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

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.

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).

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).

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.

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.