Comparison of the Q-fever complement fixation test and two commercial enzyme-linked immunosorbent assays for the detection of serum antibodies against Coxiella burnetti (Q-fever) in ruminants : recommendations for use of serological tests on imported animals in New Zealand.

AIM: To make valid recommendations on the use of serological test methods for the detection of serum antibodies in ruminants against Coxiella burnetii (Q-fever), by comparing the performance of the complement fixation test (CFT) and two ELISA, and by identifying reasons for discrepancies between the test methods. METHODS: A total of 73 serum samples from infected cattle, 69 from infected goats, and 100 samples from non-infected cattle and 57 samples from non-infected sheep, as well as 95 samples from infected cattle herds (mix of seropositive and seronegative samples), were tested using the CFT, the IDEXX ELISA (I-ELISA) and the Pourquier ELISA (P-ELISA). A mixed panel of 12 serum samples from sheep from inter-laboratory proficiency testing (proficiency panel) was also tested using the CFT and both ELISA, and further investigated using IgG- and IgM-specific ELISA. RESULTS: Generally, the two commercial ELISA were more sensitive than the CFT for the detection of infected ruminants. Good agreement between ELISA for positive and negative results was found for samples from the infected herd, while results for the positive panels varied between the two ELISA. For the total of the positive serum panels, the I-ELISA detected 95% of samples as positive or suspicious, while the P-ELISA detected only 81%. In the P-ELISA, more samples were considered suspicious (18%) than in the I-ELISA (14%). All sera from non-infected sheep and cattle tested negative in the serological test methods employed, except for one positive sample from a sheep in the P-ELISA. Further investigation revealed that a CFT-positive but ELISA-negative result was due to high IgM and low IgG reactivity. CONCLUSIONS: The two commercial ELISA were more sensitive than the CFT in all panels from infected ruminants. However, they could only detect IgG. The I-ELISA should be the serological test method of choice for cattle, sheep and goats for import testing of animals into New Zealand because it was more sensitive than the P-ELISA and was equally specific to the PELISA and the CFT. For other animal species, such as deer and camelids, the CFT should still be used since none of the ELISA has been evaluated for these species. This study has shown that the two commercial ELISA will detect the majority of infected ruminants but may miss animals that have not developed an IgG response.

Rabies virus typing--preliminary survey in Botswana.

A survey was made of the subtypes of rabies isolates made in Botswana using monoclonal antibodies. Two subtypes were detected, termed canine and mongoose. The canine subtype predominated in the north and west and appeared to be related to the distribution of both the domestic dog and wild jackal. The mongoose subtype was found in the south-east and was associated with feline and viverrid wildlife. The geographical distribution also matched reports of isolates examined in the neighbouring countries.

Response of cattle persistently infected with bovine virus diarrhoea virus to bovine leukosis virus.

Six cattle persistently infected with bovine virus diarrhoea virus (BVDV) and seronegative, and two control, virus negative seropositive cattle were inoculated with lymphocytes infected with bovine leukosis virus (BLV). The two controls produced a normal immune response to BLV, developing antibodies at four and five weeks after inoculation. Two of the six cattle persistently infected with BVDV developed a strong antibody response by six weeks after inoculation with BLV. Four developed a depressed response to BLV, characterised in three by a 'hooking' reaction in the immunodiffusion test which persisted in successive bleedings but was interspersed occasionally by a weak positive reaction. In one of these animals, a series of 'hooking' reactions was followed by a number of negative results. The fourth animal remained serologically negative until 16 weeks after inoculation when a 'hooking' reaction was observed followed by a series of negative results. BLV was isolated from all the cattle persistently infected with BVDV at 42 or 58 weeks after inoculation regardless of whether the serum samples gave negative, 'hooking', weak positive or positive reactions in the immunodiffusion test. BLV was consistently isolated from the nasal secretions of a steer which was BVDV negative but seropositive. The possibility of decreased immune responsiveness to BLV in animals persistently infected with BVDV should be considered when formulating regulations governing the testing of animals for freedom from BLV.

Outbreaks of classical swine influenza in pigs in England in 1986.

Serum samples from pig herds in Great Britain have been examined for antibodies to influenza virus since 1968. Antibodies to H3N2 virus strains have been found since 1968 and the serological data presented here suggests that H3N2 virus strains continue to persist in the pig population. An outbreak of acute respiratory disease occurred in a 400-sow unit. The outbreak was characterised by coughing, anorexia, fever, inappetence and loss of condition. The gilts and weaners were affected and the morbidity approached 100 per cent. An influenza A virus designated A/Swine/Weybridge/117316/86 (H1N1) was isolated from the herd and 28 paired serum samples from the affected animals showed increases in the haemagglutination inhibition titres to this isolate. Haemagglutinin and neuraminidase characterisation indicated that the virus is similar to H1N1 viruses isolated recently from pigs in Europe. A total of 91 herds experiencing respiratory disease were investigated, of which 42 gave positive reactions in the haemagglutination inhibition test. Antibodies to A/Port Chalmers/1/73 (H3N2) were also detected in some of the herds but it is not known whether this strain plays any role in the current respiratory disease problems in pigs.

To compare the incubation period following intratracheal and subcutaneous inoculation of bovine leukosis virus infected lymphocytes and to study their clearance from the circulation.

The migration of fluorescein isothiocyanate labelled lymphocytes through the tracheobronchial mucosa has been studied in cattle. Following intratracheal inoculation of labelled non-infected autologous lymphocytes and bovine leukosis virus (BLV) infected heterologous (presumed allogeneic) lymphocytes, the labelled lymphocytes appeared in the blood circulation between 4 and 7 days post inoculation. Following intravenous inoculation of labelled autologous lymphocytes, the cells could be detected in the circulation for 10 days post inoculation whereas BLV infected and non-infected heterologous lymphocytes could be detected for only 2 days. The migration of BLV-infected heterologous lymphocytes through the tracheobronchial mucosa caused a delay in the appearance of labelled lymphocytes in the circulation and a corresponding delay in the appearance of BLV antibodies. Comparison was made of the effect of two different routes of inoculation, subcutaneous and intratracheal on the incubation period as indicated by the detection of antibody. Subcutaneous inoculation of 1 X 10(4), 5 X 10(3), 1 X 10(3) of lymphocytes from a BLV infected cow caused seroconversion whereas 5 X 10(2) cells did not. Intratracheal inoculation of 5 X 10(3) cells caused sero-conversion. One animal did not develop BLV antibody until 30 weeks after inoculation although BLV could be isolated from the blood at 24 and 26 weeks post inoculation.

Spread of bovine syncytial virus in a dairy herd over a two year period.

During a two year period the spread of bovine syncytial virus was monitored in a closed herd of 50 to 100 milking cows. Out of a nucleus of 49 nonpregnant and pregnant heifers, six were found to be infected with bovine syncytial virus. Virus was detected only in the progeny of infected cows and not in the progeny of uninfected animals. Nineteen progeny of the bovine syncytial virus infected cows were studied in detail and virus was isolated from only four. Horizontal spread of the virus did not occur.

An attempt to protect calves against experimental bovine leukosis using allografts.

Six calves sensitised by implanting skin from a calf were later inoculated with lymphocytes from the same calf after the calf had been infected with bovine leukosis virus (BLV). Two out of 6 calves challenged did not develop BLV antibodies and BLV was not isolated from these animals, whereas all of the 5 control calves became infected with BLV.

Infectivity of enzootic bovine leukosis infected animals during the incubation period.

Steers and calves were experimentally infected with bovine leukosis virus. The virus was isolated from the blood and from the tracheal and bronchoalveolar washings before antibodies could be detected in the serum. Bovine leukosis virus was not detected during any period in the blood plasma.

Detection of bovine leukosis virus in bronchoalveolar lung washings and nasal secretions.

Cattle and sheep persistently infected with bovine leukosis virus (BLV) were studied for the presence of the virus in bronchoalveolar lung washings and nasal secretions. The virus was demonstrated in the cellular fraction of the lung washings in six out of nine cattle and in one out of six sheep. In no instance was bovine leukosis isolated from the cell-free bronchoalveolar lung washings. The virus was isolated from the nasal secretion of only one of six naturally infected milking cows despite frequent sampling; the virus-infected nasal secretion was from a sick 10-year-old cow. Bovine leukosis virus was not isolated from cellular fractions of nasal secretions.

Protection against bovine leukosis virus infection in sheep with the BL 20 bovine lymphoblastoid cell line.

The bovine lymphoblastoid BL 20 cell line derived from a case of sporadic bovine leukosis when inoculated into sheep did not induce an antibody response directed against bovine leukosis virus (BLV) structural proteins. Sheep were inoculated twice with the BL 20 cell line and then challenged with BLV infected lymphocytes. Three out of four sheep challenged four weeks after BL 20 inoculation did not develop BLV antibodies. Of the 12 sheep challenged later, three sheep did not develop BLV antibodies. BLV was isolated from all the seropositive animals and from none of the seronegative animals.

An investigation into the susceptibility of cattle to bovine leukosis virus following inoculation by various routes.

The effect of different routes of inoculation on the incubation period, as indicated by the detection of antibody and by the detection of bovine leukosis virus (BLV) in lymphocytes, were compared. None of the 12-month-old steers exposed to BLV by the oral route developed BLV infection. Intratracheal, subcutaneous and intradermal inoculations were found to be particularly effective in establishing BLV infection, which was detected three to four weeks after inoculation. In the majority of animals, serum antibody and virus were detected at the same time. One out of four in-oestrus heifers inoculated via the uterus with mixtures of BLV infected lymphocytes and semen became infected. It appears that there is an inhibitory factor in fresh semen that prevents BLV infection from becoming established. Viral antigen was detected earlier in BLV infected lymphocytes using the cocultivation method than by electron microscopy to demonstrate BLV particles in mitogen stimulated lymphocytes.