Αναγνώσιμο απόσπασμα της τεκμηρίωσης. Για όλους τους πίνακες, τις εικόνες και τις πλήρεις οδηγίες, ανοίξτε το πρωτότυπο PDF.
Σελίδα 1
Performance evaluation of the MultiStainer AT-3001 for
auramine staining and hot Ziehl-Neelsen staining.
A. Guillouzouic , C. Besson , B. Foucher , S. Raude , M. Bretaudeau , J. Caillon and P. Bémer
Bacteriology-Hygiene Department, Nantes University Hospital, France.
Objective
The search for acid-fast bacilli (AFB) in samples by microscopic examination is an essential step in the diagnosis of mycobacterial infections. Rapid auramine staining is
often performed as a first step. The Ziehl-Neelsen hot stain (ZNC) is still the standard method. The MultiStainer AT-3001 (MT, Biocentric) is the first system that can
perform both stains in an automated way. The objective was to compare the performance of the stainer (MT) to the AutoStainer AT-2000F (AT, Biocentric) for auramine
staining and to compare the performance of the ZNC staining (MT) to the cold Ziehl-Neelsen staining (ZNF).
Material and Methods
844 clinical respiratory specimens (sputum n=498, traps n=211, gastric tubes n=70, BAL n=65) were prospectively collected and decontaminated using the
MycoPrep® technique. Using the decontamination pellet obtained for each sample, 2 smears were made and stained with auramine (Biocentric) by MT stain and AT
stain. Auramine-positive smears were then stained with ZNF (Biomérieux, Tan-Thiam-Hok method) and ZNC (Biocentric, MT). Where there were discrepancies between
ZNC and ZNF staining, 6 additional smears were taken from each decontamination pellet and stained in the same manner.
Using liquid (MGIT, BD) and solid (Coletsos, Biomérieux) cultures, whether or not they were positive for mycobacteria belonging to the tuberculosis complex (MCT or
MNCT), 2 smears were made and stained with ZNC (MT) and ZNF.
The stained smears were performed and read in a blinded fashion by different experienced technicians. The results were interpreted according to WHO
recommendations for the smears, and according to a quality score based on the intensity of the BAARs and the contrast for the cultures.
Results
Auramine MultiStainer AT 3001 (MT)
Results obtained for samples stained with auramine by MT and AT Some Several
stainers None +/- Rare 1+
2++ 3+++
Σελίδα 1
2++ 3+++
Total
None 777 1 778
844 respiratory specimens analyzed (Table 1): AutoStainer +/- 12 12
AT 2000 F Rare 1+ 2 7 9
840 samples (777 negative and 63 positive) were concordant between the (AT) Some
2 stainers, i.e. 99.5% concordance. 2++
17 1 18
4 samples were not concordant, including 3 with ≤ 1-9 BAAR /100 ch. Several
27 27
3+++
Total 777 15 7 17 28 844
Results for ZNC and ZNF stained samples Concordant results Discordant results
Of the 67 auramine positive (or discordant) DEs, 64 could be stained with Table 1: Results obtained for auramine staining with the
ZNC and ZNF stainers (Table 2): AT and MT from the samples.
30 results were concordant between ZNC and ZNF ZNC (MT)
34 results were discordant Rare Some Several
None +/- Total
To resolve these discrepancies, 171 additional smears were analyzed (Table 3): 1+ 2++ 3+++
Σελίδα 1
Total
To resolve these discrepancies, 171 additional smears were analyzed (Table 3): 1+ 2++ 3+++
None 5 7 4 1 17
68 (40%) became concordant, +/- 10 4 2 3 19
103 (60%) remained discordant, of which 93 results were more sensitive with ZNF Rare 1+ 1 5 1 4 10
the ZNC and 10 more sensitive with the ZNF. Some 2++ 2 3 5
Several
12 12
Results obtained for ZNC and ZNF staining on cultures 3+++
Total 6 17 13 6 22 64
Smears could be stained with ZNF/ZNC for 105 cultures (Table 4): Table 2: Results obtained for ZNC (MT) and ZNF staining from
51 MGIT positive (MCT n=25, MAC n=11, other MNCT n=15) Auramine-positive specimens.
54 positive on Coletsos medium (MCT n=29, MAC n=8, other MNCT n=17)
ZNC (MT)
A score of 2 (strong intensity and contrast) was obtained in 86% (91/105) with ZNC None +/- Rare 1+
Some
2++
Several
3+++ Total
and 14% (15/105) with ZNF. None 12 12 1
Σελίδα 1
3+++ Total
and 14% (15/105) with ZNF. None 12 12 11 1 1 37
+/- 2 16 23 4 7 52
Rare 1+ 5 15 5 12 37
1a 2a 3a ZNF Some
3 5 17 25
2++
Several
20 20
3+++
Total 14 33 52 15 57 171
Table 3: Results obtained for the additional ZNC-stained smears
(MT) and for ZNF, with discordance between 2 ZN stains.
Cultures (n=105) ZNC (MT)
Score 0 Score 1 Score 2
Score 0 7 1
ZNF
Score 1 1 5 76
Score 2 1 14
Σελίδα 1
5 76
Score 2 1 14
score 0: no BAAR; score 1: low intensity and contrast; score 2:
1b 2b 3b high intensity and contrast
Figure 1: ZNC staining (sputum 1a, culture of M. avium 2a and M. tuberculosis 3a) Table 4: Results obtained for ZNC and ZNF staining performed on
and ZNF staining (sputum 1b, culture of M. avium 2b and M. tuberculosis 3b). the cultures.
Conclusion
The MultiStainer AT-3001 stainer is the first automated system to perform auramine staining and hot ZN staining. The AT and MT machines have comparable
performance for auramine staining. On the other hand, ZN staining is considerably improved by heat, both in terms of sensitivity from samples, and in terms of intensity
and contrast from cultures, and is becoming routinely accessible thanks to its automation.