WO2014057233A2

WO2014057233A2 - Screening method and apparatus

Info

Publication number: WO2014057233A2
Application number: PCT/GB2013/000420
Authority: WO
Inventors: Brian Gordon FORDE
Original assignee: Lancaster University Business Enterprises Ltd
Priority date: 2012-10-09
Filing date: 2013-10-08
Publication date: 2014-04-17
Also published as: GB201218089D0; WO2014057233A3

Abstract

According to the present invention there is provided an organism growth array device suitable for automated phenotypic analysis of either plants or microorganisms under aseptic conditions and comprising: a plurality of receptacles, each receptacle having a housing comprising first and second open ends and one or more longitudinal sides extending from the first open end to the second open end, wherein at least one of the longitudinal sides is flat, and wherein a portion of each receptacle is fused to the neighbouring receptacle(s) along at least 10% of its length along the longitudinal axis of the receptacle; growth medium housed within the housing extending from a first surface of the growth medium, disposed longitudinally towards the first open end of the receptacle to a second surface of the growth medium disposed towards the second open end of the receptacle, wherein generally, in use, the first surface of the growth medium is disposed above the second surface of the growth medium; wherein the open ends of each receptacle are individually accessible, preferably for the addition of different compositions; and wherein the growth medium of each receptacle is visible, and imagable across the lateral axis of the receptacle and along the longitudinal axis of the receptacle from the first surface to the second surface, and in use the growth of the organism within or into the growth medium is visible and imagable across the lateral axis of the receptacle and along the longitudinal length of the growth medium. There is also provided a method of imaging and/or analysing the development of one or more organisms.

Description

SCREENING METHOD AND APPARATUS

The present invention relates to an organism growth array device and a method of imaging one or more organisms cultured in such a device. There is also provided a method of analysing the development of one or more organisms and an assay for identifying a molecule or composition with bioactive potential or for detecting or measuring the bioactivity of a biological or environmental sample.

BACKGROUND TO THE INVENTION

Methods of imaging organisms are known, including methods of imaging the roots of a plant. For instance, US 201 1/0197509 discloses a method of imaging a root and/or aerial portion of a plant, in particular the root tip morphology of the plant to assess its development. This document discloses that such a method may be used to measure environmental conditions. EP1 154370 discloses a method of evaluating the growth of a plant automatically through digital imaging techniques.

However, there remains a long-felt need for an accurate, reproducible and high-throughput method of analysing the development of an organism such as a plant, in particular where such a method can be used to compare the effects of different variables on the development of the organism.

There is also a need for improved high-throughput methods to detect and measure antimicrobial activity (e.g. for discovery of novel antibiotics or fungicides or for measuring the levels of such bioactive compounds in biological or environmental samples). As noted in Pascual et al. (2012). ("A prescreening system for enriched selection of secondary metabolite-producing unicellular bacteria." Planta Medica 78(1 1 ): 1 135-1 136), since the inception of the modern antibiotic era, many inventive agar diffusion methods have been developed to detect and measure the antimicrobial activity of microorganisms. These methods are unified around the observation that when a compound with biological activity diffuses through an agar layer homogeneously seeded with a target organism, and after incubation and growth of the target organisms, a clear to diffuse zone of inhibition (ZOI) radiates from the point of compound application.

Agar diffusion assays remain popular because of their relative simplicity, low sample consumption and the capacity to test multiple compounds, concentrations, and mixtures from diverse sources in parallel against single or multiple microorganisms. To our knowledge, the principles of agar diffusion assays have not previously been applied in screening methods where plants are the target organism.

DESCRIPTION

Definitions

The following definitions apply throughout this document:

As used herein the term "compound" encompasses the pharmaceutically or agrochemically acceptable salts thereof.

"Growth" is used herein to refer to an increase, as in size, number, strength or expansion and includes growth measured directly (e.g. as increased size or optical density) or indirectly (e.g. using pH indicator dyes).

"Development" is used herein to refer to the act or process of growing, branching, progressing, or developing, including the progression from a simpler or lower to a more advanced, mature, or complex form or stage, i.e. the development of a plant from a seed.

As used herein the terms "development" and "growth" (which have different technical meanings) include each other.As used herein the term "plant" includes vascular and nonvascular plants, the term "root" includes rhizoid when applied to non-vascular plants and the term "shoot" includes the plant body when applied to non-vascular plants.

As used herein the term "seed" includes spores of non-vacular plants.

As used herein, "biological activity" refers to any process in the organism, other than growth, that can be detected from outside the organism (e.g. by the output of fluorescent or luminescent markers or by changes in pH of the medium).

All numerical values provided incorporate 10% less than and 10% more than the numerical value provided.

A novel micro-phenotyping technology has been developed, incorporating agar diffusion principles, which is applicable to the high-throughput screening of synthetic or natural compounds for a broad range of biological activities (including herbicides, growth regulators, stress protectants, antibiotics, fungicides etc.). The technology is applicable to both plants and microorganisms and, importantly, is amenable to automation. When used with plants the technology provides a high-content phenotypic screen that uniquely covers multiple traits of both roots and shoots while operating with low volumes of liquid (generally <0.5 ml). Because of their plasticity and phenotypic complexity, roots provide a particularly information-rich read-out for phytoactivity that is not fully exploited in other technologies. Using this technology, traits such as primary root growth, gravitropism, branching, root hair development and in vivo expression of fluorescent or luminescent markers can be visualised at high resolution and at multiple developmental stages and, with appropriate image analysis software, quantified. As currently used with Arabidopsis thaliana seedlings, the technology enables 96 compositions to be screened in the footprint of a single microtitre plate, and may be readily scaled up to 192 or more compositions in the same footprint. The technology has been validated by demonstrating its ability to distinguish the phenotypic effects of a series of plant hormones (auxin, cytokinin, gibberellic acid, ethylene etc). In a pilot experiment it has been used successfully to screen a library of over 3000 small molecules, leading to the identification of several novel compounds of scientific and potential commercial interest.

When used with microorganisms this technology may be suitably be used to detect or measure antimicrobial activity or may suitably be used for other assays for biological activity that depend for example on monitoring changes in the intensity of fluorescent or luminescent markers in the microbial cells.

A first aspect of the invention relates to an organism growth array device comprising a plurality of receptacles, each receptacle having a housing comprising first and second open ends, growth medium housed within the housing extending from a first surface of the growth medium, disposed longitudinally towards the first open end of the receptacle to a second surface of the growth medium disposed towards the second open end of the receptacle, wherein generally, in use, the first surface of the growth medium is disposed above the second surface of the growth medium;

wherein the open ends of each receptacle are individually accessible, preferably for the addition of different compositions; and wherein the growth medium of each receptacle is visible, and imagable across the lateral axis of the receptacle and along the longitudinal axis of the receptacle from the first surface to the second surface, and in use the growth of the organism within or into the growth medium is visible and imagable across the lateral axis of the receptacle and along the entire longitudinal length of the growth medium. If the organism is a plant, said first open end is generally an organism receiving end. The organism growth array device of the present invention allows any growth, or cultivation of the organism into or within the growth medium to be viewed from the side (across the lateral axis) of the receptacle. In this way, when the organism is a plant, seed is "sown" on the first surface and the entire growth structure of the organism into the growth medium can be viewed and imaged as it proliferates within the growth medium. This allows features of the growth structure to be viewed, imaged and analysed, including features of the growth structure disposed close to the first surface of the growth medium. Such features may include the degree of branching of the growth structure, the direction of growth and length of primary and/or secondary growth structures. The ability to view the entire growth, as well as the shoot, allows the development of the organism to be monitored effectively, allowing many different aspects of the development of the organism to be assessed and monitored. Abnormal development can be identified and the cause of this can be assessed. When the organism is a microorganism, the inoculum may be distributed evenly within the growth medium and growth of the organism within the growth medium from the first surface to the second surface is monitored. Changes in the development of an organism following contact with a test composition can also be identified and monitored effectively.

Each receptacle of the organism growth array device can be accessed individually, meaning that different test compositions may be added to each of the receptacles. Alternatively or additionally, the growth medium may comprise one or more test compositions. The development of organisms in each receptacle can be monitored and compared easily, meaning that test compositions which may have bioactive potential can be identified accurately. When the test compositions are applied to the second surface of the growth medium a longitudinal diffusion gradient within the receptacle is established, allowing the effects of a range of concentrations on the organism's development, or on biological activity within the organism, to be studied.

A further aspect of the present invention provides a plant growth array device comprising a plurality of receptacles, each receptacle having:

a) a housing extending from a first open ("seed receiving") end, preferably disposed substantially longitudinally from a second open end;

b) plant growth medium housed within the housing extending from a first surface of the plant growth medium disposed towards the first end of the receptacle to a second surface of the plant growth medium disposed towards the second end of the receptacle; wherein the second open end of each receptacle is individually accessible, preferably for the addition of different compositions; and wherein the plant growth medium of each receptacle is visible, and imagable across the lateral axis of the receptacle and along the longitudinal axis of the receptacle from the first surface to the second surface, and in use the root structure of a plant grown in the plant growth array device is visible and imagable across the lateral axis of the receptacle along the entire length of the plant growth medium.

A further aspect of the present invention relates to a method of imaging one or more organisms comprising the steps of:

a) culturing one or more organisms in the receptacle of the organism growth array device as described above, wherein a first aspect of the growth structure of the organism (generally the root) is within the growth medium (generally extending from the first surface of the growth medium into the growth medium), and a second aspect of the growth structure of the organism (generally the shoot) may extend onto the first surface of the growth medium and/or from the first surface of the growth medium away from the growth medium;

b) imaging the first aspect of the growth structure of the organism from the first surface of the growth medium towards the second surface of the growth medium;

c) optionally imaging the second aspect of the growth structure from the first surface of the growth medium away from the growth medium;

d) optionally imaging the second aspect of the growth structure from above.

Generally the method of imaging one or more organisms comprises the steps of:

a) where the organism is a plant, culturing one or more seedlings in the receptacle of the organism growth array device as described above, wherein the primary growth structure extends from the first surface of the growth medium into the growth medium, and the shoot extends above the first surface of the growth medium or where the organism is a microorganism, preferably filling the receptacle with growth medium containing a microbial inoculum;

b) imaging the first aspect of the growth structure of the organism from the first surface of the growth medium towards the second surface of the growth medium as well as the shoot above the surface of the growth medium, or imaging growth of the microbial inoculum along the longitudinal axis of the receptacle.

A further aspect of the invention relates to a method of analysing the development of one or more organisms comprising the steps of analysing one or more of: 1. the branch structure of the growth of the organism(s), in particular the first aspect of the growth structure of the organism (s) as described above;

2. the length of the longest primary member in the growth structure, in particular the first aspect of the growth structure of the organism (s) as described above;

3. gravitropism of the organism or growth structure;

4. phototropism of the organism or growth structure;

5. the morphology of the organism or growth structure (including root hairs);

6. biological activity of the organism or growth structure;

7. changes in the optical density of the growth medium.

A further aspect of the present invention relates to an assay for identifying a molecule or composition with bioactive potential comprising the steps of:

a) culturing one or more organisms in a first receptacle of the organism growth array device as described above;

b) contacting the growth medium of the first receptacle with a control composition;

c) analysing the development of the organism, suitably according to the method as described above;

d) growing one or more of the organism(s) in a second receptacle of the organism growth array device as described above;

e) contacting the growth medium of the second receptacle with a test composition comprising at least one molecule which may have bioactive potential;

f) analysing the development of the organism according to the method used in step c); g) comparing the development of the organisms in the first and second receptacles, where a difference in the development is indicative of the molecule of the test composition having bioactive potential.

According to a further aspect of the present invention, there is provided an assay for identifying a molecule or composition with bioactive potential comprising the steps of:

• providing a device as described herein, wherein the growth medium comprises a control composition;

• culturing one or more organisms in the first receptacle of the device;

• analysing the development of the organism, suitably according to the method as described herein;

• culturing one or more of the organism (s) in a second receptacle of the device as described herein, wherein the growth medium comprises a test composition, comprising one or more molecules which may have bioactive potential; • analysing the development of the organism(s) according to the method used in the third step of the method;

• comparing the development of the organisms in the first and second receptacles, where a difference in the development is indicative of a component of the test composition having bioactive potential.

DETAILED DESCRIPTION

Plant Growth Array Device

wherein the open ends of each receptacle are individually accessible, preferably for the addition of different compositions; and wherein the growth medium of each receptacle is visible, and imagable across the lateral axis of the receptacle and along the longitudinal axis of the receptacle from the first surface to the second surface, and in use the growth of the organism within or into the growth medium is visible and imagable across the lateral axis of the receptacle and along the entire longitudinal length of the growth medium.

According to a further aspect of the present invention there is provided an organism growth array device suitable for automated phenotypic analysis of either plants and microorganisms under aseptic conditions and comprising:

a plurality of receptacles, each receptacle having a housing comprising first and second open ends and one or more longitudinal sides extending from the first open end to the second open end, wherein at least one of the longitudinal sides is flat, and wherein a portion of each receptacle is fused to the neighbouring receptacle(s) along at least 10% of its length along the longitudinal axis of the receptacle;

growth medium housed within the housing extending from a first surface of the growth medium, disposed longitudinally towards the first open end of the receptacle to a second surface of the growth medium disposed towards the second open end of the receptacle, wherein generally, in use, the first surface of the growth medium is disposed above the second surface of the growth medium;

wherein the open ends of each receptacle are individually accessible, preferably for the addition of different compositions; and wherein the growth medium of each receptacle is visible, and imagable across the lateral axis of the receptacle and along the longitudinal axis of the receptacle from the first surface to the second surface, and in use the growth of the organism within or into the growth medium is visible and imagable across the lateral axis of the receptacle and along the longitudinal length of the growth medium.

According to one embodiment, more than one of the longitudinal sides of the receptacle are flat. Generally all longitudinal sides of the receptacle are flat.

There are two important advantages associated with such embodiments of the present invention over tubular receptacle arrangements. A curved surface causes optical problems that make image capture difficult. For tubular arrangements, the refractive index at the surface of the material forming the receptacle (generally plastic) may have to be modified, for instance through submersion of the tubes in water. In addition, a receptacle comprising one or more flat longitudinal side tends to promote, or force the growth of an organism grown therein to form a 2D structure rather than a 3D structure. This is particularly pronounced where the distance between opposite longitudinal sides of the receptacle is small relative to the root/growth structure at the first surface of the growth medium. For instance, the distance between opposite longitudinal sides of the receptacle may be the same, or up to 2 times greater than the diameter of the seed of the plant. Generally the distance between opposite longitudinal sides of the receptacle is from around 0.5 mm to around 5 mm, typically from around 1 mm to around 3 mm, suitably from 2 to 3 mm. According to one embodiment, the receptacle narrows along its longitudinal axis from the first end towards the second end additionally promoting 2D growth rather than 3D growth. It is more straightforward to extract information from images of primarily 2D root growth than from root growth which is primarily 3D. This makes it easier to extract information from the images about the growth and architecture of the organisms.

a) a housing extending from a first open ("seed receiving") end, preferably disposed substantially longitudinally from a second open end; b) plant growth medium housed within the housing extending from a first surface of the plant growth medium disposed towards the first end of the receptacle to a second surface of the plant growth medium disposed towards the second end of the receptacle;

wherein the second open end of each receptacle is individually accessible, preferably for the addition of different compositions; and wherein the plant growth medium of each receptacle is visible, and imagable across the lateral axis of the receptacle and along the longitudinal axis of the receptacle from the first surface to the second surface, and in use the root structure of a plant grown in the plant growth array device is visible and imagable across the lateral axis of the receptacle along the entire length of the plant growth medium.

According to a further aspect of the present invention there is provided a plant growth array device comprising a plurality of receptacles, each receptacle having:

a) a housing extending from a first open ("seed receiving") end, disposed substantially longitudinally from a second end, preferably a second open end and one or more longitudinal sides extending from the first open end to the second end, wherein at least one of the longitudinal sides is flat, and wherein a portion of each receptacle is fused to the neighbouring receptacle(s) along at least 10% of its length along the longitudinal axis of the receptacle;

b) plant growth medium housed within the housing extending from a first surface of the plant growth medium disposed towards the first end of the receptacle to a second surface of the plant growth medium disposed towards the second end of the receptacle;

According to one embodiment, the organism is a plant. In such embodiments, the seed of the plant is generally placed on the first surface of the growth medium. The root structure of the plant grows into the growth medium towards the second surface. The shoot portion of the plant grows away from the growth medium. The root structure provides important information about the development of the plant, and changes to the development of the growth structures of the plant provide an indication of the effects of test compounds thereon. As the device of the present invention allows the root structure to be viewed along the entire longitudinal axis of the receptacle, the majority of the root structure is visible. The device thus provides a substantially complete and accurate indication of the development of the plant, and any changes thereto.

The organism growth array device of the present invention allows the entire growth structure of the organism to be viewed, imaged and analysed. This includes the growth structure proximate to the first surface of the growth medium. The entire growth structure of an organism provides important information about the developmental impact of potentially bioactive compounds.

Prior art systems fail to provide easy access and imaging of the entire growth structure, generally concentrating on the root tip morphology. Important information is not collected or analysed using such prior art systems, meaning that a less effective method of assessing the development of an organism is provided. In particular, in such prior art systems any developmental abnormalities which occur to parts of the growth structure other than the tip of the primary root would not be observed.

Generally the organism is a plant, and the root structure of the plant is visible and imagable along the entire length of its structure. Where the organism is a plant, the first aspect of the growth structure is the root structure and the entire root structure may be easily viewed, imaged and analysed.

Alternatively the organism may be a microorganism for example a fungus, bacterium, planktonic microorganism or persister cells (dormant cells) such as those found in biofilms. In such embodiments, the organism will generally be mixed with the growth medium prior to the growth medium being housed in the receptacle. Generally, for such embodiments, the organism is substantially evenly distributed within the growth medium in the receptacle. As such, the growth of such organisms is primarily within the growth medium. The growth array device of the present invention allows growth of the organism along the entire length of the receptacle to be visible and imagable. The growth of the microorganism provides important information about the effects of test compounds on the development of the microorganism. The growth of the organism in such embodiments will primarily be within the growth medium, and the device of the present invention allows the growth to be viewed, imaged and analysed, thus providing a substantially complete and accurate indication of the development of the microorganism, and any changes thereto. The entire growth structure of an organism grown in the organism growth array device can be viewed and imaged. Particular attention may be paid to a first aspect of the growth structure of the organism within the growth medium or extending from the first surface of the growth medium into the growth medium. In general the entire first aspect of the growth structure is viewed, imaged and analysed. The growth structure of the organism, (in particular the first aspect of the growth structure) can be viewed and imaged from the side of the organism growth array device, across the lateral axis and along the longitudinal axis of the receptacle. Various and many aspects of the growth structure can be analysed and easily compared.

Generally the housing of the receptacle is transparent or semi-transparent to allow the growth medium to be viewed and imaged through the housing. Typically the housing exhibits a light transmissibility of at least 80%, suitably at least 90%, more suitably 95 to 97%.

According to one embodiment, the housing of the receptacle comprises at least one flat portion along its longitudinal axis. Typically the receptacle comprises at least one flat side. This promotes ease of imaging of the growth structure in the growth medium.

Typically the receptacle has an oval, square, rectangular, semi-circular or circular cross- section. Suitably the cross-section of the receptacle along the lateral axis is square or rectangular.

According to one embodiment, the receptacle narrows along the longitudinal axis. This promotes ease of imaging of the entire growth structure in the growth medium, minimising the visually obstructing effect of growth towards the outer edges of the growth medium.

According to one embodiment, each receptacle is of the same or similar shape and dimensions to allow easy and accurate comparison of organism development in different receptacles. Generally the shape and dimensions of the receptacles in the device of the present invention differ by less than 50%, typically less than 2%, suitably less than 1%, more suitably less than 0.1%.

Likewise, each receptacle in the device generally comprises the same growth medium, disposed at the same distances from ends of the receptacle to promote ease and accuracy of the comparison of the organism development in different receptacles. Generally the first surface of the growth medium is 5 mm or less from the first end of the receptacle, typically 2 mm or less, suitably 1 mm or less. According to one embodiment, the first surface of the growth medium is 0.1 to 0.5 mm from the top of the receptacle, or the first open end of the receptacfe.

Generally the second surface of the growth medium is 1 mm or less from the second open end of the receptacle. Typically the growth medium extends to open end of the receptacle.

Suitably the growth medium does not comprise any significant air bubbles which may distort the development of the organism.

According to one embodiment, the growth medium may comprise one or more test compounds or test compositions.

According to one embodiment, the dimensions of the receptacles ensure that the device of the present invention may be housed within the wells of a microtttre plate, wherein portions of the device of the present invention between the receptacles rest on the walls of the wells of a microtitre plate, supporting the receptacles and ensuring that the open ends of the receptacles do not rest on the base of the wells.

Typically the width/diameter of the receptacle at the first end is 5 to 15 mm, generally 7 to 10 mm, suitably 8 to 9 mm. Typically the width/diameter of the receptacle at the second open end is 1 to 5 mm, generally 1 to 3 mm, suitably 1.5 to 2 mm. Typically the length of the receptacle is 10 to 50 mm, generally 15 to 30 mm, suitably 20 to 25 mm.

According to one embodiment, the receptacle has a rectangular or oval cross-section. In such embodiments the greater width/diameter at the first end is typically 5 to 15 mm, generally 7 to 10 mm, suitably 8 to 9 mm. Generally, the smaller width/diameter at the second end is 1 to 5 mm, generally 2 to 4 mm, typically around 2 mm.

According to one embodiment the volume of the receptacle is 50 to 500 μΙ, generally 200 to 400 μΙ, suitably 250 to 350 μΙ, more suitably around 300 μΙ.

The growth medium of each receptacle is visible, and imagable across the lateral axis of the receptacle from the first surface to the second surface, and in use the growth of the organism into the growth medium is visible and imagable across the lateral axis of the receptacle along the entire length of the growth medium. Accordingly, the device of the present invention comprises one or two rows of receptacles.

According to one embodiment, the device comprises one row of 6 to 24 receptacles. According to a further embodiment, the device comprises two rows of 6 to 24 receptacles.

According to one embodiment, the receptacle has a rectangular or oval cross-section. In such embodiments the greater width/diameter at the first end is typically 6 to 30 mm, generally 7 to 20 mm, suitably around 7 mm. Generally, the smaller width/diameter at the second end is 5 to 18 mm, generally 5 to 14 mm, suitably around 6 mm.

According to one embodiment the volume of the receptacle is 100 to 1000 μΙ, generally 300 to 800 μΙ.

Typically the device comprises one or two rows of 6 to 12 receptacles, generally 6 or 8 receptacles.

The device may comprise a portion between the receptacles to provide increased rigidity and stability of the device. According to one embodiment, a portion of each receptacle is joined, typically fused to the neighbouring receptacle(s), typically along 30 to 50 % of its length along the longitudinal axis of the receptacle.

Alternatively, the receptacles may be individually detachable from the device.

The device may be moulded from a single polymeric material, suitably a plastic material such as polypropylene, or may comprise the receptacles and a separate support portion formed from a material having a high modulus, providing rigidity, typically a polymeric material, suitably a plastic material such as high density polyethylene. Generally the receptacles are formed from a transparent material, typically a polymeric material, suitably a plastic material such as polypropylene The device may include a handle portion to increase ease of handling of the device. The growth medium is generally transparent or translucent. According to one embodiment, the array device may be imaged at, for instance, UV frequencies. Typical growth medium includes compositions comprising one or more of guar gum, xantham gum, gum arabic, psyllium husk powder, chia seed powder, gellan gum, Phytagel^®, agar and petrifilm. Particular mention may be made of gellan gum. The growth medium may comprise organism nutrients. According to one embodiment, growth of the organism does not extend to the second open end of the receptacle. This provides the advantage that the test composition may be applied to the growth medium at the second open end without risk of directly contacting the organism. Typically, there is a distance of at least 1 mm between the ends of the growth structure and the second open end.

Alternatively the growth of the organism may extend past the second open end of the receptacle.

Method of Imaging

The present invention relates to a method of imaging one or more organisms comprising the steps of:

c) optionally imaging the second aspect of the growth structure from the first surface of the growth medium away from the growth medium.

According to one embodiment, more than one organism (generally a seedling) is grown in each receptacle, generally wherein each organism is of the same species. This means that an average developmental response can be generated within each receptacle. Alternatively more than one organism of different species may be grown in each receptacle to provide a developmental response for more than one species of organism.

Generally the first aspect of the growth structure is imaged using a digital still or video camera, generally with a macro lens, or a using a microscope fitted with a digital or video camera.

The images can be assessed by the human eye. Alternatively, the images can be imported into image analysis computer systems allowing quantitative data on traits such as growth, development, morphology of the organism, or the outputs of in vivo fluorescent or luminescent markers to be generated.

According to one embodiment, the entire first aspect of the growth structure is imaged, that is any growth within the growth medium is imaged. Additionally one or more of the following traits may be imaged:

• the portion of the first aspect of the growth structure proximate to the first surface of the growth medium (for instance, within a longitudinal distance of 1 to 5 mm of the first surface of the growth medium), which in one embodiment develops before application of the test compound (s);

• the morphology of the tip of the primary growth of the first aspect of the growth structure;

• the primary growth of the first aspect of the growth structure along its entire length;

• the second aspect of the growth structure.

According to one embodiment all of the traits described above may be imaged. Generally all of the traits will be imaged across the lateral axis of the receptacle and along the longitudinal axis. The traits may be imaged from both sides along the lateral axis of the receptacle. Additionally, images may also be taken from above and below the receptacle, along the longitudinal axis of the receptacle.

Generally, the growth of the organism(s) in each receptacle is imaged in the same manner, using the same angles and distances for imaging and the same levels of magnification in order to promote the ease of comparison of the images.

Typically the growth of the organism(s) is imaged without removal of the organism (s) from the device of the present invention.

Generally more than one image is made of the growth structure within or extending from each receptacle. Each image may be of a different magnification depending on the subject of the image.

Where the organism is a microorganisms, it will typically be unicellular or hyphal and will not generally have a visible growth structure. Changes in the growth and development of the microorganism may typically be imaged and analysed by assessing changes in optical density or using fluorescent markers. Method of Analysis

The present invention relates to a method of analysing the development of one or more organisms comprising the steps of analysing one or more of:

1. the branch structure of the growth of the organism(s), in particular the first aspect of the growth structure of the organism(s) as described above;

2. the length of the longest primary member in the growth structure, in particular the first aspect of the growth structure of the organism(s) as described above;

3. gravitropism of the growth structure;

4. changes in optical density of the growth medium.

Where the organism is a plant, the method typically comprises the steps of analysing one or more of:

a. the branch structure of the growth of the organism (s), in particular the root structure of the plant;

b. the length of the longest root in the root structure;

c. gravitropism of the root structure;

d. length of the shoots.

According to one embodiment, the method comprises the steps of analysing all of traits a. to d.

The method may include the step of analysing additional traits. Particular mention may be made of

• the portion of the first aspect of the growth structure proximate to the first surface of the growth medium (for instance, within a longitudinal distance of 1 to 5 mm of the first surface of the growth medium);

• the tip of the primary growth of the first aspect of the growth structure (generally the tip of the longest root), in particular the morphology of the tip of the primary growth (distortion/swelling etc.);

• number, density and dimensions of hairs on the growth structure, in particular of the first aspect of the growth structure;

« direction of growth of the first and/or second aspect of the growth structure;

o the second aspect of the growth structure, in particular shape, size and colour of the second aspect of the growth structure (generally shoots);

• hypocotyl elongation and/or phototropism. The method of analysis may include the step of analysing one or more of the features described above. According to one embodiment, the method of analysis includes the step of analysing all of the features described above. According to another embodiment, any or all of these features may be imaged at intervals during the growth period. These intervals may be from 4 to 96 hours, more suitably 24 to 48 hours.

Generally the method of analysis includes the step of analysing all of the images produced according to the method of imaging an organism as described above.

Where the organism is a plant, the method may include the step of analysing the number and colour of the leaves, the length of the stem and the direction of stem and/or leaf growth.

According to another embodiment, the organism may be a microorganism such as a bacterium or fungus (such as a yeast). In such embodiments, the growth medium composition may initially comprise the organism and generally the organism may be substantially evenly distributed throughout the growth medium. The growth of the organism may be substantially or completely within the growth medium. In such embodiments growth of the organism, generally within a diffusion gradient of the test compound emanating from the second open end of the receptacle, is typically determined by analysis of the rate of increase in optical density within the growth medium measured along the longitudinal axis of the receptacle.

According to one embodiment, fluorescent and/or luminescent markers or dyes may be introduced into the organism, and used to monitor development of the organism, or biological activity within the organism, generally by using appropriate excitation/emission wavelengths. There may also be within one receptacle more than organism or strain of organism carrying different fluorescent and/or luminescent markers. Mention may be made of green fluorescent protein, luciferase and histochemical dyes.

Assay

The present invention also relates to an assay for identifying a molecule or composition with bioactive potential comprising the steps of:

b) contacting the growth medium of the first receptacle with a control composition; c) analysing the development of the organism, suitably according to the method as described above; d) culturing one or more of the organism(s) in a second receptacle of the organism growth array device as described above;

According to a further aspect of the present invention there is provided an assay for identifying a molecule or composition with bioactive potential comprising the steps of:

1. providing the organism growth array device as described above, wherein the growth medium comprises a control composition;

2. culturing one or more organisms in the first receptacle of the device;

3. analysing the development of the organism, suitably according to the method as described above;

4. culturing one or more of the organism(s) in a second receptacle of the device as described herein, wherein the growth medium comprises a test composition, comprising one or more molecule(s) which may have bioactive potential;

5. analysing the development of the organism(s) according to the method used in step 4;

6. comparing the development of the organisms in the first and second receptacles, where a difference in the development is indicative of a component of the test composition having bioactive potential.

Each receptacle of the organism growth array device of the present invention is individually accessible meaning that a different test composition may be applied to each receptacle. This enables the effects of a high number of test molecules or compositions to be considered in a short time frame. The ability to automate the process using robotic devices thus provides a high-throughput method of screening for compounds which affect the development of the organism, and which may have bioactivity.

Each receptacle comprises a second open end. The second surface of the growth medium can be accessed at or through the second open end of the receptacle. Different compositions can be applied to the growth medium at different time points without the need for removal of the growth medium from the receptacle, and without the need for removal of the receptacle from the plant growth array device. During application of the test composition to the growth medium, the development of the organism is not affected by movement from the growth array device or movement of the receptacle. Accordingly, changes in the development of the organism can be attributed to the effects of the test composition, rather than for example the effects of changes in light or temperature through such movement.

Accessing the growth medium at or through the second open end of the receptacle means that the second aspect of the growth structure of the organism (the shoot portion where the organism is a plant) remains undisturbed. There is no need to disturb the second aspect of the growth structure to access the growth medium. The risk of inadvertently contacting the second aspect of the growth structure with the test composition is minimised accordingly. Additionally, as noted above, the first aspect of the growth structure does not generally extend to the second surface of the growth medium. Accordingly, the test/control composition may be applied to the growth medium at or through the second open end of the receptacle without disturbing the first aspect of the growth structure.

Generally the test/control composition is applied to the growth medium and is not applied directly to any portion of the organism, including the first and second aspects of the growth structure (root and shoot portions respectively where the organism is a plant).

In contrast to applying the test composition directly to the growth structure of the organism, applying the test composition to the growth medium at the second open end of the receptacle generally results in a diffusion gradient, where the concentration of the test composition in the growth medium varies along the longitudinal axis of the growth medium. The concentration of the test composition in the growth medium generally varies due to diffusion of the test composition in the growth medium. High concentrations of the test composition may be provided at or near the second open end of the receptacle, with the concentration decreasing with increasing distance from the second open end. The effects of the test composition on organism growth can be assessed at different concentrations by assessing the development of the growth structure (i.e. plant growth structure or microbial culture) at different longitudinal distances from the second surface of the growth medium. In particular, analysis of the first aspect of the growth structure (root structure) provides an indication of the effect of a test composition. For instance, growth of the organism proximate to the first surface may be observed due to low concentration of the test compound, but limited or no growth may be observed closer to the second surface of the growth medium due to higher concentrations of the test compound. In addition this diffusion gradient can allow comparison of growth traits before and after application of the test composition, where indications of growth traits before application may be provided at larger longitudinal distances from the second surface of the growth medium and indications of growth traits after application may be provided at smaller longitudinal distances from the second surface of the growth medium.

These attributes allow a greater range of tests to be run, including testing a bank of chemicals firstly for their effects on growth, but also secondly on their effects on growth when chemicals or cocktails are present in the growth medium (i.e. remediation properties).

According to one embodiment, the assay may comprise culturing one or more of the organisms in a third and optionally subsequent receptacle(s) of the device and contacting the growth medium of the third receptacle, and optionally subsequent receptacles with different test compositions (where different test compositions may vary in terms of the components therein and the concentration of the components).

In addition, the diffusion gradient of the test composition along the longitudinal axis of the receptacle provides an indication of how potent the test composition is, or how rapidly it diffuses through the growth medium from the second surface.

According to one embodiment, the growth medium comprises one or more test compounds. The test compound(s) may be substantially evenly distributed throughout the growth medium or the test compound(s) may be provided at different concentrations in the growth medium, typically through a diffusion gradient.

Alternatively or additionally, one or more further test compounds may be added to the growth medium at the second open end of the receptacle. This provides an indication of how the test compounds interact, and how this interaction affects the development of the organism. This is of particular utility where one or more of the test compounds may provide remediation of the effects of one or more of the other test compounds.

In particular the growth medium may comprise a first test compound which may affect the development of the organism, and a second test compound may be added to the growth medium at the second open end of the receptacle to assess whether the second compound provides remediation of the effects of the first compound (or vice versa).

Alternatively or additionally, the first test compound may be added to the growth medium at the second open end of the receptacle and after an interval, the second test compound may be added to the growth medium at the second open end of the receptacle (or vice versa). Most suitably the second test compound is applied 24-48 hours after application of the first test compound.

Additionally or alternatively the growth medium may comprise one or more test compounds or one or more test compounds may be added to the growth medium at the second open end of the receptacle.

The identity and structure of the test compounds may be known and they may be synthetic or natural compounds. Alternatively, the exact identity and structure of the test compounds may be unknown, for example the test composition may comprise an unrefined extract from a plant, an extract from an environmental soil or water sample or a live culture of a microorganism or mixture of microorganisms.

The test composition may comprise more than one test compound.

According to one embodiment the test/control composition includes nutrients for organism growth that may otherwise become limited in the relatively small volume of growth medium. The water in which the test compounds are dissolved also provides replacement for water lost from the growth medium through evaporation and transpiration.

Generally the test composition is the same as the control composition with the addition of the test compound(s). Typically the control/test composition may comprise one or more pharmaceutical or agrochemical excipients or carriers such as solvents, anti-adherants, binders, fillers, diluents, lubricants and preservatives.

Suitably the organisms compared are as similar as possible, for instance in terms of identity, maturity and size.

Typically the first and second receptacles are as similar as possible, as described above, in terms of, for instance, size, dimensions, amount and identity of growth medium etc.

According to one embodiment, different test compositions may be applied to the growth medium at different time periods. Test compositions may differ in terms of the test compound(s) comprised in the test composition and the concentration of the test compound (s) in the test composition. According to one embodiment, test compositions may be applied to the growth medium at different stages during the growth of the organism. This ensures that the test composition does not have to be present in the growth medium upon initial growth of the organism.

Where the organism is a plant, the assay of the present invention ensures that the test composition does not have to be present in the growth medium during germination, which is advantageous as some test compositions may be detrimental to germination.

According to one embodiment, two different test compositions may be applied sequentially, for instance a first test composition may be applied to the growth medium prior to the application of a second test composition. The first test composition may, for instance, be thought to protect organism development from the effects of the second test composition. Allowing a delay between application of the first and second test compositions may provide the first test composition with sufficient time to establish and exert its antagonistic effect to the second test composition.

Typically several organisms may be cultured in each receptacle. The effect of the test composition and the associated developmental response of several different organisms can be assessed with one application providing an indication in the developmental response of several plants with one application of the test composition.

The organisms may be cultured in the receptacles by the application of the organism to the first surface of the growth medium. The organism may be applied as a seed where the organism is a plant. Typically a seed suspension is formed comprising seeds suspended in water or a growth medium.

Where the organism is a plant, generally 1 to 20 seeds and suitably 6 to 10 seeds are applied to the first surface of the growth medium of each receptacle via the first open ("seed receiving") end.

Alternatively the organism may be applied as a composition mixed with the growth medium, for instance a composition comprising one or more microorganisms, typically one or more fungal and/or bacterial cultures. The identity of the cultures contained in the sample is generally known prior to application. According to one embodiment, the composition may comprise more than one fungal culture, typically two or three fungal cultures. According to a further embodiment, the composition may comprise more than one bacterial culture, typically two or three bacterial cultures. The microbial composition comprises at least 1 x 10² microbial colony forming units per ml, typically at least 1 x 10³, suitably at least 1 x 10⁴, more suitably 5 x 10⁴ microbial cfu per ml.

The organism may be applied to the first growth medium surface using any suitable technique, for example by pipetting, painting, spreading or spraying.

The method of the present invention includes the step of incubating the organism for an incubation period following application of the organism. The incubation period is typically from 5 to 16 days, suitably 12 days or less, more suitably up to 10 days. If the organism is a plant, a pre-incubation period of 1 to 4 days at 3-4 degrees Celcius is generally included to improve germination. The receptacle may be incubated at controlled temperatures following application of the organism. If the organism is a plant the controlled temperature may be 18 to 30 degrees Celsius, suitably 21 -25 degrees Celsius. For microorganisms, the controlled temperatures may be 25-40 degrees Celsius, suitably 30 to 37 degrees Celsius depending on the strain. Typically the incubation period is conducted under elevated humidity levels. Suitably the humidity levels are greater than 40%, typically greater than 50%. The incubation period may last until the organism has grown substantially. Where the organism is a plant, the incubation period may last until the longest root portion of the plant has grown 5 to 25 mm into the growth medium. Where the organism is a microorganism, in particular a fungus and/or a bacterium, the incubation period may last until the fungal/bacterial isolate has grown substantially, sufficient to allow measurement of significant changes in optical density. According to one embodiment, a difference in the properties tested of 10% or more is indicative of the molecule of the test composition having bioactive potential, generally 20% or more, suitably 30 % or more.

According to one embodiment, a difference in two or more of the properties tested is indicative of the molecule of the test composition having bioactive potential.

Typically steps a) to g) above may be repeated 5 to 10 times.

Generally the growth traits are assessed daily, typically every two days.

Suitably where the molecule of the test composition may have bioactive potential, steps a) to g) above may be repeated using test compositions having different concentrations of the molecule. The volume of test/control composition added to the growth medium of each receptacle is generally 40-60% of the volume of the growth medium in the receptacle, most suitably 50% of the volume.

Test compositions having bioactive potential may increase or decrease the development of the organism or may increase or decrease the intensity in vivo of fluorescence or luminescence markers.

Method of Manufacture

According to the present invention, there is provided a method of manufacturing the organism growth array device as described above comprising the steps of:

• providing a device having a plurality of receptacles each receptacle having a housing comprising a first open end and a second open end longitudinally disposed from the first open end, wherein a removable closure member is provided over the second open end;

• introducing growth medium into the housings via the first open end;

• allowing the growth medium to solidify;

• removing the removable closure members.

According to the present invention, there is provided an alternative method of manufacturing the organism growth array device as described above comprising the steps of:

• providing a device having a plurality of receptacles each receptacle having a housing extending from a first open end to a closed end longitudinally disposed from the first end;

• introducing growth medium into the housings via the first open end;

• allowing the growth medium to solidify;

• removing the closed end of the housings, typically through cutting, so as to leave a second open end.

The method may include the step of mixing the growth medium with the organism prior to introducing the growth medium into the housings, in particular where the organism is a microorganism.

As noted above, the growth medium may comprise one or more test composition(s), a control composition and/or nutrients to promote the growth and development of the organism. The present invention will now be described by way of example only with reference to the accompanying Figures in which:

Figure 1 shows a schematic diagram of a preferred organism growth array device of the present invention;

Examples

Description of the micro-phenotyping method

All operations must be performed under sterile conditions

1. Preparing the agar tubes. 300 μΐ molten 0.8% agar medium (containing nutrients for plant growth) is pipetted into each of an array of 96 tubes per plate, made of 12 strips of 8 tubes (FrameStrips™ 4ti.co.uk). The tubes are supported in a plastic box with 96 holes, spaced appropriately. At this point the tubes have a first open end and the second end is closed. Note that the agar is kept at ~50°C on a hot plate to stop it solidifying and is injected into the bottom of the tubes to prevent trapping of air bubbles.

2. Sowing the seed. Seed of Arabidopsis thaliana is sterilised in dilute bleach, rinsed thoroughly and suspended in 0.2% agar containing 20 μΜ ammonium nitrate. The seed suspension is pipetted onto the surface of the solidified agar so that there are approximately 8-10 seed per tube. If necessary to break seed dormancy, tubes can be stored at 4°C for 2 days at this stage. For larger-seeded species the tubes are wider and longer and spaced appropriately for (say) 48-well or 24-well plates, with 8 strips of 6 tubes or 6 strips of 4 tubes per plate, respectively.

3. Growth of the seedlings. The boxes of tubes are placed in the growth room, inside small clear plastic boxes to maintain sterility. These boxes are in turn placed within a standard horticultural propagator with a bed of well-moistened absorbent paper to maintain humidity. Temperature (for Arabidopsis) is 22°C and day length is 16 h. After 2 days, the second ends of the tubes are excised using a guillotine to allow additional water and nutrients to be supplied through the bottom of the tube - the excised tubes are transferred to 96-well microtitre plates with V-shaped wells containing a reservoir of 150 μΙ nutrient medium.

4. Chemical treatments: these are applied by adding a small volume of the chemical to the reservoir of nutrient medium in the microtitre plate, which then diffuses rapidly upwards through the agar.

5. Measurements:

Root growth The length of the primary root is measured, imaged and analysed. The length of the primary root may be monitored manually by observing the tubes from the side with a binocular x3.5 lens and marking on the tube with a felt-tip the position of the tip of the longest primary root in each tube. This can be done daily or less frequently, depending on the requirements of the experiment. At the end of the experiment the strips are placed on a document scanner and scanned, the images then being imported into image analysis software to provide reproducible data on root growth rates. Note that this process may be readily automated by an imaging system that records changes in length of the longest root with time as required without the need to mark the tubes.

Hypocotylelongation/phototropism. Hypocotyl elongation may be monitored automatically from the same images used to monitor root growth by determining maximum height of seedlings. If illumination is directional (say 45°) during the period of growth, then the hypocotyl angle may be used to quantify phototropism.

Other shoot traits: If required, leaf size, shape and colour are monitored by additional imaging from above, or by turning the strips through 90°.

Root gravitropism and root branching: The same images may be analysed to detect effects on root gravitropism and root branching. The tube strips are conveniently photographed in blocks of four with a 60 mm macro lens on a standard digital SLR camera. Root hairs: Using high resolution images or higher magnification (x10-20), it is possible to visualise root hairs and determine their length and density. This is currently done by viewing the strips under a binocular microscope.

Root tip morphology: The same higher power images can reveal changes in root tip morphology (e.g. distortion, swelling).

Fluorescent or luminescent markers: Using appropriate excitation/emission wavelengths It is possible to detect differences in the spatial and temporal expression of fluorescent markers (e.g. green fluorescent protein) or luminescent markers (e.g. luciferase) or differences in staining with fluorescent dyes or histochemical dyes.

Improvements and modifications may be incorporated herein without deviating from the scope of the invention.

Claims

1. An organism growth array device suitable for automated phenotypic analysis of either plants and microorganisms under aseptic conditions and comprising:

2. The device of claim 1 wherein the receptacle is transparent or semi-transparent to allow the growth medium to be viewed and imaged through the housing.

3. The device of either one of claims 1 and 2 wherein the receptacle narrows along its longitudinal axis.

4. The device of any one of claims 1 to 3 wherein each receptacle is of the same or similar shape and dimensions.

5. The device of any one of claims 1 to 4 wherein the growth medium extends to the second open end of the receptacle, or to within 0.01 to 0.5 cm from the second open end of the receptacle.

6. The device of any one of claims 1 to 5 wherein the growth medium comprises one or more test compounds or test compositions.

7. The device of any one of claims 1 to 6 comprising one or two rows of receptacles.

8. The device of any one of claims 1 to 7 wherein each receptacle is fused to the neighbouring receptacle(s) along 30 to 50 % of its length along the longitudinal axis of the receptacle.

9. A method of imaging one or more organisms comprising the steps of:

culturing one or more organisms in the receptacle of the device as claimed in any one of claims 1 to 8, wherein a first aspect of the growth structure of the organism is within the growth medium, and a second aspect of the growth structure of the organism may extend onto the first surface of the growth medium and/or from the first surface of the growth medium away from the growth medium;

imaging the first aspect of the growth structure of the organism from the first surface of the growth medium towards the second surface of the growth medium.

10. The method of imaging as claimed in claim 9 wherein more than one organism is cultured in each receptacle.

11. The method as claimed in either one of claims 9 and 10 wherein the first aspect of the growth structure is imaged using a digital still or video camera, generally with a macro lens, or a using a microscope fitted with a digital or video camera.

12. The method as claimed in any one of claims 9 to 11 wherein all growth within the growth medium is imaged.

13. The method as claimed in any one of claims 9 to 12 wherein one or more of the following traits are imaged:

• the morphology of the tip of the primary growth of the first aspect of the growth structure; • the primary growth of the first aspect of the growth structure along its entire length;

• the second aspect of the growth structure.

14. A method of analysing the development of one or more organisms comprising the steps of analysing one or more of:

a. the branch structure of the growth of the organism(s), in particular the first aspect of the growth structure of the organism (s);

b. the length of the longest primary member in the growth structure;

c. gravitropism of the growth structure

d. changes in the optical density of the growth medium.

15. The method of claim 14 wherein all of a. to c. are analysed.

16. The method of either one of claims 14 and 15 wherein one or more of the following traits are analysed:

• the tip of the primary growth of the first aspect of the growth structure, in particular the morphology of the tip of the primary growth (distortion/swelling etc);

• direction of growth of the first and/or second aspect of the growth structure;

• the second aspect of the growth structure, in particular shape, size and colour of the second aspect of the growth structure;

• hypocotyl elongation and/or phototropism.

17. An assay for identifying a molecule or composition with bioactive potential comprising the steps of:

i. culturing one or more organisms in a first receptacle of the device as claimed in any one of claims 1 to 8; ii. contacting the growth medium of the first receptacle with a control composition;

iii. analysing the development of the organism, suitably according to the method as claimed in any one of claims 14 to 16;

iv. culturing one or more of the organism(s) in a second receptacle of the organism growth array device as described above;

v. contacting the growth medium of the second receptacle with a test composition comprising at least one molecule which may have bioactive potential;

vi. analysing the development of the organism according to the method used in step iii.;

vii. comparing the development of the organisms in the first and second receptacles, where a difference in the development is indicative of the molecule of the test composition having bioactive potential.

18. An assay for identifying a molecule or composition with bioactive potential comprising the steps of:

1) providing a device as claimed in claim 1 , wherein the growth medium comprises a control composition;

2) culturing one or more organisms in the first receptacle of the device;

3) analysing the development of the organism, suitably according to the method as claimed in any one of claims 14 to 16;

4) culturing one or more of the organism (s) in a second receptacle of the device as claimed in claim 1 , wherein the growth medium comprises a test composition, comprising a molecule which may have bioactive potential;

5) analysing the development of the organism according to the method used in step 4);

6) comparing the development of the organisms in the first and second receptacles, where a difference in the development is indicative of the molecule of the test composition having bioactive potential.

19. The assay of either one of claims 17 and 18 comprising the steps of culturing one or more of the organisms in a third and optionally subsequent receptacles of the device; contacting the growth medium of the third receptacle, and optionally subsequent receptacles with different test compositions (where different test compositions may vary in terms of the components therein and the concentration of components); and analysing the development of the organism in the third receptacle, and optionally subsequent receptacles and comparing the development of the organisms in the first, second, third and optionally subsequent receptacles.

20. The assay of any one of claims 17 to 19 wherein the second surface of the growth medium at or towards the second open end of the or each receptacle is contacted with a or the test composition and a diffusion medium of the or each test composition is formed within the growth medium.

21. The assay of any one of claims 17 to 19 wherein the growth medium comprises a first test composition and a second test composition is applied to the growth medium, generally at the second surface of the growth medium at the second open end of the receptacle.

22. A plant growth array device comprising a plurality of receptacles, each receptacle having:

23. A microorganism growth array device comprising a plurality of receptacles, each receptacle having: a) a housing extending from a first open end, preferably disposed substantially longitudinally from a second open end, and one or more longitudinal sides extending from the first open end to the second open end, wherein at least one of the longitudinal sides is flat, and wherein a portion of each receptacle is fused to the neighbouring receptacle(s) along at least 10% of its length along the longitudinal axis of the receptacle;

b) microbial growth medium housed within the housing extending from a first surface of the microbial growth medium disposed towards the first end of the receptacle to a second surface of the microbial growth medium disposed towards the second end of the receptacle;

wherein the second open end of each receptacle is individually accessible, preferably for the addition of different compositions; and wherein the microbial growth medium of each receptacle is visible, and imagable across the lateral axis of the receptacle and along the longitudinal axis of the receptacle from the first surface to the second surface, and in use the growth of the microorganism within the array device is visible and imagable across the lateral axis of the receptacle along the entire length of the microbial growth medium.