2.5 Measurement of CO2 efflux rate and microbial
activity as qCO2
The CO2 efflux rate was measured by a closed-chamber
system (Seki et al., 2019) at a frequency of approximately every 2 week
in the rainy season and every month in the dry season for a total of 40
times throughout the experimental period. Polyvinyl chloride (PVC)
columns (diameter 13 cm, height 30 cm) were inserted randomly in each
plot at the end of each September or August, i.e., after FYM
application. We waited at least 1 week after the installation until
measuring the CO2 efflux rate, so as not to disturb the
plots when installing columns. Columns were re-installed within a plot
every year, as mentioned above. Since soil respiration is composed of
microbial respiration and plant-root respiration, plant-root respiration
was excluded by the trenching method (Shinjo et al., 2006), following
the detailed in Seki et al (2019). Gases were sampled at 0 min and again
40 min after the top of the column was covered with a plastic sheet, and
analyzed with an infrared CO2 analyzer (ZFP9-AA11; Fuji
Electric, Japan) equipped with a voltage capture detector (C-R8A;
Shimadzu, Japan) and N2 carrier gas (Shinjo et al.,
2006). The CO2 efflux rate was calculated based on the
increase in CO2 concentration in the column after 40
min. Two columns were installed in each plot, and we used the average
data in each plot with three replicates (plots). The CO2efflux rate was always measured between 08:00 and 11:00 am in the field.
To evaluate the microbial activity as qCO2 (generally
termed as a metabolic quotient) (Anderson & Domsch, 1985), we divided
the measured CO2 efflux rate by the MBC. In the
calculation, both CO2 efflux rate and MBC were expressed
on an area basis (µg CO2-C m−2h−1 and mg MBC m−2, respectively).