Whole-genome sequencing of Theileria parva strains provides insight into parasite migration and diversification in the african continent

The disease caused by the apicomplexan protozoan parasite Theileria parva, known as East Coast fever or Corridor disease, is one of the most serious cattle diseases in Eastern, Central, and Southern Africa. We performed whole-genome sequencing of nine T. parva strains, including one of the vaccine strains (Kiambu 5), field isolates from Zambia, Uganda, Tanzania, or Rwanda, and two buffalo-derived strains. Comparison with the reference Muguga genome sequence revealed 34 814–121 545 single nucleotide polymorphisms (SNPs) that were more abundant in buffalo-derived strains. High-resolution phylogenetic trees were constructed with selected informative SNPs that allowed the investigation of possible complex recombination events among ancestors of the extant strains. We further analysed the dN/dS ratio (non-synonymous substitutions per non-synonymous site divided by synonymous substitutions per synonymous site) for 4011 coding genes to estimate potential selective pressure. Genes under possible positive selection were identified that may, in turn, assist in the identification of immunogenic proteins or vaccine candidates. This study elucidated the phylogeny of T. parva strains based on genome-wide SNPs analysis with prediction of possible past recombination events, providing insight into the migration, diversification, and evolution of this parasite species in the African continent

Isotope or mass encoding of combinatorial libraries

Background: Combinatorial chemistry using solid-phase synthesis is a rapidly developing technology that can result in a significant reduction in the time required to find and optimize lead compounds. The application of this approach to traditional medicinal chemistry has led to the construction of libraries of small organic molecules on resin beads. A major difficulty in developing large combinatorial libraries is the lack of a facile encoding and decoding methodology to identify active compounds.Results: Several encoding schemes are described which use the ability of mass spectrometry to ascertain isotopic distributions. Molecular tags are attached to resin beads in parallel or on the linker used for chemical library synthesis. The tags are encoded via a controlled ratio of a number of stable isotopes on the tagging molecules, and range from a single to a complex isotopic distribution.Conclusions: A novel coding scheme is described that is useful for the generation of large encoded combinatorial libraries. The code can be cleaved after assay and analyzed by mass spectrometry in an automated fashion. An important element of the combinatorial discovery process is the ability to extract the structure-activity relationship (SAR) information made available by library screening. The speed and sensitivity of the mass-encoding scheme has the potential to determine the full SAR for a given library

Risk-based inspection as a cost-effective strategy to reduce human exposure to cysticerci of Taenia saginata in low-prevalence settings

Taenia saginata cysticercus is the larval stage of the zoonotic parasite Taenia saginata, with a life-cycle involving both cattle and humans. The public health impact is considered low. The current surveillance system, based on post-mortem inspection of carcasses has low sensitivity and leads to considerable economic burden. Therefore, in the interests of public health and food production efficiency, this study aims to explore the potential of risk-based and cost-effective meat inspection activities for the detection and control of T. saginata cysticercus in low prevalence settings

Improved PCR-RFLP for the Detection of Diminazene Resistance in Trypanosoma congolense under Field Conditions Using Filter Papers for Sample Storage

Animal African trypanosomiasis (AAT) is caused by different species of the pro- tozoan parasite Trypanosoma and affects a wide range of domestic animals. Trypano- soma congolense is widespread in the whole of sub-Saharan Africa and is the species causing considerable losses in livestock pro- duction, often affecting the health status of humans through endangering the food supply of rural communities. It is estimated that 50 million cattle are at risk of the disease and that the direct and indirect annual losses related to AAT reach US$4.5 billion [1]

Label-free segmentation of co-cultured cells on a nanotopographical gradient

The function and fate of cells is influenced by many different factors, one of which is surface topography of the support culture substrate. Systematic studies of nanotopography and cell response have typically been limited to single cell types and a small set of topographical variations. Here, we show a radical expansion of experimental throughput using automated detection, measurement, and classification of co-cultured cells on a nanopillar array where feature height changes continuously from planar to 250 nm over 9 mm. Individual cells are identified and characterized by more than 200 descriptors, which are used to construct a set of rules for label-free segmentation into individual cell types. Using this approach we can achieve label-free segmentation with 84% confidence across large image data sets and suggest optimized surface parameters for nanostructuring of implant devices such as vascular stents

Probing degeneracy in antigen-antibody recognition at the immunodominant site of foot-and-mouth disease virus

Antigen-antibody binding is regarded as one of the most representative examples of specific molecular recognition in nature. The simplistic view of antigenic recognition in terms of a lock-and-key mechanism is obsolete, as it is evident that both antigens and antibodies are flexible and can undergo substantial mutual adaptation. This flexibility is the source of complexities such as degeneracy and nonadditivity in antigenic recognition. We have used surface plasmon resonance to study the effects of combining multiple amino acid replacements within the sequence of the antigenic GH loop of foot-and-mouth disease virus, Our aim was 2-fold: to explore the extent to which antigenic degeneracy can be extended in this particular case, and to search for potential nonadditive effects in introducing multiple amino acid replacements. Combined Spain analysis of one such multiply substituted peptide by SPR, solution NMR and X-ray diffraction shows that antigenic degeneracy can be expected as long as residues directly interacting with the paratope are conserved and the peptide bioactive folding is unaltered

Longitudinal monitoring of Ehrlichia ruminantium infection in Gambian lambs and kids by pCS20 PCR and MAP1-B ELISA

<p>Abstract</p> <p>Background</p> <p>The epidemiology of <it>E. ruminantium </it>infection in extensively managed young animals is not adequately understood. Thus in this study, we monitored the onset (age at first infection) and kinetics of <it>E. ruminantium </it>infection and antibody response in extensively managed newborn lambs and kids at three sites in The Gambia.</p> <p>Methods</p> <p>We used a nested pCS20 PCR and MAP1-B ELISA in a longitudinal study to monitor the onset (age at first infection) and kinetics of <it>E. ruminantium </it>infection and antibody response respectively, in 77 newborn lambs and kids under a traditional husbandry system at three sites (Kerr Seringe, Keneba, Bansang) in The Gambia where heartwater is known to occur. The animals were monitored for field tick infestation and the comparative performance of the two assays in detecting <it>E. ruminantium </it>infection was also assessed.</p> <p>Results</p> <p>The infection rate detected by pCS20 PCR varied between 8.6% and 54.8% over the 162-day study period. Nineteen per cent of the animals in week 1 post-partum tested positive by pCS20 PCR with half of these infections (7/14) detected in the first 3 days after birth, suggesting that transmission other than by tick feeding had played a role. The earliest detectable <it>A. variegatum </it>infestation in the animals occurred in week 16 after birth. Antibodies detected by MAP1-B ELISA also varied, between 11.5% and 90%. Although there is considerable evidence that this assay can detect false positives and due to this and other reasons serology is not a reliable predictor of infection at least for heartwater. In contrast to the pCS20 PCR, the serological assay detected the highest proportion of positive animals in week 1 with a gradual decline in seropositivity with increasing age. The pCS20 PCR detected higher <it>E. ruminantium </it>prevalence in the animals with increasing age and both the Spearman's rank test (<it>r</it>_{<it>s </it>}= -0.1512; P = 0.003) and <it>kappa </it>statistic (-0.091 to 0.223) showed a low degree of agreement between the two assays.</p> <p>Conclusion</p> <p>The use of pCS20 PCR supported by transmission studies and clinical data could provide more accurate information on heartwater epidemiology in endemic areas and single-occasion testing of an animal may not reveal its true infection status. The view is supported because both the vector and vertical transmission may play a vital role in the epidemiology of heartwater in young small ruminants; the age range of 4 and 12 weeks corresponds to the period of increased susceptibility to heartwater in traditionally managed small ruminants.</p

High Prevalence of Drug Resistance in Animal Trypanosomes without a History of Drug Exposure

Trypanosomosis is responsible for the death of 3 million heads of cattle yearly, with 50 million animals at risk in sub-Saharan Africa. DA, a commonly used drug against the disease, was marketed decades ago. Drug resistance is reported in 21 African countries. A common argument about the origin of drug resistance is the selection by the drug of rare individuals that are naturally resistant and the propagation of those individuals in the population because of the competitive advantage they have when exposed to drug. When the drug pressure decreases, the wild-type individuals regain their supremacy. The principal objective of this study was thus to estimate the prevalence of trypanosomes resistant to DA in a population that was never exposed to the drug. Our results showing a high prevalence of drug resistance in environments free of any drug pressure is thought provoking and suggests that ceasing the use of DA will not allow for a return to a DA-sensitive population of trypanosomes. Drug resistance in animal trypanosomes thus present a pattern different from what is observed with Plasmodium sp. (causative agent of malaria) where a complete stoppage in the use of the chloroquine allows for a return to drug sensitivity

Vaccination with human anti-trastuzumab anti-idiotype scFv reverses HER2 immunological tolerance and induces tumor immunity in MMTV.f.huHER2(Fo5) mice

International audienceINTRODUCTION: Novel adjuvant therapies are needed to prevent metastatic relapses in HER2-expressing breast cancer. Here, we tested whether trastuzumab-selected single-chain Fv (scFv) could be used to develop an anti-idiotype-based vaccine to inhibit growth of HER2-positive tumor cells in vitro and in vivo through induction of long-lasting HER-specific immunity. METHODS: BALB/c mice were immunized with anti-trastuzumab anti-idiotype (anti-Id) scFv (scFv40 and scFv69), which mimic human HER2. Their sera were assessed for the presence of HER2-specific Ab1' antibodies and for their ability to reduce viability of SK-OV-3 cells, a HER2-positive cancer cell line, in nude mice. MMTV.f.huHER2(Fo5) transgenic mice were immunized with scFv40 and scFv69 and, then, growth inhibition of spontaneous HER2-positive mammary tumors, humoral response, antibody isotype as well as splenocyte secretion of IL2 and IFN-γ were evaluated. RESULTS: Adoptively-transferred sera from BALB/c mice immunized with scFv40 and scFv69 contain anti-HER2 Ab1' antibodies that can efficiently inhibit growth of SK-OV-3 cell tumors in nude mice. Similarly, prophylactic vaccination with anti-Id scFv69 fully protects virgin or primiparous FVB-MMTV.f.huHER2(Fo5) females from developing spontaneous mammary tumors. Moreover, such vaccination elicits an anti-HER2 Ab1' immune response together with a scFv69-specific Th1 response with IL2 and IFN-γ cytokine secretion. CONCLUSIONS: Anti-trastuzumab anti-Id scFv69, used as a therapeutic or prophylactic vaccine, protects mice from developing HER2-positive mammary tumors by inducing both anti-HER2 Ab1' antibody production and an anti-HER2 Th2-dependent immune response. These results suggest that scFv69 could be used as an anti-Id-based vaccine for adjuvant therapy of patients with HER2-positive tumors to reverse immunological tolerance to HER2

Human Papillomavirus-16 E7 Interacts with Glutathione S-Transferase P1 and Enhances Its Role in Cell Survival

Background:Human Papillomavirus (HPV)-16 is a paradigm for "high-risk" HPVs, the causative agents of virtually all cervical carcinomas. HPV E6 and E7 viral genes are usually expressed in these tumors, suggesting key roles for their gene products, the E6 and E7 oncoproteins, in inducing malignant transformation.Methodology/Principal Findings:By protein-protein interaction analysis, using mass spectrometry, we identified glutathione S-transferase P1-1 (GSTP1) as a novel cellular partner of the HPV-16 E7 oncoprotein. Following mapping of the region in the HPV-16 E7 sequence that is involved in the interaction, we generated a three-dimensional molecular model of the complex between HPV-16 E7 and GSTP1, and used this to engineer a mutant molecule of HPV-16 E7 with strongly reduced affinity for GSTP1.When expressed in HaCaT human keratinocytes, HPV-16 E7 modified the equilibrium between the oxidized and reduced forms of GSTP1, thereby inhibiting JNK phosphorylation and its ability to induce apoptosis. Using GSTP1-deficient MCF-7 cancer cells and siRNA interference targeting GSTP1 in HaCaT keratinocytes expressing either wild-type or mutant HPV-16 E7, we uncovered a pivotal role for GSTP1 in the pro-survival program elicited by its binding with HPV-16 E7.Conclusions/Significance:This study provides further evidence of the transforming abilities of this oncoprotein, setting the groundwork for devising unique molecular tools that can both interfere with the interaction between HPV-16 E7 and GSTP1 and minimize the survival of HPV-16 E7-expressing cancer cells. © 2009 Mileo et al