Stem Cell Bank of Andalucia (Spanish Central Node), Hospital Universitario
Virgen de las Nieves, Avda Fuerzas Armadas, 2, 18014, Granada, Spain. fernancobo@fundacionhvn.org
The transplant of cells of human origin is an increasingly complex sector of
medicine which entails great opportunities for the treatment of a range of
diseases. Stem cell banks should assure
the quality, traceability and safety of cultures for transplantation and must
implement an effective programme
to prevent contamination of the final product. In donors, the presence of
infectious micro-organisms, like human immunodeficiency virus, hepatitis B
virus, hepatitis C virus and human T cell lymphotrophic
virus, should be evaluated in addition to the possibility of other new
infectious agents (e.g. transmissible spongiform encephalopathies
and severe acute respiratory syndrome). The introduction of the nucleic
acid amplification can avoid the window period of these viral infections.
Contamination from the laboratory
environment can be achieved by routine screening for bacteria, fungi, yeast and
mycoplasma by European pharmacopoeia tests. Fastidious micro-organisms, and an
adventitious or endogenous virus, is a well-known fact that will also have to be
considered for processes involving in vitro culture of stem cells. It is
also a standard part of current good practice in stem cell banks to carry out
routine environmental microbiological monitoring of the cleanrooms where the
cell cultures and their products are prepared.
The risk of viral contamination from
products of animal origin, like bovine serum and mouse fibroblasts as a "feeder
layer" for the development of embryonic cell lines, should also be considered.
Stem cell lines should be tested for prion
particles and a virus of animal origin that assure an acceptable quality.
Division of Infectious Diseases and Food Chain Quality, Animal Sciences Group,
Wageningen University and Research Centre (WUR), P.O. Box 65, Lelystad AB8200,
The Netherlands. adriaan.antonis@wur.nl
Veterinary vaccines are usually tested
for the absence of contaminants. However, the quality control does not always
imply that vaccines are not contaminated as, for example, illustrated by the
bovine herpes virus 1 (BHV1) vaccine used in The Netherlands in 1999 that
contained a small amount of bovine viral diarrhoea
virus (BVDV1).
Thousands of cows were vaccinated with BHV1 vaccine
batches, and the question arose as to whether these small amounts of BVDV1, most
likely not detected with in vitro tests, could have infected cattle. More in
general, the question was whether the outcome of the in vitro tests, i.e. the in
vitro infectivity, was indicative for the infectivity for cattle, i.e. the in
vivo infectivity. We therefore carried out in vitro experiments to determine the
sensitivity of a BVDV1 isolation assay. In addition, we performed two animal
experiments, in which we estimated the lowest dose needed to infect calves with
BVDV1. We extrapolated the experimental in vitro and in vivo results from a
tissue culture infectious dose (TCID50) to a cattle infectious dose (CID50). We
observed a partial response in the calves inoculated with this dose: four out of
six calves turned out to be infected. In the tissue culture test, all 20 samples
tested negative. The response in vivo, however, was not significantly higher
than the in vitro response, which implies that no difference in susceptibility
was observed between the animal test and the tissue culture test. Based on the
results in our experiments, some cattle may have been infected with BVDV1 after
the application of the contaminated BHV1 vaccine during the vaccination
campaign. The question remains that how
many cattle received contaminated vaccine, and became infected with BVDV1.
School of Humanities, Faculty of Arts, Education, and Social Sciences,
University of Western Sydney, Locked Bag 1797, South Penrith Distribution
Centre, NSW 1797, Australia. chris.wilson@uws.edu.au
Virological R&D Department, Intervet International b.v., Wim de Korverstraat 35,
NL-5831 AN, Boxmeer, The Netherlands. Birgit.Makoschey@Intervet.com
A protocol to test foetal calf serum (FCS) for contamination with bovine viral
diarrhoea virus (BVDV) is described. Following this protocol, which combines
cell culture methods and detection of pestivirus RNA, seven batches of FCS were
tested. Infectious BVDV
was detected in four of those batches. One of the remaining batches contained a
relatively high number of non-infectious BVDV
particles. A sample of this batch was formulated with aluminium
hydroxide and aluminium
phosphate as adjuvant into an experimental vaccine preparation.
This
product was injected twice into BVDV seronegative cattle with a 4 week interval.
Blood samples taken 4 weeks after the second application were negative for BVDV
specific antibodies. Our data stress that detection of BVDV RNA is not
sufficient for a complete risk assessment on FCS. Discrimination between
infectious and non-infectious BVDV is essential. This can only be achieved by
cell culture methods.
Istituto Superiore di Sanita, Laboratorio di Medicina Veterinaria, Viale Regina
Elena 299, 00161 Rome, Italy.
A non-cytopathic strain of BVDV-2 was isolated from a batch of live infectious
bovine rhinotracheitis (IBR) vaccine, and inoculated intranasally into four
3-month-old calves. Severe signs of disease developed by days 4 and 6 in three
of the calves, free of BVDV and antibodies to BVDV, that had been exposed to the
virus. These calves survived the acute phase of the infection and progressively
recovered. BVDV was consistently isolated, or the respective viral RNA was
detected, in the buffy coats from blood samples collected starting from days 2
or 4 up to days 11 or 14 after the experimental infection. Viral RNA was also
detected in sera from these infected calves until the presence in the serum of
virus neutralizing antibodies was demonstrated. By contrast, the only calf
having pre-existing neutralizing antibodies to BVDV at the start of the study
was protected from the disease. No virus was detected at any time after
experimental inoculation of this calf. Genomic characterization of the BVDV-2
isolated in cell cultures, or detected in sera from the experimentally infected
animals, revealed 100%, homology in the nucleotide sequence with the BVDV-2
detected as a contaminant of the live IBR virus vaccine.
These findings provided evidence of the
infective nature of the contaminant BVDV-2 and of its potential to generate
disease outbreaks when inoculated into susceptible animals.
Official Medicines Control Laboratory Biologika and R&D Unit, Division of
Biologicals, Swiss Federal Office of Public Health, P.O. Box 3003, Bern,
Switzerland.
In view of the use of potentially
contaminated foetal
calf serum (FCS)
in cell cultures pestiviruses
may be present in live viral vaccines. Thirty-six lots of human live viral
vaccines produced by three manufacturers were tested for the presence of pestiviruses.
Bovine viral diarrhoea
virus (BVDV)
RNA was detected in 33% of the vaccine lots. All positive results were
caused by the mumps component of a single manufacturer. Partial sequences of the
5' untranslated region of BVD viral RNA were determined. The sequences were
closely related to that of the NADL strain of BVDV. The amount of BVDV RNA in
the vaccines was determined by real-time RT-PCR using the LightCycler. Between
3.3*10(2) and 6.2*10(5) RNA copies per dose were found to be present in the
vaccine samples.Additionally, culture tests were done with FCS and human diploid
cells used in the vaccine production of the manufacturer whose vaccines were
positive by PCR. All attempts to detect virus antigen in MRC-5 human diploid
cells or to infect these cells with BVDV failed. This suggests that BVDV RNA
detected in human live viral vaccines represents passive carry over of BVDV from
contaminated FCS rather than active virus replication in human diploid cells.
Our results indicate that contamination with BVDV of FCS used in vaccine
production does not appear to be of immediate concern to human health.
Furthermore, our results indicate that gamma-irradiation of FCS destroys BVDV
particles and is also effective in preventing the presence of BVDV RNA in the
vaccines. Copyright 2002 The International Association for Biologicals.
Published by Elsevier Science Ltd. All rights reserved.
Department of Pharmacy Practice and Science, College of Pharmacy, University of
Arizona, 1703 E. Mabel Street, Tucson, AZ 85721-0207, USA. erstad@pharmacy.arizona.edu
The implications of prion-induced diseases for the use of medications that
theoretically could harbor the infectious pathogens are discussed. Prions have
been identified as protein particles that lack nucleic acids. There is evidence
that prions cause the transmissible neurodegenerative diseases known as
transmissible spongiform encephalopathies. Of these diseases, bovine spongiform
encephalopathy (BSE) and the human spongiform encephalopathy to which it has
been linked, new variant Creutzfeldt-Jakob disease (CJD), have generated the
most attention. The first cases of new variant CJD appeared in Britain in the
mid-1990s. Ingestion of prion-infected beef remains the only known cause of new
variant CJD. No cases of BSE or new variant CJD have been documented in the
United States. The time from exposure to the development of clinical sequelae
appears to be about 10 years. The median duration of illness is 14 months, and
the outcome is invariably death. There is no treatment; currently the only
available approach is prevention. There is no reliable method of predicting the
number of new cases that might occur because of lack of definitive information
on the efficiency of transmission from animals to humans and the number of
people currently infected and at risk for infection. The infectivity of
medications and plasma fractionation products containing material from cattle
with BSE is unknown, but the risk is believed to be very low. No cases of such
transmission have been identified. Guidelines to keep the risk of transmission
via medications low have been promulgated by FDA, and further research is
warranted. There have been no reports of medications or plasma fractionation
products being contaminated with the prions that cause new variant CJD. Ongoing
vigilance and research are appropriate, however.
Division of Neurology, Department of Medicine, Chiang Mai University, Chiang Mai
50200, Thailand. spiyasir@mail.med.cmu.ac.th
Public tolerance to adverse reactions is minimal. Several reporting systems have
been established to monitor adverse events following immunization. The present
review summarizes data on neurologic complications followingcination, and
provides evidence that indicates whether they were directly associated with the
vaccines. These complications include autism (measles vaccine), multiple
sclerosis (hepatitis B vaccine), meningoencephalitis (Japanese encephalitis
vaccine), Guillain-Barre syndrome and giant cell arteritis (influenza vaccine),
and reactions aftermaterials)] are
discussed.
Instituut voor Dierhouderij en Diergezondheid, ID Lelystad, Postbus 65, 8200 AB
Lelystad. c.j.m.bruschke@id.wag-ur.nl
In February 1999, 12 Dutch herds were vaccinated with a live bovine herpesvirus
1 vaccine from which bovine virus diarrhea virus (BVDV) could be isolated. All
vaccine batches that were on the Dutch market and that had not yet reached the
expiry date were tested for BVDV. In
total, seven of 82 batches tested were found positive. Batch numbers
TX3607, VB3914, VB3915, VB4046, TW3391, and TV3294 were positive for BVDV type
1, and batch number WG4622 was positive for BVDV type 2. This latter batch
induced clinical signs of BVDV in an animal experiment with susceptible animals.
Abteilung Anaesthesiologie und Operative Intensivmedizin Justus-Liebig-Universitat,
Giessen. Thilo.Menges@chiru.med.uni-giessen.de
The transmissible spongiform encephalopathies (TSE) are known to affect humans
and various animals. The bovine spongiform encephalopathy (BSE) and the human
Creutzfeldt-Jacob disease (CJD) are among the most notable degenerative
disorders caused by prions. Considering the BSE epidemic and the description of
a new variant of Creutzfeldt-Jacob disease (nvCJD), which is probably related to
bovine spongiform encephalopathy, TSE have recently gained a lot of public
attention. Although the causative factors (prions, viruses) are still under
discussion, none of the present concepts are explanatory for all aspects of the
human CJD. CJD may present as a sporadic, genetic, or infectious illness and
there is now considerable concern that bovine prions may have been passed to
humans. To exclude transmission of
CJD
via medical products and instruments, the effectiveness of cleaning,
disinfection and sterilization procedures must be firmly established.
This manuscript presents an overview to anaesthesiology and intensive care
medicine of recommended inactivation procedures and assessed these procedures in
the light of the inactivation of prions.
From the Centers for Disease Control and Prevention. Public
Health Service recommendations for the use of vaccines manufactured with
bovine-derived materials.
Laboratory of Pediatrics and Respiratory Viral Diseases, Center for Biologics
Evaluation and Research, U.S. Food and Drug Administration, Bethesda, Maryland
20892, USA.
Pestiviruses
are potential contaminants of biological products produced in bovine or porcine
cells or manufactured via processes using animal-derived raw materials such as
bovine serum. In order to investigate possible contamination of products
including those manufactured and/or licensed in the US, 38 lots of viral
vaccines and five lots of interferon alpha (IFNalpha) were tested by reverse
transcriptase polymerase chain reaction (RT-PCR) for the presence of bovine
viral diarrhoea virus (BVDV). All vaccines and interferons were negative for
contaminating BVDV RNA when tested by RT-PCR, with the exception of an
experimental live viral vaccine that had been produced in BVDV contaminated
rabbit kidney cells. Cell lines commonly used to produce biological products and
vaccines were experimentally infected with the NADL strain of BVDV to determine
if they were permissive for virus replication. MRC-5 and WI-38 cells were not
infected. In contrast, Vero, CHO and CEF cells showed evidence of pestivirus
infection. Taken together these data
suggested that currently licensed viral vaccines were unlikely to be
contaminated with pestiviruses.
However, cell banks derived from non-human primate, hamster or rabbit kidney
cell lines, or cultures of primary chick embryo fibroblasts, may be infected
with BVDV
if exposed to pestivirus
contaminated raw materials during manufacture.
USDA, APHIS, VS, Center for Veterinary Biologics-Laboratory, Ames, Iowa, USA.
Infection with bovine viral diarrhoea virus (BVDV) and other viruses is frequent
in the bovine population. In utero infection leads to virus and antibody
contamination of foetal and other serum used in cell culture production. The use
of contaminated cells for vaccine production may result in contaminated
vaccines, which may lead to seroconversion or disease in the vaccinated animal.
Contaminated serum or cell cultures may also interfere with the diagnosis of
viral infections. Methods for the detection of BVDV and other viruses in serum,
cell cultures, seed viruses and vaccines at the CVB-L, and the frequency of
detection are described. Reasons for continued use of serum in cell culture
production, and the risks of using serum, are discussed.
Bovine sera used in the manufacture of biologicals: current
concerns and policies of the U.S. Food and Drug Administration regarding the
transmissible spongiform encephalopathies.
Laboratory of Method Development, Division of Viral Products, Office of Vaccines
Research and Review, Center for Biologics Evaluation and Research, Food and Drug
Administration, Rockville, MD 20852-1448, USA.
Since 1993, consistent with its statutory responsibility to ensure that
regulated products are safe, pure, and free of << extraneous organisms, >> the
United States Food and Drug Administration (FDA) has requested that, with
certain exceptions, bovine-derived materials from animals born in or residing in
countries where bovine spongiform encephalopathy has occurred, should not be
used to manufacture products intended for humans. FDA's Center for Biologics
Evaluation and Research (CBER) has specifically recommended that serum used to
produce biologicals be obtained from sources << certified to be free from
contaminants and adventitious agents, such as the agent responsible for the
production of Bovine Spongiform Encephalopathy. >> The United States Department
of Agriculture (USDA) has prohibited importation of such serum for use in
products. FDA staff are aware that bovine blood, including foetal blood, and
placental tissues and fluids that might contaminate foetal serum have not been
found to contain the infectious agent of BSE, and that those tissues are
considered by most authorities to have little risk for transmitting disease to
humans or animals. However, studies of BSE have been limited in size and
sensitivity, and several experimental studies of scrapie and CJD in rodents
found their blood to be infectious. In addition, a recent unpublished study of
BSE (requiring confirmation) reported finding infectivity in the bone marrow of
cattle. Possible transmission of BSE from cows to calves, although unlikely to
constitute a major mode for maintaining the BSE outbreak, has also not been
rigorously ruled out. Considering the special nature of biological products,
especially of vaccines intended for widespread use in children, it seems prudent
for U.S. regulatory authorities to continue current conservative policies that
discourage or prohibit the use of bovine serum from countries with BSE.
National Veterinary Services Laboratories, Animal and Plant Health Inspection
Service, USDA, Ames, IA 50010.
Bovine viral diarrhea virus (BVDV) infection is common in the bovine population.
Infection in utero leads to virus and antibody contamination of the fetal bovine
serum used in cell cultures. These contaminants can interfere with diagnosis of
viral infection. The high frequency of
virus and antibody detection in individual animal or small pool samples suggests
that any large pool of unscreened sera will be contaminated. Infection of cell
cultures with BVDV
can lead to interference with the growth of other viruses. Vaccine produced on
contaminated cells may in turn be contaminated, leading to seroconversion
or disease in the vaccine. The safety, purity, and efficacy of viral
vaccines require BVDV testing of ingredients, cell substrates and final product.
Methods for detection of BVDV in nutrient serum, cell cultures, seed viruses,
and viral vaccines, and the frequency of their detection at the National
Veterinary Services Laboratories are discussed.
Regional Institute for Veterinary Health, Gouda, The Netherlands.
Bovine viral diarrhea virus (BVDV)
was isolated from 28 animals with a history of immunization against respiratory
disease with a vaccine contaminated with BVDV.
The vaccine-derived parental virus strain and the 28 isolates were analyzed
using 10 monoclonal antibodies (MAbs) directed against different epitopes and
antigenic domains on the major envelope glycoprotein of BVDV. None of the
isolates displayed a reaction pattern identical with the parental virus.
Instead, seven different reaction patterns (#A-G) emerged. Circumstantial
evidence indicated that six of these were vaccine related whereas in one case
(pattern #F) the origin of the isolate was unclear. The
results indicated that BVDV
rapidly changed during animal passages and that the tracing of the vaccine
contaminant using Mabs
was impossible.
National Veterinary Services Laboratories, Ames, IA 50010.
The National Veterinary Services Laboratories (NVSL) routinely monitors
continuous cell lines (CCL's) used for veterinary biologicals and diagnostic
virology. All veterinary biologicals produced in CCL's must follow the master
seed concept which limits the use of the master seed CCL to up to 20 passages
beyond the passage level characterized and deposited at NVSL. All CLL's are
evaluated for the presence of adventitious agents such as mycoplasma, bovine
viral diarrhea virus, and other bacteria and viruses. Previously, CCLs were
evaluated for tumorigenicity by the Syrian hamster cheek pouch method; however,
this procedure has now been eliminated.
The adventitious agents most frequently detected in CCL's
have been bovine viral diarrhea virus and mycoplasma. Our laboratory has
consistently found that the source of bovine viral diarrhea contamination of
CCLs has been the use of contaminated fetal bovine cell culture enrichment
serum. Gamma irradiation at 2.5-3.5 megarads at -40 degrees C of carefully
screened fetal bovine serum has been used in the Diagnostic Virology Laboratory
for over 10 years. If the irradiated serum is used at a final concentration of
10 percent, there is no untoward effect on cell susceptibility for virus
propagation or cell culture growth. Gamma irradiation has also been demonstrated
to be a very efficient inactivator of mycoplasma. Specific conditions utilized
by our laboratory to preserve fetal bovine serum cell culture growth factors
while eliminating adventitious bovine viral diarrhea virus will be presented.
DISCLAIMER: All
information, data, and material contained, presented, or provided here is for
general information purposes only and is not to be construed as reflecting the
knowledge or opinions of the publisher, and is not to be construed or intended
as providing medical or legal advice. The decision whether or not to vaccinate
is an important and complex issue and should be made by you, and you alone, in
consultation with your health care provider.
