Evaluation of Weed Species for Host Status to the Root-Knot Nematodes Meloidogyne enterolobii and M. incognita Race 4.

Weeds that compete with valuable crops can also host plant-parasitic nematodes, acting as a source of nematode inoculum in a field and further damaging crops. The host status of 10 weed species commonly found in North Carolina, USA, was determined for the root-knot nematodes Meloidogyne enterolobii and M. incognita race 4 in the greenhouse. Each weed species was challenged with 5,000 eggs/plant of either M. enterolobii or M. incognita race 4, with five replicate plants per treatment in two separate greenhouse trials. Root galling severity and total number of nematode eggs per root system were recorded 60 days after inoculation. Reproduction factor (Rf = final nematode population/initial nematode population) was calculated to determine the host status of each weed species to M. enterolobii and M. incognita race 4. Four weed species (Datura stramonium, Digitaria sanguinalis, Senna obtusifolia, and Cyperus esculentus) were poor hosts (Rf < 1) to both nematode species, and roots of these weed plants did not display galling. Four weed species (Ipomoea hederacea, Amaranthus palmeri, Portulaca pilosa, and Ipomoea lacunosa) were hosts (Rf > 1) to both nematode species, and all had observable root gall formation. Sida rhombifolia and Cyperus rotundus were poor hosts to M. enterolobii but susceptible hosts to M. incognita. This study documents a differential host status of some common weeds to M. enterolobii and M. incognita race 4, and these results highlight the necessity of managing root-knot nematodes through controlling weeds in order to protect valuable crops.

First report of the barley root-knot nematode, Meloidogyne naasi Franklin, 1965 from Pennsylvania, with new host report.

Meloidogyne naasi Franklin, 1965, the barley root-knot nematode, was originally found in field crops such as cereals, grasses, and sugar beet (Beta vulgaris L.) in England and Wales, (Franklin,1965). This nematode is one of the most significant root-knot nematodes impacting grains in European countries (Santos et al. 2020). Among root-knot nematode species, M. naasi, exhibits a distinct preference for grasses, with documented impacts on turfgrasses leading to reduced growth and vigor (Skantar et al., 2023; Cook and Yeates, 1993). In September 2022, root-knot nematode females and second-stage juveniles (J2) were recovered from roots of fowl manna grass, Glyceria striata (Lam.) Hitchc., during a nematode survey on natural vegetation at the Allegheny National Forest (41°30'13.8"N 79°09'46.2"W). Second-stage juvenile specimens were recovered from soil using sugar centrifugal flotation (Jenkins, 1964). Small galls with egg masses were dissected from fowl manna grass roots originally collected from the surveyed areas. In parallel, five plants of non-infected fowl manna grass were placed in a pot in the greenhouse using naturally nematode-infested soil collected from the same forested area. Small galls and female specimens recovered from these plants were dissected and processed for further analyses. Female and J2 were fixed in 3% formaldehyde solution and processed to glycerin (Golden, 1990; Hooper, 1970). The specimens were examined by light microscopy, morphometric measurements, and molecular markers, which included the D2-D3 region of the large ribosomal subunit 28S, and the rDNA internal transcribed spacer region (ITS). The perennial pattern of five females analyzed morphologically were consistent to the patterns observed for M. naasi. The perennial patterns had coarse ridges on the cuticle in dorsal region forming broken irregular lines around anal and phasmid area. We also noted a prominent fold that covered some of the anus and showed a curved line between vulval slit and phasmids, typical of M. naasi. The area around the vulval area had a few or no striae except for a few lines radiating from the vulval slit as in the original description. Measurements of ten J2 had a body length ranged between 380 and 410 µm, stylet 11-13 µm, tail 50-70 µm long with a hyaline tail terminus between 12-22 µm in length, 4 lines in the lateral field, a and c ratio between 29.23-35.91 and 5.79-7.9 fitting the original description by Franklin, 1965 and others populations found in the USA (Skantar et al., 2023). The matrix codes for the female specimes are A32, B324, C3, D3 and for J2's A2, B21, C123, D1, E3, F12 (Subbotin et al., 2021). The amplified DNA fragments were sequenced, resulting in an 726 bp fragment flanked by the D2-D3 primers (PP097762), while for the ITS primers an 634 bp fragment was obtained (PP092043). Both generated sequences for the specimens collected in Pennsylvania revealed >99% similarity to M. naasi sequences deposited at GenBank, and therefore, validating the morphological analyses. Based on both morphological and molecular analyses the specimens collected in the state of Pennsylvania were identified as M.naasi. To our knowledge, this is the first report of this species from this state and being associated with naturally infected fowl manna grass.

Host status of melon, carrot, and Meloidogyne incognita-susceptible and -resistant cotton, cowpea, pepper, and tomato for M. floridensis from California.

The host status of carrot, melon, and susceptible and resistant cultivars of tomato, cotton, cowpea, and pepper for a California isolate of the peach root-knot nematode Meloidogyne floridensis was determined in greenhouse pot experiments. It was compared to a race 3 isolate of M. incognita. Melon was an excellent host for both isolates and roots were heavily galled after the 8-week trial. Carrot was a host for M. incognita, but a poor host for M. floridensis, although both isolates caused similar levels of galling. Susceptible cotton was a good host for M. incognita race 3, but a poor host for M. floridensis. Susceptible tomato, cowpea, and pepper were good hosts for both isolates. The M. incognita resistance in tomato and pepper was broken by M. floridensis. Resistant cowpea was a maintenance host as population levels of M. floridensis remained virtually unchanged over the trial period. We conclude that M. floridensis poses a risk to some important vegetable crops in California, as it reproduces on most vegetable crops, including some cultivars that are resistant to M. incognita. On susceptible crops, the reproduction of M. floridensis was always significantly less than that of M. incognita, and we hypothesize that in mixed species field populations, M. incognita will outcompete M. floridensis. This study demonstrates that efforts to limit the spread and prevent further introductions of M. floridensis in California are important to maintain the effectiveness of plant resistance as a nematode management strategy in vegetable crops.

Depth distribution of plant-parasitic nematodes on bentgrass golf greens in Missouri and Indiana.

Control of plant-parasitic nematodes (PPNs) on golf putting greens with nematicides is dependent on the seasonal occurrence and depth distribution of target PPN populations. This study aimed to determine if plant-parasitic nematode populations on golf course putting greens in Missouri and Indiana peaked at a targetable depth at a specific time in the year, focusing primarily on lance (Hoplolaimus spp.) and root-knot (Meloidogyne spp.) nematodes. To elucidate species diversity in the region, rDNA from a subset of lance and root-knot nematodes was sequenced and analyzed, with additional micromorphology of a lance nematode assessed in scanning electron micrographs (SEM). Soil samples were taken to a depth of 25 cm and stratified into 5 cm increments during April, June, August and October at seven sites across Missouri, three in the Kansas City metro of Kansas in 2021 and in ten sites across Indiana in 2022. Samples were stratified in five-centimeter increments and aggregated for a total of 100 cm3 of soil at each depth for each sampling. Samples were processed using a semi-automatic elutriator followed by the sucrose-flotation method, and populations were counted using a hemocytometer and recorded. For molecular characterization, rDNA was extracted and analyzed from 31 individual lance nematodes from one site in Missouri and eight sites in Indiana, and 13 root-knot nematodes from nine sites across Indiana. A significant interaction occurred between sampling month and depth for lance and ring nematodes Missouri/KS, with both PPN populations peaking at the 0-5 cm depth during October, which is well after most targeted nematicide applications are applied. Ring nematodes in Indiana did not follow this trend and were most abundant in August at a depth of 0-5 cm. No significant interaction between depth and month occurred for lance or root-knot nematodes in Indiana, or root-knot nematodes in Missouri/KS. Hoplolaimus stephanus and H. magnistylus were the lance species identified on golf greens, and Meloidogyne naasi, M. graminicola and M. marylandi were the root-knot species identified. Scanning-electron micrographs confirmed morphological characteristics unique to H. stephanus.

Response of the symbiotic microbial community of Dioscorea opposita Thunb. cv. Tiegun to root-knot nematode infection.

Yam is an important medicinal and edible dual-purpose plant with high economic value. However, nematode damage severely affects its yield and quality. One of the major effects of nematode infestations is the secondary infection of pathogenic bacteria or fungi through entry wounds made by the nematodes. Understanding the response of the symbiotic microbial community of yam plants to nematodes is crucial for controlling such a disease. In this study, we investigated the rhizosphere and endophytic microbiomes shift after nematode infection during the tuber expansion stage in the Dioscorea opposita Thunb. cv. Tiegun yam. Our results revealed that soil depth affected the abundance of nematodes, and the relative number of Meloidogyne incognita was higher in the diseased soil at a depth of 16-40 cm than those at a depth of 0-15 cm and 41-70 cm. The abundance of and interactions among soil microbiota members were significantly correlated with root-knot nematode (RKN) parasitism at various soil depths. However, the comparison of the microbial alpha diversity and composition between healthy and diseased rhizosphere soil showed no difference. Compared with healthy soils, the co-occurrence networks of M. incognita-infested soils included a higher ratio of positive correlations linked to plant health. In addition, we detected a higher abundance of certain taxonomic groups belonging to Chitinophagaceae and Xanthobacteraceae in the rhizosphere of RKN-infested plants. The nematodes, besides causing direct damage to plants, also possess the ability to act synergistically with other pathogens, especially Ramicandelaber and Fusarium, leading to the development of disease complexes. In contrast to soil samples, RKN parasitism specifically had a significant effect on the composition and assembly of the root endophytic microbiota. The RKN colonization impacted a wide variety of endophytic microbiomes, including Pseudomonas, Sphingomonas, Rhizobium, Neocosmospora, and Fusarium. This study revealed the relationship between RKN disease and changes in the rhizosphere and endophytic microbial community, which may provide novel insights that help improve biological management of yam RKNs.

Characterization of data observing Meloidogyne incognita, Neofusicoccum parvum, and Xylella fastidiosa infection effects on development of grapevine phenolic compound levels and resistance to subsequent Neofusicoccum parvum infections.

Grapevines encounter many different pathogens throughout their lifespans, including the bacterial pathogen Xylella fastidiosa, which causes Pierce's disease that results in vascular occlusion and eventual plant host death, the fungal pathogen Neofusicoccum parvum, which causes stem cankers that kill individual vines and reduce fruit yields, and the root knot nematode Meloidogyne incognita, which destroys root tissues that impacts host vigour. To date, little research has been conducted to examine how one infection could impact subsequent infections by the same or different pathogens despite this is important to ensure healthy vineyards. Therefore, grapevines initially infected with either X. fastidiosa, N. parvum, or M. incognita were subsequently infected with N. parvum eight weeks later to observe developing lesion lengths, which were assessed to determine grapevine resistance to infections. Collected data shows that when prior infections were present, the N. parvum lesions lengths were smaller. This suggests grapevines had induced resistance to combat infections. Further, defence-associated phenolics were measured by high-performance liquid chromatography to determine roles in observed resistance to the secondary N. parvum infections. Data shows that of the different phenolics examined, only stilbenoids were different due to infections, with lowered levels observed in plants that were infected compared with non-infected controls. These data provide insight into how infections by different pathogens could impact grapevine host resistance to new, subsequent pathogen infections.

First report of Meloidogyne hapla on hemp (Cannabis sativa) in Oregon.

Hemp is a crop that has gained interest in Washington and Oregon. As with other crops, hemp production faces challenges due to biotic factors, including plant-parasitic nematodes. During a survey for plant-parasitic nematodes associated with hemp, Meloidogyne sp. was found in a composite root sample collected in Oregon. Morphological characterization of second-stage juveniles identified the nematode as Meloidogyne hapla. Molecular identification confirmed the population as M. hapla. To our knowledge, this is the first report of M. hapla on hemp in the Pacific Northwest of the United States.

First Report of the Root-Knot Nematode Meloidogyne incognita on Bletilla striata Rchb.f. (Orchidaceae) in China.

Bletilla striata Rchb.f., is a perennial herbaceous bulbous plant known as the Chinese ground or hyacinth orchid classified in the Orchidaceae. It is native to southeast Asia and mainly distributed in China, Japan and northern Myanmar (He et al. 2017). It has the functions of astringent hemostasis and analgesia, and can also be used to treat traumatic bleeding, ulcers, swelling and chapped skin. Therefore, it occupies an important position in traditional Chinese medicine (Xu et al. 2019). In June 2023, three farmers in Mengzi (103.39°N, 23.21°E), Yunnan Province, China, observed that some Bletilla striata Rchb.f. plants grew poorly with small and chlorotic leaves (Figure 1 A). We suspected that these symptoms were caused by root-knot nematode infection, but the galls on the roots were small and inconspicuous (Figure 1 A). The presence of nematode females in both the galled regions and the normal roots (Figure 1 B), revealed by fuchsin staining (Byrd et al. 1983), indicated that the symptoms were probably caused by root-knot nematode infection. To estimate the incidence rates, we randomly selected 100 B. striata Rchb.f. plants from each of five fields representing a total area of 3000 m2. In these fields, the occurrence of stained root-knot nematodes were 19.3%, 17%, 18.3%, 15%, and 13%, respectively. The gall rating of the infected plants in the B striata Rchb.f. samples collected from the five fields was 2 (rating scale of 0 to 5). Females (n=20), second-stage juveniles (J2s, n=20) and egg masses (n=20) were extracted and collected from roots and soil for morphological and molecular identification. The females had a white, pyriform body and their perineal patterns exhibited a high and square dorsal arch, lacking distinct lateral line (Figure 1. C & D). Measurements of females (n = 20) were: body length (BL) = 708.64±89.6 µm (554.36 to 844.51 µm); maximum body width (BW) = 461.73±47.44 µm (365.25 to 561.49 µm); stylet length (ST) = 15.49±3.15 µm (10.55 to 19.78 µm); and distance from dorsal esophageal gland opening to the stylet knobs (DGO) = 3.33±0.27 µm (2.77 to 3.93 µm). Measurements of J2s (n=20) were BL = 417.7±47.67 µm (342.16 to 499.68 µm); BW = 15.74±2.66 µm (11.05 to 25.63 µm); ST = 12.49±1.12 µm (10.19 to 15.02 µm); DGO = 2.64±0.59 µm (40.17 to 68.74 µm); tail length = 51.93±8.55 µm (10.43 to 27.22 µm); hyaline tail terminus = 18.23±3.99 µm (1.48 to 3.98 µm). These morphological features match the description of Meloidogyne incognita (Eisenback et al. 1981). To further confirm the species, we selected three infected plants from each field for molecular identification, the ITS region amplified using the primers 18S/26S (5'-TTGATTACGTCCCTGCCCTTT-3',5'-TTTCACTCGCCGTTACTAAGG-3') (Vrain et al. 1992) . A 729 bp PCR product of ITS region (accession nos. OR463907) was obtained from all infected plants. The amplicons from 18S/26S primer pair were sequenced and the sequences showed 95.29% homology with sequences of M. incognita (accession nos. MT209948.1). Moreover, a 835 bp DNA fragment (accession nos. OR469000) was obtained using the specific primers Mi-F/Mi-R (5'-GTGAGGATTCAGCTCCCCAG-3',5'-ACGAGGAACATACTTCTCCGTCC-3') for M. incognita (Meng et al. 2004), the sequence showed 99.28% homology with sequences of M. incognita (accession nos. ON416569). The morphological features and molecular data confirmed the identification of the root-knot nematode on B. striata Rchb.f. as M. incognita. To confirm the pathogenicity, ten healthy B. striata Rchb.f. seedlings were each inoculated with 500 freshly hatched J2s isolated from field Bletilla striata Rchb.f.. Five healthy seedlings without J2 inoculation were used as controls. At 60 days after inoculation, most of the inoculated plants exhibited similar symptoms to those initially observed by farmers in the field. On average, 1532 J2s were recovered from each inoculated plant, yielding a reproductive factor of 2.1. The gall rating for these inoculated plants was 2. Fuchsin staining revealed the presence of root-knot nematode females within the roots, with an average of 17 females detected per inoculated plant. No symptoms were observed in the control plants. This is the first report of M. incognita infecting B. striata Rchb.f. in China. M. incognita can cause severe infection and damage to some crops, resulting in serious economic losses (Eisenback, 2022). The growers need to take measures to prevent the spread of this nematode.

Brown planthopper infestation on rice reduces plant susceptibility to Meloidogyne graminicola by reducing root sugar allocation.

Plants are simultaneously attacked by different pests that rely on sugars uptake from plants. An understanding of the role of plant sugar allocation in these multipartite interactions is limited. Here, we characterized the expression patterns of sucrose transporter genes and evaluated the impact of targeted transporter gene mutants and brown planthopper (BPH) phloem-feeding and oviposition on root sugar allocation and BPH-reduced rice susceptibility to Meloidogyne graminicola. We found that the sugar transporter genes OsSUT1 and OsSUT2 are induced at BPH oviposition sites. OsSUT2 mutants showed a higher resistance to gravid BPH than to nymph BPH, and this was correlated with callose deposition, as reflected in a different effect on M. graminicola infection. BPH phloem-feeding caused inhibition of callose deposition that was counteracted by BPH oviposition. Meanwhile, this pivotal role of sugar allocation in BPH-reduced rice susceptibility to M. graminicola was validated on rice cultivar RHT harbouring BPH resistance genes Bph3 and Bph17. In conclusion, we demonstrated that rice susceptibility to M. graminicola is regulated by BPH phloem-feeding and oviposition on rice through differences in plant sugar allocation.

First report of the damaging pest Meloidogyne enterolobii parasitizing mulberry (Morus alba L.) in Vietnam.

Mulberry (Morus alba L.) is highly important crop in Vietnam, playing a key role in the country's economy through sericulture, food supply, pharmaceuticals, and beverage industries (Nguyen et al., 2018; Rohela et al., 2020). Recently, many mulberry-growing areas in Lam Dong, Vietnam have reported severe symptoms associated with nematode infection, including yellowing leaves, stunted growth, and severe root galling, leading to a significant decline in mulberry productivity. From April to December 2022, twenty soil and root samples from mulberry-growing areas in Lam Dong (Da Teh: 11°28'48.11"N; 107°28'23.74"E elevation: 133m; Lam Ha 11°48'25.13"N; 108°14'7.13"E elevation: 848m) were collected to uncover the presence of Meloidogyne enterolobii parasitizing mulberry in Vietnam. One nematode population was randomly selected for characterizing in this study among analyzed nematode populations. Females were extracted from heavily galled roots (Fig. S1) from a single mulberry tree in Lam Dong, Vietnam, using a needle and forceps (Subbotin et al., 2021). The perineal patterns of adult females (n = 10) have an oval shape, with clearly visible phasmids, along with a prominently high and squared dorsal arch. The striae are smooth and coarse, while the perivulval region remains devoid of striae. The lateral lines appear indistinct, and the tail tip is easily observable. Morphometric measurements were as follows: body length = 585 ± 78 (464-724) µm, body width = 367 ± 75 (271-529) µm, neck length = 221.5 ± 30.7 (167-269.6) µm, stylet length = 13.1 ± 1.2 (11.4-15.1) µm, vulva-slit length 16.3±2.3 (10.4-18) µm, vulva-anus distance = 16.8±3.0 (11.4-18) µm, anus-tail tip distance = 10.3±2.1 (6.9-14.2) µm, interphasmidial distance = 15.9 ± 3.7 (10.3-23.4) µm. The morphology of this nematode population is highly in agreement with the original description of M. enterolobii (Yang & Eisenback, 1983). This population was also identified using the D2-D3 of 28S rRNA and 18S rRNA (Powers et al., 2017; Subbotin et al., 2006) regions. The D2-D3 of 28S rRNA sequences from this study (accession numbers: OR889633) exhibited 99.5-99.8% similarity to the sequences of M. enterolobii from GenBank (accession numbers: OR214950 and ON496981). While the 18S rRNA sequences (accession numbers: OR896547) showed 99.2-99.3% similarity to the sequences of M. enterolobii from GenBank (accession numbers: MZ955995, MZ531901, and MW488150). To carry out Koch's postulates, 2000 J2s from collected M. enterolobii egg masses (initial population) were inoculated on two-month-old plantlets of mulberry (n = 6), planted on 2L pots within a screenhouse, non-inoculated plantlets (n=6) served as negative controls. After 90 days post-inoculation, nematode reproduction factors (RF = final density (nematodes were extracted from the whole root system and corresponding soil samples (Subbotin et al., 2021)) / initial population) and root damage symptoms were evaluated. The inoculated plantlets exhibited consistent yellowing leaves, stunting, and root galling symptoms (Fig. S1), mirroring observations from the field, with an average RF of 11.5. Control plants displayed no symptoms. Root-knot nematodes extracted from the roots were identified as M. enterolobii through molecular analyses of D2-D3 of 28S and 18S rRNA regions (GenBank accession numbers: OR889634 (D2-D3 of 28S) and OR896548 (18S)), thereby confirming that mulberry acts as a host for M. enterolobii. Currently, this nematode has been reported to be associated with two different host plants, including guava (Trinh et al., 2022) and pomelo (Le et al., 2023). Our discovery marks the first documented case of Meloidogyne enterolobii parasitizing mulberry in Vietnam. While the impact on mulberry productivity remains to be really important for sericulture food supply, pharmaceuticals, and beverage industries; the aggressive nature of M. enterolobii, as observed in the field and confirmed by the screenhouse tests, raises concerns about potential economic losses in mulberry production. Therefore, further investigations are needed to assess the extent of M. enterolobii infestation in mulberry orchards and to develop effective control measures to safeguard the sustainability of mulberry cultivation in Vietnam.

Host Suitability of Summer Cover Crops and Peach Rootstocks to the Peach Root-Knot Nematode, Meloidogyne floridensis.

Greenhouse experiments were conducted to determine the host suitability of ten summer cover crops and two peach rootstocks to Meloidogyne floridensis by inoculating them with 10,000 M. floridensis eggs. Brown top millet and sunn hemp were nonhosts as they did not support nematode reproduction. Buckwheat, cowpea, pearl millet, Japanese millet, and sunflower supported more than 25,000 eggs/pot, which indicated that these crops are good hosts to M. floridensis. The crops that supported poor nematode reproduction were sesame, grain sorghum, and sorghum-sudangrass, with their reproduction ranging from 219 to 7,750 eggs/pot. In addition to having many galls on the roots, the peach rootstock Guardian had 10,100 eggs on the roots and 450 second-stage juveniles in the pot, which indicated that 'Guardian' is a good host to M. floridensis. Although the nematode reproduction on MP-29 rootstock was relatively lower, the presence of many large galls on the roots indicates MP-29 is susceptible to M. floridensis. Results from the current study suggest that the employment of nonhost cover crops and poor-host rootstocks could aid in effective nematode management programs for peaches.

Eco-friendly management of Meloidogyne incognita in cadmium-contaminated soil by using nematophagous fungus Purpureocillium lavendulum YMF1.683: Efficacy and mechanism.

Root-knot nematodes (RKNs) are distributed globally, including in agricultural fields contaminated by heavy metals (HM), and can cause serious crop damages. Having a method that could control RKNs in HM-contaminated soil while limit HM accumulation in crops could provide significant benefits to both farmers and consumers. In this study, we showed that the nematophagous fungus Purpureocillium lavendulum YMF1.683 exhibited a high nematocidal activity against the RKN Meloidogyne incognita and a high tolerance to CdCl2. Comparing to the P. lavendulum YMF1.838 which showed low tolerance to Cd2+, strain YMF1.683 effectively suppressed M. incognita infection and significantly reduced the Cd2+ uptake in tomato root and fruit in soils contaminated by 100 mg/kg Cd2+. Transcriptome analyses and validation of gene expression by RT-PCR revealed that the mechanisms contributed to high Cd-resistance in YMF1.683 mainly included activating autophagy pathway, increasing exosome secretion of Cd2+, and activating antioxidation systems. The exosomal secretory inhibitor GW4869 reduced the tolerance of YMF1.683 to Cd2+, which firstly demonstrated that fungal exosome was involved in HM tolerance. The up-regulation of glutathione synthesis pathway, increasing enzyme activities of both catalase and superoxide dismutase also played important roles in Cd2+ tolerance of YMF1.683. In Cd2+-contaminated soil, YMF1.683 limited Cd2+-uptake in tomato by up-regulating the genes of ABCC family in favor of HM sequestration in plant, and down-regulating the genes of ZIP, HMA, NRAMP, YSL families associated with HM absorption, transport, and uptake in plant. Our results demonstrated that YMF1.683 could be a promising bio-agent in eco-friendly management of M. incognita in Cd2+ contaminated soils.

Eryngium foetidum reported as a new host of Meloidogyne enterolobii in the state of Pará, in the eastern Amazon region, Brazil.

Amazon chicory (Eryngium foetidum L. [Apiaceae]), also known as culantro, is native to Tropical America and the West Indies. It belongs to the unconventional food plants (UFPs) group, and in addition to be consumed as a spice herb, it possesses a wide range of ethnomedicinal uses (Paul et al. 2011). In 2019, in the eastern Amazon region of Brazil, state of Pará, producers of E. foetidum in the municipality of Castanhal (01°15'363" S 047°10'232" W) reported the occurrence of underdeveloped plants with leaf yellowing and a large number of galls in the root system, which are typical symptoms of root-knotting nematode. Soil and root samples were collected and sent to the Nematology Laboratory (LabNema) located at the Faculty of Agrarian and Veterinary Sciences, UNESP, Jaboticabal, São Paulo, Brazil. A total of 46 second-stage juveniles (J2s) were extracted per 100 cm3 of soil, and a total of 460 eggs and J2s Meloidogyne spp. were found per gram of root. Morphological and molecular techniques were used to identify the species. The analysis of the perineal patter of ten females revealed thin striations in an oval shape with a high and semi-trapezoidal dorsal arch. No striations were observed in the perivulvar region. The labial region of the ten males analyzed exhibited a non-prominent labial disc, fused and slightly recessed submedian lips, with no apparent annulations. The morphological characteristics observed in the adults were consistent with those originally described for Meloidogyne enterolobii (Yang; Eisenback, 1983), confirming the species purity of the recovered population. Three individual nematodes had their 18S rDNA region sequenced (Holterman et al. 2006) which showed an average identity of 99.7% with other sequences of M. enterolobii available in the GenBank database. A Bayesian phylogenetic tree was constructed, providing insights into the specific relationship of M. enterolobii recovered from E. foetidum with other related nematodes. Each of the three sequenced nematodes represented a unique haplotype, resulting in their separation into distinct clades. Moreover, the obtained sequences presented polymorphisms that differed from the M. enterolobii sequences already available in the database, highlighting the genetic diversity of this species in relation to its original host (Silva et al. 2021). The species M. enterolobii was also confirmed using species-specific primers for M. incognita, M. javanica, and M. enterolobii (Zijlstra et al. 2000; Tigano et al. 2010). To confirm the pathogenicity of M. enterolobii on E. foetidum, a modified Koch Postulate was conducted. Six seedlings of E. foetidum were transplanted individually to 10-liter pots containing autoclaved soil. Each pot was then inoculated with 5 mL of a suspension containing 3,000 eggs and J2s from the original population of M. enterolobii obtained from E. foetidum. After 90 days, the inoculated plants exhibited root galls with a plentiful egg mass, in contrast to the healthy non-inoculated plants. The average number of M. enterolobii nematodes recovered from the roots of the inoculated plants was 42,040 eggs and J2s, resulting in a reproduction factor (RF) of 14.0. The importance of reporting the occurrence of M. enterolobii in E. foetidum is due to the fact that this plant species is cultivated in a crop rotation system with other vegetables such as lettuce and coriander, which are also hosts of M. enterolobii. Consequently, different crop rotation strategies and control alternatives need to be considered in areas where E. foetidum is grown. This is the first report of E. foetidum serving as a host for the root-knot nematode M. enterolobii worldwide.

Root-knot nematode infections and soil characteristics significantly affected microbial community composition and assembly of tobacco soil microbiota: a large-scale comparison in tobacco-growing areas.

Introduction: Tobacco root-knot nematode (RKN) is a highly destructive soil-borne disease worldwide. However, there is a lack of research on the relationship between RKN and tobacco root microbial community composition under large-scale geographical conditions in China. Methods: In this study, we collected 65 samples from 28 main tobacco-growing areas across 10 provinces in China and conducted 16S rDNA sequencing to investigate the dynamic microbial changes in tobacco soil infected by RKN compared to healthy tobacco soil. Based on the analysis of rhizosphere soil bacterial communities, changes after RKN infection, and soil environmental factors. Results: We found the 28 tobacco-growing areas could be divided into two distinct groups with different microbial compositions and varying responses to RKN infection. In group1 of the provinces of Anhui, Henan, Shanxi, and Heilongjiang, Vicinamibacteria dominated the bacterial community, while Acidobacteriae was present in low abundance. In contrast, group2 of the other six provinces (Yunnan, Guizhou, Chongqing, Guangxi, Hubei, and Shandong) exhibited an opposite pattern. After infected by RKN, the genera Chitinophaga increased significant in group 1, while the genera Rhodococcus in group 2 exhibited a substantial increase. Alpha-diversity analysis revealed that RKN-infected tobacco exhibited a richer and more diverse rhizosphere soil bacterial community compared to healthy tobacco in most growing areas. A total of 12 kinds of soil environmental factors were measured in healthy and RKN-infected tobacco soil, and based on the co-occurrence and correlation analysis between environmental factors and microbial species, the pH level, calcium (Ca), magnesium (Mg), phosphorus (P), iron (Fe), and sodium (Na) were identified as key environmental factors influencing the population composition of rhizosphere microorganisms during RKN infection. We observed that RKN infection further increased the pH in weakly alkaline group 1 soil, while weakly acidic group 2 soil experienced a further decrease in pH. Furthermore, we identified three genera as potential biocontrol or plant growth-promoting bacteria for tobacco. Discussion: These findings provide valuable reference data for managing RKN disease in different tobacco-growing areas and contribute to the exploration of new and effective biological control methods.

A Comparison between Meloidogyne floridensis and M. incognita from California on Susceptible and Resistant Sweetpotato Cultivars.

The reproduction and ability to cause root-galling of a California isolate of the peach root-knot nematode Meloidogyne floridensis was evaluated on seven sweetpotato (Ipomea batatas) cultivars and compared to a M. incognita race 3 and a M. incognita Mi-gene resistance-breaking isolate. The susceptible tomato (Solanum lycopersicum) cultivar 'Daniela' and the Mi-gene-carrying resistant cultivar 'Celebrity' were included as controls. Repeated trials were done in pots in a nematode-quarantine greenhouse at the University of California, Riverside. The three Meloidogyne isolates reproduced equally well on susceptible tomato. On Mi-gene resistant tomato, the reproduction and root-galling by M. floridensis was intermediate between the avirulent M. incognita race 3 and the resistance-breaking M. incognita isolate. The sweetpotato cultivars 'Beauregard' and 'Diane' were excellent hosts for all three Meloidogyne isolates. Cultivars 'Bellevue', 'Burgundy', and 'Covington' were resistant to these isolates. The cultivars 'Bonita' and 'Murasaki-29' were hosts for the M. floridensis and the resistance-breaking M. incognita isolate allowing an increase in nematode levels, while they were poor hosts resulting in a decrease in nematode levels for the M. incognita race 3 isolate. The study showed that M. floridensis can reproduce on tomato and some sweetpotato cultivars that are considered resistant to M. incognita.

Characterization of Meloidogyne enterolobii intercepted from baobab (Adansonia digitata L.) seedlings from Thailand during Japanese import plant quarantine inspection.

In April 2019, baobab (Adansonia digitata L.) seedlings from Thailand, exhibiting galls on the roots, were intercepted during an import plant quarantine inspection at Chubu Centrair International Airport, Japan. Root-knot nematodes (RKNs) were extracted from the galled roots of baobab seedlings and identified by morphological, morphometrical and molecular methods as the guava root-knot nematode, Meloidogyne enterolobii Yang & Eisenback. The morphology and morphometrics of the intercepted population were similar to those of the original and subsequent descriptions of M. enterolobii. The sequences of D2-D3 of 28S rRNA, mtDNA intergenic COII-16S rRNA and COI genes obtained in this study matched well (99-100% similarity) with each of the gene sequences of M. enterolobii deposited in GenBank. Phylogenetic analysis of these genes revealed that the intercepted population clustered with M. enterolobii and clearly differed from other RKN species. To the best of our knowledge, this is the first report of M. enterolobii from baobab.

First report of root-knot nematode, Meloidogyne javaniva, infecting Stachys byzantina on São Paulo, Brazil.

Stachys byzantina belongs to the Labiatae and is known by the names "peixinho-da-horta" (Brazil) and "lamb's ear" (USA). Its importance is associated with its medicinal properties (Bahadori et al. 2020) and nutritional aspects (Milião et al. 2022). Root-knot nematodes cause severe damage to plants and suppress production. In January 2021, plants of S. byzantina in the municipality of Jaboticabal (21°14'38.7"S, 48°17'10.6"W) showed symptoms of reduced growth, yellowed leaves and the presence of galls in the roots. Initially, samples of roots from a S. byzantina were analyzed at the Nematology Laboratory (LabNema/UNESP), Jaboticabal, Brazil, estimating 20,000 eggs and juveniles of Meloidogyne sp. in 10 g of roots. To confirm the host ability of the species, a pathogenicity test was performed using Koch's postulate. For this purpose, the test was conducted in a greenhouse where 3,000 eggs and second-stage juveniles (J2) were inoculated onto three plants (n=3) of S. byzantina. After 90 days, the inoculated plants showed the same symptoms as those observed in the field. No symptom or nematode was detected in the uninoculated plant (control). Nematodes were extracted from the roots of inoculated plants and quantified. The perineal pattern of females (n=10) (Netscher and Taylor, 1974) and the labial region of males (n=10) (Eisenback and Hirschmann, 1981) were analyzed and compared with the morphological characteristics of the original description of the species (Chitwood, 1949). For analysis based on esterase isozyme phenotype, the α-method of Esbenshade and Triantaphyllou (1990) was used, and females (n=7) were examined. To confirm identification, whole genomic DNA from an adult female (n=1) was extracted using the Qiagen DNeasy® Blood & Tissue Kit and this sample was used for both genetic sequencing and the sequence-characterized amplified region techniques (SCAR). PCR amplifications were performed for the 18s rRNA gene using primers 988F and 1912R from Holterman et al (2006). Our sequence was deposited in GenBank (NCBI) under the identifier OP422209. Finally, species-specific SCAR primers (Fjav/Rjav, Me-F/Me-R, and Finc-F/Finc-R) designed by Zijlstra (2000) were used to identify Meloidogyne spp. Koch's postulate analysis yielded the following results: (n=1) 9,280 eggs and J2 (Reproduction factor, RF = 33.09); (n=2) 111,720 eggs and J2 (RF = 37.24); (n=3) 59,700 eggs and J2 (RF = 19.9) (RF mean = 30.08). The following characteristics were observed in the perineal region of females: Low and rounded trapezoidal dorsal arch with two distinct lateral lines clearly separating the dorsal and ventral arch regions, similar to the morphological features of the species description by Chitwood (1949). Males had a convex labial plate with a non-raised labial disk joining the submedial labia, a non-rugged labial region, the basal tubercles were usually wider than high, and a rounded tail tip (Eisenback and Hirschmann 1981). The α-esterase enzyme profile showed the J3 phenotype typical of M. javanica (Rm [×100] = 46.0, 54.5, and 58.9). The 18s rRNA sequences grouped Meloidogyne sp. with species such as M. enterolobii, M. incognita, and M. javanica. A DNA fragment of about 700 bp was amplified with Mj (Fjav/Rjav) primers, but not with Me (Me-F/Me-R) and Mi (Finc-F/Finc-R) primers, which confirmed the identification of M. javanica. Accurate identification and characterization of the occurrence of new hosts of M. javanica will allow us to determine the range and geographic distribution of the species. This is the first report on the occurrence of M. javanica on S. byzantina in Brazil. This report is important so that management strategies can be applied to prevent the spread of the pest to other areas.

Transcriptome-Based Comparative Expression Profiling of Sweet Potato during a Compatible Response with Root-Knot Nematode Meloidogyne incognita Infection.

M. incognita, a root-knot nematode (RKN), infects the roots of several important food crops, including sweet potato (Ipomoea batatas Lam.), and severely reduces yields. However, the molecular mechanisms underlying infection remain unclear. Previously, we investigated differential responses to RKN invasion in susceptible and resistant sweet potato cultivars through RNA-seq-based transcriptome analysis. In this study, gene expression similarities and differences were examined in RKN-susceptible sweet potato cultivars during the compatible response to RKN infection. Three susceptible cultivars investigated in previous research were used: Dahomi (DHM), Shinhwangmi (SHM), and Yulmi (YM). Of the three cultivars, YM had the highest number of genes with altered expression in response to infection. YM was also the cultivar with the highest susceptibility to RKN. Comparisons among cultivars identified genes that were regulated in more than one cultivar upon infection. Pairwise comparisons revealed that YM and DHM shared the most regulated genes, whereas YM and SHM shared the lowest number of regulated genes. Five genes were up-regulated, and two were down-regulated, in all three cultivars. Among these, four genes were highly up-regulated in all cultivars: germin-like protein, anthranilate synthase α subunit, isocitrate lyase, and uncharacterized protein. Genes were also identified that were uniquely regulated in each cultivar in response to infection, suggesting that susceptible cultivars respond to infection through shared and cultivar-specific pathways. Our findings expand the understanding of the compatible response to RKN invasion in sweet potato roots and provide useful information for further research on RKN defense mechanisms.

A New Root-Knot Nematode species, Meloidogyne karsseni n. sp. (Nematoda: Meloidogynidae), From Mexico and a Taxonomic Update on M. paranaensis From Guatemala.

A new root-knot nematode (RKN) species, Meloidogyne karsseni n. sp., associated with sweet pepper from Mexico, and a population of M. paranaensis from Guatemala, are described using data from morphological, biochemical (isozyme enzymes), molecular, and phylogenetic analyses. Meloidogyne karsseni n. sp. can be morphologically diagnosed using the combined features of the second-stage juveniles, viz. body length (345 to 422 µm), a conical rounded head region, a post-labial annule lacking transverse striation, a thin stylet 11 to 12 µm long, rounded to oval and backwardly sloping knobs, dorsal gland orifice (DGO) at 5.2 to 6.0 µm from the knobs, a hemizonid just above the secretory-excretory (SE) pore, a tapering tail with finely rounded terminus and one or two very weak constrictions at hyaline tail tip; the female characters viz. oval-to-rounded perineal pattern with coarse striation on lateral sides around the anus, low dorsal arch with finer striations, and distinctly visible lateral lines; and the male characteristics viz. a rounded and continuous head, a post-labial annule without transverse striations, a robust stylet 20 to 24 µm long, rounded-to-oval and slightly backwardly sloping knobs, and a DGO at 2.4 to 2.9 µm from the knobs. In all the studied males of M. paranaensis, a characteristic sclerotization around the duct of SE-pore was also observed for the first time. Sequences of 18S, D2-D3 of 28S, and ITS of rDNA, and cox1 of mtDNA were generated for the two species, and in the phylogenetic trees based on these genes, both species appeared in the tropical RKN species complex clade.

Root-Knot-Nematode-Encoded CEPs Increase Nitrogen Assimilation.

C-terminally encoded peptides (CEPs) are plant developmental signals that regulate growth and adaptive responses to nitrogen stress conditions. These small signal peptides are common to all vascular plants, and intriguingly have been characterized in some plant parasitic nematodes. Here, we sought to discover the breadth of root-knot nematode (RKN)-encoded CEP-like peptides and define the potential roles of these signals in the plant-nematode interaction, focusing on peptide activity altering plant root phenotypes and nitrogen uptake and assimilation. A comprehensive bioinformatic screen identified 61 CEP-like sequences encoded within the genomes of six root-knot nematode (RKN; Meloidogyne spp.) species. Exogenous application of an RKN CEP-like peptide altered A. thaliana and M. truncatula root phenotypes including reduced lateral root number in M. truncatula and inhibited primary root length in A. thaliana. To define the role of RKN CEP-like peptides, we applied exogenous RKN CEP and demonstrated increases in plant nitrogen uptake through the upregulation of nitrate transporter gene expression in roots and increased 15N/14N in nematode-formed root galls. Further, we also identified enhanced nematode metabolic processes following CEP application. These results support a model of parasite-induced changes in host metabolism and inform endogenous pathways to regulate plant nitrogen assimilation.