Milk–Tween agar, a semiselective medium for isolation and differentiation of Pseudomonas syringae pv. syringae, Pseudomonas syringae pv. phaseolicola and Xanthomonas axonopodis pv. phaseoli

Goszczynska, Teresa; Serfontein, J. J.

doi:10.1016/s0167-7012(98)00005-0

Cited by 49 publications

(36 citation statements)

References 18 publications

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“…nov. (Goszczynska & Serfontein, 1998); 0.5 g l 21 tyrosine; and 1 % (w/v) CM-cellulose (Sigma) (Teather & Wood, 1982). Hydrolytic activities were determined by the formation of clearance zones around colonies (starch, casein, tyrosine and CM-cellulose) or turbid zones around colonies (Tweens 20,40,60 and 80).…”

Section: Marinobacterium Nitratireducens Cn44 T (Eu573965)mentioning

confidence: 99%

Endozoicomonas atrinae sp. nov., isolated from the intestine of a comb pen shell Atrina pectinata

Hyun

Shin

Kim

et al. 2014

International Journal of Systematic and Evolutionary Microbiology

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Endozoicomonas atrinae sp. nov., isolated from the intestine of a comb pen shell Atrina pectinata A novel bacterium, designated strain WP70 T , was isolated from the gut of a comb pen shell (Atrina pectinata) collected from the southern sea of Yeosu in Korea. The isolate was Gram-stainnegative, aerobic, non-motile and rod-shaped. Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain WP70 T belonged to the genus Endozoicomonas. The highest level of sequence similarity (98.4 %) was shared with Endozoicomonas elysicola MKT110 T . Optimal growth occurred in 2 % (w/v) NaCl at 30 6C and at pH 7. The major cellular fatty acids were summed feature 3 (C 16 : 1 v7c and/or C 16 : 1 v6c), summed feature 8 (C 18 : 1 v7c and/or C 18 : 1 v6c) and C 16 : 0 . The main respiratory quinone was Q-9. The polar lipids comprised phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol, three unidentified phospholipids, an unidentified aminolipid, an unidentified aminophospholipid and an unidentified lipid. The genomic DNA G+C content was 50.5 mol% and DNA-DNA hybridization values indicated ,11 % genomic relatedness to the closest species. Physiological, biochemical, chemotaxonomic and genotypic analyses indicated that strain WP70 T represents a novel species of the genus Endozoicomonas, for which the name Endozoicomonas atrinae sp. nov. is proposed. The type strain is WP70 T (5KACC 17474 T 5JCM 19190 T ).The genus Endozoicomonas belongs to the order Oceanospirillales within the class Gammaproteobacteria, and was first proposed by Kurahashi & Yokota (2007). Members of the genus Endozoicomonas are Gram-stain-negative and strictly aerobic or facultatively anaerobic rod-shaped bacteria, and at the time of writing there are five species of the genus with validly published names: Endozoicomonas elysicola (Kurahashi & Yokota, 2007), Endozoicomonas montiporae (Yang et al., 2010), Endozoicomonas numazuensis (Nishijima et al., 2013), Endozoicomonas euniceicola and Endozoicomonas gorgoniicola (Pike et al., 2013). All species of the genus Endozoicomonas were isolated from marine environments; sources include sponges (Nishijima et al., 2013), corals (Pike et al., 2013, Yang et al., 2010 and sea slugs (Kurahashi & Yokota, 2007).The comb pen shell (Atrina pectinata) belongs to the Pinnidae family, and is a large wedge-or fan-shaped suspension-feeding bivalve (Okutani, 1997), which is a valuable seafood resource in Korea, China and Japan. It is distributed throughout the coastal waters of East Asia, and its habitat ranges from muddy-to-sandy sediments to tidal flats or shallow subtidal environments up to 20 m deep (Yurimoto et al., 2003). Although three bacterial species have been isolated from Atrina pectinata-associated sources, Agromyces atrinae and Kocuria atrinae from fermented Atrina pectinata seafood (Park et al., 2010a, b) and Polaribacter atrinae from the gut of Atrina pectinata (Hyun et al., 2014), information about the microbiota within live comb pen shells is limited. Here, a novel Endozoicomonaslike ...

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Section: Marinobacterium Nitratireducens Cn44 T (Eu573965)mentioning

confidence: 99%

Endozoicomonas atrinae sp. nov., isolated from the intestine of a comb pen shell Atrina pectinata

Hyun

Shin

Kim

et al. 2014

International Journal of Systematic and Evolutionary Microbiology

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“…The colonies formed on the semi-selective medium MT are creamy white, flat and circular after four-five days of incubation (Goszczynska and Serfontein, 1998;ISF, 2006;Kurowski and Remeeus, 2008). Many colonies of Psp on MT can form a pale-blue fluorescent pigment when using UV light (ISF, 2006).…”

Section: Discussionmentioning

confidence: 99%

“…Many colonies of Psp on MT can form a pale-blue fluorescent pigment when using UV light (ISF, 2006). Goszczynska and Serfontein (1998) described the use of the semi-selective medium MT for routine seed testing in the laboratory and for the detection and distinction of all bacterial pathogens screened for in dry bean seed and plant material. This study confirmed the advantage of the MT medium, which allowed simultaneous detection of Xap and Psp on the tested bean cultivar Oplenac, where seeds were infected with both of the bacteria.…”

Section: Discussionmentioning

confidence: 99%

“…After incubating, 0.1 mL of undiluted extracts obtained from the seeds and their 10-fold dilution series (to 10 -5 ) were plated onto the surface of two semi-selective media, Modified Sucrose Peptone Agar (MSP) and Milk Tween Agar (MT), described by Mohan and Schaad (1987) and Goszczynska and Serfontein (1998), respectively. The series of dilution were prepared with the aim to get single colonies of Psp which could be morfologically differentiated from other non-targeted organisms.…”

Section: Isolation On Semi-selective Mediamentioning

confidence: 99%

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Application of semi-selective mediums in routine diagnostic testing of Pseudomonas savastanoi pv. phaseolicola on common bean seeds

Popović

Milovanović²,

Aleksić

et al. 2012

Sci. agric. (Piracicaba, Braz.)

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Halo blight, caused by Pseudomonas savastanoi pv. phaseolicola (Psp), is considered to be an important bacterial disease on common bean (Phaseolus vulgaris L.) in Serbia. Use of pathogen-free seeds is one of the most effective control measures against this disease. The aim of this study was to evaluate a detection method for Psp on untreated common bean seeds (23 genotypes) from commercial crops grown within Serbia. Detection of this pathogen was made by plating onto the modified sucrose peptone (MSP) and Milk Tween (MT) semi-selective mediums from soaked whole common bean seed. Colonies growing on the MSP medium were light yellow, convex and shiny, whereas on the MT medium, they were creamy white, flat and circular. The pathogenicity of the obtained strains was confirmed by the inoculation of germinated bean seed. The isolates recovered from the seed assay were further confirmed to be Psp by using both Enzyme-linked immunosorbent assay (ELISA) and Polymerase Chain Reaction (Nested-PCR) detection methodologies. The International Seed Testing Association (ISTA) method selected for this work was found to be effective in detecting the presence of Psp in common bean seed. The bacterium Psp was detected in only two of the 23 seed samples analyzed by this method, which shows that the bacterium is not widespread in Serbia.

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“…Plate assays consist of plating seeds or seed extracts on a semi-selective agar medium (Sheppard et al, 1989;Goszczynska & Serfontein, 1998 …”

Section: Introductionmentioning

confidence: 99%

Detection of Xanthomonas axonopodis pv. phaseoli in bean seeds by flow cytometry, immunostaining and direct viable counting

Tebaldi¹,

Peters²,

Souza³

et al. 2010

Trop. plant pathol.

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Flow cytometric analysis of immuno-stained cells (immuno-FCM) was compared to immunofluorescence microscopy (IF) and dilution plating on a semi-selective medium, for quantitative detection of Xanthomonas axonopodis pv. phaseoli (Xap) in bean seed extracts. Cell concentrations of Xap between 10 3 -10 7 CFU/mL were added to healthy bean seed extracts. A flow cytometry sorting procedure was developed to separate immuno-stained Xap cells from crude seed extracts and confirming by PCR. FCM was evaluated for direct viable counting (DVC) of Xap using combinations of propidium iodide (PI) and carboxy fluorescein diacetate (cFDA) or PI and SYTO 9 and also the combination of immuno-FCM and PI. Dilution plating and IF allowed detection of Xap in bean seed extracts in a range of 10 3 -10 6 CFU/mL and immuno-FCM from 10 4 -10 6 CFU/mL. Sorted cells could be detected in crude seed extracts by PCR without further extraction. FCM also allowed quantification of viable cells of Xap after DVC procedures; the red fluorescent dye propidium iodide was used to identify dead cells in combination with the green fluorescent dyes cFDA or SYTO 9, these identifying live cells. The combination of immuno-FCM and PI could be more promising and reliable to detect this pathogen in seeds. Key words: seed pathology, flow sorting, PCR-amplification, viability probes, immunofluorescence, bacteria. RESUMO Detecção de Xanthomonas axonopodis pv. phaseoli em sementes de feijão usando citometria de fluxo em combinação com anticorpo e sondas fluorescentes de viabilidadeA combinação do uso do citômetro de fluxo (FCM) e de anticorpo policlonal (imuno-FCM) foi comparada à microscopia de imunofluorescência (IF) e ao plaqueamento em meio de cultura semi-seletivo, para a detecção de Xanthomonas axonopodis pv. phaseoli (Xap) em sementes de feijão. Concentrações de Xap variando de 10 3 a 10 7 CFU/mL foram adicionados aos extratos de sementes. Um método de separação pelo citômetro de fluxo foi desenvolvido para a detecção de Xap em extratos de semente e posterior confirmação por PCR. Para avaliação da viabilidade das células foram usadas sondas fluorescentes, iodeto de propídio (PI)/carboxi diacetato de fluoresceína (cFDA) e PI/SYTO 9 e também, a combinação de imuno-FCM e PI. Em meio semi-seletivo e IF foram detectadas 10 3 -10 6 UFC/mL e no FCM 10 4 -10 6 UFC/mL, em extratos de sementes artificialmente infestados. Xap somente foi detectada em extratos de sementes por PCR, após o processo de separação pelo FCM. Foi possível pelo FCM a quantificação e identificação de células viáveis (verde fluorescente) e células mortas (vermelho fluorescente) de Xap, pelas sondas cFDA/SYTO 9 e PI, respectivamente. A combinação de immuno-FCM e PI poderá ser uma técnica promissora e segura para a detecção deste patógeno em sementes. Palavras-chave: patologia de sementes, PCR, sondas de viabilidade, imuno-fluorescência, bactéria. INTRODUCTIONXanthomonas axonopodis pv. phaseoli (Smith) Vauterin, Hoste, Kersters & Swinges (Xap) is a seedtransmitted plant-pathogenic bacterium w...

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Milk–Tween agar, a semiselective medium for isolation and differentiation of Pseudomonas syringae pv. syringae, Pseudomonas syringae pv. phaseolicola and Xanthomonas axonopodis pv. phaseoli

Cited by 49 publications

References 18 publications

Endozoicomonas atrinae sp. nov., isolated from the intestine of a comb pen shell Atrina pectinata

Endozoicomonas atrinae sp. nov., isolated from the intestine of a comb pen shell Atrina pectinata

Application of semi-selective mediums in routine diagnostic testing of Pseudomonas savastanoi pv. phaseolicola on common bean seeds

Detection of Xanthomonas axonopodis pv. phaseoli in bean seeds by flow cytometry, immunostaining and direct viable counting

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