Transcriptome analysis of Echinococcus granulosus sensu stricto protoscoleces reveals differences in immune modulation gene expression between cysts found in cattle and sheep.

Cystic Echinococcosis (CE), a zoonotic parasitic disease, is caused by the cestode Echinococcus granulosus sensu lato. CE inflicts severe damage in cattle, sheep, and human hosts worldwide. Fertile CE cysts are characterized by the presence of viable protoscoleces. These parasite forms are studied with minimal contamination with host molecules. Hosts, cattle and sheep, show differences in their CE cyst fertility. The effect of the host in protoscolex transcriptome is not known. We genotyped and performed transcriptomic analysis on sheep protoscoleces obtained from liver and lung CE cysts. The transcriptomic data of Echinococcus granulosus sensu stricto protoscoleces from 6 lung CE cysts and 6 liver CE cysts were Collected. For host comparison analysis, 4 raw data files belonging to Echinococcus granulosus sensu stricto protoscoleces from cattle liver CE cysts were obtained from the NCBI SRA database. Principal component and differential expression analysis did not reveal any statistical differences between protoscoleces obtained from liver or lung cysts, either within the same sheep or different sheep hosts. Conversely, there are significant differences between cattle and sheep protoscolex samples. We found differential expression of immune-related genes. In cattle, 7 genes were upregulated in protoscoleces from liver cysts. In sheep, 3 genes were upregulated in protoscoleces from liver and lung CE cysts. Noteworthy, are the differential expression of antigen B, tegument antigen, and arginase-2 in samples obtained from sheep CE cysts, and basigin in samples from cattle CE cysts. These findings suggest that the host species is an important factor involved in the differential expression of immune related genes, which in turn is possibly related to the fertility of Echinococcus granulosus sensu stricto cysts.

Response patterns in adventitial layer of Echinococcus granulosus sensu stricto cysts from naturally infected cattle and sheep.

Cystic echinococcosis is a zoonotic disease caused by the metacestode of Echinococcus granulosus sensu lato. The disease is characterized by the development of cystic structures inside viscera of the intermediate host, mainly liver and lungs. These cysts are formed by three layers: germinal, laminated, and adventitial layer, the latter being the local host immune response. Metacestodes that develop protoscoleces, the infective stage to the definitive host, are termed fertile, whereas cysts that do not produce protoscoleces are termed non-fertile. Sheep usually harbor fertile cysts while cattle usually harbor non-fertile cysts. Adventitial layers with fibrotic resolution are associated to fertile cysts, whereas a granulomatous reaction is associated with non-fertile cysts. The aim of this study was to analyze cellular distribution in the adventitial layer of fertile and non-fertile E. granulosus sensu stricto cysts found in liver and lungs of cattle and sheep. A total of 418 cysts were analyzed, 203 from cattle (8 fertile and 195 non-fertile) and 215 from sheep (64 fertile and 151 non-fertile). Fertile cysts from cattle showed mixed patterns of response, with fibrotic resolution and presence of granulomatous response in direct contact with the laminated layer, while sheep fertile cysts always displayed fibrotic resolution next to the laminated layer. Cattle non-fertile cysts display a granulomatous reaction in direct contact with the laminated layer, whereas sheep non-fertile cysts display a granulomatous reaction, but in direct contact with the fibrotic resolution. This shows that cattle and sheep cystic echinococcosis cysts have distinct local immune response patterns, which are associated to metacestode fertility.

Fasciola hepatica coinfection modifies the morphological and immunological features of Echinococcus granulosus cysts in cattle.

Polyparasitism occurs when animals harbour multiple parasites concomitantly. It is a common occurrence but is generally understudied in wild and domestic animals. Fasciola hepatica and Echinococcus granulosus, which are helminths of ungulates, frequently coinfect cattle. The effects of this particular type of polyparasitism are not well documented. The metacestode of Echinococcus granulosus is surrounded by the adventitial layer, which constitutes the host immune response to the parasite. This layer in cattle is produced by a granulomatous reaction and is involved in echinococcal cyst (EC) fertility. Due to the systemic immune-modulating abilities of Fasciola hepatica, coinfection possibly generates a favourable environment for EC growth. A total of 203 Echinococcus granulosus sensu stricto cysts were found in 82 cattle, of which 42 ECs were found in 31 animals coinfected with Fasciola hepatica. The overall infection intensity was 3 cysts per animal. Coinfection with Fasciola hepatica decreased the mean infection intensity to 1.4 cysts per animal. Regarding EC size, coinfection resulted in smaller ECs (15.91 vs 22.09 mm), especially for infertile lung cysts. The adventitial layer of ECs in coinfected animals lacked lymphoid follicles and palisading macrophages, which are generally hallmarks of the granulomatous immune response. The ECs in coinfected animals had organized laminated layers, whereas those in animals without coinfection did not. Although coinfection was not statistically associated with EC fertility, we did not find fertile cysts in the livers of coinfected animals. We concluded that coinfection with Fasciola hepatica and Echinococcus granulosus has a detrimental effect on ECs, particularly infertile cysts.

Lymphocyte Populations in the Adventitial Layer of Hydatid Cysts in Cattle: Relationship With Cyst Fertility Status and Fasciola Hepatica Co-Infection.

Cystic echinococcosis is a worldwide zoonosis caused by the cestode Echinococcus granulosus. Two types of hydatid cysts occur in intermediate hosts: fertile cysts that generate protoscoleces from the germinal layer of the cyst, and infertile cysts that do not produce protoscoleces and are unable to continue the life cycle of the parasite. The adventitial layer, a host-derived fibrous capsule surrounding the hydatid cyst, is suggested to play an important role in local immune regulation during infection and in fertility of the cysts. Fasciola hepatica, another important parasite of cattle, induces a characteristic Th2-like immune response that could modulate the immune response against E. granulosus. Natural co-infection of both parasites is common in cattle, but no reports describe the local immune response against E. granulosus with F. hepatica infection in the same host. This study analyzed the number and distribution of T and B cells in the adventitial layer of liver and lung cysts and the relationship with cyst fertility and F. hepatica co-infection. T lymphocytes were the predominant cell type in the adventitial layer of infertile hydatid cysts and were more numerous in infertile hydatid cysts. B lymphocyte numbers were not associated with hydatid cyst fertility. Mast cells were infrequent in the adventitial layer. The number of T and B cells was not associated with F. hepatica co-infection. The present study contributes to the understanding of local immune responses in bovine cystic echinococcosis.

Multiple haplotypes of Echinococcus granulosus sensu stricto in single naturally infected intermediate hosts.

Cystic echinococcosis is a disease that affects both humans and animals, caused by cryptic species complex belonging to the platyhelminth Echinococcus granulosus sensu lato (s.l.). This disease is distributed worldwide, with E. granulosus sensu stricto (s.s.) being the most widespread of the species. High genetic variability has been demonstrated within E. granulosus s.s. studying single cyst per infected animal identifying a number of different haplotypes. However, few studies have addressed the genetic diversity of this parasite within a single intermediate host with multiple Echinococcus cysts. To date, it remains unknown if specific haplotypes of E. granulosus s.s. produce differences in biological features of the cyst. Here, we use the full length of the mitochondrial gene cox1 to determine E. granulosus s.s. haplotypes in samples from both cattle and sheep which harboured more than one cyst in different areas in Chile, where this parasite is endemic. We found 16 different haplotypes in 66 echinococcal cysts from 10 animals, and both cattle and sheep can harbour up to five different haplotypes of E. granulosus s.s. in the same animal. Regarding cyst fertility, five animals had both fertile and infertile Echinococcus cysts in both single and multiple haplotype infections. There was no association between haplotype and cyst fertility, size, or adventitial layer characteristics. Sampling and sequencing every Echinococcus cyst found in the intermediate host reveals a high molecular variability. We speculate that multiple haplotype infections could also suggest that intermediate hosts come from hyperendemic areas.

Exploring Veterinary Medicine Students' Experiences with Team-Based Learning at the Universidad Andrés Bello.

This study explored the use of team-based learning (TBL) in a Clinical Anatomy course taken by third-year veterinary medicine undergraduate students at the Universidad Andrés Bello in Chile. While research has shown that active learning methodologies yield improved student learning outcomes compared to lecture-based teaching, the incorporation of new pedagogical strategies is complex and its success depends on a range of contextual factors. This study sought to understand the strengths and weaknesses of using TBL in a specific subject (anatomy), discipline (veterinary medicine), and country (Chile). Students in the course had not been previously exposed to TBL. At the end of the semester during which TBL was used, the research team collected student satisfaction survey data and conducted a focus group in order to understand students' experiences with TBL in the course. We found that overall, students were satisfied with TBL and appreciated that it increased the amount of feedback they received, reinforced key concepts, and helped them to build skills they would need in their future professions. There was also a certain level of dissatisfaction, which may have been caused by negative experiences with team members and difficulties reading the preparatory material, which was in English. Given our findings, we discuss modifications that could be made in order to improve veterinary medicine students' experiences with TBL.

Intra-articular treatment with corticosteroids increases apoptosis in human rotator cuff tears.

PURPOSE: The aim of this study is to evaluate in vivo the level of apoptosis in human rotator cuff tears and the relationship it might have with tendon degeneration. METHODS: Rotator cuff biopsies from 19 male and female patients, ages between 38 and 68 years, with and without previous corticosteroid infiltrations were collected via arthroscopy. Biopsies from seven patients with healthy rotator cuffs were used as a control group. An in situ terminal deoxynucleotidyl transferase dUTP nick end labeling assay was performed to detect the level of apoptosis, which was expressed as a percentage of apoptotic cells (PAC). RESULTS: PAC in patients with corticosteroid infiltrations was 76.97 ± 16.99 in all tendon rupture zones, in non-infiltrated patients was 35.89 ± 22.96, whereas in control patients was 14.48 ± 8.15. Likewise, the tendency of PAC reveals that apoptosis in control and non-infiltrated groups was different and dispersed in all tear zones; while in corticosteroid treated patients, the tendency was similar in all rupture sites. CONCLUSIONS: This investigation leads us to conclude that the administration of corticosteroid is associated with a higher amount of apoptosis at the insertion site of the rotator cuff (rupture edge).

Characterization of 14-3-3 isoforms expressed in the Echinococcus granulosus pathogenic larval stage.

The 14-3-3 protein family of eukaryotic regulators was studied in Echinococcus granulosus, the causative agent of cystic hydatid disease. These proteins mediate important cellular processes in eukaryotes and are expected to play important roles in parasite biology. Six isoforms of E. granulosus 14-3-3 genes and proteins (Eg14-3-3.1-6) were analyzed, and their phylogenetic relationships were established with bona fide 14-3-3 orthologous proteins from eukaryotic species. Eg14-3-3 isoforms with previous evidence of expression (Eg14-3-3.1-4) in E. granulosus pathogenic larval stage (metacestode) were cloned, and recombinant proteins were used for functional studies. These protein isoforms were detected in different components of E. granulosus metacestode, including interface components with the host. The roles that are played by Eg14-3-3 proteins in parasite biology were inferred from the repertoires of interacting proteins with each isoform, as assessed by gel overlay, cross-linking, and affinity chromatography assays. A total of 95 Eg14-3-3 protein ligands were identified by mass spectrometry. Eg14-3-3 isoforms have shared partners (44 proteins), indicating some overlapping functions; however, they also bind exclusive partners (51 proteins), suggesting Eg14-3-3 functional specialization. These ligand repertoires indicate the involvement of Eg14-3-3 proteins in multiple biochemical pathways in the E. granulosus metacestode and note some degree of isoform specialization.

Matrix metalloproteinases regulate extracellular levels of SDF-1/CXCL12, IL-6 and VEGF in hydrogen peroxide-stimulated human periodontal ligament fibroblasts.

Periodontitis is a highly prevalent infectious disease characterized by the progressive inflammatory destruction of tooth-supporting structures, leading to tooth loss. The underling molecular mechanisms of the disease are incompletely understood, precluding the development of more efficient screening, diagnostic and therapeutic approaches. We investigated the interrelation of three known effector mechanisms of the cellular response to periodontal infection, namely reactive oxygen species (ROS), matrix metalloproteinases (MMPs) and cytokines in primary cell cultures of human periodontal ligament fibroblast (hPDLF). We demonstrated that ROS increase the activity/levels of gelatinolytic MMPs, and stimulate cytokine secretion in hPDLF. Additionally, we proved that MMPs possesses immune modulatory capacity, regulating the secreted levels of cytokines in ROS-stimulated hPDLF cultures. This evidence provides further insight in the molecular pathogenesis of periodontitis, contributing to the future development of more effective therapies.

The TORC1/P70S6K and TORC1/4EBP1 signaling pathways have a stronger contribution on skeletal muscle growth than MAPK/ERK in an early vertebrate: Differential involvement of the IGF system and atrogenes.

Knowledge about the underlying mechanisms, particularly the signaling pathways that account for muscle growth in vivo in early vertebrates is still scarce. Fish (Paralichthys adspersus) were fasted for 3weeks to induce a catabolic period of strong muscle atrophy. Subsequently, fish were refed for 2weeks to induce compensatory muscle hypertrophy. During refeeding, the fish were treated daily with either rapamycin (TORC blocker), PD98059 (MEK blocker), or PBS (V; vehicle), or were untreated (C; control). Rapamycin and PD98059 differentially impaired muscle cellularity in vivo, growth performance, and the expression of growth-related genes, and the inhibition of TORC1 had a greater impact on fish muscle growth than the inhibition of MAPK. Blocking TORC1 inhibited the phosphorylation of P70S6K and 4EBP1, two downstream components activated by TORC1, thus affecting protein contents in muscle. Concomitantly, the gene expression in muscle of igf-1, 2 and igfbp-4, 5 was down-regulated while the expression of atrogin-1, murf-1, and igfbp-2, 3 was up-regulated. Muscle hypertrophy was abolished and muscle atrophy was promoted, which finally affected body weight. TORC2 complex was not affected by rapamycin. On the other hand, the PD98059 treatment triggered ERK inactivation, a downstream component activated by MEK. mRNA contents of igf-1 in muscle were down-regulated, and muscle hypertrophy was partially impaired. The present study provides the first direct data on the in vivo contribution of TORC1/P70S6K, TORC1/4EBP1, and MAPK/ERK signaling pathways in the skeletal muscle of an earlier vertebrate, and highlights the transcendental role of TORC1 in growth from the cellular to organism level.

Adaptive responses of mitochondria to mild copper deprivation involve changes in morphology, OXPHOS remodeling and bioenergetics.

Copper is an essential cofactor of complex IV of the electron transfer chain, and it is directly involved in the generation of mitochondrial membrane potential. Its deficiency induces the formation of ROS, large mitochondria and anemia. Thus, there is a connection between copper metabolism and bioenergetics, mitochondrial dynamics and erythropoiesis. Copper depletion might end in cellular apoptosis or necrosis. However, before entering into those irreversible processes, mitochondria may execute a series of adaptive responses. Mitochondrial adaptive responses (MAR) may involve multiple and diverse mechanisms for preserving cell life, such as mitochondrial dynamics, OXPHOS remodeling and bioenergetics output. In this study, a mild copper deficiency was produced in an animal model through intraperitoneal injections of bathocuproine disulfonate in order to study the MAR. Under these conditions, a new type of mitochondrial morphology was discovered in the liver. Termed the "butternut squash" mitochondria, it coexisted with normal and swollen mitochondria. Western blot analyses of mitochondrial dynamics proteins showed an up-regulation of MFN-2 and OPA1 fusion proteins. Furthermore, isolated liver mitochondria displayed OXPHOS remodeling through a decrease in supercomplex activity with a concomitant increase at an individual level of complexes I and IV, higher respiratory rates at complex I and II levels, higher oligomycin-insensitive respiration, and lower respiratory control ratio values when compared to the control group. As expected, total ATP and ATP/ADP values were not significantly different, since animal's health was not compromised. As a whole, these results describe a compensatory and adaptive response of metabolism and bioenergetics under copper deprivation.

Weighted gene co-expression network analysis reveals immune evasion related genes in Echinococcus granulosus sensu stricto.

Cystic echinococcosis (CE) is a zoonotic disease caused by the tapeworm Echinococcus granulosus sensu lato (s.l). In the intermediate host, this disease is characterized by the growth of cysts in viscera such as liver and lungs, inside of which the parasite develops to the next infective stage known as protoscoleces. There are records that the infected viscera affect the development and morphology of E. granulosus s.l. protoscolex in hosts such as buffalo or humans. However, the molecular mechanisms that drive these differences remains unknown. Weighted gene co-expression network analysis (WGCNA) using a set of RNAseq data obtained from E. granulosus sensu stricto (s.s.) protoscoleces found in liver and lung cysts reveals 34 modules in protoscoleces of liver origin, of which 12 have differential co-expression from protoscoleces of lung origin. Three of these twelve modules contain hub genes related to immune evasion: tegument antigen, tegumental protein, ubiquitin hydrolase isozyme L3, COP9 signalosome complex subunit 3, tetraspanin CD9 antigen, and the methyl-CpG-binding protein Mbd2. Also, two of the twelve modules contain only hypothetical proteins with unknown orthology, which means that there are a group of unknown function proteins co-expressed inside the protoscolex of liver CE cyst origin. This is the first evidence of gene expression differences in protoscoleces from CE cysts found in different viscera, with co-expression networks that are exclusive to protoscoleces from liver CE cyst samples. This should be considered in the control strategies of CE, as intermediate hosts can harbor CE cysts in liver, lungs, or both organs simultaneously.

Efficacy of Multivalent, Cochleate-Based Vaccine against Salmonella Infantis, S. Enteritidis and S. Typhimurium in Laying Hens.

Salmonella enterica is an important foodborne pathogen. Commercial poultry are the main reservoirs of Salmonella enterica, leading to the contamination of food and outbreaks in humans. The vaccination of chickens is one of the most important strategies to reduce the number of Salmonella in poultry farms. Unfortunately, commercial vaccines have not been fully effective in controlling the spread and do not contain all the Salmonella serovars that circulate on farms. In this study, we evaluate a new, cochleate-based, trivalent injectable vaccine against S. Enteritidis, S. Typhimurium and S. Infantis, describing the vaccine security, capacity to induce specific anti-Salmonella serovar IgY and the gene expression of immune markers related to CD4 and CD8 T-cell-mediated immunity. Efficacy was evaluated through oral challenges performed separately for each Salmonella serotype. The efficacy and safety of the trivalent vaccine was proven under controlled conditions. The vaccine has no local or systemic reactions or adverse effects on poultry performance related to the vaccine. The vaccine provided significantly increased serum IgY titer levels, significantly reduced Salmonella CFU/g present in the cecum and an increased CD4+/CD8+ ratio in vaccinated animals when challenged with S. Infantis, S. Enteritidis and S. Typhimurium. These results indicate that this new trivalent vaccine does not generate adverse effects in poultry and produces an increase in neutralizing antibodies against the three Salmonella serovars.

Bovine (Bos taurus) humoral immune response against Echinococcus granulosus and hydatid cyst infertility.

Echinococcus granulosus, the agent of hydatid disease, presents an indirect life cycle, with canines (mainly dogs) as definitive hosts, and herbivores and human as intermediary ones. In intermediary hosts fertile and infertile cysts develop, but only the first ones develop protoscoleces, the parasite form infective to definitive hosts. We report the presence of bovine IgGs in the germinal layer from infertile cysts (GLIC), in an order of magnitude greater than in the germinal layer from fertile cysts (GLFC). When extracted with salt solutions, bovine IgGs from GLIC are associated with low or with high affinity (most likely corresponding to non specific and antigen specific antibodies, respectively). Specific IgGs penetrate both the cells of the germinal layer and HeLa cultured cells and recognize parasitic proteins. These results, taken together with previous ones from our laboratory, showing induction of apoptosis in the germinal layer of infertile hydatid cysts, provide the first coherent explanation of the infertility process. They also offer the possibility of identifying the parasite antigens recognized, as possible targets for immune modulation.

A Unique Approach to Project-Based Learning (PjBL) in a Veterinary Anatomy Course.

INTRODUCTION: Project-based learning (PjBL) is a teaching methodology designed to engage students in solving real-world problems, acknowledging that students are active agents of their learning process. This methodology has historically been popular in architecture and industrial sciences; however, its use in teaching veterinary anatomy is scarcely published. METHODS: Using information and communication technologies, the PjBL methodology was implemented in a first-year veterinary anatomy course. The methodology included teamwork and the selection of a routine object in the veterinary clinic at the beginning of the academic semester. The project's goal was to analyze the object and associate it with both a domestic animal species and an anatomical region, along with making and presenting a video or a simulation model about the object. RESULTS: More than 80% of students prefer active learning classes compared to traditional classes. In addition, 66% and 86% of students indicate that PjBL allowed them to improve their understanding of theoretical content for the first and second years of post-implementation, respectively. Students' self-assessment indicates that more than 80% of the students (first and second year post-implementation) felt they were responsible for the execution of the project, able to conduct research, and able to develop autonomous learning skills. After 2 years of PjBL implementation, failure rates in the course decreased by 21%. DISCUSSION: In general, PjBL results show that veterinary students prefer active learning activities that allow them to learn in a team-based learning process as well as to develop soft skills such as self-learning, responsibility, and teamwork. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s40670-021-01205-1.

Fasciola hepatica co-infection enhances Th1 immune response in the adventitial layer of non-fertile Echinococcus granulosus cysts.

Cystic echinococcosis is a widespread zoonosis caused by the larval stage of the tapeworm Echinococcus granulosus. In intermediary hosts, two types of echinococcal cysts can be found: fertile, which produce protoscoleces, the infective form of the parasite to dogs; and infertile, that do not present protoscoleces and therefore are not able to continue with the parasite life cycle. The adventitial layer, the local immune response against the cyst, plays an important role in cyst fertility. Grazing cattle can often feature Fasciola hepatica co-infection, a parasite known to modulate the host systemic immune response. In this work the cellular Th1/Th2 immune profiles were evaluated in the adventitial layer of fertile and non-fertile cysts with and without co-infection with Fasciola hepatica. Measuring with immunohistochemistry and qPCR in adventitial layer, we report that non-fertile cysts present higher levels of Th1 cytokines (IFN-γ (P < 0.0001) and TNF-α (P < 0.05)), and fertile cysts have higher levels of Th2 cytokines (IL-4 (P < 0.001)). Co-infection with Fasciola hepatica is associated with a decrease in the expression of IL-4 (P < 0.05) and an increase in the expression of IFN-γ (P < 0.0001) in the adventitial layer of fertile cysts. Non-fertile cysts were associated with higher levels of Th1 cytokines in the adventitial layer, with IFN-γ expression enhanced by F. hepatica co-infection (P < 0.0001), confirming that polyparasitism should be considered in the treatment and control of naturally infected cattle.

Piscirickettsia salmonis induces apoptosis in macrophages and monocyte-like cells from rainbow trout.

Piscirickettsia salmonis is the etiologic agent of the salmonid rickettsial septicemia (SRS) which causes significant losses in salmon production in Chile and other and in other regions in the southern hemisphere. As the killing of phagocytes is an important pathogenic mechanism for other bacteria to establish infections in vertebrates, we investigated whether P. salmonis kills trout macrophages by apoptosis. Apoptosis in infected macrophages was demonstrated by techniques based on morphological changes and host cell DNA fragmentation. Transmission electron microcopy showed classic apoptotic characteristics and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling showed fragmented DNA. Programmed cell death type I was further confirmed by increased binding of annexin V to externalized phosphatidylserine in infected macrophages. Moreover, significant increases of caspase 3 activation were detected in infected cells and treatment with caspase inhibitor caused a decrease in levels of apoptosis. This is the first evidence that P. salmonis induces cell death in trout macrophages. This could lead to bacterial survival and evasion of the host immune response and play an important role in the establishment of infection in the host.

Cattle co-infection of Echinococcus granulosus and Fasciola hepatica results in a different systemic cytokine profile than single parasite infection.

E. granulosus is a cestode that causes Cystic Echinococcosis (CE), a zoonotic disease with worldwide presence. The immune response generated by the host against the metacestode induces a permissive Th2 response, as opposed to pro-inflammatory Th1 response. In this view, mixed Th2 and regulatory responses allow parasite survival. Overall, larval Echinococcus infections induce strong regulatory responses. Fasciola hepatica, another common helminth parasite, represents a major infection in cattle. Co-infection with different parasite species in the same host, polyparasitism, is a common occurrence involving E. granulosus and F. hepatica in cattle. 'While it is known that infection with F. hepatica also triggers a polarized Th2/Treg immune response, little is reported regarding effects on the systemic immune response of this example of polyparasitism. F. hepatica also triggers immune responses polarized to the Th2/ Treg spectrum. Serum samples from 107 animals were analyzed, and were divided according to their infection status and Echinococcal cysts fertility. Cytokines were measured utilizing a Milliplex Magnetic Bead Panel to detect IFN-γ, IL-1, IL-2, IL-4, IL-6, IL-10, IL-12 and IL-18. Cattle infected only with F. hepatica had the highest concentration of every cytokine analyzed, with both 4.24 and 3.34-fold increases in IL-10 and IL-4, respectively, compared to control animals, followed by E. granulosus and F. hepatica co-infected animals with two-fold increase in IL-10 and IL-4, compared to control animals, suggesting that E. granulosus co-infection dampens the cattle Th2/Treg immune response against F. hepatica. When considering Echinococcal cyst fertility and systemic cytokine concentrations, fertile cysts had higher IFN-γ, IL-6 and IL-18 concentrations, while infertile cysts had higher IL-10 concentrations. These results show that E. granulosus co-infection lowers Th1 and Th2 cytokine serological concentration when compared to F. hepatica infection alone. E. granulosus infections show no difference in IFN-γ, IL-1, IL-2, IL-6 and IL-18 levels compared with control animals, highlighting the immune evasion mechanisms of this cestode.

Reproductive and Behavioral Evaluation of a New Immunocastration Dog Vaccine.

Canine immunocastration development has been of interest for many years as a complementary strategy to surgical castration. The purpose of this paper was to verify the effect of a recombinant vaccine for dog immunocastration. Two tests were done, one under controlled conditions and a second under field conditions. Animals were injected with 1 mL of 500 µg GnRXG/Q recombinant protein; 500 µg of low molecular weight chitosan as adjuvant; 1 mL NaCl 0.9% q.s. In the first trial, eight Beagle male dogs between the ages of 1 and 3 comprised the sample, randomly divided into two groups: vaccinated group (n = 7) and control group (n = 2). The second trial had 32 dogs with owners. In the first controlled conditions trial, the vaccine produced specific antibodies that remained until the end of the trial (day 270), inducing reduced testosterone and spermiogram changes in the immunized animals. In a second trial, on the field, specific immunity was induced, which remained high up to day 150. The vaccine also reduced sexual agonistic and marking behaviors. This new vaccine proved to be safe, immunogenic, capable of reducing gonadal functionality, and had a positive effect on inducing reduced sexual, agonistic, and marking behavior of the animals.

Echinococcus granulosus calreticulin: molecular characterization and hydatid cyst localization.

Hydatidosis, whose etiological agent is the larval stage of the platyhelminth parasite Echinococcus granulosus, is a major zoonotic public health problem and causes great economic losses in many countries, affecting humans and livestock species. Calreticulin (CRT) is a multifunctional and conserved chaperone calcium-binding protein, present in every cell of higher organisms, except erythrocytes. In other parasites species, CRT emerges as a key modulator of several immunological aspects of their relationships with their vertebrate hosts. We report herein the cloning of two DNA segments coding for sequences of E. granulosus CRT (EgCRT). Together with another partial sequence available at the NCBI database, a complete EgCRT cDNA sequence is now proposed. EgCRT mRNA is equally expressed in fertile and infertile hydatid cysts germinal layers, as well as in protoscoleces. In hydatid cysts, EgCRT is a 50 kDa protein, expressed in germinal layer and protoscoleces, mainly in perinuclear and cytoplasmic zones.