Automated method for selecting microbial strains which can degrade or emulsify oil

Patent metadata

Field	Value
Publication number	US-10767159-B2
Application number	US-201615546771-A
Country	US
Kind code	B2
Filing date	Jan 27, 2016
Priority date	Jan 28, 2015
Publication date	Sep 8, 2020
Grant date	Sep 8, 2020

Abstract

The invention provides an automated method for selecting a microbial strain from within a microbial strain library which can degrade or emulsify a target oil substrate. An apparatus, device or system adapted to perform the method of the invention is further provided. The use of the method of the invention in a method for preparing a microbial inoculum suitable for use to degrade or emulsify a target oil substrate; in a method of treating an oil reservoir or environmental remediation; and in a method for producing a biosurfactant-like substance is also provided.

First claim

The invention claimed is:

1. An automated method for selecting a microbial strain from within a microbial strain library which can degrade or emulsify a target oil substrate, said method comprising in no particular order, unless specified:
(a) providing a plurality of receptacles adapted to receive a liquid microbial cell culture as part of a multi-well culture plate wherein the internal surface(s) of the receptacles are coated, at least in part, with a layer of the target oil substrate;
(b) applying to each of said oil-coated receptacle a sample of one of the members of the strain library, wherein a plurality of members of the strain library are applied to the plate;
(c) culturing said samples in a liquid cell culture medium and monitoring said oil layer coating at least part of the internal surface(s) of said receptacles for a change in its appearance, wherein a change in the appearance of said oil layer coating at least part of the internal surface(s) of said receptacles is indicative of degradation or emulsification of said target oil substrate;
(d) selecting those samples from step (c) which caused degradation or emulsification of the oil substrate;
(e) separately culturing either (i) all the members of the library applied in step (b) or (ii) the library members selected in step (d) in the presence of an n-alkane and monitoring for degradation of said n-alkane;
(f) separately culturing either (i) all the members of the library applied in step (b) or (ii) the library members selected in step (d) in the presence of a polycyclic aromatic hydrocarbon (PAH) and monitoring for degradation of said PAH;
(g) subjecting either (i) all the members of the library applied in step (b) or (ii) the library members selected in step (d) to environmental tolerance testing, wherein said testing comprises culturing the library members under varying environmental conditions and determining the limits of these conditions at which colony viability ceases, wherein said environmental conditions tested comprising temperature, pH and ionic concentration, and optionally further comprising O 2 concentration or pressure;
(h) optionally repeating steps (a) to (c) with the library members selected at step (d); and
(i) selecting a microbial strain on the basis of its performance in steps (c), (e), (f), (g) and optionally (h).

2. The automated method of claim 1 wherein the microbial strain selected on the basis of its performance in steps (c), (e), (f), (g) and optionally (h) is a microbial strain which causes biodegradation or emulsification of the oil substrate, degradation of said n-alkane or said PAH and which tolerates environmental conditions selected from temperature, pH and ionic concentration, and optionally O 2 concentration or pressure that expected to be encountered during its use.

3. The automated method of claim 2 wherein each microbial strain of the library is scored in order of relative performance during steps (c), (e), (f), (g) and optionally (h) and selection is based on said scores.

4. The automated method of claim 3 , wherein greater weight is given to the score for steps (c) and optionally (h) than steps (e), (f) or (g).

5. The automated method of claim 3 wherein the microbial strains of the library are ranked based on said scores and selection is of the portion of strains at the top of the rankings.

6. The automated method of claim 1 , wherein said multi-well plate is a microplate.

7. The automated method of claim 1 , wherein said oil is crude oil.

8. The automated method of claim 1 , wherein said culturing in any one of steps (c), (e) or (f) is in a minimal medium.

9. The automated method of claim 1 , wherein the n-alkane is a C16 or a C 32 alkane.

10. The automated method of claim 1 , wherein the PAH is dibenzothiophene.

11. A method for preparing a microbial inoculum suitable for use to degrade or emulsify a target oil substrate, said method comprising
(i) performing the automated method as defined in claim 1 ,
(ii) culturing a sample of said selected strain to increase the numbers of cells in the culture.

12. A method of treating an oil reservoir or environmental remediation, said method comprising
(i) performing the automated method as defined in claim 1 , and
(ii) introducing a sample of said selected microbial strain or said microbial inoculum to an oil reservoir or contacting a site or material undergoing environmental remediation with a sample of said selected microbial strain or said microbial inoculum.

13. A method for producing a biosurfactant-like substance (BLS), said method comprising
(i) performing the automated method as defined in claim 1 and selecting a strain which can emulsify the target oil substrate,
(ii) culturing a sample of said selected strain under conditions which result in BLS production.

14. A method of treating an oil reservoir or environmental remediation, said method comprising
(i) the method for preparing a microbial inoculum as defined in claim 11 , and
(ii) introducing a sample of said selected microbial strain or said microbial inoculum to an oil reservoir or contacting a site or material undergoing environmental remediation with a sample of said selected microbial strain or said microbial inoculum.

Assignees

• Equinor Energy As

Inventors

• Kjølhamar Ane
• Skarstad Anita
• Kotlar Hans-Kristian

Classifications

• C12N1/20

  Bacteria; Culture media therefor · CPC title

• C09K8/582

  characterised by the use of bacteria · CPC title

• B01F23/40

  Mixing liquids with liquids; Emulsifying · CPC title

• C12N1/26 Primary

  Processes using, or culture media containing, hydrocarbons (refining of hydrocarbon oils by using microorganisms C10G32/00) · CPC title

• B01F3/08

  Operations & Transport · mapped topic