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The ARG density determined by qPCR assay is highly related to microscopic cell count (r2 = 0.726, p < 0.001), and the limit of detection (LOD) and limit of quantification (LOQ) of the qPCR method were 0.02 pg and 0.2 pg of DNA, respectively.
However, the deletions that are detected appear to be correct, since the estimates of normal cell content are in agreement with those based on microscopic cell count.
Among three major studies, the levels of total bacteria ranged from 6 to 9 log/L (by microscopic cell count); the authors also stated that less than 1% of the collected bacteria could be cultivated on nonselective media [ 51].
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All samples were separated in two subsamples; one was used for direct microscopic cell counts by Axioscope A1 microscope (Carl Zeiss, Oberkochen, Germany).
These data were compared with traditional metrics, such as microscopic cell counts and analytical toxin measurements, and the public health implications are discussed.
Direct microscopic cell counting and spectrophotometric readings indicated that cell numbers in flight and ground biological replicate cultures differed by less than 2-fold (data not shown).
An underestimation of the amount of chromosomal DNA by microscopic cell counting can easily occur due to factors such as cell clumping, cell division or cell lysis.
The reference was estimated by microscopic cell counting and assuming a chromosomal weight calculated from the chromosome length and GC content of approximately 3.0 fg for S. aureus and 5.7 fg for E. coli, respectively.
Also, quality of the tissue digest can be checked by simple microscopic cell counting with dye viability exclusion, an assessment which was sufficient to discriminate between our two different biopsy types.
As shown by the results, the measured amount of DNA in some cases exceeded that of the reference, which was estimated by the 'gold standard' of microscopic cell counting, resulting in percentages above 100%.
Briefly, cells from overnight cultures were collected by centrifugation and were used to inoculate plain SVO, after 4 h of growth at 77°C, the culture was then subjected to microscopic cell counting (to determine the total number of cells) and was plated with plain SVO (to find out the number of viable cells).
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