Review of policy: importation of grapevine

3.1.1 Dormant cuttings

Mandatory on-arrival inspection

Mandatory on-arrival fumigation

Alternative treatments to methyl-bromide fumigation for grapevine dormant cuttings, if requested by an exporting country, will be considered by Plant Biosecurity on a case by case basis. Prior to the acceptance of an alternative treatment for grapevine dormant cuttings, Plant Biosecurity would need to assess the efficacy of that fumigant to ensure it gives an equal level of protection to methyl-bromide for all pests likely to be associated with the commodity.

Mandatory hot water treatment

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  Hot water treatment at 50 °C for 30 minutes is effective against some phytoplasmas (Caudwell et al. 1997) and in eliminating most known fungal pathogens and endophytes from grapevine cuttings, including pathogens associated with young grapevine decline (Crous et al. 2001).
  
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  After hot water treatment, dormant cuttings must be plunged into cold water to quickly lower the temperature and minimise heat damage to the tissue (Waite et al. 2005).
  

Mandatory sodium hypochlorite treatment

Mandatory culturing

Mandatory growth in PEQ facilities

It is recommended that during PEQ growth period, plants and plantlets are subjected to visual inspection, electron microscopy and active testing, including biological indexing and molecular testing.

Pathogen screening (visual screening) during growth in PEQ is recommended to continue for the detection of symptomatic pathogens. Fungal and bacterial pathogens associated with grapevines may produce distinct symptoms that make them easy to identify by visual inspection during growth period in PEQ.

The recommended pathogen testing during growth in PEQ will include active testing for quarantine pathogens, using traditional and modern techniques. Laboratory methods; including culturing, biological indicators, electron microscopy and molecular tests (PCR); may be used to detect grapevine pathogens.

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  Active pathogen testing including molecular tests for Xylella fastidiosa, in addition to hot water treatment and visual inspection is recommended.
  
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  Diagnostic tests, including culturing and microscopy, are recommended for Xanthomonas campestris pv. viticola and Xylophilus ampelinus. However, if symptoms develop during growth in PEQ, molecular testing (including PCR) for Xanthomonas campestris pv. viticola (Trindade et al. 2005) and Xylophilus ampelinus (Botha et al. 2001) is recommended.
  

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  Newly established plants (from imported propagative material) will be subject to growing season inspection and if symptoms develop during the PEQ period, further diagnostic testing; including culturing, microscopy and molecular tests; is recommended.
  

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  Newly established plants (from imported propagative material) will be subject to growing season inspection and active pathogen testing, including a generic PCR.
  

Table 3.1 Recommended screening procedures for bacteria, fungi and phytoplasma

Grapevine viruses are transmissible entities; they can be detected and identified on herbaceous and woody indicator plants. Herbaceous host indexing assays may be completed in a matter of weeks whereas woody indicator assays require a lengthier incubation period (up to two years) to complete (Rowhani et al. 2005). Herbaceous hosts are used to test for sap transmissible nepoviruses, whereas woody indicator plants are used to test for phloem limited viruses (Rowhani et al. 2005). Laboratory methods; including electron microscopy and molecular tests (PCR); can also be used to detect grapevine infecting viruses.

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  As woody indexing is time consuming, molecular tests are recommended to replace woody indexing, thereby leading to a reduction of the PEQ growth period from a minimum of 24 months to a minimum of 16 months.
  
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  Molecular tests (PCR, RT-PCR, and qPCR) target the genetic material of plant pathogens and specifically test for molecular sequences that are unique to a particular pathogen. Molecular tests can be used for the detection of grapevine pathogens because each pathogen has its own unique genetic code (Van Guilder et al. 2008). However, these molecular tests may not detect different strains or variants of a particular virus. Therefore, a combination of biological indexing and molecular tests is recommended to increase the likelihood of detecting viruses and their variants.
  

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  Electron microscopy for the identified viruses.
  
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  Herbaceous host indexing for nepoviruses (Chenopodium quinoa, Chenopodium amaranticolor, Cucumis sativus and other species may be used as herbaceous indicators).
  
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  Generic molecular tests for Ampelovirus, Ilarvirus, Maculavirus, Nepovirus and Vitivirus.
  
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  Specific RT-PCR for GVB (strains associated with corky bark).
  
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  Specific RT–PCR for GRBaV (Al Rwahnih et al. 2012a).
  

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  Detection of ampeloviruses will include, but will not be limited to, the following tests:
  

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  Mandatory generic PCR for GLRaV-6, 10, 11 using the dHSP-nest2 / LR5 clusdoL primers (Maliogka et al. 2008b); and
  
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  Mandatory specific one step RT-PCR for GLRaV-7 using the primer pair LR7-F/ LR7-R (Engel et al. 2008).
  

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  Detection of ilarviruses will include, but will not be limited to, the following tests:
  

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  Herbaceous host indexing, including Cucumis sativus or Nicotiana glutinosa (Grapevine line pattern virus); and
  
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  Mandatory genus specific nested PCR for ilarviruses (GAMV, GLPV) using the Ilar2F5/Ilar2R9 primer pair (Untiveros et al. 2010).
  

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  Detection of maculaviruses will include, but will not be limited to, the following test:
  

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  Mandatory genus specific nested PCR for maculaviruses (GAMaV, GRGV) using the primer pair RD1/RGAP (Sabanadzovic et al. 2000).
  

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  Herbaceous host indexing using a range of herbaceous indicators, that include but are not limited to:
  

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  Chenopodium quinoa (ArMV, BLMoV, CLRV, GARMV GBLV, GCMV, GDefV, GFLV, GTRSV, PRMV, RpRSV, SLRV, TBRV, ToRSV);
  

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  Chenopodium amaranticolor (ArMV, BLMoV, CLRV, GARMV, GBLV, GCMV, GDeF, GFLV, PRMV, RpRSV, SLRV, TBRV, ToRSV);
  
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  Cucumis sativus (AILV, SLRV, TBRV, ToRSV); and
  

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  Generic PCR testing for nepoviruses (Digiaro et al. 2007; Wei and Clover 2008). If nepoviruses are detected, then virus specific tests must be performed. Virus specific tests may include (but are not limited to):
  

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  ArMV and GFLV using the primer pair M2/M3 (Wetzel et al. 2002);
  
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  CLRV using the primer pair CLRV-5/CLRV-3 (Werner et al. 1997);
  
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  GARSV using the primer pair A34-1/ A34-2 (Gokalp et al. 2003);
  
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  GCMV and TBRV using the primer pair P1/P2 (Le Gall et al. 1995);
  
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  GDefV using the primer pair N66-1/ N66-2 (Cigsar et al. 2003);
  
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  PRMV using the primer pair PRMVV1/ PRMVC1 (Kheder et al. 2004);
  
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  RpRSV using the primer pair RpRSVF1/ RpRSVR1 (Ochoa-Corona et al. 2006);
  
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  SLRSV using the primer pair SLRSV-5D / SLRSV-3D (Faggioli et al. 2002); and
  
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  ToRSV using the primer pair D1/U1 (Griesbach 1995).
  

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  Detection of vitiviruses will include, but will not be limited to, the following test:
  

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  Mandatory specific RT-PCR for GVB (strains associated with corky bark) (Minafra and Hadidi 1994).
  

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  Detection of tombusviruses will include, but will not be limited to, the following tests:
  

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  Mandatory genus specific nested PCR for Tombusvirus (PetAMV) using the pairs TomCPR/TomCPR (Russo et al. 2002) or TBSVGralF1/ TBSVGralR1 (Harris et al. 2006).
  

A summary of recommended grapevine virus indexing procedures is provided in Table 3.2.

 Genus specific nested PCR for Tombusvirus (Russo et al. 2002 or Harris et al. 2006)

 Specific RT-PCR test (Yoshikawa et al. 1997; Abou Ghanem-Sabanadazovic et al. 2003; or Engel et al. 2008)

● Genus specific PCR for Ampelovirus (Maliogka et al. 2008b)

 Genus specific nested PCR for ilarviruses (Untiveros et al. 2010)

 Genus specific nested PCR for maculaviruses (Sabanadzovic et al. 2000)

Δ Strain specific PCR (Minfra and Hadidi 1994)

۩ Generic PCR test for Vitivirus (Dovas and Katis 2003)

Virus specific test for Grapevine red blotch-associated virus (Al Rwahnih et al. 2012a)

Plant Biosecurity acknowledges that advances in serological or molecular techniques is an on-going process and therefore the recommended PCR tests can be replaced when more up-to-date testing procedures are validated.