lrRNA Genes Research Articles

Complete mitochondrial genome of the earthworm Amynthas seungpanensis (Clitellata: Megascolecidae)

Amynthas seungpanensis Song and Paik, 1970 (Clitellata: Megascolecidae) is an endemic and ecologically important species in Korea; however, knowledge regarding its genetic characteristics is lacking. In this study, we sequenced the complete mitochondrial genome (mitogenome) of A. seungpanensis. The mitogenome is 15,085-bp long and contains 13 protein-coding genes (PCGs), 2 rRNA genes, 22 tRNA genes, and 1 major non-coding region, the control region. This mitogenome has an arrangement identical to that observed in the mitogenomes of most earthworms, and all 37 genes are transcribed from the same directional strand. All 13 PCGs have the same start codon (ATG). Six PCGs (COI, COII, ND6, CytB, ATP6, and ND4) end with TAA and ND4 ends with TAG, whereas the remaining genes exhibit an incomplete stop codon, T. The A/T content of the complete mitogenome is 65.8%; however, it varies among regions and genes as follows: control region, 77.8%; srRNA genes, 65.8%; lrRNA genes, 68.8%; tRNA genes, 64.2%; and PCGs, 65.0%. Results of phylogenetic analyses suggested the family Megascolecidae to be a monophyletic group with the highest nodal support, whereas the genus Amynthas, which belongs to Megascolecidae, was determined to be non-monophyletic.

First report of Meloidogyne arenaria on roots of Grona triflora in Guangdong Province, China.

Grona triflora (Desmodium triflorum), a perennial herbaceous legume, is widely distributed in southern China. G. triflora has antipyretic, antiseptic and expectorant properties and can therefore be used as a phytomedicine (Ghosal et al. 1973). In July 2020, roots of G. triflora were investigated for nodules and rhizobia collection at the Shibaluohan Mountain Forest Park of Guangzhou. Root galls induced by a root-knot nematode were observed on 90% of the G. triflora samples (in a 200 m2 plot) and the infested plants had yellow, small and withered leaves compared with the healthy ones. The galls number on a G. triflora root ranged from 43 to 92 and the population densities of second stage juveniles (J2s) ranged from 573 to 894 per 100 cm3 soil surrounding the plant. The female perineal patterns showed a low dorsal arch, with lateral field marked by forked and broken striae, no punctate markings between the anus and tail terminus, which matched with the description of Meloidogyne arenaria (Hartman and Sasser 1985). The J2s had the following morphometric characters (n = 15): body length = 501.05 ± 23.71 µm; body width = 17.14 ± 1.23 µm; DGO = 3.13 ± 0.27 µm; stylet length = 12.97 ± 1.38 µm; tail length = 58.02 ± 4.77 µm; hyaline tail terminus = 10.08 ± 0.65 µm. DNA from four female nematodes was isolated for PCR-based diagnostic analyses. A fragment between the COII and LrRNA genes of the mitochondrial DNA was amplified with primers C2F3/1108 (Powers and Harris 1993). In addition, a 28S ribosomal DNA D2/D3 region was amplified with primers MF/MR (Hu et al. 2011). The amplicons were sequenced (GenBank No. MW315989 and MW307358). Nucleotide BLAST results indicated that both sequences show 100% identity with corresponding M. arenaria sequences of isolates from various countries such as Brazil, China, Myanmar and Vietnam (e.g., MK033428, JQ446377, KY293688 and MK026624). For further confirmation, sequence characterized amplified region (SCAR) PCR was employed using the M. arenaria specific primers Far/Rar (Zijlstra et al. 2000). The amplicon was also sequenced (GenBank No. MW315990). The Nucleotide BLAST results showed >99% identity with M. arenaria isolates from Indonesia and Argentina (KP234264, KP253748 and MK015624). Greenhouse tests were conducted to analyze the capacity of M. arenaria to induce galls on G. triflora roots. The G. triflora seeds were collected from the sampling plot and germinated on 0.8% (W/V) agar plates. Then the seedlings were planted in 14 cm deep and 15 cm diam pots filled with sterilized soil from sampling plot. Every seedling was inoculated with 2,000 J2s (n = 15) and plants without J2s were used as a control. Two months later, galls were observed for inoculated roots while no galls were formed on roots of control plants. An average of 13,300 J2s and eggs of M. arenaria (reproduction factor = 6.65) were recovered from the root. Stanton and Rizo (1988) found that G. triflora was susceptible to M. javanica in Australia, and Ogbuji (1978) reported that a population of M. incognita reproduced on roots of G. triflora in Nigeria after artificial inoculation. To our knowledge, this is the first report on G. triflora parasitized by M. arenaria in Guangdong province. M. arenaria has potential to infest local, economically important plants like citrus, pomelo, sugarcane, maize and peanut. As G. triflora is widely distributed in southern China, there is the risk of spreading M. arenaria into agricultural and horticultural systems, that will cause yield loss and economic impacts.

The first mitochondrial genome of Capitulum mitella (Crustacea: Cirripedia) from China: revealed the phylogenetic relationship within Thoracica

Capitulum mitella (Crustacea: Cirripedia) is an important stalked barnacle. The first mitochondrial genome of C. mitella from China was presented, which is a circular molecule of 15,930 bp in length and AT content is 64.4%. It encodes 37 genes, including 13 PCGs, 22 tRNAs, and two rRNAs, which is consistent with most barnacles species reported. There are 15 genes encoded on the light strand and 22 genes encoded on the heavy strand. Identical to most barnacles species reported, srRNA and lrRNA genes are adjacent and separated only by trnV gene. Phylogenetic trees showed that C. mitella clustered with Pollicipes polymerus, indicating Pollicipedidae is monophyletic. However, Scalpelliformes was not monophyletic from the phylogenetic tree. From the level of order, the Lepadiformes was located at the base of the phylogenetic tree, indicating that its divergence time was earlier than Scalpelliformes. The results provided more insights into phylogenetic consideration at the genomic level within superorder Thoracica.

The complete mitochondrial genome of Conwentzia sinica (Neuroptera: Coniopterygidae)

The complete mitochondrial genome (mitogenome) of Conwentzia sinica Yang, 1974 was sequenced and analyzed. The sequenced mitogenome is 15,153 bp in size, including 13 protein-coding genes (PCGs), 22 tRNA genes, 2 rRNA genes, and one control region. Thirteen PCGs use ATN as the start codon. Most of PCGs terminate with TAA codons but ND5, ND4 with a single T residue, and ND3 terminates with TAG. The lrRNA gene is 1290 bp. The measured length of srRNA gene is 782 bp. Twenty-two tRNA genes possess the typical clover-leaf secondary structure except for tRNASer(AGN). The phylogenetic result supports the monophyly of the family Coniopterygidae and a closer relationship between Conwentzia and Coniopteryx.

Complete mitochondrial genome sequence of Acoptolabrus changeonleei Ishikawa et Kim, 1983 (Coleoptera: Carabidae)

Acoptolabrus changeonleei Ishikawa et Kim, 1983 (Coleoptera: Carabidae) has been listed as an endangered insect in South Korea. The complete mitochondrial genome (mitogenome) of the species was 16,831-bp long with a typical set of genes (13 protein-coding genes [PCGs], 2 rRNA genes, and 22 tRNA genes) and one non-coding region, with the arrangement identical to that observed in most insect genomes. Twelve PCGs had the typical ATN start codon, whereas ND1 had the atypical TTG codon. The 1,712-bp long A+T-rich region was composed of 88.6% A/T nucleotides and had no long repeat sequences. Phylogenetic analyses with concatenated sequences of the 13 PCGs and the lrRNA gene placed A. changeonleei as a sister to the within-subfamilial species Damaster mirabilissimus in Carabinae, with the highest nodal support by both analyses.

First Report of the Root-Knot Nematode Meloidogyne arenaria Infecting Noni in China.

Noni (Morinda citrifolia) is an important medicinal plant and its fruit and root are of high value. In recent years, noni has been cultivated widely on Hainan Island, China. A survey of eight commercial noni fields in four counties found that most fields had plants with symptoms consistent with damage caused by root-knot nematodes. Above-ground symptoms included reduced vigor, plant stunting, and chlorosis. Affected roots were galled, swollen, cracked, and rotten. Fruit loss associated with diseased plants was quantified as 85%. In each field, three samples were taken consisting of 15 cm wide × 20 cm deep soil (containing roots). The nematode population was extracted and quantified according to Barker (1). Nematodes were found in all soil and root samples with population densities ranging from 450 to 835 eggs and second-stage juveniles (J2s) per 200 g subsample of soil, and 685 to 985 eggs and J2s per 10 g sub-sample of fresh roots. Three single egg masses were respectively hand-picked from one sample of diseased noni roots and inoculated onto tomato plants grown with sterilized soil at 20 to 28°C in the greenhouse. After 8 weeks, nematodes were extracted from the roots of tomato plants and identified by morphology, enzyme analysis, and molecular characterization. Morphology of the female perineal patterns showed a low dorsal arch with large lateral lines separating the striae of the dorsal and ventral sectors, leading to the tail terminus; and wavy, coarse striae with forking at lateral lines and short irregular striae near the lateral lines. Enzyme analysis of the esterase phenotype was also typical of the A1 phenotype in M. arenaria. Based on the perineal pattern and esterase phenotype, the Meloidogyne species was identified as M. arenaria (Est A1) (3). Total genomic DNA was extracted from ca. 10 μl of packed second-stage juveniles (J2s) using the method of Cenis (2). The primers C2F3 (5'-GGTCAATGTTCAGAAATTTGTGG-3') and 1108 (5'-TACCTTTGACCAATCACGCT-3') (4) was used to amplify the intergenic region between COII and LrRNA genes of the mtDNA and an amplification product (1,700 bp) was obtained, similar to M. hispanica, M. incognita, and M. javanica. Root-knot nematodes (Meloidogyne spp.) have been reported to be cause disease on noni in Hawaii. However, to our knowledge, this is the first report of M. arenaria (Est A1) infecting noni in China.

Genetic Differentiation of the Giant Honey Bee (Apis dorsata) in Thailand Analyzed by Mitochondrial Genes and Microsatellites

Genetic diversity and population differentiation of the giant honey bee (Apis dorsata) in Thailand were examined. Six PCR-RFLP mitotypes were generated from digestion of the COI-COII, Cytb-tRNA(ser), ATPase6-8, and lrRNA genes with Dra I and Hin fI. Low genetic diversity (h=0.074, pi=0.032%) and a lack of genetic population differentiation between A. dorsata originating from geographically different regions were observed from mtDNA polymorphisms (P > 0.05). In contrast, microsatellite (A14, A24, and A88) polymorphisms revealed a relatively high level of genetic diversity in A. dorsata (H (o)=0.68-0.74, average number of alleles per locus=6.0-9.0). Both A24 and A88 indicated significant population differentiation between bees from the north-to-central region (north, northeast, and central regions), peninsular Thailand, and Samui Island.

Mitochondrial genomes of two luminous beetles, Rhagophthalmus lufengensis and R. ohbai (Arthropoda, Insecta, Coleoptera)

We determined the mitochondrial DNA (mtDNA) sequences of two luminous beetles (Arthropoda, Insecta, Coleoptera), Rhagophthalmus lufengensis from Yunnan, China and Rhagophthalmus ohbai from Yaeyama Island, Japan. We identified all the 37 mtDNA genes of R. lufengensis (15,982 bp) and the 34 genes of R. ohbai (15,704 bp). R. lufengensis and R. ohbai genomes have higher A + T contents than other coleopteran genomes although the gene arrangements are similar. Interestingly, in a study of the evolutionary relationship among R. lufengensis, R. ohbai and the firefly Pyrocoelia rufa, the phylogenetic tree inferred from lrRNA genes from mitochondrial genomes indicates a biogeographic relationship among the bioluminescent insects in East Asia and the phylogenetic tree inferred from luciferase-related genes from nuclear genomes shows an appropriate relationship among coleopterans, reflecting the evolutionary origin of bioluminescence. Thus, the mtDNAs of luminescent beetles can provide an insight into their evolutionary origin and biogeography.

Characterisation of Meloidogyne species from China using Isozyme Phenotypes and Amplified Mitochondrial DNA Restriction Fragment Length Polymorphism

Forty-six Meloidogyne populations from 14 provinces of China were characterised in terms of malate dehydrogenase, esterase phenotypes and HinfI restriction profiles of amplification products from the mitochondrial DNA (mtDNA) region between the COII and lrRNA genes. Isozyme phenotyping revealed that 29 of the populations were M. incognita, six were M. javanica, six were M. arenaria, three were M. hapla and two were M. enterolobii. HinfI restriction patterns of the COII-lrRNA region correlated with nematode isozyme phenotypes and enabled reliable differentiation and identification of the five root-knot nematodes occurring in China. The size and sequence of the mtDNA amplification product were determined for the first time for M. enteroloii, a potentially economically important crop pathogen. Sequence comparison showed that the sequence of the intergenic region between the COII and lrRNA genes for M. enterolobii was identical to that reported for M. mayaguensis. Together with published observations on morphology, host range and esterase phenotype of the two nominal species, the mtDNA sequence evidence suggests that M. mayaguensis could be conspecific with M. enterolobii.

Genetic differentiation of the honey bee (Apis cerana) in Thailand revealed by polymorphism of a large subunit of mitochondrial ribosomal DNA

The large subunit of the ribosomal (lr) RNA gene of Thai Apis cerana was amplified and sequenced. The average sequence divergence among geographically diverse bees was 1.13% (0.15% - 1.70%, n = 8). Based on these nucleotide sequences, genetic differentiation analysis of A. cerana was simplified to a RFLP based on Dra I. Four haplotypes of digested lrRNA gene were observed. Haplotype A was found in north, north-east and the central region (collectively called the north-to-central sample) whereas haplotype B was restricted to specimens from peninsular Thailand, Phuket and Samui Islands (collectively called the southern sample). Haplotype C was found in 47.06% of A. cerana originating from Samui Island but was not found in other geographic samples. Geographic heterogeneity analysis and FST statistics indicated the existence of population differentiation of A. cerana in Thailand (P < 0.0001). Analysis of molecular variance (AMOVA) also illustrated significant genetic differences between bees from the north-to-central region (A), peninsular Thailand and Phuket (B) and Samui Island (C) (P < 0.0001).

Mitochondrial DNA diversity and population structure of the honey bee, Apis cerana, in Thailand

SUMMARYGenetic variation and population differentiation of Apis cerana in Thailand were examined by restriction analysis of amplified mitochondrial srRNA and lrRNA genes, and the intergenic CO1-CO2 region with DraI. Twelve composite haplotypes were found from 170 colonies investigated. Large genetic differences were observed among A. cerana populations from northern and peninsular Thailand. The population from Samui Island was distinct, whereas A. cerana from Phuket Island was genetically similar to that from peninsular Thailand. Geographic heterogeneity analysis and FST estimates indicated a high level of population subdivision in this species (P < 0.0001and FST = 0.604). Using percent sequence divergence as a measure of genetic distance, three distinct groups were found using the clustering algorithm UPGMA. These were: north Thailand (A), peninsular Thailand, and Phuket (B) and Samui Islands (C).

A phylogenetic study of the squid family Onychoteuthidae (Cephalopoda: Oegopsida)

The oegopsid squid family Onychoteuthidae presently comprises six genera (Moroteuthis, Onychoteuthis, Ancistroteuthis, Kondakovia, Onykia and Chaunoteuthis) but the status of some of these is still uncertain. An interdisciplinary study was undertaken to clarify the phylogenetic relationships, which included a morphological approach (morphometric analysis), and a molecular study of a mitochondrial gene portion (l-rRNA gene, 16S). The morphometric analysis identified two groups, one including some Onychoteuthis and some Moroteuthis knipovitchi, and another including the remaining genera or species. At the intrageneric level, M. knipovitchi appears to be separated from the other species of the genus; M. robusta is closely related to M. ingens and M. pacifica. Morphometric analysis confirmed that Kondakovia longimana is different from M. ingens. Likewise, Onychoteuthis was clearly separated from the other genera, but there is neither geographical grouping nor morphometric differentiation of the Onychoteuthis species with the parameters measured. Some specimens were apparently intermediate between the Onychoteuthis and Ancistroteuthis groups. On a molecular basis, the Onychoteuthidae appeared to be monophyletic. Monophyly of the genus Moroteuthis (four species studied) was not strongly supported: M. knipovitchi was distinct from the others. The molecular analysis showed three Hawaiian species, Onychoteuthis compacta, O. sp. B and O. sp. C, to be closely related. The gene sequence for the newly created species of Onykia is clearly different from all the others, indicating that it is a true species.

Incipient mitochondrial evolution in yeasts: I. The physical map and gene order of Saccharomyces douglasii mitochondrial DNA discloses a translocation of a segment of 15,000 base-pairs and the presence of new introns in comparison with Saccharomyces cerevisiae

We have determined the physical and genetic map of the 73,000 base-pair mitochondrial genome of a novel yeast species Saccharomyces douglasii. Most of the protein and RNA-coding genes known to be present in the mitochondrial DNA of Saccharomyces cerevisiae have been identified and located on the S. douglasii mitochondrial genome. The nuclear genomes of the two species are thought to have diverged some 50 to 80 million years ago and their nucleo-mitochondrial hybrids are viable but respiratorily deficient. The mitochondrial genome of S. douglasii displays many interesting features in comparison with that of S. cerevisiae. The three mosaic genes present in both genomes are quite different with regard to their structure. The S. douglasii COXI gene has two new introns and is missing the five introns of the S. cerevisiae gene. The S. douglasii cytochrome b gene has one new intron and lacks two introns of the S. cerevisiae gene. Finally, the L-rRNA gene of S. douglasii, like that of S. cerevisiae, has one intron of which the structure is different. Another salient feature of the S. douglasii mitochondrial genome reported here is that the gene order is different in comparison with S. cerevisiae mitochondrial DNA. In particular, a segment of approximately 15,000 base-pairs including the genes coding for COXIII and S-rRNA has been translocated to a position between the genes coding for varl and L-rRNA.