Extracellular vesicles of the probiotic bacteria E. coli O83 activate innate immunity and prevent allergy in mice.

BACKGROUND: E. coli O83 (Colinfant Newborn) is a Gram-negative (G-) probiotic bacterium used in the clinic. When administered orally, it reduces allergic sensitisation but not allergic asthma. Intranasal administration offers a non-invasive and convenient delivery method. This route bypasses the gastrointestinal tract and provides direct access to the airways, which are the target of asthma prevention. G- bacteria such as E. coli O83 release outer membrane vesicles (OMVs) to communicate with the environment. Here we investigate whether intranasally administered E. coli O83 OMVs (EcO83-OMVs) can reduce allergic airway inflammation in mice. METHODS: EcO83-OMVs were isolated by ultracentrifugation and characterised their number, morphology (shape and size), composition (proteins and lipopolysaccharide; LPS), recognition by innate receptors (using transfected HEK293 cells) and immunomodulatory potential (in naïve splenocytes and bone marrow-derived dendritic cells; BMDCs). Their allergy-preventive effect was investigated in a mouse model of ovalbumin-induced allergic airway inflammation. RESULTS: EcO83-OMVs are spherical nanoparticles with a size of about 110 nm. They contain LPS and protein cargo. We identified a total of 1120 proteins, 136 of which were enriched in OMVs compared to parent bacteria. Proteins from the flagellum dominated. OMVs activated the pattern recognition receptors TLR2/4/5 as well as NOD1 and NOD2. EcO83-OMVs induced the production of pro- and anti-inflammatory cytokines in splenocytes and BMDCs. Intranasal administration of EcO83-OMVs inhibited airway hyperresponsiveness, and decreased airway eosinophilia, Th2 cytokine production and mucus secretion. CONCLUSIONS: We demonstrate for the first time that intranasally administered OMVs from probiotic G- bacteria have an anti-allergic effect. Our study highlights the advantages of OMVs as a safe platform for the prophylactic treatment of allergy. Video Abstract.

Canine antibody response against Dirofilaria repens in natural occult and microfilaremic infections.

Dirofilaria repens is the causative agent of the zoonotic canine skin condition called subcutaneous dirofilariosis. Despite being endemic throughout much of Europe, little is known about host humoral responses to D. repens. To address this, we analyzed serum immunoglobulin isotypes recognizing D. repens somatic antigens (DrSA) in naturally infected dogs. Titers of anti-DrSA IgG and IgE, but not IgM were significantly higher in dogs infected with D. repens compared to those that were negative. Moreover, microfilaremic infections were associated with higher levels of IgG1 than IgG2, while occult infections occurred with significantly higher levels of IgG2 than IgG1. Finally, the measurement of anti-DrSA IgG antibodies allowed the detection of occult subcutaneous dirofilariosis in dogs.

The Immunological Role of Vascular and Lymphatic Endothelial Cells in Filarial Infections.

The embryonic stage of filarial nematodes, or microfilariae (Mf), shows daily and seasonal periodicity that requires their migration through blood vessels into the lungs, where they are sequestered when not circulating in the peripheral blood. Therefore, Mf and the host endothelium are likely in a permanent state of hide and seek. Interestingly, filarial nematodes co-cultured in media with a murine endothelial cell line survive eight times longer than those cultured in media alone. This suggests that the endothelium is an important element of the immune response in filarial nematodes, perversely promoting their survival in the host. In this review, we will focus on potential pathways involved in the relationship between filarial nematodes and the host endothelium, including the role of endothelial ICAM/VCAM/PECAM adhesion molecules, surface markers involved in the passage of Mf through host tissue, anti-thrombolic effects caused by the presence of filarial nematodes (including plasmins), endothelial cell proliferation (VEGF), and other aspects of the immune activation of the endothelium. The aim of this review is to merge the knowledge about the cross-talk between Mf of different filarial nematode species and endothelial cells (EC), thus allowing a better understanding of the mechanism of these parasitic infections.

Hematological and Biochemical Changes in Dogs Naturally Infected With Dirofilaria repens.

Subcutaneous dirofilariosis is a zoonotic disease emerging throughout Europe caused by the filarial nematode Dirofilaria repens. Despite its increasing prevalence, there is a large gap in knowledge of the impact of the parasite on the host. Currently classified as being non-pathogenic, recent evidence suggests that skin dirofilariosis is associated with dermatological conditions including concomitant pruritus, neoplastic processes, inflammation, and even blindness in dogs and humans. The aim of this study was to determine if natural canine D. repens infection leads to biological changes in the canine host. In a real-life veterinary clinic setting, animals are often presented to clinicians for unrelated issues, and D. repens is incidentally identified during inspection. As such, we compared hematological and biochemical parameters of 218 uninfected and 197 dogs naturally infected with D. repens. Interestingly, animals infected with D. repens had lower numbers of lymphocytes (p < 0.001), red blood cells (p < 0.001), and thrombocytes (p = 0.025), decreased hematocrit (p < 0.001), and increased alkaline phosphatase (p = 0.016) and creatinine activity (p = 0.023) compared to uninfected dogs. We further selected a subpopulation of 214 dogs having prima facie hematological and biochemical results within normal reference ranges to evaluate the effect of D. repens infections in seemingly healthy dogs. Among these patients, 93 dogs infected with D. repens had lower numbers of lymphocytes (p = 0.031), red blood cells (p = 0.025), and hematocrit (p = 0.002), higher glucose levels (p = 0.023), and border line elevated alkaline phosphatase levels (p = 0.054) compared to 121 uninfected animals. Despite being categorized as asymptomatic, we have observed hematological and biochemical changes associated with D. repens infections in dogs, and our data suggest that dirofilariosis may induce a state of chronic stress. These results link the presence of skin dirofilariosis to biological changes in the canine host, suggesting a mechanism for pathogenicity and shedding new light on the host-parasite relationship.

Case Studies of Severe Microfilaremia in Four Dogs Naturally Infected With Dirofilaria repens as the Primary Disease or a Disease Complicating Factor.

Subcutaneous dirofilariosis in dogs, caused by Dirofilaria repens, is an underdiagnosed disease, now recognized for its zoonotic potential, and growing distribution and prevalence across Europe and Asia. Our understanding of the pathogenicity in human and canine host remains unclear, but case reports suggest that microfilariae (Mf) as well as adult D. repens may directly cause internal organs damage or may be a factor complicating the course of other ailments. The purpose of the study was to report high Mf in dogs and to discuss potential relevance with co-morbidity. Our data from a modified Knott's test performed on 62 infected dogs indicate that the median Mf count in D. repens infections is 675 Mf/ml and we consider microfilaremia above 10,000 Mf/ml as high intensity. This collection of case reports discusses 4 cases of high intensity D. repens microfilaremia in companion dogs; one presenting pathology from a very high intensity of adult D. repens with post-treatment complications, and 3 dogs in which high microfilaremia was detected incidentally during the management of other primary illnesses. To our knowledge this report describes the highest D. repens microfilaremia ever detected in a dog, at 178,000 Mf/ml. The issue of high microfilaremic infections in dogs is poorly studied and there is growing need to identify the presentation and understand the mechanisms of associated pathogenesis in the host-parasite relationship.