Virulence Proteins Research Articles

EhVps35, a retromer component, is involved in the recycling of the EhADH and Gal/GalNac virulent proteins of Entamoeba histolytica

The retromer is a highly conserved eukaryotic complex formed by the cargo selective complex (CSC) and the sorting nexin (SNX) dimer subcomplexes. Its function is protein recycling and recovery from the endosomes to conduct the target molecules to the trans-Golgi network or the plasma membrane. The protozoan responsible for human amoebiasis, Entamoeba histolytica, exhibits an active membrane movement and voracious phagocytosis, events in which the retromer may be fully involved. In this work, we studied the structure of EhVps35 the central member of the CSC retromeric subcomplex as it binds EhVps26 and EhVps29, the other two CSC members, allowing the position of the retromer in the membranes. We also studied the EhVps35 role in the recycling of virulence proteins, particularly those involved in phagocytosis. Confocal microscopy assays revealed that EhVps35 is located in the plasmatic and endosomal membranes and in the phagocytic cups and channels. In addition, it follows the target cell from the moment it is in contact with the trophozoites. Molecular docking analyses, immunoprecipitation assays, and microscopy studies revealed that EhVps35 interacts with the EhADH, Gal/GalNac lectin, and actin proteins. In addition, experimental evidence indicated that it recycles surface proteins, particularly EhADH and Gal/GalNac proteins, two molecules highly involved in virulence. Knockdown of the Ehvps35 gene induced a decrease in protein recycling, as well as impairments in the efficiency of adhesion and the rate of phagocytosis. The actin cytoskeleton was deeply affected by the Ehvps35 gene knockdown. In summary, our results revealed the participation of EhVps35 in protein recycling and phagocytosis. Furthermore, altogether, our results demonstrated the concert of finely regulated molecules, including EhVps35, EhADH, Gal/GalNac lectin, and actin, in the phagocytosis of E. histolytica.

Unmasking the dynamics of Mycoplasma gallisepticum: deciphering HD11 macrophage polarization for innovative infection control strategies

Mycoplasma gallisepticum (MG) is a highly contagious avian respiratory pathogen characterized by rapid spread, widespread distribution, and long-term persistence of infection. Previous studies have shown that chicken macrophage HD11 cells play a critical role in the replication and immunomodulation of MG. Macrophages are multifunctional immunomodulatory cells that polarize into different functions and morphologies in response to exogenous stimuli. However, the effect of MG infection on HD11 polarization is not well understood. In this study, we observed a time-dependent increase in both the expression of the MG-related virulence protein pMGA1.2 and the copy number of MG upon MG infection. Polarization studies revealed an upregulation of M1-type marker genes in MG-infected HD11 cells, suggesting that MG mainly induces HD11 macrophages towards M1-type polarization. Furthermore, MG activated the inflammatory vesicle NLRP3 signaling pathway, and NLRP3 inhibitors affected the expression of M1 and M2 marker genes, indicating the crucial regulatory role of the NLRP3 signaling pathway in MG-induced polarization of HD11 macrophages. Our findings reveal a novel mechanism of MG infection, namely the polarization of MG-infected HD11 macrophages. This discovery suggests that altering the macrophage phenotype to inhibit MG infection may be an effective control strategy. These findings provide new perspectives on the pathogenic mechanism and control measures of MG.

Assessing the efficacy of cinnamon compounds against H. pylori through molecular docking, MD Simulations and ADMET analyses.

Antibiotics are the drugs that are used for the management of microbial diseases. However, these conventional synthetic drugs can harmfully affect the human health. Since phytochemicals are extracted from natural sources and, are hence relatively safer for human health, they are the enticing alternatives in this regard. Cinnamon is also one of those plants which is being employed as herbal medication for centuries against certain microbial infections due its significant therapeutic effectiveness. A well-known pathogenic bacterium called H. pylori causes a wide range of illnesses in human body. This pathogen's pathogenicity is determined by certain virulent proteins. In this study, some of such proteins, which included virB4, virB8, and virB9 were selected to evaluate the therapeutic efficiency of cinnamon compounds. These proteins were identified in different isolates of H. pylori. The structural modelling of all these proteins were performed initially in order to proceed them for molecular docking analysis. While, the docking studies illustrated that one of the cinnamon compounds, cinnamyl acetate, showed significant binding interactions with virB4 and virB9. However, benzyl benzoate which is another cinnamon compound, docked well with virB8. Afterwards, the MD simulations were incorporated to explore the interaction motions and structural stability of all the docked complexes. In this regard, the resultant maps of Bfactor, eigenvalues and elastic network model, among other factors ensured the structural stabilities of all the respective complexes. After these crucial estimations, benzyl benzoate and cinnamyl acetate underwent the ADMET investigation to assess their pharmacokinetic characteristics. SwissADME and ADMETLab 2.0 server were employed for this investigation. The compiled findings these servers revealed that both, benzyl benzoate and cinnamyl acetate, exhibited a significant level of pharmacokinetic and drug-likeness conformity.

In Silico Prediction and Molecular Simulation of Antimicrobial Peptide Variants From Lactobacillus sp. Against Porphyromonas gingivalis and Fusobacterium nucleatum in Oral Squamous Cell Carcinoma

ABSTRACT Porphyromonas gingivalis and Fusobacterium nucleatum are known to contribute to a variety of tumorigenic pathways linked to the progression of oral squamous cell carcinoma (OSCC). The growing global incidence of antibiotic resistance highlights the critical need to consider the use of antimicrobial peptides (AMPs) as a viable alternative to conventional antibiotics. The current study comprehensively tested Lactobacillus sp.−derived AMPs against bacterially exacerbated OSCC. A total of 52 AMPs were obtained from various databases, and an in silico analysis determined their potent antibacterial and anticancer characteristics after a rigorous screening and pruning approach. Twelve AMPs were tested for 3D structural alignment prediction and validation, with the GH12 synthetic AMP serving as a control. These candidate peptides were thoroughly screened against six important virulence proteins of P. gingivalis and four of F. nucleatum, with the lowest energy score of the docked complexes measuring binding affinity and interactions with active residues being chosen. plpl_18 was determined as the most efficient new AMP that interacted with the virulence protein RagB of P. gingivalis and Fap2 of F. nucleatum with docking scores of −238.24 and −254.27 kcal/mol, respectively. This AMP plpl_18 was docked against selective target OSCC regulatory proteins such as cytokines, metallomatrix proteinase, MAPK, E‐cadherin, and JAK‐1 proteins. Among these proteins, it docked against matrix metalloproteinase‐9 with the highest negative docking scores of −7.5, −260.956, and −1361.9 kcal/mol using AutoDock Vina, HPEPDOCK, and ClusPro 2.0, respectively. Molecular dynamic simulation was used to perform extrapolated validation. These computational studies provide an essential foundation for anticipated laboratory and clinical investigations concerning the possibility of adapting therapeutic peptides based on probiotics to combat the proliferation of OSCC, which is accelerated by F. nucleatum and P. gingivalis.

Distinct tomato yellow leaf curl Chuxiong virus isolated from whiteflies and plants in China and its symptom determinant and suppressor of post-transcriptional gene silencing

A begomovirus isolated from whiteflies (Bemisia tabaci) and tomato, sweet potato in China was found to be representative of a distinct begomovirus species, for which the name tomato yellow leaf curl Chuxiong virus (TYLCCxV) is proposed. The results of genomic identification and sequence comparison showed that TYLCCxV shares the highest complete nucleotide sequence identity (88.3%) with croton yellow vein mosaic virus (CroYVMV), and may have originated from the recombination between synedrella leaf curl virus (SyLCV) and squash leaf curl Yunnan virus (SLCuYV). Agrobacterium-mediated inoculation showed that TYLCCxV is highly infectious for a range of plant species, producing upward leaf curling, leaf crumpling, chlorosis, distortion, and stunt symptoms in Solanum lycopersicum plants. The results of Southern blot indicated that TYLCCxV is capable of efficiently replicating two heterologous betasatellites. The inoculation of PVX::C4 on Nicotiana benthamiana induced upward leaf curling and stem elongation symptoms, suggesting that TYLCCxV C4 functions as a symptom determinant. TYLCCxV V2 is an important virulence factor that induces downward leaf curling symptoms, elicits systemic necrosis, and suppresses local and systemic GFP silencing in co-agroinfiltrated N. benthamiana and transgenic 16c plants. Considering the multifunctional virulence proteins V2 and C4, the possibility of TYLCCxV causing devastating epidemics on tomato in China is discussed.

Antibacterial Profiling of Zanthoxylum armatum Extracts: A Comprehensive Computational and Experimental Study

Objectives The current study was conducted to evaluate the antibacterial potential of leaf and fruit extracts of Zanthoxylum armatum against two pathogenic bacterial isolates, Staphylococcus aureus and Staphylococcus epidermidis. Methods Twelve commercially available antibiotics were tested S. aureus and S. epidermidis by antimicrobial susceptibility test (AST). Qualitative analysis of phytochemicals was performed to evaluate the presence of certain secondary metabolites. The activity of Z. armatum extracts against S. aureus and S. epidermidis was measured as a maximum zone of inhibition exhibited by each leaf and fruit extract. An in-silico study was conducted on flavonoids and alkaloids to show their binding affinity with the PBP2a receptor protein of S. aureus and TcaR of S. epidermidis. Results The AST revealed that S. aureus was resistant to Penicillin, Ampicillin, Clindamycin, Vancomycin, Rifampicin, Novobiocin, and Oxacillin, whereas S. epidermidis was resistant to Streptomycin, Oxacillin, Tetracycline, and Novobiocin. Qualitative analysis of phytochemicals resulted in the presence of Saponins, fixed oils, flavonoids, alkaloids, starch, and fatty acids in both leaf and fruit extracts. The maximum zone of inhibition against S. aureus was produced by methanolic leaf extracts of Z. armatum and chloroform fruit extracts. For S. epidermidis, the best activity was exhibited by benzene leaf extracts and methanolic fruit extracts. An in-silico study showed that flavonoids Nitidine and Nevadensin exhibited binding affinity with the PBP2a receptor protein higher than selected antibiotics, ie, Penicillin, Chloramphenicol, and Oxacillin. TcaR of S. epidermidis interacted with Tambuletin, followed by Nitidine and Kaempferol. Conclusion After in vitro testing, in silico analysis advised extracting and purifying the bioactive components from Z. armatum extracts that showed significant interaction with bacterial virulence proteins for use as natural antibiotics against antibiotic-resistant bacteria.

Virtual analysis of machine learning models for diseases prediction in muskmelon

Muskmelon, a crop prized for its economic potential, has a relatively brief growth cycle. Disease susceptibility during this period can have a profound impact on yields, posing challenges for farmers. Environmental conditions are pivotal in disease occurrence. Unfavorable conditions reduce the likelihood of pathogens infecting vulnerable host plants as temperature and humidity influence pathogen behavior, including toxin synthesis, virulence protein production, and reproduction. Pathogens can lie dormant in the soil until suitable conditions activate them. When the right environment and host plants align, these dormant pathogens can cause outbreaks. Disease prediction becomes possible by analyzing environmental variables. Real-time data collected via strategically placed sensors focused on viral, fungal, and bacterial infections. Results indicated that the extreme gradient boosting (XGBoost) algorithm, with a maximum tree depth of 4 and 30 trees per iteration, achieved remarkable performance, yielding an accuracy of 97%. For comparison, the XGBoost model outperformed an 8-layer Backpropagation network with 7 nodes per layer, which achieved 95% accuracy. These findings underscore XGBoost's efficacy in forecasting and mitigating muskmelon plant diseases, offering the potential for improved crop yields and agricultural sustainability.

The WY Domain of an RxLr Effector Drives Interactions with a Host Target Phosphatase to Mimic Host Regulatory Proteins and Promote Phytophthora infestans Infection.

Plant pathogens manipulate the cellular environment of the host to facilitate infection and colonization that often lead to plant diseases. To accomplish this, many specialized pathogens secrete virulence proteins called effectors into the host cell, which subvert processes such as immune signaling, gene transcription, and host metabolism. Phytophthora infestans, the causative agent of potato late blight, employs an expanded repertoire of RxLR effectors with WY domains to manipulate the host through direct interaction with protein targets. However, our understanding of the molecular mechanisms underlying the interactions between WY effectors and their host targets remains limited. In this study, we performed a structural and biophysical characterization of the P. infestans WY effector Pi04314 in complex with the potato Protein Phosphatase 1-c (PP1c). We elucidate how Pi04314 uses a WY domain and a specialized C-terminal loop carrying a KVxF motif that interact with conserved surfaces on PP1c, known to be used by host regulatory proteins for guiding function. Through biophysical and in planta analyses, we demonstrate that Pi04314 WY or KVxF mutants lose their ability to bind PP1c. The loss of PP1c binding correlates with changes in PP1c nucleolar localization and a decrease in lesion size in plant infection assays. This study provides insights into the manipulation of plant hosts by pathogens, revealing how effectors exploit key regulatory interfaces in host proteins to modify their function and facilitate disease. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

The N-terminal domain is required for cell surface localisation of VapA, a member of the Vap family of Rhodococcus equi virulence proteins.

Rhodococcus equi pneumonia is an important cause of mortality in foals worldwide. Virulent equine isolates harbour an 80-85kb virulence plasmid encoding six virulence-associated proteins (Vaps). VapA, the main virulence factor of this intracellular pathogen, is known to be a cell surface protein that creates an intracellular niche for R. equi growth. In contrast, VapC, VapD and VapE are secreted into the intracellular milieu. Although these Vaps share very high degree of sequence identity in the C-terminal domain, the N-terminal domain (N-domain) of VapA is distinct. It has been proposed that this domain plays a role in VapA surface localization but no direct experimental data provides support to such hypothesis. In this work, we employed R. equi 103S harbouring an unmarked deletion of vapA (R. equi ΔvapA) as the genetic background to express C-terminal Strep-tagged Vap-derivatives integrated in the chromosome. The surface localization of these proteins was assessed by flow cytometry using the THE2122;-NWSHPQFEK Tag FITC-antibody. We show that VapA is the only cell surface Vap encoded in the virulence plasmid. We present compelling evidence for the role of the N-terminal domain of VapA on cell surface localization using fusion proteins in which the N-domain of VapD was exchanged with the N-terminus of VapA. Lastly, using an N-terminally Strep-tagged VapA, we found that the N-terminus of VapA is exposed to the extracellular environment. Given the lack of a lipobox in VapA and the exposure of the N-terminal Strep-tag, it is possible that VapA localization on the cell surface is mediated by interactions between the N-domain and components of the cell surface. We discuss the implications of this work on the light of the recent discovery that soluble recombinant VapA added to the extracellular medium functionally complement the loss of VapA.

Essential roles of ferric reductase-like proteins in growth, development, stress response, and virulence of the filamentous entomopathogenic fungus Beauveria bassiana

In yeasts, ferric reductase catalyzes reduction of ferric ion to ferrous form, which is essential for the reductive iron assimilation system. However, the physiological roles of ferric reductases remain largely unknown in the filamentous fungi. In this study, genome-wide annotation revealed thirteen ferric reductase-like (Fre) proteins in the filamentous insect pathogenic fungus Beauveria bassiana, and all their functions were genetically characterized. Ferric reductase family proteins exhibit different sub-cellular distributions (e.g., cell periphery and vacuole), which was due to divergent domain architectures. Fre proteins had a synergistic effect on fungal virulence, which was ascribed to their distinct functions in different physiologies. Ten Fre proteins were not involved in reduction of ferric ion in submerged mycelia, but most proteins contributed to blastospore development. Only two Fre proteins significantly contributed to B. bassiana vegetative growth under the chemical-induced iron starvation, but most Fre proteins were involved in resistance to osmotic and oxidative stresses. Notably, a bZIP-type transcription factor HapX bound to the promoter regions of all FRE genes in B. bassiana, and displayed varying roles in the transcription activation of these genes. This study reveals the important role of BbFre family proteins in development, stress response, and insect pathogenicity, as well as their distinctive role in the absorption of ferric iron from the environment.

Identification of a new export signal that targets early subunits to the flagellar type III secretion export machinery.

Many bacterial pathogens utilize T3SS to inject virulence proteins (effectors) into host cells or to assemble flagella on the bacterial cell surface. Bacterial flagella present a paradigm for how cells build and operate complex cell-surface "nanomachines." Efficient subunit targeting from the bacterial cytosol to type III secretion systems is essential for rapid assembly and secretion by T3SSs. Subunits are thought to dock at the export machinery before being unfolded and translocated into the export channel. However, little is known about how subunits dock at the export machinery and the events that occur post docking. Here, we identified a new export signal within the C-termini of subunits that is essential for targeting of subunits to the type III export machinery. We show that this new export signal and previously identified export signals are recognized separately and sequentially, revealing a pathway for subunit transit through the type III export machinery in which sequential recognition events carry out different roles at major steps in the export pathway.

Rice false smut virulence protein subverts host chitin perception and signaling at lemma and palea for floral infection.

The flower-infecting fungus Ustilaginoidea virens causes rice false smut, which is a severe emerging disease threatening rice (Oryza sativa) production worldwide. False smut not only reduces yield, but more importantly produces toxins on grains, posing a great threat to food safety. U. virens invades spikelets via the gap between the 2 bracts (lemma and palea) enclosing the floret and specifically infects the stamen and pistil. Molecular mechanisms for the U. virens-rice interaction are largely unknown. Here, we demonstrate that rice flowers predominantly employ chitin-triggered immunity against U. virens in the lemma and palea, rather than in the stamen and pistil. We identify a crucial U. virens virulence factor, named UvGH18.1, which carries glycoside hydrolase activity. Mechanistically, UvGH18.1 functions by binding to and hydrolyzing immune elicitor chitin and interacting with the chitin receptor CHITIN ELICITOR BINDING PROTEIN (OsCEBiP) and co-receptor CHITIN ELICITOR RECEPTOR KINASE1 (OsCERK1) to impair their chitin-induced dimerization, suppressing host immunity exerted at the lemma and palea for gaining access to the stamen and pistil. Conversely, pretreatment on spikelets with chitin induces a defense response in the lemma and palea, promoting resistance against U. virens. Collectively, our data uncover a mechanism for a U. virens virulence factor and the critical location of the host-pathogen interaction in flowers and provide a potential strategy to control rice false smut disease.

Identifying virulence factors using graph transformer autoencoder with ESMFold-predicted structures

With the increasing resistance of bacterial pathogens to conventional antibiotics, antivirulence strategies targeting virulence factors (VFs) have become an effective new therapy for the treatment of pathogenic bacterial infections. Therefore, the identification and prediction of VFs can provide ideal candidate targets for the implementation of antivirulence strategies in treating infections caused by pathogenic bacteria. Currently, the existing computational models predominantly rely on the amino acid sequences of virulence proteins while overlooking structural information. Here, we propose a novel graph transformer autoencoder for VF identification (GTAE-VF), which utilizes ESMFold-predicted 3D structures and converts the VF identification problem into a graph-level prediction task. In an encoder-decoder framework, GTAE-VF adaptively learns both local and global information by integrating a graph convolutional network and a transformer to implement all-pair message passing, which can better capture long-range correlations and potential relationships. Extensive experiments on an independent test dataset demonstrate that GTAE-VF achieves reliable and robust prediction accuracy with an AUC of 0.963, which is consistently better than that of other structure-based and sequence-based approaches. We believe that GTAE-VF has the potential to emerge as a valuable tool for assessing VFs and devising antivirulence strategies.

Pins Gene Table v2.0: An Online Genome Database of 37 Pythium insidiosum Strains for Gene Content Exploration and Phylogenomic Analysis.

Unlike most pathogenic oomycetes, Pythium insidiosum infects humans and animals instead of plants. P. insidiosum has three clinically relevant genotypes/clades that cause a severe disease called pythiosis. To develop strategies for infection control, it is necessary to understand the biology and pathogenesis of this pathogen. Investigating the evolutionary mechanisms behind the host-specific adaptation is vital, and comparative genomic analysis can help with this. To facilitate genomic analysis, an online bioinformatics tool called P. insidiosum (Pins) Gene Table v2.0 was developed. This tool includes genomic data from 37 genetically diverse P. insidiosum strains and four related species. The database contains 732,686 genes, grouped into 80,061 unique clusters and further divided into core and variable categories at genus, species, and genotype levels. A high-resolution phylogenomic relationship among P. insidiosum strains and other oomycetes was projected through hierarchical clustering and core gene analyses. 3156 P. insidiosum-specific genes were shared among all genotypes and may be responsible for causing disease in humans and animals. After comparing these species-specific genes to the MvirDB database, 112 had significant matches with 66 known virulence proteins, some of which might be involved in vascular occlusion, which is a pathological feature of pythiosis. The correlation of genotypes, geographic origins, and affected hosts of P. insidiosum suggests that clade-I strains are more specific to animals, while clade-II/III strains are more specific to humans. The clade-specific genes might link to host preference. In summary, Pins Gene Table v2.0 is a comprehensive genome database accessible to users with minimal bioinformatics experience for the analysis of P. insidiosum genomes.

The Role of Proline-Proline-Glutamic Acid (PPE) Proteins in Mycobacterium tuberculosis Virulence: Mechanistic Insights and Therapeutic Implications

The Role of Proline-Proline-Glutamic Acid (PPE) Proteins in Mycobacterium tuberculosis Virulence: Mechanistic Insights and Therapeutic Implications

Deciphering Campylobacter jejuni DsbA1 protein dynamics in the presence of anti-virulent compounds: a multi-pronged computer-aided approach

The current study aims to evaluate Asinex library compounds against Campylobacter jejuni DsbA1 protein, a thiol disulfide oxidoreductase enzyme that plays a major role in the oxidative folding of bacterial virulence proteins, making it a promising anti-viral drug target. By employing several techniques of computer-aided drug design, BDC25697459, BDD33601083, and BDC30129064 were identified with binding energy scores of −8.8 kcal/mol, −8.8 kcal/mol, and −8.3 kcal/mol, respectively. However, the control molecule, tetraethylene glycol, exhibited a binding energy score of −7.0 kcal/mol. The control, BDD33601083, and BDC30129064 were unveiled to bind the same co-crystallized binding site (pocket 1), while BDC25697459 interacted with a new binding pocket (pocket 2) adjacent to the control binding region. The molecular dynamics simulation showed that complexes exhibit stable dynamics without significant global or residue-level fluctuations. The average RMSD values were in the range of 2.07 Å–2.45 Å. Similarly, mean RMSF was recorded between 1.30 and 1.42 Å. The C. jejuni DsbA1 was also observed as compact in the presence of the compounds, showing a mean RoG value in the range of 16.42 Å–16.55 Å. In terms of MM/PBSA binding energy, the BDC30129064 complex was ranked top with −44.88 ± 4.14 kcal/mol, whereas the positive control molecule exhibited −22.22 ± 3.33 kcal/mol. From a pharmacokinetic perspective, the compounds are suitable candidates for clinical trial investigation. Preliminary computational analysis of these virtual hits indicates that these compounds have a low potential for ADME and toxicity-associated liabilities. In summary, the compounds displayed a high affinity for the C. jejuni DsbA1 protein, indicating potential efficacy that requires further investigation. Communicated by Ramaswamy H. Sarma

Revolutionizing the virulent protein Internalin a in Listeria monocytogenes and designing multi epitope-based vaccine via immunoinformatic approaches

ABSTRACT This study addresses the crucial protein Internalin A in the development of a possible vaccination against Listeria monocytogenes. A meticulous in silico strategy was used which included physiochemical analysis, epitope prediction, and molecular interaction studies. Computational techniques shed light on potential development of a vaccine candidate against Listeria, with a focus on the Internalin A protein along with its immune simulations. Insights into molecular interactions and physiochemical features aid in understanding of prospective Listeria vaccine candidates. Ultimately, the computational analysis in this study sets the groundwork for the creation of a possible vaccination against Listeria. The discovered epitopes and insights into molecular interactions are useful for experimental validation. In vitro and in vivo investigations should be conducted in the future to confirm the effectiveness of these anticipated epitopes. These findings advanced our understanding of vaccine design by providing insights into battling Listeria infections and perhaps guiding tactics for other diseases.

P05 Identifying novel drug targets of the ESKAPE pathogen Pseudomonas aeruginosa

Abstract The Gram-negative ESKAPE pathogen Pseudomonas aeruginosa is currently listed top of the priority one pathogens by the WHO that require urgent research and development of new antimicrobials. Currently, microbiologists and clinicians face an ever-growing challenge in treating MDR bacterial infections, for this, and other pathogens, using conventional antibiotics. With a limited number of antibiotics currently in clinical trials and a general stagnation in the discovery of promising antimicrobials, new drug targets are desperately needed, and recent investigations include targeting bacterial virulence factors. Targeting bacterial virulence factors should reduce the pathogenesis of an organism, slow its infection progression and prevent vital virulence processes such as biofilm formation as well as potentially increasing a strain’s susceptibility to antibiotics. However, given the incredible diversity of bacteria, virulence mechanisms vary massively, meaning that identifying a broad-spectrum virulence target presents a major challenge. This important point has brought researchers to a fascinating group of enzymes that are ubiquitous in nature, and possess a plethora of biological roles, including acting as virulence factors. The macrophage infectivity potentiators (Mips) belong to the ubiquitous FK506 binding protein (FKBPs) family of peptidyl-prolyl cis/trans isomerases (PPIases). FKBPs are found in all bacteria serving as general house-keeping proteins involved in protein folding and chaperoning. However, a small subset has been identified as virulence proteins. Mips have been characterized in several Gram-negative pathogens including Legionella pneumophila and Burkholderia pseudomallei and were shown to be essential for invasion of macrophages, both in vitro and in murine-infection models, and for other virulence determinants. Pseudomonas aeruginosa also possess three Mip virulence factors, PaMip1, PaMip2 and PaMip3, which are required for the full virulence of P. aeruginosa in in vivo and ex vivo models. Virulence can be addressed in a pharmacological manner using bacterial natural products.

Shigella virulence protein VirG is a broadly protective antigen and vaccine candidate

Diarrhea caused by Shigella has been associated with high morbidity and mortality in young children worldwide. There are no licensed vaccines, and those clinically advanced have restricted coverage as they elicit serotype-specific immunity while disease is caused by multiple circulating serotypes. Our group had previously reported a close association between serum antibodies to the Shigella virulence factor VirG (or IcsA) and clinical protection in infected individuals. VirG is highly conserved among Shigella strains and appealing as a broad-spectrum vaccine candidate. In this study, we investigated the immunogenicity and protective capacity of VirG as a subunit vaccine in mice. The surface-exposed alpha (α) domain of VirG (VirGα) was produced as a recombinant protein. This region has almost identical immune reactivity to full-length VirG. Administered intramuscularly with alum, VirGα elicited robust immune responses and high protective efficacy against S. flexneri 2a and S. sonnei. Almost complete protection was afforded by VirGα given intranasally with the E. coli double mutant heat-labile toxin (dmLT). VirGα-specific antibodies recognized VirG expressed on live Shigella, and blocked Shigella adhesion and invasion to human colonic cells. These results show for the first time that VirGα is a promising cross-protective vaccine candidate to prevent Shigella infection.

Proteins of the food-borne pathogen <i>Listeria monocytogenes</i> strain F2365 relevant to lethal acidic stress and during rapid inactivation

<abstract> <p><italic>Listeria monocytogenes</italic>, which causes human listeriosis after consumption of contaminated food, can adapt and survive under a wide range of physiological and chemical stresses. In this study, the overall proteomic response of the <italic>L. monocytogenes</italic> strain F2365—a strain with mutations limiting its ability to tolerate acidic conditions—to progressive non-thermal acidic inactivation was investigated. The challenge process was investigated in the early stationary growth phase where F2365 cultures were acidified (pH 3.0, HCl) at 5 min, 1 h, and 2 h, generating pH 4.8, pH 4.1, and pH 3.5, respectively, with protein abundance measured using iTRAQ. Approximately 73 proteins increased in abundance and 8 declined when acidic stress became non-growth-permissive (pH < 4.1) and inactivation accelerated to approximately 2 log units/h. The functional categories of responding proteins were broad but the proteins involved were specific in nature and did not include whole pathways. Many responses likely accentuate energy conservation and compensate vital metabolic processes. For example, further repression of FlaA, normally repressed under acidic stress, occurs accompanied by an increase in quinol oxidase subunit QoxA and glycerol kinase GlpK. Proteins maintaining cell wall integrity, such as Iap and CwlO, manifested the overall largest abundance increase trend. Virulence proteins were also induced, including InlA, InlC, Hyl, Mpl, PlcA, and PlcB, suggesting that acidification may have mimicked conditions inducing some host survival traits. The overall suite of proteins affected appears to be the "last ditch" responses to non-thermal inactivation above and beyond the standard protections afforded in the stationary-growth phase. The array of proteins found here may provide a deeper understanding of the physiological responses of this pathogen during non-thermal inactivation.</p> </abstract>