Munro E S, Gahlawat S K, Acosta F, Ellis A E
Fisheries Research Services, Marine Laboratory, Aberdeen, UK.
J Fish Dis. 2006 Jan;29(1):43-8. doi: 10.1111/j.1365-2761.2005.00680.x.
The level of infection by infectious pancreatic necrosis virus (IPNV) of kidney macrophages from 12 asymptomatic carrier Atlantic salmon post-smolts was studied. Kidney leucocytes were fractionated on 34/51% Percoll gradients, allowed to adhere to plastic wells overnight, washed to remove non-adherent cells and cultured for up to 7 days with or without renewal of medium on day 3. On day 1, supernatants were harvested, macrophages were counted, lysed and IPNV in the supernatants and lysates was titred in chinook salmon embryo (CHSE-214) cells. The multiplicity of infection ranged between 1:2.2 and 1:7.4 (virus:macrophages). On day 3, the titres of IPNV in macrophage lysates decreased and in wells where the medium was renewed on day 3, IPNV was no longer detectable on day 7. In the supernatants, one fish was positive for IPNV on day 1, four fish on day 3 but none were detectably positive on day 7. In parallel wells in which the medium was not renewed, on day 7 IPNV was detected in macrophage lysates of three fish and the supernatants were also IPNV positive in two of these fish. This suggests that virus might be shed from infected macrophages and then reinfect other macrophages. When macrophages were serially diluted in wells and cultured for 24 h, IPNV could be cultured from macrophage lysates of wells containing between two and 70 macrophages. These results indicate that a very high proportion of the adherent kidney macrophages must be infected with very few non-replicating virions.
对12尾无症状携带传染性胰腺坏死病毒(IPNV)的大西洋鲑鱼后幼鱼的肾脏巨噬细胞的感染水平进行了研究。肾脏白细胞在34/51%的Percoll梯度上进行分级分离,使其在塑料孔中过夜贴壁,洗涤以去除未贴壁细胞,并在第3天更换或不更换培养基的情况下培养长达7天。在第1天,收集上清液,对巨噬细胞进行计数、裂解,并在奇努克鲑鱼胚胎(CHSE-214)细胞中对上清液和裂解物中的IPNV进行滴定。感染复数范围在1:2.2至1:7.4之间(病毒:巨噬细胞)。在第3天,巨噬细胞裂解物中IPNV的滴度下降,并且在第3天更换培养基的孔中,第7天不再能检测到IPNV。在上清液中,1尾鱼在第1天IPNV呈阳性,4尾鱼在第3天呈阳性,但在第7天均未检测到阳性。在未更换培养基的平行孔中,第7天在3尾鱼的巨噬细胞裂解物中检测到IPNV,其中2尾鱼的上清液中IPNV也呈阳性。这表明病毒可能从受感染的巨噬细胞中释放出来,然后再感染其他巨噬细胞。当巨噬细胞在孔中连续稀释并培养24小时时,在含有2至70个巨噬细胞的孔的巨噬细胞裂解物中可以培养出IPNV。这些结果表明,附着的肾脏巨噬细胞中很大比例必定被极少数非复制性病毒粒子感染。