Bacterial extracellular vesicles (EVs) perform various biological functions, including those that are critical to microbes. Determination of EVs composition allows for a deep understanding of their role in the bacterial community and communication among them. Cutibacterium acnes, formerly Propionibacteriumacnes, are commensal bacteria responsible for various infections, e.g., prosthesis, sarcoidosis, soft-tissue infections, and the most known but still controversial—acnes lesion. In C. acnes, three major phylotypes represented variable disease associations. Herein, for the first time, we present a comparative analysis of EVs obtained from three C. acnes phylotypes (IA1, IB, and II) to demonstrate the existence of differences in their protein and lipid composition. In the following work, the morphological analysis of EVs was performed, and the SDS-PAGE protein profile and the lipid profile were presented using the TLC and MALDI-TOF MS methods. This study allowed us to show major differences between the protein and lipid composition of C. acnes EVs. This is a clear indication that EVs released by different phylotypes of the one species are not identical to each other in terms of composition and should be separately analyzed each time to obtain reliable results.

Actinobacteria are abundant in soil and other environmental ecosystems and are also an important part of the human microbiota. Hence, they can also be detected in indoor environments and on building materials, where actinobacterial proliferation on damp materials can indicate moisture damage. The aim of this study was to evaluate the matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) for the identification of 28 environmental strains of Actinobacteria isolated from building materials and indoor and outdoor air samples, mainly collected in the context of moisture damage investigations in buildings in Finland. The 16S rRNA gene sequencing and chemotaxonomic analyses were performed, and results were compared with the MALDI-TOF MS Biotyper identification. Using 16S rRNA gene sequencing, all isolates were identified on the species or genus level and were representatives of Streptomyces, Nocardia, and Pseudonocardia genera. Based on MALDI-TOF MS analysis, initially, 11 isolates were identified as Streptomyces spp. and 1 as Nocardia carnea with a high identification score. After an upgrade in the MALDI-TOF MS in-house database and re-evaluation of mass spectra, 13 additional isolates were identified as Nocardia , Pseudonocardia, and Streptomyces . MALDI-TOF MS has the potential in environmental strain identification; however, the standard database needs to be considerably enriched by environmental Actinobacteria representatives. IMPORTANCE: The manuscript addresses the challenges in identifying environmental bacteria using matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) Biotyper-based protein profiling. The matter of the studies—actinobacterial strains—has been isolated mostly from building materials that originated from a confirmed moisture-damaged situation. Polyphasic taxonomy, 16S RNA gene sequencing, and MALDI-TOF mass spectrometry were applied for identification purposes. In this experimental paper, a few important facts are highlighted. First, Actinobacteria are abundant in the natural as well as built environment, and their identification on the species and genus levels is difficult and time-consuming. Second, MALDI-TOF MS is an effective tool for identifying bacterial environmental strains, and in parallel, continuous enrichment of the proteomics mass spectral databases is necessary for proper identification. Third, the chemical approach aids in the taxonomical inquiry of Actinobacteria environmental strains.

Cutibacterium are part of the human skin microbiota and are opportunistic microorganisms that become pathogenic in immunodeficient states. These lipophilic bacteria willingly inhabit areas of the skin where sebaceous glands are abundant; hence, there is a need to thoroughly understand their metabolism. Lipids are no longer considered only structural elements but also serve as signaling molecules and may have antigenic properties. Lipidomics remains a major research challenge, mainly due to the diverse physicochemical properties of lipids. Therefore, this study aimed to perform a large comparative lipidomic analysis of eight representatives of the Cutibacterium genus, including four phylotypes of C. acnes and two strains of C. granulosum , C. avidum , and C. namnetense . Lipidomic analysis was performed by liquid chromatography‒mass spectrometry (LC-MS) in both positive and negative ion modes, allowing the detection of the widest range of metabolites. Fatty acid analysis by gas chromatography‒mass spectrometry (GC-MS) corroborated the lipidomic data. As a result, 128 lipids were identified, among which it was possible to select marker compounds, some of which were characteristic even of individual C. acnes phylotypes. These include phosphatidylcholine PC 30:0, sphingomyelins (SM 33:1, SM 35:1), and phosphatidylglycerol with an alkyl ether substituent PG O-32:0. Moreover, cardiolipins and fatty acid amides were identified in Cutibacterium spp. for the first time. This comparative characterization of the cutibacterial lipidome with the search for specific molecular markers reveals its diagnostic potential for clinical microbiology. IMPORTANCE: Cutibacterium (previously Propionibacterium ) represents an important part of the human skin microbiota, and its role in clinical microbiology is growing due to opportunistic infections. Lipidomics, apart from protein profiling, has the potential to prove to be a useful tool for defining the cellular fingerprint, allowing for precise differentiation of microorganisms. In this work, we presented a comparative analysis of lipids found in eight strains of the genus Cutibacterium , including a few C. acnes phylotypes. Our results are one of the first large-scale comprehensive studies regarding the bacterial lipidome, which also enabled the selection of C. acnes phylotype-specific lipid markers. The increased role of lipids not only as structural components but also as diagnostic markers or potential antigens has led to new lipid markers that can be used as diagnostic tools for clinical microbiology. We believe that the findings in our paper will appeal to a wide range of researchers.

Cutaneous tuberculosis (CTB) is a very rare disease and accounts for only 1–2% of cases of extrapulmonary tuberculosis (EPTB). Due to the variety of its clinical manifestations, the uncharacteristic appearance of its lesions, resembling other dermatoses in the early stages, and the limited experience of clinicians due to the rarity of CTB, diagnosis is very difficult. Particularly noteworthy is the fact that most cases of EPTB, including skin tuberculosis (TB), can be a manifestation of systemic involvement. In this paper, we present a case of an immunocompromised patient who was diagnosed with CTB almost a year after the first dermatological lesions were located on the lower extremities. At the same time, due to respiratory symptoms, a diagnosis of pulmonary TB (PTB) was made, and radiological and microbiological confirmations were obtained.

Cutibacterium acnes represents one of the predominant bacterial taxa colonizing human skin. Its impact on host physiology is strongly phylotype-dependent: certain phylotypes are associated with the initiation and progression of acnes lesions, whereas others contribute to the stability and homeostasis of the cutaneous microbiota. Moreover, phylotype II C. acnes has been increasingly implicated in the etiology of chronic prostatitis, suggesting a broader pathogenic potential. The growing incidence of antimicrobial resistance in C. acnes underscores the need to identify novel antigenic determinants and molecular candidates for next-generation vaccine strategies. Notably, C. acnes , like many Gram-positive bacteria, produces extracellular vesicles (EVs) that function as carriers of enzymes, metabolites, immunogenic components and lipid structures. In the present study, four glycolipids (GL1–GL4) isolated from distinct C. acnes phylotypes were subjected to comprehensive structural characterization using GLC-MS, MALDI-TOF MS and NMR spectroscopy. These GL1-GL4 compounds are glycoglycerolipids composed of one or two glucose residues, a glycerol moiety and different numbers of fatty acid residues (mainly 15:0–18:0). The presence of GL1 within EVs fractions obtained from four C. acnes phylotypes has been confirmed. Subsequently, the effects of EVs and the glycolipids GL1 and GL3 on the viability of human prostate epithelial and fibroblast cell lines were assessed. None of the tested preparations exhibited significant cytotoxicity. However, EVs derived from phylotypes IA1 and II elicited markedly elevated release of IL-6 and TNF-α, demonstrating their substantial pro-inflammatory capacity and supporting their potential role in host-pathogen interactions.