Crowding of white shrimp Litopenaeus vananmei depresses their immunity to and resistance against Vibrio alginolyticus and white spot syndrome virus

Abstract

The full-length sequence of α2-macroglobulin (α2-M) was cloned from the haemocytes of scalloped spiny lobster Panulirus homarus by reverse transcription polymerase chain reaction (RT-PCR), cloning and sequencing of overlapping PCR and the rapid amplification of cDNA ends (RACE) method. The α2-M cDNA consists of 4,809 bp with an open reading frame (ORF) of 4,380 bp, a 72-bp 5-untranslated region (UTR), and a 357-bp 3-UTR. The predicted molecular mass of the translated protein (1,460 aminoacids) is 161.2 kDa, with an estimated pI of 5.73. The P. homarus α2-M sequence contains putative functional domains including a bait region (678-784), a GCGEQNM (979-985) thioester domain, and a receptorbinding domain (1219-1460). Sequence comparison and phylogenetic analyses show that α2-M deduced amino acid sequence of P. homarus has similarity to that of freshwater prawn Macrobrachium rosenbergii (63.4%), tiger shrimp Penaeus monodon (62.0%), white shrimp Litopenaeus vannamei (62.0%), kuruma shrimp Marsupenaeus japonicus (62.1%), mud crab Scylla serrata (46.4%), and American horseshoe crab Limulus polyphemus (43.8%), and soft tick Ornithodoros moubata (44.1%), respectively. The α2-M transcript was mainly expressed in haemocytes. Quantitative real-time RT-PCR analysis showed that α2-M mRNA transcript in haemocytes of P. homarus increased significantly in 6, 12, and 24 h, then slightly declined, but still maintained significantly higher in 48 and 72 h post Vibrio alginolyticus injection indicating an activation of innate response.