| 1. |
- Arai, M, et al.
(författare)
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Both mosquito-derived xanthurenic acid and a host blood-derived factor regulate gametogenesis of Plasmodium in the midgut of the mosquito
- 2001
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier. - 0166-6851 .- 1872-9428. ; 116:1, s. 17-24
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Tidskriftsartikel (refereegranskat)abstract
- Gametogenesis of Plasmodium in vitro can be induced by the combined stimulus of a 5 degrees C fall in temperature and the presence of xanthurenic acid (XA). In-vitro experiments showed that P. gallinaceum (EC(50)=80 nM) is much more sensitive to XA than P. berghei (9 microM), P. yoelii (8 microM), and P. falciparum (2 microM). However, in the mosquito vector, we do not know whether the temperature shift and XA are the only gametocyte-activating factors (GAF), nor do we know with certainty the true source(s) of XA in the mosquito blood meal. Previous studies indicate that XA is the only source of GAF in the mosquito. By defining, and then contrasting, the ability of an XA-deficient mutant of Aedes aegypti, with the wild-type mosquito to support exflagellation and ookinete formation in vivo, we determined the roles of parasite-, mosquito- and host blood-derived GAF in the regulation of gametogenesis of P. gallinaceum. Removal of both host and vector sources of GAF totally inhibited both exflagellation and ookinete production, whilst the lack of either single source resulted in only a partial reduction of exflagellation and ookinete formation in the mosquito gut. Both sources can be effectively replaced/substituted by synthetic XA. This suggests (1) both mosquito- and vertebrate-derived factors act as GAF in the mosquito gut in vivo; (2) the parasite itself is unable to produce any significant GAF activity. Studies are underway to determine whether vertebrate-derived GAF is XA. These data may form the basis of further studies of the development of new methods of interrupting malarial transmission.
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| 2. |
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| 3. |
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| 4. |
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| 5. |
- Fusai, T, et al.
(författare)
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Characterisation of the chondroitin sulphate of Saimiri brain microvascular endothelial cells involved in Plasmodium falciparum cytoadhesion
- 2000
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Ingår i: Molecular and biochemical parasitology (Print). - 0166-6851 .- 1872-9428. ; 108:1, s. 25-37
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Tidskriftsartikel (refereegranskat)abstract
- Cytoadhesion of Plasmodium falciparum-infected erythrocytes (IRBC) to chondroitin-4-sulphate (CSA) is inhibited by soluble CSA in vitro on Saimiri brain microvascular endothelial cells (SBEC) and in vivo in P. falciparum-infected Saimiri monkeys. We tested whether the SBEC model was appropriate for studying CSA-binding IRBC using four cell lines. All SBEC expressed a chondroitin sulphate (CS), with a composition of CSA. The mean sizes of these CSA were 20.5, 22, 23, 32.5 and 36 kDa for SBEC 3A and C2, CHO, SBEC 1D and 17, respectively. We found that cytoadhesion of the Palo-Alto (FUP)1 CSA-binding phenotype, selected by panning on SBEC 17, was specifically inhibited in a dose-dependent manner by all the purified CSA. The extent of inhibition depended on the cellular origin of the tested CSA. SBEC 17 CSA was 33 times more efficient than CHO-CSA and 21 times more efficient than the 50 kDa commercial bovine trachaea CSA. Dynabeads coated with a total extract of SBEC 1D CS-proteoglycans interacted with CSA- but not with CD36- or ICAM-1-binding IRBC. These Dynabeads also interacted specifically with the PfEMP1 DBL-3 domain, on the surface of CHO transfectants, but not with the CIDR-1 domain. Thrombomodulin was involved in IRBC adhesion to all SBEC whereas CD44 was only expressed by SBEC 1D and 17. These two CSA-proteoglycans have also been detected at the surface of human endothelial cells. Thus, the two homologous models, SBEC/Saimiri sciureus, are useful and reliable tools for the evaluation of new anti-CSA adhesion treatments and anti-disease vaccines for pregnant women.
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| 6. |
- Henriksson, Jan, et al.
(författare)
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Chromosome specific markers reveal conserved linkage groups in spite of extensive chromosomal size variation in Trypanosoma cruzi
- 1995
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Ingår i: Molecular and biochemical parasitology (Print). - 0166-6851 .- 1872-9428. ; 73:1-2, s. 63-74
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Tidskriftsartikel (refereegranskat)abstract
- The karyotypes of three cloned stocks, CL Brener (CL), CA I/72 (CA) and Sylvio X10/7 (X10), of Trypanosoma cruzi were studied by pulsed-field gel electrophoresis followed by ethidium bromide staining and hybridization with 35 different probes, 30 of which identified single chromosomes. The chromosome-specific probes identified between 26 and 31 chromosomal bands in the three cloned stocks, corresponding to 20 unique chromosomes in CL and 19 in CA and X10. Considering the DNA content of the parasite, it was predicted that the markers recognise at least half of all T. cruzi chromosomes. A majority of identified chromosomes showed large differences in size among different strains, in some cases by up to 50%. Interestingly, CL had in general larger chromosomes than the two other studied cloned stocks. Several of the markers showed linkage and nine different linkage groups were identified, each comprising 2-4 markers. The linkage between the markers was maintained in 8 of the 9 linkage groups when a panel comprising 26 different T. cruzi strains representing major T. cruzi populations was tested. One linkage group was found to be maintained in some strains but not in others. This result shows that chromosomal rearrangements occur in the T. cruzi genome, albeit with a low frequency. Repetitive DNA, both non-coding and in one case coding, was more abundant in the cloned stock CL Brener than in CA and X10. The information presented will make it possible to select chromosomes for the construction of physical chromosomal maps required for the T. cruzi genome project.
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| 7. |
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| 8. |
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| 9. |
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| 10. |
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| 11. |
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| 12. |
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| 13. |
- Agüero, Fernán, et al.
(författare)
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Purification, cloning, and expression of the mitochondrial malate dehydrogenase (mMDH) from protoscolices of Echinococcus granulosus
- 2004
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 137:2, s. 207-214
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Tidskriftsartikel (refereegranskat)abstract
- Protoscolices of the parasitic helminth Echinococcus granulosus contain two malate dehydrogenases (EC 1.1.1.37), one cytosolic and one mitochondrial. The latter has been separated from the other isoform and purified to protein homogeneity. Sequencing of tryptic peptides by Edman degradation allowed the design of oligonucleotide primers for PCR, leading to the cloning and sequencing of a full length cDNA. The encoding gene is present as a single copy per haploid genome and codes for a protein with high sequence identity (56-58%) with the similar enzymes from mammals, Caenorhabditis elegans and yeast. Active recombinant mitochondrial malate dehydrogenase was expressed in Escherichia coli, as protein fusions with glutathione S-transferase or a poly-His tail. The purified recombinant enzymes had a kinetic behaviour similar to that of the native enzyme, being inhibited by excess of the substrate oxaloacetate and unaffected by excess L-malate. The results indicate that E. granulosus contains two typical eukaryotic malate dehydrogenases, with relative levels quite different from those present in mammalian tissues like heart, in good agreement with the predominantly fermentative metabolism of the protoscolices.
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| 14. |
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| 15. |
- Birkeland, Shanda R., et al.
(författare)
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Transcriptome analyses of the Giardia lamblia life cycle
- 2010
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 174:1, s. 62-65
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Tidskriftsartikel (refereegranskat)abstract
- We quantified mRNA abundance from 10 stages in the Giardia lamblia life cycle in vitro using Serial Analysis of Gene Expression (SAGE). 163 abundant transcripts were expressed constitutively. 71 transcripts were upregulated specifically during excystation and 42 during encystation. Nonetheless, the transcriptomes of cysts and trophozoites showed major differences. SAGE detected co-expressed clusters of 284 transcripts differentially expressed in cysts and excyzoites and 287 transcripts in vegetative trophozoites and encysting cells. All clusters included known genes and pathways as well as proteins unique to Giardia or diplomonads. SAGE analysis of the Giardia life cycle identified a number of kinases, phosphatases, and DNA replication proteins involved in excystation and encystation, which could be important for examining the roles of cell signaling in giardial differentiation. Overall, these data pave the way for directed gene discovery and a better understanding of the biology of G. lamblia.
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| 16. |
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| 17. |
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| 18. |
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| 19. |
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| 20. |
- Chen, QJ, et al.
(författare)
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Untitled
- 2004
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Ingår i: MOLECULAR AND BIOCHEMICAL PARASITOLOGY. - : Elsevier BV. - 0166-6851. ; 134:1, s. 1-1
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)
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| 21. |
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| 22. |
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| 23. |
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| 24. |
- Ferella, Marcela, et al.
(författare)
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Gene expression changes during Giardia-host cell interactions in serum-free medium
- 2014
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 197:1-2, s. 21-23
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Tidskriftsartikel (refereegranskat)abstract
- Serial Analysis of Gene Expression (SAGE) was used to quantify transcriptional changes in Giardia intestinalis during its interaction with human intestinal epithelial cells (IECs, HT-29) in serum free M199 medium. Transcriptional changes were compared to those in trophozoites alone in M199 and in TYI-S-33 Giardia growth medium. In total, 90 genes were differentially expressed, mainly those involved in cellular redox homeostasis, metabolism and small molecule transport but also cysteine proteases and structural proteins of the giardin family. Only 29 genes changed their expression due to IEC interaction and the rest were due to M199 medium. Although our findings generated a small dataset, it was consistent with our earlier microarray studies performed under different interaction conditions. This study has confined the number of genes in Giardia to a small subset that specifically change their expression due to interaction with IECs.
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| 25. |
- Fernandez-Calero, Tamara, et al.
(författare)
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Profiling of small RNA cargo of extracellular vesicles shed by Trypanosoma cruzi reveals a specific extracellular signature.
- 2015
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Ingår i: Molecular and Biochemical Parasitology. - : Elsevier BV. - 1872-9428 .- 0166-6851. ; 199:1-2, s. 19-28
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Tidskriftsartikel (refereegranskat)abstract
- Over the last years, an expanding family of small regulatory RNAs (e.g. microRNAs, siRNAs and piRNAs) was recognized as key players in novel forms of post-transcriptional gene regulation in most eukaryotes. However, the machinery associated with Ago/Dicer-dependent small RNA biogenesis was thought to be either entirely lost or extensively simplified in some unicellular organisms including Trypanosoma cruzi, Saccharomyces cerevisiae, Leishmania major and Plasmodium falciparum. Although the biogenesis of small RNAs from non-coding RNAs represent a minor fraction of the normal small RNA transcriptome in eukaryotic cells, they represent the unique small RNA pathways in Trypanosoma cruzi which produce different populations of small RNAs derived from tRNAs, rRNAs, sn/snoRNAs and mRNAs. These small RNAs are secreted included in extracellular vesicles and transferred to other parasites and susceptible mammalian cells. This process represents a novel form of cross-kingdom transfer of genetic material suggesting that secreted vesicles could represent new relevant pieces in life cycle transitions, infectivity and cell-to-cell communication. Here, we provide for the first time a detailed analysis of the small RNA cargo of extracellular vesicles from T. cruzi epimastigotes under nutritional stress conditions compared to the respective intracellular compartment using deep sequencing. Compared with the intracellular compartment, shed extracellular vesicles showed a specific extracellular signature conformed by distinctive patterns of small RNAs derived from rRNA, tRNA, sno/snRNAs and protein coding sequences which evidenced specific secretory small RNA processing pathways.
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| 26. |
- Flick, K, et al.
(författare)
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var genes, PfEMP1 and the human host
- 2004
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Ingår i: Molecular and biochemical parasitology. - : Elsevier BV. - 0166-6851. ; 134:1, s. 3-9
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Tidskriftsartikel (refereegranskat)
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| 27. |
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| 28. |
- Garcia-Silva, Maria Rosa, et al.
(författare)
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A population of tRNA-derived small RNAs is actively produced in Trypanosoma cruzi and recruited to specific cytoplasmic granules
- 2010
-
Ingår i: Molecular and Biochemical Parasitology. - : Elsevier BV. - 1872-9428 .- 0166-6851. ; 171:2, s. 64-73
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Tidskriftsartikel (refereegranskat)abstract
- Over the last years an expanding family of small RNAs (i.e. microRNAs, siRNAs and piRNAs) was recognized as key players in diverse forms of gene silencing and chromatin organization. Effectors functions of these small RNAs are achieved through ribonucleoprotein (RNP) complexes containing at their center an Argonaute/Piwi protein. Although these proteins and their small RNA-associated machinery can be traced back to the common ancestor of eukaryotes, this machinery seems to be entirely lost or extensively simplified in some unicellular organisms including Trypanosoma cruzi, which are unable to trigger RNAi related phenomena. Speculating about the presence of alternate small RNA-mediated pathways in these organisms, we constructed and analyzed a size-fractionated cDNA library (20-35 nt) from epimastigotes forms of T. cruzi. Our results showed the production of an abundant class of tRNA-derived small RNAs preferentially restricted to specific isoacceptors and whose production was more accentuated under nutritional stress. These small tRNAs derived preferentially from the 5' halves of mature tRNAs and were recruited to distinctive cytoplasmic granules. Our data favor the idea that tRNA cleavage is unlikely to be the consequence of non-specific degradation but a controlled process, whose biological significance remains to be elucidated. (C) 2010 Elsevier B.V. All rights reserved.
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| 29. |
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| 30. |
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| 31. |
- Holmfeldt, Per, et al.
(författare)
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The Schistosoma mansoni protein SM16/SmSLP/SmSPO-1 is a membrane-binding protein that lacks the proposed microtubule-regulatory activity
- 2007
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 156:2, s. 225-234
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Tidskriftsartikel (refereegranskat)abstract
- Sm16/SmSLP/SPO-1 (Sm16) has been identified as a developmentally regulated protein that is released from specific glands of the Schistosoma mansoni parasite during skin penetration. Sm16 has been ascribed both anti-inflammatory activities and a functional similarity with the conserved cytosolic tubulin-binding protein stathmin/Op18. Here we used a cell line to confirm signal peptide-dependent secretion and to define the secreted form of Sm16 for production in E. coli. We present evidence from both in vitro experiments and studies on transfected human cells that refute any functional similarity with stathmin/Op18. Instead of an Op18-like activity, we found that targeting of Sm16 to the cytosol of human cells, which was achieved by ectopic expression of Sm16 lacking the signal peptide, results in a caspase-dependent apoptotic response. Interestingly, by analysis of recombinant preparations we found that the secreted form of Sm16 is a lipid bilayer-binding protein that efficiently binds to the surface of diverse cell types by a polyanion-independent mechanism, which results in uptake by endocytosis. While the significance of the pro-apoptotic activity exerted by cytosolic Sm16 remains unclear, the present findings on cell-surface-binding properties of Sm16 seems likely to be of functional relevance during skin penetration of the parasite.
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| 32. |
- Johnson, Paul C D, et al.
(författare)
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Abundant variation in microsatellites of the parasitic nematode Trichostrongylus tenuis and linkage to a tandem repeat.
- 2006
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 148:2
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Tidskriftsartikel (refereegranskat)abstract
- An understanding of how genes move between and within populations of parasitic nematodes is important in combating the evolution and spread of anthelmintic resistance. Much has been learned by studying mitochondrial DNA markers, but autosomal markers such as microsatellites have been applied to only a few nematode species, despite their many advantages for studying gene flow in eukaryotes. Here, we describe the isolation of 307 microsatellites from Trichostrongylus tenuis, an intestinal nematode of red grouse. High levels of variation were revealed at sixteen microsatellite loci (including three sex-lined loci) in 111 male T. tenuis nematodes collected from four hosts at a single grouse estate in Scotland (average He = 0.708; mean number of alleles = 12.2). A population genetic analysis detected no deviation from panmixia either between (F(ST) = 0.00) or within hosts (F(IS) = 0.015). We discuss the feasibility of developing microsatellites in parasitic nematodes and the problem of null alleles. We also describe a novel 146-bp repeat element, TteREP1, which is linked to two-thirds of the microsatellites sequenced and is associated with marker development failure. The sequence of TteREP1 is related to the TcREP-class of repeats found in several other trichostrongyloid species including Trichostrongylus colubriformis and Haemonchus contortus.
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| 33. |
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| 34. |
- Lauwaet, Tineke, et al.
(författare)
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Protein phosphatase 2A plays a crucial role in Giardia lamblia differentiation
- 2007
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 152:1, s. 80-89
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Tidskriftsartikel (refereegranskat)abstract
- The ability of Giardia lamblia to undergo two distinct differentiations in response to physiologic stimuli is central to its pathogenesis. The giardial cytoskeleton changes drastically during encystation and excystation. However, the signal transduction pathways mediating these transformations are poorly understood. We tested the hypothesis that PP2A, a highly conserved serine/threonine protein phosphatase, might be important in giardial differentiation. We found that in vegetatively growing trophozoites, gPP2A-C protein localizes to basal bodies/centrosomes, and to cytoskeletal structures unique to Giardia: the ventral disk, and the dense rods of the anterior, posterior-lateral, and caudal flagella. During encystation, gPP2A-C protein disappears from only the anterior flagellar dense rods. During excystation, gPP2A-C localizes to the cyst wall in excysting cysts but is not found in the wall of cysts with emerging excyzoites. Transcriptome and immunoblot analyses indicated that gPP2A-C mRNA and protein are upregulated in mature cysts and during the early stage of excystation that models passage through the host stomach. Stable expression of gPP2A-C antisense RNA did not affect vegetative growth, but strongly inhibited the formation of encystation secretory vesicles (ESV) and water-resistant cysts. Moreover, the few cysts that formed were highly defective in excystation. Thus, gPP2A-C localizes to universal cytoskeletal structures and to structures unique to Giardia. It is also important for encystation and excystation, crucial giardial transformations that entail entry into and exit from dormancy.
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| 35. |
- Liu, Jingyi, et al.
(författare)
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Cleavage specificity of recombinant Giardia intestinalis cysteine proteases : Degradation of immunoglobulins and defensins
- 2019
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 227, s. 29-38
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Tidskriftsartikel (refereegranskat)abstract
- Giardia intestinalis is a protozoan parasite and the causative agent of giardiasis, a common diarrheal disease. Cysteine protease (CP) activities have been suggested to be involved in Giardia's pathogenesis and we have recently identified and characterized three secreted Giardia CPs; CP14019, CP16160 and CP16779. Here we have studied the cleavage specificity of these CPs using substrate phage display and recombinant protein substrates. The phage display analyses showed that CP16160 has both chymase and tryptase activity and a broad substrate specificity. This was verified using recombinant protein substrates containing different variants of the cleavage sites. Phage display analyses of CP14019 and CP16779 failed but the substrate specificity of CP14019 and CP16779 was tested using the recombinant substrates generated for CP16160. CP16160 and CP14019 showed similar substrate specificity, while CP16779 has a slightly different substrate specificity. The consensus sequence for cleavage by CP16160, obtained from phage display analyses, was used in an in silico screen of the human intestinal proteome for detection of potential targets. Immunoglobulins, including IgA and IgG and defensins (α-HD6 and β-HD1) were predicted to be targets and they were shown to be cleaved by the recombinant CPs in vitro. Our results suggest that the secreted Giardia CPs are key players in the interaction with host cells during Giardia infections since they can cleave several components of the human mucosal defense machinery.
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| 36. |
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| 37. |
- Mok, Bobo W., et al.
(författare)
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Comparative transcriptornal analysis of isogenic Plasmodium falciparum clones of distinct antigenic and adhesive phenotypes
- 2007
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 151:2, s. 184-192
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Tidskriftsartikel (refereegranskat)abstract
- Antigenic variation is a survival mechanism developed by the malaria parasite Plasmodium falciparum in order to allow for the establishment of a chronic infection. Here we have studied clonal differences in the transcriptomes of two isogenic P. falciparum clones (3D7S8.4 and 3D7AH1S2) of distinct adhesive and antigenic phenotypes employing a P. falciparum 70-mer oligonucleolide microarray. Fifteen transcripts were highly differentially expressed (greater than a 5-fold change) with five transcripts upregulated in 3D7AH1S2 compared to 3D7S8.4, and ten downregulated. Identified genes encode apical organellar (Gbph2, GBP-related antigen), cell cycle and DNA/RNA processing (SERA-5, RNA-methylase), cell-rescue, defense/virulence (RESA-2, RIFIN, PfEMP1) and hypothetical proteins (PFB0115w, PFI1445w, MAL13P1.121). A number of short and full-length var transcripts were differentially expressed between the clones but one full-length transcript was dominant in both rings and trophozoites (PFD0630c versus PFF0845c). Distinct members of two other variant gene families (phist-a and rif-like), scattered over the subtelomeric areas of the 14 chromosomes, were also found to be clonally and developmentally expressed. Three sibling-clones of 3D7AH1S2 (3D7AH1S1, -S3, -S4) were further studied for the expression of transcripts upregulated in 3D7AH1S2 compared to 3D7S8.4. Individual var and phist-a genes were found expressed in all of the clones while the expression of a rif-like gene and gbph2 varied in-between the clones. The present data provides evidence for complex transcriptional differences between closely related isogenic R falciparum of distinct adhesive and antigenic characteristics.
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| 38. |
- Morrison, David
(författare)
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Microarray analyses of mouse responses to infection by Neospora caninum identifies disease associated cellular pathways in the host response
- 2010
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Ingår i: Molecular and Biochemical Parasitology. - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 174, s. 117-127
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Tidskriftsartikel (refereegranskat)abstract
- Neospora caninum is a coccidian cyst-forming parasite found in a wide range of host species such as mice, dogs and Cattle. The development of methods such as vaccines to prevent abortion and fetal loss due to neosporosis would be greatly assisted by further knowledge on immunity and host responses to infection. In this study we used microarray technology to investigate the protective host responses occurring at 611 post infection in the spleen of mice infected with a prototype live N caninum vaccine. Naive non-pregnant mice were infected with the NC-Nowra isolate as such infections are known to induce protective host responses that will prevent transplacental transmission of a challenge given using pregnancy. The expression data was analysed by SAM (significance of microarrays). ANOVA and clustering methods. Gene lists were investigated for enrichment of gene ontology terms by functional annotation using hypergeometric tests. The results show that Qs and BALB/c mice infected with NC-Nowra differ in their transcriptional responses to infection and these affect a wide range of biological and molecular processes. Transcriptional changes in the Jak-STAT signaling pathway (as well as Id and other IFN-gamma regulated molecules such as GTPases) confirmed the influence of IFN-gamma in the mouse response to N. caninum. Gene ontology analyses also assigned some of the molecules involved to well known disease pathways associated with cancer, Parkinson's and Alzheimer's diseases, which were linked to the cell cycle, mitochondrial electron transport chain and coupled proton transport pathways amongst others Although infection of mice with NC-Nowra causes little or no signs of clinical disease, the molecular functions, processes and pathways identified through these studies clearly warrant further investigation for their-role in the development of protective Immunity as well as pathogenesis. These studies therefore provide new, exciting leads by which to study neosporosis. (C) 2010 Elsevier B.V. All rights reserved,
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| 39. |
- Ochaya, Stephen, et al.
(författare)
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Characterization of a Trypanosoma cruzi acetyltransferase : cellular location, activity and structure
- 2007
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 152:2, s. 123-131
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Tidskriftsartikel (refereegranskat)abstract
- Trypanosomatids are widespread parasites that cause three major tropical diseases. In trypanosomatids, as in most other organisms, acetylation is a common protein modification that is important in multiple, diverse processes. This paper describes a new member of the Trypanosoma cruzi acetyltransferase family. The gene is single copy and orthologs are also present in the other two sequenced trypanosomatids, Trypanosoma brucei and Leishmania major. This protein (TcAT-1) has the essential motifs present in members of the GCN5-related acetyltransferase (GNAT) family, as well as an additional motif also found in some enzymes from plant and animal species. The protein is evolutionarily more closely related to this group of enzymes than to histone acetyltransferases. The native protein has a cytosolic cellular location and is present in all three life-cycle stages of the parasite. The recombinant protein was shown to have autoacetylation enzymatic activity.
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| 40. |
- Palinauskas, Vaidas, et al.
(författare)
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Experimental evidence for hybridization of closely related lineages in Plasmodium relictum
- 2017
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Ingår i: Molecular and Biochemical Parasitology. - : Elsevier BV. - 0166-6851. ; 217, s. 1-6
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Tidskriftsartikel (refereegranskat)abstract
- Over 50 avian Plasmodium species have been described. However, PCR-based information shows much broader diversity of genetic lineages in these parasites. This discrepancy indicates insufficient knowledge about taxonomic diversity and boundaries of a single species in avian Plasmodium species. In recent taxonomy, most of genetically closely related lineages that share the same morphology and development patterns are attributed to the same biological species, but there is no information if these lineages are able to cross. This information is crucial to understand if these lineages form single or multiple evolutionary units. Due to presence of sexual process and sporogonic development of Plasmodium parasites in mosquitoes, self and cross-fertilization can occur and be identified during the oocyst stage. We initiated in vivo hybridization experiments of two widespread Plasmodium relictum lineages (pSGS1 and pGRW11) in experimentally infected Culex pipiens pipiens form molestus mosquitoes. To study putative hybrid oocysts, we used a laser microdissection technique together with PCR-based analyses of mitochondrial and nuclear genes. We demonstrate that both pSGS1 and pGRW11 lineages develop in infected mosquitoes in parallel, but also form hybrid oocysts of these two lineages. Our results are in accord to a recent global phylogeographic study of P. relictum that suggested that cross-fertilization between pSGS1 and pGRW11 might occur. This information helps to understand population structure, gene flow and the evolutionary process of haemosporidian parasites.
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| 41. |
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| 42. |
- Parussini, Fabiola, et al.
(författare)
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Characterization of a lysosomal serine carboxypeptidase from Trypanosoma cruzi.
- 2003
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Ingår i: Mol Biochem Parasitol. - 0166-6851. ; 131:1, s. 11-23
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Tidskriftsartikel (refereegranskat)abstract
- Trypanosoma cruzi, the flagellate protozoan which is the causative agent of the American trypanosomiasis, Chagas disease has carboxypeptidase activity. The enzyme has been purified to protein homogeneity, and shown to be a lysosomal monomeric glycoprotein with a molecular mass of about 54kDa. The enzyme has an optimum acidic pH (4.5 with furyl acryloyl-Phe-Phe as substrate), is highly specific for hydrophobic C-terminal amino acid residues, and is strongly inhibited by 3,4-dichloroisocoumarin (IC(50) value 0.3 microM). The enzyme is encoded by a number of genes arrayed in head-to-tail tandems; one of these genes has been cloned and sequenced. Sequence comparisons indicate that the enzyme belongs to the C group of serine carboxypeptidases, within the S10 serine peptidase family, and shows the higher similarity to plant and yeast enzymes. The residues involved in catalysis and most of those involved in substrate binding are conserved in the T. cruzi enzyme as well as 8 out of 10 Cys residues known to be involved in disulfide bridges in the yeast enzyme. This is the first report of an S10 family enzyme in trypanosomatids. The presence of serine carboxypeptidases is not restricted to T. cruzi, being possibly a general character of trypanosomatids.
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| 43. |
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| 44. |
- Raabe, Andreas C, et al.
(författare)
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Quantitative assessment of DNA replication to monitor microgametogenesis in Plasmodium berghei
- 2009
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier/North-Holland Biomedical Press. - 0166-6851 .- 1872-9428. ; 168:2, s. 172-176
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Tidskriftsartikel (refereegranskat)abstract
- Targeting the crucial step of Plasmodium transition from vertebrate host to mosquito vector is a promising approach to eliminate malaria. Uptake by the mosquito activates gametocytes within seconds, and in the case of male (micro) gametocytes leads to rapid DNA replication and the release of eight flagellated gametes. We developed a sensitive assay to monitor P. berghei microgametocyte activation based on [(3)H]hypoxanthine incorporation into DNA. Optimal pH range and xanthurenic acid concentrations for gametocyte activation were established and the kinetics of DNA replication investigated. Significance of the method was confirmed using P. berghei mutants and the assay was applied to analyse the effect of protease inhibitors, which revealed differences regarding their inhibitory action. The developed method thus appears suitable for reproducible determination of microgametocyte activation, medium-throughput drug screenings and deeper investigation of early blocks in gametogenesis and will facilitate the analysis of compounds for transmission blocking activities.
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| 45. |
- Rattanachuen, Woraphol, et al.
(författare)
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Probing the roles of non-homologous insertions in the N-terminal domain of Plasmodium falciparum hydroxymethylpterin pyrophosphokinase-dihydropteroate synthase
- 2009
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 168:2, s. 135-142
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Tidskriftsartikel (refereegranskat)abstract
- Plasmodium falciparum bifunctional hydroxymethylpterin pyrophosphokinase-dihydropteroate synthase (pfHPPK-DHPS) is a crucial enzyme in the de novo folate biosynthesis pathway. The crystal structure is not yet available for this enzyme, however, homology model of the enzyme reported previously revealed the presence of parasite-specific insertions. Alignment of pfHPPK-DHPS with HPPK and DHPS sequences from other microorganisms reveals two insertions relative to the corresponding enzyme in other organisms, i.e. HPPK-1 and HPPK-2. The former encompasses amino acid residues 66-162, while the latter covers residues 213-311. In order to investigate the roles of the two insertions, we constructed a number of mutants in which parts of these two insertions were deleted. Characterization of the mutationally altered proteins revealed that deletions of residues 74-80 in the HPPK-1 sequence of the pfHPPK-DHPS, but not that of the monofunctional pfHPPK, decreased the HPPK activity. A longer deletion (residues 74-86) in the HPPK-1 sequence of the bifunctional pfHPPK-DHPS completely inactivated both HPPK and DHPS activities. However, deletion in the HPPK-2 sequence from residues 247-306 did not disrupt the activities of HPPK and DHPS, but the kinetic properties of the recombinant proteins were slightly changed. The importance of HPPK-1 sequence on the catalytic activities of HPPK and DHPS in the bifunctional pfHPPK-DHPS could have implications for development of inhibitors targeting the non-catalytic region of this chemotherapeutically important enzyme.
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| 46. |
- Ribacke, Ulf, et al.
(författare)
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Genome wide gene amplifications and deletions in Plasmodium falciparum
- 2007
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 155:1, s. 33-44
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Tidskriftsartikel (refereegranskat)abstract
- The extent to which duplications and deletions occur in the Plasmodium falciparum genome, outside of the subtelomeres, and their contribution to the virulence of the malaria parasite is not known. Here we show the presence of multiple genome wide copy number polymorphisms (CNPs) covering 82 genes, the most extensive spanning a cumulative size of 110 kilobases. CNPs were identified in both laboratory strains and fresh clinical isolates using a 70-mer oligonucleotide microarray in conjunction with fluorescent in situ hybridizations and real-time quantitative PCR. The CNPs were found on all chromosomes except on chromosomes 6 and 8 and involved a total of 50 genes with increased copy numbers and 32 genes with decreased copy numbers relative to the 3D7 parasite. The genes, amplified in up to six copies, encode molecules involved in cell cycle regulation. cell division, drug resistance, erythrocyte invasion, sexual differentiation and unknown functions. These together with previous findings, suggest that the malaria parasite employs gene duplications and deletions as general strategies to enhance its survival and spread. Further analysis of the impact of discovered genetic differences and the underlying mechanisms is likely to generate a better understanding of the biology and the virulence of the malaria parasite.
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| 47. |
- Ringqvist, Emma, et al.
(författare)
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Release of metabolic enzymes by Giardia in response to interaction with intestinal epithelial cells
- 2008
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Ingår i: Molecular and biochemical parasitology (Print). - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 159:2, s. 85-91
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Tidskriftsartikel (refereegranskat)abstract
- Giardia lamblia, an important cause of diarrheal disease, resides in the small intestinal lumen in close apposition to epithelial cells. Since the disease mechanisms underlying giardiasis are poorly understood, elucidating the specific interactions of the parasite with the host epithelium is likely to provide clues to understanding the pathogenesis. Here we tested the hypothesis that contact of Giardia lamblia with intestinal epithelial cells might lead to release of specific proteins. Using established co-culture models, intestinal ligated loops and a proteomics approach, we identified three G. lamblia proteins (arginine deiminase, ornithine carbamoyl transferase and enolase), previously recognized as immunodominant antigens during acute giardiasis. Release was stimulated by cell-cell interactions, since only small amounts of arginine deiminase and enolase were detected in the medium after culturing of G. lamblia alone. The secreted G. lamblia proteins were localized to the cytoplasm and the inside of the plasma membrane of trophozoites. Furthermore, in vitro studies with recombinant arginine deiminase showed that the secreted Giardia proteins can disable host innate immune factors such as nitric oxide production. These results indicate that contact of Giardia with epithelial cells triggers metabolic enzyme release, which might facilitate effective colonization of the human small intestine.
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| 48. |
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| 49. |
- Tyden, Eva, et al.
(författare)
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Differential expression of beta-tubulin isotypes in different life stages of Parascaris spp after exposure to thiabendazole
- 2016
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Ingår i: Molecular and Biochemical Parasitology. - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 205, s. 22-28
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Tidskriftsartikel (refereegranskat)abstract
- Anthelmintic resistance (AR) to macrocyclic lactones (ML) has been described in Parascaris of horses world-wide. In contrast, benzimidazoles (BZ) are still effective, although reduced efficacy to this drug class was recently reported. The mode of action of BZ is binding to beta-tubulin, which prevents polymerisation of microtubules. In this study, beta-tubulin gene expression of isotypes 1 and 2 was investigated at seven time points (0, 6, 24, 72, 96 and 120h) during embryogenesis and in adult worms. In addition, an in ovo larval developmental test was developed to study beta-tubulin gene expression of both isotypes in parasacaris eggs after exposure to different concentrations of thiabendazole (TBZ) for five days at 25 degrees C. A strong pattern of differential expression of beta-tubulin and isotype 1 was observed in all stages, while isotype 2 expression was mainly found at an early phase of the embryogenesis. For isotype 1, a 5-fold increase was observed during the first 48 h, but gene expression gradually decreased after 72, 96 and 120 h. Isotype 2 was only expressed during the first 24 h, followed by a 130-fold decrease at (time points) 72, 96 and 120 h. The in ovo larval developmental test, in which we exposed initially unembryonated eggs to increased concentrations of TBZ, did affect isotype I gene expression but not isotype 2. This assumes that each isotype has specific functions in different life stages. This is in agreement with the 'multi-tubulin' hypothesis, which states that different tubulin isotypes are required for specialised microtubule functions. Isotype I is the most likely drug target for BZs, as isotype 2 was only expressed at very low levels later in development. Increasing concentrations of TBZ altered beta-tubulin isotype 1 gene expression after exposure of the eggs for five days, but this was not seen for isotype 2. (C) 2016 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
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| 50. |
- Tyden, Eva, et al.
(författare)
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Gene expression of ABC transporters in Cooperia oncophora after field and laboratory selection with macrocyclic lactones
- 2014
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Ingår i: Molecular and Biochemical Parasitology. - : Elsevier BV. - 0166-6851 .- 1872-9428. ; 198, s. 66-70
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Tidskriftsartikel (refereegranskat)abstract
- The most widespread helminth parasites of grazing cattle in northern Europe are the gastrointestinal nematodes Ostertagia ostertagi and Cooperia oncophora. Heavy reliance on the use of macrocyclic lactone (ML) in cattle has led to world-wide emergence of resistance to this drug class in C. oncophora. There is evidence that members of the ATP-binding cassette (ABC) transporter family, such as P-glycoproteins (P-gp) and multidrug-resistant proteins (MRP), play a role in resistance to ML In this study gene expression of Con-pgp9, Con-pgp11, Con-pgp12, Con-pgp16 and Con-mrp1 was examined in two isolates of C oncophora sharing the same genetic background but exposed to ML differently. For isolate one (Laboratory-selected), adult worms were recovered before and after treatment with ML in vivo. For isolate two (Field-selected), adult worms were collected from tracer animals that had never received anthelmintics themselves. One group grazed together with untreated animals and one group grazed with animals that received suppressive prophylactic treatment with ML at monthly intervals for up to two consecutive grazing seasons. Real-time PCR data demonstrated differences in gene expression after ML selection, with the highest constitutive expression levels for Con-pgp16 and Con-mrp1. Remarkably, the same pattern of increasing expression levels of the ABC transport genes was observed in both Laboratory- and Field-selected isolates, despite the Field-selected isolate not being directly exposed to ML The higher expression levels of ABC transporters observed in the Field-selected isolate was thus not a response to direct exposure to ML, but rather appeared to reflect a genetic characteristic inherited from worms in the previous generation which had survived exposure to ML in the co-grazing treated animals. (C) 2015 Elsevier B.V. All rights reserved.
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