
外部圧力の印加により、細菌コロニーは密度が高くなり、細菌間の機械的相互作用が強化される。一方で、剛体の閉じ込めとは異なり、外部圧力は軟らかい制約として作用し、ほぼ線形の長期的な個体数増加とコロニーサイズの拡大を可能にする。


coremsg

3次元コロニー内の応力応答性細菌の成長と圧力抑制

title_rewrite


細菌集団内の一部の細菌は、薬剤排出ポンプの発現増加、外膜小胞の分泌、膜の変化などの応答を示すことで、抗菌ペプチドの蓄積を抑え、表現型耐性を示す。


Antimicrobial resistance threatens the viability of modern medical interventions. There is a dire need of developing novel approaches to counter resistance mechanisms employed by starved or slow-growing pathogens that are refractory to conventional antimicrobial therapies. Antimicrobial peptides have been advocated as potential therapeutic solutions due to low levels of genetic resistance observed in bacteria against these compounds. However, here we show that subpopulations of stationary phase Escherichia coli and Pseudomonas aeruginosa survive tachyplesin treatment without genetic mutations. These phenotypic variants induce efflux, outer membrane vesicles secretion and membrane modifications in response to tachyplesin exposure, sequestering the peptide in their membranes where it cannot exert its antimicrobial activity. We discovered that formation of these phenotypic variants could be prevented by administering tachyplesin in combination with sertraline, a clinically used antidepressant, suggesting a novel approach for combatting antimicrobial-refractory stationary phase bacteria.

### Competing Interest Statement

The authors have declared no competing interest.

Heterogeneous efflux pump expression underpins phenotypic resistance to antimicrobial peptides

細菌集団内の異質な薬剤排出ポンプ発現が抗菌ペプチドに対する表現型耐性の基盤となる


細菌の III型分泌装置トランスロコンタンパク質EseBが、NLRC4/NAIPインフラマソームと直接結合し、ホスト細胞のピロプトーシスを誘導する。この機能は、EseBホモログを持つ様々な病原細菌で保存されている。


Type III secretion system (T3SS) is a virulence apparatus existing in many bacterial pathogens. Structurally, T3SS consists of the base, needle, tip, and translocon. The NLRC4 inflammasome is the major receptor for T3SS needle and basal rod proteins. Whether other T3SS components are recognized by NLRC4 is unclear. In this study, using Edwardsiella tarda as a model intracellular pathogen, we examined T3SS−inflammasome interaction and its effect on cell death. E. tarda induced pyroptosis in a manner that required the bacterial translocon and the host inflammasome proteins of NLRC4, NLRP3, ASC, and caspase 1/4. The translocon protein EseB triggered NLRC4/NAIP-mediated pyroptosis by binding NAIP via its C-terminal region, particularly the terminal 6 residues (T6R). EseB homologs exist widely in T3SS-positive bacteria and share high identities in T6R. Like E. tarda EseB, all of the representatives of the EseB homologs exhibited T6R-dependent NLRC4 activation ability. Together these results revealed the function and molecular mechanism of EseB to induce host cell pyroptosis and suggested a highly conserved inflammasome-activation mechanism of T3SS translocon in bacterial pathogens.

### Competing Interest Statement

The authors have declared no competing interest.

T3SS translocon induces pyroptosis by direct interaction with NLRC4/NAIP inflammasome

細菌の-iii型分泌装置トランスロコンが-nlrc4-naip-インフラマソームと直接相互作用し-ピロプトーシスを誘導する


細菌の III型分泌装置トランスロコンが NLRC4/NAIP インフラマソームと直接相互作用し、ピロプトーシスを誘導する



細菌バイオフィルムにおいて、細胞接着の過程で上昇する環状ジグアノシン一リン酸(c-di-GMP)が、毒素タンパク質HipHの発現と活性を制御することで、遺伝体の安定性と抗生物質耐性を決定する。


Biofilms are complex bacterial communities characterized by a high persister prevalence, which contributes to chronic and relapsing infections. Historically, biofilm persister formation has been linked to constraints imposed by their dense structures. However, we observed an elevated persister frequency accompanying the stage of cell adhesion, marking the onset of biofilm development. Subsequent mechanistic studies uncovered a distinctive type of toxin-antitoxin (TA) module triggered by cell adhesion, which is responsible for this elevation. In this module, the toxin HipH acts as a genotoxic deoxyribonuclease, inducing DNA double strand breaks and genome instability. While the second messenger c-di-GMP functions as the antitoxin, exerting control over HipH expression and activity. The dynamic interplay between c-di-GMP and HipH levels emerges as a crucial determinant governing genome stability and persister generation within biofilms. These findings unveil a unique TA system, where small molecules act as the antitoxin, outlining a biofilm-specific molecular mechanism influencing genome stability and antibiotic persistence, with potential implications for treating biofilm infections.

### Competing Interest Statement

The authors have declared no competing interest.

Cyclic di-GMP as an Antitoxin Regulates Bacterial Genome Stability and Antibiotic Persistence in Biofilms

細菌バイオフィルムにおける抗毒素としての環状ジグアノシン一リン酸-c-di-gmp-が遺伝体の安定性と抗生物質耐性に与える影響


細菌バイオフィルムにおける抗毒素としての環状ジグアノシン一リン酸(c-di-GMP)が遺伝体の安定性と抗生物質耐性に与える影響



ブラウンリポタンパク質は、ペプチドグリカンへの固定化と遊離の間で動的平衡を保っている。


Peptidoglycan (PG) is a giant macromolecule that completely surrounds bacterial cells and prevents lysis in hypo-osmotic environments. This net-like macromolecule is made of glycan strands linked to each other by two types of transpeptidases that form either 4→3 (PBPs) or 3→3 (LDTs) cross-links. Previously, we devised a heavy isotope-based PG full labeling method coupled to mass spectrometry to determine the mode of insertion of new-subunits into the expanding PG network ([Atze et al., 2022][1]). We showed that PG polymerization operates according to different modes for the formation of the septum and of the lateral cell walls, as well as for bacterial growth in the presence or absence of β-lactams in engineered strains that can exclusively rely on LDTs for PG cross-linking when drugs are present. Here, we apply our method to the resolution of the kinetics of the reactions leading to the covalent tethering of the Braun lipoprotein (Lpp) to PG and the subsequent hydrolysis of that same covalent link. We find that Lpp and disaccharide-peptide subunits are independently incorporated into the expanding lateral cell walls. Newly synthesized septum PG appears to contain small amounts of tethered Lpp. LDTs did mediate intense shuffling of Lpp between PG stems leading to a dynamic equilibrium between the PG-tethered and free forms of Lpp.

### Competing Interest Statement

The authors have declared no competing interest.

 [1]: #ref-3

Peptidoglycan-tethered and free forms of the Braun lipoprotein are in dynamic equilibrium in Escherichia coli

大腸菌におけるブラウンリポタンパク質の固定化型と遊離型の動的平衡


偽膜性肺炎桿菌PAO1株のタイプIV線毛の高分解能クライオ電子顕微鏡構造を明らかにし、他の細菌のタイプIV線毛との比較を行った。


Type IV pili (T4Ps), which are abundant in many bacterial and archaeal species, have been shown to play important roles in both surface sensing and twitching motility, with implications for adhesion, biofilm formation and pathogenicity. While Type IV pilus (T4P) structures from other organisms have been previously solved, a high-resolution structure of the native, fully assembled T4P of Pseudomonas aeruginosa, one of the major human pathogens, is not available. Here, we report a 3.2 Å-resolution structure of the P. aeruginosa PAO1 T4P determined by electron cryomicroscopy (cryo-EM). PilA subunits constituting the T4P exhibit a classical pilin fold featuring an extended N-terminal α-helix linked to a C-terminal globular β-sheet-containing domain, which are packed tightly along the pilus. The N-terminal helices constitute the pilus core where they stabilise the tubular assembly via hydrophobic interactions. The α-helical core of the pilus is surrounded by the C-terminal globular domain of PilA that coats the outer surface of the pilus, mediating interactions with the surrounding environment. Comparison of the P. aeruginosa T4P with T4P structures from other organisms, both at the level of the pilin subunits and the fully assembled pili, allows us to enumerate key differences, and detect common architectural principles in this abundant class of prokaryotic filaments. This study provides a structural framework for understanding the molecular and cell biology of these important cellular appendages mediating interaction of prokaryotes to surfaces.

### Competing Interest Statement

The authors have declared no competing interest.

Structure of the Pseudomonas aeruginosa PAO1 Type IV pilus

偽膜性肺炎桿菌pao1株のタイプiv線毛の構造


偽膜性肺炎桿菌PAO1株のタイプIV線毛の構造



A. baumannii の細胞表層多糖は、自身の VI 型分泌システムの活性を抑制することで、細菌間競争において防御的な役割を果たす。


The type VI secretion system (T6SS) is a sophisticated, contact-dependent nanomachine involved in interbacterial competition. To function effectively, the T6SS must penetrate the membranes of both attacker and target bacteria. Structures associated with the cell envelope, like polysaccharides chains, can therefore introduce spatial separation and steric hindrance, potentially affecting the efficacy of the T6SS. In this study, we examined how the capsular polysaccharide (CPS) of Acinetobacter baumannii affects T6SS’s antibacterial function. Our findings show that the CPS confers resistance against T6SS-mediated assaults from rival bacteria. Notably, under typical growth conditions, the presence of the surface-bound capsule also reduces the efficacy of the bacterium’s own T6SS. This T6SS impairment is further enhanced when CPS is overproduced due to genetic modifications or antibiotic treatment. Furthermore, we demonstrate that the bacterium adjusts the level of the T6SS inner tube protein Hcp according to its secretion capacity, by initiating a degradation process involving the ClpXP protease. Collectively, our findings contribute to a better understanding of the dynamic relationship between T6SS and CPS and how they respond swiftly to environmental challenges.

### Competing Interest Statement

The authors have declared no competing interest.

Capsular Polysaccharide Restrains Type VI Secretion in Acinetobacter baumannii

a-baumannii-の細胞表層多糖が-vi-型分泌システムの活性を抑制する


A. baumannii の細胞表層多糖が VI 型分泌システムの活性を抑制する



細菌の細胞間シグナル物質2-アミノアセトフェノンは、免疫細胞のPGC-1α/ERRα経路を阻害することで、ミトコンドリアのピルビン酸取り込みと ATP 産生を低下させ、感染に対する耐性を誘導する。


How bacterial pathogens exploit host metabolism to promote immune tolerance and persist in infected hosts remains elusive. To achieve this, we show that Pseudomonas aeruginosa (PA), a recalcitrant pathogen, utilizes the quorum sensing (QS) signal 2-aminoacetophenone (2-AA). Here, we unveil how 2-AA-driven immune tolerization causes distinct metabolic perturbations in macrophages’ mitochondrial respiration and bioenergetics. We present evidence indicating that these effects stem from a decrease in pyruvate transport into mitochondria. This reduction is attributed to decreased expression of the mitochondrial pyruvate carrier (MPC1), which is mediated by diminished expression and nuclear presence of its transcriptional regulator, estrogen-related nuclear receptor alpha (ERRα). Consequently, ERRα exhibits weakened binding to the MPC1 promoter. This outcome arises from the impaired interaction between ERRα and the peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α). Ultimately, this cascade results in diminished pyruvate influx into mitochondria and consequently reduced ATP production in tolerized macrophages. Exogenously added ATP in infected macrophages restores the transcript levels of MPC1 and ERR-α and enhances cytokine production and intracellular bacterial clearance. Consistent with the in vitro findings, murine infection studies corroborate the 2-AA-mediated long-lasting decrease in ATP and acetyl-CoA and its association with PA persistence, further supporting this QS signaling molecule as the culprit of the host bioenergetic alterations and PA persistence. These findings unveil 2-AA as a modulator of cellular immunometabolism and reveal an unprecedent mechanism of host tolerance to infection involving the PGC-1α/ERRα axis in its influence on MPC1/OXPHOS-dependent energy production and PA clearance. These paradigmatic findings paving the way for developing treatments to bolster resilience to pathogen-induced damage. Given that QS is a common characteristic of prokaryotes, it is likely that 2-AA-like molecules with similar functions may be present in other pathogens.

### Competing Interest Statement

L.G.R. has a financial interest in Spero Therapeutics, a company developing therapies to treat bacterial infections. L.G.R. financial interests are reviewed and managed by Massachusetts General Hospital and Partners Health Care in accordance with their conflict-of-interest policies. No funding was received from Spero Therapeutics, and it had no role in study design, data collection, analysis, interpretation, or the decision to submit the work for publication. The remaining authors declare no competing interests.

The Bacterial Quorum-Sensing Signal 2-Aminoacetophenone Rewires Immune Cell Bioenergetics through the PGC-1α/ERRα Axis to Mediate Tolerance to Infection

pa由細菌の細胞間シグナル物質2-アミノアセトフェノンが免疫細胞のバイオエネルギー代謝を-pgc-1α-errα経路を介して変化させ-感染に対する耐性を誘導する


PA由細菌の細胞間シグナル物質2-アミノアセトフェノンが免疫細胞のバイオエネルギー代謝を PGC-1α/ERRα経路を介して変化させ、感染に対する耐性を誘導する



細菌由来ATPは局所および全身性の炎症反応を抑制し、敗血症の重症化に寄与する。


Sepsis causes millions of deaths per year worldwide and is a current global health priority declared by the WHO. Sepsis-related deaths are a result of dysregulated inflammatory immune responses indicating the need to develop strategies to target inflammation. An important mediator of inflammation is extracellular adenosine triphosphate (ATP) that is released by inflamed host cells and tissues, and also by bacteria in a strain-specific and growth-dependent manner. Here, we investigated the mechanisms by which bacteria release ATP. Using genetic mutant strains of Escherichia coli ( E. coli ), we demonstrate that ATP release is dependent on ATP synthase within the inner bacterial membrane. In addition, impaired integrity of the outer bacterial membrane notably contributes to ATP release and is associated with bacterial death. In a mouse model of abdominal sepsis, local effects of bacterial ATP were analysed using a transformed E. coli bearing an arabinose-inducible periplasmic apyrase hydrolyzing ATP to be released. Abrogating bacterial ATP release shows that bacterial ATP suppresses local immune responses, resulting in reduced neutrophil counts and impaired survival. In addition, bacterial ATP has systemic effects via its transport in outer membrane vesicles (OMV). ATP-loaded OMV are quickly distributed throughout the body and upregulated expression of genes activating degranulation in neutrophils, potentially contributing to the exacerbation of sepsis severity. This study reveals mechanisms of bacterial ATP release and its local and systemic roles in sepsis pathogenesis.

### Competing Interest Statement

The authors have declared no competing interest.

Released Bacterial ATP Shapes Local and Systemic Inflammation during Abdominal Sepsis

腹腔内敗血症における細菌由来atpの局所および全身性炎症への影響


腹腔内敗血症における細菌由来ATPの局所および全身性炎症への影響



腸内細菌は血清中のL-セリンを化学受容体Tsrを介して認識し、血清に強く引き付けられる。この「細菌の吸血行動」は、腸出血病変への定着と増殖を促進し、敗血症リスクを高める可能性がある。


Bacteria of the family Enterobacteriaceae are associated with gastrointestinal (GI) bleeding and bacteremia and are a leading cause of death, from sepsis, for individuals with inflammatory bowel diseases. The bacterial behaviors and mechanisms underlying why these bacteria are prone to bloodstream entry remains poorly understood. Herein, we report that clinical isolates of non-typhoidal Salmonella enterica serovars, Escherichia coli , and Citrobacter koseri are rapidly attracted toward sources of human serum. To simulate GI bleeding, we utilized a custom injection-based microfluidics device and found that femtoliter volumes of human serum are sufficient to induce the bacterial population to swim toward and aggregate at the serum source. This response is orchestrated through chemotaxis, and a major chemical cue driving chemoattraction is L-serine, an amino acid abundant in serum that is recognized through direct binding by the chemoreceptor Tsr. We report the first crystal structures of Salmonella Typhimurium Tsr in complex with L-serine and identify a conserved amino acid recognition motif for L-serine shared among Tsr orthologues. By mapping the phylogenetic distribution of this chemoreceptor we found Tsr to be widely conserved among Enterobacteriaceae and numerous World Health Organization priority pathogens associated with bloodstream infections. Lastly, we find that Enterobacteriaceae use human serum as a source of nutrients for growth and that chemotaxis and the chemoreceptor Tsr provides a competitive advantage for migration into enterohaemorrhagic lesions. We term this bacterial behavior of taxis toward serum, colonization of hemorrhagic lesions, and the consumption of serum nutrients, as “bacterial vampirism” which may relate to the proclivity of Enterobacteriaceae for bloodstream infections.

### Competing Interest Statement

A.B. owns Amethyst Antimicrobials, LLC.

Bacterial vampirism mediated through taxis to serum

腸内出血を引き起こす細菌の化学走性による吸血行動