Validation of an immunohistochemical assay for bovine cysticercosis, with comparison to a standard histological method

The larval stage (syn Cysticercus bovis) of the human tapeworm Taenia saginata causes cysticercosis in cattle, which has both aesthetic and food safety implications to consumers of beef. A monoclonal antibody-based immunohistochemical (IHC) assay developed to improve postmortem diagnosis of this parasite and a standard histological method were assessed to determine their fitness for intended use. Sections from 169 known-positive specimens of T. saginata from experimentally or naturally infected cattle, and from 30 known-negative specimens and lesions of various etiologies from non-infected cattle, were tested. The IHC assay identified significantly more known positive bovine cysticerci than the histological method (91.7% and 38.5%, respectively). Positive IHC staining occurred on sections from other cestode species, but should not affect the diagnostic specificity of this assay for bovine cysticercosis, due to the different host and/or tissue preferences amongst these parasites. Use of the IHC assay should improve the reliability of diagnosing lesions caused by degenerated cysticerci, facilitating more effective and efficient control of bovine cysticercosis.     

Evaluation of an enzyme-linked immunosorbent assay (ELISA) for detection of Taenia saginata cysticercosis in cattle.

Serum IgG response of cattle with cysticercosis caused by Taenia saginata was studied in an enzyme-linked immunosorbent assay (ELISA) where a T. saginata metacestode surface extract was used as antigen. In experimentally infected calves, a sharp rise in specific antibody levels was found 3-4 weeks after the infection followed by a logical level of detection corresponded to about 25 cysts. The ELISA was employed in cattle herds where cysticercosis outbreaks had occurred and also in supposedly uninfected herds. Significantly increased antibody levels were found in the herds with massive cysticercosis cases. The test was not adapted for individual diagnosis as some animals of the uninfected herds, especially within the older age groups, had elevated antibody values. The ELISA was, however, useful in the investigation of outbreaks to determine the extent and pattern of the infection in the herd. The rate of decline in antibody levels in these herds was studied by follow up sampling. The increased antibody levels in the infected herds were also reflected in colostrum-fed calves. This observation was employed to estimate the time of infection.     

An outbreak of cysticercosis in feedlot cattle

An outbreak of cysticercosis (infestation with the larvae of Taenia saginata) occurred in feedlot cattle in Ontario in 1986. Two hundred and thirty-three of 271 steers were confirmed histologically to be positive for cysticerci. Nineteen (8.2%) animals had viable cysticerci, 87 (37.3%) had degenerated cysticerci, 77 (33.0%) had mineralized cysticerci, and 50 (21.5%) steers had lymphoid granulomas consistent with cysticercosis. Three viable cysticerci were partly evaginated and one degenerate cysticercus was fully evaginated.     

Evaluation of a 'dipstick' immunoassay to detect cysticercosis in experimentally infected cattle

A 'dipstick' immunoassay for bovine cysticercosis, using an antigen isolated from Taenia hydatigena cyst fluid, was evaluated in cattle experimentally infected with Taenia saginata. The assay correctly identified six out of seven infected cattle, including an animal in which only 12 living cysticerci were found. Cattle became seropositive as early as 3 weeks post-infection. A false-negative reaction was found for one very lightly infected animal, from which only four living cysticerci were recovered at necropsy. The assay was also used to detect circulating antibodies in experimentally infected cattle before and after therapeutic treatment with anthelminthics. The results suggest that praziquantel-treated animals gradually revert to being seronegative after the cysticerci are killed.     

Epidemiological survey of swine cysticercosis in two rural communities of West-Cameroon

To determine the prevalence of porcine cysticercosis, a survey was carried out in 27 villages belonging to two rural communities of West-Cameroon (Bafou and Bamendou). Between January and August, 2000, a total of 707 pigs were examined serologically and by tongue inspection. Serum samples were examined for circulating parasite antigen using a monoclonal antibody-based sandwich enzyme-linked-immunosorbent assay (Ag-ELISA) and for antibodies against cysticerci (Ab-ELISA). Seventy eight samples (11.0%) were found positive in the Ag-ELISA and 154 (21.8%) in the Ab-ELISA, while by tongue inspection on the same animals cysticerci were detected in 43 pigs (6.1%). Gibbs sampling using results of these three tests indicated that the estimated prevalence of porcine cysticercosis was 10.9%. Analysis of the Ag-ELISA results demonstrated that adult pigs showed a significantly higher seroprevalence (15%) than young ones (8.4%). There was no statistical difference in cysticercosis prevalence in pigs raised in households with or without a latrine. Animals that were reported to be usually confined were significantly less infected (9.9%) than free-roaming pigs (16.2%). Infection rates were significantly higher in pigs that had access to human faeces (13.8%) than those which did not have access (9.1%). This study has identified some community behavioural and environmental practices that should be modified to prevent continuous transmission of porcine cysticercosis.     

The incidence of Dictyocaulus viviparus infections in cattle in The Netherlands. I. The Enzyme Linked Immunosorbent Assay as a diagnostic tool

The Enzyme Linked Immunosorbent Assay (ELISA) provides a very efficient technique for detecting antibodies against Dictyocaulus vivparus in calves. Although low level cross reactions were found in animals with gastrointestinal nematodes, the specificity and sensitivity of the technique are sufficient for herd diagnosis of lungworm infections and for survey work. This conclusion is reached on the basis of artificially and naturally infected calves. ELISA titres correlate well with Indirect Haemagglutination titres, parasitological findings, and clinical observations.     

Taenia solium porcine cysticercosis: viability of cysticerci and persistency of antibodies and cysticercal antigens after treatment with oxfendazole

The aim of this study was to assess the effect of treating Taenia solium infected pigs with oxfendazole (OFZ) on viability and clearance of cysticerci and the corresponding persistence of specific antibody isotypes (IgG(total), IgG1, IgG2 and IgA) and circulating cysticercal antigen (CCA). Antibody isotypes and CCA responses were measured by antibody-ELISA (Ab-ELISA) and antigen ELISA (Ag-ELISA), respectively. Correlations were made between antibodies, CCA and the total number of cysticerci enumerated at necropsy. Forty pigs with cysticercosis were randomly allocated into two groups: Treatment group (n=20) was treated with OFZ at 30 mg/kg orally while the treatment control group (n=20) was not treated. Five uninfected pigs served as negative controls. Pigs were killed at 1, 4, 8 and 26 weeks post-treatment (wkpt). Overall, the mean total cyst count in treated pigs was 2904+/-5397 (mean+/-S.D.) while in the controls it was 6235+/-6705. Mean cyst viability was 5+/-11% (mean+/-S.D.) and 97+/-4% in treated and control pigs, respectively. Results showed that OFZ killed muscular cysticerci over a period of 4 weeks but failed to kill cerebral cysticerci. Antibodies, CCA responses and clearance of dead cysts from the meat, depended on the cyst intensity of individual pigs at time of treatment since both antibody and CCA correlated with intensity of cysticerci at necropsy (r=0.441, P=0.005; r=0.654, P<0.001), respectively. IgG1 responses were the best indicator of treatment efficacy because they were predominant in both infected treated and control pigs and disappeared early after treatment. Both Ab/Ag-ELISA failed to detect cysts in the brain. Though dead cysticerci took some time (26 wkpt) to clear from the meat, treatment of porcine cysticercosis with OFZ should, in combination with other intervention measures be considered as an important, cost-effective measure in the control of taeniosis/cysticercosis.     

Sero-epidemiological study of Taenia saginata cysticercosis in Belgian cattle

A sero-epidemiological survey of Taenia saginata cysticercosis was carried out to determine the prevalence of the infection in cattle presented for slaughter in Belgium. Between November 1997 and June 1998, a total of 1164 serum samples were collected in 20 export abattoirs. Meat inspection was routinely carried out by veterinary inspectors. Serum samples were examined for circulating parasite antigen using a monoclonal antibody-based sandwich enzyme-linked-immunosorbent assay (Ag-ELISA). Thirty six serum samples (3.09%) were found positive in the Ag-ELISA, while by meat inspection on the same animals cysticerci were detected in only three carcasses (0.26%). Sero-prevalence was positively correlated with the age of the animals. The sero-prevalence found in this study was more than 10 times higher than the annual prevalence (0.26%) reported by the Institute for Veterinary Inspection. This study clearly indicates that the classical meat inspection techniques detect only a minor fraction of the carcasses infected with cysticerci.     

Risk factors associated with porcine cysticercosis in selected districts of Eastern and Southern provinces of Zambia

To determine the risk factors associated with Taenia solium transmission in humans and pigs in the rural areas of Eastern and Southern provinces of Zambia, a questionnaire was administered in 788 households from 155 villages. Pigs were examined from 800 households. Tongue examination and enzyme-linked immunosorbent assay (Ag-ELISA) for the detection of circulating antigens of T. solium cysticerci were used to measure infection in pigs. A snowballing technique was utilised to select households with pigs. Prevalence of households with pigs infected with T. solium on tongue examination by district ranged from 12.7% to 32.1% with Ag-ELISA having a range of 30.0-51.7%. Of the total number of households visited, 18.8% and 37.6% had at least one pig positive for porcine cysticercosis on tongue examination and Ag-ELISA, respectively. Risk factors associated with T. solium infection were lack of pork inspection at slaughter (96.7%), consumption of pork with cysts (20.1%), selling of pork infected with T. solium cysticerci (18.3%), free-range husbandry system (83.2%) and absence of latrines (58.0). Free-range husbandry system (OR=1.68; 95% CI=1.36-2.07) was a significant risk factor for porcine cysticercosis in the surveyed areas. The result that pigs were mostly kept on free-range and semi-intensive husbandry systems may have permitted them to have access to eating human faeces that could be contaminated with tapeworm eggs. This study has shown that T. solium infection poses a high public health risk in the study areas and urban areas as well. We recommend that a human survey be conducted to verify the human exposure to taeniasis and/or cysticercosis in Zambia.     

Development of a copro-antigen capture ELISA for detecting Ostertagia ostertagi infections in cattle

The present study reports on the development of a copro-antigen capture ELISA for detecting Ostertagia ostertagi infections in cattle. The ELISA was based on polyclonal rabbit antibodies, which recognize O. ostertagi excretory/secretory antigens (ES). ES antigens are released by the metabolic active stages of the parasite in the abomasum, and passed in the faeces of the host. The detection limit of pure ES material was 30 ng ml(-1) in sample buffer and 125 ng ml(-1) in faecal extract. The test was evaluated using a follow up from six artificially infected calves. Elevated levels of Ostertagia coproantigens could be measured from 21 days after infection, indicating that only the presence of adult parasites can be detected. To evaluate the capacity of the assay to measure levels of infection, three groups of cattle were tested: 38 artificially infected calves, 17 naturally infected first grazing season calves and 16 naturally infected adult dairy cows. Optical densities were significantly correlated to the worm burdens of the animals and the ELISA had an overall sensitivity of 91% and a specificity of 45%. The test gave negative readings for faeces of animals carrying patent mono-infections with Cooperia oncophora.     

The incidence of Dictyocaulus viviparus infections in cattle in the Netherlands

Summary

The Enzyme Linked Immunosorbent Assay (ELISA) provides a very efficient technique for detecting antibodies against Dictyocaulus vivparus in calves. Although low level cross reactions were found in animals with gastrointestinal nematodes, the specificity and sensitivity of the technique are sufficient for herd diagnosis of lungworm infections and for survey work. This conclusion is reached on the basis of artificially and naturally infected calves. ELISA titres correlate well with Indirect Haemagglutination titres, parasitological findings, and clinical observations.     

Performance of light vs heavy steers grazing Plains Old World bluestem at three stocking rates

Live weight gains of light and heavy calves grazing Plains Old World bluestem at three stocking rates were evaluated during the summers of 1997 and 1998. Initial weights of mixed-breed light-weight steers (LHT) were 141 SD = 17 kg (n = 214) in 1997 and 160 SD = 23 kg (n = 193) in 1998. Initial weights of mixed-breed heavy steers (HWT) were 265 SD = 17 kg (n = 115) in 1997 and 248 SD = 13 kg (n = 126) in 1998. Initial stocking rates for both sizes of steers were as follows: light, 392 kg of live weight/ha; moderate, 504 kg of live weight/ha (increased to 616 kg live weight/ha in 1998); and heavy, 840 kg of live weight/ ha. Averaged gain and gain/hectare are reported as stocking rate by steer type within year. Heavy steers had greater ADG than LHT steers during both years. Forage intake, expressed as a percentage of BW, was greater (P = 0.05) for LHT (3.1%) than for HWT (2.8%) calves. Grazing time (min/d; 1998 only) was greater (P = 0.05) for LHT (665) than for HWT (624) steers. Forage CP and in vivo digestible organic matter (DOM) were slightly greater (P < 0.05) in pastures grazed by HWT vs LHT cattle. Gain/hectare was greater (P < 0.05) for LHT than for HWT calves at all three stocking rates during both years. A linear decline in ADG was observed (P < 0.07) as stocking rates increased for HWT steers in 1997 and LHT steers in 1998. However, ADG did not decline with increasing stocking rate for LHT calves during 1997 or HWT calves during 1998. Forage intake was not different among stocking rates in either 1997 or 1998. Grazing time was greatest (P < 0.05) for steers in the moderate and heavy stocking rates. Forage in vivo DOM decreased (P < 0.05) as stocking rate increased. Both LHT and HWT steers had lower (P < 0.05) ADG at all three stocking rates during 1998 compared with 1997. Despite lower ADG, LHT steers had greater gain/hectare than HWT steers during both 1997 and 1998.     

Evaluation of diagnostic tests for Johne's disease in young cattle

OBJECTIVE: To investigate the development of immune responses in calves experimentally and naturally infected with Mycobacterium paratuberculosis and to evaluate the potential for diagnostic tests to detect infected calves. DESIGN: Sequential testing of four treatment groups of calves over a 2 year period. PROCEDURE: Twenty-nine calves were allocated to four groups. Group D calves were orally dosed with M paratuberculosis, group N calves naturally exposed to M paratuberculosis, group V calves vaccinated for M paratuberculosis, and group C were control calves (not infected or vaccinated). Blood and faecal specimens were collected from each calf at monthly intervals to 18 months of age and then every 2 months until they were slaughtered between the ages of 21 and 29 months. Specimens were tested using absorbed EIA, IFN-gamma EIA and faecal culture. The infection status of the calves was confirmed by extensive histopathological examination and tissue culture. RESULTS: M paratuberculosis infection was confirmed in 10 calves, comprising six of eight orally dosed calves, three of five naturally exposed calves and one of nine vaccinated calves. The six artificially infected calves and one naturally infected calf were detected shedding M paratuberculosis in their faeces. Results with positive absorbed EIA were obtained from one artificially infected calf, one naturally infected calf and three vaccinated calves. All calves including controls had positive results on at least one occasion using the IFN-gamma EIA. In addition, seven calves had positive bovine tuberculosis results using the IFN-gamma EIA, even though bovine tuberculosis has been eradicated from Australia. CONCLUSION: Detection of M paratuberculosis infection in young cattle continues to be difficult using current tests.     

Development and preliminary assessment of a polyclonal antibody-based enzyme immunoassay for the detection of Tritrichomonas foetus antigen in breeding cattle

More sensitive tests are required for the diagnosis of Tritrichomonas foetus infection in cattle and an antigen-detecting enzyme immunoassay (EIA) has been applied to this purpose. An affinity purified immunoglobulin fraction obtained from rabbits immunised with cultured T. foetus served as both capture antibody and as biotinylated indicator antibody. While highly sensitive in the detection of antigen derived from cultured organisms, the assay showed poor sensitivity in the detection of antigen in the cervico-vaginal mucus of artificially infected heifers, with only 75% of culture-positive samples being considered positive for antigen. In a direct comparison, 23/122 samples from a naturally infected dairy herd gave positive cultures, while only 10/122 samples were considered antigen positive by EIA.     

The prevalence of porcine cysticercosis in Eastern and Southern provinces of Zambia

The objective of this study was to determine the prevalence and importance of porcine cysticercosis in rural areas of Zambia. The study involved an abattoir survey of 1316 pigs at a slaughter slab in Lusaka and two field surveys in villages in Southern and Eastern provinces. Lingual examination of live pigs and visual inspection of their carcass as well as blood sampling for measuring circulating parasite antigen by enzyme-linked immunosorbent assay (Ag-ELISA) were used as parameters to measure infection. In the field surveys, a questionnaire was administered to every household whose pigs were examined to obtain information on pig husbandry practices and to study risk factors for the infection. Out of the 1316 pigs examined at the slaughter slab, 143 (10.9%) and 271 (20.6%) were positive by lingual examination and meat inspection, respectively. Most of the pigs were very heavily infected with predominantly live cysts. The field surveys revealed that eight (8.2%) out of 98 pigs from Southern province and eight (5.2%) out of 151 pigs from Eastern province were positive for cysticercosis by tongue palpation. Using the Ag-ELISA 20 (20.8%) and 14 (9.3%) pigs were positive in Southern and Eastern provinces, respectively. The questionnaire survey revealed poor pig husbandry practices, absence of meat inspection and control, poor knowledge of the disease and poor sanitation in the surveyed villages. The prevalence of pig cysticercosis found in this study ranks among the highest in the southern African region, in Africa and in the world. The current study suggests the presence of human tapeworm carriers and a high risk of human cysticercosis in the surveyed areas as well as in urban centres where pigs from rural areas are increasingly sold, slaughtered and consumed.     

Attempted infection of calves with Taenia crocutae cysticerci and their subsequent serological response

The susceptibility of naive cattle to infection with cysticerci of Taenia crocutae was tested using three six- to nine-month-old Ayrshire bull calves, previously unexposed to infection with taeniid eggs. One calf was given 10,000 T crocutae eggs orally, another 5000 hatched unactivated oncospheres orally and the third 5000 hatched and activated oncospheres by intravenous injection. None of the calves contained viable cysticerci at post mortem examination 15 to 17 weeks later. All three calves contained small numbers of lesions in the liver and lesions were also present in the lungs of the calf which received oncospheres intravenously. All the calves developed an antibody response which was most pronounced in the calf given hatched unactivated oncospheres orally.     

Investigation of an outbreak of mucosal disease in a beef cattle herd in southwestern Saskatchewan.

This study describes the epidemiological investigation of an outbreak of mucosal disease that occurred on a ranch in southwestern Saskatchewan. Over a six-month period during the fall and winter of 1991-1992, in a herd of 515 beef cattle and 96 bison, 20 yearling cattle from a group of 105 housed in one feedlot pen died from mucosal disease. A further eight yearlings were slaughtered for salvage because they were at risk of dying from mucosal disease. Mucosal disease mortalities were the first observed evidence of fetal infections with bovine viral diarrhea virus in this herd. Animals that died from mucosal disease exhibited signs of ill thrift prior to death. Deaths from mucosal disease were confined to the progeny of one herd of beef cows. Following an outbreak of fetal infection with bovine viral diarrhea virus during 1989-1990, at least 28 (22%) of the 128 calves born from this herd of cows in the spring of 1990 were persistently infected with bovine viral diarrhea virus. However, only one calf born from this herd in 1991, and five calves born from all herds in 1992 were persistently infected. Of the five persistently infected calves born in 1992, three were born to persistently infected replacement heifers born in 1990. These heifers calved without assistance in 1992, but only one of their calves survived past three days of age, and it was persistently infected. In January 1992, 82% of the total herd had reciprocal antibody titers to bovine viral diarrhea virus of > or = 1024 which suggested a high level of herd immunity to bovine viral diarrhea virus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Detection of calves persistently infected with bovine pestivirus in a sample of dairy calves in south-eastern Queensland

Objective To determine the proportion and incidence of calves persistently infected with bovine pestivirus in calves (n = 1521) supplied to the Tick Fever Research Centre and to assess the test regime to detect calves persistently infected with bovine pestivirus.
Design Calves, 1 to 6 weeks old, selected for use in the production of the tick fever vaccine were collected from 21 properties in 56 separate groups between October 1990 and December 1996. Each group was examined for the presence of calves persistently infected with bovine pestivirus.
Procedure All calves were routinely tested for antibody to bovine pestivirus and bovine pestivirus antigen using a serum neutralisation test and an antigen-capture ELISA, respectively. Pooled lymphocyte samples from calves were also monitored for bovine pestivirus by inoculation of sheep. Whole herd testing was carried out in eight herds, using a serum neutralisation test as a screen test followed by an antigen-capture ELISA of cattle with a serum neutralisation test titre of less than 32.
Results Fourteen of the 1521 calves tested (0.9%), were detected as persistently infected and the incidence ranged from 0.0 to 3.0 % per year over 6 years. Persistently infected calves were found in 13 of the 59 groups and originated from 7 of the 21 herds used. In whole herd testing on the properties of origin, cattle persistently infected with bovine pestivirus were detected in four of the eight herds tested
Conclusions The proportion of calves persistently infected with bovine pestivirus is similar to that in other countries and indicates that bovine pestivirus could be a significant cause of economic loss in Australian cattle herds. In detecting calves persistently infected with bovine pestivirus, the combination of sheep inoculation, paired antigen-capture ELISA and serum neutralisation tests appeared to be highly sensitive and specific.     

猪囊虫病基因工程疫苗用于免疫治疗的研究

应用猪囊虫病基因工程疫苗分别对实验性猪囊虫病和自然感染的猪囊虫病进行了分组治疗,以探讨该疫苗的免疫治疗作用。１０头人工复制的囊虫病猪,随机分成３组。第１组４头猪,于感染于１个月注射该疫苗,连续３次,间隔１５天;第２组４头猪,于感染后２个月注射疫苗,同上连续３次;第３组２头猪,不注苗。第１组注射疫苗后１个月循环抗原滴度开始降低,２．０、２．５及３．５个月各有１头猪ＣＡ转阴,５个月ＯＤ值分别为０．１４、０．０９、０．１７、０．３８。第２组和第３组各猪血清ＯＤ值一直维持在１．０以上。感染后５个月剖检,第１组４头猪均未检出囊虫;第２组４头猪在咬肌、膈肌、腰肌、股内侧肌、肩胛外侧肌等部位均检测到囊虫,检测部位４０ｃｍ＾２面积发现囊虫３～７个,有部分虫体已钙化,胆汁卵化率为１５．４％;第３组各猪也在各检测部位检测到囊虫,４     

Factors affecting the natural transmission of bovine leukaemia virus infection in Queensland dairy herds

Natural transmission of bovine leukaemia virus (BLV) infection in south-eastern Queensland dairy herds was slow in 2 herds with a low to moderate (13 to 22%) prevalence of infection. Infection spread much more rapidly in a herd that had a higher prevalence (42%) when first tested. In a 13 month study of this herd, the cumulative incidence of infection was 24%. In one herd new infections were confined almost entirely to calves of uninfected dams. Following the end of feeding bulk milk to calves, a common practice in dairy herds, no more calves in this herd became infected. In laboratory experiments, neither prolonged housing of susceptible calves with infected cattle, consumption of drinking water contaminated with infected blood, nor inoculation of sheep with saliva from infected cattle resulted in transmission of BLV infection. Sheep were infected by subcutaneous inoculation of a suspension of purified lymphocytes from an infected heifer. The minimum infective dose was 10(3) lymphocytes, equivalent to the number of lymphocytes in approximately 0.1 microliter blood. Thus, procedures involving the transfer of a very small volume of blood from animal-to-animal have the potential to transmit infection.