Search Results (31)

Streptococcus suis is an important zoonotic pathogen responsible for severe infections in pigs and humans. Its capacity to survive within phagocytic cells is considered a key virulence mechanism that contributes to dissemination and persistence in host tissues. This study employed comparative proteomic profiling to investigate intracellular adaptation of S. suis serotypes 2 (SS2) and 14 (SS14) during infection of human U937 macrophages. Five isolates originating from humans and pigs were analyzed using gel electrophoresis with liquid chromatography–tandem mass spectrometry (GeLC–MS/MS), revealing 118 differentially expressed proteins grouped into 11 functional categories. Translation-related proteins represented the largest group (48%), including upregulated ribosomal subunits (30S: S2, S5, S7, S8, S12, S15; 50S: L1, L5, L18, L22, L24, L33, L35) and translation factors such as GidA/TrmFO and RimP. Enrichment of carbohydrate metabolism and DNA replication proteins, including phosphoenolpyruvate carboxylase (PEP), UDP-N-acetylglucosamine pyrophosphorylase (GlmU), and ATP-dependent DNA helicase RuvB, indicated metabolic reprogramming and stress adaptation under intracellular conditions. Stress-response proteins such as molecular chaperone DnaK were also induced, supporting their multifunctional, “moonlighting” roles in virulence and host interaction. Comparative analysis showed that SS2 expressed a broader range of adaptive proteins than SS14, consistent with its higher virulence potential. These findings reveal conserved intracellular responses centered on translation, energy metabolism, and stress tolerance, which enable S. suis to survive within human macrophages. Integration of these intracellular proteomic signatures with previous exoproteomic, peptidomic, and network-based studies highlights translational and metabolic proteins—particularly DnaK, enolase, elongation factor EF-Tu, and GlmU—as multifunctional candidates linking survival and immunogenicity. This work establishes a comparative proteomic foundation for understanding S. suis intracellular adaptation and highlights potential targets for future vaccine or therapeutic development against this zoonotic pathogen. Full article

Valorization of agri-food residues has garnered significant interest for obtaining value-added compounds such as enzymes or bioactive molecules. Rice milling by-products, such as rice bran, have limited commercial value and may pose environmental challenges. Filamentous fungi are recognized for their ability to grow on residues and for their capacity to produce large amounts of metabolites and enzymes of industrial interest. Here, we used filamentous fungi to produce enzyme cocktails from rice bran, which, due to its polysaccharide composition, serves as an ideal substrate for the growth of fungi producing cellulases and xylanases. To this end, sixteen fungal strains were isolated from rice bran and identified at the species level. The species belonged to the genera Aspergillus, Penicillium, and Mucor. The Aspergillus species displayed the highest efficiency in cellulase and xylanase activities, especially A. niger var. phoenicis and A. amstelodami. A. terreus, A. tritici, and A. montevidensis stood out as xylanolytic isolates, while P. parvofructum exhibited good cellulase activity. A. niger var. phoenicis followed by A. terreus showed the highest specific enzymatic activities of α- and β-D-galactosidase, α-L-arabinofuranosidase, α- and β-D-glucosidase, and β-D-xylosidase. Additionally, proteomic analysis of A. terreus, A. niger var. phoenicis, and P. parvofructum exoproteomes revealed differences in enzyme production for rice bran degradation. A. niger var. phoenicis had the highest levels of xylanases and cellulases, while P. parvofructum excelled in proteases, starch-degrading enzymes, and antifungal proteins. Finally, two Penicillium isolates were notable as producers of up to three different antifungal proteins. Our results demonstrate that filamentous fungi can effectively valorize rice bran by producing enzyme cocktails of industrial interest, along with bioactive peptides, in a cost-efficient manner, aligning with the circular bio-economy framework. Full article

The basidiomycete Crucibulum laeve strain LE-BIN1700 (Agaricales, Nidulariaceae) is able to grow on agar media supplemented with individual components of lignocellulose such as lignin, cellulose, xylan, xyloglucan, arabinoxylan, starch and pectin, and also to effectively destroy and digest birch, alder and pine sawdust. C. laeve produces a unique repertoire of proteins for the saccharification of the plant biomass, including predominantly oxidative enzymes such as laccases (family AA1_1 CAZymes), GMC oxidoreductases (family AA3_2 CAZymes), FAD-oligosaccharide oxidase (family AA7 CAZymes) and lytic polysaccharide monooxygenases (family LPMO X325), as well as accompanying acetyl esterases and loosenine-like expansins. Metabolomic analysis revealed that, specifically, monosaccharides and carboxylic acids were the key low molecular metabolites in the C. laeve culture liquids in the experimental conditions. The proportion of monosaccharides and polyols in the total pool of identified compounds increased on the sawdust-containing media. Multiple copies of the family AA1_1, AA3_2, AA7 and LPMOs CAZyme genes, as well as eight genes encoding proteins of the YvrE superfamily (COG3386), which includes sugar lactone lactonases, were predicted in the C. laeve genome. According to metabolic pathway analysis, the litter saprotroph C. laeve can catabolize D-gluconic and D-galacturonic acids, and possibly other aldonic acids, which seems to confer certain ecological advantages. Full article

White rot fungi, especially representatives of the genus Trametes spp. (Polyporaceae), are effective destructors of various xenobiotics, including oestrogens (phenol-like steroids), which are now widespread in the environment and pose a serious threat to the health of humans, animals and aquatic organisms. In this work, the ability of the white rot fungus Trametes hirsuta LE-BIN 072 to transform oestrone (E1) and 17β-oestradiol (E2), the main endocrine disruptors, was shown. More than 90% of the initial E1 and E2 were removed by the fungus during the first 24 h of transformation. The transformation process proceeded predominantly in the direction of the initial substrates’ detoxification, with the radical oxidative coupling of E1 and E2 as well as their metabolites and the formation of less toxic dimers in various combinations. A number of minor metabolites, in particular, less toxic estriol (E3), were identified by HPLC-MS. The formation of E1 from E2 and vice versa were shown. The exoproteome of the white rot fungus during the transformation of oestrogens was studied in detail for the first time. The contribution of ligninolytic peroxidases (MnP5, MnP7 and VP2) to the process of the extracellular detoxification of oestrogens and their possible metabolites is highlighted. Thus, the studied strain appears to be a promising mycodetoxicant of phenol-like steroids in aquatic environments. Full article

Within the genus Corynebacterium, six species are potential carriers of the tox gene, which encodes the highly potent diphtheria exotoxin: Corynebacterium diphtheriae, Corynebacterium belfantii, Corynebacterium rouxii, Corynebacterium ulcerans, Corynebacterium pseudotuberculosis and Corynebacterium silvaticum. Based on their potential to infect different host species and cause either human infections, zoonotic diseases or infections of economically important animals, these bacteria are of high scientific and economic interest and different research groups have carried out proteome analyses. These showed that especially the combination of MS-based proteomics with bioinformatic tools helped significantly to elucidate the functional aspects of corynebacterial genomes and to handle the genome and proteome complexity. The combination of proteomic and bioinformatic approaches was also used to discover new vaccine and drug targets. In addition, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry has been established as a fast and precise tool for the identification of these bacteria. Full article

Being an abundant renewable source of aromatic compounds, lignin is an important component of future bio-based economy. Currently, biotechnological processing of lignin through low molecular weight compounds is one of the conceptually promising ways for its valorization. To obtain lignin fragments suitable for further inclusion into microbial metabolism, it is proposed to use a ligninolytic system of white-rot fungi, which mainly comprises laccases and peroxidases. However, laccase and peroxidase genes are almost always represented by many non-allelic copies that form multigene families within the genome of white-rot fungi, and the contributions of exact family members to the overall process of lignin degradation has not yet been determined. In this article, the response of the Trametes hirsuta LE-BIN 072 ligninolytic system to the presence of various monolignol-related phenolic compounds (veratryl alcohol, p-coumaric acid, vanillic acid, and syringic acid) in culture media was monitored at the level of gene transcription and protein secretion. By showing which isozymes contribute to the overall functioning of the ligninolytic system of the T. hirsuta LE-BIN 072, the data obtained in this study will greatly contribute to the possible application of this fungus and its ligninolytic enzymes in lignin depolymerization processes. Full article

Leishmaniasis, a vector-borne parasitic protozoan disease, is among the most important neglected tropical diseases. In the absence of vaccines, disease management is challenging. The available chemotherapy is suboptimal, and there are growing concerns about the emergence of drug resistance. Thus, a better understanding of parasite biology is essential to generate new strategies for disease control. In this context, in vitro parasite exoproteome characterization enabled the identification of proteins involved in parasite survival, pathogenesis, and other biologically relevant processes. After 2005, with the availability of genomic information, these studies became increasingly feasible and revealed the true complexity of the parasite exoproteome. After the discovery of Leishmania extracellular vesicles (EVs), most exoproteome studies shifted to the characterization of EVs. The non-EV portion of the exoproteome, named the vesicle-depleted exoproteome (VDE), has been mostly ignored even if it accounts for a significant portion of the total exoproteome proteins. Herein, we summarize the importance of total exoproteome studies followed by a special emphasis on the available information and the biological relevance of the VDE. Finally, we report on how VDE can be studied and disclose how it might contribute to providing biologically relevant targets for diagnosis, drug, and vaccine development. Full article

Diazotrophic bacteria isolated from the rhizosphere of a wild wheat ancestor, grown from its refuge area in the Fertile Crescent, were found to be efficient Plant Growth-Promoting Rhizobacteria (PGPR), upon interaction with an elite wheat cultivar. In nitrogen-starved plants, they increased the amount of nitrogen in the seed crop (per plant) by about twofold. A bacterial growth medium was developed to investigate the effects of bacterial exudates on root development in the elite cultivar, and to analyze the exo-metabolomes and exo-proteomes. Altered root development was observed, with distinct responses depending on the strain, for instance, with respect to root hair development. A first conclusion from these results is that the ability of wheat to establish effective beneficial interactions with PGPRs does not appear to have undergone systematic deep reprogramming during domestication. Exo-metabolome analysis revealed a complex set of secondary metabolites, including nutrient ion chelators, cyclopeptides that could act as phytohormone mimetics, and quorum sensing molecules having inter-kingdom signaling properties. The exo-proteome-comprised strain-specific enzymes, and structural proteins belonging to outer-membrane vesicles, are likely to sequester metabolites in their lumen. Thus, the methodological processes we have developed to collect and analyze bacterial exudates have revealed that PGPRs constitutively exude a highly complex set of metabolites; this is likely to allow numerous mechanisms to simultaneously contribute to plant growth promotion, and thereby to also broaden the spectra of plant genotypes (species and accessions/cultivars) with which beneficial interactions can occur. Full article

Histomonas meleagridis, a poultry-specific intestinal protozoan parasite, is histomonosis’s etiological agent. Since treatment or prophylaxis options are no longer available in various countries, histomonosis can lead to significant production losses in chickens and mortality in turkeys. The surfaceome of microbial pathogens is a crucial component of host–pathogen interactions. Recent proteome and exoproteome studies on H. meleagridis produced molecular data associated with virulence and in vitro attenuation, yet the information on proteins exposed on the cell surface is currently unknown. Thus, in the present study, we identified 1485 proteins and quantified 22 and 45 upregulated proteins in the virulent and attenuated strains, respectively, by applying cell surface biotinylation in association with high-throughput proteomic analysis. The virulent strain displayed upregulated proteins that could be linked to putative virulence factors involved in the colonization and establishment of infection, with the upregulation of two candidates being confirmed by expression analysis. In the attenuated strain, structural, transport and energy production proteins were upregulated, supporting the protozoan’s adaptation to the in vitro environment. These results provide a better understanding of the surface molecules involved in the pathogenesis of histomonosis, while highlighting the pathogen’s in vitro adaptation processes. Full article

White-rot basidiomycete fungi are a unique group of organisms that evolved an unprecedented arsenal of extracellular enzymes for an efficient degradation of all components of wood such as cellulose, hemicelluloses and lignin. The exoproteomes of white-rot fungi represent a natural enzymatic toolbox for white biotechnology. Currently, only exoproteomes of a narrow taxonomic group of white-rot fungi—fungi belonging to the Polyporales order—are extensively studied. In this article, two white-rot fungi, Peniophora lycii LE-BIN 2142 from the Russulales order and Trametes hirsuta LE-BIN 072 from the Polyporales order, were compared and contrasted in terms of their enzymatic machinery used for degradation of different types of wood substrates—alder, birch and pine sawdust. Our findings suggested that the studied fungi use extremely different enzymatic systems for the degradation of carbohydrates and lignin. While T. hirsuta LE-BIN 072 behaved as a typical white-rot fungus, P. lycii LE-BIN 2142 demonstrated substantial peculiarities. Instead of using cellulolytic and hemicellulolytic hydrolytic enzymes, P. lycii LE-BIN 2142 primarily relies on oxidative polysaccharide-degrading enzymes such as LPMO and GMC oxidoreductase. Moreover, exoproteomes of P. lycii LE-BIN 2142 completely lacked ligninolytic peroxidases, a well-known marker of white-rot fungi, but instead contained several laccase isozymes and previously uncharacterized FAD-binding domain-containing proteins. Full article

The fungal cell wall occupies a central place in the interaction between fungi and their environment. This study focuses on the role of the putative polysaccharide synthase Cps1 in the physiology, development and virulence of the grey mold-causing agent Botrytis cinerea. Deletion of the Bccps1 gene does not affect the germination of the conidia (asexual spores) or the early mycelial development, but it perturbs hyphal expansion after 24 h, revealing a two-phase hyphal development that has not been reported so far. It causes a severe reduction of mycelial growth in a solid medium and modifies hyphal aggregation into pellets in liquid cultures. It strongly impairs plant penetration, plant colonization and the formation of sclerotia (survival structures). Loss of the BcCps1 protein associates with a decrease in glucans and glycoproteins in the fungus cell wall and the up-accumulation of 132 proteins in the mutant’s exoproteome, among which are fungal cell wall enzymes. This is accompanied by an increased fragility of the mutant mycelium, an increased sensitivity to some environmental stresses and a reduced adhesion to plant surface. Taken together, the results support a significant role of Cps1 in the cell wall biology of B. cinerea. Full article

Bacterial extracellular proteins participate in the host cell communication by virtue of the modulation of pathogenicity, commensalism and mutualism. Studies on the microbiome of cervical mucus of the water buffalo (Bubalus bubalis) have shown the occurrence of Staphylococcus pasteuri and that the presence of this bacterium is indicative of various physiological and reproductive states in the host. Recently, S. pasteuri has been isolated from the cervical mucus of the buffalo during the different phases of estrous cycle, and has proved to be much more pronounced during the estrus phase. The basis underlying the availability of a significantly increased S. pasteuri population, specifically during the estrus phase, is not known. Consequently, it is important to determine the significance of the specific abundance of S. pasteuri during the estrus phase of the buffalo host, particularly from the perspective of whether this bacterial species is capable of contributing to sexual communication via its extracellular proteins and volatiles. Therefore, the relevance of S. pasteuri exoproteome in the buffalo cervical mucus during the estrus phase was analyzed using LC-MS/MS. As many as 219 proteins were identified, among which elongation factor Tu (EF-Tu), 60-kDa chaperonin (Cpn60), enolase, fructose-bisphosphate aldolase class 1 (FBP aldolase), enoyl-[acyl-carrier-protein] reductase [NADPH] (ENR) and lipoprotein (Lpp) were the functionally important candidates. Most of the proteins present in the exoproteome of S. pasteuri were those involved in cellular–metabolic functions, as well as catalytic- and binding activities. Moreover, computational studies of Lpp have shown enhanced interaction with volatiles such as acetic-, butanoic-, isovaleric- and valeric acids, which were identified in the cervical mucus S. pasteuri culture supernatant. The present findings suggest that S. pasteuri extracellular proteins may play an important role in buffalo sexual communication during the estrus phase. Full article

The expansion of multiple drug resistant (MDR) strains of Klebsiella pneumoniae presents an immense threat for public health. Annually, this microorganism causes thousands of lethal nosocomial infections worldwide. Currently, it has been shown that certain strains of lactic acid bacteria (LAB) can efficiently inhibit growth of K. pneumoniae and the formation of its biofilms; however, the active principle of such action remains unknown. In the current article, the growth inhibition of MDR K. pneumoniae by two LAB—Limosilactobacillus reuteri LR1 and Lacticaseibacillus rhamnosus F—is demonstrated, and the nature of this inhibition studied at the level of exoproteome. This article shows that the exoproteomes of studied LAB contains both classically and non-classically secreted proteins. While for L. reuteri LR1 the substantial portion of classically secreted proteins was presented by cell-wall-degrading enzymes, for L. rhamnosus F only one out of four classically secreted proteins was presented by cell-wall hydrolase. Non-classically secreted proteins of both LAB were primarily metabolic enzymes, for some of which a possible moonlighting functioning was proposed. These results contribute to knowledge regarding antagonistic interaction between LAB and pathogenic and opportunistic microorganisms and set new perspectives for the use of LAB to control the spread of these microorganisms. Full article

Lactobacillus sp. have long been studied for their great potential in probiotic applications. Recently, proteomics analysis has become a useful tool for studies on potential lactobacilli probiotics. Specifically, proteomics has helped determine and describe the physiological changes that lactic acid bacteria undergo in specific conditions, especially in the host gut. In particular, the extracellular proteome, or exoproteome, of lactobacilli contains proteins specific to host– or environment–microbe interactions. Using gel-free, label-free ultra-high performance liquid chromatography tandem mass spectrometry, we explored the exoproteome of the probiotic candidate Lactobacillus mucosae LM1 subjected to bile treatment, to determine the proteins it may use against bile stress in the gut. Bile stress increased the size of the LM1 exoproteome, secreting ribosomal proteins (50S ribosomal protein L27 and L16) and metabolic proteins (lactate dehydrogenase, phosphoglycerate kinase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate dehydrogenases, among others) that might have moonlighting functions in the LM1 bile stress response. Interestingly, membrane-associated proteins (transporters, peptidase, ligase and cell division protein ftsH) were among the key proteins whose secretion were induced by the LM1 bile stress response. These specific proteins from LM1 exoproteome will be useful in observing the proposed bile response mechanisms via in vitro experiments. Our data also reveal the possible beneficial effects of LM1 to the host gut. Full article

The swine pathogen Streptococcus suis is a Gram-positive bacterium which causes infections in pigs, with an impact in animal health and in the livestock industry, and it is also an important zoonotic agent. During the infection process, surface and secreted proteins are essential in the interaction between microorganisms and their hosts. Here, we report a comparative proteomic analysis of the proteins released to the extracellular milieu in six human clinical isolates belonging to the highly prevalent and virulent serotype 2. The total secreted content was precipitated and analyzed by GeLC-MS/MS. In the six strains, 144 proteins assigned to each of the categories of extracellular or surface proteins were identified, as well as 680 predicted cytoplasmic proteins, many of which are putative moonlighting proteins. Of the nine predicted signal peptide-I secreted proteins, seven had relevant antigenic potential when they were analyzed through bioinformatic analysis. This is the first work comparing the exoproteome fraction of several human isolates of this important pathogen. Full article