Environmental factors associated with Dictyocaulus viviparus and Fasciola hepatica prevalence in dairy herds from Costa Rica.

Dictyocaulosis and fasciolosis are parasitic diseases that cause considerable economic losses for owners of farm animals worldwide, with special relevance on fasciolosis because it is an emerging zoonosis. Indirect diagnosis of these diseases through analyses of bulk milk tank samples has allowed carrying out large-scale prevalence studies, while the use of geographical information systems has helped to visualize and determine those variables that affect distribution of these pathogens. This study is intended to describe the spatial distribution of Dictyocaulus viviparus and Fasciola hepatica in dairy herds from Costa Rica, as well as their associated environmental factors. Bulk milk tank samples from 526 dairy herds in the three most important dairy regions of Costa Rica were analyzed using enzyme immunoassays. Results from the farms were subjected to spatial analyses using Holdridge's life zones, relief and soil type environmental layers. Of the total bulk milk tank samples analyzed, 3.8% (n=20) and 3.6% (n=19) were positive for D. viviparus and F. hepatica, respectively. Moran's I analysis revealed the existence of potential cluster (Moran's I=1.789, z=12.726 p<0.05) for D. viviparus. Consequently, Getis-Ord General G analysis showed that the spatial distribution of positive farms in the dataset was clustered (Observed General G=0.015, variance=0.000001, z=12.823, p<0.05). No significant positive spatial autocorrelation (Moran's I=0.038, z=0.286, p>0.0.5) was observed for F. hepatica. Furthermore, a significant difference was detected in the spatial locations of both parasites (latitude p<0.05, longitude p<0.05), and about the spatial distribution of both D. viviparus negative and positive farms (latitude p<0.05, longitude p<0.05), as well as in F. hepatica negative and positive farms regarding on latitude (p<0.05), but not on longitude (p>0.05). In the case of environmental factors, significant differences were found for D. viviparus and F. hepatica with respect to types of soil, precipitation, altitudinal belts, life zones, biotemperature, and elevation.

Detection of antibodies against Chlamydophila abortus in Costa Rican sheep flocks.

A total of 359 sheep samples from 15 flocks were analyzed for the presence of antibodies against Chlamydophila abortus using a commercial Enzyme linked Immunosorbent Assay (ELISA). Antibodies were detected in 19 (5.29%) sheep from 12 (80%) flocks. Seropositive animals were found in all analyzed regions (Central, Chorotega, Atlantic Huetar, North Huetar and Central Pacific) determining prevalence between 0.28% and 4.4%, and intra-flock positivity between 3.7% and 25.0%. The survey revealed two risk factors associated with seropositivity; introducing animals (males and females), embryos, or semen from other farms or from abroad without any sanitary certification, and flocks not having quarantine areas or separated boxes for diseased animals. No clinical signs of disease were observed in positive seroreactors. C. abortus seems to be present in Costa Rica in a very low prevalence in sheep flocks. Further studies, to isolate the bacteria are required. Finally, implementation of control measures to prevent the spread of C. abortus is recommended.

Serosurveillance of infectious agents in equines of the Central Valley of Costa Rica.

Blood samples from 181 equines from the Central Valley of Costa Rica were collected in the year 2012 to determine the presence of antibodies against selected infectious agents in horses and to determine the risk factors associated with these agents. The presence of antibodies against Equine Infectious Anemia Virus (EIAV), Equine Herpes Virus 1 and 4 (EHV-1 and EHV-4), West Nile Virus (WNV), Influenza A Virus (IAV), Equine Viral Arteritis Virus (EVAV), Babesia caballi, Theileria equi, Neospora caninum and Chlamydia abortus was determined using commercial assays, and risk factors associated with seropositivity to the different infectious agents was established. The most seroprevalent agent detected was EHV-4 (96.7%), followed by WNV (44.2%), and IAV (41.8%). Horses >3 years, used for work or sports, and with access to pastures, had significantly increased probability to be seropositive to WNV, whereas horses used for breeding and recreational purposes, being stabled, and without access to pastures, had significantly greater probability to be seropositive to IAV. Seroprevalence to B. caballi (19.9%) was lower than to T. equi (38.1%). For B. caballi, access to pastures was determined as a risk factor, whereas being older than 3 years was established as a risk factor for T. equi. Low seroprevalences were determined for EHV-1 (5.0%), EVAV (5.0%), C. abortus (4.8%), and N. caninum (4.4%). Mares having history of abortion were more likely to be seropositive to EHV-1, whereas horses >3 years, used for work and sports, and mares having multiple parturitions, were more likely to be seropositive to N. caninum. None of the horses were seropositive to EIAV. Earlier, only diseases caused by EIAV, WNV and piroplasmosis were reported in Costa Rica. The present study however, determined the presence of carriers for EHV-1, EHV-4, and EIAV.

Chlamydia psittaci genotype B in a pigeon (Columba livia) inhabiting a public place in San José, Costa Rica.

Human chlamydiosis is a zoonotic disease of avian origin caused by Chlamydia psittaci. The highest infection rates have been detected in parrots (Psittacidae) and pigeons (Columbiformes), the latter most frequently carry the genotypes B and E. These genotypes have been shown to also infect humans. Because pigeons (Columba livia) cohabit with humans in urban areas, C. psittaci present in the dust from dry feces of infected pigeons may be transmitted by inhalation and represent a significant public health problem. Between 2012 and 2013 a total of 120 fecal samples were collected from pigeons at four public places (Plaza de la Cultura, Parque Morazán, Parque Central de Guadalupe, Plaza de las Garantías Sociales) in San José, Costa Rica. A nested polymerase chain reaction (PCR) was used to amplify a region of the outer membrane protein A gene of C. psittaci. Only one sample was positive in PCR and the positive sample was further subjected to sequencing and genotyping. Sequencing identified this sample as C. psittaci genotype B. This study is the first report to show the presence of this organism in pigeons of Costa Rica, and shows that the infected pigeons may represent a significant risk for humans who visit public places that are inhabited by pigeons.

First isolation and molecular characterization of Ehrlichia canis in Costa Rica, Central America.

The present study investigated Ehrlichia species in blood samples from dogs suspected of clinical ehrlichiosis, using molecular and isolation techniques in cell culture. From a total of 310 canine blood samples analyzed by 16S rRNA nested PCR, 148 (47.7%) were positive for Ehrlichia canis. DNA from Ehrlichia chaffeensis or Ehrlichia ewingii was not detected in any sample using species-specific primers in separated reactions. Leukocytes from five PCR-positive dogs were inoculated into DH82 cells; successful isolation of E. canis was obtained in four samples. Partial sequence of the dsb gene of eight canine blood samples (including the five samples for in vitro isolation) was obtained by PCR and their analyses through BLAST showed 100% of identity with the corresponding sequence of E. canis in GenBank. This study represents the first molecular diagnosis, isolation, and molecular characterization of E. canis in dogs from Costa Rica.

A cross-sectional survey of gastrointestinal parasites with dispersal stages in feces from Costa Rican dairy calves.

A cross-sectional study was carried out to determine the prevalence of gastrointestinal parasites and lungworm nematodes in dairy calves from five different ecoclimatic areas of Costa Rica. Also intensity of infection of nematodes was determined. In order to describe management practices and anthelmintic control, a questionnaire was applied in 73 farms. The influence of area, farm, host (breed, age) and ecological factors (low and high rainfall period) upon eggs per gram feces (epg) of gastrointestinal nematodes (GIN) and first larval stage counts (L1) of Dictyocaulus viviparus were investigated. Furthermore, association of host, ecological and management risk factors to the prevalence of gastrointestinal parasites and D. viviparus were analyzed. The most prevalent GIN, cestodes and protozoan identified in dairy farms were similar in all areas studied. Strongylidae was the most prevalent parasite group detected, represented mainly by Haemonchus spp. and Cooperia spp., whereas Ostertagia spp. and Mecistocirrus digitatus were barely found. The most prevalent protozoan was Eimeria spp. The questionnaire applied to producers revealed the following management practices: weaning age of calves 1-4 months (52.1%), semi-confinement of calves upon 5-8 months of age (41.1%), number of paddocks used for calves <10 (57.5%), first deworming of calves at ages ≥15 days (74.70%) and deworming of calves at intervals >60 days (52.1%). Anthelmintic products were changed in 56.1% of the farms at intervals between 13 and 24 months. Although 91.8% of the farms had veterinary assistance, the majority performed parasite control regimes according to the criteria of the producers (66.7%). Common practices were the dispersion of animal feces on the pastures (64.4%) and use of disinfectant in the milking room (63.4%). The analyses of variance showed significant influence (p<0.05) of age, rainfall period, interaction of rainfall period on area (rainfall period×area) and nested effect of farm within area [farm (area)] on epg of Strongylidae; age, area, rainfall period×area and [farm (area)] on epg of Strongyloides papillosus; age, rainfall period and farm (area) on epg of Trichuris spp.; rainfall period, rainfall period×area and [farm (area)] on L1 of D. viviparus. The logistic regression analyses determined area, semi-confinement, management of feces, use of disinfectant in the milking room as risk factors for the presence of Strongylidae, S. papillosus and Trichuris spp; rainfall, age, paddock numbers for D. viviparus; and area, age, veterinary assistance, deworming program, age at first deworming and use of disinfectant in the milking room for Eimeria spp. and Buxtonella sulcata.

Dictyocaulus viviparus seroprevalence and epidemiology in Costa Rican dairy cattle.

A cross-sectional serological survey of Dictyocaulus viviparus was carried out to determine the prevalence of lungworm infections in 28 dairy cattle farms distributed in five selected areas from Costa Rica. The influence of area, farm, host (breed, age and lactation number) and ecological factors (altitude and life zones) on the presence of lungworm infection was analyzed. A sub-sample of 924 sera collected between September 1998 and July 1999 was processed by ELISA (Ceditest). A total of 162 (17.5%) animals from 26 (93.0%) farms showed antibodies against D. viviparus. The overall seroprevalence detected among areas was Poás 25.0%, Cartago 24.3%, Tilarán 22.0%, Alfaro Ruiz 12.0% and San Carlos 12.1%. Using analysis of variance no significant influence of area and host factors on D. viviparus infections was determined, whereas the variable farm within area was highly significant (p<0.001). However, altitude and life zones showed significant association to seropositive animals, when a Chi-square test was applied. In altitudes of 1000-2000 m (p<0.001) and life zones of Lower Montane moist forest and Montane moist forest (p<0.001) D. viviparus infections in bovines were significantly higher. The results obtained in this study indicate a high D. viviparus seroprevalence in the analyzed farms and that the factors farm, altitude and life zones were significantly related to lungworm infections.

Fecal and serological survey of Neospora caninum in farm dogs in Costa Rica.

To detect oocysts of Neospora caninum in dog feces and to determine the excretion pattern in dogs from specialized dairy farms in Costa Rica, a total of 265 fecal samples from 34 dogs were collected at intervals from February to August 2005. Fecal samples were examined for N. caninum-like oocysts microscopically, by DNA detection using the polymerase chain reaction (PCR), and by bioassay. N. caninum DNA was detected by PCR in four fecal samples, twice from one dog, but oocysts were not detected microscopically in these dogs. Sera of 31 of 34 dogs were tested for antibodies to N. caninum by a competitive-inhibition ELISA (VMRD). Fifteen (48.4%) of 31 dogs had antibodies to N. caninum by ELISA. Seroconversion was not found in 28 dogs that were bled twice, 4 months apart (March and July 2005). Only one dog tested positive to N. caninum by both ELISA and PCR. This is the first report of finding N. caninum DNA in feces of naturally infected dogs in Costa Rican dairy farms.

Prevalence of Toxocara spp., Toxascaris leonina and ancylostomidae in public parks and beaches in different climate zones of Costa Rica.

This epidemiological study was conducted in different regions of Costa Rica to determine the prevalence of the developmental stages of potential zoonotic intestinal helminths of dogs and cats in public places. Samples were collected within three main climate zones including rural and urban areas during both the rainy and the dry season. Faecal and environmental samples were taken from 69 parks and beaches. Of the faecal samples 3% contained Toxascaris spp. eggs, 7% Toxocara spp. eggs and 55% contained ancylostomidae eggs. Of the soil samples, 2% contained ancylostomidae eggs and 0.8% contained ascarid eggs. Significant differences in the presence of parasites were found in faecal samples of dry, moist and wet climate zones and between the dry and rainy seasons. Significant differences in the presence of eggs and larvae were also found in the grass samples in the dry, the moist and the wet climate zones and between the different seasons. No significant differences were found between rural and urban areas.

Dynamics of infections with gastrointestinal parasites and Dictyocaulus viviparus in dairy and beef cattle from Costa Rica.

A longitudinal survey was carried out to determine and describe the prevalence and intensity of gastrointestinal parasite infections and Dictyocaulus viviparus in a dairy and a beef cattle farm of two different ecological zones in Costa Rica. The influence of anthelmintic treatment, age and meteorological factors (rainfall, minimum and maximum temperatures) on gastrointestinal nematodes and D. viviparus counts was determined. Calves were subjected to monthly sampling of feces and blood between April 2002 and March 2003. Coprological techniques were used to detect gastrointestinal helminthes, protozoan and D. viviparus. Blood samples were analyzed for antibodies to D. viviparus by ELISA. The most prevalent gastrointestinal parasites detected on both farms (dairy cattle, A; beef cattle, B) were Eimeria spp. (94.7%, 93.7%), Strongylidae (75.0%, 81.4%), Buxtonella sulcata (38.0%, 21.6%) and Strongyloides papillosus (29.8%, 31.7%), whereas Moniezia benedeni (4.8%, 9.1%), Trichuris spp. (7.3%, 13.2%), Toxocara vitulorum (0.0%, 1.8%) and Entamoeba bovis (2.5%, 1.1%) were less prevalent. Mean fecal egg counts (FEC) showed highest values of Strongylidae in April, May and July (>335.3 eggs/g feces) on farm A, and April, May and August (>304.3 eggs/g feces) on farm B. S. papillosus presented low FEC throughout the year on farm A, on farm B the highest values were obtained in April (303.0 eggs/g feces). Trichuris spp. presented maximum FEC values in May (328.6 eggs/g feces) on farm A and in June (157.5 eggs/g feces) on farm B. Treatment and age had significant influence on infection intensity of Strongylidae (farms A and B), S. papillosus (farms A and B) and Trichuris spp. (farm A). Rainfall had significant effect on S. papillosus (farms A and B) and Trichuris spp. (farm B). Maximum temperature showed significant effect on S. papillosus (farm A) and Trichuris spp. (farms A and B). Minimum temperature had significant influence on Strongylidae (farm A), S. papillosus (farms A and B) and Trichuris spp. (farm B). Haemonchus spp. (57%, 66%) and Cooperia spp. (30.0%, 30.7%) were the most prevalent genera identified by coproculture on both farms, in contrast, Trichostrongylus spp. and Oesophagostomum spp. were less frequent. Patent lungworm infections were low on both farms (10.8%, 1.8%). On farm A, high prevalence of antibodies against D. viviparus was determined only at the beginning of the study, in contrast, on farm B the seroprevalence fluctuated throughout the year. Treatment, age and maximum temperature had significant effect on D. viviparus counts on farm A, but not on farm B.

Effect of neosporosis on productive and reproductive performance of dairy cattle in Costa Rica.

A cross-sectional study was conducted to assess the effect of neosporosis on productive and reproductive parameters in dairy cows. Cows (n=2743) from 94 farms located in the most important dairy areas in Costa Rica were used in the study. The size of the herds ranged from 32 to 379 females (mean=110, median=125). An indirect ELISA was used to determine the serostatus of the cows towards Neospora caninum. The effect of neosporosis on milk production was analysed by a mixed linear model. In addition, the effects on calving interval (days) and calving to conception interval (days) were analysed by survival analysis. The risk of abortion in relation to N. caninum serostatus was assessed by logistic regression, with herd as a random effect. Overall, 1185 of 2743 cows (43.3%) were seropositive for Neospora. Eighty-nine of 94 (94.7%) farms were classified as Neospora-seropositive. It was estimated that cows seronegative to Neospora produced an additional 84.7L of milk (P=0.6). Serostatus did not have a significant effect on the length of the calving interval in the Cox proportional hazard survival analysis (Hazard ratio=1.2, 95% CI: 0.9, 1.4). The logistic regression model had a weak positive association between serostatus and abortion (OR 1.7, 95% CI: 0.8, 3.9), but did not show a strong association between serostatus and the number of services per conception (OR=0.95, 95% CI: 0.7, 1.3). In conclusion, there were no significant effects of Neospora serostatus on production and reproductive performance in this study.

Factors associated with Neospora caninum serostatus in cattle of 20 specialised Costa Rican dairy herds.

Twenty-five specialised Costa Rican dairy farms (located in the Poás area) were used to determine neosporosis seroprevalence and the association of seropositivity with environmental and management factors. The farms involved were selected intentionally and all of them use VAMPP 5.1 (Veterinary Automated Management and Production Control Programme) as management-information system. Holstein-Friesian, Jersey and crosses between them were the most-frequent breeds in these herds. The number of females per farm varied from 41 to 296. Our cross-sectional study had two phases. In the first phase, we determined the presence or absence of seropositivity at herd level. For the second phase, all females in 20 seropositive farms were bled. Serum samples were tested for antibodies to Neospora caninum using an indirect enzyme-linked immunosorbent assay (ELISA). A questionnaire with factors mentioned in the literature was administered to the farmers. Logistic regression (LR with herd as random effect) was used to assess the relationships of the serostatus at the individual level with characteristics of the cows and environmental factors. In the first phase all herds had >20% seropositive females; therefore, all herds were eligible for the second phase. In the second phase, the overall prevalence was 39.7% (1191/3002), and within-herd prevalences were between 25.0 and 70.5%. Age 3-6 years, parity < or =2 of the dam of the cow, Jersey breed and lack of purposive sampling to diagnose abortive infectious disease were associated with positive serostatus; other management and environmental factors did not show significant associations. The lack of association between management and environmental factors with serostatus might be because all farms were exposed to a considerable number of potential factors. That all herds of this study were seropositive for neosporosis and the within-herd prevalence was considerable raises questions about how far the infection is spread in other dairy areas of Costa Rica.

Serological status for Chlamydophila psittaci, Newcastle disease virus, avian polyoma virus, and Pacheco disease virus in scarlet macaws (Ara macao) kept in captivity in Costa Rica.

From 1998 to 1999, a total of 128 blood samples were collected from scarlet macaws (Ara macao), kept in captivity in 11 different aviaries located in six provinces of Costa Rica. The sera were examined for antibodies directed against Chlamydophila psittaci, Newcastle disease virus (NDV), avian polyoma virus (APV), and Pacheco disease virus (PDV). Testing by enzyme-linked immunosorbent assay (ELISA), showed 16 (12.39%) of the samples (n = 129) exhibited antibodies directed against C. psittaci. Employing haemagglutination inhibition tests for NDV antibodies, all of the samples were found to be negative. The prevalence of antibodies specific for APV was tested with a blocking ELISA and serum neutralization tests (SNT) and 12 of 128 samples (9.37%) were found to be positive with both tests. In SNT, two out of 128 samples (1.56%) were positive for PDV. This is the first description of the serological status in scarlet macaws in captivity in Costa Rica. The study demonstrates the absence of NDV antibodies in the birds investigated on one hand, but also indicates a health hazard for numerous avian species due to the risk of infections with C. psittaci, APV or PDV.

Comparison of agar gel immunodiffusion test, enzyme-linked immunosorbent assay and western blotting for the detection of BLV antibodies.

An indirect enzyme-linked immunosorbent assay (ELISA) for the diagnosis of bovine leukaemia virus (BLV) infection was developed and compared with the agar gel immunodiffusion test (AGIDT). Western blotting (WB) was used as confirmatory test. ELISA and AGIDT had specificities that were comparable with that of WB, however, ELISA showed a higher sensitivity than AGIDT. The ELISA was useful for screening a large number of samples, whereas WB was important for detecting the antibody response against the individual BLV-proteins. Different types of positive serological reactions were discerned in WB, that correlated with reactions of sera in AGIDT and ELISA. The most important antigen in WB and ELISA was the BLV protein p24, whereas the BLV glycoproteins gp51 and gp30 were of special importance in AGIDT. The relevance of repeatedly testing the antibody response in BLV-infected herds for control and eradication programmes using assays with higher sensitivity than AGIDT was demonstrated.

Bovine leukaemia-virus infection in Costa Rica.

The geographical distribution of bovine leukaemia-virus (BLV)-infected herds in Costa Rica and the isolation of BLV from cases of enzootic bovine leukosis are presented. It was found that BLV is prevalent throughout the country, affecting mostly dairy cattle (Bos taurus) located in the highlands around the central valley of Costa Rica. By contrast, beef cattle (Bos indicus) raised in the lowlands were affected to a lesser extent. One out of four isolates of BLV obtained from local cattle with lymphosarcomatous tumours was analysed and had similar physical, serological and biological characteristics to a reference strain of BLV isolated in the USA. However, different patterns of recognition of BLV-protein p24 were observed among naturally infected cattle using the western-blotting technique.

[Progressive tibial lengthening in children (author's transl)]. / L'allongement progressif du tibia chez l'enfant. A propos de 48 cas.

The authors have studied 38 "simple" tibial lengthenings and 10 "iterative" tibial lengthenings in children, with the use of an external rigid distractor. Four out of five of these lengthenings have been performed in cases of constitutional shortness. The authors got an average lengthening of 4,2 cm (that is 17,5 p. 100) for first procedure and 5,5 cm (16,5 p 100) for iterative ones. Consolidation has been obtained after about 3 months and free walking was authorized after 6 months for simple lenghtenings. These delays are longer (5 months and 6 and a half months) for iterative lengthenings. The bone and articular complications observed are closely linked to the technic used. The authors recommended the use of an external distractor of Wagner type, an oblique osteotomy with ample oblique section and extensive tenotomies and aponevrotomies. Out of the author's own experience, the most frequent complications were: delay in consolidations, especially in acquired shortness, and foot complications, especially frequent on those patients with previous alterations at that level. Progressive tibial lengthenings must be, of course, part of a complete program of length equalization of the inferior limbs and must however be preceded by the necessary surgery to obtain rightness and stabilisation of the knee, leg, ankle and foot before starting the required lengthening.