Table 1.
Comparison of FECPAKG2 standard operating procedures for analyzing animal and human stool.
Fig 1.
Materials needed to perform the FECPAKG2 method.
(A) The FECPAKG2 sedimenter with indications of the water line and the saline lines used to analyse faeces of cattle or sheep and horses. For analysis of human samples, the saline line for sheep and horses is applied. (B) The filtration unit with cylinder base, cylinder top and two filter frames. (C) The unit fits together and can be disassembled for cleaning. The two filter frames have different mesh sizes (425μm and 250μm) and are designed to be used together. Samples are collected from the filtration unit using a pipette with special extension tip designed to fit into the filtration unit and the top of the cassette (D). The FECPAKG2 cassette with its two counting wells with central light rods is shown in side view (E) and from the top containing a sample (F). A filled cassette is fed into a portal of the MICRO-I device (G) and then pulled through automatically for imaging of each well. (H) A standard image that is produced by the FECPAKG2 and presented to the user. Some debris and numerous STH eggs are visible. Visible Ascaris lumbricoides eggs (n = 118) are marked with green dots, the Trichuris trichiura eggs are marked with yellow dots (n = 29) and the hookworm eggs are marked with a red dot (n = 5). After mark-up, this information is stored and remains available for review by authorized users.
Fig 2.
The FECPAKG2 standard operating protocol.
Three grams of fresh human stool is weighed in a Fill-FLOTAC device (step 1), water is added and the stool is homogenized using the Fill-FLOTAC device (Step 2 & 3). The fecal suspension is then transferred to a FECPAKG2 sedimenter (step 4) and water is added to the water line (step 5 & 6). The supernatant is removed by tilting the sedimenter towards the A-side (step 7) and floatation solution is added to the retained slurry until the saline line is reached (step 8). This mixture is transferred into a FECPAKG2 filtration unit by pouring from the B-side of the sedimenter (step 9). The content is mixed by inverting the filtration unit 3 times and a sample is extracted from the inside of the inner sieve using a pipette with special tip. Samples are pipetted into both wells of a FECPAKG2 cassette (step 10 & 11). The filled cassette is then placed on a level surface to allow accumulation of the helminth eggs (step 12). Finally, the cassette is placed in the MICRO-I device and both wells are imaged (step 13). Images, together with accompanying sample information is stored and uploaded to the cloud.
Fig 3.
Schematic representation of the experiment designed to evaluate the optimal sedimentation time.
Fig 4.
Schematic representation of the experiment designed to evaluate the optimal accumulation time.
Fig 5.
Line plots representing the percentage of STH eggs recovered in the slurry of the sedimenter at different sedimentation times (10, 30, 60 minutes and overnight (ON)).
The average recovery percentage over the different samples is indicated in red. Significant increases in egg recovery percentages between subsequent sedimentation times are indicated (* P < 0.05; ** P < 0.01, *** P < 0.001).
Fig 6.
Line plots showing the average relative amount of STH eggs counted in FECPAKG2 cassettes over increasing accumulation times.
The average relative percentage of visible eggs in the different samples over the different time points is indicated in red. Panel A shows the results obtained from the accumulation experiment performed with 10 samples from Ethiopia where samples were imaged for 20 minutes. Panel B shows the combined results obtained from the accumulation experiment performed with 30 additional samples (10 from Brazil, 10 from Laos and 10 from Tanzania). These samples were imaged for a total of 30 minutes. A grey horizontal dotted line is drawn at the height of 80% of maximum number eggs counted in the cassette wells.