Microorganism transfer device

Abstract

The invention relates to a device for transferring microorganisms. It comprises: - a tube (12), having a first end (14) engaged through a stopper (16) and a second end (15); - a first piston (30) having an orifice (36) and a second piston (32) engaged inside said tube (12) and connected by a rod (26) forming a chamber (34) suitable for receiving microorganisms; - a bell (40) having a finger (44) mounted to engage with said stopper (16) so that said finger (44) extends opposite said first piston (30); Said bell (40) is adapted to be driven so that said finger (44) comes to rest against said first piston (30) and to drive said second piston (32) in translation out of said tube (12) so as to free said second free end (15). Figure to be published with the abbreviation: Fig. 2

Description

Title of the invention: Microorganism transfer device

[0001] The present invention relates to a device for transferring microorganisms in a dormant state to a culture medium.

[0002] One envisaged field of application is that of the production of microorganisms to promote the growth of agricultural plant production.

[0003] It is known to apply agrochemicals such as fertilizers or plant protection products to plant crops to promote their growth and protect them from phytopathogenic and predatory agents.

[0004] On the other hand, the large-scale use of fertilizers and agrochemical plant protection products leads to significant emissions of greenhouse gases including carbon dioxide and nitrous oxide; it also contributes to the pollution of soils and subsoils, groundwater, watercourses, oceans and air and affects the biodiversity of ecosystems.

[0005] Consequently, it was conceived to implement more natural, and above all less polluting, fertilizing and / or phytosanitary elements.

[0006] Alternative solutions, at least partial, to these agrochemicals aim to implement plant growth-promoting bacteria. They are commonly referred to by the English acronym PGPB for "plant growth-promoting bacteria". Yeasts, fungi, microalgae, or other types of microorganisms can also be used.

[0007] Thus these microorganisms can be applied as a seed coating, in the crop furrow or by foliar spraying.

[0008] However, microorganisms are living organisms requiring very specific production and storage conditions before they can be introduced, while still alive, into the soil and into planted areas, in contrast to conventional mineral or synthetic fertilizers, which can be easily stored under certain conditions for extended periods. Furthermore, these fertilizers retain their simple chemical formula and constant activity over a long period.

[0009] Thus, a problem which arises and which the present invention aims to solve is to provide a transfer device which allows the microorganisms to be stored and transported in order to be transferred into a culture medium, and then, after in situ proliferation, to be easily and efficiently associated with plant cultures.

[0010] In order to solve this problem, and according to a first object, a sealed device for transferring microorganisms is proposed, comprising:

[0011] - a tube having a first end mounted axially in contact through a sealing element and a second free end opposite said first end;

[0012] - a first piston having an axial orifice, engaged inside said tube near of said first end and a second piston engaged inside said tube near said second free end, forming, between the two pistons, a chamber suitable for receiving microorganisms, said two pistons being linked together in translation by a rod extending inside said tube; and,

[0013] - a bell comprising a finger extending axially inside said bell, said bell being mounted in contact with said stopper so that said finger extends opposite said first piston.

[0014] And said bell is adapted to be driven in motion relative to said sealing member so that said finger comes to rest against said first piston and to drive said second piston in translation out of said tube so as to release said second free end, whereby said microorganisms contained inside said chamber escape from said tube.

[0015] Thus, a feature of the invention lies in the implementation of a transfer device comprising a chamber for holding microorganisms and a sealing element adapted to be installed in the neck of a sealed container so as to allow the microorganisms to be transferred into the container aseptically, protected from air. In other words, the transfer device allows the microorganisms to be stored in a dormant state after their production and transported to their culture and processing location.

[0016] The microorganisms are trapped in the tube between the two pistons, and they are released from the second end of the tube when the two pistons are driven in translation by means of the bell.

[0017] In other words, microorganisms can be transferred simply, in a culture medium, efficiently and in aseptic conditions.

[0018] This transfer can be carried out by a non-specialized operator, for example, a farmer, and not solely by a microbiologist under laboratory conditions within a biosafety cabinet. With a simple transfer device according to the invention, the operator can then trigger their proliferation and the decentralized production of a larger quantity of microorganisms near the point of use. For example, the proliferation can take place directly on a farm, near it, or on several farms, in environments containing many types of germs. Shortly thereafter, the newly amplified microorganisms can be used directly on-site.

[0019] Being produced extemporaneously, their effectiveness is much greater than that of microorganisms formulated industrially to be stable in the long term and consequently, more or less dormant at the time of their agricultural implementation.

[0020] Overall, the transfer device according to the invention promotes the economy of the process.

[0021] According to the invention, said bell is preferably screw-mounted onto said capping member. Consequently, the release of microorganisms occurs by rotating the bell relative to the capping member. In doing so, the finger, bearing against the first piston, moves it in translation, as does the second piston, via the rod.

[0022] According to a particularly advantageous embodiment of the invention, said finger is hollow to form a passage extending from said axial orifice of said first piston and opening at the top of said bell. Thanks to the hollow finger, extended opposite the axial orifice of the first piston, after the microorganisms have been poured into the container, the latter is supplied with oxygen from the air to allow their growth.

[0023] Preferably, said bell comprises a filter membrane mounted across said passageway, said filter membrane having pores with a diameter of less than 0.5 micrometers, preferably less than 0.2 micrometers. In this way, the membrane filters most of the germs contained in the ambient air, and in particular all bacteria. Consequently, the proliferation of microorganisms is not hindered by pathogenic germs.

[0024] According to a particularly advantageous embodiment of the invention, said bell comprises a lid located at the top of said bell to seal said passageway. Thus, during transport of the transfer device, the lid makes the chamber containing the microorganisms completely airtight with respect to the outside. This prevents any contamination and protects the filter membrane.

[0025] The lid is then removed after the microorganisms have been released from the chamber to be introduced into the container in which they will proliferate.

[0026] Advantageously, the transfer device includes a breakable ring mounted around said stopper, to prevent movement of said bell relative to said stopper. Movement of the bell relative to the stopper is only permitted when the ring is broken, thereby releasing the bell from the stopper.

[0027] The breakable ring makes it possible to prevent the relative movement of the bell and the sealing element during transport and thus, to prevent the release of microorganisms in an untimely manner.

[0028] According to a particularly advantageous embodiment of the invention, the rod connecting the two pistons is hollow and has a first open end protruding near the first piston and a second open end protruding near the second piston. The first and second ends of the hollow rod are preferably radially open.

[0029] According to one embodiment, the rod consists of a split hollow cylinder having a C-shaped cross-section and mounted coaxially inside the tube, forming a slide. The two pistons are mounted at the two ends of the hollow cylinder. The microorganisms are then stored inside the hollow cylinder. When the second piston is driven out of the tube, the microorganisms escape through the slot in the cylinder.

[0030] Advantageously, said rod has a wall that bears against the inner wall of said tube. In other words, the outer wall of the hollow cylinder bears against the inner wall of the tube. And when the bell causes the first piston to move in translation, the cylinder is simultaneously moved in translation inside the tube along with the second piston.

[0031] According to another object, a microorganism culture assembly is proposed comprising a sealed container ending in a neck and a transfer device as described above and engaged inside said neck so that said sealing member closes said sealed container.

[0032] The leak-proof container is, for example, a conical flask of the "Erlenmeyer" or "Fembach" type, or simply a bottle of substantially cylindrical shape, with a capacity greater than 500 mL, for example greater than 1,000 mL or greater than 10,000 mL. According to a particularly advantageous embodiment, the conical flask has a capacity of 5,000 mL.

[0033] The culture assembly also includes a liquid culture medium contained within said sealed container. The liquid culture medium comprises the nutrients necessary for the growth of microorganisms, and in particular sugars as a carbon source.

[0034] Preferably, the culture assembly further comprises a device for moving said liquid culture medium. For example, it comprises a magnetic stir bar installed inside the container, which is then mounted on a block including a permanent magnet driven in rotation by a motor so as to drive the magnetic stir bar and, consequently, the liquid culture medium with the microorganisms. Such agitation allows the microorganisms to be dispersed throughout the culture medium and thus promotes their multiplication. The agitation also ensures aeration of the medium since the filter membrane is permeable to gases.

[0035] Also, the container can be installed on the table of an orbital shaker as will be explained in more detail below.

[0036] Other features and advantages of the invention will become apparent from the following description of particular embodiments of the invention, given by way of example but not limitation, with reference to the accompanying drawings in which:

[0037] [Fig-1] is a schematic axial cross-sectional view of the object of the present application in a first state; and,

[0038] [Fig.2] is a schematic view of the object of [Fig.1] associated with another element and in a second state.

[0039] Figure 1 shows, in axial section, a transfer device 10 according to the invention. It comprises a tube 12 made here of a transparent polymer material, for example polycarbonate. The tube 12 extends longitudinally over a height H and transversely has a diameter D. The height H is, for example, between 5 cm and 15 cm, while the diameter D is between 1 cm and 5 cm.

[0040] The tube 12 has a first end 14, or upper end, mounted in contact with a stopper 16, adapted to form a stopper. At the opposite end, the tube 12 has a second end 15, or lower end, which is free.

[0041] The sealing member 16 has an upper part forming a crown 18 and a lower part forming a first skirt 20, extending from the crown 18. The first end 14 of the tube 12 is precisely fitted through the crown 18.

[0042] The lower part 20 of the sealing member 16 has cylindrical symmetry and its diameter is greater than that of the tube 12, so as to form a free annular space 22 between the first skirt 20 and the tube 12. For example, the diameter of the first skirt is greater than 1.5 times the diameter D of the tube 12. Advantageously, the diameter of the first skirt 20 is substantially equal to twice the diameter D.

[0043] In addition, the first skirt 20 has an internal wall 24 provided with helical threads for fixing by screwing as will be explained below.

[0044] The transfer device 10 comprises a hollow slotted cylinder 26, mounted to slide coaxially inside the tube 12. The cylinder 26 has a C-shaped cross-section defining an axial opening 28.

[0045] The transfer device 10 further includes an upper piston 30, or first piston, mounted to slide tightly inside the first end 14 of the tube 12, and opposite, a lower piston 32, or second piston, mounted to slide tightly inside the second end 15 of the tube 12.

[0046] The upper piston 30 and the lower piston 32 are respectively fixed to the two opposite ends of the hollow cylinder 26. In addition, the upper piston 30 and lower piston 32 seal the two opposite ends 14, 15 of the hollow cylinder 26 respectively so as to form a slide.

[0047] In other words, the two pistons 30, 32 are linked to each other in translation by means of the hollow split cylinder 26. And the hollow split cylinder 26 defines a receiving chamber 34.

[0048] The receiving chamber 34 defines a volume between 1 cm3 and 100 cm3. And it is intended to receive a strain of microorganisms in a dormant state, prior to the insertion of the hollow cylinder 26 inside the tube 12. For example, 15 cm3 of bacteria of the species Azospirillum brasilense are placed inside the receiving chamber 34.

[0049] Furthermore, the upper piston 30 has an axial orifice 36 through which a conduit 38 is engaged which connects the receiving chamber 34 and the outside of the plugging member 16.

[0050] The transfer device 10 further comprises a bell 40 having a cylindrical wall 42 surmounted by a peak 43 and a hollow finger 44, extending axially inside the cylindrical wall 42 along the circular axis of symmetry of the cylindrical wall 42. The hollow finger 44 has a channel 45 opening at each of its ends. The width of the channel 45 illustrated in [Fig. 1] is not limited. The channel 45 can be wider to allow greater air circulation, as will be explained below. The bell 40 is mounted in contact with the sealing member 16 at the upper part forming a ring 18.

[0051] More specifically, the cylindrical wall 42 is screwed onto the sealing member, while the hollow finger 44 comes coaxially to bear against the conduit 38. Also, the cylindrical wall 42 has an edge 46, which bears axially against a breakable ring 48 mounted integrally with the sealing member 16.

[0052] Preferably, the cylindrical wall 42 has a collar, not shown, extending at the edge 46, while the breakable ring 48 has a rim with a return that engages in said collar to trap the bell 40.

[0053] The bell 40 is thus held in a fixed position so that the hollow finger 44 comes to be applied in a watertight manner on the edge of the conduit 30 in order to put the receiving chamber 34 and the channel 45 into communication.

[0054] Furthermore, the apex 43 of the bell 40 has a central recess 50 extending around the end of the channel 45. A filter membrane 52 is then bonded into the recess 50 across the end of the channel 45. The filter membrane 52 has pores with a diameter of less than 0.5 µm. Such a membrane is designed to prevent the passage of pathogens.

[0055] In addition, the top 43 of the bell and the membrane 52 are covered by a lid 54. This lid is sealed to the top 43 and it allows the conduit 45 to be sealed airtight. According to another embodiment, not shown, the top of the bell and the filter membrane are covered by a plug screwed onto the bell 40.

[0056] Thus, such a transfer device 10 is intended to be loaded with a given quantity of microorganisms at an industrial microorganism production site. It can then be stored and subsequently transported to a farm for use, as will be explained below with reference to Figure 2.

[0057] Previously, it was noted that microorganisms can be dispersed in a liquid. Therefore, to prevent the liquid from saturating the filter membrane during transport of the transfer device, a thin, locally sealed film is interposed between the free end of the finger 44 and the conduit 38 to seal the latter. The thin film thus protects the filter membrane from the liquid medium in which the microorganisms are immersed. The thin film can then be torn and automatically removed from the conduit when the bell 40, and consequently the finger 44, are rotated relative to the sealing element 16 and the conduit 38.

[0058] The objective is to be able to produce large quantities of microorganisms in situ in a simple way so that they can then be spread and brought into contact with cultivated plants.

[0059] To achieve this, necked containers are used, for example a conical flask 56 as shown in [Fig. 2]. Its maximum volume is 3 liters here. However, conical flasks of smaller or larger volumes can be used.

[0060] Furthermore, and advantageously, containers will be used whose surface is mostly opaque but which has a transparent window allowing the passage of light for an optical measurement of the proliferation of microorganisms while avoiding stray light which could disrupt said measurement.

[0061] Also, a conical flask with an external screw neck 58, compatible with the stopper 16, will be chosen.

[0062] In addition, beforehand, an aqueous culture medium 60 comprising mannose as a carbon source and also potassium phosphate, magnesium sulfate, sodium chloride, calcium chloride, ferric chloride and sodium molybdate is injected into the conical flask 56.

[0063] A magnetic bar 62 is also inserted there. Its role will be explained below.

[0064] After the transfer device 10 has been installed on the conical flask 56, otherwise said, after the inner wall 24 of the first skirt 20 has been screwed onto the neck 58 of the conical flask 56, the microorganisms will be released from the receiving chamber 34, so that they can move by gravity into the culture medium 60.

[0065] To do this, the breakable ring 48 is first broken so as to be able to release the bell 40.

[0066] The cylindrical wall 42 of the bell 40 being screwed onto the stopper 16, the bell 40 is screwed even further onto the stopper 16 so as to to drive it in translation. By the same token, the finger 44, resting against the conduit 38, simultaneously drives in translation the two pistons 30, 32 and the hollow cylinder 26.

[0067] In so doing, the lower piston 32, initially engaged in the tube 12 at its second end 15, is drawn in translation out of the tube 12. And in addition, part of the axial opening 28 of the hollow cylinder 26 is also drawn out of the tube 12 by releasing a window 64.

[0068] In this way, the microorganisms contained in the receiving chamber 34 escape and end up in the culture medium 60.

[0069] Next, the operculum 54 appearing on [Fig. 1] is removed to allow the passage of air through the filter membrane 52. The air thus filtered can then travel through the channel 45, and the conduit 38 to reach the inside of the conical flask 56 by passing through the hollow cylinder 26.

[0070] Then, the conical flask 56 is placed on a magnetic stirrer 66 adapted to drive the magnetic bar 62 in rotation to cause agitation of the culture medium 60 including the microorganisms.

[0071] Other methods of agitating the culture medium and microorganisms are envisaged. The conical flask 56 can be mounted on an orbital shaker, which moves the flask itself in a horizontal plane.

[0072] Such an orbital shaker also has the advantage of being able to accommodate a plurality of conical flasks of the same type including a culture medium and microorganisms.

[0073] According to another embodiment, cylindrical containers with a neck and a capacity greater than 5 liters are used, for example with a capacity of 10 or 20 liters.

[0074] Thus, thanks to the transfer device according to the invention, microorganisms can be easily cultured and multiplied in situ in a farm by a non-specialized operator and under satisfactory aseptic conditions.

[0075] We then have freshly amplified microorganisms, which are much more efficient in terms of activity than microorganisms formulated to be stable in the long term and even more or less dormant.

[0076] In addition to the production of plants, the transfer device according to the invention can be implemented, in the agricultural field, for the treatment of residues from animal production, for example for the treatment of bovine slurry.

[0077] Another use concerns the inclusion of microorganisms in animal feed. These microorganisms, then usually called probiotics, can confer various properties of interest such as, in the example of cattle production: improved milk or meat yield; relative protection against certain diseases such as mastitis; reduction of methane production, for example if these microorganisms use hydrogen and reduce its availability to methanogenic microorganisms present in the rumen.

[0078] For example, microorganisms are introduced into corn silage before fermentation.

Claims

Demands

1. A sealed microorganism transfer device characterized in that it comprises: - a tube (12) having a first end (14) mounted axially engaged through a sealing member (16) and a second free end (15) opposite said first end; - a first piston (30) having an axial orifice (36), engaged inside said tube (12) near said first end (14) and a second piston (32) engaged inside said tube near said second free end (15) forming, between the two pistons, a chamber (34) suitable for receiving microorganisms, said two pistons (30, 32) being linked together in translation by a rod (26) extending inside said tube; - a bell (40) comprising a finger (44) extending axially inside said bell, said bell being mounted in contact with said plugging member (16) so that said finger (44) extends opposite said first piston (30);and in that said bell (40) is adapted to be driven in motion relative to said sealing member (16) so that said finger (44) comes to rest against said first piston (30) and to drive said second piston (32) in translation out of said tube (12) so as to release said second free end (15), whereby said microorganisms contained within said chamber (34) escape from said tube.;

2. Transfer device according to claim 1, characterized in that said bell (40) is screw-mounted on said plugging member (16).

3. Transfer device according to claim 1 or 2, characterized in that said finger (44) is hollow to form a passageway (45) extending from said axial orifice of said first piston and opening at the top (43) of said bell.

4. Transfer device according to claim 3, characterized in that said bell (40) comprises a filter membrane (52) mounted across said passageway (45), said filter membrane having pores of a diameter less than 0.5 micrometer.

5. Transfer device according to claim 3 or 4, characterized in that said bell (40) comprises a lid (54) located at the top (43) of said bell to close said passageway (45).

6. Transfer device according to any one of claims 1 to 5, characterized in that it comprises a breakable ring (48) mounted around said sealing member (16), to prohibit the movement of said bell (40) relative to said sealing member.

7. Transfer device according to any one of claims 1 to 6, characterized in that said rod (26) connecting together the two pistons (30, 32) is hollow and in that it has a first open end opening in the vicinity of said first piston (30) and a second open end opening in the vicinity of said second piston (32).

8. Transfer device according to claim 7, characterized in that said rod (26) has a wall that applies itself against the inner wall of said tube (12).

9. Microorganism culture assembly comprising a sealed container (56) terminated by a neck (58) and a transfer device (10) according to any one of claims 1 to 8, engaged inside said neck (58) so that said sealing member (16) seals said sealed container (56).

10. Culture assembly according to claim 9, characterized in that it further comprises a liquid culture medium (60) contained inside said sealed container (56).

11. Culture assembly according to claim 10, characterized in that it further comprises a moving drive device (62) for said liquid culture medium.

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