Search

Background: The number of periprosthetic joint infections (PJI) after total knee arthroplasty (TKA) is increasing annually. Animal models have been used to clarify their clinical characteristics and the infection mechanism of pathogenic bacteria, However, since the prosthesis design of animal models is not uniform, it is difficult to simulate the environment of clinical PJI. Objectives: To retrospect the progress on the prosthesis design of animal models of PJI after TKA and to summarize the criteria for evaluating a clinically representative model of PJI. Methods: This systematic review was reported on the basis of Systematic Reviews and Meta-Analyzes (PRISMA). Pubmed, EMbase, Cochrane Library, Web of Science, Wanfang Data and China National Knowledge Infrastructure were researched for animal models of PJI after TKA from database establishment to April 2019 according to Chinese and English retrieval words, including “periprosthetic joint infections and total knee arthroplasty,” “periprosthetic joint infections and model,” “periprosthetic joint infections and biofilm,” and “total knee arthroplasty and model.” Results: A total of 12 quantitative studies were enrolled in our study finally: 8 representative studies described prosthesis designs used in PJI animal models, 4 studies described prosthesis designs in non-infected animal models which were suitable for infection models. The major problems need to be dealed with were prosthesis, installation location, material, the function of separating the articular and medullary cavity, fixation manner, and the procedure of preserving the posterior cruciate ligament. Conclusion: A highly representative design of the animal prosthesis of PJI should meet the following criteria: the surface of the prosthesis is smooth with the formation of biofilm, composed of titanium-6Al-4V or cobalt-chromium-molybdenum alloy; prosthesis can bear weight and is highly stable; and it can connect the joint cavity and medullary cavity simultaneously. To reach a more reliable conclusion, further experiments and improvements are required. [ABSTRACT FROM AUTHOR]

Mayaro virus (MAYV) is an arbovirus that circulates in Latin America and is emerging as a potential threat to public health. Infected individuals develop Mayaro fever, a severe inflammatory disease characterized by high fever, rash, arthralgia, myalgia and headache. The disease is often associated with a prolonged arthralgia mediated by a chronic inflammation that can last months. Although the immune response against other arboviruses, such as chikungunya virus (CHIKV), dengue virus (DENV) and Zika virus (ZIKV), has been extensively studied, little is known about the pathogenesis of MAYV infection. In this study, we established models of MAYV infection in macrophages and in mice and found that MAYV can replicate in bone marrow-derived macrophages and robustly induce expression of inflammasome proteins, such as NLRP3, ASC, AIM2, and Caspase-1 (CASP1). Infection performed in macrophages derived from Nlrp3–/–, Aim2–/–, Asc–/–and Casp1/11–/–mice indicate that the NLRP3, but not AIM2 inflammasome is essential for production of inflammatory cytokines, such as IL-1β. We also determined that MAYV triggers NLRP3 inflammasome activation by inducing reactive oxygen species (ROS) and potassium efflux. In vivo infections performed in inflammasome-deficient mice indicate that NLRP3 is involved with footpad swelling, inflammation and pain, establishing a role of the NLRP3 inflammasome in the MAYV pathogenesis. Accordingly, we detected higher levels of caspase1-p20, IL-1β and IL-18 in the serum of MAYV-infected patients as compared to healthy individuals, supporting the participation of the NLRP3-inflammasome during MAYV infection in humans. [ABSTRACT FROM AUTHOR]

Klebsiella pneumoniae (Kp), one of the most common causes of healthcare-associated infections, increases patient morbidity, mortality, and hospitalization costs. Kp must acquire nutrients from the host for successful infection; however, the host is able to prevent bacterial nutrient acquisition through multiple systems. This includes the innate immune protein lipocalin 2 (Lcn2), which prevents Kp iron acquisition. To identify novel Lcn2-dependent Kp factors that mediate evasion of nutritional immunity during lung infection, we undertook an InSeq study using a pool of >20,000 transposon mutants administered to Lcn2+/+ and Lcn2-/- mice. Comparing transposon mutant frequencies between mouse genotypes, we identified the Kp citrate synthase, GltA, as potentially interacting with Lcn2, and this novel finding was independently validated. Interestingly, in vitro studies suggest that this interaction is not direct. Given that GltA is involved in oxidative metabolism, we screened the ability of this mutant to use a variety of carbon and nitrogen sources. The results indicated that the gltA mutant has a distinct amino acid auxotrophy rendering it reliant upon glutamate family amino acids for growth. Deletion of Lcn2 from the host leads to increased amino acid levels in bronchioloalveolar lavage fluid, corresponding to increased fitness of the gltA mutant in vivo and ex vivo. Accordingly, addition of glutamate family amino acids to Lcn2+/+ bronchioloalveolar lavage fluid rescued growth of the gltA mutant. Using a variety of mouse models of infection, we show that GltA is an organ-specific fitness factor required for complete fitness in the spleen, liver, and gut, but dispensable in the bloodstream. Similar to bronchioloalveolar lavage fluid, addition of glutamate family amino acids to Lcn2+/+ organ lysates was sufficient to rescue the loss of gltA. Together, this study describes a critical role for GltA in Kp infection and provides unique insight into how metabolic flexibility impacts bacterial fitness during infection. [ABSTRACT FROM AUTHOR]

Chronic wound infections are increasingly recognized to be dynamic and polymicrobial in nature, necessitating the development of wound models which reflect the complexities of infection in a non-healing wound. Wound slough isolated from human chronic wounds and transferred to mice was recently shown to create polymicrobial infection in mice, and there is potential this tool may be improved by cryogenic preservation. The purpose of this study was to investigate the application of cryogenic preservation to transferring polymicrobial communities, specifically by quantifying the effects of cryopreservation and wound microbiome transplantation. Slough from an established murine polymicrobial surgical excision model and five patients were subjected to three preservation strategies: refrigeration until infection, freezing in liquid nitrogen, or freezing in liquid nitrogen with glycerol solution prior to infection in individual mice. Four days following inoculation onto mice, wound microbiota were quantified using either culture isolation or by 16s rRNA gene community profiling and quantitative PCR. Cryogenic preservation did not significantly reduce bacterial viability. Reestablished microbial communities were significantly associated with patient of origin as well as host context (i.e., originally preserved from a patient versus mouse infection). Whereas preservation treatment did not significantly shape community composition, the transfers of microbiomes from human to mouse were characterized by reduced diversity and compositional changes. These findings indicated that changes should be expected to occur to community structure after colonization, and that compositional change is likely due to the rapid change in infection context as opposed to preservation strategy. Furthermore, species that were present in higher relative abundance in wound inoculate were more likely to colonize subsequent wounds, and wound inoculate with higher bacterial load established wound communities that were more compositionally similar. Results inform expectations for the complementation of chronic wound in vivo modeling with cryogenic preservation archives. [ABSTRACT FROM AUTHOR]

Buruli Ulcer (BU) is a cutaneous disease caused by Mycobacterium ulcerans. The pathogenesis of this disease is closely related to the secretion of the toxin mycolactone that induces extensive destruction of the skin and soft tissues. Currently, there are no effective measures to prevent the disease and, despite availability of antibiotherapy and surgical treatments, these therapeutic options are often associated with severe side effects. Therefore, it is important to develop alternative strategies for the treatment of BU. Endolysins (lysins) are phage encoded enzymes that degrade peptidoglycan of bacterial cell walls. Over the past years, lysins have been emerging as alternative antimicrobial agents against bacterial infections. However, mycobacteria have an unusual outer membrane composed of mycolylarabinogalactan-peptidoglycan. To overcome this complex barrier, some mycobacteriophages encode a lipolytic enzyme, Lysin B (LysB). In this study, we demonstrate for the first time that recombinant LysB displays lytic activity against M. ulcerans isolates. Moreover, using a mouse model of M. ulcerans footpad infection, we show that subcutaneous treatment with LysB prevented further bacterial proliferation, associated with IFN-γ and TNF production in the draining lymph node. These findings highlight the potential use of lysins as a novel therapeutic approach against this neglected tropical disease. [ABSTRACT FROM AUTHOR]

Over recent decades, Salmonella infection research has predominantly relied on murine infection models. However, in many cases the infection phenotypes of Salmonella pathovars in mice do not recapitulate human disease. For example, Salmonella Typhimurium ST313 is associated with enhanced invasive infection of immunocompromised people in Africa, but infection of mice and other animal models with ST313 have not consistently reproduced this invasive phenotype. The introduction of alternative infection models could help to improve the quality and reproducibility of pathogenesis research by facilitating larger-scale experiments. To investigate the virulence of S. Typhimurium ST313 in comparison with ST19, a combination of avian and insect disease models were used. We performed experimental infections in five lines of inbred and one line of outbred chickens, as well as in the alternative chick embryo and Galleria mellonella wax moth larvae models. This extensive set of experiments identified broadly similar patterns of disease caused by the African and global pathovariants of Salmonella Typhimurium in the chicken, the chicken embryo and insect models. A comprehensive analysis of all the chicken infection experiments revealed that the African ST313 isolate D23580 had a subtle phenotype of reduced levels of organ colonisation in inbred chickens, relative to ST19 strain 4/74. ST313 isolate D23580 also caused reduced mortality in chicken embryos and insect larvae, when compared with ST19 4/74. We conclude that these three infection models do not reproduce the characteristics of the systemic disease caused by S. Typhimurium ST313 in humans. [ABSTRACT FROM AUTHOR]

Pathogens and hosts require rapid modulation of virulence and defense mechanisms at the infection axis, but monitoring such modulations is challenging. In studying the human fungal pathogen Cryptococcus neoformans, mouse and rabbit infection models are often employed to shed light on the disease mechanisms but that may not be clinically relevant. In this study, we developed an animal infection model using the non-human primate cynomolgus monkey Macaca fascicularis. In addition, we systematically profiled and compared transcriptional responses between the infected mice and the cynomolgus monkey, using simultaneous or dual RNA next-generation sequencing. We demonstrated that there are shared but distinct transcriptional profiles between the two models following C. neoformans infection. Specifically, genes involved in immune and inflammatory responses are all upregulated. Osteoclastogenesis and insulin signaling are also significantly co-regulated in both models and disrupting an osteoclastogenesis-associated gene (OC-STAMP) or the insulin-signaling process significantly altered the host tolerance to C. neoformans. Moreover, C. neoformans was shown to activate metal sequestration, dampen the sugar metabolism, and control cell morphology during infection. Taking together, we described the development of a non-human primate model of cryptococcosis that allowed us to perform an in-depth analysis and comparison of transcriptome profiles during infections of two animal models and conceptually identify host genes important in disease responses. This study provides new insights in understanding fungal pathogenesis mechanisms that potentially facilitate the identification of novel drug targets for the treatment of cryptococcal infection. [ABSTRACT FROM AUTHOR]

Escherichia coli infection of the female reproductive tract is a significant cause of disease in humans and animals, but simple animal models are lacking. Here we report that vaginal inoculation of uropathogenic E. coli strains UTI89 and CFT073 in non-pregnant, estrogen-treated mice resulted in robust colonization of the vagina and uterine horns, whereas titers of the lab strain MG1655 were significantly lower. Non-estrogenized mice also became colonized, but there was more variation in titers. A dose of 104 colony-forming units (CFU) UTI89 was sufficient to result in colonization in all estrogenized mice, and we also observed bacterial transfer between inoculated and uninoculated estrogenized cage mates. UTI89 infection led to inflammation and leukocyte infiltration into the uterine horns as evidenced by tissue histology. Flow cytometry experiments revealed that neutrophil, monocyte and eosinophil populations were significantly increased in infected uterine horns. This model is a simple way to study host-pathogen interactions in E. coli vaginal colonization and uterine infection. There are immediate implications for investigators studying urinary tract infection using mouse models, as few E. coli are required to achieve reproductive colonization, resulting in an additional, underappreciated mucosal reservoir. [ABSTRACT FROM AUTHOR]

Ancestral state reconstruction models use genetic data to characterize a group of organisms’ common ancestor. These models have been applied to salmonellosis outbreaks to estimate the number of transmissions between different animal species that share similar geographical locations, with animal host as the state. However, as far as we are aware, no studies have validated these models for outbreak analysis. In this study, salmonellosis outbreaks were simulated using a stochastic Susceptible-Infected-Recovered model, and the host population and transmission parameters of these simulated outbreaks were estimated using Bayesian ancestral state reconstruction models (discrete trait analysis (DTA) and structured coalescent (SC)). These models were unable to accurately estimate the number of transmissions between the host populations or the amount of time spent in each host population. The DTA model was inaccurate because it assumed the number of isolates sampled from each host population was proportional to the number of individuals infected within each host population. The SC model was inaccurate possibly because it assumed that each host population's effective population size was constant over the course of the simulated outbreaks. This study highlights the need for phylodynamic models that can take into consideration factors that influence the characteristics and behavior of outbreaks, e.g. changing effective population sizes, variation in infectious periods, intra-population transmissions, and disproportionate sampling of infected individuals. [ABSTRACT FROM AUTHOR]

Dengue is the most widespread vector-borne viral disease caused by dengue virus (DENV) for which there are no safe, effective drugs approved for clinical use. Here, by using sequential antigen panning of a yeast antibody library derived from healthy donors against the DENV envelop protein domain III (DIII) combined with depletion by an entry defective DIII mutant, we identified a cross-reactive human monoclonal antibody (mAb), m366.6, which bound with high affinity to DENV DIII from all four DENV serotypes. Immunogenetic analysis indicated that m366.6 is a germline-like mAb with very few somatic mutations from the closest VH and Vλ germline genes. Importantly, we demonstrated that it potently neutralized DENV both in vitro and in the mouse models of DENV infection without detectable antibody-dependent enhancement (ADE) effect. The epitope of m366.6 was mapped to the highly conserved regions on DIII, which may guide the design of effective dengue vaccine immunogens. Furthermore, as the first germline-like mAb derived from a naïve antibody library that could neutralize all four DENV serotypes, the m366.6 can be a tool for exploring mechanisms of DENV infection, and is a promising therapeutic candidate. [ABSTRACT FROM AUTHOR]

Background: The species of the Rickettsia genus are separated into four groups: the ancestral group, typhus group, transitional group and spotted fever group. Rickettsia parkeri, a spotted fever group Rickettsia, has been reported across the American continents as infecting several tick species and is associated with a relatively mild human disease characterized by eschar formation at the tick feeding site, regional lymphadenopathy, fever, myalgia and rash. Currently, there are several mouse models that provide good approaches to study the acute lethal disease caused by Rickettsia, but these models can only be performed in an animal biosafety level 3 laboratory. We present an alternative mouse model for acute lethal rickettsial disease, using R. parkeri Atlantic Rainforest strain and C3H/HeN mice, with the advantage that this model can be studied in an animal biosafety level 2 laboratory. Principal findings: In the C3H/HeN mouse model, we determined that infection with 1x106 and 1x107 viable R. parkeri Atlantic Rainforest strain organisms produced dose-dependent severity, whereas infection with 1x108 viable bacteria resulted in a lethal illness. The animals became moribund on day five or six post-infection. The lethal disease was characterized by ruffled fur, erythema, labored breathing, decreased activity, and hunched posture, which began on day three post-infection (p.i.) and coincided with the peak bacterial loads. Significant splenomegaly (on days three and five p.i.), neutrophilia (on days three and five p.i.), and thrombocytopenia (on days one, three and five p.i.) were observed. Significance: Since R. parkeri is used at biosafety level 2, the greatest advantage of this inbred mouse model is the ability to investigate immunity and pathogenesis of rickettsiosis with all the tools available at biosafety level 2. [ABSTRACT FROM AUTHOR]

Zika virus (ZIKV) is a mosquito-borne flavivirus associated with microcephaly and other neurological disorders in infants born to infected mothers. Despite being declared an international emergency by the World Health Organization, very little is known about the mechanisms of ZIKV pathogenesis or the long-term consequences of maternal ZIKV infection in the affected offspring, largely due to the lack of appropriate rodent models. To address this issue, our lab has developed a working model of prenatal ZIKV infection in rats. In this study, we infected immune competent pregnant female rats with 105–107 PFU of ZIKV (PRVABC59, Puerto Rico/Human/Dec 2015) in order to examine its pathogenesis in the dams and pups. We examined the febrile response and sickness behavior in the dams, in addition to neonatal mortality, microglia morphology, cortical organization, apoptosis, and brain region-specific volumes in the offspring. Here, we demonstrate that pregnant and non-pregnant female rats have a distinct febrile response to ZIKV infection. Moreover, prenatal ZIKV infection increased cell death and reduced tissue volume in the hippocampus and cortex in the neonatal offspring. For the first time, we demonstrate the efficacy and validity of an immunocompetent rat model for maternal ZIKV infection that results in significant brain malformations in the neonatal offspring. [ABSTRACT FROM AUTHOR]

Bourbon virus (BRBV) is an emerging tick-borne RNA virus in the orthomyxoviridae family that was discovered in 2014. Although fatal human cases of BRBV have been described, little is known about its pathogenesis, and no antiviral therapies or vaccines exist. We obtained serum from a fatal case in 2017 and successfully recovered the second human infectious isolate of BRBV. Next-generation sequencing of the St. Louis isolate of BRBV (BRBV-STL) showed >99% nucleotide identity to the original reference isolate. Using BRBV-STL, we developed a small animal model to study BRBV-STL tropism in vivo and evaluated the prophylactic and therapeutic efficacy of the experimental antiviral drug favipiravir against BRBV-induced disease. Infection of Ifnar1-/- mice lacking the type I interferon receptor, but not congenic wild-type animals, resulted in uniformly fatal disease 6 to 10 days after infection. RNA in situ hybridization and viral yield assays demonstrated a broad tropism of BRBV-STL with highest levels detected in liver and spleen. In vitro replication and polymerase activity of BRBV-STL were inhibited by favipiravir. Moreover, administration of favipiravir as a prophylaxis or as post-exposure therapy three days after infection prevented BRBV-STL-induced mortality in immunocompromised Ifnar1-/- mice. These results suggest that favipiravir may be a candidate treatment for humans who become infected with BRBV. [ABSTRACT FROM AUTHOR]

Candida albicans, a species of fungi, can thrive in diverse niches of its mammalian hosts; it is a normal resident of the GI tract and mucosal surfaces but it can also enter the bloodstream and colonize internal organs causing serious disease. The ability of C. albicans to thrive in these different host environments has been attributed, at least in part, to its ability to assume different morphological forms. In this work, we examine one such morphological change known as white-opaque switching. White cells are the default state of C. albicans, and most animal studies have been carried out exclusively with white cells. Here, we compared the proliferation of white and opaque cells in two murine models of infection and also monitored, using specially constructed strains, switching between the two states in the host. We found that white cells outcompeted opaque cells in many niches; however, we show for the first time that in some organs (specifically, the heart and spleen), opaque cells competed favorably with white cells and, when injected on their own, could colonize these organs. In environments where the introduced white cells outcompeted the introduced opaque cells, we observed high rates of opaque-to-white switching. We did not observe white-to-opaque switching in any of the niches we examined. [ABSTRACT FROM AUTHOR]

Chagas disease, caused by the parasite Trypanosoma cruzi, develops into chronic Chagas’ cardiomyopathy in ~30% of infected individuals, characterized by conduction disorders, arrhythmias, heart failure, and even sudden cardiac death. Current anti-parasitic treatments are plagued by significant side effects and poor efficacy in the chronic phase of disease; thus, there is a pressing need for new treatment options. A therapeutic vaccine could bolster the protective TH1-mediated immune response, thereby slowing or halting the progression of chronic Chagas’ cardiomyopathy. Prior work in mice has demonstrated therapeutic efficacy of a Tc24 recombinant protein vaccine in the acute phase of Chagas disease. However, it is anticipated that humans will be vaccinated therapeutically when in the chronic phase of disease. This study investigates the therapeutic efficacy of a vaccine prototype containing recombinant protein Tc24, formulated with an emulsion containing the Toll-like receptor 4 agonist E6020 as an immunomodulatory adjuvant in a mouse model of chronic T. cruzi infection. Among outbred ICR mice vaccinated during chronic T. cruzi infection, there is a significant increase in the number of animals with undetectable systemic parasitemia (60% of vaccinated mice compared to 0% in the sham vaccine control group), and a two-fold reduction in cardiac fibrosis over the control group. The vaccinated mice produce a robust protective TH1-biased immune response to the vaccine, as demonstrated by a significant increase in antigen-specific IFNγ-production, the number of antigen-specific IFNγ-producing cells, and IgG2a antibody titers. Importantly, therapeutic vaccination significantly reduced cardiac fibrosis in chronically infected mice. This is a first study demonstrating therapeutic efficacy of the prototype Tc24 recombinant protein and E6020 stable emulsion vaccine against cardiac fibrosis in a mouse model of chronic T. cruzi infection. [ABSTRACT FROM AUTHOR]

Infections of humans and livestock with African trypanosomes are treated with drugs introduced decades ago that are not always fully effective and often have severe side effects. Here, the trypanosome haptoglobin-haemoglobin receptor (HpHbR) has been exploited as a route of uptake for an antibody-drug conjugate (ADC) that is completely effective against Trypanosoma brucei in the standard mouse model of infection. Recombinant human anti-HpHbR monoclonal antibodies were isolated and shown to be internalised in a receptor-dependent manner. Antibodies were conjugated to a pyrrolobenzodiazepine (PBD) toxin and killed T. brucei in vitro at picomolar concentrations. A single therapeutic dose (0.25 mg/kg) of a HpHbR antibody-PBD conjugate completely cured a T. brucei mouse infection within 2 days with no re-emergence of infection over a subsequent time course of 77 days. These experiments provide a demonstration of how ADCs can be exploited to treat protozoal diseases that desperately require new therapeutics. [ABSTRACT FROM AUTHOR]

Baloxavir marboxil (BXM) is an orally available small molecule inhibitor of cap-dependent endonuclease (CEN), an essential enzyme in the initiation of mRNA synthesis of influenza viruses. In the present study, we evaluated the efficacy of BXM against influenza virus infection in mouse models. Single-day oral administration of BXM completely prevented mortality due to infection with influenza A and B virus in mice. Moreover, 5-day repeated administration of BXM was more effective for reducing mortality and body weight loss in mice infected with influenza A virus than oseltamivir phosphate (OSP), even when the treatment was delayed up to 96 hours post infection (p.i.). Notably, administration of BXM, starting at 72 hours p.i. led to significant decrease in virus titers of >2-log10 reduction compared to the vehicle control within 24 hours after administration. Virus reduction in the lung was significantly greater than that observed with OSP. In addition, profound and sustained reduction of virus titer was observed in the immunocompromised mouse model without emergence of variants possessing treatment-emergent amino acid substitutions in the target protein. In our immunocompetent and immunocompromised mouse models, delayed treatment with BXM resulted in rapid and potent reduction in infectious virus titer and prevention of signs of influenza infection, suggesting that BXM could extend the therapeutic window for patients with influenza virus infection regardless of the host immune status. [ABSTRACT FROM AUTHOR]

Background: We demonstrated therapeutic nonequivalence of “bioequivalent” generics for meropenem, but there is no data with generics of other carbapenems. Methods: One generic product of imipenem-cilastatin was compared with the innovator in terms of in vitro susceptibility testing, pharmaceutical equivalence, pharmacokinetic (PK) and pharmacodynamic (PD) equivalence in the neutropenic mouse thigh, lung and brain infection models. Both pharmaceutical forms were then subjected to analytical chemistry assays (LC/MS). Results and conclusion: The generic product had 30% lower concentration of cilastatin compared with the innovator of imipenem-cilastatin. Regarding the active pharmaceutical ingredient (imipenem), we found no differences in MIC, MBC, concentration or potency or AUC, confirming equivalence in terms of in vitro activity. However, the generic failed therapeutic equivalence in all three animal models. Its Emax against S. aureus in the thigh model was consistently lower, killing from 0.1 to 7.3 million less microorganisms per gram in 24 hours than the innovator (P = 0.003). Against K. pneumoniae in the lung model, the generic exhibited a conspicuous Eagle effect fitting a Gaussian equation instead of the expected sigmoid curve of the Hill model. In the brain infection model with P. aeruginosa, the generic failed when bacterial growth was >4 log10 CFU/g in 24 hours, but not if it was less than 2.5 log10 CFU/g. These large differences in the PD profile cannot be explained by the lower concentration of cilastatin, and rather suggested a failure attributable to the imipenem constituent of the generic product. Analytical chemistry assays confirmed that, besides having 30% less cilastatin, the generic imipenem was more acidic, less stable, and exhibited four different degradation masses that were absent in the innovator. [ABSTRACT FROM AUTHOR]

The emergence of Zika virus (ZIKV) in the New World has led to more than 200,000 human infections. Perinatal infection can cause severe neurological complications, including fetal and neonatal microcephaly, and in adults there is an association with Guillain-Barré syndrome (GBS). ZIKV is transmitted to humans by Aedes sp. mosquitoes, yet little is known about its enzootic cycle in which transmission is thought to occur between arboreal Aedes sp. mosquitos and non-human primates. In the 1950s and ‘60s, several bat species were shown to be naturally and experimentally susceptible to ZIKV with acute viremia and seroconversion, and some developed neurological disease with viral antigen detected the brain. Because of ZIKV emergence in the Americas, we sought to determine susceptibility of Jamaican fruit bats (Artibeus jamaicensis), one of the most common bats in the New World. Bats were inoculated with ZIKV PRVABC59 but did not show signs of disease. Bats held to 28 days post-inoculation (PI) had detectable antibody by ELISA and viral RNA was detected by qRT-PCR in the brain, saliva and urine in some of the bats. Immunoreactivity using polyclonal anti-ZIKV antibody was detected in testes, brain, lung and salivary glands plus scrotal skin. Tropism for mononuclear cells, including macrophages/microglia and fibroblasts, was seen in the aforementioned organs in addition to testicular Leydig cells. The virus likely localized to the brain via infection of Iba1+ macrophage/microglial cells. Jamaican fruit bats, therefore, may be a useful animal model for the study of ZIKV infection. This work also raises the possibility that bats may have a role in Zika virus ecology in endemic regions, and that ZIKV may pose a wildlife disease threat to bat populations. [ABSTRACT FROM AUTHOR]

Several Zika virus (ZIKV) vaccines designed to elicit protective antibody (Ab) responses are currently under rapid development, but the underlying mechanisms that control the magnitude and quality of the Ab response remain unclear. Here, we investigated the CD4+ T cell response to primary intravenous and intravaginal infection with ZIKV. Using the LysMCre+Ifnar1fl/fl (myeloid type I IFN receptor-deficient) C57BL/6 mouse models, we identified six I-Ab-restricted ZIKV epitopes that stimulated CD4+ T cells with a predominantly cytotoxic Th1 phenotype in mice primed with ZIKV. Intravenous and intravaginal infection with ZIKV effectively induced follicular helper and regulatory CD4+ T cells. Treatment of mice with a CD4+ T cell-depleting Ab reduced the plasma cell, germinal center B cell, and IgG responses to ZIKV without affecting the CD8+ T cell response. CD4+ T cells were required to protect mice from a lethal dose of ZIKV after infection intravaginally, but not intravenously. However, adoptive transfer and peptide immunization experiments showed a role for memory CD4+ T cells in ZIKV clearance in mice challenged intravenously. These results demonstrate that CD4+ T cells are required mainly for the generation of a ZIKV-specific humoral response but not for an efficient CD8+ T cell response. Thus, CD4+ T cells could be important mediators of protection against ZIKV, depending on the infection or vaccination context. [ABSTRACT FROM AUTHOR]

Enterohemorrhagic Escherichia coli (EHEC) colonize intestinal epithelium by generating characteristic attaching and effacing (AE) lesions. They are lysogenized by prophage that encode Shiga toxin 2 (Stx2), which is responsible for severe clinical manifestations. As a lysogen, prophage genes leading to lytic growth and stx2 expression are repressed, whereas induction of the bacterial SOS response in response to DNA damage leads to lytic phage growth and Stx2 production both in vitro and in germ-free or streptomycin-treated mice. Some commensal bacteria diminish prophage induction and concomitant Stx2 production in vitro, whereas it has been proposed that phage-susceptible commensals may amplify Stx2 production by facilitating successive cycles of infection in vivo. We tested the role of phage induction in both Stx production and lethal disease in microbiome-replete mice, using our mouse model encompassing the murine pathogen Citrobacter rodentium lysogenized with the Stx2-encoding phage Φstx2dact. This strain generates EHEC-like AE lesions on the murine intestine and causes lethal Stx-mediated disease. We found that lethal mouse infection did not require that Φstx2dact infect or lysogenize commensal bacteria. In addition, we detected circularized phage genomes, potentially in the early stage of replication, in feces of infected mice, confirming that prophage induction occurs during infection of microbiota-replete mice. Further, C. rodentium (Φstx2dact) mutants that do not respond to DNA damage or express stx produced neither high levels of Stx2 in vitro or lethal infection in vivo, confirming that SOS induction and concomitant expression of phage-encoded stx genes are required for disease. In contrast, C. rodentium (Φstx2dact) mutants incapable of prophage genome excision or of packaging phage genomes retained the ability to produce Stx in vitro, as well as to cause lethal disease in mice. Thus, in a microbiome-replete EHEC infection model, lytic induction of Stx-encoding prophage is essential for lethal disease, but actual phage production is not. [ABSTRACT FROM AUTHOR]

Background: The mortality from cryptococcal meningitis remains high, despite the availability of antiretroviral therapy (ART) and amphotericin-based fungal regimens. The role of neutrophils in cryptococcosis is controversial. Our objective was to examine the association between blood neutrophil counts and outcomes in terms of mortality, the incidence of bacterial infections (including Mycobacterium tuberculosis) and hospitalization among HIV-infected patients presenting with cryptococcal meningitis. Methods: We used data from participants from the Cryptococcal Optimal ART Timing (COAT) trial (2010–2012; Uganda and South Africa) and the Adjunctive Sertraline for Treatment of Cryptococcal Meningitis (ASTRO-CM) trial (2013–2017; Uganda). We estimated 30-day mortality risk with Cox proportional hazards models by baseline neutrophil counts (a) on a continuous scale and (b) with indicators for both relatively high (> 3,500 cells/mm3) and low (≤ 1,000 cells/mm3) counts. Follow-up neutrophil counts from the COAT trial were used to examine the time-dependent association of neutrophil counts with 12-month mortality and rehospitalization. Results: 801 participants had an absolute neutrophil value at meningitis diagnosis. The median baseline absolute neutrophil count was 2100 cells/mm3 (IQR, 1400 to 3300 cells/mm3). Baseline neutrophil count was positively associated with 30-day mortality (adjusted hazard ratio = 1.09, 95%CI, 1.04–1.13, per 1000 cells/mm3 increase; p<0.001). Baseline absolute neutrophil counts ≤ 1000 cells/mm3 did not have increased risk of 30-day mortality compared to those with baseline neutrophils of 1001–3500 cells/mm3; however, baseline >3500 cells/mm3 had significantly increased risk, with an adjusted hazard ratio of 1.85(95%CI, 1.40–2.44; p<0.001). Among the COAT participants with follow-up neutrophil data, there was a strong association between time-updated neutrophil count and 12-month mortality (adjusted hazard ratio = 1.16, 95% CI 1.09–1.24; p<0.001. Conclusion: Higher blood neutrophil counts in HIV-infected patients with cryptococcal meningitis were associated with mortality. Neutrophils role requires further investigation as to whether this may be a mediator directly contributing to mortality or merely a marker of underlying pathologies that increase mortality risk. [ABSTRACT FROM AUTHOR]

Clostridium difficile is a Gram-positive, spore-forming anaerobic bacterium that infects the colon, causing symptoms ranging from infectious diarrhea to fulminant colitis. In the last decade, the number of C. difficile infections has dramatically risen, making it the leading cause of reported hospital acquired infection in the United States. Bacterial toxins produced during C. difficile infection (CDI) damage host epithelial cells, releasing erythrocytes and heme into the gastrointestinal lumen. The reactive nature of heme can lead to toxicity through membrane disruption, membrane protein and lipid oxidation, and DNA damage. Here we demonstrate that C. difficile detoxifies excess heme to achieve full virulence within the gastrointestinal lumen during infection, and that this detoxification occurs through the heme-responsive expression of the eme ctivated ransporter system (HatRT). Heme-dependent transcriptional activation of hatRT was discovered through an RNA-sequencing analysis of C. difficile grown in the presence of a sub-toxic concentration of heme. HatRT is comprised of a TetR family transcriptional regulator (hatR) and a major facilitator superfamily transporter (hatT). Strains inactivated for hatR or hatT are more sensitive to heme toxicity than wild-type. HatR binds heme, which relieves the repression of the hatRT operon, whereas HatT functions as a heme efflux pump. In a murine model of CDI, a strain inactivated for hatT displayed lower pathogenicity in a toxin-independent manner. Taken together, these data suggest that HatR senses intracellular heme concentrations leading to increased expression of the hatRT operon and subsequent heme efflux by HatT during infection. These results describe a mechanism employed by C. difficile to relieve heme toxicity within the host, and set the stage for the development of therapeutic interventions to target this bacterial-specific system. [ABSTRACT FROM AUTHOR]

Chronic livestock diseases cause large financial loss and affect animal health and welfare. Controlling these diseases mostly requires precise information on both individual animal and population dynamics to inform the farmer’s decisions, but even successful control programmes do by no means assure elimination. Mathematical models provide opportunities to test different control and elimination options rather than implementing them in real herds, but these models require robust parameter estimation and validation. Fitting these models to data is a difficult task due to heterogeneities in livestock processes. In this paper, we develop an infectious disease modeling framework for a livestock disease (paratuberculosis) that is caused by Mycobacterium avium subsp. paratuberculosis (MAP). Infection with MAP leads to reduced milk production, pregnancy rates, and slaughter value and increased culling rates in cattle and causes significant economic losses to the dairy industry. These economic effects are particularly important motivations in the control and elimination of MAP. In this framework, an individual-based model (IBM) of a dairy herd was built and MAP infection dynamics was integrated. Once the model produced realistic dynamics of MAP infection, we implemented an evaluation method by fitting it to data from three dairy herds from the Northeast region of the US. The model fitting exercises used least-squares and parameter space searching methods to obtain the best-fitted values of selected parameters. The best set of parameters were used to model the effect of interventions. The results show that the presented model can complement real herd statistics where the intervention strategies suggest a reduction in MAP prevalence without elimination. Overall, this research not only provides a complete model for MAP infection dynamics in a dairy herd but also offers a method for estimating parameters by fitting IBM models. [ABSTRACT FROM AUTHOR]

Foot-and-mouth disease (FMD) is a highly infectious transboundary disease that affects domestic and wild cloven-hoofed animal species. The aim of this review was to identify and critically assess some modelling techniques for FMD that are well supported by scientific evidence from the literature with a focus on their use in African countries where the disease remains enzootic. In particular, this study attempted to provide a synopsis of the relative strengths and weaknesses of these models and their relevance to FMD prevention policies. A literature search was conducted to identify quantitative and qualitative risk assessments for FMD, including studies that describe FMD risk factor modelling and spatiotemporal analysis. A description of retrieved papers and a critical assessment of the modelling methods, main findings and their limitations were performed. Different types of models have been used depending on the purpose of the study and the nature of available data. The most frequently identified factors associated with the risk of FMD occurrence were the movement (especially uncontrolled animal movement) and the mixing of animals around water and grazing points. Based on the qualitative and quantitative risk assessment studies, the critical pathway analysis showed that the overall risk of FMDV entering a given country is low. However, in some cases, this risk can be elevated, especially when illegal importation of meat and the movement of terrestrial livestock are involved. Depending on the approach used, these studies highlight shortcomings associated with the application of models and the lack of reliable data from endemic settings. Therefore, the development and application of specific models for use in FMD endemic countries including Africa is encouraged. [ABSTRACT FROM AUTHOR]

Although Blastocystis spp. infect probably more than 1 billion people worldwide, their clinical significance is still controversial and their pathophysiology remains poorly understood. In this study, we describe a protocol for an efficient and reproducible model of chronic infection in rats, laying the groundwork for future work to evaluate the pathogenic potential of this parasite. In our experimental conditions, we were unable to infect rats using vacuolar forms of an axenically cultivated ST4 isolate, but we successfully established chronic infections of 4 week-old rats after oral administration of both ST3 and ST4 purified cysts isolated from human stool samples. The infection protocol was also applied to 4 week-old C57BL/9, BALB/C and C3H mice, but any mouse was found to be infected by Blastocystis. Minimal cyst inoculum required for rat infection was higher with ST3 (105) than with ST4 (102). These results were confirmed by co-housing experiments highlighting a higher contagious potential of ST4 in rats compared to ST3. Finally, experiments mimicking fecal microbiota transfer from infected to healthy animals showed that Blastocystis spp. could easily infect a new host, even though its intestinal microbiota is not disturbed. In conclusion, our results provide a well-documented and robust rat model of Blastocystis chronic infection, reproducing “natural” infection. This model will be of great interest to study host parasite interactions and to better evaluate clinical significance of Blastocystis. [ABSTRACT FROM AUTHOR]

Severe fever with thrombocytopenia syndrome (SFTS), caused by SFTS virus (SFTSV), is a viral hemorrhagic fever with a high case fatality rate. Favipiravir was reported to be effective in the treatment of SFTSV infection in vivo in type I interferon receptor knockout (IFNAR−/−) mice at treatment dosages of both 60 mg/kg/day and 300 mg/kg/day for a duration of 5 days. In this study, the efficacy of favipiravir at dosages of 120 mg/kg/day and 200 mg/kg/day against SFTSV infection in an IFNAR−/− mouse infection model was investigated. IFNAR−/− mice were subcutaneously infected with SFTSV at a 1.0 × 106 50% tissue culture infectious dose followed by twice daily administration of favipiravir, comprising a total dose of either 120 mg/kg/day or 200 mg/kg/day. The treatment was initiated either immediately post infection or at predesignated time points post infection. Neutralizing antibodies in the convalescent-phase mouse sera was examined by the pseudotyped VSV system. All mice treated with favipiravir at dosages of 120 mg/kg/day or 200 mg/kg/day survived when the treatment was initiated at no later than 4 days post infection. A decrease in body weight of mice was observed when the treatment was initiated at 3–4 days post infection. Furthermore, all control mice died. The body weight of mice did not decrease when treatment with favipiravir was initiated immediately post infection at dosages of 120 mg/kg/day and 200 mg/kg/day. Neutralizing antibodies were detected in the convalescent-phase mouse sera. Similar to the literature-reported peritoneal administration of favipiravir at 300 mg/kg/day, the oral administration of favipiravir at dosages of 120 mg/kg/day and 200 mg/kg/day to IFNAR−/− mice infected with SFTSV was effective. [ABSTRACT FROM AUTHOR]

Background: Zika virus (ZIKV) infection causes diseases ranging from acute self-limiting febrile illness to life-threatening Guillain–Barré Syndrome and other neurological disorders in adults. Cumulative evidence suggests an association between ZIKV infection and microcephaly in newborn infants. Given the host-range restrictions of the virus, a susceptible animal model infected by ZIKV must be developed for evaluation of vaccines and antivirals. In this study, we propose a convenient mouse model for analysis of neurological disorders caused by ZIKV. Methodology: Six-day-old immunocompetent ICR suckling mice were used in the experiment. Different inoculum virus concentrations, challenge routes, and challenge times were assessed. Viremic dissemination was determined in the liver, spleen, kidney, and brain through Western blot assay, plaque assay, absolute quantification real-time PCR, and histological observation. Azithromycin, a well-characterized anti-ZIKV compound, was used to evaluate the ICR suckling mouse model for antiviral testing. Conclusions: Signs of illness and neurological disease and high mortality rate were observed in mice injected with ZIKV intracerebrally (102 to 105) and intraperitoneally (103 to 105). Viremic dissemination was observed in the liver, spleen, kidney, and brain. ZIKV transmitted, rapid replicated, and induced monocyte infiltration into the brain approximately 5 to 6 days post inoculum. Azithromycin conferred protection against ZIKV-caused neurological and life-threatening diseases. The developed model of ZIKV infection and disease can be used for screening drugs against ZIKV and discovering the underlying mechanism of ZIKV pathogenesis. [ABSTRACT FROM AUTHOR]

The whooping cough agent Bordetella pertussis coordinately regulates the expression of its virulence factors with the two-component system BvgAS. In laboratory conditions, specific chemical modulators are used to trigger phenotypic modulation of B. pertussis from its default virulent Bvg+ phase to avirulent Bvg- or intermediate Bvgi phases, in which no virulence factors or only a subset of them are produced, respectively. Whether phenotypic modulation occurs in the host remains unknown. In this work, recombinant B. pertussis strains harboring BvgS variants were tested in a mouse model of infection and analyzed using transcriptomic approaches. Recombinant BP-BvgΔ65, which is in the Bvgi phase by default and can be up-modulated to the Bvg+ phase in vitro, could colonize the mouse nose but was rapidly cleared from the lungs, while Bvg+-phase strains colonized both organs for up to four weeks. These results indicated that phenotypic modulation, which might have restored the full virulence capability of BP-BvgΔ65, does not occur in mice or is temporally or spatially restricted and has no effect in those conditions. Transcriptomic analyses of this and other recombinant Bvgi and Bvg+-phase strains revealed that two distinct ranges of virulence gene expression allow colonization of the mouse nose and lungs, respectively. We also showed that a recombinant strain expressing moderately lower levels of the virulence genes than its wild type parent was as efficient at colonizing both organs. Altogether, genetic modifications of BvgS generate a range of phenotypic phases, which are useful tools to decipher host-pathogen interactions. [ABSTRACT FROM AUTHOR]

Zika virus (ZIKV) has gained worldwide attention since it emerged, and a global effort is underway to understand the correlates of protection and develop diagnostics to identify rates of infection. As new therapeutics and vaccine approaches are evaluated in clinical trials, additional effort is focused on identifying the adaptive immune correlates of protection against ZIKV disease. To aid in this endeavor we have begun to dissect the role of CD4+T cells in the protection against neuroinvasive ZIKV disease. We have identified an important role for CD4+T cells in protection, demonstrating that in the absence of CD4+T cells mice have more severe neurological sequela and significant increases in viral titers in the central nervous system (CNS). The transfer of CD4+T cells from ZIKV immune mice protect type I interferon receptor deficient animals from a lethal challenge; showing that the CD4+T cell response is necessary and sufficient for control of ZIKV disease. Using a peptide library spanning the complete ZIKV polyprotein, we identified both ZIKV-encoded CD4+T cell epitopes that initiate immune responses, and ZIKV specific CD4+T cell receptors that recognize these epitopes. Within the ZIKV antigen-specific TCRβ repertoire, we uncovered a high degree of diversity both in response to a single epitope and among different mice responding to a CD4+T cell epitope. Overall this study identifies a novel role for polyfunctional and polyclonal CD4+T cells in providing protection against ZIKV infection and highlights the need for vaccines to develop robust CD4+T cell responses to prevent ZIKV neuroinvasion and limit replication within the CNS. [ABSTRACT FROM AUTHOR]

Mycoplasma gallisepticum is a serious pathogen for poultry that causes chronic respiratory disease in chickens. Increased embryonic mortality, as well as reduced weight gain and egg production have been found in infected chickens, which can lead to considerable economic losses in poultry production. Increased antibiotic resistance compromises the use of tetracyclines, macrolides and quinolones in the farm environment. In the present study, danofloxacin concentrations were simulated below the MIC99, between the MIC99 and MPC (the mutant prevention concentration), and above the MPC in an in vitro dynamic model against M. gallisepticum. The relationship between the simulated danofloxacin pharmacokinetics, pharmacodynamics (PK/PD) parameters and development of resistance for M. gallisepticum was explored based on the available data obtained from various dosing regimens in the in vitro model. Danofloxacin concentration, counts of viable cell and susceptibility were determined during the experiment. The mutations in gyrA, gyrB, parC and parE as well as efflux pumps were examined. The MIC of danofloxacin against M. gallisepticum was increased when drug concentrations were between the lower and upper boundaries of the mutant selection window. The upper boundary of the selection window in vitro was estimated as a Cmax/MPC value of 1. The lower boundary was estimated as Cmax/MPC value of 0.05. Both in terms of the MIC and resistance frequency, M. gallisepticum resistance was developed when danofloxacin concentrations fell inside the mutant selection window (ratios of Cmax to MPC between 0.05 and 1). The single mutation in gyrA (Ser-83→Arg) was found in all mutants, while double mutations in gyrA and parC (Ala-64→Ser) were observed only in the mutant with the highest MIC. In addition, no change of susceptibility in the mutants was observed in the presence of reserpine and carbonyl cyanide 3-chlorophenylhydrazone (CCCP). This suggested that ATP-binding cassette superfamily (ABC transporter) and major facilitator superfamily (MFS transporter) did not play a role in danofloxacin efflux. [ABSTRACT FROM AUTHOR]

Clostridium difficile is a Gram-positive spore-former bacterium and the leading cause of nosocomial antibiotic-associated diarrhea that can culminate in fatal colitis. During the infection, C. difficile produces metabolically dormant spores, which persist in the host and can cause recurrence of the infection. The surface of C. difficile spores seems to be the key in spore-host interactions and persistence. The proteome of the outermost exosporium layer of C. difficile spores has been determined, identifying two cysteine-rich exosporium proteins, CdeC and CdeM. In this work, we explore the contribution of both cysteine-rich proteins in exosporium integrity, spore biology and pathogenesis. Using targeted mutagenesis coupled with transmission electron microscopy we demonstrate that both cysteine rich proteins, CdeC and CdeM, are morphogenetic factors of the exosporium layer of C. difficile spores. Notably, cdeC, but not cdeM spores, exhibited defective spore coat, and were more sensitive to ethanol, heat and phagocytic cells. In a healthy colonic mucosa (mouse ileal loop assay), cdeC and cdeM spore adherence was lower than that of wild-type spores; while in a mouse model of recurrence of the disease, cdeC mutant exhibited an increased infection and persistence during recurrence. In a competitive infection mouse model, cdeC mutant had increased fitness over wild-type. Through complementation analysis with FLAG fusion of known exosporium and coat proteins, we demonstrate that CdeC and CdeM are required for the recruitment of several exosporium proteins to the surface of C. difficile spores. CdeC appears to be conserved exclusively in related Peptostreptococcaeace family members, while CdeM is unique to C. difficile. Our results sheds light on how CdeC and CdeM affect the biology of C. difficile spores and the assembly of the exosporium layer and, demonstrate that CdeC affect C. difficile pathogenesis. [ABSTRACT FROM AUTHOR]

There are very few articles in the literature describing continuous models of bacterial infections that mimic disease pathogenesis in humans and animals without using separate cohorts of animals at each stage of disease. In this work, we developed bioluminescent mouse models of partial-thickness scald wound infection and sepsis that mimic disease pathogenesis in humans and animals using a recombinant luciferase-expressing Staphylococcus aureus strain (Xen29). Two days post-scald wound infection, mice were treated twice daily with a 2% topical mupirocin ointment for 7 days. For sepsis experiments, mice were treated intraperitoneally with 6 mg/kg daptomycin 2 h and 6 h post-infection and time to moribund monitored for 72 h. Consistent bacterial burden data were obtained from individual mice by regular photon intensity quantification on a Xenogen IVIS Lumina XRMS Series III biophotonic imaging system, with concomitant significant reduction in photon intensities in drug-treated mice. Post-mortem histopathological examination of wounds and bacterial counts in blood correlated closely with disease severity and total flux obtained from Xen29. The bioluminescent murine models provide a refinement to existing techniques of multiple bacterial enumeration during disease pathogenesis and promote animal usage reduction. The models also provide an efficient and information-rich platform for preclinical efficacy evaluation of new drug classes for treating acute and chronic human and animal bacterial infections. [ABSTRACT FROM AUTHOR]

Streptococcus suis serotype 2 is an important porcine bacterial pathogen and zoonotic agent responsible for sudden death, septic shock, and meningitis. However, serotype 2 strains are heterogeneous, composed of a multitude of sequence types (STs) whose distribution greatly varies worldwide. Of the virulence factors presently described for S. suis, the capsular polysaccharide (CPS) is a critical factor implicated in a multitude of functions, including in impairment of phagocytosis and innate immune cell activation by masking underlying bacterial components. However, these roles have been described using Eurasian ST1 and ST7 strains, which greatly differ from North American ST25 strains. Consequently, the capacity of the CPS to mask surface antigens and putative virulence factors in non-Eurasian strains remains unknown. Herein, the role of the S. suis serotype 2 CPS of a prototype intermediate virulent North American ST25 strain, in comparison with that of a virulent European ST1 strain, with regards to interactions with dendritic cells, as well as virulence during the systemic phase of infection, was evaluated. Results demonstrated that the CPS remains critical for virulence and development of clinical disease regardless of strain background, due to its requirement for survival in blood. However, its role in the interactions with dendritic cells is strain-dependent. Consequently, certain key characteristics associated with the CPS are not necessarily applicable to all S. suis serotype 2 strains. This indicates that though certain factors may be important for S. suis serotype 2 virulence, strain background could be as determining, reiterating the need in using strains from varying backgrounds in order to better characterize the S. suis pathogenesis. [ABSTRACT FROM AUTHOR]

Background: The lumpy skin disease virus (LSDV) is a dsDNA virus belonging to the Poxviridae family and the Capripoxvirus genus. Lumpy skin diseases (LSD) is a highly contagious transboundary disease in cattle producing major economic losses. In 2014, the disease was first reported in the European Union (in Cyprus); it was then reported in 2015 (in Greece) and has spread through different Balkan countries in 2016. Indirect vector transmission is predominant at small distances, but transmission between distant herds and between countries usually occurs through movements of infected cattle or through vectors found mainly in animal trucks. Methods and principal findings: In order to estimate the threat for France due to the introduction of vectors found in animal trucks (cattle or horses) from at-risk countries (Balkans and neighbours), a quantitative import risk analysis (QIRA) model was developed according to the international standard. Using stochastic QIRA modelling and combining experimental/field data and expert opinion, the yearly risk of LSDV being introduced by stable flies (Stomoxys calcitrans), that travel in trucks transporting animals was between 6 x 10−5 and 5.93 x 10−3 with a median value of 89.9 x 10−5; it was mainly due to the risk related to insects entering farms in France from vehicles transporting cattle from the at-risk area. The risk related to the transport of cattle going to slaughterhouses or the transport of horses was much lower (between 2 x 10−7 and 3.73 x 10−5 and between 5 x 10−10 and 3.95 x 10−8 for cattle and horses, respectively). The disinsectisation of trucks transporting live animals was important to reduce this risk. Conclusion and significance: The development of a stochastic QIRA made it possible to quantify the risk of LSD being introduced in France through the import of vectors that travel in trucks transporting animals. This tool is of prime importance because the LSD situation in the Balkans is continuously changing. Indeed, this model can be updated to process new information on vectors and the changing health situation, in addition to new data from the TRAde Control and Expert System (TRACES, EU database). This model is easy to adapt to different countries and to other vectors and diseases. [ABSTRACT FROM AUTHOR]

Campylobacter infections are among the leading bacterial causes of diarrhea and of ‘environmental enteropathy’ (EE) and growth failure worldwide. However, the lack of an inexpensive small animal model of enteric disease with Campylobacter has been a major limitation for understanding its pathogenesis, interventions or vaccine development. We describe a robust standard mouse model that can exhibit reproducible bloody diarrhea or growth failure, depending on the zinc or protein deficient diet and on antibiotic alteration of normal microbiota prior to infection. Zinc deficiency and the use of antibiotics create a niche for Campylobacter infection to establish by narrowing the metabolic flexibility of these mice for pathogen clearance and by promoting intestinal and systemic inflammation. Several biomarkers and intestinal pathology in this model also mimic those seen in human disease. This model provides a novel tool to test specific hypotheses regarding disease pathogenesis as well as vaccine development that is currently in progress. [ABSTRACT FROM AUTHOR]

Burkholderia pseudomallei, a gram-negative intracellular bacillus, is the causative agent of a tropical infectious disease called melioidosis. Bacterial ATP-binding cassette (ABC) transporters import and export a variety of molecules across bacterial cell membranes. At present, their significance in B. pseudomallei pathogenesis is poorly understood. We report here characterization of the BPSL1039-1040 ABC transporter. B. pseudomallei cultured in M9 medium supplemented with nitrate, demonstrated that BPSL1039-1040 is involved in nitrate transport for B. pseudomallei growth under anaerobic, but not aerobic conditions, suggesting that BPSL1039-1040 is functional under reduced oxygen tension. In addition, a nitrate reduction assay supported the function of BPSL1039-1040 as nitrate importer. A bpsl1039-1040 deficient mutant showed reduced biofilm formation as compared with the wild-type strain (P = 0.027) when cultured in LB medium supplemented with nitrate under anaerobic growth conditions. This reduction was not noticeable under aerobic conditions. This suggests that a gradient in oxygen levels could regulate the function of BPSL1039-1040 in B. pseudomallei nitrate metabolism. Furthermore, the B. pseudomallei bpsl1039-1040 mutant had a pronounced effect on plaque formation (P < 0.001), and was defective in intracellular survival in both non-phagocytic (HeLa) and phagocytic (J774A.1 macrophage) cells, suggesting reduced virulence in the mutant strain. The bpsl1039-1040 mutant was found to be attenuated in a BALB/c mouse intranasal infection model. Complementation of the bpsl1039-1040 deficient mutant with the plasmid-borne bpsl1039 gene could restore the phenotypes observed. We propose that the ability to acquire nitrate for survival under anaerobic conditions may, at least in part, be important for intracellular survival and has a contributory role in the pathogenesis of B. pseudomallei. [ABSTRACT FROM AUTHOR]

The cellular prion protein, designated PrPC, is a membrane glycoprotein expressed abundantly in brains and to a lesser extent in other tissues. Conformational conversion of PrPC into the amyloidogenic isoform is a key pathogenic event in prion diseases. However, the physiological functions of PrPC remain largely unknown, particularly in non-neuronal tissues. Here, we show that PrPC is expressed in lung epithelial cells, including alveolar type 1 and 2 cells and bronchiolar Clara cells. Compared with wild-type (WT) mice, PrPC-null mice (Prnp0/0) were highly susceptible to influenza A viruses (IAVs), with higher mortality. Infected Prnp0/0 lungs were severely injured, with higher inflammation and higher apoptosis of epithelial cells, and contained higher reactive oxygen species (ROS) than control WT lungs. Treatment with a ROS scavenger or an inhibitor of xanthine oxidase (XO), a major ROS-generating enzyme in IAV-infected lungs, rescued Prnp0/0 mice from the lethal infection with IAV. Moreover, Prnp0/0 mice transgenic for PrP with a deletion of the Cu-binding octapeptide repeat (OR) region, Tg(PrPΔOR)/Prnp0/0 mice, were also highly susceptible to IAV infection. These results indicate that PrPC has a protective role against lethal infection with IAVs through the Cu-binding OR region by reducing ROS in infected lungs. Cu content and the activity of anti-oxidant enzyme Cu/Zn-dependent superoxide dismutase, SOD1, were lower in Prnp0/0 and Tg(PrPΔOR)/Prnp0/0 lungs than in WT lungs. It is thus conceivable that PrPC functions to maintain Cu content and regulate SOD1 through the OR region in lungs, thereby reducing ROS in IAV-infected lungs and eventually protecting them from lethal infection with IAVs. Our current results highlight the role of PrPC in protection against IAV infection, and suggest that PrPC might be a novel target molecule for anti-influenza therapeutics. [ABSTRACT FROM AUTHOR]

The molecular mechanisms underlying susceptibility to severe respiratory syncytial virus (RSV) infection remain poorly understood. Herein, we report on the role of osteopontin (OPN) in regulation of RSV infection in human epithelial cells and how interleukin-1 beta (IL-1β), a cytokine secreted soon after RSV infection, when persistently expressed can induce OPN expression leading to increased viral infection. We first compared OPN expression in two human epithelial cell lines: HEK-293 and HEp-2. In contrast to HEp-2, HEK-293 expresses low levels of pro-caspase-1 resulting in decreased IL-1β expression in response to RSV infection. We found a correlation between low IL-1β levels and a delay in induction of OPN expression in RSV-infected HEK-293 cells compared to HEp-2. This phenomenon could partially explain the high susceptibility of HEp-2 cells to RSV infection versus the moderate susceptibility of HEK-293 cells. Also, HEK-293 cells expressing low levels of pro-caspase-1 exhibit decreased IL-1β expression and delayed OPN expression in response to RSV infection. HEK-293 cells incubated with human rIL-1β showed a dose-dependent increase in OPN expression upon RSV infection. Also, incubation with rOPN increased RSV viral load. Moreover, HEp-2 cells or mice infected with a mucogenic RSV strain RSV-L19F showed elevated levels of OPN in contrast to mice infected with the laboratory RSV strain rA2. This correlated with elevated levels of OPN following infection with RSV-L19F compared to rA2. Together, these results demonstrate that increased OPN expression is regulated in part by IL-1β, and the interplay between IL-1β and OPN signaling may play a pivotal role in the spread of RSV infection. [ABSTRACT FROM AUTHOR]

Bacterial spores play an important role in disease initiation, transmission and persistence. In some species, the exosporium forms the outermost structure of the spore and provides the first point of contact between the spore and the environment. The exosporium may also be involved in spore adherence, protection and germination. Clostridium sordellii is a highly lethal, spore forming pathogen that causes soft-tissue infections, enteritis and toxic-shock syndrome. Despite the importance of C. sordellii spores in disease, spore proteins from this bacterium have not been defined or interrogated functionally. In this study, we identified the C. sordellii outer spore proteome and two of the identified proteins, CsA and CsB, were characterised using a genetic and phenotypic approach. Both proteins were essential for the correct formation and positioning of the C. sordellii spore coat and exosporium. The absence of CsA reduced sporulation levels and increased spore sensitivity to heat, sodium hydroxide and hydrochloric acid. By comparison, CsB was required for normal levels of spore adherence to cervical, but not vaginal, cells, with csB mutant spores having increased adherence properties. The establishment of a mouse infection model of the gastrointestinal tract for C. sordellii allowed the role of CsA and CsB to be interrogated in an infected host. Following the oral administration of spores to mice, the wild-type strain efficiently colonized the gastrointestinal tract, with the peak of bacterial numbers occurring at one day post-infection. Colonization was reduced by two logs at four days post-infection. By comparison, mice infected with the csB mutant did not show a reduction in bacterial numbers. We conclude that C. sordellii outer spore proteins are important for the structural and functional integrity of spores. Furthermore, outer spore proteins are required for wild-type levels of colonization during infection, possibly as a result of the role that the proteins play in spore structure and morphology. [ABSTRACT FROM AUTHOR]

Zika virus (ZIKV) infection during human pregnancy may cause diverse and serious congenital defects in the developing fetus. Previous efforts to generate animal models of human ZIKV infection and clinical symptoms often involved manipulating mice to impair their Type I interferon (IFN) signaling, thereby allowing enhanced infection and vertical transmission of virus to the embryo. Here, we show that even pregnant mice competent to generate Type I IFN responses that can limit ZIKV infection nonetheless develop profound placental pathology and high frequency of fetal demise. We consistently found that maternal ZIKV exposure led to placental pathology and that ZIKV RNA levels measured in maternal, placental or embryonic tissues were not predictive of the pathological effects seen in the embryos. Placental pathology included trophoblast hyperplasia in the labyrinth, trophoblast giant cell necrosis in the junctional zone, and loss of embryonic vessels. Our findings suggest that, in this context of limited infection, placental pathology rather than embryonic/fetal viral infection may be a stronger contributor to adverse pregnancy outcomes in mice. Our finding demonstrates that in immunocompetent mice, direct viral infection of the embryo is not essential for fetal demise. Our immunologically unmanipulated pregnancy mouse model provides a consistent and easily measurable congenital abnormality readout to assess fetal outcome, and may serve as an additional model to test prophylactic and therapeutic interventions to protect the fetus during pregnancy, and for studying the mechanisms of ZIKV congenital immunopathogenesis. [ABSTRACT FROM AUTHOR]

Few animal models of Zika virus (ZIKV) infection have incorporated arthropod-borne transmission. Here, we establish an Aedes aegypti mosquito model of ZIKV infection of mice, and demonstrate altered vector competency among three strains, (Orlando, ORL, Ho Chi Minh, HCM, and Patilas, PAT). All strains acquired ZIKV in their midguts after a blood meal from infected mice, but ZIKV transmission only occurred in mice fed upon by HCM, and to a lesser extent PAT, but not ORL, mosquitoes. This defect in transmission from ORL or PAT mosquitoes was overcome by intrathoracic injection of ZIKV into mosquito. Genetic analysis revealed significant diversity among these strains, suggesting a genetic basis for differences in ability for mosquito strains to transmit ZIKV. The intrathoracic injection mosquito-mouse transmission model is critical to understanding the influence of mosquitoes on ZIKV transmission, infectivity and pathogenesis in the vertebrate host, and represents a natural transmission route for testing vaccines and therapeutics. (152 words) [ABSTRACT FROM AUTHOR]

Backgrounds: HY12 viruses are enteroviruses recently isolated from cattle characterized by severe respiratory and digestive disease with high morbidity and mortality in China. While the viruses exhibit unique biological and molecular characters distinct from known enterovirus E, the pathogenicity and viral pathogenesis remains largely unknown. Methods: Neonatal mice of Balb/C, ICR, and Kunming strain are infected with HY12 to determine the susceptible mouse strain. The minimal infection dose, the virus infection routes, the pathogenicity and tissue tropism for HY12 were determined by infecting susceptible mice with HY12 viruses, and confirmed by different approaches including virus isolation and recovery, virus detection, histopathology, and immunohistochemistry. Results: A murine model for HY12 infection was successfully established and employed to investigate the pathogenicity of HY12 viruses. ICR mouse strain is the most susceptible strain for HY12 infection with a minimal infective dose as 2×106TCID50/mouse. HY12 viruses have the capability of infecting ICR suckling mice via all infection routes including intranasal administration, oral administration, intraperitoneal injection, subcutaneous injection, and intramuscular injection, which are confirmed by the isolation and recovery of viruses from HY12-infected mice; detection of viruses by RT-PCR; observations of pathological lesions and inflammatory cell infiltrations in the intestine, lung, liver, and brain; uncovering of HY12 virus antigens in majority of tissues, especially in intestine, lung, and infected brain of mice by immunohistochemistry assay. Conclusions: A neonatal murine model for HY12 infection is successfully established for determining the susceptible mouse strain, the minimal infective dose, the infection route, the viral pathogenicity and the tropism of HY12, thus providing an invaluable model system for elucidating the pathogenesis of HY12 viruses and the elicited immunity. [ABSTRACT FROM AUTHOR]

Dengue virus (DENV) infection imposes enormous health and economic burden worldwide with no approved treatment. Several small molecules, including lovastatin, celgosivir, balapiravir and chloroquine have been tested for potential anti-dengue activity in clinical trials; none of these have demonstrated a protective effect. Recently, based on identification and characterization of cross-serotype neutralizing antibodies, there is increasing attention on the potential for dengue immunotherapy. Here, we tested the ability of VIS513, an engineered cross-neutralizing humanized antibody targeting the DENV E protein domain III, to overcome antibody-enhanced infection and high but brief viremia, which are commonly encountered in dengue patients, in various in vitro and in vivo models. We observed that VIS513 efficiently neutralizes DENV at clinically relevant viral loads or in the presence of enhancing levels of DENV immune sera. Single therapeutic administration of VIS513 in mouse models of primary infection or lethal secondary antibody-enhanced infection, reduces DENV titers and protects from lethal infection. Finally, VIS513 administration does not readily lead to resistance, either in cell culture systems or in animal models of dengue infection. The findings suggest that rapid viral reduction during acute DENV infection with a monoclonal antibody is feasible. [ABSTRACT FROM AUTHOR]

Hyperbaric oxygen therapy (HBO) is applied very successfully in treatment of various diseases such as chronic wounds. It has been already suggested as adjunctive treatment option for osteitis by immune- and fracture modulating effects. This study evaluates the importance of HBO in an early implant-associated localized osteitis caused by Staphylococcus aureus (SA) compared to the standard therapy. In a standardized murine model the left femur of 120 BALB/c mice were osteotomized and fixed by a titanium locking plate. Osteitis has been induced with a defined amount of SA into the fracture gap. Debridément and lavages were progressed on day 7, 14, 28 and 56 to determine the local bacterial growth and the immune reaction. Hyperbaric oxygen (2 ATA, 90%) was applied for 90 minutes on day 7 to 21 for those mice allocated to HBO therapy. To evaluate the effect of HBO therapy the following groups were analyzed: Two sham-groups (12 mice / group) with and without HBO therapy, two osteotomy groups (24 mice / group) with plate osteosynthesis of the femur with and without HBO therapy, and two osteotomy SA infection groups (24 mice / group) with and without HBO therapy. Fracture healing was also quantified on day 7, 14, 28 and 56 by a.p. x-ray and bone healing markers from blood samples. Progression of infection was assessed by estimation of colony-forming units (CFU) and immune response was analyzed by determination of polymorphonuclear neutrophils (PMN), Interleukin (IL) - 6, and the circulating free DNA (cfDNA) in lavage samples. Osteitis induced significantly higher IL-6, cfDNA- and PMN-levels in the lavage samples (on day 7 and 14, each p < 0.05). HBO-therapy did not have a significant influence on the CFU and immune response compared to the standard therapy (each p > 0.05). At the same time HBO-therapy was associated with a delayed bone healing assessed by x-ray radiography and a higher rate of non-union until day 28. In conclusion, osteitis led to significantly higher bacterial count and infection parameters. HBO-therapy neither had a beneficial influence on local infection nor on immune response or fracture healing compared to the standard therapy in an osteitis mouse model. [ABSTRACT FROM AUTHOR]

HIV synergy with sexually transmitted co-infections is well-documented in the clinic. Co-infection with Neisseria gonorrhoeae in particular, increases genital HIV shedding and mucosal transmission. However, no animal model of co-infection currently exists to directly explore this relationship or to bridge the gap in understanding between clinical and in vitro studies of this interaction. This study aims to test the feasibility of using a humanized mouse model to overcome this barrier. Combining recent in vivo modelling advancements in both HIV and gonococcal research, we developed a co-infection model by engrafting immunodeficient NSG mice with human CD34+ hematopoietic stem cells to generate humanized mice that permit both systemic HIV infection and genital N. gonorrhoeae infection. Systemic plasma and vaginal lavage titres of HIV were measured in order to assess the impact of gonococcal challenge on viral plasma titres and genital shedding. Engrafted mice showed human CD45+ leukocyte repopulation in blood and mucosal tissues. Systemic HIV challenge resulted in 104−105 copies/mL of viral RNA in blood by week 4 post-infection, as well as vaginal shedding of virus. Subsequent gonococcal challenge resulted in unchanged plasma HIV levels but higher viral shedding in the genital tract, which reflects published clinical observations. Thus, human CD34+ stem cell-transplanted NSG mice represent an experimentally tractable animal model in which to study HIV shedding during gonococcal co-infection, allowing dissection of molecular and immunological interactions between these pathogens, and providing a platform to assess future therapeutics aimed at reducing HIV transmission. [ABSTRACT FROM AUTHOR]

The efficacies of many new T cell vaccines rely on generating large populations of long-lived pathogen-specific effector memory CD8 T cells. However, it is now increasingly recognized that prior infection history impacts on the host immune response. Additionally, the order in which these infections are acquired could have a major effect. Exploiting the ability to generate large sustained effector memory (i.e. inflationary) T cell populations from murine cytomegalovirus (MCMV) and human Adenovirus-subtype (AdHu5) 5-beta-galactosidase (Ad-lacZ) vector, the impact of new infections on pre-existing memory and the capacity of the host’s memory compartment to accommodate multiple inflationary populations from unrelated pathogens was investigated in a murine model. Simultaneous and sequential infections, first with MCMV followed by Ad-lacZ, generated inflationary populations towards both viruses with similar kinetics and magnitude to mono-infected groups. However, in Ad-lacZ immune mice, subsequent acute MCMV infection led to a rapid decline of the pre-existing Ad-LacZ-specific inflating population, associated with bystander activation of Fas-dependent apoptotic pathways. However, responses were maintained long-term and boosting with Ad-lacZ led to rapid re-expansion of the inflating population. These data indicate firstly that multiple specificities of inflating memory cells can be acquired at different times and stably co-exist. Some acute infections may also deplete pre-existing memory populations, thus revealing the importance of the order of infection acquisition. Importantly, immunization with an AdHu5 vector did not alter the size of the pre-existing memory. These phenomena are relevant to the development of adenoviral vectors as novel vaccination strategies for diverse infections and cancers. (241 words) [ABSTRACT FROM AUTHOR]

Background: Cases of acute respiratory tract infection caused by Pteropine orthoreovirus (PRV) of the genus Orthoreovirus (family: Reoviridae) have been reported in Southeast Asia, where it was isolated from humans and bats. It is possible that PRV-associated respiratory infections might be prevalent in Southeast Asia. The clinical course of PRV is not fully elucidated. Methods: The virulence, pathology, and pathogenesis of two PRV strains, a human-borne PRV strain (isolated from a patient, who returned to Japan from Bali, Indonesia in 2007) and a bat-borne PRV (isolated from a bat [Eonycteris spelaea] in the Philippines in 2013) were investigated in BALB/c mice using virological, pathological, and immunological study methods. Results: The intranasal inoculation of BALB/c mice with human-borne PRV caused respiratory infection. In addition, all mice with immunity induced by pre-inoculation with a non-lethal dose of PRV were completely protected against lethal PRV infection. Mice treated with antiserum with neutralizing antibody activity after inoculation with a lethal dose of PRV showed a reduced fatality rate. In this mouse model, bat-borne PRV caused respiratory infection similar to human-borne PRV. PRV caused lethal respiratory disease in an animal model of PRV infection, in which BALB/c mice were used. Conclusions: The BALB/c mouse model might help to accelerate research on the virulence of PRV and be useful for evaluating the efficacy of therapeutic agents and vaccines for the treatment and prevention of PRV infection. PRV was shown for the first time to be a causative virus of respiratory disease on the basis of Koch’s postulations by the additional demonstration that PRV caused respiratory disease in mice through their intranasal inoculation with PRV. [ABSTRACT FROM AUTHOR]

The matrix proteins of Staphylococcus aureus biofilm have not been well defined. Previous efforts to identify these proteins were performed using in vitro systems. Here we use a proteomic approach to identify biofilm matrix proteins directly from infected bone implants using a rat model of orthopedic implant-associated S. aureus infection. Despite heavy presence of host proteins, a total of 28 and 105 S. aureus proteins were identified during acute infection and chronic infection, respectively. Our results show that biofilm matrix contains mostly intracellular cytoplasmic proteins and, to a much less extent, extracellular and cell surface-associated proteins. Significantly, leukocidins were identified in the biofilm matrix during chronic infection, suggesting S. aureus is actively attacking the host immune system even though they are protected within the biofilm. The presence of two surface-associated proteins, Ebh and SasF, in the infected bone tissue during acute infection was confirmed by immunohistochemistry. In addition, a large number of host proteins were found differentially expressed in response to S. aureus biofilm formed on bone implants. [ABSTRACT FROM AUTHOR]

Background: Opisthorchiasis is a parasitic infection caused by the liver flukes of the Opisthorchiidae family. Both experimental and epidemiological data strongly support a role of these parasites in the etiology of the hepatobiliary pathologies and an increased risk of intrahepatic cholangiocarcinoma. Understanding a functional link between the infection and hepatobiliary pathologies requires a detailed description a host-parasite interaction on different levels of biological regulation including the metabolic response on the infection. The last one, however, remains practically undocumented. Here we are describing a host response on Opisthorchiidae infection using a metabolomics approach and present the first exploratory metabolomics study of an experimental model of O. felineus infection. Methodology and Principal findings: We conducted a Nuclear Magnetic Resonance (NMR) based longitudinal metabolomics study involving a cohort of 30 animals with two degrees of infection and a control group. An exploratory analysis shows that the most noticeable trend (30% of total variance) in the data was related to the gender differences. Therefore further analysis was done of each gender group separately applying a multivariate extension of the ANOVA—ASCA (ANOVA simultaneous component analysis). We show that in the males the infection specific time trends are present in the main component (43.5% variance), while in the females it is presented only in the second component and covers 24% of the variance. We have selected and annotated 24 metabolites associated with the observed effects and provided a physiological interpretation of the findings. Conclusions: The first exploratory metabolomics study an experimental model of O. felineus infection is presented. Our data show that at early stage of infection a response of an organism unfolds in a gender specific manner. Also main physiological mechanisms affected appear rather nonspecific (a status of the metabolic stress) the data provides a set of the hypothesis for a search of the more specific metabolic markers of the Opisthorchiidae infection. [ABSTRACT FROM AUTHOR]