Comparative analysis of the interactions of different Streptococcus suis strains with monocytes, granulocytes and the complement system in porcine blood.

Streptococcus suis (S. suis) is an important porcine pathogen causing meningitis, arthritis, and septicemia. Serotypes 2 and 14 are the most common zoonotic ones worldwide, whereas serotypes 2, 9, and 7 are very important in pigs in Europe. To cause invasive infections S. suis needs to enter the bloodstream. Consequently, the immune response in blood represents an important line of defense and bacteremia plays a key role in the pathogenesis of invasive S. suis infections. We investigated the working hypothesis that S. suis strains of the same serotype but different clonal complex (CC) might exhibit substantial differences in the interaction with components of the immune system in porcine blood. The experimental design of this study includes comparative analysis of 8 virulent strains belonging to 4 serotypes with strains of the same serotype being genetically not closely related. Significant differences between two strains of the same serotype but different clonal complex were recorded in the flow cytometric analysis of association with different leukocytes for serotype 9 and 14. Our results demonstrate that the serotype 9 strain of CC94 shows significantly increased association with monocytes and survival in porcine blood of conventional piglets as well as a tendency towards decreased composition of C3 in plasma of these piglets in comparison to the serotype 9 strain of CC16. Correlation analysis of C3 deposition on the bacterial surface and survival in respective blood samples of 8-week-old piglets demonstrated a negative correlation indicating that C3 deposition is a crucial step to limit bacterial survival and proliferation of different S. suis pathotypes in the blood of these piglets. In summary, our results indicate that the capsule composition of a S. suis strain is not alone sufficient to determine association with leukocytes, activation of complement, induction of proinflammatory cytokines, oxidative burst, and bacterial survival in porcine blood. In this study, substantial differences in these host-pathogen interactions were observed between strains of the same serotype. Therefore, a more comprehensive characterization of the field isolates, including at least MLST analysis to determine the sequence type/clonal complex, is recommended.

A distinct variant of the SzM protein of Streptococcus equi subsp. zooepidemicus recruits C1q independent of IgG binding and inhibits activation of the classical complement pathway.

Streptococcus equi subsp. zooepidemicus (SEZ) is a major equine pathogen that causes pneumonia, abortion, and polyarthritis. It can also cause invasive infections in humans. SEZ expresses the M-like protein SzM, which recruits host proteins such as fibrinogen to the bacterial surface. Equine SEZ strain C2, which binds only comparably low amounts of human fibrinogen in comparison to human SEZ strain C33, was previously shown to proliferate in equine and human blood. As the expression of SzM_C2 was necessary for survival in blood, this study investigated the working hypothesis that SzM_C2 inhibits complement activation through a mechanism other than fibrinogen and non-immune immunoglobulin binding. Loss-of-function experiments showed that SEZ C2, but not C33, binds C1q via SzM in IgG-free human plasma. Furthermore, SzM C2 expression is necessary for recruiting purified human or equine C1q to the bacterial surface. Flow cytometry analysis demonstrated that SzM expression in SEZ C2 is crucial for the significant reduction of C3b labelling in human plasma. Addition of human plasma to immobilized rSzM_C2 and immobilized aggregated IgG led to binding of C1q, but only the latter activated the complement system, as shown by the detection of C4 deposition. Complement activation induced by aggregated IgG was significantly reduced if human plasma was pre-incubated with rSzM_C2. Furthermore, rSzM_C2, but not rSzM_C33, inhibited the activation of the classical complement pathway in human plasma, as determined in an erythrocyte lysis experiment. In conclusion, the immunoglobulin-independent binding of C1q to SzM_C2 is associated with complement inhibition.

Immunoglobulin M-degrading enzyme of Streptococcus suis (Ide Ssuis ) impairs porcine B cell signaling.

Streptococcus suis (S. suis) is an important porcine pathogen, causing severe disease like meningitis and septicemia primarily in piglets. Previous work showed that the IgM-degrading enzyme of S. suis (Ide Ssuis ) specifically cleaves soluble porcine IgM and is involved in complement evasion. The objective of this study was to investigate Ide Ssuis cleavage of the IgM B cell receptor and subsequent changes in B cell receptor mediated signaling. Flow cytometry analysis revealed cleavage of the IgM B cell receptor by recombinant (r) Ide Ssuis _homologue as well as Ide Ssuis derived from culture supernatants of S. suis serotype 2 on porcine PBMCs and mandibular lymph node cells. Point-mutated rIde Ssuis _homologue_C195S did not cleave the IgM B cell receptor. After receptor cleavage by rIde Ssuis _homologue, it took at least 20 h for mandibular lymph node cells to restore the IgM B cell receptor to levels comparable to cells previously treated with rIde Ssuis _homologue_C195S. B cell receptor mediated signaling after specific stimulation via the F(ab')2 portion was significantly inhibited by rIde Ssuis _homologue receptor cleavage in IgM+ B cells, but not in IgG+ B cells. Within IgM+ cells, CD21+ B2 cells and CD21- B1-like cells were equally impaired in their signaling capacity upon rIde Ssuis _homologue B cell receptor cleavage. In comparison, intracellular B cell receptor independent stimulation with tyrosine phosphatase inhibitor pervanadate increased signaling in all investigated B cell types. In conclusion, this study demonstrates Ide Ssuis cleavage efficacy on the IgM B cell receptor and its consequences for B cell signaling.

Evaluation of proline-rich antimicrobial peptides as potential lead structures for novel antimycotics against Cryptococcus neoformans.

Background: Cryptococcosis and cryptococcal meningitis, caused by Cryptococcus neoformans infections, lead to approximately 180,000 deaths per year, primarily in developing countries. Individuals with compromised immune systems, e.g., due to HIV infection (AIDS) or chemotherapy, are particularly vulnerable. Conventional treatment options are often limited and can cause severe side effects. Therefore, this study aimed to investigate the antifungal effect of insect-derived proline-rich antimicrobial peptides (PrAMPs) against C. neoformans. These peptides are known for their low toxicity and their high efficacy in murine infection models, making them a promising alternative for treatment. Results: A preliminary screening of the minimal inhibitory concentrations (MICs) of 20 AMPs, including the well-known PrAMPs Onc112, Api137, and Chex1Arg20 as well as the cathelicidin CRAMP against the C. neoformans strains 1841, H99, and KN99α revealed promising results, with MICs as low as 1.6 µmol/L. Subsequent investigations of selected peptides, determining their influence on fungal colony-forming units, confirmed their strong activity. The antifungal activity was affected by factors such as peptide net charge and sequence, with stronger effects at higher net charges probably due to better intracellular uptake confirmed by confocal laser scanning microscopy. Inactive scrambled peptides suggest a specific intracellular target, although scanning electron microscopy showed that PrAMPs also damaged the cell exterior for a low proportion of the cells. Possible pore formation could facilitate entry into the cytosol.

Road traffic noise impacts sleep continuity in suburban residents: Exposure-response quantification of noise-induced awakenings from vehicle pass-bys at night.

Nocturnal traffic noise has been associated with adverse health outcomes in exposed residents. Precise quantification of traffic noise effects on sleep is thus of great importance. Here we establish an exposure-response relationship for the awakening probability due to intermittent road traffic noise in suburban residents. We conducted a field study in residential areas where road traffic was the dominant noise source, and noise events were attributable to separate vehicle pass-bys. Forty healthy participants underwent polysomnography for five consecutive nights at their homes. A total of 11,003 road traffic noises derived from simultaneous acoustic measurements at the sleepers' ears were included in an event-related analysis of awakenings. Logistic regression analysis revealed that the awakening probability due to road traffic noise increased with the maximum sound pressure level (SPL) and the maximum slope of the increasing SPL of a vehicle pass-by, as well as the age of the exposed individual. Compared to sleep stage 2, the awakening probability was higher in rapid eye movement sleep (REMS) and lower in slow wave sleep (SWS). The protective effect of both stage 2 and SWS against awakenings decreased with age, whereas no age-dependent change was observed for REMS. When adjusting for other contributing factors, the probability of a noise-induced awakening ranged from 0% at a maximum SPL of 27.1 dB(A) to 2.0% at 70 dB(A). Road traffic noise at night - even in suburban areas with moderate traffic density - negatively impacts residents' sleep continuity. Exposure-response quantification for traffic noise-induced awakenings may serve as a basis for noise protection efforts by regulators and policy makers.

Traffic noise-induced changes in wake-propensity measured with the Odds-Ratio Product (ORP).

Nocturnal traffic noise can disrupt sleep and impair physical and mental restoration, but classical sleep scoring techniques may not fully capture subtle yet clinically relevant alterations of sleep induced by noise. We used a validated continuous measure of sleep depth and quality based on automatic analysis of physiologic sleep data, termed Wake Propensity (WP), to investigate temporal changes of sleep in response to nocturnal noise events in 3-s epochs. Seventy-two healthy participants (mean age 40.3 years, range 18-71 years, 40 females, 32 males) slept for 11 nights in a laboratory, during which we measured sleep with polysomnography. In 8 nights, participants were exposed to 40, 80 or 120 road, rail and/or aircraft noise events with maximum noise levels of 45-65 dB LAS,max during 8-h sleep opportunities. We analyzed sleep macrostructure and event-related change of WP during noise exposure with linear mixed models. Nocturnal traffic noise led to event-related shifts towards wakefulness and less deep, more unstable sleep (increase in WP relative to pre-noise baseline ranging from +29.5% at 45 dB to +38.3% at 65 dB; type III effect p < 0.0001). Sleep depth decreased dynamically with increasing noise level, peaking when LAS,max was highest. This change in WP was stronger and occurred more quickly for events where the noise onset was more rapid (road and rail) compared to more gradually time-varying noise (aircraft). Sleep depth did not immediately recover to pre-noise WP, leading to decreased sleep stability across the night compared to quiet nights, which was greater with an increasing number of noise events (standardized ß = 0.053, p = 0.003). Further, WP was more sensitive to noise than classical arousals. Results demonstrate the usefulness of WP as a measure of the effects of external stimuli on sleep, and show WP is a more sensitive measure of noise-induced sleep disruption than traditional methods of sleep analysis.

Identification of Disease-Associated Cryptococcal Proteins Reactive With Serum IgG From Cryptococcal Meningitis Patients.

Cryptococcus neoformans, an opportunistic fungal pathogen ubiquitously present in the environment, causes cryptococcal meningitis (CM) mainly in immunocompromised patients, such as AIDS patients. We aimed to identify disease-associated cryptococcal protein antigens targeted by the human humoral immune response. Therefore, we used sera from Colombian CM patients, with or without HIV infection, and from healthy individuals living in the same region. Serological analysis revealed increased titers of anti-cryptococcal IgG in HIV-negative CM patients, but not HIV-positive CM patients, compared to healthy controls. In contrast, titers of anti-cryptococcal IgM were not affected by CM. Furthermore, we detected pre-existing IgG and IgM antibodies even in sera from healthy individuals. The observed induction of anti-cryptococcal IgG but not IgM during CM was supported by analysis of sera from C. neoformans-infected mice. Stronger increase in IgG was found in wild type mice with high lung fungal burden compared to IL-4Rα-deficient mice showing low lung fungal burden. To identify the proteins targeted by human anti-cryptococcal IgG antibodies, we applied a quantitative 2D immunoproteome approach identifying cryptococcal protein spots preferentially recognized by sera from CM patients or healthy individuals followed by mass spectrometry analysis. Twenty-three cryptococcal proteins were recombinantly expressed and confirmed to be immunoreactive with human sera. Fourteen of them were newly described as immunoreactive proteins. Twelve proteins were classified as disease-associated antigens, based on significantly stronger immunoreactivity with sera from CM patients compared to healthy individuals. The proteins identified in our screen significantly expand the pool of cryptococcal proteins with potential for (i) development of novel anti-cryptococcal agents based on implications in cryptococcal virulence or survival, or (ii) development of an anti-cryptococcal vaccine, as several candidates lack homology to human proteins and are localized extracellularly. Furthermore, this study defines pre-existing anti-cryptococcal immunoreactivity in healthy individuals at a molecular level, identifying target antigens recognized by sera from healthy control persons.

iSens: A Fiber-Based, Highly Permeable and Imperceptible Sensor Design.

Embedded sensors are key to optimizing processes and products; they collect data that allow time, energy, and materials to be saved, thereby reducing costs. After production, they remain in place and are used to monitor the long-term structural health of buildings or aircraft. Fueled by climate change, sustainable construction materials such as wood and fiber composites are gaining importance. Current sensors are not optimized for use with these materials and often act as defects that cause catastrophic failures. Here, flexible, highly permeable, and imperceptible sensors (iSens) are introduced that integrate seamlessly into a component. Their porous substrates are readily infused with adhesives and withstand harsh conditions. In situ resistive temperature measurements and capacitive sensing allows monitoring of adhesives curing as used in wooden structures and fiber composites. The devices also act as heating elements to reduce the hardening time of the glue. Results are analyzed using numerical simulations and theoretical analysis. The suggested iSens technology is widely applicable and represents a step towards realizing the Internet of Things for construction materials.

Removal of Trace Organic Contaminants by Parallel Operation of Reverse Osmosis and Granular Activated Carbon for Drinking Water Treatment.

In response to increasingly stringent restrictions for drinking water quality, a parallel operation of two common technologies, low-pressure reverse osmosis (LPRO) and activated carbon filtration (ACF), was investigated in a comprehensive five-month pilot study for the removal of 32 typical trace organic contaminants (TrOCs) from Rhine bank filtrates employing a semi- technical plant. TrOCs have been divided into three groups: polyfluorinated aliphatic compounds; pharmaceuticals, pesticides and metabolites; in addition to volatiles, nitrosamines and aminopolycarboxylic acids, which were also examined. The net pressure behavior, normalized salt passage and rejection of TrOCs by LPRO were investigated and compared with ACF operation. In addition, autopsies from the leading and last membrane modules were performed using adenosine triphosphate (ATP), total organic carbon (TOC), ICP-OES and SEM-EDX techniques. Generally, rather stable LPRO membrane performance with limited membrane fouling was observed. TrOCs with a molecular weight of ≥ 150 Da were completely retained by LPRO, while the rejection of di- and trichloro compounds improved as the filtration progressed. ACF also showed significant removal for most of the TrOCs, but without desalination. Accordingly, the ACF and LPRO can be operated in parallel such that the LPRO permeate and the ACF-treated bypass can be mixed to produce drinking water with adjustable hardness and significantly reduced TrOCs.

Construction of a 3D brain extracellular matrix model to study the interaction between microglia and T cells in co-culture.

Neurodegenerative disorders are characterised by the activation of brain-resident microglia cells and by the infiltration of peripheral T cells. However, their interplay in disease has not been clarified yet. It is difficult to investigate complex cellular dynamics in living animals, and simple two-dimensional (2D) cell culture models do not resemble the soft 3D structure of brain tissue. Therefore, we developed a biomimetic 3D in vitro culture system for co-cultivation of microglia and T cells. As the activation and/or migration of immune cells in the brain might be affected by components of the extracellular matrix, defined 3D fibrillar collagen I-based matrices were constructed and modified with hyaluronan and/or chondroitin sulphate, resembling aspects of brain extracellular matrix. Murine microglia and spleen-derived T cells were cultured alone or in co-culture on the constructed matrices. Microglia exhibited in vivo-like morphology and T cells showed enhanced survival when co-cultured with microglia or to a minor degree in the presence of glia-conditioned medium. The open and porous fibrillar structure of the matrix allowed for cell invasion and direct cell-cell interaction, with stronger invasion of T cells. Both cell types showed no dependence on the matrix modifications. Microglia could be activated on the matrices by lipopolysaccharide resulting in interleukin-6 and tumour necrosis factor-α secretion. The findings herein indicate that biomimetic 3D matrices allow for co-cultivation and activation of primary microglia and T cells and provide useful tools to study their interaction in vitro.

Thermomechanical and microhardness data of melamine-formaldehyde-based self-healing resin film able to undergo reversible crosslinking via Diels-Alder reaction.

The data presented in this article characterize the thermomechanical and microhardness properties of a novel melamine-formaldehyde resin (MF) intended for the use as a self-healing surface coating. The investigated MF resin is able to undergo reversible crosslinking via Diels Alder reactive groups. The microhardness data were obtained from nanoindentation measurements performed on solid resin film samples at different stages of the self-healing cycle. Thermomechanical analysis was performed under dynamic load conditions. The data provide supplemental material to the manuscript published by Urdl et al. 2020 (http://doi.org/10.1016/j.eurpolymj.2020.109601, [1]) on the self-healing performance of this resin, where a more thorough discussion on the preparation, the properties of this coating material and its application in impregnated paper-based decorative laminates can be found [1].

Group 2 Innate Lymphoid Cells (ILC2) Suppress Beneficial Type 1 Immune Responses During Pulmonary Cryptococcosis.

Cryptococcus neoformans is an opportunistic fungal pathogen preferentially causing disease in immunocompromised individuals such as organ-transplant-recipients, patients receiving immunosuppressive medications or, in particular, individuals suffering from HIV infection. Numerous studies clearly indicated that the control of C. neoformans infections is strongly dependent on a prototypic type 1 immune response and classical macrophage activation, whereas type 2-biased immunity and alternative activation of macrophages has been rather implicated in disease progression and detrimental outcomes. However, little is known about regulatory pathways modulating and balancing immune responses during early phases of pulmonary cryptococcosis. Here, we analyzed the role of group 2 innate lymphoid cells (ILC2s) for the control of C. neoformans infection. Using an intranasal infection model with a highly virulent C. neoformans strain, we found that ILC2 numbers were strongly increased in C. neoformans-infected lungs along with induction of a type 2 response. Mice lacking ILC2s due to conditional deficiency of the transcription factor RAR-related orphan receptor alpha (Rora) displayed a massive downregulation of features of type 2 immunity as reflected by reduced levels of the type 2 signature cytokines IL-4, IL-5, and IL-13 at 14 days post-infection. Moreover, ILC2 deficiency was accompanied with increased type 1 immunity and classical macrophage activation, while the pulmonary numbers of eosinophils and alternatively activated macrophages were reduced in these mice. Importantly, this shift in pulmonary macrophage polarization in ILC2-deficient mice correlated with improved fungal control and prolonged survival of infected mice. Conversely, adoptive transfer of ILC2s was associated with a type 2 bias associated with less efficient anti-fungal immunity in lungs of recipient mice. Collectively, our date indicate a non-redundant role of ILC2 in orchestrating myeloid anti-cryptococcal immune responses toward a disease exacerbating phenotype.

Prominent Binding of Human and Equine Fibrinogen to Streptococcus equi subsp. zooepidemicus Is Mediated by Specific SzM Types and Is a Distinct Phenotype of Zoonotic Isolates.

Streptococcus equi subsp. zooepidemicus is an important pathogen in horses that causes severe diseases such as pneumonia and abortion. Furthermore, it is a zoonotic agent, and contact with horses is a known risk factor. In this study, we investigated the working hypothesis that the zoonotic potential varies among S. equi subsp. zooepidemicus strains in association with differences in M-like protein-mediated binding of host plasma proteins. We demonstrate via in-frame deletion mutagenesis of two different S. equi subsp. zooepidemicus strains that the M-like protein SzM is crucial for the binding of fibrinogen to the bacterial surface and for survival in equine and human blood. S. equi subsp. zooepidemicus isolates of equine and human origins were compared with regard to SzM sequences and binding of equine and human fibrinogens. The N-terminal 216 amino acids of the mature SzM were found to exhibit a high degree of diversity, but the majority of human isolates grouped in three distinct SzM clusters. Plasma protein absorption assays and flow cytometry analysis revealed that pronounced binding of human fibrinogen is a common phenotype of human S. equi subsp. zooepidemicus isolates but much less so in equine S. equi subsp. zooepidemicus isolates. Furthermore, binding of human fibrinogen is associated with specific SzM types. These results suggest that SzM-mediated binding of human fibrinogen is an important virulence mechanism of zoonotic S. equi subsp. zooepidemicus isolates.

Data on production and characterization of melamine-furan-formaldehyde particles and reversible reactions thereof.

The data present in this article affords insides in the characterization of a newly described bi-functional furan-melamine monomer, which is used for the production of monodisperse, furan-functionalized melamine-formaldehyde particles, as described in https://doi.org/10.1016/j.eurpolymj.2019.04.006 Urdl et al., 2019. In the related research article Urdl et al., 2019 data interpretations can be found. The furan-functionalization of particles is necessary to perform reversible Diels-Alder reactions with maleimide (BMI) crosslinker to form thermoreversible network systems. To understand the reaction conditions of Diels-Alder (DA) reaction with a Fu-Mel monomer and a maleimide crosslinker, model DA reaction were performed and evaluated using dynamic FT-IR measurements. During retro Diels-Alder (rDA) reactions of the monomer system, it was found out that some side reaction occurred at elevated temperatures. The data of evaluating the side reaction is described in one part of this manuscript. Additional high resolution SEM images of Fu-Mel particles are shown and thermoreversible particle networks with BMI2 are shown. The data of different Fu-Mel particle networks with maleimide crosslinker are presented. Therefore, the used maleimide crosslinker with different spacer lengths were synthesized and the resulting networks were analyzed by ATR-FT-IR, SEM and DSC.

Characterization of phosphonate-based antiscalants used in drinking water treatment plants by anion-exchange chromatography coupled to electrospray ionization time-of-flight mass spectrometry and inductively coupled plasma mass spectrometry.

In drinking water production, phosphonates are frequently applied as antiscalants in order to prevent the precipitation of salts on reverse osmosis membranes. Whereas the nominal constituents are defined, phosphonate-based antiscalants were found to contain significant amounts of undeclared phosphorous contaminants, particularly noticeable technical diethylenetriamine penta(methylene phosphonic acid) (DTPMP). Following a literature review on impurities of phosphonic acids, separation by anion exchange chromatography coupled with ICP-MS- and ESI-TOF-detection allowed the comprehensive characterization of technical phosphonates. After the identification of synthesis by-products, intermediates and degradation products, the quantification of the phosphorus compounds was accomplished by complementary phosphorus-sensitive ICP-MS detection. The contents of the nominal constituents as well as the organophosphorous impurities of 24 technical antiscalants were determined and compared with the manufacturer's declaration. Of all compounds detected within the technical formulations, 86% were confirmed with reference compounds. Phosphorous impurities were found to contribute to the total phosphorus content with close to 20% (in ATMP- and PBTC-based products) and 38%-65% (in DTPMP-based products). Correspondingly, the nominal compounds were found to contribute with 78%-80% (based on 6 ATMP products) and with 81.5%-83% (based on 2 PBTC products), whereas in 10 out of 12 DTPMP products the nominal compound was found to contribute with 34%-44% to the total phosphorus. Only in two products, elevated DTPMP proportions of 55% and 62% with regard to total phosphorus were determined. The screening for aminomethylphosphonic acid (AMPA) revealed contents between 1.9 mg/L and 157 mg/L. This study represents the first in-depth characterization of phosphonate-based antiscalant products and stock solutions used for drinking water production.

Orf virus (ORFV) infection in a three-dimensional human skin model: Characteristic cellular alterations and interference with keratinocyte differentiation.

ORF virus (ORFV) is the causative agent of contagious ecthyma, a pustular dermatitis of small ruminants and humans. Even though the development of lesions caused by ORFV was extensively studied in animals, only limited knowledge exists about the lesion development in human skin. The aim of the present study was to evaluate a three-dimensional (3D) organotypic culture (OTC) as a human skin model for ORFV infection considering lesion development, replication of the virus, viral gene transcription and modulation of differentiation of human keratinocytes by ORFV. ORFV infection of OTC was performed using the ORFV isolate B029 derived from a human patient. The OTC sections showed a similar structure of stratified epidermal keratinocytes as human foreskin and a similar expression profile of the differentiation markers keratin 1 (K1), K10, and loricrin. Upon ORFV infection, OTCs exhibited histological cytopathic changes including hyperkeratosis and ballooning degeneration of the keratinocytes. ORFV persisted for 10 days and was located in keratinocytes of the outer epidermal layers. ORFV-specific early, intermediate and late genes were transcribed, but limited viral spread and restricted cell infection were noticed. ORFV infection resulted in downregulation of K1, K10, and loricrin at the transcriptional level without affecting proliferation as shown by PCNA or Ki-67 expression. In conclusion, OTC provides a suitable model to study the interaction of virus with human keratinocytes in a similar structural setting as human skin and reveals that ORFV infection downregulates several differentiation markers in the epidermis of the human skin, a hitherto unknown feature of dermal ORFV infection in man.

Orf virus infection of human keratinocytes and dermal fibroblasts: Limited virus detection and interference with intercellular adhesion molecule-1 up-regulation.

Orf virus (Parapoxvirus ovis, ORFV) is a dermatotropic virus causing pustular dermatitis in small ruminants and humans. We analysed isolated human primary keratinocytes (KC) and dermal fibroblasts (FB) for cell death and virus replication by infection with a patient-derived ORFV isolate. ORFV infection was associated with rapid induction of cell death in KC allowing for considerable virus removal. Upon infection with ORFV, KC and FB harboured intracytoplasmic ORFV and showed viral protein presence; however, missing virus spread indicated an abortive infection. Upon ORFV exposure, KC but not FB secreted the pro-inflammatory cytokine interleukin (IL)-6. ORFV infection enhanced the frequency of KC expressing intercellular adhesion molecule (ICAM)-1 which was independent of IL-6. Interestingly, ORFV inhibited ICAM-1 up-regulation on infected but not on non-infected KC. Even interferon-γ, a potent inducer of ICAM-1, up-regulated ICAM-1 only on non-infected KC. Transfer of ORFV-free supernatant from infected to non-infected KC induced ICAM-1 on non-infected KC pointing to the involvement of soluble mediator(s). Similarly as in KC, in FB interference with ICAM-1 up-regulation by ORFV infection was also observed. In conclusion, we shed light on epidermal and dermal defense mechanisms to ORFV infection and point to a novel ICAM-1-related immune evasion mechanism of ORFV in human skin.

IgM cleavage by Streptococcus suis reduces IgM bound to the bacterial surface and is a novel complement evasion mechanism.

Streptococcus suis (S. suis) causes meningitis, arthritis and endocarditis in piglets. The aim of this study was to characterize the IgM degrading enzyme of S. suis (IdeSsuis) and to investigate the role of IgM cleavage in evasion of the classical complement pathway and pathogenesis. Targeted mutagenesis of a cysteine in the putative active center of IdeSsuis abrogated IgM cleavage completely. In contrast to wt rIdeSsuis, point mutated rIdeSsuis_C195S did not reduce complement-mediated hemolysis indicating that complement inhibition by rIdeSsuis depends on the IgM proteolytic activity. A S. suis mutant expressing IdeSsuis_C195S did not reduce IgM labeling, whereas the wt and complemented mutant showed less IgM F(ab')2 and IgM Fc antigen on the surface. IgM cleavage increased survival of S. suis in porcine blood ex vivo and mediated complement evasion as demonstrated by blood survival and C3 deposition assays including the comparative addition of rIdeSsuis and rIdeSsuis_C195S. However, experimental infection of piglets disclosed no significant differences in virulence between S. suis wt and isogenic mutants without IgM cleavage activity. This work revealed for the first time in vivo labeling of S. suis with IgM in the cerebrospinal fluid of piglets with meningitis. In conclusion, this study classifies IdeSsuis as a cysteine protease and emphasizes the role of IgM cleavage for bacterial survival in porcine blood and complement evasion though IgM cleavage is not crucial for the pathogenesis of serotype 2 meningitis.

Comparative analysis of humoral immune responses and pathologies of BALB/c and C57BL/6 wildtype mice experimentally infected with a highly virulent Rodentibacter pneumotropicus (Pasteurella pneumotropica) strain.

BACKGROUND: Mice are a natural host for Rodentibacter (R.) pneumotropicus. Despite specific monitoring, it is still one of the most important infectious agents in laboratory animals. The objective of this study was to determine the virulence of a prevalent pathotype of R. pneumotropicus and characterize the host response in a new animal model. RESULTS: Intranasal infection of C57BL/6 and BALB/c mice with a R. pneumotropicus strain (JF4Ni) bearing the genes of the three known repeats in toxin (RTX) toxins resulted in an unprecedented high mortality and morbidity above 50 and 80%, respectively. Morbidity was associated with severe weight loss as well as conjunctivitis and dyspnea. A main pathology was a catarrhal purulent to necrotic bronchopneumonia. Specific immune globuline (Ig) A was detected in tracheonasal lavages of most surviving mice which were still colonized by R. pneumotropicus. Furthermore, all surviving animals showed a distinct production of IgG antibodies. To differentiate T-helper cell (Th) 1 and Th2 immune responses we used subclasses of IgGs as indicators. Mean ratios of IgG2b to IgG1 were below 0.8 in sera drawn from both mice strains prior infection and from BALB/c mice post infection. In contrast, C57BL/6 mice had a mean IgG2b/IgG1 ratio of 1.6 post infection indicating a Th1 immune response in C57BL/6 versus a Th2 response in BALB/c mice associated with a tenfold higher bacterial load in the lung. In accordance with a Th1 response high antigen-specific IgG2c titers were detected in the majority of surviving C57BL/6 mice. CONCLUSIONS: R. pneumotropicus JF4Ni is a highly virulent strain causing severe pneumonia and septicemia after intranasal infection of C57BL/6 and BALB/c mice. Persisting infections in the two mice strains are associated with Th1 and Th2 immune responses, respectively, and differences in the bacterial burden of the lung. The described model is ideally suited for future vaccination studies using the natural host.