Dpto. Anatomía y Anatomía Patológica Comparadas y Toxicología
Universidad de Córdoba
Pedro López-López, María A. Risalde, María Casares-Jiménez, Javier Caballero-Gómez, Andrés Martín-Gómez, Javier Martínez-Blasco, Irene Agulló-Ros, Mario Frías, Ignacio García-Bocanegra, José C. Gómez-Villamandos,et al. Elsevier BV
Irene Agulló-Ros, Marisa Andrada, Marta Pérez-Sancho, Álvaro Roy, Javier Bezos, Thomas Bonnet, Inmaculada Moreno, Yania Paz-Sánchez, Mercedes Domínguez, José C. Gómez-Villamandos,et al. Elsevier BV
Pedro López‐López, Mario Frias, Angela Camacho, Isabel Machuca, Javier Caballero‐Gómez, María A. Risalde, Ignacio García‐Bocanegra, Ignacio Pérez‐Valero, Jose C. Gomez‐Villamandos, Antonio Rivero‐Juárez,et al. Wiley
The aim of our study was to evaluate HEV antibody kinetics in HIV/HCV-coinfected patients with cirrhosis. A longitudinal retrospective study was designed. Patients were followed up every 6 months, anti-HEV IgG and IgM antibodies levels and HEV-RNA by qPCR were analyzed. The prevalence and incidence of every HEV infection marker were calculated. The kinetics of anti-HEV IgG and IgM during the follow-up were evaluated. Seventy-five patients comprised the study population. The seroprevalence observed was 17.3%. None showed IgM antibodies or HEV-RNA at baseline. None showed detectable HEV viral load during the study period. After a median follow-up of 5.1 years, 2 of 62 seronegative patients (3.2%) seroconverted to IgG antibody. The incidence for IgM was 2.7%. Of the 13 patients with IgG seropositivity at baseline, 5 (38.5%) seroreverted. Meanwhile, of the 2 patients who exhibited IgM positivity during the study, 1 (50%) showed intermittent positivity. We found that HEV seropositivity is common in HIV/HCV-coinfected cirrhotic patients. A remarkable rate of IgG seroreversions and IgM intermittence was found, limiting the use of antibodies for the diagnosis of HEV infection in this population. This article is protected by copyright. All rights reserved.
Antonio Rivero‐Juárez, Alejandro Dashti, Mónica Santín, Pamela C. Köster, Pedro López‐López, María A. Risalde, Ignacio García‐Bocanegra, José Carlos Gómez‐Villamandos, Javier Caballero‐Gómez, Mario Frías,et al. Wiley
Enteropathogenic parasites can infect a wide range of mammals, including humans, supposing an important zoonotic risk. Hepatitis E virus (HEV) is an emerging foodborne pathogen of increasing public health relevance, affecting both human and animal populations. Because both microorganisms share faecal-oral transmission route they may constitute an excellent model to evaluate the interplay between them. Thus, we aim to evaluate the viral-parasite interactions at the enteric interface in swine. We included pigs of two different breeds farming in South Spain under different production systems. We compared the HEV prevalence by the presence of Giardia duodenalis, Cryptosporidium spp., Balantioides coli, Blastocystis sp., and Enterocytozoon bieneusi in faecal samples. The HEV prevalence was 13.1 (62 out 475, 95% CI: 10.2-16.4). Those pigs infected with Cryptosporidium spp. showed a higher prevalence of HEV (30.8% vs. 12%; p = 0.012). In the same way, animals bearing E. bieneusi seem to have a higher rate of HEV infection (24.2% vs. 12.2%; p = 0.06). According to their location in the gut, animals bearing intracellular enteroparasites showed a higher HEV prevalence than those uninfected (29.6% vs. 12.7%; p = 0.038), meanwhile those carrying extracellular enteroparasites had a lower likelihood to be infected by HEV than those uninfected (12.1% vs. 23.1%; p = 0.071). Those animals bearing both type of enteroparasites showed a similar prevalence of HEV infection than those exhibiting negative for both (20.8% vs. 26.1%; p = 0.763). Our study provides evidence that intracellular and extracellular enteroparasites modulate the susceptibility to HEV infection in pigs. Meanwhile, the presence of extracellular enteroparasites shows a protective effect on the risk of HEV acquisition in swine, whereas intracellular enteroparasites seems to have the opposite effect, favouring the HEV infection. This article is protected by copyright. All rights reserved.
Alejandro Dashti, Antonio Rivero‐Juárez, Mónica Santín, Nadja S. George, Pamela C. Köster, Pedro López‐López, María A Risalde, Ignacio García‐Bocanegra, Jose Carlos Gómez‐Villamandos, Javier Caballero‐Gómez,et al. Wiley
Numerous protist species are shared between humans and pigs. Among those, Giardia duodenalis, Cryptosporidium spp., and Balantioides coli have a clear public and animal health significance. For other such as Enterocytozoon bieneusi and Blastocystis sp., their impact in animal health has not been fully stablished. Little information is currently available on the molecular diversity of these protists in swine populations. To fill this gap, we molecularly assessed G. duodenalis, Cryptosporidium spp., B. coli, Blastocystis sp., and E. bieneusi in faecal samples from Iberian and Large White pigs raised under different (intensive and/or extensive) management systems in southern Spain. A total of 151 extensively raised Iberian pigs, 140 intensively raised Iberian pigs, and 184 intensively raised Large White pigs were investigated. Blastocystis sp. was the agent most prevalently found (47.8%), followed by B. coli (45.5%), G. duodenalis (10.7%), E. bieneusi (6.9%), and Cryptosporidium spp. (5.5%). Blastocystis sp. was significantly less prevalent in intensively raised Iberian pigs (22.9%) than in their extensively raised counterparts (51.0%) or in intensively raised Large White pigs (64.1%). A significant higher prevalence was found for G. duodenalis), Cryptosporidium spp., and E. bieneusi in Large White pigs than Iberian pigs. Balantioides coli was similarly distributed (40.0-51.1%) in all three investigated swine populations. Sequence analyses revealed the presence of G. duodenalis assemblage E, two Cryptosporidium species (Cryptosporidium scrofarum and Cryptosporidium suis), B. coli (genotypes A and B), Blastocystis sp. (ST1, ST3, and ST5), E. bieneusi (EbpA, EbpC, EbpD, O, and a novel genotype named PigSpEb2). Novel genotype PigSpEb2 was found alone or in combination with EbpA. Data suggest a widespread exposure to protist enteroparasites in domestic pig populations irrespectively of breed and raising management system. Many of the species/genotype identified have zoonotic potential and might represent a public health concern. This article is protected by copyright. All rights reserved.
Maria A Risalde, Ana Mª Molina, Antonio J Lora, Nahum Ayala, Jose C Gómez-Villamandos, and Mª Rosario Moyano Elsevier BV
Bisphenol A (BPA) is used to produce plastic and plastic derived products in multitude of daily utensils, being one of the industrial compounds most widely used. This endocrine disrupting chemical (EDCs) is a well-known environmental pollutant released into the aquatic environment from industrial wastewater, sewage sludge or landfill leachate. Aromatases are considered potential targets of EDCs with characteristics that make them suitable biomarkers of exposure to their effects. The main objective of our study was to evaluate the expression of cyp19a aromatase as a toxicological endpoint after BPA exposure through the identification and assessment of alterations of the main cells responsible for cyp19a1a and cyp19a1b expression in the zebrafish ovary and brain using different concentrations of BPA in water. Immunohistochemistry was used to analyze the expression of these enzymes in female zebrafish exposed and not exposed to different concentrations of BPA (1, 10, 100 and 1000 μg / L) in water (n = 6/group) for 14 days. The results obtained in this study showed that the cyp19a aromatase system, involved in the synthesis of steroid compounds, is specially located in distinct oocyte stages in the ovary (cyp19a1a) and in radial glial cells of the brain (cyp19a1b). An overexpression of these aromatases was observed after BPA exposure in zebrafish, peaking from a concentration of 10 µg/L and showing to be good biomarkers of exposure to identify the early effects of low BPA concentrations. To our knowledge, this study is the first to localize and quantify the expression of cyp19a1a and cyp19a1b in the cells of brain and ovary after fish exposure to different BPA concentrations in water.
Maria A Risalde, Antonio Rivero‐Juarez, Mario Frias, Israel Olivas, Pedro Lopez‐Lopez, Ignacio García‐Bocanegra, Teresa Brieva, Javier Caballero‐Gómez, Angela Camacho, Vicente Fernández‐Molera,et al. Wiley
Background Identifying pig farms infected with hepatitis E virus (HEV) is a key aspect to implement surveillance programmes for this emerging zoonotic agent. Detection of HEV in blood has several drawbacks, including animal handling, economic costs and animal stress. The objective of this study was to evaluate the effectiveness of a non-invasive screening approach for determining the HEV status of pig farms under different management systems. Methods Forty stool samples randomly collected from the pen floor of 17 intensive pig farms and the yard of nine extensive ones were tested for HEV RNA. The invasive method used to confirm the HEV status of the farm was HEV RNA analysis of serum samples randomly collected from 40 animals on each farm. Results Twenty-one HEV-positive farms were detected by invasive and non-invasive methods. No positive serum or stool samples were detected on five intensive farms. A high intertest agreement (K=1; P<0.00001) was observed between both methodologies, showing the stool screening approach a 100 per cent of sensitivity and specificity with respect to the invasive method. Likewise, a significant negative relationship was observed between the HEV within-farm prevalence and the number of the first HEV-positive stool sample found (Spearman’s rho=−0.64; P=0.0004). This negative relationship was higher in intensively managed farms. Conclusion This non-invasive screening approach could be reliably applied in a large-scale surveillance programme for determining the HEV status of pig farms under different management systems.
María A. Risalde, Fernando Romero-Palomo, Cristina Lecchi, Fabrizio Ceciliani, Chiara Bazzocchi, Stefano Comazzi, Martina Besozzi, Jose C. Gómez-Villamandos, and Camilla Luzzago Elsevier BV
Bovine viral diarrhea virus (BVDV) has been detected in peripheral blood mononuclear cells (PBMCs) of immunocompetent animals, not being clear whether the development of a specific humoral immune response can prevent BVDV infection. The aim of this study was to evaluate the ability of non-cytopathic BVDV to replicate and produce infectious virus in PBMCs from calves pre-infected with BVDV and to elucidate the immunomodulatory effect of BVDV on these cells in an in vitro model. Quantification of virus was by quantitative PCR, while its replicative capacity and shedding into the extracellular environment was evaluated by viral titration. Apoptosis was assessed by flow cytometry analysis of annexin V and propidium iodide, and by expression of caspase-3/7. Flow cytometry was used to analyze the expression of CD14/CD11b/CD80, CD4/CD8/CD25, MHC-I/MHC-II and B-B2 markers. Our results showed that PBMCs from cattle naturally infected with BVDV were more susceptible to in vitro BVDV infection and showed a more severe apoptosis response than those from naïve animals. Non-cytopathic BVDV in vitro infection also resulted in a lack of effect in the expression of antigen presentation surface markers. All these findings could be related to the immunosuppressive capacity of BVDV and the susceptibility of cattle to this infection.
Ignacio García‐Bocanegra, Antonio Rivero, Javier Caballero‐Gómez, Pedro López‐López, David Cano‐Terriza, Mario Frías, Saul Jiménez‐Ruiz, Maria A. Risalde, Jose C. Gómez‐Villamandos, and Antonio Rivero‐Juarez Wiley
Hepatitis E (HE) is an important emerging disease in European countries. To analyse the role of equids as potential reservoirs for HE virus (HEV), we determined the prevalence of HEV infection in 861 equines from 464 herds in Spain. HEV RNA in serum was detected in 0.4% (3/692) of horses, 1.2% (1/86) of donkeys and 3.6% (3/83) of mules. Phylogenetic analysis identified the zoonotic genotype 3 as being closely related to viral human and swine strains. In this first report on HEV in equids in Europe, we confirm the susceptibility of horses, donkeys and mules to HEV infection. The low prevalence detected indicates that equids may be considered spillover hosts rather than true reservoirs.
Alejandro Núñez, Pedro J. Sánchez-Cordón, Miriam Pedrera, Jose C. Gómez-Villamandos, and Librado Carrasco Wiley
Classical swine fever (CSF) is a highly contagious and often fatal viral disease of domestic pigs and wild boar. Pulmonary oedema and haemorrhages in lung parenchyma are common lesions in the acute forms of CSF that may compromise pig survival and whose pathogenetic mechanisms remain unclear. The appearance of pulmonary lesions in pigs infected with Alfort/187 strain of classical swine fever virus (CSFV) euthanized between 2 and 17 days postinfection (dpi) and the role played by cytokines secreted by different pulmonary macrophage populations in the evolution of lesions was evaluated in this study. Microscopic changes of alveolar septal thickening along with oedema and haemorrhages became more severe at middle-late stages of the experiment. A significant increase in the number of pulmonary macrophages, mainly pulmonary intravascular macrophages (PIMs), was observed coinciding with the onset of alveolar septal thickening from Day 4 pi. PIMs were the main target of CSFV from initial stages of infection while the presence of infected pulmonary alveolar macrophages (PAMs) was scarce and late. Initial infection of PIMs induced phagocytic and biosynthetic activation with subsequent release of chemotactic cytokines. TNFα and, to a lesser extent, IL-1α secreted by PIMs were the major cytokines involved, while IL-6 played only a minor role. On the contrary, results suggested only a secondary role of PAMs as source of cytokines. The presence of vascular changes from Day 9 pi coincided with the highest levels of infected PIMs and the highest number of pro-inflammatory cytokines secreting PIMs. Activation and phagocytosis of platelets were observed in the lungs of infected pigs from early stages, also coinciding with the expression of cytokines with a proven procoagulant activity. The existence of intravascular coagulation phenomena in lung was ruled out.
Pedro Lopez-Lopez, Maria de los Angeles Risalde, Mario Frias, Ignacio García-Bocanegra, Teresa Brieva, Javier Caballero-Gomez, Angela Camacho, Vicente Fernández-Molera, Isabel Machuca, Jose Carlos Gomez-Villamandos,et al. Elsevier BV
Pigs are considered important reservoirs of HEV and so constitute a major risk of transmission to humans, either via direct contact or by consuming raw or undercooked contaminated pork products. Once the scale of this disease on European pig farms has been estimated, the identification of risk factors associated with HEV infection in these species could help determine contingency strategies to minimize the risk of transmission to humans. Our objective was to evaluate risk factors associated with HEV in pigs under different production systems. We included 1040 pigs from 26 farms. The prevalence of HEV infection in the study population, evaluated by RT-qPCR, was calculated, then studied according to animal and farm characteristics. Factors associated with HEV infection were analyzed by multivariate analysis. One hundred and seventy-two pigs were infected by HEV, which gave an individual prevalence of 16.5% (95% CI: 14.4%-18.9%). Factors associated with higher prevalence of HEV infection were: extensive farming [23.9%; OR = 2.239 (1.036-4.837)], absence of sanitary ford [33.8%; OR = 3.597 (1.649-7.850)], no quarantine period [20.8%; OR = 2.723 (1.450-5.112)], and contact with domestic species [24.5%; OR = 3.893 (1.453-10.431)]. Our evidence showed that pigs reared on extensive farms are at a higher risk of HEV infection than those reared intensively. The use of control measures could reduce the risk of HEV infection in pigs and minimize the risk of zoonotic transmission.
A. Rivero-Juarez, M. Frias, P. Lopez-Lopez, A. Martinez-Peinado, M. Á. Risalde, T. Brieva, I. Machuca, Á. Camacho, I. García-Bocanegra, J. C. Gomez-Villamandos,et al. Wiley
Diagnosis of acute hepatitis E virus (HEV) infection is established by detection of anti‐HEV IgM antibodies by ELISA or by amplification of serum viral RNA. Here, we evaluate the diagnostic value of testing HEV RNA in saliva to identify patients with acute HEV infection. Prospective proof‐of‐concept study including patients with acute hepatitis. Whole blood and neat saliva samples were obtained from all patients. Saliva samples were processed and analysed for HEV RNA by RT‐PCR within 2 hr after collection. A total of 34 patients with acute hepatitis and 12 healthy donors were included in the study. HEV RNA in serum was confirmed by RT‐PCR in eight of these patients (23.5%; 95% CI: 12.2%–40.2%). HEV was isolated in the saliva of eight of 34 patients (23.5%; 95% CI: 12.2%–40.2%). All patients with HEV RNA amplified in saliva had detectable HEV RNA in serum. HEV was isolated neither in the saliva of any of the 26 patients without detectable HEV RNA in serum nor in healthy donors. Our study suggests that acute HEV infection could be diagnosed by assessing viral load in saliva.
Antonio Rivero-Juarez, María A. Risalde, Mario Frias, Ignacio García-Bocanegra, Pedro Lopez-Lopez, David Cano-Terriza, Angela Camacho, Saul Jimenez-Ruiz, Jose C. Gomez-Villamandos, and Antonio Rivero Springer Science and Business Media LLC
BackgroundIt has been shown that wildlife can serve as natural reservoirs of hepatitis E virus (HEV). The wild boar (Sus scrofa) is probably the main natural reservoir of HEV and could therefore represent an important route of transmission in Europe, especially in regions where game meat is widely consumed. We evaluated the prevalence of HEV infection in wild boar in the south of Spain, with the aim of identifying associated risk factors. A cross-sectional study that included hunted wild boar was carried out during the 2015/2016 hunting season (October 15 to February 15) in Andalusia (southern Spain). The outcome variable was HEV infection, defined as amplification of HEV RNA in serum by RT-PCR.ResultsA total of 142 animals, selected from 12 hunting areas, were included and formed the study population. Thirty-three wild boars (23.2%; 95% CI: 16.8%–30.7%) were positive for HEV infection. Prevalence peaked in October and November, then gradually declined until the end of December. After multivariate analysis, only hunting date was independently associated with HEV infection across sex and age.ConclusionsOur study found a relatively high prevalence of HEV infection in wild boar in the south of Spain, suggesting that prevalence may depend on the season when the animal is hunted. In consequence, the potential risk of zoonotic transmission could fluctuate.
María A. Risalde, Antonio Rivero-Juárez, Fernando Romero-Palomo, Mario Frías, Pedro López-López, David Cano-Terriza, Ignacio García-Bocanegra, Saúl Jiménez-Ruíz, Ángela Camacho, Isabel Machuca,et al. Public Library of Science (PLoS)
Hepatitis E virus (HEV) is an emerging zoonotic pathogen with pigs and wild boar serving as reservoirs for human infection through direct contact with infected animals or the consumption of raw or undercooked pork products. The liver is considered the main target site of HEV replication in swine and an important organ in the pathogenesis of the disease. The aim of this study was to characterize the target liver cells for HEV entry in naturally infected wild boar and to evaluate the type and severity of the pathological changes in order to reach a better understanding of the hepatic pathogenic mechanisms involved in hepatitis E. In total, 58 livers from hunted wild boar were histopathologically evaluated. The presence of specific HEV antibodies in serum was determined by indirect ELISA. Immunohistochemistry was used for the detection of HEV antigen and Real time RT-PCR to detect HEV RNA in liver and serum. HEV seroprevalence in these animals was of 5.197% (CI95%: 1.77–14.14). By Real time RT-PCR, HEV was detected in the liver tissue of four wild boar (6.8%; CI95%: 2.7–16.4) and only one animal was also positive in serum (1.7%; CI95%: 0.3–9.1). The non-viremic animals naturally infected with HEV presented evidence of liver infection, mainly in Kupffer cells and liver sinusoidal endothelial cells, without apparent associated hepatitis lesions. This study supports the hypothesis that low viral titers may persist in the liver of non-viremic individuals, giving thus the possibility of consumption of contaminated liver of animals diagnosed as HEV-negative in serum. Further immunopathogenic studies are necessary to elucidate the mechanisms responsible for this process and to evaluate the protocols of HEV diagnosis in animals destined for human consumption.
A. Rivero-Juarez, M. Frias, A. Martinez-Peinado, M. A. Risalde, D. Rodriguez-Cano, A. Camacho, I. García-Bocanegra, F. Cuenca-Lopez, J. C. Gomez-Villamandos, and A. Rivero Wiley
An HIV‐infected patient was diagnosed with acute hepatitis E infection in our hospital. An epidemiological inquiry was performed to collect demographic, food and animal exposure variables in order to identify the potential route of transmission. The patient reported that his family traditionally hunted wild boar for food. All family members were analysed for hepatitis E virus infection. Additionally, route of transmission by wild boar meat consumption and prevalence of HEV infection among wild boar from the same hunting area were investigated. In all‐family members (n = 8), HEV‐RNA was amplified. Two wild boar meat slices consumed was analysed, showing the presence of HEV. The virus isolated was consistent with genotype 3, revealing 100% homology between family members and meat. Additionally, we tested nine wild boar hunted in the same hunting area. All of them were RNA‐HEV positive, isolating the same HEV genotype 3 viral strain. We demonstrated by phylogenetic analysis zoonotic transmission of HEV by wild boar meat consumption. The prevalence of HEV infection among wild boar found in our study suggests that this species is an important route of transmission to human.
Antonio Rivero-Juarez, Mario Frias, Pedro Lopez-Lopez, María de Los Angeles Risalde, Teresa Brieva, Isabel Machuca, Angela Camacho, Antonio Martinez-Peinado, Jose Carlos Gomez-Villamandos, and Antonio Rivero Oxford University Press (OUP)
Although hepatitis E virus (HEV) is regarded as a self-limiting infection and anti-HEV antibodies seem to protect against reinfection, its pathogenesis is not well established. We describe 2 cases of acute symptomatic HEV infection after hepatitis C therapy in patients carrying anti-HEV immunoglobulin G antibodies, raising 2 major questions: reactivation or reinfection?
F. Romero-Palomo, M. A. Risalde, and J. C. Gómez-Villamandos Wiley
The aim of this work was to investigate the effect of pre-infection with bovine viral diarrhoea virus (BVDV) on thymus immune cells from calves challenged with bovine herpesvirus 1 (BHV-1). Twelve Friesian calves, aged 8 to 9 months, were inoculated with non-cytopathic BVDV-1. Ten of them were subsequently challenged with BHV-1 and euthanized in batches of two at 1, 2, 4, 7 or 14 dpi with BHV-1. The other two calves were euthanized prior to the second inoculation and were used as BVDV-infected controls. A further 10 calves were inoculated solely with BHV-1 and euthanized at the same time points. Two calves were not inoculated with any agent and were used as negative controls. Quantitative changes in immune cells were evaluated with immunohistochemical methods to compare coinfected calves and calves challenged only with BHV-1. The results of this study pointed out BVDV as responsible for the thymic lesions observed in the experiment as well as for the majority of immunopathologic changes, including a downregulation of Foxp3 lymphocytes and TGFβ, which reverted as BVDV was cleared, and an overexpression of medullary CD8+ T cells. However, despite not inducing evident lesions in the thymus, BHV-1 seemed to prompt some immune alterations. Collectively, these data contribute to the knowledge on the immunopathologic alterations of the thymus during BVDV infections, and its importance in the development of secondary infections.
M. A. Risalde, V. Molina, P. J. Sánchez-Cordón, F. Romero-Palomo, M. Pedrera, and J. C. Gómez-Villamandos SAGE Publications
The aim of this work was to study the interstitial aggregates of immune cells observed in pulmonary parenchyma of calves preinfected with bovine viral diarrhea virus and challenged later with bovine herpesvirus 1. In addition, the intent of this research was to clarify the role of bovine viral diarrhea virus in local cell-mediated immunity and potentially in predisposing animals to bovine respiratory disease complex. Twelve Friesian calves, aged 8 to 9 months, were inoculated with noncytopathic bovine viral diarrhea virus genotype 1. Ten were subsequently challenged with bovine herpesvirus 1 and euthanized at 1, 2, 4, 7, or 14 days postinoculation. The other 2 calves were euthanized prior to the second inoculation. Another cohort of 10 calves was inoculated only with bovine herpesvirus 1 and then were euthanized at the same time points. Two calves were not inoculated with any agent and were used as negative controls. Pulmonary lesions were evaluated in all animals, while quantitative and biosynthetic changes in immune cells were concurrently examined immunohistochemically to compare coinfected calves and calves challenged only with bovine herpesvirus 1. Calves preinfected with bovine viral diarrhea virus demonstrated moderate respiratory clinical signs and histopathologic evidence of interstitial pneumonia with aggregates of mononuclear cells, which predominated at 4 days postinoculation. Furthermore, this group of animals was noted to have a suppression of interleukin-10 and associated alterations in the Th1-driven cytokine response in the lungs, as well as inhibition of the response of CD8+ and CD4+ T lymphocytes against bovine herpesvirus 1. These findings suggest that bovine viral diarrhea virus preinfection could affect the regulation of the immune response as modulated by regulatory T cells, as well as impair local cell-mediated immunity to secondary respiratory pathogens.
F. Romero-Palomo, M.A. Risalde, V. Molina, S. Lauzi, M.J. Bautista, and J.C. Gómez-Villamandos Elsevier BV
Since the thymus is a target organ for the bovine viral diarrhea virus (BVDV), our experiment aimed to understand its relationship with the immunosuppressive effect by studying the consequences of a previous infection with BVDV on the thymus of calves challenged with bovine herpesvirus 1.1 (BHV-1). For this purpose, 12 animals were inoculated intranasally with non-cytopathic BVDV-1; 12 days later, 10 of them were coinfected intranasally with BHV-1. These animals were euthanized in batches of two at 0, 1, 2, 4, 7 or 14 dpi with BHV-1. Another 10 calves were inoculated solely with BHV-1 and euthanized in batches of two at 1, 2, 4, 7 or 14 dpi with BHV-1; two uninoculated calves were used as negative controls. Thymus samples from these animals were processed for viral detection and histopathological, immunohistochemical, and ultrastructural studies focused on BVDV/BHV-1 antigens, cortex:medulla ratio, apoptosis (TUNEL and caspase-3), collagen deposition, and factor VIII endothelial detection. Our study revealed the immunohistochemical presence of BVDV antigen in all animals in the BVDV-infected group, unlike BHV-1 detection, which was observed in animals in both infection groups only by molecular techniques. BVDV-preinfected animals showed severe atrophic changes associated with reduced cortex:medulla ratio, higher presence of cortical apoptosis, and increased collagen deposition and vascularization. However, calves solely infected with BHV-1 did not show atrophic changes. These findings could affect not only the numbers of circulating and local mature T cells but also the T cell-mediated immunity, which seems to be impaired during infections with this virus, thus favoring pathogenic effects during secondary infections.
P.J. Sánchez-Cordón, A.C. Pérez de Diego, J.C. Gómez-Villamandos, J.M. Sánchez-Vizcaíno, F.J. Pleguezuelos, B. Garfia, P. del Carmen, and M. Pedrera Elsevier BV
Protective immunity in sheep with bluetongue virus (BTV) infection as well as the role of BTV-induced cytokines during immune response remains unclear. Understanding the basis immunological mechanisms in sheep experimentally infected with serotypes 1 and 8 (BTV-1 and -8) was the aim of this study. A time-course study was carried out in order to evaluate cell-mediated immune response and serum concentrations of cytokines (IL-1β, TNFα, IL-12, IFNγ, IL-4 and IL-10) with inflammatory and immunological functions. Depletion of T cell subsets (mainly CD4(+), γδ and CD25(+)) together with the absence of cytokines (IFNγ and IL-12) involved in the regulation of cell-mediated antiviral immunity at the first stage of the disease suggested that both BTV-1 and BTV-8 might impair host's capability against primary infections which would favor viral replication and spreading. However, cellular immune response and cytokines elicited an immune response in sheep that efficiently reduced viremia in the final stage of the experiment. Recovery of T cell subsets (CD4(+) and CD25(+)) together with a significant increase of CD8(+) T lymphocytes in both infected groups were observed in parallel with the decrease of viremia. Additionally, the recovery of CD4(+) T lymphocytes together with the significant increase of IL-4 serum levels at the final stage of the experiment might contribute to humoral immune response activation and neutralizing antibodies production against BTV previously described in the course of this experiment. These results suggested that both cellular and humoral immune response may contribute to protective immunity against BTV-1 and BTV-8 in sheep. The possible role played by IL-10 and CD25(+) cells in controlling inflammatory and immune response in the final stage of the experiment has also been suggested.
A. I. Raya, J. C. Gomez-Villamandos, and M. J. Bautista SAGE Publications
Thymic epithelial cells could play an important role in lymphoid depletion during bovine viral diarrhea virus (BVDV) infection. To evaluate this hypothesis, we examined proliferation of lymphocytes, expression of cytokeratins by thymic epithelial cells, and ultrastructural features at sequential time points after experimental infection of colostrum-deprived calves with the noncytopathogenic BVDV1 strain 7443. Ten clinically healthy Friesian calves were used. Eight were inoculated with the virus, and 2 were used as uninfected controls. Calves were sedated and euthanized in batches between 3 and 14 days postinoculation. At necropsy, thymus samples were collected for structural, immunohistochemical, and ultrastructural study. Thymic lymphoid depletion was accompanied by a decrease in lymphocyte proliferation and immunohistochemical and ultrastructural changes in thymic epithelial cells. Immunohistochemical and ultrastructural results reflect a disturbance of the thymic epithelial cell network, which may explain the decrease in lymphocyte proliferation by defective thymocyte-epithelial cell interactions.
J.M. Sánchez-Vizcaíno, L. Mur, J.C. Gomez-Villamandos, and L. Carrasco Elsevier BV
African swine fever (ASF) is one of the most important infectious diseases of swine and has major negative consequences for affected countries. ASF is present in many sub-Saharan countries, Sardinia and several countries of eastern and central Europe, where its continuous spread has the swine industry on heightened alert. ASF is a complex disease for which no vaccine or treatment is available, so its control is based on early detection and rapid control of spread. For a robust and reliable early detection programme it is essential to be able to recognize the clinical signs and pathological changes of ASF, keeping in mind that in most cases the first introductions don't show high mortality nor characteristic clinical signs or lesions, but fever and some hemorrhagic lymph nodes. Knowledge of the main characteristics of this infection, including its current distribution and routes of transmission, is also essential for preventing and controlling ASF. This review addresses each of these topics and aims to update knowledge of the disease in order to improve early detection of ASF in the field and allow implementation of public health programmes.
V. Molina, M. A. Risalde, P. J. Sánchez-Cordón, F. Romero-Palomo, M. Pedrera, B. Garfia, and J. C. Gómez-Villamandos Wiley
Acute infections with bovine viral diarrhoea virus (BVDV), a major pathogen of cattle, are often asymptomatic or produce only mild clinical symptoms. However, they may play an important role in the bovine respiratory disease complex by exerting a marked immunosuppressive effect, as a result of the death of the immunocompetent cell populations involved in controlling innate and adaptive immune responses, together with a marked reduction of both cytokine expression and co-stimulatory molecule synthesis. Although experimental research and field studies have shown that acute BVDV infection enhances susceptibility to secondary infection, the precise mechanism involved in BVDV-induced immunosuppression remains unclear. The present study is aimed at measuring a range of blood parameters in a single group of fourteen calves infected with non-cytopathic BVDV-1. Focus has been put on those related to the cell-mediated immune response just as leucocyte populations and lymphocyte subpopulations, serum concentrations of cytokines (IL-1β, TNF-α, IFN-γ, IL-12, IL-4 and IL-10) and acute phase proteins [haptoglobin, serum amyloid A (SAA), fibrinogen and albumin], as well as BVDV-specific antibodies and viremia. After non-cytopathic BVDV-1 infection, clinical signs intensity was never more than moderate coinciding with the presence of viremia and leucocyte and lymphocyte depletion. An early increase in TNF-α, IFN-γ and IL-12 levels in contrast to IL-1β was observed in line with a raise in haptoglobin and SAA levels on the latest days of the study. As regards IL-4 levels, no evidence was found of any changes. However, a slight increase in IL-10 was observed, matching up the TNF-α decline during the acute phase response. These findings would help to increase our knowledge of the immune mechanisms involved in acute infection with non-cytopathic BVDV-1 strains, suggesting the existence of a clear tendency towards a type 1 immune response, thereby enhancing resistance against viral infections.
F. Romero-Palomo, M. A. Risalde, V. Molina, P. J. Sánchez-Cordón, M. Pedrera, and J. C. Gómez-Villamandos SAGE Publications
Dendritic cells (DCs) are “professional” antigen-presenting cells with a critical role in the regulation of innate and adaptive immune responses and thus have been considered of great interest in the study of a variety of infectious diseases. The objective of this investigation was to characterize the in vivo distribution of DCs in bovine tissues by using potential DC markers to establish a basis for the study of DCs in diseased tissues. Markers evaluated included MHCII, CD208, CD1b, CD205, CNA.42, and S100 protein, the latter 2 being expressed by follicular dendritic cells whose origin and role are different from the rest of hematopoietic DCs. Paraffin wax–embedded tissues from 6 healthy Friesian calves were subjected to the avidin-biotin-peroxidase method, and the most appropriate fixatives, dilutions, and antigen retrieval pretreatments were studied for each of the primary antibodies. The most significant results included the localization of CD208-positive cells not only in the T zone of lymphoid organs but also within lymphoid follicles; CD1b-positive cells were mainly found in thymus and interfollicular areas of some lymph nodes; cells stained with anti–CD205 antibody were scarce, and their location was mainly in nonlymphoid tissues; and CNA.42- and S100 protein–positive cells localized in primary lymphoid follicles and light zones of germinal centers, although showing differences in the staining pattern. Furthermore, MHCII was established as one of the most sensitive markers for any DC of hematopoietic origin. These results increase our understanding of DC immunolabeling and will help in future DC studies of both healthy and diseased tissues.