Seroprevalence of Neospora caninum, Toxoplasma gondii and Sarcocystis sp. in llamas (Lama glama) from Jujuy, Argentina

Llamas (Lama glama) are South American camelids described as intermediate hosts of Neospora caninum, Toxoplasma gondii and Sarcocystis aucheniae. Due to the potential role of these protozoan infections as a cause of economic losses, the aim of this study was to determine the seroprevalence for T. gondii, N. caninum and Sarcocystis sp. in llamas from Argentina. Serum samples from 308 llamas (>2 years old) were collected between 2005 and 2007. A total of 55 farms located in six departments of Jujuy province, Argentina were sampled. Presence of antibodies to N. caninum, T. gondii and Sarcocystis sp. was determined by the indirect fluorescent antibody test (IFAT). For Sarcocystis, 2 different bradyzoites-based antigens were prepared using S. aucheniae and S. cruzi. Sera were tested at dilutions 1:25 and 1:50. Antibodies to N. caninum were found in 4.6% serum samples. Fifty percent of departments and 14.5% of farms had positive animals. Antibodies to T. gondii were found in 30% of samples, distributed in 66% of departments and 43.6% of farms. Antibodies to Sarcocystis sp. were detected in 96% of samples and all departments and farms had positive animals, suggesting frequent contact between llamas and canids. Co-infection with N. caninum, T. gondii and Sarcocystis sp. was also recorded. Low seroprevalence of N. caninum in llamas detected in this study could be related to climatic and geographical conditions that limit cattle breeding activity, reducing the source of infection for definitive hosts. Seroprevalence of T. gondii and the positive animal distribution suggest frequent contamination of grass with felid faeces. In conclusion, this is the first report of combined seroprevalence for N. caninum, T. gondii and Sarcocystis sp. in llamas. Further studies are needed to determine the potential role of these protozoan infections as cause of abortion in Argentina as well as presence of these protozoans in llama meat used for human consumption.     

Toxoplasma gondii infection in adult llamas (Lama glama) and vicunas (Vicugnavicugna) in the Peruvian Andean region

The present study was designed to investigate Toxoplasma gondii infection in adult llamas (Lama glama) and vicunas (Vicugna vicugna) in the Peruvian Andean region, for which to date no information has been available. Serum samples from 43 llamas (L. glama) and 200 vicunas were tested by IFAT detecting titres of 1:50 or higher in 55.8% (33.9-70.9%) and 5.5% (2.8-9.6%), respectively. IFAT titres ranged from 1:50 to 1:6400. In order to avoid cross reactions with closely related coccidian parasites and to confirm the existence of T. gondii specific antibodies, IFAT positive sera from both ruminant species were also analysed by western blot. T. gondii specific antigens were recognised by IFAT positive sera, although different IFAT cut-off points could be selected for llamas (1:200) and vicunas (1:50) meaning seroprevalence of 44.2% (29.1-60.1%) and 5.5% (2.8-9.6%), respectively. Based on the frequency and intensity of tachyzoite antigen recognition, at least three immunodominant antigens with apparent molecular weights of 22-24, 30, and 38-40 kDa were detected, together with other minor protein fractions located in the 18-73 kDa range. This study documents for the first time the presence of T. gondii infection and reports the target T. gondii antigens in adult llamas and vicunas in Peru.     

Suckling behavior of llamas (Lama glama)

In the present study the suckling behaviour of six Ilama crias was analysed. A total of 3398 suckling bouts were observed and evaluated. The crias mostly suckled in a reverse parallel position with no side preference. The cria initiated suckling in most suckling bouts whereas the dams usually ended them. The total number of suckling bouts could be divided into two types: suckling bouts of short duration and suckling bouts of long duration. There were no significant differences in duration per suckling bout among the different ages of crias. But the animals suckled less frequently and spent less total time suckling as they grow older. The age related differences and evidence for a dual function (nutrition, social contact) of suckling behaviour are discussed.     

Pharmacokinetics of ivermectin in llamas (Lama glama)

The pharmacokinetic behaviour of ivermectin was investigated in adult llamas (Lama glama) by using high performance liquid chromatography with a lower limit of quantification of 2 ng/ml to measure its concentration in serum. Llamas were treated with one of three commercial formulations (injectable, pour-on or oral paste) at dosages recommended by the manufacturer, or with an experimental injectable sustained-release formulation. In five llamas given 1 per cent ivermectin subcutaneously at 200 microg/kg, the median peak serum concentration (Cmax) was 3 ng/ml and the area under the serum concentration-time curve (AUC) was 13.5 ng x day/ml. In six llamas treated topically with 0.5 per cent ivermedin pour-on at 500 microg/kg, Cmax was 2.5 ng/ml or less and the AUC was 7.75 ng x day/ml or less. In seven llamas with measurable concentrations of ivermedin, the median times to peak serum concentration (tmax) were six days after subcutaneous injection and seven days after treatment with the pour-on formulation. In six llamas, the serum concentration of ivermectin remained less than 2 ng/ml for 124 hours after treatment with a 1.87 per cent oral paste at 200 microg/kg. In five llamas treated subcutaneously with 25 per cent ivermectin sustained-release microspheres at 1500 microg/kg, the median Cmax was 5 ng/ml and the median AUC was 224 ng x day/ml.     

Detection of specific antibodies to Neospora caninum and Toxoplasma gondii in naturally infected alpacas (Lama pacos), llamas (Lama glama) and vicunas (Lama vicugna) from Peru and Germany

Sera of an experimentally Neospora caninum infected llama and a non-infected control llama were used to establish an immunoblot, an ELISA and an IFAT to detect antibodies against N. caninum tachyzoites. Subsequently, serum samples collected from a total of 871 South American Camelids (SAC: Lama glama, Lama pacos, Lama vicugna) of two farms in Peru and from 32 SAC of a farm in central Germany were examined for antibodies against N. caninum and Toxoplasma gondii. Based on the recognition of specific bands in the immunoblot, sera of SAC from Peru were differentiated into N. caninum-positive (n = 18) and T. gondii-positive (n = 30) samples and into samples negative or inconclusive for both parasites. Using the immunoblot results as the reference, a modified version of the p38-ELISA and the IFAT were evaluated for detecting N. caninum antibodies in SAC sera. Applying a cut-off as determined by two graph-receiver operating characteristic analysis both, the ELISA and the IFAT, exhibited a sensitivity and specificity of about 95% in the SAC sera from Peru. Serological testing confirmed that SAC may become infected with N. caninum under field conditions in Peru. In addition to alpacas and llamas also 114 wild living vicunas had been examined for antibodies against N. caninum. However, only the alpacas and llamas but no vicunas were found N. caninum-positive. In contrast, T. gondii-seropositive animals were detected in all three SAC species. The lack of N. caninum-seropositive vicunas indicates that in the study area in Peru wild canids might not serve as definitive hosts of N. caninum while for T. gondii a life cycle including wild felids is likely. On the German farm no N. caninum- but only T. gondii-seropositive SAC (n = 14) were detected. The seroprevalence of T. gondii infection was significantly higher in adult SAC (alpacas in Peru, llamas in Germany) than in crias (i.e. < 12 months old foals) indicating that the predominant route of infection is post natal. Since the present study was restricted to a few farms, the seroprevalences determined are not representative. However, our results confirm natural infections with N. caninum and T. gondii in SAC. Whether these infections are linked to any disease, e.g. reproductive losses, has to be clarified in further studies.     

Intraocular pressure in normal llamas (Lama glama) and alpacas (Lama pacos)

Objective: To determine the mean intraocular pressure in llamas ( Lama glama ) and alpacas ( Lama pacos ) using applanation tonometry. Animals studied: Ten llamas and 10 alpacas. Procedures: Intraocular pressure (IOP) was measured with a Tono-Pen™ XL (Mentor Ophthalmics, Inc., Norwell, MA, USA). Three values, with 5% variance, were recorded for each eye. Least-squares means were determined for IOP for each eye of llamas and alpacas. Controlling for age, differences between left and right eye were analyzed using anova . Two age groups were established, less than 5 years and greater than 5 years. The effect of age on IOP within each group was analyzed by linear regression. Probability values of less than 0.05 were considered significant. Results: Comparison of mean IOP between right ( n  = 20) and left eyes ( n  = 20), independent of species type, showed no differences in IOPs for llamas and alpacas. Mean IOP declined with increasing age in llamas and alpacas. Mean IOPs for 20 eyes in 10 llamas was 16.96 ± 3.51 mmHg. Mean IOP for 20 eyes in 10 alpacas was 16.14 ± 3.74 mmHg. Mean IOP for all eyes ( n  = 40), independent of species, was 16.55 ± 3.55 mmHg. The range of IOP in normal llamas and alpacas within 2 SD (95% of the population) was 14.89±18.21 mmHg. Conclusions: There was no significant difference in IOP between alpacas and llamas. Mean IOP in both species decreased with increased age.     

Fetal membranes and ancillary structures of llamas (Lama glama)

The placenta of llamas is epitheliochorial, with patchy areas of dense folded papillation serving as the placentome. The amnion of the full-term placenta is closely adhered to either the allantois or the chorion and remains with these structures at the time of parturition. Llamas and alpacas, like dromedaries, have an extra fetal membrane that is derived from the epidermis of the fetus. In association with the watery amniotic fluid of llamas, the epidermal membrane is slippery, facilitating delivery of the fetus.     

Serological survey of viral antibodies in llamas (Lama glama) in Argentina.

This study analysed sera from 390 llamas (Lama glama) from nine farms located in three different Argentine provinces: Buenos Aires, Cordoba and Jujuy. The samples were tested for antibodies against 8 virus known to infect cattle: bovine herpesvirus type 1 (BHV-1), bovine viral diarrhea virus (BVDV), bovine adenovirus (BAdV III), bovine enterovirus (BEV), bovine rotavirus (BRV), bluetongue virus (BTV), bovine leukaemia virus (BLV), and foot-and-mouth virus (FMDV) by conventional methods such as seroneutralization, immunoperoxidase staining, and agar gel immunodiffusion. The antibody prevalences detected in llamas were: BHV-1 in 0.77% (3/390), BVDV in 2.05% (8/390), BAdV III in 5.13% (20/390), BEV in 4.10% (16/390), BRV in 87.69% (342/390). No antibodies against BTV, BLV and VIAA (FMDV infection associated antigen) were detected.     

Plasma progesterone concentrations in pregnant and non-pregnant llamas (Lama glama)

Female llamas ovulate in response to copulation, and progesterone secretion by the corpus luteum indicates recent ovulation (mating) and, or, pregnancy. The plasma progesterone concentration was 0.9 to 1.4 ng/ml in five non-pregnant llamas and 7.4 to 9.2 ng/ml in three llamas in the last month of pregnancy. After ovulation had been induced in nine of 10 llamas by a single intramuscular injection of 500 or 750 iu of human chorionic gonadotrophin, the plasma progesterone concentration increased after two days from 0.5 to 1.2 ng/ml to 4.6 to 10.3 ng/ml after six to nine days and returned to basal values after 10 to 13 days, reflecting the life-span of a corpus luteum in the absence of conception. After a male llama had been introduced into a group of 13 females, 10 matings which resulted in eight conceptions occurred in the first 11 days, and 11 of the llamas became pregnant. The llamas' progesterone concentrations increased after mating and remained high if conception had occurred: 6 to 12 ng/ml in months one to four, and 5 to 9 ng/ml in months five to nine of the 11-month gestation. Two of the 13 llamas had high concentrations of progesterone although they did not become pregnant.     

Evaluation of normal triiodothyronine and tetraiodothyronine concentrations in llamas (Lama glama)

Basal serum triiodothyronine (T3) and tetraiodothyronine (T4) concentrations have not been established for the llama (Lama glama). In addition, changes in T3 and T4 concentrations in response to thyroid-stimulating hormone (TSH) administration have not been determined, making clinical evaluation of problems referable to thyroid dysfunction difficult. In study 1, basal T3 and T4 concentrations were determined in serum samples collected from 132 clinically healthy llamas. The llamas were allotted to 3 groups: mature intact or neutered males (group I, n = 25), nonpregnant sexually mature females (group II, n = 21), and pregnant females (group III, n = 86). A mean concentration and a 95% confidence interval were computed for each group. An analysis of variance (ANOVA) indicated that a single confidence interval range (0.45 to 4.18, mean = 1.37 ng T3/ml) adequately defined the normal T3 concentrations for all groups. An ANOVA indicated that the T4 concentrations for the female populations (groups II and III) could be combined with a normal confidence interval range of 39 to 204 ng/ml (mean = 88 ng/ml), whereas a separate range (70 to 220 ng/ml, mean = 124 ng/ml) was determined for the male population. An ANOVA indicated that a single confidence interval range (0.0066 to 0.0321, mean = 0.0146) adequately defined the normal T3/T4 ratio for all groups. In study 2, T3 and T4 concentrations were evaluated in 10 healthy llamas immediately preceding and at 2, 4, 6, 8, and 24 hours after the IV administration of 3 IU of TSH/44 kg of body weight. The T3 and T4 concentrations were significantly higher by 2 hours after TSH administration in both groups.(ABSTRACT TRUNCATED AT 250 WORDS)     

Morphometric evaluation of growth in llamas (Lama glama) from birth to maturity.

Normal growth characteristics of llamas (Lama glama) were evaluated from conception until 1 week after parturition in pregnant females (study 1) and from 1 month after birth to maturity (study 2). In study 1, pregnant multiparous llamas (n = 10) were measured at monthly intervals from conception until 1 week after parturition; in study 2, llamas (n = 270) were measured once. Body weight of pregnant llamas (study 1) did not increase significantly until after the eighth month of pregnancy. Llamas of study 2 reached mature height, length, thoracic circumference, and weight at 18, 24, 36, and 36 months of age, respectively. From 1 month of age to maturity, the growth characteristics of males, nonpregnant females, and females during the first 8 months of pregnancy did not differ. Correlations (r2) between height, length, and thoracic circumference related to body weight for all but the pregnant llamas during the last 3 months of pregnancy were 0.822, 0.834, and 0.948, respectively. The equation describing thoracic circumference as a predictor of body weight was: Weight (kg) = (1.005 x 10(-3)) x circumference (cm)2.424.     

Results of Schirmer tear test in clinically normal llamas (Lama glama)

Purpose To determine the normal reference range for Schirmer tear test (STT) values in clinically normal llamas (Lama glama) Animals Nine captive llamas (Lama glama) (seven females and two males) were used in this study. Procedure Complete ophthalmic examinations were performed without chemical restraint. STT I values were evaluated in both eyes of all llamas using a commercial STT strip of a single lot number (Schirmer‐Tränentest^®, Germany). STT II value was also measured in both eyes of seven female llamas. Results No statistically significant differences among ages or between right and left eyes were found for any of the results. The mean ± SD STT I of 18 eyes of nine llamas was 17.3 ± 1.1 mm/min (Range 15–19 mm/min). The mean ± SD STT II of 14 eyes of seven llamas was 15.4 ± 1.7 mm/min (Range 12.5–17.5 mm/min). A paired samples t‐test demonstrated that there was a significant difference between the STT I and II values (P = 0.001). Conclusion This study provides novel data for normal reference ranges of STT I and II values in healthy llamas. Results of this study may assist veterinarians in the diagnosis of ocular surface disease and syndromes affecting the tear film in these species.     

Plasma evaluation for ivermectin in llamas (Lama glama) after standard subcutaneous dosing.

Plasma levels of the parasiticide ivermectin were studied by high-performance liquid chromatography in five llamas (Lama glama) after single 200 microg/kg s.c. injections. Ivermectin levels were undetectable in plasma samples drawn up to 4 wk after injection, suggesting that the dosage used was insufficient to reach therapeutic concentrations in this species.