Natural immunity or resistance to pathogens most often relies on the genetic make-up of the host. In a LEW rat model of refractoriness to toxoplasmosis, we previously identified on chromosome 10 the Toxo1 locus that directs toxoplasmosis outcome and controls parasite spreading by a macrophage-dependent mechanism. Now, we narrowed down Toxo1 to a 891 kb interval containing 29 genes syntenic to human 17p13 region. Strikingly, Toxo1 is included in a haplotype block strictly conserved among all refractory rat strains. The sequencing of Toxo1 in nine rat strains (5 refractory and 4 susceptible) revealed resistant-restricted conserved polymorphisms displaying a distribution gradient that peaks at the bottom border of Toxo1, and highlighting the NOD-like receptor, Nlrp1a, as a major candidate. The Nlrp1 inflammasome is known to trigger, upon pathogen intracellular sensing, pyroptosis programmed-cell death involving caspase-1 activation and cleavage of IL-1β. Functional studies demonstrated that the Toxo1-dependent refractoriness in vivo correlated with both the ability of macrophages to restrict T. gondii growth and a T. gondii-induced death of intracellular parasites and its host macrophages. The parasite-induced cell death of infected macrophages bearing the LEW-Toxo1 alleles was found to exhibit pyroptosis-like features with ROS production, the activation of caspase-1 and IL1-β secretion. The pharmacological inactivation of caspase-1 using YVAD and Z-VAD inhibitors prevented the death of both intravacuolar parasites and host non-permissive macrophages but failed to restore parasite proliferation. These findings demonstrated that the Toxo1-dependent response of rat macrophages to T. gondii infection may trigger two pathways leading to the control of parasite proliferation and the death of parasites and host macrophages. The NOD-like receptor NLRP1a/Caspase-1 pathway is the best candidate to mediate the parasite-induced cell death. These data represent new insights towards the identification of a major pathway of innate resistance to toxoplasmosis and the prediction of individual resistance. Toxoplasmosis is a ubiquitous parasitic infection causing a wide spectrum of diseases. It is usually asymptomatic but can lead to severe ocular and neurological disorders. The host factors that determine natural resistance to toxoplasmosis are yet poorly characterized. Among the animal models to study susceptibility to toxoplasmosis, rats develop like humans a subclinical chronic infection. The finding of a total resistance in the LEW rat strain has allowed genetic studies leading to the identification of Toxo1, a unique locus that controls the outcome of toxoplasmosis. In this report, a panel of recombinant inbred rat strains was used to genetically reduce the Toxo1 locus, on chromosome 10, to a limited region containing 29 genes. This locus is highly conserved among five resistant, by comparison to four susceptible, rat strains, indicating that refractoriness to toxoplasmosis could be predicted. The Toxo1-controlled refractoriness depends on the ability of macrophages to restrict parasite proliferation and the rapid death of both T. gondii and host macrophages in vitro. The NOD-like receptor NLRP1a/Caspase-1 pathway is the best candidate to mediate the parasite-induced cell death. Our data represent new insights towards the identification of a major pathway of innate immunity that protects from toxoplasmosis.

TheChlamydialesorder is composed of nine families of strictly intracellular bacteria. Among them,Chlamydia trachomatis,C. pneumoniae, andC. psittaciare established human pathogens, whereasWaddlia chondrophilaandParachlamydia acanthamoebaehave emerged as new pathogens in humans. However, despite their medical importance, their biodiversity and ecology remain to be studied. Even if arthropods and, particularly, ticks are well known to be vectors of numerous infectious agents such as viruses and bacteria, virtually nothing is known about ticks and chlamydia. This study investigated the prevalence ofChlamydiaein ticks. Specifically, 62,889Ixodes ricinusticks, consolidated into 8,534 pools, were sampled in 172 collection sites throughout Switzerland and were investigated using pan-Chlamydialesquantitative PCR (qPCR) for the presence ofChlamydialesDNA. Among the pools, 543 (6.4%) gave positive results and the estimated prevalence in individual ticks was 0.89%. Among those pools with positive results, we obtained 16S rRNA sequences for 359 samples, allowing classification ofChlamydialesDNA at the family level. A high level of biodiversity was observed, since six of the nine families belonging to theChlamydialesorder were detected. Those most common wereParachlamydiaceae(33.1%) andRhabdochlamydiaceae(29.2%). “UnclassifiedChlamydiales” (31.8%) were also often detected. Thanks to the huge amount ofChlamydialesDNA recovered from ticks, this report opens up new perspectives on further work focusing on whole-genome sequencing to increase our knowledge aboutChlamydialesbiodiversity. This report of an epidemiological study also demonstrates the presence ofChlamydia-related bacteria withinIxodes ricinusticks and suggests a role for ticks in the transmission of and as a reservoir for these emerging pathogenicChlamydia-related bacteria.

Ticks are vectors of several microorganisms responsible for infectious diseases in human and animals, such as Anaplasma phagocytophilum and Coxiella burnetii. In this study, we investigated the prevalence of these two bacteria in 62 889 Ixodes ricinus ticks in selected regions covering all Switzerland. A high prevalence of 11.9% of A. phagocytophilum DNA was observed by real-time PCR on 8534 pools of ticks. This pool prevalence corresponds to an estimated prevalence of 1.71% in individual tick. A total of 144 of the 171 collection sites (84.2%) were positive for the presence of A. phagocytophilum, and these sites were homogenously distributed throughout Switzerland. Such prevalence and geographical distribution underline the risk of human and animal exposure to A. phagocytophilum and highlight the need to assess the epidemiology and clinical diagnosis of human and animal anaplasmosis in Switzerland. However, DNA of C. burnetii was never found in any tick pool. This absence suggests a very low role of I. ricinus ticks as vector and reservoir of C. burnetii in Switzerland, and it supports previous reports demonstrating the role of sheep and goats in the epidemiology of Q fever. However, considering its pathogenic potential, it is necessary to keep monitoring for the possible reemergence of this bacterium in ticks in the future.

Hydrogels (gels) are attractive tools for tissue engineering and regenerative medicine due to their potential for drug delivery and ECM-like composition. In this study, we use rheology to characterize GelMA/alginate gels loaded with human platelet lysate (PL). We then characterize these gels from a physicochemical perspective and evaluate their ability to transport PL proteins, their pore size, and their rate of degradation. Finally, their biocompatibility is evaluated. We describe how alginate changes the mechanical behavior of the gels from elastic to viscoelastic after ionic (calcium-mediated) crosslinking. In addition, we report the release of ~90% of PL proteins from the gels and relate it to the degradation profile of the gels. Finally, we evaluated the biocompatibility of the gels. Thus, the developed gels represent attractive substrates for both cell studies and as bioactive materials.

Filopodia are finger-like actin-rich protrusions that extend from the cell surface and are important for cell–cell communication and pathogen internalization. The small size and transient nature of filopodia combined with shared usage of actin regulators within cells confounds attempts to identify filopodial proteins. Here, we used phage display phenotypic screening to isolate antibodies that alter the actin morphology of filopodia-like structures (FLS) in vitro. We found that all of the antibodies that cause shorter FLS interact with SNX9, an actin regulator that binds phosphoinositides during endocytosis and at invadopodia. In cells, we discover SNX9 at specialized filopodia in Xenopus development and that SNX9 is an endogenous component of filopodia that are hijacked by Chlamydia entry. We show the use of antibody technology to identify proteins used in filopodia-like structures, and a role for SNX9 in filopodia.

Waddlia chondrophila, an obligate intracellular bacterium belonging to the Chlamydiales order, is considered as an emerging pathogen. Some clinical studies highlighted a possible role of W. chondrophila in bronchiolitis, pneumonia and miscarriage. This pathogenic potential is further supported by the ability of W. chondrophila to infect and replicate within human pneumocytes, macrophages and endometrial cells. Considering that W. chondrophila might be a causative agent of respiratory tract infection, we developed a mouse model of respiratory tract infection to get insight into the pathogenesis of W. chondrophila. Following intranasal inoculation of 2 x 10⁸ W. chondrophila, mice lost up to 40% of their body weight, and succumbed rapidly from infection with a death rate reaching 50% at day 4 post-inoculation. Bacterial loads, estimated by qPCR, increased from day 0 to day 3 post-infection and decreased thereafter in surviving mice. Bacterial growth was confirmed by detecting dividing bacteria using electron microscopy, and living bacteria were isolated from lungs 14 days post-infection. Immunohistochemistry and histopathology of infected lungs revealed the presence of bacteria associated with pneumonia characterized by an important multifocal inflammation. The high inflammatory score in the lungs was associated with the presence of pro-inflammatory cytokines in both serum and lungs at day 3 post-infection. This animal model supports the role of W. chondrophila as an agent of respiratory tract infection, and will help understanding the pathogenesis of this strict intracellular bacterium.