The effect of magnetic water on macadamia plants' physiology and phenological characteristics

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The effect of magnetic water on

macadamia plants' physiology and

phenological characteristics

NA Boogaers

orcid.org 0000-0003-2519-6962

Dissertation submitted in fulfilment of the requirements for

the degree

Masters of Science in Botany

at the

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ACKNOWLEDGMENTS

First and most importantly, I am grateful to the Almighty God for my good health and perseverance that was needed to complete this thesis on a part time basis.

I would like to express my sincere gratitude to my supervisors Dr. Jacques M. Berner, and Dr. Tilla van der Westhuizen for their continuous support of my Masters study as well as their patience and critical guidance throughout. They guided me through the research project and provided support during the writing of this thesis. Without funding, I would not have been able to complete my project either, for this I would like to acknowledge Mr. Robin Haussman and Mr. Mark Hassenkamp of Red Sun Hortitech for their partial financial support and for providing the necessary project area with material included so that I could conduct a part of my research there. Furthermore, I would like to thank Mr. Jason Matozzo of Fractal Water USA for his generous donation of Imploders to be able to magnetize the water needed for this study.

I would also like to give thanks to the North West University for my study bursary and for the opportunity to complete my Masters under their respected institutional name. Besides my supervisors and funders, I would also like to acknowledge Christine Grobbelaar, Claudia Pretorius, Charne Malan, Velesia Lesch, WC Heppel, and Mr. Lindsay Tredgold for their advice, support, and help during the project setup, late night measurements, guidance as well as important referrals without which this project would not succeed. I would like to express a very special thanks to Renier van Wyk who has supported me throughout my studies emotionally and financially, without his support I would not have been able to complete a part time Masters project within the normal time frame.

Last but not the least; I would like to express my extremely profound gratitude to my parents (Mr. and Mrs. Boogaers) and brother Michael for supporting me throughout my years of study and through the process of researching and writing this thesis. My father always used to say:

“If you do something, do it right and to the best of your ability the first time, because tomorrow holds its own new challenges that will demand your best once more”.

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ABSTRACT

The biggest obstacle in the macadamia industry is both slow germination and growth rates. Several research studies have been conducted on magnetized water and its effect on plant growth. Magnetizing water can increase the number of hydrogen bonds by 0.34% leading to increased nutrient uptake in plants. The induction of biochemical changes within a plant species and the stimulation of several growth-related reactions have been known to be triggered by Magnetic fields.Two separate studies had been conducted during 2016 to 2018 at the North-West University in Potchefstroom (greenhouse trial 1); and at Red Sun Hortitech in Tzaneen region of Limpopo, South Africa (nursery trial 2). The aim was to investigate the influence of magnetic water on Macadamia plants’ physiology and phenological characteristics. It was hypothesized that magnetized water would increase the germination and growth rates of macadamias during propagation as well as improve overall plant vitality.

Macadamia integrifolia seeds were propagated under standard macadamia nursery conditions

during which germination and growth rates were measured. The plants were irrigated with magnetized water induced by the Imploder (a magnetic treatment device manufactured by Fractal Water). The photosynthetic efficiency and chlorophyll levels of the plants were measured in vivo during different growth stages. Analysis of the soil, water and nutrient status were also made after growth measurements were taken monthly. The results indicated that irrigation with magnetized water culminated a positive (P=<0.001) effect on seed germination with a higher success rate as compared to the control. In addition, magnetic water treatment improves plant growth (P=<0.001), root systems (P=<0.001) and leads to an earlier grafting age (P=<0.001). It also significantly (P=<0.001) increased the chlorophyll content levels of the plants. The overall plant vitality and photosynthetic activity of macadamias was improved (P=<0.001) with magnetized water which led to a more successful propagation. Magnetized water influences the ion charges in the soil to improve overall soil quality as it had a better water retainability with increased nutrient levels. Magnetic water treatment also decreased the total amount of nutrients that leached out of the planting bags during irrigation.

Keywords:

Chlorophyll a fluorescence; Chlorophyll content, Germination; Growth; Hydrogen

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LIST OF TABLES

Table 2-1: The importance of certain nutrients for the growth and health of macadamias ... 4

Table 2-2: Summary of the JIP-test formulas as described by Strasser et al., (2004). 17

Table 3-1: Plant material used during the study (Trial 2) at Red Sun Hortitech. ... 24

Table 4.-1: The germination success of macadamias grown under greenhouse conditions compared to local macadamia farms in the Mpumalanga province. (MWT – Magnetic Water Treatment) ... 33

Table 4-2: The accumulative germination over time between the control and magnetic water treatment of trial 1, grown under greenhouse conditions. ... 35

Table 4-3: Growth success of macadamia juveniles of both treatments measured at the age of 3 months during trial 1 compared to the growth success reported by local macadamia farmers in Mpumalanga province. ... 35

Table 4-4: The propagation success during trial 1 (germination and growth) of macadamias of both treatments compared to the reported propagation success of local macadamia farms in the Mpumalanga province. ... 36

Table 4-5: The average stem length (cm) grown over time of the magnetic water treated plants and the control during their different growth stages of trial 2 from June (germination) until February (after grafting). ... 47

Table 4-6: The grafting age of macadamias of the control as well as the magnetic water treated plants from October when the first plants were ready until December when the last plants were grafted during trial 2. ... 48

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Table 4-7: The overall graft and propagation success of macadamias in the two test groups (control and magnetic water treatment) throughout the study of trial 2 grown under nursery conditions. ... 49

Table 4-8: The average root and stem length of the macadamia plants after 10 months of growth for both the control and the magnetic water treated plants (Trial 2). ... 50

Table 4-9: The average plants’ biomass (dried) for both test groups after 10 months of growth during trial 2, grown under nursery conditions. ... 50

Table 4-10: The average Chlorophyll content (in SPAD units) of macadamia leaves of both treatments during different growth stages (Nursery study). ... 51

Table 4-11: Nutrient information of the macadamia growth mediums after ten months for both treatments (as given by the ARC Rustenburg). ... 53

Table 4-12: The nutrient values inside the water source before irrigation (clean water) and after irrigation (soil drainage) of macadamia plants’ soil bags during trial 2. ... 55

Table 4-13: Nutrient analyses of dry biomass of macadamias under both MWT and Control after ten months of growth during trial 2. ... 56

Table 4-14: Optimum nutrient range levels for Macadamia leaves (De Villiers and Joubert, 2003) compared to the levels in the magnetic water treatment plants. ... 57

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LIST OF FIGURES

Figure 2-1: Illustration of the observation of fluorescence as described by Strasser et

al. (2004). ... 11

Figure 2-2: Schematic illustration of PSII structure and the origin of fluorescence at the proximal antenna (Strasser et al., 2004). ... 12

Figure 2-3: Schematic illustration of the relation between primary photochemistry and chlorophyll a fluorescence (Strasser et al., 2004)... 12

Figure 2-4: An example of a graph illustration of the Kautsky effect (Govindjee, 1995). ... 13

Figure 2-5: An illustration of a typical chlorophyll a fluorescence increase that shows the accumulation of reduced QA (Strasser et al., 2004). ... 14

Figure 2-6: Schematic model for the energy flow of PSII as described by Strasser et

al., (2004) where ABS = photons absorbed through antennae pigments, TR = Exciton

flow (flux) to the RC and ET = electron transport further than QA. ... 15

Figure 2.7: The Z-scheme of electron transport in photosynthesis as described by Govindjee (1995). ... 16

Figure 3-1: A diagram depicting the greenhouse study experimental design at the North West University in Potchefstroom. ... 19

Figure 3-2: A diagram depicting the experimental design of trial 2 in the growth tunnel at Red Sun Hortitech’s nursery premises in Tzaneen (Note: Not drawn to scale). ... 25

Figure 4-1: Accumulative line graph indicating the difference in number of germinated seeds (% germinated over time) between the treatment and the control over 2 months during trial 1 (Data taken from table 5). ... 34

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Figure 4-3: (A) Average growth rate of macadamia juveniles’ stems of both treatments (magnetic water treatment and control) over six weeks, grown under greenhouse conditions during trial 1, and (B) the average root length of 6 weeks old juveniles of both treatments. ... 37

Figure 4-5: Phenomenological fluxes and performance indexes within the O-J-I-P transient with magnetic water treatment (MWT) normalized in terms of the control macadamias. ... 40

Figure 4-6: Difference in relative variable fluorescence of the chlorophyll a fluorescence transients at their specific JIP time (in ms) of PSII (680 nm) exhibited by dark-adapted leaves of 3 month old macadamia juveniles propagated under greenhouse conditions and grown under different treatments (magnetic water treatment (MWT) and non-magnetic treated). Transients are presented as kinetics of relative variable fluorescence (VI=(FI-F0)/(FM-F0) with the magnetic water treatment

normalized in terms of the control (non-magnetic treatment). ... 41

Figure 4-7: Line graph of the Macadamia stem growth rate over 10 months during trial 2, plants grown under nursery conditions at Red Sun Hortitech. ... 47

Figure 4-8: Chlorophyll content (in SPAD units) of macadamia leaves from both treatments over ten months of growth since first signs of developed leaves. ... 52

Figure 4-10: The OJIP Chl a fluorescence transient curve (log time scale) in macadamia leaves under MWT and Control, Where (A) is the O-J-I-P curve after grafting had occurred, (B) is at grafting age, (C) is 1 month before grafting and (D) is at first signs of developed leaves. ... 59

Figure 4-11: Difference in relative variable fluorescence of the chlorophyll a fluorescence transients at their specific JIP time (in ms) of PSII (680 nm) exhibited by

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LIST OF ABBREVIATIONS

Abbreviation

Meaning

ARC- Agricultural Research Council

ATP- Adenosine triphosphate

MW- Magnetized Water

MWT- Magnetic Water Treatment

NADPH- Nicotinamide adenine dinucleotide phosphate

NUE- Nitrogen Use Efficiency

NWU- North West University

PAR- Photo synthetically Active Radiation

ppm- Part per million

PS- Photosystem

PSI- Photosystem one

PSII- Photosystem two

QA-- Quinone A

QB-- Quinone B

SPAD- Soil Plant Analysis Development

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CHAPTER 1: INTRODUCTION

1.1 General introduction

For many years, the effect of static magnetic fields on plant growth has been a topic of scientific research. A few academic research findings have been published on the positive influence of magnetically induced water on the metabolism and growth of different plant species. Different experiments have been conducted, which prove that seedlings that are exposed to magnetic influence over time, have a faster germination period and better durability than seedlings under non-magnetic influence. There have also been reports of longer, stronger root systems (Bogatin et al., 1999). However, from the literature it could be concluded that most of this research was conducted on fast growing plants such as lettuce, peas and tomatoes (Harari and Lin, 1989; Lin and Yotvat, 1990; Govoroon et al., 1992; Hilal and Hilal, 2000; Moon and Chung, 2000; Belyavskaya, 2001; Reina et al., 2001; Maheshwari and Grewal, 2009; Shabrangi and Majd, 2009; Qados and Hozayn, 2010). That would make this study the only one conducted on macadamias, and the first of its kind on the effects of magnetically induced water on photosynthetic efficiency. This means that many of the explanations are still theoretical and much more research will have to be conducted to prove the theories as viable facts. It can also be noted that a lot of the research conducted is old and outdated.

Water treated by a magnetic field, or passed through a magnetic treatment device, refers to magnetized water. These devices are described as environmentally friendly, with no extra energy requirements and no needed maintenance after installation. Magnetized water can be used to increase crop yield and induce seed germination. Magnetic water treatment (MWT) is currently used all over the world for these purposes for the last 40 years (Gehr et al., 1995; Qados and Hozyan, 2010).

According to Marks and Szecówka (2010), the chances of positive influences of magnetically induced water on plants - that is increase in growth rate and durability of macadamia plants as well- are highly probable and worth researching. Due to the shortage of information

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1.1.1 Problem statement

The germination period of macadamia seeds is between one to four months, and thereafter, farmers wait an approximate of two years (11-18 months) before trees are grafted and ready for planting. Germination mostly takes place at nurseries and plants are later sold to farmers (De Villiers and Joubert, 2003). Personal communication with macadamia growers explained, that it is economically more viable to order juvenile plants from large nurseries than propagating their own, due to the low propagation success (less than 60%). If there was a way to increase the slow propagation rates of macadamias, whilst increasing propagation success without weakening product quality, development of the macadamia industry would increase.

1.1.2 Aim and Objectives

The aim of this study was to investigate and determine the effect of magnetic water on macadamia propagation.

The objectives for the greenhouse study were (1) to determine the influence of magnetic water on the germination rate and success of macadamia seeds; (2), to determine the influence of magnetic water on the growth rates and success of macadamia juveniles by investigating stem, and root growth; and (3) to evaluate the effect of magnetic water treatment on the photosynthetic efficiency of macadamia plants.

The objectives for the nursery study were (1) to evaluate the influence of magnetic water treatment on the growth rates and overall propagation success by measuring stem and root length, dry biomass and total successfully established trees; (2) to determine the effect of magnetized water on the chlorophyll content on fully developed leaves; (3) to investigate the influence of magnetic water on the grafting rate and success of macadamia juveniles; (4) to measure the level of influence of magnetic water treatment on the photosynthetic efficiency of macadamia plants; and (5) to evaluate the influence of magnetic water on a nutrient level by conducting soil, water and leaf analysis after successful grafting.

1.1.3 Hypothesis

It was hypothesized that magnetized or restructured water would improve macadamia plants’ growth, photosynthetic activity and overall vitality.

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CHAPTER 2: LITERATURE REVIEW

2.1 Macadamia plant development

Macadamia nut production is on the rise worldwide due to its rich nutrient content and health benefits. Macadamia nuts are described as a rich source of nutrients with minerals such as K, P, Mg, Zn, Cu and Mn. It is furthermore suggested that macadamias have cardio-protective potential and they are recommended as a part of a heart-healthy diet (Mereles et al., 2017).

2.1.1 Macadamia propagation

With regards to seed selection, Boogaers (2014) explained that both, Macadamia tetraphylla (rough shell) and Macadamia integrifolia (smooth shell) macadamia species grow in South Africa, although many hybrids occur as a result of crossbreeding. This makes it nearly impossible to obtain seeds that are specifically M. tetraphylla. The seed of any macadamia can be planted if they germinate well and produce uniform seedlings (Boogaers, 2014). Seedlings are usually fully propagated 15 - 18 months after being planted, when they are roughly 10 mm thick and at a height of 300-400 mm. Macadamia seeds are harvested when they fall from the tree or when their green husks split open, and it is important that the seeds are removed from their husks and planted immediately, otherwise dormancy would be initiated and the longer a seed is dormant, the longer it would take to initiate germination (De Villiers and Joubert, 2003). According to Boogaers (2014), heavier seeds produce stronger seedlings than lighter seeds. In order to distinguish between the heavy and lighter seeds, the seeds should be placed in water; the seeds that sink to the bottom are used for propagation purposes while those that float are discarded. The lighter seeds that float could possibly have kernels that were damaged by insects or diseases. If the seeds rattle or shake when handled, they should also be discarded. Seeds that cannot be planted immediately should be stored at room temperature, away from light. It should also be noted that seeds will still germinate, even after twelve months of storage, but it will germinate much slower than freshly used seeds. Boogaers (2014), has also stated that seeds rapidly lose their viability after four months of storage.

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2003; Boogaers, 2014). Through this crack, the taproot grows in the opposite direction of the shoot. For this reason, it is beneficial to plant the seeds with the white spot facing sideways or downwards. By soaking the seed in water before planting, the absorption of water into the kernel is enhanced, which in turn stimulates germination (De Villiers and Joubert, 2003).

Macadamia seeds are planted at a depth of 20 mm, and 80 mm apart, in well-drained seed beds that are lightly shaded (60%-70% sunlight exposure) and frequently watered. Germination may take between one to three months, depending on soil temperature, seed age and the origin of the seeds. Thus, it is important to note that, to maintain the fastest germination rate, seeds need to be soaked after harvest and planted without being stored. The soil quality also plays a significant role in germination and root development.

2.1.2 Mineral requirements of macadamia trees

According to De Villiers and Joubert (2003), Table 2-1 summarises the importance of specific nutrients to macadamia plant development.

Table 2-1: The importance of certain nutrients for the growth and health of macadamia

Nutrient Advised cation saturation ratio (me/100 g soil)

Effected by/ influences Notes

Ca 70-75% Too high Mg and K can cause a deficiency

Mg 15-20% Too high Ca or K can cause a deficiency

Important for photosynthesis, influences chlorophyll

K 5-8% Too high or too low Ca and Mg can cause a deficiency

Known as the nut quality nutrient

N Important for

vegetative/reproductive balance

Lower levels favour reproduction

P Deficiencies cause stunted root growth

Required for all energy processes in plants & cell

division

Zn Deficiency leads to chlorosis Participates in chlorophyll formation and photochemical

reactions

Fe Availability restricted by high or low soil pH, high P levels cause deficiencies

Very immobile in plants, needed in chlorophyll and photosynthesis & enzyme reaction centres

B Influences the uptake of Ca, Mg, and K. Too high or low pH limits uptake

Toxicity is easily induced if levels are too high

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2.2 Nutrient, water & soil quality and the influence of ion charges

Water plays a big role in the biogeochemical processes as well as chemical reactions of soil, and according to Duckworth et al., (2014), these chemical reactions that control soil formation, occur in liquid water. The hydrogen bonds that are created from the electrostatic interactions between the more positive hydrogen atoms and the more negative oxygen atoms in adjacent molecules, are relatively strong intermolecular forces (Duckworth et al., 2014).

The consistent use of saline water as irrigation during crop production increases soil salinization which in turn causes an accumulation of high soluble salts in the soil. This can significantly decrease the productivity of agricultural lands. One of the main agriculture practice limitations in developing or under developed countries such as South Africa and Iran, is the poor water quality and high salinity thereof (Mostafazadeh-Fard et al., 2011). Furthermore Higashitani et al., (1993), explained that magnetising water leads to a change in the physical characteristics of water that include physical properties such as salt solution capacity, density, and the deposition ratio of solid particles. They explained it further by using water containing CaCO3. They elaborated that as the ions of the calcium and carbon enter the

magnetising process, they are pushed into opposite directions caused by their difference in charges and after all of the same ions have been pushed to one side, they tend to collide with one another and stick together forming a solid form of CaCO3 also known as aragonite. In

other words, during the magnetisation process, water is restructured as anions and cations start to vibrate and later stick together causing the electrical charges of these particles to decrease. Mostafazadeh-Fard et al., (2011) concluded that by irrigating soil with magnetised water, one creates better soil conditions for plant growth to occur.

By measuring the pH of soil, one determines the ionised hydrogen (H+_{) activity in the soil}

solution; the higher the ionised hydrogen, the lower the pH (more acidic) in the soil. According to Hopkins and Hüner (2009), the two significant soil parameters to take note of are inorganic and organic soil particles also known as colloids. Colloids are described by Hopkins and Hüner (2009), as particles that are small enough to stay in suspension but too big to dissolve into a true solution. The soil colloids serve as nutrient reservoirs in the soil that retain soluble

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these surfaces carry a large amount of charges. It is important to note that these charged surfaces bind a lot of charged ions, especially positively charged cations from the soil solution because these colloids are predominantly negatively charged.

Ions are hydrated, meaning they are surrounded by shells of water molecules (Hopkins and Hüner, 2009). Electrostatic rules are modulated by the relative hydrated size of the ion according to Hopkins and Hüner (2009), and ions of smaller hydrated size can approach the colloids more closely leading to them being more tightly bound than the other ions. This process of exchange between adsorbed ions and ions in solution is defined as ion exchange (Hopkins and Hüner, 2009).

Due to the negative charge of soil colloids, anions tend not to bind that easily to nutrient reservoirs and, therefore agricultural practices have to apply more anion fertilizer than cations. Hopkins and Hüner (2009), explained that the anions would leach from the soil much easier and a plant can have anion nutrient deficiencies easier than cation deficiencies if the nutrients are not continuously re-supplied. A good example of this would be nitrogen in the nitrate form (NO3).

The charges within soils and amongst different nutrients, play a large role in what is available for plants’ roots to absorb, and what is available in the soil for absorption. It is known that by magnetizing water, the charges within the water’s nutrients and solids are changed. According to Nasher (2008), magnetised water can increase the levels of CO2 and H+ in soils,

comparable to the addition of fertilizers. This would play a role in the availability of nutrients available in the soil for macadamias during propagation where no additional fertilizer was added, and nutrients leach out from the planting bag during irrigation. If one could rearrange the charges of nutrients so that the colloids available in the soil retain more nutrients, the possibility of nutrient losses and soil depleting would decrease. This in turn could enhance nutrient uptake by plants and improve overall plant health.

It is known that most plants require small amounts of nutrient elements to complete their life cycle, and the ones required are known as essential elements which means that the plant would not be able to complete its life cycle in the absence of such nutrients. Hopkins and Hüner (2009) explained that there are 2 categories for essential elements to be divided into namely macro- (required in large amounts) and micro-nutrients (required in small amounts). They concluded that macro-nutrients are mostly needed for molecular structuring whereas the micro-nutrients are more specifically for catalytic and regulatory roles such as enzyme activators.

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2.3 Magnetized water

2.3.1 Water quality and Magnetized water

The hydrogen-bonded environment of water attributes to its known properties. These hydrogen bonds are affected by magnetic fields during which physical and chemical properties can be altered (Nasher, 2008). Several research studies have been conducted with magnetized water and its effect on plant growth and development (Reina et al., 2001; Racuciu

et al., 2008; Grewal and Maheshwari, 2011; Ahmed and Hassan, 2015). There is an

abundance of evidence supporting a positive effect of magnetic water treatment (MWT) on improved water productivity (Bogatin et al., 1999), salt solubility (Hilal and Hilal, 2000), as well as alkalinity (Hilal and Hilal, 2000). Changes in pH was also observed with the application of a magnetic field to water (Busch et al., 1986).

2.3.2 What is magnetized water

When water passes through a magnetic field, it becomes magnetized (Gehr et al., 1995, Qados and Hozayn, 2010). Magnetic water treatment works with magneto hydrodynamics where electric energy is added to charged particles in water that contain ions and small solid particles with electrostatic charges by a magnetic field. Energy is produced by the particles’ momentum and it stays attached to the particles as surface energy (Gehr et al., 1995). Magnetized water causes a redistribution of the energy flow due to a momentum change of charged particles (Chang and Weng, 2006).

Natural water contains micro and macro particles (organic and inorganic) as well as different ions, zoo- and phytoplankton, and micro bubbles (Bogatin et al., 1999). When water passes through a magnetic water softener, a Lorentz force is exerted on each ion in the opposite direction. The redirection of the particles increases the frequency of collisions between ions on opposite sides, combining to form a mineral precipitate or insoluble compound (Gholizadeh

et al., 2008). Calcium carbonate precipitates out of the solution as a sludge and can be easily

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(Bogatin et al., 1999). Regardless, Magnetic water treatment is considered a “green technology”. Magnetic treatment technologies can reduce energy consumption, chemical resources, and water usages, thus leading to cost saving and environmental improvement. Devices for creating magnetic water is generally compact, with permanent magnets that are created from clean and safe technology (Otsuka and Ozeki, 2006).

2.3.3 Determined benefits of magnetized water

Physical benefits

Salinity and Alkalinity

Magnetizing water causes changes in the amount of salt crystallisation centres in the water (Shercliff, 1965). The quick change of the magnetic field in a properly designed magnetic water softener, loosens the hydrate layers plus films in a moving liquid, enabling coagulation and coalescence (Bogatin et al., 1999). Magnetic water treatment of saline water can be used as a method of soil desalinization (Kney and Parsons, 2006). Field experiments conducted in Egypt, indicated that sandy loam soil pots that were irrigated with normal saline water of an electrical conductivity value of 8.2 mmohs/cm, retained more salts, as compared to pots irrigated with magnetised saline water (Hilal and Hilal, 2000). The study showed that magnetic water increased the leaching of excess soluble salts, lowered soil alkalinity as well as dissolved slightly soluble salts (Hilal and Hilal, 2000).

Changes in pH of the water to more suitable levels had been observed when Joshi and Kamat (1966) as well as Busch et al., (1986) applied a magnetic field to water. Parsons et al. (1997) confirmed a decrease in pH during an investigation where magnetic treatments were applied to the solution after sodium hydroxide was used to stabilize the pH at 8.5. In their study, the magnetically treated water required up to 2.5 times more sodium hydroxide, as compared to the controls in order to stabilize the pH level. The pH has been shown to decrease from 9.2 to 8.5 after magnetic treatment in a system with Ca(OH)2, where the degree of the reduction

depended on the strength of the magnetic treatment (Ellingsen and Kristiansen,1979). Busch

et al., (1986) showed an initial decrease in pH, from 7.0 to 6.5, that was followed by an

increase in pH from 7.5 – 8.0.

Soil Permeability, water use efficiency and scale reduction

Applying a magnetic field to natural water can enhance degassing by 25-30%. This is caused by local dehydration of surface micro-bubble films, as well as a decrease in pressure in the

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centre of vortices, resulting in an increase in free gas bubbles (Bogatin et al., 1999). Degassing can increase permeability in the soil, resulting in an increase in irrigation efficiency because more water can cypher through the soil and be absorbed by plants

It is believed that magnetised water, when used for irrigation, can improve water productivity (Duarte Diaz et al., 1997), conserving water supplies for the expected future global water scarcity. This improved water usage has theoretically decreased the daily use of water with 20% (Duarte Diaz et al., 1997). For an arid environment such as South Africa, suffering from frequent droughts, this could have an influence on the future of their water recourses.

The precipitation of CaSO4 was investigated by Gehr et al., (1995) and it was found that

magnetic treatment induced precipitation of gypsum crystals (CaSO4 H2O2). It was further

established that, if magnetic treatment were to be an effective treatment for scale prevention, it would most likely reduce precipitation on solid surfaces and encourage crystallization,with an efficiency rating of 20-40%, (Kronenberg, 1993).

The U.S. Department of Energy reported that a thin film (1/32 inch) of scale in a heat exchange surface can increase energy consumption by 8.5%, and scale build-up of up to 1-inch increases energy consumption by 25% (U.S. DOE, 1998). It was estimated that scale removal in Britain cost £ 1 billion per year in the early 1990’s (Smith et al., 2003). The installation of magnetic treatment devices for the removal or prevention of scale build-up has been shown to generate significant energy savings, even with the initial capital to invest in the technology (Kronenberg, 1993).

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Biological benefits

Germination rate, germination success and crop yield

The application of a magnetic field induced seed germination and increased the percentage of germinated seeds when Moon and Chung (2000) treated tomato seeds with a magnetic field. They found that germination rates were accelerated by an estimated 1.1–2.8 times, when compared to the control. Pinus tropicalis, a tree species endemic to western Cuba, typically experiences a <50% germination rate, yet when seeds were treated with magnetic water as part of a 2007 study, germination occurred in 70-81% of the seeds (Morejón et al., 2007). Germination of broad bean seeds was found to take place two to three days earlier when seeds underwent magnetic treatment (Podleoney et al., 2004).

Lin and Yotvat (1990) indicated an increase in crop yield, size and sugar content of melons grown with magnetised water. Maheshwari and Grewal (2009) reported statistically significant increase in the yield and water productivity of snow peas and celery. Harari and Lin (1989) conducted a study that demonstrated that the size of muskmelons, the number of fruits as well as the sugar content significantly increased with magnetic water treatment. Lentils irrigated with magnetized water displayed a significant increase in growth, (Qados and Hozayn, 2010). Magnetised water has also been reported to triple seedling emergence of wheat (Hilal and Hilal, 2000). Reina et al., (2001) reported a significant increase in the rate of water absorption and an increase in total mass of lettuce.

2.4 Plants’ photosynthetic activity and how to measure it

During photosynthesis, sunlight (in the form of energy) absorbed via the chlorophyll a pigment, can follow three different routes. The first is for photosynthesis to occur, the second route is where energy can be released or lost in the form of heat, or thirdly, it can be used as chlorophyll a fluorescence (Misra et al, 2012). According to Hopkins and Hüner, (2009), these three routes are in constant competition with each other and as the Law of Conservation of Energy explains, the total amount of energy in the system is constant. Thus, if there is a rise in the amount of energy in the form of fluorescence, there will be a decline in energy at the other two routes.

Chlorophyll a fluorescence is defined as the loss of partial exit energy after the antennae has absorbed the chlorophyll light. This happens in PSII (Photosystem II) through the radiation of red-light energy with a wavelength of 680 nm. In the case where no energy is exited, damage could be done to the leaves resulting in a negative influence on photosynthesis. The

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fluorescence intensity is a measurement of the number of absorbed photons that were not used for photosynthesis and is also an indicator of the amount of stress in a plant. This is done by visually observing the plants.

PSII consists of 31 sub-units that each play a role in the transportation of electrons as well as the uptake of photons from the shorter wavelength of light that was absorbed (Figure 2-1). PSII consists of a Light Harvesting Complex that absorbs light and lets it flow through the antennae of the chlorophyll to the reaction complex. From here light (photons) move further down the membrane to the cytochrome complex and photosystem I. Temperature has an influence on the absorption of photons and thus on the electron transport in PSII. Water has an influence on the plants’ temperature regulation and thus an influence on PSII (Misra et al., 2012). The absorption of a photon through a leaf, causes the chlorophyll α molecule to rise to its lowest active singlet. Misra et al. (2012) further explained that there are three ways for the molecule to end at the ground phase. They are fluorescence, internal transformation and intersystem crossings. Fluorescence originates at the antennas of photosystem II (PSII) as seen in the Figure 2-2: Fluorescence occurs when the molecule returns to the ground state with emission of radiation.

Figure 2-1: Illustration of the observation of fluorescence as described by Strasser et

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Internal transformation is when the energy of the molecule is transformed into a vibration energy, and intersystem crossings refers thereafter to the singlet phase being transformed to a triplet phase (Misra et al., 2012). Relations between primary photochemistry and chlorophyll fluorescence are described by the law of conservation of energy: Exiting tempo = sum of the de-exiting tempos (Figure 2-3).

Figure 2-2: Schematic illustration of PSII structure and the origin of fluorescence at the proximal antenna (Strasser et al., 2004).

Figure 2-3: Schematic illustration of the relation between primary photochemistry and chlorophyll a fluorescence (Strasser et al., 2004).

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The Kautsky effect

When a leaf, that is adapted to darkness, is exposed to light, a characteristic change is noted in fluorescence emission over a period of a few minutes, until a constant value is reached. This is known as fluorescence induction, or the Kautsky effect (Strasser et al., 2004).

The Kautsky effect was described in 1931 by Dr. Hans W. Kautsky as a description of the fluorescence producing plants and other chlorophyll organisms when they are exposed to light. During the Kautsky curve, there is a sample containing chlorophyll that is exposed to a constant light source. The fluorescence intensity increases with time and normally reaches a maximum of 300 – and 500 ms (milliseconds) as seen in Figure 2-4 (Govindjee, 1995):

The JIP-test consists of the measurement and analysis of the fluorescence inclination (J-I-P) as described by Strasser et al., (2004). The original chlorophyll a fluorescence at the O-step, is called the minimum fluorescence and occurs when all the plastoquinone (QA)

molecules are in an oxidized state. At the end of the curve, at the P-step, all the QA molecules

are reduced. The J- and I-steps occur between the O- and P-steps. The steps from O to J are the reductions of QA to QA- and this goes with the photochemical reactions of PSII. The

intermediate step that occurs, the I-step, together with the end of the curve, P, shows the existence of the slow and quick reducing PQ centres. The O-J-I-P curve shows the measurement of the PQ-pool inside the leaf tissue and that the curve will change due to environmental influences (Strasser et al., 2004).

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The fluorimeter with a high exiting light intensity, quick time resolution and big data collecting capacity, shows the O-J-I-P steps on a logarithmic timescale. The chlorophyll a fluorescence increase shows the accumulation of reduced QA (when the reaction complex closes). This is

the net result of the reduction of Q-_{through PSII and the re-oxidation thereof through PSI.}

The abbreviations refer to the fluorescence data that was used by the JIP-test for the calculations of the different parameters quantified by the PSII structure and function. PEA plus software was used to complete the calculations (Strasser et al., 2004). The meanings of the abbreviations are as follows: The fluorescence intensity F0 (at 50 µs); the fluorescence

intensity FJ (at 2 ms); the fluorescence intensity FI (at 30 ms); and the maximum fluorescence

intensity Fp = Fm at 300 ms.

The JIP-test serves as an in vivo measurement for the behaviour of the photosynthesis mechanisms, the PSII-function in specific. This serves as a vitality measurement of a plant. From the fluorescence curve, the following data points are used for calculations:

(i) Fluorescence intensity at 50 μs, 100 μs, 300 μs, 2 ms (shown as FJ), 30 ms and

the maximum Fm.

(ii) FO, minimum fluorescence (calculated by the M-PEA instrument).

(iii) tFmax, refers to the time needed to reach Fm.

(iv) Mo, start-up gradient, calculated by using the values at 100 μs and 300 μs. (v) Sm, surface above the fluorescence curve.

Figure 2-5: An illustration of a typical chlorophyll a fluorescence increase that shows the accumulation of reduced QA (Strasser et al., 2004).

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The specific energy flow (per RC) at t= 0, namely, ABS/RC, TR0/RC and ET0/RC are calculated

from the experimental values. The relation of two energy flows result in the: 1. Quantum increase of primary photochemistry (TR0/ABS)

2. Success of changing exit energy to electron transport (ET0/TR0), and

3. The probability that an absorbed photon will move into an electron in the ET-chain (ET0/ABS).

The phenomenological energy flow (per CS) is calculated by multiplying the specific energy flow with the density of the RC’s as described in Figure 2-6 (RC/CS) (Strasser et al., 2004).

Chl*

RC

ABS

TR

ET

F

Q

_A A

-(N-1)

N

e-t t ABS RC RC RC TR₀ ET₀ ET₀ TR₀ TR₀ ABS = P0 = 0 E0 = ET₀ ABS

Figure 2-6: Schematic model for the energy flow of PSII as described by Strasser et al., (2004) where ABS = photons absorbed through antennae pigments, TR = Exciton flow (flux) to the RC and ET = electron transport further than QA.

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Strasser et al., (2004) explained that chlorophyll a fluorescence can be divided into four important steps known as (1) Absorption at the FO-step (0.03 ms), (2) Trapping of electron

energy at the FJ-step (2 ms), (3) Electron transport from the FI-step to the FP-step (30 ms),

and (4) The reduction of NADP+_{to NADPH at the F}

P-step (300 ms) of the OJIP-curve.

The Z-scheme in Figure 2-7, is an indication of which step occurs in the photosynthesis process of the light dependant part of photosynthesis. At these four steps as well as the FK

-step at 0.3 ms, as explained by the formulas in Table 2-2 below by Strasser et al., (2004), five important parameters’ information can be extracted from the OJIP-curve. These parameters are known as quantum efficiencies and are calculated by specific formulas seen in Table 2-2. Information about the absorption and translocation of energy can be found at 0.03 ms on the OJIP-curve. Information about the ability to be able to split water molecules as well as the reduction success of QA to QA- can be found at the 0.3 and 2 ms graph points respectively.

The FI-step of 30 ms indicates the electron transport efficiency, and information about how

well NADP+ is reduced to NADPH can be extracted at the FJ-step of 300 ms on the

OJIP-graph seen in Figure 2-7 (Strasser et al., 2004).

Figure 2.7: The Z-scheme of electron transport in photosynthesis as described by Govindjee (1995).

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Table 2-2: Summary of the JIP-test formulas as described by Strasser et al., (2004).

Extracted and Technical Fluorescence Parameters

F0 F100μs F300 μs FJ FI FM t_FM VJ (dV / dt)0 = M0 = = = = = = = = = F50μs , fluorescence intensity at 50μs fluorescence intensity at 100μs fluorescence intensity at 300μs

fluorescence intensity at the J-step (at 2ms) fluorescence intensity at the I-step (at 30ms) maximal fluorescence intensity

time to reach FM , in ms

relative variable fluorescence at the J-step = (F2ms - F0) / (FM - F0)

fractional rate of PS II reaction centre closure = 4 . (F300 - F0)/(FM - F0)

Quantum Efficiencies or Flux Ratios or yields φPo = TR0 / ABS φEo = ET0 / ABS φRo = RE/ABS Ψ0 = ET0 / TR0 = = = = [1 - (F0 / FM)] = FV / FM [1 - (F0 / FM)] . Ψ0 (1-VI)/(1-VJ). φEo (1 - VJ)

Phenomenological Fluxes or Phenomenological Activities

ABS/CS TR0 /CS ET0 /CS DI0 / CS = = = =

ABS / CSChl = Chl / CS or ABS / CS0 F0 or ABS / CSM FM

φPo . (ABS / CS)

φPo . Ψ0 . (ABS/CS)

(ABS / CS) - (TR0 / CS)

Density of Reaction Centres (Gamma RC)

RC / CS = φPo . (VJ / M0) . ABS / CS

Performance Indexes

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CHAPTER 3: MATERIAL AND METHODS

The research project consisted of a greenhouse and nursery investigation. The greenhouse study was conducted in 2016 at the North-West University, Potchefstroom and is hereafter known as trial 1. The nursery study was much more extensive than trial 1 and was completed at Red Sun Hortitech in Tzaneen, Limpopo during 2017 and 2018. The nursery study is hereafter referred as trial 2.

3.1 Greenhouse study (Trial 1)

A greenhouse study was conducted to test if this project would carry any validation. Seeds were germinated under hessian and planted over into 5 L planting bags with decomposed bark soil. During this study several differences were noted such as increased germination success and increased germination rates, improved plant health as well as faster growth rates. This greenhouse study served to prove the worth of conducting a full-scale research project in this topic during further macadamia propagation until grafting stage.

The macadamia propagation method that was used for this trial, was adapted from De Villiers and Joubert (2003). The greenhouse study conducted, focused on seed germination rates and germination success as well as initial growth rates. These results are given in the result section.

3.1.1 Plant material

Three hundred Macadamia integrifolia (smooth shell) seeds of the same age and origin were collected and de-husked. Seeds of the 788 Beaumont cultivar were used as they revealed to be the best germinating seeds during the greenhouse study. The tree from which the seeds were harvested was 10 years old with a Beaumont root stock variety. These seeds are described as integrated as opposed to a hybrid species and they have a large nut size with medium growth speed. After de-husking, the seeds were divided into two groups, firstly the experimental group for magnetized irrigation, and secondly the control group of non-magnetized irrigation.

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3.1.2 Experimental design

The experimental area was set up inside a greenhouse under controlled conditions. The temperature during the day was set at 26 °C; and the temperature at night was set at 15 °C and lights were used to maintain a 10 hours day length.

The experimental area was divided into four equal rectangular areas called grids (Figure 3-1). Within each grid, the treatments were plotted in accordance with a randomized design principle that plots are very likely to vary less within the smaller area of a grid than over the whole experimental area. Two grids were used for germination and the other two for irrigation during growth. As seen in Figure 3-1, one germination grid (grid A) and one growth grid (grid C) were connected to magnetically induced irrigation with the imploder installed, and the other two (grids B and D) were connected to municipal water irrigation. Grids A and B were allocated for the hessian germination period. These two grids were subdivided in the same RCBD design as grids C and D in the greenhouse, just on a smaller scale by using tape to form a grid. Within these smaller scaled plots of grid, A and B, the germinating seeds were separated from those that indicated rot or damage.

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3.1.3 Preparations

Planting bags (5 L) were filled with composted bark soil and placed on grids C and D, each with its own white tag for marking the date planted. Glass bottles (2 L) were sterilized and filled with water (according to the treatment), either magnetically induced or municipal water.

3.1.4 Water magnetization

Water was magnetically induced by using the Imploder, a magnetic treatment device manufactured by Fractal Water; which was connected to the irrigation system. The machine consists of a 100 mm pipe section with its internal diameter at 22 mm. Inside the piping there are two magnets arranged that the poles are opposite of one another. The imploder has a magnetic array with a unique directional nozzle. This nozzle allows the water to only be passed through the machine once to be magnetized. The magnetic field intensity was at 5070 Gauss and with a magnetic field in the range of 3.5 mT – 136 mT. The combination of these forces creates “implosion”, which refers to the sorting and phase locking of the plasmic forces of water. This creates centripetal intense flux lines, resulting in a dynamic spin rate as well as a smaller water cluster size.

3.1.5 Soaking of seeds, germination and planting

Seeds were soaked for 24 hours in water using glass bottles, according to the treatment i.e. magnetically soaked (150), and the control (150). After soaking, 90 seeds in total were removed due to rot or infections (caused by insects). There were 105 seeds per treatment left for germination. Seeds were placed into the misting beds of grids A and B consisting of hessian material. The irrigation method used was misting, with a scheduling of three times a day for 10 minutes. This continued until first signs of germination were noticed. Germinating seeds were then taken to the growing grids C or D for planting.

As soon as germination signs occurred, the germinated seeds were planted over into planting bags and tagged according to the date planted. A ruler was used to measure about 2 cm deep into the soil for each seedling to be planted with the micropyle facing to the side. Bags were moved within their grid once a week to ensure a randomized block design. The soil consisted of one-part sand, two parts compost (decomposed bark medium). The compost was well-rotted plant material, and the mixture had a pH of 6 to 7.

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3.1.6 Data collection and analyses

3.1.6.1 Statistical analyses

Statistical analysis was done by subjecting all data to a one-way ANOVA analysis of variance (complete randomized design) to compare treatments at 5% significance level (Kruskal-Wallis and Tukey Test). SigmaPlot v12.0 software was used.

3.1.6.2 Water data analyses

The magnetically induced water as well as the municipal water was tested. This was done with the Lovibond Multimeter which measures pH, salt conductivity and oxygen content. The water samples were also analyzed for mineral content. Water was sampled in marked falcon test tubes. Samples were tested twice and compared to one another. For mineral content testing, water was tested immediately after being magnetized as well as after 24 hours of being magnetized to determine if the change in water due to the magnetization was temporary or permanent.

3.1.6.3 Germination and growth measurements

Germination rates were monitored by noting the number of seeds germinated per week. After germination was completed, the germination success was determined by subtracting the amount of successfully germinated seeds from the initial count of 105 and the percentage was calculated. This was done for both the treatments to compare results. Thereafter, plant growth rates were monitored by measuring the length of the plant stem with a ruler (cm) once a week for six weeks. The growth success was also determined by subtracting the amount of successfully grown juveniles from the number of seeds geminated and planted. This too was calculated in percentage for both the treatments.

Propagation, in macadamia farming, refers to the process from seed germination until successful juvenile growth or grafting age. Thus, by calculating both the germination and

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treatments as well as other nurseries. Photographs were taken for visual observation and comparisons.

3.1.6.4 Photosynthetic efficiency measurements (JIP-Test):

The Multi-Function-Plant-Efficiency-Analyser (M-PEA) was used to make in vivo measurements of the following parameters simultaneously: (1) The kinetics of the polyphasic prompt fluorescence rise (PF) with which the OJIP-test could be conducted and (2) the modulated reflection change near 820 nm (MR).

Chlorophyll a fluorescence of the macadamia plants was measured on the adaxial leaf surface of dark-adapted leaf discs in the glasshouse. The measurements were conducted after sunset between 20:00 and 04:00 before dawn, whilst ensuring the plants were not exposed to any external light source. The reason for measuring 1 hour after sunset is to eliminate all the extra electrons available in the chlorophyll of the plant. The photosynthetic rate needed to be calculated, and this cannot be done if there are already electrons in the system driving photosynthesis. A red LED-light of 625 nm wavelength with an intensity of 5000 µmol photons m-2_s-1_{was used for the PF-signal detection system of the M-PEA (Strasser et al., 2000).}

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3.2 Nursery study (Trial 2)

After the successful germination during the greenhouse study, a nursery macadamia propagation study was conducted to evaluate the effects of magnetized irrigation until after grafting age when plants are sent to farmers.

This study was conducted from 2017 to 2018 at one of the largest macadamia nurseries in South Africa called Red Sun Hortitech, situated in Tzaneen, Limpopo. The propagation methods were conducted according to their protocol and standards, and in one of their growth tunnels specifically designed for macadamia propagation. Due to seeds being germinated in soil beds on a large scale; and considering the success of the pilot study; this study was initiated after germinated seedlings were replanted into soil bags and placed into the propagation tunnel.

Detailed propagation methods have not been disclosed in this dissertation as it is the intellectual property of Red Sun Hortitech.

3.2.1 Plant material

The plant material starts at test group 5 as seen in Table 3-1. This is because tests 1-4 were indigenous trees tested by Red Sun Hortitech under the same conditions as the macadamia tests for their own comparisons.

The test groups can be divided into 2 sets where set 1 is rootstock germinated from seeds; and set 2 is cuttings from mother plants. Each set can be subdivided into the days on which the seeds were sown, or the cuttings were cut from their mother plants.

All the test groups were transplanted or weaned into the growth tunnel where the magnetic water treatment was initiated within the same week in June 2017.

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Table 3-1: Plant material used during the study (Trial 2) at Red Sun Hortitech. Test group Rootstock/ Cutting Date sown/cut Date transplanted/ weaned

Total Traceability Graft cutting

5 695 Beaumont Rootstock 08/11/2016 sown 01/06/2017 transplanted 1120 K.B.F. 842 cutting grafted on top of rootstock, A4 graft 6 695 Beaumont Rootstock 29/11/2016 sown 02/06/2017 transplanted 1120 K.B.F 842 cutting grafted on top of rootstock, A4 graft 7 695 Beaumont Rootstock 29/11/2016 sown 05/06/2017 transplanted 512 K.B.F & LNR/ARC A4 grafting 8 695 Beaumont Cutting 02/03/2016 cut 08/05/2017 Weaned 07/06/2017 grown 80 Aquamatrix N. A 9 695 Beaumont Cutting 02/03/2016 cut 08/05/2017 Weaned 05/06/2017 grown 1520 LNR/ARC N. A

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3.2.2 Experimental design

The study area can be seen in Figure 3-2 where the area was divided into 2 sections namely the magnetic water treatment MWT (Left) with the imploder implemented onto the irrigation line, and the control (right) with a clean irrigation line. Both irrigation lines originate from the same water source and irrigated at the same rate and quantity. Each section was divided into the test group (TG) blocks (1-9) and distributed equally below the aerial sprinkler irrigation system.

Figure 3-2: A diagram depicting the experimental design of trial 2 in the growth tunnel at Red Sun Hortitech’s nursery premises in Tzaneen (Note: Not drawn to scale).

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3.2.3 Water magnetization

Water was magnetically induced by using the imploder, a magnetic treatment device manufactured by Fractal Water; which was connected to the irrigation system in the grow tunnel at Red Sun Nursery for the magnetic water treatment (MWT) side of the study. The same magnetization machine and method was used as during trial 1.

A water meter was added to both irrigation lines, the imploder line as well as the control line. This meter ensured that each treatment received the exact same amount of water with the same flow rate. Irrigation occurred daily and manually as the grower deemed fit. This way overwatering was avoided as macadamia plants do not like ‘wet feet’.

3.2.4 Planting, growth and grafting

Seed germination & Planting

Seeds were germinated in shallow sand banks after which the successfully germinated seeds were replanted into composted bark soil planting bags and moved to the growth tunnel.

Cuttings and planting of cuttings

Red Sun Hortitech have their own Mother blocks with plants from which they make cuttings to plant over. The technique behind propagating from cuttings is very specific to each nursery and protected as their intellectual property. A basic method can be found in De Villiers and Joubert (2003). After the cuttings have been weaned and were successfully growing, they were moved to the same growth tunnel as the germinated seeds.

Growth

The growth tunnel consisted of a rectangular area covered by shade net (60%). Plants were packed onto a grid elevated about 15 cm above ground to allow ample water drainage through the soil. The ground floor was covered with plastic to avoid any possible diseases spreading. Another hygiene precaution was the chemical troughs at each tunnel entrance for decontamination of shoes before entering the growth area. Plants were cared for daily to remove sick plants, irrigate when needed, and identify plant health deficits or pests that might cause permanent damage. Plants were grown in the tunnel until grafting age, and after grafting until they were strong enough to be planted in the field.

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Grafting

As explained by De Villiers and Joubert (2003), grafting of rootstock occurs when the stem is about pencil thickness (10 mm thick) and the plant is in good health. Grafting is done by hand by a specially trained team using cutters, tape and fresh cuttings taken from mother plants. Strict hygiene protocol played a big role in the successful grafting of cuttings onto root stock. Grafting is done onto root stock grown from seed, not from cuttings that have been propagated. According to De Villiers and Joubert (2003), grafting occurs when the rootstock plants are about 15 to 18 months old (in some cases as early as 10 months of age).

3.2.5 Data collection & analyses

Most data collection occurred at the study area and the analyses thereof occurred at the North West University.

3.2.5.1 Statistical analyses of data

Statistical analysis of the photosynthetic (O-J-I-P) parameters and variable fluorescence were conducted. This was done by subjecting all data to a one-way ANOVA on Ranks analysis of variance (complete randomized design) to compare treatments at 5% significance level (Kruskal-Wallis and the Tukey Test). SigmaPlot v12.0 software (Systat Software, Inc., San Jose California USA) was used.

3.2.5.2 Plant growth

The growth analyses of plants were conducted by taking monthly measurements of the stem lengths above ground with a ruler (in cm). The average size grown per month was then calculated and grafted to determine which group (Magnetic water treatment or Control) grew faster. The root growth analyses were done at the end of the study (after successful grafting). Fifty plants per group (Magnetic water treatment and Control) were selected (5 per test group) to be de-rooted. The soil was rinsed off, photos were taken, and the roots’ lengths were

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was used to determine a propagation success percentage per test group. The same concept was used to calculate the grafting success after grafting took place. Another measurement taken at grafting was the age of the plants when they were ready for grafting as this plays a big role in the economic value of the project. Plants that became sick, or died, were thrown out and counted as waste to avoid disease or sickness spreading.

3.2.5.4 Growth characteristics (Biomass, root growth etc.)

Growth characteristics were determined and analysed by using the plant material collected during the root analyses. After the soil was washed off, the plant material was bagged and sent to the Unit for Environmental Sciences and Management, North West University,. One full plant was bagged (leaves, stem and roots) per paper bag. This sampling took place once at the end of the study when the plants were developed and successfully grafted upon. Fifty plants per group (Magnetic water treatment and Control) were selected (5 per test group) to be de-rooted (100 plants in total). The plant material was air dried in hot air oven at 75 °C for 3 days. After the plants were completely dry, each bag was weighed to determine the full dry biomass per plant and thereafter average biomass was calculated per test group.

3.2.5.5 Chlorophyll content

The second set of leaves per plant were selected to measure the chlorophyll content of each plant. The reason for the second set of leaves is to ensure that the youngest fully developed leaves were used. A fully developed leaf’s chlorophyll will also be fully functional whereas a developing leaf’s chlorophyll content will not be consistent.

Twenty plants per test group were used to measure the chlorophyll content. Two leaves per plant were selected, the youngest fully developed leaves. Three measurements per leaf were taken (6 measurements per plant) after which an average per test group was calculated. The chlorophyll content was measured with a hand-held chlorophyll content meter (Model CCM 300 from Opti-science, USA). To estimate the leaf chlorophyll content, SPAD values were generated, otherwise known as Dimensionless Soil Plant Analyses Development. Measurements were taken by placing the leaf clip at a central point on the leaf between the midrib and the margin of the leaves.

The plants that were measured were marked with stickers to avoid measuring the same plants again during the next month’s measurements.

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3.2.5.6 Nutrient analyses of the growth mediums and water resource

Soil sampling

When the plant material was sampled for the root and biomass analyses, the soil that was removed from the plants were bagged and labelled for further testing. Soil samples of 5 plants per test group, 50 plants per group (control and Magnetic water treatment) and 100 plants in total were collected. The soil was sent to Ms. K.P Ngwato at the ARC Rustenburg to be analysed on 09/03/2018.

The analysing method followed included compressing the soil as 100 ml substrate samples for 60 seconds at a pressure of 500 g/cm2_{. The 100 mL samples were extracted and filtered}

by using deionised water at a ratio of 1:1,5. The 100 mL extract was analysed and correlated according to the water volume inside the soil sample. The results were displayed in table format in milligram per litre.

Water sampling

Water sampling took place the same day the plant material and soil sampling took place. The sampling for water was divided into two sets:

Set 1 was clean water collected from the (a) control irrigation source (after going through the sprinkler system) and (b) the magnetic water treatment irrigation source (after going through the Imploder and the sprinkler system). This set’s sampling was done by hanging empty containers from the sprinkler line and catching up the needed water. Five litre water was collected per (a) and (b).

Set 2 was soil drained water from (a) the control’s irrigated plants and (b) the Magnetic water treatment’s (MWT) irrigated plants. The samples were collected by placing several containers underneath the plant bags (spread over the whole study area) to collect all the water draining through the soil after irrigation has occurred. Five litre water of each (a) and (b) were collected

The effect of magnetic water on macadamia plants' physiology and phenological characteristics