FUNGI ASSOCIATED TO PLATYPUS CYLINDRUS FAB. (COLEOPTERA: PLATYPODIDAE) IN CORK OAK
FUNGOS ASSOCIADOS AO INSECTO PLATYPUS CYLINDRUS FAB. (COLEOPTERA: PLATYPODIDAE) EM SOBREIRO
Joana Henriques, Maria de Lurdes Inácio, Edmundo Sousa
Abstract Platypus cylindrus é uma praga que desde Platypus cylindrus is a pest that since the os anos 80 do século passado tem sido refe- 80’s of the last century has been considered rida como agente de mortalidade do sobrei- a cork oak mortality agent in Portugal. It ro em Portugal. É um insecto ambrósia que is an ambrosia beetle that establishes com- estabelece simbioses complexas com fungos plex symbioses with fungi whose role in the cujo papel não está completamente esclare- insect-fungus-host interaction has not been cido na interacção insecto-fungo-sobreiro. completely clarified. In order to characterize Com o objectivo de caracterizar a micoflora P. cylindrus associated micoflora in Portugal, associada a P. cylindrus em Portugal foram fungi were isolated from different beetle or- efectuados isolamentos a partir de diferen- gans and from its galleries in cork oak trees. tes órgãos do insecto e suas galerias em so- Fungi of the genera Acremonium, Aspergillus, breiro. Identificaram-se fungos dos géneros Beauveria, Botrytis, Chaetomium, Fusarium, Acremonium, Aspergillus, Beauveria, Bo- Geotrichum, Gliocladium, Nodulisporium, trytis, Chaetomium, Fusarium, Geotrichum, Paecilomyces, Penicillium, Raffaelea, Scyta- Gliocladium, Nodulisporium, Paecilomyces, lidium, Trichoderma and of the order Muco- Penicillium, Raffaelea, Scytalidium, Tricho- rales were identified. An actinomycete of the derma e da ordem Mucorales. Foi igualmen- genus Streptomyces was also identified. Some te identificado um actinomiceta do género of these genera were related for the first time Streptomyces. Alguns destes géneros são to this interaction. In the present work the iso- referidos pela primeira vez nesta interacção. lated fungi are characterized and their contri- No presente trabalho caracterizam-se os fun- bution for beetle population establishment and gos isolados e discute-se a sua contribuição tree weakness is discussed. para o estabelecimento das populações do insecto e enfraquecimento das árvores. Key-words: ambrosia beetle, decline, inte- raction, mycoflora,Quercus suber. Palavras-chave: insecto ambrósia, declínio, interacção, micoflora,Quercus suber.
Resumo INTRODUÇÃO
Instituto Nacional de Recursos Biológicos, I.P. Scolytidae and Platypodidae are among the Edifício da ex-Estação Florestal Nacional, Quinta do Marquês, 2780-159 Oeiras most successful wood-inhabiting beetles cau- [email protected]; lurdes.inacio@efn. sing damage of economic significance to tre- com.pt; [email protected] es and timber (Cassier et al., 1996). Platypus cylindrus Fab. is a cork oak (Quercus suber L.) Recepção/Reception: 2009.01.15 pest that has come to assume an increasing im- Aceitação/Acception: 2009.05.14 portance in Portugal and Mediterranean basin FUNGI ASSOCIATED TO PLATYPUS CYLINDRUS FAB. (COLEOPTERA: PLATYPODIDAE) IN CORK OAK 57 countries (Ferreira & Ferreira, 1989; Chakali when excavating tunnels. Auxiliary ambrosia et al., 2002; Riziero et al., 2002; Sousa et al., fungi are transitory, non-specific with respect 2005). Until recently, damages produced by to symbiont insect and may appear after in- this insect were limited to dead or weakened sect development. They may not be present trees. The understanding of recent population in larval cradles or in adult beetles; and their outbreaks, mainly in Portuguese cork oak habitat and distribution range are unrestric- stands, can be based on three assumptions: (i) ted and unrelated to that of the ambrosia be- gradual changes of the cork oak stand dyna- etles. mics, (ii) development of more specific host Several fungi have already been isolated colonization mechanisms and (iii) changes on from P. cylindrus and from galleries in Quer- the insects and their natural enemies’ popula- cus spp. (Baker, 1963; Cassier et al., 1996; tion dynamics (Sousa & Inácio, 2005). Within Sousa et al., 1997; Morelet, 1998; Henriques the host colonization strategies, it’s essential et al., 2006). Their constant presence asso- to consider the fungal symbiosis that may ciated to the insect allows us to presume that contribute to the host weakness and create bet- they play an important role in the symbiosis. ter conditions for the establishment of insects Besides the implication in insect feeding, am- (Henriques et al., 2006). brosia fungi might also be involved in other In deed, P. cylindrus, as almost all members processes such as host weakness, through of the Platypodidae, is denominated an am- pathogenic action; decomposition of ligno- brosia beetle because larvae and adults feed cellulolitic compounds, which helps gallery mainly on fungi (ambrosia fungi) that cover construction and enables fungi colonization; the gallery walls (Batra, 1963). This insect- and/or antagonism that controls the growth fungi relation is expressed in an ectosymbio- of other fungi (Sousa & Inácio, 2005; Henri- sis in which the fungi live outside the insects’ ques et al., 2006). body but are temporarily stored in special ec- The aim of the present work is to charac- todermical organs for dissemination purposes terize the micoflora associated toP. cylindrus (Francke-Grosmann, 1967). Insects carry via- and discuss its contribution for beetle popu- ble inoculum in sac-like structures, called my- lations’ establishment and host weakness. cangia, located in the prothorax; the inoculum is protected from desiccation during the entire life of the beetle and is disseminated into new Material and methods breeding sites at the time of tunnel excavation (Batra, 1963; Sousa & Inácio, 2005). Four infested logs of cork oak trees exhi- Ambrosia fungi definition includes a set biting decline symptoms were collected from of concepts whose interception allows the three of the main Portuguese cork oak produc- classification of several fungi as ambrosia: i) tive regions: Chamusca (Ribatejo province), direct participation in insect feeding; ii) pre- Montemor and Grândola (Alentejo province). sence inside insect galleries in the host; iii) The logs were maintained in the laboratory in dimorphism, meaning ability to grow both order to capture 100 P. cylindrus insects, males as yeast and mycelia; iv) possible specificity and females, as they emerged. Samplings were in the insect-fungi-host relationship (Batra, repeated during 2005, 2006 and 2007. The in- 1963; Beaver, 1989). sects were aseptically dissected in mycangia, Batra (1985) grouped ambrosia fungi as intestine and parts of the exoskeleton (elytra). primary and auxiliary. Primary ambrosia fun- The logs were cut in order to identify the diffe- gi are highly insect species specific and their rent insect gallery sections: cork, inner-bark, distribution correspond to those of insect pre-parental section, larval section and gallery symbionts. They are present and dominant end. One complete gallery was observed from in the tunnels and isolated regularly from the each log and six samples (fragments with 1 mycangia of the beetles in the flight stage or cm2) of each section were collected. 58 REVISTA DE CIÊNCIAS AGRÁRIAS
The samples were surface sterilized with Results a sodium hypochlorite solution (1%) for 1 min, rinsed with distilled sterilized water and Fifteen genera were isolated from the di- then plated in malt extract agar (MEA, Dif- fferent insect parts and from all cork oak gal- co, USA) added with streptomycin (500 mg/l) lery sections (Table 1). The identified fungi and MEA added with cycloheximide (500 are classified in very distinct orders including mg/l). The cultures were incubated at 24±1ºC the Ascomycota (Eurotiales, Heliotioales, in darkness. Pure cultures of each fungi isolate Hypocreales, Ophiostomatales, Saccharomy- were obtained and identified as genus based cetales, Sordiales and a genus not assigned on morphological features according to Ellis to any order) and Basidiomycota (Xylariales) (1971, 1976), Lanier et al. (1978), Kiffer & although all the obtained genera, except Cha- Morelet (1997) and Barnett & Hunter (1998). etomium, were isolated in the mitosporic sta- Cultural description of colonies was made on te. Also, an actinomycete was isolated from potato dextrose agar (PDA, Difco, USA). the insect (Streptomycetales).
Table 1 – Genera isolated from Platypus cylindrus intestine, exoskeleton and mycangia and from the different sections of insects galleries on cork oak.
* Streptomyces (order Actinomycetales) is an actinomycete, the other genera described belong to Fungi.
The isolated genera were described ac- isolated, probably belonging to different spe- cording to the morphological features ba- cies. Their microscopic features are: hyphae sed on optical microscopic observations septate and branched, conidiophores macro- and literature guides. In table 2 the main nematous, mononematous, erect, simple, often cultural features for all isolated genera are with a foot cell, with a terminal vesicle bearing described. short branches or phialides radiating from the From the genus Aspergillus several individu- entire surface. When present, the branches are als with different cultural characteristics were in one or several series and the terminal ones FUNGI ASSOCIATED TO PLATYPUS CYLINDRUS FAB. (COLEOPTERA: PLATYPODIDAE) IN CORK OAK 59
Table 2 – Cultural characteristics on PDA of the isolated genera.
always bear phialides. Conidiogenic cells mo- Penicillium produces conidiophores ma- nophialidic, discrete, several arising together cronematous, mononematous, simple or at the end of terminal branches or over the sur- branched, penicilated, ending in a group of face of the vesicles, mostly determinate, rarely phialides. The walls of conidiophores may be percurrent, ampulliform or lageniform, colla- smooth and thin or variously roughened, with rettes sometimes present. Conidia are phialos- aerial portions appearing delicately echinula- poric, unicellular, dry, smooth, rugose, echinu- te, granular or asperulate. The penicillus co- late, globose and hyaline but colored in mass, vers all the branching system and can be mo- disposed in long basiptal chains (Figure 1a). noverticillate or biverticillate, symmetrical 60 REVISTA DE CIÊNCIAS AGRÁRIAS or asymmetrical. Conidia are phialosporic, gularly grouped, inflated in the base and ta- unicellular, dry, globose or ovoid, hyaline to pered in the apical fertile portion that appears green (Figure 1b). sinusoidal after conidia production. Conidia Paecilomyces produces conidiophores are simpodulosporic, unicellular, dry, hyali- macronematous, mononematous, arising as nes and ovoid with small denticles, giving upright branches from hyphal ropes or ae- the conidiogenous cells a spiny appearance rial mycelium, smooth-walled, usually with (Figure 1g). several stages of irregular branching, and Gliocladium presents conidiophores ma- frequently bearing secondary branches, with cronematous and mononematous, erect, sep- divergent penicilated phialides, cylindrical to tate and branched, ending in a branched sys- ellipsoidal in the lower part, usually narro- tem of phialides disposed in tight penicilate wing abruptly into a long cylindrical neck. structure with three phialides per metulae. Conidia phialosporic, unicellular, hyaline, Phialides narrowly cylindrical to sub-ulate, ovoid, smooth-walled, highly variable in size taper slightly towards the tip, smooth-walled. produced in long, strongly divergent chains Conidia are phialosporic, unicellular, hyali- (Figure 1c). ne, ovoid or cylindrical and aggregated in Several Raffaelea isolates were obtained conidial masses, slimy to watery, whitish or showing a great macro and microscopic va- light-yellow, never forming imbricate chains riability, probably corresponding to different (Figure 1i). species. This genus produces hyphae hyali- Acremonium produces hyphae hyaline sep- ne and septate that bound together forming tate that sometimes bound together by anas- compact hyphae ropes. Superficial sporodo- tomoses. Conidiophores are macronematous chia, effuse, white to brownish. Conidiopho- and mononematous, erect, solitary to weakly res are macronematous and mononematous, branched, straight, tapered to the apex, with erect, septate, slender with a tapered apex, basal septum to separate the conidiopho- producing simpodulosporic conidiae that re from the vegetative hyphae. Conidia are leave cicatricial scars in the conidiogenous phialosporic, unicellular and ovoid, aggrega- cells. Conidiae are unicellular and hyaline, ted in a slimy drop (Figure 1k). with variable forms (triangular, oval, allan- Geotrichum produces conidiophores mi- toid, fusiform or truncated claviform) and cronematous and mononematous producing dimensions (Figure 1d). arthrosporic conidiae, hyaline, unicellular, Fusarium produces conidiophores macro- dry, smooth, short cylindrical with truncated nematous and mononematous, variable, slen- bases, resulting from fragmentation of undi- der, and simple or stout, short, septate and fferentiated hyphae by fission through dou- branched that originates phialosporic coni- ble septum, (Figure 1l). dia, hyaline, canoe-shaped, with 5 to 6 trans- Botrytis has conidiophores macronema- versal septa and collected in a slimy drop. No tous, mononematous, straight or flexuous, microconidia were observed (Figure 1e). smooth, brown, slender, irregularly bran- Trichoderma conidiophores macronema- ched with enlarged apical polyblastic cells tous and mononematous, hyaline and highly where botryoblatosporic conidia are produ- ramified, disposed in pyramidal structures ced simultaneously; conidia solitary, simple, with inserted phialides in 90º angles. Conidia smooth, hyaline to grey in mass, unicellular, are phialosporic, enteroblastic and monoblas- ovoid with a denticule. Mycelium immersed tic, unicellular, green, globose to subglobose or superficial, with brownish course hyphae. and remain grouped in the top of the phiali- Sclerotia frequently present (Figure 1h). des (Figure 1f). Chaetomium produces superficial perithe- Beauveria produces conidiophores micro- cia, brown, single, globose, covered with nematous and mononematous, simple, irre- different sized brown filaments; asci clavate, FUNGI ASSOCIATED TO PLATYPUS CYLINDRUS FAB. (COLEOPTERA: PLATYPODIDAE) IN CORK OAK 61 pedunculate with evanescent walls. Ascospo- ticilated. Conidiae sympodulosporic, acro- res unicellular, light olive-brown and lemo- pleurogenous, unicellular, hyaline or bro- niform, smooth, often pushed out of ostiole wn to olive in mass, ellipsoidal or obovoid, in a cirrhus (Figure 1j). smooth or roughened, with a small frill when Streptomyces produces filaments that ori- detached (Figure 1n). ginate very small spores by fragmentation Scytalidium produces sparse thick that remain disposed in helical chains (Figu- hyphae, septate, hardly branched, hyaline or re 1m). light-brown, disposed in parallel; conidio- Nodulisporium produces mycelium partly phores micronematous and mononematous immersed and partly superficial, conidio- with terminal conidiogenic cell originating phores macronematous and mononematous, arthroconidiae by holothalic fragmentation. arising laterally from the brownish vegetati- Conidia are catenulated, schizogenous, uni ve hyphae, with principal axis erect, septate, or bicellular, hyaline, rectangular. It also branched, hyaline to light brown, slightly forms terminal or interpolate chlamydospo- rugouse, conidiogenic cells poliblastic and res, with thick wall, brown and ellipsoidal sympodial, slender or short and thick, ver- (Figure 1o).
b c
a
g h
d e f
i
k o i
l m n
Figure 1 – Reproductive structures of isolated genus: a) Aspergillus (x1000); b) Penicillium (x1000); c) Paecilomyces (x600); d) Raffaelea (x600); e) Fusarium (x600); f) Trichoderma (x600); g) Beauveria (x1000); h) Botrytis (x600); i) Gliocladium (x400); j) Chaetomium (x1000); k) Acremonium (x600); l) Geotrichum; m) Streptomyces (x1000); n) Nodulisporium (x600); o) Scytalidium (x600).
In P. cylindrus galleries on cork oak, gre- observed. Penicillium, Raffaelea and Asper- ater fungi variety were presented than in the gillus were the most frequent genus in the insects’ body. In both cases were isolated three sampled organs and for both sexes. cosmopolite fungi and others more specific Beauveria and Streptomyces were only iso- for insect-fungi relation. Concerning isola- lated from the insect, the former only from tes from insects, it was in the exoskeleton the exoskeleton. that was found the major diversity of genus, Along P. cylindrus galleries in cork oak, followed by mycangia and then intestine. the majority of the isolated genera were Among sexes, no relevant differences were found in all sections and, in general, in si- 62 REVISTA DE CIÊNCIAS AGRÁRIAS milar proportions, highlighting cosmopolite isolated only from the galleries but in very and saprophytic fungi such as Penicillium, low percentages and distribution. Trichoderma, Gliocladium and Scytalidium, In Figures 2 and 3 are represented the dis- but also Raffaelea was isolated in a conside- tribution of the isolated fungi both in P. cylin- rable percentage in all sections. Botrytis, Fu- drus body and along its galleries in cork oak, sarium, Geotrichum and Chaetomium were respectively.
50 % F M 40
e 30 in
st 20 te
In 10
0
50
40 n
to 30 le 20 ke
os 10 Ex 0 50
40 ia 30
20 Mycang 10
0
a a m m es es ri um illu s rm ri yc yc lliu m nium ve rg aelea adiu ci po ff alidiu ode om au ni mo pe ilom h pt io cl Ra ic Be As Pe ec Scyt re Gl Tr Acre odulis St Pa N Ge nus
Figure 2 – Percentage of isolated genera in the different body parts (intestine, exoskeleton and mycangia) of Platypus cylindrus females (F) and males (M).
Figure 3 – Percentage of isolated genera from the different Platypus cylindrus galleries on cork oak (cork, inner bark, pre-parental section, larval section and gallery end). FUNGI ASSOCIATED TO PLATYPUS CYLINDRUS FAB. (COLEOPTERA: PLATYPODIDAE) IN CORK OAK 63
DISCUSSION galleries on cork oak and other oaks (Baker, 1963; Sousa et al., 1997). Nodulisporium and From all isolated genera, Raffaelea is hi- Scytalidium were found in mycangia and gal- ghlighted because of its known importance leries on cork oak, the last one only in the as ambrosia fungi and both its frequency host (Sousa et al., 1997). and distribution (Beaver, 1989). This genus Four genera were newly associated with includes twelve species, the majority asso- this interaction: Beauveria, Geotrichum, ciated with ambrosia beetles (Kubono & Ito, Chaetomium and Streptomyces. Beauveria is 2002; Bisby et al., 2006). Two species (R. an entomopathogenic fungus and two species ambrosiae and R. montetyi) are identified as (B. bassiana and B. brongniartii) are alrea- P. cylindrus primary ambrosia fungi (Arx & dy known as harmful to Platypus spp. (Glare Hennebert, 1965; Morelet, 1998) and have et al., 2002). Several species of Geotrichum been isolated both from the insect organs have been identified in association with in- and from its galleries in the host (Sousa et sects (Suh & Blackwell, 2006) and the genus al., 1997; Morelet, 1998). Although Raffa- Streptomyces act as an antimicrobial defense elea sexual state is lacking, observations in the termite-fungi association (Mueller & of conidial development support its place- Gerardo, 2002). Chaetomium is a very com- ment within the Ophiostomatales (Gebhardt mon genus in soil and plant debris, in par- & Oberwinkler, 2005) and, 18S ribosomal ticular in wood (Hawksworth, 1995) and it DNA sequence analysis, shows a monophy- might be a Scytalidium teleomorph (Halin, letic lineage which forms a sister group to 1997). species of the genus Ophiostoma (Jones & The role of each isolated genus in the in- Blackwell, 1998). The Ophiostomatales are sect-fungi-host interaction is discussed. Most economically important sapstaining fungi of them are considered cosmopolite and sa- occurring world-wide on hardwoods and prophytic fungi (Hawksworth et al., 1995; commercially produced pines, and are in Kiffer & Morelet, 1997) which can be invol- some cases, already known as pathogenic to ved just in a commensalistic relation or might oaks (Degreef, 1992). The effect of Raffaelea play some other action due to their isolation spp. on cork oak remains unknown, but in Ja- frequency and distribution among samples. pan, R. quercivora involved in an interaction Following P. cylindrus host colonization with P. quercivorus has proven pathogenici- process, symbiotic fungi may start to act as ty to several oak trees (Kubono & Ito, 2002; wood degrading, thus facilitating galleries’ Kinnura & Kobayashi, 2006). excavation. Trichoderma, as a great extroli- Other isolated fungi were already related te producer such as lignocellulolictic enzy- to P. cylindrus. Sousa et al. (1997) isolated mes, might be an active intervenient in that A. carbonarius from mycangia and cork oak process (Samuels, 1996). Then, to overco- galleries. Pa. variotii has been isolated from me host defense reaction, the insect might galleries in cork oak and other oaks (Baker, inoculate phytopathogenic fungi. Nodulis- 1963). Also Penicillium was found both on porium might be relevant in this part as the the insect and in oak galleries (Baker, 1963; ITS rDNA analysis proved that it is a Bis- Cassier et al., 1996; Sousa et al., 1997). Bo- cogniauxia mediterranea anamorph, which trytis was isolated from galleries in oak, as is responsible for cork oak charcoal disease well as Acremonium and Fusarium (Baker, (Collado et al., 2001). Other isolated gene- 1963). Acremonium sp. and F. solani, Glio- ra might also interfere in this phase: Acre- cladium roseum and G. solani has also been monium which might be Nectria anamorph, isolated by Sousa et al. (1997) from P. cylin- thus being a potential pathogenic to several drus organs and galleries in cork oak. Cas- woody plants and Fusarium also pathoge- sier et al. (1996) isolated Trichoderma from nic to several plant organs (Kiffer & Mo- the insect and T. viride was isolated from relet, 1997). Within the galleries, fungi that 64 REVISTA DE CIÊNCIAS AGRÁRIAS will be nourishment for larvae develop in Bibliographic references a controlled manner by the permanent care of parental insects but probably also due to Arx, J.A. von & Hennebert, G.L. (1965) - the antagonistic action of fungi over others. Deux champignons ambrosia. Mycopatho- Gliocladium and Trichoderma are known logia et mycologia applicata 25: 309-315. by their antagonistic activity being used as Baker, J.M. (1963) - Ambrosia beetle and biocontrol agents (Papavizas, 1985). Strep- their fungi, with particular reference to tomyces and Scytalidium might act in the Platypus cylindrus Fab. Symposia of the fungi colonies management within the gal- Society for General Microbiology 13: 323- leries. Also in the insect body this manage- 354. ment might be essential to achieve ambrosia Barnett, H.L. & Hunter, B.B. (1988) - Illus- fungi transport. The final role of the isola- trated Genera of Imperfect Fungi. APS ted fungi, which is the base of this insect- Press, Minnesota, USA, 218 pp. fungi interaction, is the larvae nourishment. Batra, L.R. (1963) - Ecology of ambrosia According to definition, ambrosia fungi are fungi and their dissemination by beetles. eaten by the insects and thus are the ones Transactions of the Kansas Academy of found in their intestine. Nevertheless, whe- Science 66: 213-236. reas larvae eat exclusively ambrosia fungi, Batra, L.R. (1985) - Ambrosia beetle and the adults might also eat wood, which ex- their associated fungi: Research trends and plains all the cosmopolite and saprophytic techniques. Proceedings of the Indian Aca- fungi found in their intestine that were alre- demy of Sciences 49: 137-148. ady contaminating tree wood. Also, in my- Beaver, R.A. (1989) - Insect-fungus rela- cangia, where specific fungi for this inte- tionships in the bark and ambrosia beetles. raction are transported, ambrosia fungi are In: Wilding, N.; Collins, N.M.; Hammond, expected to be present. In both cases, intes- P.M. & Webber, J.F. (Eds.) Insect-Fungus tine and mycangia, Raffaelea was the most Interactions. Academic Press, London, pp. frequent genus, which supports the fact that 121-143. some species of this genus are P. cylindrus Bisby, F.A.; Ruggiero, M.A.; Roskov, primary ambrosia fungi. The other isolated Y.R.;Cachuela-Palacio, M.; Kimani, S.W.; fungi might be considered auxiliary ambro- Kirk, P.M.; Soulier-Perkins, A. & van sia fungi as sustained by Batra (1985) or Hertum, J. (2006) - Species 2000 & ITIS simply be worldwide saprobes that are fre- Catalogue of Life: 2006 Annual Checklist. quently present in host tissues. Available in
and effect of different ecological factors on Kiffer, E. & Morelet, M. (1997) - Les distribution of the fungus. Mycologia 93: Deutèromycetes – classification et clés 875-886. d´identification générique. INRA Editions, Degreef, J. (1992) - Isolation of Ophiostoma Paris, 306 pp. querci (Georgev.) Nannfeldt from decli- Kinuura, H. & Kobayashi, M. (2006) - Death ning oaks in Belgium: selection techniques of Quercus crispula by inoculation with and pathogenicity test. In: Proceedings of adult Platipus quercivorus (Coleoptera: an International Congress “Recent Ad- Platypodidae). Applied Entomology and vances in Studies on oak decline”. Selva Zoology 41, 1: 123-128. di Fasano (Brindisi), Italy, pp. 471-473. Kubono, T. & Ito, S. (2002) - Raffaelea quer- Ellis, M.B. (1971) - Dematiaceous Hyphomy- civora sp. nov. associated with mass mor- cetes. CAB, England, 608 pp. tality of Japanese oak, and the ambrosia Ellis, M.B. (1976) - More Dematiaceous beetle (Platypus quercivorus). Mycoscien- Hyphomycetes. CAB, England, 507 pp. ce 43: 255-260. Ferreira, M.C. & Ferreira, G.W.S. (1989) - Lanier, L.; Joly, P.; Bondoux, P. & Bellemè- Platypus cylindrus F. (Coleoptera: Platipo- re; A. (1978) - Mycologie et pathologie didae) Plaga de Quercus suber. Boletin de forestières. Tome I - Mycologie forestière. Sanidade Vegetal Plagas 4: 301-305. Masson, Paris. 487 pp. Francke-Grosmann, H. (1967) - Ectosym- Morelet, M. (1998) - Une espèce nouvelle de biosis in wood-inhabiting insects. In: S.M. Raffaelea, isolée de Platypus cylindrus, co- Henry (Ed.) Symbiosis, its physical and léoptère xylomycétophage des chênes. Ex- biochemical significance. Academic Press, trait des Annales de la Société des Science New York, pp. 141-203. Naturelles et d’Archéologie de Toulon et Gebhardt, H. & Oberwinkler, F. (2005) - Co- du Var 50: 185-193. nidial development in selected ambrosial Mueller, U.G. & Gerardo, N. (2002) - Fun- species of the genus Raffaelea. Antoine gus-farming insects: Multiple origins and van Leeuwenhoek 88: 61-66. diverse evolutionary histories. Proceedin- Glare, T.R.; Placet, C.; Nelson, T.L. & Reay, gs of the National Academy of Sciences 99, S.D. (2002) - Potencial of Beauveria and 24: 15247-15249. Metarhizium as control agents of pinhole Papavizas, G.C. (1985) - Trichoderma and borers (Platypus spp.). New Zealand Plant Gliocladium: biology, ecology and poten- Protection 55: 73-79. tial for biocontrol. Annual Review Phyto- Halin, R.T. (1997) - Ilustrated Genera of As- pathology 23: 23-54. comycetes. Vol I., APS Press, Minnesota, Riziero, T.; Ragazzi A.; Marianelli, L.; Sab- 263 pp. batini, P. & Roversi, P.F. (2002) - Insects Hawksworth, D.L.; Kirk, P.M.; Sutton, B.C. and fungi involved in oak decline in Italy, & Pegler, D.N. (1995) - Ainsworth & IOBC/wprs Bulletin 25, 5: 67-74. Bisby’s Dictionary of the Fungi. CAB In- Samuels, G.J. (1996) - Trichoderma: a review ternational, UK, 616 pp. of biology and systematics of the genus. Henriques, J.; Inácio, M.L. & Sousa, E. Mycological Research 100, 8: 923-935. (2006) - Ambrosia fungi in the insect-fungi Sousa, E. & Inácio, M.L. (2005) - New As- symbiosis in relation to cork oak decline. pects of Platypus cylindrus Fab. (Coleop- Revista Iberoamericana Micologia 23: tera: Platypodidae) Life History on Cork 185-188. Oak Stands in Portugal. In: F. Lieutier & Jones, K.G. & Blackwell, M. (1998) - Phylo- D. Ghaioule (Eds.) Entomological Rese- genetic analysis of ambrosial species in arch in Mediterranean Forest Ecosystems. the genus Raffaelea based on 18S rDNA INRA Editions, Paris, 280 pp sequences. Mycologycal Research 102: Sousa, E.; Inácio, M.L.; El Antry, S.; Bakry, 661-665. M. & Kadiri, Z.A. (2005) - Comparaison 66 REVISTA DE CIÊNCIAS AGRÁRIAS
de la bio-écologie et du comportement de dae). Phytopathologia Medierranea. 36: l’insecte Platypus cylindrus Faber. (Colé- 145:153. optère, Platypodidae) dans les subéraies Suh, S.O. & Blackwell, M. (2006) - Three Portugaises et Marocaines. IOBC/wprs new asexual asthroconidial yeasts, Geotri- Bulletin 28: 137-144. chum carabidarum sp. nov., Geotrichum Sousa, E., Tomaz, I. L.; Moniz, F.A. & histeridarum sp. nov., and Geotrichum Basto, S. (1997) - La répartition spatia- cucujoidarum sp. nov., isolated from the le dês champignons associés à Platypus gut of insects. Mycological Research 110: cylindrus Fab. (Coleoptera: Platypodi- 220-228.