Species of Bursaphelenchus Fuchs, 1937 (Nematoda: Parasitaphelenchidae) associated with maritime pine in Portugal

Abstract

Summary -Species of Bursaphelenchus associated with maritime pine, Pinus pinaster, from Portugal -within and outside the quarantine restricted demarcated zone of B. xylophilus -are described and characterised both morphologically (LM and SEM) and with the use of molecular biology (ITS-RFLP). A new staining method for spicules is proposed. Species include B. hellenicus, B. hylobianum, B. leoni, B. pinophilus, B. sexdentati, B. tusciae, B. teratospicularis, B. xylophilus and Bursaphelenchus sp. 1. Bursaphelenchus hylobianum was collected from the insect Hylobius sp. The most frequent species in the demarcated zone, besides B. xylophilus, was Bursaphelenchus sp. 1. Morphological characterisation is compared with the original descriptions and discussed. The differentiation between B. pinophilus and B. sexdentati is not clear in the literature and is discussed. Since differentiation of B. xylophilus (mucronate form) from B. mucronatus, and B. pinophilus from B. sexdentati, as well as their juvenile forms, is almost im possible on the basis of morphological features, a molecular approach based on ITS-RFLPs was used. Ribosomal DNA containing the 5.8S gene, the internal transcribed spacer region 1 and 2, and partial regions of 18S and 28S gene were amplified by PCR. Restriction profiles of the amplified products generated species-specific differences, leading to the unambiguous identification of isolates belonging to B. xylophilus, B. mucronatus, B. sexdentati, B. tusciae and B. hylobianum.

The pinewood nematode, Bursaphelenchus xylophilus, in Madeira Island

“…The ITS-RFLP patterns for the Madeira Island isolates were specific to the species B. xylophilus ( Fig. 3) and identical to restriction patterns revealed by other PWN isolates from Continental Portugal and other parts of the world (Mota et al, 1999;Penas et al, 2004;Burgermeister et al, 2009;EPPO, 2009). The ITS restriction pattern with HhaI revealed a pattern (Fig.…”

“…Some females presented a terminal nipple-like extension resembling a mucro. Variation in the shape of the female tails, from round, digitate to mucronate has been detected in PWN isolates from Continental Portugal (Penas et al, 2004;Fonseca et al, 2008a;Penas et al, 2008). Mucronate tailed females of PWN have also been reported in Japan and North America (Wingfield et al, 1983;Bolla & Boschert, 1993).…”

Bursaphelenchus Fuchs, 1937 (Nematoda: Parasitaphelenchidae) species associated with Pinus species in northern Turkey

“…2B) vs. the absence of cucullus in the male spicules. However, these isolates show the same morphological differences cited for other B. sexdentati isolates collected in Greece (Lange et al, 2006), Italy (Ambrogioni & Caroppo, 1998) and Portugal (Penas et al, 2004). Previously, isolates A-14, A-58 and T-47 were also classified as Bursaphelenchus sp.…”

“…On the other hand, B. sexdentati is one of the most wide spread species among the European continent, being the most frequently encountered Bursaphelenchus species in southern European countries. Regarding several studies where B. sexdentati has been reported (Ambrogioni & Caroppo, 1998;Penas et al, 2004;Lange et al, 2006), this species shows a significant morphological variability when compared with other species of the genus (e.g. the shape of the female tail).…”

“…In the case of B. pinophilus the morphological data obtained in this study follow the original description of Brzeski & Baujard (1997), and the same ITS-RFLP patterns published for this species (Burgermeister et al, 2005;Lange et al, 2006). The known distribution of this species is quite limited, reported only in Germany (Braasch, 2001), Poland (Brzeski & Baujard, 1997) and Portugal (Penas et al, 2004). Although it has been reported to be associated with pine species (P. sylvestris and P. pinaster), it had never been associated with black pine (P. nigra).…”