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Brazilian Journal of Microbiology

versão impressa ISSN 1517-8382

Braz. J. Microbiol. vol.43 no.1 São Paulo jan./mar. 2012 



Winter prevalence of obligate aphid pathogen Pandora neoaphidis mycosis in the host Myzus persicae populations in southern China: modeling description and biocontrol implication



Xiang Zhou

Key Laboratory of Forest Protection, College of Forestry and Biotechnology, Zhejiang Agricultural and Forestry University, Lin'an 311300, People's Republic of China




Pandora neoaphidis overwintering had been investigated by monitoring its prevalence in Myzus persicae populations in open fields. Cabbage plants in field plots were weekly taken after mycosis initiation, to count and examine the living and dead aphids infected by P. neoaphidis. Based on the field data, infection levels (I) varied with field temperature (T), relative humidity (RH) and aphid count (numbers of living aphids per plant, N) over days (D), fitting well to the modified logistic equation I=0.91/[1+exp(8.5+(2.0HTHRH-20.2NI0)D)] (r2=0.897), where HT indicated daily hours of low temperature (<4°C), HRH daily hours of high air humidity (>90% RH) and I0 primary infection level. The model demonstrated the abiotic and biotic factors influencing P. neoaphidis mycosis development in winter, and also verifies the fungal overwintering by infecting available host aphids without a resting stage. Ultimately, P. neoaphidis mycosis reduced 81.4% of aphid populations, presenting great potential for biocontrol.

Key words: Entomopathogenic fungi, Aphid, Epizootic modeling, Entomophthorales, Microbial control.




Green peach aphid (Myzus persicae Sulzer), a worldwide euryphagous pest of crucifer vegetables and ornamental plants, infests cool-season crops such as Brassica oleracea in southern China even in the wintertime (2). Its natural enemy Pandora (=Erynia) neoaphidis (Remaudière & Hennebert) Humber in the order of Entomophthorales widely disseminated via host flight is recorded from over 70 species of aphids on annual and perennial crops and weeds in temperate regions (10, 14). This entomopathogenic fungus actively discharges numerous conidia from mycosed dead aphids to infect nearby hosts and frequently prevails over aphid cohorts in nature, thus P. neoaphidis is considered as a promising agent for aphid control (8, 14, 15).

Many species of entomophthoralean fungi have limited epizootic potential, because of frequently forming resting spores in vivo particularly under harsh surroundings as in winter (16, 23, 24). Resting spores forcibly interrupt infection cycles and stay quiescence for months or years irrespective of host availability (7). P. neoaphidis proves lacking this type of spore, and probably overwinters by either leaving conidia on humid and cool topsoil layers or forming thick-walled conidia or hypha within cadavers in situ or reproducing in alternate hosts (4, 5, 11, 13, 17). Continuously invading available hosts in winter favors controlling aphids on cool-season crops which are widely cultivated in southern China, but whether this is the prior overwintering mode and the detailed performance of P. neoaphidis affecting host population dynamics are unclear for integrated pest management.

Accordingly, the modeling description and analysis on the seasonal occurrence of P. neoaphidis are required for evaluating the fungal potential in controlling aphid populations. But modeling studies on mycosis prevalence of Entomphthorales in the host populations are scarce, especially in semi-natural habitats or open fields, due to the effects of the biotic and environmental factors and their interactions are complicated in natural epizootic development (9, 14, 19). Epizootic outbreak necessitates high relative humidity (> 90% RH) lasting several days, due to fungal sporulation, conidial germination and infection requiring near-saturated air humidity (12, 22). High (> 30°C) or low (< 4°C) temperatures inactivate most Entomophthorales, and in between, temperature significantly influences the span of infection cycle period, e.g., P. neoaphidis-killed aphids appear in 4.2, 6.9 and 13.6 days after maintained under fluctuating summer, autumn and winter temperatures in UK respectively (1, 12). Light regime probably contributes to death of mycosed hosts occurring in late afternoon for mycosis diffusion on humid nights (12), and ultraviolet radiation otherwise shortens fungal survival duration (6). Besides, host-related factors such as host density and other biotic factors including fungal infection level and actions of contaminated predators affect mycosis transmission, due to the efficiency of transmission dependent on contact probability of infectious conidia and healthy hosts (18, 20).

The present study aimed to reveal P. neoaphidis prevalence potential in winter for aphid control. This was achieved by describing the fungal dynamics in M. persicae populations with a modified logistic model based on field data. In the model, the associations between P. neoaphidis epizootic development and the abiotic (field temperature and relative humidity) and biotic (host density and primary infection level) factors were analyzed.



Aphid and plant culture

Myzus persicae colonies were cultured for later field colonization, from the beginning of vigorous apterae collected in fields in September 2007. All the colonies were maintained on cabbage plants in a growth chamber at 25°C under a photoperiod of 12 h light and 12 h dark. Meanwhile, cool-season vegetable crops of cabbage (Brassica oleracea) were planted for infestation of M. persicae in the field experiment. Cabbage seeds germinated on layers of water-saturated filter paper at 20°C and light/dark 12h:12h, and then the seedlings were planted in walk-in 0.5-mm-mesh cages (2´2´2 m) standing in a field of the university campus farm for six weeks before transferred into open field plots.

Aphid infestation for observation

Healthy cabbage plants with four or five leaves were transferred into field plots in 20-cm plant spacing in late October. Four square field plots located separately on the university campus farm (119.729° E, 30.258° N) were prepared as replicates. Successively, five vigorous apterae of M. persicae with visually similar size taken from plants in the chamber were colonized onto each plant in the plots, to form a defined aphid population. The aphid cohorts infesting on the plants were used for monitoring natural Pandora neoaphidis prevalence level. Throughout the experimental period, all plants were grown conventionally, and no insecticide or fungicide was used.

Sampling and counting

P. neoaphidis mycosis seasonally occurs in this experimental planting zone, probably initiated by infected host immigrants or air-borne conidia (3, 14). Sampling started as soon as white-tan dead aphids held tightly to the plants by rhizoids were observed. On each weekly sampling occasion, 12 plants were taken arbitrarily from the four plots (3 per each) and transferred to laboratory for counting living and dead aphids per plant. Temperature and relative humidity (RH) in the field were recorded two-hourly using an electronic hydrothermometer (Zheda Electric Apparatus, Inc., Hangzhou, Zhejiang, China) placed at the ground level.

Examining and evaluating the level of P. neoaphidis infection

All dead aphids on the 12 plants were separately maintained on coverslips for 12 hours at 20°C and 100% RH, for examining mycelial outgrowth and sporulation. To figure out the proportion of P. neoaphidis-killed aphids in cadavers, conidia ejected from mycosed cadavers were microscopically examined, and the identity of P. neoaphidis was confirmed based on the description of Humber (10). Besides, to figure out the P. neoaphidis-infected proportion in living aphids, thirty living aphids per plant (instars 3-4 without symptoms of infection and other diseases) were arbitrarily selected (total 360 capita examined on each sampling), and monitored for six days at 20°C and 12h light:12h dark using leaf discs; cadavers and conidia were examined as above. Leaf discs that need neither leaf changes nor aphid transference during the aphid monitoring period, were prepared by embedding the upside blades of detached cabbage within 1.5% agar in 90-mm Petri dishes and exposing the undersides for aphid raising (3).

Modeling description for winter prevalence of P. neoaphidis

The abiotic/biotic factors of low-temperature duration per day (weekly mean hours of <4°C, denoted as HT hereafter), high-RH (>90% RH) duration per day (HRH), aphid count (number of living aphids per plant, N) and the primary infection level (computed from the first-sampling data, I0) regulating infection level (I) trend over the days (D) after mycosis initiated were investigated. Before modeling analysis, values of HT, HRH, N and D were log (x+1) transformed and those of I0, I were arcsine square-root transformed for reducing heterogeneity of variances. I-D observations were then fitted to a modified logistic equation I=K/[1+exp(a+(b1HTHRH+
b2NII)D)], the binomial b1HTHRH+b2NII presenting the increase rate of I over D, K a theoretically maximal I, a an intercept for the equation. The modeling analysis was conducted using an updated version of DPS software (21).



Field observation

Myzus persicae colonies were well established on plants, living aphid density reaching the peak of average 840 aphids per plant on Dec. 23 and successively declining to the lowest point of 107 aphids per plant on Mar. 17 (Figure 1 a). Sampling started on Dec. 16 when M. persicae killed by Pandora neoaphidis were first observed, six weeks after the colonization by M. persicae. In the subsequent weeks, the peak of numbers of cadavers per plant was 482 recorded on Feb. 10 (Figure 1 a). Weather-wise, daily mean temperature and RH in the field fluctuated from -1.1 to 16.9°C (average 7.2°C) and from 54 to 99.8% RH (average 82.4% RH), respectively. Daily duration (³ 10 hours) of >90% RH occurred in eight weeks and the weekly mean span of <4°C never exceeded 16.3 hours per day (Table 1), these environmental conditions favoring the occurrence and prevalence of P. neoaphidis mycosis.

Assessing Pandora neoaphidis prevalence level in fields

The proportions of cadavers killed by P. neoaphidis mycosis fluctuated in a range of 0.277-0.915, and the infected proportions in living aphids spanned from 0.001 to 0.610 (Figure 1 b). Based on these proportions, the weekly infection level of P. neoaphidis was computed as the sum of fungus-infected and -killed ones divided by the total of living and dead aphids, in a range of 0.024-0.654 (Table 1). Infection level increased slowly during the first four weeks, mostly attributing to the slight initial level of mycosis and environmental conditions. And the level soon over 0.5 meant the fungus-affected M. persicae in excess of mycosis-free ones in populations later. Finally, P. neoaphidis-killed cadavers accounted for 81.4% of the 18,384 cadavers collected, greater than that of dead aphids parasitized by Hymenoptera or killed by other causes, i.e., P. neoaphidis mycosis contributed to 81.4% of the reduction of aphid population irrespective of migrants, and was the key factor acting on local aphid dynamics in winter.

Modeling description

Winter epizootic development in P. neoaphidis-affected M. periscae populations over time after mycosis initiation was well fitted to the abiotic/biotic-factors-modified logistic model I=0.91/[1+exp(8.5+(2.0HTHRH-20.2NI0)D)], (r2=0.897, F3, 10=28.99, P=0.0001), all estimated parameters in modeling significantly (Student's t test: P<0.05 for all parameters). The estimate of K equaled to the value (0.62) of theoretically maximal I, approaching that of 0.65 observed in the fields. The factors and their interaction including field temperature-RH and aphid density-primary infection level shaped the trend of the P. neoaphidis epizootic development, and the rate of I over D was fitted as 2.0HTHRH-20.2NI0. Thus, the resultant model was biologically robust enough to describe the mycosis occurrence of P. neoaphidis in winter quantitatively in relation to the variables considered (Figure 2). Variables we did not measure included the influence of sunlight, host transference and other natural enemies, thus a complete explanation is impossible here, warranting further studies.

Concluding remarks and implications

Based on the results presented above, winter field temperatures in southern China are not consistently low enough (rarely below 0°C) to prevent the activities of P. neoaphidis, and infection continuously occurred in this field experiment, demonstrating the fungus tending to invade available hosts without a resting stage for overwintering. In the previous report (5), winter conditions in Switzerland (temperature constantly below 0°C) killing host pea aphids (Acyrthosiphon pisum) force P. neoaphidis to survive as primary spores in soil (a resting stage), and the fungus infects hosts recurred in spring. The distinct results attributes to the difference of host systems and environmental conditions, and also implies that P. neoaphidis prefers to infect hosts if available, i.e., active infection is the prior strategy of the fungal overwintering. But winter field conditions drastically prolong infection cycle period, four weeks required for infection level increasing from 0.024 to 0.534 in this study compared with 9 days for Neozygites fresenii-infected proportion in Aphis Gossypii rising from 0.12 to 0.75 in summer (1, 12). Considering entomophthoralean epizootics always occurring too late to control pest populations below economic damage levels, mycosis development in winter is still vital for the timing of epizootic outbreaks (4, 14, 18, 20). That is, no pesticide is required to use on cool-season crops in winter, due to maintaining a high infection level of P. neoaphidis in fields favoring pest management in aphid-infested spring, in agreement with suggestions in some studies (1, 15) such as keeping plants in field margins to support fungal survival in alternate nonpest hosts between crop cycles for improving early season multiplication and dispersal to crop aphids.



This study was supported by the grants from Zhejiang Agricultural and Forestry University (2010FR065) and Zhejiang Education Department (Y201119481).



1. Baverstock, J.; Clark, S.J., Pell, J.K. (2008). Effect of seasonal abiotic conditions and field margin habitat on the activity of Pandora neoaphidis inoculum on soil. J. Invertebr. Pathol. 97, 282-290.         [ Links ]

2. Blackman, R.L.; Eastop, V.F. (2001). Aphids on the world's crops. In: Blackman, R.L.; Eastop, V.F. (eds) John Wiley & Sons. Ltd, New York. p. 1-466.         [ Links ]

3. Feng, M.G.; Chen, C.; Chen, B. (2004). Wide dispersal of aphid-pathogenic Entomophthorales among aphids relies upon migratory alates. Environ. Microbiol. 6, 510-516.         [ Links ]

4. Feng, M.G.; Johnson, J.B.; Halbert, S.E. (1991). Natural control of cereal aphids (Homoptera: Aphididae) by entomopathogenic fungi (Zygomycetes: Entomophthorales) and parasitoids (Hymenoptera: Braconidae and Encyrtidae) on irrigated spring wheat in southwestern Idaho. Environ. Entomol. 20, 1699-1710.         [ Links ]

5. Fournier, A.; Widmer, F.; Enkerli, J. (2010). Assessing winter-survival of Pandora neoaphidis in soil with bioassays and molecular approaches. Biol. control 54, 126-134.         [ Links ]

6. Furlong, M.J.; Pell, J.K. (1997). The influence of environmental factors on the persistence of Zoophthora radicans conidia. J. Invertebr. Pathol. 69, 223-233.         [ Links ]

7. Hajek, A.E. (1997). Ecology of terrestrial fungal entomopathogens. In: Jones (ed.) Advances in microbial ecology. Vol15. Plenum press, New York. p.193-249.         [ Links ]

8. Hajek, A.E.; Delalibera, Jr. I. (2010). Fungal pathogens as classical biological control agents against arthropods. BioControl 55, 147-158.         [ Links ]

9. Hesketh, H.; Roy, H.E.; Eilenberg, J.; Pell, J.K.; Hails, R.S. (2010). Challenges in modeling complexity of fungal entomopathogens in semi-natural populations of insects. BioControl 55, 55-73.         [ Links ]

10. Humber, R.A. (1989). Synopsis of a revised classification for the Entomophthorales (Zygomycotina). Mycotaxon 34, 441-460.         [ Links ]

11. McLeod, P.J.; Steinkraus, D.C.; Correll, J.C.; Morelock, T.E. (1998). Prevalence of Erynia neoaphidis (Entomophthorales: Entomophthoraceae) infections of green peach aphid (Homoptera: Aphididae) on spinach in the Arkansas River Valley. Environ. Entomol. 27, 796-800.         [ Links ]

12. Milner, R.J.; Bourne, J. (1983). Influence of temperature and duration of leaf wetness on infection of Acyrthosiphon kondoi with Erynia neoaphidis. Ann. Appl. Biol. 102, 19-27.         [ Links ]

13. Nielsen, C.; Hajek, A.E.; Humber, R.A.; Bresciani, J.; Eilenberg, J. (2003). Soil as an environment for winter survival of aphid-pathogenic Entomophthorales. Biol. Control 28, 92-100.         [ Links ]

14. Pell, J.K.; Eilenberg, J.; Hajek, A.E.; Steinkraus, D.C. (2001). Biology, ecology and pest management potential of Entomophthorales. In: Butt, T.M.; Jackson, C.; Magan, N. (eds), Fungi as Biocontrol Agents. CABI Publishing, Wallingford. p. 71–153.         [ Links ]

15. Pell, J.K.; Hannam, J.J.; Steinkraus, D.C. (2010). Conservation biological control using fungal entomopathogens. BioControl 55, 187-198.         [ Links ]

16. Pilarska, D.; McMaus, M.; Pilarski, P.; Georgiv, G.; Mirchev, P.; Linde, A. (2006). Monitoring the establishment and prevalence of the fungal entomopathogen Entomophaga maimaiga in two Lymantria dispar L. populations in Bulgaria. J. Pest Sci. 79, 63-67.         [ Links ]

17. Scorsetti, A.C.; Humber, R.A.; Garcia, J.J.; López-Lastra, C.C. (2007). Natural occurrence of entomopathogenic fungi (Zygomycetes: Entomophthorales) of aphid (Hemiptera: Aphididae) pests of horticultural crops in Argentina. BioControl 52, 641-655.         [ Links ]

18. Scorsetti, A.C.; Maciá, A.; Steinkraus, D.C.; López-Lastra, C.C. (2010). Prevalence of Pandora neoaphidis (Zygomycetes: Entomophthorales) infecting Nasonovia ribisnigri (Hemiptera: Aphididae) on Lettuce crops in Argentina. Biol. Control 52, 46-50.         [ Links ]

19. Steinkraus, D.C.; Boys, G.O.; Rosenheim, J.A. (2002). Classical biological control of Aphis gossypii (Homoptera: Aphididae) with Neozygites fresenii (Entomophthorales: Neozygitaceae) in California cotton. Biol. Control 25, 297-304.         [ Links ]

20. Steinkraus, D.C. (2006). Factors affecting transmission of fungal pathogens of aphids. J. Invertebr. Pathol. 92, 125-131.         [ Links ]

21. Tang, Q.Y.; Feng, M.G. (2007). DPS Data Processing System: Experimental Design, Statistical Analysis and Data Mining. Science Press, Beijing, China.         [ Links ]

22. Wilding, N. (1969). Effect of humidity on the sporulation of Entomophthora aphidis and E. thaxteriana. T. Brit. Mycol. Soc. 53, 126-130.         [ Links ]

23. Zhang, G.Z.; Feng, M.G.; Chen, C; Ying, S.H. (2007). Opportunism of Conidiobolus obscurus stems from depression of infection in situ to progeny colonies of host alatae as disseminators of the aphid-pathogenic fungus. Environ. Microbiol. 9, 859-868.         [ Links ]

24. Zhou, X.; Feng, M.G. (2010). Improved sporulation of alginate pellets entrapping Pandora nouryi and millet powder and their potential to induce an aphid epizootic in field cages after release. Biol. Control 54, 153-158.         [ Links ]



Submitted: March 31, 2011; Approved: August 15, 2011.



*Corresponding Author. Mailing address: Key Laboratory of Forest Protection, College of Forestry and Biotechnology, Zhejiang Agricultural and Forestry University, Lin'an 311300, People's Republic of China.; E-mail:

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