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1. Introduction

     In the EPG technique, an insect with piercing mouthparts and a plant are made part of an electrical circuit. Two electrodes to the plant and the insect connect them to that electronic measuring circuit, called the primary circuit (Fig. 1). In general, the insect may be any 'organism' with piercing mouthparts, such as a mite or a thick, and the plant can be any substrate such as a prey or an artificial substrate behind a membrane. Essential is that the electrical mouthpart contact is not electrically short-circuited circuit by legs or any other body parts.

     The recorded EPG signal is a fluctuating voltage which is caused by two different sources. One source is a fluctuating voltage generated by the insect, the plant or their interactions. This is called the ´electromotive force [emf] component´ of the signal. Additionally, another fluctuating voltage is caused by electrical resistance fluctuations in plant and insect. This is called the ´resistance component´ of the signal (chapter 3). The voltage of resistance component is due to voltage division by the fluctuating insect-plant resistance and the constant input resisttance of a certain voltage supplied to the primary circuit. Originally (McLean and Kinsey, 1964,1965), the supplied voltage was an oscillating AC voltage (alternating current) the amplitude of which fluctuated due to the insect-plant resistance fluctuation. Later (Tjallingii, 1985, 1988, 2000), the supplied voltage was a DC voltage the amplitude of which fluctuated by the insect-plant resistance.

     EPG systems, AC, DC, and combined AC/DC systems are discussed here, as well as their use with different circuit properties and signal processing principles.

   Experimental set up and operating instructions are given in the download manuals on this site.

 

2. Primary circuit

   The primary circuit properties determine what is recorded and what is not. It incorporates a voltage supply Vs, the plant electrode in the plant itself, in potting or outside soil, or cut off in water, the insect its electrode, and the input resistor Ri shown in the figure below. The input resistor is positioned between the pre-amplifier input and ground. The pre-amplifier is connected to the circuit at measuring point (M) but is not a part of the primary circuit as such, and does not affect the voltages in the primary circuit.  As soon as the insect stylets penetrate the plant, the circuit is completed and a fluctuating voltage at the measuring point - the EPG signal - is amplified and recorded on a computer hard disk. The voltage fluctuations appear in a number of distinct patters with respect to amplitude, frequency, and voltage level, which are referred to as 'EPG waveforms'. For a number of insects thesewaveforms have been correlated experimentally with the insect's stylet penetration (probing) activities and stylet tip positions in the plant tissues and cells.

 

 Fig. 1. The primary EPG circuit

2. The complete experimental setup

In the complete experimental setup the pre-amplifier is incorporated in a separate unit, the EPG probe. The probe is the most sensitive part of the setup and located close to the insect in a Faraday cage to shielding it from external electro-magnetic noise, mainly coming from 110 or 220VAC power cables at 60 or 50Hz. It comprises the input (measuring point) and the first stage 50x amplifier. The plant and insect should be located in the Faraday cage too. The probe is enveloped by conductive material that is grounded as the Fararday cage is as well. The control box can stay outside the cage.

Fig. 2. Experimental EPG configuration. Only one of the 8 probes shown

The newest Giga-8dd 2020 features are specified in availability.pdf in free the manuals for hard and software on the Downloads page.

 

3. The electrical origin or EPG signal sources

The fluctuating voltages originate from two different physical sources in the insect-plant combination:  1) 'generated' voltages, called electromotive force (emf) and 2) fluctuating electrical resistance (R). The signal components of these sources are referred to as emf- and R-components, respectively. The R-components are mainly due to 'valve' activities and positions in the food and salivary stylet canals and resistance changes at the stylet tips inside the plant. The emf-components originate mainly from 'membrane potentials' of plant cells when punctured by the stylet tips and also, from 'streaming potentials' caused by the fluid movements in the two capillary stylet canals. Muscle and neural potentials in the insect do to contribute to the EPG signal. Both, R- and emf-components include biological information on the insect's activities and the stylet tip positions in the plant. Both, R and emf components are measured as voltage fluctuations. The insect resistance fluctuations modulate the stable DC voltage in the primary voltage as well as the amplitude of the emf component voltage, which makes the EPG signal not a simple sum of the two components. The two components concurrently occur and are mixed at the measuring point in the circuit.

 

4. Different EPG measuring systems

The first EPG system - called feeding monitor - was introduced by McLean and Kinsey (1964) supplying an alternating current (AC) voltage source (Vs). This source was in fact the carrier wave of which the amplitude was modulated by resistance fluctuations of the insect. This fluctuating amplitude was processed in the same way as AM radio signals. The demodulated signal reflected exactly the resistance fluctuations of the insect. This first monitor has been called an AC system but as this AM signal was only sensitive to resistance fluctuations it can also be referred to as an 'R-EPG system'. Later, using a different system design with a DC voltage supply it appeared that the EPG signal also contains electromotive force (emf) originated components, i.e. fluctuating voltages at the measuring point that are generated by the insect activities and active electrical components in the plant, mainly cell membrane potentials (Tjallingii, 1978, 1988). Similar to the carrier wave amplitude modulation in the R-EPG system here the DC voltage was amplitude modulated. In this EPG system design thus there is a DC AM signal and an emf originated signal. This system was called a DC EPG system, but as its signal is sensitive to both R and emf components it can also be referred to as an ´R+emf EPG system. This has become more or less the standard EPG system as shown in this website. In addition to the R-EPG and the standard R+emf EPG systems, a third EPG system has been constructed: which can be referred to as and emf-EPG system; except for the much higher Ri value (with Ri ≥ 1013 Ohm, = 10 TΩ) the design is identical to the standard DC system (Tjallingii, 1988. In the present Giga-8dd version a remote controlled switch the device can switched between normal mode (standard R+emf system) and emf mode (emf EPG sysyem). The much higher input resistor value is making the device insensitive to resistance fluctuations of the insect or plant.

Originally, aphids were mainly used in both systems. The input resistor (Fig. 1, Ri) of the standard DC system has a value of 109 Ohm (1 GΩ) since 1988, This was selected because the aphid average resistance seemed to fluctuate around this 1 GΩ resistance value.

 The newest Giga-8dd (2020) has a digitally operated (remote) switch between the standard Ri of 1 GΩ normal mode and the 10 TΩ for emf mode. In emf-mode the system is especially suitable in plant physiology to measure plant cell membrane potentials and their de- and re-polarizations of stylet punctured plant cells, phloem cells in particular (Salvador-Recatalá et al, 2014).

Summary  There are in fact 3 main EPG system principles: 1) the AC R-EPG system recording R-components only, 2) the DC R+emf EPG system recording both R- and emf-components, and 3) the DC based emf-EPG system recording emf-components only. The standard DC EPG system contains the widest range of biological information.

 

Recently the present DC EPG system, such as the Basic EPG Systems on this website, has increasingly been criticized by Backus et al. Here a summary of their main objections, criticisms, and claims followed a rebuttal (RE:).

1) The Basic EPG system would not supply an accurate DC voltage (Pearson et al., 2014). When the voltage supply (Vs) on the device was set to 0Volt an offset was observed and in older EPG systems (Giga-4, for example) when the voltage supply knob was set to 0Volt  no accurate 0Volt was measured at the voltage supply output socket.

RE: This criticism is correct, but completely irrelevant: In the measuring (primary) circuit, the insect and plant (soil) electrodes represent two metal/electrolyte interfaces, together acting as a galvanic element (battery). These voltages are called electrode potentials. These potentials have an unpredictable value and sign; up to more than 100 MV and positive or negative. They contribute to the DC voltage in the primary circuit level and the voltage supply Vs is used to compensate for them. This can only be done effectively after a recording has started, during one of the first stylet penetrations when the primary circuit is completed, regardless of any set initial Vs value (0Volt according to Pearson et al.). The initial Vs value before the primary circuit is completed is inappropriate thus.

2) For EPG recording of large insects an AC EPG system should be used and for small insects a DC EPG system (Backus et al., 2018) because the AC and DC supplied voltages in these systems would specifically interfere with feeding behavior of insects of these sizes. The underlying property would be the insect size related ´inherent electrical resistance'

RE: Although large insect have a lower average electrical resistance in general, due to their wider capillary stylet canals, this seems no reason as such to the supposed higher sensitivity to DC for large insects or to AC for small insects. There is no theoretical basis for this view. Additionally, the experimental evidence shown by Backus (2018) for this point of view is not convincing: For insects with the AC treatment group the DC voltage supply was set to 0Volt before recording and in the DC treatment group (without AC voltage supply) a DC voltage was set at different voltage levels before recording. This is a wrong experimental procedure because the unpridictable electrode potentials always present in the measuring circuit are not compensated by the DC voltage supply. Thus both a 0 Volt or any other voltage set before recording will be completely overruled by these electrode potentials thus both treatment groups had an unreliable DC voltage seriously interfering with the goal of Backus' (2018) experiments. Moverover the numbers of replicates were very low in these experiments. Thus experimentally and statiscally these results are unreliable.

3) The operational amplifiers (OpAmps) used in the DC EPG system are inferior since these would be 'notoriuos' for drift.

RE: The OpAmps in all DC EPG systems are all high quality and similar to those used in the Backus' device primary circuit. Moreover, if a small drift might occur it will be negligible in relation to electrode potentials and their possible polarization (which is a still not well studied aspect).

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