Keywords:

Fungus

Black weevil

Musaceae

Ecacy

Control

Biocontrol of Cosmopolites sordidus using entomopathogenic fungi under laboratory conditions,

Ecuador

Biocontrol de Cosmopolites sordidus mediante el uso de hongos entomopatógenos en condiciones

de laboratorio, Ecuador

Biocontrole de Cosmopolites sordidus usando fungos entomopatogênicos em condições de

laboratório, Ecuador

Danilo Ramiro Valdez Rivera

Simón Ezequiel Farah Asang

1,2

Winston Carlos Espinoza Morán

1,2

Freddy Fernando Veliz Piguave

Henry Paul Villon Leoro

Edwin Stalin Hasang Moran

Liliana Herrera Espinoza

Rev. Fac. Agron. (LUZ). 2025, 42(2): e254217

ISSN 2477-9407

DOI: https://doi.org/10.47280/RevFacAgron(LUZ).v42.2.I

Crop production

Associate editor: Dr. Francisco Osorio

Postgraduate College, Veracruz Campus

United Mexican States

Universidad Agraria del Ecuador. Facultad de Ciencias

Agrarias “Dr. Jacobo Bucaram Ortiz”, Av. 25 de Julio y Pio

Jaramillo. Guayaquil-Ecuador. P.O. BOX 09-04-100.

Universidad Agraria del Ecuador. Escuela de Posgrado

“Ing. Jacobo Bucaram Ortiz PhD”

Received: 01-12-2024

Accepted: 27-02-2025

Published: 30-03-2025

Abstract

Cosmopolites sordidus known as the black weevil, is an insect

pest of economic importance for Musaceae crops such as bananas

and plantain, resulting in production losses due to its damage. One of

the alternatives for the management of the black weevil is biological

control using entomopathogens that regulate the insect populations

in Musaceae plantations. The objective of the research was to

determine the action of the entomopathogenic fungi Beauveria

bassiana, Metarhizium anisopliae, and Verticillium lecanii on

adults of the black banana weevil under controlled laboratory

conditions. A completely randomized design was implemented

with 8 treatments with the application of the entomopathogens

individually and in combinations, as well as a control treatment.

The results showed that the treatment with the combination of B.

bassiana + M. anisopliae showed the best control of the insect

pest with 100 % mortality 10 days after inoculation, followed by

B. bassiana with 88 % and 24 % control with M. anisopliae, while

the treatment with the fungus V. lecanii achieved 32 % control

of the black weevil. Entomopathogens are an alternative for the

management of the insect pest of Musaceae C. sordidus.

This scientic publication in digital format is a continuation of the Printed Review: Legal Deposit pp 196802ZU42, ISSN 0378-7818.

Rev. Fac. Agron. (LUZ). 2025, 42(2): e254217 April-June. ISSN 2477-9409.

2-5 |

Resumen

Cosmopolites sordidus conocido como picudo negro, es un

insecto plaga de importancia económica para los cultivos de

musáceas como banano y plátano, teniendo como resultado por su

daño perdidas en producción. Entre unas de las alternativas para el

manejo del picudo negro se encuentran el control biológico mediante

el uso de entomopatógenos que regulen las poblaciones del insecto

en las plantaciones de musáceas. El objetivo de la investigación fue

determinar la acción de los hongos entomopatogenos Beauveria

bassiana, Metarhizium anisopliae y Verticillium lecanii en el control

del picudo negro del banano C. sordidus bajo condiciones controladas

de laboratorio. Se implementó un diseño completamente al azar con

8 tratamientos con la aplicación de los entomopatógenos de forma

individual y en combinaciones, al igual que un tratamiento testigo.

Los resultados mostraron que el tratamiento con la combinación de

B. bassiana + M. anisopliae mostró el mejor control sobre el insecto

plaga con un 100 % en la mortalidad a los 10 días de la inoculación,

seguido de B. bassiana con 88 % y un 24 % en el control con M.

anisopliae, mientras que para el tratamiento con el hongo V. lecanii se

alcanzó un 32 % en el control del picudo negro. Los entomopatógenos

son una alternativa para el manejo del insecto plaga de musáceas C.

sordidus.

Palabras claves: hongos, picudo negro, musáceas, ecacia, control.

Resumo

O Cosmopolites sordidus, conhecido como gorgulho da palmeira-

preta, é um inseto praga de importância econômica para as culturas

de musáceas, como a banana e o plátano, resultando em perdas de

produção devido a seus danos. Uma das alternativas para o manejo do

gorgulho da palmeira-preta é o controle biológico por meio do uso de

entomopatógenos que regulam as populações do inseto nas plantações

de musáceas. O objetivo da pesquisa foi determinar a ação dos fungos

entomopatogênicos Beauveria bassiana, Metarhizium anisopliae e

Verticillium lecanii sobre os adultos do gorgulho da palmeira preta

em condições controladas de laboratório. Um projeto completamente

aleatório com 8 tratamentos foi implementado com a aplicação dos

entomopatógenos individualmente e em combinações, bem como um

tratamento de controle. Os resultados mostraram que o tratamento

com a combinação de B. bassiana + M. anisopliae apresentou o

melhor controle do inseto-praga, com 100 % de mortalidade 10 dias

após a inoculação, seguido por B. bassiana com 88 % e 24 % de

controle com M. anisopliae, enquanto o tratamento com o fungo V.

lecanii obteve 32 % de controle do gorgulho da palmeira preta. Os

entomopatógenos são uma alternativa para o manejo do inseto-praga

C. sordidus

Palavras-chave: fungos, black weevil, musaceae, ecácia, controle.

Introduction

The cultivation of bananas (Musa x paradisiaca L.) in Ecuador

has a transcendence and importance at the socioeconomic level at the

national level, ranking the rst place within the exports of agricultural

products in the country and worldwide occupies the rst place in

exports of Musacea; representing 2 % of the total gross domestic

product (GDP) and 35 % of agricultural GDP in the last six years, and

annual production has ranged between 6,023,390 and 6,583,477 tons

with economic revenues ranging from 2,000 million to 3,600 million

dollars (Zambrano and Gallardo, 2022). Cosmopolites sordidus or

black weevil is considered the most important pest of banana and

plantain cultivation (Tresson et al., 2022). The insect pest causes

damage to the plant and aects plant species of the Musaceae family

(banana, plantain, abacá). It is native to Southeast Asia and has spread

to all countries that export bananas. In the larval stage, it bores into

the corm, damages the root and the vascular system, reducing the

absorption of water and nutrients; and weakening the stability of

the plant: in early attacks, it can cause crop loss (Gold and Tinzaara,

2008).

There are more than 750 species of fungi considered biopesticides

used to control insects, among the most important and commercially

used are Metarhizium anisopliae and Beauveria bassiana. The use of

entomopathogens in agriculture focuses on identifying native strains

with potential commercial use (Hernández et al., 2019; Pacheco et

al., 2019). Studies carried out by Silva et al., (2020), determined the

anity and ecacy of B. bassiana and M. anisopliae for the control of

insect pests in the cultivation of cabbage and lettuce, in an aquaponics

system, where it was evidenced that B. bassiana and M. anisopliae

controlled whiteies and aphids. Likewise, Castro and Martínez.

(2019) provided that in an integrated management scheme for the

control of Trialeurodes vaporariorum, Chrysoperla externa together

with B. bassiana and M. anisopliae can be used in tomato cultivation.

Membang et al. (2021) indicated that the success of bioinsecticides

based on entomopathogenic fungi depends on favorable properties

in three successive stages in the development of entomopathogenic

fungi, conidia germination, mycelial growth and conidiogenesis,

and their response to environmental variables such as the range of

temperatures to which the fungus will be exposed.

Fungi are an important part of the biopesticide market for the

biological control of arthropods. Entomopathogenic fungi produce

spores that can attach to the cuticle of arthropods, germinate,

penetrate, and cause infection. On the other hand, studies demonstrate

another approach to the use of entomopathogenic fungi: the study

of the biocontrol potential of the enzymes produced by these fungi

(Motta and Murcia, 2011). Nowadays, the biological control of

pests has increased since the impact of the application of synthetic

pesticides is negative for both people and the environment, in addition,

these entomopathogenic fungi can produce lytic enzymes, esterase,

oxidoreductase and cytochrome P450, reacting with pesticides and

decomposing into non-toxic substances (Kannan et al., 2024). The

present research aims to determine the action of the entomopathogenic

fungi Beauveria bassiana, Metarhizium anisopliae, and Verticillium

lecanii in the control of the black weevil of the banana C. sordidus

under controlled laboratory conditions

Materials and methods

Study area

This study was conducted in the entomology laboratory of the

INIAP (National Institute of Agricultural Research - South Litoral

Experimental Station) geographical coordinates 2°15’07”S and

79°38’36”W, km 26 via Durán-Tambo, Guayaquil, Ecuador (National

Institute of Meteorology and Hydrology [INAMHI], 2018). The

specimens of live adults of Cosmopolites sordidus were obtained

from the “Angela María” banana farm located in the province of

Guayas, Ecuador. For the bioassay with entomopathogens and their

action on C. sordidus, a completely randomized statistical design was

This scientic publication in digital format is a continuation of the Printed Review: Legal Deposit pp 196802ZU42, ISSN 0378-7818.

Valdez et al. Rev. Fac. Agron. (LUZ). 2025, 42(2): e254217

3-5 |

performed with 8 treatments and 5 replications, with a total of 40

experimental units (Table 1).

Table 1. Treatments with entomopathogens for the control of

Cosmopolites sordidus.

Treatments Combinations

Beauveria bassiana

Metarhizium anisopliae

Verticillium lecanii

Beauveria bassiana + Metarhizium anisopliae

Beauveria bassiana + Verticillium lecanii

Verticillium lecanii + Metarhizium anisopliae

Beauveria bassiana + Verticillium lecanii +

Metarhizium anisopliae

T8 Control

Collection of adults of Cosmopolites sordidus

The adult specimens of C. sordidus were obtained in the eld,

using plastic traps (20 L containers) with two openings in the upper

part of 8 cm wide by 12 cm long. Pieces of pineapples, molasses

(225 cc), and water (75 cc) were used as attractants. Ethological traps

made from pseudostems of harvested banana plants were also used,

the sandwich traps were randomly distributed, placing one every 9

plants at a distance of 9m x 9m (Figure 1), following the methodology

of Farah et al. (2022).

Figure 1. Sandwich-type trap for capturing Cosmopolites

sordidus.

Fungi Beauveria bassiana, Metarhizium anisopliae and

Verticillium lecanii

The entomopathogens for the assay were received from the

Microbiolab laboratory. For the preparation of the solution to be

applied in the treatments, 2 grams for each of the entomopathogens B.

bassiana, M. anisopliae, and V. lecani were weighed on an electronic

balance and mixed or diluted in sterilized water to obtain a solution

of 1x10

spores. mL

-1

Bioassays of entomopathogenic fungi on C. sordidus.

To carry out the bioassays, live insects of C. sordidus in the adult

stage were used, which were previously captured in the traps placed in

the eld, from where they were removed and taken to the entomology

laboratory. The specimens used were those that showed greater

biological activity, guaranteeing their viability during the execution

period of the assay; they were disinfected with 0.5 % sodium

hypochlorite solution applied with an atomizer and subsequently

rinsed with disinfected deionized water. For the assays, 40 Petri dishes

were used to conne the inoculated specimens (Figure 2). Food was

provided with small pieces of banana pseudostems, changing them

every day, and maintaining them at an average temperature of 27

and 75 % relative humidity. In each Petri dish, 5 specimens of C.

sordidus were placed, totaling 25 insects per treatment. These were

inoculated with the entomopathogens using a sprayer or atomizer

with the solution corresponding to each treatment.

Figure 2. Experimental units in control bioassay for Cosmopolites

sordidus.

To evaluate the mortality of the specimens and the eciency

of the treatments, monitoring began after the installation of the

assay on day 6, giving time for the development of the inoculated

microorganism in the host and its entomopathogenic action inside the

insect. Subsequent evaluations were periodic (daily) until the tenth

day with the culmination of the bioassay, completing a total of 5

evaluations. The dead Coleoptera in each evaluation were separated

in Petri dishes individually and placed in wet chambers to observe

the progress of the disease and consequently the sporulation of

the fungus, with this the death of the insect by entomopathogenic

microorganisms was conrmed.

Figure 3. Insect infected with the fungus, Beauveria bassiana +

Metarhizium anisopliae.

Data processing and analysis

For the processing and analysis of the dependent variable

number of dead insects and the independent variable based on

entomopathogenic fungi, the non-parametric analysis of Kruskal

Wallis at 5 % error was used, this was tabulated with the InfoStat

This scientic publication in digital format is a continuation of the Printed Review: Legal Deposit pp 196802ZU42, ISSN 0378-7818.

Rev. Fac. Agron. (LUZ). 2025, 42(2): e254217 April-June. ISSN 2477-9409.

4-5 |

program (2017). It is worth mentioning that the normality and

homoscedasticity tests were signicant (p>0.065).

Results and discussion

Mortality of C. sordidus in the application of entomopathogens

According to the results obtained in the assay of entomopathogens

on the insect pest C. sordidus, the treatment of B. bassiana + M.

anisopliae (T4) showed the highest mortality with a mean of 5.00

specimens on day ten of inoculation, followed by treatment with

B. bassiana (T1) with a mean of 4.4 dead specimens, being the

entomopathogenic activity and mycelial growth higher than in the

other treatments under study, with T4 showing the beginning of C.

sordidus mortality at 6 days after inoculation, and T1 at 8 days (Table

2). In studies carried out by Maharaj and Khan (2017), spray applications

of mineral oil and water as spore carriers of B. bassiana were evaluated

to improve virulence against C. sordidus at concentrations of 1 x 10

1 x 10

, 1 x 10

and 1 x 10

pores.mL

-1

, where the highest

mortality was evidenced in the formulation of mineral oil, and B.

bassiana 1 x 10

pores.mL

-1

from 13 to 16 days after inoculation.

Table 2. Average mortality of Cosmopolites sordidus.

Treatments

Specimen mortality

Day 6 Day 7 Day 8 Day 9 Day 10

B. bassiana+ M. ani-

sopliae

0.4 0.4 2.0b 4.0c 5.0c

T1 B. bassiana 0.0 0.0 1.2b 2.2bc 4.4bc

B. bassiana + V.

lecanii+ M.anisopliae

0.0 0.0 0.0a 0.6a 1.6ab

T3 V. lecanii 0.0 0.0 0.4a 0.8ab 1.6ab

T5 B. bassiana+ V. lecanii 0.0 0.0 0.2ab 0.4ab 1.6a

T2 M. anisopliae 0.0 0.0 0.2a 0.6ab 1.2a

V. lecanii+ M. aniso-

pliae

0.0 0.0 0.0a 0.0a 0.8a

T8 Control 0.0 0.0 0.0a 0.0a 0.0a

p-Value N/A N/A 0.0009 0.0004 0.0003

Mortality percentage

According to the treatments analyzed, it can be pointed out

that T4 and T1, at the concentrations of 1x10

spores.mL

-1

of the

entomopathogens, exerted greater control in adults of C. sordidus,

where T4 obtained 100 % mortality on day 10 after its application,

followed by T1 with 88 % mortality, T3, T7, and T5 treatments

obtained mortality of 32 % of specimens. For the T2 and T6

treatments, there was mortality of 24 % and 16 %, respectively, and

for the control treatment (T8) there was no mortality of specimens 10

days after the installation of the assay in the control of the insect pest

under laboratory conditions (Table 3). These results are consistent

with Membang et al. (2021) who established that the combination

of B. bassiana + M. anisopilae and B. bassiana individually act as

promising biopesticides for commercial use to control C. sordidus

in banana crops. However, in laboratory research conducted by

Armendáriz et al., (2016) the B. bassiana strain was ineective on

adults of C. sordidus, even though Biryol et al. (2021) evaluated

eight isolates of Indigenous fungi, four of B. bassiana and four of

Metarhizium brunneum, with an oil base at maximum concentrations

of 1 x 10

spore.mL

-1

against nymphs and adults of the target pest

Orosanga japonica, causing mortality against nymphs and adults of

97 % under eld conditions, showing greater ecacy than commercial

insecticides.

Table 3. Mortality percentage of Cosmopolites sordidus.

Treatments

Specimen mortality (%)

Day 6 Day 7 Day 8 Day 9 Day 10

B. bassiana+ M.

anisopliae

8.0 8.0 40.0b 80.0c 100.0c

T1 B. bassiana 0.0 0.0 24.0b 44.0bc 88.0bc

T5 B. bassiana+ V. lecanii 0.0 0.0 4.0a 8.0a 32.0ab

B. bassiana + V.

lecanii+ M. anisopliae

0.0 0.0 0.0a 12.0ab 32.0ab

T3 V. lecanii 0.0 0.0 8.0ab 16.0ab 32.0a

M. anisopliae

0.0 0.0 4.0a 12.0ab 24.0a

V. lecanii+ M. aniso-

pliae

0.0 0.0 0.0a 0.0a 16.0a

T8 Control 0.0 0.0 0.0a 0.0a 0.0a

p-Value N/A N/A 0.0009 0.0004 0.0003

Ahmed (2010) conducted an investigation with some strains of

Verticiliun lecanii and M. anisopliae for the control of Sitophilus

zeamais Motsch (Coleoptera) in stored corn kernel, where the

insects treated with a higher concentration of 1 x 10

spores.mL

-1

had

greater control and mortality, compared to the other treatments. The

estimates of dead Coleoptera were veried after application, that is, at

5, 10, and 15 days after infecting with entomopathogenic fungi in the

various treatments, presenting the highest percentage of mortality in

the application of 1x10

of conidia, reaching 1 to 5 dead individuals in

the period of 5 to 10 days, while the other treatments in their dierent

concentrations took longer. However, studies conducted by Kisaakye

et al. (2021), used 8 isolates of B. bassiana ICIPE 273, ICIPE 648,

and ICIPE 660, which were the most lethal, exhibited high levels of

mortality and low median lethal time against the body weight of C.

sordidus in the laboratory.

Regarding the statistical result, the control with the treatment

of B. bassiana obtained 88 % mortality of the black weevil, while

M. anisopliae obtained 24 %; considering that, by joining the two

entomopathogens agents, 100 % control over C. sordidus was

achieved, this considers that the action of entomopathogens when

used separately tends to reduce their percentage of control of the

insect pest. In this sense, Pelizza et al. (2015), provided that the

comparative eects of the insecticides Rynaxypyr, Lufenuron, and

Methoxyfenozide together with B. bassiana and M. anisopliae,

were the ones that produced higher mortalities in the control of

grasshoppers in eld cage conditions. Likewise, González et al.

(2022) determined that the mortality of C. sordidus reached 16 % and

7 %, using M. anisopliae, B. bassiana, separately and in a mixture of

the two entomopathogenic fungi, reached 78 % in 24 days after the

application of the biopesticides.

Conclusions

Regarding the results obtained in the entomopathogenic action

of the microbial agents under analysis, it can be concluded that the

combination of B. bassiana plus M. anisopliae and the treatment based

on B. bassiana are more eective than when applied individually,

This scientic publication in digital format is a continuation of the Printed Review: Legal Deposit pp 196802ZU42, ISSN 0378-7818.

Valdez et al. Rev. Fac. Agron. (LUZ). 2025, 42(2): e254217

5-5 |

having high mortality in adults of C. sordidus, being a viable option

in the management of the pest under study.

The implementation of treatments on adults of C. sordidus did

not cause damage to the insect’s integrity during the initial evaluation

phases, but later on, mycelial growth was subsequently observed

on the insect’s body, showing the colonization of the fungi in the

inoculated specimens.

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