INTERNATIONAL JOURNAL O F
SYSTEMATIC
BACTERIOLOGY
Vol. 18, No. 1
J a n u a r y 1968
pp. 51- 66
Copyright 1968, Iowa State University P r e s s
PROPOSAL FOR THE RECOGNITION OF TWO SPECIES
IN THE GENUS CHLAMYDIA JONES,
RAKE, AND STEARNS, 1945'
L.A. P a g e Z
National Animal D i s e a s e L a b o r a t o r y , Animal D i s e a s e and
P a r a s i t e R e s e a r c h Division, A g r i c u l t u r a l R e s e a r c h S e r v i c e ,
United S t a t e s Department of A g r i c u l t u r e , A m e s , Iowa
ABSTRACT. T h e b a c t e r i a w h i c h c a u s e t h e p s i t t a c o s i s - t r a c h o m a g r o u p of d i s e a s e s a n d w h i c h
a r e assigned to the genus Chlamydia Jones,
Rake, and Stearns, 1945, can be separated
conveniently and logically into two species.
This separation is based on relatively stable
morphological and chemical characteristics
of t h e o r g a n i s m s r a t h e r t h a n o n t h e i r p r e sumed host o r tissue preferences o r on the
s p e c i f i c s e r o l o g y of t h e i r c e l l w a l l a n t i g e n s .
The type species Chlamydia trachomatis (Busacca) Rake 1957 is differentiated f r o m the
second species
Chlamydia psittaci (Lillie
1930) comb. nov., by a s i m p l e key using t e s t s
t h a t can be p e r f o r m e d r e a d i l y i n t h e l a b o r a tory.
E a c h species is defined, and the laboratory tests are described.
- - - - - - - - - - - INTRODUCTION
In 1965 it w a s a p p a r e n t that the n u m e r o u s taxonomic and
n o m e n c l a t u r a l proposals m a d e o v e r two decades by A m e r i can, R u s s i a m a n d F r e n c h w o r k e r s had failed t o s t a b i l i z e the
scientific naming of the group of r e l a t e d o r g a n i s m s that
c a u s e p s i t t a c o s i s , t r a c h o m a , and o t h e r d i s e a s e s . P r i o r t o
This p r o p o s a l h a s been reviewed and approved by a m a j o r ity of the m e m b e r s of the Subcommittee on the Chlamydiac e a e , Taxonomy Committee, A m e r i c a n Society f o r Microbiology, October 1967.
S e c r e t a r y , Subcommittee on the Chlamydiaceae, Taxonomy
Committee, A m e r i c a n Society f o r Microbiology.
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
52
INTERNATIONAL JOURNAL
1965, A m e r i c a n s y s t e m a t i s t s a d h e r e d t o R a k e ' s p l a c e m e n t
of t h e s e o r g a n i s m s into two g e n e r a , Chlamydia Rake 1956
and Miyagawanella B r u m p t 1938, although m a n y w o r k e r s
f a v o r e d , f o r h i s t o r i c a n d phonetic r e a s o n s t h e g e n e r i c n a m e
Bedsonia, which w a s belatedly p r o p o s e d by M e y e r (1953).
Intervening w e r e the validly published p r o p o s a l s of Moshkovs ky ( 1945), Ryzhkov ( 1 950), Zhdanov a n d K o r e n b l i t ( 1950),
and Zhdanov(l953) of t h e genus n a m e s Chlamydozoon f o r t h e
organisms causing trachoma and Ehrlichia o r Rickettsiaf o r m i s f o r the o r g a n i s m s c a u s i n g p s i t t a c o s i s .
The Russian
a u t h o r s c o n s i d e r e d t h e m t o be v i r u s e s .
Lastly, Levaditi,
R o g e r , and D e s t o m b e s p r o p o s e d i n 1964 t h a t t h e s e o r g a n i s m s
be s p l i t a m o n g f o u r g e n e r a : Chlamydia Rake, Miyagawanella
B r u m p t , Bedsonia M e y e r , and R a k e i a L e v a d i t i gt 21.
T h e q u e s t i o n s r a i s e d by t h e s e v a r i o u s p r o p o s a l s w e r e
e x a m i n e d f o r t a x o n o m i c m e r i t i n t h e light of r e c e n t c h l a m y d i a l r e s e a r c h , and evaluated a c c o r d i n g t o t h e I n t e r n a t i o n a l
Code of N o m e n c l a t u r e of B a c t e r i a (1958) f o r n o m e n c l a t u r a l
l e g i t i m a c y ( P a g e 1966).
T h e conclusions of t h i s study,
r e a c h e d after consultation with Arne r i c a n s y s t e m a t i s t s a n d
microbiologists,l were:
( 1 ) T h a t t h e evidence a c c u m u l a t e d by M o u l d e r ( 1 964, 1966)
and o t h e r s e s t a b l i s h e d t h a t t h e etiologic a g e n t s of t r a c h o m a ,
psittacosis, and related diseases w e r e procaryotic m i c r o o r g a n i s m s , o r b a c t e r i a in t h e b r o a d s e n s e , a n d w e r e fundamentally distinct f r o m the viruses.
( 2 ) T h a t t h e c h a r a c t e r i s t i c s u s e d t o unite t h e s e o r g a n i s m s
i n t o a s i n g l e genus (morphology, d e v e l o p m e n t a l cycle, a n d
g r o u p a n t i g e n ) w e r e of g r e a t e r t a x o n o m i c significance t h a n
t h e c h a r a c t e r i s t i c s u s e d t o s e p a r a t e t h e m i n t o two o r f o u r
g e n e r a ( p r e s u m e d h o s t o r t i s s u e p r e f e r e n c e s on t h e p a r t of
t h e o r g a n i s m s , and p a t r o n y m i c s on t h e p a r t of h i s t o r y m i n d e d humans). T h e r e f o r e , it w a s a p p r o p r i a t e t o c l a s s i f y
the g r o u p of o r g a n i s m s into one genus.
( 3 ) T h a t , a c c o r d i n g t o t h e r u l e s of the I n t e r n a t i o n a l Code
of N o m e n c l a t u r e of B a c t e r i a , t h e e a r l i e s t validly published,
l e g i t i m a t e and c o r r e c t n a m e f o r t h i s genus w a s C h l a m y d i a ,
J o n e s , R a k e and S t e a r n s , 1945.
These authors also were
t h e f i r s t t o e x p r e s s t h e concept t h a t a l l of the a g e n t s c a u s i n g
either trachoma, psittacosis o r related diseases were m e m b e r s of a s i n g l e g e n u s . A l l o t h e r g e n e r i c n a m e s f o r c h l a m y d i a e e i t h e r w e r e invalidly published o r w e r e l a t e r s y n o n y m s
of Chlamydia.
R. E. Buchanan, C. B. P h i l i p , J. W. M o u l d e r , F. B. Gordon,
E. J a w e t z , a n d M. P o l l a r d .
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
SYSTEMATIC
BACTERIOLOGY
53
T h e publication by P a g e (1966) did not p r o p o s e any new
n a m e s , but i t c l a r i f i e d t h e t a x o n o m i c and n o m e n c l a t u r a l
position of p r e v i o u s l y p r o p o s e d n a m e s and e m p h a s i z e d t h e
c o r r e c t s c i e n t i f i c name. Its p u r p o s e w a s t o s i m p l i f y and t o
s t a b i l i z e chlamydia1 n o m e n c l a t u r e .
R e a s o n s f o r t h e Recognition a t P r e s e n t of
Two S p e c i e s i n t h e Genus Chlamydia
In t h e p a s t , c l a s s i f i c a t i o n s y s t e m s p r o p o s e d f o r c h l a m y d i a e have u s e d p r e s u m e d t i s s u e o r h o s t affinities of t h e o r g a n i s m s a s heavily weighted t a x o n o m i c c r i t e r i a . T h i s is
u n d e r s t a n d a b l e b e c a u s e l e s s w a s known about t h e o r g a n i s m s
t h e m s e l v e s t h a n about the d a m a g e t h e y c a u s e d t o t h e i r h o s t s .
Moshkovsky ( 1945) e m p h a s i z e d h o s t c e l l o r i g i n and i n t r o duced t h e t e r m s " m e s o d e r m o s i s " f o r p s i t t a c o s i s , lymphog r a n u l o m a v e n e r e u m , a n d a n u m b e r of o t h e r d i s e a s e s , and
" e c t o d e r m o s i s " f o r t r a c h o m a and v a r i o u s conjunctival inflammations.
He c l a s s i f i e d t h e etiologic a g e n t s of t h e s e
d i s e a s e s on t h e b a s i s of t h e i r p r e s u m e d - t i s s u e c e l l affinities.
The c l a s s i f i c a t i o n s y s t e m of R a k e in B e r g e y ' s 6th
(1948) and 7th ( 1 957) editions a l s o r e f l e c t e d a tissue-affinity/
h o s t p r e f e r e n c e outlook. E x a m p l e s a r e :
Disease
Trachoma
Inclus ion c onjunct i viti s
Lymphogranuloma venereum
Psittacosis
P s i t t a c o s i s i n nonpsittacine birds
Enzootic abortion in ewes
S p o r a d i c bovine
e n c e pha l o m ye l i ti s
Calf e nte r i t i s
S pe c i f i c e t i 010g y
Chlamydia t r a c h o m a t i s
Chlamydia oculogenitale
Miyagawanella
lymphog r a n u l o m a t o s i s
Miyagawanella ps i t t a c i
Miyagawanella o r n i t h o s i s
Miyagawanella ovis
Mi yagawanella p e c o r i s
Miyagawanella bovis
On t h e o t h e r hand, L e v a d i t i e t &. (1964) s u g g e s t e d t h a t
t h e s e o r g a n i s m s should be s e p a r a t e d on t h e b a s i s of a p r e s u m e d l y unique epidemiology.
Thus, the agents associated
with t r a c h o m a belonged i n t h e genus Chlamydia, with l y m p h o g r a n u l o m a v e n e r e u m i n Miyagawanella, with p s i t t a c o s i s
i n b i r d s and m a n i n Bedsonia, and with d i s e a s e s of d o m e s t i c
h e r b i v o r e s i n Rakeia.
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
54
INTERNATIONAL
JOURNAL
In t h e light of r e c e n t evidence, the u s e of s u c h c r i t e r i a i n
t h e c l a s s i f i c a t i o n of chlamydiae is c o n t r a i n d i c a t e d ; f o r , when
knowledge of the c l i n i c a l s y n d r o m e of the h o s t i s not availa b l e , Chlamydia t r a c h o m a t i s i n Rake's s c h e m a cannot be
s a t i s f a c t o r i l y distinguished by l a b o r a t o r y t e s t s f r o m C h l a m &a
oculogenitale o r f r o m Miyagawanella lymphogranulomat o s i s . If the o r g a n i s m s a r e i s o l a t e d f r o m the g e n i t a l t r a c t
of a p e r s o n with no evidence of d i s e a s e , they cannot. be a s signed with c e r t a i n t y t o any of R a k e ' s s p e c i e s . Dunlop and
h i s colleagues (1966) r e c e n t l y r e p o r t e d the i s o l a t i o n of
chlamydiae f r o m t h e u r o g e n i t a l t r a c t s and conjunctiva of
patients with c l i n i c a l u r e t h r i t i s , opththalmia neonatorurn, o r
t r a c h o m a . A l l of t h e i s o l a t e s w e r e m i c r o b i o l o g i c a l l y i n d i s tinguishable. T h e s e a u t h o r s s t a t e d t h a t t h e r e w a s "no diff e r e n c e between the i s o l a t e f r o m t h e e y e and t h a t f r o m the
genital t r a c t in e a c h case."
In a n o t h e r c a s e , t h e s e a u t h o r s
r e p o r t e d t h a t "an i s o l a t e of what i s probably a TRIG* agent
h a s been obtained f r o m the r e c t u m of a w o m a n with o c u l a r
T h e difficulty in differentiating
infection due t o the agent..
s u c h c a s e s f r o m lymphogranuloma v e n e r e u m ( L G V ) , and
TRIC agent f r o m t h e agent of LGV, i s a p p a r e n t . "
O t h e r e x a m p l e s i l l u s t r a t e t h e inadvisability of a s c r i b i n g
a s p e c i e s designation t o a s t r a i n of o r g a n i s m s p r e s u m e d t o
be the s o l e c a u s e of a p a r t i c u l a r d i s e a s e . S c h a c h t e r (1967)
r e p o r t e d the i s o l a t i o n f r o m the t i s s u e of a patient with clinic a l lymphogranuloma v e n e r e u m of a n o r g a n i s m t h a t w a s
morphologically, pathogenically, a n d c h e m i c a l l y indistinguishable f r o m s t r a i n s n o r m a l l y r e c o v e r e d f r o m p s i t t a c i n e
birds. Was the patient with c l i n i c a l L G V infected with Miyagawanella l y m p h o g r a n u l o m a t o s i s o r with ,M. p s i t t a c i ?
P a g e ( 1967) r e p o r t e d t h a t a chlamydia1 agent i m p l i c a t e d
in a c a s e of bovine e n t e r i t i s and in a case of fatal p s i t t a c o s i s
w a s i d e n t i c a l in "pathotype" and-m o r p h o l o g i c a l a n d c h e m i c a l
c h a r a c t e r i s t i c s t o a s t r a i n t y p i c a l of t h o s e f r e q u e n t l y r e c o v e r e d f r o m pigeons. However, i t w a s quite d i f f e r e n t f r o m
a n o t h e r s t r a i n w h i c h had c a u s e d a n epizootic of bovine a b o r t i o n s . W e r e t h e bovines and t h e human s u b j e c t infected with
M. p s i t t a c i , _M. o r n i t h o s i s , &l.
bovis, o r x . p e c o r i s ? W a s
t h e patient who w a s affected by t h e bovine agent suffering
f r o m psittacosis, ornithosis, o r m a m m a l o s i s ?
Unfortunately, s e r o t y p i n g of the s t r a i n s i s o l a t e d f r o m
v a r i o u s h o s t s with differing d i s e a s e s y n d r o m e s h a s not r e -
-
..
* TRIC
= TRachoma Inclusion Conjunctivitis.
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
SYSTEMATIC
BACTERIOLOGY
55
solved the difficulties. While numerous s e r o t y p e s can be
distinguished on the basis of the specificityof cell wall antigens by using toxin-antitoxin reactions, growth-inhibition in
t i s s u e culture, o r complement fixation t e s t s , this approach
h a s failed t o produce a p a t t e r n suitable f o r taxonomic c l a s sification ( F r a s e r and B e r m a n 1965). P a s t experience indic a t e s that specific serology as a b a s i s f o r s p e c i e s s e p a r a tion leads t o long l i s t s of n a m e s which a s s i s t epidemiology
but d e t r a c t f r o m taxonomy, especially when e m b a r r a s s i n g
c r o s s reactions occur. DNA homology t e s t s a l s o have not
aided clsssification because of the high d e g r e e of homology
between t h e DNA's of different s t r a i n s (Gerloff e t g. 1966).
A solution t o this nomenclatural and taxonomic dilemma
was suggested by the work of Gordon and Quan (1965) who
reported that a number of chlamydia1 s t r a i n s r e c o v e r e d f r o m
a variety of d i s e a s e d t i s s u e s and hosts could be s e p a r a t e d
into two m a j o r groups on t h e b a s i s of morphological and
chemical c h a r a c t e r i s t i c s of the o r g a n i s m s alone. Gordon
and Quan observed (as had o t h e r s previously) t h a t o r g a n i s m s
a s s o c i a t e d with t r a c h o m a , inclusion conjunctivitis, lymphogranuloma venereum, and m o u s e pneumonitis developed
( inclusions i n
compact intracytoplasmic microcolonies
t i s s u e c e l l s ) which produced sufficient quantities of glycogen
detectable by staining with iodine solution. Lin and Moulder
(1966) confirmed these observations and added another apparently stable c h a r a c t e r i s t i c . They found that the multiplication in chicken e m b r y o s of the chlamydiae that f o r m e d
compact microcolonies and produced glycogen was inhibited
by sodium sulfadiazine.
The s t r a i n s in the nonglycogen
producing, sulfadiazine, r e s i s t a n t group w e r e o r g a n i s m s
frequently a s s o c i a t e d with psittacosis, meningopneumonitis,
guinea pig conjunctivitis, bovine encephalomyelitis, feline
pneumonitis, o r caprine pneumonitis.
The s e morphological and chemical c h a r a c t e r is t i c s we r e
t e s t e d recently and confirmed by the author f o r a group of
15 s t r a i n s of chlamydiae isolated f r o m m a n , t u r k e y s , a
pigeon, a s p a r r o w , p a r a k e e t s , a gull, a m o u s e , cattle, sheep
and a m u s k r a t . Representative s t r a i n s f r o m both biological
groups mentioned by Gordon and Quan w e r e a l s o tested. All
s t r a i n s fit the expected pattern ( s a v e the well known "6BC"
p a r a k e e t s t r a i n which i s glycogen negative but sensitive t o
sulfadiazine) and w e r e readily s e p a r a t e d into one o r the
other m a j o r group ( P a g e 1967, unpublished observations).
T h e r e s u l t s of Gordon and Quan and Lin and Moulder e s tablished the f i r s t n a t u r a l grouping of t h e s e o r g a n i s m s based
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
56
INTERNATIONAL JOURNAL
on r e l a t i v e l y s t a b l e m o r p h o l o g i c a l a n d c h e m i c a l c h a r a c t e r i s t i c s of t h e o r g a n i s m s t h e m s e l v e s , r a t h e r t h a n on v a r i a b l e
r e s p o n s e s of a n i m a l h o s t s t o infection o r on a m u l t i t u d e of
s p e c i f i c c e l l w a l l antigens.
Moreover, the c h a r a c t e r i s t i c s
u s e d a r e o n e s t h a t a r e s u b j e c t t o t e s t i n any l a b o r a t o r y
equipped t o i s o l a t e and study chlamydiae. T h e s e a r e condit i o n s that g r e a t l y a s s i s t t h e t a s k of taxonomy.
It is e x p e c t e d t h a t o t h e r r e l a t i v e l y s t a b l e c h a r a c t e r i s t i c s
of c h l a m y d i a e w i l l be r e c o g n i z e d and u t i l i z e d t o s u b s t a n t i a t e
e i t h e r t h e p r e s e n t dichotomous grouping or t o justify t h e
introduction of new groupings. Such i s the d y n a m i s m and
flexibility of taxonomy.
T h e r e f o r e , i t is p r o p o s e d t h a t two s p e c i e s be r e c o g n i z e d
f o r the o r g a n i s m s of t h e genus Chlamydia ( J o n e s , R a k e a n d
S t e a r n s 1945) which include t h e c a u s a t i v e a g e n t s of t r a c h o m a ,
p s i t t a c o s i s and r e l a t e d d i s e a s e s .
T h e s e two s p e c i e s a r e
Chlamydia t r a c h o m a t i s ( B u s a c c a ) Rake 1957, which is t h e
p r e v i o u s l y e s t a b l i s h e d type s p e c i e s , a n d Chlamydia p s i t t a c i
( L i l l i e ) comb. nov.
A definition of t h e genus and of e a c h
s p e c i e s , a s i m p l e key f o r t h e d i f f e r e n t i a t i o n of t h e s p e c i e s ,
a n d a n explanation of the l a b o r a t o r y t e s t s u s e d f o r t h e diffe r e n t i a t i o n , follow.
GENERAL LIMITS O F T H E ORGANISMS I N THE GENUS
CHLAMYDIA JONES, RAKE AND STEARNS 1945
Morphology a n d Mode of Reproduction. T h e o r g a n i s m s
a r e s p h e r o i d a l , bounded by -a r i g i d c e l l w a l l when m a t u r e ,
and nonmotile. T h e y have a unique, obligately i n t r a c e l l u l a r
d e v e l o p m e n t a l cycle c h a r a c t e r i z e d by t h e change of t h e p r i m a r y infectious o r g a n i s m i n the c y t o p l a s m of t h e h o s t c e l l
t o a r e t i c u l a t e thin walled s p h e r o i d , 0. 9-1.0 p i n d i a m e t e r ,
which divides by fission.
Division continues, while s o m e
daughter cells gradually reduce in s i z e t o become dense
s p h e r u l e s , 0. 3 p in d i a m e t e r , which a r e t h e f i n a l infectious
f o r m of t h e o r g a n i s m .
Intracytoplasmic microcolonies, o r
p l a q u e s , 2-12 p i n d i a m e t e r , m a y contain all s t a g e s of init i a l , i n t e r m e d i a t e , and m a t u r e f o r m s of the o r g a n i s m . T h e
o r g a n i s m s c a n be s e p a r a t e d i n t o two g r o u p s b a s e d onwhethe r t h e m i c r o c o l o n y i s r i g i d and c o m p a c t , o r w h e t h e r i t is
diffused throughout the h o s t c e l l ' s cytoplasm. When t h e h o s t
c e l l i s d a m a g e d , t h e infectious s p h e r u l e s a r e r e l e a s e d t o
r e p e a t the cycle in o t h e r h o s t c e l l s , although s e c o n d a r y
m i c r o c o l o n i e s m a y be e s t a b l i s h e d in a s i n g l e h o s t cell.
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
SYSTEMATIC
BACTERIOLOGY
57
C h e m i s t r y of t h e O r g a n i s m s . Both deoxyribo- and ribon u c l e i c a c i d s have been d e m o n s t r a t e d i n the o r g a n i s m s a t
a l l s t a g e s of growth. T h e i r c e l l w a l l s a r e s i m i l a r i n g e n e r a l
c h e m i c a l composition t o t h o s e of G r a m - n e g a t i v e b a c t e r i a .
T h e o r g a n i s m s have a negative G r a m r e a c t i o n and s t a i n
r e a d i l y by G i e m s a ' s , Macchiavello's, o r C a s t e n a d a ' s m e t h ods. S t r a i n s that f o r m c o m p a c t m i c r o c o l o n i e s c h a r a c t e r i s t i c a l l y produce glycogen t h a t i s d e m o n s t r a b l e by staining
t h e m i c r o c o l o n y with a solution of iodine-potassium iodide.
S t r a i n s that f o r m diffuse m i c r o c o l o n i e s fail t o p r o d u c e det e c t a b l e glycogen, O r g a n i s m s of a l l s t r a i n s have m e t a b o l i c
capabilities independent of the h o s t c e l l , s u c h as t h e catabolism of glucose t o c a r b o n dioxide when provided with e s s e n tial i n o r g a n i c and o r g a n i c c o f a c t o r s . T h e i n t r a c e l l u l a r m u l tiplication of o r g a n i s m s of e v e r y s t r a i n (except e x p e r i m e n t a l
m u t a n t s ) i s inhibited by t e t r a c y c l i n e antibiotics. T h e m u l t i pli c a t i on of glycogen- producing s t r a i n s i s cha ract e r is ti cally
inhibited by s o d i u m sulfadiazine.
E v e r y s t r a i n contains
a n antigeni cally s i m i l a r lipo-protein- c a rbohyd r a t e c o m p l e x
which is s t a b l e a t 100°C. S t r a i n - s p e c i f i c a n t i g e n s a r e found
i n t h e c e l l walls.
N a t u r a l Habitat and Cultivation. T h e o r g a n i s m s a r e p a r a s i t e s of t i s s u e c e l l s of v e r t e b r a t e s , although they m a y be
found in o r on e c t o p a r a s i t i c a r a c h n i d s . T h e y a r e t r a n s m i t t e d
between v e r t e b r a t e h o s t s by t h e inhalation' of infectious exc r e m e n t s of b i r d s and mammals, by contamination of conjunctiva o r o t h e r m u c o u s m e m b r a n e with infectious u r o g e n i
t a l exudates during b i r t h o r v e n e r e a l contact. I n t r a v e n o u s
t r a n s m i s s i o n m a y t a k e place m e d i a t e d by sanguivorous
a r t h r o p o d s . T h e o r g a n i s m s may p r o d u c e g e n e r a l i z e d infections i n a given h o s t , but s o m e s t r a i n s l o c a l i z e and c a u s e
pronounced c e l l u l a r changes i n one o r m o r e t i s s u e s o r o r g a n s , s u c h a s , the conjunctiva, e y e s , lungs, liver, s p l e e n ,
v a s c u l a r endothelium, joints , u r e t h r a , vagina, lymph nodes,
f e t a l and placental m e m b r a n e s , m e n i n g e s , o r brain. A f t e r
a p p r o p r i a t e adaptation, all of the s t r a i n s m a y be p r o p a g a t e d
i n t h e yolk s a c of chicken e m b r y o s o r i n c u l t u r e s of v e r t e b r a t e t i s s u e s . T o d a t e , e x t r a c e l l u l a r propagation of the o r g a n i s m s h a s not been achieved.
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
58
INTERNATIONAL JOURNAL
DEFINITION O F THE TYPE SPECIES AND ONE
ADDITIONAL SPECIES I N THE GENUS CHLAMYDIA
1. Chlamydia t r a c h o m a t i s ( B u s a c c a ) Rake 1957, 958.
(Rickettsia t r a c h o m a e (sic) Busacca 1935,567; R i c k e t t s i a
t r a c h o m a t i s F o l e y and P a r r o t 1937,230; Chlamydozoon
t r a c h o m a t i s Moshkovsky 1945,18; Chlamydazoon t r a c h o m i a t i
( s i c ) Ryzhkov 1950,17. )
T h e following s p e c i e s previously d e s c r i b e d i n B e r g e y ' s
7th Edition and e l s e w h e r e a r e h e r e c o n s i d e r e d putative synonyms of _C. t r a c h o m a t i s ( B u s a c c a ) Rake:
Chlamydia oculogenitalis (Moshkovsky, 1945) Rake 1957;
Chlamydozoon oculopenitale Moshkovsky op. cit. ;
Miyagawanella lymphogranulomatosis, Brumpt, Ann. de
P a r a s i t o l . 16, 1938, 155;
E h r l i c h i a (Miyagawanella) lymphogranulomatosis Moshkovsky, op. cit. ;
Ricketts i a f o r m i s lymphogranulomatis Zhdanov and
Korenblit, op. cit. ;
Chlamydozoon lymphophilus , Ryzhkov, op. cit. ;
Miyagawanella bronchopneumoniae (Moshkovs ky, 1945)
Rake 1948, 1118;
E h r l i c h i a bronchopneumoniae Moshkovsky, op. cit. ;
Cystidium bronchopneumoniae m u r i s Ruska (Ruska and
Poppe 1947);
Cystidium gonne rtianum, Rus ka, op. cit. ;
R i c k e t t s i a f o r m i s m u r i s , Zhdanov and Korenblit, op. cit. ;
Chlamydozoon m u r i n u s Ryzhkov, op. cit.
tra. cho'ma. tis. Gr. n. t r a c h o m a roughness; M. L. n.
t r a c h o m a t r a c h o m a ; M. L. gen. n. t r a c h o m a t i s of t r a c h o m a .
Spheroidal, nonmotile o r g a n i s m s whose d i a m e t e r s range
f r o m 0. 3 t o 0. 9 p depending upon t h e i r s t a g e of development
in a n obligately i n t r a c e l l u l a r multiplication cycle. During
the f i r s t two t o t h r e e d a y s of growth, the o r g a n i s m s f o r m
rigid , compact i n t r a c y t o p l a s m i c m i c r o c o l o n i e s which m a y
contain o r g a n i s m s i n all s t a g e s of development. In t i m e ,
s e v e r a l hundred m a t u r e infectious f o r m s , which a r e small,
d e n s e s p h e r u l e s 0.3 p i n d i a m e t e r , m a y e m e r g e f r o m rupt u r e d plaques t o f i l l t h e h o s t cell's cytoplasm. S e c o n d a r y
m i c r o c o l o n i e s m a y be formed. T h e s p h e r u l e s a r e r e l e a s e d
t o infect o t h e r c e l i s when the h o s t c e l l is damaged. F o r a
limited period (two o r t h r e e d a y s ) glycogen is a c c u m u l a t e d
i n the i n t r a c e l l u l a r plaque i n sufficient quantities t o be
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
SYSTEMATIC
BACTERIOLOGY
59
demonstrable microscopically a f t e r fixing the iflfected host
cells i n methyl alcohol and then staining t h e m with an alcoholic solution of W0.iodine-potas s i u m iodide. The o r g a n i s m s
a r e Gram-negative and can be readily s e e n microscopically
a f t e r staining them by G i e m s a ' s method. In cell c u l t u r e s ,
the o r g a n i s m s may be s e e n without any staining i f phase
contrast optics a r e used.
Multiplication of the o r g a n i s m s in the yolk s a c m e m b r a n e
of the chick e m b r y o is inhibited at l e a s t one hundred fold by
The d e g r e e of inhibition is
1 m g of sodium sulfadiazine.
determined by a dual t i t r a t i o n of a suspension of o r g a n i s m s
containing a t l e a s t one million e m b r y o lethal doses 50% p e r
ml i n chicken embryos by the yolk s a c route i n the p r e s e n c e
and absence of 1 mg of sodium sulfadiazine (inoculated separately into the yolk s a c ) p e r embryo. Intracellular multiplication of the o r g a n i s m s is inhibited by tetracycline also.
The o r g a n i s m s contain a lipo-protein-carbohydrate complex that i s antigenically s i m i l a r to a complex found i n all
s t r a i n s of both s p e c i e s of the genus. The antigen is stable
a t 100°C and r e a c t s with antibodies against any other s t r a i n
t o fix complement.
Various s e r o t y p e s , however, can be
distinguished on the basis of the antigenic specificity of components of the o r g a n i s m s ' c e l l walls.
The o r g a n i s m s a r e p a r a s i t e s of t i s s u e c e l l s of v e r t e b r a t e s
and m a y be t r a n s m i t t e d between v e r t e b r a t e hosts, especially
man, by t r a n s f e r of ophthalmic o r urogenital exudates f r o m
m e m b r a n e s of the susceptible host. T r a n s m i s s i o n o c c u r s
through various f o r m s of contact contamination such a s that
of a fetus during birth o r that of mucous m e m b r a n e s during
v e n e r e a l contact. In man, the o r g a n i s m s m a y localize i n
one o r m o r e t i s s u e s o r organs and cause pronounced cellular changes i n the conjunctiva, cornea, synovial m e m b r a n e s ,
u r e t h r a and adjoining lymph nodes, meninges, or brain.
S t r a i n s of ig.' t r a c h o m a t i s a r e usually responsible f o r various d i s e a s e s o r syndromes in m a n called t r a c h o m a , inclusion conjunctivitis , lymphogranuloma venereum, and m a y
include a r t h r i t i s and meningitis. One s t r a i n c a u s e s endemic
pneumonitis i n colonies of m i c e and is probably t r a n s m i t t e d
via the r e s p i r a t o r y route.
The o r g a n i s m s m a y be isolated and propagated in the
yolk s a c of chicken embryos incubated at 35 to 37OC. S e r i a l
blind passage of the culture in chicken e m b r y o s may be nec e s s a r y before any effects on the e m b r y o can be observed.
Some s t r a i n s have a stenothermal r e q u i r e m e n t growing only
a t 35°C 1OC. They m a y be propagated in cultures of v e r -
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
60
INTERNATIONAL JOURNAL
t e b r a t e t i s s u e s . S t r a i n s t h a t a r e f r e q u e n t l y a s s o c i a t e d with
o p h t h a l m i c d i s e a s e s of m a n m a y be p r o p a g a t e d i n t h e lungs
of m i c e and guinea pigs, and t h e s k i n of guinea pigs. In subhuman p r i m a t e s they readily cause ophthalmic abnormalities
a n d i n s o m e c a s e s m a y d i s s e m i n a t e throughout the body via
t h e blood s t r e a m . O t h e r s t r a i n s c a n be p r o p a g a t e d i n t h e
lungs o r b r a i n s of young m i c e a f t e r inoculation by t h e app r o p r i a t e route.
2. Chlamydia p s i t t a c i ( L i l l i e ) comb. nov.
(Rickettsia paittaci, Lillie, 1930,773; Microbacterium
m u l t i f o r m e p s i t t a c o s i s , Levinthal, 1930,523; E h r l i c h i a
p s i t t a c i Moshkovsky, 1956,18; R i c k e t t s i a f o r m i s p s i t t a c o s i s
Zhdanov and K o t e n b l i t , 1950,43; Chlamydozoon p s i t t a c i
Ryzhkov, 1950,17. )
H e r e i n c o n s i d e r e d a s putative s y n o n y m s a r e :
Miyagawanella p s i t t a c i , M. o r n i t h o s i s , 3. pneumoniae,
M. f e l i s , ,M. l o u i s i a n a e , M. i l l i n i i , M. o p o s s u m i , &I.
M. b o v i s , a n d M . p e c o r i s which Rake placed i n t h e genus
Miyanawanella i n t h e 7th edition of B c r g e y ' s Manual ( 1 957);
R i c k e t t s i a f o r m i s o r n i t h o s i s , 3.p n e u m o n i a e , _R. m e n i n g o p n e u m o n i a e , and &. f a s , Zhdanov a n d K o r e n b l i t , op. cit.
a n d R . c o l u m b a r u m , 5. o p o s s u m , &. bovis, 3. o v i s y
Zhdanov, 1953; Chlamydozoon S
i
, ,C. f e l i s ,
mening o p h i l u s , ,C. c r i c e t i s , ,C. h o m i n i s , ,C. o s t e o p h i l u s , G.
o p o s s u m i , and C. bovis, Ryzhkov, op. cit.
a,
-
c.
psit'ta. ci. G r . n. p s i t t a c u s a p a r r o t ; M. L. gen. n.
p s i t t a c i of a p a r r o t .
S p h e r o i d a l , nonmotile o r g a n i s m s , 0.3 t o 1. 0 p i n d i a m e t e r , r e s e m b l i n g t h o s e of C h l a m y d i a t r a c h o m a t i s . D u r i n g
t h e f i r s t day of obligate, i n t r a c e l l u l a r growth, t h e o r g a n i s m s
f o r m l o o s e l y bound int r a c y t o p l a s m i c m i c r o c o l o n i e s which
give t h e o r g a n i s m s t h e a p p e a r a n c e of being d i s t r i b u t e d about
c y t o p l a s m of t h e h o s t cell.
In t h i s r e s p e c t t h e y d i f f e r f r o m
C h l a m y d i a t r a c h o m a t i s , but t h e m o d e of m u l t i p l i c a t i o n i s the
s a m e . T h e m i c r o c o l o n i e s d o not f o r m any glycogen d e t e c t a b l e by iodine staining.
The organisms a r e Gram-negative
and c a n be r e a d i l y s e e n m i c r o s c o p i c a l l y after s t a i n i n g by
G i e m s a ' s , M a c c h i a v e l l o ' s o r C a s t e n a d a ' s m e t h o d s , , o r without s t a i n i n g i f p h a s e c o n t r a s t o p t i c s a r e used. .
Multiplication of t h e o r g a n i s m s i n t h e yolk s a c of chicken
e m b r y o s is inhibited by t e t r a c y c l i n e a n t i b i o t i c s but not by
s o d i u m sulfadiazine.
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
SYSTEMATIC
BACTERIOLOGY
61
T h e o r g a n i s m s contain a l i p o - p r o t e i n - c a r b o h y d r a t e c o m plex t h a t i s antigenically s i m i l a r t o that found in a l l s t r a i n s
of Chlamydia. T h e antigen c a n be used t o d e t e c t c o m p l e m e n t fixing antibodies t o t h i s c o m p l e x s t i m u l a t e d by any
other strain.
N u m e r o u s s e r o t y p e s , h o w e v e r , can be d i s tinguished on the b a s i s of the antigenic s p e c i f i c i t y of the
components of the c e l l w a l l s of the o r g a n i s m s . T h e s e antigens c a n be s e p a r a t e d when r e a c t e d s e p a r a t e l y with homologous and h e t e r o l o g o u s antibodies i n toxin p r o t e c t i o n t e s t s i n
m i c e , growth inhibition t e s t s i n t i s s u e c u l t u r e s and complem e n t fixation t e s t s .
T h e o r g a n i s m s a r e p a r a s i t e s of . t i s s u e c e l l s of v e r t e brates.
T h e y m a y be t r a n s m i t t e d between v e r t e b r a t e h o s t s
by inhalation of a e r o s o l s of n a s a l o r f e c a l e x c r e m e n t s of inf e c t e d b i r d s o r m a m m a l s , by v a r i o u s f o r m s of contact cont a m i n a t i o n possibly including the bite of s a n g u i v o r o u s a r a c h n i d s , and by t r a n s o v a r i a n p a s s a g e t o t h e fetus i n m a m m a l s .
T h e o r g a n i s m s g e n e r a l l y p r o d u c e s y s t e m i c infection i n a
b r o a d r a n g e of a v i a n and m a m m a l i a n h o s t s , although s o m e
s t r a i n s a p p e a r t o have a n a r r o w h o s t s p e c t r u m . R e s p i r a t o r y
t r a n s m i s s i o n of the o r g a n i s m s is r e a d i l y a c c o m p l i s h e d when
a s u s c e p t i b l e h o s t i n h a l e s fine d u s t containing d r i e d n a s a l o r
cloaca1 e x c r e m e n t of infected b i r d s .
T h i s m e t h o d h a s been
r e s p o n s i b l e f o r e p i d e m i c s of p s i t t a c o s i s i n m a n and o t h e r
animals.
T h e o r g a n i s m s have been d e t e c t e d i n o v e r 100
s p e c i e s of wild and d o m e s t i c b i r d s , m o s t d o m e s t i c a t e d animals, and i n m a n y wild mammals. S o m e of t h e d i s e a s e s
c a u s e d by v a r i o u s s t r a i n s of Chlamydia p s i t t a c i have been
d e s c r i b e d as: p s i t t a c o s i s , o r n i t h o s i s , meningopneumonitis;
feline, bovine , ovine , p o r c i n e , c a p r i n e , o r human pneumon i t i s ; o p o s s u m e n c e p h a l i t i s ; s p o r a d i c bovine encephalomyel i t i s ; ovine, bovine , o r p o r c i n e a r t h r i t i s ; epizootic bovine
a b o r t i o n ; enzootic a b o r t i o n of e w e s ; guinea pig conjunctivitis;
f a t a l epizootic d i s e a s e of h a r e s and m u s k r a t s .
In g e n e r a l i z e d infections, t h e o r g a n i s m s m a y m u l t i p l y
i n i t i a l l y i n e p i t h e l i a l c e l l s of s e v e r a l o r g a n s , e s p e c i a l l y the
l o w e r r e s p i r a t o r y t r a c t , and then a r e d i s t r i b u t e d throughout
t h e body via the blood s t r e a m . D a m a g e t o v a s c u l a r endothelia
c a u s e s s p r e a d of the o r g a n i s m s and p l a s m a t o t h e s u r f a c e s
of m a n y o r g a n s w h e r e f i b r i n o u s i n f l a m m a t o r y e x u d a t e s a c c u m u l a t e r e s u l t i n g i n c h a r a c t e r i s t i c l e s i o n s . C o m m o n findings accompanying pneumonitis a r e p e r i c a r d i t i s , p e r i h e p a t i t i s , p e r i t o n i t i s , a s c i t e s , hepatomegaly, s p l e n o m e g a l y , and
o t h e r s i g n s of g e n e r a l i z e d infection.
The organisms cause
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
62
INTERNATIONAL
JOURNAL
a wide range of e f f e c t s depending upon a l l of the c l a s s i c
f a c t o r s affecting h o s t - p a r a s i t e r e l a t i o n s h i p s . Many h o s t s of
v a r i o u s s p e c i e s b e c o m e a s y m p t o m a t i c c a r r i e r s of Chlamyd i a p s i t t a c i t h e r e b y perpetuating t h e o r g a n i s m s in n a t u r e .
S o m e s t r a i n s localize in c e r t a i n t i s s u e s o r o r g a n s of the
h o s t c a u s i n g splenomegaly, o r t r a n s i e n t pneumonitis, a r t h r i t i s , f e t a l o r placental inflammation r e s u l t i n g i n abortion.
T h e o r g a n i s m s a r e r e a d i l y i s o l a t e d and propagated i n t h e
yolk s a c s of chicken e m b r y o s incubated a t 37 t o 40'C. Many
s t r a i n s r e c o v e r e d f r o m mammals with high n o r m a l body
t e m p e r a t u r e s have o p t i m a l growth t e m p e r a t u r e s of 3 9 ° C o r
above. Most s t r a i n s c a n be propagated i n t h e lungs, b r a i n s ,
o r p e r i t o n e a l cavity of l a b o r a t o r y m i c e o r guinea pigs.
S t r a i n s c a n be grouped a c c o r d i n g t o t h e i r d i s e a s e producing
c a p a b i l i t i e s in m i c e , guinea pigs and pigeons.
T h e o r g a n i s m s c a n be propagated in c u l t u r e s of v e r t e b r a t e t i s s u e s , and s o m e s t r a i n s produce d e s t r u c t i o n of h o s t
cells after s e r i a l passage.
All s t r a i n s c a n be p r e s e r v e d indefinitely a f t e r propagation in chicken e m b r y o s by s t o r i n g infected yolk s a c s a t
-70°C;h o w e v e r , the l a r g e a m o r p h o u s f o r m of t h e o r g a n i s m
which develops i n the e a r l y s t a g e s of multiplication i s inact i v a t e d by f r e e z i n g .
Key t o the s p e c i e s of Chlamydia
I. I n t r a c y t o p l a s m i c m i c r o c o l o n y is rigid and
compact
ranging f r o m 2-10 )I i n diameter. T h e plaque contains
glycogen detectable by staining with iodine. Growth i n
t h e yolk sac of t h e chicken e m b r y o is inhibited by
s o d i u m sulfadiazine (1 m g p e r e m b r y o ) .
Chlamydia t r a c h o m a t i s ( B u s a c c a ) Rake
......
LI. I n t r a c y t o p l a s m i c m i c r o c o l o n y is nonrigid, i r r e g u l a r
in s h a p e and u p t o 12 p wide. T h e m i c r o c o l o n y does
not contain glycogen d e t e c t a b l e by staining with iodine.
Growth i n the yolk s a c of t h e chicken e m b r y o is not
inhibited by s o d i u m sulfadiazine ( 1 m g p e r e m b r y o ) .
Chlamydia p s i t t a c i ( L i l l i e ) comb. nov.
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
SYSTEMATIC
BACTERIOLOGY
63
T e s t s f o r the Identification and Differentiation
of Chlamydiae
New i s o l a n t s of c h l a m y d i a e c a n be conclusively identified
a s Chlamydia t r a c h o m a t i s o r Chlamydia p s i t t a c i by t h r e e
tests. These are:
( 1 ) Identification of t h e o r g a n i s m s as m e m b e r s of the
genus Chlamydia.
A p u r e c u l t u r e of t h e o r g a n i s m s which have b e e n shown t o
be unable t o colonize on l i f e l e s s b a c t e r i o l o g i c m e d i a , e. g.,
blood a g a r , i s propagated i n the yolk s a c of chicken e m b r y o s a t a p p r o p r i a t e incubation t e m p e r a t u r e s ( 3 5 - 3 7 " C f o r
human urogenital t r a c t s t r a i n s ; 37-39°C f o r strains f r o m
a n i m a l s with n o r , m a l body t e m p e r a t u r e s t h a t a r e h i g h e r than
3 7 ° C ) . At t h e t i m e of death o r obvious r e t a r a a t i o n of growth
of infected e m b r y o s , t h e yolk s a c s a r e r e m o v e d , t r i t u r a t e d
with s a n d and a buffered diluent (pH 7. 0 ) t o m a k e a 20 t o 30%
suspension. T h e s u s p e n s i o n i s boiled f o r 30 m i n u t e s , cooled,
a n d a q u e o u s 5% phenol is a d d e d t o m a k e a f i n a l c o n c e n t r a t i o n
of 0. 5T0 phenol. One day l a t e r , t h e s u s p e n s i o n i s t i t r a t e d i n
a c o m p l e m e n t fixation t e s t a g a i n s t a c o n s t a n t dilution of
s e r u m p r e v i o u s l y shown t o contain 8 c o m p l e m e n t fixing units
of c h l a m y d i a l g r o u p antibodies p e r unit volume. T h e boiled
and phenolized s u s p e n s i o n is a l s o t i t r a t e d a g a i n s t s e r u m
p r e v i o u s l y shown not t o contain s u c h antibodies. F i x a t i o n of
c o m p l e m e n t a t dilutions of antigen beyond any a n t i c o m p l e m e n t a r y l e v e l i s conclusive evidence that the yolk s a c s cont a i n e d c h l a m y d i a l o r g a n i s m s . Usually a m i n i m u m of 1 0 , 0 0 0
o r g a n i s m s i s sufficient t o p r o d u c e c o m p l e t e fixatiori of two
e x a c t units of c o m p l e m e n t i n t h e p r e s e n c e of one C F unit of
antibody.
Antigens p r e p a r e d by boiling and phenolizing c h l a m y d i a l
o r g a n i s m s grown in c u l t u r e s of v e r t e b r a t e t i s s u e s a l s o m a y
be u s e d in s i m i l a r t e s t s f o r detection of the c h l a m y d i a l
g r o u p antigen, but s u c h a n t i g e n s a r e usually of l o w e r t i t e r .
( 2 ) T e s t f o r t h e p r e s e n c e of glycogen in i n t r a c y t o p l a s m i c
m i c r ocolonie s
.
T h e c h l a m y d i a e a r e p r o p a g a t e d in c u l t u r e s of v e r t e b r a t e
tissues.
The tissue cultures a r e examined microscopically
at 1, 2 and 3 d a y s a f t e r inoculation t o a s c e r t a i n t h e p r e s e n c e
of i n t r a c y t o p l a s m i c c l u s t e r s of c h l a m y d i a e o r m i c r o c o l o n i e s .
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
64
INTERNATIONA
L JOURNAL
T h e f o r m a t i o n of glycogen by t h e m i c r o c o l o n y is a t r a n s i e n t
phenomenon t h a t o c c u r s f r o m t h e f i r s t through t h e t h i r d day
of growth. During t h i s period, infected t i s s u e c e l l s growing
on c o v e r s l i p s a r e r e m o v e d intact and fixed in a b s o l u t e m e t h y l alcohol f o r a m i n i m u m of four h o u r s .
The c o v e r s l i p s
a r e then i m m e r s e d in a 1:l m i x t u r e of 5. 0% iodine in 100’?70
e t h y l alcohol and aqueous 5.0% p o t a s s i u m iodide f o r f o u r
h o u r s . T h e c o v e r s l i p s a r e t h e n placed c e l l s i d e down on a
d r o p of the iodine m i x t u r e in 50% g l y c e r i n on a m i c r o s c o p e
slide.
At magnifications g r e a t e r t h a n 200X, m i c r o c o l o n i e s
containing glycogen w i l l a p p e a r brown a g a i n s t a light t a n
background. Nonglycogen f o r m i n g m i c r o c o l o n i e s cannot be
distinguished f r o m o t h e r c e l l u l a r m a t e r i a l by t h i s iodine
staining method. It is i m p o r t a n t however t h a t t h e r e be c o r a l l a r y proof that growth and m u l t i p l i c a t i o n of the o r g a n i s m s
a c t u a l l y o c c u r r e d i n t h e t i s s u e c u l t u r e s before a s t r a i n of
chlamydiae i s d e c l a r e d glycogen negative. Infected t i s s u e
c u l t u r e s m u s t be incubated at t e m p e r a t u r e t h a t i s o p t i m a l
f o r growth of t h e o r g a n i s m s .
( 3 ) T e s t f o r s u s c e p t i b i l i t y of chlamydia1 s t r a i n s t o
s o d i u m sulfadiazine.
D e c i m a l dilutions of a s u s p e n s i o n of c h l a m y d i a e p r e viously propagated i n 6-day-old chicken e m b r y o s a r e inoculated i n t o two groups of chicken e m b r y o s .
One g r o u p of t h e
e m b r y o s is inoculated via . t h e yolk sac r o u t e with 1 m g of
s o d i u m sulfadiazine in a diluent buffered a t pH 7.0.
The
second g r o u p of e m b r y o s is inoculated via t h e yolk sac with
a n e q u a l volume of the diluent without sulfadiazine. The
e m b r y o s a r e candled twice daily and d e a t h s r e c o r d e d f o r 14
d a y s , at which t i m e a calculation of the LDSo/unit volume of
t h e o r i g i n a l s u s p e n s i o n is m a d e f o r both s e t s of e m b r y o s .
F o r p u r p o s e s of taxonomic differentiation, s u s c e p t i b i l i t y t o
sulfadiazine is defined a s a hundredfold o r m o r e reduction
i n t h e n u m b e r of e m b r y o l e t h a l d o s e s 50% in a s u s p e n s i o n of
one m i l l i o n o r m o r e c h l a m y d i a e p e r unit volume t i t r a t e d in
chicken e m b r y o s i n the p r e s e n c e and a b s e n c e of 1 m g of
s o d i u m sulfadiazine. T h i s m e a n s that t h e ELDSO d e t e r m i n a t i o n s f o r the t i t r a t i o n s should d i f f e r by a f a c t o r of 100 o r
more.
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
SYSTEMATIC
BACTERIOLOGY
65
REFERENCES
B r u m p t , E. 1938. R i c k e t t s i a i n t r a c e l l u l a i r e s t o m a c a l e
( R i c k e t t s i a c u l i c i s n. sp. ) de Culex fatigans. Ann. d e
P a r a s i t o l o g i e 16:153-158.
B u s a c c a , A. 1935. Un g e r m e a u x c a r a c t e r e s de r i c k e t t s i e s
( R i c k e t t s i a t r a c h o m a e ) d a n s l e s tis s u s t r a c h o m a t e u x .
A r c h . Ophthal., P a r i s 52:567-572.
Dunlop, E . M. C . , L A . H a r p e r , M. Khalaf a l - H u s s a i n i , J.
A. G a r l a n d , J . D . T r e h a r n e , D. J . M . Wright, and B.R.
J o n e s . 1966. Relation of t r i c agent t o "non-specific"
genital infection. Brit. J. Ven. Dis. %:77-87.
F o l e y , H. a n d L. P a r r o t . 1937. S u r l a R i c k e t t s i a du t r a chome. C. R. SOC. Biol. ( P a r i s ) =4:230-232.
F r a s e r , C. E . 0. and D. B e r m a n . 1965. Type s p e c i f i c antigens i n the psittacosis-lymphogranuloma v e n e r e u m g r o u p
of o r g a n i s m s . J. Bact. 89:943-948.
G e r l o f f , R . K . , D. B. R i t t e r , and R.O. Watson. 1966. DNA
homology between t h e meningopneumonitis agent and
r e l a t e d m i c r o o r g a n i s m s . J. Inf. Dis. =:197-202.
Gordon, F. B. and A. L. Quan. 1965. O c c u r r e n c e of glycogen i n inclusions of t h e psittacosis-lymphogranuloma
v e n e r e u m - t r a c h o m a agents. J. Inf. Dis. 115:186-196.
J o n e s , H., G. Rake a n d B. S t e a r n s . 1945. Studies on lymphogranuloma v e n e r e u m . LZI. T h e action of t h e sulfonam i d e s on t h e a g e n t of lymphogranuloma venereum.
J. Inf. Dis. 2 : 5 5 - 6 9 .
Levaditi, J . C . , F. R o g e r , a n d P. Destombes. 1964. T e n t a t i v e de c l a s s i f i c a t i o n d e s C h l a m y d i a c e a e (Rake, 1955)
t e n a n t e c o m p t e d e l e u r s affinites t i s s u l a i r e s et d e l e u r
epidemiologie. Ann. Inst. P a s t e u r =:656-662.
Levinthal, A. 1930. Die Aetiologie d e r P s i t a k o s . 1st
Intern. Congr. Microbiol. P a r i s L:523-528.
L i l l i e , R. D. 1930. P s i t t a c o s i s - r i c k e t t s i a - l i k e inclusions i n
m a n a n d i n e x p e r i m e n t a l a n i m a l s . Publ. Hlth. Rep.
Wash. 2 : 7 7 3 - 7 7 8 .
L i n , H. and J. W. Moulder. 1966. P a t t e r n s of r e s p o n s e t o
sulfadiazine, D - c y c l o s e r i n e and D-alanine in m e m b e r s
of t h e p s i t t a c o s i s group. J. Inf. Dis. L6:372-376.
M e y e r , K.F. 1953. P s i t t a c o s i s Group. Ann. N. Y. Acad.
Sci. 2 : 5 4 5 - 5 5 6 .
Moulder, J. W. 1964. T h e p s i t t a c o s i s g r o u p a s b a c t e r i a .
J. Wiley and Sons, New York. pp. 1-95.
1966. The r e l a t i o n of t h e p s i t t a c o s i s g r o u p (Chlamyd i a e ) t o b a c t e r i a and v i r u s e s . Ann. Rev. Microbiol. 2 0 :
107- 130.
-.
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19
66
INTERNATIONAL JOURNAL
Moshkovs ky , S. D. 1945. T h e cytatropic a g e n t s of infections
and t h e position of t h e r i c k e t t s i a e i n t h e s y s t e m of
Chlamydozoa. (in R u s s i a n ) Advances in Modern Biology
( i n Russian). Uspekhi Souremennoi Biol. l9:1-44.
P a g e , L. A. 1966. Revision of t h e family Chlamydiaceae
Rake ( R i c k e t t s i a l e s ) ; unification of t h e p s i t t a c o s i s lymphogranuloma v e n e r e u m - t r a c h o m a g r o u p of o r g a n i s m s
i n t h e genus Chlamydia J o n e s , Rake and S t e a r n s , 1945.
Int. J. Syst. Bact. 5 : 2 2 3 - 2 5 2 .
1967. C o m p a r i s o n of "pathotypes'' among chlamydia1
( p s i t t a c o s i s ) s t r a i n s r e c o v e r e d f r o m diseas'ed b i r d s and
mammals. Bull. Wildl. Dis. Assoc. 3:166-175.
Rake. G. 1948. F a m i l y Chlamydiaceae Moshkovsky.
Bergey's Manual of Determinative Bacteriology. 6th ed.
pp. 1119-1120. W i l l i a m s and Wilkins Co. , B a l t i m o r e , Md.
1957. F a m i l y 11. Chlamydiaceae Rake fam. nov. In:
B e r g e y ' s Manual of Determinative Bacteriology, 7th ed.
pp. 957-.968. W i l l i a m s and Wilkins, B a l t i m o r e , Md.
Ruske, H. and K. 'Poppe. 1947. E l e k t r o n e n m i k r o s k o p i s c h e
Untersuchungen z u r Morphologie d e r S e i f f e r t s c h e n Mikroo r g a n i s m e n und des E r r e g e r s d e r Lungenseuche d e s
Rindes. Z e i t s c h r . f. Hygiene =7:201-215.
Ryzhkov, V. L. 1950. Study on s y s t e m a t i c of v i r u s e s . (in
Russian. ) Voprosy Meditsinskoi Virusol. Moskau 2:9-19.
S c h a c h t e r , J. 1967. A bedsonia i s o l a t e d f r o m a patient with
c l i n i c a l lymphogranuloma vene reum. Am. J. Ophthomol.
6 3 ~ 1 0 4 9 -1052.
Zhdanov, V. and R. S . Korenblit. 1950. S y s t e m a t i c s and
n o m e n c l a t u r e of v i r u s e s . Z hur. Mi k r obiol. Epidemiol.
Immunobiol. 9:40-44.
( R e p o r t at scientific conference
Ukrainian Inst. i m e n i Mechnikov i n Kharkov 11 October
In:
.
.
-
1949).
.
1953. Guide t o the human and a n i m a l v i r u s e s . (in
R u s s i a n ) Akad. Med. Nauk. SSSR M O X C O Wpp.
~ 1-348.
Downloaded from www.microbiologyresearch.org by
IP: 88.99.165.207
On: Sat, 17 Jun 2017 22:27:19