Pathogen-associated Molecular Patterns Research Articles

Pattern-Triggered Immunity and Effector-Triggered Immunity: crosstalk and cooperation of PRR and NLR-mediated plant defense pathways during host-pathogen interactions.

The elucidation of the molecular basis underlying plant-pathogen interactions is imperative for the development of sustainable resistance strategies against pathogens. Plants employ a dual-layered immunological detection and response system wherein cell surface-localized Pattern Recognition Receptors (PRRs) and intracellular Nucleotide-Binding Leucine-Rich Repeat Receptors (NLRs) play pivotal roles in initiating downstream signalling cascades in response to pathogen-derived chemicals. Pattern-Triggered Immunity (PTI) is associated with PRRs and is activated by the recognition of conserved molecular structures, known as Pathogen-Associated Molecular Patterns. When PTI proves ineffective due to pathogenic effectors, Effector-Triggered Immunity (ETI) frequently confers resistance. In ETI, host plants utilize NLRs to detect pathogen effectors directly or indirectly, prompting a rapid and more robust defense response. Additionally epigenetic mechanisms are participating in plant immune memory. Recently developed technologies like CRISPR/Cas9 helps in exposing novel prospects in plant pathogen interactions. In this review we explore the fascinating crosstalk and cooperation between PRRs and NLRs. We discuss epigenomic processes and CRISPR/Cas9 regulating immune response in plants andrecent findings that shed light on the coordination of these defense layers. Furthermore, we alsohave discussed the intricate interactions between the salicylic acid and jasmonic acid signalling pathways in plants, offering insights into potential synergistic interactions that would be harnessed for the development of novel and sustainable resistance strategies against diverse group of pathogens.

Recombinant CD5 and CD6 Ectodomains Induce Antiparasitic and Immunomodulatory Effects in Secondary Cystic Echinococcosis.

Scavenger receptors participate in a wide range of biological functions after binding to multiple non-self or altered self-ligands. Among them, CD5 and CD6 are lymphocyte scavenger receptors known to interact with different microbial-associated molecular patterns, and the administration of the recombinant soluble ectodomains of human CD5 (rshCD5) and/or CD6 (rshCD6) has shown therapeutic/prophylactic potential in experimental models of fungal, bacterial and echinococcal infections. The latter is a zoonosis caused by the larval stage of the cestode parasite Echinococcus granulosus sensu lato, which in humans can induce secondary cystic echinococcosis (CE) after the spillage of protoscoleces contained within fertile cysts, either spontaneously or during surgical removal of primary hydatid cysts. Herein, we have analysed the mechanisms behind the significant protection observed in the mouse model of secondary CE following prophylactic administration of rshCD5 or rshCD6. Our results show that both molecules exhibit intrinsic antiparasitic activities in vitro, as well as immunomodulatory functions during early secondary CE, mainly through Th1/Th17 cytokine bias and promotion of peritoneal polyreactive antibodies. These data support the relevance of the parasite components bound by rshCD5 and rshCD6, as well as the potential of their prophylactic administration as a useful strategy to reduce secondary CE in patients.

Bivalent histone modifications: how phytopathogens evade plant immunity

Bivalent histone modifications regulate gene expression during development, but little is known about their function in plant–microbe interactions. In a recent report, Zhao et al. showed that expression of bivalent chromatin-marked gene 1 (BCG1), containing a pathogen-associated molecular pattern (PAMP) motif, is epigenetically regulated by trimethylation of lysine 4 (H3K4me3) and lysine 27 (H3K27me3) of histone H3 to evade plant immunity.

Fish ELOVL7a is involved in virus replication via lipid metabolic reprogramming

The elongation of very long chain fatty acids (ELOVL) proteins are key rate-limiting enzymes that catalyze fatty acid synthesis to form long chain fatty acids. ELOVLs also play regulatory roles in the lipid metabolic reprogramming induced by mammalian viruses. However, little is known about the roles of fish ELOVLs during virus infection. Here, a homolog of ELOVL7 was cloned from Epinephelus coioides (EcELOVL7a), and its roles in red-spotted grouper nervous necrosis virus (RGNNV) and Singapore grouper iridovirus (SGIV) infection were investigated. The transcription level of EcELOVL7a was significantly increased upon RGNNV and SGIV infection or other pathogen-associated molecular patterns stimulation in grouper spleen (GS) cells. Subcellular localization analysis showed that EcELOVL7a encoded an endoplasmic reticulum (ER) related protein. Overexpression of EcELOVL7a promoted the viral production and virus release during SGIV and RGNNV infection. Furthermore, the lipidome profiling showed that EcELOVL7a overexpression reprogrammed cellular lipid components in vitro, evidenced by the increase of glycerophospholipids, sphingolipids and glycerides components. In addition, VLCFAs including FFA (20:2), FFA (20:4), FFA (22:4), FFA (22:5) and FFA (24:0), were enriched in EcELOVL7a overexpressed cells. Consistently, EcELOVL7a overexpression upregulated the transcription level of the key lipid metabolic enzymes, including fatty acid synthase (FASN), phospholipase A 2α (PLA 2α), and cyclooxygenases −2 (COX-2), LPIN1, and diacylglycerol acyltransferase 1α (DGAT1α). Together, our results firstly provided the evidence that fish ELOVL7a played an essential role in SGIV and RGNNV replication by reprogramming lipid metabolism.

Olfactory receptors impact pathophysiological processes of lung diseases in bronchial epithelial cells

BackgroundTherapeutic options for steroid-resistant non-type 2 inflammation in obstructive lung diseases are limited. Bronchial epithelial cells are key in the pathogenesis by releasing the central proinflammatory cytokine interleukine-8 (IL-8). Olfactory receptors (ORs) are expressed in various cell types. This study examined the drug target potential of ORs by investigating their impact on associated pathophysiological processes in lung epithelial cells. MethodsExperiments were performed in the A549 cell line and in primary human bronchial epithelial cells. OR expression was investigated using RT-PCR, Western blot, and immunocytochemical staining. OR-mediated effects were analyzed by measuring 1) intracellular calcium concentration via calcium imaging, 2) cAMP concentration by luminescence-based assays, 3) wound healing by scratch assays, 4) proliferation by MTS-based assays, 5) cellular vitality by Annexin V/PI-based FACS staining, and 6) the secretion of IL-8 in culture supernatants by ELISA. ResultsBy screening 100 potential OR agonists, we identified two, Brahmanol and Cinnamaldehyde, that increased intracellular calcium concentrations. The mRNA and proteins of the corresponding receptors OR2AT4 and OR2J3 were detected. Stimulation of OR2J3 with Cinnamaldehyde reduced 1) IL-8 in the absence and presence of bacterial and viral pathogen-associated molecular patterns (PAMPs), 2) proliferation, and 3) wound healing but increased cAMP. In contrast, stimulation of OR2AT4 by Brahmanol increased wound healing but did not affect cAMP and proliferation. Both ORs did not influence cell vitality. ConclusionORs might be promising drug target candidates for lung diseases with non-type 2 inflammation. Their stimulation might reduce inflammation or prevent tissue remodeling by promoting wound healing.

Interleukin-6 receptor (IL-6R) is involved in the innate immune response of olive flounder (Paralichthys olivaceus) to resist pathogens

The immunity of bony fish depends on the conserved components and functional complexity of immune cytokines. Herein, we used the pleiotropic cytokine IL-6 receptor as a target gene to study the defense response of PoIL-6R resist pathogens in different immune tissues of olive flounder, we found that PoIL-6R is widely expressed in these tissues studied and shows a significant immune response after stimulation with bacteria in vivo. In addition, we found a broad-spectrum immune response of PoIL-6R to bacterial and pathogen-associated molecular patterns through in vitro stimulation of olive flounder embryonic cell lines (FEC), and further revealed the function of PoIL-6R in positively regulating the natural immunity of olive flounder through activation of the NF-κB signaling pathway by overexpression of PoIL-6R in FEC. Collectively, these results demonstrate for the involvement of interleukin-6 receptors in the innate immune response of olive flounder to resist pathogens, and provides new perspectives and theoretical foundations for assessing the evolutionary mechanisms of adaptive immunity in vertebrates.

A Review of Temporomandibular Joint (TMJ) Synovitis and the Important Role of the Nuclear Protein, High Mobility Group Box 1 (HMGB1)

Studies identifying TMJ synovitis biomarkers are few and far between. During the late 1990s, Chuck Milam’s oxidative stress and re-perfusion models elucidated the molecular mechanisms of TMJ synovitis which centrally involves the cytokines TNF-α, IL-1β, and IL-6. However, a new understanding of TMJ synovitis has been developed given the fields current knowledge on pattern recognition receptors (PRRs), damage-associated molecular patterns (DAMPs), pathogen-associated molecular patterns (PAMPs) and most notably, high mobility group box 1 (HMGB1). Among these molecular markers, the oxidation of HMGB1 plays a key role in promoting pathological inflammation. In this perspective, we explore the role of HMGB1 during TMJ synovitis and how a traumatized TMJ responds to different HMGB1 post translational modifications (PTMs). Specifically, synovial inflammation will be explored in the context of how different PTMs of HMGB1 directs leukocytes to produce chemokines or cytokines. We will also look at the different modifications of HMGB1 molecule via Reduction Oxygenation Reactions. These recently identified mechanisms provide a suitable addition to the currently understood molecular actions of TMJ synovitis noted by Milam and others in the late 1990s and early 2000s. We will then conclude with a discussion on the use of antibodies to the HMGB1 molecule for a variety of conditions: cancers, sepsis, liver disease, traumatic brain injuries and early intervention for joint synovitis along with the use of different delivery modalities.

Interplay of Cytokines and Chemokines in Aspergillosis.

Aspergillosis is a fungal infection caused by various species of Aspergillus, most notably A. fumigatus. This fungus causes a spectrum of diseases, including allergic bronchopulmonary aspergillosis, aspergilloma, chronic pulmonary aspergillosis, and invasive aspergillosis. The clinical manifestations and severity of aspergillosis can vary depending on individual immune status and the specific species of Aspergillus involved. The recognition of Aspergillus involves pathogen-associated molecular patterns (PAMPs) such as glucan, galactomannan, mannose, and conidial surface proteins. These are recognized by the pathogen recognition receptors present on immune cells such as Toll-like receptors (TLR-1,2,3,4, etc.) and C-type lectins (Dectin-1 and Dectin-2). We discuss the roles of cytokines and pathogen recognition in aspergillosis from both the perspective of human and experimental infection. Several cytokines and chemokines have been implicated in the immune response to Aspergillus infection, including interferon-γ (IFN-γ), tumor necrosis factor-α (TNF-α), CCR4, CCR17, and other interleukins. For example, allergic bronchopulmonary aspergillosis (ABPA) is characterized by Th2 and Th9 cell-type immunity and involves interleukin (IL)-4, IL-5, IL-13, and IL-10. In contrast, it has been observed that invasive aspergillosis involves Th1 and Th17 cell-type immunity via IFN-γ, IL-1, IL-6, and IL-17. These cytokines activate various immune cells and stimulate the production of other immune molecules, such as antimicrobial peptides and reactive oxygen species, which aid in the clearance of the fungal pathogen. Moreover, they help to initiate and coordinate the immune response, recruit immune cells to the site of infection, and promote clearance of the fungus. Insight into the host response from both human and animal studies may aid in understanding the immune response in aspergillosis, possibly leading to harnessing the power of cytokines or cytokine (receptor) antagonists and transforming them into precise immunotherapeutic strategies. This could advance personalized medicine.

Regulation of renal aquaporin water channels in acute pyelonephritis.

Acute pyelonephritis (APN) is most frequently caused by uropathogenic Escherichia coli (UPEC), which ascends from the bladder to the kidneys during a urinary tract infection. Patients with APN have been reported to have reduced renal concentration capacity under challenged conditions, polyuria, and increased aquaporin-2 (AQP2) excretion in the urine. We have recently shown increased AQP2 accumulation in the plasma membrane in cell cultures exposed to E. coli lysates and in the apical plasma membrane of inner medullary collecting ducts in a 5-day APN mouse model. This study aimed to investigate if AQP2 expression in host cells increases UPEC infection efficiency and to identify specific bacterial components that mediate AQP2 plasma membrane insertion. As the transepithelial water permeability in the collecting duct is codetermined by AQP3 and AQP4, we also investigated whether AQP3 and AQP4 localization is altered in the APN mouse model. We show that AQP2 expression does not increase UPEC infection efficiency and that AQP2 was targeted to the plasma membrane in AQP2-expressing cells in response to the two pathogen-associated molecular patterns (PAMPs), lipopolysaccharide and peptidoglycan. In contrast to AQP2, the subcellular localizations of AQP1, AQP3, and AQP4 were unaffected both in lysate-incubated cell cultures and in the APN mouse model. Our finding demonstrated that cellular exposure to lipopolysaccharide and peptidoglycan can trigger the insertion of AQP2 in the plasma membrane revealing a new regulatory pathway for AQP2 plasma membrane translocation, which may potentially be exploited in intervention strategies.NEW & NOTEWORTHY Acute pyelonephritis (APN) is associated with reduced renal concentration capacity and increased aquaporin-2 (AQP2) excretion. Uropathogenic Escherichia coli (UPEC) mediates changes in the subcellular localization of AQP2 and we show that in vitro, these changes could be elicited by two pathogen-associated molecular patterns (PAMPs), namely, lipopolysaccharide and peptidoglycan. UPEC infection was unaltered by AQP2 expression and the other renal AQPs (AQP1, AQP3, and AQP4) were unaltered in APN.

Abstract 2179: Microbiome-derived metabolites mediate carcinogenic alterations of breast tissue in the context of obesity

Abstract Obesity increases the relative risk for breast cancer incidence. Multiple molecular mechanisms linked to obesity drive breast cancer progression. Obesity shifts the gut microbiome in ways that may increase breast cancer risk. Microbial-associated molecular pattern (MAMP)-proteins and metabolites could directly affect breast epithelial cell signaling. In this study, we focused on lipopolysaccharide (LPS), a structural component of the outer membranes of gram-negative bacteria and a toll-like receptor 4 (TLR4)-agonist. Systemic LPS levels are known to increase in obesity. We hypothesize that chronic low-grade inflammation caused by LPS contributes to breast cancer initiation. To test this hypothesis, we first quantified levels of LPS, along with a panel of adipokines/cytokines, in serum samples from women (n= 33; BMI range= 18.3-45.3 Kg/m2; Age range= 18-51 years). Women with elevated serum LPS had significantly higher BMI, leptin, and leptin/adiponectin ratio, confirming a link between systemic LPS and metabolic markers of obesity. Other biomarkers of breast cancer risk include DNA damage. Analyses of non-cancerous breast tissue sections from matched donors revealed higher densities of DNA double-strand breaks (estimated based on 53BP1 foci counts) in women with high serum LPS. Experiments with HMT-3522 S1 breast acini showed that LPS increases DNA break frequencies and oxidative stress levels in 3D structures of mammary epithelium. Moreover, LPS increases oxidative lesions in the DNA (estimated based on 8-oxoguanine fluorescence in MCF-10A cells) that may lead to increased LPS-mediated DNA damage. Patient obesity status was associated with alterations in oxidative modifications in the breast proteome. We also found that LPS disrupts epithelial polarity, a hallmark of breast tissue homeostasis. In mammary epithelium, LPS activates the nuclear factor-kappa B (NFκB) pathway by binding to the TLR4 receptor, leading to an increased expression of inflammatory cytokines such as tumor necrosis factor-alpha (TNF-α). Interestingly, TLR4 blockade prevented the loss of apical polarity and DNA damage induced by LPS. Last, but not least, we show an association between LPS immunogenicity (anti-LPS IgA concentrations) and participant BMI in non-cancerous breast tissue samples. These novel findings show that LPS is a key systemic and local mediator of the obesity-modulated microbiome that increases breast cancer risk. The outcomes of our study underscore the importance of considering the microbiome in the prevention of breast cancer. Citation Format: Mohamed Gaber, Kayla Neary, Julia Holmes, Adam S. Wilson, Adam J. Katz, Pierre-Alexandre Vidi, Katherine L. Cook. Microbiome-derived metabolites mediate carcinogenic alterations of breast tissue in the context of obesity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2179.

Pathophysiology of severe burn injuries: new therapeutic opportunities from a systems perspective.

Severe burn injury elicits a profound stress response with the potential for high morbidity and mortality. If polytrauma is present, patient outcomes appear to be worse. Sex-based comparisons indicate females have worse outcomes than males. There are few effective drug therapies to treat burn shock and secondary injury progression. The lack of effective drugs appears to arise from the current treat-as-you-go approach rather than a more integrated systems approach. In this review, we present a brief history of burns research and discuss its pathophysiology from a systems' perspective. The severe burn injury phenotype appears to develop from a rapid and relentless barrage of damage-associated molecular patterns (DAMPs), pathogen-associated molecular patterns (PAMPs) and neural afferent signals, which leads to a state of hyperinflammation, immune dysfunction, coagulopathy, hypermetabolism and intense pain. We propose that if the central nervous system (CNS) control of cardiovascular function and endothelial-glycocalyx-mitochondrial coupling can be restored early, these secondary injury processes may be minimized. The therapeutic goal is to switch the injury phenotype to a healing phenotype by reducing fluid leak and maintaining tissue O2 perfusion. Currently, no systems-based therapies exist to treat severe burns. We have been developing a small-volume fluid therapy comprising adenosine, lidocaine and magnesium (ALM) to treat hemorrhagic shock, traumatic brain injury and sepsis. Our early studies indicate that the ALM therapy holds some promise in supporting cardiovascular and pulmonary functions following severe burns. Future research will investigate the ability of ALM therapy to treat severe burns with polytrauma and sex disparities, and potential translation to humans.

A bacterial toxin co-opts caspase-3 to disable active gasdermin D and limit macrophage pyroptosis

During infections, host cells are exposed to pathogen-associated molecular patterns (PAMPs) and virulence factors that stimulate multiple signaling pathways that interact additively, synergistically, or antagonistically. The net effect of such higher-order interactions is a vital determinant of the outcome of host-pathogen interactions. Here, we demonstrate one such complex interplay between bacterial exotoxin- and PAMP-induced innate immune pathways. We show that two caspases activated during enterohemorrhagic Escherichia coli (EHEC) infection by lipopolysaccharide (LPS) and Shiga toxin (Stx) interact in a functionally antagonistic manner; cytosolic LPS-activated caspase-11 cleaves full-length gasdermin D (GSDMD), generating an active pore-forming N-terminal fragment (NT-GSDMD); subsequently, caspase-3 activated by EHEC Stx cleaves the caspase-11-generated NT-GSDMD to render it nonfunctional, thereby inhibiting pyroptosis and interleukin-1β maturation. Bacteria typically subvert inflammasomes by targeting upstream components such as NLR sensors or full-length GSDMD but not active NT-GSDMD. Thus, our findings uncover a distinct immune evasion strategy where a bacterial toxin disables active NT-GSDMD by co-opting caspase-3.

Abstract 6739: Efficacy and toxicity of microbial mimetic mesoporous silica nanoparticles in a mouse model of ovarian cancer

Abstract Ovarian cancer is predominately left undetected and untreated until the later stages of the disease where it becomes incredibly difficult to treat. Preclinical studies demonstrate that immunotherapy can be highly effective. Our lab has been working on an immunotherapy approach to treat women with ovarian cancer that utilizes mesoporous silica nanoparticles(MSN). MSN that present pathogen-associated molecular patterns (PAMPS) are able to significantly reduce tumor burden in mouse models of serous epithelial ovarian cancer when administered by intraperitoneal injection. These microbial mimetic particles are decorated with polyethyleneimine (PEI), CpG oligonucleotide, and monophosphoryl lipid A (MPLA). When in the presence of antigen presenting cells, they bind to Toll-like receptors (TLR) present on the cell surface and within the endosome. To better understand mechanisms underlying their efficacy, we injected fluorescent MSN-PEI-CpG-MPLA (100 or 500 nm) or unmodified MSN into mice with advanced ovarian cancer. The particles, regardless of the presence of TLR-agonists, were rapidly internalized by myeloid cells, with trafficking almost exclusively to immune clusters located in tumor-burdened adipose tissues when administered by intraperitoneal injection. Conversely, intravenous injection of MSN-PEI-CpG-MPLA led to predominate accumulation in liver, with no particles found in tertiary lymphoid clusters 24 hours post injection. Furthermore, unlike intraperitoneal injection, intravenous injection of MSN-PEI-CpG-MPLA was without therapeutic effect. Currently we are studying MSN safety and the impact of administration route and dose on efficacy and cytotoxicity. In summary, MSN, with and without TLR agonists, are trafficked by myeloid cells to tumor-burdened fat-associated lymphoid clusters when administered by intraperitoneal injection. Intraperitoneal injection of TLR-agonist modified MSN effectively reduces tumor burden in mice with ovarian cancer. Citation Format: Lorel Y. Medina, Rita E. Serda, Benjamin Marwedel, Melanie S. Jun, Josh Adogola. Efficacy and toxicity of microbial mimetic mesoporous silica nanoparticles in a mouse model of ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6739.

Joint application of plant immunity-inducing elicitors and fungicides to control Phytophthora diseases

Phytophthora are destructive plant pathogens that pose a serious threat to crop production. Traditional control methods rely heavily on chemical fungicides, which are harmful to the environment and human health. Currently, effective green prevention and control methods for Phytophthora pathogens are lacking. Plants rely primarily on their innate immune system to resist pathogens. Plant cells perceive pathogen invasion and activate immune responses by recognizing specific pathogen-derived molecules, called elicitors, which mainly include pathogen-associated molecular patterns (PAMPs) and microbial effector proteins. PAMPs, which are conserved molecular features of microbes and recognized by plant cell surface-localized pattern-recognition receptors (PRRs), activate mild and broad-spectrum disease resistance. However, there are few reports on elicitor proteins that induce broad resistance against Phytophthora pathogens. In this study, we identified BcIEB1, a fungal-derived PAMP, which activated plant immune responses in a BAK1- and SOBIR1-dependent manner. BcIEB1 could induce plant resistance to various Phytophthora pathogens, including P. capsici, P. infestans, and P. parasitica. We further found that the combination of lower concentrations of BcIEB1 with fungicides, such as pyraclostrobin, azoxystrobin, and metalaxyl-M, could enhance the effect on Phytophthora disease control while reducing the dependence on fungicides, thereby reducing environmental pollution. This study identified a novel, less toxic strategy for controlling Phytophthora diseases.

Mitogen-activated protein kinase phosphatase 1 controls broad spectrum disease resistance in Arabidopsis thaliana through diverse mechanisms of immune activation.

Arabidopsis thaliana Mitogen-activated protein Kinase Phosphatase 1 (MKP1) negatively balances production of reactive oxygen species (ROS) triggered by Microbe-Associated Molecular Patterns (MAMPs) through uncharacterized mechanisms. Accordingly, ROS production is enhanced in mkp1 mutant after MAMP treatment. Moreover, mkp1 plants show a constitutive activation of immune responses and enhanced disease resistance to pathogens with distinct colonization styles, like the bacterium Pseudomonas syringae pv. tomato DC3000, the oomycete Hyaloperonospora arabidopsidis Noco2 and the necrotrophic fungus Plectosphaerella cucumerina BMM. The molecular basis of this ROS production and broad-spectrum disease resistance controlled by MKP1 have not been determined. Here, we show that the enhanced ROS production in mkp1 is not due to a direct interaction of MKP1 with the NADPH oxidase RBOHD, nor is it the result of the catalytic activity of MKP1 on RBHOD phosphorylation sites targeted by BOTRYTIS INDUCED KINASE 1 (BIK1) protein, a positive regulator of RBOHD-dependent ROS production. The analysis of bik1 mkp1 double mutant phenotypes suggested that MKP1 and BIK1 targets are different. Additionally, we showed that phosphorylation residues stabilizing MKP1 are essential for its functionality in immunity. To further decipher the molecular basis of disease resistance responses controlled by MKP1, we generated combinatory lines of mkp1-1 with plants impaired in defensive pathways required for disease resistance to pathogen: cyp79B2 cyp79B3 double mutant defective in synthesis of tryptophan-derived metabolites, NahG transgenic plant that does not accumulate salicylic acid, aba1-6 mutant impaired in abscisic acid (ABA) biosynthesis, and abi1 abi2 hab1 triple mutant impaired in proteins described as ROS sensors and that is hypersensitive to ABA. The analysis of these lines revealed that the enhanced resistance displayed by mkp1-1 is altered in distinct mutant combinations: mkp1-1 cyp79B2 cyp79B3 fully blocked mkp1-1 resistance to P. cucumerina, whereas mkp1-1 NahG displays partial susceptibility to H. arabidopsidis, and mkp1-1 NahG, mkp1-1 aba1-6 and mkp1-1 cyp79B2 cyp79B3 showed compromised resistance to P. syringae. These results suggest that MKP1 is a component of immune responses that does not directly interact with RBOHD but rather regulates the status of distinct defensive pathways required for disease resistance to pathogens with different lifestyles.

Активность липополисахарида типового штамма Azospirillum palustre B2 в отношении проростков пшеницы (Triticum aestivum L.)

Lipopolysaccharide is the main structural component of the outer membrane of Gram-negative bacteria, which can also be a part of extracellular polymeric substances. Lipopolysaccharides of bacteria that stimulate the growth and development of plants belong to the group of molecules that form a microbe-associated molecular pattern (MAMP). These glycoconjugates of both symbiotic and phytopathogenic bacteria induce the activation of immune responses in plants. However, the level of plant response under the influence of symbiont lipopolysaccharides differs significantly, also due to their structural features, which make it possible to bypass or weaken the reactions of innate autoimmunity. In this paper, we present the results of the analysis of the reactions of wheat seedlings Triticum aestivum L. after incubation with lipopolysaccharide of associative bacteria Azospirillum palustre B2(T). Incubation of wheat seedlings in the presence of A. palustre B2 lipopolysaccharide for three days led to the activation of plant growth processes, namely an increase in the length of shoots, roots, the area of the first leaf, and a change in the content of pigments in the leaves.

Chimeric NOD2 Mincle Agonists as Vaccine Adjuvants.

There is a need for improved vaccine adjuvants to augment vaccine efficacy. One way to address this is by targeting multiple immune cell pathogen recognition receptors (PRRs) using chimeric pathogen-associated molecular patterns (PAMPs). Conjugation of the PAMPs will ensure codelivery of the immunostimulatory molecules to the same cell, enhancing adjuvant activity. The macrophage inducible C-type lectin (Mincle) is a promising PRR for adjuvant development; however, no effective chimeric Mincle adjuvants have been prepared. We addressed this by synthesizing Mincle adjuvant conjugates, MDP-C18Brar and MDP-C18Brar-dilipid, which contain PAMPs recognized by Mincle and the nucleotide-binding oligomerization domain 2 (NOD2). The two PAMPs are joined by a pH-sensitive oxyamine linker which, upon acidification at lysosomal pH, hydrolyzed to release the NOD2 ligands. The conjugates elicited the production of Th1 and Th17 promoting cytokines in vitro, and when using OVA as a model antigen, exhibited enhanced T-cell-mediated immune responses and reduced toxicity in vivo, compared to the coadministration of the adjuvants.

Adverse Environmental Perturbations May Threaten Kelp Farming Sustainability by Exacerbating Enterobacterales Diseases.

Globally kelp farming is gaining attention to mitigate land-use pressures and achieve carbon neutrality. However, the influence of environmental perturbations on kelp farming remains largely unknown. Recently, a severe disease outbreak caused extensive kelp mortality in Sanggou Bay, China, one of the world's largest high-density kelp farming areas. Here, through in situ investigations and simulation experiments, we find indications that an anomalously dramatic increase in elevated coastal seawater light penetration may have contributed to dysbiosis in the kelp Saccharina japonica's microbiome. This dysbiosis promoted the proliferation of opportunistic pathogenic Enterobacterales, mainly including the genera Colwellia and Pseudoalteromonas. Using transcriptomic analyses, we revealed that high-light conditions likely induced oxidative stress in kelp, potentially facilitating opportunistic bacterial Enterobacterales attack that activates a terrestrial plant-like pattern recognition receptor system in kelp. Furthermore, we uncover crucial genotypic determinants of Enterobacterales dominance and pathogenicity within kelp tissue, including pathogen-associated molecular patterns, potential membrane-damaging toxins, and alginate and mannitol lysis capability. Finally, through analysis of kelp-associated microbiome data sets under the influence of ocean warming and acidification, we conclude that such Enterobacterales favoring microbiome shifts are likely to become more prevalent in future environmental conditions. Our study highlights the need for understanding complex environmental influences on kelp health and associated microbiomes for the sustainable development of seaweed farming.

Differential expression of immune-related biomarkers in primary cultures from Atlantic salmon (Salmo salar) exposed to processed Paecilomyces variotii with or without inactivated Moritella viscosa

Paecilomyces variotii (a filamentous fungus), is a promising novel protein source in fish feeds due to its high nutritional value. Also, P. variotii has Microbial-Associated Molecular Patterns (MAMPs) such as glucans and nucleic acids that could modulate the host's immune response. To understand the potential bioactive properties of this fungus in Atlantic salmon (Salmo salar), our study was conducted to evaluate the gene expression of immune-related biomarkers (e.g., cytokines, effector molecules and receptors) on primary cultures from salmon head kidney (HKLs) and spleen leukocytes (SLs) exposed to either UV inactivated or fractions from P. variotii with or without inactivated Moritella viscosa (a skin pathogen in salmonids). Moreover, the effect of the fermentation conditions and down-stream processing on the physical ultrastructure and cell wall glucan content of P. variotii was characterized. The results showed that drying had a significant effect on the cell wall ultrastructure of the fungi and the choice of fermentation has a significant effect on the quantity of β-glucans in P. variotii. Furthermore, stimulating Atlantic salmon HKLs and SLs with P. variotii and its fractions induced gene expression related to pro-inflammatory (tnfα, il1β) and antimicrobial response (cath2) in HKLs, while response in SLs was related to both pro-inflammatory and regulatory response (tnfα, il6 and il10). Similarly, the stimulation with inactivated M. viscosa alone led to an up-regulation of genes related to pro-inflammatory (tnfα, il1β, il6) antimicrobial response (cath2), intra-cellular signalling and recognition of M. viscosa (sclra, sclrb) and a suppression of regulatory response (il10) in both HKLs and SLs. Interestingly, the co-stimulation of cells with P. variotii and M. viscosa induced immune homeostasis (il6, tgfβ) and antimicrobial response (cath2) in SLs at 48h. Thus, P. variotii induces immune activation and cellular communication in Atlantic salmon HKLs and SLs and modulates M. viscosa induced pro-inflammatory responses in SLs. Taken together, the results from physical and chemical characterization of the fungi, along with the differential gene expression of key immune biomarkers, provides a theoretical basis for designing feeding trials and optimize diets with P. variotii as a functional novel feed ingredient for Atlantic salmon.

Characterization of the ADP-β-d-manno-heptose biosynthetic enzymes from two pathogenic Vibrio strains

ADP-activated β-d-manno-heptoses (ADP-β-d-manno-heptoses) are precursors for the biosynthesis of the inner core of lipopolysaccharide in Gram-negative bacteria. Recently, ADP-d-glycero-β-d-manno-heptose (ADP-d,d-manno-heptose) and its C-6′′ epimer, ADP-l-glycero-β-d-manno-heptose (ADP-l,d-manno-heptose), were identified as potent pathogen-associated molecular patterns (PAMPs) that can trigger robust innate immune responses. Although the production of ADP-d,d-manno-heptose has been studied in several different pathogenic Gram-negative bacteria, current knowledge of ADP-β-d-manno-heptose biosynthesis in Vibrio strains remains limited. Here, we characterized the biosynthetic enzymes of ADP-d,d-manno-heptose and the epimerase that converts it to ADP-l,d-manno-heptose from Vibrio cholerae (the causative agent of pandemic cholera) and Vibrio parahaemolyticus (non-cholera pathogen causing vibriosis with clinical manifestations of gastroenteritis and wound infections) in comparison with their isozymes from Escherichia coli. Moreover, we discovered that β-d-mannose 1-phosphate, but not α-d-mannose 1-phosphate, could be activated to its ADP form by the nucleotidyltransferase domains of bifunctional kinase/nucleotidyltransferases HldEVC (from V. cholerae) and HldEVP (from V. parahaemolyticus). Kinetic analyses of the nucleotidyltransferase domains of HldEVC and HldEVP together with the E. coli–derived HldEEC were thus carried out using β-d-mannose 1-phosphate as a mimic sugar substrate. Overall, our works suggest that V. cholerae and V. parahaemolyticus are capable of synthesizing ADP-β-d-manno-heptoses and lay a foundation for further physiological function explorations on manno-heptose metabolism in Vibrio strains.Key points• Vibrio strains adopt the same biosynthetic pathway as E. coli in synthesizing ADP-β-d-manno-heptoses.• HldEs from two Vibrio strains and E. coli could activate β-d-mannose 1-phosphate to ADP-β-d-mannose.• Comparable nucleotidyltransfer efficiencies were observed in the kinetic studies of HldEs.