30 STUDIES ON DAHI : III. TAXONOMY OF THE LACTIC ACID, ETC. [Vol. XXII, Part I,
to base the classification of the species on the most consistent and easily differentiable
characteristics of the organisms reported by different workers and to reduce the
number of independent species by assigning species rank only to such organisms
which have been described fully and whose characteristics have been well established.
In recent years some workers have suggested the classification of lactic acid bacteria
on the basis of their specific nutritional requirements [Hucker et al., 1944; Dunn
et al., 1947 ; Hoff-Jorgensen et al., 1947]. Although a knowledge of the vitamin
and amino acid requirements may help in differentiating species which are closely
related to each other, there is a possibility of encountering several anamolous results
by this method. For example, L. delbruckii and L. casei, which are quite different
from each other in their physiological as well as biochemical reactions, were considered
to be no more than varieties of the same species because their nutritional requirements
were similar [Dunn et al., 1947]. The former is a non-lactose fermenting organism
occurring in molasses and growing at high temperatures while the latter is a lactose
fermenting lactobacillus commonly found in dairy products and able to grow only
at temperatures below 45°C. The nutritional requirements of an organism can,
therefore, be considered useful only in providing supplementary evidence regarding
its identity. It would thus appear that the taxonomical classification of lactic
acid bacteria will have to be based only on such cultural, and physiological character-
istics as have been considered to be stable and typical of the groups or species under
question, having regard to the source from which the isolates were obtained and
the conditions of their subsequent maintenance.
A summary of the typical, well-established characteristics of different species
of lactic acid bacteria which have been described by various workers [Orla Jensen,
1942; Sherman, 1937; Breed et al., 1948] and which have been considered to be
important in the taxonomical classification of the organisms in this study is given
in Table I.
EXPERIMENTAL
Isolation of lactic acid bacteria
The original samples of dahi as well as stab cultures of the organisms made
from dahi at various centres, secured in the manner explained in Part II [Laxmi-
narayana et al., 1952] of this series, were used for the isolation of lactic acid bacteria.
As soon as the samples were received, a microscopic examination was carried
out to determine the predominant types of organisms present. Small amounts
of well mixed samples were then transferred to sterile yeast dextrose litmus milk
tubes. One set of tubes was incubated at 45°C., one at 37°C., and a third set at
room temperature (24° to 30°C.). When the inoculated samples had coagulated
or shown acid production, they were plated either immediately or after further
transfers in order to eliminate yeasts and encourage the growth of specific types of
lactic acid organisms. Immediate plating gave only the most active types of rods
or cocci present in dahi. Repeated transfers resulted in obtaining pure cultures
of organisms having their optimum near the respective temperatures of incubation.
Rapid transfers of the cultures soon after curdling were helpful in securing a pre-
dominance of streptococci over lactobacilli.