ORIGINAL ARTICLE
Comparison of methods used in the recovery of Phylloplane bacteria: a case study of Pseudomonas savastanoi pv. savastanoi applied to the Phylloplane of Olea europaea sub-species
 
More details
Hide details
1
Department of Science and Technology for Agriculture, Forestry, Nature and Energy (DAFNE), Tuscia University, Via San Camillo de Lellis, 01100 Viterbo, Italy
2
Fruit Development Directorate (FDD), Kirtipur, Kathmandu, Nepal
CORRESPONDING AUTHOR
Jay Ram Lamichhane
Department of Science and Technology for Agriculture, Forestry, Nature and Energy (DAFNE), Tuscia University, Via San Camillo de Lellis, 01100 Viterbo, Italy
Submission date: 2013-04-20
Acceptance date: 2014-01-14
 
Journal of Plant Protection Research 2014;54(1):22–27
KEYWORDS
TOPICS
ABSTRACT
An efficient and accurate method of sampling, recovery, and enumeration of epiphytic bacterial populations are of fundamental importance for their precise estimation. In this study, effectiveness and reliability of processing methods, sampling type, sample storage, and plating techniques, for the recovery of the epiphytic bacterial populations, were evaluated. Pseudomonas savastanoi pv. savastanoi, the causal agent of olive knot disease, and two olive sub-species were used. Bulk-leaf sampling allowed for a higher number of the bacterial recovery. The use of a lab blender was the most effective and reliable method among the four commonly used processing methods. A short storage of leaf samples was possible through freezing. Bacterial survival was not influenced as long as the samples were processed within 5 days and frozen without a buffer. No difference was observed among the drop and spread platings, suggesting that one of them can be used and the choice depends on the cost and time required.
CONFLICT OF INTEREST
The authors have declared that no conflict of interests exist.
 
REFERENCES (25)
1.
Balestra G.M., Lamichhane J.R., Kshetri M.B., Mazzaglia A., Varvaro L. 2009. First report of Pseudomonas savastanoi pv. savastanoi on olive in Nepal. Plant Pathol. 58 (2): 393–393.
 
2.
Beattie G.A. 2002. Leaf surface waxes and the process of leaf colonization by microorganisms. p. 3–26. In: “Phyllosphere Microbiology” (S.E. Lindow, E.I. Hecht-Poinar, V.J. Elliott, eds.). APS Press, Minnesota, 408 pp.
 
3.
Conlin K.C., McCarter S.M. 1983. Effectiveness of selected chemicals in inhibiting Pseudomonas syringae pv. tomato in vitro and in controlling bacterial speck. Plant Dis. 67 (6): 639–644.
 
4.
Crosse J.E. 1959. Bacterial canker of stone fruits: IV. Investigation of a method for measuring the inoculums potential of cherry trees. Ann. Appl. Biol. 47 (2): 306–317.
 
5.
Donegan K., Matyac C., Seidler R., Porteous A. 1991. Evaluation of methods for sampling, recovery and enumeration of bacteria applied to the phylloplane. Appl. Environ. Microbiol. 57 (1): 51–56.
 
6.
Ercolani G.L. 1971. Presenza epifitica di Pseudomonas savastanoi (E.F. Smith) Stevens sull’Olivo, in Puglia. Phytopathol. Mediterr. 10: 130–132.
 
7.
Ercolani G.L. 1978. Pseudomonas savastanoi and other bacteria colonizing the surface of olive in the field. J. Gen. Microbiol. 109 (2): 245–257.
 
8.
Ercolani G.L. 1979. Distribuzione di Pseudomonas savastanoi sulle foglie dell’olivo. Phytopathol. Mediterr. 18: 85–88.
 
9.
Ercolani G.L. 1991. Distribution of epiphytic bacteria on olive leaves and the influence of leaf age and sampling time. Microb. Ecol. 21 (1): 35–48.
 
10.
Hirano S.S., Nordheim E.V., Arny D.C., Upper C.D. 1982. Lognormal distribution of epiphytic bacterial populations on leaf surfaces. Appl. Environ. Microbiol. 44 (3): 695–700.
 
11.
Jacques M.A. 1996. The effect of leaf sge and position on the dynamics of microbial populations on aerial plant surfaces. p. 233–248. In: “The Aerial Plant Surface Microbiology”( C.E. Morris, C. Nguyen, P.C. Nicot, eds.). NetLibrary, Incorporated, Netherlands, 307 pp.
 
12.
Knudsen G.R., Walter M.V., Porteous L.A., Prince V.J., Armstrong J.L., Seidler R.J. 1988. Predictive model of conjugative plasmid transfer in the rhizosphere and phyllosphere. Appl. Environ. Microbiol. 54 (2): 343–347.
 
13.
Lamichhane J.R. 2011. Phytobacteriological studies related to the introduced and native olive sub-species in Nepal. Ph.D. Thesis, Tuscia, University Viterbo, Italy, 99 pp.
 
14.
Lamichhane J.R., Varvaro L. 2013. Epiphytic Pseudomonas savastanoi pv. savastanoi can infect and cause olive knot disease on Olea europaea subsp. cuspidata. Aust. Plant Pathol. 42 (2): 219–225.
 
15.
Lamichhane J.R., Balestra G.M., Varvaro L. 2010a. Phytobacteriological investigation on Olea spp. in different districts of Nepal. Petria 20 (2): 147–148.
 
16.
Lamichhane J.R., Balestra G.M., Varvaro L. 2010b. Responses of tomato cultivars largely cultivated in Nepal to Pseudomonas syringae p v. tomato. Phytopathol. Mediterr. 49: 406–413.
 
17.
Lindow S.E., Brandl M.T. 2003. Microbiology of the phyllosphere. Appl. Environ. Microbiol. 69 (4): 1875–1883.
 
18.
Lindow S.E., Arny D.C., Upper C.D. 1978. Distribution of ice nucleation active bacteria on plants in nature. Appl. Environ. Microbiol. 36 (6): 831–838.
 
19.
Lindow S.E., Knudsen G.R., Seidler R.J., Walter M.V., Lambou V.W. 1988. Aerial dispersal and epiphytic survival of Pseudomonas syringae during a pretest for the release of genetically engineered strains into the environment. Appl. Environ. Microbiol. 54 (6): 1557–1563.
 
20.
Meynell G.G., Meynell E. 1965. Theory and Practice in Experimental Bacteriology. Cambridge University Press, USA, 288 pp.
 
21.
Surico G. 1993. Scanning electron microscopy of olive and oleander leaves colonized by Pseudomonas syringae subsp. savastanoi. J. Pytopathol. 138 (1): 31–40.
 
22.
Quesada J.M., Garcia A., Bertolini E., Lopez M.M., Penyalver R. 2007. Recovery of Pseudomonas savastanoi pv. savastanoi from symptomless shoots of naturally infected olive trees. Int. Microbiol. 10 (2): 77–84.
 
23.
Quesada J.M., Penyalver R., Perez-Panades J., Salcedo C.I., Carbonell E.A., Lopez M.M. 2010. Comparison of chemical treatments for reducing epiphytic Pseudomonas savastanoi pv. savastanoi populations and for improving subsequent control of olive knot disease. Crop Prot. 29 (12): 1413–1420.
 
24.
Varvaro L., Surico G. 1987. Multiplication of wild types of Pseudomonas Savastanoi pv. savastanoi (Smith) Young et al. and their indolacetic deficient mutants in olive tissues. p. 556–565. In: “Plant Pathogenic Bacteria” (E.L. Civerolo, A. Collmer, R.E. Davis, A.G. Gillaspie, eds.). Springer, Dordrecht.
 
25.
Zagory D., Lindow S.E., Parameter J.R. 1983. Toxicity of smoke to epiphytic ice nucleation active bacteria. Appl. Environ. Microbiol. 46 (1): 114–119.
 
eISSN:1899-007X
ISSN:1427-4345