Development and application of serological tests for specific detection of mouse parvovirus infection

利用小鼠细小病毒（MPV/UT）和小鼠微小病毒（MMV）的核苷酸序列在大肠杆菌中表达重组蛋白。建立了以重组主要衣壳蛋白（rVP2）和重组主要非结构蛋白（rNS1）为抗原的酶联免疫吸附试验，并评价了它们在鼠细小病毒感染血清学检测中的性能。MPV-rVP2和MMV-rVP2 elisa分别与抗mpv和抗mmv小鼠血清特异性反应。MMV- rns1抗原与抗血清对MPV、MMV、Kilham rat virus （KRV）、H-1病毒等鼠细小病毒有广泛的反应范围。所有经口鼻感染MPV的小鼠，在感染后4周，用MPV- rvp2和MMV- rns1抗原进行ELISA筛查，血清均呈阳性，而用全MMV抗原进行常规ELISA筛查，血清均呈阴性。接触传播实验显示，MPV在感染后4周仍可传播，所有笼内同伴均经MPV- rvp2和MMV-rNS1酶联免疫吸附检测呈血清阳性。这些结果表明，MPV- rvp2和MMV-rVP2是区分MPV和MVM感染的特异性ELISA抗原，而MMV-rNS1抗原可用于多种啮齿动物细小病毒的ELISA通用抗原。MPV- rvp2酶联免疫吸附试验在检测口腔感染小鼠和笼内配偶血清向MPV转化的灵敏度高于传统酶联免疫吸附试验，这表明MPV- rvp2酶联免疫吸附试验在实验室小鼠MPV感染的检疫和微生物学监测中是有用的。此外，我们将这些重组蛋白用于荧光微珠试验（FMA）。FMA的灵敏度等于或高于ELISA，且FMA在一次反应中检测到多种抗原的抗体而不降低其特异性和敏感性。

Nucleotide sequences of mouse parvovirus (MPV/UT) and mouse minute virus (MMV) were used for expressing recombinant proteins in E. coli. ELISA tests using recombinant major capsid protein (rVP2) and recombinant major non-structural protein (rNS1) as antigens were developed and their performance in serologic detection of rodent parvovirus infection was assessed. MPV-rVP2 and MMV-rVP2 ELISAs reacted specifically with anti-MPV and anti-MMV mouse sera, respectively. MMV-rNS1 antigen had a wide reaction range with antisera to rodent parvoviruses including MPV, MMV, Kilham rat virus (KRV) and H-1 virus. All mice oronasally infected with MPV were seropositive at 4 weeks post-infection in screening by ELISAs using MPV-rVP2 and MMV-rNS1 antigens, but were negative by conventional ELISA using whole MMV antigen. A contact transmission experiment revealed that transmission of MPV occurred up to 4 weeks post-infection, and all cage mates were seropositive in screening with MPV-rVP2 and MMV-rNS1 ELISAs. These results indicate that MPV-rVP2 and MMV-rVP2 are specific ELISA antigens which distinguish between MPV and MVM infection, while MMV-rNS1 antigen can be used in generic ELISA for a variety of rodent parvoviruses. The higher sensitivity of MPV-rVP2 ELISA than conventional ELISA for detecting seroconversion to MPV in oronasally infected mice as well as in cage mates suggests the usefulness of MPV-rVP2 ELISA in quarantine and microbiological monitoring of MPV infection in laboratory mice. In addition, we used these recombinant proteins in fluorescent microbead assay (FMA). The sensitivity of the FMA was equal to or higher than that of ELISA, and FMA detected antibodies to multiplex antigens in a single reaction without decreasing its specificity and sensitivity.