WO2015120156A1

WO2015120156A1 - Continuous flow organism extractor

Info

Publication number: WO2015120156A1
Application number: PCT/US2015/014633
Authority: WO
Inventors: Ted MOSKAL
Original assignee: Vivione Biosciences, LLC
Priority date: 2014-02-05
Filing date: 2015-02-05
Publication date: 2015-08-13

Abstract

An apparatus for the continuous capture of microorganisms in a liquid reservoir using immunological methods of capture are disclosed. Further, methods of using the apparatus are disclosed.

Description

CONTINUOUS FLOW ORGANISM EXTRACTOR

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to United States Provisional Application 61/935,934 filed on February 5, 2015 which is incorporated by reference herein in its entirety.

FIELD

[0002] The embodiments disclosed herein relate generally to microbiology. More particularly the embodiments disclosed herein relate to extracting microorganisms from a microorganism containing liquid media for analysis.

BACKGROUND

[0003] Infectious diseases are the second largest cause of death worldwide. Water discharged by home toilets, restaurant dishwashers, agricultural runoff, manufacturing processes, and other large-scale industrial activities contain harmful microorganisms.

[0004] To prevent infection of humans or other animals due to biologically contaminated water, or to treat humans or other animals after exposure, a physician needs to know how best to treat the infection. The best treatment will depend on identification of the microorganism in question.

[0005] In the case of microorganisms, screening can be performed in clinical laboratories but, despite the high global human and financial cost of infectious diseases, many tests for pathogenic microorganisms still use the labor-intensive methods developed by Pasteur in the 19th century. In these tests, microorganisms from the patient are grown in specialized culture media until they reach sufficient numbers to be seen by the human eye. This culture step takes 24-48 hours and, once the bacteria have been identified, it takes a further 24 hours before antibiotic susceptibility can be determined.

[0006] From taking a microorganism sample to identifying an appropriate antimicrobial therapy therefore takes 2 to 3 days. Current infectious disease diagnostics therefore have little impact on patient management. As a physician cannot afford to wait 3 days before starting treatment, patients are subjected to 'best guess' antimicrobial therapy, and this typically involves expensive broad- spectrum antimicrobials which can be unnecessary or inappropriate, and there is also an increased risk that antimicrobial resistance will develop.

[0007] Upon completion of the diagnostic test, they are generally used to confirm whether the 'best guess' was appropriate rather than being used to educate or inform clinical choices. In particular, it is an object to provide systems for both rapid identification and rapid antimicrobial sensitivity testing of clinically-important bacteria, direct from environmental samples.

[0008] In biologically contaminated water, the bacteria, while still posing a risk to human or animal health, is sometimes too diluted to quickly analyze by microscope. A method of continuously extracting microorganisms directly from the water in a manner wherein they are concentrated over a period of time and leaving the bulk of the water behind would be a valuable tool in microorganism diagnostics.

SUMMARY

[0009] Certain embodiments of the invention herein pertain to a microorganism extraction device comprising: an inlet tube defining a tube axis, the inlet tube having a proximal end and a distal end, the distal end having a closure and the proximal end having an opening, wherein the closure has a distal side oriented away from the proximal end of the inlet tube and a proximal side orientated toward the proximal end of the inlet tube. Still further, in this embodiment, the device comprises a pump motor with a proximal end being in physical connection with the distal side of the closure, the proximal end having a magnetic interface capable of exerting a rotational force; a rotatable pump inside the elongated tube and oriented along the tube axis, the rotatable pump having a proximal end and a distal end, the distal end abutting the proximal side of the closure and magnetically interfacing with the pump motor, the rotatable pump being capable of rotation within the inlet tube, wherein the rotation is at an angle which is perpendicular to the tube axis; a side port opening in the inlet tube, the side port opening being distal to the proximal end of the inlet tube; and an outlet tube oriented parallel to the tube axis, the outlet tube having a proximal end attached to an outlet tube proximal end closure and at least one outlet port positioned distally to the outlet tube proximal end closure, wherein the at least one outlet port is in fluid connection with the proximal end of the inlet tube through the side port opening. In the aforementioned embodiment, the outlet tube proximal end closure comprises an immunological trap for capturing microorganisms through antibody-ligand interaction.

[0010] In further embodiments, the device is capable of drawing liquid from the proximal end of the elongated tube, through the side port opening, into the outlet tube and out the at least one outlet port.

[0011] In further embodiments, the device further comprises a fluid ejector nozzle positioned within the outlet tube, and wherein the fluid ejector nozzle has a distal end with an outer diameter substantially the same as the inner diameter of the outlet tube and a proximal end with an outer diameter less than the diameter of the distal end of the fluid ejector nozzle. In further embodiments, the side port is in fluid connection with the outlet port through the fluid ejector nozzle. In such embodiments the side port is in fluid connection with the outlet port through the fluid ejector nozzle. Still further, the proximal end of the fluid ejector nozzle is proximal to the at least one outlet port.

[0012] In embodiments of the invention concerning the pump of the device, in certain instances, the pump has a speed controlled by rotational force exerted by the pump motor.

[0013] In certain embodiments concerning the device, the immunological trap is an immuno- capture cone. Still further, the immune-capture cone comprises an immuno-screen extending distally into the outlet tube from the outlet tube proximal end closure. In further embodiments, the immuno-screen has a proximal side adapted to receive an immuno-screen holder. Further regarding the immuno-screen holder, the holder is capable of being removed as the immuno- screen is adapted to receive a pipette tip when the immuno-screen holder is removed.

[0014] In other embodiments, the immunological trap is an affinity chromatography column.

[0015] Other embodiments of the invention relate to a method of collecting bacteria from a reservoir containing a liquid, the method comprising: 1) inserting the proximal end of the inlet tube of the aforementioned device into the reservoir; 2) turning on the motor of the device; 3) pumping liquid from the proximal end of the inlet tube into the fluid ejector nozzle; 4) exposing the immunological trap to the liquid exiting the fluid ejector nozzle; and wherein bacteria are captured by the immunological trap, and wherein after exposure of the immunological trap to the liquid, the liquid exits the device through the one or more outlet port.

BRIEF DESCRIPTION OF THE DRAWINGS

[0016] In order that the manner in which the above-recited and other enhancements and objects of the invention are obtained, we briefly describe a more particular description of the invention briefly rendered by reference to specific embodiments thereof which are illustrated in the appended drawings. Understanding that these drawings depict only typical embodiments of the invention and are therefore not to be considered limiting of its scope, we herein describe the invention with additional specificity and detail through the use of the accompanying drawings in which:

[0017] Fig. 1 is an embodiment of the microorganism capture device of the present invention; and

[0018] Fig. 2 is an illustration of the immuno-capture cone in greater detail.

List of Reference Numerals

[0019] 10 Inlet tube

[0020] 20 Inlet tube closure

[0021] 30 Pump motor

[0022] 40 Battery pack

[0023] 50 Pump

[0024] 60 Side port

[0025] 70 Outlet tube

[0026] 80 Outlet port

[0027] 90 Fluid ejector nozzle [0028] 100 Immune capture cone

[0029] 110 Immuno-screen

[0030] 120 Immuno-screen holder

[0031] 130 Screen

DETAILED DESCRIPTION

[0032] We show the particulars shown herein by way of example and for purposes of illustrative discussion of the preferred embodiments of the present invention only. We present these particulars to provide what we believe to be the most useful and readily understood description of the principles and conceptual aspects of various embodiments of the invention. In this regard, we make no attempt to show structural details of the invention in more detail than is necessary for the fundamental understanding of the invention. We indent that the description should be taken with the drawings. This should make apparent to those skilled in the art how the several forms of the invention are embodied in practice.

[0033] We mean and intend that the following definitions and explanations are controlling in any future construction unless clearly and unambiguously modified in the following examples or when application of the meaning renders any construction meaningless or essentially meaningless. In cases where the construction of the term would render it meaningless or essentially meaningless, we intend that the definition should be taken from Webster's Dictionary

3^rd Edition.

[0034] As used herein, the term "tube" means and refers to a physical structure surrounding a fluid flow path.

[0035] As used herein, the term "fluidly connected or in fluid connection" means and refers to a connection for the transfer of fluid between one fluid flow path to another fluid flow path.

[0036] As used herein, the term "fluid" refers to a non-solid material such as a gas, a liquid or a colloidal suspension capable of being transported through a pipe, line or conduit. Examples of fluids include by way of non-limiting examples the following: air, mud, water, blood and the like.

[0037] As used herein, the term "attached," or any conjugation thereof describes and refers to the at least partial connection of two items.

[0038] As used herein, the term "proximal" refers to a direction away the pump motor.

[0039] As used herein, the term "distal" refers to a direction towards the pump motor.

[0040] The embodiments herein pertain to a device to continuously capture microorganisms through a pump mechanism inserted into a liquid reservoir containing said microorganisms.

[0041] Certain embodiments of the invention concern the inlet tube and the outlet tube of the present invention wherein liquid is pumped or drawn into the inlet tube and released through the outlet tube. In such embodiments, the tubes can be any shape as long as they are able to function for their intended purpose of drawing or pumping liquid into the inlet tube and disposing of said liquid through the outlet tube. In this manner, the tubes can be cylindrical, oval, triangular, rectangular, octagonal and the like.

[0042] In certain further embodiments of the invention concerning the inlet tube and the outlet tube, the tubes can be made of any substance as long as they are able to function for their intended purpose of drawing or pumping liquid into the inlet tube and disposing of said liquid through the outlet tube. Still further, the substance should be capable of allowing a magnetic interface between the pump and the pump motor. Examples of such substances include glass, acrylic, polyvinyl chloride, steel, poly-propylene and the like.

[0043] Certain further embodiments of the invention pertain to the pump. The pump is a pump capable of rotation in the inlet tube. In certain embodiments, the pump is physically connected to the motor. In other embodiments, the pump is magnetically interfaced to the motor. In certain embodiments, the pump is a lift pump. In certain other embodiments the pump is an impeller.

[0044] Further embodiments of the invention pertain to the motor. In certain embodiments, the motor is an electric motor. In other embodiments, the motor is a gas powered motor. In embodiments wherein the motor is an electric motor, the motor can be a direct current powered motor or an alternating current powered motor. In embodiments wherein the motor is a direct current powered motor, the motor is electrically connected to a battery pack. In still other embodiments, the pump is powered manually.

[0045] Certain further embodiments concern the fluid ejector nozzle. In certain embodiments, the nozzle is replaceable with a nozzle having a different diameter proximal end or a different length. In such embodiments, the proximal end of the outlet tube can be opened and the nozzle is replaced. In other embodiments, the nozzle is not interchangeable. The nozzle, like the inlet tube and the outlet tube, is made of a material that does not deviate from the purpose of device. Examples of materials suitable for the fluid ejector nozzle include, but are not limited to: glass, rubber, plastic, metal and ceramic.

[0046] In certain further embodiments, the proximal end of the outlet tube, and accordingly, the immuno-capture cone, can be moved in a proximal or distal direction by using a proximal end with distal facing side walls of different lengths. As a non-limiting example, a user can think of this proximal end as a cap with sides protruding distally around the center of the cap. In this respect, the cap "wall" would surround the outlet tube and if pushed in all the way, would position the immuno-capture cone closer to the fluid ejector nozzle. Likewise, if the cap "wall" is pulled out in a proximal direction, the immuno-capture cone would be further from the fluid ejector nozzle.

[0047] Other embodiments of the invention concern the immuno-capture cone. In such embodiments the immuno-capture cone comprises an immuno-screen. The immuno-screen is attached to antibodies or antibody fragments which are able to bind to ligands on the surface of bacteria. The immuno-screen is further adapted to receive a pipette tip on its proximal side when an immuno-screen holder is not in place.

[0048] In implementation, a user places the proximal end of the inlet tube of the device into a liquid reservoir in which microorganisms are thought to be present. The proximal end of the inlet tube is submerged, however the proximal end of the outlet tube is not submerged. The motor is turned on. As the motor is magnetically coupled to the pump, the motor exerts a rotational force that in turn causes the pump to rotate within the inlet tube. The rotation of the pump results in the pump pulling liquid from the reservoir into the device. As the liquid rises distally in the inlet tube, it flows into a side port and into the outlet tube. The liquid flows through the fluid ejector nozzle and onto the immuno-cone.

[0049] The speed and pressure of the pump can be adjusted by adjusting the speed of the motor. In such instances, the device can deliver an amount of liquid to the immuno-cone that fosters antibody-ligand binding but does not wash away collected microorganisms due to nozzle pressure. This can also be adjusted in certain embodiments by changing the diameter or length of the fluid ejector nozzle or by changing the distance from the nozzle to the immuno-cone by moving the proximal end of the outlet tube closer or further from the nozzle. In this way, the liquid can become static near the immuno-cone so as not to wash away bacteria. Further, to prevent pressure from washing away the bacteria, the outlet ports are distal to the proximal end of the fluid ejector nozzle. This allows the immuno-cone to be submerged in liquid while bacteria within the liquid are exposed to the cone.

[0050] Further in implementation, the immuno-cone can be exchanged for another immuno- cone with different antigen binding properties. For example, the antigen binding of one cone could be focused on recovery of Yersinia pestis, while another cone could be focused on E. coli.

[0051] Upon exposure to the immuno-cone, which comprises the immuno-screen and is plugged from the outside of the device by the immuno-screen holder, the liquid rises in the outlet tube until it exits the device through outlet ports. As the liquid exits while other liquid enters, the device can operate indefinitely until the user feels that the amount of microorganism present for analysis is sufficient.

[0052] When the user feels that the amount of microorganism present for analysis is sufficient, the user turns off the motor, which stops the pump. The user is then able to remove the pump from the liquid reservoir. Upon removal, the immuno-screen holder is removed and a pipette tip is inserted into the immuno-screen. The pipette tip is able to draw microorganism which were immunologically trapped for further analysis.

EXAMPLES

[0053] The following examples are included to demonstrate preferred embodiments of the invention. It should be appreciated by those of skill in the art that the techniques disclosed in the examples which follow represent techniques discovered by the inventors to function well in the practice of the invention, and thus can be considered to constitute preferred modes for its practice. However, those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made in the specific embodiments which are disclosed and still obtain a like or similar result without departing from the spirit or scope of the invention. The following Examples are offered by way of illustration and not by way of limitation.

[0054] Depicted in Fig. 1 is an embodiment of the microorganism capture device of the present invention. As can be seen, the device has an inlet tube with a proximal end, which would be inserted into a liquid reservoir. In this embodiment, the proximal end has a screen fitted over the end such that particulate matter will not be sucked into the extraction device. Other filters are envisioned to exclude certain specific microorganisms. For example, if a user of the device were to focus on extraction of coliform bacteria, a filter that could prevent introduction of protozoa could be employed.

[0055] Still further as depicted in Fig. 1, within the inlet tube is situated a pump which draws liquid through the screen and up the inlet tube until it reaches the side port. At the side port, the liquid is pushed down, by pump pressure or forced down by gravity, through the fluid ejector nozzle which is situated in the outlet tube. The liquid is then able to drip or exude onto the immuno-capture cone at the sealed proximal end of the outlet tube. The immuno-capture cone retains microorganisms through an antibody ligand interaction. The liquid then flows distally up the outlet tube and outside of the fluid ejector nozzle and exits the device through the outlet ports.

[0056] As can be further seen in Fig. 1, the pump, in this embodiment, extends the length of the inlet tube and the distal end of the pump abuts the inlet tube closure. Here the pump is isolated from the outside of the device, yet is magnetically coupled to the pump motor on the distal side of the inlet tube closure such that liquid does not contact the pump motor. Still further, the motor in this embodiment is a DC motor and is powered by a battery pack.

[0057] Fig. 2 is an illustration of the immuno-capture cone in greater detail. As can be seen, the immuno-capture cone is positioned at the proximal end of the outlet tube, in an outlet tube cap, thereby sealing the proximal end of the outlet tube. The immuno-capture cone is expanded in Fig. 2 to illustrate its two components. The most distal component is the immuno-screen. This fits inside the outlet tube. The screen is adapted to receive the immuno-screen holder. With the holder in place, liquid is prevented from exiting the proximal end of the outlet tube. When the immuno-screen holder is removed, the configuration of the immuno-screen is such that it is adapted to receive a pipette tip to draw off captured bacteria for analysis.

[0058] From the foregoing description, one of ordinary skill in the art can easily ascertain the essential characteristics of this disclosure, and without departing from the spirit and scope thereof, can make various changes and modifications to adapt the disclosure to various usages and conditions. For example, we do not mean for references such as above, below, left, right, and the like to be limiting but rather as a guide for orientation of the referenced element to another element. A person of skill in the art should understand that certain of the above-described structures, functions, and operations of the above-described embodiments are not necessary to practice the present disclosure and are included in the description simply for completeness of an exemplary embodiment or embodiments. In addition, a person of skill in the art should understand that specific structures, functions, and operations set forth in the above-described referenced patents and publications can be practiced in conjunction with the present disclosure, but they are not essential to its practice.

[0059] The invention can be embodied in other specific forms without departing from its spirit or essential characteristics. A person of skill in the art should consider the described embodiments in all respects only as illustrative and not restrictive. The scope of the invention is, therefore, indicated by the appended claims rather than by the foregoing description. A person of skill in the art should embrace, within their scope, all changes to the claims which come within the meaning and range of equivalency of the claims. Further, we hereby incorporate by reference, as if presented in their entirety, all published documents, patents, and applications mentioned herein.

Claims

CLAIMS What is claimed is:

1. A microorganism extraction device comprising: a. an inlet tube defining a tube axis, the inlet tube having a proximal end and a distal end, the distal end having a closure and the proximal end having an opening, wherein the closure has a distal side oriented away from the proximal end of the inlet tube and a proximal side orientated toward the proximal end of the inlet tube; b. a pump motor with a proximal end being in physical connection with the distal side of the closure, the proximal end having an interface capable of exerting a rotational force; c. a rotatable pump inside the elongated tube and oriented along the tube axis, the rotatable pump having a proximal end and a distal end, the distal end abutting the proximal side of the closure and interfacing with the pump motor, the rotatable pump being capable of rotation within the inlet tube; d. a side port opening in the inlet tube, the side port opening being distal to the proximal end of the inlet tube; e. an outlet tube oriented at an angle other than perpendicular to the tube axis, the outlet tube having a proximal end attached to an outlet tube proximal end closure and at least one outlet port positioned distally to the outlet tube proximal end closure, wherein the at least one outlet port is in fluid connection with the proximal end of the inlet tube through the side port opening; and wherein the outlet tube proximal end closure comprises a immunological trap for capturing microorganisms through antibody-ligand interaction.

2. The device of claim 1, wherein the device is capable of drawing liquid from the proximal end of the elongated tube, through the side port opening, into the outlet tube and out the at least one outlet port.

3. The device of claim 1, wherein the rotatable pump is a lift pump.

4. The device of claim 1, further comprising a filter proximal to the proximal end of the elongated tube.

5. The device of claim 1, wherein the outlet tube has an internal diameter and wherein the device further comprises a fluid ejector nozzle positioned within the outlet tube, and wherein the fluid ejector nozzle has a distal end with an outer diameter substantially the same as the inner diameter of the outlet tube and a proximal end with an outer diameter less than the diameter of the distal end of the fluid ejector nozzle.

6. The device of claim 5, wherein the side port is in fluid connection with the outlet port through the fluid ejector nozzle.

7. The device of claim 6, wherein the proximal end of the fluid ejector nozzle is proximal to the at least one outlet port.

8. The device of claim 1, wherein the pump has a speed controlled by rotational force exerted by the pump motor.

9. The device of claim 1, wherein the pump motor is powered by a battery

10. The device of claim 1, wherein the immunological trap is an immuno-capture cone.

11. The device of claim 10, wherein the immune-capture cone comprises an immuno-screen extending distally into the outlet tube from the outlet tube proximal end closure.

12. The device of claim 11, wherein the immuno-screen has a proximal side adapted to receive an immuno-screen holder.

13. The device of claim 12, wherein the immuno- screen holder is capable of being removed, and wherein the immuno-screen is adapted to receive a pipette tip when the immuno- screen holder is removed.

14. The device of claim 1, wherein the immunological trap is an affinity chromatography column.

15. A method of collecting bacteria from a reservoir containing a liquid, the method comprising: a. inserting the proximal end of the inlet tube of the device of claim 6 into the reservoir; b. turning on the motor of the device; c. pumping liquid from the proximal end of the inlet tube into the fluid ejector nozzle; d. exposing the immunological trap to the liquid exiting the fluid ejector nozzle; and wherein bacteria are captured by the immunological trap, and wherein after exposure of the immunological trap to the liquid, the liquid exits the device through the one or more outlet port.

16. The method of claim 15, wherein the immunological trap is an immuno-capture cone comprising an immuno-screen extending distally into the outlet tube from the outlet tube proximal end closure and wherein the immuno-screen has a proximal side receiving an immuno-screen holder.

17. The method of claim 16, wherein after a period of time wherein the immunological trap is exposed to liquid, the motor is turned off and the device is removed from the reservoir.

18. The method of claim 17, wherein upon removal of the device from the reservoir, the immuno-screen holder is removed and a pipette tip is inserted in its place, the pipette tip drawing bacteria captured by the immuno-screen into a pipette for further analysis of the bacteria.