Primary circuit

In the EPG technique, an aphid (or another insect with piercing mouthparts) and a plant are made part of an electrical circuit by inserting a wire into the soil of a potted plant, and attaching a very thin wire to the insect. The circuit also incorporates an electrical resistor (Ri) and a voltage source (V), as illustrated below. As soon as the aphid stylets penetrate the plant, the circuit is completed and a fluctuating voltage, called the 'EPG signal', occurs at the measuring point which is then 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 these waveforms 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

 

Electrical origin of the signal

The fluctuating voltages oringinate from two different sources in the insect-plant combination:  1) fluctuating electrical resistance (R) and 2) fluctuating 'generated' voltages, called electromotive force (emf). The signal components of the sources are referred to as the R-components and emf-components, respectively. The two components concurrently occur and are superimposed at the measuring point in the circuit. The R-components are mainly due to activities of the valves 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 stylets and 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 important biological information on the insect's activities and the stylet tip positions in the plant.

EPG systems

The first EPG system was introduced by McLean and Kinsey (1964) using an alternating current (60Hz AC) as a voltage source, the voltage amplitude of which was modulated by resistance fluctuations in the insect, similar to signal processing in AM radio circuits. However, this design appeared to be not sensitive to the emf-components in the primary circuit. They were lost in the´AC-EPG system´ and therefore, it is better to indicate it as the 'R-EPG system'. In the later 'DC-EPG system' (Tjallingii, 1978, 1088) the voltage source was replaced by a direct current (DC) source. The input resistor (Fig. 1, Ri) has a value of 109 Ohm (1 GΩ) is selected because the insect's resistance fluctuates around the same resistance value as an average, a ratio of about 1:1 thus. This results in a system sensitivity to changes in resistance as well as to emf changes of the insect and plant in the measuring circuit and the EPG signal. The DC-EPG system is the regular system now (Giga-8d, Products page) and since it records both signal components, R and emf, it is better to indicate this as the 'full-', 'regular-', or 'normal-EPG system'.   

In addition to the R-EPG and normal-EPG systems, a third EPG system has been constructed: the emf-EPG system, based of the DC design too. This system is using a much higher input resistor value (with Ri ≥ 1012 Ohm [≥1 TΩ]*, Tjallingii 1988) making resistance fluctuations of the insect or plant negligible, thus only recording the emf-components. So in summary, there are in fact 3 EPG systems: 1) the full or normal EPG system (DC based) that records both signal components, 2) the R-EPG system (AC based) that only records the R-components, and 3) the emf-EPG system (DC based) that only records the emf-components. The EPG from the normal EPG system is more ´complicated´ but contains the widest range of biological information.

Combined systems Some signal details in the normal-EPG may be hidden (masked): R-components may be masked by emf-components and vice versa. A few details visible in the R-EPG (Jiang and Walker, 2001) or in the emf-EPG (Tjallingii, 1985. 1988, 2000) are not or difficult to distinguish in signals from the normal-EPG but it is has not been investigated so far what biological information these details represent. In order to study these signals details combined systems have been developed. The combined normal-/R-EPG system with two outputs, one for the normal and one for the simultaneous R-EPG has been used, for example, in recording thrips and aphid EPGs (Kindt et al, 2006; Tjallingii et al, 2010). Also, the normal EPG circuit (R+emf) and the emf-EPG circuit have been combined. However, simultaneous recording of the normal and emf-EPG is not possible because the primary circuit can have only one input resistance value at the time, either 1 GΩ or 1TΩ. Therefore, this device - now vailable as the 'probe with emf switch (see below and on Products page 'Other items') - has a switch to alter between normal EPG (R+emf) and emf-EPG mode. In the This emf-mode the system is especially suitable in plant physiological experiments for accurate measurements of membrane potentials and depolarisations in plant cells while punctured by the aphid stylets (Salvador-Recatalá et al, 2014). Another combined device that should be mentioned here is the 'AC-DC correlation monitor' (Backus and Bennett, 2009), which the authors also indicate as the '3rd generation' EPG system. This system not only combines simultaneous normal and R-EPG recording but in addition, several EPG mixtures of normal and R/EPG. However, these mixtures have signals with normal (R + emf) added to R-components: R+emf + R components; a strange over-representation of the R-components thus. Moreover, the electronics of this device produces unreliable signals due to natural offset changes that normally occur during each EPG recording. The offset knob that was added later to this device should be adjust for these natural offset changes again and again, therefore. The authors claim that this device would be suitable for beginners seems completely irrelevant in fact. Only experienced user will be aware of the differences between the signals. This also holds for the selection possibilty of different input resistance values in that device, which will result in signal changes that are complicated to interprete and should only be used by experts familiar with electronics and electrophysiology. Moreover, the analysis of more than one output signal is more time consuming and therefore, not recommended for regular EPG experiments. Thus this 'versatile AC-DC correlation monitor' cannot be recommended and this 3rd generation rather seems a ´lost generation´.

 

The complete experimental setup

 

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


  EPG probe 4 channel AD convertor 8 channel AD convertor Giga-4 Giga-8 The available EPG system is the Giga-8d, a digital controlled device with 8 channels, suitable for simultaneous recording of up to 8 insects. In addition to the Giga-8d control unit the complete system comprises 8 probes, small pre-amplifiers that should be mounted inside a Faraday cage (Fig. 2). 'Giga' refers to the 1 GΩ (109 Ω) input resistance in the probe (see previous text sections). EPG recording should be done in the laboratory using potted plants or detached plant parts. There is no field recording possible with this system*. 

* FUTURE developments: (Feel free to ask for more details)

  •  Field EPG system To record EPGs in the filed a one channel prototype has been tested successfuly; no regular production so far.

  •  EMF system Probes with a switch to make EPG recording suitable for accurate measurements of plant potentials, such as plant potentials (leaf surface potentials, long distance phloem propagated electrical signals, etc. These emf probes can be switched from emf recording to normal EPG recording (this option will become available in the 2017 series).

Technical specifications of the Giga-8d system and operation instructions are available in free manuals for hard and software on the Downloads page.

 

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