Wilps, H. et al. (1996): Ecotoxicology studies (2) - Summary report on the field research in Mauritania, 1993-199...

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Table 5a. Summary of bioassays with IGRs

Active ingredient and formulation (g a.i./l or kg)	Taxon	Stage	*Max. concentr. per animal or substrate (no of applications )**	Application and exposure	Endpoint	Results	Comments	Rating
Triflumuron	Spiders
a) 50 ULV b) 480 SC	Thanatus sp. Philodromidae)	subadult	a) 25 µg b) > 15 µg (3)	a) topical b) oral	a, b) mortality and moult	a, b) no effects		harmless
a) 50 ULV b) 480 SC	Peucetia viridis (Oxyopidae)	subadult	a) 50 µg b) > 50 µg (7)	a) topical b) oral	a, b) mortality and moult	a, b) no effects		harmless
	Insects
a) 50 ULV b) 480 SC	Coranus arenaceus (Heteroptera: Reduviidae)	adult	a) 25 µg b) > 18 µg (3)	a) topical b) oral	a, b) mortality b) fecundity	a) LD₅₀ = 8.1 µg a.i./bug (95%- CI: 5.7 -11.4 µg) b) reversible reduction in fertility and fecundity with sublethal dose	a) LD₅₀ > E_max (~ 0.25 µg)	moderately toxic
50 ULV	Coranus arenaceus	larval (L4)	25 µg	topical	mortality and moult	• LD₅₀ = 0.3 µg a.i./nymph (95%- CI: < 0.1 - 20.9 µg)	• LD₅₀ > E_max (~ 0.1 µg) • wide 95%- CI, rating therefore provisional	(highly toxic)
50 ULV	Pimelia subquadrata chudeaul (Coleoptera: Tenebrionidae)	adult	> 0.4 µg (25)	oral	mortality	• no effects	• chronic exposure over 25 days	harmless
Teflubenzuron	Spiders
50 ULV	Thanatus sp.	subadult	25 µg	topical	mortality and moult	• no effects
	Insects
50 ULV	Coranus arenaceus	adult	25 µg	topical	mortality	• LD₅₀= 5.8 µg a.i./bug (95%- CI: 1.7 -19.3 µg)	• LD₅₀ > E_max (~ 0.25 µg)	moderately toxic
50 ULV	Coranus arenaceus	larval (L4)	25 µg	topical	mortality and moult	• LD₅₀ = 0.6 µg a.i./nymph (95%- CI: > 0.1 - 31.4 µg)	• LD₅₀ > E_max (~ 0.1 µg) • wide 95%- CI, rating, therefore provisional	(highly toxic)
	P. subquadrata chudeaui	adult	> 0.4 µg (25)	oral	mortality		• chronic exposure (sublethal) over 25 days	harmless
Diflubenzuron	Insects
a) 60 OF b) 250 WP	Pharoscymnus anchorago (Coleoptera: Coccinellidae)	larval (L4)	a) 0.6 µg / cm² b) 2 µg / cm²	a, b) residual**	development to imago	a) only 14% of larvae develop into imagoes b) EC₅₀ = 0.61 µg a.i./ cm² (95%- CI: 0.36 - 0.84 µg) EC₉₅ = 1.62 µg a.i./ cm² (95%- CI: 1.11 - 4.3 µg)	b) EC₅₀ ~ E_max (0.6 µg) EC95 > E_max • Due to the high remanence of diflubenzuron and thus exposure time the hazard in the field can be higher than in the laboratory test	highly toxic

Abbreviations: a.i. = active ingredient; E_max = estimated maximum exposure relative to dorsal surface of the respective organism or surface of the substrate; 95%-CI = 95%-Confidence interval
* multiple treatments of test organisms
** two days' exposure on treated leaves

Table 5b. Summary of bioassays with entomopathogenic fungi

Fungal species (both in ULV-formulation)	Taxon	Stage	Max. concentr. per animal or substrate	Application and exposure	Endpoint	Results	Comments	Rating
Beauveria bassiana	Spiders
(Conidia, strain GHA 1991)	Peucetia viridis	subadult	5 x 10⁵ SP	topical	mortality and infection	• 90% mortality and infection		moderately harmful
	Insects
	Coranus arenaceus	adult	a) 1.25 x 10⁵ SP b) 2.5 x 10⁵ SP/ cm²	a) topical b) residual*	"	a) 100% mortality and infection b) 89% mortality and infection		harmful
	Pimelia subquadrata chudeaui	adult	a) 3.75 x 10⁵ SP b) 2.5 x 10⁵ SP/ cm²	a) topical b) residual**	"	a) 20% mortality and infection b) no effects		harmless
	Trachyderma hispida (Coleoptera: Tenebrionidae)	adult	5 x 10⁵ SP	topical		a) 30% mortality and infection		harmless
Metarhizium flavoviride	insects only
(Conidia, strain IMI 330189)	Cosmopleurus sp. (Heteroptera: Lygaeidae)	adult	1 x 10⁵ SP/ cm²	residual**		• no infection, but high mortality	• acute toxicity of carrier substance (kerosene) at 8 l/ha (= 2 x 10¹³ SP/ha) instead of recommended 2 l/ha (= 5 x 10¹² SP/ha)	harmless
	Hyles livornica (Lepidoptera: Sphingidae)	larval	1 x 10⁵ SP	topical		• no effects		harmless
Metarhizium flavoviride	Spiders
(Blastospores strain DSM 11336)	Thanatus sp.	subadult	1 x 10⁴ SP	topical		• no effects		harmless
	Insects
	Palpares tesselatus (Neuroptera: Myrmeleontidae)	larval	-	indirect^§		• no effects		harmless
	Trachyderma hispida	adult	5 x 10⁵ SP	topical		• no effects		harmless
	Pharoscymnus anchorago	larval (L4)	2.5 x 10⁴ SP/ cm²	residual^§§		• no effects		harmless

Abbreviation: SP = spores; * permanent exposure via treated food or prey ** permanent exposure via treated plant foods
§ two-time exposure via infected prey (S. gregaria), §§ two days' exposure on treated leaves

Table 5c. Summary of bioassays with Melia volkensii

Formulation (g a.i./l)	Taxon	Stage	Max. concentr. per animal or substrate	Application and exposure	Endpoint	Results	Comments	Rating
	Insects only
Ethanolic extract, a.i. content not known precisely	Coranus arenaceus	larval (L4)	5 nl	topical	development to imago	• retardation and arrest of development (age of permanent larvae up to > 15 days) • ED₅₀ = 0.0025 nl a.i./nymph (95%-CI: < 0.001 -0.05 nl)	• ED₅₀ < E_max (~ 0.025 nl) • wide 95%•CI, rating not possible	?
	Pharoscymnus anchorago	larval (L4)	5 µg / cm²	residual*	development to imago	• retardation and arrest of development (age of permanent larvae [Dauerlarvae] up to > 45 days) • EC₅₀ = 1.14 µg a.i./ cm² (95%-CI: 0.44 - 2.38 µg)	• EC₅₀ < E_max (~ 0.1 µg)	moderately toxic

Abbreviation: 95-% CI = 95%-confidence interval
* = two days' exposure on treated leaves

Table 5d. Summary of bioassays with conventional products

Active Ingredient and formulation (g a.i./l)	Taxon	Stage	Max. concentr. per animal or substrate	Application and exposure	Endpoint	*Results (acute toxicity, 96 h )**	Comments	Rating
Profenofos Cypermethrin	Spiders
198 + 22 ULV	Thanatus sp.	subadult	5 µg	topical	mortality	• LD₅₀ = 0.28 µg a.i./spider (95%-CI: 0.16 - 0.5 µg) LD₉₅ = 3.08 µg a.i./spider (95%-CI: 0.79 - 12.07 µg)	• LD₅₀ < E_max (~ 1.1 µg) LD₉₅ > E_max	highly toxic
	Insects
198 + 22 ULV	Coranus arenaceus	adult	5 µg	topical	mortality	LD₅₀ = 0.23 µg a.i./bug (95%-CI: 0.14 - 0.37 µg) LD₉₅ = 1.06 µg a.i./bug (95%-CI: 0.48 - 2.35 µg)	• LD₅₀ < E_max (~ 1.1 µg) LD₉₅ ~ E_max	highly toxic
Fenitrothion + Esfenvalerate	Insects
245 + 5 ULV	P. subquadrata chudeaui a) weight < 0.8 g b) weight > 0.8 g	adult	a, b) 16 µg	topical	mortality	a) LD₅₀ =2.03 µg a.i./beetle (95%- CI:1.11- 2.97 µg) LD95 = 4.06 µg a.i./beetle (95%-CI: 2.82 - 17.41 µg) b) LD₅₀ =3.49 µg a.i./beetle (95%- CI: 1.71 - 5.07 µg) LD95 = 6.96 µg a.i./beetle (95%-CI: 4.84 - 26.72 µg)	a) LD₅₀ < E_max (~ 3.75 µg) LD₉₅ ~ E_max b) LD₅₀ < E_max (~ 4 µg) LD₉₅ > E_max	moderately toxic
245 + 5 ULV	T. hispida a) weight < 1 g b) weight > 1 g	adult	a, b) 32 µg	topical	mortality	a) LD₅₀ = 6.07 µg a.i./beetle (95%-CI: 4.72 - 7.23 µg) b) LD₅₀ = 9.0 µg a.i./beetle (95%- CI: 7.0 -11.33 µg)	a) LD₅₀ > E_max (~ 5 µg) b) LD₅₀ > E_max (~ 6.25 µg) LD₉₅ > E_max • on mild overdose slightly harmful	moderately toxic
245 + 5 ULV	Pharoscymnus anchorago	larval (L4)	6.25 µg / cm²	residual**	mortality	• LC₅₀ = 0.35 µg a.i./ cm² (95%-CI: 0.11 - 0.73 µg) LC₉₅ = 5.72 µg a.i./ cm² (95%-CI: 2.72 -18.75 µg)	• LC₅₀ < E_max (~ 2.5 µg) LC₉₅ > E_max	highly toxic

Abbreviations: a.i. = active ingredient, E_max = estimated maximum exposure relative to the dorsal surface of the respective organism or the surface of the substrate; 95%- CI = 95%-confidence interval
* with P. anchorago after 48 h
** two days' exposure on treated leaves

2.1.3.2 Field tests

Two products were investigated in field tests in the Akjoujt region, triflumuron (Alsystin 050 UL) and M. flavoviride (IMI 330I89). The investigations focused on the terrestrial arthropod fauna, which were collected and examined using pitfall traps and direct methods of collection.

Methods and risk classification

As a rule, the highest recommended dosage is tested in field tests. In Mauritania a different approach was taken, which involved testing one low and one high dosage for each product. In the case of triflumuron, the dosages were 25 and 100 g a.i./ha. The first value represents the lower limit of the recommended field rates, whereas the second represents a "worst case scenario". M. flavoviride was also tested in a single (5 x 10¹² conidiospores/ha) and a quadruple dose (2 x 10¹³ conidiospores/ha). The plot size was I to 2 ha. These were isolated islands of vegetation, covered with relatively dense stands of annual plants such as S. thebaica, Fagonia olivieri and Hyoscyamus muticus (degree of coverage 40 - 90%), and surrounded by largely vegetation-free sandy or stony desert. The natural isolation of these microbiotopes prevents a rapid exchange of fauna with other biotopes, at least with regard to terrestrial arthropods incapable of flight. The duration of the tests was approximately 6 weeks (2 weeks before and 4 weeks after application). During this period, the arthropod fauna were examined at intervals of two days. The effects were calculated by comparing the relative abundance of the various taxa before and after application with the relative abundance of the same taxa in control plots which were also under investigation at the same time. A distinction was made here between acute effects immediately after application (approximately 10 days) and late effects (> 10 days).

Risk classification was performed on an analogous basis to the laboratory trials, using a four-point scale as proposed inter alia by the BBA (1992):

Risk class	Effect*	Rating
I	< 25 %	harmless
II	> 25 %	slightly harmful
III	> 50 %	moderately harmful
IV	> 75 %	harmful

* e.g. mortality, reduction in the parasitisation or predation rate, etc.

Compared to the scale for evaluation of the field tests, the boundaries of the risk classes here have been shifted downward by a considerable margin, as exposure in laboratory trials is as a rule considerably higher than under field conditions. It should be noted in this context, however, that field trials are also not evaluated on a standardised basis. In addition, methodological and biological factors make it more difficult to clearly distinguish between coincidental and insecticide-induced effects, in which case the effects can be either positive (increase in relative abundance) or negative (reduction).

Results

The results of the field trials are summarised in Table 5e. The various arthropods were allocated to risk classes II - IV only when the relative increase in abundance was > 25% and statistically significant. Non-significant reductions > 25%, as well as significant and non-significant increases, were interpreted as risk class I (harmless).

I. Triflumuron
Cicadellids were affected severely by triflumuron at both low and high doses (class IV). The effect was especially pronounced during the later phase of the trial. The delayed effect indicated that predominantly nymphs were exposed and - like the similarly hemimetabolous locusts - died during the next moult. At 100 g a.i./ha the spider Thanatus sp. was also severely reduced. Further investigations showed that juvenile spiders in particular were affected, and that they were affected less by the active ingredient itself than by residues of the formulation agents of low volatility. In addition, slight hazards (class II) were observed in calliphorids (100 g a.i./ha) which was, however, interpreted as a chance result. Caterpillars of Heliothis peltigera sustained moderate hazard (class III) at the higher dosage. Since IGRs are applied primarily to control lepidopteran pests, this effect had been anticipated. All other taxa remained unaffected by triflumuron.

II. Metarhizium flavoviride
Apart from beetles (class II), no significant side-effects of the fungal preparation were observed. Since a significant reduction in abundance was recorded only with the single, but not with the quadruple dosage, it cannot be concluded that the effect was an insecticidal one. One striking observation was the extraordinarily sharp increase in flying insects (Diptera, Hymenoptera) in the treated plots. It was not possible to establish whether the large number of animals caught was due to hyperactivity, hatching or immigration. The one observation crucial for the evaluation, however, was the fact that there was no significant reduction in abundance.

III. Comparative evaluation of the two control agents
For both preparations, a single dose produced minor side-effects on terrestrial arthropods. Whilst cicadellids were severely affected by triflumuron, this family in the traditional sense is in fact considered more a target than a non-target organism. The same applies to H. peltigera. On the other hand, the application of triflumuron must be assumed to pose a threat in principle to all butterfly fauna. Application of 100g a.i./ha triflumuron also produced severe harmful side-effects on the spider Thanatus sp., which is numbered among the beneficial fauna. Since fewer toxic solvents are now being used in more recent ULV formulations, however, there need be no reservations concerning the use of this product in locust breeding areas away from stagnant waters. It should be emphasised that the applications for entomopathogenic fungi and IGRs are entirely different. Whilst the former are appropriate essentially as biological contact insecticides for direct control of nymphs - and thus compete with the acutely toxic carbamate, organophosphate and pyrethroid insecticides - the latter are appropriate for barrier application within the scope of preventive control.

Table 5e. Side-effects of triflumuron and M. flavoviride on arthropods: summary of the results of 4 field trials (frequent taxa only).

Triflumuron

Metarhizium flavoviride

Taxa

25 g a.i./ha

100 g a.i./ha

5 x 10¹² spores/ha

2 x 10¹³ spores/ha

Effect/risk class

E (%), acute

E (%), late

Risk class

E (%), acute

E (%), late

Risk class

E (%), acute

E (%), late

Risk class

E (%), acute

E (%), late

Risk class

Spiders: Thanatus sp.

+ 7

-51**

-113**

n.a.

Insects:

Homoptera, Cicadellidae

-20*

-94**

-26*

-76**

-18**

-22*

-19

Heteroptera, Reduviidae: Coranus aranaceus

-25

-27

-18

-66

n.a.

Coleoptera

-20

-32

-21

-36*

+13

-10

Coleoptera, Tenebrionidae

-17

-1

-48

-11

-4

-1

Tenebrionidae: Mesostena angustata

-14

-22

-5

-81

n.a.

Tenebrionidae: Zophosis quadrilineata

+13

-1

+39

n.a.

Coleoptera, Anthicidae: Notoxus sp.

-29

-36

n.a.

-54

-13

+41

-24

Hymenoptera, Apoidea

-1

-16

-2

-28

+10

Hymenoptera, Sphecidae

+27*

-36

+44

-43

+206*

+445*

+46*

Hymenoptera, Formicidae

-10

-18

-1

+23

n.a.

Hymenopt., Form.: Monomorium arenophilum

n.a.

+36

-15

-9

Diptera

-6

+26

-13

+111**

+22*

+95*

Diptera, Calliphoridae

-22

-9

-2

-37*

n.a.

Diptera, Tachinidae

n.a.

+ 6

+332*

+14

+145

Lepidopt., Noctuidae: Heliothis peltigera (larv.)

-33

+19

-25*

-61**

III

n.a.

Microlepidoptera (adult)

n.a.

-20

+32

-9

Abbreviations: a.i. = active ingredient; n.a. = not available (taxa which either were not distinguished or which occurred only in small numbers); E = effect (= relative increase/decrease in abundance after treatment); level of significance: p < 0.05 (*) and p < 0.01 (**); See text for risk classes

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