Non-infected Animals Research Articles

Rosuvastatin revert memory impairment and anxiogenic-like effect in mice infected with the chronic ME-49 strain of Toxoplasma gondii.

The aim of this study was to investigate the effect of rosuvastatin treatment on memory impairment, and anxiogenic-like effects in mice chronically infected with Toxoplasma gondii. For this, Balb/c mice were infected orally with chronic ME-49 strain of Toxoplasma gondii. Oral treatment with rosuvastatin (40mg/kg/day) started on the 51st day post-infection and was performed daily for 21 days. After completion of treatment, anxiety-like effects and locomotion were investigated in the open field (OF) test, whereas novel object recognition (NOR) test was used for evaluation of short- and long-term memory. At the end of the experiments, the brain was collected for Toxoplasma gondii DNA quantification and histopathological analysis. Infection with ME-49 strain decreased the time spent in the center of OF, indicating an anxiogenic effect, without affecting total and peripheral locomotion. Rosuvastatin treatment inhibited the change in the center time. Besides, pharmacological treatment increased total and central locomotion in both non-infected and infected animals. Infection also impaired both short- and long-term memory in the NOR test, and these effects were reverted by rosuvastatin treatment. In addition to effects in behavioral changes, rosuvastatin also reduced parasite load in the brain and attenuated signs of brain inflammation such as perivascular cuffs, inflammatory cell infiltration and tissue damage. These findings indicate for the first time the efficacy of rosuvastatin in treatment of memory impairment and anxiogenic effect evoked by infection with Toxoplasma gondii. These effects might be mediated by reduced cyst load, which in turn decrease inflammation and damage in the brain.

Tri-Model Therapy: Combining Macrocyclic Lactone, Piperazine Derivative and Herbal Preparation in Treating Humpsore in Cattle.

Stephanofilariasis or humpsore is a chronic parasitic dermatitis of cattle. Various treatment regimens were attempted in the past but were found to be partially effective. Here, we claim a successful treatment regime using an FDA-approved macrocyclic lactone, a piperazine derivative, and an herbal preparation. Twenty-four cattle (18 affected and 6 unaffected) were selected and divided into Gr 1: positive control (infected without treatment; n = 6), Gr 2: treatment group (infected with treatment with ivermectin; n = 6), Gr 3: treatment group (infected with treatment with tri-model therapy including ivermectin, diethylcarbamazine citrate, and an herbal ointment, n = 6), and Gr 4: negative control (non-infected animals; n = 6). In Gr 2 and Gr 3, treatment to the ailing animals were given for 30 days. Lesion was significantly reduced in day 15 of post-treatment and completely healed on day 30 of post-treatment in Gr 3. Tri-model therapy recorded significant improvement in the surface area of the sore as compared to ivermectin administration alone. Antioxidants were increased and malondialdehyde (MDA) and cortisol concentrations were decreased significantly (p < 0.05) in Gr 3 than in untreated control group at day 14, 21 and 28. Histopathological changes in infected animals were characterized by parakeratotic hyperkeratosis along with presence of nucleated keratinocytes. There were infiltrations of polymorphonuclear cells specially eosinophils along with a few monomorphonuclear cells. Microfilarial organism was observed beneath the epidermis, which was surrounded by fibrocytes and infiltrated cells. In the tri-model-treated animal after recovery, the skin revived a normal architecture. Therefore, tri-model therapy has the potential to cure humpsore.

Relationship of Porphyromonas gingivalis and Alzheimer's disease: a systematic review of pre-clinical studies.

This study aimed to analyze the following PICO question: Are animals infected with Porphyromonas gingivalis (P. gingivalis) or bacterial lipopolysaccharide (Pg-LPS) more affected by neurodegeneration, similar to the pathogenesis generated by Alzheimer's disease(AD), compared with non-infected animals? Databases PubMed, Lilacs, SciELO, Science Direct, Scopus, Web of Science, and Cochrane were searched for pre-clinical in vivo studies in which mice were infected with P. gingivalis or received Pg-LPS, in order to assess the brain tissue and cognitive impairment. No limit for date or publication language was imposed and this study was registered at the International Prospective Register of Systematic Reviews (PROSPERO), with nine articles included. Syrcle's protocol was used to evaluate bias in the selected studies. Nine articles were included. Infection by P. gingivalis or the administration of Pg-LPS increased the production of the inflammatory mediators, TNF-α(tumor necrosis factor-alpha), IL-6(interleukin-6), and IL-1β(interleukin-1beta), augmented Aβ(amyloid beta) production, and activated the complement system, causing inflammation, brain tissue degeneration, and cognitive impairment, consistent with the damage in AD. Infection by P. gingivalis and Pg-LPS administration appears to be in relation with the pathogenesis of AD by activating the complement cascade, increasing Aβ production and augmenting pro-inflammatory cytokine expression, causing age-dependent brain inflammation, neuroinflammation, and neurodegeneration. Taking into account the importance of holistic treatment in the dental office, this study focuses on identifying highly prevalent oral diseases, such as periodontal disease, as risk factors for the aggravation of degenerative diseases in the elderly population.

PLASMA VITAMIN C LEVELS IN DERMATOPHILOSIS INFECTED AND ON-INFECTED CATTLE

&#x0D; &#x0D; &#x0D; &#x0D; In two separate experiments, blood samples from dermatophilosis infected and non-infected cattle were analyzed for their vitamin C content. In the first experiment using abattoir samples, streptothricosis infected animals showed significantly lower lever (P &lt; 0.01) Of ascorbic acid in mg per 100 ml of plasma (0.56) than non-infected ones (1.11). In the second experiment using samples from live animals, a non- significant decrease in the concentration of Vitamin in C was also observed in infected (0.94) compared with non-infected animals (1.05)&#x0D; &#x0D; &#x0D; &#x0D;

Brain-derived neurotrophic factor is down regulated after bovine alpha-herpesvirus 5 infection in both wild-type and TLR3/7/9 deficient mice.

Neurotrophic factors have been implicated in the control of neuronal survival and plasticity in different brain diseases. Meningoencephalitis caused by bovine alpha-herpesvirus 5 (BoHV-5) infection is a frequent neurological disease of young cattle, being the involvement of apoptosis in the development of neuropathological changes frequently discussed in the literature. It’s well known that Toll-like receptors (TLRs) can activate neuroinflammatory response and consequently lead to neuronal loss. However, there are no studies evaluating the expression of neurotrophic factors and their association with brain pathology and TLRs during the infection by BoHV-5. The current study aimed to analyze brain levels of neurotrophic factors along with neuropathological changes during acute infection by BoHV-5 in wild-type (WT) and TLR3/7/9 (TLR3/7/9−/−) deficiency mice. The infection was induced by intracranial inoculation of 1 × 104 TCID50 of BoHV-5. Infected animals presented similar degrees of clinical signs and neuropathological changes. Both infected groups had meningoencephalitis and neuronal damage in CA regions from hippocampus. BoHV-5 infection promoted the proliferation of Iba-1 positive cells throughout the neuropil, mainly located in the frontal cortex. Moreover, significant lower levels of brain-derived neurotrophic factor (BDNF) were detected in both BoHV-5 infected WT and TLR3/7/9 deficient mice, compared with non-infected animals. Our study showed that BDNF down regulation was associated with brain inflammation, reactive microgliosis and neuronal loss after bovine alpha-herpesvirus 5 infection in mice. Moreover, we demonstrated that combined TLR3/7/9 deficiency does not alter those parameters.

Modified chitosan microparticles to molecular diagnostic of Chagas disease

Background: Chagas disease is caused by the protozoan Trypanosoma cruzi (T.cruzi), is a major public health problem in Latin America and an emerging threat in worldwide. Direct diagnosis is mainly based on the microscopic observation of the parasite but has low sensitivity mainly during the chronic phase; molecular techniques increase the sensitivity, but still limited. Nanobiotechnology is latest advance in biomedicine, recently the innovation of natural polymers is has been explored. The novel use of micro and/or nanoplatforms can be used to increase the sensitivity. The objective of the study was to develop different modified chitosan microparticles with capacity of adsorption of deoxyribonucleic acid (DNA) for the molecular diagnosis of Chagas disease. Methods and materials: Different microparticles of chitosan were synthesized and characterized, the DNA adsorption procedure was optimized evaluating parameters (type, pH of the medium, concentration, time and the desorption method) The limit of detection of plasmid DNA and T. cruzi DNA was determined. The sensitivity and specificity in the detection of kinetoplast DNA (kDNA) and satellite DNA (satDNA) was determined in urine samples of experimentally infected animals obtained at 14 and 45 d post-infection (dpi). Results: The results showed the glutaraldehyde crosslinked chitosan microparticles has the highest DNA adsorption. Using these microparticles the best conditions were obtained with: 1% glutaraldehyde, pH at 6.9, contact time of 60 m and EDTA-SDS as a desorption buffer. The detection limit of plasmid DNA was 101 copies/mL and T.cruzi DNA was 10-1 parasites/mL. Higher sensitivity was obtained with kDNA (71%) compared to satDNA (25%). The sensitivity in urine samples of infected animals at 14 dpi was 42% using ADNsat target and 78% with kDNA target; at 45 dpi with ADNsat target was 7% and with the ADNk target was 64%. High specificity (100%) was obtained for both targets in non-infected animals. Conclusion: Chitosan microparticles is innovative method and could be used to capture and concentrate T.cruzi DNA in urine (Tr-DNA).The detection of Tr-DNA could be used for diagnosis of Chagas disease.

Enhanced Free Fatty Acid Uptake Via CD36 Promotes a Metabolic Switch to B-Oxidation within Hematopoietic Stem Cells in Response to Acute Infection

Acute infection is known to promote rapid differentiation of hematopoietic stem cells (HSC) and expansion of leukocytes. Metabolic changes in the HSC underpin the mammalian response to pathogenic stimuli however, knowledge of how this occurs is not fully understood. Here we investigate the immunometabolic changes which facilitate HSC expansion in response to acute infection. We created a novel in vivo transplant model to understand if the immune response to infection involves the acquisition of free fatty acids (FFA) by HSCs in the bone marrow (BM). We transduced CD45.1 lineage negative, c-kit positive cells with firefly luciferase and transplanted into CD45.2 animals. Post engraftment animals were treated with lipopolysaccharide (LPS). Live animal imaging confirmed activation of luciferase in the BM, demonstrating in vivo, uptake of long chain FFA by haematopoietic cells in response to LPS. To confirm the specific haematopoietic stem and/or progenitor cells (HSPC) with increased lipids during infection, mice were treated with S.typhimurium for 72h then sacrificed Analysis of LSK, MPP and HSC populations (LN-, CD117+, Sca1+, CD48-, CD150+ and CD34+) showed increased intracellular BODIPY 493/503 neutral lipid dye and uptake of Bodipy FL-C12 (FFA linked to bodipy) at 72 h compared to control non-infected animals. Together, these experiments show that HSC, multipotent progenitor (MPP) and LSK (LN-, CD117+, Sca1+) cells all acquire FFA in response to bacterial infection. Seahorse mitochondrial stress test confirmed increased oxygen consumption levels in HSC from LPS (16 hours) and S.typhimurium (72 hours) treated C57BL/6. LSK metabolic activity for different substrates was assessed LSKs from LPS treated animals had an increased dependency on β-oxidation when compared to control cells. Moreover, the β-oxidation inhibitor etomoxir blocks LSK basal metabolism and reduced HSC expansion as measured by Ki-67 staining in LPS and S.typhimurium treated animals. To understand the importance of β-oxidation in HSC expansion in response to infection we first measured CPT1A expression in HSC. RNA analysis confirmed CPT1A was increased in sorted HSCs in response to LPS. Next we transplanted WTor CPT1A KD cells into WT animals, and treated with LPS. HSC expansion as measured by Ki-67 staining was inhibited in the CPT1A KD cells in response to LPS, thus confirming the importance of β-oxidation based metabolism in HSC expansion. To determine how FFA are transported into the HSC we analysed the expression of known genes to be involved in lipid uptake. HSC sorted from LPS and S.typhimurium treated animals had increased CD36 mRNA expression compared to control. C57BL/6 mice were pre-treated with the CD36 inhibitor sulfosuccinimidyl oleate (SSO) before injection with LPS. HSCs had had less lipids, lower maximal respiration and reduced cycling when compared to animals treated with LPS alone. In a similar way CD36 KO animals were treated with LPS for 16 h, no increased FFA uptake or increased lipid content or HSC cycling was observed. Seahorse mitochondrial stress test confirmed CD36-/- LSK have increased basal ECAR but no change in basal OCR in response to S.typhimurium infection. Moreover, CD36-/- LSK have reduced dependency on β-oxidation. To determine if the uptake of FFA response is specific to HSC we transplanted CD36+/+ (CD45.1) LK into CD36-/- (CD45.2) animals. We found that CD36 expression was elevated in CD36+/+ HSC transplanted into CD36-/- treated with LPS, normal uptake of FFA, an increase in lipid content and cycling of HSCs. In addition, transplanted WT CD36+/+ (CD45.1) LSK into CD36-/- (CD45.2) animals reversed the metabolic phenotype observed in CD36-/- in response to LPS. We transplanted WT(CD36+/+) or CD36KO (WT(CD36-/-)) LSK into WT animals infected with S. typhimurium for 4 days. WT(CD36-/-) transplanted animals showed enhanced mortality, increased weight loss and increased liver injury. We also observed less HSC cycling and a reduced lipid content in HSC from WT(CD36-/-) compared to WT(CD36+/+). These findings expand the mechanistic understanding of the interplay between HSC and the bone marrow microenvironment, and in doing so explains how immunometabolic reprogramming of HSCs during infection supports the metabolic demands of HSCs in response to a pathogenic challenge without which results in increased susceptibility to infection. Disclosures Bowles: AbbVie: Research Funding; Janssen: Research Funding. Rushworth:AbbVie: Research Funding; Janssen: Research Funding.

Gas chromatography-mass spectrometry (GC/MS) reveals urine metabolites associated to light and heavy infections by Schistosoma mansoni in mice.

High-throughput profiling of metabolites has been used to identify metabolic changes in murine models as a response to the infection by the parasitic trematode Schistosoma. These investigations have contributed to our understanding on the pathogenesis of this tropical neglected disease, with a potential of helping diagnosis. Here, our study aimed to investigate the application of gas chromatography-mass spectrometry (GC/MS) on the profiling of urine metabolites from mice carrying infections by Schistosoma mansoni. Two larval infection doses created distinctive infection intensities in mice, whereby the heavily infected animals were found to release 25 times more eggs in faeces than lightly infected animals. Over 200 urine metabolites were identified from these animals by GC/MS, following two complementary derivatisation methods. A list of 14 individual metabolites with altered relative abundances between groups were identified. Most of the altered metabolites showed a trend of increased abundances in response to infection intensity, indicating host-specific metabolic alterations as a result of the disease. Hippurate, a metabolite which concentration is intimately modulated by the gut microbiota, was found to be highly correlated to infection intensity. Our study showed that urine metabolic profiling by GC/MS can distinguish non-infected animals from those carrying light and heavy infections by S. mansoni, revealing metabolites associated to the infection and providing insights on the pathogenesis of schistosomiasis.

Assessment of Acute-Phase Protein Response Associated with the Different Pathological Forms of Bovine Paratuberculosis.

Simple SummaryParatuberculosis (PTB) is a chronic debilitating disease caused by Mycobacterium avium subspecies paratuberculosis (Map) that affects ruminants worldwide. Many aspects related to the pathogenesis of this disease are still unknown, including the inflammatory acute-phase response developed during the course of the infection. To clarify this, serum levels of haptoglobin and serum amyloid A, two positive acute-phase proteins, were evaluated in a total of 190 cows, among which 59 were healthy control animals and 131 cows were diagnosed post-mortem with different types of lesion associated with Map-infection. The results reflect a significant increase of these proteins’ levels in the infected animals and, more specifically, in those animals with types of lesion characterized by a low bacterial load and with predominance of a cell-mediated immune response. This suggests that these molecules would play a certain role in the pathogenesis of the PTB and a possible utility as biomarkers of different stages of the disease.In this study, the concentrations of two acute-phase proteins (APPs), haptoglobin (Hp) and serum amyloid A (SAA), were quantitatively assessed in serum samples from cattle naturally infected with paratuberculosis (PTB). APP profiles were compared across 190 animals classified according to the different pathological forms associated with infection: uninfected (n = 59), with focal lesions (n = 73), multifocal lesions (n = 19), and diffuse paucibacillary (n = 11) and diffuse multibacillary lesions (n = 28). Our results showed a significant increase in both APPs in infected animals compared to the control group, with differences depending on the type of lesion. Hp and SAA levels were increased significantly in all infected animals, except in cows with diffuse multibacillary lesions that showed similar values to non-infected animals. The expression pattern of both APPs was similar and negatively correlated with the antibody levels against PTB. These results indicate that the release of Hp and SAA is related to the presence of PTB lesions associated with a high cell-mediated immune response and a lower bacterial load, suggesting that the pro-inflammatory cytokines that are associated with these forms are the main stimulus for their synthesis. These molecules could show some potential to be used as putative biomarkers of PTB infection, particularly for the identification of subclinical animals showing pathological forms related to latency or resistance to the development of advanced lesions.

Regulation of Liver Fibrosis during Murine Schistosomiasis Mansoni

Background: The main pathology of Schistosoma mansoni (S. mansoni) infection is induced by a granulomatous tissue reaction against the parasite eggs. Unfortunately, no therapy has been proven to prevent the progression of hepatic fibrosis associated with granulomatous hypersensitivity to parasite eggs. Accumulating evidence has demonstrated a critical pathogenic role for both interleukin 17 (IL-17) and interleukin 13 (IL-13) in organ fibrosis. Objective: The present study investigated the role of IL-17 and IL-13 in the pathogenesis of liver fibrosis during S. mansoni infection. Material and Methods: Thirty female C57BL/6 mice were divided into three groups, normal, infected, and anti- mouse IL-17 treated groups. The infected and anti-mouse IL-17 groups were infected with 40±5 cercariae of S. mansoni per mouse. Neutralizing rat anti-mouse IL-17 mAb or an isotype-matched rat IgG mAb was first administered intraperitoneally 3 weeks after S. mansoni infection (62.5μg per mouse) then at the same dose every third day until 2 days before killing. Serum IL-17, IL-13, and proinflammatory cytokines levels were determined by ELISA. Liver granulomas were measured by an ocular micrometer. Results: Serum level of IL-17 was significantly higher in infected mice compared with non-infected animals. Reducing IL-17 activity using anti-IL-17 monoclonal antibodies improves liver functions and reduces the size of the liver granulomas. Meanwhile, Th-2 profibrogenic cytokine IL-13 was also decreased in infected/anti-IL-17 mAB- treated mice. IL-17-induced proinflammatory mediators (IL-1Land TNF-that involved in liver fibrosis were markedly reduced in anti-IL-17 mAB-treated mice. Conclusion: IL-17 and IL-13 contribute to granulomatous inflammatory and fibrosing reactions in murine schistosomiasis.

Regulation of Liver Fibrosis during Murine Schistosomiasis Mansoni

Background: The main pathology of Schistosoma mansoni (S. mansoni) infection is induced by a granulomatous tissue reaction against the parasite eggs. Unfortunately, no therapy has been proven to prevent the progression of hepatic fibrosis associated with granulomatous hypersensitivity to parasite eggs. Accumulating evidence has demonstrated a critical pathogenic role for both interleukin 17 (IL-17) and interleukin 13 (IL-13) in organ fibrosis. Objective: The present study investigated the role of IL-17 and IL-13 in the pathogenesis of liver fibrosis during S. mansoni infection. Material and Methods: Thirty female C57BL/6 mice were divided into three groups, normal, infected, and anti- mouse IL-17 treated groups. The infected and anti-mouse IL-17 groups were infected with 40±5 cercariae of S. mansoni per mouse. Neutralizing rat anti-mouse IL-17 mAb or an isotype-matched rat IgG mAb was first administered intraperitoneally 3 weeks after S. mansoni infection (62.5μg per mouse) then at the same dose every third day until 2 days before killing. Serum IL-17, IL-13, and proinflammatory cytokines levels were determined by ELISA. Liver granulomas were measured by an ocular micrometer. Results: Serum level of IL-17 was significantly higher in infected mice compared with non-infected animals. Reducing IL-17 activity using anti-IL-17 monoclonal antibodies improves liver functions and reduces the size of the liver granulomas. Meanwhile, Th-2 profibrogenic cytokine IL-13 was also decreased in infected/anti-IL-17 mAB- treated mice. IL-17-induced proinflammatory mediators (IL-1Land TNF-that involved in liver fibrosis were markedly reduced in anti-IL-17 mAB-treated mice. Conclusion: IL-17 and IL-13 contribute to granulomatous inflammatory and fibrosing reactions in murine schistosomiasis.

Detection of latent forms of Mycobacterium avium subsp. paratuberculosis infection using host biomarker-based ELISAs greatly improves paratuberculosis diagnostic sensitivity.

Bovine paratuberculosis (PTB) is a chronic granulomatous enteritis, caused by Mycobacterium avium subsp. paratuberculosis (MAP), responsible for important economic losses in the dairy industry. Current diagnostic methods have low sensitivities for detection of latent forms of MAP infection, defined by focal granulomatous lesions and scarce humoral response or MAP presence. In contrast, patent infections correspond to multifocal and diffuse types of enteritis where there is increased antibody production, and substantial mycobacterial load. Our previous RNA-Seq analysis allowed the selection of five candidate biomarkers overexpressed in peripheral blood of MAP infected Holstein cows with focal (ABCA13 and MMP8) and diffuse (FAM84A, SPARC and DES) lesions vs. control animals with no detectable PTB-associated lesions in intestine and regional lymph nodes. The aim of the current study was to assess the PTB diagnostic potential of commercial ELISAs designed for the specific detection of these biomarkers. The ability of these ELISAs to identify animals with latent and/or patent forms of MAP infection was investigated using serum from naturally infected cattle (n = 88) and non-infected control animals (n = 67). ROC analysis revealed that the ABCA13-based ELISA showed the highest diagnostic accuracy for the detection of infected animals with focal lesions (AUC 0.837, sensitivity 79.25% and specificity 88.06%) and with any type of histological lesion (AUC 0.793, sensitivity 69.41% and specificity 86.57%) improving on the diagnostic performance of the popular IDEXX ELISA and other conventional diagnostic methods. SPARC and MMP8 showed the highest diagnostic accuracy for the detection of animals with multifocal (AUC 0.852) and diffuse lesions (AUC 0.831), respectively. In conclusion, our results suggest that quantification of ABCA13, SPARC and MMP8 by ELISA has the potential for implementation as a diagnostic tool to reliably identify MAP infection, greatly improving early detection of MAP latent infections when antibody responses and fecal shedding are undetectable using conventional diagnostic methods.

Effect of bisphosphonates on healing of tooth extraction wounds in infectious osteomyelitis of the jaw.

Antiresorptive agent-related osteonecrosis of the jaw (ARONJ) and infectious osteomyelitis of the jaw (OMJ) in antiresorptive-naïve patients are different disease entities. Although osteoclast inhibition is at the center of the pathogenesis of ARONJ, the role of osteoclast inhibition in infectious OMJ is unknown. The objective of this study was to determine the effect of bisphosphonate osteoclast inhibition in infectious OMJ. Osteomyelitis was induced in mice by S. aureus inoculation. The establishment of OMJ was verified by the culture of bone marrow samples obtained from the mandible. Infected animals received either zoledronic acid (ZA) or saline starting at week-2. Treated animals along with non-infected animals underwent tooth extractions at week-4 post-infection. Healing was assessed every week using in vivo micro-computed tomography and intraoral photos. Animals were euthanized at week-8 and cervical lymph nodes were assessed for lymphatic and blood vessels. Tooth extraction wounds did not heal in animals with OMJ. These wounds were characterized by incomplete soft tissue coverage, sporadic bone fill in the sockets, and inflammatory cell accumulation in the connective tissue at 4weeks after tooth extractions. Conversely, the majority of tooth extraction wounds in the infected animals treated with ZA had improved healing with better bone fill than even non-infected control animals. Consistently, atrophic lymphatic vessels were noted in the draining lymph nodes in animals with OMJ. However, infected animals treated with ZA had lymphatic vessels that were unaltered and showed a similar appearance to those in non-infected control animals. ZA treatment promoted wound healing in the jaw with infectious osteomyelitis. Clearly, antiresorptive therapy is contraindicated in patients with ARONJ. However, our finding suggests that osteoclast inhibition is potentially an effectual remedy for infectious OMJ in antiresorptive-naïve patients.

How much does it cost to prevent and control visceral leishmaniasis in Brazil? Comparing different measures in dogs

Visceral leishmaniasis (VL) is an important zoonosis in Brazil. Dogs are considered the main domestic reservoirs of the disease in the country; hence, control measures are focused on these reservoirs. Despite efforts to prevent and control VL, important reductions in disease prevalence and incidence have not been identified, stimulating the development and application of new strategies. The choice and implementation of new control strategies can benefit from the application of mathematical models that allow the simulation of different strategies in different scenarios. Selecting the best strategy to be implemented is also supported by cost-effectiveness studies. Here we used the results of a mathematical model in which scenarios, including isolated use of the vaccine and insecticide-impregnated collar (IIC), both at different coverage rates, were simulated to conduct a cost-effectiveness study. The costs were calculated for each scenario considering a simulation period of four years. Collar application in both infected and non-infected animals was the most cost-effective strategy. For example, to reduce the prevalence in humans and dogs by approximately 70%, the costs ranged from $250,000 and $550,000 for the IICs and vaccination, respectively. Even in the scenario with 40% loss/replacement of IICs, this measure was more advantageous in terms of cost-effectiveness than vaccination. If the vaccine were applied with culling of seropositive tested dogs, then the measure became more effective with a reduced cost compared with the vaccine alone. The use of the three first consecutive vaccine doses had the greatest impact on the cost of the vaccination strategy. The advantage of using IICs is that there is no need for a prior diagnosis, unlike vaccination, reducing costs and facilitating implementation. The present study aims to contribute to strategies to reduce hosts infected with VL by reducing public expenditure.

Combined Transcriptome Sequencing of Mycoplasma hyopneumoniae and Infected Pig Lung Tissue Reveals Up-Regulation of Bacterial F1-Like ATPase and Down-Regulation of the P102 Cilium Adhesin in vivo.

Mycoplasma hyopneumoniae (M. hyopneumoniae) causes enzootic pneumonia in pigs but it is still largely unknown which host-pathogen interactions enable persistent infection and cause disease. In this study, we analyzed the host and bacterial transcriptomes during infection using RNA sequencing. Comparison of the transcriptome of lung lesion tissue from infected pigs with lung tissue from non-infected animals, identified 424 differentially expressed genes (FDR < 0.01 and fold change > 1.5LOG2). These genes were part of the following major pathways of the immune system: interleukin signaling (type 4, 10, 13, and 18), regulation of Toll-like receptors by endogenous ligand and activation of C3 and C5 in the complement system. Besides analyzing the lung transcriptome, a sampling protocol was developed to obtain enough bacterial mRNA from infected lung tissue for RNA sequencing. This was done by flushing infected lobes in the lung, and subsequently enriching for bacterial RNA. On average, 2.2 million bacterial reads were obtained per biological replicate to analyze the bacterial in vivo transcriptome. We compared the in vivo bacterial transcriptome with the transcriptome of bacteria grown in vitro and identified 22 up-regulated and 30 down-regulated genes (FDR < 0.01 and fold change > 2LOG2). Six out of seven genes in the operon encoding the mycoplasma specific F1-like ATPase (MHP_RS02445-MHP_RS02475) and all genes in the operon MHP_RS01965-MHP_RS01990 with functions related to nucleotide metabolism, spermidine transport and glycerol-3-phoshate transport were up-regulated in vivo. Down-regulated in vivo were genes related to glycerol uptake, cilium adhesion (P102), cell division and myo-inositol metabolism. In addition to providing a novel method to isolate bacterial mRNA from infected lung, this study provided insights into changes in gene expression during infection, which could help development of novel treatment strategies against enzootic pneumonia caused by M. hyopneumoniae.

Helicobacter pylori Colonization Drives Urokinase Receptor (uPAR) Expression in Murine Gastric Epithelium During Early Pathogenesis.

(1) Background: Persistent Helicobacter pylori infection is the most important risk factor for gastric cancer. The urokinase receptor (uPAR) is upregulated in lesions harboring cancer invasion and inflammation. Circumstantial evidence tends to correlate H. pylori colonization with increased uPAR expression in the human gastric epithelium, but a direct causative link has not yet been established in vivo; (2) Methods: In a mouse model of H. pylori-induced gastritis, we investigated the temporal emergence of uPAR protein expression in the gastric mucosa in response to H. pylori (SS1 strain) infection; (3) Results: We observed intense uPAR immunoreactivity in foveolar epithelial cells of the gastric corpus due to de novo synthesis, compared to non-infected animals. This uPAR induction represents a very early response, but it increases progressively over time as do infiltrating immune cells. Eradication of H. pylori infection by antimicrobial therapy causes a regression of uPAR expression to its physiological baseline levels. Suppression of the inflammatory response by prostaglandin E2 treatment attenuates uPAR expression. Notwithstanding this relationship, H. pylori does induce uPAR expression in vitro in co-cultures with gastric cancer cell lines; (4) Conclusions: We showed that persistent H. pylori colonization is a necessary event for the emergence of a relatively high uPAR protein expression in murine gastric epithelial cells.

SAT-LB131 Examination of Hepatic Gene Expression Following Developmental Exposure to Dieldrin in Trachemys Scripta and Discovery of a Novel Hepacivirus

The Massachusetts Military Reservation (MMR) is a Superfund site where ground water has been contaminated by a mixture of pollutants. Exposure to these chemicals is a public health concern and reproductive impairments have been observed in a population of turtles (Chrysemys picta) endemic to this site. We hypothesize that developmental exposure to endocrine disrupting compounds originating from the MMR might lead to abnormalities seen in adult animals. Upon examination of egg yolk from turtles at the impacted site, we found the presence of dieldrin and p,p’-DDE. Turtles from a reference site were also found to have p,p’-DDE present in the yolk. In order to investigate these chemicals in the laboratory we used a closely related turtle (Trachemys scripta) and applied vehicle, dieldrin, or p,p’-DDE to the eggshells. Absorption of p,p’-DDE through the eggshell was limited. Although there were variations in absorbance, we were able to achieve levels of dieldrin in the yolk similar to what was seen in animals from the impacted site. Following in ovo exposure to dieldrin, we used RNAseq to examine hepatic gene expression in neonates and found that several transcripts were repressed at least 1.5-fold in the dieldrin-treated animals. QPCR was carried out to confirm differences in gene expression. We found that hepatic gene expression (fold ± SEM) of Gamma-Butyrobetaine Hydroxylase 1 (Bbox1) was higher in vehicle-treated animals (1 ± 0.60) compared to dieldrin-treated animals (0.29 ± 0.12). Bbox1 catalyzes the last step in the L-carnitine biosynthetic pathway, which is necessary for mitochondrial beta-oxidation. Dieldrin-induced reduction in L-carnitine production may be a critical factor in adult reproductive function. Further, Protein-L-Isoaspartate (D-Aspartate) O-Methyltransferase Domain Containing Protein 1 (Pcmtd1) was higher (1± 0.76) in vehicle-treated animals compared to dieldrin-treated animals (0.13 ± 0.044), and DENN Domain Containing 5B (Dennd5b) was also found to be higher in vehicle-treated (1 ± 0.39) compared to dieldrin-treated (0.28 ± 0.10) turtles. We found that dieldrin exposure did not alter gene expression of Cytochrome p450 1a (Cyp1a) a marker of aryl hydrocarbon receptor signaling, or Vitellogenin 2 (Vtg2) a marker of estrogen signaling. In our RNAseq analysis we unexpectedly discovered a hepacivirus infecting T. scripta. In the dieldrin-treated group, we used QPCR to examine gene expression of potential markers of hepacivirus infection. Neither Interleukin 1 Beta (Il-1β), SMAD Family Member 6 (Smad6), C-C Motif Chemokine Ligand 5 (Ccl5), or TNF Receptor Superfamily Member 9 (Tnfrsf9) was found to differ in turtles carrying the virus, compared to non-infected animals. In conclusion, we found that developmental dieldrin exposure of T. scripta slightly reduces neonatal expression of several gene transcripts which may be correlated to adult reproductive fitness.

Sendai Virus, a Strong Inducer of Anti-Lentiviral State in Ovine Cells

Small ruminant lentiviruses (SRLVs) are widely spread in the ovine and caprine populations, causing an incurable disease affecting animal health and production. Vaccine development is hindered owing to the high genetic heterogeneity of lentiviruses and the selection of T-cell and antibody escape mutants, requiring antigen delivery optimization. Sendai virus (SeV) is a respiratory paramyxovirus in mice that has been recognized as a potent inducer of innate immune responses in several species, including mouse and human. The aim of this study was to stimulate an innate antiviral response in ovine cells and evaluate the potential inhibitory effect upon small ruminant lentivirus (SRLV) infections. Ovine alveolar macrophages (AMs), blood-derived macrophages (BDMs), and skin fibroblasts (OSFs) were stimulated through infection with SeV encoding green fluorescent protein (GFP). SeV efficiently infected ovine cells, inducing an antiviral state in AM from SRLV naturally-infected animals, as well as in in vitro SRLV-infected BDM and OSF from non-infected animals. Supernatants from SeV-infected AM induced an antiviral state when transferred to fresh cells challenged with SRLV. Similar to SRLV, infectivity of an HIV-1-GFP lentiviral vector was also restricted in ovine cells infected with SeV. In myeloid cells, an M1-like proinflammatory polarization was observed together with an APOBEC3Z1 induction, among other lentiviral restriction factors. Our observations may boost new approximations in ameliorating the SRLV burden by stimulation of the innate immune response using SeV-based vaccine vectors.

Wound healing, antimicrobial and antioxidant efficacy of Amaranthus spinosus ethanolic extract on rats

The present study was aimed to evaluate wound healing activity of ethanolic exact of Amaranthus spinosus on rats infected by microbes and its antioxidant potential for fast recovery from wounds. The whole plant extraction was performed using 65% ethanol followed by phytochemical screening to determine secondary metabolites present in the plant. In addition, HPLC analysis of plant extract was conducted to identify the active constituents. Wound healing activity of A. spinosus was determined using wound excision mode on rats. Animals were treated with two concentrations (5% and 10% w/w ointment) of A. spinosus and soframycin was used as standard. Antimicrobial activity against bacterial and fungal strains was evaluated using agar disc diffusion method. Antioxidant activity was evaluated by measuring oxidative stress parameters in rat tissues. Results showed that A. spinosus significantly restored wound tissues in both infected and non-infected animals. Complete found contraction was achieved on day 18th due to treatment of 10% ointment of A. spinosus . In addition, epithelization period was reduced to 12.16 days as compared to control group i.e. 24.66 days. A. spinosus extract was effective against all selected microbial strains; however, maximum zone of inhibition (21.8 ± 0.76 mm) was noted against Staphylococcus epidermidis . The antioxidant potential of A. spinosus extract significant restored the level of oxidative free radicals in rats and helped in fast recovery of wounds. The study concluded that A. spinosus extract may be an alternative therapy for healing both infected- and non-infected wounds.

Lipophosphoglycan-3 recombinant protein vaccine controls hepatic parasitism and prevents tissue damage in mice infected by Leishmania infantum chagasi

AimsIn this work, we aimed to evaluate the effects of the Leishmania infantum chagasi infection on the liver of vaccinated mice, considering parameters of tissue damage and the inflammatory response elicited by vaccination. Main methodsWe used recombinant LPG3 protein (rLPG3) as immunogen in BALB/c mice before challenge with promastigote forms of L. infantum chagasi. The animals were separated into five groups: NI: non-infected animals; NV: non-vaccinated; SAP: treated with saponin; rLPG3: immunized with rLPG3; rLPG3 + SAP: immunized with rLPG3 plus SAP. The experiment was conducted in replicate, and the vaccination protocol consisted of three subcutaneous doses of rLPG3 (40 μg + two boosters of 20 μg). The mice were challenged two weeks after the last immunization. Key findingsOur results showed that rLPG3 + SAP immunization decreased the parasite burden in 99 %, conferring immunological protection in the liver of the infected animals. Moreover, the immunization improved the antioxidant defenses, increasing CAT and GST activity, while reducing the levels of oxidative stress markers, such as H2O2 and NO3/NO2, and carbonyl protein in the organ. As a consequence, rLPG3 + SAP immunization preserved tissue integrity and reduced the granuloma formation, inflammatory infiltrate and serum levels of AST, ALT, and ALP. SignificanceTaken together, these results showed that rLPG3 vaccine confers liver protection against L. infantum chagasi in mice, while maintaining the liver tissue protected against the harmful inflammatory effects caused by the vaccine followed by the infection.