Blood redistribution preferentially protects vital organs under hypoxic stress in Pelteobagrus vachelli.

Blood redistribution occurs in mammals under hypoxia but has not been reported in fish. This study investigated the tissue damage, hypoxia-inducible factor (HIF) activation level, and blood flow changes in the brain, liver, and muscle of Pelteobagrus vachelli during the hypoxia process for normoxia-hypoxia-asphyxia. The results showed that P. vachelli has tissue specificity in response to hypoxic stress. Cerebral blood flow increased with less damage than in the liver and muscle, suggesting that P. vachelli may also have a blood redistribution mechanism in response to hypoxia. It is worth noting that severe hypoxia can lead to a sudden increase in the degree of brain tissue damage. In addition, higher dissolved oxygen levels activate HIF and may have contributed to the reduced damage observed in the brain. This study provides basic data for investigating hypoxic stress in fish.

Examination of organs and tissues of adult sheep grazed in an area with possible intoxication with rocket fue, Kazakhstan

The authors have conducted experiments to study the pathoanatomical and histological pattern of organs and tissues of adult sheep affected by unsymmetric dimethylhydrazine (UDMH). This highly toxic fuel was spilled on the territory of the Karsakpay and Ulytau districts of Karaganda region, Kazakhstan, because of the fall of the rocket 'Proton­M' after an unsuccessful launch from the Baikonur cosmodrome in 2007. In the experiment, the study group was consisted of 7 adult sheep that grazed in the area of possible intoxication with rocket fuel UDMH. The main objects of the study were histological preparations obtained from fixed structures. As the structures have a flat contrast and are poorly detected in the ordinary light microscope, the specially processed preparations were used. After preparing, the authors studied organs and tissues using a microscope, which allowed to reveal in detail the level of damage caused by intoxication and to establish the negative effect of UDMH on the internal organs. The group of sheep showed a high index of macroscopic signs of interstitial pneumonia (85.7 ± 14.3%), and histologically quite high index was granulomatous inflammation of liver (71.4 ± 18.4%). Kidneys also showed a high level of abnormalities.

Analysis of Morphological Changes in the CNS and Internal Organs of Macaca mulatta Monkeys after Intracerebral Injection of a Low-Attenuated Rubella Virus Strain.

We studied the localization and severity of morphological changes in CNS and internal organs of animals intacerebrally infected with a low-attenuated rubella virus strain "Orlov-14". The data obtained can be used as morphological criteria reflecting low level of attenuation of rubella virus strains to improve the control of the safety of attenuated strains of live rubella vaccines.

Effect of sample size and length of observation period on the reliability of apparent pig organ lesion prevalence.

Comprehensive identification of on-farm animal-health issues still requires extensive efforts so that in practice such monitoring is applied only sparsely. An appealing approach to improve on-farm animal health and welfare monitoring is the application of organ lesion scoring data from the abattoir as such is instantly available for every commercial farm in Europe. Unfortunately, it is also well-known that organ lesion scoring is often unreliable because results are altered by several non-health-related factors, diluting the validity of lesion scoring prevalence as a proxy for on-farm animal health. However, it is theoretically possible to improve prevalence reliability a-posteriori by application of time-series smoothing. The aim of this paper was therefore to analyse whether it is practically possible to increase apparent prevalence estimation reliability retrospectively using a running average, and, if so, which window length and smallest sample size should be preferred in such an approach. Because no gold standard for direct evaluation of lesion reliability is available for field-data, apparent prevalence reliability had to be approximated using prevalence agreement over time. Results indicate that by raising the number of lesion scores per prevalence estimate, apparent prevalence agreement over time can in general be considerably increased. Based on findings presented, a reasonable threshold for prevalence estimation is given by at least n = 50 lesions per farm/abattoir/time-series segment. Results further suggest that it is necessary to consider differences in prevalence sample size for future monitoring purposes, because prevalences that are estimated on a continuum of different sample sizes put together in one evaluation may induce substantial error in prevalence estimates.

Pesticides and transgenerational inheritance of pathologies: Designing, analysing and reporting rodent studies.

Single-centre studies examining the transgenerational inheritance of pathologies in rodents exposed to pesticides have not always taken important design and analysis issues into account. This paper examines these methodological and statistical issues in detail. Its particular focus is on the estimation of 'litter effects': the tendency for rodents within a litter to be more alike than rodents in different litters. Appropriate statistical models were fitted to published data from a series of widely reported studies carried out at Washington State University. These studies were amalgamated into a single dataset in order to estimate these litter effects and associated treatment effects. Litter effects varied by outcome and were often substantial. Consequently, the effective sample size was often substantially less than the number of observations with implications for the power of the studies. Moreover, the reported precision of the estimates of treatment effects was too low. These problems are exacerbated by unexplained missing data across generations. Researchers in the life sciences could be more cognisant of the guidelines established in medicine for reporting randomised controlled trials, particularly cluster randomised trials. More attention should be paid to the design and analysis of multi-generational rodent studies; their imperfections have important implications for assessments of the evidence relating to the risks of pesticides for public health.

3,4-Dimethoxychalcone induces autophagy through activation of the transcription factors TFE3 and TFEB.

Caloric restriction mimetics (CRMs) are natural or synthetic compounds that mimic the health-promoting and longevity-extending effects of caloric restriction. CRMs provoke the deacetylation of cellular proteins coupled to an increase in autophagic flux in the absence of toxicity. Here, we report the identification of a novel candidate CRM, namely 3,4-dimethoxychalcone (3,4-DC), among a library of polyphenols. When added to several different human cell lines, 3,4-DC induced the deacetylation of cytoplasmic proteins and stimulated autophagic flux. At difference with other well-characterized CRMs, 3,4-DC, however, required transcription factor EB (TFEB)- and E3 (TFE3)-dependent gene transcription and mRNA translation to trigger autophagy. 3,4-DC stimulated the translocation of TFEB and TFE3 into nuclei both in vitro and in vivo, in hepatocytes and cardiomyocytes. 3,4-DC induced autophagy in vitro and in mouse organs, mediated autophagy-dependent cardioprotective effects, and improved the efficacy of anticancer chemotherapy in vivo. Altogether, our results suggest that 3,4-DC is a novel CRM with a previously unrecognized mode of action.

Comparative evaluation of pathogenicity of three isolates of vesicular stomatitis virus (Indiana serotype) in pigs.

Vesicular stomatitis (VS) is a notifiable disease of livestock affecting cattle, horses, pigs and humans. Vesicular stomatitis virus (VSV) serotypes Indiana and New Jersey are endemic to Central America; however, they also cause sporadic and scattered outbreaks in various countries in South and North America, including the USA. In order to develop an effective experimental challenge model for VSV, we compared the pathogenicity of three VSV serotype Indiana isolates in 36 4-5 week-old pigs. Two bovine isolates of Central American origin and one equine isolate from the USA were used for the experimental infections. Each pig was inoculated with a single isolate by both the intradermal and intranasal routes. Clinical signs of VSV infection were recorded daily for 10 days post-inoculation (days p.i.). Nasal and tonsillar swab samples and blood were collected to monitor immune responses, virus replication and shedding. Post-challenge, characteristic signs of VS were observed, including vesicles on the nasal planum and coronary bands, lameness, loss of hoof walls and pyrexia. Pigs inoculated with the Central American isolates showed consistently more severe clinical signs in comparison to the pigs infected with the USA isolate. Genomic RNA was isolated from the original challenge virus stocks, sequenced and compared to VSV genomes available in GenBank. Comparative genome analysis demonstrated significant differences between the VSV isolate from the USA and the two Central American isolates. Our results indicate that the Central American isolates of VSV serotype Indiana used in this study are more virulent in swine than the USA VSV serotype Indiana isolate and represent good candidate challenge strains for future VSV studies.

Antimicrobial activity of the antibacterial peptide PMAP-36 and its analogues.

The growing problem of antibiotic resistance has attracted people's attention; thus, the search for new antibacterial agents is imminent. In this study, a series of antimicrobial peptides (AMPs) based on the porcine antibacterial peptide PMAP-36 were designed by amino acid substitution to develop peptide analogues as new classes of antimicrobial agents. By extending the α-helix and increasing the positive charge, two peptide analogues, PMAP-36PW and PMAP-36PK, were synthesized. The antibacterial activities of PMAP-36 and its peptide analogues were detected in vitro and in vivo. The results showed that PMAP-36PW and PMAP-36PK had a broadened antibacterial spectrum compared to that of PMAP-36. After the modification, PMAP-36PW and PMAP-36PK exhibited antibacterial activities on swine Escherichia coli K88, while PMAP-36 did not. PMAP-36, PMAP-36PW and PMAP-36PK did not have antibacterial activities against Enterococcus faecium B21. PMAP-36 PW had significant antibacterial activity against seven bacterial strains compared to PMAP-36, and PMAP-36PK had significant antibacterial activity against five bacterial strains compared to PMAP-36. Furthermore, PMAP-36PW exhibited enhanced pH stability. Moreover, in the in vivo efficacy assessment of mice infected with Salmonella choleraesuis C78-1 and Listeria monocytogenes CICC 21533, the peptide analogues exhibited an impressive therapeutic effect by reducing bacterial gene copies and decreasing inflammatory damage in mouse livers and lungs, resulting in a reduction in mouse mortality. This study provides reference data for the design of clinically effective antibacterial peptides.

Accurate diagnosis of lesions suspected of being caused by Taenia solium in body organs of pigs with naturally acquired porcine cysticercosis.

The definitive method for diagnosis of porcine cysticercosis is the detection of cysticerci at necropsy. Cysts are typically located in the striated muscle and brain. Until recently Taenia solium cysticerci have not been definitively identified in other tissue locations, despite several comprehensive investigations having been undertaken which included investigation of body organs other than muscle and brain. Recently a study conducted in Zambia reported 27% infection with T. solium in the liver of pigs with naturally acquired porcine cysticercosis, as well as some T. solium infection in the lungs and spleen of some animals. We investigated the cause of lesions in sites other than the muscle or brain in a total of 157 pigs from T. solium endemic regions of Uganda and Nepal which were subjected to extensive investigations at necropsy. Lesions which had the potential to be caused by T. solium were characterised by macroscopic and microscopic examination, histology as well as DNA characterisation by PCR-RFLP and sequencing. Lesions were confirmed as being caused by Taenia hydatigena (both viable and non-viable), by T. asiatica and Echinococcus granulosus (in Nepal) and nematode infections. No T. solium-related lesions or cysticerci were identified in any tissue other than muscle and brain. It is recommended that future evaluations of porcine cysticercosis in aberrant tissue locations include DNA analyses that take appropriate care to avoid the possibility of contamination of tissue specimens with DNA from a different tissue location or a different animal. The use of appropriate control samples to confirm the absence of cross-sample contamination is also recommended.

Susceptibility of Chickens to Porcine Deltacoronavirus Infection.

Porcine deltacoronavirus (PDCoV) is a novel swine enteropathogenic coronavirus with worldwide distribution. PDCoV belongs to the Deltacoronavirus (DCoV) genus, which mainly includes avian coronaviruses (CoVs). PDCoV has the potential to infect human and chicken cells in vitro, and also has limited infectivity in calves. However, the origin of PDCoV in pigs, the host range, and cross-species infection of PDCoV still remain unclear. To determine whether PDCoV really has the ability to infect chickens in vivo, the three lines of chicken embryos and specific pathogen free (SPF) chickens were inoculated with PDCoV HNZK-02 strain to investigate PDCoV infection in the current study. Our results indicated that PDCoV can infect chicken embryos and could be continuously passaged on them. Furthermore, we observed that PDCoV-inoculated chickens showed mild diarrhea symptoms and low fecal viral RNA shedding. PDCoV RNA could also be detected in multiple organs (lung, kidney, jejunum, cecum, and rectum) and intestinal contents of PDCoV-inoculated chickens until 17 day post-inoculation by real-time quantitative PCR (qRT-PCR). A histology analysis indicated that PDCoV caused mild lesions in the lung, kidney, and intestinal tissues. These results prove the susceptibility of chickens to PDCoV infection, which might provide more insight about the cross-species transmission of PDCoV.

Epizootic Infection by Trypanosoma vivax in Cattle from the State of Minas Gerais, Brazil.

Trypanosomiasis is caused by a pathogenic protozoan of the genus Trypanosoma, being Trypanosoma vivax the most important agent for cattle. The aim of the present study was to demonstrate the expansion of T. vivax infection in different mesoregions of Minas Gerais, Brazil, and describe the clinicopathological findings of trypanosomiasis in cattle. The diagnosis was based on visualization of the parasite in blood smears and DNA detection of T. vivax in the blood of live cows and tissues of necropsied animals by the polymerase chain reaction (PCR). Thirty suspected herds were tested, of which 11 were positive for T. vivax. The most frequent clinical signs were anemia, apathy, drop in milk production, weight loss, reproductive disorders, and nervous signs. Concomitant diseases, such as malignant edema, pneumonia and increased cases of mastitis were associated with T. vivax infection. Three cows were necropsied and the most significant findings were low body condition score, pale mucous and spleen with white pulp hyperplasia. The results demonstrated the expansion of T. vivax infection in Minas Gerais, that PCR-associated blood smears are promising for diagnosis, and that other diseases often occur concomitantly to T. vivax infection in regions with trypanosomiasis in cattle.

Comprehensive Kinetic Survey of Intestinal, Extra-Intestinal and Systemic Sequelae of Murine Ileitis Following Peroral Low-Dose Toxoplasma gondii Infection.

We have recently shown that following peroral low-dose Toxoplasma gondii infection susceptible mice develop subacute ileitis within 10 days. Data regarding long-term intestinal and extra-intestinal sequelae of infection are scarce, however. We therefore challenged conventional C57BL/6 mice with one cyst of T. gondii ME49 strain by gavage and performed a comprehensive immunopathological survey 10, 36, and 57 days later. As early as 10 days post-infection, mice were suffering from subacute ileitis as indicated by mild-to-moderate histopathological changes of the ileal mucosa. Furthermore, numbers of apoptotic and proliferating/regenerating epithelial cells as well as of T and B lymphocytes in the mucosa and lamina propria of the ileum were highest at day 10 post-infection, but declined thereafter, and were accompanied by enhanced pro-inflammatory mediator secretion in ileum, colon and mesenteric lymph nodes that was most pronounced during the early phase of infection. In addition, subacute ileitis was accompanied by distinct shifts in the commensal gut microbiota composition in the small intestines. Remarkably, immunopathological sequelae of T. gondii infection were not restricted to the intestines, but could also be observed in extra-intestinal tissues including the liver, kidneys, lungs, heart and strikingly, in systemic compartments that were most prominent at day 10 post-infection. We conclude that the here provided long-term kinetic survey of immunopathological sequalae following peroral low-dose T. gondii infection provides valuable corner stones for a better understanding of the complex interactions within the triangle relationship of (parasitic) pathogens, the host immunity and the commensal gut microbiota during intestinal inflammation. The low-dose T. gondii infection model may be applied as valuable gut inflammation model in future pre-clinical studies in order to test potential treatment options for intestinal inflammatory conditions in humans.

Experimental infection in mice with Erwinia persicina.

Erwinia persicinus (E. persicina) is a plant pathogenic bacterial species that was previously isolated from a case of human infection. This study aimed to create an experimental infection protocol for E. persicina in laboratory mice. Seventy-two adult mice were divided into four groups (18 animal/group): the control group (G1), the group infected with E. persicina (G2), the group immune-suppressed with cyclophosphamide (G3) and the group immune-suppressed with cyclophosphamide and infected with E. persicina (G4). G2 and G4 were injected with 200 µL of (1 × 1013 cfu/ml) concentration intraperitoneally. Clinical signs, such as diarrhoea, apathy and mortality were observed only in G2 and G4 animals. E. persicina was not detected in blood. Necropsies of the G2 and G4 animals showed lesions in the intestine, liver, kidney and lung tissue. These lesions were characterized by infiltration of inflammatory cells, hyperaemia and focal areas of tissue necrosis in the liver. The results of the pro-inflammatory cytokines analysis revealed a significant increase in the levels of TNF-α and IL1-ß in the liver tissue of the G4 group. E. persicina is an emerging bacterium that can cause pathological lesions into mammalian tissue, which warrants further investigation.

In vitro/vivo Mechanism of Action of MP1102 With Low/Nonresistance Against Streptococcus suis Type 2 Strain CVCC 3928.

Streptococcosis is recognized as a leading infectious disease in the swine industry. Streptococcus suis serotype 2 is regarded as the most virulent species, which threatens human and pig health and causes serious economic losses. In this study, multiple in vitro and in vivo effects of MP1102 on multidrug resistant S. suis was studied for the first time. MP1102 exhibited significant antibacterial activity against S. suis (minimum inhibitory concentration, MIC = 0.028-0.228 µM), rapid bacteriocidal action, a longer postantibiotic effect than ceftriaxone, and a synergistic or additive effect with lincomycin, penicillin, and ceftriaxone (FICI = 0.29-0.96). No resistant mutants appeared after 30 serial passages of S. suis in the presence of MP1102. Flow cytometric analysis and electron microscopy observations showed that MP1102 destroyed S. suis cell membrane integrity and affected S. suis cell ultrastructure and membrane morphology. Specifically, a significantly wrinkled surface, intracellular content leakage, and cell lysis were noted, establishing a cyto-basis of nonresistance to this pathogen. DNA gel retardation and circular dichroism analysis indicated that MP1102 interacted with DNA by binding to DNA and changing the DNA conformation, even leading to the disappearance of the helical structure. This result further supported the mechanistic basis of nonresistance via interaction with an intracellular target, which could serve as a means of secondary injury after MP1102 is transported across the membrane. Upon treatment with 2.5-5.0 mg/kg MP1102, the survival of mice challenged with S. suis was 83.3-100%. MP1102 decreased bacterial translocation in liver, lung, spleen, and blood; inhibited the release of interleukin-1ß and tumor necrosis factor-α; and relieved the lung, liver, and spleen from acute injury induced by S. suis. These results suggest that MP1102 is a potent novel antibacterial agent for the treatment of porcine streptococcal disease.

Velogenic newcastle disease virus tissue tropism and pathogenesis of infection in chickens by application of in situ PCR, immunoperoxase staining and HE staining.

Limited deep studies are available in the field of early stages of pathogenesis of Newcastle disease virus (NDV) infection and tissue tropism of NDV. In this study, 24 specific pathogen free (SPF) chickens of white leghorn breed were infected with Newcastle disease (ND) by intranasal administration of 105 50% EID50/0.1 mL of velogenic NDV (vNDV). A second group of 15 chickens were kept as a control group. Chickens were monitored every day to record clinical signs. Infected chickens were euthanized by cervical dislocation at successive times, namely at hours (hrs) 2, 4, 6, 12, days 1, 2, 4, and 6 post-inoculation (pi). Whereas, control group chickens were euthanized on days 0, 1, 2, 4, and 6 pi. Tissues of brain, trachea, lung, caecal tonsil, liver, kidney, spleen, heart, proventriculus, intestine, and thymus were collected, fixed in 10% buffered formalin, embedded in paraffin, and sectioned. HS staining, immunoperoxidase staining (IPS) and in situ PCR were applied. It was concluded that at hr 2 pi, virus seemed to be inclined to trachea and respiratory tract. Meanwhile, it attacked caecal tonsils, intestine and bursa of Fabricus. While primary viraemia was ongoing, virus created footing in kidney and thymus. At hr 4 pi, proventriculus, liver, and spleen were attacked. However, at hr 6 pi, brain and heart were involved. Secondary viraemia probably started as early as hr 12 pi since all collected tissues were positive. Tissue tropism was determined in trachea, caecal tonsil, liver, bursa of Fabricius, intestine, proventriculus, lung, spleen, thymus, kidney, heart, and brain.

Infectious bursal disease virus tissue tropism and pathogenesis of the infection in chickens by application of in situ PCR, immunoperoxase and HE staining.

Infectious bursal disease is one of an OIE list of notifiable diseases. Chicken is the only host that manifests clinical signs and its pathogenicity is correlated with the distribution of antigens in organs. This study was conducted to determine disease pathogenesis and virus tissue tropism by in situ PCR, immunoperoxidase staining (IPS), and HE staining. Twenty four chickens were infected with very virulent Infectious Bursal Disease Virus (vvIBDV). Fifteen chickens were kept as a control group. Infected chickens were sacrificed at hrs 2, 4, 6, 12, days 1, 2, 4, and 6 post-inoculation (pi). While, control chickens were euthanized on days 0, 1, 2, 4, and 6 pi. Different tissues were collected, fixed in 10% buffered formalin, and processed. At hr 2 pi, virus was detected in intestinal, junction of the proventriculus and gizzard, cecal tonsil, liver, kidney, and bursa of Fabricius. At hr 4 pi, virus reached spleen, and at hr 6 pi, it entered thymus. At hr 12 pi, virus concentration increased in positive tissues. The latest invaded tissue was muscle on day 1 pi. Secondary viraemia occurred during 12-24 h pi. In situ PCR was the most sensitive technique to highlight obscure points of infection in this study.

A Synthetic Receptor as a Specific Antidote for Paraquat Poisoning.

Accidental or suicidal ingestion of the world's most widely used herbicide, paraquat (PQ), may result in rapid multi-organ failure with a 60% fatality rate due to the absence of an effective detoxification solution. Effective, specific antidotes to PQ poisoning have been highly desired. Methods: The binding constant of PQ and a synthetic receptor, cucurbit[7]uril (CB[7]), was first determined in various pH environments. The antidotal effects of CB[7] on PQ toxicity were firstly evaluated with in-vitro cell lines. With in-vivo mice models, the pharmacokinetics and the biodistribution of PQ in major organs were determined to evaluate the influence of CB[7] on the oral bioavailability of PQ. Major organs' injuries and overall survival rates of the mice were systemically examined to evaluate the therapeutic efficacy of CB[7] on PQ poisoning. Results: We demonstrate that CB[7] may complex PQ strongly under various conditions and significantly reduce its toxicity in vitro and in vivo. Oral administration of PQ in the presence of CB[7] in a mouse model significantly decreased PQ levels in the plasma and major organs and alleviated major organs' injuries, when compared to those of mice administered with PQ alone. Further studies indicated that oral administration of CB[7] within 2 h post PQ ingestion significantly increased the survival rates and extended the survival time of the mice, in contrast to the ineffective treatment by activated charcoal, which is commonly recommended for PQ decontamination. Conclusion: CB[7] may be used as a specific oral antidote for PQ poisoning by strongly binding with PQ and inhibiting its absorption in the gastrointestinal tracts.

Direct in vivo application of induced pluripotent stem cells is feasible and can be safe.

Increasing evidence suggests the consensus that direct in vivo application of induced pluripotent stem cells (iPSCs) is infeasible may not be true. Methods: Teratoma formation and fate were examined in 53 normal and disease conditions involving brain, lung, liver, kidney, islet, skin, hind limb, and arteries. Results: Using classic teratoma generation assays, which require iPSCs to be congregated and confined, all mouse, human, and individualized autologous monkey iPSCs tested formed teratoma, while iPSC-derived cells did not. Intravenously or topically-disseminated iPSCs did not form teratomas with doses up to 2.5×108 iPSCs/kg and observation times up to 18 months, regardless of host tissue type; autologous, syngeneic, or immune-deficient host animals; presence or absence of disease; disease type; iPSC induction method; commercial or self-induced iPSCs; mouse, human, or monkey iPSCs; frequency of delivery; and sex. Matrigel-confined, but not PBS-suspended, syngeneic iPSCs delivered into the peritoneal cavity or renal capsule formed teratomas. Intravenously administered iPSCs were therapeutic with a dose as low as 5×106/kg and some iPSCs differentiated into somatic cells in injured organs. Disseminated iPSCs trafficked into injured tissue and survived significantly longer in injured than uninjured organs. In disease-free animals, no intravenously administered cell differentiated into an unwanted long-lasting cell or survived as a quiescent stem cell. In coculture, the stem cell medium and dominant cell-type status were critical for iPSCs to form cell masses. Conclusion: Teratoma can be easily and completely avoided by disseminating the cells. Direct in vivo iPSC application is feasible and can be safe.

The untranslated regions of EV-A71 contribute to its pathogenicity and virulence.

Enterovirus A71 (EV-A71) is known for its manifestation as hand foot and mouth disease (HFMD), which has caused countless large-scale epidemic outbreaks throughout the world. However, the molecular pathogenesis of EV-A71 infection is still elusive. Previous studies found that the biological characteristics of a mild EV-A71 strain (SDLY1) and a severe EV-A71 strain (SDLY107) are significantly different, and sequence analysis showed that there are several differences in nucleotide sites of UTRs (88 nt, 123 nt, 143 nt, 154 nt, 187 nt, 241 nt, 243 nt, 253 nt, 291 nt, 438 nt, 440 nt, 571 nt, 579 nt, 602 nt, 658 nt, 664 nt, 690 nt, 696 nt, 7328 nt, 7335 nt, 7367 nt, and 7395 nt). The aim of this study was to determine whether these amino sites in UTRs are associated with the pathogenesis of EV-A71 and are responsible for different clinical manifestations. Based on the reverse genetics technology, we rescued two chimeric viruses SDLY107(1-5'UTR) and SDLY107(1-3'UTR) by replacing 5'UTR/3'UTR gene fragments of an infectious cDNA clone. Replication kinetics and cytotoxicity assays showed that the virulence of the two chimeric strains significantly changed in vitro. The viral loads of the two chimeric strains in infected ICR mice were reduced and pathological damage in the brains, lungs, intestinal tissues, and muscles were lightened. Our findings suggest that some nucleotide sites in UTRs may have a function in the pathogenicity and virulence of EV-A71.

The importance of Toll-like receptor 4 during experimental Sporothrix brasiliensis infection.

Here we investigated the importance of Toll-like receptor 4 (TLR-4) in innate immune response to Sporothrix brasiliensis, a virulent fungus of Sporothrix spp. In vitro assays, using C57Bl/6 (wild type [WT]) bone marrow-derived macrophages (BMDMs), and TLR-4 knockout (TLR-4-/-) showed that the absence of TLR-4 resulted in impaired phagocytosis and lower levels of tumor necrosis factor α (TNF-α), interleukin (IL)-6, and nitric oxide. In vivo assays were also performed, and the mice (WT and TLR-4-/-) were intraperitoneally infected with S. brasiliensis yeast ATCC MyA-4831 and euthanized on days 7, 14, and 28 postinfection, with the following parameters evaluated: fungal burden in liver, spleen, kidney, and brain, and the production of cytokines interferon γ (IFN-γ), TNF-α, IL-2, IL-4, IL-6, and IL-10. The results demonstrate the macrophages dependency on TLR-4 for inflammatory activation and in the absence of TLR-4 during experimental S. brasiliensis infection enhanced dissemination occurred after 14 and 28 days. These data show that TLR-4 signals are important for the recognition of S. brasiliensis by macrophages, and their absence promotes the persistence of the infection.