Oxidative stress in wild boars naturally and experimentally infected with mycobacterium bovis

This is an open access article distributed under the terms of the Creative Commons Attribution License.-- et al.Reactive oxygen and nitrogen species (ROS-RNS) are important defence substances involved in the immune response against pathogens. An excessive increase in ROS-RNS, however, can damage the organism causing oxidative stress (OS). The organism is able to neutralise OS by the production of antioxidant enzymes (AE); hence, tissue damage is the result of an imbalance between oxidant and antioxidant status. Though some work has been carried out in humans, there is a lack of information about the oxidant/antioxidant status in the presence of tuberculosis (TB) in wild reservoirs. In the Mediterranean Basin, wild boar (Sus scrofa) is the main reservoir of TB. Wild boar showing severe TB have an increased risk to Mycobacterium spp. shedding, leading to pathogen spreading and persistence. If OS is greater in these individuals, oxidant/antioxidant balance in TB-affected boars could be used as a biomarker of disease severity. The present work had a two-fold objective: i) to study the effects of bovine TB on different OS biomarkers (namely superoxide dismutase (SOD), catalasa (CAT), glutathione peroxidase (GPX), glutathione reductase (GR) and thiobarbituric acid reactive substances (TBARS)) in wild boar experimentally challenged with Mycobacterium bovis, and ii) to explore the role of body weight, sex, population and season in explaining the observed variability of OS indicators in two populations of free-ranging wild boar where TB is common. For the first objective, a partial least squares regression (PLSR) approach was used whereas, recursive partitioning with regression tree models (RTM) were applied for the second. A negative relationship between antioxidant enzymes and bovine TB (the more severe lesions, the lower the concentration of antioxidant biomarkers) was observed in experimentally infected animals. The final PLSR model retained the GPX, SOD and GR biomarkers and showed that 17.6% of the observed variability of antioxidant capacity was significantly correlated with the PLSR X's component represented by both disease status and the age of boars. In the samples from free-ranging wild boar, however, the environmental factors were more relevant to the observed variability of the OS biomarkers than the TB itself. For each OS biomarker, each RTM was defined as a maximum by one node due to the population effect. Along the same lines, the ad hoc tree regression on boars from the population with a higher prevalence of severe TB confirmed that disease status was not the main factor explaining the observed variability in OS biomarkers. It was concluded that oxidative damage caused by TB is significant, but can only be detected in the absence of environmental variation in wild boar.This work was funded by the PAIDI Research Group RNM18 from Junta de Andalucía. ES was supported by the postdoctoral program (SFRH/BPD/96637/2013) of the Fundação para a Ciência ea Tecnologia, Portugal. We would like to thank the University of Aveiro (Department of Biology) and FCT/MEC for the financial support to CESAM RU (UID/AMB/50017) through national funds and, where applicable, co-financed by the FEDER, within the PT2020 Partnership Agreement.Peer reviewe

Protective Effect of Intestinal Helminthiasis Against Tuberculosis Progression Is Abrogated by Intermittent Food Deprivation

Background: Tuberculosis (TB) is still a major challenge for humankind. Because regions with the highest incidence also have a high prevalence of helminthiasis and nutritional scarcity, we wanted to understand the impact of these on TB progression. Methods: We have developed an experimental murine model for active TB in C3HeB/FeJ, coinfected with Trichuris muris and Heligmosomoides polygyrus nematodes, and exposed to an environmental mycobacterium (M. manresensis) and intermittent fasting. Cause-effect relationships among these factors were explored with Partial Least Squares Path modelling (PLSPM). Results: Previous parasitization had a major anti-inflammatory effect and reduced systemic levels of ADA, haptoglobin, local pulmonary levels of IL-1β, IL-6, TNF-α, CXCL-1, CXCL-5 and IL-10. Oral administration of heat-killed M. manresensis resulted in a similar outcome. Both interventions diminished pulmonary pathology and bacillary load, but intermittent food deprivation reduced this protective effect increasing stress and inflammation. The PLSPM revealed nematodes might have protective effects against TB progression. Conclusions: Significantly higher cortisol levels in food-deprivation groups showed it is a stressful condition, which might explain its deleterious effect. This highlights the impact of food security on TB eradication policies and the need to prioritize food supply over deworming activities.The project leading to these results has received funding from la Caixa Foundation (ID 100010434), under agreement LCF/ PR/GN16/10290002. This work was supported by the Plan Nacional I + D + I co-financed by ISCIII-Subdirección General de Evaluación and Fondo-EU de Desarrollo Regional (FEDER) through PC contract IFI14/00015. LA was supported by the European Commission Horizon 2020 research and innovation program under grant agreement TBVAC2020 No. 64338. The Experimental Tuberculosis Unit is accredited by the Catalan Agency for Management of University and Research Grants (AGAUR) with code 2017 SGR500 and the IGTP is a member of the CERCA network of institutes. ES and AT are supported by the SpanishMinisterio de Ciencia Innovación y Universidades (MICINN) through a Ramon y Cajal agreements (RYC-2016-21120; RYC-2017-22992). The research activities of CG-A are supported by a research fellow linked to the RYC-2016-21120

Seasonal diet composition of Pyrenean chamois is mainly shaped by primary production waves

In alpine habitats, the seasonally marked climatic conditions generate seasonal and spatial differences in forage availability for herbivores. Vegetation availability and quality during the growing season are known to drive life history traits of mountain ungulates. However, little effort has been made to understand the association between plant phenology and changes in the foraging strategies of these mountain dwellers. Furthermore, this link can be affected by the seasonal presence of livestock in the same meadows. The objective of this work was to study the seasonal changes in diet composition of Pyrenean chamois (Rupicapra p. pyrenaica) and its relationship to primary production trends in a Mediterranean alpine environment. Moreover, diet composition in two populations with contrasting livestock pressure was compared in order to study the effect of sheep flocks on the feeding behaviour of chamois. From 2009 to 2012, monthly diet composition was estimated by cuticle microhistological analysis of chamois faeces collected in the eastern Pyrenees. The primary production cycle was assessed by remote sensing, using the Normalized Difference Vegetation Index. Additionally, the diet of sheep sharing seasonally the subalpine and alpine meadows with chamois was analysed. Diet selection of chamois and sheep and their overlap was also assessed. Our results show an intra-annual variation in the diet composition of Pyrenean chamois and demonstrate a strong relationship between plant consumption dynamics and phenology in alpine areas. In addition, Calluna vulgaris, Cytisus spp. and Festuca spp., as well as forbs in the summer, are found to be key forage species for Pyrenean chamois. Furthermore, this study couldn't detect differences between both chamois populations despite the presence of sheep flocks in only one area. However, the detection of a shift in the diet of chamois in both areas after the arrival of high densities of multi-specific livestock suggest a general livestock effect. In conclusion, Pyrenean chamois are well adapted to the variations in the seasonal availability of plants in alpine habitats but could be disturbed by the seasonal presence of livestock. Due to the key plants in their diet, we suggest that population management programmes should focus on the preservation of mixed grasslands composed of patches of shrubs and herbs. The effects of climate change and shrub expansion should be studied as they may potentially affect chamois population dynamics through changes in habitat composition and temporal shifts in forage availability

Zoonotic "Enterocytozoon bieneusi" genotypes in free-ranging and farmed wild ungulates in Spain

Microsporidia comprises a diverse group of obligate, intracellular, and spore-forming parasites that infect a wide range of animals. Among them, Enterocytozoon bieneusi is the most frequently reported species in humans and other mammals and birds. Data on the epidemiology of E. bieneusi in wildlife are limited. Hence, E. bieneusi was investigated in eight wild ungulate species present in Spain (genera Ammotragus, Capra, Capreolus, Cervus, Dama, Ovis, Rupicapra, and Sus) by molecular methods. Faecal samples were collected from free-ranging (n = 1058) and farmed (n = 324) wild ungulates from five Spanish bioregions. The parasite was detected only in red deer (10.4%, 68/653) and wild boar (0.8%, 3/359). Enterocytozoon bieneusi infections were more common in farmed (19.4%, 63/324) than in wild (1.5%, 5/329) red deer. A total of 11 genotypes were identified in red deer, eight known (BEB6, BEB17, EbCar2, HLJD-V, MWC_d1, S5, Type IV, and Wildboar3) and three novel (DeerSpEb1, DeerSpEb2, and DeerSpEb3) genotypes. Mixed genotype infections were detected in 15.9% of farmed red deer. Two genotypes were identified in wild boar, a known (Wildboar3) and a novel (WildboarSpEb1) genotypes. All genotypes identified belonged to E. bieneusi zoonotic Groups 1 and 2. This study provides the most comprehensive epidemiological study of E. bieneusi in Spanish ungulates to date, representing the first evidence of the parasite in wild red deer populations worldwide. Spanish wild boars and red deer are reservoir of zoonotic genotypes of E. bieneusi and might play an underestimated role in the transmission of this microsporidian species to humans and other animal

The physiological cost of male-biased parasitism in a nearly monomorphic mammal

Background: Even though male-biased parasitism is common in mammals, little effort has been made to evaluate whether higher parasitic burden in males results in an extra biological cost, and thus a decrease in fitness. Body condition impairment and the augmentation of oxidative stress can be used as indicators of the cost of parasite infections. Here, we examined relationships between gastrointestinal and respiratory helminths, body condition and oxidative stress markers (glutathione peroxidase, paraoxonase-1) in 28 Pyrenean chamois (Rupicapra p. pyrenaica) sampled in autumn. Results: Only male chamois showed a reduction in body condition and higher oxidative stress due to parasite infection, likely because of the extremely high parasite burdens observed in males. Conclusions: This study made evident a disparity in the physiological cost of multiple parasitism between sexes in a wild mammal, mainly due to parasitic richness. Because of the similar life expectancy in male and female chamois, we suggest that males may have developed natural mechanisms to compensate for higher parasite loads during the rut

Male-biased gastrointestinal parasitism in a nearly monomorphic mountain ungulate

Background: Pyrenean chamois (Rupicapra pyrenaica pyrenaica) is a nearly monomorphic mountain ungulate with an unbiased sex-specific overwinter adult survival. Few differences in gastrointestinal parasitism have been reported by coprology as yet. This study aims to assess diversity, prevalence, intensity of infection and aggregation of gastrointestinal nematodes in male and female adult chamois. We expect no differences in the parasite infection rates between sexes. Findings: Gastrointestinal tracts of 28 harvested Pyrenean chamois in the Catalan Pyrenees (autumn 2012 and 2013) were necropsied and sexual differences in the diversity and structure of parasite community, prevalence, intensity of infection, and richness were investigated. We found 25 helminth species belonging to 13 different genera. Conclusions: Contrary to our expectations, male chamois showed different parasite communities, higher prevalence, intensity of infection and richness than females. Such sexual differences were clear irrespective of age of individuals. Hence, male chamois must cope with a more diverse and abundant parasite community than females, without apparent biological cost. Further research will be required to confirm this hypothesis

Management of a caseous lymphadenitis outbreak in a new Iberian ibex (Capra pyrenaica) stock reservoir

Background: In 2010, an Iberian ibex (Capra pyrenaica hispanica) stock reservoir was established for conservation purposes in north-eastern Spain. Eighteen ibexes were captured in the wild and housed in a 17 hectare enclosure. Once in captivity, a caseous lymphadenitis (CLA) outbreak occurred and ibex handlings were carried out at six-month intervals between 2010 and 2013 to perform health examinations and sampling. Treatment with a bacterin-based autovaccine and penicillin G benzatine was added during the third and subsequent handlings, when infection by Corynebacterium pseudotuberculosis was confirmed. Changes in lesion score, serum anti-C. pseudotuberculosis antibodies and haematological parameters were analyzed to assess captivity effects, disease emergence and treatment efficacy. Serum acute phase proteins (APP) Haptoglobin (Hp), Amyloid A (SAA) and Acid Soluble Glycoprotein (ASG) concentrations were also determined to evaluate their usefulness as indicators of clinical status.Once in captivity, 12 out of 14 ibexes (85.7%) seroconverted, preceding the emergence of clinical signs; moreover, TP, WBC, eosinophil and platelet cell counts increased while monocyte and basophil cell counts decreased. After treatment, casualties and fistulas disappeared and both packed cell volume (PCV) and haemoglobin concentration significantly increased. Hp, SAA and ASG values were under the limit of detection or showed no significant differences. Conclusions: A role for captivity in contagion rate is suggested by the increase in antibody levels against C. pseudotuberculosis and the emergence of clinical signs. Although boosted by captivity, this is the first report of an outbreak of caseous lymphadenitis displaying high morbidity and mortality in wild ungulates. Treatment consisting of both vaccination and antibiotic therapy seemed to prevent mortality and alleviate disease severity, but was not reflected in the humoural response. Haematology and APP were not useful indicators in our study, perhaps due to the sampling frequency. Presumably endemic and irrelevant in the wild, this common disease of domestic small ruminants is complicating conservation efforts for the Iberian ibex in north-eastern Spain

Oxidative Stress in Wild Boars Naturally and Experimentally Infected with Mycobacterium bovis

Reactive oxygen and nitrogen species (ROS-RNS) are important defence substances involved in the immune response against pathogens. An excessive increase in ROS-RNS, however, can damage the organism causing oxidative stress (OS). The organism is able to neutralise OS by the production of antioxidant enzymes (AE); hence, tissue damage is the result of an imbalance between oxidant and antioxidant status. Though some work has been carried out in humans, there is a lack of information about the oxidant/antioxidant status in the presence of tuberculosis (TB) in wild reservoirs. In the Mediterranean Basin, wild boar (Sus scrofa) is the main reservoir of TB. Wild boar showing severe TB have an increased risk to Mycobacterium spp. shedding, leading to pathogen spreading and persistence. If OS is greater in these individuals, oxidant/antioxidant balance in TB-affected boars could be used as a biomarker of disease severity. The present work had a two-fold objective: i) to study the effects of bovine TB on different OS biomarkers (namely superoxide dismutase (SOD), catalasa (CAT), glutathione peroxidase (GPX), glutathione reductase (GR) and thiobarbituric acid reactive substances (TBARS)) in wild boar experimentally challenged with Mycobacterium bovis, and ii) to explore the role of body weight, sex, population and season in explaining the observed variability of OS indicators in two populations of free-ranging wild boar where TB is common. For the first objective, a partial least squares regression (PLSR) approach was used whereas, recursive partitioning with regression tree models (RTM) were applied for the second. A negative relationship between antioxidant enzymes and bovine TB (the more severe lesions, the lower the concentration of antioxidant biomarkers) was observed in experimentally infected animals. The final PLSR model retained the GPX, SOD and GR biomarkers and showed that 17.6% of the observed variability of antioxidant capacity was significantly correlated with the PLSR X's component represented by both disease status and the age of boars. In the samples from free-ranging wild boar, however, the environmental factors were more relevant to the observed variability of the OS biomarkers than the TB itself. For each OS biomarker, each RTM was defined as a maximum by one node due to the population effect. Along the same lines, the ad hoc tree regression on boars from the population with a higher prevalence of severe TB confirmed that disease status was not the main factor explaining the observed variability in OS biomarkers. It was concluded that oxidative damage caused by TB is significant, but can only be detected in the absence of environmental variation in wild boar

Effects of helminth co-infections on tuberculosis status in a wildlife reservoir

En aquesta tesi s’ha estudiat des d’aspectes bàsics de parasitologia del porc senglar (Sus scrofa), fins l'estudi més ecològic de les seves poblacions de patògens, les co-infeccions que en deriven i els efectes sobre el seu estatus sanitari. En concret s’ha portat a terme sis investigacions, on les tres primeres van ser orientades a revisar aspectes de la parasitofauna del senglar, que inclou una clau per identificar les 5 espècies de nematodes pulmonars més comuns (Capítol 1), les prevalences de l'acantocèfal Macracanthorhynchus hirudinaceus confirmant la seva subestimació (Capítol 2) i els usos i limitacions del recompte d'ous en femtes de senglar per avaluar la càrrega parasitària dels mateixos (Capítol 3). D’altra banda s’ha avaluat els nivells d'estrès oxidatiu en una població de porcs senglars infectats tant de forma natural com de forma experimental per M. bovis (Capítol 4) i s’ha abordat l'efecte de la co-infecció, entre els helmints i la tuberculosi, en termes de salut en una població de senglars naturalment infectada per M. bovis (Capítol 5). Finalment s’ha estudiat l'efecte de la desparasitació sobre l'estructura de la comunitat d’helmints i la tuberculosi (Capítol 6).En esta tesis se ha estudiado desde aspectos básicos de parasitología del jabalí (Sus scrofa), hasta aspectos más ecológicos de sus poblaciones de patógenos, las co-infeciones que padecen y los efectos sobre su estatus sanitario. En concreto, se han llevado a cabo seis investigaciones, siendo las tres primeras orientadas a revisar aspectos de la parasitofauna del jabalí. En concreto, se ha desarrollado una clave para identificar las 5 especies de nematodos pulmonares más comunes (Capítulo 1), se han estudiado las prevalencias del acantocéfalo Macracanthorhynchus hirudinaceus confirmando su subestimación (Capítulo 2) y se ha revisado los usos y limitaciones del conteo de huevos en heces de jabalí para evaluar la carga parasitaria de los mismos (Capítulo 3). Por otro lado, se han evaluado los niveles de estrés oxidativo tanto en una población de jabalíes infectados de forma natural como experimentalmente por M. bovis (Capítulo 4) y se ha abordado el efecto sobre la salud de la co-infección, entre los helmintos y la tuberculosis, en una población de jabalíes naturalmente infectada por MTC (Capítulo 5). Finalmente, se ha estudiado el efecto de la desparasitación sobre la estructura de la comunidad de helmintos y la tuberculosis (Capítulo 6).This thesis uses basic parasitology studies of wild boar (Sus scrofa) to focus on a more ecological view of their pathogen populations, co-infections that arise and the effects on their health status. Specifically, six investigations were conducted being the first three aimed at reviewing aspects of the wild boar parasitofauna In particular, keys to identify the five most common species of lung nematodes (Chapter 1), the real prevalence of the Acanthocephala Macracanthorhynchus hirudinaceus denouncing the underestimation of this species (Chapter 2) and a review of the uses and limitations of faecal egg count for assessing worm burden in wild boars (Chapter 3) are presented. Furthermore, the levels of oxidative stress in a population of feral pigs infected naturally and experimentally by M. bovis (Chapter 4) and the effect of co-infection between helminths and tuberculosis in terms of physiological cost of health in a wild boar population naturally infected by MTC (Chapter 5) were addressed. Finally, the effect of an antiparasite drug treatment on the helminth community structure and on the outcome of disease infection in a wild boar population naturally infected by M. bovis (Chapter 6) was studied

Leishmania Infection in Wild Lagomorphs and Domestic Dogs in North-East Spain

Leishmania infantum is a zoonotic protozoan parasite distributed worldwide that is transmitted by phlebotomine sandflies. Dogs are the main reservoir for human infections. However, in recent years, the capacity of lagomorphs to contribute to Leishmania transmission has been confirmed. The present study aimed to assess Leishmania spp. exposure and infection in lagomorphs and sympatric domestic dogs in NE Spain. Sera from European hares, European rabbits, and rural dogs were tested for antibodies against L. infantum using an in-house indirect ELISA. PCR analysis targeting Leishmania spp. was performed in spleens from L. europaeus. Antibodies against Leishmania spp. were detected in all the species analyzed. Total sample prevalence was significantly higher in O. cuniculus (27.9%) than in L. europaeus (2.0%). Results of the PCR were all negative. The present study expands knowledge about Leishmania infections in free-ranging lagomorphs in the Iberian Peninsula, suggesting a more important role of O. cuniculus in the study area. Given the strong correlation between lagomorph densities and human leishmaniasis outbreaks in Spain, the high rabbit and human densities in NE Spain, and the high Leishmania spp. seroprevalence in rabbits, it becomes imperative to establish surveillance programs for lagomorphs in this region.X. F. Aguilar was funded through the Maria Zambrano post-doctoral program (ID-709248). M. P. Ribas was funded through the 2021 FI Scholarship, Departament de Recerca i Universitats, Generalitat de Catalunya, Spain (FI_B 00171).info:eu-repo/semantics/publishedVersio