Analysis of extracellular proteins of two Perkinsus spp. isolated from the softshell clam Mya arenaria in vitro

McLaughlin, Shawn; Elsayed, Ehab; Faisal, Mohamed

doi:10.1016/s0305-0491(00)00231-5

Cited by 17 publications

(27 citation statements)

References 20 publications

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“…cultures (Gauthier & Vasta 1993, Kleinschuster et al 1994, Dungan & Hamilton 1995, McLaughlin & Faisal 1998). In the current study, microbial contamination of cultures initiated from clam haemolymph was more frequent than in cultures initiated from gills or hypnospores.…”

Section: Discussionmentioning

confidence: 99%

“…cultures have been established from a variety of tissues including hearts, visceral ganglia and gills (Kleinschuster & Swink 1993, La Peyre et al 1993, McLaughlin & Faisal 1998, Coss et al 2001a) but there is little information on the optimal source of parasites to use. The success rate in establishing continuous cultures can be used as a criterion for selecting a tissue as a source of parasites.…”

Section: Discussionmentioning

confidence: 99%

“…These procedures varied greatly among researchers in the type of medium used, the culture conditions and the method used for the establishment of P. marinus primary cultures (reviewed in La Peyre 1996). P. marinus and P. chesapeaki were propagated in the culture medium JL-ODRP-1 originally formulated to resemble the composition of eastern oyster plasma (La Peyre et al 1993, McLaughlin & Faisal 1998. Commercial media including DMEM:Ham's F-12 and Leibowitz's L-15 medium, with various nutritional supplements, were used to culture P. marinus and P. andrewsi (Gauthier & Vasta 1993, Kleinschuster & Swink 1993, Kleinschuster et al 1994, Dungan & Hamilton 1995, Coss et al 2001a).…”

Section: Abstract: Perkinsus Atlanticus · Tapes Decussatus · In Vitrmentioning

confidence: 99%

“…Since the original culture of P. marinus, there have been reports of at least 2 additional Perkinsus species, P. chesapeaki and P. andrewsi, being propagated in vitro. McLaughlin & Faisal (1998) cultured 2 protozoa from the gill (G117) and haemolymph (H49) of an infected softshell clam (Mya arenaria). The gill isolate G117 was further characterised as a new Perkinsus species, P. chesapeaki (Kotob et al 1999, McLaughlin et al 2000.…”

Section: Introductionmentioning

confidence: 99%

“…Cultures of the 3 Perkinsus spp. have been established from various sources including hearts, visceral ganglia and haemolymph of eastern oysters, gills and haemolymph of a softshell clam, and heart and haemolymph of baltic clams (Gauthier & Vasta 1993, Kleinschuster & Swink 1993, La Peyre et al 1993, McLaughlin & Faisal 1998, Coss et al 2001a. Parasite hypnospores isolated from infected eastern oyster tissues incubated in Ray's fluid thioglycollate medium (RFTM) were used to quickly establish P. marinus continuous cultures from different geographical regions , Bushek & Allen 1996b.…”

Section: Introductionmentioning

confidence: 99%

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Continuous in vitro culture of the carpet shell clam Tapes decussatus protozoan parasite Perkinsus atlanticus

Casas¹,

Peyre²,

Reece³

et al. 2002

Dis. Aquat. Org.

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Continuous in vitro cultures of the clam Tapes decussatus parasite Perkinsus atlanticuswere established from infected gill fragments, infected haemolymph and parasite hypnospores isolated from infected gill fragments following incubation in Ray's fluid thioglycollate medium (RFTM). No continuous cultures could be initiated from P. atlanticus zoospores. Cultures initiated from hypnospores yielded the highest percentage of continuous cultures (100%, 6/6), followed by cultures initiated from gill fragments (93%, 43/46) and from haemolymph (30%, 3/10). Failures to establish continuous cultures were due to microbial contamination. The source of parasite influenced the success rate, the time taken to establish cultures and the size of cultured cells. In vitro proliferation of parasite cells was mainly by vegetative multiplication. Zoosporulation, yielding motile biflagellated zoospores, was observed at a low frequency (<1% of dividing cells) in every culture. Morphology of cultured cells examined with light and transmission electron microscopy corresponded to that of P. atlanticus found in clam tissues. Cultured cells enlarged in RFTM and stained blue-black with Lugol's solution, which are characteristics of the Perkinsus species cells. DNA sequences of the internal transcribed spacer (ITS) region of the ribosomal RNA gene complex matched those of P. atlanticus. All cultures were established in a medium designated JL-ODRP-2A that was similar in composition to the culture medium JL-ODRP-1 originally used to propagate Perkinsus marinus in vitro. Proliferation of P. atlanticus in vitro could be supported by the commercial culture medium (1:2 v/v) DME:Ham's F-12 with fetuin. KEY WORDS: Perkinsus atlanticus · Tapes decussatus · In vitro culture · Clam parasite · Ribosomal RNA gene complex · ITS · UltrastructureResale or republication not permitted without written consent of the publisher Dis Aquat Org 52: [217][218][219][220][221][222][223][224][225][226][227][228][229][230][231] 2002 spacer (ITS) region of the rRNA gene complex (Goggin 1994, de la Herrán et al. 2000, Robledo et al. 2000, Casas et al. 2002. Recently, Ordás & Figueras (1998) reported the in vitro culture of P. atlanticus from the haemolymph of an infected carpet shell clam. Identification of the cultured cells was based solely on their morphology (light and electron microscopy). Sequence analysis of the SSU small subunit rRNA gene, however, showed that those cultured cells did not correspond to a Perkinsus organism (Figueras et al. 2000). Continuous cultures of P. atlanticus, therefore, still need to be established.The development of continuous cultures of Perkinsus species can lead to a better understanding of this group of parasites . Cultures of the eastern oyster pathogen Perkinsus marinus, for example, have been used in a wide range of studies to address the parasite's environmental tolerance (Burreson et al. 1994, Dungan & Hamilton 1995, virulence , Bushek & Allen 1996a, Volety & Chu 1997, genetic composition (Reece et al. 1997, Reece et...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%