Impact of Subolesin and Cystatin Knockdown by RNA Interference in Adult Female Haemaphysalis longicornis (Acari: Ixodidae) on Blood Engorgement and Reproduction

Abstract

Currently, multi-antigenic vaccine use is the method of choice for the strategic control of ticks. Therefore, determining the efficacy of combined antigens is a promising avenue of research in the development of anti-tick vaccines. The antigen responsible for blood intake and reproduction has proven suitable as a vaccine antigen. It has been shown to silence Haemaphysalis longicornis salivary cystatin (HlSC-1) and subolesin by RNA interference. Adult unfed female ticks were injected with double-stranded RNA of (A) subolesin, (B) cystatin, (C) subolesin plus cystatin, and (D) injection buffer, then fed alongside normal unfed males up to spontaneous drop-down. The percentage of knockdowns was determined by real-time polymerase chain reaction. Sixty-three percent and 53% knockdown rates were observed in subolesin and cystatin double-stranded RNA-injected ticks respectively, while 32 and 26% knockdown rates of subolesin and cystatin transcript were observed in subolesin plus cystatin double-stranded RNA-injected ticks. Subolesin and/or cystatin knockdown causes a significant (p < 0.05) reduction in tick engorgement, egg mass weight, and egg conversion ratio. Most importantly, combined silencing did not act synergistically, but caused a similarly significant (p < 0.05) reduction in tick engorgement, egg mass weight, and egg conversion ratio. Therefore, the elucidation of multiple antigens may be helpful in the future of vaccines.

Characterization of a Novel Cysteine Protease Inhibitor from Poultry Red Mites: Potential Vaccine for Chickens

“…While no difference in the SR was observed in the first experiment, the RC of PRMs that fed on the immune plasma was significantly decreased. These data indicate that Dg-Cys plays a crucial role in the development and/or hatching of eggs, as previously reported in cystatins in ticks [37,38], and that vaccinating with Dg-Cys-his may control the population growth of PRMs. In the second experiment, no significant differences were observed in the MR.…”

Functional Evolution of Subolesin/Akirin

“…Subolesin/Akirin are also involved in the Relish/NF-kB independent gene regulation (Figure 4 ), thus playing a role in various biological processes in addition to the immune response (Figure 5 ). These processes include animal reproduction and development, causing lethal embryonic or reduced growth phenotypes in knockout mice, fruit fly, ticks, and nematodes ( Maeda et al, 2001 ; de la Fuente et al, 2006a ; Goto et al, 2008 ; Carpio et al, 2013 ; Qu et al, 2014 ), metazoan myogenesis ( Marshall et al, 2008 ; Salerno et al, 2009 ; Macqueen et al, 2010a ; Mobley et al, 2014 ; Sun et al, 2016 ), Xenopus neural development ( Liu et al, 2017 ), meiosis/carcinogenesis ( Komiya et al, 2008 ; Macqueen et al, 2010b ; Clemons et al, 2013 ), tick stress response, feeding, growth and reproduction ( Almazán et al, 2003 , 2005 ; de la Fuente et al, 2006a , 2008 ; Smith et al, 2009 ; Busby et al, 2012 ; Rahman et al, 2018 ), pathogen infection and transmission in ticks ( de la Fuente et al, 2006b , 2016 , 2017a ; Zivkovic et al, 2010a , b ; Busby et al, 2012 ; Hajdušek et al, 2013 ) and turbot ( Yang et al, 2011 ), human glioblastoma cell apoptosis ( Krossa et al, 2015 ), cattle marbling ( Sasaki et al, 2009 ; Watanabe et al, 2011 ; Kim et al, 2013 ), and mouse mitogen-induced B cell cycle progression and humoral immune responses ( Tartey et al, 2015 ). For example, as previously reported in I. scapularis and other tick species ( de la Fuente and Kocan, 2006 ; de la Fuente et al, 2006a , b , 2011 , 2013 ; Merino et al, 2013a ; de la Fuente and Contreras, 2015 ), Subolesin appears to function in multiple biological processes such as tick response to infection, feeding, reproduction, development and stress response (Figure 6 ).…”

Molecular cloning, identification, transcriptional analysis, and silencing of enolase on the life cycle of Haemaphysalis longicornis (Acari, Ixodidae) tick

“…The enolase PCR product was joined to a T7 promotor sequence using T7 promotor-linked (at both 5′ and 3′ ends) as described previously [ 13 ] with oligonucleotide primers ( Supplementary Table S1 ). T7 promoter-linked oligonucleotide primers, forward primer DsE-F (double-stranded RNA enolase forward) and reverse primer DsE-R (double-stranded RNA enolase reverse) for the enolase gene were used to generate the template for double-stranded RNA (dsRNA; Supplementary Table S1 ).…”

“…Group A (20 female ticks) was injected with 500 ng/tick of enolase dsRNA. Group B was injected with injection buffer (10 mM Tris–HCl, pH 7.0 and 1 mM EDTA) as a control [ 13 ]. Ticks were injected with dsRNA as previously described [ 16 , 17 ] using a Hamilton 33-gauge needle.…”