Total numbers of culturable protozoa were estimated by a
most-probable number (MPN) method using agar plates
(Stotzky et al., 1993). The medium (5 g NaCl, 10 g agar,
and 1 l tap water; autoclaved for 15 min at 1218C) was
aseptically dispensed into sterile Petri plates and allowed
to solidify and dry overnight. The plates were then cored
with the open end of a ¯amed test tube (25-mm OD), so that
islands of agar, 25 mm dia., were physically separated from
the remaining agar. A loopful of a suspension of mixed
bacteria isolated from the soil and grown on soil extract
agar (SEA; 0.2 g KH2PO4, 1 g dextrose, 15 g agar, 100 ml
soil extract, and 900 ml tap water) was spread over each
island to serve as a food source for the protozoa. Soil was
serially diluted, and 100 ml each of three dilutions was added to the center of ®ve replicate islands,
and the plates were incubated in the dark at 24 ^ 28C. After
5±7 days, each island was examined under low-power
magni®cation (100 £ ) for the presence of protozoa. Total
numbers of protozoa were calculated from MPN tables.