Skip to main content

Advertisement

Egyptian Journal of Biological Pest Control
Egyptian Journal of Biological Pest Control Cover Image

Use of the entomopathogenic fungi Beauveria bassiana (Hyphomycetes: Moniliales) and Isaria fumosorosea (Hypocreales: Cordycipitaceae) to control Diaphorina citri Kuwayama (Hemiptera: Liviidae) under laboratory and semi-field conditions

Egyptian Journal of Biological Pest Control volume 28, Article number: 75 (2018) Cite this article

Article metrics

  • 737 Accesses

  • 1 Citations

Abstract

The Asian citrus psyllid (ACP), Diaphorina citri Kuwayama (Hemiptera: Liviidae), is the vector of the pathogen, Candidatus Liberibacter asiaticus that causes Huanglongbing (HLB) or citrus greening disease, the serious threats to citrus industry worldwide. The objective of this study was to evaluate the role of the entomopathogenic fungi (EPF) Beauveria bassiana (Hyphomycetes: Moniliales) and Isaria fumosorosea (Hypocreales: Cordycipitaceae) in controlling the adult citrus psyllid of D. citri under laboratory and semi-field conditions. Bioassays were performed by exposing the adults to the EPF at the concentration of (1 × 108 conidia/ml). The results showed that both EPF had the potential to control the adult citrus psyllid, giving 64–74% mortality rate in the laboratory and 61–72% under semi-filed conditions. These results support the use of both EPF as effective biopesticides for integrated management of the Asian citrus psyllid, D. citri.

Background

The Asian citrus psyllid (ACP), Diaphorina citri Kuwayama (Hemiptera: Psyllidae), is an economically important insect pest of citrus that disperses throughout the citrus growing regions, including tropical and subtropical Asia, Middle East, Central America, South America, Mexico, and Brazil (Grafton-Cardwell et al. 2006; Hall et al. 2012). Primarily, it attacks young flushes of citrus and can also attack the stressed citrus trees, if the population is high (Hussain et al. 2018). Both nymphs and adult psyllid feed directly on phloem of tissues causing the distorted growth of plants as well as shoot dieback. It is also a vector of the phloem-limited bacteria, Candidatus Liberibacter asiaticus and the causal agent of citrus greening disease known as Huanglongbing (HLB) (Halbert et al. 2000; Bové 2006). HLB is the most deadly disease of Citrus spp. (Sapindales: Rutaceae) because the infected trees usually survive for 5–8 years with a poor fruit quality in both taste and color (Kamble et al. 2017). Citrus psyllids also excrete honeydews on leaf surface that promotes the sooty mold development and impairs the photosynthetic rate of plants (Reynolds 1999). Husain and Nath (1927) recorded the infestation of ACP in Pakistan’s citrus orchards, and now, it has become the most destructive and well-established pest to citrus crop in the country (Mahmood et al. 2014). To date, application of synthetic insecticides is one of the tactics being engaged in psyllid management programs (Naeem et al. 2016), but it requires the repeated applications throughout the crop season (Tiwari et al. 2011).

However, the insecticides’ applications are expected to increase the selection pressure for resistant psyllid population; residual effects on fruits and disruption of the natural enemies associated with the pest (Tiwari et al. 2011). To minimize these risks, there is a need to develop a sustainable management technique for both the vector and HLB disease. Entomopathogenic fungi (EPF) are noticeable and promising tools for effective management of insect pests due to easily mass production as well infecting various life stages of insect host though its cuticle (Subandiyah et al. 2000). To mitigate the resistance problem in ACP and its effective control, the EPF are considered one of the best options as alternatives to synthetic chemicals. The EPF Isaria javanica (Friederichs & Bally) Samson & Hywel-Jones, I. fumosorosea (Wize), Hirsutella citriformis Speare, Lecanicillium lecanii Zimm., Beauveria bassiana (Bals.-Criv.), Cladosporium sp. nr. oxysporum, and Capnodium citri Berk. and Desm. (Subandiyah et al. 2000; Meyer et al. 2007; Stauderman et al. 2012) have been reported earlier as effective bioagents against ACP. Similarly, Metarhizium anisopliae (Metschn.) is also very effective that has a wide range of hosts (de la Cruz et al. 2013).

The objective of this study was to evaluate the efficacy of the EPF, Beauveria bassiana and Isaria fumosorosea, against adult citrus psyllid, D. citri, under laboratory and semi-field conditions.

Materials and methods

Sampling of D. citri

Stem tap samplings of the psyllid population were obtained from two mature Citrus reticulata orchards: the first (5 ha) was selected from Citrus Research Institute (CRI), 32°06′45.0″N 72°40′53.2″E, and the second (2 ha) from research area at College of Agriculture (CA), 32°07′48.5″N 72°41′08.1″E, University of Sargodha, Pakistan. Each orchard was further divided into three blocks, and ten trees were randomly selected from each block. Four shoots (1 from each cardinal direction) were selected from each tree for tap sampling. A white metal pan (20 × 20 × 10 cm) was held under each selected branch and forcefully tapping the branch three times, using PVC pipe. All adult psyllids fallen on the white pan were counted immediately, and the population of ACP was recorded at weekly intervals during March 2018.

Insect colony

Adult citrus psyllids were collected from C. reticulata orchards, using aspirator, and were reared on orange jasmine, Murraya paniculata (L.) seedlings inside ventilated Plexiglas cages (40 × 40 × 40) under the environmental conditions of 23–28 °C, 55–70% RH, and a 14:10 h (L:D) photoperiod. M. paniculata was used as the preferred host for oviposition and feeding of ACP (Chow et al. 2016).

Entomopathogenic fungi

The commercial formulations of EPF Beauveria bassiana NCIM 1216 ATCC 26851 (AgriLife, India) and Isaria fumosorosea IF-171201 (AgriLife, India) were used against ACP in this study. Both EPF were tested at a feasible conidial concentration of (1 × 108 conidia/ml) (Dal Bello et al. 2018). The quality control of the conidia was determined by counting the conidial concentration in a Neubauer chamber, hemacytometer. The conidial germination was determined on PDA, and the conidial viability was measured based on the counts of 200 random conidia per plate 18 h post incubation at 25 ± 2 °C (Ayala-Zermeño et al. 2015).

Laboratory bioassay

Adult psyllids (n = 20; sex ratio 1:1) were collected from the rearing culture, and each treatment was sprayed with 400 μl. The experimental arena was glass vials, containing young seedling of M. paniculata. The control group was treated by distilled water. The mortality data were recorded after 10 days of incubation at 25 ± 2 °C, 65–80% RH, and 14-h photo period. The experiment was performed as a randomized experimental design, with three replications, and the test was repeated three times on different dates.

Semi-field bioassay

The efficacy of EPF against ACP, reared on C. reticulata plants, was assessed under a greenhouse (as semi-field conditions) (Ribeiro et al. 2015). The plants measuring, approximately 60 cm were placed on white plastic trays. Two branches from each plant were covered by mesh cages (net-like fabric bags), and 20 adults of ACP were released on each branch. The infested plants were placed in a greenhouse. Five plants were selected for each treatment, and the experiment was performed under a randomized design, with three replications and the whole trial was repeated three times on different dates. Fungal suspensions of both fungi were prepared as in the laboratory bioassay and were sprayed, at the rate of 15 ml per plant, using a handheld sprayer. The distilled water was applied in the control treatments. Ten days post branches were collected into bags and transferred to the laboratory to record the mortality data. The cadavers were separated and placed in an incubator (25 °C and 90% RH) for 3 days to check the fungal development and sporulation.

Statistical analysis

To determine the status of ACP population, data from tap sampling were analyzed by keeping the block as the main factor for each location. Similarly, the mortality data were analyzed, using one-way ANOVA technique by keeping the treatments as the main factor. Means were separated by the Least Significance Difference (LSD) test. All the analyses were performed using SPSS 20.0 software.

Results and discussion

Stem tap sampling was used to follow the population of ACP in the two different experimental locations of Sargodha district. The results showed that there was a significant (F = 7.72, P = 0.0007) difference in ACP populations sampled from the three blocks in CRI. However, in the second location CA, insignificant difference (F = 1.24, P = 0.294) was found among the three blocks. The highest ACP population averaged 5.42 adults/tap per sample in CRI and 3.85 in CA location (Fig. 1). Survey results from the research sites, carried out during March 2018, clearly indicated the developed population of ACP on C. reticulata, the most growing and economically important cultivar in Pakistan. A large number of young flushes with abundant number of psyllids at both sampled locations were found. There was a significant difference in ACP population in the three blocks of orchard in Citrus Research Institute; however, no difference was found in the orchard at College of Agriculture. It could be due to non-uniformity of young flushes across a block of trees having higher numbers of adult psyllids. The average number of adults per tap sample was (3–5.5 adults/trap) in both sampling sites, during March 2018, indicating the well-established population of the psyllids in these locations. The psyllids’ population has increased in Sargodha region since the last couple of years, and the increase in population was expected with increasing the availability of young flushes for psyllid development and oviposition. Flush growth quality, especially in term of physical characteristics and its nutritional value, had an important influence on the fitness of citrus psyllids (Nava et al. 2007; Tsagkarakis and Rogers 2010; Alves et al. 2014). Citrus mandarin’s fitness effects were more significant for the development of citrus psyllids compared to Rangpur lime (Citrus limonia Osbeck), sour orange (C. aurantium L.), or orange jasmine (Murraya exotica L.) (Nava et al. 2007).

Fig. 1
figure1

Number (means ± SE) of adult psyllids per tap sample per location: CRI (Citrus Research Institute) and CA (College of Agriculture). Means sharing similar letters for each location are insignificantly different at (P > 0.05)

Full size image

The results showed that both B. bassiana and I. fumosorosea were significantly (F = 172, P < 0.001) effective against the adult psyllids by direct contact, resulting in higher mortality reaching 64.4 and 73.8%, respectively, compared to the control (8.3%) under laboratory conditions. A total number of psyllid cadavers, showing fungal development and sporulation, was insignificantly different (F = 0.06, P = 0.834) after B. bassiana and I. fumosorosea applications. B. bassiana caused 56.1% sporulation, while I. fumosorosea caused 57.6% under laboratory conditions (Fig. 2). Similarly at semi-field conditions, both EPF showed significant (F = 1189, P < 0.001) mortality rates of adult psyllids. I. fumosorosea caused higher mortality was 72.1%, followed by B. bassiana that showed 61.2%. In the control treatment, the mortality rate was 5.6%. Similarly, sporulation percentage was also significantly (F = 34.9, P < 0.001) different between B. bassiana and I. fumosorosea. The sporulation percentage was 52.6% in case of I. fumosorosea and 38.4% in B. bassiana application (Fig. 3).

Fig. 2
figure2

Percent (means ± SE) mortality of adult psyllids and sporulation from cadavers after application of Isaria fumosorosea (Ifr) and Beauveria bassiana (BB) under laboratory conditions. Means sharing similar letters are insignificantly different (P > 0.05)

Full size image
Fig. 3
figure3

Percent (means ± SE) mortality of adult psyllids and sporulation from cadavers after application of Isaria fumosorosea (Ifr) and Beauveria bassiana (BB) under semi-field conditions. Means sharing different letters are significantly different at (P < 0.05)

Full size image

Both microbial insecticides have shown encouraging results in controlling adult psyllids. Less number of sporulation under semi-filed conditions could be strongly affected by climatic conditions, temperature, and RH, but interestingly, the mortality level was satisfactory. I. fumosorosea induced (9.4–10.9%) higher mortality rate of psyllid adults under laboratory and semi-field conditions. The efficacy of fungal species against ACP has been reported earlier (Hoy et al. 2010; Ferreira Pinto et al. 2012; Stauderman et al. 2012). The results are in accordance to those of Ferreira Pinto et al. (2012), Lezama-Gutiérrez et al. (2012), and Ausique et al. (2017). The entomopathogens cause the direct effect on insect pests by infection; however, Avery et al. (2011) reported that I. fumosorosea decreased the feeding activity of ACP before their death and led to a reduction in the transmission of the bacterial pathogen responsible for citrus greening disease in citrus. The efficacy of EPF in open field with respect to environmental conditions of Pakistan needs to be studied because the environmental factors have great influence on the development of diseases induced by entomopathogenic fungi. However, Ausique et al. (2017) reported that environmental factors, mainly temperature and RH, strongly affected the sporulation on the fungal-killed citrus psyllids but the mortality level was satisfactory even at the drier conditions of the year.

Conclusions

The microbial pesticides based on the fungi I. fumosorosea and B. bassiana had a potential in controlling the citrus psyllids’ population. Yet, further studies are needed to check their efficacy and compatibility against psyllids under different field climatic conditions of Pakistan.

Abbreviations

ACP:

Asian citrus psyllid

CA:

College of Agriculture

CRI:

Citrus Research Institute

EPF:

Entomopathogenic fungi

HLB:

Huanglongbing disease

References

  1. Alves GR, Diniz AJF, Para JPR (2014) Biology of the Huanglongbing vector Diaphorina citri (Hemiptera: Psyllidae) on different host plants. J Econ Entomol 107:691–696

  2. Ausique JJS, D’Alessandro CP, Conceschi MR, Mascarin GM, Ju’nior I.D. (2017) Efficacy of entomopathogenic fungi against adult Diaphorina citri from laboratory to field applications. J Pest Sci. https://doi.org/10.1007/s10340-017-0846-z

  3. Avery PB, Wekesa VW, Hunter WB, Hall DG, Mckenzie CL, Osborne LS, Powell CA, Rogers ME (2011) Effects of the fungus Isaria fumosorosea (Hypocreales: Cordycipitaceae) on reduced feeding and mortality of the Asian citrus psyllid, Diaphorina citri (Hemiptera: Psyllidae). Biocontrol Sci Tech 21:1065–1078

  4. Ayala-Zermeño MA, Gallou A, Berlanga-Padilla AM, Serna-Domínguez MG, Arredondo-Bernal HC, Montesinos-Matías R (2015) Characterization of entomopathogenic fungi used in the biological control programme of Diaphorina citri in Mexico. Biocontrol Sci Tech 25:1192–1207

  5. Bové JM (2006) Huanglongbing: a destructive, newly emerging, century-old disease of citrus. J Plant Pathol 88:7–37

  6. Chow A, Dunlap CA, Jackson MA, Flores D, Pratt JM, Sétamou M (2016) Oviposition behavior and survival of Tamarixia radiata (Hymenoptera: Eulophidae), an ectoparasitoid of the Asian citrus psyllid, Diaphorina citri (Hemiptera: Liviidae), on hosts exposed to an entomopathogenic fungus, Isaria fumosorosea (Hypocreales: Cordycipitaceae), under laboratory conditions. J Econ Entomol 109:1995–2005

  7. Dal Bello GM, Fusé CB, Pedrini N, Padín SB (2018) Insecticidal efficacy of Beauveria bassiana, diatomaceous earth and fenitrothion against Rhyzopertha dominica and Tribolium castaneum on stored wheat. Int J Pest Manag 64:279–286

  8. de la Cruz T, Cortez-Madrigal MM, Ortiz HGCF, Cappello GS, de la-Cruz PA (2013) Characterization of native isolates of Metarhizium anisopliae and their pathogenicity against Aeneolamia postica in Tabasco. Mex Rev Colomb Entomol 39:40–46

  9. Ferreira Pinto AP, Batista FA, Almeida JEM, Wenzel IM (2012) Patogenicidade de Beauveria bassiana ao psilı’deo Diaphorina citri e compatibilidade do fungo com produtos fitossanita’rios. Pesq Agrop Brasileira 47:1673–1680

  10. Grafton-Cardwell EE, Godfrey KE, Rogers ME, Childers CC, Stansly PA (2006) Asian citrus psyllid. Univ. Calif. Div. Agr. Natl. Res. Publ, Oakland, p 8. Available at: http://anrcatalog.ucdavis.edu/pdf/8205.pdf

  11. Halbert SE, Sun X, Dixon WN (2000) Asian citrus psyllid and citrus greening disease. Citrus Ind 91:22–24

  12. Hall DG, Hentz MG, Meyer JM, Kriss AB, Gottwald TR, Boucias DG (2012) Observations on the entomopathogenic fungus Hirsutella citriformis attacking adult Diaphorina citri (Hemiptera: Psyllidae) in a managed citrus grove. BioControl 57:663–675

  13. Hoy MA, Sing R, Rogers ME (2010) Evaluations of a novel isolate of Isaria fumosorosea for control the Asian citrus psyllid, Diaphorina citri (Hemiptera: Psyllidae). Fla Entomol 93:24–32

  14. Husain MA, Nath D (1927) The citrus psylla (Diaphorina citri Kuw.) (Psyllidae: Homoptera). Memoirs of the Department of Agriculture in India. Entomol Ser 10:5–27

  15. Hussain M, Akutse KS, Lin Y, Chen S, Huang W, Zhang J, Idrees A, Qiu D, Wang L (2018) Susceptibilities of Candidatus Liberibacter asiaticus-infected and noninfected Diaphorina citri to entomopathogenic fungi and their detoxification enzyme activities under different temperatures. MicrobiologyOpen:e607. https://doi.org/10.1002/mbo3.607

  16. Kamble SG, Ghutukade KS, Patil NP, Yamgar SV, Bulbule SV (2017) Responses of different genotypes of citrus to Huanglongbing (citrus greening) under field condition. J Pharmacogn Phytochem 6:207–211

  17. Lezama-Gutiérrez R, Molina-Ochoa J, Chávez-Flores O, Ángel-Sahagún CA, Skoda SR, Reyes-Martínez G, Foster JE (2012) Use of the entomopathogenic fungi Metarhizium anisopliae, Cordyceps bassiana and Isaria fumosorosea to control Diaphorina citri (Hemiptera: Psyllidae) in Persian lime under field conditions. Int J Tropic Insect Sci 32:39–44

  18. Mahmood R, Rehman A, Ahmad M (2014) Prospects of biological control of citrus insect pests in Pakistan. J Agric Res 2:229–244

  19. Meyer JM, Hoy MA, Boucias DG (2007) Morphological and molecular characterization of a Hirsutella species infecting the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae). J Invertebr Pathol 95:101–109

  20. Naeem A, Freed S, Jin FL, Akmal M, Mehmood M (2016) Monitoring of insecticide resistance in Diaphorina citri Kuwayama (Hemiptera: Psyllidae) from citrus groves of Punjab, Pakistan. Crop Prot 86:62–68

  21. Nava DE, Torres MLG, Rodrigues MDL, Bento JMS, Parra JRP (2007) Biology of Diaphorina citri (Hemiptera, Psyllidae) on different hosts and at different temperatures. J Appl Entomol 131:709–715

  22. Reynolds DR (1999) Capnodium citri: the sooty mold fungi comprising the taxon concept. Mycopathologia 148(3):141–147

  23. Ribeiro LDP, Santos MS, Gonçalves GLP, Vendramim JD (2015) Toxicity of an acetogenin-based bioinsecticide against Diaphorina citri (Hemiptera: Liviidae) and its parasitoid Tamarixia radiata (Hymenoptera: Eulophidae). Fla Entomol 98:835–842

  24. Stauderman K, Avery P, Aristizábal L, Steven A (2012) Evaluation of Isaria fumosorosea (Hypocreales: Cordycipitaceae) for control of the Asian citrus psyllid, Diaphorina citri (Hemiptera: Psyllidae). Biocontrol Sci Tech 22(7):747–761

  25. Subandiyah S, Nlkoh N, Sato H, Wagiman F, Tsyuymyu S, Fukatsu T (2000) Isolation and characterization of two entomopathogenic fungi attacking Diaphorina citri (Homoptera, Psylloidea) in Indonesia. Mycoscience 41:509–513

  26. Tiwari S, Mann RS, Rogers ME, Stelinski LL (2011) Insecticide resistance in field populations of Asian citrus psyllid in Florida. Pest Manag Sci 67:1258–1268

  27. Tsagkarakis AE, Rogers ME (2010) Suitability of coleopteran mandarin as a host plant for Diaphorina citri (Hemiptera: Psyllidae). Fla Entomol 93:451–453

Download references

Acknowledgements

The authors are thankful to the authorities of Department of Horticulture, University of Sargodha for providing citrus plants for this experiment.

Availability of data and materials

Data and materials will be shared if needed.

Author information

Affiliations

  1. Department of Entomology, University of Sargodha, Sargodha, 40100, Pakistan

    • Muhammad Irfan Ullah
    • , Muhammad Arshad
    • , Asad Abdullah
    •  & Syed Muhammad Ali Zahid

  2. Department of Environmental Sciences, COMSATS Institute of Information Technology, Vehari, Pakistan

    • Samina Khalid

  3. Department of Plant Pathology, University of Sargodha, Sargodha, 40100, Pakistan

    • Yasir Iftikhar

Authors

  1. Search for Muhammad Irfan Ullah in:

  2. Search for Muhammad Arshad in:

  3. Search for Asad Abdullah in:

  4. Search for Samina Khalid in:

  5. Search for Yasir Iftikhar in:

  6. Search for Syed Muhammad Ali Zahid in:

Contributions

MIU and MA designed and conducted the study. SK and YI helped in reviewing, proof reading, and data analyses. AA and SMAZ helped in data collection and analysis. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Muhammad Irfan Ullah.

Ethics declarations

Ethics approval and consent to participate

Not applicable

Consent for publication

Not applicable

Competing interests

The authors declare that they have no competing interests.

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Ullah, M.I., Arshad, M., Abdullah, A. et al. Use of the entomopathogenic fungi Beauveria bassiana (Hyphomycetes: Moniliales) and Isaria fumosorosea (Hypocreales: Cordycipitaceae) to control Diaphorina citri Kuwayama (Hemiptera: Liviidae) under laboratory and semi-field conditions. Egypt J Biol Pest Control 28, 75 (2018) doi:10.1186/s41938-018-0071-y

Download citation

Keywords

  • Diaphorina citri
  • Citrus
  • Entomopathogenic fungi
  • Microbial control