ubiF was impaired to grow at low pH and other abiotic stresses including 5 mM ascorbate and 0.500 mM Zn2+. Although the ubiF mutant retained the ability to nodulate alfalfa and Phaseolus vulgaris, it was unable to compete with the R. favelukesii LPU83 wild-type strain for nodulation in Medicago sativa and P. vulgaris, suggesting that ubiF is important for competitiveness. Here, we report for the first time an ubiF homolog being essential for nodulation competitiveness and tolerance to specific stresses in rhizobia.

Analysis of SARS-CoV-2 synonymous codon usage evolution throughout the COVID-19 pandemic.

SARS-CoV-2, the seventh coronavirus known to infect humans, can cause severe life-threatening respiratory pathologies. To better understand SARS-CoV-2 evolution, genome-wide analyses have been made, including the general characterization of its codons usage profile. Here we present a bioinformatic analysis of the evolution of SARS-CoV-2 codon usage over time using complete genomes collected since December 2019. Our results show that SARS-CoV-2 codon usage pattern is antagonistic to, and it is getting farther away from that of the human host. Further, a selection of deoptimized codons over time, which was accompanied by a decrease in both the codon adaptation index and the effective number of codons, was observed. All together, these findings suggest that SARS-CoV-2 could be evolving, at least from the perspective of the synonymous codon usage, to become less pathogenic.

The phylogeny of the genus Azohydromonas supports its transfer to the family Comamonadaceae.

The genus Azohydromonas encompasses five validly described species belonging to the betaproteobacterial class. Recognized for their potential biotechnological uses, they were first described as belonging to the genus Alcaligenes. The phylogeny of the 16S rRNA gene of the original strains as well as newly described species led to a description of these strains within a new bacterial genus, Azohydromonas. However, the phylogenetic position of this genus remains described as part of the family Alcaligenaceae, even those some authors have placed it within the order Burkholderiales. To unravel the precise position of the genus Azohydromonas, a wide phylogenomic analysis was performed. The results of 16S rRNA gene phylogeny, as well as those obtained by the multilocus analysis of homologous proteins and overall genome relatedness indices, support the reclassification of Azohydromonas in the Rubrivivax-Ideonella lineage of the family Comamonadaceae, so the transfer of this genus is proposed.

BioF is a novel B2 metallo-ß-lactamase from Pseudomonas sp. isolated from an on-farm biopurification system.

Antimicrobial resistance represents a major global health concern and environmental bacteria are considered a source of resistance genes. Carbapenems are often used as the last antibiotic option to treat multidrug-resistant bacteria. Metallo-ß-lactamases (MBLs) are able to render resistance to almost all ß-lactam antibiotics, including carbapenems. Unfortunately, there are no inhibitors against MBLs for clinical use. Subclass B2 MBLs are the only enzymes working as strict carbapenemases, under-represented, encoded in chromosome genes and only functional as mono-zinc enzymes. Despite current efforts in MBLs inhibitor development, B2 carbapenemase activity is especially difficult to suppress, even in vitro. In this study we characterized BioF, a novel subclass B2 MBL identified in a new environmental Pseudomonas sp. strain isolated from an on-farm biopurification system (BPS). Although blaBioF is most likely a chromosomal gene, it is found in a genomic island and may represent a step previous to the horizontal transmission of B2 genes. The new B2 MBL is active as a mono-zinc enzyme and is a potent carbapenemase with incipient activity against some cephalosporins. BioF activity is not affected by excess zinc and is only inhibited at high metal chelator concentrations. The discovery and characterization of B2 MBL BioF as a potent carbapenemase in a BPS bacterial isolate emphasizes the importance of exploring antibiotic resistances existing in the environmental microbiota under the influence of human activities before they could emerge clinically.

Easy identification of insertion sequence mobilization events in related bacterial strains with ISCompare.

Bacterial genomes are composed of core and accessory genomes. The first is composed of housekeeping and essential genes, while the second is highly enriched in mobile genetic elements, including transposable elements (TEs). Insertion sequences (ISs), the smallest TEs, have an important role in genome evolution, and contribute to bacterial genome plasticity and adaptability. ISs can spread in a genome, presenting different locations in nearly related strains, and producing phenotypic variations. Few tools are available which can identify differentially located ISs (DLISs) on assembled genomes. Here, we introduce ISCompare, a new program to profile IS mobilization events in related bacterial strains using complete or draft genome assemblies. ISCompare was validated using artificial genomes with simulated random IS insertions and real sequences, achieving the same or better results than other available tools, with the advantage that ISCompare can analyze multiple ISs at the same time and outputs a list of candidate DLISs. ISCompare provides an easy and straightforward approach to look for differentially located ISs on bacterial genomes.

Comparative Analysis of Three Bradyrhizobium diazoefficiens Genomes Show Specific Mutations Acquired during Selection for a Higher Motility Phenotype and Adaption to Laboratory Conditions.

Microbial genomes are being extensively studied using next-generation sequencing technologies in order to understand the changes that occur under different selection regimes. In this work, the number and type of mutations that have occurred in three Bradyrhizobium diazoefficiens USDA 110T strains under laboratory conditions and during selection for a more motile phenotypic variant were analyzed. Most of the mutations found in both processes consisted of single nucleotide polymorphisms, single nucleotide deletions or insertions. In the case of adaptation to laboratory conditions, half of the changes occurred within intergenic regions, and around 80% were insertions. When the more motile phenotypic variant was evaluated, eight single nucleotide polymorphisms and an 11-bp deletion were found, although none of them was directly related to known motility or chemotaxis genes. Two mutants were constructed to evaluate the 11-bp deletion affecting the alpha subunit of 2-oxoacid:acceptor oxidoreductase (AAV28_RS30705-blr6743). The results showed that this single deletion was not responsible for the enhanced motility phenotype. IMPORTANCE The genetic and genomic changes that occur under laboratory conditions in Bradyrhizobium diazoefficiens genomes remain poorly studied. Only a few genome sequences of this important nitrogen-fixing species are available, and there are no genome-wide comparative analyses of related strains. In the present work, we sequenced and compared the genomes of strains derived from a parent strain, B. diazoefficiens USDA 110, that has undergone processes of repeated culture in the laboratory environment, or phenotypic selection toward antibiotic resistance and enhanced motility. Our results represent the first analysis in B. diazoefficiens that provides insights into the specific mutations that are acquired during these processes.

Aminoguanidine Prevents the Oxidative Stress, Inhibiting Elements of Inflammation, Endothelial Activation, Mesenchymal Markers, and Confers a Renoprotective Effect in Renal Ischemia and Reperfusion Injury.

Oxidative stress produces macromolecules dysfunction and cellular damage. Renal ischemia-reperfusion injury (IRI) induces oxidative stress, inflammation, epithelium and endothelium damage, and cessation of renal function. The IRI is an inevitable process during kidney transplantation. Preliminary studies suggest that aminoguanidine (AG) is an antioxidant compound. In this study, we investigated the antioxidant effects of AG (50 mg/kg, intraperitoneal) and its association with molecular pathways activated by IRI (30 min/48 h) in the kidney. The antioxidant effect of AG was studied measuring GSSH/GSSG ratio, GST activity, lipoperoxidation, iNOS, and Hsp27 levels. In addition, we examined the effect of AG on elements associated with cell survival, inflammation, endothelium, and mesenchymal transition during IRI. AG prevented lipid peroxidation, increased GSH levels, and recovered the GST activity impaired by IRI. AG was associated with inhibition of iNOS, Hsp27, endothelial activation (VE-cadherin, PECAM), mesenchymal markers (vimentin, fascin, and HSP47), and inflammation (IL-1ß, IL-6, Foxp3, and IL-10) upregulation. In addition, AG reduced kidney injury (NGAL, clusterin, Arg-2, and TFG-ß1) and improved kidney function (glomerular filtration rate) during IRI. In conclusion, we found new evidence of the antioxidant properties of AG as a renoprotective compound during IRI. Therefore, AG is a promising compound to treat the deleterious effect of renal IRI.

Glutathione S-Transferase and Clusterin, New Players in the Ischemic Preconditioning Renal Protection in a Murine Model of Ischemia and Reperfusion.

BACKGROUND/AIMS: Renal ischemia and reperfusion injury (IRI) involves oxidative stress, disruption of microvasculature due to endothelial cell damage, loss of epithelial cell polarity secondary to cytoskeletal alterations, inflammation, and the subsequent transition into a mesenchymal phenotype. Ischemic preconditioning (IPC) has been proposed as a therapeutic strategy to avoid/ameliorate the IRI. Since previous results showed that IPC could have differential effects in kidney cortex vs. kidney medulla, in the present study we analyzed the effectiveness and molecular mechanisms implicated in IPC in both kidney regions. METHODS: We evaluated 3 experimental groups of BALB/c male mice: control (sham surgery); renal ischemia (30 min) by bilateral occlusion of the renal pedicle and reperfusion (48 hours) (I/R); and renal IPC (two cycles of 5 min of ischemia and 5 min of reperfusion) applied just before I/R. Acute kidney injury was evaluated by glomerular filtration rate (GFR), Neutrophil Gelatinase-Associated Lipocalin (NGAL) blood level, and histologic analysis. Oxidative stress was studied measurement the Glutathione S-Transferase (GST) activity, GSH/GSSG ratio, and lipoperoxidation levels. Inflammatory mediators (IL-1ß, IL-6, Foxp3, and IL-10) were quantified by qRT-PCR. The endothelial (PECAM-1), epithelial (AQP-1), mesenchymal (Vimentin, Fascin, and Hsp47), iNOS, clusterin, and Hsp27 expression were evaluated (qRT-PCR and/or Western blot). RESULTS: The IPC protocol prevented the decrease of GFR, reduced the plasma NGAL, and ameliorated morphological damage in the kidney cortex after I/R. The IPC also prevented the downregulation of GST activity, lipoperoxidation and ameliorated the oxidized glutathione. In addition, IPC prevented the upregulation of vimentin, fascin, and Hsp47, which was associated with the prevention of the downregulation of AQP1 after I/R. The protective effect of IPC was associated with the upregulation of Hsp27, Foxp3, and IL-10 expression in the renal cortex. However, the upregulation of iNOS, IL-1ß, IL-6, and clusterin by I/R were not modified by IPC. CONCLUSION: IPC conferred better protection in the kidney cortex as compared to the kidney medulla. The protective effect of IPC was associated with amelioration of oxidative stress, tubular damage, and the induction of markers of Treg lymphocytes activity in the cortical region. Further studies are needed to evaluate if lower tubular cell stress/damage after I/R may explain the preferential induction of Treg response in the kidney cortex induced by IPC.

Phylogenomic analysis supports the reclassification of Burkholderia novacaledonica as Caballeronia novacaledonica comb. nov.

Burkholderia novacaledonica is a Betaproteobacterial species isolated from ultramafic soils in New Caledonia. The characterization and classification of this species into the Burkholderia genus was done simultaneously with the proposal of the new genus Caballeronia, initially composed of closely related Burkholderia glathei-like species. Thereafter, some reports based on the use of phylogenetic marker genes suggested that B. novacaledonica forms part of Caballeronia genus. Lacking a formal validation, and with the availability of its genome sequence, a genome-based phylogeny of B. novacaledonica was obtained to unravel its taxonomic position in Burkholderia sensu lato. A partial gyrB gene phylogeny, extended multilocus sequence typing on homologous protein sequences, and genomic distance-based phylogeny, all support the placement of this species in the Caballeronia genus. Therefore, the reclassification of B. novacaledonica to Caballeronia novacaledonica comb. nov. is proposed.

The two-component system ActJK is involved in acid stress tolerance and symbiosis in Sinorhizobium meliloti.

The nitrogen-fixing α-proteobacterium Sinorhizobium meliloti genome codifies at least 50 response regulator (RR) proteins mediating different and, in many cases, unknown processes. RR-mutant library screening allowed us to identify genes potentially implicated in survival to acid conditions. actJ mutation resulted in a strain with reduced growth rate under mildly acidic conditions as well as a lower capacity to tolerate a sudden shift to lethal acidic conditions compared with the parental strain. Mutation of the downstream gene actK, which encodes for a histidine kinase, showed a similar phenotype in acidic environments suggesting a functional two-component system. Interestingly, even though nodulation kinetics, quantity, and macroscopic morphology of Medicago sativa nodules were not affected in actJ and actK mutants, ActK was required to express the wild-type nitrogen fixation phenotype and ActJK was necessary for full bacteroid development and nodule occupancy. The actJK regulatory system presented here provides insights into an evolutionary process in rhizobium adaptation to acidic environments and suggests that actJK-controlled functions are crucial for optimal symbiosis development.

Codon Usage Optimization in the Prokaryotic Tree of Life: How Synonymous Codons Are Differentially Selected in Sequence Domains with Different Expression Levels and Degrees of Conservation.

Prokaryote genomes exhibit a wide range of GC contents and codon usages, both resulting from an interaction between mutational bias and natural selection. In order to investigate the basis underlying specific codon changes, we performed a comprehensive analysis of 29 different prokaryote families. The analysis of core gene sets with increasing ancestries in each family lineage revealed that the codon usages became progressively more adapted to the tRNA pools. While, as previously reported, highly expressed genes presented the most optimized codon usage, the singletons contained the less selectively favored codons. The results showed that usually codons with the highest translational adaptation were preferentially enriched. In agreement with previous reports, a C bias in 2- to 3-fold pyrimidine-ending codons, and a U bias in 4-fold codons occurred in all families, irrespective of the global genomic GC content. Furthermore, the U biases suggested that U3-mRNA-U34-tRNA interactions were responsible for a prominent codon optimization in both the most ancestral core and the highly expressed genes. A comparative analysis of sequences that encode conserved (cr) or variable (vr) translated products, with each one being under high (HEP) and low (LEP) expression levels, demonstrated that the efficiency was more relevant (by a factor of 2) than accuracy to modeling codon usage. Finally, analysis of the third position of codons (GC3) revealed that in genomes with global GC contents higher than 35 to 40%, selection favored a GC3 increase, whereas in genomes with very low GC contents, a decrease in GC3 occurred. A comprehensive final model is presented in which all patterns of codon usage variations are condensed in four distinct behavioral groups.IMPORTANCE The prokaryotic genomes-the current heritage of the most ancient life forms on earth-are comprised of diverse gene sets, all characterized by varied origins, ancestries, and spatial-temporal expression patterns. Such genetic diversity has for a long time raised the question of how cells shape their coding strategies to optimize protein demands (i.e., product abundance) and accuracy (i.e., translation fidelity) through the use of the same genetic code in genomes with GC contents that range from less than 20 to more than 80%. Here, we present evidence on how codon usage is adjusted in the prokaryotic tree of life and on how specific biases have operated to improve translation. Through the use of proteome data, we characterized conserved and variable sequence domains in genes of either high or low expression level and quantitated the relative weight of efficiency and accuracy-as well as their interaction-in shaping codon usage in prokaryotes.