Utilisation of Actiphage in combination with IS900 qPCR as a diagnostic tool for rapid determination of paratuberculosis infection status in small ruminant herds

Examination of fresh environmental samples for the presence of Mycobacterium avium subsp. paratuberculosis (MAP) using an Actiphage–IS900 qPCR assay, direct IS900 qPCR, and a conventional phage amplification assay (PA)

Sample no.	Sample characteristics	Direct IS900 qPCR (copy/g)	Conventional PA (PFU/g)*	Actiphage–IS900 qPCR (copy/g)
1	Soil	5.42 × 10²	4.80 × 10²	–
2	Soil	1.09 × 10²	2.18 × 10²	–
3	Soil	1.02 × 10²	2.88 × 10²	–
4	Soil	4.20 × 10²	–	–
5	Cobweb	1.20 × 10²	–	–
6	Soil/plants	1.64 × 10⁰	7.54 × 10³	1.84 × 10⁰
7	Soil	5.88 × 10⁰	3.06 × 10²	–
8	Soil	6.56 × 10⁰	1.09 × 10²	–
9	Soil	2.40 × 10⁰	4.36 × 10¹	–

1 PFU – plaque-forming units; – denotes a negative result;

1* – plaques occurred during the conventional PA, but MAP was not confirmed by plaque PCR in any of the samples (five plaques per sample were tested)

Table 2.

Examination of frozen environmental samples for the presence of Mycobacterium avium subsp. paratuberculosis (MAP) using an Actiphage–IS900 qPCR assay, direct IS900 qPCR, a conventional phage amplification assay (PA) and culture

Sample no.	Sample characteristics	Time frozen	Direct IS900 qPCR (copy/g)*	Conventional PA (PFU/g)†	Culture*	Actiphage–IS900 qPCR
1	Soil	11 years	6.43 × 10³	6.98 × 10²	–	–
2	Soil	11 years	1.18 × 10³	8.72 × 10²	–	–
3	Soil	11 years	1.16 × 10²	2.88 × 10³	–	–
4	Soil	11 years	2.11 × 10²	1.92 × 10³	–	–
5	Soil	11 years	1.30 × 10²	2.01 × 10²	–	–
6	Soil	11 years	7.94 × 10²	2.62 × 10²	–	–
7	Soil	11 years	3.48 × 10²	7.85 × 10²	–	–
8	Roots	11 years	1.07 × 10²	3.49 × 10²	–	–
9	Soil	11 years	7.39 × 10²	2.62 × 10²	–	–
10	Roots	11 years	1.68 × 10²	3.49 × 10²	–	–
11	Soil	11 years	1.61 × 10²	2.62 × 10²	–	–
12	Roots	11 years	4.36 × 10²	6.98 × 10²	–	–
13	Roots	11 years	6.41 × 10²	2.27 × 10²	–	–
14	Soil	11 years	9.81 × 10²	6.10 × 10²	–	–
15	Soil	11 years	4.09 × 10²	1.74 × 10²	–	–
16	Roots	11 years	1.57 × 10²	5.58 × 10²	–	–
17	Soil	8 years	4.40 × 10²	4.36 × 10²	–	–
18	Detritus	8 years	1.25 × 10²	4.36 × 10¹	–	–
19	Feed residues	8 years	1.94 × 10²	–	–	–
20	Soil	11 years	2.67 × 10¹	8.72 × 10¹	–	–
21	Soil	11 years	7.26 × 10¹	1.13 × 10³	–	–
22	Soil	11 years	8.93 × 10¹	5.67 × 10²	–	–
23	Soil	11 years	6.11 × 10¹	3.49 × 10³	–	–
24	Soil	11 years	7.95 × 10¹	1.74 × 10²	–	–
25	Roots	11 years	6.44 × 10¹	5.23 × 10¹	–	–
26	Soil	11 years	7.90 × 10¹	8.72 × 10⁰	–	–
27	Soil	11 years	2.70 × 10¹	1.40 × 10²	–	–
28	Roots	11 years	9.91 × 10¹	1.74 × 10²	–	–

1 PFU – plaque-forming units; – denotes a negative result;

1* – the analysis was performed immediately after sample collection;

1† – plaques occurred during the conventional PA, but MAP was not confirmed by plaque PCR in any of the samples (five plaques per sample were tested)

References

Authors

Metrics

Articles in this issue

DOI: https://doi.org/10.2478/jvetres-2023-0041 | Journal eISSN: 2450-8608 (formerly 2300-3235)

Journal RSS Feed

Language: English

Page range: 347 - 352

Submitted on: Feb 20, 2023

Accepted on: Jul 4, 2023

Published on: Sep 20, 2023

Published by: National Veterinary Research Institute in Pulawy

In partnership with: Paradigm Publishing Services

Keywords:

Actiphage-qPCR,

environmental samples,

Mycobacterium avium subsp. paratuberculosis,

phage amplification assay,

small ruminants.

Related subjects:

Life sciences,

Molecular biology,

Microbiology and virology,

Life sciences, other,

Medicine,

Veterinary medicine

© 2023 Monika Beinhauerova, Iva Slana, published by National Veterinary Research Institute in Pulawy
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Volume 67 (2023): Issue 3 (September 2023)