el nematodo del pino, bursaphelenchus xylophilus
TRANSCRIPT
EL NEMATODO DEL PINO,,BURSAPHELENCHUS XYLOPHILUS
Alfonso NavasAlfonso NavasMuseo Nacional de Ciencias Naturales. CSIC
5º Congreso Forestal EspañolSeminario 11: Situación y medidas ante el nematodo de la madera del pino
(24 de septiembre de 2009, Ávila)
Anisakis simplex
Male tail is curved and pointed with short, oval caudal alae at tail tip -hence the genus name.
spicules with prominent disc expansions at distal endspicules with prominent disc expansions at distal end
A very similar but non-pathogenic species, B. mucronatus, was described by Mamiya & Enda (1979) differing morphologically only indescribed by Mamiya & Enda (1979), differing morphologically only in minor respects from B. xylophilus and most obviously by the presence in the female of a caudal mucro (finger-like projection) in the former species which was absent in the latter. However, p ,populations of B. xylophilus were subsequently discovered in the USA which also carried a mucro on the tail. There has thus been much discussion about the taxonomic relationships between these ptwo species, and also with B. fraudulentus Rühm, a nematode from deciduous trees in central Europe.
Sin mucrón
Pequeño mucrón
There are some 75 described species of
Bursaphelenchus. p
Most species have a phoretic relationship with insects, especially bark
beetles and wood borers and are associated with dead or dying conifers.
All species feed on fungi.
B. mucronatus
Bursaphelenchus xylophilus
Bursaphelenchus xylophilus (Steiner and Buhrer, 1934) Nickle, 1970
Criterios morfológicos Criterios filogenéticosMagentti (1983) y De Coninck (1965) De Ley and Blaxter (2002, 2004)
Clase: SECERNENTEA CHROMODOREA
Subclase: DIPLOGASSTERIA CHROMODORIA
Orden: TYLENCHIDA RHABDITIDA
S b d APHELENCHINA TYLENCHINASuborden: APHELENCHINA TYLENCHINA
Infraorden: TYLENCHOMORPHA
S perfamilia APHELENCHOIDEASuperfamilia: APHELENCHOIDEA
Familia: APHELENCHOIDIDAE APHELENCHOIDIDAE
Subfamilia: BURSAPHELENCHINAE BURSAPHELENCHINAESubfamilia: BURSAPHELENCHINAE BURSAPHELENCHINAE
Sinónimos: Aphelenchoides xylophilus Steiner & Buhrer 1934 (USA)p y p ( )Bursaphelenchus lignicolus Mamiya & Kiyohara, 1972 (Japón)
Notes on taxonomy and nomenclature: Biochemical studies of several populations of B. xylophilus, B. mucronatus and B. fraudulentus have clearly confirmed the distinctness of the three species (Webster et al., 1990; Abad et al., 1991). B. xylophilus is a native of North America, whereas the other two are Palaearctic species, one colonizing coniferous trees, the other deciduous. B. xylophilus found in Japan and other Asian countries is
b i l i t d ti f N th A i A thobviously an introduction from North America. Another apparently closely related species, Bursaphelenchus kolymensis Korenchenko, has been described from Larix from the far east of the former USSR (Korenchenko 1980);from the far east of the former USSR (Korenchenko, 1980); this nematode has not been studied extensively but is possibly synonymous with B. mucronatus. A population of B. mucronatus has been detected in the Province of Quebecmucronatus has been detected in the Province of Quebec, Canada (Harmey & Harmey, 1993), but it is not known if this is an introduction.
method for discriminating
ITS-RFLP patterns of a Bursaphelenchus xylophilus isolate compared with a B. mucronatus isolate. Restriction fragments were obtained by digestion of the amplified rDNA containing the ITS
) ) S Sregions with the restriction enzymes indicated. M) 100 bp marker, 1) ITS-Amplicon ITS-Amplicon, 2) RsaI, 3) HaeIII, 4) MspI, 5) HinfI, 6) AluI.
ITS-RFLP patterns ITS-RFLP patterns
M 1 2 3 4 5 6 M M 1 2 3 4 5 6 M
B. xylophilus B. mucronatus
ITSITS--RFLPRFLP patterns of patterns of BursaphelenchusBursaphelenchus speciesspecies(Aphelenchoididae Nematoda)(Aphelenchoididae Nematoda)(Aphelenchoididae, Nematoda)(Aphelenchoididae, Nematoda)
Braasch, Metge & Burgermeister (1999), Mota et al (1999), Braasch, Burgermeister & Metge (2001), Braasch & Burgermeister (200Braasch, Metge & Burgermeister (1999), Mota et al (1999), Braasch, Burgermeister & Metge (2001), Braasch & Burgermeister (2002)2)
juveniles adultos
DNA satélite
Monochamus galloprovincialis Monochamus galloprovincialis (Oliver, 1795)(Oliver, 1795)
Familia: Cerambycidae
Subfamilia: Lamiinae
Tribus: Monochamini
Genus: Monochamus
El t d d l Pi (PWN) t itid lEl nematodo de los Pinos (PWN) es transmitido a las coníferas por varias especies del género Monochamus, Coleópteros de la familia Ceramb cidaeCerambycidae.
Otras especies de Otras especies de MonochamusMonochamusvectores de vectores de B. xylophilusB. xylophilus
Especies de MEspecies de Monocham sonocham s Distrib ción geográficaDistrib ción geográfica HospedadorHospedador VectorVectorEspecies de MEspecies de Monochamusonochamus Distribución geográficaDistribución geográfica HospedadorHospedador VectorVector
NorteaméricaNorteaméricaM. caroliniensisM. caroliniensis OlivierOlivier EE.UU., Canadá y MéxicoEE.UU., Canadá y México PinusPinus ++
M. marmoratorM. marmorator kirbykirby “ ““ “ Abies, PiceaAbies, Picea ++
M. mutatorM. mutator LeConteLeConte “ ““ “ PinusPinus ++
M. obtususM. obtusus CaseyCasey “ ““ “ Abies, Abies, Pinus pseudotsugaPinus pseudotsuga
++Pinus pseudotsugaPinus pseudotsuga
M. scutellatusM. scutellatus scutellatusscutellatus SaySay Noreste EE.UU. y MéxicoNoreste EE.UU. y México Abies, Larix Picea, PinusAbies, Larix Picea, Pinus ++
M. scutellatusM. scutellatus oregonensisoregonensis Costa Oeste de EE.UU. y Costa Oeste de EE.UU. y PiceaPicea ++scute atusscute atus o ego e s so ego e s sLeConteLeConte
Costa Oeste de UU yCosta Oeste de UU yCanadáCanadá
ceacea
M.titillatorM.titillator (Fabricius)(Fabricius) EE.UU. (Centro, Este,Sudeste)EE.UU. (Centro, Este,Sudeste)y Canadáy Canadá
Abies, Picea PinusAbies, Picea Pinus ++
Otras especies de Otras especies de MonochamusMonochamus vectoresvectoresRegión Paleartica Región Paleartica M. alternatus M. alternatus (Hope)(Hope) Japón, Corea, Taiwan, Japón, Corea, Taiwan, Pinus, Cedrus, Abies,Pinus, Cedrus, Abies, ++
Hong Kong, Laos y ChinaHong Kong, Laos y China Picea, LarixPicea, LarixM. saltuarius M. saltuarius (Eschscholz)(Eschscholz)
Japón, China, Siberia, Japón, China, Siberia, Lituania Alpes, Europa Lituania Alpes, Europa Central y Este hasta ItaliaCentral y Este hasta Italia
PiceaPicea ++
Central y Este hasta ItaliaCentral y Este hasta ItaliaM. galloprovincialisM. galloprovincialis(Olivier)(Olivier)
Norte de África, Europa Norte de África, Europa hasta Siberiahasta Siberia
PinusPinus ++
M sartorM sartor (Fabricius)(Fabricius) Europa (PirineosEuropa (Pirineos Picea PinusPicea PinusM. sartor M. sartor (Fabricius)(Fabricius) Europa (Pirineos Europa (Pirineos ––Ucrania), China, Siberia y Ucrania), China, Siberia y RusiaRusia
Picea, PinusPicea, Pinus --
M. sutorM. sutor (Linnaeus)(Linnaeus) China, Siberia, Rusia,China, Siberia, Rusia, Larix, Picea y PinusLarix, Picea y Pinus --M. sutorM. sutor (Linnaeus)(Linnaeus) China, Siberia, Rusia, China, Siberia, Rusia, Países nórticos, Alpes Países nórticos, Alpes hasta los Pirineoshasta los Pirineos
Larix, Picea y PinusLarix, Picea y Pinus
Distribution of Monochamus spp.
Geographic distribution of Monochamus speciesGeographic distribution of Monochamus species
M. sartor
M. galloprovincialis
M. sutor
M. Urussovi and M. saltuarius (eastern species)
BIOLOGY of Bursaphelenchus xylophilus
two different modes in its life cycle, a propagative mode and a dispersal mode (Wingfield, 1983).two different modes in its life cycle, a propagative mode and a dispersal mode (Wingfield, 1983).
In both cases, the nematodes are transmitted from one host to the next by species of Monochamus.
In the propagative life cycle fourth-stage larvae of the nematodes are transmitted to recently dead orIn the propagative life cycle, fourth-stage larvae of the nematodes are transmitted to recently dead ordying trees during oviposition by the female vector.
The nematodes leave the beetle and enter the tree through the hole in the bark cut by the vector to lay its eggs.
Within the wood the nematodes feed on the hyphae of fungi (usually Ceratocystis spp.) also transmitted to the wood by ovipositing beetlestransmitted to the wood by ovipositing beetles.
Immediately on entering the wood the larval nematodes moult to adult and begin laying eggs. The population is composed of males, females and four larval stages in this propagative stage of rapid multiplication.
NEMATODO DE LOS PINOS: Bursaphelenchus xylophilusRelación forética entre Monochamus galloprovincialis (Coleóptero: cerambícido)
y Bursaphelenchus xylophylusy Bursaphelenchus xylophylus
Los huéspedes resistentes provocan
la muerte de losAlimentación en lo alto de los árboles
(Wingfield, 1983)
Adultos emergidos
Transportan nematodos
Ovoposición
la muerte de los nematodos
Árboles
Ovoposición
TRANSMISIÓN MEDIANTE
ALIMENTACIÓNÁrboles
TRANSMISIÓN
Los nematodos (larvas Dauer) migran hacia las pupas justo antes de que emerja el adulto
senescentes o muertos
ALIMENTACIÓN
Los nematodos penetran por
sanos
MEDIANTE OVOPOSICIÓN
Los nematodos se alimentan de
las heridas
Huésped sensibleNematodos ponen huevos se alimentan de
células de la madera o de hongos
Los nematodos se reproducen en el
xilema de los
en hendiduras y en la corteza y penetran por las hendiduras
La larva del coleóptero se desarrolla en la madera
xilema de los árboles provocando la muerte del árbol
1 mes de evolución
16 / X / 09
ModeloModelo
M. Galloprovincialis B. xylophilus Bacteria
¿? ¿Bacterias?Athrophalus ferus
Aphelenchoididae
Orthotomicus erosus
Proyecto UE (María Arias)Proyecto UE (María Arias)
Objetivos españoles
¿Que especies de Bursaphelenchus existe en españa?españa?
Otras especies associadas con insectos vectores
Estudiar patogenicidad. Origen de la patogenicidad ej nematodo bacteria o nematodo+bacterias delej nematodo, bacteria o nematodo+bacterias del género pseudamonas
¿Que bacterias hay asociada con los nematodes cuyos vectores son insectos?
Pine wilt disease in EuropePine wilt disease in Europe Nematode surveysNematode surveys
BursaphelenchusBursaphelenchus species reported in: species reported in:
P lP l ii FF GG ii UKUKPortugalPortugal SpainSpain FranceFrance GermanyGermany AustriaAustria UKUKB. hellenicusB. hellenicus B. eggersiB. eggersi B. leoniB. leoni B. borealisB. borealis B. abietinusB. abietinus ¿?¿?
B. hofmanniB. hofmanni B. fungivorusB. fungivorus B. mucronatusB. mucronatus B. eggersiB. eggersi B. eggersiB. eggersiB. hofmanniB. hofmanni B. fungivorusB. fungivorus B. mucronatusB. mucronatus B. eggersiB. eggersi B. eggersiB. eggersiB. hylobianumB. hylobianum B. hylobianumB. hylobianum B. naujaciB. naujaci B. eremusB. eremus B. hofmanniB. hofmanniB. leoniB. leoni B. leoniB. leoni B. pinasteriB. pinasteri B. fungivorusB. fungivorus B. hylobianumB. hylobianumB mucronatusB mucronatus B mucronatusB mucronatus B pinophilusB pinophilus B leoniB leoni B leoniB leoniB. mucronatusB. mucronatus B. mucronatusB. mucronatus B. pinophilusB. pinophilus B. leoniB. leoni B. leoniB. leoniB. pinasteriB. pinasteri B. pinasteriB. pinasteri B. sexdentatiB. sexdentati B. mucronatusB. mucronatus B. mucronatusB. mucronatusB. pinophilusB. pinophilus B. pinophilusB. pinophilus B. silvestrisB. silvestris B. pinasteriB. pinasteri B. paracorneolusB. paracorneolusB d iB d i B d iB d i B li hiB li hi B i iB i iB. sexdentatiB. sexdentati B. sexdentatiB. sexdentati B. poligraphiB. poligraphi B. pinasteriB. pinasteriB. teratospicularisB. teratospicularis B. teratospicularisB. teratospicularis B. sexdentatiB. sexdentati B. sexdentatiB. sexdentatiB. tusciaeB. tusciae B. silvestrisB. silvestrisB. xylophilusB. xylophilus B. teratospicularisB. teratospicularis
B. thailandaeB. thailandaeB. vallesianusB. vallesianus
PHRAME Pine Wilt Disease Technical Meeting PHRAME Pine Wilt Disease Technical Meeting –– Portugal 2006Portugal 2006
B. willibaldiB. willibaldi
Pine wilt disease in EuropePine wilt disease in Europe Nematode surveysNematode surveys
Insect / Insect / BursaphelenchusBursaphelenchus associationsassociationsInsectInsect NematodeNematode RefRefInsectInsect NematodeNematode Ref.Ref.Hylobius sp.Hylobius sp. Bursaphelenchus sp.Bursaphelenchus sp. Penas Penas et alet al. (2006). (2006)Hylurgus ligniperdaHylurgus ligniperda B. hellenicusB. hellenicus ““y g g py g g p
B. tusciaeB. tusciae ““Ips sexdentatusIps sexdentatus B. hellenicusB. hellenicus ““
B. sexdentatiB. sexdentati ““Monochamus galloprovincialisMonochamus galloprovincialis B. mucronatusB. mucronatus ““
B xylophilusB xylophilus ““B. xylophilusB. xylophilusOrthotomicus erosusOrthotomicus erosus B. fungivorusB. fungivorus Arias Arias et alet al. (2005). (2005)
B. sexdentatiB. sexdentati Penas Penas et alet al. (2006). (2006)B. teratospicularis B. teratospicularis ““
Pityogenes sp.Pityogenes sp. B. leoni ?B. leoni ? Penas Penas et alet al. (2006). (2006)
PHRAME Pine Wilt Disease Technical Meeting PHRAME Pine Wilt Disease Technical Meeting –– Portugal 2006Portugal 2006
Tomicus piniperdaTomicus piniperda B. hellenicusB. hellenicus ““
Killing trees is unusual for this Killing trees is unusual for this ggnematode, except outside its native nematode, except outside its native
rangerangerange.range.How does it do this?How does it do this?
Secuencia de interacciónSecuencia de interacciónSecuencia de interacciónSecuencia de interacción
E t dE t d i d ñ di d ñ d1.1. EntradaEntrada: via daño causado por : via daño causado por alimentación en ramas vivasalimentación en ramas vivas
2.2. MovimientoMovimiento: ¿donde va el nematodo : ¿donde va el nematodo despues de la entrada inicial ?despues de la entrada inicial ?pp
3.3. ReproduciónReprodución: donde se reproduce el : donde se reproduce el nematodo?nematodo?nematodo?nematodo?
4.4. Respuesta del arbolRespuesta del arbol: ¿respuestas : ¿respuestas fisiológicas? porq é alg nos m erenfisiológicas? porq é alg nos m erenfisiológicas? ¿porqué algunos mueren y fisiológicas? ¿porqué algunos mueren y otros no?otros no?
EntradaEntradaNematodos en forma larvas dauer
é d h id bientran a través de heridas abiertas. Las larvas mudan hasta conventirse en adulto (en unas 24 horas)en adulto (en unas 24 horas)
Invasión por nematodosInvasión por nematodosMonochamus se alimenta de los tejidos hasta llegar al xylema ocasioando un herida que permite la introducciónxylema, ocasioando un herida que permite la introducción del nematodo dentro del árbol.
Rutas Potenciales para la dispersión Rutas Potenciales para la dispersión dentro del arbol.dentro del arbol.
P i t t t hl d lPrimary transport systems are phloem and xylem
Xylema: Upward flowPhloema: Downward flow Xylema: Upward flow, primarily water transport system
Phloema: Downward flow, primarily nutrient transport system
Outside
InsideCross section
Cavitación debido a la actividad d l t d l d lde los nematodos en la zona del cambium.
Movimiento en las canales de resina
Canales de resina con nematodos
Canales de resina con t dnematodos
Reproducion masiva del nematodo y cavitación
Mueren las celulas del xilema y parenquima axialp q
Nematodos virulentos y Nematodos virulentos y avirulentosavirulentos
Indicativo de agresividad / virulencia?
- Razas fisiológicas o patotipospatotipos
WP 6 Pathogenicit testWP 6 Pathogenicit test Pin s nigraPin s nigraWP 6: Pathogenicity test, WP 6: Pathogenicity test, Pinus nigraPinus nigra
Class 1
1 25 %
Class 2
26 50 %
Class 3
51 75 %
Class 4
76 99 %1 – 25 % 26 – 50 % 51 – 75 % 76 – 99 %Federal Biological Research Centre for Agriculture and Forestry T. SchröderDepartment of National and International Plant Health
WP 6 Pathogenicit testWP 6 Pathogenicit test Pin s nigraPin s nigraWP 6: Pathogenicity test, WP 6: Pathogenicity test, Pinus nigraPinus nigra
C l Cl 5Control Class 5
Federal Biological Research Centre for Agriculture and Forestry T. SchröderDepartment of National and International Plant Health
PatogeneicidadPatogeneicidad Pin s sppPin s sppPatogeneicidad Patogeneicidad –– Pinus sppPinus spp
80
90
100
%
50
60
70
mor
talit
y in
%
10
20
30
40
tree
Pinus nigra
us cembra
sylvestri
s
trobusste
rta Control
USAChina
Portugal0
10
PPinus
Pinus sylv
Pinus stro
Pinus pinaste
Pinus radiata
Pinus mugo
Pinus pinea
Pinus halepensis
Control
WP 6 Pathogenicit testWP 6 Pathogenicit testWP 6: Pathogenicity test WP 6: Pathogenicity test –– B. xylophylus populationsB. xylophylus populations
Pi i d d t (03 J l 06 O t 2003)Pinus nigra , dead trees (03. July - 06. Oct. 2003)
90
100
P t l
60
70
80
90
s in
%
PortugalChinaUSA
30
40
50
60
ad tr
ees USA
Control
0
10
20
30
dea
00 2 4 6 8 10 12 14
weekly examination
Federal Biological Research Centre for Agriculture and Forestry T. SchröderDepartment of National and International Plant Health
WP 6 Pathogenicit testWP 6 Pathogenicit testWP 6: Pathogenicity test WP 6: Pathogenicity test -- B. xylophylus populationsB. xylophylus populations
Pi l i d d (09 J l 08 O 2003)Pinus sylvestris , dead trees (09. July - 08. Oct. 2003)
100
70
80
90
s in
%
PortugalChinaUSA
30
40
50
60
ad tr
ees USA
Control
0
10
20
30
dea
00 2 4 6 8 10 12
weekly examination
Federal Biological Research Centre for Agriculture and Forestry T. SchröderDepartment of National and International Plant Health
WP 6 Pathogenicit testWP 6 Pathogenicit testWP 6: Pathogenicity test WP 6: Pathogenicity test -- B. xylophylus populationsB. xylophylus populations
Pinus pinaster , dead trees (14. July - 15. Oct. 2003)
90
100
Portugal
60
70
80
s in
%
PortugalChinaUSA
30
40
50
ead
tree
s
Control
0
10
20de
00 2 4 6 8 10 12
weekly examination
Federal Biological Research Centre for Agriculture and Forestry T. SchröderDepartment of National and International Plant Health
WP 6: Pathogenicity testWP 6: Pathogenicity test -- B xylophylus populationsB xylophylus populationsWP 6: Pathogenicity test WP 6: Pathogenicity test B. xylophylus populationsB. xylophylus populations
Pi d d t (21 J l 21 O t 2003)Pinus mugo , dead trees (21. July - 21. Oct. 2003)
90
100
Portugal
60
70
80
es in
%
PortugalChinaUSA
30
40
50
ead
tree Control
0
10
20
30
de
00 2 4 6 8 10 12
weekly examination
Federal Biological Research Centre for Agriculture and Forestry T. SchröderDepartment of National and International Plant Health
WP 6: Pathogenicity test WP 6: Pathogenicity test -- other other g yg yspeciesspecies
80
90
100
50
60
70
20
30
40
50
dua riUSA
ChinaPortugal
0
10
20
Larix decidua
Larix kaempferi
Abies alba
Picea abies
Control
Federal Biological Research Centre for Agriculture and Forestry T. SchröderDepartment of National and International Plant Health
WP 6 Pathogenicit testWP 6 Pathogenicit testWP 6: Pathogenicity test WP 6: Pathogenicity test -- B. xylophylus populationsB. xylophylus populations
L i d id d d t (17 J l 16 O t 2003)Larix decidua, dead trees (17. July - 16. Oct. 2003)
90
100
Portugal
60
70
80
es in
%
PortugalChinaUSAC t l
30
40
50
ead
tree Control
0
10
20
30
de
00 2 4 6 8 10 12
weekly examination
Federal Biological Research Centre for Agriculture and Forestry T. SchröderDepartment of National and International Plant Health
WP 6 Pathogenicit testWP 6 Pathogenicit testWP 6: Pathogenicity test WP 6: Pathogenicity test -- B. xylophylus populationsB. xylophylus populations
Lari kaempferi d d t (23 J l 20 O t 2003)Larix kaempferi , dead trees (23. July - 20. Oct. 2003)
90
100
Portugal
60
70
80
es in
%
PortugalChinaUSAC t l
30
40
50
ead
tree Control
10
20
30
de
00 2 4 6 8 10 12
weekly examination
Federal Biological Research Centre for Agriculture and Forestry T. SchröderDepartment of National and International Plant Health
Objetivos futurosObjetivos futuros
Análisis filogenéticos y detección de razas o biotipos
Modelización y prognosis de su distribuciónModelización y prognosis de su distribución
C id i b l i d B. Consideraciones sobre el origen de B. xylophilus encontrado en España
¡¡¡Muchas gracias!!!¡¡¡ g
M í A iMaría Arias
Lee Robertson
Miguel Escuer
Antonio Bello