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Porphyromonas, Tannerella, and Prevotella species found in severe periodontitis use the Type IX Secretion System (T9SS) to load their outer membrane surface with an array of virulence factors. These virulence factors are then released on outer membrane vesicles (OMVs), which penetrate the host to dysregulate the immune response to establish a positive feedback loop of chronic, inflammatory destruction of the tooth's supporting tissues. In this review, we present the latest information on the molecular architecture of the T9SS and provide mechanistic insight into its role in secretion and attachment of cargo proteins to produce a virulence coat on cells and OMVs. The recent molecular structures of the T9SS motor comprising PorL and PorM as well as the secretion pore Sov, together with advances in the overall interactome, have provided insight into the possible mechanisms of secretion. We propose the presence of PorL/M motors arranged in a circle at the inner membrane with bent periplasmic rotors interacting with the PorN protein. At the outer membrane, we envisage a slide carousel model where the PorN protein is driven around a circular track composed of PorK. Cargo proteins are transported by PorN to PorW and the Sov translocon just as slides are rotated to the projection window. Secreted proteins are proposed to then be shuttled along highways consisting of the PorV shuttle protein to an array of attachment complexes distributed around the cell. The cell surface attachment of cargo is a hallmark of the T9SS, and in Porphyromonas gingivalis and Tannerella forsythia, this attachment is achieved via covalent bonding to a linking sugar synthesized by the Wbp/Vim pathway. The cell-surface attached cargo are enriched on OMVs, which are then released from the cell. [ABSTRACT FROM AUTHOR]

Many chemoreceptors contain a C-terminal pentapeptide at the end of a linker. In Escherichia coli, this pentapeptide forms a high-affinity binding site for CheR and phosphorylated CheB, and its removal interferes with chemoreceptor adaptation. Analysis of chemoreceptors revealed significant variation in their pentapeptide sequences, and bacteria often possess multiple chemoreceptors with differing pentapeptides. To assess whether this sequence variation alters CheR affinity and chemotaxis, we used Pectobacterium atrosepticum SCRI1043 as a model. SCRI1043 has 36 chemoreceptors, with 19 of them containing a C-terminal pentapeptide. We show that the affinity of CheR for the different pentapeptides varies up to 11-fold (K D 90 nM to 1 μM). Pentapeptides with the highest and lowest affinities differ only in a single amino acid. Deletion of the cheR gene abolishes chemotaxis. The replacement of the pentapeptide in the PacC chemoreceptor with those of the highest and lowest affinities significantly reduced chemotaxis to its cognate chemoeffector, L-Asp. Altering the PacC pentapeptide also reduced chemotaxis to L-Ser, but not to nitrate, which are responses mediated by the nontethered PacB and PacN chemoreceptors, respectively. Changes in the pentapeptide sequence thus modulate the response of the cognate receptor and that of another chemoreceptor., (© 2024 John Wiley & Sons Ltd.)

Purines and their derivatives control intracellular energy homeostasis and nucleotide synthesis, and act as signaling molecules. Here, we combine structural and sequence information to define a purine-binding motif that is present in sensor domains of thousands of bacterial receptors that modulate motility, gene expression, metabolism, and second-messenger turnover. Microcalorimetric titrations of selected sensor domains validate their ability to specifically bind purine derivatives, and evolutionary analyses indicate that purine sensors share a common ancestor with amino-acid receptors. Furthermore, we provide experimental evidence of physiological relevance of purine sensing in a second-messenger signaling system that modulates c-di-GMP levels., (© 2024. The Author(s).)

The type IX secretion system (T9SS) is a large multi-protein transenvelope complex distributed into the Bacteroidetes phylum and responsible for the secretion of proteins involved in pathogenesis, carbohydrate utilization or gliding motility. In Porphyromonas gingivalis, the two-component system PorY sensor and response regulator PorX participate to T9SS gene regulation. Here, we present the crystal structure of PorX Fj , the Flavobacterium johnsoniae PorX homolog. As for PorX, the PorX Fj structure is comprised of a CheY-like N-terminal domain and an alkaline phosphatase-like C-terminal domain separated by a three-helix bundle central domain. While not activated and monomeric in solution, PorX Fj crystallized as a dimer identical to active PorX. The CheY-like domain of PorX Fj is in an active-like conformation, and PorX Fj possesses phosphodiesterase activity, in agreement with the observation that the active site of its phosphatase-like domain is highly conserved with PorX., (© 2024. The Author(s).)

Purpose: Physical activity (PA) provides multiple health-related benefits in children and adolescents, however, at present, the majority of young people are insufficiently physically active. The aim of this study was to evaluate if neighborhood walkability and/or socio-economic status (SES) could affect the practice of walking, play outdoors and sports practice in a representative sample of Spanish children and adolescents., Methods: A sample of 4092 youth (aged 8-16 years old) from 245 primary and secondary schools in 121 localities from each of the 17 Spanish autonomous communities participated in the study. Walk Score was used to evaluate walkability of the neighborhood and household income was used as an indicator of SES. A 7-item self-reported validated questionnaire, was used to assess PA levels, and in a subsample of 10% of the participants, randomly selected from the entire sample, PA was objectively measured by accelerometers., Results: Youth from more walkable areas reported more minutes walking per day compared with those from less walkable neighborhoods (51.4 vs 48.8 minutes, respectively). The lowest average minutes spent in playing outdoors was found among participants from low-SES and low-walkable neighborhoods. Neighborhood SES influenced on the participation in team sports during the weekend, being this participation higher in high SES neighborhoods., Conclusion: Providing high walkable environments seems a good strategy to promote PA regardless SES levels. It seems that improving the walkability is a key component to partially overcome the SES inequalities, especially in urban areas with low SES. High-SES environments can offer better sports facilities and more organized physical activities than low-SES ones., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Aznar et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Introduction: The aims of this study were to compare several countermovement jump (CMJ) kinetic variables between professional (PRO) and semi-professional (SEMI-PRO) futsal players and examine the differences amongst playing positions., Methods: CMJ performance from 56 male futsal players (25.2 ± 4.8 years; weight: 74.4 ± 6.4 kg) was analysed. Players were separated into PRO ( n = 29; 27.0 ± 4.4 years; 75.4 ± 6.0 kg) and SEMI-PRO ( n = 27; 22.7 ± 4.3 years; 73.1 ± 6.8 kg), and according to playing position: defenders ( n = 16; 25.4 ± 3.7 years; 75.2 ± 6.0 kg), wingers ( n = 26; 23.5 ± 4.5 years; 72.0 ± 6.9 kg), and pivots ( n = 14; 28.0 ± 5.6 years; 77.8 ± 4.3 kg). Linear mixed models and effect sizes were used for the analyses based on the mean of two jumps for each variable., Results: PRO players presented a deeper center of mass (COM) displacement ( p = 0.002, ES = 0.83), greater eccentric (Ecc) absolute ( p = 0.019, ES = 0.61) and relative peak power ( p = 0.046, ES = 0.52), and achieved greater Ecc peak velocities ( p = 0.004, ES = 0.76) when compared to SEMI-PRO. Non-significant and trivial-to-small differences were observed in all the other CMJ variables according to the competitive level and playing position., Discussion: Ecc capabilities (i.e., deeper COM displacement, greater Ecc absolute and relative peak power, and peak velocity) during vertical jump seem to differentiate PRO and SEMI-PRO players. However, CMJ variables do not discriminate amongst playing positions in futsal players., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (© 2024 Spyrou, Alcaraz, Martínez-Serrano, Marín-Cascales, Ferioli, Contreras, Ribeiro, Travassos and Freitas.)

This study aimed to describe the current practices in futsal regarding a variety of topics related to performance and injury risk mitigation. Thirty-seven coaches from Spain and Portugal completed a questionnaire consisting of 28 closed questions organized in four categories: a) background information; b) training load (TL) monitoring and assessment of players' physical qualities; c) strength training (ST) practices; and d) recovery (REC) methods. The results showed that coaches varied in experience (1-8 years) and age (from 20 years to >50 years). Overall, 97.3% of the participants declared monitoring TL, with rating of perceived exertion, heart rate monitors, and wearable technology being used by 86.5%, 40.5%, and 37.8%, respectively. Neuromuscular and strength testing are the most common practices to evaluate performance and fatigue during the season. ST is a significant component of futsal, being performed 3 times/week during the pre- and in-season. ST is prescribed via %1RM - XRM (59.5%), velocity-based training (21.7%), repetitions in reserve (18.9%), until failure (10.8%), and circuit training (2.7%). 'Better Monitoring', 'More Individualized', 'Better Facilities', 'More Staff', and 'More Time' were the main aspects to improve ST. Multiple post-match REC strategies are used, with durations ranging from 0-15 to 16-30 min independently of game location.

In transenvelope multiprotein machines such as bacterial secretion systems, protein-protein interactions not only occur between soluble domains but might also be mediated by helix-helix contacts in the inner membrane. Several assays have been therefore developed to test homotypic and heterotypic interactions between transmembrane α-helices in their native membrane environment. Here, we provide detailed protocols for two genetic assays, TOXCAT and GALLEX, which are based on the reconstitution of dimeric regulators allowing the control of expression of reporter genes., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Proteases are enzymes that catalyze the hydrolytic degradation of other proteins into peptides or amino acids through the digestion of the peptide bond. Promiscuous proteases that target a wide range of proteins are distinguished from specific proteases that have a narrow range of substrates. In terms of activity, endoproteases cleave their substrates at specific residues within the target proteins, whereas exoproteases cleave from one extremity and may have processive activities. Proteases are therefore very useful tools to study proteins, notably their structure or conformation. In addition, proteases can be used to probe the topology of bacterial membrane proteins. Here, we describe limited protease accessibility assays to define inner membrane protein topology and conformational changes based on digestion profiles., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Most bacterial secretion systems are large machines that cross the cell envelope to deliver effectors outside the cell or directly into target cells. The peptidoglycan layer can therefore represent a physical barrier for the assembly of these large machines. Secretion systems and their counterparts such as type IV pili, flagella, and conjugation machines have therefore evolved or hijacked enzymes with peptidoglycan degradation activity. These enzymes are usually glycoside hydrolases that cleave the glycan chains of the peptidoglycan. Their activities are spatially controlled to avoid cell lysis and to create local rearrangement of the cell wall. In addition, peptidoglycan hydrolases may not be only required for the proper assembly of the secretion systems but may directly participate to the release of the effectors. Finally, several antibacterial effectors possess peptidoglycan degradation activity that damage the cell wall once delivered in the target cell. Here, we describe protocols to test the peptidoglycan degradation activity of these proteins in vitro and in solution., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Importance: Bacteria use weapons to deliver effectors into target cells. One of these weapons, the type VI secretion system (T6SS), assembles a contractile tail acting as a spring to propel a toxin-loaded needle. Due to its size and mechanism of action, the T6SS was intuitively thought to be energetically costly. Here, using a combination of mutants and growth measurements in liquid medium, on plates, and in competition experiments, we show that the T6SS does not entail a growth cost to enteroaggregative Escherichia coli ., Competing Interests: The authors declare no conflict of interest.

Background: Maintaining a healthy lifestyle is crucial for safeguarding the well-being and quality of life perception, appropriate growth, and development of children and adolescents, while also mitigating the risk of future adult-onset diseases., Objective: To assess associations between perceived quality of life and healthy lifestyle and related outcomes in Spanish children and adolescents., Methods: Cross-sectional analysis of 8-16-year-old children and adolescents (n = 3534) were included in the nationwide study of Physical Activity, Sedentarism, and Obesity in Spanish Youth (PASOS). Data were collected through (1) questionnaires on health-related quality of life (HRQoL), healthy lifestyle outcomes (dietary intake, physical fitness, sleep, and screen time), and (2) anthropometric measurements for weight status assessment. Data were analysed by logistic regression, using the health-related quality of life (HRQoL) as the grouping variable., Results: Participants with a lower HRQoL were those with a lower adherence to the MedDiet and lower achievement of the recommended daily intake of fruit and vegetables. They were also less likely to follow the recommendations for screen time and sleep (with the exception of the weekend) compared to participants with a higher HRQoL. Participants with a lower HRQoL showed a lower healthy weight status and poorer physical fitness than those with a higher HRQoL., Conclusions: Healthy eating habits, healthy weight status (normal weight), appropriate sleep time, physical fitness, and limited screen time play a crucial role in the perceived quality of life in children and adolescents.

Background: Youth is a vulnerable period. To classify lifestyle behaviors and its relationship with health-related outcomes of Spanish children and adolescents., Methods: Cross-sectional study including 3261 children aged 7.5-17.5 y (52.8% females). Physical activity (PA), screen-time, sleep time, adherence to Mediterranean diet (MD), weight status (WS) by validated methods. Cluster analysis was run considering chronological age., Results: Six clusters were identified: C1: high screen time, low adherence to MD and sleep time (n = 431,13.20%); C2: high WS, medium adherence to MD,high sleep time, and low screen time (n = 466,14.30%); C3: young group with low screen time and high PA, adherence to MD and sleep (n = 537,16.40%); C4: worst profile regarding adherence to MD, PA, WS and sleep time (n = 609,18.70%); C5: low screen time and PA, high sleep time (n = 804,24.70%); C6: high PA and screen time, low WS (n = 414,12.70%). Mean absolute values were statistically different among PA levels, screen and sleep time, adherence to MD, age, and WS (all p < 0.001)., Conclusions: The most prevalent pattern was low levels of PA, MD, and screen time, and high sleep time. The second most prevalent was characterized by very low levels of PA, sleep time, and adherence to MD, and high screen time, and WS in adolescents., Impact Statement: The main identified lifestyle behavior was poor physical activity, low adherence to Mediterranean Diet and high screen and sleep time. Children should increase physical activity levels, adherence to Mediterranean diet, decrease screen and sleep the appropriate hours per day. Families, schools, and medical communities must work together to gloss over present and future diseases. Sleep time had not been previously included in cluster analysis with physical activity, sedentary behaviors, obesity, and nutritional status, thus the present data open a new perspective in Spanish population. Health policies should focus on promoting physical activity, Mediterranean diet, adequate sleep and reducing screen time., (© 2023. The Author(s), under exclusive licence to the International Pediatric Research Foundation, Inc.)

The type VI secretion system (T6SS) is a contractile nanomachine widespread in Gram-negative bacteria. The T6SS injects effectors into target cells including eukaryotic hosts and competitor microbial cells and thus participates in pathogenesis and intermicrobial competition. Pseudomonas fluorescens MFE01 possesses a single T6SS gene cluster that confers biocontrol properties by protecting potato tubers against the phytopathogen Pectobacterium atrosepticum (Pca). Here, we demonstrate that a functional T6SS is essential to protect potato tuber by reducing the pectobacteria population. Fluorescence microscopy experiments showed that MFE01 displays an aggressive behaviour with an offensive T6SS characterized by continuous and intense T6SS firing activity. Interestingly, we observed that T6SS firing is correlated with rounding of Pectobacterium cells, suggesting delivery of a potent cell wall targeting effector. Mutagenesis coupled with functional assays then revealed that a putative T6SS secreted amidase, Tae3 Pf , is mainly responsible for MFE01 toxicity towards Pca. Further studies finally demonstrated that Tae3 Pf is toxic when produced in the periplasm, and that its toxicity is counteracted by the Tai3 Pf inner membrane immunity protein., (© 2023 The Authors. Environmental Microbiology published by Applied Microbiology International and John Wiley & Sons Ltd.)

Polymorphic toxins (PTs) are a broad family of toxins involved in interbacterial competition and pathogenesis. PTs are modular proteins that are comprised of a conserved N-terminal domain responsible for its transport, and a variable C-terminal domain bearing toxic activity. Although the mode of transport has yet to be elucidated, a new family of putative PTs containing an N-terminal MuF domain, resembling the Mu coliphage F protein, was identified in prophage genetic elements. The C-terminal toxin domains of these MuF PTs are predicted to bear nuclease, metallopeptidase, ADP-ribosyl transferase and RelA&#95;SpoT activities. In this study, we characterized the MuF-RelA&#95;SpoT toxin associated with the temperate phage of Streptococcus pneumoniae SPNA45. We show that the RelA&#95;SpoT domain has (p)ppApp synthetase activity, which is bactericidal under our experimental conditions. We further determine that the two genes located downstream encode two immunity proteins, one binding to and inactivating the toxin and the other detoxifying the cell via a pppApp hydrolase activity. Finally, based on protein sequence alignments, we propose a signature for (p)ppApp synthetases that distinguishes them from (p)ppGpp synthetases., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Importance: Chemotaxis of motile bacteria has multiple physiological functions. It enables bacteria to locate optimal ecological niches, mediates collective behaviors, and can play an important role in infection. These multiple functions largely depend on ligand specificities of chemoreceptors, and the number and identities of chemoreceptors show high diversity between organisms. Similar diversity is observed for the spectra of chemoeffectors, which include not only chemicals of high metabolic value but also bacterial, plant, and animal signaling molecules. However, the systematic identification of chemoeffectors and their mapping to specific chemoreceptors remains a challenge. Here, we combined several in vivo and in vitro approaches to establish a systematic screening strategy for the identification of receptor ligands and we applied it to identify a number of new physiologically relevant chemoeffectors for the important opportunistic human pathogen P. aeruginosa . This strategy can be equally applicable to map specificities of sensory domains from a wide variety of receptor types and bacteria., Competing Interests: The authors declare no conflict of interest.

Background and Aims: Hepatitis B virus (HBV) infection affects 300 million individuals worldwide, representing a major factor for the development of hepatic complications. Although existing antivirals are effective in suppressing replication, eradication of HBV is not achieved. Therefore, a multi-faceted approach involving antivirals and immunomodulatory agents is required. Non-human primates are widely used in pre-clinical studies due to their close evolutionary relationship to humans. Nonetheless, it is fundamental to identify the differences in immune response between humans and these models. Thus, we performed a transcriptomic characterization and interspecies comparison of the early immune responses to HBV in human and cynomolgus macaques., Methods: We characterized early transcriptomic changes in human and cynomolgus B cells, T cells, myeloid and plasmacytoid dendritic cells (pDC) exposed to HBV ex vivo for 2 hours. Differentially-expressed genes were further compared to the profiles of HBV-infected patients using publicly-available single-cell data., Results: HBV induced a wide variety of transcriptional changes in all cell types, with common genes between species representing only a small proportion. In particular, interferon gamma signaling was repressed in human pDCs. At the gene level, interferon gamma inducible protein 16 ( IFI16 ) was upregulated in macaque pDCs, while downregulated in humans. Moreover, IFI16 expression in pDCs from chronic HBV-infected patients anti-paralleled serum HBsAg levels., Conclusion: Our characterization of early transcriptomic changes induced by HBV in humans and cynomolgus macaques represents a useful resource for the identification of shared and divergent host responses, as well as potential immune targets against HBV., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Roca Suarez, Planel, Grand, Couturier, Tran, Porcheray, Becker, Reynier, Delgado, Cascales, Peyrot, Tamellini, Saliou, Elie, Baum, Vuong, Testoni, Roques, Zoulim, Hasan and Chemin.)

Background: Non-communicable chronic diseases are associated with a low-quality diet, low physical activity, and sedentary behavior., Objective: To assess how parents' diet and physical activity habits were associated with their offsprings' lifestyles., Study Design: A cross-sectional analysis of 8-16-year-old children and adolescents (n = 2539; 51.9% girls) was carried out within the frame of the first edition of the Physical Activity, Sedentarism, Lifestyles, and Obesity in Spanish Youth study (PASOS-2019). Data on adherence to the Mediterranean Diet (MedDiet), daily moderate-vigorous physical activity (MVPA), and screen time per day (television, computer, video games, and mobile phone) were collected from children and adolescents, and data on parents' diet quality and physical activity were compiled. Logistic regression models were used to assess the association between parents' lifestyles and those of children and adolescents., Results: High diet quality of parents was associated with higher adherence to the MedDiet of children and adolescents, as well as high consumption of fruit, vegetables, fish, nuts, and legumes. The high physical activity level of parents was associated with the low consumption of fast foods, sweets, and candies in children and adolescents. Children with high levels of physical activity were those whose parents showed better diet quality and physical activity levels., Conclusions: Parents' high diet quality and physical activity were associated with healthy lifestyles, higher adherence to the MedDiet, and physical activity of their offspring, mainly in adolescents.

In the environment, bacteria compete for niche occupancy and resources; they have, therefore, evolved a broad variety of antibacterial weapons to destroy competitors. Current laboratory techniques to evaluate antibacterial activity are usually labor intensive, low throughput, costly, and time consuming. Typical assays rely on the outgrowth of colonies of prey cells on selective solid media after competition. Here, we present fast, inexpensive, and complementary optimized protocols to qualitatively and quantitively measure antibacterial activity. The first method is based on the degradation of a cell-impermeable chromogenic substrate of the β-galactosidase, a cytoplasmic enzyme released during lysis of the attacked reporter strain. The second method relies on the lag time required for the attacked cells to reach a defined optical density after the competition, which is directly dependent on the initial number of surviving cells. Key features First method utilizes the release of β-galactosidase as a proxy for bacterial lysis. Second method is based on the growth timing of surviving cells. Combination of two methods discriminates between cell death and lysis, cell death without lysis, or survival to quasi-lysis. Methods optimized to various bacterial species such as Escherichia coli, Pseudomonas aeruginosa , and Myxococcus xanthus . Graphical overview., Competing Interests: Competing interestsThe authors declare that they have no competing interests. Ethics approval was not required for this study., (©Copyright : © 2023 The Authors; This is an open access article under the CC BY-NC license.)

Bacterial signal transduction systems are typically activated by the binding of signal molecules to receptor ligand binding domains (LBDs), such as the NIT LBD. We report here the identification of the NIT domain in more than 15,000 receptors that were present in 30 bacterial phyla, but also in 19 eukaryotic phyla, expanding its known phylogenetic distribution. The NIT domain formed part of seven receptor families that either control transcription, mediate chemotaxis or regulate second messenger levels. We have produced the NIT domains from chemoreceptors of the bacterial phytopathogens Pectobacterium atrosepticum (PacN) and Pseudomonas savastanoi (PscN) as individual purified proteins. High-throughput ligand screening using compound libraries revealed a specificity for nitrate and nitrite binding. Isothermal titration calorimetry experiments showed that PacN-LBD bound preferentially nitrate ( K D  = 1.9 μM), whereas the affinity of PscN-LBD for nitrite ( K D  = 2.1 μM) was 22 times higher than that for nitrate. Analytical ultracentrifugation experiments indicated that PscN-LBD is monomeric in the presence and absence of ligands. The R182A mutant of PscN did not bind nitrate or nitrite. This residue is not conserved in the NIT domain of the Pseudomonas aeruginosa chemoreceptor PA4520, which may be related to its failure to bind nitrate/nitrite. The magnitude of P. atrosepticum chemotaxis towards nitrate was significantly greater than that of nitrite and pacN deletion almost abolished responses to both compounds. This study highlights the important role of nitrate and nitrite as signal molecules in life and advances our knowledge on the NIT domain as universal nitrate/nitrite sensor module., (© 2023 The Authors. Molecular Microbiology published by John Wiley & Sons Ltd.)

Unhealthy dietary habits determined during childhood may represent a risk factor to many of the chronic non-communicable diseases (NCDs) in adulthood. Mediterranean Diet (MD) adherence in children and adolescents (8-16 years) living in Spain was investigated using the KIDMED questionnaire in a comparative analysis of two cross-sectional nationwide representative studies: enKid (1998-2000, n = 1001) and PASOS (2019-2020, n = 3540). Taking into account the educational level of pupils, as well as the characteristics of the place of living, a significant association was found between a KIDMED score ≥ 8 (optimal MD adherence) and primary education as well as residency in an area of <50,000 inhabitants, while living in the southern regions was associated with non-optimal MD adherence ( p < 0.001). Participants of the 2019-2020 study showed an increase in the consumption of dairy products (31.1% increase), pasta/rice (15.4% increase), olive oil (16.9% increase), and nuts (9.7% increase), as well as a decreased sweets and candies intake (12.6% reduction). In contrast, a significantly lower MD adherence was found when comparing the 2019-2020 (mean ± SE: 6.9 ± 0.04) and the 1998-2000 study (7.37 ± 0.08); p < 0.001), due to less consumption of fish (20.3% reduction), pulse (19.4% reduction), and fruits (14.9% reduction), and an increased intake of commercial goods/pastries or fast-food intake (both 19.4% increase). The lowest adherence was recorded for adolescents also in the most recent study, where 10.9% of them presented a KIDMED score ≤ 3. This study shows that eating habits are deteriorating among Spanish children and adolescents. Such findings point out the urgency of undertaking strong measures to promote the consumption of healthy, sustainable, and non-ultra-processed food, such as those available in an MD, not only at a scientific and academic level, but also at a governmental one.

Abstract: Spyrou, K, Alcaraz, PE, Marín-Cascales, E, Herrero-Carrasco, R, Cohen, DD, and Freitas, TT. Neuromuscular performance changes in elite futsal players over a competitive season. J Strength Cond Res 37(5): 1111-1116, 2023-A professional futsal season imposes a great amount of physiological and mechanical stress on players. The main aim of this study was to examine the changes in neuromuscular performance qualities across the season. Ten professional male players performed a 10-m sprint, standing long jumps (SLJs), and countermovement jumps (CMJs) during the competitive season (i.e., every ∼5 weeks from September to January). A one-way repeated measures ANOVA with post hoc pairwise comparisons and effect sizes (ESs) were used to analyze potential differences among these assessments. A significant and large decline was found in concentric peak power ( p = 0.040; ES = 1.24). A nonsignificant and moderate decrease was observed in sprint ability ( p = 0.155; ES = 1.03), CMJ height ( p = 0.175; ES = 1.00), and SLJ distance ( p = 0.164; ES = 1.03). Regarding other CMJ kinetic variables, nonsignificant and moderate changes were found. In summary, considering the neuromuscular performance tests and variables assessed, only concentric peak power in CMJ decreased significantly across the season; however, nonsignificant decrements were observed in sprinting time, SLJ, CMJ height, and other kinetic metrics. CMJ variables during the jump-land cycle should be incorporated alongside more traditional measures (e.g., jump height) to monitor performance during the season., (Copyright © 2023 National Strength and Conditioning Association.)

Childhood obesity is a public health problem worldwide. An important determinant of child and adolescent obesity is socioeconomic status (SES). However, the magnitude of the impact of different SES indicators on pediatric obesity on the Spanish population scale is unclear. The aim of this study was to assess the association between three SES indicators and obesity in a nationwide, representative sample of Spanish children and adolescents. A total of 2791 boys and girls aged 8 to 16 years old were included. Their weight, height, and waist circumference were measured. SES was assessed using two parent/legal guardian self-reported indicators (educational level -University/non-University- and labor market status -Employed/Unemployed-). As a third SES indicator, the annual mean income per person was obtained from the census section where the participating schools were located (≥12.731€/<12.731€). The prevalence of obesity, severe obesity, and abdominal obesity was 11.5%, 1.4%, and 22.3%, respectively. Logistic regression models showed an inverse association of both education and labor market status with obesity, severe obesity, and abdominal obesity (all p < 0.001). Income was also inversely associated with obesity ( p < 0.01) and abdominal obesity ( p < 0.001). Finally, the highest composite SES category (University/Employed/≥12.731€ n = 517) showed a robust and inverse association with obesity (OR = 0.28; 95% CI: 0.16-0.48), severe obesity (OR = 0.20; 95% CI: 0.05-0.81), and abdominal obesity (OR = 0.36; 95% CI: 0.23-0.54) in comparison with the lowest composite SES category (Less than University/Unemployed/<12.731€; n = 164). No significant interaction between composite SES categories and age and gender was found. SES is strongly associated with pediatric obesity in Spain.

The type VI secretion system (T6SS) is a multiprotein weapon evolved by Gram-negative bacteria to deliver effectors into eukaryotic cells or bacterial rivals. The T6SS uses a contractile mechanism to propel an effector-loaded needle into its target. The contractile tail is built on an assembly platform, the baseplate, which is anchored to a membrane complex. Baseplate-membrane complex interactions are mainly mediated by contacts between the C-terminal domain of the TssK baseplate component and the cytoplasmic domain of the TssL inner membrane protein. Currently, the structural details of this interaction are unknown due to the marginal stability of the TssK-TssL complex. Here we conducted a mutagenesis study based on putative TssK-TssL contact pairs identified by co-evolution analyses. We then evaluated the impact of these mutations on T6SS activity, TssK-TssL interaction and sheath assembly and dynamics in enteroaggregative Escherichia coli. Finally, we probed the TssK-TssL interface by disulfide cross-linking, allowing to propose a model for the baseplate-membrane complex interface., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The type VI secretion system (T6SS) delivers enzymatic effectors into target cells to destroy them. Cells of the same strain protect themselves against effectors with immunity proteins that specifically inhibit effectors. Here, we report the identification and characterization of a Tle3 phospholipase effector and its cognate immunity protein Tli3-an outer membrane lipoprotein from adherent-invasive Escherichia coli (AIEC). Enzymatic assays demonstrate that purified Tle3 AIEC has a phospholipase A1, and not A2, activity and that its toxicity is neutralized by the cognate immunity protein Tli3 AIEC . Tli3 AIEC binds Tle3 in a 1:1 stoichiometric ratio. Tle3 AIEC , Tli3 AIEC and the Tle3 AIEC -Tli3 AIEC complex were purified and subjected to crystallization. The Tle3 AIEC -Tli3 AIEC complex structure could not be solved by SeMet phasing, but only by molecular replacement when using an AlphaFold2 prediction model. Tle3 AIEC exhibits an α/β-hydrolase fold decorated by two protruding segments, including a N-terminus loop. Tli3 AIEC displays a new fold of three stacked β-sheets and a protruding loop that inserts in Tle3 AIEC catalytic crevice. We showed, experimentally, that Tle3 AIEC interacts with the VgrG AIEC cargo protein and AlphaFold2 prediction of the VgrG AIEC -Tle3 AIEC complex reveals a strong interaction between the VgrG AIEC C-terminus adaptor and Tle3 AIEC N-terminal loop.

Rearrangement hot spot (Rhs) proteins are members of the broad family of polymorphic toxins. Polymorphic toxins are modular proteins composed of an N-terminal region that specifies their mode of secretion into the medium or into the target cell, a central delivery module, and a C-terminal domain that has toxic activity. Here, we structurally and functionally characterize the C-terminal toxic domain of the antibacterial Rhsmain protein, TreTu, which is delivered by the type VI secretion system of Salmonella enterica Typhimurium. We show that this domain adopts an ADP-ribosyltransferase fold and inhibits protein synthesis by transferring an ADP-ribose group from NAD+ to the elongation factor Tu (EF-Tu). This modification is specifically placed on the side chain of the conserved D21 residue located on the P-loop of the EF-Tu G-domain. Finally, we demonstrate that the TriTu immunity protein neutralizes TreTu activity by acting like a lid that closes the catalytic site and traps the NAD+., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Type VIIb secretion systems (T7SSb) were recently proposed to mediate different aspects of Firmicutes physiology, including bacterial pathogenicity and competition. However, their architecture and mechanism of action remain largely obscure. Here, we present a detailed analysis of the T7SSb-mediated bacterial competition in Bacillus subtilis, using the effector YxiD as a model for the LXG secreted toxins. By systematically investigating protein-protein interactions, we reveal that the membrane subunit YukC contacts all T7SSb components, including the WXG100 substrate YukE and the LXG effector YxiD. YukC's crystal structure shows unique features, suggesting an intrinsic flexibility that is required for T7SSb antibacterial activity. Overall, our results shed light on the role and molecular organization of the T7SSb and demonstrate the potential of B. subtilis as a model system for extensive structure-function studies of these secretion machineries. IMPORTANCE Type VII secretion systems mediate protein extrusion from Gram-positive bacteria and are classified as T7SSa and T7SSb in Actinobacteria and in Firmicutes , respectively. Despite the genetic divergence of T7SSa and T7SSb, the high degree of structural similarity of their WXG100 substrates suggests similar secretion mechanisms. Recent advances revealed the structures of several T7SSa cytoplasmic membrane complexes, but the molecular mechanism of secretion and the T7SSb architecture remain obscure. Here, we provide hints on the organization of T7SSb in B. subtilis and a high-resolution structure of its central pseudokinase subunit, opening new perspectives for the understanding of the T7SSb secretion mechanism by using B. subtilis as an amenable bacterial model.

Bacteria have evolved many different signal transduction systems to sense and respond to changing environmental conditions. Signal integration is mainly achieved by signal recognition at extracytosolic ligand-binding domains (LBDs) of receptors. Hundreds of different LBDs have been reported, and our understanding of their sensing properties is growing. Receptors must function over a range of environmental pH values, but there is little information available on the robustness of sensing as a function of pH. Here, we have used isothermal titration calorimetry to determine the pH dependence of ligand recognition by nine LBDs that cover all major LBD superfamilies, of periplasmic solute-binding proteins, and cytosolic LBDs. We show that periplasmic LBDs recognize ligands over a very broad pH range, frequently stretching over eight pH units. This wide pH range contrasts with a much narrower pH response range of the cytosolic LBDs analyzed. Many LBDs must be dimeric to bind ligands, and analytical ultracentrifugation studies showed that the LBD of the Tar chemoreceptor forms dimers over the entire pH range tested. The pH dependences of Pseudomonas aeruginosa motility and chemotaxis were bell-shaped and centered at pH 7.0. Evidence for pH robustness of signaling in vivo was obtained by Förster Resonance Energy Transfer (FRET) measurements of the chemotaxis pathway responses in Escherichia coli. Bacteria have evolved several strategies to cope with extreme pH, such as periplasmic chaperones for protein refolding. The intrinsic pH resistance of periplasmic LBDs appears to be another strategy that permits bacteria to survive under adverse conditions. IMPORTANCE Demonstration of the pH robustness of extracytoplasmic sensing reveals a previously undescribed evolutionary mechanism that enables bacteria to monitor environmental changes under changing conditions. This mechanism includes the maintenance of the dimeric state of four-helixbundle ligand-binding domains (LBDs). The construction of biosensors is a rapidly growing field of research, and their use to monitor the progression of the COVID-19 pandemic has impressively demonstrated their usefulness. LBDs represent an enormous reservoir of binding modules that can be used to create novel biosensors. Among ligands recognized by LBDs are neurotransmitters, hormones, and quorum-sensing signals. The demonstration that extracytosolic LBDs bind their signals over a wide range of pH values will facilitate the design of biosensors that function under highly variable conditions of acidity and alkalinity.

Foliar bacterial pathogens have to penetrate the plant tissue and access the interior of the apoplast in order to initiate the pathogenic phase. The entry process is driven by chemotaxis towards plant-derived compounds in order to locate plant openings. However, information on plant signals recognized by bacterial chemoreceptors is scarce. Here, we show that the perception of GABA and l-Pro, two abundant components of the tomato apoplast, through the PsPto-PscC chemoreceptor drives the entry of Pseudomonas syringae pv. tomato into the tomato apoplast. The recognition of both compounds by PsPto-PscC caused chemoattraction to both amino acids and participated in the regulation of GABA catabolism. Mutation of the PsPto-PscC chemoreceptor caused a reduced chemotactic response towards these compounds which in turn impaired entry and reduced virulence in tomato plants. Interestingly, GABA and l-Pro levels significantly increase in tomato plants upon pathogen infection and are involved in the regulation of the plant defence response. This is an example illustrating how bacteria respond to plant signals produced during the interaction as cues to access the plant apoplast and to ensure efficient infection., (© 2022 The Authors. Molecular Plant Pathology published by British Society for Plant Pathology and John Wiley & Sons Ltd.)

The Porphyromonas gingivalis type IX secretion system (T9SS) promotes periodontal disease by secreting gingipains and other virulence factors. By in situ cryoelectron tomography, we report that the P. gingivalis T9SS consists of 18 PorM dimers arranged as a large, caged ring in the periplasm. Near the outer membrane, PorM dimers interact with a PorKN ring complex of ∼52 nm in diameter. PorMKN translocation complexes of a given T9SS adopt distinct conformations energized by the proton motive force, suggestive of different activation states. At the inner membrane, PorM associates with a cytoplasmic complex that exhibits 12-fold symmetry and requires both PorM and PorL for assembly. Activated motors deliver substrates across the outer membrane via one of eight Sov translocons arranged in a ring. The T9SSs are unique among known secretion systems in bacteria and eukaryotes in their assembly as supramolecular machines composed of apparently independently functioning translocation motors and export pores.

Motile bacteria usually rely on external apparatus like flagella for swimming or pili for twitching. By contrast, gliding bacteria do not rely on obvious surface appendages to move on solid surfaces. Flavobacterium johnsoniae and other bacteria in the Bacteroidetes phylum use adhesins whose movement on the cell surface supports motility. In F. johnsoniae, secretion and helicoidal motion of the main adhesin SprB are intimately linked and depend on the type IX secretion system (T9SS). Both processes necessitate the proton motive force (PMF), which is thought to fuel a molecular motor that comprises the GldL and GldM cytoplasmic membrane proteins. Here, we show that F. johnsoniae gliding motility is powered by the pH gradient component of the PMF. We further delineate the interaction network between the GldLM transmembrane helices (TMHs) and show that conserved glutamate residues in GldL TMH2 are essential for gliding motility, although having distinct roles in SprB secretion and motion. We then demonstrate that the PMF and GldL trigger conformational changes in the GldM periplasmic domain. We finally show that multiple GldLM complexes are distributed in the membrane, suggesting that a network of motors may be present to move SprB along a helical path on the cell surface. Altogether, our results provide evidence that GldL and GldM assemble dynamic membrane channels that use the proton gradient to power both T9SS-dependent secretion of SprB and its motion at the cell surface., Competing Interests: The authors have declared that no competing interests exist.