Vaccination with cathepsin L mimotopes of Fasciola hepatica in goats reduces worm burden, morphometric measurements, and reproductive structures.

Fasciolosis is a worldwide emergent zoonotic disease that significantly constrains the productivity of livestock. We conducted an experimental trial with four groups of five goats each, vaccinated with 1 × 1013 phage particles of cathepsin L1 (CLI: DPWWLKQ), CL1 (SGTFLFS), and CL2 (PPIRNGK) mimotopes combined with Quil A adjuvant. Animals received a booster four weeks later. The control group received phosphate-buffered saline. All animals were infected with 200 Fasciola hepatica metacercariae at week six and euthanised 16 weeks later. The percentage of significant worm reduction in CL1 (DPWWLKQ), CL1 (SGTFLFS), and CL2 (PPIRNGK) compared to the control group were 55.40%, 70.42% (P < 0.05), and 32.39%, respectively. Vaccinated animals showed a significant reduction in faecal egg production and egg viability. A significant reduction in the total biomass of parasites recovered was observed in the CL1 (DPWWLKQ) and CL1 (SGTFLFS) groups. In goats vaccinated with CL2 (PPIRNGK), fluke length and width were smaller than those in the control group. Furthermore, animals receiving CL mimotopes showed a significant reduction in the total area of reproductive structures. Goats immunised with phage-displayed mimotopes produced significantly high titres of specific IgG1 and IgG2 isotypes, indicating a mixed Th1/Th2 response. The liver fluke burdens in goats vaccinated with CL1 (DPWWLKQ) and CL1 (SGTFLFS) were significantly correlated with IgG and IgG1 levels.

Vaccination with cathepsin L phage-exposed mimotopes, single or in combination, reduce size, fluke burden, egg production and viability in sheep experimentally infected with Fasciola hepatica.

Fascioliasis is a worldwide emergent zoonotic disease that significantly constrains the productivity of livestock. In this study, fluke burdens, liver fluke size and biomass, faecal eggs counts, serum levels of hepatic enzymes and immune response were assessed in sheep vaccinated with peptide mimotopes of cathepsin L and infected with metacercariae. A total of 25 sheep were allocated randomly into five groups of five animals each, and experimental groups were immunised with 1 × 1013 filamentous phage particles of cathepsin L1 (CL1) (TPWKDKQ), CL2 (YGSCFLR) and mixtures of CL1 + CL2 mimotopes, in combination with Quil A adjuvant, and wild-type M13KE phage in a two-vaccination scheme on weeks 0 and 4. The control group received phosphate-buffered saline. All groups were challenged with 300 metacercariae two weeks after the last immunisation and euthanised 16 weeks later. The CL1 vaccine was estimated to provide 57.58% protection compared with the control group; no effect was observed in animals immunised with CL2 and CL1 + CL2 (33.14% and 11.63%, respectively). However, animals receiving CL2 had a significant reduction in parasite egg output. Vaccinated animals showed a significant reduction in fluke length and width and wet weights. In the CL1 group, there was a significant reduction in the total biomass of parasites recovered. Egg development was divided into seven stages: dead, empty, unembryonated, cell division, eyespot, hatched and hatching. The highest percentage of developmental stages was detected for vaccinated sheep administered CL1 + CL2 with cell division, and the lowest percentage was observed in the hatching stage. Furthermore, a significant difference in all developmental stages was observed between vaccinated animals and the control group (P < 0.01). The levels of anti-phage total IgG in immune sera increased significantly at four weeks after immunisation and were always significantly higher for cathepsin L vaccine group than in the challenged control group. Total IgG was inversely and significantly correlated with worm burden in the CL1 group.

Phage display-based vaccine with cathepsin L and excretory-secretory products mimotopes of Fasciola hepatica induces protective cellular and humoral immune responses in sheep.

Fasciolosis is a foodborne zoonotic disease that affects grazing animals and causes substantial economic losses worldwide. Excretory/secretory (E/S) products and cathepsin L mimotopes from Fasciola hepatica were used to immunise experimentally infected sheep against liver flukes. The level of protection was measured in terms of fluke burden, morphometric measurements and faecal egg counts, as well as the humoral and cellular immune responses elicited. Five groups of 5 sheep each were immunised with 1 × 1013 phage particles of cathepsin L1 (group 1: SGTFLFS), cathepsin L1 (group 2: WHVPRTWWVLPP) and immunodominant E/S product (group 3) mimotopes with Quil A adjuvant, and wild-type M13KE phage (group 4) at the beginning and as a booster two weeks later. The control group received phosphate-buff ;ered saline. All groups were challenged with 300 metacercariae at week four and slaughtered 18 weeks later. The mean fluke burdens after challenge were reduced by 52.39 % and 67.17 % in sheep vaccinated with E/S products (group 3) and cathepsin L1 (group 1: SGTFLFS), respectively; no eff ;ect was observed in animals inoculated with cathepsin L1 (group 2: WHVPRTWWVLPP). Animals vaccinated showed a significant reduction in fluke length and width, wet weights and egg output Sheep immunised with phage-displayed mimotopes induced the development of specific IgG1 and IgG2, indicating a mixed Th1/Th2 immune response. Measurement of cytokine levels revealed higher levels of IFN-γ as well as lower production of IL-4 in sheep vaccinated with the mimotope peptide of F. hepatica. Fluke-specific production of IFN-γ in immunised animals was significantly correlated with fluke burden (P < 0.01). As helminth infection progressed, increased levels of IL-4 were evident in the wild-type M13KE phage (group 4) and the control groups (group 5), accompanied by a downregulation of IFN-γ production. Vaccinated animals with cathepsin L1 (group 1: SGTFLFS) showed that amino acids located in the middle (64SG65) of the linear sequence and C-terminal end (314TFLFS318) were associated with significant protection.

Echocardiographic follow-up to right ventricular modifications in secondary pulmonary hypertension to diabetes in rats.

Clinical studies suggest that diabetes is a risk factor in the development of pulmonary arterial hypertension. The increase in blood pressure in the pulmonary area is characterized by the increase in the afterload and hypertrophy of the right ventricle. The objective of this study was to conduct a longitudinal follow-up of the morphological and functional changes in the right ventricle in a rat model with pulmonary arterial hypertension secondary to diabetes. Male Sprague Dawley rats were randomly divided into a control group (saline solution) and a diabetic group (60 mg/kg with streptozotocin). For 12 weeks, an echocardiography for longitudinal (in vivo) image analysis of the pulmonary pressure was performed at the same time as the evaluation of myocardial remodeling and right ventricular. After this period, the pulmonary pressure was measured by means of a pulmonary artery catheterization, and the presence of hypertrophy was determined by means of the Fulton index. The plasma concentration of brain natriuretic peptide was measured by means of the ELISA technique. It was found that the diabetic rats showed an increase in pressure in the pulmonary arteries, an increase in the Fulton index, and an increase in brain natriuretic peptide. The echocardiographic follow-up showed that the diabetic rats presented an increase in the pulmonary artery from the fourth week, while hypertrophy and right ventricular systolic dysfunction occurred until the twelfth week. In conclusion, pulmonary arterial hypertension induced by experimental diabetes generated hypertrophy and systolic dysfunction of the right ventricle.

Prevalence and risk factors of gastrointestinal nematode infestation of horses, donkeys and mules in tropical, dry and temperate regions in Mexico.

The aims of this study were to determine the prevalence of gastrointestinal nematodes (GIN) in horses, donkeys and mules and its associations with age, sex and climatic factors (derived from satellite data) to identify the potential risk factors of different climate regions in four Mexican states. From May 2017 to April 2018, a total of 560 farm owners or managers answered the questionnaire, and the data were used to establish three Köppen climate classes (tropical, dry, temperate). The overall prevalence of GIN parasites in equines was 77.9% (436 out of 560). The highest percentage of GIN was detected in mules (87.5%). In addition, the highest prevalence among the climate regions was found in the temperate climate (79.9%). The identified nematodes were strongyle type (77.9%), Parascaris equorum (5.7%), Strongyloides (0.7%) and Oxyuris equi (2.9%). The highest mean value of strongylid eggs per g faeces was found in the dry climate (632.6 ± 96.8), followed by that in the tropical climate (518.3 ± 49.7) and temperate climate (383.8 ± 30.2); however, a similar prevalence was observed between different climate regions. Age and sex were identified as risk factors for high shedding of strongyle eggs: the odds ratios for higher shedding intensities were 3.858 for geldings compared to mares, 2.602 for 6-10-year-old equines and 3.597 for ≤16-year-olds compared to young equines (≤5 years old).

The prevalence and risk factors of liver fluke infection in Mexican horses, donkeys and mules in tropical and temperate regions.

The objective of the present study was to determine the seroprevalence of Fasciola hepatica infection in horses, donkeys and mules from different climate regions in two states of Mexico. A total of 594 serum samples were analysed for immunoglobulin G (IgG) antibodies using enzyme-linked immunosorbent assay (ELISA), with excretory-secretory (E/S) products as the antigen. The diagnostic sensitivity and specificity of serum IgG ELISA were 100% and 97.2%, respectively. We collected data using a questionnaire. The overall prevalence of the parasite in equids between May 2018 and April 2019 was 13.1% (78 out of 594). The highest prevalence was found in mules (17.9%), followed by that in donkeys (13.9%) and horses (10.4%). In addition, the highest percentage of positive equines was detected in samples from the tropical climate (17.5%). The identification of risk factors was assessed by bivariate logistic regression analysis. Mules had 8.608 times higher risk for F. hepatica infections as compared with horses (odds ratio (OR) 8.608; 95% confidence interval (CI): 2.809-26.377), while the OR for 6-10-year-old equines was 93.375 compared with that of young equines (≤ 5 years old) (OR 93.375; 95% CI: 11.075-787.239). Likewise, tropical climate equines had 83.7% increased odds for fasciolosis (OR 1.83).

Prevalence of Toxoplasma gondii parasite in captive Mexican jaguars determined by recombinant surface antigens (SAG1) and dense granular antigens (GRA1 and GRA7) in ELISA-based serodiagnosis.

Toxoplasma gondii is an obligate intracellular protozoan parasite that infects almost all warm-blooded animals, including humans, causing serious public health problems. In this study, the seroprevalence of T. gondii in captive jaguars in 10 Mexican zoos was determined using single and mixtures of recombinant surface antigens (SAG1) and dense granular antigens (GRA1 and GRA7) in immunoglobulin G (IgG) enzyme-linked immunosorbent assays (ELISAs). Their efficacy was compared with the tachyzoite lysate antigen. All recombinant antigens were characterised by high sensitivity (92.5-97.5%); the specificity of the IgG ELISAs was variable (83.3-91.6%). Mixtures of the two recombinant proteins were generally more reactive than single antigens. GRA7 + SAG1 showed the highest sensitivity (97.5%) and specificity (91.6%), almost perfect agreement (96.2%), and a kappa value of 0.89. An area under the curve value of 0.998 represented a highly accurate test with a cutoff value of 0.8. The seroprevalence of anti-T. gondii IgG antibodies in the single and mixed recombinant antigen ELISAs was 75.0-76.9%. This study shows that GRA7 + SAG1 can be successfully used to diagnose T. gondii infection in jaguars for effective monitoring of prevalence and for devising control methods and prevention strategies against toxoplasmosis.

High prevalence, potential economic impact, and risk factors of Fasciola hepatica in dairy herds in tropical, dry and temperate climate regions in Mexico.

The aims of this study were to determine the prevalence of fasciolosis in cattle herds and its influence on production in order to identify potential risk factors in different climate regions and risk indices in three states of Mexico. The levels of anti-F. hepatica IgG1 antibodies were determined using an indirect ELISA. A total of 837 farm owners or managers answered the questionnaire. The total prevalence of the parasite over the period of January to March 2017 was 63.56%. The highest percentages of positive herds for the categories were found in Hidalgo state (88.33%), the no-risk classification (69.55%) and the temperate climate (76.53%). Overall, the production losses were approximately 0.51 to 1.00 kg of milk per parasitized cow per day. The annual costs of milk production loss per farm were estimated to range from US$2218.39 to US$6424.51, followed by anthelmintic treatment for young stock (US$67.68) and adult cows (US$209.47). Using multivariate logistic regression analysis, six final models were constructed for potential association with the ELISA results and were supported by the climatic, environmental and management factors. The results indicate that different factors were significantly associated with an increased risk of infection for each model.

Bulk tank milk prevalence and production losses, spatial analysis, and predictive risk mapping of Ostertagia ostertagi infections in Mexican cattle herds.

This study investigated the prevalence, production losses, spatial clustering, and predictive risk mapping in different climate zones in five states of Mexico. The bulk tank milk samples obtained between January and April 2015 were analyzed for antibodies against Ostertagia ostertagi using the Svanovir ELISA. A total of 1204 farm owners or managers answered the questionnaire. The overall herd prevalence and mean optical density ratio (ODR) of parasite were 61.96% and 0.55, respectively. Overall, the production loss was approximately 0.542 kg of milk per parasited cow per day (mean ODR = 0.92, 142 farms, 11.79%). The spatial disease cluster analysis using SatScan software indicated that two high-risk clusters were observed. In the multivariable analysis, three models were tested for potential association with the ELISA results supported by climatic, environmental, and management factors. The final logistic regression model based on both climatic/environmental and management variables included the factors rainfall, elevation, land surface temperature (LST) day, and parasite control program that were significantly associated with an increased risk of infection. Geostatistical kriging was applied to generate a risk map for the presence of parasite in dairy cattle herds in Mexico. The results indicate that climatic and meteorological factors had a higher potential impact on the spatial distribution of O. ostertagi than the management factors.

Production of aldosterone in cardiac tissues of healthy dogs and with dilated myocardiopathy.

BACKGROUND AND AIM: Aldosterone is a hormone, belonging to the group of mineralocorticoids, mainly synthesized in the adrenal cortex, basically its function is to regulate blood pressure and sodium-potassium levels in the body; high levels of this hormone have harmful effects in the organism and mainly in the heart in chronic form. Dilated cardiomyopathy is a progressive disease of heart muscle that is characterized by ventricular chamber enlargement and contractile dysfunction, is one of the most common cardiac conditions in dogs of medium and large breeds. The aim of the study was to determine and quantify if a dog's cardiac cells possess the capacity to synthesize aldosterone, as well as, the differences that appear between a healthy heart and with dilated myocardiopathy (DMC). MATERIALS AND METHODS: Cardiac tissues were used from six healthy dogs and six with DMC. Enzyme-linked immunosorbent assay was performed to determine if the dog's heart cells synthesized this mineralocorticoid in a similar way to rat, rabbit, and human tissues, as well as quantitative differences between the healthy heart and DMC. RESULTS: In healthy dog hearts, aldosterone values were 62.5 pG for both the atria and right ventricle and 125 pG for the left ventricle. As for dog hearts' with DMC, results were 125 pG in all four cavities. CONCLUSION: Both the healthy and DMC dog hearts produce aldosterone in all four cavities, observing that production increases in the atria and right ventricle of those hearts with DMC, as an intrinsic mechanism of cardiac remodeling.

Expression and biochemical characteristics of two different aldosterone receptors in both healthy and dilated cardiomyopathy dog heart tissue.

Recently, replicates of the aldosterone receptor expression have been done in healthy heart dog tissues through immunohistochemistry, showing an apparent heterogeneous distribution in the four chambers. Recent studies have also identified immediate effects of aldosterone, suggesting aldosterone also produces non-genomic effects caused by an unidentified receptor. In order to study the molecular and quantitative expression characteristics of aldosterone binding receptors in the canine heart, we conducted studies, using Western Blot, in the heart from both healthy animals and animals with dilated cardiomyopathy. The results show the presence and distribution of two aldosterone receptors; one of 110/120 kDa molecular weight, suggested as cytosolic/nuclear and the other of undetermined location with a 250 kDa molecular weight.

Owned dog ecology and demography in Villa de Tezontepec, Hidalgo, Mexico.

Dog overpopulation in developing countries has negative implications for the health and safety of people, including the transmission of zoonotic diseases, physical attacks and intimidation to humans and animals, as well as impacts on canine welfare. Understanding the ecology and demographic characteristics of a dog population can help in the planning and monitoring of canine population control programs. Little data exist regarding demography and dynamics of domestic dog populations in semi-urban areas in Mexico. A cross-sectional study was carried out between October 21 and November 7, 2015, to characterize the dog ecology and demography in Villa de Tezontepec, Hidalgo, Mexico. A face-to-face survey was used to collect data from randomly selected households in four contiguous communities using stratified two-stage cluster sampling. Within each household, adults answered questions related to their dogs and their experiences with dog bites and aggression. A total of 328 households were interviewed, representing a participation rate of 90.9% (328/361) and 1,450 people. Approximately 65.2% of the households owned one or more dogs, with a mean of 1.3 (SD=1.5) and 2.0 (SD=1.5) owned dogs in all participant households and dog-owning households, respectively. The human: owned dog ratio for all participant households was 3.4:1 (1450/428), and for the dog-owning households was 2.3:1 (984/428). The owned dog male: female ratio was 1.4:1 (249/179). Approximately 74.4% (95.0% CI=69.8% - 78.7%) of the owned dogs were older than one year (mean age: 2.9 years; SD=2.5). The mean age of owned female dogs at first litter was 1.9 years (SD=1.2) and the mean litter size was 4.2 puppies (SD=2.1). Approximately 36.9% (95.0% CI=31.8% - 46.4%) of the females were spayed, and 14.1% (95.0% CI=10.7% - 19.7%) of the males were neutered. Only 44.9% (95.0% CI=40.1% - 49.7%) were always confined when unsupervised. Approximately 84.4% (95.0% CI=80.6% - 87.7%) were reported to have been vaccinated against rabies in 2015. The knowledge of owned dog demography and ecology provided by this study can inform local government planning of dog population control interventions, and could serve as a baseline for the development of agent-based models to evaluate the effects of different dog population control strategies on dog demography.

Productive response of lambs fed Crescentia alata and Guazuma ulmifolia fruits in a tropical region of Mexico.

In vitro gas production with and without polyethylene glycol (PEG) of the fruits of Crescentia alata and Guazuma ulmifolia was evaluated, the degradation kinetics of lamb diets with added fruit of the tree was determined, and the ration intake and growth rate of lambs fed these diets were measured. Twenty-five entire male lambs of 23.5 ± 0.44 kg body weight were used and distributed in treatments: T0 (control without fruit); T1 and T2, 15 and 30 % of the fruit of C. alata; and T3 and T4, 15 and 30 % of the fruit of G. ulmifolia. Data variables chemical composition, fermentation kinetic, and digestibility in vitro were analyzed by a completely randomized design and data production response factorials design of five treatments by three evaluation periods. The total phenolic content (TP) (23.0 g/kg DM) was higher (P < 0.01) in the fruits of G. ulmifolia. The addition of PEG increased (P < 0.05) in vitro gas production (156.6 mL/g DM) in fruits of G. ulmifolia. In the fermentation kinetics, the total gas volume was higher (P < 0.01) at T0 (b = 293 mL/g DM), and the rate of degradation (c) but Lag time (t lag) was not different. In animal response, total dry matter intake was higher in lambs that received T4 (1.35 kg), and the daily weight gain and feed conversion did not differ (P > 0.05) among lambs receiving the treatments. Thirty percent G. ulmifolia fruit added in the diet increased dry matter intake and improved feed conversion but did not increase weight gain.

Cathepsin L1 mimotopes with adjuvant Quil A induces a Th1/Th2 immune response and confers significant protection against Fasciola hepatica infection in goats.

Thirty goats were randomly allocated in five groups of six animals each, for immunization with 1 × 10(14) phage particles of clones 11, 13, and 13 with Quil A adjuvant and wild-type M13KE phage at the beginning and 4 weeks later. The control group received phosphate-buffered saline. All groups were challenged with 200 metacercariae at week 6 and slaughtered 14 weeks later. The mean worm burdens after challenge were reduced by 46.91% and 79.53% in goats vaccinated with clones 13 and 13 with Quil A (P < 0.05), respectively; no effect was observed in animals immunized with clone 11 and M13KE phage. Animals receiving clones 11, 13, and 13 with Quil A showed a significant reduction in eggs output. Vaccinated animals produced parasite-specific total IgG antibody which were boosted after challenge with metacercariae of F. hepatica. Furthermore, levels of anti-phage total IgG increased rapidly within 2 weeks of the first vaccination and were always significantly higher in all vaccinated goats than in the infected control group. The fluke burden of goats immunized with clones 13 and 13 with Quil A was significantly correlated with IgG2 and total IgG. Goats vaccinated with phage clones produced significantly high titres of IgG1 and IgG2 antibodies indicating a mixed Th1/Th2 response. These data indicate that cathepsin L1 mimotopes has a potential as a vaccine candidate against Fasciola hepatica, whose efficacy will be evaluated in other host species, including those of veterinary importance.