Real-time loop-mediated isothermal amplification: an early-warning tool for quarantine plant pathogen detection

Aglietti, Chiara; Luchi, Nicola; Pepori, Alessia Lucia; Bartolini, Paola; Pecori, Francesco; Raio, Aida; Capretti, Paolo; Santini, Alberto

doi:10.1186/s13568-019-0774-9

AMB Express

Table 1 List of isolates used in this study

From: Real-time loop-mediated isothermal amplification: an early-warning tool for quarantine plant pathogen detection

Species	Isolate code	Group^b	Host	Origin	Collector^c	Molecular assay^d
						LAMP^e		qPCR^f
						t_amp (min:s)	Ta (°C)	Detection
X. fastidiosa assay
Xylella fastidiosa subsp. pauca	Co.Di.Ro^a	T	Olea europaea	Italy	M. Saponari	7:15	88.98	+
Xylella fastidiosa subsp. fastidiosa	Xff^a	T	Prunus dulcis	USA	J. Chen	14:20	88.78	+
Xylella fastidiosa subsp. multiplex	Xfm^a	T	Liquidambar styraciflua	USA	S. Russell	7:00	88.83	+
Pseudomonas savastanoi pv. savastanoi	ITM05^a	CHC	Olea europea	Italy	G. Marchi	–	–	−
Pantoea agglomerans	PaFL1^a	CHC	Olea europea	Italy	G. Marchi	–	–	−
Pseudomonas fluorescens	KL218^a	CHC	Actinidia deliciosa	Italy	G. Marchi	–	–	−
Xanthomonas arboricola pv. pruni	Xap	PR	Prunus laurocerasus	Italy	A. Raio	–	–	−
Pseudomonas savastanoi pv. nerii	Ps.sav	CHC	Nerium oleander	Italy	A. Raio	–	–	−
Pseudomonas koreensis	KL217^a	NP, O	Actinidia deliciosa	Italy	G. Marchi	–	–	−
Pseudomonas syringae	KL34^a	O	Actinidia deliciosa	Italy	G. Marchi	–	–	−
Pseudomonas syringae	KL32^a	O	Actinidia deliciosa	Italy	G. Marchi	–	–	−
Pseudomonas viridiflava	KL24^a	O	Actinidia deliciosa	Italy	G. Marchi	–	–	−
Pseudomonas mediterranea	C5P1rad1^a	O	Chrysanthemum sp.	Italy	M. Fiori	–	–	−
Pseudomonas corrugata	C2P1 rad^a	O	Chrysanthemum sp.	Italy	M. Fiori	–	–	−
Pseudomonas syringae pv. photiniae	CFBP2899^a	O	Photinia glabra	Japan	CFBP	–	–	−
Pectobacterium carotovorum	C24^a	O	Zantedeschia aethiopica	Italy	G. Marchi	–	–	−
Pectobacterium carotovorum	C6^a	O	Zantedeschia aethiopica	Italy	G. Marchi	–	–	−
Pantoea agglomerans pv. gypsophilae	Ehg824-1^a	O	Gypsophila paniculata	Israel	G. Marchi	–	–	−
Pseudomonas syringae pv. actinidiae	KL103^a	O	Actinidia deliciosa	Italy	G. Marchi	–	–	−
Pseudomonas syringae pv. tabaci	GSPB1209^a	O	Nicotiana sp.	Italy	G. Marchi	–	–	−
Pseudomonas syringae pv. viburni	CFBP1702^a	O	Viburnum sp.	USA	CFBP	–	–	−
Sphingomonas sp.	KVPT7FA^a	NP, CHC	Actinidia deliciosa	Italy	G. Marchi	–	–	−
Stenotrophomonas malthopylia	St	NP, PR	Capsicum annum	Italy	A. Raio	–	–	−
Xanthomonas campestris pv. campestris	Xcc	PR	Brassica spp.	Italy	A. Raio	–	–	−
Xanthomonas arboricola pv. juglandis	Xaj	PR	Juglans regia	Italy	A. Raio	–	–	−
Erwinia amylovora	Ea12	O	Pyrus communis	Italy	C. Cainelli	–	–	−
Agrobacterium tumefaciens	LMG37	O	Prunus spp.	USA	BCCM/LMG	–	–	−
Agrobacterium vitis	CFBP5523	O	Vitis vinifera	Australia	CFBP	–	–	−
C. platani assay
Ceratocystis platani	CBS117355	T	Platanus sp.	France	CBS-KNAW	10:08	88.14	+
Ceratocystis platani	Cp3	T	Platanus × acerifolia	Italy	IPSP-CNR	12:15	88.55	+
Ceratocystis platani	Cp6	T	Platanus × acerifolia	Italy	IPSP-CNR	9:08	88.30	+
Ceratocystis platani	G160	T	Platanus × acerifolia	Turkey	T. Dogmus	16:58	88.93	+
Ceratocystis platani	DB203	T	Platanus × acerifolia	Turkey	T. Dogmus	14:08	88.78	+
Ceratocystis platani	CBS115162	T	Platanus occidentalis	USA	CBS-KNAW	8:00	88.83	+
Ceratocystis platani	Cp24	T	Platanus × acerifolia	Italy	IPSP-CNR	8:00	88.43	+
Ceratocystis fimbriata	CBS 115167	PR	Syngonium sp.	USA	CBS-KNAW	10:05	88.45	−
Ceratocystis fimbriata	CBS 118126	PR	Syzgium aromaticum	Sulawesi	CBS-KNAW	15:10	88.70	−
Ceratocystis fimbriata	CBS 115175	PR	Mangifera indica	Brazil	CBS-KNAW	9:30	88.20	−
Ceratocystis fimbriata	CBS 115174	PR	Eucalyptus sp.	Brazil	CBS-KNAW	14:00	88.50	−
Ceratocystis fimbriata	CBS 115171	PR	Colocasia esculenta	Brazil	CBS-KNAW	10:13	88.60	−
Ceratocystis fimbriata	CBS 74040	PR	Crotolaria juncea	Brazil	CBS-KNAW	13:38	88.00	−
Sarcodontia pachyodon	Sp5	O, CHC	Platanus × acerifolia	Italy	IPSP-CNR	–	–	−
P. ramorum assay
Phytophthora ramorum	Pram^a	T	Rhododendron sp.	Greece	P. Tsopelas	7:58	88.68	+
P. ramorum	LSVM123^a	T	Rhododendron sp.	France	R. Ioos-N. Schenck	7:07	88.48	+
P. ramorum	LSVM362^a	T	Rhododendron sp.	France	R. Ioos-N. Schenck	6:15	88.73	+
P. ramorum	LSVM386^a	T	Rhododendron sp.	France	R. Ioos-N. Schenck	6:72	88.68	+
P. ramorum	LSVM390^a	T	Rhododendron sp.	France	R. Ioos-N. Schenck	7:00	88.53	+
P. ramorum	LSVM391^a	T	Leucothoe sp.	France	R. Ioos-N. Schenck	6:30	88.53	+
P. ramorum	LSVM401^a	T	Rhododendron sp.	France	R. Ioos-N. Schenck	7:07	88.53	+
P. ramorum	LSVM402^a	T	Rhododendron sp.	France	R. Ioos-N. Schenck	7:00	88.53	+
P. ramorum	LSVM405^a	T	Rhododendron sp.	France	R. Ioos-N. Schenck	7:15	88.48	+
P. lateralis	Plat^a	PR	Chamaecyparis lawsoniana	France	C. Robin	9:00	88.43	+
P. alni subsp. uniformis	Ph68	PR	Alnus cordata	Italy	G. P. Barzanti	23:27	90.47	−
P. cactorum	PCA1^a	PR	Aesculus hippocastanum	Germany	J. Schumacher	–	–	−
P. × cambivora	Ph21^a	PR	Castanea sativa	Italy	A. Vettraino	25:22	90.67	−
P. cinnamomi	28SA	PR	Laurus nobilis	Italy	IPSP-CNR	16:37	89.62	−
P. cinnamomi	Ncfc^a	PR	Unknown	Italy	IPSP-CNR	–	–	−
P. cryptogea	13SA	PR	Prunus laurocerasus	Italy	IPSP-CNR	22:07	89.78	−
P. citricola	51RC	PR	Viburnum lucidum	Italy	IPSP-CNR	–	–	−
P. citricola	Pcl1^a	PR	Unknown	Germany	T. Jung	27:27	89.82	−
P. citrophthora	33SB	PR	Euonymus spp.	Italy	IPSP-CNR	20:05	89.23	−
P. citrophthora	Ph9^a	PR	Convolvolus sp.	Italy	S. O. Cacciola	18:15	89.58	−
P. europaea	PE1^a	PR	Unknown	Germany	T. Jung	15:12	90.42	−
P. foliorum	2015–1454^a	PR, CHC	Rhododendron	UK	A. Pérez-Sierra	11:15	89.08	−
P. gonapodyides	PG7^a	PR	Quercus robur	Germany	S. Leonhard	–	–	−
P. gonapodyides	IHTM	PR	Alnus cordata	Italy	IPSP-CNR	–	–	−
P. megasperma	Ph78	PR	Prunus avium	Italy	G. P. Barzanti	–	–	−
P. megasperma	PMI^a	PR	Quercus robur	Germany	S. Leonhard	–	–	−
P. nicotianae	1RB	PR	Myrtus communis	Italy	IPSP-CNR	–	–	−
P. palmivora	44RC	PR	Prunus laurocerasus	Italy	IPSP-CNR	–	–	−
P. quercina	PQ4^a	PR	Quercus robur	Germany	S. Leonhard/J. Schumacher	–	–	−
P. syringae	Psy2^a	PR	Unknown	Germany	J. Schumacher	17:45	89.48	−
Elongisporangium anandrum	PYA^a	O	Quercus robur	Germany	S. Leonhard	–	–	−
Phytopythium litorale	40SB	O	Prunus laurocerasus	Italy	IPSP-CNR	–	–	−
Elongisporangium undulatum	76SB	O	Cupressus sempervirens	Italy	IPSP-CNR	–	–	−
Mortariella sp.	26RA	O	Arbutus unedo	Italy	IPSP-CNR	–	–	−
Diplodia mutila	Dm	O	Quercus spp.	Italy	IPSP-CNR	–	–	−
D. pinea	128	O	Pinus resinosa	USA	M. A. Palmer	–	–	−
D. scrobiculata	124	O	Pinus resinosa	USA	M. A. Palmer	–	–	−
D. seriata	UCD 352	O	Vitis vinifera	USA	J. R. Urbez-Torres	–	–	−
D. seriata	WP-J10	O	Vitis vinifera	Australia	S. Savocchia	–	–	−
Geosmithia pallida	IVV7	O	Ulmus spp.	Italy	IPSP-CNR	–	–	−
Ophiostoma novo ulmi subsp. americana	HI72	O	Ulmus spp.	USA	IPSP-CNR	–	–	−

^aSamples provided as DNA
^bFor each molecular assay developed in this study different groups of isolates were tested: target species (T); phylogenetically related species (PR), CHC = common host colonizers species (CHC); out-group species (O)
^cCBS Centraalbureau voor Schimmelcultures (CBS) Fungal Biodiversity Centre—Royal Netherlands Academy of Arts and Sciences (KNAW), Utrecht, The Netherlands; CFBP French Collection of Plant Pathogenic Bacteria, INRA, France; IPSP-CNR Institute for Sustainable Plant Protection—National Research Council, Firenze, Italy; BCCM/LMG Bacteria Collection Laboratorium voor Microbiologie Universiteit Gent Bacteria Collection, Laboratorium voor Microbiologie, Universiteit Gent, Belgium
^dMolecular assays are referred to LAMP and qPCR described in Table 2
^et_amp amplification time, T_a annealing temperature, – not detected
^f+ positive, − negative

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