55 research outputs found
Investigation into limiting dilution and tick transmissibility phenotypes associated with attenuation of the S24 vaccine strain
Abstract
Background
Babesia bovis is the causal agent of Asiatic redwater, transmitted by the pandemic tick Rhipicephalus (Boophilus) microplus. Disease control may target the tick vector using acaricides or anti-tick vaccines, or the parasite using chemoprophylaxis or anti-parasite vaccines. Current anti-parasite vaccines comprise live blood vaccines using attenuated B. bovis strains. Attenuation is attained by rapid passage that may result in different phenotypes such as reduced virulence, non-transmissibility by the tick vector, inability to sequester in the host (lack of limiting dilution) and limited genetic diversity. Attenuation and phenotypes may be linked to selection of subpopulations during rapid passage. The South African B. bovis S24 vaccine strain comprise a subpopulation that present low virulence, non-transmissibility, lack of limiting dilution phenotype and the presence of a single A558 Bv80 allele. The S24 strain could be co-transmitted with a field strain (05-100) suggesting sexual recombination. The present study investigated the change in phenotype for the S24 vaccine strain during rapid passage and co-transmission.
Methods
Vaccine phenotype change during passage as well as co-transmissibility was monitored using Bv80 allele specific PCR, limiting dilution and Illumina-based genome sequencing.
Results
The S24 population could not be rescued from the S16 passage as previously attained suggesting that selection of the S24 vaccine strain was a serendipitous and stochastic event. Passage from S16 to S24 also resulted in loss of the limiting dilution phenotype. Genome sequencing indicated sexual recombination during co-transmission with the 05-100 field strain. Analysis of the recombinant strain indicate that VESA1, smORF and SBP2 family members are present and may be responsible for the limiting dilution phenotypes, while various regions may also be responsible for the tick transmission phenotype.
Conclusions
The molecular basis for tick transmission and limiting dilution phenotypes may be defined in future using selection based on these traits in combination with sexual recombination
Geographic distribution of Theileria sp. (buffalo) and Theileria sp. (bougasvlei) in Cape buffalo (Syncerus caffer) in southern Africa : implications for speciation
Strict control measures apply to movement of buffalo in South Africa including testing for Theileria parva, the causative
agent of Corridor disease in cattle. The official test is a real-time hybridization PCR assay that amplifies the 18S rRNA V4
hyper-variable region of T. parva, T. sp. (buffalo) and T. sp. (bougasvlei). Mixed infections with the latter organisms affect
diagnostic sensitivity due to PCR suppression. While the incidence of mixed infections in the Corridor disease endemic
region of South Africa is significant, little information is available on the specific distribution and prevalence of T. sp.
(buffalo) and T. sp. (bougasvlei). Specific real-time PCR assays were developed and a total of 1211 samples known to
harbour these parasites were screened. Both parasites are widely distributed in southern Africa and the incidence of mixed
infections with T. parva within the endemic region is similar (25–50%). However, a significant discrepancy exists in
regard to mixed infections of T. sp. (buffalo) and T. sp. (bougasvlei) (10%). Evidence for speciation between T. sp.
(buffalo) and T. sp. (bougasvlei) is supported by phylogenetic analysis of the COI gene, and their designation as different
species. This suggests mutual exclusion of parasites and the possibility of hybrid sterility in cases of mixed infections.The Theileria Diagnostics
project of Onderstepoort Veterinary Institute (project
number 15/08/1P01) and a contract grant from the
Department of Agriculture and Fisheries (grant number
OV21/03/C148).http://journals.cambridge.org/action/displayJournal?jid=PARam201
Gene duplication and protein evolution in tick-host interactions
Ticks modulate their hosts' defense responses by secreting a biopharmacopiea of hundreds to thousands of proteins and bioactive chemicals into the feeding site (tick-host interface). These molecules and their functions evolved over millions of years as ticks adapted to blood-feeding, tick lineages diverged, and host-shifts occurred. The evolution of new proteins with new functions is mainly dependent on gene duplication events. Central questions around this are the rates of gene duplication, when they occurred and how new functions evolve after gene duplication. The current review investigates these questions in the light of tick biology and considers the possibilities of ancient genome duplication, lineage specific expansion events, and the role that positive selection played in the evolution of tick protein function. It contrasts current views in tick biology regarding adaptive evolution with the more general view that neutral evolution may account for the majority of biological innovations observed in ticks.The Economic Competitive
Support Programme (30/01/V010) and the National
Research Foundation (NRF) Incentive Funding
(IFR2011032400016) for Rated Researchers (NRF-Mans).
MdC was supported by an NRF/Department of Science
and Technology—Professional Development Program
(NRF/DST-PDP) studentship.http://www.frontiersin.org/Cellular_and_Infection_Microbiologyam2017Veterinary Tropical Disease
Genotypic diversity in Babesia bovis field isolates and vaccine strains from South Africa
Genotypic diversity in Babesia bovis (cause of Asiatic redwater in cattle) vaccine strains and field isolates from South Africa were investigated using the Bv80 gene as well as microsatellites. The S11 vaccine strain possessed both A and B alleles of the Bv80 gene, as well as genotypic diversity within each allele type as defined by repeat variation resulting in different amplicon sizes. Rapid serial passage of vaccine strain from passage S10 to S24 resulted in loss of genotypic diversity that yielded a single allele A genotype with an amplicon size of 558 bp. This suggested that clonal selection occurred during rapid passaging. Extensive genotypic diversity exists in 44 field isolates characterized with both Bv80 A and B alleles, but can be readily distinguished from the S24 vaccine strain using either the Bv80 allele specific PCR assays or using multi-locus micro-satellite typing. This indicated that no recent documented clinical cases of Asiatic redwater were caused by the reversion to virulence of the current vaccine strain.Red Meat Research andDevelopment South Africa (Project: OV14/06/C226).http://www.elsevier.com/locate/vetparhb2016Veterinary Tropical Disease
The host preferences of Nuttalliella namaqua (Ixodoidea : Nuttalliellidae) : a generalist approach to surviving multiple host-switches
Nuttalliella namaqua has been described as a “living fossil” and the closest extant species to the ancestral tick lineage. It was previously proposed that the Nuttalliella lineage parasitized reptile-like mammals in the Permian and had to switch hosts several times due to mass or host lineage extinctions. Extant hosts include girdled lizards and murid rodents. The current study extends knowledge on the extant host range of N. namaqua using gut meal analysis of field collected specimens. Nymphs and females can parasitize a variety of reptiles that includes skinks, geckos and girdled lizards. Blood-meal from a hyrax was also detected in a specimen suggesting that N. namaqua could parasitize a broader range of mammals than the previously suggested murid rodents. Rather than being host specific, N. namaqua is proposed to be a generalist and the ability to switch and parasitize multiple hosts allowed it to survive multiple mass and host lineage extinctions.The Joy Liebenberg Trust (21/19/JT02) allocated to BM, an incentive funding for rated researchers grant from the National Research Foundation of South Africa (NRF-Mans 76499) and a South African National Research Foundation grant allocated to AL (NRF-Spain).http://link.springer.com/journal/10493hb2014ab201
Investigating the diversity of the 18S SSU rRNA hyper-variable region of Theileria in cattle and Cape buffalo (Syncerus caffer) from southern Africa using a next generation sequencing approach
Molecular classification and systematics of the Theileria is based on the analysis of the 18S rRNA gene. Reverse line blot or conventional sequencing approaches have disadvantages in the study of 18S rRNA diversity and a next-generation 454 sequencing approach was investigated. The 18S rRNA gene was amplified using RLB primers coupled to 96 unique sequence identifiers (MIDs). Theileria positive samples from African buffalo (672) and cattle (480) from southern Africa were combined in batches of 96 and sequenced using the GS Junior 454 sequencer to produce 825711 informative sequences. Sequences were extracted based on MIDs and analysed to identify Theileria genotypes. Genotypes observed in buffalo and cattle were confirmed in the current study, while no new genotypes were discovered. Genotypes showed specific geographic distributions, most probably linked with vector distributions. Host specificity of buffalo and cattle specific genotypes were confirmed and prevalence data as well as relative parasitemia trends indicate preference for different hosts. Mixed infections are common with African buffalo carrying more genotypes compared to cattle. Associative or exclusion co-infection profiles were observed between genotypes that may have implications for speciation and systematics: specifically that more Theileria species may exist in cattle and buffalo than currently recognized. Analysis of primers used for Theileria parva diagnostics indicate that no new genotypes will be amplified by the current primer sets confirming their specificity. T. parva SNP variants that occur in the 18S rRNA hypervariable region were confirmed. A next generation sequencing approach is useful in obtaining comprehensive knowledge regarding 18S rRNA diversity and prevalence for the Theileria, allowing for the assessment of systematics and diagnostic assays based on the 18S gene.The Theileria diagnostics project ARC-OVI (OV15/08/1P01) and the Department of Agriculture, Forestry and Fisheries (OV21/03/C148).http://www.elsevier.com/locate/ttbdis2017-07-31hb2016Veterinary Tropical Disease
Mixed Theileria infections in free-ranging buffalo herds : implications for diagnosing Theileria parva infections in Cape buffalo (Syncerus caffer)
Buffalo-adapted Theileria parva causes Corridor disease in cattle. Strict control measures therefore apply to the movement
of buffalo in South Africa and include mandatory testing of buffalo for the presence of T. parva. The official test is a realtime
hybridization PCR assay that amplifies the V4 hypervariable region of the 18S rRNA gene of T. parva, T. sp. (buffalo)
and T. sp. (bougasvlei). The effect that mixed T. parva and T. sp. (buffalo)-like infections have on accurate T. parva
diagnosis was investigated in this study. In vitro mixed infection simulations indicated PCR signal suppression at 100 to
1000-fold T. sp. (buffalo) excess at low T. parva parasitaemia. Suppression of PCR signal was found in field buffalo with
mixed infections. The T. parva-positive status of these cases was confirmed by selective suppression of T. sp. (buffalo)
amplification using a locked nucleic acid clamp and independent assays based on the p67, p104 and Tpr genes. The incidence
of mixed infections in the Corridor disease endemic region of South Africa is significant, while the prevalence in buffalo
outside the endemic area is currently low. A predicted increase of T. sp. (buffalo)-like infections can affect future diagnoses
where mixed infections occur, prompting the need for improvements in current diagnostics.This project was funded by OVI project numbers 15/08/
1P01 and OV21/03/C148 (Department of Agriculture,
Forestry and Fisheries contract).http://journals.cambridge.org/action/displayJournal?jid=pa
The Hybrid II assay : a sensitive and specific real-time hybridization assay for the diagnosis of Theileria parva infection in Cape buffalo (Syncerus caffer) and cattle
Corridor disease is an acute, fatal disease of cattle caused by buffalo-adapted Theileria parva. This is a nationally controlled disease in South Africa and strict control measures apply for the movement of buffalo, which includes mandatory testing for the presence of T. parva and other controlled diseases. Accurate diagnosis of the T. parva carrier state in buffalo using the official real-time hybridization PCR assay (Sibeko et al. 2008), has been shown to be affected by concurrent infection with T. sp. (buffalo)-like parasites.We describe the Hybrid II assay, a real-time hybridization PCR method, which compareswell with the official hybridization assay in terms of specificity and sensitivity. It is, however, not influenced by mixed infections of T. sp. (buffalo)-like parasites and is as such a significant improvement on the current hybridization assay.This project was funded by OVI project numbers 15/08/
1P01 and OV21/03/C148 (Department of Agriculture,
Forestry and Fisheries contract).http://journals.cambridge.org/action/displayJournal?jid=parab201
Nuttalliella namaqua (Ixodoidea: Nuttalliellidae) : first description of the male, immature stages and re-description of the female
Nuttalliella namaqua is the only species of the enigmatic third tick family. Females possess features of hard and soft ticks
and have been designated as the ‘‘missing link’’ between the main tick families. Its position at the base of the tick tree
suggests that some of the features unique to hard and soft ticks were present in the ancestral tick lineage. Larvae, nymphae
and males have not been described to date and questions regarding their biological affinities to the main tick families
remain unclear. The current study addressed these questions via the description of larvae, nymphae and males and resolved
issues pertaining to female morphology. Field collected as well as laboratory-engorged females laid eggs and viable larvae
subsequently hatched. The larvae possess morphological structures not present in subsequent stages: namely, a sclerotized
scutum, pores on the dorsal surface of legs and a dentate anal plate. The last two characters are not present in ixodids and
argasids. N. namaqua larvae and nymphae show a similar morphology to females: a unique hypostomal structure i.e.,
bluntly rounded apically in nymphae and females and ball-like in the larvae. A re-description of some structures in female N.
namaqua has resolved differences in the original descriptions, namely that N. namaqua have 4 palpal segments as found in
ixodids and argasids and posthypostomal setae. The male was discovered for the first time and described. Characteristic
male features include: a pseudoscutum over most of the dorsum, an outgrowth on the chelicerae forming a unique rod-like
structure similar to a spematodactyl in mites and medial extension of palpal segment 2 forming a large ventral crib for
segment 4. All life stages possess some features found in hard and soft ticks and its status as the ‘‘missing link’’ between the
tick families remains.The Joy Liebenberg Trust (21/19/JT02) allocated to BM, a South African National Research Foundation grant allocated to AL
(NRF-Spain).http://www.plosone.orgab201
Next-generation sequencing as means to retrieve tick systematic markers, with the focus on Nuttalliella namaqua (Ixodoidea : Nuttalliellidae)
Nuclear ribosomal RNA (18S and 28S rRNA) and mitochondrial genomes are commonly used in tick systematics. The ability to retrieve these markers using next-generation sequencing was investigated using the tick Nuttalliella namaqua. Issues related to nuclear markers may be resolved using this approach, notably, the monotypic status of N. namaqua and its basal relationship to other tick families. Four different Illumina datasets (∼55 million, 100 bp reads each) were generated from a single tick specimen and assembled to give 350k-390k contigs. A genome size of ∼1 Gbp was estimated with low levels of repetitive elements. Contigs (>1000 bp, >50-fold coverage) present in most assemblies (n=69), included host-derived 18S and 28S rRNA, tick and host-derived transposable elements, full-length tick 18S and 28 rRNA, the mitochondrial genome in single contig assemblies and the histone cassette. Coverage for the nuclear rRNA genes was above 1000-fold confirming previous sequencing errors in the 18S rRNA gene, thereby maintaining the monotypic status of this tick. Nuclear markers for the soft tick Argas africolumbae were also retrieved from next-generation data. Phylogenetic analysis of a concatenated 18S-28S rRNA dataset supported the grouping of N. namaqua at the base of the tick tree and the two main tick families in separate clades. This study confirmed the monotypic status of N. namaqua and its basal relationship to other tick families. Next-generation sequencing of genomic material to retrieve high quality nuclear and mitochondrial systematic markers for ticks is viable and may resolve issues around conventional sequencing errors when comparing closely related tick species.The Joy Liebenberg Trust (21/19/JT02) allocated to BM,
incentive funding for rated researchers grant from the National Research Foundation
of South Africa (NRF-Mans 76499), and the ARC Tick Vaccine project
(30/01/V010).http://www.elsevier.com/locate/ttbdishb2017Veterinary Tropical Disease
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