Mannheimia haemolytica OmpH binds fibrinogen and fibronectin and participates in biofilm formation.

Mannheimia haemolytica is the causal agent of the shipping fever in bovines and produces high economic losses worldwide. This bacterium possesses different virulence attributes to achieve a successful infection. One of the main virulence factors expressed by a pathogen is through adhesion molecules; however, the components participating in this process are not totally known. The present work identified a M. haemolytica 41 kDa outer membrane protein (Omp) that participates in bacterial adhesion. This protein showed 100% identity with the OmpH from M. haemolytica as determined by mass spectrometry and it interacts with sheep fibrinogen. The 41 kDa M. haemolytica OmpH interacts with bovine monocytes; a previous incubation of M. haemolytica with a rabbit hyperimmune serum against this Omp diminished 45% cell adhesion. The OmpH was recognized by serum from bovines affected by acute or chronic pneumonia, indicating its in vivo expression; moreover, it showed immune cross-reaction with the serum of rabbit infected with Pasteurella multocida. The OmpH is present in biofilms and previous incubation of M. haemolytca with rabbit serum against this protein diminished biofilm, indicating this protein's participation in biofilm formation. M. haemolytica OmpH is proposed as a relevant immunogen in bovine pneumonia protection.

Extracellular traps involved in invertebrate immune mechanisms.

The invertebrate immune system possesses a mechanism named extracellular traps (ETs), it has been identified that this mechanism immobilizes and kills pathogens. ETs formation induces modification of histones, chromatin decondensation, and mixes with granule molecules, releasing them into the extracellular space as a defense mechanism. In the present review, we provide an overview on the identification of triggering stimuli such as pathogens, PAMPs, DAMPs, and chemical stimuli, discuss the participation of potential signaling pathways involving MAPK, PI3K, PKC, and ERK molecules that lead to NADPH oxidase or mitochondrial ROS production, and explore the potential relationship with several proteins such as myeloperoxidase, heat sock proteins, peroxinectin, elastase, and apolipoproteins. Furthermore, we also discuss the association of ETs with other immune mechanisms that could collaborate in the elimination of pathogens.

In vivo administration of LPS and ß-glucan generates the expression of a serum lectin and its cellular receptor in Cherax quadricarinatus.

In crustaceans, it has been suggested that specific protection against pathogens could be triggered by vaccines and biological response modifiers; although the specific mechanisms of this protection have not been clarified yet. In the crayfish Cherax quadricarinatus, a humoral lectin (CqL) binds its own granular hemocytes through a specific receptor (CqLR) and increases the production of reactive oxygen species (ROS). In the present study, we challenged in vivo crayfishes with immunostimulants, ß-glucan (200 µg/kg) or LPS (20 µg/kg), and identified the participation of cellular and humoral mechanisms. The stimulants generated a complex modification in the total hemocytes count (THC), as well as in the proportion of hemocyte subsets. At 2 h after the challenge, the largest value in THC was observed in either challenged crayfishes. Furthermore, at the same time, hyaline hemocytes were the most abundant subset in the hemolymph; after 6 h, granular hemocytes (GH) were the most abundant hemocyte subset. It has been observed that a specific subset of GH possesses a CqLR that has been related to ROS production. After 2 and 6 h of the ß-glucan challenge, a significant increase in CqLR expression was observed in the three circulating hemocyte subsets; also, an increased expression of CqL was detected in a granular hemocytes sub-population. After 2 and 6 h of stimulation, the specific activity of the serum lectin challenged with ß-glucan was 250% and 160% higher than in the LPS-treated-group, respectively (P < 0.05). Hemocytes from challenged crayfishes were stimulated ex vivo with CqL, ROS production was 180% higher in hemocytes treated with ß-glucan + CqL than in hemocytes treated with LPS + CqL (P < 0.05). The results evidence the effectivity of immune stimulators to activate specific crayfish defense mechanisms, the participation of CqL and its receptor (CqLR) could play an important role in the regulation of immune cellular functions, like ROS production, in Cherax quadricarinatus.

The effect of the lectin from Cherax quadricarinatus on its granular hemocytes.

In crustaceans, lectins and hemocytes of the innate immune system provide the first line of defense. Although evidence points to the potential role of lectins in regulating hemocyte activity, the processes underlying the lectin activation have not been evaluated. In the present study, the receptor for CqL, a humoral lectin from Cherax quadricarinatus specific for galactose/sialic acid, was identified in a granular subset of hemocytes. The CqL receptor (CqLR) is a 490-kDa glycoprotein, composed of four identical 120-kDa subunits. As shown by immunohistochemistry, CqL at 7.5 µg/mL as optimal dose, after 2 min, induced, specifically on granular hemocytes, increased phosphorylation of serine (152%), threonine (192%), and tyrosine (242%) as compared with non-treated hemocytes; moreover, CqL induced increased generation of reactive oxygen species (ROS). Specific kinase inhibitors showed inhibition (P < 0.001) of ROS production induced by CqL. These results strongly suggest that CqL actively participated in the generation of ROS through kinases induced by a CqLR in a subset of granular hemocytes of the crayfish C. quadricarinatus. The results provide strong evidence that CqL activates, through specific granular hemocytes, receptors that modulate cellular functions in C. quadricarinatus.