PROFESSOR STEFAN S. NICOLAU (1896-1967)
A FOUNDER OF VIROLOGY IN ROMANIA*
COSTIN CERNESCU
“St. S. Nicolau “ Institute of Virology
285, sos Mihai Bravu, Bucharest 030304
cernescu@valhalla.racai.ro
Abstract. Forty years have passed since Professor Stefan S. Nicolau deceased. He was the founder of the Institute
of Virology in to Romanian Academy’ structure. As assistant of outstanding scientist Constantin Levaditi (18741953), Nicolau was trained at the Pasteur Institute. Their collaboration was exceptionally fruitful, producing new
concepts in virology. Between their most cited contributions was the concept of viral oncolysis. Oncolytic viruses
are capable of selective replication in malignant cells and therefore offer levels of potency and specificity that are
potentially far higher than conventional treatments for tumour. Viral oncolytic therapy is to day under intense
investigation as a novel anticancer strategy, both alone and in combination with other conventional treatment
modalities. Nicolau was between the first to observe a possible analogy between viruses and gene, and hence to
anticipate the future importance of molecular biology. In Romania, it was in his Institute were people from
different specialities, biologists, biochemists, and doctors were debated the implications of genetic engineering
long before it was in practice possible that science fiction became fact, as the use of DNA synthesis, restriction
enzymes and plasmids.
Key words: Stefan S. Nicolau, ultravirus, viral oncolysis, Borna disease.
* In commemoration of 40 years since the time of Stefan S. Nicolau death
1
to isolate and cultivate as true microbes.
INTRODUCTION
Levaditi and Nicolau coined the term of
In
virology,
ultravirus instead the label of “filterable
circumscribed to the first half of a
viruses” or “invisible viruses”, the label
twentieth
the
used throughout the 19th century in France
scientists
as a blanket term for new, unorthodox,
Constantin Levaditi and Stefan S. Nicolau
infectious agents. In fact, Levaditi and
played an important role. As faithful
Pierre Lepine published the first textbook
followers of Pasteur they assumed that
of virology in 1938: “Les ultravirus des
new filterable agents of infectious diseases
maladies humaines”.2 It is to be noted that
discovered by botanists (D.I. Ivanovski –
the term “virology” became established in
Tobacco
by
its own right only after the World War II
veterinarians (F. Loeffler, and P. Frosch –
with the apparition of the S. E. Luria’s
foot and mouth disease) or by physicians
classic manual “General Virology” (1953)
(W. Reed, and J. Carroll – yellow fever;
and of the first issue of the journal
K. Landsteiner, and C. Levaditi1 -
Virology (1955).
contribution
the
history
century
of
of
sequence,
Romanian
Mosaic
Disease),
poliomyelitis) were small bacteria unable
Fig.1. Jubilant “Nicolau” Medal awarded for excellence in PhD works at the Institute of
Virology, Bucharest (a craftwork of outstanding Romanian sculptor Ion Jalea).
2
Nicolau was a natural organizer and,
relationships), Elena Oprescu (respirtory
together with Radu Portocalã and Nicolae
viral
Cajal, he began to recruit people to work
(biochemistry of the viral nucleic acids),
on isolation and cultivation of human and
Dan Sarateanu (ornithoses), Constantin
animal viruses. He developed a group of
Surdan (Rickettsia), Nicolae Cajal - his
devotees and students who adopted his
succesor at the Institute and Chair of
view points on the important problems of
virology direction. The Institute bridges
virus diseases of humans and domesticated
the gap between clinical investigation and
animals in parallel with research in
the application of basic science in order to
theoretical and experimental virology.
understand
infections),
the
Radu
pathogenesis
Portocala
of
the
infectious diseases, their diagnosis and
Stefan S. Nicolau founded the
optimal
Institute of Inframicrobiology of the
control.
management,
prevention
and
Romanian Academy. The Institute has its
origin in the nucleus of the Chair of
SHORT BIOGRAPHIC NOTE
Virology, which began its activity in 1942
in the framework of the Faculty of
Stefan S. Nicolau obtained in
Medicine in Bucharest. For the first time
1920 the MD degree at the Faculty of
in the world, in the higher medical
Medicine in Cluj, where he began this
education, a separate discipline was
collaboration with professor Moldoveanu.
devoted to the study of the ultrafiltrable
From his student years in Cluj, Nicolau
infectious agents. Afterwards, a team of
showed his tendency towards laboratory
inframicrobiology of the Academy was
research. After having spent one year in
created, which developed into the actual
Cluj, he began to work at Levaditi’s
“Nicolau” Institute of Virology. The
laboratory in the Pasteur Institute. Nicolau
«Stefan S. Nicolau» Institute of Virology,
studied for two decades at The Pasteur
which chronologically is one of the first
Institute in Paris (1920-1939). During this
institutions in this speciality in Europe,
interval he passed his PhD at Sorbonne
has
50th
(1925) and visited National Institute for
anniversary of its foundation. Many
Medical Research – London (1927-1931).
disciples
by
He received many awards from Paris
distinguished
Academy of Sciences and also important
themselves by significant contributions:
prizes: Prize Bellion, 1926, Prize Breant,
Adelina
viral
1930, Prize Montyon, 1935. Out of the
infections), Elisabeta Nastac (virus-cancer
country until 1939, Nicolau authored
celebrated
professor
of
the
in
1999
school
Nicolau
Derevic
the
created
(respiratory
3
arbovirus infection7; the introduction of
pioneering studies regarding viral diseases
such
as:
herpes3,
rabies4,
poxvirus
the synchronophylaxis concept, a fact
5,
,
which promotes him as a, precursor of the
disease7,
interference phenomenon description8; the
Aujeszky’s disease, «looping illness» and
discovery of the photodynamic action of
others.
stains (which is nowadays widely used in
infections
aphthous
(especially
fever,
Among
vaccinia)
Borna
the
6
worldwide
acknowledged priorities we mention: the
viral
mutagenesis
research
on);
the
first detection of inclusions in the yellow
description of the septic neuritis9 and
fever – a diagnostic criterion of this
others.
.
Fig 2. The office block of “Nicolau” Institute of Virology (www.virology.ro)
In 1939, Nicolau returned in
Iasi Medical School, and in 1942, as
Romania as professor of Microbiology at
professor
4
of
Inframicrobiology
at
Bucharest
Medical
School.
In
the
these objects by newly available electron
microscope in the early 1940’.
recognition of his outstanding publications
he was elected member of Romanian
In a broad prospective view of the
Academy of Medical Sciences and of
Romanian Academy.
pathogenesis of viral infections, Levaditi
He was director of the Institute of
and Nicolau underlined the cell-mediated
Inframicrobiology in Bucharest (to day,
character of the antiviral immunity. Beside
“St. S. Nicolau” Institute of Virology),
the viral cytocidal effect, Nicolau revealed
where he have the opportunity to create
the participation of the cellular immune
many independent laboratories in which
response of the host in the triggering of the
his co-workers served national public
symptomatology.
In a critical review of the literature
health and medical school. As a teacher he
was the editor of remarkable manuals
on
the
antigenic
translated in French, Russian, Chinese etc.
ultraviruses,
properties of
Nicolau
and
the
co-workers
(1928)11 called attention to the fact that
these agents of disease, as far as they have
EARLY WORKS
been studied, seem to exhibit antigenic
In 1922, Levaditi and Nicolau
properties which suggest placing them in
published a series of articles in which new
the general group of bacteria and other
properties of the viral agents were
more
demonstrated:
ultrafiltrability
(passage
bodies. They observed that vaccinia virus
through
pores
collodion
inoculated into the brain of an immunized
membrane), antigenic properties of killed
rabbit could not be demonstrated two
and replicative viruses etc. Nicolau and
hours after the injection. While vaccinia
Levaditi elaborated experiments on the
virus persisted in the circulation of a
filterability of viruses with a very original
susceptible rabbit for as long as 8 days
and more constructive interpretation. They
following
opposed to the concept of non-particulate
disappeared
infective organism. In the debate about
immune rabbit within 4 to 6 hours.
whether the viruses were living or
Nicolau and Kopciowska made a similar
nonliving, or perhaps were the primordial
observation
stuff from which life itself originated,
introduced into the brain of immunized
Nicolau underscored the macromolecular
rabbits7.
nature of viruses and the significance of
The same as the classic of Romanian
their tiny component - the genomic nucleic
medicine – Victor Babeº - Nicolau’ early
acid. The structure of viruses was soon
research works were connected with
amply confirmed by direct observation of
rabies. He described extremely important
the
of
a
5
highly
organized
intravenous
from
proteinaceous
infection,
circulation
regarding
herpes
of
it
an
virus
lesions in the central nervous system of
Between the most cited Nicolau’
animal dead of rabies or of other
publications were papers referring the role
neurotropic viruses. Although described
of
by Victor Babeº as far back as 1886, the
neurobehavioural diseases. Borna disease
cytoplasmic inclusions in rabies were
virus (BDV) is the prototype of the family
given the name of “Negri bodies”.11 The
Bornaviridae, genus Bornavirus, within
priority of Babeº was recognized owing to
the
the well documented works of St. S.
ribonucleic acid (RNA) viruses (order
Nicolau and Mircea Babeº. Nicolau was
Mononegavirales). Nicolau and Galloway
the editor of two volumes of V. Babeº
were the firsts to conduct experimental
selected works and of many monographs
study of Borna disease in sheep and cattle
which created a true virology library in
and describe the characteristics of this
Romanian. (see books at the references)
enzootic
Borna
virus
nonsegmented
infection
in
negative-strand
encephalo-myelitis.13
This
neurotropic virus appears to be distributed
In regard to the immunological
worldwide and has the potential to infect
conditions existing in herpes disease, his
most, if not all, warm-blooded hosts. The
experiments have reaffirmed that herpes
name Borna refers to the city of Borna,
virus can be neutralized with the serum of
Germany, the site of an equine epidemic in
actively immunized animals and have
1895–1896
offered an explanation for the irregularity
cavalry.
of the results of others. Nicolau et al.
reported primarily in Europe, but recent
found that brain extracts from inoculated
reports also include North America and
rabbits possess some virus-neutralizing
parts of Asia (Japan, Israel and Iran).
power, but considerably less than the
Reports
serum
control
infected animals suggest that the virus
animals. It has been shown that active
may be more widespread than previously
immunity can be attained only when some
appreciated. Reports of BDV nucleic acid
degree of reaction to the living virus has
and
occurred.
mononuclear cells also indicate a potential
of
the
corresponding
Rabbits
which
survived
neutralized serum-virus mixtures did not
that
crippled
Natural
of
proteins
infection
the
has
asymptomatic,
in
Saxon
peripheral
been
naturally
blood
for haematogenous transmission.
acquire immunity nor did those treated
Although there is consensus that
with virus phenolized to the extent of
humans are likely to be susceptible to
actual destruction.
9
BDV infection, the epidemiology and
clinical consequences of human infection
remain controversial. After 1990, German
THE MOST CITED WORKS
scientists reported that Borna infectious
virus has been isolated from humans. 14, 15
6
There have been no large controlled
of general pathology. Thus, morphological
prevalence
data supported the viral etiology of some
studies
but
methods
for
diagnosis of human infection were reliable
cardiovascular
diseases
suggesting an association between BDV
myocarditides)21
and neuropsychiatric disorders, including
chlamydia,
unipolar depression, bipolar disorder or
herpesviruses in the etiopathogenesis of
schizophrenia. BDV has also been linked
some
to chronic fatigue syndrome,
spontaneous abortions.22
and
the
rickettsia
congenital
(arteritides,
role
and
of
some
malformations
and
An innovative concept developed
I shall detail my remarks to
by Levaditi and Nicolau was viral
Nicolau’ papers on the relations between
oncolysis
the
viruses and cancer, published in the period
neurovaccinal virus was inoculated into
from 1960 to 1967, which I personally
in
grafted tumours.
studies
16
where
Sinkovics J.C., and
consider highly significant. The author
Horwath J17. (Viruses and cancer. Medical
summarises the general properties of
Hypotheses 1995, 44, 359-368.) quoted
human
from the paper of Romanian scientists:
development
“Tumours are more susceptible to viruses
microbiology and proposes guidelines that
than normal cells and tumours act as a
might help to determine the role of viruses
sponge attracting viral replication". This
in human cancer. The work on the role of
hypothesis is now in the centre of interest
viruses in the pathogenesis of human
for many biotechnology companies who
cancer was hampered by the fact that
enlarge cancer virotherapy models.
virus-associated cancer develops only in a
tumour
viruses,
of
causal
reviews
the
thinking
in
Stefan S. Nicolau made obvious
small minority of infected subjects, which
the theory of viral etiology of some
implies that, if the virus does play a role in
cancers. With a remarkable intuition
the pathogenesis of the malignancy, other
Nicolau promoted the hypothesis of the
factors must be also involved. Naturally
oncogenic potential of the viral nucleic
oncolytic
acids. He developed the infravirus model –
competent viruses that have an innate
the autonomized form of viral nucleic
ability to selectively infect and kill tumour
acids which, after its integration in the
cells. Despite being used in the original
host cell genome, contribute to the viral
attempts to treat cancer with live viruses
transformation and the occurrence of
five decades ago, interest in naturally
various morbid states.
oncolytic viruses has lagged behind the
The school of professor Nicolau
viruses
are
replication-
support for engineered adenoviruses and
provided significant arguments for the
herpesviruses
involvement of viruses in the other fields
Recently,
7
as
cancer
however,
there
therapeutics.
has
been
renewed interest in the high potency and
carcinoma); and animal T-cell Leukemia
selectivity of these naturally occurring
Viruses
(bovine,
19, 20
agents.
leukemias).
used
in
murine
T-cell
The criteria most often
determining
causality
were
Levaditi and Nicolau were the first
consistency of the association, either
to study the action of a neurotropic strain
epidemiologic or on the molecular level,
of vaccinia virus on tumours of mice and
and oncogenicity of the agent in animal
rats. They found epitheliomas of these
models or cell cultures.21,
animals to provide an excellent culture
most of the tumour viruses the viral
medium for the virus in contrast to
genome persists in an integrated or
sarcoma. These findings were in accord
episomal form with a subset of viral genes
with their previous investigations where
expressed in the tumour cells, some agents
neurovaccinia
(Hepatitis B virus) are not inherently
ectodermal
was
rather
shown
than
to
have
mesodermal
oncogenic,
tropism.
but
transformation
Studying tumours infected with
means.23,
24
22
Whereas for
infection
of
cells
leads
by
to
indirect
For some malignancies the
neurovaccinia in immune animals Levaditi
viral agent appears to serve as a cofactor
and Nicolau showed that virus did not
(Burkitt's lymphoma-EBV; mesothelioma
survive in the immune host. If a portion of
- SV(40)). For others the association is
the tumour were transplanted into non-
inconsistent (Hodgkin's Disease, breast
immune animal, it would support growth
cancer - EBV) and may either define
of the infectious agent. The proliferative
subsets of these malignancies, or the virus
power of the transplantable tumour was
may act to modify phenotype of an
also interfered and is not always possible
established tumour, contributing to tumour
to make successful transplants from
progression
tumours carrying the vaccinia virus.
tumour.
Nicolau
and
his
co workers
In
association
rather
than
many
with
other
causing
the
cases
the
malignancy
is
less
developed investigations among many
consistent or still emerging. For example,
animal and human viruses that contribute
despite the potent oncogenic properties of
to a variety of malignancies.18 Between
some strains of human adenovirus in tissue
most studied viruses linked to cancers
culture and animals the virus has not been
were human papillomaviruses (cervical
linked with any human cancers. Finally it
carcinoma);
polyomaviruses
is likely that more agents, most likely
(mesotheliomas, brain tumours); Epstein-
viruses, both known and unidentified,
Barr virus (B-cell lymphoproliferative
have yet to be implicated in human cancer.
diseases and nasopharyngeal carcinoma);
In the meantime study of tumourigenic
hepatitis
human
viruses
(hepatocellular
8
5.
infectious agents will continue to elucidate
Levaditi C., and Nicolau S., Vaccine et
neoplasmes. Ann. Inst. Pasteur, 1923, 37,
molecular oncogenic processes.
443-447.
6.
Viruses have been used for cancer
Levaditi C., Nicolau S., Sur le culture du
virus vaccinal dans les neoplasmes
treatment for over a century. From the
epitheliaux. Compt. Rend. Soc. Biol.
early clinical studies with various wild-
(Paris), 1929, 101, 334.
7.
type viruses, to the modern trials with
engineered
emerged
viruses,
as
a
virotherapy
Nicolau S. S. & Kopciowska L A new
stain method. Compt. Rend. Soc. Biol.
has
(Paris) 1929, 101, 655-7
promising therapeutic
8.
Nicolau S.S. & Kopciowska L.
strategy.19 In a recent review, which also
Persistence of rabies virus in vaccinated
mentions Levaditi and Nicolau priority,
animals. Compt. Rend. Soc. Biol. (Paris),
japanese authors discuss the progress and
1932, 110, 348-351.
challenges
associated
with
9.
oncolytic
note on the experimental study of enzootic
virotherapeutic agents, summarizing the
encephalo-myelitis (Borna disease) Br. J.
data from clinical reports, and the
Exp. Pathol. 1927, 8, 336-341.
10. Levaditi C. and, Nicolau S. S., Virus et
implications of this data for future
virotherapy
development.25
Nicolau S.S., and Galloway I. Preliminary
tumeurs. Ann Inst Pasteur 1923, 37, 1-3.
Beyond
11. Nicolau S., Kopciowska L., Balmus G.
historical perspective on the development
Inclusions cytoplasmiques simulant les
of oncolytic virus the reviewers present
corps de Negri dans le cerveau de la
the encouraging results of recent clinical
souris normale. . Compt. Rend. Soc. Biol.
trials (e.g., an modified herpesvirus has
(Paris) 1933, 113, 851-855.
12. Nicolau S.S., Galloway I.A., Dimancesco-
been tested in clinical trial of nearly 250
Nicolau O. « Étude sur les septinévrites à
patients and approved for human use by
ultravirus neurotropes » Masson, Paris,
the Chinese FDA).
1929.
13. Nicolau S.S., and Galloway I. Borna
REFERENCES
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disease and enzootic encephalomyelitis of
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sheep and cattle. Spec. Rep. Ser. Med.
transmission de la paralysie infantile aux
Res. Council 1928, 121, 7-90.
14. Bode L., Dürrwald R., Rantam F.A.,
singes. Compt. Rend. Soc. Biol. (Paris).
2.
3.
4.
1909:592-594.
Ferszt R. and Ludwig H. First isolates of
Levaditi C., Lépine P., Blanc G., (eds)
infectious human Borna disease virus
« Les ultravirus des maladies humaines ».
from patients with mood disorders.
Librairie Maloine Paris, France, 1938.
Molecular Psychiatry 1996, 1, 200–212.
15. Dittrich W., Bode L., Ludwig H., Kao M.
Levaditi C., and Nicolau S., Affinite du
virus herpetique pour les neoplasmes
and Schneider K. Learning deficiencies in
epitheliaux. Compt. Rend. Soc. Biol.
Borna disease virus-infected but clinically
(Paris), 1922, 87, 498-500.
healthy rats. Biological Psychiatry 1989,
26, 818–828.
Nicolau S., and Kopciowska L. Compt
16. St. S. Nicolau “Cancer and viruses”,
Rend Soc Biol (Paris) 1922, 86, 563.
1955, Editura Academiei, Bucharest
9
17. Sinkovics J.C., and Horwath J., Viruses
into an immunogenic virus under the
and cancer. Medical Hypotheses 1995,
action of copper. Acta Biol Med Ger.
44, 359-368.
1962, 8, 219-29.
22. Nicolau S. S. et al. Study on the rickettsial
18. Nicolau S.S., Cajal N., Schächter A., S
r þeanu D., Cepleanu M., Sorodoc Y.
etiology of various angiopathies. Rev Sci
Les relations entre l'ornithose et la
Med. 1963, 8, 69-73.
23. St. S. Nicolau “Viral hepatitis” 1960,
maladie de Hodgkin. Archives of
Editura Academiei, Bucharest.
Virology ( Springer Wien) 1963, 13, 1432.
24. Nicolau S.S. Inframicrobial epidemic
19. Nicolau S.S., The oncolytic action of
certain viruses. Stud Cercet
hepatitis, the virus of the disease. Rom
Inframicrobiol. 1960, 11, 529-48.
Med Rev. 1961, 5, 19-23.
25. Kasuya H., Takeda S., Shimoyama
Romanian.
S., Shikano T., Nomura N., Kanazumi
20. St. S. Nicolau “Basic Inframicrobiology”
(Elemente de Inframicrobiologie,
N., Nomoto S., Sugimoto H., Nakao, A.
Romanian, Russian, Chinese, 1956, 1962,
Current Cancer Drug Targets 2007,
Editura Medicalã, Bucharest.
7, 123-125.
21. Nicolau S. S. et al. Experimental
investigations on the antirabies vaccine
prepared under the action of copper ions.
Contributions concerning the mechanism
of transformation of the pathogenic virus
Books
1.
2.
3.
4.
5.
6.
7.
Şt.
S.
Nicolau
HEPATITE
INFRAMICROBIENE (“Viral hepatitis”)
- Ed. Academiei R.P.R., Bucharest, 1954.
V. BABEŞ (OPERE ALESE) („Selected
works of Victor Babeº”) Vol. I – (Şt. S.
Nicolau, M. Babeş, Fl. Horodniceanu,
eds), Ed. Academiei R.P.R., Bucharest,
1954.
V. BABEŞ (OPERE ALESE) (Selected
works of Victor Babeº”) Vol. II - Şt. S.
Nicolau, M. Babeş, Fl. Horodniceanu,
eds), Ed. Academiei R.P.R., Bucharest,
1959.
Şt. S. Nicolau CANCER ŞI VIRUSURI.
CÎTEVA
CONSIDERAŢII
PRIVITOARE LA IPOTEZA VIROTICĂ
A CANCERULUI („Cancer and Viruses –
remarks on the cancer viral etiology”), Ed.
Academiei R.P.R., Bucharest, 1955.
Şt. S. Nicolau ELEMENTE DE
INFRAMICROBIOLOGIE GENERALĂ
(„Basic
inframicrobiology”),
Ed.
Academiei R.P.R., Bucharest, 1956.
Şt. S. Nicolau (editor) PROBLEME DE
INFRAMICROBIOLOGIE („Progress in
inframicrobiology”)- coautori: Şt. S.
Nicolau, N. Cajal, A. Derevici, C. Iacob,
G. Marinescu, S. Mateescu, R. Portocală,
D. Sărăţeanu, J. Schwartz, Ed. Medicală,
Bucharest, 1956.
Şt.
S.
Nicolau
HEPATITELE
INFECŢIOASE INFRAMICROBIENE
8.
9.
10.
11.
12.
13.
10
(„Virus infections of the liver”)- Şt. S.
Nicolau,
Ed.
Academiei
R.P.R.,
Bucharest, 1957.
Şt. S. Nicolau, Al. Rădulescu, N. M.
Constantinescu, N. Cajal, G. Marinescu,
POLIOMIELITA – („Poliomyelitis”), Ed.
Academiei R.P.R., Bucharest, 1961.
Şt. S. Nicolau, N. M. Constantinescu, N.
Cajal. TURBAREA („Rabies”) - Ed.
Academiei R.P.R., Bucharest, 1962.
Şt. S. Nicolau (ed) ELEMENTE DE
INFRAMICROBIOLOGIE SPECIALĂ
(„Infections of inframicrobial etiology”) –
Ed. Medicală, Bucharest, 1962.
Şt. S. Nicolau, N. Constantinescu
RICKETTSII
ŞI
RICKETTSIOZE
(„Infections of rickttsial etiology”-, Ed.
Academiei R.S.R., Bucharest, 1965.
Şt. S. Nicolau, Elisabeta Nastac,
VIRUSURI ŞI TUMORI („Viruses and
tumours”) - Ed. Academiei R.S.R.,
Bucharest, 1968.
Şt.
S.
Nicolau,
N.
Drăgănescu
HERPESUL („Hepres viruses”) - Ed.
Academiei R.S.R., Bucharest, 1968.