Methodology to Test Control Agents and Insecticides Against the Coffee Berry Borer Hypothenemus hampei

Summary

A method using green coffee fruits (GFs) was developed to test the toxicity of insecticides against the coffee berry borer (CBB). Insecticides or toxic substances were applied to disinfected GFs before or after CBB infestation. Insect mortality, repellency, and reproductive capacity, in addition to other parameters, were evaluated.

Abstract

Prior to recommending insecticides to treat the coffee berry borer (CBB) Hypothenemus hampei, it is valuable to know the mortality and repellency of these insecticides against adult insects or their impact on reproductive output. However, currently available methods assess adult mortality only, limiting the selection of novel insecticides with a different mode of action. In this work, different experimental methods were examined to identify the diverse effects on the CBB under laboratory conditions. For this, green coffee fruits (GFs) were collected and disinfected by immersion in sodium hypochlorite solution followed by UV light irradiation. In parallel, CBB adults from a colony were disinfected by immersion in sodium hypochlorite solution. To assess fruit protection (preinfestation), the fruits were placed in plastic boxes, and the insecticides were applied. Then, the CBB adults were released at a rate of two CBBs per GF. The GFs were left under controlled conditions to evaluate CBB infestation and survival after 1, 7, 15, and 21 days. To evaluate insecticide efficacy after CBB infestation (postinfestation), CBB adults were released to the GFs in a 2:1 ratio for 3 h at 21 °C. Infested fruits showing CBB adults with their abdomens partially exposed were selected and placed in 96-well racks, and the CBBs boring into the fruits were treated directly. After 20 days, the fruits were dissected, and the CBB biological stages inside each fruit were recorded. The GFs served as substrates that mimic natural conditions to evaluate toxic, chemical, and biological insecticides against the CBB.

Introduction

The coffee berry borer (CBB), Hypothenemus hampei, was first detected in 1988 in Colombia and has since become the most important pest species of the coffee crop. CBB females leave the natal fruit already fertilized, seeking new fruits guided by the volatile chemicals that they emit^1,2. A complete cycle is fulfilled within 23 days³ at a temperature of 25 °C. The cycle starts with the founder female penetrating the seed and laying eggs in the fruit endosperm. The eclosed larvae eat the seed. If the fruits are dissected at this point, it would be possible to observe both the....

Protocol

NOTE: This protocol addresses different methods to identify different effects on the CBB under laboratory conditions.

1. Fruit collection

1. Pick GFs with a developmental age of ~120-150 days after flowering from trees in a coffee plantation early in the morning.

2. Fruit disinfection²⁰

1. Bring around 300 GFs to the laboratory. Select uniformly sized and healthy GFs and withdraw the peduncles.
2. Dip the GFs into a soap solution (2 mL of liquid dishwashing soap in 998 mL of tap water), followed by rubbing to wash the GFs. T....

Results

The results showed that the CBB females recognized the fruits, and depending on the characteristics of the fruit surface and the emitted odors, the CBB females started to penetrate or bore the fruits within 3 h at 21 °C.

The effect of an insecticide on the CBB when applied to the coffee fruits (preinfestation procedure) after 24 h and over time is shown in Figure 4. The two insecticides (alkaloid emulsion at 5% and 6%) caused high insect mortality on Day 20 (

Discussion

In this protocol, disinfection of the fruits as well as the insects are critical steps. When fruits from the field are used in the laboratory, they frequently show high contamination and dehydration since microorganisms and mites are present in the epidermis^7,15,16. Therefore, using fruits or insects that are not disinfected will cause insect death due to contamination caused by microorganisms, such as bacteria or fungi, thus in.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Beaker with spout, low form 500 mL	BRAND PP	BR87826
Benchtop Shaker	New Brunswick Scientific Innova 4000 Incubator Shaker
Dishwashing liquid soap-AXION	Colgate-Palmolive	AXION
Hood; Horizontal Laminar Flow Station	Terra Universal		Powder-Coated Steel, 1930 mm W x 1118 mm D x 1619 mm H, 120 V (https://www.terrauniversal.com/hood-horizontal-laminar-flow-station-9620-64a.html)
Insects CBB	BIOCAFE		(http://avispitas.blogspot.com/p/biocafe.html).
Multi Fold White paper towels	Familia	73551
Preval Spray unit	Preval Merck	Z365556-1KT	https://www.sigmaaldrich.com/CO/es/product/sigma/z365556?gclid=Cj0KCQiAweaNBhDEARIsAJ 5hwbfZOy1TWGj6huatFtRQt AzOyHe5-oBiKnOUK2T1exuuk WwJLdvxkvsaAjoYEALw_wcB
Reversible Racks 96-Well	heathrowscientific	HEA2345A	https://www.heathrowscientific.com/reversible-racks-96-well-i-hea2345a
Scalpel blades N 11	Merck	S2771-100EA
Scalpel handles N3	Merck	S2896-1EA
Sodium Hypochloride	The clorox company	Clorox
Stereo Microscope	Zeiss	Stemi 508	https://www.zeiss.com/microscopy/int/products/stereo-zoom-microscopes/stemi-508.html