FACILE IN VITRO GLASS SLIDE LIGHT MICROSCOPY METHOD USING TETRACYCLINE TO VISUALIZE A REPETITIVE PATTERN IN AERIAL PLANT ROOT TIPS FILAMENTOUS NETWORK

© 2020 The Author(s). This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. 155 FACILE IN VITRO GLASS SLIDE LIGHT MICROSCOPY METHOD USING TETRACYCLINE TO VISUALIZE A REPETITIVE PATTERN IN AERIAL PLANT ROOT TIPS FILAMENTOUS NETWORK


INTRODUCTION
The deleterious effect of Tetracycline on the human hair follicle soft tissue was recently demonstrated in vitro [1]. In that paper, in toto human hairs were immersed in drops of diluted powder of a commonly used antibiotic, namely Tetracycline (TE). The adhesive property of TE was proposed as one factor in damaging the human follicle's metabolism, as well as a myriad of soft tissue cells. Images of the follicle's keratin architecture were displayed ( Fig  1).
The present manuscript introduces a deleterious effect of liquid TE to aerial plant root tip tissue. Fresh harvested orchid hanging aerial root tips were exposed to liquid TE by via a previously describe technique dubbed a Single Slide Preparation (SSP) [2]. The images obtained show a damaging effect of TE on external root epidermal and internal trichoblats cells, also documented are detailed images of a continuous filamentous veil seen covering the interior tissue. The outer epidermal cells failed to exhibit a filamentous covering network.

Methods
Tetracycline capsules (500 mg) were pierced at one end and the golden yellow powder transferred to a clean white glass plate. The power was divided by a double edge razor blade into 10 piles each estimated to contain 100 mg/pile. The subdivision progressed until piles of estimated 5 mgs each were reached (Exhibit I below).

THE LIQUID TETRACYCLINE
On a separate glass slide approximate a very small amount of TE powder, such as the minimal amount adhering to a toothpick was placed on the slide. Using a medicine dropper three small demineralized drinking bottle water drops were delivered and the powder stirred for a few seconds. For the purpose of the experiments, this aqueous solution was dubbed "liquid Tetracycline" (LT).

THE ROOT TISSUE IMMERSION IN LIQUID TETRACYCLINE
Freshly harvested orchids areal root tips were placed on a glass slide. A sharp blade was used to perform a longitudinal thin cut of the distal end as shown in Figure 1 below. Using small tweezers, the previously thin distal aerial root cut sample was placed in the LT. The LT drops were gently dispersed on the slide, this in order to stabilize root tissue drift towards the drop's edges. Once stabilized, the preparation was placed on the optical video-microscope platform, and allowed to dry. Control images and video recordings after LT evaporation recorded and saved for downloading into a McBrook Apple Computer photo application.

RESULTS
To show the results of the effect of LT on plant root tissue, we need to refer the reader to an image showing the effect of LT in human soft tissue tissue of the human hair follicle a.k.a. root (1) and Figure 2 below.      Figure 7 above. Using landmarks as the raphide bundle (RB) and the unidentified filamentous structure (x), the observer could appreciate a filamentous repetitive pattern as demonstrated in Figure  6.

DISCUSSION
In this manuscript, in vitro experiments done on aerial plant root tissue introduces a facile method utilizing a commonly used antibiotic (Tetracycline) in aqueous form in direct contact with aerial plant root tissue able to uncover details of an intercellular filamentous veil. Of relevancy, also documented for the first time repetitive growth pattern of filamentous network in aerial plant roots (Figures 3,6 ,7,8,9).
A literature search failed to show the aforementioned repetitive filamentous pattern in plant roots as shown in these experiments; instead, a paper was found and it behooves corroboration by institutional affiliated researchers with a greater access to published data.
The Two Faces of Repetition Periodicity and the described phenomena of repetitive motion of DNA parts in cell division could induce or stunt growth of cellular structures, and could occur at many levels during tissue growth formation in both plants and animal tissue [3], [4]. On the other hand, a repetitive protein is essential for filament structure and function in parasites [5]. Since the antibiotic Tetracycline and its derivatives have also been reported to impact on the physiology and biochemistry in tissues of plants and animals [6], it could be stated that it has a deleterious impact on plant and animal cells, to the point of sort of eliminating barriers in light microscopy fields, thus allowing for the visualization of the repetitive patterns herein introduced (Figs 4,5 plus video recording) supporting this principle.

SUMMARY AND CONCLUSSIONS
I leave you with words from a renowned biologist "Orchids of many kinds have also adopted this high life. They lack the ponds that sustain the bromeliads, so they must collect their nourishment in other ways. Some dangle their roots in the air, absorbing moisture from the humid atmosphere and rely on the tiny number of nutriments it might have dissolved on its descent through the forest vegetation. Others spread their roots over the surface of the branches and collect the water that has trickled through the leaves and dripped from branch to branch, gathering a little nutriment on the way." [7], [8] Keratin as part of Aerial Root Filaments Regardless of the physics behind light microscopy imaging, the fact remains that the antibiotic Tetracycline when in contact with aerial plant roots appears to adhere albeit sparing root filaments from damage ( Figure 10). This finding is in support of an additional substance such as keratin as part of the root tip filament base [9], plus LT has been previously documented adhering to keratin in human hair follicle filaments ( Figure 2); furthermore in vitro experiments have documented "the explicit features of plant keratin intermediate filaments is a 24-25 nm periodic structural repeat alone the axis of both the 10 nm filaments and protofilarnents" [10]. The function of the repetitive filamentous network is theorized to be of physical support to roots inner tissue.
Plant Cell Walls and Liquid Tetracycline Aerial plant root tip epidermal cells walls are dissolved, thus pointing towards a cellulose base.
The facile method herein introduced could facilitate root function researchers in obtaining a rapid preliminary display of supporting root tissue filaments. The novel facile rapid method herein introduced could be used to obtain preliminary; or perhaps final images of plants and human hair roots filamentous anatomy.  Figure 2 showing Tetracycline adhesion to keratin in human hair filaments.

SOURCES OF FUNDING
This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.