Transcriptional changes in Teladorsagia circumcincta upon encountering host tissue of differing immune status

SUMMARYThe aim of this study was to elucidate transcriptional changes in the parasitic nematodeTeladorsagia circumcinctaupon encountering either naïve or immune ovine hosts. Pools of 100 000 exsheathed 3rd- stageT. circumcinctalarvae were exposedin vitroto either an immune or naïve ovine abomasal environment, RNA was extracted from the larvae and sequenced using the Roche 454 platform. Each sample produced approximately 82 000 reads that assembled to give approximately 5500 Isotigs (contigs). The two sequence datasets were clustered together to give a total of 6969 clusters of which 18 were differentially expressed (P&lt;0·001) between the two groups. Clusters with a predominance of reads in larvae exposed to the immune abomasal environment encoded homologues of peptidyl-glycine alpha-amidating monooxygenase, heat shock-protein 16-2 and IDA-1, a tyrosine phosphatase-like receptor protein. Clusters with a predominance of reads in the naïve environment encoded homologues of cytochrome b, EGg Laying defective family member 21 and NADH dehydrogenase subunit 5. Gene ontology analyses indicated that larvae exposed to the immune environment showed an increase in expression of genes involved in ‘carbon utilization’, ‘response to stimulus’ and ‘developmental process’. These data suggest thatT. circumcinctamodulates gene expression in response to the immune status of the host.</jats:p

Stage-specific gene expression in Teladorsagia circumcincta (Nematoda : Strongylida) infective larvae and early parasitic stages

Suppression subtractive hybridisation was used to enrich genes expressed in a stage-specific manner in infective, exsheathed L3s (xL3) versus early L4s of the ovine nematode, Teladorsagia circumcincta prior to gene expression profiling by microarray. The 769 cDNA sequences obtained from the xL3-enriched library contained 361 unique sequences, with 292 expressed sequence tags (ESTs) being represented once ("singletons") and 69 sequences which were represented more than once (overlapping and non-overlapping "contigs"). The L4-enriched EST dataset contained 472 unique sequences, with 314 singletons and 158 contigs. Of these 833 sequences, 85% of the xL3 sequences and 86% of the L4 sequences exhibited homology to known genes or ESTs derived from other species of nematode. Quantitative differential expression (P &lt;0.05) was demonstrated for 563 (68%) of the ESTs by microarray. Within the U-specific dataset, more than 30% of the transcripts represented the enzyme, guanosine-5'-triphosphate (GTP)-cyclohydrolase, which is the first and rate-limiting enzyme of the tetrahydrobiopterin synthesis pathway and may be involved in critical elements of larval development. In L4s, proteolytic enzymes were highly up-regulated, as were collagens and a number of previously characterised secretory proteins, reflecting the rapid growth of these larvae in abomasal glands. (C) 2007 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.</p

Proteomic and genomic analysis reveals novel Campylobacter jejuni outer membrane proteins and potential heterogeneity

AbstractGram-negative bacterial outer membrane proteins play important roles in the interaction of bacteria with their environment including nutrient acquisition, adhesion and invasion, and antibiotic resistance. In this study we identified 47 proteins within the Sarkosyl-insoluble fraction of Campylobacter jejuni 81-176, using LC–ESI-MS/MS. Comparative analysis of outer membrane protein sequences was visualised to reveal protein distribution within a panel of Campylobacter spp., identifying several C. jejuni-specific proteins. Smith–Waterman analyses of C. jejuni homologues revealed high sequence conservation amongst a number of hypothetical proteins, sequence heterogeneity of other proteins and several proteins which are absent in a proportion of strains

Stage-specific gene expression in Teladorsagia circumcincta (Nematoda : Strongylida) infective larvae and early parasitic stages

Suppression subtractive hybridisation was used to enrich genes expressed in a stage-specific manner in infective, exsheathed L3s (xL3) versus early L4s of the ovine nematode, Teladorsagia circumcincta prior to gene expression profiling by microarray. The 769 cDNA sequences obtained from the xL3-enriched library contained 361 unique sequences, with 292 expressed sequence tags (ESTs) being represented once ("singletons") and 69 sequences which were represented more than once (overlapping and non-overlapping "contigs"). The L4-enriched EST dataset contained 472 unique sequences, with 314 singletons and 158 contigs. Of these 833 sequences, 85% of the xL3 sequences and 86% of the L4 sequences exhibited homology to known genes or ESTs derived from other species of nematode. Quantitative differential expression (P &lt;0.05) was demonstrated for 563 (68%) of the ESTs by microarray. Within the U-specific dataset, more than 30% of the transcripts represented the enzyme, guanosine-5'-triphosphate (GTP)-cyclohydrolase, which is the first and rate-limiting enzyme of the tetrahydrobiopterin synthesis pathway and may be involved in critical elements of larval development. In L4s, proteolytic enzymes were highly up-regulated, as were collagens and a number of previously characterised secretory proteins, reflecting the rapid growth of these larvae in abomasal glands. (C) 2007 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.</p

Genetic variability of strains assessed by PCR-RFLP analysis of polymorphic membrane protein-encoding genes

International audienceThis study used PCR-RFLP to investigate the genetic variability of -encoding genes from fifty-two () strains originating from abortion cases from various geographical regions and host species. Six primer pairs were used to PCR-amplify DNA fragments encoding eighteen pmps. PCR products were digested using four restriction endonucleases and Bayesian methodologies were used to compare RFLP profiles and assign strains to a RFLP genotype. Strains could be assigned to 2 genotypes in the region encoding , 3 genotypes in the regions encoding , and , 4 genotypes in the region encoding and 5 genotypes in the region encoding -. In all regions, the majority of strains (88.4%-96.1%) had the same genotype as the reference strain S26/3. No correlation could be made between genotype, host species or geographical origin except for the two variant Greek strains, LLG and POS, which formed a discrete genotype in all -encoding regions except . Relative rates of evolution calculated for each -encoding gene locus suggest that differing selective pressures and functional constraints may exist on polymorphic membrane proteins. These findings suggest that although intraspecies heterogeneity of pmp-encoding genes in is low, the sequence heterogeneity should be an important consideration when using pmps as the basis for novel diagnostics or vaccine development

Genetic and proteomic analysis of the MHC class I repertoire from four ovine haplotypes

Immunity to livestock diseases can be studied directly in the target animal, but its elucidation is often constrained by the lack of major histocompatibility complex (MHC)-defined animals. To address this issue, we have established an MHC-defined sheep resource flock generated around four diverse MHC haplotypes. Initial characterisation of the repertoire of transcribed MHC class I genes identified three class I transcripts associated with each haplotype. Nucleotide sequence, transcript abundance and phylogenetic analysis indicated that they represent alleles at up to four polymorphic loci that vary in number between the different haplotypes. The functional significance of each of these genes is evaluated here using complementary molecular genetic and proteomic approaches. We determine which genes give rise to proteins that localise to the surface of transfected cells. In addition, we provide data to support the generation of expressed products, based on immunoprecipitation of class I products from animals homozygous for each of the four MHC haplotypes followed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. This provides a clearer picture of the number of MHC class I loci in sheep and allows more rational prediction of their classical (class Ia) or non-classical (class Ib) nature. On the basis of the cellular localisation, phylogenetic and transcriptional analyses, we propose that the ovine MHC comprises a minimum of eight class I loci, with considerable variation between haplotypes

Complete Genome Sequence of Corynebacterium pseudotuberculosis Strain 1/06-A, Isolated from a Horse in North America

Corynebacterium pseudotuberculosis causes disease in several animal species, although distinct biovars exist that appear to be restricted to specific hosts. In order to facilitate a better understanding of the differences between biovars, we report here the complete genome sequence of the equine pathogen Corynebacterium pseudotuberculosis strain 1/06-A.FAPESPA - Fundação Amazônia de Amparo a Estudos e PesquisasCAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível SuperiorFAPEMIG - Fundação de Amparo à Pesquisa do Estado de Minas GeraisCERDEIRA, L. T.; SILVA, A. L. C.; BOL, Erick J. M.; LOPES, T. S.; BARBOSA, M. S. R.; CARNEIRO, A. R.; RAMOS, R. T. J.; SCHNEIDER, M. P. C. Universidade Federal do Par

Complete genome sequences of Corynebacterium pseudotuberculosis Strains 3/99-5 and 42/02-A, isolated from sheep in Scotland and Australia, respectively

Here, we report the whole-genome sequences of two ovine-pathogenic Corynebacterium pseudotuberculosisisolates: strain 3/99-5, which represents the first C. pseudotuberculosis genome originating from the United Kingdom, and 42/02-A, the second from Austra lia. These genome sequences will contribute to the objective of determining the global pan-genome of this bacterium.FAPESPA - Fundação Amazônia de Amparo a Estudos e PesquisasCAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível SuperiorFAPEMIG - Fundação de Amparo à Pesquisa do Estado de Minas GeraisCERDEIRA, L. T.; SILVA, A. L. C.; BOL, Erick J. M.; LOPES, T. S.; BARBOSA, M. S. R.; CARNEIRO, A. R.; RAMOS, R. T. J.; SCHNEIDER, M. P. C. Universidade Federal do Par