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Entomophthorales on cereal aphids

6. Virulence of Pandora neoaphidis against Sitobion avenae and Rhopalosiphum padi

6.1 Bioassay methodology

Pathogenicity of an insect pathogen is defined as the ability to produce disease in insects (Lacey, 1997). The proof of pathogenicity is the first step towards studies on virulence. The virulence of an insect pathogen is defined as the quality or property of being virulent (Lacey, 1997). Assessment of the virulence of an insect pathogen requires quantitative studies on dose-response relationships.

In studies of dose-response relationships, the terms LC₅₀ and LD₅₀ are the most common expressions of virulence. LC₅₀ is the concentration of a given insect pathogen required to kill 50 % of the test insect population within a given period of time, whereas LD₅₀ expresses the dose required to kill 50% of the population. With respect to fungi from Entomophthorales, LC₅₀ is the appropriate term since the methodology only allows an estimate of the concentration used and not the dose actually received by the test insects (Papierok & Hajek, 1997).

The term LT₅₀ is defined as the time period required to kill 50 % of the test insect population when subjected to a given concentration or dose of an insect pathogen. The term is often used a quantitative expression of the virulence of fungi from Entomophthorales. The shorter the lethal time documented for a fungus against the test insect, the higher the virulence. Furthermore, the term LT₅₀ provides much of the information needed to understand the dispersal of the disease in the insect population and the dynamics of the host-pathogen system. In several cases, both LC₅₀ and LT₅₀ were measured for a host-pathogen system using a fungus from Entomophthorales as the pathogen and aphids as the test insect (Papierok & Hajek, 1997).

Bioassays with Entomophthorales can be performed in different ways. The most common method is the ‘conidia shower’ method (Papierok & Hajek, 1997, Eilenberg, 1999). The test insects are subjected to a conidial shower (sporulating cadavers or conidia from in vitro cultures). The concentration of conidia is expressed as number of conidia per mm² or similar. Using insect cadavers, less precise expressions such as number of cadavers are sometimes used to express the concentration of conidia. Conidial showers from cadavers or in vitro cultures do in any case reflect the natural route of infection, although studies using Entomophthorales may suffer from limited replicability.

Injections into the insect haemolymph of protoplasts or hyphal bodies have also been used to assess virulence (Papierok & Hajek, 1997). This method gives high replicability but does not reflect the natural route of infection.

6.2 Bioassays against Sitobion avenae and Rhopalosiphum padi

LC₅₀ of P. neoaphidis against S. avenae

Our experimental determination of the LC₅₀ of P. neoaphidis against S. avenae was carried out using an in vitro isolate of P. neoaphidis and the conidia shower method. Cohorts of ten aphids from two clones (green or brown alates or apterous) were subjected to a conidial shower. We varied the time under conidial shower (0, 5, 10, 15, 25, 35, 50, 80, 120, 150 and 180 minutes) to obtain different concentrations of conidia, and the number of conidia per mm² was calculated. Incubation took place at 18^oC and mortality was recorded after seven days.

Data are shown in table 6.1. There was no significant difference between the two clones. For both clones the LC₅₀ values for alates were significantly lower than for apterae.

Table 6.1
Susceptibility of morphs and clones of Sitobion avenae to infection of Pandora neoaphidis (Isolate KVL-634), expressed by LC₅₀ values

LT₅₀ of P. neoaphidis against S. avenae

The experiments to assess LT₅₀ were performed using a similar methods as in the LC₅₀ studies. All test insects were however subjected to conidia for 60 minutes before being placed in different temperatures. Data from the experiments using green alates are shown in figure 6.1.

Figure 6.1 Look here!
Lethal time (LT₅₀) of alates of the green Sitobion avenae clone (H1) infected with in vivo material of Pandora neoaphidis (Isolate KVL-634).

As seen in figure 6.1, lethal time is highly dependent on temperature. At 25^oC, the lethal time is approximately 4 days, while incubation at 5^oC results in a lethal time of 12-14 days.

A comparison of the lethal time at 18^oC for S. avenae subjected to P. neoaphidis is seen in table 6.2. The calculated lethal times varied between 6.6 and 7.5 days between the morphs and clones, though the differences were not significant.

Table 6.2
Lethal time at 18^oC of morphs and clones of Sitobion avenae infected with in vivo material of Pandora neoaphidis (Isolate KVL-634).

Virulence of P. neoaphidis against R. padi

Similar experiments were carried out using R. padi as the receptor. These data are included in table 6.3.

Table 6.3
LT₅₀ data with aphids and Entomophthorales from our experiments and from the literature.

Table 6.3 shows our data on LT₅₀ along with the literature data for P. neoaphidis or other fungi from Entomophthorales and different aphid species. The three fungus species are the most common species on cereal aphids. As can be seen in the table, the generel tendency is that at incubation temperatures above 20^oC the lethal time is 3-5 days, whereas it is much higher at lower temperatures. Our data on S. avenae give precise information on this relationship and also contributes to knowledge concerning the general understanding of aphid/fungus relationships.

6.3 Conclusions

Discussíon and conclusions

The bioassay methodologies used for Entomophthorales were modified in order to obtain precise data on S. avenae and R. padi. The method allowed a comparison between different morphs and clones for the aphids, and the data obtained were satisfactory with respect to reproducibility.

A comparison of our data with the literature data (table 6.3) demonstrated that aphids are quickly killed by fungi from Entomophthorales, but there are differences in the measured LT₅₀ data depending on aphid species, aphid morph, fungus species or even isolate and incubation temperature.

In summary, we may conclude:

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