INTERNATIONAL JOURNAL O F
SYSTEMATIC BACTERIOLOGY
A p r i l 1969
pp. 183-187
Vol. 19, No. 2
Copyright 1969, Iowa State University P r e s s
MENAQUINONES IN THE CLASSIFICATION O F
MICROCOCCACEAE, WITH OBSERVATIONS
ON THE APPLICATION O F LYSOZYME AND
NOVOBIOCIN SENSITIVITY TESTS
Leonard Jeff ri e s
Beecham R e s e a r c h L a b o r a t o r i e s
Vitamins Research Station, Tadworth, Surrey, England
Menaquinones, vitamins of t h e K, group, a r e 2 methyl
1, 4-naphthoquinones with unsaturated polyisoprenoid side
chains ( r e f e r r e d to a s "normal menaquinones, I f o r MK);
partially hydrogenated f o r m s a r e a l s o found with at l e a s t
one isoprene unit s a t u r a t e d ("hydrogenated menaquinones, I '
o r MK(H)). Isoprenologues a r e designated by t h e number
(n) of i s o p r e n e units in the side chain (Fig. 1).
0
0
F i g u r e 1.
One hundred and twenty-nine s t r a i n s of a e r o b i c M i c r o coccaceae w e r e obtained f r o m culture collections; values
for guanine t cytosine i n DNA (GC r a t i o s ) w e r e available on
about a t h i r d of t h e s e s t r a i n s , including all the pigmented
m i c r o cocci Staphylococci, m i c r o c oc c i and s t r a i n s i n t e r
mediate in GC content between t h e s e g e n e r a w e r e a l l found
t o contain e i t h e r n o r m a l o r hydrogenated menaquinones;
however, the amounts e x t r a c t e d v a r i e d f r o m about 400 ug/g
wet weight i n Staphylococcus a u r e u s to a few ug in m a r i n e cocci and e x t r e m e halophiles.
.
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184
INTERNATIONAL JOURNAL
Table 1.
Distribution of menaquinones in Microcococcaceae, sensitivity to
lysozyme and novobiocin.
X, ratio
3.
of
w
3
2
5
4
4
-
4
S.aureus(Bmm e t ol,l%7.acetoin negative bovine strains)
Miemcoccus
Baird-Parker.1965. group 2
subgroup 1
"
"
3
2
2
3
2
"
'I
"
5
3
"(Staphylococcus laotis, NCTC 189)
1
6
4
3
"
"
"
?(norr-piwnted)
18
1 1 ( ~
" "
"
)I
1
(8tephylocoocus.
NCTC 7292)
(I8
.
11
,mc8!+56)
I'
,CCY 678)
1
1
13
(yeUow-pigaented,Roaypal et al(U66),aubgroup l a )
n
n
n
11
II
It
I1
Rosypal et &1%6
1
9,
proposed type st&
Y. luteus,ATCC 398)
B(pinlr-pi~nted, roaeua ROSmal e t sl(l966)subgmup lb
Emh&ek e t al(1" ( p M r - p i ~ n t e d , M . c c ~ ~ ~ , A T966)
CC
70.&73.3
69.0-7l.q
69.0:
s u b p u p Za(yellow-pigpented)
Ja( grey-violet p i e n t e d , hi. cyaneu8,CCY 856)
n
~ ( C C M23,strpin ~f w e r t e i n classification)
67.5-69.5
uicrococcua morrhuae
57.1-61.4
67.0;-
56.4f!
Flanccoccua
Yicrococcua eucinetua
llicrocooous spp.
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l 7
1
4
1
1
4
2
SYSTEMATIC BACTERIOLOGY
(Table 1 cont., legends shown on following page)
rx-9
-
U(3-29
0
0
4(0-2 )
0
0
0
0
0
0
0
0
0
0
0
0
85
0
0
0
0
-
lZ(0-25)
0
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185
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INTERNATIONAL JOURNAL
Legends for Table 1.
Mean values for the number of strains shown, with ranges
in parenthesis,
MK-n(H) indicates a tentative identification of those compounds a s menaquinones with partially hydrogenated side
chains.
Minimum inhibitory concentrations in nutrient agar:
HR, > 5,000 ug/ml; R, > 300"5,000 ug/ml; MR, >25-300
ug/ml; S, <12-25 ug/ml; NT, not tested.
Minimum inhibitory concentrations in nutrient agar:
R, > 2.5 ug/ml; MR, > 0.6-2.5 ug/ml; S, 0.07-0.6 ug/ml;
NT, not tested.
-
- - - - - - - -
Distinct menaquinone patterns, formed by the percentages of individual isoprenologues, characterize certain species, groups o r subgroups within this family, including the
divisions among the yellow-pigmented micrococci proposed
by Rosypal, Rosypalovg and Hosejz (1966). These studies
have, however, also revealed heterogeneity within certain
subgroups, notably Micrococcus subgroup 7 of Baird-Parker
(1965). Whereas normal menaquinones were found in strains
extending over a wide range of GC ratios, hydrogenated
menaquinones were restricted to strains with high values;
according to the type of menaquinone produced the aerobic
Micrococeaceae- thus fall into two groups. The first and
broader group comprises strains yielding normal menaquinones. This includes staphylococci, nonpigmented strains
classified a s micrococci, according to Baird-Parker (1965),
planococci (BohgEek, Kocur and Martinec 1967a) and other
strains, e. g. Micrococcus morrhuae, of uncertain taxonomic
p o s i t i o n , a g of which have GC ratios lower than 61.4%. The
second group, producing hydrogenated menaquinones, contains pigmented micrococci and their nonpigmented variants,
allwith GC ratios in the range 66.3-73. 3oJ0,a s suggested for
(1966).
the genus Micrococcus by Rosypal
Two nonpigmented strains studied, although classified in
Baird-Parker' s Micrococcus subgroup 7 on biochemical and
physiological characters have, unlike pigmented micrococci,
low GC ratios and produce normal menaquinones. Thus
ATCC 8456 (GC 36.4%. BohgEek, Kocur and Martinec 1967b)
andCCM 678 (GC 59.670, M. Kocur, personal communication)
a r e anomalous in this respect. It should be borne in mind,
however, that GC ratios have been determined on many pig-
et g .
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SYSTEMATIC
BACTERIO LOGY
187
mented strains of micrococci but apparently on few nonpigmented strains. It i s suggested that this imbalance should
be corrected, so that it may be ascertained whether o r not
the nonpigmented strains in general have GC ratios lower
than 66.3% and produce normal menaquinones, thus separating them f r o m the pigmented micrococci.
As shown in Table 1, agar incorporation t e s t s for determination of sensitivity to novobiocin and to lysozyme confirm the homogeneity of strains classified as Staphylococcus
on biochemical and physiological characters.
The application of these tests to micrococci and also to strains of uncertain taxonomic position in respect of both GC ratio (e. g.
strain CCM 250) and glucose metabolism (strains NET?
7292 and 189, Mortensen and Kocur 1967) shows results of
potential value in classification.
REFERENCES
Baird-Parker, A. C. (1965), J. gen. Microbiol. 38:363. Bohgsek, J., M. Kocur and T. Martinec (1967a)FJ. gen.
Microbiol. 46:369.
and
(l967b), Spisy p r i r . Fak. Univ.
Brne, K40 ( 4 8 3 2 3 7 (1967/5).
Brown, R. W. , 0. Sandvik, R. K. Scherer, and D. L. Rose
( 1967), J. gen. Microbiol. 47 :27 3.
Mortensen, N. and M. Kocur (1967), Acta path. rnicrobiol.
s cand, 69 :445.
Rosypal, S T A. Rosypalova, and J. Hosejg (1966), J. gen.
Microbiol. 44 :28 1.
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