The virulence of bacterial outer membrane vesicles (OMVs) contributes to innate microbial defense. Limited data report their role in interspecies reactions. There are no data about the relevance of OMVs in bacterial-yeast communication. We hypothesized that model Moraxella catarrhalis OMVs may orchestrate the susceptibility of pathogenic bacteria and yeasts to cationic peptides (polymyxin B) and serum complement. Using growth kinetic curve and time-kill assay we found that OMVs protect Candida albicans against polymyxin B-dependent fungicidal action in combination with fluconazole. We showed that OMVs preserve the virulent filamentous phenotype of yeasts in the presence of both antifungal drugs. We demonstrated that bacteria including Haemophilus influenza, Acinetobacter baumannii, and Pseudomonas aeruginosa coincubated with OMVs are protected against membrane targeting agents. The high susceptibility of OMV-associated bacteria to polymyxin B excluded the direct way of protection, suggesting rather the fusion mechanisms. High-performance liquid chromatography-ultraviolet spectroscopy (HPLC-UV) and zeta-potential measurement revealed a high sequestration capacity (up to 95%) of OMVs against model cationic peptide accompanied by an increase in surface electrical charge. We presented the first experimental evidence that bacterial OMVs by sequestering of cationic peptides may protect pathogenic yeast against combined action of antifungal drugs. Our findings identify OMVs as important inter-kingdom players.

The Annotated Germs for Automated Recognition (AGAR) dataset is an image database of microbial colonies cultured on agar plates. It contains 18 000 photos of five different microorganisms as single or mixed cultures, taken under diverse lighting conditions with two different cameras. All the images are classified into countable, uncountable, and empty, with the countable class labeled by microbiologists with colony location and species identification (336 442 colonies in total). This study describes the dataset itself and the process of its development. In the second part, the performance of selected deep neural network architectures for object detection, namely Faster R-CNN and Cascade R-CNN, was evaluated on the AGAR dataset. The results confirmed the great potential of deep learning methods to automate the process of microbe localization and classification based on Petri dish photos. Moreover, AGAR is the first publicly available dataset of this kind and size and will facilitate the future development of machine learning models. The data used in these studies can be found at https://agar.neurosys.com/.

Moraxella catarrhalis is a common human respiratory tract pathogen. Its virulence factors associated with whole bacteria or outer membrane vesicles (OMVs) aid infection, colonization and may induce specific antibodies. To investigate pathogen-host interactions, we applied integrated bioinformatic and immunoproteomic (2D-electrophoresis, immunoblotting, LC-MS/MS) approaches. We showed that OMV proteins engaged exclusively in complement evasion and colonization strategies, but not those involved in iron transport and metabolism, are major targets for cross-reacting antibodies produced against phylogenetically divergent M. catarrhalis strains. The analysis of 31 complete genomes of M. catarrhalis and other Moraxella revealed that OMV protein-coding genes belong to 64 orthologous groups, five of which are restricted to M. catarrhalis. This species showed a two-fold increase in the number of OMV protein-coding genes relative to its ancestors and animal-pathogenic Moraxella. The appearance of specific OMV factors and the increase in OMV-associated virulence proteins during M. catarrhalis evolution is an interesting example of pathogen adaptation to optimize colonization. This precisely targeted cross-reactive immunity against M. catarrhalis may be an important strategy of host defences to counteract this phenomenon. We demonstrate that cross-reactivity is closely associated with the anti-virulent antibody repertoire which we have linked with adaptation of this pathogen to the host.

Neuropeptides such as substance P (SP) and calcitonin gene-related peptide (CGRP) play both pro- and anti-inflammatory activities and are produced during infection and inflammation. Moraxella catarrhalis is one of the leading infectious agents responsible for inflammatory exacerbation in chronic obstructive pulmonary disease (COPD). Since the airway inflammation in COPD is connected with activation of both epithelial cells and accumulated neutrophils, in this study we determined the in vitro effects of neuropeptides on the inflammatory potential of these cells in response to M. catarrhalis outer membrane vesicle (OMV) stimulant. The various OMV-mediated proinflammatory effects were demonstrated. Next, using hBD-2-pGL4[luc2] plasmid with luciferase reporter gene, SP and CGRP were shown to inhibit the IL-1β-dependent expression of potent neutrophil chemoattractant, hBD-2 defensin, in transfected A549 epithelial cells (type II alveolar cells) upon OMV stimulation. Both neuropeptides exerted antiapoptotic activity through rescuing a significant fraction of A549 cells from OMV-induced cell death and apoptosis. Finally, CGRP caused an impairment of specific but not azurophilic granule exocytosis from neutrophils as shown by evaluation of gelatinase-associated lipocalin (NGAL) or CD66b expression and elastase release, respectively. Concluding, these findings suggest that SP and CGRP mediate the dampening of proinflammatory action triggered by M. catarrhalis OMVs towards cells engaged in lung inflammation in vitro.

Understanding how immunity to pathogens develops is crucial for progress in the quest for effective vaccines. Intraspecies and interspecies cross-reacting antibodies are produced in high frequency against immune-relevant and shared microbial epitopes. It has been confirmed that cross-reactive antigens may have a crucial role in natural epidemiology to a particular infection and that cross-protection may influence the outcome of natural infections. On the other hand, the action of cross-reactive antibodies may be very harmful for the host. In this review we discuss both the defensive and offensive capabilities of cross-reactive antibodies. The defensive properties are discussed with regard to the beneficial cross-protective interaction of these antibodies against various microorganisms including viruses, bacteria, fungi and protozoan parasites. We summarize the current knowledge of numerous effector functions of these antibodies such as agglutination, neutralization of infectivity, complement activation, phagocytosis enhancement, and antibody-dependent cellular cytotoxicity. We also discuss the offensive action of cross-reactive antibodies including their detrimental effects in exacerbation of the infective diseases, as well as autoimmune diseases and allergy as a result of inappriopriate or deleterious inflammatory response associated with host tissue destruction. The factors influencing cross-protective capacity of antibodies are also presented.