Collecting Citrus and Grapevine Samples for Nematode Analysis

Note Number: AG1198
Date Published: October 1999
Date Reviewed: July 2011

Roots of most plant species are usually infected by one or more species of plant-parasitic nematodes. Most plant-parasitic nematodes are too small to be seen with the naked eye. Under favourable environmental conditions and suitable hosts, populations of nematodes can build up to cause economic crop loss. Symptoms caused by nematodes are not easily diagnosed in the field. These symptoms may be confused with other diseases, pests, nutritional or environmental factors. To diagnose nematode damage, it is usually necessary to test soil or plant material to get an accurate estimate of nematode numbers in the field. It is therefore very important to follow the correct sampling procedures when collecting samples for nematode analysis.

Citrus and Grapevine Nematodes

The most important plant-parasitic nematodes of citrus and grapevine are citrus nematodes (Tylenchulus semipenetrans) (Figures 1 and 2) and root-knot nematodes (Meloidogyne spp) (Figure 3). Other species of nematodes recorded on these crops include root-lesion (Pratylenchus spp) (Figure 4), stubby-root (Paratrichodorus spp) (Figure 5), dagger (Xiphinema spp), pin (Paratylenchus spp) (Figure 6), sheath (Hemicycliophora spp) (Figure 7), ring (Macroposthonia spp) (Figure 8) and stunt (Tylenchorhynchus spp) nematodes.

Figure 1. Tylenchulus semipenetrans  embedded in citrus root (Jackie Nobbs,  Plant parasitic nematodes of Australia)	Figure 2. Citrus infested with embedded in citrus root Jackie Nobbs, Tylenchulus semipenetrans
Figure 3. Grapevine infested with Meloidogyne sp.	Figure 4. Female and juvenile Pratylenchus thornei (Jackie Nobbs, Plant parasitic nematodes of Australia)
Figure 5. Anterior(right) and posterior sections of male(left) Paratrichodorus sp.	Figure 6. Female Paratylenchus sp. (Jackie Nobbs, Plant parasitic nematodes of Australia)

Sampling Time

Populations of citrus nematode do not undergo significant seasonal fluctuations. Therefore, samples for citrus nematode analysis can be collected throughout the year. But the ideal time to collect samples for root-knot nematode assessment is during September and October. The young root-knot nematode larvae hatch out from eggs in early spring, when they are present in their highest numbers.

Sampling Equipment

Figure 9. Soil sampling probe

• Clean bucket for collecting samples
• Soil probe (Figure 9) or shovel/spade
• Plastic bags to hold 500 g soil
• Labels
• Waterproof marker

Sampling Method

For Pre-plant soil test

Divide the field into 1 -2 hectare blocks. It may be necessary to divide the field based on soil type or previous crop history. Take 20 30 cores/sub-samples at a depth of 15 -20 cm. Collect these sub-samples with intervals of 10 -30 metres in a ‘Zigzag’ or ‘W’ pattern. Place these sub-samples in a clean bucket, mix thoroughly but gently with hands and take a 500 g final sample in a labelled plastic bag to be submitted for nematode analysis.

For Existing crop

Citrus

Soil samples should be collected from just inside the canopy of the tree. Collect six sub-samples to make a total of 500 g of soil, and 10 g of fine feeder roots. Place soil and root samples together in the same labelled plastic bag. Damage caused by a citrus nematode infestation depends on the age and vigour of the tree, density of the nematode population, and susceptibility of the rootstock. Mature trees can tolerate a considerable number of these nematodes before showing lack of vigour and decline symptoms. An examination of citrus roots can indicate the presence of citrus nematodes. Roots infected by significant numbers of the citrus nematode may appear coarse and dirty, as soil particles and debris adhere to egg masses, but this feature is not diagnostically reliable. The identity of the nematode still requires confirmation by microscopic examination.

Grapevine

Take samples within 60 cm around the root zone, as nematodes are most abundant in this area. Remove surface debris and dry soil from around the base of the vine. Use a soil probe or spade to a depth of at least 20 cm. Collect at least six sub-samples around the vine to make up a total of 500 g of soil, and 10 g or a handful of fine feeder roots. Careful examination of young vine roots can reveal root damage caused by root-knot or root-lesion nematodes. The presence of root-knot nematodes can be noticed on roots as they have small swellings or galls on young feeder roots or secondary rootlets. Larger galls may result from multiple infections. When the galls are broken apart, the tiny, white bodies of mature females can often be detected with the aid of a hand lens.

In the case of a single plant or an area of uniform growth, only one sample is required for nematode assessment. Different in soil type, plant variety and/or rootstock may justify the collection of separate samples. Select the majority of sub-samples from the edge of any poor patch or from plants showing symptoms of decline.

Storage and Transport

All samples should be kept in sealed plastic bags to prevent drying out. Do not refrigerate samples. Keep them away from hot sun. Do not leave samples in closed vehicles on hot days. Samples should be submitted for testing promptly after collecting.

Label and information

Label samples with identification numbers and provide the following information on a separate sheet of paper:

• Name and address of the grower as well as sender
• Plant variety, rootstock
• Planting age, plant vigour, total acreage, symptoms
• Fertilisers, pesticides and herbicides applied etc It is necessary to provide the above information so that the results of the analysis can be interpreted correctly and satisfactorily.

Results

Nematode populations in soil samples are classified as low and high:

Where to send the samples

For nematode analysis, send the samples to the Crop Health Services laboratory address given below: Crop Health Services, 621 Burwood Highway, Knoxfield 3180. Or Postal Address: Crop Health Services, Private Bag 15, Ferntree Gully Delivery Centre, Vic 3156.

Further References

Nicol, J. M., Stirling, G. R., Rose, B. J., May, P. and Van Heeswijck, R. (1999), Impact of nematodes on grapevine growth and productivity: current knowledge and future directions, with special reference to Australian Viticulture. Australian Journal of Grape and Wine Research 5, 109 – 127.

Whiteside, J.O., Garnsey, S. M and Timmer, L.W. (1989). Compendium of Citrus Diseases. The American Phytopathological Society.

Person, R. C. and Goheen, A. C. (1998) Compendium of Grape Diseases. The American Phytopathological Society.

Contact/Services available from DPI

For further information, phone Crop Health Services on (03) 9210 9222 or fax (03) 9800 3521.

Acknowledgements

Assistance received from Neville Fernando, Farm Services Victoria, in preparation of this is appreciated.

This Agnote was developed by Megan Edwards, DPI in September 1994. It was reviewed by Lila Nambier , Crop Health Services in July 2011.