- From: www.whale.to/a/nanobacteria.html
-
- Fetal Bovine Serum - Varicella Vax, some oral polio vax,
Merck reported it was in MMR to Ray Gallup , and in the former Rotavirus
vaccine.......
-
- "As reported May 23rd, 2001at the 101st General
Meeting of the American Society for Microbiology, Nanobacteria has been
found to be a contaminant in previously assumed-to-be-sterile medical products,
specifically IPV Polio Vaccine. Most human biologicals and vaccines are
made in fetal bovine serum, a medium that is known to be contaminated with
nanobacteria. In order to prevent this problem in the future, human biological
products must be made in Nano-Free Culture medium (filtered first through
20 nanometer filters,Gamma-Irradiated with 150 megarads and then heated
to 90 degrees Centigrade for at least an hour to kill any nanobacteria
present)"
-
- http://www.nanobaclabs.com/PageDisplay.asp?p1=6578
-
- excerpts......
-
- "The term 'Nanobacteria' is short for it's scientific
genus and species name 'Nanobacterium sanguineum' - a Latin scientific
term for blood nanobacteria. Nanobacteria are 'nano'-sized in that they
are from 20-200 nanometers in size (a nanometer is 1 billionth of a meter.
A nanometer is the width of ten hydrogen atoms side-to-side!) and are the
smallest known self-replicating bacteria. They are from the Archaea Family
of bacteria, known for their primitive pleomorphic lifestyles."
-
- "Nanobacteria infection by Nanobacterium sanguineum
is an 'emerging infectious disease' meaning that it is newly-discovered
and that the diseases it cause are being researched and further described.
Its DNA, RNA and Lipopolysaccaride profiles have been accurately mapped
by multiple scientific researchers at many universities worldwide. Nanobacteria
are not nice bugs and have absolutely no known positive benefits to humans.
The discoverers of nanobacteria, Drs. Ciftcioglu & Kajander developed
antigen and antibody diagnostic blood testing for nanobacterial infections
that we offer as the "NanobacTEST". NanobacLabs has developed
safe and effective nanobiotic prescription treatments"
-
- "Nanobacteria are extremely small, slowly growing
bacteria that can be cultured from the blood of humans and mammals. Their
size is 20-200 nanometers....when compared to 'regular' bacteria, Nanobacteria
are 1/100 to 1/1,000 the size, allowing them to easily move around into
other cells and invade them. Nanobacteria cause apoptosis (cell death)
when exposed to human cells or other bacteria. They can cause alteration
of RNA and DNA gene-expression patterns of cells they infect.....this can
lead to genetic alteration, abnormal cell growth and proliferation. When
compared to other bacteria, Nanobacteria grow very, very slowly, only reproducing
every 3 days..where 'regular' bacteria reproduce in minutes or hours. Nanobacteria
cannot be grown in standard culture media and can only be grown in mammalian
blood or serum.
-
- Nanobacteria were discovered in 1988 by Nobel Prize Nominees
Dr. Neva Ciftcioglu, PhD and Olavi Kajander, MD, PhD as a 'contaminant'
in mammalian cell cultures that kept killing the mammalian cells (apoptosis)
in their mammalian cell culture research. They have been researching nanobacterial
pathophysiology for nearly 14 years now and are the worldwide experts on
nanobacterial basic science. They are currently guiding and teaching researchers
all over the world. NanobacLabs is the world leader in the research and
development of prescription NANOBIOTICS that eradicate pathological calcification
and nanobacterial infections"
-
- "As reported May 23rd, 2001 at the 101st General
Meeting of the American Society for Microbiology, Nanobacteria has been
found to be a contaminant in previously assumed-to-be-sterile medical products,
specifically IPV Polio Vaccine. Most human biologicals and vaccines are
made in fetal bovine serum, a medium that is known to be contaminated with
nanobacteria. In order to prevent this problem in the future, human biological
products must be made in Nano-Free Culture medium (filtered first through
20 nanometer filters,Gamma-Irradiated with 150 megarads and then heated
to 90 degrees Centigrade for at least an hour to kill any nanobacteria
present)."
-
-
- Nanobacteria Threat To Blood Supply And Blood Products
-
- http://www.uku.fi/~kajander/ http://www.uku.fi/~kajander/fatal.html
-
- Mol. Biol. Cell, Suppl., Vol. 7, (1996): 517a Fatal (fetal)
bovine serum: discovery of Nanobacteria E. Olavi Kajander, Ilpo Kuronen
and Neva Ciftcioglu Department of Biochemistry and Biotechnology, University
of Kuopio, FIN-70211 Kuopio, Finland
-
- A new potential threat for blood and blood prouducts,
cell culture, cell and tissue banking and biotechnology has been discovered:
Nanobacterium sanguineum gen. et sp. nov.. These self-replicating ultrafilterable
bacteria were isolated from over 80% of commercial ÔsterileÕ
fetal and newborn bovine sera and are thus the most common contaminant
present in cell cultures. Growth occured in vitro under cell culture conditions
(with or without mammalian cells) but not under anaerobic conditions. Their
doubling time was 1-5 days. They were culturable also in protein and lipid-free
medium beyond three years with monthly passages. Colony formation on solid
media was poor. The agent multiplied in culture with serum in a logarithmic
mode that could be prevented with aminoglycoside antibiotics, EDTA, cytosine
arabinoside and gamma irradiation. They showed procaryotic structure with
specific antigens detectable by monoclonal antibodies, were generally mobile,
coccoid with a diameter of 200 to 300 nm in serum, stained poorly with
bacteriological stains, were very resistant to antibiotics and passed through
100 but not 50 nm filters. Their aminoterminal protein sequences were novel
and reproducible. Considerable evidence suggested presence of nontraditional
DNA. They incorporated radiolabelled uridine into DNA. 16S rRNA gene sequence
results place them in alpha-2 subgroup of Proteobacteria which includes
Brucella, also pathogens of mammalians with preference to the fetus. This
new agent causes cytotoxicity and senescence in many cultured cell lines
by apoptotic cell death and growth arrest.
-
- *******
-
- Nanobacteria Threats In Vaccines
-
- http://www.uku.fi/~kajander/threat.html (pictures and
graphs at the site)
-
- Vaccines 97, Brown & al ed.,Cold Spring Harbor Laboratory
Press, New York, 1997 A New Potential Threat in Antigen and Antibody Products:
Nanobacteria Neva Ciftcioglu, Ilpo Kuronen, Kari Akerman, Erkki Hiltunen,
Jukka Laukkanen and E. Olavi Kajander Department of Biochemistry and Biotechnology,
University of Kuopio, FIN-70211 Kuopio, Finland.
-
- Several vaccines are currently being produced by using
cultured mammalian cells. Microbiological sterility of such vaccines is
of great importance since several examples indicate potential safety hazards
in vaccines contaminated with unknown organisms. Fetal bovine serum (FBS)
used as a supplement in cell culture is a known safety risk (Hodgson, 1995).
Obviously, not all of the risk factors of FBS are yet known and thus cannot
be controlled. It is commonly known that only about 10% of FBS batches
support cell cloning well (Liddel and Cryer, 1991) but the reasons for
this have remained unclear. As with many other cell culturers, we faced
a problem about 10 years ago of poorly thrivingo cells not attributable
to any known contaminant. In this report, we describe the discovery of
a new bacterium from mammalian blood and blood products, tentatively named
as Nanobacterium sanguineum gen. et sp. nov., and show that this agent
is common and harmful.
-
- DISCUSSION
-
- Culture and Diagnosis of Nanobacteria
-
- The discovery of Nanobacteria came about because we had
a problem with cell cultures namely vacuolized cells (Fig. 1A) and poorly
thriving cultures without any contaminant detectable by standard methods.
Transmission electron microscopy (TEM) made from these poorly thriving
cell cultures indicated the presence of internalized procaryotic organisms
(Fig. 1B). That their source was the commercial "sterile" FBS
was proven by gamma-irradiating all the culture components (Table 1). This
experiment also indicated that sterile culture media for detection of new
organisms can be made by using gamma-irradiated serum as a supplement.
The new organisms passed through 100 nm (but not 50 nm) filters and were
called nanobacteria, since no other bacteria are known that can pass through
filters with such small pores. This ability to pass through such small-pore
filters was most remarkable since they were shown to have a cell wall and
yet were able to surpass the filterablity of cell-wall-less bacteria. They
were unculturable in microbiological media but could be cultured under
cell culture conditions (with or without mammalian cells, CO2 5-10%). These
minute generally coccoid organisms had a diameter of 200 to 300 nm in serum,
and their size increased during the culture due to the production of a
very thick cell envelope (Fig. 1C, D). The thick and calcified envelope
made them visible even by light microscopy. The doubling time of nanobacteria
was 1-5 days (Fig. 2). Their multiplication could be detected by specific
ELISA, optical density, microscopic counting, SDS-PAGE or methionine and
uridine incorporation, and the multiplication could be prevented with high
doses of aminoglycoside antibiotics, EDTA, cytosine arabinoside and gamma-irradiation.
Considerable evidence suggested the presence of nontraditional DNA. 16S
rRNA gene sequence results (data will be published elsewhere) placed them
into the alpha-2 subgroup of Proteobacteria which includes Brucella(which
are also pathogens of mammalians with preference to the fetus) and Bartonella.
Nanobacteria were isolated from more than 80% of commercial FBS and newborn
bovine sera and are the most common contaminant present in cell cultures.
In addition, we isolated nanobacteria from the blood of 4% of medical students
at our university. Positive identification of nanobacteria involved growth
in cell culture medium with typical growth rate and optical properties,
specific stainability with Hoechst 33258 using the high dye concentration
and positive immunoassay results with immunofluorescence and/or ELISA using
monoclonal anti-nanobacteria antibodies.
-
- Cytotoxicity of Nanobacteria
-
- Nanobacteria are cytopathic in cell cultures and invade
mammalian cells in a distinctive manner: They trigger cells that are not
normally phagocytic to engulf them. These novel organisms are one of the
causes for cell vacuolization, poor thriving and unexpected cell lysis,
problems often encountered in mammalian cell culture. Several mammalian
fibroblast lines were cultured in MEM medium as described previously (Kajander
et al., 1990), and were infected with nanobacteria. Electron microscopy
and FITC staining with specific monoclonal antibodies indicated that nanobacteria
were bound on the surface of the fibroblasts (Fig. 1E-G). We concluded
that they were internalized either by receptor-mediated endocytosis or
by a closely related pathway. After the internalization, fibroblasts showed
apoptotic abnormalities and died if subjected to a high dose (>100 nanobacteria/cell).
-
- Different Growth Phases of Nanobacteria
-
- Washed nanobacteria added to serum-free medium grew very
slowly as evidenced by increase in their numbers and protein level and
were firmly attached to the culture plates. These cultures progressed to
large multicellular formations covered by layers of a firm protective material
several micrometers thick (Fig. 1H). After addition of sterile serum, the
layer disappeared, with typical small coccoid nanobacteria later appearing
in the same cultures with the mobile, larger D-shaped ones (Fig. 1I). Specific
monoclonal antibodies indicated the presence of the same antigenic sites
in both D-shaped and coccoid nanobacteria, and their 16S rRNA gene sequences
were 98% identical.
-
- How can Cell Culture be Possible with Nanobacteria-contaminated
Fetal Bovine Serum?
-
- Although more than 80% of cell culture serum batches
are contaminated with nanobacteria, many cell culturers have not faced
this problem with their cell cultures. We have experienced a major problem
with nanobacteria in cell culture only when they are present at high concentrations
relative to cells. This can occure typically in cell cloning and in long-term
experiments where mammalian cells do not multiply. Internalization of numerous
nanobacteria by a cell results in cytotoxicity. Importantly, most cell
lines multiply faster than nanobacteria. Thus, cytotoxic concentrations
may be avoided.
-
- Why Is Nanobacteria A Potential Threat?
-
- Nanobacteria can cause a chronic infection in laboratory
animals and in humans. The agent could be isolated from blood of one peron
for 5 years despite the presence of antibody. When nanobacteria were injected
into rabbits, the agent was initially isolated from urine and then from
cerebrospinal fluid after one year. Nanobacteria multiply very slowly and,
if pathogenic in humans, might cause slow chronic autoimmune-like disorders
(compare with leprosy or brucellosis). Sofar, there are no chronic bacteraemia
known that would not be harmful. Thus, the posibility that nanobacteria
may be present in vaccines made with cell culture, or in gammaglobulin
or other antibody preparations, must be controlled.
-
- SUMMARY AND CONCLUSIONS
-
- Nanobacteria are novel microorganisms that are not detectable
with present sterility testing methods, but they are detectable with new
culture and immunomethods. They are commonly present in bovine and blood
products and thus in cell cultures and antigens, including vaccines derived
therefrom, and may be present in antibody and gammaglobulin products. Nanobacteria
are a potential risk because of their cytotoxic properties and ability
to infect fetuses, and thus their pathogenicity should be scrutinized.
-
- ACKNOWLEDGMENTS
-
- We thank E. Tahvanainen, H. Martikainen, A. Pelttari
and T. Ojanen for their valuable help in microbiological, microscopic,
and chemical techniques. P. Mäenpää, T. Eloranta, J. Kärjä
and O. Lindqvist contributed valuable ideas in discussions. The work was
supported by the Academy of Finland, University of Kuopio, Finland Technology
Center, Juho Vainio Foundation and Savo High Technology Foundation.
-
- REFERENCES
-
- Hodgson, J. 1995. To treat or not to treat: That is the
question for serum.
- BioTechnology 13: 333.
- Kajander, E. O., R. J. Harvima, L. Kauppinen, K.K. Akerman,
H. Martikainen,
- R. L. Pajula, and S. O. Kärenlampi. 1990. Effects
of selenomethionine on
- cell growth and on S-adenosylmethionine metabolism in
cultured malignant
- cells. Biochem. J. 267: 767.
- Liddel, J. E., and A. Cryer. 1991. in A practical guide
to monoclonal
- antibodies, p. 25. Wiley, New York.
- Figure 1. Ultrastructure of nanobacteria and their interaction
with
- fibroblasts.
- (A) Perinuclear vacuolization of an infected 3T6 cell
under phase-contrast
- microscope;
- (B) TEM image of a nanobacterium engulfed by a BHK cell;
- (C) cultured coccoid nanobacteria (bars 200 nm).
- (D) SEM image of nanobacteria attached to culture vessel;
- (E) nanobacteria attached to a fibroblast surface (arrow
shows the surface
- of the fibroblast; bars 1 µm).
- (F) Indirect immunofluorescence staining of cultured
healthy 3T6 cells with
- a monoclonal antibody (8/0) against nanobacteria;
- (G) 3T6 cells inoculated with nanobacteria;
- (H) TEM of a nanobacterial population in a serum-free
culture (arrow shows
- a D-shaped nanobacterium in this population);
- (I) D-shaped nanobacteria after culture in serum-containing
medium (bars 1
- µm).
-
- Figure 2. Growth-curve of nanobacteria. As a control,
gamma-irradiated FBS
- was used. At each time point, samples from triplicate
incubations were
- taken, frozen and analyzed by turbidometer at the end
of the experiment.
- Turbidometer units are means of three measurements from
1/6 dilutions of
- cultures.
-
- Table 1. The Effect of 60Co Gamma-Irradiation of Culture
Components on
- Multiplication of Nanobacteria
-
- Culture Multiplication
- FBS
- RPMI +
- FBS
- *RPMI +
- *FBS
- RPMI -
- *FBS
- RPMI
- nanobacteria or FBS +
- *FBS
- RPMI
- *nanobacteria or * FBS -
-
- The material marked with asterisk (*) received a sterilization
dose of 3
- megarads during 16 h at room temperature. Cultures were
established using
- 10 % serum and nanobacterial counts were followed for
4 weeks.
-
- ********
- http://www.idreview.co.uk/pdf/pdf-1/11E1.PDF
-
- "Another intriguing subject is that of the putative
nanobacteria studied by a Finnish group. Present inhuman and bovine sera,
they might have contaminated many biological preparations and have spread
in human populations. As they induce deposition of calcium salts,they may
be involved in diseases involving such depositions, such as renal lithiasis
and atherosclerotic plaques, or even neuro-degenerative diseases. Their
minimal size (200 nM) precludes conventional packaging of a length of DNA
sufficient to code for an autonomous microorganism. But it is possible
that their genetic information is encoded in a modified, more compact DNA.In
conclusion, our fight against emerging diseases has just begun. We should
always be vigilant against the resurgence of known infectious germs and
the emergence of new agents. More than ever, a worldwidenetwork of sentinel
laboratories and a coordinated multidisciplinary effort in biomedical research
are required for our survival. "
-
- http://noshots.homestead.com/ingredients.html
-
- Patricia A. Doyle, PhD
- Please visit my "Emerging Diseases" message
board at:
- http://www.clickitnews.com/emergingdiseases/index.shtml
- Zhan le Devlesa tai sastimasa
- Go with God and in Good Health
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