dihydrofolate reductase renders malaria parasites insensitive toWR99210 but does not affect the intrinsic activity of proguanil. Proc.Natl. Acad. Sci. U. S. A. 94, 10931–10936

30 Walter, R. etal. (1991) Pyrimethamine-resistantPlasmodiumfalciparumlack cross-resistance to methotrexate and 2,4-diamino-5-(substitutedbenzyl) pyrimidines. Parasitol. Res. 77, 346–350

31 Cunningham, R.F. et al. (1981) Clinical pharmacokinetics of probene-cid. Clin. Pharmacokinet. 6, 135–151

1471-4922/$ - see front matter q 2003 Elsevier Ltd. All rights reserved.doi:10.1016/j.pt.2003.12.005

|Letters

Africanized honeybees have unique tolerance toVarroa mites

Stephen J. Martin1 and Luis M. Medina2

1Laboratory of Apiculture and Social Insects, Department of Animal and Plant Sciences, University of Sheffield, Western Bank,

Sheffield, S10 2TN, UK2Departamento de Apicultura, Facultad de Medicina Veterinaria y Zootecnia, Universidad Autonoma de Yucatan,

Apartado Postal 4-116, CP 97100, Merida, Yucatan, Mexico

Varroa destructor is an ectoparasitic mite of the adulthoneybee, which parasitizes the bee brood. This mite haskilled millions of honeybee Apis mellifera colonies, world-wide, eliminating wild populations throughout Europeand North America [1], and resulting in the loss of billionsof dollars in agricultural production. The Africanizedhoneybee (AHB) has a unique tolerance to V. destructorthat is not present in the A. mellifera European honeybee(EHB), from which the AHB hybrid was derived [2]. Thisunexpected tolerance mechanism provides a valuableinsight into the evolution of host–parasite interactions.

Varroa mites feed solely on the hemolymph of honey-bees, and their entire reproductive cycle is completedwithin a sealed honeybee brood cell [1]. In Apis cerana(the original host of Varroa), mite reproduction occurs onlyin the small number of sealed male (drone) honeybee broodcells. Consequently, mite populations within an A. ceranacolony are low (,800) and no adverse effects are seen. InA. mellifera EHB colonies, V. destructor also reproduces inthe much more numerous worker brood cells [1], enablingmite populations to increase up to 2000-fold annually [3],causing colony death within one year [4]. However, mitepopulations in similar-sized AHB colonies stabilize at1000–3000 mites per colony [3,5], allowing colonies tosurvive indefinitely (L.M. Medina, PhD Thesis, Universityof Sheffield, 2003), although the resistance mechanism,until now, has remained elusive.

Varroa kills host colonies indirectly by providing a newtransmission route for a few naturally occurring honeybeeviruses such as the deformed wing virus (DWV) [6,7].Therefore, for a viral epidemic to kill a honeybee colonycomprising 20 000 honeybees, a minimum number ofvectors (mites) are required. The number of mites requiredto sustain a viral epidemic varies with viral virulence andhoneybee longevity because both of these factors affect thenumber of honeybees acting as viral reservoirs [6,7].

The daily changes in mite populations reproducing insimulated A. cerana, AHB and EHB colonies were

investigated using a honeybee–mite simulation model(L.M. Medina, op. cit.) [6]. Mite population growth in EHBsand AHBs is determined largely by the average number ofmated female mite offspring produced in worker broodcells per reproductive cycle (Wr). For the Korean haplotypeof V. destructor reproducing in EHB in the UK and SouthAfrica, the Wr is 0.92, whereas in AHB in Mexico andBrazil, the Wr is 0.73 and 0.64, respectively [8]. Bychanging only the value of Wr, the model generated eithera stable mite population in AHB and A. cerana colonies, oran increasing mite population in EHB colonies [6](Figure 1a). The working hypothesis is therefore, at lowmite populations in all colonies, mites reproducing indrone brood cells, which have a high reproductive success,contributed mainly to the initial mite population growth.However, because mites also show a tenfold preference toreproduce in drone cells (which comprises only 1–5% of allthe honeybee brood), they soon become overcrowded as themite population increases. This leads to inter-mitecompetition for the limited food and space, causing anincrease in mite mortality [9] and resulting in negativereproductive success for mites entering these overcrowdeddrone cells. Thus, mite population growth in drone broodcells is limited by a density-dependent mechanism.Although this occurs in all colony types, it is the normallyoverlooked reproductive ability in worker brood cells thathas the crucial role in determining whether the mitebecomes a pest. In A. cerana (Figure 1b), no mitereproduction occurs in worker brood cells per reproductivecycle (Wr ¼ 0) and the mite population stabilizes at a lowlevel (,800 mites per colony). In AHB (Figure 1c), limitedmite reproduction can occur in worker brood cells(Wr ¼ 0.7) and the mite population stabilizes at a higherlevel (1000–3000 mites). However, in EHB (Figure 1d),mite reproduction in worker brood cells (Wr ¼ 0.9) is morethan sufficient to compensate for losses as a result ofovercrowding in drone brood cells, allowing the mitepopulation to increase until the colony dies. The shortadult longevity of AHB (21 days versus 25–180 days forEHB) as a result of the tropical or sub-tropical climateCorresponding author: Stephen J. Martin (s.j.martin@sheffield.ac.uk).

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Figure 1. Effect of mite fertility in honeybee worker cells on mite population growth. (a) Predicted growth curves for mite populations in Africanized Apis mellifera (AHB)

(Wr ¼ 0.7), European A. mellifera (EHB) (Wr ¼ 0.9) and Apis cerana (Wr ¼ 0) colonies. The daily relative change in mite number in worker (black line) and drone (red line)

brood in A. cerana (b), AHB (c) and EHB (d) colonies are indicated. The gray region represents the period when the overall mite population is increasing. Data obtained

from S.J. Martin and L.M. Medina (2003) Varroa tolerance in Africanized honeybees explained, Abstract no. 274, XXXVIIIth Apimondia International Apicultural Congress,

held 24–29 August 2003 in Ljubljana, Slovenia. Abbreviations: AHB, Africanized honeybee; EHB, European honeybee; Wr, average number per reproductive cycle of mated

female mite offspring produced in worker sealed cells.

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indicates that .12 000 mites are needed to kill an AHBcolony [7]. Therefore, although DWV is present in AHBand A. cerana colonies, mite populations stabilize at levelswell below that required to kill the colony.

It is unlikely that AHB evolved Varroa tolerance afterthe AHB hybrid was created as a result of increasedhygienic behaviour or brood attractiveness [10] becausesuch factors are unlikely to lead to a stabilized mitepopulation. Instead, tolerance has probably resulted frompre-existing resistance characteristics fortuitously comingtogether in the hybrid. That is a high level of mite offspringmortality in worker brood and a short life span in the adulthoneybee.

References

1 Webster, T.C. and Delaplane, K.S. (2001) Mites of the Honey Bee,Dadant publication, Illinois

2 Martin, S.J. and Kryger, P. (2002) Reproduction of Varroa destructor inSouth African honey bees: does cell space influence Varroa malesurvivorship? Apidologie (Celle) 33, 51–61

3 Vandame, R. et al. (2000) Levels of compatibility in a new

host–parasite association: Apis mellifera Varroa jacobsoni. Can.J. Zool. 78, 2037–2044

4 Martin, S.J. et al. (1998) A scientific note on Varroa jacobsoniOudemans and the collapse of Apis mellifera colonies in the UnitedKingdom. Apidologie (Celle) 29, 369–370

5 Medina, L.M. et al. (2002) Reproduction of Varroa destructor in workerbrood of Africanized honey bee (Apis mellifera). Exp. Appl. Acarol. 27,79–88

6 Martin, S.J. (2001) The role of Varroa and viral pathogens in thecollapse of honey bee colonies: a modelling approach. J. Appl. Ecol. 38,1082–1093

7 Sumpter, D. and Martin, S.J. (2004) The dynamics of virus epidemicsin Varroa infested honey bee colonies. J. Anim. Ecol. 73, 51–63

8 Correa-Marques, M.H. et al. (2003) Comparing data on the reproduc-tion of Varroa destructor. Genet. Mol. Res. 2, 1–6

9 Donze, D. and Guerin, P.M. (1997) Time-activity budgets andspace structuring by the different life stages of Varroa jacobsoni incapped brood of the honey bee Apis mellifera. J. Insect Behav. 10,371–393

10 Guzman-Novoa, E. et al. (1999) Susceptibility of European andAfricanized honey bees (Apis mellifera) to Varroa destructor in Mexico.Apidologie (Celle) 30, 173–182

1471-4922/$ - see front matter q 2004 Elsevier Ltd. All rights reserved.doi:10.1016/j.pt.2004.01.001

Free-living endohelminths: the influence of multiplefactors

Neil J. Morley and John W. Lewis

School of Biological Sciences, Royal Holloway, University of London, Egham, Surrey, TW20 0EX, UK

In a recent review by Pietrock and Marcogliese [1], aninterpretation of laboratory data on the survival andinfectivity of free-living stages of endohelminths and theinfluence of environmental factors, especially toxic pollu-tants, was undertaken. Although we agree with theirbasic interpretations, there has, however, been an over-simplification of the significance of such data, and theimportance of biotic factors to free-living endohelminthshas been overlooked.

In our recent review of laboratory and field studies onthe transmission of larval digeneans in polluted conditions[2], we emphasized that the influence of toxicants ondigenean transmission is highly complex, with much of theobserved effects in the laboratory often masked by otherfactors in the field. In particular, the mobility ofvertebrates as target and source hosts for free-livingendohelminths can represent a major factor in prevalence,as demonstrated by Siddall et al., who found that inlaboratory studies the survival of miracidia and cercariaeof Zoogonoides viviparus was reduced in the presence ofsewage sludge [3]. Field studies demonstrated that theprevalence of Z. viviparus in its molluscan intermediatehost had a reduced gradient in parasitism towards asewage dumpsite [4] and appeared to confirm thelaboratory findings. However, the prevalence of Z. viviparusin the definitive fish host, Hippoglossoides platessoides,

revealed no differences between control and polluted sites[5]. This fish is highly mobile, and it was considered thatintermixing within its population masked the pollutioneffects demonstrated in the intermediate host. Conflictingresults are also apparent in some coastal bird–parasitesystems [2], suggesting that caution is necessary whenconsidering the impact of environmental factors on free-living helminth ‘transmission success’ under isolatedlaboratory conditions.

The influence of biotic factors on free-living endohel-minths is as important as abiotic factors. The physiologicalstatus of the host can influence the functional biology offree-living stages. For example, the rate of hatchingof Schistosoma mansoni eggs is related to host age andintensity of adult worm infections [6], whereas boththe survival and infectivity of cercariae are linked to thehealth of the mollusc from which they emerge [7,8]. Thesusceptibility of a target host to infection by free-livingstages is often dependent not only on host age [7,9], butalso by the distribution [10] and density [11] of thetarget host population. Complex interactions can occurin multi-species host communities, with hosts of lowsusceptibility acting as decoys to reduce the prevalence ofinfection in species of high susceptibility [12], and a rangeof invertebrates could interfere with the host-findingprocess [13].

Therefore, all investigators must avoid implying thatsimplified laboratory studies can accurately reflectCorresponding author: Neil J. Morley (n.morley@rhul.ac.uk).

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