Molecular characterization and genetic variation of root - knot nematode (Meloidogyne spp.,) in selected legume production areas of Eastern Kenya
Kavuluko, Jacinta Muthini
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Doot-knot nematodes (RKN) are sedentary endoparasites of plant roots and the primary ~ematodepathogens of most crop species worldwide, including many cultivatedlegumes. RKN of theMeloidogynespecies have a wide host range that includes at least1,700 plant species. Most previous studies on the diversity of Meloidogynespp. Havefocused on morphology (for example: perineal patterns, styletStructure, body length) andthe response of the populations to differential host test. Morphological differences mayhowever be absent or difficult to observe thus making identification difficult for the nonspecialist,but distinguishing them is important for utilizing appropriate crop rotations,managing resistance effectively and for plant quarantine requirements. Cowpea (VignaunguiculataL. Walp) is a food and fodder legume of significant economic importanceespecially in semi-arid regions of Africa. Root- Knot Nematodes (RKN) of the genusMeloidogyneis the most economically important nematode pests affecting cowpea production and has a diverse host range. Pigeon pea (Cajanuscajan(L) Millsp.) is animportant grain legume crop of rain-fed agriculture in the semi-arid tropics. Pigeonpea is produced as a vegetable or export grain crop in southern and eastern Africa. Forthis study; esterases (EST) and malate dehydrogenase (Mdh) were used to characterizedifferent species of Meloidogyneaffecting selected legume production in Mbeeredistrict.Esterase activity was the most useful in the identification of the different species. Thespecies identity of ninety RKNs affecting cowpeas and pigeon peas in Mbeere wasrevealed using isozymeanalysis. The two species, Meloidogynejavanica, and M arenaria.' ishableisozymephenotypes. Meloidogynejavanicahad the phenotype N 1-13ha~ dlsutmguaria had NI-A2. Species specific phenotype NI-13 was detected in 64.4%while sa.marelens from Gachoka, Mwea and Siakago while phenotype NI-A2 was found inof the P .ifi d d 1 f id ifyi601 The esterase phenotypes are species-speerc an are a goo too or I enting3R5K.N/s0.. small subunit Ribosomal DNA (SSUrDNA) has been a popular target, because. . hi'ghly conserved sequences are interspersed with less conserved regions, enablingIthISlogenetic studies at various taxonomic levels. PCR amplifications of the extracted pu~fied DNA were carried out using primers specific for 5S and 18S rDNAcistrons. The~bosomal primers successfully amplified fragments of the expected size from extracted DNA. The size of the PCR products obtained following amplification of the intergenicspacer region between the 5S and 18S ribosomal genes was about 720 bp. Purified PCRproducts were sequenced and thirteen 5S-18S rDNA sequences obtained. The sequenceswere aligned using CLUSTALW2, Sequence statistics, pairwise differences, and estimatesof divergence were determined with MEGA5. Nucleotide diversities were estimated inDnaSPv5. Phylogenetic tree was drawn using Phylowinand edited in MEGA5. Fromthe findings of the study it has been established that root knot nematodes affecting thecowpea and pigeon pea in Mbeeredistrict are M javanica, M incognita and M arenaria. Sequences have not evolved with the same pattern of substitution, as judged from the extent of differences in base composition biases between sequences. Sequences from the species under study were closely related to sequences retrieved from sequences databases especially those sequences which were less divergent due to less substitutions, deletions and insertions. It can be concluded that SSUrDNAare useful in identification, inferring genetic diversity and phylogenetic relationships between the is~lated root knot nematodes.