Adaptation of a Madin–Darby canine kidney cell line to suspension growth in serum-free media and comparison of its ability to produce avian influenza virus to Vero and BHK21 cell lines

Wielink, R. van; Kant-Eenbergen, H.C.M.; Harmsen, Michiel M.; Martens, Dirk E.; Wijffels, René H.; Coco-Martin, J.M.

doi:10.1016/j.jviromet.2010.09.029

Cited by 43 publications

(39 citation statements)

References 28 publications

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“…15,22,47 The SFM-adapted MDCK cells used in the present study clearly have higher SA 2,6 and lower SA 2,3 receptor levels than traditional MDCK cells. In contrast to the SFM-adapted MDCK cells, the allantoic cells of ECE are known to be rich in SA 2,3 but low in SA 2,6, 22 which likely contributes to the improved recovery of swine-origin FLUAV strains with SFM-adapted MDCK cells.…”

Section: Discussionmentioning

confidence: 58%

Comparative effectiveness of isolation techniques for contemporary Influenza A virus strains circulating in exhibition swine

et al. 2012

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Abstract. The current study sought to compare the effectiveness of 2 virus isolation methods for the recovery of contemporary Influenza A virus (FLUAV) strains circulating in swine at agricultural exhibitions. Following the emergence of the influenza A (H1N1)pdm09 virus, increased surveillance of FLUAV strains among swine was recommended for early detection of emerging strains that threaten animal and human health. The increase in genetic drift and genomic reassortment among FLUAV strains infecting swine since 1998 necessitates that detection protocols be periodically validated for contemporary FLUAV strains. During 2009, nasal swabs were collected from 221 clinically healthy pigs at 12 agricultural exhibitions in Ohio. Nasal swabs were tested in parallel for the presence of FLUAV strains using 3 methodologies: 2 passages through Madin-Darby canine kidney (MDCK) cells adapted to serum-free medium (SFM), 2 passages through embryonated chicken eggs (ECEs), and real-time reverse transcription polymerase chain reaction (real-time RT-PCR). Of the 221 samples, 40 (18.1%) were positive for FLUAV recovery in MDCK cell culture and 13 (5.9%) were positive in ECEs (P = 0.015). All samples positive in ECEs were also positive in MDCK cell culture. MDCK cell culture virus isolation results were in perfect agreement with results of the realtime RT-PCR. Hemagglutinin and neuraminidase combinations of the recovered isolates were H1N2 and H3N2, which were consistent with FLUAV strains circulating in U.S. pigs. Effectiveness and cost savings justify the use of SFM-adapted MDCK cell culture over ECEs for the recovery of contemporary FLUAV strains from exhibition swine.

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Section: Discussionmentioning

confidence: 58%

Comparative effectiveness of isolation techniques for contemporary Influenza A virus strains circulating in exhibition swine

et al. 2012

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“…MDCK-SFS (serum-free suspension) cells (47) were grown in suspension in SFM4BHK21 medium (HyClone, Waltham, MA), supplemented with 8 mM glutamine, 5 mg/liter phenol red, and 1.5 g/liter sodium bicarbonate, or were grown adherent in serum-free UltraMDCK medium (Lonza Biowhittaker, Basel, Switzerland) supplemented with 4 mM glutamine. Adherent NS1Bon2 MDCK cells (46) were also grown in UltraMDCK medium, additionally supplemented with 200 g/ml G418 (Promega, Fitchburg, WI) and 100 g/ml hygromycin B (Clontech, Mountain View, CA).…”

Section: Methodsmentioning

confidence: 99%

“…Note that in the comparison of reassortant virus replication, protein expression, apoptosis, and IFN induction, a nonadapted delNS1 virus strain was used, to which we refer as delNS1. The infectious influenza virus titer was measured by determining the tissue culture infective dose required to infect 50% (TCID 50 ) of MDCK cells, as previously described (47). All virus strains were propagated on the NS1-expressing NS1Bon2 MDCK cell line to generate virus seed stocks with high infectious virus titers (Ͼ7 log 10 TCID 50 /ml) prior to further viral characterization.…”

Section: Methodsmentioning

confidence: 99%

Mutations in the M-Gene Segment Can Substantially Increase Replication Efficiency of NS1 Deletion Influenza A Virus in MDCK Cells

Wielink

Harmsen

Martens

et al. 2012

J Virol

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bInfluenza viruses unable to express NS1 protein (delNS1) replicate poorly and induce large amounts of interferon (IFN). They are therefore considered candidate viruses for live-attenuated influenza vaccines. Their attenuated replication is generally assumed to result from the inability to counter the antiviral host response, as delNS1 viruses replicate efficiently in Vero cells, which lack IFN expression. In this study, delNS1 virus was parallel passaged on IFN-competent MDCK cells, which resulted in two strains that were able to replicate to high virus titers in MDCK cells due to adaptive mutations especially in the M-gene segment but also in the NP and NS gene segments. Most notable were clustered U-to-C mutations in the M segment of both strains and clustered A-to-G mutations in the NS segment of one strain, which presumably resulted from host cell-mediated RNA editing. The M segment mutations in both strains changed the ratio of M1 to M2 expression, probably by affecting splicing efficiency. In one virus, 2 amino acid substitutions in M1 additionally enhanced virus replication, possibly through changes in the M1 distribution between the nucleus and the cytoplasm. Both adapted viruses induced levels of IFN equal to that of the original delNS1 virus. These results show that the increased replication of the adapted viruses is not primarily due to altered IFN induction but rather is related to changes in M1 expression or localization. The mutations identified in this paper may be used to enhance delNS1 virus replication for vaccine production.

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“…The cells can be easily adapted to and be grown in serum-free media, and in suspension, as well as on various microcarriers maintained under various bioreactor conditions. 93,[101][102][103][104][105][106][107][108] Subclones of MDCK cells adopted to grow in suspension and support robust virus production have also been described, 91,108,109 although adherent MDCK cells appear to support more robust virus production than suspension MDCK cells. 110 Influenza vaccines derived from MDCK cells are also safe and immunogenic.…”

Section: Mdck Cell-derived Influenza Vaccines: the Final Joust?mentioning

confidence: 99%

Cell culture-based influenza vaccines: A necessary and indispensable investment for the future

Hegde

2015

Human Vaccines & Immunotherapeutics

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Adaptation of a Madin–Darby canine kidney cell line to suspension growth in serum-free media and comparison of its ability to produce avian influenza virus to Vero and BHK21 cell lines

Cited by 43 publications

References 28 publications

Comparative effectiveness of isolation techniques for contemporary Influenza A virus strains circulating in exhibition swine

Comparative effectiveness of isolation techniques for contemporary Influenza A virus strains circulating in exhibition swine

Mutations in the M-Gene Segment Can Substantially Increase Replication Efficiency of NS1 Deletion Influenza A Virus in MDCK Cells

Cell culture-based influenza vaccines: A necessary and indispensable investment for the future

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