Molecular diagnostics of the pigeon pea cyst nematode, Heterodera cajani Koshy, 1967, using real-time PCR

Summary The pigeon pea cyst nematode, Heterodera cajani, is an important nematode pest of pigeon pea that is present in all major growing regions of this crop in India and reported from Pakistan, Egypt and Myanmar. In this study, a new real-time PCR assay for detection of H. cajani using a species-specific primer and a TaqMan probe was developed. The primers and a probe were designed to amplify the COI gene fragment. The specificity of the primer-probe set was tested in singleplex or multiplex reactions against target and non-target nematodes. In multiplex real-time PCR experiments with the specific and universal primer-probe sets, the signals were simultaneously observed for COI and D3 of 28S rRNA target genes. The results showed that the real-time PCR assay with species-specific primer and probe was sensitive enough to detect H. cajani DNA extracted from 0.003 egg or second-stage juvenile.

Distribution of Pigeon Pea Cyst Nematode and Root-Knot Nematodes in Major Sesame Growing Areas in Myanmar

The pigeon pea cyst nematode, Heterodera cajani, is an economically important plant-parasitic nematode in sesame production. The objective of the study was to investigate distribution of H. cajani in the major sesame producing areas in Myanmar. Before cultivation, soil samples were collected in 96 local farmer’s fields: (19 fields in Magway, 20 fields in Mandalay, 14 fields in Sagaing and 43 in Nay Pyi Taw), and nematode density was quantified by real-time PCR. The cyst nematode was detected in 63.2% of the fields in Magway and the mean density was 22.6 eggs equivalent (20 g soil)−1. In Mandalay and Nay Pyi Taw, the cyst nematode was detected in 40.0% and 18.6% of the fields with the mean densities of 9.9 and 21.0 eggs equivalent (20 g soil)−1, respectively. By contrast, the cyst nematode was not detected in Sagaing fields, in which sesame is rotated with paddy rice. Infestation of root-knot nematodes (RKN, Meloidogyne spp.) was also evaluated, because sesame is generally rotated with rice and pulses, which are attacked by RKN. The results showed 25.6% infestation of RKN with a mean density of 34.3 eggs equivalent (20 g soil)−1 only in Nay Pyi Taw, while there was no infestation in Magway, Mandalay, and Sagaing. The relationships between the nematode infestation level and the cropping patterns in each sesame growing area in Myanmar were discussed.

Survey of Wilt (Fusarium udum) and the Cyst Nematode (Heterodera cajani) Incidence on Pigeonpea of Prayagraj District

A survey was conducted in selected villages of Prayagraj district to evaluate the disease incidence of Fusarium (Fusarium udum) wilt and cyst nematode (Heterodera cajani) of pigeon pea (Cajanus cajan). Soil and roots were collected from the rhizosphere of infested pigeonpea plants. The maximum wilt incidence was recorded (57.6%) in Dari village followed by Pump House (56%), Pahlu ka Purwa (51.2%) and the lowest incidence of wilt was recorded in Chaka (12.8%), Naini (11.2%) and Jhusi (9.6%) in the year 2016. For cyst nematode, soil samples was collected from selected fields, the soil was mixed thoroughly and 500 g of the soil was processed following Cobb’s sieving and decanting method where cyst and J2’s were separated in separate mesh size sieves. It was observed that infestation of Heterodera cajani was very high in Jari (51%) followed by Naini (41%). The minimum infestation of Heterodera cajani was found in Meza village which was responsible for wilt disease in pigeonpea concluding that Fusarium wilt of pigeonpea is a major problem in Prayagraj district of U.P. India.

Development of real-time PCR primers for the quantification of Meloidogyne graminicola, Hirschmanniella oryzae and Heterodera cajani, pests of the major crops in Myanmar

The rice root-knot nematode,Meloidogyne graminicola, and the rice root nematode,Hirschmanniella oryzae, are soil-borne pests causing damage to rice, and the pigeon pea cyst nematode,Heterodera cajani, is a pest to beans and sesame. Real-time PCR primers were designed for quantification. Relationships between the threshold cycle (Ct:y) values and number (no.) of nematodes inoculated () were:M. graminicola: ;H. oryzae: ; andH. cajani: .Meloidogyne graminicolaandH. oryzaewere detected in 25 and 38 out of 50 soils, collected from different fields in the lowland and central area of Myanmar, and their densities ranged from 1.0 to 4779 and from 0.4 to 787 (20 g soil)−1, respectively, whileH. cajaniwas detected only in two fields (3 and 268 (20 g soil)−1). The DNA-based method enables rapid and reliable quantification of the nematodes in soil.

In vitro nematicidal activity of a terrestrial cyanobacterium, Synechococcus nidulans, towards plant-parasitic nematodes

The nematicidal activity of a terrestrial cyanobacterium, Synechococcus nidulans, was investigated. Extracts of S. nidulans cultures collected at weekly intervals for 5 weeks were sonicated and tested against second-stage juveniles (J2) of Meloidogyne incognita. Extracts of 2-week-old cultures caused the maximum immobility (94.2%) and mortality (29.3%) of J2, compared with controls (medium and water). This extract was tested in vitro against infective stages and hatch of M. graminicola, Heterodera cajani, H. avenae and Rotylenchulus reniformis. Extracts of sonicated S. nidulans caused a mean immobility in the range of 91.3-98.4% in infective stages of the nematodes, with no significant difference with an increase in exposure time from 24 to 72 h. The greatest mean percentage mortality was observed in M. graminicola (31.5%) followed by M. incognita (29.3%), H. avenae (20.9%), and R. reniformis and H. cajani (both 17.4%) with a significant increase with the period of exposure from 24 to 72 h. No significant differences in mortality were observed between M. graminicola and M. incognita and between H. avenae and H. cajani. The percentage hatch inhibition over control (water) was greatest in M. incognita (94.2%), followed by H. avenae (91.6%), H. cajani (72.3%) and M. graminicola (70.6%), and least in R. reniformis (58.6%).