Aphelenchoides varicaudatus (Nematoda: Aphelenchoididae) and Helicotylenchus erythrinae (Nematoda: Hoplolaimidae) from Garlic Plantation in Magelang, Central Java, Indonesia
Many nematodes can be associated to garlic plants. Twenty genera of nematodes have been recorded to be associated with the roots of garlic plants in Yemen; Antarctenchus, Aphelenchoides, Aphelenchus, Basiria, Boleodorus, Ditylenchus, Helicotylenchus, Heterodera, Hoplolaimus, Meloidogyne, Microposthonia, Nothanguina, Pratylenchus, Rotylenchulus, Rotylenchus, Scutellonema, Tetylenchus, Tylenchorhynchus, Tylenchus, and Zygotylenchus found in garlic (Mohamed, 2015). While in Indonesia, from 2019 to 2021, several plant parasitic nematodes have been found accompanying garlic plants in Central Java and East Java, Indonesia, such as Helicotylenchus, Aphelenchoides, Rotylenchulus, Aphelenchus, Criconemoides, and Tylenchus (Wulandari & Indarti, 2019; Wulandari et al., 2021; and Kusuma et al., 2020). Several species of plant parasitic nematodes have wide host range, such as Ditylenchus dipsaci (Musyarofah & Indarti, 2019). Amritha and Budijastuti (2018) stated that Ditylenchus dipsaci, Aphelenchoides sp. and Xiphinema sp. were found in imported and local garlic in East Java. Therefore, identification of the most common plant parasitic nematode associated with garlic is needed for further management and avoid yield losses on garlic.
The most common nematode genera associated with garlic plantations in Magelang, Central Java, were Aphelenchoides and Helicotylenchus (Wulandari et al., 2021). The taxonomic determination of Aphelenchoides at the suborder level is quite controversial between “Tylenchid” and “Aphelenchid” (Sanchez-Monge et al., 2015). Many species are not fully described and do not have molecular data for taxonomic studies (Zhao, 2006; Sanchez-Monge et al., 2015). Some species of Helicotylenchus have very similar diagnostic characters, and boundaries between species are not well established (Subbotin et al., 2015). The existence of complex species in the genera makes them morphologically almost indistinguishable but may be quite distant from each other by their phylogeny (Subbotin et al., 2015). The combination of morphological and molecular identification shown to be most effective method for species identification (De Ley et al., 2005)
Nucleotides data for Aphelenchoides and Helicotylenchus genera are mostly limited in The National Center for Biotechnology Information (NCBI). At the time this research was conducted, there were only 1237 and 758 nucleotides records for Aphelenchoides and Helicotylenchus, consecutively (NCBI accessed in November 2021). However, molecular data at species level are obviously rare. Several molecular studies on Aphelenchoides were reported in Indonesia, such as A. besseyi on rice (Rahman et al., 2018; Sembiring et al., 2019) and A. varicaudatus on garlic plants (Kusuma et al., 2020), but there was no information about the molecular study of Genus Helicotylenchus in Indonesia. This study aimed to characterizing the most dominant nematodes from garlic plants in Magelang, Central Java, Indonesia, based on morphological characters and DNA sequences.
Materials and Methods
Nematodes collection
The nematodes were extracted from garlic roots, bulbs, and soil obtained from garlic field in Kaliangkrik, Magelang, Central Java, Indonesia. Two main fields used as collection sites: the first site was planted Lumbu Kuning Variety and second site was planted Tawangmangu Baru Variety. Samples were collected using the direct sampling method on symptomatic plants (purposive sampling method). The symptoms shown on garlic plants were yellowing and wilted leaves, rotting bulbs, and disturbed plant growth.
Nematode extraction was carried out by directly soaking the plant's materials in the water. Without being washed, the roots and bulbs were cut into smaller parts. Then, the roots and bulbs were transferred to the petri-dish filled with sterile water and let sit for about 6 hours at 18 – 27°C. The water was filtered to remove the plant materials and collected to a becker glass (Ajri et al., 2021). Nematode extraction from soil was carried out using the Oostenbrink dishes method (EPPO, 2013).
Nematode identification
Morphological identification
Morphological identification to genera was carried out by comparing the specimen to literature by Mai and Mullin (1996). The species identification observation compared with Uzma et al. (2015), Wouts and Yeats (1994) for Helicotylenchus identification and IPPC (2016) and Huang et al. (2012) for Aphelenchoides identification. Characters such as head region, stylet, tail, and the posisition of vulva were observed under a binocular microscope (Olympus CX31).
Molecular identification
DNA extraction
DNA extraction was perfomed using the CTAB method (Cseke et al., 2003) with some modification. A total of 20 – 30 nematodes were cut into pieces using a needle in 25 μL of CTAB buffer on a glass object under a binocular microscope then transferred into the extraction tube. The tubes were incubated at 65°C for 30 minutes, inverted every 10 minutes followed by centrifugation for 5 minutes at 5000 RPM. The DNA suspension was added with chloroform: isoamyl alcohol (CIAA 24:1), then shaken until homogeneous (1 – 3 minutes). Samples were centrifuged at 10000 RPM for 15 minutes at room temperature. The supernatant was taken slowly with a micropipette and transferred to a new tube. Cold absolute ethanol was added with the proportion 1:2 for supernatant: ethanol. Samples were incubated at −20°C for 24 hours. Then, the samples were centrifuged at 10000 RPM for 15 minutes at room temperature. The supernatant was removed and then the pellets were added with 70 % cold ethanol. The solution was homogenized and then centrifuged for 15 minutes at 10.000 RPM. The ethanol solution was discarded and left to dry. The DNA pellets were added with 20 μL of TE buffer. The DNA suspension was stored at −20°C for molecular identification.
Polymerase chain reaction
Molecular characterization was conducted by polymerase chain reaction (PCR) and sequencing nucleotides using universal primer targeting the 28S rRNA gene (D2A/D3B). PCR was carried out by Bio-Rad T100 Thermal Cycler in a total volume of 25 μL (12.5 μL MyTaq HS RedMix BioLine, 2.5 μL of each primer (10 pmol/μL), 5 μL DNA template, 2.5 μL nuclease-free water). The D2–D3 expansion region of LSU was amplified with forward primer D2A (5′-ACA AGT ACC GTG AGG GAA AGT TG-3′) and the reverse primer D3B (5′-TCG GAA GGAACC AGC TAC TA-3′) (Nunn, 1992). Initial denaturation was carried out at 94°C for 2 minutes, followed by 35 cycles with the following steps: denaturation at 94°C for 15 seconds, annealing at 50°C for 30 seconds, extension at 68°C for 60 seconds. The final synthesis was carried out at 68°C for 5 minutes with a final temperature of 4°C. PCR products were analyzed on 2 % agarose gel electrophoresis and visualized on a UV transilluminator.
Phylogenetic tree
Amplified D2–D3 expansion region were sequenced to obtain DNA base sequence data. Sequence data were edited by BioEdit 7.2 and submitted into the BLAST-N program through the NCBI (National Center for Biotechnology Information) website (https://www.ncbi.nlm.nih.gov). Several sequences within species and other groups were selected as a comparison and aligned using BioEdit 7.2. The phylogenetic tree was constructed by selecting the best fit model using Mega X program.
Ethical Approval and/or Informed Consent
For this study formal consent is not required.
Results
Nematode population
Garlic plants associated with some nematodes were looked smaller and wilted than other healthy plants. Their leaves turned yellow (Fig. 1.A). The root area looked unhealthy with rotting bulbs when the plants were uprooted. Bulbs rot usually comes with fungi (white rot) that causes the soil cover up the bulb (Fig. 1.B).
Fig. 1.
Plant damaged symptoms. (A) plant symptoms on the ground (B) rot bulb associated with fungi (C) root rot damaged.
In this study, Aphelenchoides and Helicotylenchus were the most found population of nematodes genera on garlic plantations, potentially suppressing the plant's yield. The nematode population is shown in Table 1.
The nematode population found extracted from root, soil and bulb of garlic plants.
Nematodes Genera
Site 1 (Lumbu Kuning Variety)
Site 2 (Tawangmangu Baru Variety)
Roots (0.5 g)
Bulbs (3 g)
Soil (100 g)
Roots (0.5 g)
Bulbs (3 g)
Soil (100 g)
Aphelenchoides
50.8
38
2.5
8
0.8
0
Helicotylenchus
0.6
0
7.5
0.2
0
19.7
Nematode identification
Morphological character
Based on the identification to the nematode genera (Mai & Mullin, 1996), two most found genera were identified as genus Aphelenchoides and Helicotylenchus (Fig. 2 and Fig. 3). Morphological and morphometric data lead to Aphelenchoides varicaudatus and Helicotylenchus erythrinae (Fig. 2, 3 and Tables 2, 3).
Fig. 2.
A. varicaudatus (A–C) anterior region (D–F) tail shape variation (G) entire body with large metacorpus and vulva at ~60% of body length (H) lips, stylet, and large metacorpus (I) dorsally convex-conoid tail with little mucro (Scale bars: A–F, G–I: 50 μm; G: 100 μm)
Fig. 3.
H. erythrinae (A–C) spiral shape variation (C) entire body, vulva at ±60% of body length (D–F) anterior region, lips and stylet with knob flat or slightly anterior projection (G–I) conoid tail with variation of mucro (Scale bars: 100 μm)
Measurements of female A. varicaudatus. All measurements are in μm except for V in % and in the form mean±s.d (range).
The first genus identified as Aphelenchoides (Fig. 2, Table 2.) from its large metacorpus which clearly shown at low magnification and conoid tail. Vulva at ±60 % body length (Fig. 2.G), the lip region rounded and set off, metacorpus clearly shown round to square-shaped, stylet with small basal swelling (Fig. 2.H), dorsally convex-conoid tail with terminating step like mucro at end (Fig. 2.D–F,I). The second genus population identified as H. erythrinae (Fig. 3, Table 3.). The general characteristic of the Genus Helicotylenchus is a ventrally curved or becoming spiral shape at rest. Helicotylenchus population from Magelang, Central Java had various spiral-shaped when it relaxed (Fig. 3.A–C). The spiral form did not reach 1.5 spiral and sometimes in C shapes. Vulva at ±60 % of the body (Fig. 3.C), lip region rounded/hemispherical (not truncate), stylet clearly observed with basal knob flat to intended anteriorly (Fig. 3.D–F), tail asymmetrical and dorsally curved with a slight terminal projection to quite pronounced but not longer than the width of the nematode anal portion (Fig. 3.G–I)
Molecular Characterization
PCR electrophoresis result showed that the Helicotylenchus and Aphelenchoides population from Magelang, Central Java was successfully amplified at ~780 bp (Fig. 4). The expansion region of D2–D3 28S rDNA was sequenced for molecular analysis. The BLAST-N results for the Aphelenchoides sample showed high identity to Aphelenchoides varicaudatus from Yunnan, China (HQ283353) with 99.47 % identity, while Helicotylenchus sample showed 95.22 % identity to H. erythrinae from Colombia (MT321739). Sequence results for the Aphelenchoides sample from Magelang Indonesia were submitted to GenBank with accession number OK055306, while OL757653 for Helicotylenchus sample from Magelang Indonesia. Multiples sequence alignment of Aphelenchoides species showed that A. varicaudatus Magelang (OK055306) and A. varicaudatus Yunnan (HQ283353) had 0.58 % differences. The differences were showed at length of 130, 212, 530, 628. While, differences with A. varicaudatus Tegal (MN587128) were only 0.29 % at length of 212 and 530. Moreover, the closest species, A. xui (FJ643488) had more than 100 different nucleotides from our sample. Multiple sequence alignments for these species are presented in Figure 6.
Fig. 4.
Visualization PCR electrophoresis (M) Marker (DNA ladder 100 bp; (1) H. erythrinae; (2) A. varicaudatus; (3) negative control; (4) positive control (Meloidogyne graminicola)
The phylogenetic analysis of D2–D3 28S rRNA gene showed that our sample (OK055306) Aphelenchoides from Magelang, Indonesia clustered well with the species A. varicaudatus from Yunnan, China (HQ283353) and A. varicaudatus from Tegal, Indonesia (MN587128). A. varicaudatus Magelang, Central Java, Indonesia separated from other Aphelenchoides species. Phylogenetic relationship for species representatives within Aphelenchoidea Superfamily from partial 28S rRNA gene sequences are presented in Figure 5.
Fig. 5.
Maximum-likelihood phylogenetic tree with Hasegawa-Kishino-Yano model of Aphelenchoides spp. and other represented species based on 28S rDNA gene sequences. Substitution models with the lowest Bayesian Information Criterion (BIC) scores. Bootstrap values are shown at the nodes.
Fig. 6.
Multiple sequences alignment of Aphelenchoides varicaudatus species and Aphelenchoides xui from D2–D3 expansion region.
Multiple sequences alignment for species representatives within Genus Helicotylenchus are presented in Figure 8. Nucleotides at 37, 39, 47,53, 56, 57, 58, 59, 60, 61, 63, 64, 65, 66, 78, 79, 80, 83, 85, 86, 87, 88, 106,118, 176,177, 182,183, 231, 232, 235, 236, 237, 257, 259, 278, 281, 323, 325, 328, and 495 showed specific differences from other species, which separated them from other species and clustered well with H. erythrinae (MT321739). However, Helicotylenchus from Magelang Indonesia (OL757653) had 28 different nucleotides from 569 nucleotides length (4.9 %) with H. erythrinae (MT321739). Different nucleotides were observed more for other species compared.
Phylogenetic tree for Helicotylenchus showed Helicotylenchus from Magelang, Indonesia (OL757653) separated well from other species within genus except for H. erythrinae (MT321739). Phylogenetic relationships for species representatives within Helicotylenchus genus from the partial 28S rRNA gene sequences are presented in Figure 7.
Fig. 7.
Maximum-likelihood phylogenetic tree general time-reversible model and gamma-shaped distribution (GTR+G) of Helicotylenchus spp. and other represented species based on 28S rRNA gene sequences. Substitution models with the lowest Bayesian Information Criterion (BIC) scores. Bootstrap values are shown at the nodes.
Fig. 8.
Multiple sequences alignment of Helicotylenchus species within genus from D2–D3 expansion region.
Discussion
Common symptoms caused by nematodes include yellowing, stunted growth, and accompanied by a decrease in yield. Plant damage due to nematodes alone is difficult to observe because the aboveground symptoms are similar to symptoms of nutrient deficiency and they also can be associated with other pathogens. It is necessary to carry out laboratory tests to see the presence of important plant-parasitic nematodes that can potentially reduce yields (Schmitt & Sipes, 2000; William et al., 2018). Aphelenchoides found in each extracted part (roots, bulbs, soil) in each field surveyed. The highest population for Aphelenchoides was found in the first field, which showed severe bulb rot (causing soil cover up the bulbs) with the presence of white fungi. The presence and interaction between nematodes and fungi in plants could be antagonistic interactions or synergistic interactions. Antagonistic interactions can occur through competition for space and food. The synergistic interactions can result in more severe damage to the host plant (Zhang et al., 2020). In other hand, Helicotylenchus mostly found from the soil for both field.
The Aphelenchoides samples had characters as described by Huang et al. (2012) as A. varicaudatus. This species with a rounded set-off head, multireflexed or outstretched ovary, posterior ovary with V%=±60, conoid convex tail with variable tail terminus shape, one or two mucronate structures on the tail terminus (Huang et al., 2012). Aphelenchoides sample from Magelang have the same body length range as A. varicaudatus Yunnan, China.
The molecular analysis supported Aphelenchoides Magelang Indonesia as A. varicaudatus species. All sequences comparison in this study were picked from large subunit (LSU) 28S rRNA gene region. This region has conserved domains and expansion segments, such as the D2–D3 expansion region. This region is useful for designing of diagnostics tool for the identification of nematodes to the species level (Cunha et al., 2018). In this study, A. varicaudatus Magelang Indonesia separated from unidentified Aphelenchoides species (Aphelenchoides sp.) from various country based on its 28S gene region. A. varicaudatus from Yunnan, China is closely related to Aphelenchoides sp. from ginger plant (Zingiber sp.) in Indonesia from its small subunit (SSU) and very different to A. besseyi and A. fragariae (Huang et al., 2012). The phylogenetic tree in this study also revealed that A. varicaudatus separated from other species of Aphelenchoides such as A. besseyi, A. fragariae, A. ritzemabosi based on 28S region as Huang et al. (2012) reported.
Morphology and molecular identification of Aphelenchoides from garlic plants in Magelang showed the species indicated to A. varicaudatus. A. varicaudatus is not a common plant-parasitic nematode reported species from Genus Aphelenchoides like A. besseyi, A. fragariae, and A. ritzemabozi. The status of A. varicaudatus was still unclear. However, study about this species is also important, remembering that this species was also found in the rose root and pine xylem (Huang et al., 2012; Ibrahim & Hooper, 1994). A study by Santos et al. (2021) revealed that Aphelenchoides pseudogoodeyi, which also has unclear information about its pathogenicity, is an opportunistic pathogen as the injury was required to induce symptoms in Bird's-Nest Fern and Oriental Lily. It was showed that a mainly mycophagus Aphelenchoides could become phytophagous under stress conditions (Santos et al., 2021). A. varicaudatus was first described by Ibrahim and Hooper (1994) from rose roots plants. Furthermore, A. varicaudatus was found in Yunnan, China from Simao Pine plant (P. kesiya var. langbianensis) (Huang et al., 2012). A. varicaudatus could be found in bark/moss/lichens and able to grow on fungi media (Sanchez-Monge et al., 2015). This indicated that A. varicaudatus has a wide distribution and different host associations (Huang et al., 2012). The species in Aphelenchoides are very diverse and have different types of feeding. Most species in Aphelenchoides are mycophagous (Kanzaki, 2012). There are 13 other species known as plant parasites. Most of them are generalists because they have a broad host range (Sanchez-Monge et al., 2015). Only a few species are economically important (Nickel & Hopper, 1991).
The morphological characters of Helicotylenchus are quite complicated to observe between species. Our sample had descriptions as H. erythrinae in Uzma et al. (2015) based on lips and tail observations and also description by Riascos-Ortiz et al. (2020). Based on morphometrical data, our specimens close to H. erythrinae from Calarcá, Quindío, Colombia by Riascos-Ortiz et al. (2020) with high variability observed in some characters. The existence of intraspecific variation and limited character information makes identification difficult. However, no male nematodes were found in this study. According to Riascos-Ortiz et al. (2020), female H. erythrinae species have functional spermatheca with sperm in it and there are male nematodes. Species H. erythrinae has a loose spiral post-mortem habitus, a hemispherical lip region with an indented or flattened anterior knob, a tail with long ventral projections (Riascos-Ortiz et al., 2020) which these morphological characteristics were found in our specimens.
The phylogenetic tree supported that Helicotylenchus from Magelang (OL757653) clustered well with H. erythrinae (MT321739) from Colombia. Percent identification with this species was only 95.66 %, indicating the Magelang specimen had some nucleotide variations with H. erythrinae (MT321739). However, there was only one sequence submitted for H. erythrinae out of a total of 758 submitted sequences in the Genus Helicotylenchus (data accessed in November, 2021). Nucleotide sequence observations showed specific characters with H. erythrinae (MT321739) at some orders of nucleotides length, such as at 56 – 66, and 78 – 89 of 569 nucleotides compared with other representative species in the Genus Helicotylenchus.
Based on morphological observations and molecular characters, Helicotylenchus found from garlic plantations in Magelang, Central Java, refers to the H. erythrinae species. Molecularly, H. erythrinae was first reported by Riascos-Ortiz et al. (2020) from banana plantain in Colombia. H. erythrinae has a fairly wide host range, including dadap (Sher, 1966), pepper, ginger (Koshi et al., 2005), tea (Nalini et al., 2005), cocoa, olive (El-Borai & Duncan, 2005), cassava, rice (Bridge et al., 2005). In addition to being found in some cultivated plants, H. erythrinae was also found in uncultivated jungle soils (Sauer & Winoto, 1975). H. erythrinae, H. dihystera and three undescribed Helicotylenchus species were found from Java (Sher, 1966). Systematics within the genus of Helicotylenchus is in a very unsatisfactory state and identification to species is difficult (Wouts & Yeates, 1994).
Future study for A. varicaudatus and H. erythrinae found from garlic plantation is needed since the sequences data in NCBI for both species were limited. Molecular characterization from other DNA genes target is needed to perform more comprehensive molecular characterization.
Plant damaged symptoms. (A) plant symptoms on the ground (B) rot bulb associated with fungi (C) root rot damaged.
Fig. 2.
A. varicaudatus (A–C) anterior region (D–F) tail shape variation (G) entire body with large metacorpus and vulva at ~60% of body length (H) lips, stylet, and large metacorpus (I) dorsally convex-conoid tail with little mucro (Scale bars: A–F, G–I: 50 μm; G: 100 μm)
Fig. 3.
H. erythrinae (A–C) spiral shape variation (C) entire body, vulva at ±60% of body length (D–F) anterior region, lips and stylet with knob flat or slightly anterior projection (G–I) conoid tail with variation of mucro (Scale bars: 100 μm)
Fig. 4.
Visualization PCR electrophoresis (M) Marker (DNA ladder 100 bp; (1) H. erythrinae; (2) A. varicaudatus; (3) negative control; (4) positive control (Meloidogyne graminicola)
Fig. 5.
Maximum-likelihood phylogenetic tree with Hasegawa-Kishino-Yano model of Aphelenchoides spp. and other represented species based on 28S rDNA gene sequences. Substitution models with the lowest Bayesian Information Criterion (BIC) scores. Bootstrap values are shown at the nodes.
Fig. 6.
Multiple sequences alignment of Aphelenchoides varicaudatus species and Aphelenchoides xui from D2–D3 expansion region.
Fig. 7.
Maximum-likelihood phylogenetic tree general time-reversible model and gamma-shaped distribution (GTR+G) of Helicotylenchus spp. and other represented species based on 28S rRNA gene sequences. Substitution models with the lowest Bayesian Information Criterion (BIC) scores. Bootstrap values are shown at the nodes.
Fig. 8.
Multiple sequences alignment of Helicotylenchus species within genus from D2–D3 expansion region.
Measurements of female A. varicaudatus. All measurements are in μm except for V in % and in the form mean±s.d (range).
Location Publication
Magelang. Indonesia (This Study)
Tegal. Indonesia (Kusuma et al.. 2020)
Yunnan. China (Huang et al.. 2012)
Number of Specimen
5
7
20
Body Length
645.20 ± 54.44 (565.39 – 710.57)
567.35 ± 74.74 (494.51 – 704.59)
780 ± 56.70 (634 – 900)
Stylet Length
17.14 ± 0.53 (16.67 – 18.05)
12.39 ± 1.01 (11.17 – 14.01)
14.20 ± 0.50 (13.50 – 15.00)
Tail Length
55.66 ± 3.13 (52.02 – 59.85)
36.63 ± 2.96 (32.61 – 42.34)
49.00 ± 2.90 (43.00 – 53.00)
V (%)
61.74 ± 1.25 (60.30 – 63.18)
69.72 ± 1.15 (68.32 – 71.2)
69.20 ± 0.75 (68.20 – 71.20)
a
22.30 ± 2.08 (20.02 – 25.03)
28.71 ± 3.98 (24.29 – 35.8)
30.40 ± 1.68 (26.50 – 33.20)
c
11.59 ± 0.68 (10.87 – 12.65)
15.47 ± 1.26 (13.81 – 16.90)
16.11 ± 0.58 (15.00 – 17.10)
Measurements of female H. erythrinae. All measurements are in μm except for V in % and in the form mean±s.d (range).
Location
Magelang. Indonesia
Palmerston. New Zealand
Haast. New Zealand
Calarcá. Quindío. Colombia
La Celia. Risaralda. Colombia
Publication
(This Study)
(Wouts & Yeates. 1994)
(Riascoz-Ortiz et al.. 2020)
(Uzma. et al.. 2015)
Number of Specimen
7
10
10
15
25
Body Length
605.35 ± 68.33 (500.30 – 690.54)
873 ± 63.90 (761 – 938)
690 ± 49.80 (631 – 758)
530 ± 100 (400 – 700)
600 ± 100 (460 – 680)
480 – 610
Stylet Length
28.19 ± 2.57 (22.64 – 29.87)
28.30 ± 1.23 (26.00 – 30.00)
27.90 ± 0.97 (25.50 – 29.00)
22.70 ± 1.00 (21.00 – 25.00)
22.80 ± 0.83 (21.00 – 24.00)
(23 – 26)
Tail Length
23.69 ± 6.22 (19.39 – 30.57)
20.60 ± 2.80 (16.00 – 24.00)
14.70 ± 2.71 (9 – 18)
20.53 ± 2.20 (17.00 – 25.00)
20.65 ± 2.39 (17.00 – 26.00)
V (%)
66.55 ± 4.08 (62.95 – 72.87)
62 ± 2.85 (58 – 68)
63 ± 2.22 (59 – 65)
63.80 ± 1.80 (60.10 – 67.60)
63.70 ± 2.16 (60.82 – 68.54)
(60 – 65)
a
21.79 ± 1.64 (18.86 – 23.64)
29.70 ± 1.64 (27.00 – 32.00)
28.20 ± 1.87 (24.00 – 30.00)
25.03 ± 2.00 (22.70 – 29.70)
25.90 ± 2.20 (22.86 – 30.91)
23 – 26
c
23.73 ± 2.92 (18.79 – 27.13)
43 ± 5.99 (31 – 51)
48 ± 10.55 (35 – 74)
25.93 ± 4.10 (19.20 – 31.50)
28.09 ± 3.00 (21.82 – 32.94)
27 – 34
The nematode population found extracted from root, soil and bulb of garlic plants.
Ajri, M., Indarti, S., Soffan, A., Huu, N.N. (2021): Morphological and phylogenetic characteristics of Ditylenchus dipsaci among garlic plants. Jordan J. Biol. Sci, 14(4): 769 – 773. DOI: 10.54319/jjbs/140418AjriM.IndartiS.SoffanA.HuuN.N.2021Morphological and phylogenetic characteristics of Ditylenchus dipsaci among garlic plantsJordan J. Biol. Sci14476977310.54319/jjbs/140418Abierto DOISearch in Google Scholar
Amritha, M.L., Budijastuti, W. (2018): Tingkat serangan nematoda parasit pada bawang putih (Allium sativum) impor dan lokal di Jawa Timur [The Infection Rate Parasitic Nematode On Import And Local Garlic In East Java]. LenteraBio, 7(3): 214 – 220 (In Indonesian)AmrithaM.L.BudijastutiW.2018Tingkat serangan nematoda parasit pada bawang putih (Allium sativum) impor dan lokal di Jawa Timur [The Infection Rate Parasitic Nematode On Import And Local Garlic In East Java]LenteraBio73214220(In Indonesian)Search in Google Scholar
Bridge, J., Plowright, R.A., Peng, D. (2005): Nematode Parasites of Cereals. In: Luc, M., Sikora, R.A., Bridge, J. Plant Parasitic Nematodes in Subtropical and Tropical Agriculture 2nd Edition. Wallingford, CAB International.BridgeJ.PlowrightR.A.PengD.2005Nematode Parasites of CerealsIn:LucM.SikoraR.A.BridgeJ.Plant Parasitic Nematodes in Subtropical and Tropical Agriculture2nd EditionWallingfordCAB InternationalSearch in Google Scholar
Cseke, L.J., Kaufman, P.B., Podila, G.K., Tsai, C.J. (2003): Handbook of Molecular and Cellular Methods in Biology and Medicine. Second Edition. CRC Press, United States of America.CsekeL.J.KaufmanP.B.PodilaG.K.TsaiC.J.2003Handbook of Molecular and Cellular Methods in Biology and MedicineSecond EditionCRC PressUnited States of AmericaSearch in Google Scholar
Da Cunha, T.G., Visotto, L.E., Lopes, E.A., Oliviera, C.M.G., God, P.I.V.G. (2018): Diagnostic methods for identification of root-knot nematodes species from Brazil. Crop Protection, Cienc. Rural, 48(02). DOI: 10.1590/0103-8478cr20170449Da CunhaT.G.VisottoL.E.LopesE.A.OlivieraC.M.G.GodP.I.V.G.2018Diagnostic methods for identification of root-knot nematodes species from BrazilCrop Protection, Cienc. Rural480210.1590/0103-8478cr20170449Abierto DOISearch in Google Scholar
De Ley, P., De Ley, I.T, Morris, K., Abebe, E., Mundo-Ocampo, M., Yoder, M., Heras, J., Waumann, D., Rocha-Olivares, A., Jay Burr, A.H., Baldwin, J.G., Thomas, W.K. (2005): An integrated approach to fast and informative morphological vouchering of nematodes for applications in molecular barcoding. Philos Trans R Soc B, 360: 1945 – 1958. DOI: 10.1098/rstb.2005.1726De LeyP.De LeyI.TMorrisK.AbebeE.Mundo-OcampoM.YoderM.HerasJ.WaumannD.Rocha-OlivaresA.Jay BurrA.H.BaldwinJ.G.ThomasW.K.2005An integrated approach to fast and informative morphological vouchering of nematodes for applications in molecular barcodingPhilos Trans R Soc B3601945195810.1098/rstb.2005.1726Abierto DOISearch in Google Scholar
El-Borai, F.E., Duncan, L.W. (2005): Nematode Parasites of Subtropical and Tropical Fruit Tree Crops. In: Luc, M., Sikora, R.A. Bridge, J. Plant Parasitic Nematodes in Subtropical and Tropical Agriculture 2nd Edition. Wallingford, CAB International.El-BoraiF.E.DuncanL.W.2005Nematode Parasites of Subtropical and Tropical Fruit Tree CropsIn:LucM.SikoraR.A.BridgeJ.Plant Parasitic Nematodes in Subtropical and Tropical Agriculture2nd EditionWallingfordCAB InternationalSearch in Google Scholar
Huang, Ren-E., Ye, W., Liang, J., Lu, Q., Zhang, X. (2012): Tylaphelenchus jiaae n. sp. and Aphelenchoides varicaudatus (Nematoda: Aphelenchoidinae) from Simao pine in Yunnan Province, China. Nematology, 14(1): 93 – 108. DOI: 10.1163/138855411X579418HuangRen-E.YeW.LiangJ.LuQ.ZhangX.2012Tylaphelenchus jiaae n. sp. and Aphelenchoides varicaudatus (Nematoda: Aphelenchoidinae) from Simao pine in Yunnan Province, ChinaNematology1419310810.1163/138855411X579418Abierto DOISearch in Google Scholar
Ibrahim, S.K., Hooper, D.J. (1994): Aphelenchoides varicaudatus sp. n. (Nematoda: Aphelenchoididae). AfroAsian J Nematol, 4: 210 – 214IbrahimS.K.HooperD.J.1994Aphelenchoides varicaudatus sp. n. (Nematoda: Aphelenchoididae)AfroAsian J Nematol4210214Search in Google Scholar
IPPC (2016): DP 17: Aphelenchoides besseyi, A. fragariae and A. ritzemabosi. Retrieved from https://www.ippc.int/static/media/files/publication/en/2016/11/DP_17_2016_En_2016-11-01_iaK6Hls.pdfIPPC2016DP 17: Aphelenchoides besseyi, A. fragariae and A. ritzemabosiRetrieved from https://www.ippc.int/static/media/files/publication/en/2016/11/DP_17_2016_En_2016-11-01_iaK6Hls.pdfSearch in Google Scholar
Kanzaki, N., Giblin-Davis, R. (2012): Chapter 7: Aphelenchoidea. In: Manzanilla-Lopez, R. Mendoza, N. Practical Plant Nematology. Biblioteca Básica de Agricultura, Guadalajara, México.KanzakiN.Giblin-DavisR.2012Chapter 7: AphelenchoideaIn:Manzanilla-LopezR.MendozaN.Practical Plant NematologyBiblioteca Básica de AgriculturaGuadalajara, MéxicoSearch in Google Scholar
Kusuma, M.D., Supramana, Giyanto (2020): Phytonematodes community and polyphasic character of Aphelenchoides varicaudatus on garlic plants in Tegal Regency, Central Java. Jurnal Perlindungan Tanaman Indonesia, 24(2):216 – 223. DOI: 10.22146/jpti.49779KusumaM.D.SupramanaGiyanto2020Phytonematodes community and polyphasic character of Aphelenchoides varicaudatus on garlic plants in Tegal Regency, Central JavaJurnal Perlindungan Tanaman Indonesia24221622310.22146/jpti.49779Abierto DOISearch in Google Scholar
Mai, W.F., Mullin, P.G. (1996): Plant-Parasitic Nematodes: A Pictorial Key to Genera. Comstock Pub. Associates, Cornell University Press. ISBN: 08-014-31166MaiW.F.MullinP.G.1996Plant-Parasitic Nematodes: A Pictorial Key to GeneraComstock Pub. Associates, Cornell University PressISBN: 08-014-31166Search in Google Scholar
Mohamed, R.M. Saeed (2015): Plant–parasitic nematodes associated in garlic in Yemen. Egypt. J. Agronematol, 14(1): 37 – 44. DOI: 10.21608/ejaj.2015.57489MohamedR.M. Saeed2015Plant–parasitic nematodes associated in garlic in YemenEgypt. J. Agronematol141374410.21608/ejaj.2015.57489Abierto DOISearch in Google Scholar
Musyarofah, A., Indarti, S. (2020): Host Range of Stem and Bulb Rot Parasite Nematode (Ditylenchus dipsaci). In: Tutik D.R., Roto, R., Rohana, A., Laurent, C., Kuwat, T., Indriana, K., Julius, M., Dwi, S. (Eds) Key Engineering Materials. Trans Tech Publications Ltd, Switzerland. Vol. 840: 137 – 141. DOI: 10.4028/www.scientific.net/KEM.840.137MusyarofahA.IndartiS.2020Host Range of Stem and Bulb Rot Parasite Nematode (Ditylenchus dipsaci)In:TutikD.R.RotoR.RohanaA.LaurentC.KuwatT.IndrianaK.JuliusM.DwiS.(Eds)Key Engineering MaterialsTrans Tech Publications LtdSwitzerland84013714110.4028/www.scientific.net/KEM.840.137Abierto DOISearch in Google Scholar
Nalini, C.G., Mohotti, K.M. (2005): Nematode Parasites of Tea. In: Luc, M., Sikora, R.A. Bridge, J. (Eds) Plant Parasitic Nematodes in Subtropical and Tropical Agriculture 2nd Edition. Wallingford, CAB International.NaliniC.G.MohottiK.M.2005Nematode Parasites of TeaIn:LucM.SikoraR.A.BridgeJ.(Eds)Plant Parasitic Nematodes in Subtropical and Tropical Agriculture2nd EditionWallingfordCAB InternationalSearch in Google Scholar
Nickle, W.R., Hooper, D.J. (1991): The Aphelenchina: Bud, Leaf and Insect Nematodes. In: Nickle, W.R. (Ed) Manual of agricultural Nematology, New York, NY, USA, Marcel Dekker.NickleW.R.HooperD.J.1991The Aphelenchina: Bud, Leaf and Insect NematodesIn:NickleW.R.(Ed)Manual of Agricultural NematologyNew York, NY, USAMarcel DekkerSearch in Google Scholar
Nunn, G. (1992): Nematode molecular evolution. An investigation of evolutionary patterns among nematodes based upon DNA sequences. Ph.D. dissertation, University of Nottingham, UK.NunnG.1992Nematode molecular evolution. An investigation of evolutionary patterns among nematodes based upon DNA sequencesPh.D. dissertation,University of NottinghamUKSearch in Google Scholar
Rahman, R.M., Munif, A., Kurniawati, F. (2018): Deteksi dan Identifikasi Nematoda Aphelenchoides besseyi dari Benih Padi [Detection and identification of Aphelenchoides besseyi from rice seeds]. Jurnal Fitopatologi Indonesia, 14(2): 39 – 46. DOI: 10.14692/jfi.14.2.39RahmanR.M.MunifA.KurniawatiF.2018Deteksi dan Identifikasi Nematoda Aphelenchoides besseyi dari Benih Padi [Detection and identification of Aphelenchoides besseyi from rice seeds]Jurnal Fitopatologi Indonesia142394610.14692/jfi.14.2.39Abierto DOISearch in Google Scholar
Riascos-Ortiz, D., Mosquera-Espinosa, A.T., De Agudelo, F.V., De Oliveira, C., Muñoz-Florez, J.E. (2020): An integrative approach to the study of Helicotylenchus (Nematoda: Hoplolaimidae) Colombian and Brazilian populations associated with Musa crops. J Nematol, 52, 1 – 19. DOI: 10.21307/jofnem-2020-054Riascos-OrtizD.Mosquera-EspinosaA.T.De AgudeloF.V.De OliveiraC.Muñoz-FlorezJ.E.2020An integrative approach to the study of Helicotylenchus (Nematoda: Hoplolaimidae) Colombian and Brazilian populations associated with Musa cropsJ Nematol5211910.21307/jofnem-2020-054Abierto DOISearch in Google Scholar
Sanchez-Monge, A., Flores, L., Salazar, L., Hockland, S., Bert, W. (2015): An updated list of the plants associated with plant-parasitic Aphelenchoides (Nematoda: Aphelenchoididae) and its implications for plant-parasitism within this genus. Zootaxa 4013(2):207. DOI: 10.11646/zootaxa.4013.2.3Sanchez-MongeA.FloresL.SalazarL.HocklandS.BertW.2015An updated list of the plants associated with plant-parasitic Aphelenchoides (Nematoda: Aphelenchoididae) and its implications for plant-parasitism within this genusZootaxa4013220710.11646/zootaxa.4013.2.3Abierto DOISearch in Google Scholar
Santos, E., Silva, I., Rosa, R., Oliveira, C., Buonicontro, D. (2021): Pathogenicity of Aphelenchoides pseudogoodeyi on the ornamental plants Asplenium nidus and Lilium speciosum in Brazil. Helminthologia, 58(1): 74 – 84. DOI: 10.2478/helm-2021-0003SantosE.SilvaI.RosaR.OliveiraC.BuonicontroD.2021Pathogenicity of Aphelenchoides pseudogoodeyi on the ornamental plants Asplenium nidus and Lilium speciosum in BrazilHelminthologia581748410.2478/helm-2021-0003Abierto DOISearch in Google Scholar
Sauer, M.R., Winoto, R. (1975): The genus Helicotylenchus Steiner 1945 in West Malaysia. Nematologica, 341 – 350. DOI: 10.1163/187529275x00077SauerM.R.WinotoR.1975The genus Helicotylenchus Steiner 1945 in West MalaysiaNematologica34135010.1163/187529275x00077Abierto DOISearch in Google Scholar
Schmitt, D.P., Sipes, B.S. (2000): Plant-Parasitic Nematodes and Their Management. In: Silva, J.A., Uchida, R. (Eds) Plant Nutrient Management in Hawaii's Soils, Approaches for Tropical and Subtropical Agriculture. College of Tropical Agriculture and Human Resources, University of Hawaii at Manoa.SchmittD.P.SipesB.S.2000Plant-Parasitic Nematodes and Their ManagementIn:SilvaJ.A.UchidaR.(Eds)Plant Nutrient Management in Hawaii's Soils, Approaches for Tropical and Subtropical AgricultureCollege of Tropical Agriculture and Human Resources, University of Hawaii at ManoaSearch in Google Scholar
Sembiring, A.I.M., Kurniawati, F., Supramana. (2019): Karakterisasi internal transcribed spacer (ITS) rDNA nematoda pucuk putih (Aphelenchoides besseyi Christie) [(Characterization of the internal transcribed spacer (ITS) rDNA of white tip nematode (Aphelenchoides besseyi Christie)]. Agrovigor,12(1): 16 – 25. DOI: 10.21107/agrovigor.v12i1.5085 (In Turkish)SembiringA.I.M.KurniawatiF.Supramana2019Karakterisasi internal transcribed spacer (ITS) rDNA nematoda pucuk putih (Aphelenchoides besseyi Christie) [(Characterization of the internal transcribed spacer (ITS) rDNA of white tip nematode (Aphelenchoides besseyi Christie)]Agrovigor121162510.21107/agrovigor.v12i1.5085(In Turkish)Abierto DOISearch in Google Scholar
Sher, S.A. (1966): Revision of the Hoplolaiminae (Nematoda) VI. Helicotylenchus Steiner, 1945. Nematologica, 12: 1 – 56SherS.A.1966Revision of the Hoplolaiminae (Nematoda) VI. Helicotylenchus Steiner, 1945Nematologica12156Search in Google Scholar
Subbotin, S.A., Volvas, N., Yeates, G.W., Hallmann, J., Kiewnich, S., Chizhov, V.N., Manzanilla-Lopez, R.H., Inserra, R.N.,Castaillo, P. (2015): Morphological and molecular characterization of Helicotylenchus pseudorobustus (Steiner, 1914) Golden, 1956 and related species (Tylenchida: Hoplolaimidae) with a phylogeny of the genus. Nematology, 17: 27 – 52. DOI: 10.1163/15685411-00002850SubbotinS.A.VolvasN.YeatesG.W.HallmannJ.KiewnichS.ChizhovV.N.Manzanilla-LopezR.H.InserraR.N.CastailloP.2015Morphological and molecular characterization of Helicotylenchus pseudorobustus (Steiner, 1914) Golden, 1956 and related species (Tylenchida: Hoplolaimidae) with a phylogeny of the genusNematology17275210.1163/15685411-00002850Abierto DOISearch in Google Scholar
Uzma, I., Nasira, K., Firoza, K., Shahina, F. (2015): Review of the genus Helicotylenchus Steiner, 1945 (Nematoda: Hoplolaimidae) with updated diagnostic compendium. Pak J Nematol, 33: 115 – 160. DOI: 10.18681/2015.V33.I02.P01201507310001UzmaI.NasiraK.FirozaK.ShahinaF.2015Review of the genus Helicotylenchus Steiner, 1945 (Nematoda: Hoplolaimidae) with updated diagnostic compendiumPak J Nematol3311516010.18681/2015.V33.I02.P01201507310001Abierto DOISearch in Google Scholar
William-A.M., Collins, A., Esker, P. D. (2018): Plant Parasitic Nematode Explained. College of Agricultural Science The Pennsylvania State University. Retrieved from https://extension.psu.edu/plant-parasitic-nematodes-explainedWilliamA.M.CollinsA.EskerP. D.2018Plant Parasitic Nematode ExplainedCollege of Agricultural Science The Pennsylvania State UniversityRetrieved from https://extension.psu.edu/plant-parasitic-nematodes-explainedSearch in Google Scholar
Wouts, W.M., Yeates, G.W. (1994): Helicotylenchus species (Nematoda: Tylenchida) from native vegetation and undisturbed soils in New Zealand. N Z J Zool, 21: 213 – 224. DOI: 10.1080/03014223.1994.9517988WoutsW.M.YeatesG.W.1994Helicotylenchus species (Nematoda: Tylenchida) from native vegetation and undisturbed soils in New ZealandN Z J Zool2121322410.1080/03014223.1994.9517988Abierto DOISearch in Google Scholar
Wulandari, A.S., Indarti, S. (2020): Distribution and abundance of a new pest “root and bulb parasitic nematode” at different elevation levels and soil abiotic factors garlic growing centres in Central Java. In: Tutik, D.W., Roto, R., Rohana, A., Laurent, C., Kuwat, T., Indriana, K., Julius, M., Dwi, S. (Eds) Key Engineering Materials. Vol. 840. Trans Tech Publications Ltd, Switzerland, pp. 124 – 130. DOI: 10.4028/www.scientific.net/KEM.840.124WulandariA.S.IndartiS.2020Distribution and abundance of a new pest “root and bulb parasitic nematode” at different elevation levels and soil abiotic factors garlic growing centres in Central JavaIn:TutikD.W.RotoR.RohanaA.LaurentC.KuwatT.IndrianaK.JuliusM.DwiS.(Eds)Key Engineering Materials840Trans Tech Publications LtdSwitzerland12413010.4028/www.scientific.net/KEM.840.124Abierto DOISearch in Google Scholar
Wulandari, A.S., Indarti, S., Massadeh, M.I., Van Minh, N.V. (2021): The effect of abiotic factors and elevation on the diversityof plant parasitic nematodes in garlic on Central Java, Indonesia. Sarhad J. Agric., 37 (Special issue 1): 75 – 83. DOI: 10.17582/journal.sja/2021/37.s1.75.83WulandariA.S.IndartiS.MassadehM.I.Van MinhN.V.2021The effect of abiotic factors and elevation on the diversityof plant parasitic nematodes in garlic on Central Java, IndonesiaSarhad J. Agric.37Special issue 1758310.17582/journal.sja/2021/37.s1.75.83Abierto DOISearch in Google Scholar
Zhang, Y., Li, S., Li, H., Wang, R., Zhang, K., Xu, J. (2020): Fungi-Nematode Interactions: Diversity, Ecology, and Biocontrol Prospects in Agriculture. J Fungi (Basel), 6(206): 1 – 24. DOI: 10.3390/jof6040206ZhangY.LiS.LiH.WangR.ZhangK.XuJ.2020Fungi-Nematode Interactions: Diversity, Ecology, and Biocontrol Prospects in AgricultureJ Fungi (Basel)620612410.3390/jof6040206Abierto DOISearch in Google Scholar
Zhao, Z.Q. (2006): Occurrence, taxonomy, biology and pathogenicity of aphelenchid nematodes associated with conifers in south-eastern Australia. Ph.D. Thesis, The University of Adelaide, South Australia.ZhaoZ.Q.2006Occurrence, taxonomy, biology and pathogenicity of aphelenchid nematodes associated with conifers in south-eastern AustraliaPh.D. Thesis,The University of AdelaideSouth AustraliaSearch in Google Scholar
