TY - JOUR A1 - Abda, Ebrahim M. A1 - Krysciak, Dagmar A1 - Krohn-Molt, Ines A1 - Mamat, Uwe A1 - Schmeisser, Christel A1 - Förstner, Konrad U. A1 - Schaible, Ulrich E. A1 - Kohi, Thomas A. A1 - Nieman, Stefan A1 - Streit, Wolfgang R. T1 - Phenotypic Heterogeneity Affects Stenotrophomonas maltophilia K279a Colony Morphotypes and \(\beta\)-Lactamase Expression JF - Frontiers in Microbiology N2 - Phenotypic heterogeneity at the cellular level in response to various stresses, e.g., antibiotic treatment has been reported for a number of bacteria. In a clonal population, cell-to-cell variation may result in phenotypic heterogeneity that is a mechanism to survive changing environments including antibiotic therapy. Stenotrophomonas rnaltophilia has been frequently isolated from cystic fibrosis patients, can cause numerous infections in other organs and tissues, and is difficult to treat due to antibiotic resistances. S. maltophilia K279a produces the Li and L2 beta-lactamases in response to beta-lactam treatment. Here we report that the patient isolate S. rnaltophilia K279a diverges into cellular subpopulations with distinct but reversible morphotypes of small and big colonies when challenged with ampicillin. This observation is consistent with the formation of elongated chains of bacteria during exponential growth phase and the occurrence of mainly rod-shaped cells in liquid media. RNA-seq analysis of small versus big colonies revealed differential regulation of at least seven genes among the colony morphotypes. Among those, bleu and bla(L2) were transcriptionally the most strongly upregulated genes. Promoter fusions of b/a(L1) and b/a(L2) genes indicated that expression of both genes is also subject to high levels of phenotypic heterogeneous expression on a single cell level. Additionally, the comE homolog was found to be differentially expressed in homogenously versus heterogeneously bla(L2) expressing cells as identified by RNA(seq) analysis. Overexpression of cornE in S. maltophilia K279a reduced the level of cells that were in a bla(L2)-ON mode to 1% or lower. Taken together, our data provide strong evidence that S. maltophilia K279a populations develop phenotypic heterogeneity in an ampicillin challenged model. This cellular variability is triggered by regulation networks including b/a(L1), b/a(L2), and comE. KW - xanthomonas maltophilia KW - gram-negative bacteria KW - RNA-seq KW - pseudomas aeruginosa KW - antibiotic resistance KW - colony morphotypes KW - beta-lactamases KW - K279a KW - Stenotrophomonas maltophilia KW - phenotypic heterogeneity KW - persister cells KW - streptococcus pneumoniae KW - nosocomial pathogen KW - membrane vesicles KW - sinorhizobium fredii NGR234 KW - red fluorescent protein KW - escherichia coli Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-136446 VL - 6 IS - 1373 ER - TY - JOUR A1 - von Bohl, Andreas A1 - Kuehn, Andrea A1 - Simon, Nina A1 - Nkwouano Ngongang, Vanesa A1 - Spehr, Marc A1 - Baumeister, Stefan A1 - Przyborski, Jude M. A1 - Fischer, Rainer A1 - Pradel, Gabriele T1 - A WD40-repeat protein unique to malaria parasites associates with adhesion protein complexes and is crucial for blood stage progeny JF - Malaria Journal N2 - Background During development in human erythrocytes, Plasmodium falciparum parasites display a remarkable number of adhesive proteins on their plasma membrane. In the invasive merozoites, these include members of the PfMSP1 and PfAMA1/RON complexes, which facilitate contact between merozoites and red blood cells. In gametocytes, sexual precursor cells mediating parasite transmission to the mosquito vector, plasma membrane-associated proteins primarily belong to the PfCCp and 6-cys families with roles in fertilization. This study describes a newly identified WD40-repeat protein unique to Plasmodium species that associates with adhesion protein complexes of both merozoites and gametocytes. Methods The WD40-repeat protein-like protein PfWLP1 was identified via co-immunoprecipitation assays followed by mass spectrometry and characterized using biochemical and immunohistochemistry methods. Reverse genetics were employed for functional analysis. Results PfWLP1 is expressed both in schizonts and gametocytes. In mature schizonts, the protein localizes underneath the merozoite micronemes and interacts with PfAMA1, while in gametocytes PfWLP1 primarily accumulates underneath the plasma membrane and associates with PfCCp1 and Pfs230. Reverse genetics failed to disrupt the pfwlp1 gene, while haemagglutinin-tagging was feasible, suggesting a crucial function for PfWLP1 during blood stage replication. Conclusions This is the first report on a plasmodial WD40-repeat protein associating with cell adhesion proteins. Since WD40 domains are known to mediate protein–protein contact by serving as a rigid scaffold for protein interactions, the presented data suggest that PfWLP1 supports the stability of adhesion protein complexes of the plasmodial blood stages. KW - PfCCp protein KW - Pfs230 KW - PfAMA1 KW - WD40 KW - gametocyte KW - microneme KW - merozoite KW - plasmodium falciparum KW - malaria Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-139728 VL - 14 IS - 435 ER - TY - JOUR A1 - Sass, Andrea M. A1 - Van Acker, Heleen A1 - Förstner, Konrad U. A1 - Van Nieuwerburgh, Filip A1 - Deforce, Dieter A1 - Vogel, Jörg A1 - Coenye, Tom T1 - Genome-wide transcription start site profiling in biofilm-grown Burkholderia cenocepacia J2315 JF - BMC Genomics N2 - Background: Burkholderia cenocepacia is a soil-dwelling Gram-negative Betaproteobacterium with an important role as opportunistic pathogen in humans. Infections with B. cenocepacia are very difficult to treat due to their high intrinsic resistance to most antibiotics. Biofilm formation further adds to their antibiotic resistance. B. cenocepacia harbours a large, multi-replicon genome with a high GC-content, the reference genome of strain J2315 includes 7374 annotated genes. This study aims to annotate transcription start sites and identify novel transcripts on a whole genome scale. Methods: RNA extracted from B. cenocepacia J2315 biofilms was analysed by differential RNA-sequencing and the resulting dataset compared to data derived from conventional, global RNA-sequencing. Transcription start sites were annotated and further analysed according to their position relative to annotated genes. Results: Four thousand ten transcription start sites were mapped over the whole B. cenocepacia genome and the primary transcription start site of 2089 genes expressed in B. cenocepacia biofilms were defined. For 64 genes a start codon alternative to the annotated one was proposed. Substantial antisense transcription for 105 genes and two novel protein coding sequences were identified. The distribution of internal transcription start sites can be used to identify genomic islands in B. cenocepacia. A potassium pump strongly induced only under biofilm conditions was found and 15 non-coding small RNAs highly expressed in biofilms were discovered. Conclusions: Mapping transcription start sites across the B. cenocepacia genome added relevant information to the J2315 annotation. Genes and novel regulatory RNAs putatively involved in B. cenocepacia biofilm formation were identified. These findings will help in understanding regulation of B. cenocepacia biofilm formation. KW - persistence KW - genomic islands KW - pathogen KW - identification KW - bacteria KW - small RNAs KW - translation initiation KW - cepedia complex KW - global gene expression KW - SEQ KW - resistance KW - burkholderia cenocepacia KW - biofilms KW - dRNA-Seq KW - transcription start site KW - antisense RNA Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-139748 VL - 16 IS - 775 ER - TY - JOUR A1 - Fan, Ben A1 - Li, Lei A1 - Chao, Yanjie A1 - Förstner, Konrad A1 - Vogel, Jörg A1 - Borriss, Rainer A1 - Wu, Xiao-Qin T1 - dRNA-Seq Reveals Genomewide TSSs and Noncoding RNAs of Plant Beneficial Rhizobacterium Bacillus amyloliquefaciens FZB42 JF - PLoS One N2 - Bacillus amyloliquefaciens subsp. plantarum FZB42 is a representative of Gram-positive plant-growth-promoting rhizobacteria (PGPR) that inhabit plant root environments. In order to better understand the molecular mechanisms of bacteria-plant symbiosis, we have systematically analyzed the primary transcriptome of strain FZB42 grown under rhizospheremimicking conditions using differential RNA sequencing (dRNA-seq). Our analysis revealed 4,877 transcription start sites for protein-coding genes, identified genes differentially expressed under different growth conditions, and corrected many previously mis-annotated genes. We also identified a large number of riboswitches and cis-encoded antisense RNAs, as well as trans-encoded small noncoding RNAs that may play important roles in the gene regulation of Bacillus. Overall, our analyses provided a landscape of Bacillus primary transcriptome and improved the knowledge of rhizobacteria-host interactions. KW - gene expression KW - subtilis genome KW - enterica serovar thphimurium KW - small regulatory RNAs KW - binding protein HFQ KW - escherichia coli KW - messenger RNA KW - transcriptional landscape KW - mycobacterium tuberculosis KW - listeria monocytogenes Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-138369 VL - 10 IS - 11 ER - TY - THES A1 - Frank, Benjamin T1 - Untersuchungen zur Autophagieinduktion in Leishmania major-infizierten Knochenmarksmakrophagen T1 - Analyses of autophagy induction in Leishmania major-infected bone marrow-derived macrophages N2 - Die von der WHO zu den 17 wichtigsten NTDs gezählte Leishmaniose wird durch intrazelluläre Parasiten der Gattung Leishmania hervorgerufen. Der Lebenszyklus der Parasiten besteht aus zwei Phasen. Die länglichen und beweglichen Promastigoten kennzeichnen die Phase in der Sandmücke – der Vektor der Leishmaniose. Hingegen ist die Phase im Säugerwirt durch runde unbewegliche Amastigoten charakterisiert. Aufgrund des Mangels an potenten antileishmanialen Therapien wurde in der vorliegenden Arbeit die Interaktion zwischen L. m. Parasiten und der Hauptwirtszelle, der Makrophage, v. a. in Hinblick auf autophage Prozesse in den infizierten Makrophagen näher untersucht, um demgemäß neue Erkenntnisse zu gewinnen, welche bei der Herstellung zukünftiger anti-leishmanialer Medikamente helfen könnten. Bei der Autophagie handelt es sich um einen katabolen Prozess, wodurch Zellen bei Nahrungsmangel oder zellulärem Stress ihre Homöostase erhalten können. Durch diesen Prozess können überflüssige oder beschädigte Organellen recycelt werden, um die Funktionen der Zelle aufrechtzuerhalten. Daneben übernimmt Autophagie auch eine essenzielle Rolle bei der Abwehr von ins Zytosol eindringenden Pathogenen. Mittels des neu etablierten totalen Autophagiescore konnte festgestellt werden, dass Autophagie in L. m.-infizierten BMDM induziert wird. Die intrazellulären Amastigoten werden durch Autophagie in den BMDM verdaut. Die erhöhte autophage Aktivität konnte zudem durch Western-Blot-Analysen der autophagierelevanten Proteine ATG5, LC3B und UB bestätigt werden. Die molekulargenetischen Untersuchungen von L. m.-infizier-ten BMDM mithilfe von Affymetrix Microarrays führten zu einem Netzwerk aus autophagierelevanten und infektionsspezifischen Genen, welches als LISA bezeichnet worden ist. Hier hat sich ebenfalls eine starke Verknüpfung von autophagierelevanten Genen und den Genen der Glykolyse, einem zweiten katabolen Prozess, gezeigt. Zudem konnten zwei weitere autophagierelevante und infektionsspezifische Gene außerhalb von LISA identifiziert werden, nämlich Bnip3 und Ctse, welche im Anschluss genauer untersucht worden sind. Bei beiden Genen konnte auf Proteinebene gezeigt werden, dass sie in L. m.-infizierten BMDM signifikant erhöht sind. Durch siRNA-Analysen konnte überdies beobachtet werden, dass beide für die erfolgreiche Elimination der Amastigoten essenziell sind. Somit konnte mit den Proteinen BNIP3 und CTSE zwei potenzielle neue Ansatzpunkte für mögliche zukünftige antileishmaniale Therapien gefunden werden. Auch die in LISA enthaltenen Gene stellen prinzipiell vielversprechende Ziele für künftige Medikamente gegen Leishmaniose dar. Durch all diese Untersuchungen kommt man dem Ziel einer neuen, gezielten und nebenwirkungsärmeren Behandlung der Leishmaniose einen Schritt näher. N2 - Leishmaniasis, listed by the WHO to be one of the 17 most important NTDs, is caused by intracellular parasites of the genus Leishmania. The life cycle of the parasites consists of two stages. The oblong and motile promastigotes characterize the stage in the sand fly, the vector of leishmaniasis. However, the stage in the vertebrate host is characterized by round immotile amastigotes. Due to a lack of capable antileishmanial therapies, the interaction between L. m. parasites and their main host cell, the macrophage, was investigated in the present work, huge focus on autophagic processes in infected macrophages. Our goal was to get new insights for the future production of antileishmanial drugs. Autophagy is a catabolic process whereby cells are able to maintain their homeostasis in times of starvation or cellular stress. During to this process, redundant or damaged organelles are recycled in order to sustain cellular viability. Furthermore, autophagy has an essential role in the defense of pathogens invading the cytosol. The newly established total autophagy score showed an autophagy induction in L. m.-infected BMDM. Intracellular amastigotes are digested by autophagy in BMDM. The increased autophagic activity could also be confirmed by western-blot analyses of the autophagy-relevant proteins ATG5, LC3B, and UB. Molecular genetic investigations of L. m.-infected BMDM by Affymetrix microarrays led to a network of autophagy-relevant and infection-specific genes, which was called LISA. Additionally, it showed a strong connection between autophagy-relevant genes and genes of the glycolysis, a second catabolic process. Moreover, we identified and further characterized two additional autophagy-relevant genes, Bnip3 and Ctse, which were not included in LISA. Both genes were significantly overexpressed on protein level in L. m.-infected BMDM. By siRNA analyses we also demonstrated their importance for successful elimination of amastigotes. Therefore, both proteins, BNIP3 and CTSE, could be new potential targets for possible future antileishmanial therapies. In addition, the genes included in LISA might be promising targets for future drugs against leishmaniasis. Due to all these investigations we are one step closer to our goal of a targeted and safe therapy of leishmaniasis. KW - Autophagie KW - Leishmania major KW - Cathepsin E KW - BNIP3 KW - Elektronenmikroskopie KW - Autophagie KW - Leishmania major KW - Cathepsin E KW - BNIP3 KW - Elektronenmikroskopie KW - Microarray KW - siRNA KW - Autophagiescore Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-137277 ER - TY - JOUR A1 - Beiss, Veronique A1 - Spiegel, Holger A1 - Boes, Alexander A1 - Scheuermayer, Matthias A1 - Reimann, Andreas A1 - Schillberg, Stefan A1 - Fischer, Rainer T1 - Plant expression and characterization of the transmission-blocking vaccine candidate PfGAP50 JF - BMC Biotechnology N2 - Background: Despite the limited success after decades of intensive research and development efforts, vaccination still represents the most promising strategy to significantly reduce the disease burden in malaria endemic regions. Besides the ultimate goal of inducing sterile protection in vaccinated individuals, the prevention of transmission by so-called transmission blocking vaccines (TBVs) is being regarded as an important feature of an efficient malaria eradication strategy. Recently, Plasmodium falciparum GAP50 (PfGAP50), a 44.6 kDa transmembrane protein that forms an essential part of the invasion machinery (glideosome) multi-protein complex, has been proposed as novel potential transmission-blocking candidate. Plant-based expression systems combine the advantages of eukaryotic expression with a up-scaling potential and a good product safety profile suitable for vaccine production. In this study we investigated the feasibility to use the transient plant expression to produce PfGAP50 suitable for the induction of parasite specific inhibitory antibodies. Results: We performed the transient expression of recombinant PfGAP50 in Nicotiana benthamiana leaves using endoplasmatic reticulum (ER) and plastid targeting. After IMAC-purification the protein yield and integrity was investigated by SDS-PAGE and Western Blot. Rabbit immune IgG derived by the immunization with the plastidtargeted variant of PfGAP50 was analyzed by immune fluorescence assay (IFA) and zygote inhibition assay (ZIA). PfGAP50 could be produced in both subcellular compartments at different yields IMAC (Immobilized Metal Affinity Chromatography) purification from extract yielded up to 4.1 mu g/g recombinant protein per fresh leaf material for ER-retarded and 16.2 mu g/g recombinant protein per fresh leave material for plasmid targeted PfGAP50, respectively. IgG from rabbit sera generated by immunization with the recombinant protein specifically recognized different parasite stages in immunofluorescence assay. Furthermore up to 55 % inhibition in an in vitro zygote inhibition assay could be achieved using PfGAP50-specific rabbit immune IgG. Conclusions: The results of this study demonstrate that the plant-produced PfGAP50 is functional regarding the presentation of inhibitory epitopes and could be considered as component of a transmission-blocking malaria vaccine formulation. KW - PFS25 KW - plastid targeting KW - plant-made vaccines KW - agroinfiltration KW - gametes KW - sexual stage KW - plasmodium falciparum KW - membrane KW - antibodies KW - immunization KW - RTS,S/AS01 malaria vaccine KW - recombinant proteins KW - cost-effectiveness KW - purification Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-137327 VL - 15 IS - 108 ER - TY - JOUR A1 - Okoro, Chinyere K. A1 - Barquist, Lars A1 - Connor, Thomas R. A1 - Harris, Simon R. A1 - Clare, Simon A1 - Stevens, Mark P. A1 - Arends, Mark J. A1 - Hale, Christine A1 - Kane, Leanne A1 - Pickard, Derek J. A1 - Hill, Jennifer A1 - Harcourt, Katherine A1 - Parkhill, Julian A1 - Dougan, Gordon A1 - Kingsley, Robert A. T1 - Signatures of adaptation in human invasive Salmonella Typhimurium ST313 populations from sub-Saharan Africa JF - PLoS Neglected Tropical Diseases N2 - Two lineages of Salmonella enterica serovar Typhimurium (S. Typhimurium) of multi-locus sequence type ST313 have been linked with the emergence of invasive Salmonella disease across sub-Saharan Africa. The expansion of these lineages has a temporal association with the HIV pandemic and antibiotic usage. We analysed the whole genome sequence of 129 ST313 isolates representative of the two lineages and found evidence of lineage-specific genome degradation, with some similarities to that observed in S. Typhi. Individual ST313 S. Typhimurium isolates exhibit a distinct metabolic signature and modified enteropathogenesis in both a murine and cattle model of colitis, compared to S. Typhimurium outside of the ST313 lineages. These data define phenotypes that distinguish ST313 isolates from other S. Typhimurium and may represent adaptation to a distinct pathogenesis and lifestyle linked to an-immuno-compromised human population. KW - genome sequence KW - infection KW - pathogenicity KW - children KW - disease KW - adults KW - identification KW - Escherichia coli KW - virulence Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-143779 VL - 9 IS - 3 ER - TY - JOUR A1 - Berg, Stefan A1 - Schelling, Esther A1 - Hailu, Elena A1 - Firdessa, Rebuma A1 - Gumi, Balako A1 - Erenso, Girume A1 - Gadisa, Endalamaw A1 - Mengistu, Araya A1 - Habtamu, Meseret A1 - Hussein, Jemal A1 - Kiros, Teklu A1 - Bekele, Shiferaw A1 - Mekonnen, Wondale A1 - Derese, Yohannes A1 - Zinsstag, Jakob A1 - Ameni, Gobena A1 - Gagneux, Sebastien A1 - Robertson, Brian D A1 - Tschopp, Rea A1 - Hewinson, Glyn A1 - Yamuah, Lawrence A1 - Gordon, Stephen V A1 - Aseffa, Abraham T1 - Investigation of the high rates of extrapulmonary tuberculosis in Ethiopia reveals no single driving factor and minimal evidence for zoonotic transmission of Mycobacterium bovis infection JF - BMC Infectious Diseases N2 - Background: Ethiopia, a high tuberculosis (TB) burden country, reports one of the highest incidence rates of extra-pulmonary TB dominated by cervical lymphadenitis (TBLN). Infection with Mycobacterium bovis has previously been excluded as the main reason for the high rate of extra-pulmonary TB in Ethiopia. Methods: Here we examined demographic and clinical characteristics of 953 pulmonary (PTB) and 1198 TBLN patients visiting 11 health facilities in distinct geographic areas of Ethiopia. Clinical characteristics were also correlated with genotypes of the causative agent, Mycobacterium tuberculosis. Results: No major patient or bacterial strain factor could be identified as being responsible for the high rate of TBLN, and there was no association with HIV infection. However, analysis of the demographic data of involved patients showed that having regular and direct contact with live animals was more associated with TBLN than with PTB, although no M. bovis was isolated from patients with TBLN. Among PTB patients, those infected with Lineage 4 reported "contact with other TB patient" more often than patients infected with Lineage 3 did (OR = 1.6, CI 95% 1.0-2.7; p = 0.064). High fever, in contrast to low and moderate fever, was significantly associated with Lineage 4 (OR = 2.3; p = 0.024). On the other hand, TBLN cases infected with Lineage 4 tended to get milder symptoms overall for the constitutional symptoms than those infected with Lineage 3. Conclusions: The study suggests a complex role for multiple interacting factors in the epidemiology of extra-pulmonary TB in Ethiopia, including factors that can only be derived from population-based studies, which may prove to be significant for TB control in Ethiopia. KW - zoonotic KW - Mycobacterium KW - Ethiopia KW - tuberculosis KW - Bovis KW - pulmonary KW - extrapulmonary KW - lymphadenitis Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-143935 VL - 15 IS - 112 ER - TY - JOUR A1 - Rodriguez, Héctor A1 - Rico, Sergio A1 - Yepes, Ana A1 - Franco-Echevarría, Elsa A1 - Antoraz, Sergio A1 - Santamaría, Ramón I. A1 - Díaz, Margerita T1 - The two kinases, AbrC1 and AbrC2, of the atypical two-component system AbrC are needed to regulate antibiotic production and differentiation in Streptomyces coelicolor JF - Frontiers in Microbiology N2 - Two-component systems (TCSs) are the most important sensing mechanisms in bacteria. In Streptomyces, TCSs-mediated responses to environmental stimuli are involved in the regulation of antibiotic production. This study examines the individual role of two histidine kinases (HKs), AbrC1 and AbrC2, which form part of an atypical TCS in Streptomyces coelicolor. gRT-PCR analysis of the expression of both kinases demonstrated that both are expressed at similar levels in NB and NMMP media. Single deletion of abrC1 elicited a significant increase in antibiotic production, while deletion of abrC2 did not have any clear effect. The origin of this phenotype, probably related to the differential phosphorylation ability of the two kinases, was also explored indirectly, analyzing the toxic phenotypes associated with high levels of phosphorylated RR. The higher the AbrC3 regulator phosphorylation rate, the greater the cell toxicity. For the first time, the present work shows in Streptomyces the combined involvement of two different HKs in the response of a regulator to environmental signals. Regarding the possible applications of this research, the fact that an abrC1 deletion mutant overproduces three of the S. coelicolor antibiotics makes this strain an excellent candidate as a host for the heterologous production of secondary metabolites. KW - halstedii JM8 KW - biosynthesis KW - expression mutants KW - domain genes A3(2) KW - two-component systems KW - Streptomyces KW - antibiotic production KW - histidine kinases KW - heterologous production KW - activation KW - response regulator KW - PCR Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-143048 VL - 6 IS - 450 ER - TY - JOUR A1 - Afonso-Grunz, Fabian A1 - Hoffmeier, Klaus A1 - Müller, Sören A1 - Westermann, Alexander J. A1 - Rotter, Björn A1 - Vogel, Jörg A1 - Winter, Peter A1 - Kahl, Günter T1 - Dual 3'Seq using deepSuperSAGE uncovers transcriptomes of interacting Salmonella enterica Typhimurium and human host cells JF - BMC Genomics N2 - Background: The interaction of eukaryotic host and prokaryotic pathogen cells is linked to specific changes in the cellular proteome, and consequently to infection-related gene expression patterns of the involved cells. To simultaneously assess the transcriptomes of both organisms during their interaction we developed dual 3'Seq, a tag-based sequencing protocol that allows for exact quantification of differentially expressed transcripts in interacting pro-and eukaryotic cells without prior fixation or physical disruption of the interaction. Results: Human epithelial cells were infected with Salmonella enterica Typhimurium as a model system for invasion of the intestinal epithelium, and the transcriptional response of the infected host cells together with the differential expression of invading and intracellular pathogen cells was determined by dual 3'Seq coupled with the next-generation sequencing-based transcriptome profiling technique deepSuperSAGE (deep Serial Analysis of Gene Expression). Annotation to reference transcriptomes comprising the operon structure of the employed S. enterica Typhimurium strain allowed for in silico separation of the interacting cells including quantification of polycistronic RNAs. Eighty-nine percent of the known loci are found to be transcribed in prokaryotic cells prior or subsequent to infection of the host, while 75% of all protein-coding loci are represented in the polyadenylated transcriptomes of human host cells. Conclusions: Dual 3'Seq was alternatively coupled to MACE (Massive Analysis of cDNA ends) to assess the advantages and drawbacks of a library preparation procedure that allows for sequencing of longer fragments. Additionally, the identified expression patterns of both organisms were validated by qRT-PCR using three independent biological replicates, which confirmed that RELB along with NFKB1 and NFKB2 are involved in the initial immune response of epithelial cells after infection with S. enterica Typhimurium. KW - complete genome sequence KW - secretion systems KW - RNA-Seq KW - deepSuperSAGE KW - transcriptome KW - gene expression KW - serovar Typhimurium KW - human macrophages KW - epithelial cells KW - infection KW - SuperSAGE KW - receptors KW - Dual 3'seq KW - MACE KW - tag based KW - simultaneous KW - genome wide KW - gene expression profiling KW - host pathogen interaction KW - Salmonella enterica Typhimurium strain SL1344 Y1 - 2015 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-143230 VL - 16 IS - 323 ER -