730 H. Y. Samaliev and M. Mohamedova
Bulgarian Journal of Agricultural Science, 17 (No 6) 2011, 730-735
Agricultural Academy
PLANT-PARASITIC NEMATODES ASSOCIATED WITH STRAWBERRY
(FRAGARIA AIIANASSA DUCH.) IN BULGARIA
H. Y. SAMALIEV* and M. MOHAMEDOVA
Department of Entomology, Agricultural University, BG - 4000 Plovdiv, Bulgaria
Abstract
SAMALIEV, H. Y. and M. MOHAMEDOVA, 2011. Plant-parasitic nematodes associated with
strawberry (Fragaria aiianassa Duch.) in Bulgaria. Bulg. J. Agric. Sci., 17: 730-735
A survey was conducted in five major strawberry growing regions in Bulgaria (Plovdiv, Blagoevgrad, Ruse,
Vratza and Sofia) to study the occurrence of plant-parasitic nematodes associated with strawberry (Fragaria ananassa
Duch.). A total 205 soil and 109 foliage samples containing mixed populations of 10 genera and 15 species
belonging to order Tylenchida and Dorylaimida were analyzed. The identified species were: Aphelenchoides
fragariae, Aphelenchoides ritzemabosi, Aphelenchoides besseyi, Criconema nutabile, Diylenchus dipsaci, Helicotylenchus
dihystera, Longidorus caespiticola, Longidorus elongates, Meloidogyne arenaria, Meloidogyne
hapla, Pratylenchus penetrans, Pratylenchus microdorus, Paratylenchus pseudoparietinus, Tylenchorhynchus
claytoni and Xiphinema pachtaicum. Many of the observed species are phytopathologically important parasites
of strawberry and some are also vectors of plant viruses. Frequency and density of each species were highly variable
from field to field and within the field.
Key words: Plant-parasitic nematodes, strawberry, distribution frequency, population density, Bulgaria
*E-mail: h.y.samaliev@abv.bg
Introduction
Strawberry (Fragaria ananassa Duch.) is an
important agri cultural commodity in Bulgaria. Its
cultivation covers 5 800 da with average yield of
907 кg/da (Ministry of Agriculture and Forestry,
Bulgaria, 2009). During recent years interest has
increased in strawberry cultivation as an alternative
cash crop including small family farms in our
country. Plant-parasitic nematodes are known to
be pests of fruit and vegetable crops (Samaliev
and Stoyanov, 2007). In the sixties and seventies,
nematode counts in soil and foliage samples
indicated that Aphelenchoides fragariae (Ritzema
Bos) Christie, and some other plant-parasitic nematodes
(Aphelenchoides ritzemabosi (Schwartz)
Steiner, Aphelenchoides subtensis (Coobb) Steiner
et Buhrer, Meloidogyne hapla Chitwood), were
spread in strawberry fields in producing regions
of Sofia (Stoyanov, 1961, 1975) and Blagoevgrad
(Gateva and Budurova, 1975) and at the beginning
of 2000 also in Plovdiv region (Samaliev and
Stoyanov, 2007). Information on distribution,
frequency of occurrence and population density
of plant-parasitic nematodes associated with
strawberry in Bulgaria is lacking.
Therefore, the objective of this study was to
conduct a survey to determine and document the
731 Plant-Parasitic Nematodes Associated with Strawberry (Fragaria Aiianassa Duch.) in Bulgaria
occurrence, distribution, density and prevalence
of plant-parasitic nematodes in strawberry fields
major strawberry producing regions in Bulgaria.
Material and Method
A survey was conducted in different locations in
the major strawberry growing regions of Plovdiv,
Blagoevgrad, Ruse, Vratza and Sofia. Sampling
was conducted during April-June 2008-2009. Soil,
root, and foliage (leaves, stem and flower-buds)
samples were collected randomly from the most
fields of each region (Figure 1). Each soil sample
was compo site of 10-15 soil cores from the same
field, collected randomly to a depth of 20-cm with
a hand shovel. Samples were placed in plastic
bags, sealed and brought back to the laboratory
and stored at 5°C until processed for nematode
extraction and were examined in a Laboratory of
Nematology at Agricultural University, Plovdiv.
Root-knot nematodes were extracted from roots
which were washed free of soil and examined
for galling and root-knot infection. Root-knot
nematodes were isolated from galled roots and
identified by the examination of perinea patterns
of adult females as well as the characters of the
second-stage juveniles (Eisenback et al., 1981).
Vermiform nematodes were recovered from
two 100 cm3 sub samples (after thorough mixing
of the soil from each sample) by Cobb’s sieving
and decanting technique followed by a modified
Baermann funnel method (Hooper, 1986).
An incubation method was used to extract the
nematodes from strawberry roots (Young, 1954).
Strawberry foliage samples were randomly
chosen from each region. Within 24 h of arrival
at laboratory, all the stems, buds and leaves in
each sample were cut into small pieces (0.5-1
cm) placed in bags and mixed. A two 15-g sub
sample from each sample was used for nematode
extraction using Baermann funnel for 3 days.
Isolated nematodes were killed at 65°C and fixed in
4% formalin and placed in viols. Prior to counting,
solution containing nematodes were agitated
thoroughly and 3 ml poured to a counting dish.
Nematodes were counted using a stereo binocular
microscope. Counting of root-knot nematodes
based on second stage juveniles (J2S) only. Selected
specimens for each of the recorded species were
processed for dehydration by Seinhorst’s (1959)
rapid glycerin method and mounted on plastic
slides in anhydrous glycerin. Identification of
species of plant parasitic nematodes was based on
the morphology and measurements of adults (Perry
et al., 1959; Esser, 1973; Handoo and Golden,
1989; Handoo, 2000; Siddiqi, 2000).
Simple statistics (frequency and density) were
performed on nematode counts using SPSS-12
program.
Results
Nematodes in 10 genera and 15 species belonging
to order Tylenchida (Filipjev) Thorne and
Dorylaimida Pearce, were found in association
with strawberry (Fragaria ananassa Duch.) from
total 205 soil and 109 and foliage samples (Table
1 and Table 2).
Fig. 1. Map of Bulgaria; showing sampling sites
in the major of strawberry production region,
Bulgaria
732 H. Y. Samaliev and M. Mohamedova
Table 1
Occurrence frequency and density of plant-parasitic nematodes in strawberry fields (in 100 cm3 soil) \
in major strawberry growing regions, Bulgaria
Nematode
species
Strawberry growing regions / number of samples collected
Plovdiv / 35 Blagoevgrad / 30 Vratzca / 40 Ruse / 70 Sofia / 30
RF* MD** RF MD RF MD RF MD RF MD
Aphelenchoides
fragariae - - - - 5 7 ±1.7 - - - -
Aphelenchoides
ritzemabosi - - - - 7.5 9 ±2.2 - - - -
Aphelenchoides
besseyi 2.8 5 ±1.7 - - - - - - - -
Criconema
nutabile - - - - 12.5 10 ±14.6 17.1 32 ±19.5 3.3 41 ±33.6
Ditylenchus
dipsaci 11.4 8 ±4.1 13.3 9 ±3.4 - 15.7 16 ±5.7 10 13 ±4.2
Helicotylenchus
dihystera - - 11.1 35±16.9 30 21 ±7.8 21.4 18 ±5.6 33.3 29
±10.5
Longidorus
caespiticola 5.7 10 ±3.6 - - 5 6±4.6 8.6 5±2.6 - -
Longidorus
elongatus 2.8 5 ±1.1 - - 7.5 9 ±3.5 - - - -
Meloidogyne
arenaria 31 61 ±22.1 26.6 49 ±17.4 22.5 19 ±13.1 20 14 ±10.4 20 16 ±13.1
Meloidogyne
hapla 26 55 ±35.4 36.6 70 ±47.8 42.5 111±48.3 41.4 125 ±56.0 30 89
±55.4
Pratylenchus
penetrans 37 28 ±16.1 33.3 19 ±10.4 26 15 ±13.1 45.7 28 ±15.7 20 21 ±10.2
Pratylenchus.
microdorus - - - - 12.5 17±9.6 15.7 25 ±13.5 - -
Paratylenchus
pseudoparietinus - - - - 10 76±24.6 - - 13.3 65 ±25.5
Tylenchorhynchus
claytoni - - 10 21±8.4 5 31 ±11.8 7.1 21 ±7.8 - -
Xiphinema
pachtaicum 8.6 10±3.8 6.6 16 ±3.8 - - 10 22 ±6.8 - -
*RF = Relative frequency of occurrence (percentage of samples in which species was found); **MD = Mean
density ± standard deviation of nema todes in 100 cm3 soil.
Plant-parasitic nematodes on soil samples
Root-knot northern Meloidogyne hapla Chitwood
and peanut M. arenaria (Neal) Chitwood
nematodes, associated with strawberry, were the
733 Plant-Parasitic Nematodes Associated with Strawberry (Fragaria Aiianassa Duch.) in Bulgaria
Table 2
Occurrence frequency and population density of Aphelenchoides fragariae, Aphelenchoides ritzemabosi,
Aphelenchoides besseyi and Ditylenchus dipsaci in strawberry plant tissues
Nematode species
Strawberry growing regions / number of samples collected
Plovdiv / 17 Blagoevgrad /15 Vratza / 20 Ruse / 42 Sofia / 15
Aphelenchoides fragariae
Occurrence frequency* 35.3 26.6 60 64.2 53.3
Nematodes densily** 34 ±7.5 42 ±14.5 70±20.1 98 ±43.8 61 ±10.6
Aphelenchoides ritzemabosi
Occurrence frequency* 41.1 53.3 25 28.6 26.6
Nematodes densily** 70 ±15.6 66 ±18.5 23±9.4 29 ±13.8 31 ±12.6
Aphelenchoides besseyi
Occurrence frequency* 11.7 - - 7.1 6.7
Nematodes densily** 8 ±1.6 - - 11 ±2.8 9.0 ±1.8
Ditylenchus dipsaci
Occurrence frequency* 11.7 13.3 10 14.3 20
Nematodes densily** 14 ±5.6 8 ±2.5 13±4.4 16 ±5.8 11 ±3.6
* Percentage of samples in which nematode was found; **Mean ± standard deviation of nematodes per 15 g
fresh tissue of strawberry plant.
most conspicuous plant-parasitic nematodes of
this group, collected from all surveyed regions.
However, their occurrence frequency and density
were highly variable, as the highest frequency
and density for M. hapla was recorded in Vratza
(42.5% and 111 nematodes/100 cm3 soil) and
Ruse (41.4% and 125 nematodes/100 cm3 soil),
and for M. arenaria was in Plovdiv (31% and
61 nematodes/100 cm3 soil) and Blagoevgrad
(26.6% and 49 nematodes/100 cm3 soil), Table 1.
The lowest frequency and density for M. hapla
was in Plovdiv (26% and 55 nematodes/100 cm3
soil) and Sofia (30% and 89 nematodes/100 cm3
soil), and for M. arenaria – Ruse (20% and 14
nematodes/100 cm3 soil) and Sofia (20% and
16 nematodes/100 cm3 soil), Table 1. Of the soil
samples infested with Meloidogyne spp., 48.0%
had M. hapla, 27% had M. arenaria and 25% had
both species.
The other proven parasite of strawberry
found in this study was root-lesion nematodes
Pratylenchus penetrans Cobb and P. microdorus
Andrassy. Pratylenchus penetrans was the predominat
species and was found of the samples at
the all regions (Plovdiv, Blagoevgrad, Vratza, Ruse
and Sofia) and were present in 37, 33.3, 26, 47.5
and 20% of sampled fields with density 28, 19, 15,
28 and 21 nematodes/100 cm3 soils, respectively.
The other species P. microdorus Andrassy was
collected only from Vratza and Ruse regions in
12.5 and 15.7% of sampled fields (density 17 and
25 nematodes/100 cm3 soil) , respectively (Table
1) in mix population with P. penetrans.
The pin nema tode Paratylenchus pseudoparietinus
(Micoletzky) Micoletzky, was plant-parasitic
nematode with the highest density in Vratza and
Sofia in tested strawberry fields. No information
is presently available on the pathogenicity of this
ectoparasitic genus on strawberry plants.
Six other plant-parasitic nematode species
on roots, Criconema nutabile (Taylor) Raski -
Luc, Helicotylenchus dihystera (Cobb) Sher,
734 H. Y. Samaliev and M. Mohamedova
Longidorus elongatus (de Man) Thorne et Swanger,
L. caespiticola Hooper, Tylenchorhynchus claytoni
Steiner and Xiphinema pachtaicum (Tulaganov)
Kirjanova, were also found occasionally and in
low density (Table 1).
Plant-parasitic nematodes on foliage
(leaves, stem and flower-buds)
Strawberry bud Aphelenchoides fragariae
(Ritzema Bos) Christie and chrysanthemum
Aphelenchoides ritzemabosi (Schwartz) Steiner
nematodes, were predominant species from aboveground
plant parts collected from all surveyed regions
(Table 2). Occurrence frequency and density
of both species were highly variable in different
investigated regions. For instance, the highest frequency
and density of Aph. fragariae was recorded
in Ruse (64.2% and 98 nematodes/15 g fresh tissue),
Vratza (60% and 70 nematodes/15 g fresh
tissue) and Sofia (53.3% and 61 nematodes/15 g
fresh tissue) and the lowest was in Blagoevgrad
(26.6% and 42 nematodes/15 g fresh tissue) and
Plovdiv (35.3% and 34 nematodes/15 g fresh tissue)
(Table 2). Vice versa for Aph. ritzemabosi,
the highest frequency and density was recorded
in Blagoevgrad and Plovdiv (53.3% and 41.1%
and 70 and 66 nematodes/15 g fresh tissue) and
the lowest – Ruse, Vratca and Sofia (Table 2). Of
the foliage samples infested with Aphelenchoides
spp., 22% had Aph. fragariae, 12% had Aph.
ritzemabosi, and 66% had both species.
Two other plant-parasitic nematode species on
foliage Aphelenchoides besseyi Christi and stem
nematode Ditylenchus dipsaci (Kfflin) Filipjev,
were also found occasionally.
Discussion
This survey provides information on occurrence
and density of plant-parasitic nematodes
associated with straw berry crops in the major
strawberry growing regions in Bulgaria. Some
nematodes identified in this survey are of economic
importance and are considered serious
pests of strawberry species and were found in
the samples of all strawberry growing regions
investigated - Plovdiv, Blagoevgrad, Ruse, Vratza
and Sofia. These nematodes were Aph. fragariae,
Aph. ritzemabosi, M. hapla, M. arenaria and P.
penetrans. Aphelenchoides fragariae and Aph.
ritzemabosi are also known as foliar nematodes
are parasites of above-ground plant part and may
be endo- or ectoparasitic, produce small curled or
crinkled leaves (crimp), deformed buds and flowers.
Aphelenchoides fragariae and Aph. ritzemabosi
were found in soils from Vratca only. They
might have disseminated through rain water, the
infected buds or leaves which may have fallen a
few days before sampling. In most of samples both
species are in mixed population. This widespread
distribution of Aphelenchoides spp. in all major
strawberry regions is consistent with results from
earlier surveys of strawberry crops in Sofia and
Blagoevgrad region (Stoyanov, 1975; Gateva and
Budurova, 1975). Meloidogyne hapla and M.
arenaria were found in the soil or as sedentary
endoparasites in roots and induce root galls. Earlier
surveys (Gateva and Baycheva, 1976; Stoyanov,
1980; Samaliev and Stoyanov, 2007) also showed
a widespread distribution of M. hapla and M.
arenaria on tobacco in Southwestern and Cen tral
Bulgaria and M. hapla localized in single fields on
strawberry in Blagoevgrad (Gateva and Budurova,
1975) and Plovdiv (Samaliev and Stoyanov, 2007)
regions. Knowl edge, of which species are present
in a field, will enable the grower to implement
the proper rotation sequence in addition to other
control practices, thereby suppressing root-knot
nematode populations to increase strawberry fruit
quality. Pratylenchus penetrans is a migratory
endoparasite of roots, and its infection makes fine
reddish-brown lesions on rootlets. It is also known
to enhance the severity of Gnomonia comari (fungus)
(Kurppa and Vrain, 1989) and Rhizoctonia
fragariae (LaMondia, 2003) on strawberry roots.
It is reported that P. penetrans and Aph. fragariae
have been most frequently associated with strawberry
damage in Blagoevgrad region (Gateva and
735 Plant-Parasitic Nematodes Associated with Strawberry (Fragaria Aiianassa Duch.) in Bulgaria
Budurova, 1975).
Presence of Aph. fragariae, Aph. ritzemabosi,
M. hapla, M. arenaria and P. penetrans nematodes
in straw berry fields indicates a risk of damage, and
recommended management strategies should be
taken by growers to prevent them from increasing
and spreading.
Strawberries, in this investigation, are also host
of D. dipsaci, Logidorus sp. and Xiphinema sp.
All of these nematodes are potential patho gens to
strawberry and their identification in strawberry
plantings or in land to be planted to strawberries
should be cause of concern (Samaliev and Stoyanov,
2007).
Other nematode species identified in this
survey are ectoparasites of roots and have not been
documented as dangerous pests of strawberry to
date. They include H. dihystera and T. claytoni.
The species of Criconemoides, Helicotylenchus,
Tylenchorhynchus and Longidorus have also been
commonly found in samples from virus vectors
they can be damaging at very low population
levels.
Conclusion
The results of this survey indicate that plantparasitic
nematodes on strawberry fields are widely
distributed in Bulgaria. Several of the recorded
species are known pathogens of strawberry and are
capable of causing damage to the crop. Knowl edge
of which species and race is present in a field is
important to determine the possible threat to strawberry
and its control. The damage thresholds are
also needed to be established in the soil types and
under the growing conditions of different regions
of Bulgaria. Particularly the population dynamics
of Aphelenchoides spp., Meloidogyne spp. and
Pratylenchus spp. on strawberry and their interactions
with other soil bourn disease organisms deserve
further studies. Our results provide essential
background information needed for planning and
practicing nematode management strategies in
Bulgarian strawberry fields.
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Received January, 5, 2011; accepted for printing September, 23, 2011.
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