Probe matrix-based device for identifying microorganisms
View Patent ↗Device for identifying microbial organisms using a collection of polynucleotide probes, each probe having binding specificity for the ribosomal nucleic acids of at least one microbe. The collection of probes is organized into a series of “addresses” that provide information about the presence or absence of one or more polynucleotide sequences in the biological sample. The system is particularly suited to automated analysis.
1. A device for detecting nucleic acids, comprising:
a solid support; and
a plurality of addresses disposed on the solid support, each address comprising at least one detectably labeled probe capable of hybridizing to ribosomal nucleic acids from at least one microbial species under high stringency conditions, said plurality of addresses comprising
a first address comprising one or more detectably labeled probes capable of hybridizing to ribosomal nucleic acids of more than one microbial species, and
a second address comprising a mixture of
(a) a first detectably labeled probe capable of hybridizing to ribosomal nucleic acids from a subset of species having ribosomal nucleic acids that hybridize at the first address, and
(b) a second detectably labeled probe capable of hybridizing to ribosomal nucleic acids from a microbial species which does not have ribosomal nucleic acids that hybridize at the first address, but not capable of hybridizing to ribosomal nucleic acids from any microbial species having ribosomal nucleic acids that hybridize at the first address.
2. The device of claim 1 , wherein said subset of species having ribosomal nucleic acids that hybridize at the first address comprises at least one bacterial species having ribosomal nucleic acids that hybridize at the first address.
3. The device of claim 2 , wherein said microbial species which does not have ribosomal nucleic acids that hybridize at the first address comprises a bacterial species which does not have ribosomal nucleic acids that hybridize at the first address.
4. The device of claim 2 , wherein said microbial species which does not have ribosomal nucleic acids that hybridize at the first address comprises a fungal species which does not have ribosomal nucleic acids that hybridize at the first address.
5. The device of claim 1 , wherein said subset of species having ribosomal nucleic acids that hybridize at the first address comprises at least one fungal species having ribosomal nucleic acids that hybridize at the first address.
6. The device of claim 5 , wherein said microbial species which does not have ribosomal nucleic acids that hybridize at the first address comprises a bacterial species which does not have ribosomal nucleic acids that hybridize at the first address.
7. The device of claim 5 , wherein said microbial species which does not have ribosomal nucleic acids that hybridize at the first address comprises a fungal species which does not have ribosomal nucleic acids that hybridize at the first address.
8. The device of claim 1 , wherein each of said at least one detectably labeled probe capable of hybridizing to ribosomal nucleic acids is a detectably labeled soluble probe capable of hybridizing to ribosomal nucleic acids.
9. The device of claim 8 , wherein each of said at least one detectably labeled soluble probe capable of hybridizing to ribosomal nucleic acids comprises a label selected from the group consisting of a chemiluminescent label, a redox-active electron transfer label, and a fluorescent label.
10. The device of claim 9 , wherein each of said at least one detectably labeled soluble probe capable of hybridizing to ribosomal nucleic acids comprises a chemiluminescent label.
11. The device of claim 1 , wherein said first address is selected from the group consisting of:
(a) a pan-bacterial address that hybridizes to ribosomal nucleic acids from a plurality of species of Gram(+) bacteria, a plurality of species of bacteria in the family Enterobacteriaceae, a plurality of species of bacteria in the genus Enterococcus , a plurality of species of bacteria in the genus Staphylococcus , and a plurality of species of bacteria in the genus Campylobacter,
(b) a pan-fungal address that hybridizes to ribosomal nucleic acids from a plurality of fungal species,
(c) a Gram(+) address that hybridizes to ribosomal nucleic acids of a plurality of Gram(+) bacteria, wherein said plurality of Gram(+) bacteria comprise a plurality of bacteria in the genus Staphylococcus,
(d) a family Enterobacteriaceae address that hybridizes to ribosomal nucleic acids from a plurality of bacteria in the family Enterobacteriaceae, and
(e) a Staphylococcus genus address that hybridizes to ribosomal nucleic acids from a plurality of species in the Staphylococcus genus.
12. The device of claim 2 , wherein said at least one bacterial species having ribosomal nucleic acids that hybridize at the first address comprises a species selected from the group consisting of Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, and Streptococcus pneumoniae.
13. The device of claim 12 , wherein said first address is a family Enterobacteriaceae address that specifically hybridizes ribosomal nucleic acids from a plurality of bacteria in the family Enterobacteriaceae, and wherein said at least one bacterial species having ribosomal nucleic acids that hybridize at the first address comprises Escherichia coli.
14. The device of claim 2 , wherein said first address is a Gram(+) address that specifically hybridizes ribosomal nucleic acids of a plurality of Gram(+) bacteria, and wherein said microbial species which does not have ribosomal nucleic acids that hybridize at the first address comprises Escherichia coli.
15. The device of claim 3 , wherein said first address is a Staphylococcus genus address that hybridizes ribosomal nucleic acids from a plurality of species in the Staphylococcus genus.
16. The device of claim 15 , wherein said at least one bacterial species having ribosomal nucleic acids that hybridize at the first address comprises Staphylococcus aureus.
17. The device of claim 4 , wherein said fungal species which does not have ribosomal nucleic acids that hybridize at the first address comprises Candida albicans.
18. The device of claim 5 , wherein said at least one fungal species having ribosomal nucleic acids that hybridize at the first address comprises Candida albicans.
19. The device of claim 6 , wherein said bacterial species which does not have ribosomal nucleic acids that hybridize at the first address comprises a species selected from the group consisting of Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, and Streptococcus pneumoniae.
20. The device of claim 7 , wherein said first detectably labeled probe is capable of hybridizing to ribosomal nucleic acids from Candida albicans but not to the ribosomal nucleic acids of other species in the genus Candida.