Publications by authors named "Matthias Lenk"

13 Publications

  • Page 1 of 1

Evaluation of ELISA and VNT for sheeppox virus antibody detection and development of an immunoenzymatic quantitative method.

J Immunol Methods 2022 Jan 13;502:113226. Epub 2022 Jan 13.

Laboratory of Research and Development virology, MCI Animal Health, Lot. 157, Zone Industrielle Sud-Ouest (ERAC) B.P: 278, Mohammedia 28810, Morocco.

Vaccination against sheep pox (SPV) is the most efficient tool to control spread of the disease and virus neutralization test (VNT) is the gold standard for vaccination monitoring. In the presented study, we evaluated the use of ELISA and VNT for quantification of SPV humoral response post vaccination. Results confirmed that VNT is more sensitive since ELISA did not detect 22% of positive tested sera, and VNT weak positive sera were either negative or doubtful by ELISA. The most sensitive cells to perform VNT were ESH-L instead of Lamb primary cells. We also investigated immunoperoxidase IPMA and immunofluorescence IFA assays for detection of SPV specific antibodies and IPMA showed higher antibody titers comparatively to IFA. VNT using ESH-L cells with immune-enzymatic revelation provide specific quantitative SPV antibody titers, easier to read in shorter incubation time.
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http://dx.doi.org/10.1016/j.jim.2022.113226DOI Listing
January 2022

Replication of Rift Valley Fever Virus in Amphibian and Reptile-Derived Cell Lines.

Pathogens 2021 May 31;10(6). Epub 2021 May 31.

Institute of Novel and Emerging Infectious Diseases, Friedrich-Loeffler-Institut, Insel Riems, 17493 Greifswald, Germany.

Rift Valley fever phlebovirus (RVFV) is a zoonotic arthropod-borne virus, which has led to devastating epidemics in African countries and on the Arabian Peninsula. Results of in-vivo, in-vitro and field studies suggested that amphibians and reptiles may play a role as reservoir hosts of RVFV, promoting its maintenance during inter-epidemic periods. To elucidate this hypothesis, we examined two newly established reptile-derived cell lines (Egyptian cobra and Chinese pond turtle) and five previously generated reptile- and amphibian-derived cell lines for their replicative capacity for three low- and high-pathogenic RVFV strains. At different time points after infection, viral loads (TCID), genome loads and the presence of intracellular viral antigen (immunofluorescence) were assessed. Additionally, the influence of temperatures on the replication was examined. Except for one cell line (read-eared slider), all seven cell lines were infected by all three RVFV strains. Two different terrapin-derived cell lines (Common box turtle, Chinese pond turtle) were highly susceptible. A temperature-dependent replication of RVFV was detected for both amphibian and reptile cells. In conclusion, the results of this study indicate the general permissiveness of amphibian and reptile cell lines to RVFV and propose a potential involvement of terrapins in the virus ecology.
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http://dx.doi.org/10.3390/pathogens10060681DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8228813PMC
May 2021

Comparative sensitivity study of primary cells, vero, OA3.Ts and ESH-L cell lines to lumpy skin disease, sheeppox, and goatpox viruses detection and growth.

J Virol Methods 2021 07 14;293:114164. Epub 2021 Apr 14.

Laboratory of Research and Development virology, MCI Animal Health, Lot. 157, Zone Industrielle Sud-Ouest (ERAC) B.P: 278, 28810 Mohammedia, Morocco.

Lumpy skin disease virus (LSDV), sheeppox virus (SPPV) and goatpox (GTPV) virus have been usually grown on primary cells for diagnosis, production and titration purposes. The use of primary cells present several inconvenient, heavy preparation, heterogeneous cell population, non-reproducible viral titration and presence of potential endogenous contaminants. Therefore investigating sensitivity of candidate continuous cell lines is needed. In this study, we compared the above Capripox viruses (CaPVs) sensitivity of primary cells of four origin (heart, skin, testis and kidney), with three cell lines (Vero, OA3.Ts and ESH-L). We tested sensitivity for virus isolation, replication cycle and titration, revealed by cytopathic effect (CPE), immunoenzymatic staining and immunofluorescence. Our results show that ESH-L cells and primary fetal heart cells present the highest sensitivity for CaPVs growth and detection. Vero cells can replicate those viruses but without showing any CPE while the titer obtained on OA3.Ts is lower than primary and ESH-L cells. ESH-L cells are an effective alternative to primary cells use for growing Capripoxviruses and their diagnosis.
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http://dx.doi.org/10.1016/j.jviromet.2021.114164DOI Listing
July 2021

Common vole (Microtus arvalis) and bank vole (Myodes glareolus) derived permanent cell lines differ in their susceptibility and replication kinetics of animal and zoonotic viruses.

J Virol Methods 2019 12 9;274:113729. Epub 2019 Sep 9.

Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Novel and Emerging Infectious Diseases, Südufer 10, 17493 Greifswald - Insel Riems, Germany; German Center for Infection Research (DZIF), Partner site Hamburg-Lübeck-Borstel-Insel Riems, Germany. Electronic address:

Pathogenesis and reservoir host adaptation of animal and zoonotic viruses are poorly understood due to missing adequate cell culture and animal models. The bank vole (Myodes glareolus) and common vole (Microtus arvalis) serve as hosts for a variety of zoonotic pathogens. For a better understanding of virus association to a putative animal host, we generated two novel cell lines from bank voles of different evolutionary lineages and two common vole cell lines and assayed their susceptibility, replication and cytopathogenic effect (CPE) formation for rodent-borne, suspected to be rodent-associated or viruses with no obvious rodent association. Already established bank vole cell line BVK168, used as control, was susceptible to almost all viruses tested and efficiently produced infectious virus for almost all of them. The Puumala orthohantavirus strain Vranica/Hällnäs showed efficient replication in a new bank vole kidney cell line, but not in the other four bank and common vole cell lines. Tula orthohantavirus replicated in the kidney cell line of common voles, but was hampered in its replication in the other cell lines. Several zoonotic viruses, such as Cowpox virus, Vaccinia virus, Rift Valley fever virus, and Encephalomyocarditis virus 1 replicated in all cell lines with CPE formation. West Nile virus, Usutu virus, Sindbis virus and Tick-borne encephalitis virus replicated only in a part of the cell lines, perhaps indicating cell line specific factors involved in replication. Rodent specific viruses differed in their replication potential: Murine gammaherpesvirus-68 replicated in the four tested vole cell lines, whereas murine norovirus failed to infect almost all cell lines. Schmallenberg virus and Foot-and-mouth disease virus replicated in some of the cell lines, although these viruses have never been associated to rodents. In conclusion, these newly developed cell lines may represent useful tools to study virus-cell interactions and to identify and characterize host cell factors involved in replication of rodent associated viruses.
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http://dx.doi.org/10.1016/j.jviromet.2019.113729DOI Listing
December 2019

Profiling host ANP32A splicing landscapes to predict influenza A virus polymerase adaptation.

Nat Commun 2019 07 30;10(1):3396. Epub 2019 Jul 30.

Institute of Medical Virology, University of Zurich, Winterthurerstrasse 190, 8057, Zurich, Switzerland.

Species' differences in cellular factors limit avian influenza A virus (IAV) zoonoses and human pandemics. The IAV polymerase, vPol, harbors evolutionary sites to overcome restriction and determines virulence. Here, we establish host ANP32A as a critical driver of selection, and identify host-specific ANP32A splicing landscapes that predict viral evolution. We find that avian species differentially express three ANP32A isoforms diverging in a vPol-promoting insert. ANP32As with shorter inserts interact poorly with vPol, are compromised in supporting avian-like IAV replication, and drive selection of mammalian-adaptive vPol sequences with distinct kinetics. By integrating selection data with multi-species ANP32A splice variant profiling, we develop a mathematical model to predict avian species potentially driving (swallow, magpie) or maintaining (goose, swan) mammalian-adaptive vPol signatures. Supporting these predictions, surveillance data confirm enrichment of several mammalian-adaptive vPol substitutions in magpie IAVs. Profiling host ANP32A splicing could enhance surveillance and eradication efforts against IAVs with pandemic potential.
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http://dx.doi.org/10.1038/s41467-019-11388-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6667478PMC
July 2019

Detection of Herpesvirus anguillae (AngHV-1) in European eel Anguilla anguilla (L.) originating from northern Poland-assessment of suitability of selected diagnostic methods.

J Fish Dis 2017 Nov 24;40(11):1717-1723. Epub 2017 Aug 24.

Department of Meat Sciences, West Pomeranian University of Technology in Szczecin, Szczecin, Poland.

The Community Action Plan requests EU member states to implement measures that ensure the recovery of the severely depleted European eel stocks. One of the main threats is posed by Anguillid herpesvirus 1 (AngHV-1) leading to increased mortality in both wild and farmed eels. Following recommendations of the OIE to minimize the risk of obtaining false-negative results, the main aim of the study was to optimize diagnostic methods for AngHV-1 detection using conventional PCR, nested PCR and in situ hybridization assay. While 53.3% of the individual organ samples were tested positive for AngHV-1 by PCR, the additional virus analysis via nested PCR revealed that the actual prevalence was 93.3%. In the cell cultivation passages, a cytopathic effect was hardly found in the first two rounds. In the third passage onto cell cultures, a lytic CPE was detected. The identification and confirmation of the viruses obtained from cell cultures as well as directly from the organ tissues were proceeded by PCR, nested PCR and sequencing of the PCR products. While no positive signal was detectable in the first round by PCR using samples from the third cell culture passages, the nested PCR provided weak but visible positive signals.
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http://dx.doi.org/10.1111/jfd.12689DOI Listing
November 2017

A bovine cell line that can be infected by natural sheep scrapie prions.

PLoS One 2015 7;10(1):e0117154. Epub 2015 Jan 7.

Institute of Novel and Emerging Infectious Diseases at the Friedrich-Loeffler-Institut, Greifswald-Isle of Riems, Germany.

Cell culture systems represent a crucial part in basic prion research; yet, cell lines that are susceptible to prions, especially to field isolated prions that were not adapted to rodents, are very rare. The purpose of this study was to identify and characterize a cell line that was susceptible to ruminant-derived prions and to establish a stable prion infection within it. Based on species and tissue of origin as well as PrP expression rate, we pre-selected a total of 33 cell lines that were then challenged with natural and with mouse propagated BSE or scrapie inocula. Here, we report the successful infection of a non-transgenic bovine cell line, a sub-line of the bovine kidney cell line MDBK, with natural sheep scrapie prions. This cell line retained the scrapie infection for more than 200 passages. Selective cloning resulted in cell populations with increased accumulation of PrPres, although this treatment was not mandatory for retaining the infection. The infection remained stable, even under suboptimal culture conditions. The resulting infectivity of the cells was confirmed by mouse bioassay (Tgbov mice, Tgshp mice). We believe that PES cells used together with other prion permissive cell lines will prove a valuable tool for ongoing efforts to understand and defeat prions and prion diseases.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0117154PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4286239PMC
December 2015

More novel hantaviruses and diversifying reservoir hosts--time for development of reservoir-derived cell culture models?

Viruses 2014 Feb 26;6(3):951-67. Epub 2014 Feb 26.

Institute for Novel and Emerging Infectious Diseases, Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Südufer 10, 17493 Greifswald-Insel Riems, Germany.

Due to novel, improved and high-throughput detection methods, there is a plethora of newly identified viruses within the genus Hantavirus. Furthermore, reservoir host species are increasingly recognized besides representatives of the order Rodentia, now including members of the mammalian orders Soricomorpha/Eulipotyphla and Chiroptera. Despite the great interest created by emerging zoonotic viruses, there is still a gross lack of in vitro models, which reflect the exclusive host adaptation of most zoonotic viruses. The usually narrow host range and genetic diversity of hantaviruses make them an exciting candidate for studying virus-host interactions on a cellular level. To do so, well-characterized reservoir cell lines covering a wide range of bat, insectivore and rodent species are essential. Most currently available cell culture models display a heterologous virus-host relationship and are therefore only of limited value. Here, we review the recently established approaches to generate reservoir-derived cell culture models for the in vitro study of virus-host interactions. These successfully used model systems almost exclusively originate from bats and bat-borne viruses other than hantaviruses. Therefore we propose a parallel approach for research on rodent- and insectivore-borne hantaviruses, taking the generation of novel rodent and insectivore cell lines from wildlife species into account. These cell lines would be also valuable for studies on further rodent-borne viruses, such as orthopox- and arenaviruses.
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http://dx.doi.org/10.3390/v6030951DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3970132PMC
February 2014

Replicative Capacity of MERS Coronavirus in Livestock Cell Lines.

Emerg Infect Dis 2014 Feb;20(2):276-9

Replicative capacity of Middle East respiratory syndrome coronavirus (MERS-CoV) was assessed in cell lines derived from livestock and peridomestic small mammals on the Arabian Peninsula. Only cell lines originating from goats and camels showed efficient replication of MERS-CoV. These results provide direction in the search for the intermediate host of MERS-CoV.
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http://dx.doi.org/10.3201/eid2002.131182DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3901466PMC
February 2014

Identification of a gene for an ancient cytokine, interleukin 15-like, in mammals; interleukins 2 and 15 co-evolved with this third family member, all sharing binding motifs for IL-15Rα.

Immunogenetics 2014 Feb 26;66(2):93-103. Epub 2013 Nov 26.

Institute for Comprehensive Medical Science, Fujita Health University, Dengakugakubo 1-98, Toyoake, Aichi, 470-1192, Japan,

Interleukins 2 and 15 (IL-2 and IL-15) are highly differentiated but related cytokines with overlapping, yet also distinct functions, and established benefits for medical drug use. The present study identified a gene for an ancient third IL-2/15 family member in reptiles and mammals, interleukin 15-like (IL-15L), which hitherto was only reported in fish. IL-15L genes with intact open reading frames (ORFs) and evidence of transcription, and a recent past of purifying selection, were found for cattle, horse, sheep, pig and rabbit. In human and mouse the IL-15L ORF is incapacitated. Although deduced IL-15L proteins share only ~21 % overall amino acid identity with IL-15, they share many of the IL-15 residues important for binding to receptor chain IL-15Rα, and recombinant bovine IL-15L was shown to interact with IL-15Rα indeed. Comparison of sequence motifs indicates that capacity for binding IL-15Rα is an ancestral characteristic of the IL-2/15/15L family, in accordance with a recent study which showed that in fish both IL-2 and IL-15 can bind IL-15Rα. Evidence reveals that the species lineage leading to mammals started out with three similar cytokines IL-2, IL-15 and IL-15L, and that later in evolution (1) IL-2 and IL-2Rα receptor chain acquired a new and specific binding mode and (2) IL-15L was lost in several but not all groups of mammals. The present study forms an important step forward in understanding this potent family of cytokines, and may help to improve future strategies for their application in veterinarian and human medicine.
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http://dx.doi.org/10.1007/s00251-013-0747-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3894449PMC
February 2014

A novel porcine in vitro model of the blood-cerebrospinal fluid barrier with strong barrier function.

PLoS One 2012 22;7(6):e39835. Epub 2012 Jun 22.

Pediatric Infectious Diseases, Department of Pediatrics, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany.

Epithelial cells of the plexus choroideus form the structural basis of the blood-cerebrospinal fluid barrier (BCSFB). In vitro models of the BCSFB presenting characteristics of a functional barrier are of significant scientific interest as tools for examination of BCSFB function. Due to a lack of suitable cell lines as in vitro models, primary porcine plexus epithelial cells were subjected to a series of selective cultivation steps until a stable continuous subcultivatable epithelial cell line (PCP-R) was established. PCP-R cells grow in a regular polygonal pattern with a doubling time of 28-36 h. At a cell number of 1.5×10(5) in a 24-well plate confluence is reached in 56-72 h. Cells are cytokeratin positive and chromosomal analysis revealed 56 chromosomes at peak (84th subculture). Employing reverse transcription PCR mRNA expression of several transporters and components of cell junctions could be detected. The latter includes tight junction components like Claudin-1 and -3, ZO-1, and Occludin, and the adherens junction protein E-cadherin. Cellular localization studies of ZO-1, Occludin and Claudin-1 by immunofluorescence and morphological analysis by electron microscopy demonstrated formation of a dense tight junction structure. Importantly, when grown on cell culture inserts PCP-R developed typical characteristics of a functional BCSFB including high transepithelial electrical resistance above 600 Ω×cm(2) as well as low permeability for macromolecules. In summary, our data suggest the PCP-R cell line as a suitable in vitro model of the porcine BCSFB.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0039835PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3382175PMC
November 2012

Rapid characterisation of cell cultures by matrix-assisted laser desorption/ionisation mass spectrometric typing.

J Virol Methods 2010 Mar 24;164(1-2):116-21. Epub 2009 Nov 24.

Institute of Molecular Biology, Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Südufer 10, 17493 Greifswald-Insel Riems, Germany.

Misidentification or cross-contamination of cultured cell lines used for scientific or diagnostic purposes are a continuing challenge for laboratories and tissue culture repositories. Institutions dedicated to veterinary virology are particularly affected since the variety of viruses under investigation often requires the parallel maintenance of numerous different cell lines from different host species. To provide rapid but yet exact characterisation of cell cultures, matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) mass spectrometric typing was applied to 66 cell culture samples representing 34 species from insects to primates. A reference spectra library was generated that allows unambiguously the identification of all 66 cell lines. Spectrum-based phylogenetic analysis showed that clustering was mainly driven by taxonomy and allows the species determination of unknown samples.
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http://dx.doi.org/10.1016/j.jviromet.2009.11.022DOI Listing
March 2010

Functional analysis of the pseudorabies virus UL51 protein.

J Virol 2005 Mar;79(6):3831-40

Institute of Molecular Biology, Friedrich-Loeffler-Institut, Boddenblick 5A, Greifswald-Insel Riems D-17493, Germany.

Homologs of the UL51 protein of herpes simplex virus have been identified in all herpesvirus subfamilies, but until now, no function has been assigned to any of them. To investigate function of the UL51 gene product of the alphaherpesvirus pseudorabies virus (PrV), we isolated and analyzed a mutant lacking the major part of the open reading frame, PrV-DeltaUL51F, and a rescuant. One-step growth analysis of PrV-DeltaUL51F revealed only slightly reduced titers, but plaque size was notably diminished and reached only approximately 30% the plaque size of wild-type PrV. Ultrastructurally, intracytoplasmic capsids were found in large numbers either without envelope or in different stages of envelopment, indicating that secondary envelopment in the cytoplasm was less efficient. However, neuroinvasion in the mouse trigeminal pathway after intranasal infection was only slightly delayed. A PrV UL11 mutant also showed a defect in secondary envelopment (M. Kopp, H. Granzow, W. Fuchs, B. G. Klupp, E. Mundt, A. Karger, and T. C. Mettenleiter, J. Virol. 77:5339-5351, 2003). Since both proteins are part of the viral tegument and are predicted to be membrane associated, they may serve similar, possibly redundant functions during viral morphogenesis. Therefore, we also isolated a mutant simultaneously lacking UL51 and UL11. This mutant exhibited further reduced plaque size compared to the single-deletion mutants, but viral titers were comparable to those for the UL11 mutant. In electron microscopic analyses, the observed defect in secondary envelopment was similar to that found in the UL11 single-deletion mutant. In conclusion, both conserved tegument proteins, either singly or in combination, are involved in virion morphogenesis in the cytoplasm but are not essential for viral replication in vitro and in vivo.
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http://dx.doi.org/10.1128/JVI.79.6.3831-3840.2005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1075737PMC
March 2005
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