The bachelor thesis consists of the theoretical introduction into pressing the nuts, familiarization with used machines followed by testing and evaluating of outcomes. Pressing the nuts is tested for torque and pulling force. In the practical part of the thesis chosen press nuts will be tested. Objective of this thesis is to find out suitable pressing parameters, which can substitute present unsuitable practices

Daniel Růžek (351096)

Hana Tykalová (3551945)

John Fazakerley (3551942)

José de la Fuente (35632)

Julia Loecherbach (3551939)

Lesley Bell-Sakyi (33822)

Libor Grubhoffer (420445)

Margarita Villar (420749)

Marina Popara (525781)

Mick Watson (533792)

Sabine Weisheit (792366)

Parasites & Vectors

PubMed

TBEV infection levels in mock-infected and TBEV-infected tick cells. Numbers of copies of TBEV NS5 were determined by qRT-PCR using NS5 primers and the linearised plasmid pJET-NS5 to create a standard curve. Copy numbers were normalised to 1 μg of total RNA. The limit of detection was derived from the number of NS5 copies in the highest dilution which was still detectable with a variance less than one Ct and was normalised to 1 μg of total RNA. (A) IDE8 infected and mock-infected (control) at days 2 (2d) and 6 (6d) p.i. (B) IRE/CTVM19 infected and mock-infected (control) at days 2 and 6 p.i.. Error bars are standard deviations. Samples marked with + passed both RNA and protein quality checks and were used in transcriptomic and proteomic analyses

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Additional file 3: of 
                     Ixodes scapularis and Ixodes ricinus tick cell lines respond to infection with tick-borne encephalitis virus: transcriptomic and proteomic analysis

BACKGROUND: Ixodid ticks are important vectors of a wide variety of viral, bacterial and protozoan pathogens of medical and veterinary importance. Although several studies have elucidated tick responses to bacteria, little is known about the tick response to viruses. To gain insight into the response of tick cells to flavivirus infection, the transcriptomes and proteomes of two Ixodes spp cell lines infected with the flavivirus tick-borne encephalitis virus (TBEV) were analysed. METHODS: RNA and proteins were isolated from the Ixodes scapularis-derived cell line IDE8 and the Ixodes ricinus-derived cell line IRE/CTVM19, mock-infected or infected with TBEV, on day 2 post-infection (p.i.) when virus production was increasing, and on day 6 p.i. when virus production was decreasing. RNA-Seq and mass spectrometric technologies were used to identify changes in abundance of, respectively, transcripts and proteins. Functional analyses were conducted on selected transcripts using RNA interference (RNAi) for gene knockdown in tick cells infected with the closely-related but less pathogenic flavivirus Langat virus (LGTV). RESULTS: Differential expression analysis using DESeq resulted in totals of 43 and 83 statistically significantly differentially-expressed transcripts in IDE8 and IRE/CTVM19 cells, respectively. Mass spectrometry detected 76 and 129 statistically significantly differentially-represented proteins in IDE8 and IRE/CTVM19 cells, respectively. Differentially-expressed transcripts and differentially-represented proteins included some that may be involved in innate immune and cell stress responses. Knockdown of the heat-shock proteins HSP90, HSP70 and gp96, the complement-associated protein Factor H and the protease trypsin resulted in increased LGTV replication and production in at least one tick cell line, indicating a possible antiviral role for these proteins. Knockdown of RNAi-associated proteins Argonaute and Dicer, which were included as positive controls, also resulted in increased LGTV replication and production in both cell lines, confirming their role in the antiviral RNAi pathway. CONCLUSIONS: This systems biology approach identified several molecules that may be involved in the tick cell innate immune response against flaviviruses and highlighted that ticks, in common with other invertebrate species, have other antiviral responses in addition to RNAi

Weisheit, S

Villar, M

Tykalova, H

Popara, M

Loecherbach, J

Watson, M

Ruzek, D

Grubhoffer, L

de la Fuente, J

Fazakerley, JK

Bell-Sakyi, L

University of Melbourne Institutional Repository

Ixodes scapularis and Ixodes ricinus tick cell lines respond to infection with tick-borne encephalitis virus: transcriptomic and proteomic analysis

Drápal, Michal

National Repository of Grey Literature

Pressing the Nuts to Sheet Metal Blanks

Edinburgh Research Explorer

Abstract Background Ixodid ticks are important vectors of a wide variety of viral, bacterial and protozoan pathogens of medical and veterinary importance. Although several studies have elucidated tick responses to bacteria, little is known about the tick response to viruses. To gain insight into the response of tick cells to flavivirus infection, the transcriptomes and proteomes of two Ixodes spp cell lines infected with the flavivirus tick-borne encephalitis virus (TBEV) were analysed. Methods RNA and proteins were isolated from the Ixodes scapularis-derived cell line IDE8 and the Ixodes ricinus-derived cell line IRE/CTVM19, mock-infected or infected with TBEV, on day 2 post-infection (p.i.) when virus production was increasing, and on day 6 p.i. when virus production was decreasing. RNA-Seq and mass spectrometric technologies were used to identify changes in abundance of, respectively, transcripts and proteins. Functional analyses were conducted on selected transcripts using RNA interference (RNAi) for gene knockdown in tick cells infected with the closely-related but less pathogenic flavivirus Langat virus (LGTV). Results Differential expression analysis using DESeq resulted in totals of 43 and 83 statistically significantly differentially-expressed transcripts in IDE8 and IRE/CTVM19 cells, respectively. Mass spectrometry detected 76 and 129 statistically significantly differentially-represented proteins in IDE8 and IRE/CTVM19 cells, respectively. Differentially-expressed transcripts and differentially-represented proteins included some that may be involved in innate immune and cell stress responses. Knockdown of the heat-shock proteins HSP90, HSP70 and gp96, the complement-associated protein Factor H and the protease trypsin resulted in increased LGTV replication and production in at least one tick cell line, indicating a possible antiviral role for these proteins. Knockdown of RNAi-associated proteins Argonaute and Dicer, which were included as positive controls, also resulted in increased LGTV replication and production in both cell lines, confirming their role in the antiviral RNAi pathway. Conclusions This systems biology approach identified several molecules that may be involved in the tick cell innate immune response against flaviviruses and highlighted that ticks, in common with other invertebrate species, have other antiviral responses in addition to RNAi

Sabine Weisheit

Margarita Villar

Hana Tykalová

Marina Popara

Julia Loecherbach

Mick Watson

Daniel Růžek

Libor Grubhoffer

José de la Fuente

John K. Fazakerley

Lesley Bell-Sakyi

Directory of Open Access Journals

This article is distributed under the terms of the Creative Commons Attribution 4.0
International License.-- et al.[Background]: Ixodid ticks are important vectors of a wide variety of viral, bacterial and protozoan pathogens of medical and veterinary importance. Although several studies have elucidated tick responses to bacteria, little is known about the tick response to viruses. To gain insight into the response of tick cells to flavivirus infection, the transcriptomes and proteomes of two Ixodes spp cell lines infected with the flavivirus tick-borne encephalitis virus (TBEV) were analysed. [Methods]: RNA and proteins were isolated from the Ixodes scapularis-derived cell line IDE8 and the Ixodes ricinus-derived cell line IRE/CTVM19, mock-infected or infected with TBEV, on day 2 post-infection (p.i.) when virus production was increasing, and on day 6 p.i. when virus production was decreasing. RNA-Seq and mass spectrometric technologies were used to identify changes in abundance of, respectively, transcripts and proteins. Functional analyses were conducted on selected transcripts using RNA interference (RNAi) for gene knockdown in tick cells infected with the closely-related but less pathogenic flavivirus Langat virus (LGTV). [Results]: Differential expression analysis using DESeq resulted in totals of 43 and 83 statistically significantly differentially-expressed transcripts in IDE8 and IRE/CTVM19 cells, respectively. Mass spectrometry detected 76 and 129 statistically significantly differentially-represented proteins in IDE8 and IRE/CTVM19 cells, respectively. Differentially-expressed transcripts and differentially-represented proteins included some that may be involved in innate immune and cell stress responses. Knockdown of the heat-shock proteins HSP90, HSP70 and gp96, the complement-associated protein Factor H and the protease trypsin resulted in increased LGTV replication and production in at least one tick cell line, indicating a possible antiviral role for these proteins. Knockdown of RNAi-associated proteins Argonaute and Dicer, which were included as positive controls, also resulted in increased LGTV replication and production in both cell lines, confirming their role in the antiviral RNAi pathway. [Conclusions]: This systems biology approach identified several molecules that may be involved in the tick cell innate immune response against flaviviruses and highlighted that ticks, in common with other invertebrate species, have other antiviral responses in addition to RNAi.SW and MP were Early Stage Researchers supported by the POSTICK ITN (Post-graduate training network for capacity building to control ticks and tick-borne diseases) within the FP7- PEOPLE – ITN programme (EU Grant No. 238511). The research was supported by: grants BFU2011-23896 from the Ministerio de Economía y Competitividad, Spain and the European Union FP7 ANTIGONE project number 278976 (MV, JF); the Czech Science Foundation (GACR) (15-03044S) and by Institutional support RVO: 60077344 from Biology Centre ASCR, Institute of Parasitology (HT, LG); the Czech Science Foundation (projects nos P502/11/2116 and GA14-29256S) and the MEYS of the Czech Republic (project LO1218) under the NPU I programme (DR); the United Kingdom Biotechnology and Biological Sciences Research Council’s National Capability Grant to the Pirbright Institute (LBS).Peer Reviewe

Weisheit, Sabine

Villar, Margarita

Popara, Marina

Fuente, José de la

Bell-Sakyi, Lesley

Digital.CSIC

Discussion of additional transcripts and proteins that may be involved in innate immunity and cell stress [154-181].(DOCX 16 kb

Additional file 6: of 
                     Ixodes scapularis and Ixodes ricinus tick cell lines respond to infection with tick-borne encephalitis virus: transcriptomic and proteomic analysis

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Pressing the Nuts to Sheet Metal Blanks

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