Larval Mortality Factors of Spodoptera Littoralis in the Azores

Larval mortality factors of Spodoptera littoralis

in the Azores

Tiago MARTINS, Luısa OLIVEIRA* and Patrıcia GARCIADepartamento de Biologia, Universidade dos Acores, Rua da Mae de Deus, 9501-801Ponta Delgada, S. Miguel-Acores, Portugal

*Author for correspondence; e-mail: [email protected]

Received 27 May 2004; accepted in revised form 16 December 2004

Abstract. Mortality among larval developmental stages of Spodoptera littoralis (Bois-

duval) (Lepidoptera: Noctuidae), was determined by weekly sampling on weeds in apasture on Sao Miguel Island (Azores, Portugal), from August to December, over a3-year period (1999–2001). In all the years surveyed, larvae of S. littoralis usually

appeared in pastures after the third week of August, with higher abundances inSeptember and the beginning of October. Three different factors causing larvalmortality were identified: one fungal pathogen, Furia virescens (Thaxter) Humber

(Zygomycetes: Entomophthoraceae), two nucleopolyhedroviruses and one larval par-asitoid, Meteorus communis (Cresson) (Hymenoptera: Braconidae). The percentages ofdead larvae infected by virus or fungus were significantly higher than the other causes ofmortality, regardless of the year. Furthermore, the percentage of larvae that died due to

virus contamination was generally higher than the percentage of larvae infected byfungus. Significant correlations between the environmental factors and the percentage oflarvae infected by virus or by fungus, were only observed during 2001. In 2001, the

prevalence of fungal infection was negatively correlated with that of viral infectionalthough prevalences of these two agents were positively correlated in both 1999 and2000. These results show that virus and fungus are potential biological control agents

for S. littoralis in Azores.

Key words: biological control, Entomophthorales, Meteorus communis, nucleopolyhe-

drovirus, natural enemies

Introduction

Spodoptera littoralis (Boisduval) (Lepidoptera: Noctuidae) is a seriouspest of several important crops, such as cotton, tobacco and corn inMediterranean and Asian countries (Balachowsky, 1972; Sneh et al.,1981; Sannino et al., 1996). In Azores islands (Portugal), S. littoralis hasbeen known since the first half of this century, but only during these last

BioControl (2005) 50:761–770 � Springer 2005DOI 10.1007/s10526-004-7731-4

years has it been considered as a pest in tobacco, sugar beet and corn(Carneiro, 1979; Carvalho et al., 1999). High larval densities have beenobserved in pastures, especially in Trifolium sp., Leodonton sp., andBromus spp., and in weeds, such as Rumex sp., Amaranthus sp., Daturastramonium and Solanum nigrum (Martins, 2000). The larval infestationsoccur mainly in summer and early autumn, rarely in winter and spring,causing considerable ecological and economic impacts (Martins, 2000).

Numerous natural enemies of S. littoralis are known in different partsof theworld and canbe classified into three groups: predators, parasitoids,and entomopathogens. Three groups represent predators of S. littoralis:Neuroptera (Chysopa sp. and Chrysoperla carnea), Hemiptera (Orius sp.and Deraeocoris pallens) and Coleoptera (Coccinella sp.) (El-Wakeil,1997; Ghavami et al., 1998). The following hymenopterous parasitoidshave been reported from S. littoralis: Chelonus inanitus, Microplitis rufi-ventris, Telenomus remus and Meteorus sp. (Hegazi et al., 1988; Gross-niklaus-Burgin et al., 1994; El-Wakeil, 1997; Hegazi et al., 1997).Regarding the entomopathogens, some strains of Bacillus thuringiensisare referred to be as potential microbial control agents of larvae of S.littoralis (Sneh et al., 1981;Kalfon andBarjac, 1985;Hassani et al., 1998),as well as the fungus Nomuraea rileyi (Ignoffo 1981), a nucleopolyhe-drovirus (NPV) (Santiago-Alvarez andOsuna, 1985; Sannino et al., 1996)and nematodes of the family Steinernematidae (Abbas and Saleh, 1998).However, nothing is known of the presence of natural enemies of S. lit-toralis in theAzores Islands. To improve a biological control programme,a survey for both the pest and its natural enemy population densitiesshould be performed (Gullan and Cranston, 1994), and their relationshipwith climatic parameters should be analysed.

The purposes of this work were to evaluate the abundance S. litto-ralis larvae during a three-year period (1999–2001), in one pasture onSao Miguel Island, and to determine the most important larval mor-tality factors (fungi, viruses or parasitoids) that could potentially beused in biological control programmes.

Material and methods

Sampling for larval abundance

Our study was performed at Sao Miguel Island, Azores, Portugal(37�30¢ N and 25�30¢ W). Since S. littoralis is generally found at lowaltitude, between 50 and 150 m (Martins, 2000), larvae were sampledweekly in the same pasture (±1 ha), located at Relva (100 m altitude).

TIAGO MARTINS ET AL.762

Sampling larval populations of S. littoralis was carried out during theyears 1999, 2000 and 2001. The number of larvae was assessed weekly intwenty 0.25 m2 sample areas, randomly distributed in the pasture,throughout the seasons and across years.

Larval mortality factors

To determine causes of mortality, 20–110 S. littoralis larvae were col-lected each week, in the above-mentioned pasture. A total of 489, 1613and 1659 larvae were collected from the field in 1999, 2000 and 2001,respectively. Larvae were hand picked using a broad point forceps. Eachcollected larva was immediately isolated in a Petri dish and brought tothe laboratory to be reared. Larvae were fed with small portions (1 cm3)of artificial diet (Poitout and Bues, 1970; modified by Oliveira, 1991),renewed every two days for the first 4 instars and every day for the 5thand 6th instars or until larvae died.

When a larva died, the cause of death was then determined, checkingand quantifying the occurrence of fungus, virus or parasitoids. Larvaewere considered as naturally infected by fungus or virus if deathoccurred within a time frame of 12 or 7 days, respectively, after beingcollected from the field. Dead larvae suspected of being infected withfungus were examined with a dissecting microscope to describe the grossmorphology of the fungus on the host. Discharge of conidia or thepresence of resting spores within the cadaver was used to diagnose aninfection by Entomophthorales. Dead larvae infected with the funguswere light to medium grey in colour, and primary conidia were forciblydischarged from a dense fungal hymenium which covered the cadaver.This characteristic is typical of infections by Entomophthorales, whichwith the sole exception of the genus Massospora are distinguished fromall other Zygomycotina by the presence of forcibly discharged conidia(King and Humber, 1981). Resting spores, when produced, wereembedded within cadavers. The presence of virus within the larvae wasdetermined through the observation of the fat body using a phasecontrast light microscope. Larvae having fat body with polyhedralocclusion bodies were considered as infected by baculovirus (Evans andShapiro, 1997). Parasitoids were identified following their emergencefrom the host larvae, using morphology.

Weather variables

Meteorological data were obtained from the agro-climatic stationslocated at Relva (50 m altitude), provided by the Institute of

LARVAL MORTALITY FACTORS OF S. LITTORALIS 763

Meteorology/Azores. Calendar week averages for temperature, relativehumidity and precipitation, resulting from 12:00 GMT daily readings,were used.

Statistical analysis

Data and larval abundance for each year were transformed by�(x + 0.5) and compared by an analysis of variance (ANOVA), fol-lowed by a LSD test at the p £ 0.05 level (Zar, 1996). Data for all causesof mortality, for each year, were transformed by arcsine�(x), andcompared using a 2-way ANOVA (with year and mortality factors asmain effects), followed by LSD tests at the p £ 0.05 level (Zar, 1996).Pearson correlation analyses were also performed among larval mor-tality caused by each factor (weekly percentages) and the weathervariables (weekly averages), for each year. All analyses were performedusing SPSS 10.0 Windows (SPSS Inc., 1999).

Results

Larval abundance

In all the years surveyed, larvae of S. littoralis usually appeared inpastures after the third week of August, with higher abundances inSeptember and the beginning of October. Afterwards, the populationdecreased until the end of December (Figure 1). During the period ofsampling over the 3 years studied, S. littoralis had two peaks of larvalabundance, which probably correspond to two generations (Figure 1).

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Figure 1. Spodoptera littoralis larval density per 5m2 at Relva (mean number of larvaeeach week), from August to December, during the years 1999–2001.


Larval abundance was significantly higher (F ¼ 3.884, df ¼ 2, 57,p ¼ 0.026) during 2001, compared to 1999 and 2000.

Larval mortality factors

During this study we identified three different causes of mortality inlarvae of S. littoralis: one fungus, identified as Furia virescens (Thaxter)Humber (Zygomycetes: Entomophthoraceae), two nucleopolyhedrovi-ruses, which cause distinct types of pathologies in infected larvae,(T. Martins, unpubl.data) and the parasitoid Meteorus communis(Hymenoptera: Braconidae). A low number of larvae died by a nonidentified cause (Table 1).

Analyses of larval mortality data for each year show a significantdifference between years (F ¼ 12.17, df ¼ 2, 213, p < 0.01, 2-wayANOVA) and between causes of larval mortality (F ¼ 51.61, df ¼ 3,212; p < 0.01, 2-way ANOVA). The interaction between these twofactors was also significant (F ¼ 3.96, df ¼ 6, 209, p < 0.01, 2-wayANOVA). The LSD tests revealed that the larval mortality in 1999 wassignificantly less than in 2000 (p < 0.05) and 2001 (p < 0.05). Thisresult could be due to the significantly lower prevalence of fungalinfections during 1999 (F ¼ 12.88, df ¼ 2, 51, p < 0.001, 1-wayANOVA), when compared to the other years. Overall, the percentage oflarvae that died due to virus infection was significantly higher than othercases of mortality, followed by the percentage of larvae that died due tofungal infections (p < 0.001, LSD-tests). Larval mortality caused byM.communis was very low, less than 1% per year, and similar to the per-centage of larvae that died due to unidentified causes during the threeyears studied (p ¼ 0.77, LSD-test) (Table 1, Figure 2a, b and c).

Table 1. Percentages (mean ± SD) of Spodoptera littoralis larvae dying (infected by

fungus or virus, or parasitized by Meteorus communis) or healthy, observed during theyears 1999–2001

Cause of dead 1999 2000 2001

Fungus 2.50 ± 5.17 14.80 ± 16.44 28.15 ± 24.54

Virus 19.83 ± 25.30 29.83 ± 20.31 24.02 ± 18.21

Parasitoids 0.09 ± 0.37 0.67 ± 0.89 0.21 ± 0.41

Not identified 0.00 ± 0.00 0.44 ± 0.66 1.01 ± 1.01

Healthy 77.58 ± 28.93 54.27 ± 32.19 46.62 ± 22.85

n 489 1613 1650

n = total number of larvae collected.


When climatic factors were analysed, no significant differences werefound among years for each environmental parameter (p > 0.05, 1-wayANOVA). Nevertheless, in 2001, precipitation was generally higher thanin the other years, from August to October and in December (Figure 3),while the other climatic factors had similar patterns.

A significant positive correlation was found between the percentageof dead larvae infected by fungus and the relative humidity (R2 ¼ 0.51,df ¼ 34, p ¼ 0.03) and precipitation (R2 ¼ 0.54, df ¼ 34, p ¼ 0.02)during 2001. The percentage of larvae infected with virus was negativelycorrelated with precipitation (R2 ¼ )0.53, df ¼ 34, p ¼ 0.02), relativehumidity (R2 ¼ )0.56, df ¼ 34, p ¼ 0.02) and temperature (R2 ¼ )0.79,df ¼ 34, p < 0.01) during 2001.

The percent fungal infection was positively correlated with percentviral infection during 1999 (R2 ¼ 0.63, df ¼ 34, p ¼ 0.005) and 2000(R2 ¼ 0.52, df ¼ 34, P ¼ 0.027), while a negative correlation betweenthese mortality factors was observed for 2001 (R2 ¼ )0.48, df ¼ 34,p ¼ 0.045).

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Figure 2. Variation in the prevalence of virus, fungus and parasitoids over the years1999 (a), 2000 (b) and 2001 (c).


Discussion

According to Martins (2000), the theoretical number of S. littoralisgenerations using temperature data from Relva and the number ofdegree-days necessary for development from egg to adult, is four gen-erations per year. According to our results, S. littoralis can have twogenerations at Relva, between the second half of August and the end ofDecember. Higher larval abundances were observed in September andbeginning of October in pastures, especially on weeds.

Results show that the percentages of dead larvae infected by virus orfungus were significantly higher than the other mortality causes,regardless of the year. Furthermore, the percentage of larvae that dieddue to virus infection was generally higher than the percentage of larvaeinfected by fungus. However, in the year 2001, there was an increase inthe number of S. littoralis larvae infected by fungus, especially betweenAugust and the beginning of October and during December. The per-centages of larvae killed by fungus in this year was correlated withprecipitation, confirming that fungus infections can be most successfulwhen humidity (75–92%) and/or precipitation (5–15 l/m2) are high, asexpected for such organisms (Hajek and Soper, 1992; Weseloh andAndreadis, 1992; Smitley et al., 1995; Lacey et al., 2001). In addition,only in the year 2001 these two mortality factors peaked at differenttimes, with higher virus infections overlapping the periods of lowerprecipitation (i.e., between October and the beginning of December).

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Figure 3. Variation in the precipitation (weekly means) over the years 1999, 2000 and2001.


The levels of larval parasitism were very low, never exceeding 1%.Only one species of parasitoid was detected, the generalistM. communis,that is also found in pastures as a larval parasitoid of Pseudaletia uni-puncta (Lepidoptera: Noctuidae) (Anunciada, 1983; Medeiros, 1996).Medeiros (1996) also found that the level of parasitism of P. unipunctalarvae by M. communis was very low (less than 1%) in Azores.

Our results show that due to the low level of larval parasitism, viraland fungal pathogens seem to be the best prospective biological controlagents for S. littoralis in the Azores. Within this context, several morestudies concerning the evaluation of the suitability of these fungal andviral agents for of control S. littoralis need to be performed.

Acknowledgements

The authors thank Pedro Mata from the Institute of Meteorology/Azores for providing meteorological data. We are grateful to NelsonSimoes and Jorge Medeiros for identification of the NPV and AntonioMartins for the identification of the fungus.

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Larval Mortality Factors of Spodoptera Littoralis in the Azores

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