Comparison of various reproductive status in Sika deer (Cervus nippon) using fecal steroid analysis

The feasibility of fecal steroid analysis for pregnancy diagnosis and sex determination were tested in sika deer (Cervus nippon). Feces were collected from captive sika deer in June (non-breeding season and late-pregnancy period) and October (breeding season), and also from the rectum of 24 female sika deer (19 pregnant and 5 non-pregnant females) shot as part of programs for population control in February and March (mid-pregnancy period). In mid- and late-pregnancy periods, fecal progesterone concentrations were significantly higher in pregnant female than in male and non-pregnant female deer. In October, fecal testosterone concentrations were higher in adult male deer, and no difference was found between young males and females. These results suggest that fecal steroid analysis would be a useful means for estimating pregnancy status and for detecting adult male among wild deer.     

Comparison of different commercial DNA extraction kits to detect Toxoplasma gondii oocysts in cat faeces

The cat is the definitive host of Toxoplasma gondii and plays an important role in the transmission of this and other coccidian parasites, e.g. Hammondia hammondi, a protozoon closely related and morphologically similar to T. gondii. A number of techniques to detect T. gondii nucleic acids in feline faeces are described and several extraction kits for isolating pathogen DNA from faeces or soil are commercially available. To compare the performance of such kits with regard to isolating oocyst DNA, a feline sample that had tested negative for coccidian parasites including T. gondii and H. hammondi was spiked with 10(4), 10(3), 10(2), 50 and 10 H. hammondi oocysts. Several ready-to-use stool or soil kits and an in-house method were then used to extract parasite DNA from these spiked faecal samples. Of six kits tested, two were found suitable for the detection of H. hammondi oocysts DNA by the polymerase chain reaction (PCR) in faecal samples with a detection limit of 250 oocysts per 1 g of faecal sample. These two kits revealed a similar, even slightly lower detection limit (50 oocysts per 1 g of sample) when tested with faecal samples spiked with T gondii oocysts.     

Fecal progestagens to detect and monitor pregnancy in captive female cheetahs (Acinonyx jubatus)

The purposes of the present study were to establish a noninvasive monitoring assay of fecal progestagen measurement to detect pregnancy and to identify the components of fecal progestagens in early, middle and late pregnancy in cheetahs. Feces were collected from 7 female cheetahs and analyzed from 30 days before the last copulation to parturition in 9 pregnancies. Blood was collected from one cheetah. Fecal progestagen and serum progesterone concentrations were determined by enzyme immunoassay (EIA). The profiles of the fecal progestagen concentrations were similar to the serum progesterone profile. Fecal progestagen and serum progesterone concentrations remained at the baseline until copulation. In the mean fecal progestagen profile during pregnancy (92.8 ± 0.4 days; from the last copulation to parturition), the concentrations increased 3-4 days after the last copulation and remained high until parturition. To investigate changes in the components of progestagen metabolites in the tripartite periods of gestation, fecal progestagens were analyzed by HPLC-EIA. Marked immunoreactive peaks consistent with 5α-pregnan-3α/β-ol-20-one and 5α-pregnan-3,20-dione and small peaks consistent with 5β-pregnan-3α/β-ol-20-one were detected. There were no distinct difference in the components of progestagens among the first, second and third trimesters of pregnancy. The hormone assay, as an indicator of fecal 5α-reduced pregnanes, is useful for detecting pregnancy and monitoring pregnant luteal activity in cheetahs.     

Development and evaluation of an indirect enzyme immunoassay for detection of porcine antibodies to pseudorabies virus.

An indirect enzyme immunoassay is described for detection of porcine serum antibody to pseudorabies virus. The analytical sensitivity of the enzyme immunoassay was found to be approximately 4.5 log 4 X 10 (5120 times) greater than the serum neutralization test, based on parallel end point titrations. The diagnostic sensitivity of the enzyme immunoassay was comparable or superior to that of the serum neutralization, based on the earliest detectable antibody after infection of swine with pseudorabies virus by intranasal or intrauterine routes or by contact with infected pigs. The enzyme immunoassay, at a screening dilution of 1:20, gave 100% agreement with ELISA results provided with a U.S. Department of Agriculture-Animal and Plant Health Inspection Service proficiency panel of 40 sera. One serum having demonstrable antibody by the enzyme immunoassay was seronegative by the serum neutralization test.     

A competitive enzyme immunoassay for the detection of bovine antibodies to Brucella abortus using monoclonal antibodies

A competitive enzyme immunoassay (EIA) for the detection of circulating bovine antibodies to Brucella abortus has been developed using horseradish peroxidase conjugated monoclonal antibodies (MAb) raised against B. abortus cell surface antigens. Antibodies present in the serum of either vaccinated or infected cattle can apparently displace the conjugated MAb from the lipopolysaccharide antigen (LPS) in a quantitatively different manner allowing an assessment of immune status of the animal. The results from a panel of sera from animals with a known status of vaccination or infection indicated that the test was more selective in the detection and discrimination of infected from uninfected or immunized animals, than conventional complement fixation, agglutination or indirect enzyme immunoassay procedures.     

Serologic detection of antibodies to Newcastle disease virus in water fowl using the hemagglutination inhibition test and enzyme immunoassay]

8410 samples from Moscovy duck, Pekin duck and geese were incorporated into examinations of antibodies against the Newcastle Disease virus. A new enzyme-immuno-assay (EIA) for antibody detection in Moscovy duck and Pekin duck was developed using purified antigen from NDV-strain "La Sota". The epidemiology as well as the relation of incidence of the Newcastle Disease in waterfowl was discussed.     

Assessment of pregnancy in sheep by analysis of plasma progesterone using an amplified enzyme immunoassay technique

An amplified enzyme immunoassay kit for progesterone analysis was used to diagnose pregnancy in a flock of 130 mule ewes. An accuracy of 100 per cent was obtained after the analysis of progesterone in plasma samples taken 15 to 16 days after mating. In mule ewes a plasma progesterone level greater than 5.9 nmol/litre was indicative of pregnancy. In the validation of the technique, duplicate ewe plasma samples and progesterone standards were compared with a radio immunoassay technique; the regression coefficient between the two techniques was r = 0.82.     

Development and evaluation of a rapid absorbed enzyme immunoassay test for the diagnosis of Johne''s disease in cattle

An absorbed enzyme immunoassay (EIA) test for Johne's disease in cattle was developed in which absorption of cross-reacting antibodies occurred as a rapid reaction in solution rather than overnight with whole organisms and a subsequent centrifugation step. Total test time was reduced to less than 2 h with a minimum of manipulations. The test was evaluated in cattle herds from Johne's disease-endemic and Johne's disease-free regions of Australia. Specificity was 99.8%. Calculations of sensitivity were affected by the history of the herd under test. However, the EIA detected in excess of 80% of animals before onset of clinical disease and 65% of faecal shedders were EIA positive on, or before, first detection of Mycobacterium paratuberculosis in their faeces. The test should aid epidemiological studies and be a useful tool in the management and control of Johne's disease.     

A review of enzyme immunoassay for detection of antibody to Brucella abortus in cattle

Enzyme immunoassay has gained wide acceptance for serological diagnosis of bovine brucellosis because of its ability to detect antibody of all isotypes unlike the conventional tests. The indirect enzyme immunoassay, however, presents several parameters that require careful analysis. These parameters include the choice of antigen and antiglobulin-enzyme conjugate reagents for use in the assay, dealing with the large amount of data the semi-automatic or automatic assay can generate and the inter- and intralaboratory standardization and quality control. This review considers the various methods described in the literature and, briefly, how some of the problems have been overcome or how they might be dealt with.     

Use of a new and rapid milk progesterone assay to monitor reproductive activity in the cow

A new and rapid enzyme-amplified immunoassay (AELIA) has been developed for the measurement of progesterone in milk. The AELIA system is a non-isotopic method that gives results within 35 minutes. Milk progesterone concentrations measured in 10 cows sampled daily at various stages of the reproductive cycle were very similar to those recorded by a validated radioimmunoassay. The results show that the speed and sensitivity of the AELIA system would make it possible to diagnose pregnancy rapidly at about 24 days after insemination, to predict the onset of behavioural oestrus from decreasing progesterone values during the third week after a preceding oestrus, and to obtain a daily record of milk progesterone levels in animals treated for infertility of ovarian origin.     

The development of a simple fecal immunoreactive progestin assay to monitor reproductive function in swine.

The objectives of the study were to (a) develop a simple fecal progestin extraction and radioimmunoassay method to measure immunoreactive progestin in porcine feces and (b) to characterize fecal progestin profiles during the estrous cycle and postpartum. A simple extraction method was developed in trial 1 and the mean (+/- SD) progestin recovery of the method was 84.3 +/- 3.5%. Progesterone levels measured at five different spiked concentrations (50, 100, 200, 400, and 500 ng/0.5 g feces) showed no systematic error. The sensitivity of the assay was 0.16 nmol/L of the extract. Trial 2 involved collecting fecal samples from six cycling sows every second or third day, beginning on the day of estrus (day 0) and continuing until day 22. The mean (+/- SD) fecal progestin concentrations of these sows determined by the above assay during days 0-5, days 6-10, days 11-15, and days 16-21 were 87.1 +/- 17.5, 262.6 +/- 102.1, 1188.2 +/- 454.1, and 897.3 +/- 274.1 x 10(-3) nmol/g feces, respectively. In trial 3, fecal samples from six postpartum sows were collected at weekly intervals beginning from day 7 after farrowing until day 50. The mean (+/- SD) fecal progestin concentrations were 111.0 +/- 61.1, 74.1 +/- 21.3, 66.5 +/- 26.1, 122.7 +/- 58.8 and 533.5 +/- 244.2 x 10(-3) nmol/g feces, during days 7-10, days 11-20, days 21-30, days 31-40, and days 41-50 postpartum, respectively. The results indicate that simple fecal progestin extraction and assay are feasible alternatives to the standard blood progesterone assays for monitoring reproductive function in swine.     

Use of an enzyme immunoassay to determine concentrations of progesterone in caprine plasma and milk

Concentrations of progesterone (P4) were determined using enzyme immunoassay kits on plasma and milk samples obtained on the same days from 18 lactating dairy goats. Progesterone profiles documenting anestrus, short estrous cycles, normal estrous cycles, a prolonged follicular phase, and prolonged luteal phases were established. When plasma P4 concentrations were used as an accurate indication of the presence or absence of functional luteal tissue, milk P4 concentrations agreed with plasma determinations in 79.4% of the 465 samples tested. Milk samples could not be used to make a definitive decision because of marginal values in 11.2% of the determinations. Milk P4 concentrations were high when plasma P4 concentrations were low in 6.2% of the paired samples, especially those obtained around the time of estrus when peripheral P4 concentrations were changing rapidly. The remaining 3.2% of milk samples had low milk P4 concentrations when plasma P4 concentrations were high. Composite milk from 8 does in estrus or 8 does in the luteal phase was not consistently different from strippings in butterfat percentage or P4 concentration.     

A microplate enzyme immunoassay for detecting and measuring antibodies to Babesia bovis in cattle serum

A microplate enzyme immunoassay (EIA) is described for measuring IgG antibody to Babesia bovis in cattle serum. B. Bovis antibody status (whether positive or negative) and the amount of B. Bovis antibody (EIA score), were measured by comparison with reference serums. The EIA was shown to be specific for B. Bovis, and EIA score correlated well with EIA titre. Comparison of EIA with the Indirect Fluorescent Antibody Test (IFAT) showed more than 95% agreement between the methods and disagreement in only 1.6% of serum samples tested. The remaining 3.2% were positive by EIA and suspected positive by IFAT. The EIA was shown, by titrating positive serums, to be more sensitive than IFAT, which explained its tendency to detect more positive serums than IFAT. EIA detected B. bovis antibody in experimentally infected cattle by day 14 post infection (pi) and for at least 268 days pi. EIA score for B. bovis antibody in immune cattle increased significantly (p less than 0.05) following heterologous strain challenge.     

The use of milk progesterone analysis to monitor reproductive activity in the camel (Camelus dromedarius)

Milk progesterone profiles were used to monitor the ovarian changes during the post-partum period of five she-camels. Milk samples were collected daily for 60 days from four suckling camels (1-4) and from one animal whose calf died on day 3. Progesterone was determined using the Ovucheck Bovine progesterone kits. The first increase of progesterone level in the five animals occurred on days 28, 26, 21, 24 and 20 respectively. At least two progesterone peaks of 6 +/- 2 SD days' duration occurred during the 60 days. An early one day peak was also noticed in three animals.