With the introduction of African swine fever virus into China, the virus has diversified, and the proportion of low-virulence strains has been rising, and it is difficult to detect early by clinical symptoms alone. Biosecurity measures, combined with pathogen detection and antibody monitoring, are the most effective means to prevent and control non-plague.
It is necessary to clarify the window period for non-plague virus pathogen detection and antibody detection for the prevention and control of non-plague virus.
Jutta Pikalo, a scientist from Germany, used a variety of methods to diagnose experimental animals infected with African swine fever, and it is not difficult to find from the results of the study that there is an obvious gap between the three methods of clinical symptom diagnosis, pathogen detection and antibody detection in the early warning of African swine fever. The results of the study have been Xi and shared as follows:
1. Experimental protocol
2. Clinical symptoms:
All animals from post-inoculationDay 4Non-specific clinical symptoms such as depression, anorexia, reluctance to stand, decreased mobility, tremors, redness, cyanosis of the ears and nose, hunched back, dyspnea begin;
After infectionDay 4Fever began, peaking on days 8-9 post-infection, and peak on days 7-8 post-infection (Figure 1), but no elevation in body temperature was observed in animal No. 3.
Fig.1 Body temperature curve of experimental animals3. Pathogen detection
After infection3 daysNon-pestiflor virus was detected in 3 pigs, and non-pestivirus was detected in the others 7 days after infection, but the pigs with ear tag 3 were detected with non-pestiflor pathogens on the 10th day after infection;
On days 7-14 after infection, the viral load reaches its maximum of 129×10 3-6.8 10 4 copies of the reaction (see table for results).
Table 1 Real-time PCR results of animal blood from 0 to 18dpi after infection.
Note: ND indicates undetected, NA indicates no pathogen of interest, numerical value indicates the number of genomic copies detected per PCR reaction (5 L), and color intensity indicates the relative level of viral load (darker colors indicate higher viral load, from pale yellow to dark red).
Table 2 Real-time PCR results of blood, serum, and tissue samples of experimental animals on the day of euthanasia.
Note: ND indicates undetected, the numeric value indicates the number of genomic copies detected per PCR reaction (5 L), and the color intensity indicates the relative level of viral load (darker colors indicate higher viral load, from pale yellow to dark red).
4. Antibody testing
Ingenasa ELISA kit (detection of p72 antibody): post-infectionDay 10, 2 pigs turned positive for antibodies, and 3 pigs were suspicious (Figure 2);
IDVET competes for ELISA kit (detection of p32 antibody): all antibodies turned positive 10 days after infection, and one of them had suspicious antibodies on the 7th day after infection;
IDVET indirect ELISA kit: On day 10 post-infection, 3 were suspicious, 1 was negative, and 1 was positive. On the 14th day after infection, all the antibodies of the suspected pigs and the negative pigs turned positive
Indirect immune horseradish peroxidase assay: post-infectionDay 7, the serum semi-quantitative results shifted, and the antibody titers of the surviving animals ranged from 2560 to 5120 on day 18 post-infection (semi-quantitative results).
Fig.2 Detection of seroconversion in experimental animals using Ingenasa ELISA products Note: The cutoff value of ELISA is 50%, which is represented by a dotted line. Summary:
Experimental animals showed clinical symptoms on day 4, pathogens were detectable on day 3 of qPCR, and antibodies were detectable on day 7 after infection in ELISA. Guanmu viewpoint:
The prevention and control of attenuated strains of African swine fever requires a comprehensive upgrade of the detection system to ensure early detection and timely removal of the virus before it spreads on a large scale
Screening of high-sensitivity, well-stabilized four-channel fluorescence PCR instrument.
Screening nucleic acid extractors and supporting nucleic acid extraction kits with high sensitivity, strong anti-interference ability, high nucleic acid extraction purity, and strong anti-pollution ability.
Screening of dual-target fluorescent PCR reagents with internal standard monitoring to improve detection sensitivity and accuracy.
Strengthen the detection of people, vehicles, and objects to prevent viruses from entering.
Abnormal pigs collected mouth, nose and blood swabs qPCR to detect pathogens.
EDTA anticoagulant blood qPCR was collected regularly to monitor the pathogen, and ELISA was monitored for antibodies.
Infection is found and screened, and it is quickly and accurately removed.
References: Pikalo J, Schoder M E, Sehl-Ewert J, et al towards efficient early warning: pathobiology of african swine fever virus “belgium 2018/1” in domestic pigs of different age classes[j]. animals, 2021, 11(9): 2602.