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Piglets, were orally infected with an inoculum suspension (containing 1.5E8 CFU/ml ETEC F4ac) at day 0, 1 and 2 of the study.
For each PGPM treatment, 2 mL inoculum suspension was added.
To start the fermentation, the inoculum suspension was added to the bioreactor.
Panicles were spray-inoculated with a freshly prepared inoculum suspension (approximately 1.7 to 2 ml per panicle) by a small sprayer.
To maintain a concentrated NH4 +-depot, the inoculum suspension applied over the NH4 +-depot was more concentrated than the one drenched over the seed-hole.
After incubation, the inoculum suspension was decanted and samples were rinsed three times by adding sterile 1xPBS to the dish, rotating for 10 seconds on an orbital shaker set to 80 rpm, and decanting the rinsate.
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Following treatment, and after a brief interval to allow inoculum suspensions to dry, the segments were stored in a constant temperature room at 17°C under moist or ambient conditions for 6 weeks.
Inoculum suspensions of filamentous fungi and commercial antifungal agents were prepared by the method of National Committee for Clinical Laboratory Standards (NCCLS) M38-A (NCCLS, 2002) [27].
Pairs of cells, as well as rare triplets and quadruplets, were counted as single units, to maximize compatibility between hemocytometer counts and viable counts, set up from the final inoculum suspensions and enumerated the following day.
We tried thorough vortexing with glass beads to obtain homogenous inoculum suspensions.
Inoculum suspensions were prepared by selecting colonies from overnight growth on nutrient agar plates.
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