US 2022/0186293 A1 · USE OF PROBES TO DETECT TOXINOGENIC CYANOBACTERIA, DETECTION METHOD AND CORRESPONDING KITS

Abstract

Probes for the detection of toxinogenic cyanobacteria, and the use of at least one pair of these probes in a method for the detection of the toxinogenic cyanobacteria in a sample likely to contain the toxinogenic cyanobacteria. Also, corresponding kits including at least one pair probes specific to toxinogenic cyanobacteria.

Claims21

Dependent claims

1-21. (canceled)

A method for detecting toxinogenic cyanobacteria in a sample likely to contain at least one toxinogenic cyanobacteria of the genus Microcystis using at least one pair of probes specific to toxinogenic cyanobacteria of the genus Microcystis, the sequences of the said probes being chosen from x elements of one of the following sets:

(SEQ ID NO:
1, SEQ ID NO:
2, SEQ ID NO:
3 and SEQ ID NO:
4)
(SEQ ID NO:
5, SEQ ID NO:
6 and SEQ ID NO:
7)
(SEQ ID NO:
8, SEQ ID NO:
9, SEQ ID NO:
10 and SEQ ID NO:
11)
(SEQ ID NO:
12, SEQ ID NO:
13 and SEQ ID NO:
14)
(SEQ ID NO:
15, SEQ ID NO:
16 and SEQ ID NO:
17)
(SEQ ID NO:
18 and SEQ ID NO:
19)
x being 2, 3 or 4
or the sequences of said probes having at least 92% identity with said sequences SEQ ID NO:
1, SEQ ID NO:
2, SEQ ID NO:
3, SEQ ID NO:
4, SEQ ID NO:
5, SEQ ID NO:
6, SEQ ID NO:
7, SEQ ID NO:
8, SEQ ID NO:
9, SEQ ID NO:
10, SEQ ID NO:
11, SEQ ID NO:
12, SEQ ID NO:
13, SEQ ID NO:
14, SEQ ID NO:
15, SEQ ID NO:
16, SEQ ID NO:
17, SEQ ID NO:
18, SEQ ID NO:
19,
one probe of said pair being a capture probe linked to at least one attachment molecule positioned at 3′ or 5′ of its sequence and the other probe of said pair being a signal probe linked to at least one marking molecule positioned at 3′ or 5′ of its sequence,
said capture probe and said signal probe being capable of hybridizing with the ribosomal nucleic acid of the toxinogenic cyanobacteria of the genus Microcystis optionally present in said sample to form a complex,
the minimum detection threshold of the toxinogenic cyanobacteria of the genus Microcystis being
estimated from 10 to 575 active living cells per millilitre of sample (cells/mL), or
less than or equal to 1.0 ng of ribosomal RNA per millilitre of sample, and in particular from 0.02 ng to 0.7 ng of ribosomal RNA per millilitre of sample and preferably from 0.02 ng to 0.1 ng of ribosomal RNA per millilitre of sample,
in particular, the sequences of the probes of the said pairs being as follows:
(SEQ ID NO:
1 and SEQ ID NO:
2), (SEQ ID NO:
1 and SEQ ID NO:
3), (SEQ ID NO:
1 and SEQ ID NO:
4), (SEQ ID NO:
2 and SEQ ID NO:
3), (SEQ ID NO:
3 and SEQ ID NO:
4)
(SEQ ID NO:
5 and SEQ ID NO:
6), (SEQ ID NO:
5 and SEQ ID NO:
7), (SEQ ID NO:
6 and SEQ ID NO:
7)
(SEQ ID NO:
8 and SEQ ID NO:
9), (SEQ ID NO:
8 and SEQ ID NO:
10), (SEQ ID NO:
8 and SEQ ID NO:
11), (SEQ ID NO:
9 and SEQ ID NO:
10), (SEQ ID NO:
9 and SEQ ID NO:
11), (SEQ ID NO:
10 and SEQ ID NO:
11)
(SEQ ID NO:
12 and SEQ ID NO:
13), (SEQ ID NO:
12 and SEQ ID NO:
14), (SEQ ID NO:
13 and SEQ ID NO:
14)
(SEQ ID NO:
15 and SEQ ID NO:
16), (SEQ ID NO:
15 and SEQ ID NO:
17), (SEQ ID NO:
16 and SEQ ID NO:
17)
(SEQ ID NO:
18 and SEQ ID NO:
19).

97 more elements

A pair of probes for the detection of toxinogenic cyanobacteria whose sequences are chosen from x elements of one of the following sets:

(SEQ ID NO:
1, SEQ ID NO:
2, SEQ ID NO:
3 and SEQ ID NO:
4)
(SEQ ID NO:
5, SEQ ID NO:
6 and SEQ ID NO:
7)
(SEQ ID NO:
8, SEQ ID NO:
9, SEQ ID NO:
10 and SEQ ID NO:
11)
(SEQ ID NO:
12, SEQ ID NO:
13 and SEQ ID NO:
14)
(SEQ ID NO:
15, SEQ ID NO:
16 and SEQ ID NO:
17)
(SEQ ID NO:
18 and SEQ ID NO:
19)
x being 2, 3 or 4
or the sequences of said probes having at least 92% identity with the abovementioned sequences SEQ ID NO:
1, SEQ ID NO:
2, SEQ ID NO:
3, SEQ ID NO:
4, SEQ ID NO:
5, SEQ ID NO:
6, SEQ ID NO:
7, SEQ ID NO:
8, SEQ ID NO:
9, SEQ ID NO:
10, SEQ ID NO:
11, SEQ ID NO:
12, SEQ ID NO:
13, SEQ ID NO:
14, SEQ ID NO:
15, SEQ ID NO:
16, SEQ ID NO:
17, SEQ ID NO:
18, SEQ ID NO:
19,
for the detection of toxinogenic cyanobacteria of the genus Microcystis, or pair of probes whose sequences are chosen from x elements of one of the following sets:
(SEQ ID NO:
20, SEQ ID NO:
21 and SEQ ID NO:
22)
(SEQ ID NO:
23 and SEQ ID NO:
24)
x being 2 or 3,
or the sequences of said probes having at least 92% identity with the abovementioned sequences SEQ ID NO:
20, SEQ ID NO:
21, SEQ ID NO:
22, SEQ ID NO:
23, SEQ ID NO:
24,
for the detection of toxinogenic cyanobacteria of the genus Aphanizomenon, or pair of probes whose sequences are chosen from x elements of one of the following sets:
(SEQ ID NO:
25 and SEQ ID NO:
26)
x being 2,
or the sequences of said probes having at least 92% identity with the aforementioned sequences SEQ ID NO:
25, SEQ ID NO:
26,
for the detection of toxinogenic cyanobacteria of the genus Dolichospermum, or pair of probes whose sequences are chosen from x elements of one of the following sets:
(SEQ ID NO:
27, SEQ ID NO:
28 and SEQ ID NO:
29)
(SEQ ID NO:
30, SEQ ID NO:
31 and SEQ ID NO:
32)
(SEQ ID NO:
33, SEQ ID NO:
34 and SEQ ID NO:
35)
(SEQ ID NO:
36, SEQ ID NO:
37, SEQ ID NO:
38 and SEQ ID NO:
39)
(SEQ ID NO:
40, SEQ ID NO:
41 and SEQ ID NO:
42)
(SEQ ID NO:
43, SEQ ID NO:
44 and SEQ ID NO:
45)
(SEQ ID NO:
46, SEQ ID NO:
47 and SEQ ID NO:
48)
(SEQ ID NO:
49, SEQ ID NO:
50 and SEQ ID NO:
51)
(SEQ ID NO:
52, SEQ ID NO:
53 and SEQ ID NO:
54)
(SEQ ID NO:
55 and SEQ ID NO:
56)
(SEQ ID NO:
57, SEQ ID NO:
58 and SEQ ID NO:
59)
(SEQ ID NO:
60, SEQ ID NO:
61 and SEQ ID NO:
62)
x being 2, 3 or 4,
or the sequences of said probes having at least 92% identity with the abovementioned sequences SEQ ID NO:
27, SEQ ID NO:
28, SEQ ID NO:
29, SEQ ID NO:
30, SEQ ID NO:
31, SEQ ID NO:
32, SEQ ID NO:
33, SEQ ID NO:
34, SEQ ID NO:
35, SEQ ID NO:
36, SEQ ID NO:
37, SEQ ID NO:
38, SEQ ID NO:
39, SEQ ID NO:
40, SEQ ID NO:
41, SEQ ID NO:
42, SEQ ID NO:
43, SEQ ID NO:
44, SEQ ID NO:
45, SEQ ID NO:
46, SEQ ID NO:
47, SEQ ID NO:
48, SEQ ID NO:
49, SEQ ID NO:
50, SEQ ID NO:
51, SEQ ID NO:
52, SEQ ID NO:
53, SEQ ID NO:
54, SEQ ID NO:
55, SEQ ID NO:
56, SEQ ID NO:
57, SEQ ID NO:
58, SEQ ID NO:
59, SEQ ID NO:
60, SEQ ID NO:
61, SEQ ID NO:
62,
for the detection of toxinogenic cyanobacteria of the genus Anabaena, or pair of probes whose sequences are chosen from x elements of one of the following sets:
(SEQ ID NO:
63, SEQ ID NO:
64, SEQ ID NO:
65 and SEQ ID NO:
66)
(SEQ ID NO:
67, SEQ ID NO:
68 and SEQ ID NO:
69)
x being 3 or 4,
or the sequences of said probes having at least 92% identity with the abovementioned sequences SEQ ID NO:
63, SEQ ID NO:
64, SEQ ID NO:
65, SEQ ID NO:
66, SEQ ID NO:
67, SEQ ID NO:
68, SEQ ID NO:
69,
for the detection of toxinogenic cyanobacteria of the genus Planktothrix.

173 more elements

Kit for the detection of toxinogenic cyanobacteria of the genus Microcystis, said kit containing:

a) at least one pair of probes specific to toxinogenic cyanobacteria of the genus Microcystis, the sequences of said probes being selected from x elements of one of the following sets:
(SEQ ID NO:
1, SEQ ID NO:
2, SEQ ID NO:
3 and SEQ ID NO:
4)
(SEQ ID NO:
5, SEQ ID NO:
6 and SEQ ID NO:
7)
(SEQ ID NO:
8, SEQ ID NO:
9, SEQ ID NO:
10 and SEQ ID NO:
11)
(SEQ ID NO:
12, SEQ ID NO:
13 and SEQ ID NO:
14)
(SEQ ID NO:
15, SEQ ID NO:
16 and SEQ ID NO:
17)
(SEQ ID NO:
18 and SEQ ID NO:
19)
x being 2, 3 or 4
or the sequences of said probes having at least 92% identity with the abovementioned sequences SEQ ID NO:
1, SEQ ID NO:
2, SEQ ID NO:
3, SEQ ID NO:
4, SEQ ID NO:
5, SEQ ID NO:
6, SEQ ID NO:
7, SEQ ID NO:
8, SEQ ID NO:
9, SEQ ID NO:
10, SEQ ID NO:
11, SEQ ID NO:
12, SEQ ID NO:
13, SEQ ID NO:
14, SEQ ID NO:
15, SEQ ID NO:
16, SEQ ID NO:
17, SEQ ID NO:
18, SEQ ID NO:
19,
one probe of said pair being a capture probe linked to at least one attachment molecule positioned at 3′ or 5′ of its sequence and the other probe of said pair being a signal probe linked to at least one marking molecule positioned at 3′ or 5′ of its sequence, said capture probe and said signal probe being capable of hybridizing with the ribosomal nucleic acid of a toxinogenic cyanobacteria of the genus Microcystis,
b) optionally a hybridization solution
c) optionally a washing solution
d) optionally one or more revelation solutions.

48 more elements

Dependencies are inferred from the claim text.

Use of probes to detect toxinogenic cyanobacteria, detection method and corresponding kits

Life of this patent

Abstract

Claims21

Use of probes to detect toxinogenic cyanobacteria, detection method and corresponding kits

Life of this patent

Abstract

Claims21