D. Hopkins. Xylella. in Laboratory guide for identification of by guy22

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                                       Xylella (2-1-06)
Much of information in this section extracted from:

D. Hopkins. Xylella. in: Laboratory guide for identification of plant pathogenic bacteria, 3 rd edition.

Alphaproteobacteria – Rhizobiales Rhizobiaceae (Agrobacterium)
                              Sphingomonadales Sphingomonadaceae (Rhizomonas)

Betaproteobacteria – Burkholderiales            Burkholderiaceae (Burkholderia, Ralstonia)
                                                      Comamonadaceae (Acidovorax)
                                                      Oxalobacteraceae (Herbaspirillum)

                                                        Genera incertae sedis (Xylophilus)

Gammaproteobacteria –           Pseudomonadales - Pseudomonadaceae
                                               (Pseudomonas, Rhizobacter)

                                        Xanthomonadales – Xanthomonadaceae
                                                   (Xanthomonas, Xylella)

                                        Enterobacteriales - Enterobacteriaceae
                                                      (Erwinia, Brenneria, Pectobacterium,
                                                      Pantoea)


Xylella fastidiosa

Xylella fastidiosa - 1987

Causal agent first observed in 1973 by Hopkins and Mollenhauer

First grown in axenic culture 1977 (Davis et al.)

First plant pathogenic bacterium sequenced in Brazil (CVC strain)

          Gram-negative, xylem-limited bacteria were first discovered in association with Pierce’s
disease of grapevine in 1973 and were called rickettsia-like bacteria. Previously, Pierce’s disease
and phony peach disease were believed to be caused by xylem-inhabiting viruses. Since then these
bacteria have been found to cause many other plant diseases that had no known causal agent. The
first of these xylem-limited bacteria was grown in axenic culture in 1978 (Davis et al.). In a
taxonomic study, the name Xylella fastidiosa was proposed to establish a new genus with one
species to include all of the known strains of fastidious, Gram-negative, xylem-limited bacteria.
Cells of X. fastidiosa are very narrow and cannot be seen by bright field microscopy, but only by



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phase contrast or dark field microscopy. Because the cells are small (0.2-0.4 by 1.0-4.0 m) their
size becomes very useful in their identification.

         This bacterium causes economic losses in many agriculturally important plants, including
grapevine, peach, plum, coffee, and citrus. It also causes leaf scorch and declines in many urban
shade trees, such as elm, oak, and sycamore. Some strains of X. fastidiosa have very wide host
ranges; however, many of the hosts may be symptomless. The bacterium is transmitted by sucking
insects that feed on xylem sap, such as the sharpshooter leafhoppers, but is not transmitted
mechanically from plant to plant.

         Although X. fastidiosa strains are normally not host specific, they are cell or tissue
specific. In the plant, they are restricted to xylem vessels or intercellular spaces of the xylem.
Bacteria also tend to accumulate in specific plant parts and the pattern of bacterial distribution often
is related to the type symptoms produced in that host.

Plant symptoms include:
- leaf scorch
- leaf chlorosis
- stunting
- dieback
- general decline

X. fastidiosa strains are difficult to isolate and grow in axenic culture. They do not grow on
most common bacterial media, but require specialized media, such as PD2 (Davis et al, 1980),
PW (Davis et al, 1981), CS20 (Chang and Walker, 1988), or BCYE (Wells et al (1981), Table 1).
Growth is slow on these specialized media with doubling times of 9 h to 2.0 days. This makes
contamination by faster growing bacteria a serious problem in the isolation of X. fastidiosa from
plant tissue. An important factor in the successful isolation is the selection of the host tissue for
isolation that contains the largest populations of the bacterium.

Serological techniques

                1)       Immunofluorescence can be used for rapid presumptive identification of
                culture of the phony peach strains of Xyella fastidiosa. In a few cases, regular
                direct- or indirect-immunofluorescence can be used to detect X. fastidiosa in plant
                extracts. However, the X. fastidiosa concentration is usually too low and the plant
                debris background too high for accurate diagnosis.

                2)      Membrane entrapment immunofluorescence of plant extracts (De
                        Lima et al

                3)      Dot immunobinding assay (DIBA) (Lee et al, 1992)

                4)      Enzyme-linked immunosorbent assay (ELISA)




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Causes diseases that result in significant losses in economically important crops
such as grape, citrus, peach and plum.

Diseases include:
       Pierce’s Disease of Grape – Limits grape production in Florida
       Phony Disease of Peach
       Leaf Scorch of Almond, Plum, Elm, Sycamore, etc.
       Wilt of Periwinkle
       Stunting of Ragweed
       Citrus Variegated Chlorosis (CVC)
       Sumatra Disease of Clove
       Crinkle Leaf Disease of Kenaf
         DNA techniques have been useful in grouping strains according to
         host of origin. Indicates host specialization.




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            Transmission
           Insect vectors (xylem-feeding insects) and includes members of
            leafhopper family known as sharpshooters and members of spittlebug
            family. Leafhopper vectors have wide host ranges. Transmission of
            bacterium requires no latent period (unusual for leafhopper transmitted
            procaryotic diseases which require latent period)

         Grafting
         Mechanical means in which inoculum is introduced into xylem
Phytotoxin production

      Major symptoms such as leaf marginal necrosis, leaf abscission, dieback,
delayed growth in spring and decline in vigor leading to death of plant. Toxin
implicated in the leaf scorch symptoms.

Host range - Wide (28 families of mono- and dicots. Host specificity of strains
exists.

Pierce’s disease Disease Cycle:
Pierce's disease is caused by Xylella fastidiosa, a gram-negative bacterium that survives and multiplies
within the water conductive system (xylem) of its plant hosts. X. fastidiosa has a diverse natural host
range with over 100 herbaceous and woody plant species. Many of these plant species are thought to be
symptomless hosts, yet may serve as reservoirs of inoculum for later insect transmission. Many of the
reservoir hosts in California
(www.cnr.berkeley.edu/xylella/temp/hosts.htm) are indigenous to NC and the Southeast. Fifty of the plant
host species reported in California occur in the Southeast, and different species in 43 additional genera
reported also occur here. Sharpshooter leafhoppers (Cicadellidae) and spittle-bugs
(Cercopidae) acquire and transmit the bacterium as they feed in the xylem of plants. In California, four
sharpshooter species have been shown to transmit the bacterium: blue-green sharpshooter, green
sharpshooter, red-headed sharpshooter, and the glassy-winged sharpshooter. The current epidemic in
California is likely related to the introduction of the glassy-winged sharpshooter, which has a wider host
range and can fly farther than other species of sharpshooters present. The major disease transmitting
insects of X. fastidiosa have not yet been identified in North Carolina, but are currently under
investigation.
http://www.cals.ncsu.edu/plantpath/ExtensionPro/grapes/pestdescriptions/dise




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Pierce’s Disease of Grape           Glassy Winged sharpshooter -
                                    vector




               Citrus Variegated Chlorosis




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