The porcine reproductive and respiratory syndrome (PRRS), caused by the PRRS virus (PRRSV), is responsible for significant economic losses worldwide [1]. The PRRSV is a single strand RNA virus with high genetic variation. Two major subtypes of the virus have been described, the European genotype (type 1) and the North American genotype (type 2) [1],[2]. Highly pathogenic strains that are a sub-lineage of the PRRSV type 2 were isolated in Asia [3],[4]. An assessment of risk factors as well as the establishment of monitoring and surveillance programs are necessary to prevent losses due to PRRS [5]. In order to control the disease, one possible initiative is to regain a stable status in PRRSV positive herds, for instance by herd closure or mass vaccination [6],[7]. Another option is the eradication of PRRSV in pig herds [8] or even in larger geographic regions [9],[10]. On the other hand it is essential to maintain the status of PRRSV negative herds, for instance boar studs. Continuous and reliable monitoring of the PRRSV status of a pig herd is required in order to observe the success of the taken measures. Test systems with a high specificity and sensitivity are thus needed [11]. Several PCR methods have been established and are widely used for early diagnosis of an infection [12],[13].

One cost effective method is the serological detection of antibodies against PRRSV by ELISA. Several ELISAs have recently been developed, most of them detecting antibodies against both PRRSV type 1 and type 2 [14]-[17]. Some ELISAs, however, are intended to be able to differentiate between type 1 and 2 antibodies [16]. The IDEXX PRRS X3 Ab Test (IDEXX, Westbrook, USA) with a sensitivity of 98.8% and a specificity of 99.9%, according to the manufacturer, is the most often cited test [1],[6],[14] and is generally reckoned to be the de facto gold standard of the ELISAs for detection of antibodies against PRRSV [14],[15],[17].

The objective of the study was to test three different commercial ELISAs for the detection of antibodies against PRRSV in serum and to evaluate their specificity and sensitivity in comparison to the IDEXX PRRS X3 Ab Test.

In this study, three commercial ELISAs were compared to the IDEXX PRRS X3 Ab Test (ELISA I in this study). Two of the ELISAs (ELISA II and III) were recently developed. No published data exists concerning the specificity and sensitivity or data of agreement for these ELISAs. One recent study is available that refers to ELISA IV in comparison to ELISA I in experimentally infected pigs and 205 samples of pigs from PRRSV negative herds [16]. However, no current data for this ELISA are available for seroconversion in vaccinated pigs, pigs from naturally infected herds or wild boars.

ELISA I, II and III detected a seroconversion in all three pigs of group 1 at day 12 after vaccination with an attenuated PRRSV live vaccine. ELISA I and III, however, were able to find PRRSV antibodies in one pig as early as day 9 after vaccination. Seroconversion can usually be first detected between day 9 and 15 after vaccination or inoculation [15],[16],[22]-[25]. This was confirmed in our study with ELISA I, II and III. One pig with positive results at three occasions was found in ELISA IV which is, according to manufacturer, detecting type 1 antibodies. Since no PRRSV type 1-RNA could be detected in the pigs of group 1, this is probably a cross reaction of the ELISA. It would be useful to have more defined vaccinated or inoculated pigs to see how often this cross reaction occurs.

According to the kappa coefficient, an almost perfect agreement (κ > 0.80) was found between ELISA I on one hand and ELISA II and III on the other hand. No published reference data are available for ELISA II and III, but the newly developed ELISA II seems to be much more precise than the former INgezim PRRS Universal (Ingenasa) [26]. ELISA IV had a substantial agreement (κ = 0.71) with ELISA I that is caused by a lower specificity and sensitivity.

In our study, ELISA I as reference test had a specificity of 100%, which agrees with the manufacturer’s declaration (99.9%) and was also found in another study [16]. Other studies found only few false positive results in this ELISA [15],[17]. In our study, a very high specificity (99%) was found in ELISA II as well. More false positive samples were detected in ELISA III and IV. A lower specificity of ELISA IV has also been described in other studies (93.3% and 92.5%) [16],[26]. It has to be considered, however, that for determination of specificity, only group 3 was used in our study. ELISA IV tested ten out of 12 samples from Southeast Asia as PRRSV type 1 antibody positive that in PCR were found positive only for highly pathogenic PRRSV type 2. In Southeast Asia, the dominant genotype is highly pathogenic PRRSV type 2 [4]. No occurrence of PRRSV type 1 in the region of sample origin has been reported until now. Cross reactions of the antibodies between PRRSV type 1 and type 2 could be the reason for the positive results by ELISA IV. According to Stadejek et al. [2], ELISAs with coated antigens of either PRRSV type 1 or type 2 positive pigs preferentially, but not exclusively, detect antibodies of this type. The lower specificity of ELISA IV is also reflected in the lower positive predictive value, accuracy and kappa coefficient of this ELISA.

In group 4 on the other hand, some positive results were found with ELISA II and some more with ELISA III and IV, whereas ELISA I remained negative but with S/P values in some samples slightly beneath the cut-off. Since no anamnestic information is available on the herds, a PRRSV infection or previous vaccination cannot be excluded in farms of group 4 by a single testing for PRRSV antibodies. Since the sensitivity of ELISA I is, according to manufacturer, 98.9%, it may well be that the sensitivity of ELISA II and III is higher than this. On the other hand, antibody response detectable by ELISA after a PRRSV infection has a duration of at least 120 [11] to 137 days post infection [25]. Antibodies against PRRSV, detectable by ELISA, are present in a high percentage of older fatteners and sows of a positive farm [27]. In our study, the samples found positive in group 4 did not correspond between the ELISAs and concerned every farm of the group.

A slightly lower sensitivity (91.5%) of ELISA IV was found in our study, which is in agreement to the results of other studies [16],[26], although the sensitivity found in these studies was even lower (66.7% and 47.1%).

Blood samples of wild boars are usually of poor quality. Therefore, the sensitivity, but more so the specificity of the ELISAs must be very high to produce reliable results. In our study, only a few wild boar samples in ELISA I, II and III were PRRSV antibody positive. This confirms the usually low seroprevalence between 0% and 3.8% in European wild boars that was shown in other studies [28]-[30] and confirms the high specificity and robustness of ELISAs I, II and III. ELISA IV, however, detected more wild boar samples as PRRSV antibody positive. This also agrees with the lower specificity that was found in the other groups in this study.

This study shows that the newly developed ELISAs tested in this study are able to reliably detect antibodies against PRRSV in vaccinated and naturally infected pigs. The agreement to the reference ELISA I was almost perfect. ELISA II stood out due to a high specificity (99%) and sensitivity (96%). Occasional false positive results, however, need to be considered in all of the tested ELISAs, even in ELISA I that was used as reference method in this study. ELISA III has an especially high sensitivity (100%) that is probably superior to the reference ELISA I, but a lower specificity (95.1%). In ELISA IV that is on the market for several years, a substantial agreement to ELISA I was found. Both sensitivity and specificity of this ELISA were lower than in the other ELISAs.