Andoni Ramirez-Garcia

The rise of antimicrobial resistance is reducing the effectiveness of current therapies and highlights the need for new antimicrobial agents. Marine environments, rich in microorganisms adapted to extreme conditions, represent a promising reservoir of bioactive compounds. The marine bacterium Marinomonas mediterranea secretes, among others, lysine oxidase (LodA) and glycine oxidase (GoxA), enzymes that generate hydrogen peroxide and exhibit antibacterial activity. Based on this mechanism, we hypothesized that modifying specific experimental con-ditions could enhance the antimicrobial potential of the secreted enzymes and additional com-pounds, broadening their activity spectrum. In this study, culture conditions were optimized to promote the production of inhibitory molecules, and the antimicrobial activity of the M. medi-terranea secretome was assessed against clinically relevant microorganisms. The concentrated secretome obtained from growth in Marine Broth exhibited antimicrobial activity against the Gram-negative and Gram-positive bacteria analyzed and several pathogenic fungi, an effect at-tributable to some proteins present in it. Among them, LodA and GoxA, previously associated with bacterial growth inhibition, also appear to contribute to antifungal activity. Moreover, valyl-tRNA synthetase emerged as a potential new antimicrobial factor. These findings high-light the biotechnological potential of marine microorganisms and support further exploration of their secretomes for the development of new antimicrobial agents.

The Granulin motif is an ancient extracellular growth determinant in eukaryotes but not in fungi. The motif consists of 12 cysteines that give the protein a unique tertiary structure of four stacked β-hairpins. In humans, this domain is related to survival, growth modulation, migration, inflammation, wound repair, and diseases like frontotemporal dementia, Alzheimer's disease, and cancer. The protein encoded by the gene Afu6g07200 of Aspergillus fumigatus shows a domain with predicted granulin structure and is overexpressed during spore germination. Despite low conservation of its amino acid sequence, this domain exhibits the same predicted 3D structure as several human granulins, with the highest confidence (95.4%) being observed with human GRNA. Moreover, this protein is conserved among the Pezizomycotina subdivision. In this study, we demonstrate that this A. fumigatus protein is involved in proliferation, polarization, cell wall integrity, cytoskeletal organization, stress response, and sensing. Fungal granulin is implicated in maintaining the periplasmic space, is related to polarity markers and actin, and plays an essential role in activating MAPK cascades. Furthermore, complementation with human GRNA fulfilled the activities of the fungal protein at morphological and molecular levels. Therefore, this is the first description of a granulin domain in filamentous fungi based on 3D structure and function homology.

Candidozyma auris (Candida auris) is an emerging pathogenic yeast of global concern due to its persistence on abiotic and biotic surfaces and the difficulty of treating the severe infections it causes, which are frequently associated with high mortality rates because of its extensive antifungal resistance. Thus, new therapeutic strategies are urgently needed to complement or replace current antifungal drugs. In this study, we evaluated the efficacy of Ca37, a monoclonal antibody (mAb) targeting the alcohol dehydrogenase (Adh) protein of Candida albicans, against C. auris both in vitro and in vivo. Protein electrophoresis and Western Blot analyses demonstrated immunoreactivity of Ca37 mAb with C. auris total protein and cell wall-associated protein extracts, among which Adh was identified. In vitro, incubation with Ca37 mAb significantly reduced the growth of several C. auris strains and enhanced the phagocytic activity of RAW 264.7 murine macrophages. In vivo, Ca37 mAb treatment increased the survival of Galleria mellonella larvae. In a murine model of systemic infection, treated mice displayed improved clinical condition, along with a greater number and larger area of immune-associated foci in the kidneys, suggesting enhanced fungal recognition. These findings support the potential of Ca37 mAb as an antifungal immunotherapy, although further studies in murine models are necessary to establish optimal dosing, efficacy, and mechanisms of action.

Airborne fungi are ubiquitous microorganisms in the environment, and some of them are known opportunistic pathogens. In recent years, azole resistance, which can have a clinical or environmental origin, has become a critical issue. Four environmental samplings were performed to assess the prevalence and diversity of potentially azole‐resistant fungi in three areas (hospital surrounding, rural and urban) from the Basque Country. The microbial concentration varied from 40 to 3670 CFU/m ³ , depending on the location and sampling. The CFU/m ³ count on plates incubated with voriconazole at 37°C was only three times lower than that of plates incubated without the antifungal, suggesting many 37°C‐growing fungi might be triazole‐resistant. Three hundred and twenty one potentially resistant isolates were identified, belonging to 21 genera and 55 species. Alternaria (62.31%) and Talaromyces (18.69%) were the predominant genera, with Alternaria infectoria (47.19%) being the most abundant species. Overall, the two coastal provinces (Bizkaia and Gipuzkoa) showed the most similarity. The rural area exhibited the highest alpha diversity values for each province, whereas samples from this area were more alike in terms of beta diversity. PCoA analysis indicated that sampling points were clustered by samplings or provinces. This study provides the first characterisation of the outdoor environment from the Basque Country and highlights the importance of determining the prevalence of potentially azole‐resistant isolates.

The human body is colonized by diverse microorganisms, with bacteria being the most extensively studied. However, fungi, collectively known as “the mycobiota,” are increasingly recognized as integral components of the microbiota, inhabiting nearly all mucosal surfaces. Commensal fungi influence host immunity similarly to bacteria and contribute to other essential functions, including metabolism. This emerging understanding positions fungi as potential biomarkers for the diagnosis and prognosis of various diseases. In this review, we explore the dual roles of fungi as both commensals and pathogens, and the potential of antifungal antibodies to serve as diagnostic and prognostic tools, especially in chronic immune-inflammatory non-communicable diseases, including inflammatory bowel disease, rheumatoid arthritis, multiple sclerosis, and neurodegenerative disorders. Finally, we address current challenges and outline future perspectives for leveraging fungal biomarkers in clinical practice.

Recent studies have increasingly focused on the role of fungi, including Candida albicans , in carcinogenesis. Since C. albicans is a component of the human microbiota, particularly on the skin, we investigated its effect on the phenotype and signalling pathways of melanoma cells. Assays for migration, adhesion, angiogenesis, and hepatic metastasis showed that C. albicans promotes a more malignant phenotype in melanoma cells. At the transcriptomic level, C. albicans increased the expression of VEGF ( Vegfa ), and genes associated with MAPK and HIF-1 signalling pathways, and with aerobic glycolysis. Further in vitro analysis revealed that TLRs and EphA2 receptors are involved in the recognition of live C. albicans , stimulating VEGF secretion and expression of the AP-1 transcription factor component c-Fos through p38-MAPK and HIF-1α. These pathways also regulate the expression of other AP-1 constituents such as Atf3 , Jun , and Jund . Moreover, p38-MAPK regulates glycolytic genes like Hk2 , Slc2a1 , and Eno2 . In conclusion, C. albicans activates the p38-MAPK/c-Fos/AP-1 and HIF-1/HIF-1α/c-Fos/AP-1 pathways in melanoma cells, promoting a pro-angiogenic environment and metabolic reprogramming. Therefore, this study clarifies the impact of C. albicans on melanoma cells, which can lead to the use of antifungal therapies as complementary to traditional treatments for melanoma.

The difficulty of accurately identifying Candida auris and the high resistance rates presented have increased the concern in the healthcare setting. Due to this, the aim of this study was to analyse the fungal response to oxidative stress. To achieve this goal, gene and protein expression were examined using qPCR and two-dimensional electrophoresis, respectively, peroxiredoxin Tsa1b being discovered to be overexpressed under oxidative stress. Besides, its antigenicity was also confirmed by western blotting. Subsequently, the significance of Tsa1b was next investigated by creating and characterizing the C. auris ΔTSA1B and C. auris ΔTSA1B::TSA1B strains using CRISPR-Cas9. The findings demonstrated that the ΔTSA1B strain was more susceptible to oxidative and cell wall stressors than the wild-type strain, which was consistent with an increase in the cell wall β-glucan amounts when grown in the presence of oxidative stress. Furthermore, the ΔTSA1B strain was also more vulnerable to the presence of dendritic cells and bone marrow-derived macrophages. Finally, in vivo infections performed in Galleria mellonella and mice showed a slower progression of the disease in those animals infected with the mutant strain. In conclusion, the peroxiredoxin Tsa1b has been identified as an important protein for the C. auris response to oxidative stress and as a virulence factor, allowing for a more thorough knowledge of the pathobiology of this yeast. This study points out the potential that this protein may have for the development of new diagnostic and therapeutic approaches. HIGHLIGHTS: Several metabolic proteins are implicated in C. auris response to oxidative stress C. auris response to oxidative stress is influenced by the Tsa1b peroxiredoxin Lack of Tsa1b generates more susceptibility to stresses and an altered cell wall C. auris Tsa1b is involved in the fungal interaction with host immune cells The Tsa1b of C. auris contributes to the progression of the infection in vivo

In inflammatory bowel diseases (IBDs), such as Crohn’s disease (CD) and ulcerative colitis (UC), the immune system relentlessly attacks intestinal cells, causing recurrent tissue damage over the lifetime of patients. The etiology of IBD is complex and multifactorial, involving environmental, microbiota, genetic, and immunological factors that alter the molecular basis of the organism. Among these, the microbiota and immune cells play pivotal roles; the microbiota generates antigens recognized by immune cells and antibodies, while autoantibodies target and attack the intestinal membrane, exacerbating inflammation and tissue damage. Given the altered molecular framework, the analysis of multiple molecular biomarkers in patients proves exceedingly valuable for diagnosing and prognosing IBD, including markers like C reactive protein and fecal calprotectin. Upon detection and classification of patients, specific treatments are administered, ranging from conventional drugs to new biological therapies, such as antibodies to neutralize inflammatory molecules like tumor necrosis factor (TNF) and integrin. This review delves into the molecular basis and targets, biomarkers, treatment options, monitoring techniques, and, ultimately, current challenges in IBD management.

Immune disorders arise from complex genetic and environmental factors, which lead to dysregulation at the cellular and inflammatory levels and cause tissue damage. Recent research highlights the crucial role of reactive antibodies in autoimmune diseases and graft rejection, but their complex determination poses challenges for clinical use. Therefore, our study aimed to ascertain whether the presence of reactive antibodies against membrane antigens in tissues from both animal models and humans could serve as biomarkers in patients with autoimmune disorders. To address this issue, we examined the binding profile of serological antibodies against a diverse panel of cell membranes from the spleen, liver, and kidney tissues of monkeys, rats, and humans. After developing the cell membrane microarrays, human sera were immunologically assayed. The study was first conducted on sera from two groups, healthy subjects and patients with inflammatory and autoimmune disorders, and then optimized for kidney transplant patient sera. A significant increase in antibody reactivity against specific monkey kidney and spleen membranes was observed in the serum of patients with lupus nephritis, while kidney transplant patients showed a significant enhancement against human tissues and human embryonic kidney 293 cells. These results show the potential importance for clinical and basic research purposes of studying the presence of specific IgG against membrane antigens in patients’ serum as potential biomarkers of immune disorders. However, it is important to note that these results need to be verified in further studies with a larger sample size to confirm their relevance.