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Recombinant Rhipicephalus appendiculatus gut (Ra86) and salivary gland cement (Trp64) proteins as candidate antigens for inclusion in tick vaccines: protective effects of Ra86 on infestation with adult R. appendiculatus
Saimo M,Odongo DO,Mwaura S,Vlak JM
Vaccine: Development and Therapy , 2011,
Abstract: Margaret Saimo1,2,*, David O Odongo3,4,*, Stephen Mwaura3, Just M Vlak1, Anthony J Musoke5, George W Lubega2, Richard P Bishop3, Monique M van Oers11Laboratory of Virology, Wageningen University, Wageningen, The Netherlands; 2School of Veterinary Medicine, Makerere University, Kampala, Uganda; 3International Livestock Research Institute, Nairobi, Kenya; 4School of Biological Sciences, University of Nairobi, Nairobi, Kenya; 5Onderstepoort Veterinary Institute, Onderstepoort, Pretoria, South Africa *These two authors made an equal contribution to this workAbstract: Rhipicephalus appendiculatus gut protein Ra86 (variants Ra85A and Ra92A) and the salivary gland cement protein (Trp64) were expressed in the baculovirus-insect cell system. The recombinant gut proteins expressed as soluble proteins and the recombinant cement protein, as insoluble inclusion bodies, were used to immunize rabbits, which were then challenged with larval, nymphal, and adult stages of R. appendiculatus ticks. High tick mortality (23.3%) occurred on adult ticks that fed on rabbits vaccinated with the gut proteins, compared with 1.9% mortality in ticks that fed on unvaccinated na ve control rabbits. The mean weight of engorged female ticks was significantly reduced by 31.5% in rabbits vaccinated with the Ra86 recombinant protein compared with controls, as was egg production. Marked effects on these parameters were also observed in adult ticks as a result from vaccination using Trp64, but these were not statistically significant. For both antigens, there was no demonstrable effect on larval or nymphal ticks. This study demonstrates for the first time the protective efficacy of a homolog of Boophilus microplus Bm86 in reducing tick infestation by the adult stage of the three-host tick R. appendiculatus. The results demonstrate the potential of Ra86 for vaccine development against this tick and for the control of East Coast fever.Keywords: baculovirus, Ra85A, Ra92A, Boophilus decoloratus, Boophilus microplus
Heterogeneous Host Susceptibility Enhances Prevalence of Mixed-Genotype Micro-Parasite Infections
Wopke van der Werf ,Lia Hemerik ,Just M. Vlak,Mark P. Zwart
PLOS Computational Biology , 2011, DOI: 10.1371/journal.pcbi.1002097
Abstract: Dose response in micro-parasite infections is usually shallower than predicted by the independent action model, which assumes that each infectious unit has a probability of infection that is independent of the presence of other infectious units. Moreover, the prevalence of mixed-genotype infections was greater than predicted by this model. No probabilistic infection model has been proposed to account for the higher prevalence of mixed-genotype infections. We use model selection within a set of four alternative models to explain high prevalence of mixed-genotype infections in combination with a shallow dose response. These models contrast dependent versus independent action of micro-parasite infectious units, and homogeneous versus heterogeneous host susceptibility. We specifically consider a situation in which genome differences between genotypes are minimal, and highly unlikely to result in genotype-genotype interactions. Data on dose response and mixed-genotype infection prevalence were collected by challenging fifth instar Spodoptera exigua larvae with two genotypes of Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV), differing only in a 100 bp PCR marker sequence. We show that an independent action model that includes heterogeneity in host susceptibility can explain both the shallow dose response and the high prevalence of mixed-genotype infections. Theoretical results indicate that variation in host susceptibility is inextricably linked to increased prevalence of mixed-genotype infections. We have shown, to our knowledge for the first time, how heterogeneity in host susceptibility affects mixed-genotype infection prevalence. No evidence was found that virions operate dependently. While it has been recognized that heterogeneity in host susceptibility must be included in models of micro-parasite transmission and epidemiology to account for dose response, here we show that heterogeneity in susceptibility is also a fundamental principle explaining patterns of pathogen genetic diversity among hosts in a population. This principle has potentially wide implications for the monitoring, modeling and management of infectious diseases.
Evolutionary Trajectory of White Spot Syndrome Virus (WSSV) Genome Shrinkage during Spread in Asia
Mark P. Zwart,Bui Thi Minh Dieu,Lia Hemerik,Just M. Vlak
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0013400
Abstract: White spot syndrome virus (WSSV) is the sole member of the novel Nimaviridae family, and the source of major economic problems in shrimp aquaculture. WSSV appears to have rapidly spread worldwide after the first reported outbreak in the early 1990s. Genomic deletions of various sizes occur at two loci in the WSSV genome, the ORF14/15 and ORF23/24 variable regions, and these have been used as molecular markers to study patterns of viral spread over space and time. We describe the dynamics underlying the process of WSSV genome shrinkage using empirical data and a simple mathematical model.
In silico identification of putative promoter motifs of White Spot Syndrome Virus
Hendrik Marks, Xin-Ying Ren, Hans Sandbrink, Mari?lle CW van Hulten, Just M Vlak
BMC Bioinformatics , 2006, DOI: 10.1186/1471-2105-7-309
Abstract: The collective information shows that the upstream region of early WSSV genes, containing a TATA box and an initiator, is similar to Drosophila RNA polymerase II core promoter sequences, suggesting utilization of the cellular transcription machinery for generating early transcripts. The alignment of the 5' ends of known well-established late genes, including all major structural protein genes, identified a degenerate motif (ATNAC) which could be involved in WSSV late transcription. For these genes, only one contained a functional TATA box. However, almost half of the WSSV late genes, as previously assigned by microarray analysis, did contain a TATA box in their upstream region.The data may suggest the presence of two separate classes of late WSSV genes, one exploiting the cellular RNA polymerase II system for mRNA synthesis and the other generating messengers by a new virus-induced transcription mechanism.White Spot Syndrome Virus (WSSV), type species of the virus family Nimaviridae (genus whispovirus), is a pathogen of major economic importance in cultured penaeid shrimp [1,2]. Histopathological studies on WSSV infected shrimp have shown that the virus mainly infects tissues of ectodermal and mesodermal origin, such as the stomach, gills, heart, gut, muscle tissue and hematopoietic tissue [3-5]. Infected cells within these tissues are characterized by the appearance of homogeneous hypertrophied nuclei and chromatin margination [1,5,6]. WSSV particles have been mainly detected in the nuclei of infected cells, indicating that transcription, replication and virion assembly probably occur in the nucleus [5-8]. It is not clear how the virions are released from the nucleus of an infected cell, but this most likely occurs by budding or by rupture of the nuclear envelope and/or the cell membrane.The circular ds DNA genome of three WSSV isolates, originating from Taiwan (WSSV-TW), China (WSSV-CN) and Thailand (WSSV-TH), have been completely sequenced [9-11]. The genome of W
Functional analysis of the Autographa californica nucleopolyhedrovirus IAP1 and IAP2
XianDong Zeng,Fang Nan,ChangYong Liang,JianHua Song,Qian Wang,Just M. Vlak,XinWen Chen
Science China Life Sciences , 2009, DOI: 10.1007/s11427-009-0105-5
Abstract: The Autographa californica nucleopolyhedrovirus (AcMNPV) contains three apoptosis suppressor genes: p35, iap1 and iap2. AcMNPV P35 functions as a pancaspase inhibitor, but the function of IAP1 and IAP2 has not been entirely resolved. In this paper, we analyze the function of IAP1 and IAP2 in detail. AcMNPV with p35-deletion inhibited the apoptosis of BTI-Tn-5B1-4 (Tn-Hi5) cells induced by a Helicoverpa armigera single nucleocapsid NPV (HearNPV) infection and rescued the replication of HearNPV and BV production in these cells. Transient-expression experiments indicated that both IAP1 and IAP2 suppress apoptosis of Tn-Hi5 cells during HearNPV infection. Recombinant HearNPVs expressing AcMNPV iap1, iap2 and p35, respectively, not only prevented apoptosis but also allowed HearNPV to replicate in Tn-Hi5 cells. However, the iap1, iap2 and p35 genes when expressed in HearNPV were unable to rescue BV production. These results indicate that both AcMNPV iap1 and iap2 function independently as apoptosis inhibitors of and are potential host range factors.
苜蓿银纹夜蛾核多角体病毒iap1和iap2基因功能分析
曾宪东,南方,梁昌镛,宋建华,王倩,Just M. Vlak,陈新文
中国科学 生命科学 , 2009,
Abstract: 苜蓿银纹夜蛾核多角体病毒(Autographa californica nucleopolyhedrovirus,AcMNPV)基因组含有3个细胞凋亡抑制基因,即p35,iap1和iap2.其中,p35作为一个有效的依赖于天冬氨酸的半胱氨酸蛋白酶(caspase)抑制因子,能够抑制多种因素诱发细胞凋亡,而iap1和iap2的功能仍未完全明晰,本研究对IAP1和IAP2的功能进行了详细分析.缺失了p35的AcMNPV仍可抑制棉铃虫核多角体病毒(Helicoverpa armigera single nucleocapsid NPV,HearNPV)诱导的BTI-Tn-5B1-4(Tn-Hi5)细胞凋亡并挽救HearNPV在Tn-Hi5细胞中复制及HearNPV出芽型病毒粒子的产生.进一步构建了瞬时表达质粒以及分别表达AcMNPV的p35,iap1和iap2基因的重组HearNPV,转染瞬时表达的IAP1和IAP2对HearNPV感染诱导的Tn-Hi5细胞凋亡有抑制效果,而重组病毒感染Tn-Hi5细胞也可抑制其凋亡并在其中复制,然而重组HearNPV表达的p35,iap1和iap2并未能挽救出芽型病毒粒子的产生....
Functional analysis of the Autographa californica nucleopolyhedrovirus IAP1 and IAP2

XianDong Zeng,Fang Nan,ChangYong Liang,JianHua Song,Qian Wang,Just M Vlak,XinWen Chen,

中国科学C辑(英文版) , 2009,
Abstract: The Autographa californica nucleopolyhedrovirus (AcMNPV) contains three apoptosis suppressor genes: p35, iap1 and iap2. AcMNPV P35 functions as a pancaspase inhibitor, but the function of IAP1 and IAP2 has not been entirely resolved. In this paper, we analyze the function of IAP1 and IAP2 in de-tail. AcMNPV with p35-deletion inhibited the apoptosis of BTI-Tn-5B1-4 (Tn-Hi5) cells induced by a Helicoverpa armigera single nucleocapsid NPV (HearNPV) infection and rescued the replication of HearNPV and BV production in these cells. Transient-expression experiments indicated that both IAP1 and IAP2 suppress apoptosis of Tn-Hi5 cells during HearNPV infection. Recombinant HearNPVs ex-pressing AcMNPV iap1, iap2 and p35, respectively, not only prevented apoptosis but also allowed HearNPV to replicate in Tn-Hi5 cells. However, the iap1, iap2 and p35 genes when expressed in HearNPV were unable to rescue BV production. These results indicate that both AcMNPV iap1 and iap2 function independently as apoptosis inhibitors of and are potential host range factors.
苜蓿银纹夜蛾核多角体病毒iap1和iap2基因功能分析
曾宪东,南方,梁昌镛,宋建华,王倩,Just,M.,Vlak,陈新文
中国科学 生命科学 , 2009,
Abstract: 苜蓿银纹夜蛾核多角体病毒(Autographacalifornicanucleopolyhedrovirus,AcMNPV)基因组含有3个细胞凋亡抑制基因,即p35,iap1和iap2.其中,p35作为一个有效的依赖于天冬氨酸的半胱氨酸蛋白酶(caspase)抑制因子,能够抑制多种因素诱发细胞凋亡,而iap1和iap2的功能仍未完全明晰,本研究对IAP1和IAP2的功能进行了详细分析.缺失了p35的AcMNPV仍可抑制棉铃虫核多角体病毒(HelicoverpaarmigerasinglenucleocapsidNPV,HearNPV)诱导的BTI-Tn-5B1-4(Tn-Hi5)细胞凋亡并挽救HearNPV在Tn-Hi5细胞中复制及HearNPV出芽型病毒粒子的产生.进一步构建了瞬时表达质粒以及分别表达AcMNPV的p35,iap1和iap2基因的重组HearNPV,转染瞬时表达的IAP1和IAP2对HearNPV感染诱导的Tn-Hi5细胞凋亡有抑制效果,而重组病毒感染Tn-Hi5细胞也可抑制其凋亡并在其中复制,然而重组HearNPV表达的p35,iap1和iap2并未能挽救出芽型病毒粒子的产生.结果表明,AcMNPV的iap1和iap2基因表达产物作为细胞凋亡抑制因子是有功能的.
Recombinant Rhipicephalus appendiculatus gut (Ra86) and salivary gland cement (Trp64) proteins as candidate antigens for inclusion in tick vaccines: protective effects of Ra86 on infestation with adult R. appendiculatus
Saimo M, Odongo DO, Mwaura S, Vlak JM, Musoke AJ, Lubega GW, Bishop RP, van Oers MM
Vaccine: Development and Therapy , 2011, DOI: http://dx.doi.org/10.2147/VDT.S20827
Abstract: ombinant Rhipicephalus appendiculatus gut (Ra86) and salivary gland cement (Trp64) proteins as candidate antigens for inclusion in tick vaccines: protective effects of Ra86 on infestation with adult R. appendiculatus Original Research (1844) Total Article Views Authors: Saimo M, Odongo DO, Mwaura S, Vlak JM, Musoke AJ, Lubega GW, Bishop RP, van Oers MM Published Date November 2011 Volume 2011:1 Pages 15 - 23 DOI: http://dx.doi.org/10.2147/VDT.S20827 Margaret Saimo1,2,*, David O Odongo3,4,*, Stephen Mwaura3, Just M Vlak1, Anthony J Musoke5, George W Lubega2, Richard P Bishop3, Monique M van Oers1 1Laboratory of Virology, Wageningen University, Wageningen, The Netherlands; 2School of Veterinary Medicine, Makerere University, Kampala, Uganda; 3International Livestock Research Institute, Nairobi, Kenya; 4School of Biological Sciences, University of Nairobi, Nairobi, Kenya; 5Onderstepoort Veterinary Institute, Onderstepoort, Pretoria, South Africa *These two authors made an equal contribution to this work Abstract: Rhipicephalus appendiculatus gut protein Ra86 (variants Ra85A and Ra92A) and the salivary gland cement protein (Trp64) were expressed in the baculovirus-insect cell system. The recombinant gut proteins expressed as soluble proteins and the recombinant cement protein, as insoluble inclusion bodies, were used to immunize rabbits, which were then challenged with larval, nymphal, and adult stages of R. appendiculatus ticks. High tick mortality (23.3%) occurred on adult ticks that fed on rabbits vaccinated with the gut proteins, compared with 1.9% mortality in ticks that fed on unvaccinated na ve control rabbits. The mean weight of engorged female ticks was significantly reduced by 31.5% in rabbits vaccinated with the Ra86 recombinant protein compared with controls, as was egg production. Marked effects on these parameters were also observed in adult ticks as a result from vaccination using Trp64, but these were not statistically significant. For both antigens, there was no demonstrable effect on larval or nymphal ticks. This study demonstrates for the first time the protective efficacy of a homolog of Boophilus microplus Bm86 in reducing tick infestation by the adult stage of the three-host tick R. appendiculatus. The results demonstrate the potential of Ra86 for vaccine development against this tick and for the control of East Coast fever.
Effective Chikungunya Virus-like Particle Vaccine Produced in Insect Cells
Stefan W. Metz,Joy Gardner,Corinne Geertsema,Thuy T. Le,Lucas Goh,Just M. Vlak,Andreas Suhrbier,Gorben P. Pijlman
PLOS Neglected Tropical Diseases , 2013, DOI: 10.1371/journal.pntd.0002124
Abstract: The emerging arthritogenic, mosquito-borne chikungunya virus (CHIKV) causes severe disease in humans and represents a serious public health threat in countries where Aedes spp mosquitoes are present. This study describes for the first time the successful production of CHIKV virus-like particles (VLPs) in insect cells using recombinant baculoviruses. This well-established expression system is rapidly scalable to volumes required for epidemic responses and proved well suited for processing of CHIKV glycoproteins and production of enveloped VLPs. Herein we show that a single immunization with 1 μg of non-adjuvanted CHIKV VLPs induced high titer neutralizing antibody responses and provided complete protection against viraemia and joint inflammation upon challenge with the Réunion Island CHIKV strain in an adult wild-type mouse model of CHIKV disease. CHIKV VLPs produced in insect cells using recombinant baculoviruses thus represents as a new, safe, non-replicating and effective vaccine candidate against CHIKV infections.
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