The interpretation and use of Snyder tests
and lactobacillus counts
W. Sim s, MS, PhD, FDS, London
These tw o c a rie s -a ctivity tests are means of e s ti­
m a tin g th e num ber of v ia b le lactobaci 11i in a given
volum e of saliva. The size of the c o u n t is d ire c tly
related to th e num ber of stagnation areas o f the
m outh where th ic k plaque accum ulates and caries
can occur. The te sts are useful fo r m o n ito rin g the
progress o f d ie t therapy in preventive d e n tis try a fte r
oral hygiene has been raised to the h ig h est a tta in ­
able standard.
T w o recent articles concerned with preventive den­
tistry1 and a m odified Snyder test2 have generated
much interest, particularly among dental practi­
tioners. H ow ever, the resulting correspondence
and discussion have revealed considerable con fu ­
sion regarding the nature and value o f lactobacillus
counts and Snyder tests. This paper endeavors to
clarify the situation by considering what inform a­
tion these tests provide and what use can be made
o f it.
Similarity of Snyder tests and
lactobacillus counts
A lthough m any papers and textbooks say that the
Snyder test m easures the acid production o f all
the m icroorganism s present in the inoculated sa­
liva, this is, in fact, untrue. T he acidity o f Snyder
m edium (pH 4 .8 -5 .1 ) is such that all the acid-pro­
ducing organism s o f any consequence, except the
lactobacilli, are inhibited. A part from lactobacilli,
the only oral m icroorganism s that have been found
capable o f grow ing on Snyder m edium (and many
hundreds o f strains have been tested) are p ed iococ-
ci and som e strains o f yeast. Pediococci rarely o c­
cur in the m outh3 and m ost o f the yeasts, although
able to grow, fail to produce enough acid to cause
the indicator to change color.
T he Snyder test m easures the tim e required for
sufficient acid to be produced to change the color
o f the brom cresol green indicator from green to
yellow . Since only lactobacilli grow in the m edium ,
the test m easures the tim e required for the lacto­
bacilli in the inoculated saliva to produce this d e­
gree o f acidity. Further, because the rate at which
bacteria produce acid in a given volu m e o f m edia
is directly proportional to the number o f bacteria
inoculated, it follow s that the lactobacillus count
and Snyder test are essentially identical tests. They
are sim ply different m ethods o f estim ating the
sam e thing, that is, the num ber o f viable lactoba­
cilli in a given volum e o f saliva. It is hardly sur­
prising, therefore, that in all studies w here agree­
m ent between the tw o tests has been measured,
excellen t correlations have been found.4-5
Significance of the number of
lactobacilli in saliva
H aving estim ated the num ber o f lactobacilli in the
saliva o f a patient, w e m ust next consider the m ean­
ing o f the variable num bers o f these organism s.
In practically all situations in nature w here lac­
tobacilli are found, streptococci are found also.
1315
M oreover, the lactobacilli only com e to prom i­
nen ce in these situations when acid conditions,
created by the streptococci, selectively favor them.
T h is is exactly w hat our know ledge o f these organ­
ism s and studies in the fields o f bacterial genetics
and population growth would lead us to expect.
T he streptococci, provided the acidity o f the en ­
vironm ent is not greater than a pH o f about 5.5,
h ave a shorter generation tim e than the lactoba­
cilli and w ill proliferate at the latter’s expense.
H ow ever, the streptococci, like the lactobacilli,
can only obtain energy by the inefficient m echan­
ism o f breaking down sugars to lactic acid, so an
inevitable con sequ en ce o f their proliferation is an
increase in acidity. A s the pH o f the environm ent
steadily falls, the generation tim e o f the strepto­
cocci proportionately lengthens until it equals,
and then exceeds, that o f the lactobacilli. Since
the lactobacilli now have the shorter generation
tim e, they begin to m ultiply at the expense o f the
streptococci and go on replicating quite rapidly
w hen con d itions have becom e so acid that the
growth o f the streptococci is com pletely inhibited.
T he foregoing description applies to closed sys­
tem s w here there are sufficient nutrients and sugar
to support streptococcal proliferation and where
the acid end product accum ulates. A human ex ­
am ple is afforded by the vagina which harbors
streptococci as part o f the normal flora from soon
after birth. W hen, at puberty, ferm entable carbo­
hydrate becom es available in the form o f glycogen
deposits in the vaginal epithelium , acid conditions
b ecom e established and, thereafter, lactobacilli
can be readily isolated.6-8
T he m outh, in contrast, is effectually an open
system because saliva is continually being secreted
and sw allow ed. Thus, although the normal oral
flora contains large numbers o f streptococci and
cariogenic diets supply an abundance o f sugar, the
organism s, the sugar, and the acid end products
are being repeatedly swallowed, and the saliva,
(or the oral cavity itself) never becom es sufficiently
acid to favor the selective proliferation o f lacto­
bacilli. T he necessary acid conditions can only be
created at sites in the mouth w here the streptococci
are in such high concentration that acid is p ro­
duced rapidly. T hen, despite neutralization, d ilu ­
tion, and rem oval o f acid by saliva, hydrogen ions
still accum ulate and the pH inexorably falls. Such
sites are, o f course, the stagnation regions o f the
mouth w here dental plaque accum ulates. T he acid­
ity o f thick dental plaque, as was shown by S te­
phan9 and confirm ed by m any subsequent workers,
can fall to pH 5.0 within a few m inutes after a
1316 ■ JADA, Vol. 80, June 1970
ferm entable carbohydrate is ingested. Steinle,
M adonia, and B ah n 10 have shown that oral lacto­
bacilli are strictly localized to sites on the teeth
w here caries occurs, or has occurred, and the size
o f the salivary lactobacillus count is directly re­
lated to the number o f such sites w hich were har­
boring lactobacilli at the tim e the count was made.
B ecau se saliva is supersaturated with calcium
and phosphate ions with respect to hydroxyapatite,
a pH o f 5.5 or low er is required before enamel
w ill d issolve in it. It is at acidities o f about pH 5.5
that the streptococci begin to be inhibited and the
proliferation o f lactobacilli facilitated. Since
acidities o f this order can arise only in stagnation
areas and it is only in such areas that caries occurs,
the link between dental caries and increasing num ­
bers o f oral lactobacilli is, surely, self-evident.
A ll tooth surfaces that are not directly abraded
during m astication are covered with a film o f bac­
teria. E ven enam el p ellicle floated o ff teeth which
have been vigorously scrubbed contains large
num bers o f organism s em bedded in it.11 O nly in
regions not subjected to m echanical cleansing can
this film thicken into what w e w ould call dental
plaque. T o produce oral conditions sufficiently
acid to favor the lactobacilli, high concentrations
o f streptococci are required. Electron m icrographs
show that organism s in the dental plaque are
tightly packed together, and so the only w ay to in­
crease the concentration o f bacteria covering a
given area o f tooth surface is for the plaque to b e­
com e thicker. F or plaque to produce the required
degree o f acidity fairly rapidly it has to thicken
into m acroscopically v isib le am ounts that are
readily susceptible to m echanical reduction or re­
m oval.9-12 H ence, the value o f oral hygiene in
the prevention o f dental caries is evident.
T he salivary lactobacillus count then, indicates
the num ber o f stagnation areas in the mouth where
thick plaque has form ed and acid conditions of
sufficient degree to favor the lactobacilli are peri­
odically, or continuously, occurring.
Relationship between the salivary
lactobacillus count and dental caries
W hen a representative sam ple o f saliva is obtained
and a bacterial count is perform ed, the practition­
er should bear in m ind the scope for error in as­
sessing the incidence o f dental caries. It is, and
always was, unrealistic to expect the agreement
betw een lactobacillus counts and dental caries to
be other than approxim ate. T he notion that the
num bers o f lactobacilli and dental caries should
be associated originated with the view that lacto­
bacilli w ere the specific cau se o f this disease. F ew
h old this view today. N evertheless, m ost o f the
m isunderstanding regarding these tests results
from the literal expectation, unfortunately encour­
aged by som e papers, that a certain lactobacillus
count w ill always indicate a certain num ber o f
new carious lesion s in a given period.
W e have seen that acid conditions which pro­
duce caries favor lactobacilli. That there is, in
general, a correlation betw een the lactobacillus
count and caries activity is beyond dispute.4-5 The
m ain reason w hy the correlation betw een the la c­
tobacillus count and the subsequent in cid en ce o f
dental caries m ay vary between good and non­
existent, is that whereas all sites o f acidity in the
m outh favor the m ultiplication o f lactobacilli,
caries m ay either have occurred already or sim ply
not be p ossible at one or m ore, or even all o f these
sites. C arious dentin is alm ost alw ays acid 13 and
lactobacilli abound in it. I f all carious lesion s are re­
stored, the lactobacillus count falls n oticeab ly.14-15
E xam p les o f sites w here thick acid-producing
plaques m ay be present but no caries can occur,
are plaques on restorations o f all types, dentures,
and orthodontic appliances. T he extrem e exam ple
o f this is the com plete denture. Shklair and M az-
zarella16 have shown that the lactobacillus count
falls to nearly zero w hen all teeth are extracted.
It rem ains at this low level until dentures are in ­
serted, and it then tends to return to the preextrac­
tion level. That com plete-denture w earers may
have high lactobacillus counts has been confirm ed
in the author’s laboratory. B y u se o f the m ethods
o f Steinle, M adonia, and B ah n 10 I have found
that lactobacilli on dentures, as in the natural den­
tition, are located in stagnation areas. If com plete
dentures are rem oved and thoroughly scrubbed,
the lactobacillus count falls to zero. A lthough lac­
tobacilli certainly proliferate in carious lesions,
the finding o f high lactobacillus counts in co m ­
plete-denture wearers entirely refutes the view that
the lactobacillus count indicates the existence o f
dental caries.
A t the other end o f the spectrum from the co m ­
plete-denture wearer is the individual with an in ­
tact natural dentition. In these individuals, a cari­
ous lesion w ill result in every situation where co n ­
ditions sufficiently acid to favor the lactobacilli
prevail, and the agreem ent between the lacto­
bacillus count and caries incidence w ill b e good.
In betw een the extrem es o f the intact dentition
Sims: SNYDER TESTS AND LACTOBACILLUS COUNTS ■ 1317
and com plete dentures co m e an alm ost infinite
variety o f com binations o f thick plaques on tooth
surfaces w here caries can result, and plaques on
restorations or in carious lesions, where, although
the plaque contributes to the lactobacillus count,
no new caries can occur. F or each different set o f
oral conditions the sam e lactobacillus count w ill
lead to com p letely different numbers o f new
carious lesions. T h e Snyder test and lactobacillus
count m easure the potential for producing dental
caries prevailing in a given m outh. E xactly how
m any new lesions w ill, in fact, develop depends
entirely on the scope for realizing this potential
existing in that m outh.
T he m ost notable feature o f the original d e­
scriptions o f the lactobacillus cou n t17-18 and S ny­
der test19 w as the excellen t agreem ent betw een a
zero count, or negative Snyder reading, and the
absence o f caries activity. Practically all su b se­
quent work, although disagreeing sharply with
other aspects o f the tests, has thoroughly co n ­
firm ed this feature o f the original studies. A zero
or very low lactobacillus count m eans there are
no sites in the mouth sufficiently acid to favor se ­
lective lactobacillus m ultiplication. W hile this
condition persists, dental caries cannot occur. A
negative Snyder test result, or a lactobacillus count
o f less than a few hundred per m illiliter o f saliva
m eans, incontrovertibly, no caries activity. O f
course, there are cases recorded in the literature
w here a zero lactobacillus count is alleged to be
associated with high caries activity. I can only
com m ent that in all such cases investigated in this
laboratory, these assertions have proved false.
The use of the Snyder test and
lactobacillus count
I f the result o f a caries-activity test is to be sensibly
interpreted and usefully applied, the oral co n d i­
tion at the tim e when the saliva specim en is c o l­
lected is o f crucial im portance. F ew dentists seem
to be aware o f this and, consequently, m ost
caries-activity tests are sim ply a fruitless ex p en ­
diture o f laboratory tim e and m oney. T he p rinci­
pal use o f caries-activity tests is to m onitor the
progress o f diet therapy in preventive dentistry.
F or the tests to be o f use in this regard, the essen ­
tial prerequisite before com m encing therapy is
reduction o f oral stagnation to the absolute m in i­
mum. If this condition is not m et, the lactobacillus
count w ill sim ply indicate the degree o f total oral
stagnation, instead o f that sm all part o f it which
m ust b e controlled by diet therapy because it can­
not be elim inated in any other way.
A ll procedures which rem ove plaque or elim ­
inate stagnation areas w ill produce a correspond­
ing fall in the lactobacillus count. Thus, restoring
all carious lesio n s,14-15 extracting all teeth ,16 im ­
proving the oral hygiene,20 and even having the
patient ch ew w ax pellets to provide successive
saliva sp ecim ens,21 all decidedly reduce the lacto­
bacillus cou n t or Snyder test reading. Conversely,
increasing the num ber o f sites w here plaques m ay
form by fitting orthodontic bands22 or inserting
orthodontic appliances23 or dentures16 greatly in ­
creases the lactobacillus count. B efore em barking
on diet therapy and caries-activity tests it is essen­
tial to restore all carious lesions, replace all leaky
restorations, and bring the oral hygiene, including
the cleansing o f any oral appliances, to the high­
est attainable standard. A sid e from caries-activity
tests, th ese m easures are norm al, sound dental
treatm ent and are necessary for preservation o f the
supporting structures o f the teeth.
■ C a ries-a ctivity tests in d ie t therapy: T he lacto­
bacillus count, in patients who have a naturally
intact or com p letely restored dentition that is
kept thoroughly clean, is in dicative o f the num ­
ber o f acid sites which can be eradicated only by
d iet therapy. In m ost such individuals the count
w ill be low and the am ount o f carbohydrate re­
striction required to reduce it to zero w ill be small
enough to b e acceptable to any patient w ho gen ­
uinely desires to preserve h is dental health. It has
been said elsew h ere,1 but w ill certainly bear rep­
etition here, that to attem pt to reduce the lacto­
bacillus cou n t by dietary m ethods in the presence
o f carious lesions, leaky fillings, and poor oral
hygiene is futile. T he required diets are so austere
that they prove unacceptable to patients. Further­
m ore, m any dentists who b elieve, quite erroneous­
ly, that on ly such grossly restrictive diets are o f
value in caries prevention, have abandoned diet
therapy altogether. Persuading patients to alter
their dietary habits can p rove a form idable task.
D ie t therapy should b e used to control caries only
in those sites w here good dentistry and the tooth­
brush w ill not suffice.
If, w hen the foregoing conditions have been
m et, the Snyder test result is p ositive or the lacto­
bacillus cou n t is m ore than a few hundred, co n d i­
tions for producing caries are still prevailing in the
m outh. T he size o f the lactobacillus count, or the
rapidity o f the color change in the Snyder test,
1318 ■ JADA, Vol. 80, June 1970
are indicative o f the m agnitude o f the potential
for producing caries. I f the dental con d ition is
such that there is scope for this potential to be
realized in the form o f new carious lesions, diet
therapy is used to reduce the count or render the
Snyder test negative.
T h e negative Snyder test result or zero la cto ­
bacillus count is all-im portant in caries p reven ­
tion. A p ositive result m eans that the necessary
conditions for producing dental caries exist in the
patient’s m outh, a fact which is usually obvious,
or soon becom es so, and w hich can b e taken for
granted in m ost patients. F or the reasons already
discussed, future caries incidence cannot be p re­
dicted by the degree o f positivity o f these tests.
H ow ever, a negative result m eans, unequivocally,
that if the present oral condition is m aintained no
further carious lesions w ill occur. In fact, the real
valu e o f the Snyder test and lactobacillus count is
to determ ine caries inactivity. Since there is no
w ay o f establishing that a mouth is caries inactive
except by m eans o f a caries-activity test, these
tests are indispensable in preventive dentistry as
indicators that the goal o f preventive therapy has
been achieved. T he m odified Snyder test2 is an
entirely feasible test to carry out in the dental o f ­
fice; it cheaply and reliably provides the inform a­
tion required to m onitor d iet therapy and estab­
lish when the aim o f such therapy has been
achieved. T he test result can be shown to, and
discussed with, the patient. C onsequently, as he
realizes that his response to dietary ad vice is being
m easured, the test becom es a potent factor in se ­
curing his collaboration.
■ C a ries-a ctivity tests in clin ica l stu d ies: The
other principal use o f caries-activity tests is in
clinical trials and field studies w here agents or
factors b elieved to affect the caries rate are being
investigated. In all these studies the aim is to have,
in the control and study groups, sim ilar numbers
o f the sam e types o f teeth exposed to the sam e
degree o f caries attack. O nly when this condition
is m et can any observed differences in caries rate
between the tw o groups be justifiably attributed
to the treatm ent or agent under trial.
W hen the study involves a large num ber o f sub­
jects o f sim ilar age distribution w ho are from the
sam e geographical location and have been ran­
dom ly assigned to control and study groups, it can
safely be assum ed that the caries activity o f both
groups will be sim ilar and that caries-activity tests
are superfluous.
With small groups, or if there is any reason to
suspect that the caries activity o f the tw o groups
m ay be dissim ilar, it is essential that the caries ac­
tivity be m easured. If caries-activity tests are not
used and the groups do differ in caries activity,
then the valu e o f an agent under trial m ay be e x ­
aggerated or a w orthless agent m ay appear to have
som e effect. C onversely, the effectiveness o f an
agent m ay be underestim ated or obscured alto­
gether. O nly a low lactobacillus count or negative
Snyder test reading has an unequivocal interpreta­
tion for an individual patient. On a group basis
there is a good correlation between the size o f the
m ean lactobacillus count, or degree o f positivity
o f the Snyder test, and the subsequent caries in ­
cidence. T h e reason is that individual variations in
dental condition are com pensated and balanced
out by other m em bers o f the group. I f tw o groups
are reasonably com parable in other respects, but
have a statistically significant difference in mean
lactobacillus count, they w ill exhibit correspond­
ingly different caries rates.
Doctor Sim s is reader in m icrobiology in relation to den­
tis try at th e School o f Dental Surgery, Royal Dental Hospi­
tal, U niversity o f London, London, WC2, England.
1. Sims, W. Preventive de n tistry fo r the dental p ra c titio n ­
er. Dent Pract (B ristol) 18:309 May 1968.
2. Sims, W. A m odified Snyder test fo r caries-activity in
humans. Arch Oral Biol 13:853 Aug 1968.
3. Sims, W. The isolation o f pediococci from human sa­
liva. Arch Oral Biol 11:967 Oct 1966.
4. Snyder, M.L., and others. Evaluation o f laboratory tests
fo r estim ation of caries activity. JADA 65:30 Ju ly 1962.
5. Snyder, M.L., and others. Evaluation o f laboratory tests
fo r the estim ation of caries activity. C orrelation w ith spe­
c ific surfaces. Arch Oral Biol 8:541 July 1963.
6. C ruickshank, R., and Sharman, A. Biology o f the vagina
in the human subject; glycogen in vaginal epithelium and
its relation" to ovarian activity. J Obstet Gynaec B rit Comm
41:190 A p ril 1934.
Sim s: SNYDER TESTS AND LACTOBACILLUS COUNTS ■ 1319
7. Cruickshank, R., and Sharman, A. The biology o f the
vagina in the human subject; bacterial flo ra and secretion
of vagina a t various age-periods and th e ir relation to g lyco ­
gen in the vaginal epithelium . J Obstet Gynaec B rit Comm
41:208 A pril 1934.
8. Rogosa, M., and Sharpe, M.E. Species d iffe re n tia tio n
of human vaginal lactobacilli. J Gen M icrobiol 2 3 :1 9 7 Aug
1960.
9. Stephen, R.M. Intra-oral hydrogen-ion concentrations
associated with dental caries a ctivity. J Dent Res 23:257
Aug 1944.
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examinations. Caries Res 2:294, No. 4 1968.
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