Agglutination processes of activated sludge cultures induced by extracellular lectins

We examine the agglutinating ability of five compounds, namely, A1, A2, A3, A4 and BS1, isolated from activated sludge on selective media typical of a number of dominant microbial cultures that contribute to the formation of microbial aggregates. The morphological properties of the isolates and their lectin activity, as well as the physiological and biochemical properties of individual isolates were studied; microorganisms in their composition were identified. We assessed the capacity of the isolates under study to synthesize an exopolysaccharide matrix, as well as the sedimentation of activated sludge under the action of the native solution and culture liquid of the BS1 isolate. Based on their capacity to agglutinate, the BS1 and A2 isolates were selected for further research as producers of extracellular lectins and objects of agglutination, respectively. The biophysiochemical properties and molecular-genetic identification of the BS1 isolate allowed the degree of identity with r. Bacillus to be defined (96.19%); for the A2 isolate, 92.93% identity with p. Shigella and p. Escherichia was determined. To assess the capacity to synthesize a biofilm matrix, the BS1 and A2 isolates were cultivated on an agar nutrient solution using Congo Red dye. According to the obtained results, the isolates are capable of synthesizing an exopolysaccharide matrix, the main component of bacterial biofilms. The research results on the sedimentation of activated sludge induced by the native solution and culture liquid of BS1 showed the following. The sedimentation rate of activated sludge increased significantly at the beginning of the process upon adding a BS1 cell suspension, while the introduction of the native solution of BS1 intensified the process following 5 minutes of contact. The obtained experimental data suggest that the media containing extracellular bacterial lectins can be effectively used as a coagulant (flocculant) for the sedimentation of activated sludge.

Proteome-wide prediction of bacterial carbohydrate-binding proteins as a tool for understanding commensal and pathogen colonisation of the vaginal microbiome

Bacteria use carbohydrate-binding proteins (CBPs), such as lectins and carbohydrate-binding modules (CBMs), to anchor to specific sugars on host surfaces. CBPs in the gut microbiome are well studied, but their roles in the vagina microbiome and involvement in sexually transmitted infections, cervical cancer and preterm birth are largely unknown. We established a classification system for lectins and designed Hidden Markov Model (HMM) profiles for data mining of bacterial genomes, resulting in identification of >100,000 predicted bacterial lectins available at unilectin.eu/bacteria. Genome screening of 90 isolates from 21 vaginal bacterial species shows that those associated with infection and inflammation produce a larger CBPs repertoire, thus enabling them to potentially bind a wider array of glycans in the vagina. Both the number of predicted bacterial CBPs and their specificities correlated with pathogenicity. This study provides new insights into potential mechanisms of colonisation by commensals and potential pathogens of the reproductive tract that underpin health and disease states.

The conserved core component EsaA mediates bacterial killing by the type VIIb secretion system

Type VII secretion systems (T7SS) are employed by Gram-positive bacteria to infect the host or to intoxicate bacterial competitors. Bacterial killing mediated by other secretion systems involves the assembly of large, extracellular structures promoting toxin delivery into bacterial competitors. However, such extracellular structures are not known for T7SS nor is it known whether the secreted toxins have cell-penetrating properties. Here, we used a multi-drug resistant Staphylococcus aureus, one of the most relevant hospital-associated bacterial pathogens, to study the mechanism of bacterial killing by the T7SSb. We report the proteolysis-resistant, extracellular structure of the EsaA core component, which is an integral part of the membrane-embedded T7SSb secretion machine. The extracellular structure of EsaA has an elongated antenna-like shape providing an explanation how the T7SSb spans the staphylococcal cell wall. We demonstrate that the extracellular structure of EsaA has an essential function in secretion and contributes to the killing of prey bacteria. We show that the bactericidal activity of EsaA can be attributed to a membrane-damaging activity that leads to the permeabilization of phospholipid bilayers and could facilitate the delivery of antibacterial toxin into bacterial competitors. The structural similarity of the EsaA tip domain to bacterial lectins and human integrins suggests an additional function of EsaA in cell adhesion during infection or in targeting of bacterial competitors.

The conserved core component EsaA mediates bacterial killing by the type VIIb secretion system

Type VII secretion systems (T7SS) are employed by Gram-positive bacteria to infect the host or to intoxicate bacterial competitors. Bacterial killing mediated by other secretion systems involves the assembly of large, extracellular structures promoting toxin delivery into bacterial competitors. However, such extracellular structures are not known for T7SS nor is it known whether the secreted toxins have cell-penetrating properties. Here, we used a multi-drug resistant Staphylococcus aureus, one of the most relevant hospital-associated bacterial pathogens, to study the mechanism of bacterial killing by the T7SSb. We report the proteolytic resistant, extracellular structure of the EsaA core component, which is an integral part of the membrane-embedded T7SSb secretion machine. The extracellular structure of EsaA has an elongated antenna-like shape providing an explanation how the T7SSb spans the staphylococcal cell wall. We demonstrate that the extracellular structure of EsaA has an essential function in secretion and contributes to the killing of prey bacteria. We show that the bactericidal activity of EsaA can be attributed to a membrane-damaging activity that leads to the permeabilization of phospholipid bilayers and could facilitate the delivery of antibacterial toxin into bacterial competitors. The structural similarity of the EsaA tip domain to bacterial lectins and human integrins suggests an additional function of EsaA in cell adhesion during infection or in targeting of bacterial competitors.

Proteome-wide prediction of bacterial carbohydrate-binding proteins as a tool for understanding commensal and pathogen colonisation of the vaginal microbiome

Bacteria use protein receptors called lectins to anchor to specific host surface sugars. The role of lectins in the vaginal microbiome, and their involvement in reproductive tract pathophysiology is poorly defined. Here we establish a classification system based on taxonomy and protein 3D structure to identify 109 lectin classes. Hidden Markov Model (HMM) profiles for each class were used to search bacterial genomes, resulting in the prediction of >100 000 bacterial lectins available at unilectin.eu/bacteria. Genome screening of 90 isolates from 21 vaginal bacterial species showed that potential pathogens produce a larger variety of lectins than commensals indicating increased glycan-binding potential. Both the number of predicted bacterial lectins, and their specificities for carbohydrates correlated with pathogenicity. This study provides new insights into potential mechanisms of commensal and pathogen colonisation of the reproductive tract that underpin health and disease states.

Nutrition and gut health: the impact of specific dietary components – it's not just five-a-day

The health benefits of fruit, vegetables and dietary fibre have been promoted for many years. Much of the supporting evidence is circumstantial or even contradictory and mechanisms underlying health benefits of specific foods are poorly understood. Colorectal cancer shows marked geographical differences in incidence, probably linked with diet, and explanations for this require knowledge of the complex interactions between diet, microbiota and the gut epithelium. Dietary fibres can act as prebiotics, encouraging growth of saccharolytic bacteria, but other mechanisms are also important. Some but not all soluble fibres have a ‘contrabiotic’ effect inhibiting bacterial adherence to the epithelium. This is particularly a property of pectins (galacturonans) whereas dietary fructans, previously regarded as beneficial prebiotics, can have a proinflammatory effect mediated via toxic effects of high butyrate concentrations. This also suggests that ulcerative colitis could in part result from potentially toxic faecal butyrate concentrations in the presence of a damaged mucus layer. Epithelial adherence of lectins, either dietary lectins as found in legumes, or bacterial lectins such as the galactose-binding lectin expressed by colon cancer-associated Fusobacterium nucleatum, may also be important and could be inhibitable by specific dietary glycans. Conversely, emulsifiers in processed foods may increase bacterial translocation and alter the microbiota thus promoting inflammation or cancer. Focusing on one condition is of limited value although in developing public health messages and growing evidence for impacts of dietary components on all-cause mortality is gaining more attention. We are only just starting to understand the complex interactions between food, the microbiota and health.

Carbohydrate-dependent B cell activation by fucose-binding bacterial lectins

Bacterial lectins are typically multivalent and bind noncovalently to specific carbohydrates on host tissues to facilitate bacterial adhesion. Here, we analyzed the effects of two fucose-binding lectins, BambL fromBurkholderia ambifariaand LecB fromPseudomonas aeruginosa, on specific signaling pathways in B cells. We found that these bacterial lectins induced B cell activation, which, in vitro, was dependent on the cell surface expression of the B cell antigen receptor (BCR) and its co-receptor CD19, as well as on spleen tyrosine kinase (Syk) activity. The resulting release of intracellular Ca2+was followed by an increase in the cell surface abundance of the activation marker CD86, augmented cytokine secretion, and subsequent cell death, replicating all of the events that are observed in vitro upon canonical and antigen-mediated B cell activation. Moreover, injection of BambL in mice resulted in a substantial, BCR-independent loss of B cells in the bone marrow with simultaneous, transient enlargement of the spleen (splenomegaly), as well as an increase in the numbers of splenic B cells and myeloid cells. Together, these data suggest that bacterial lectins can initiate polyclonal activation of B cells through their sole capacity to bind to fucose.

Lectins ofMycobacterium tuberculosis– rarely studied proteins

The importance of bacterial lectins for adhesion, pathogenicity, and biofilm formation is well established for many Gram-positive and Gram-negative bacteria. However, there is very little information available about lectins of the tuberculosis-causing bacterium,Mycobacterium tuberculosis(Mtb). In this paper we review previous studies on the carbohydrate-binding characteristics of mycobacteria and relatedMtbproteins, discussing their potential relevance toMtbinfection and pathogenesis.