Cytochrome C Oxidase Subunit II Gene Research Articles

First Report of the Root-Knot Nematode, Meloidogyne floridensis, on Tomato in Georgia, USA.

Meloidogyne floridensis, also known as the peach root-knot nematode (RKN), is a new emerging species found to break crop host-resistance to M. incognita (Stanley et al. 2009). It was first described from Florida (Handoo et al. 2004) parasitizing M. incognita-resistant rootstock cultivars of peach (Prunus persica), and tomato (Solanum lycopersicum) (Church 2005). The nematode has recently been reported in California's almond orchards (Westphal et al. 2019) and peach rootstock (cv. Guardian) in South Carolina (Reighard et al. 2019). In a 2018 survey of vegetable fields sampled randomly in South Georgia, RKN was found with a high density (5,264 second-stage juveniles (J2)/100 cm3 of soil) from a tomato field in Ware County, GA. The soil sample consist of 30 soil cores sampled at 20-cm depth across the field in a zig-zag motion. To perform Koch's postulate, 2,000 eggs from a single egg-mass culture were inoculated into deepots filled with mixture of sand and sterilized field soil (1:1 v/v) and grown with tomato cv. Rutgers for 60 days in the greenhouse. A reproduction factor of 21.1 ± 6.1 was obtained confirming the nematode parasitism on tomato (Fig. 1S). For molecular identification, DNA was extracted by smashing three individual females isolated from the galled roots in 50 µl sterile distilled water, followed by a freeze-thaw (95°C, 1 min). Results of PCR analyzes by species-specific primers (Fjav/Rjav, Finc/Rinc and Far/Rar) did not detect the nematode species (Zijlstra et al. 2000). PCR products were obtained and sequenced from two primer sets consisting of the forward NAD5F2 (5'-TATTTTTTGTTTGAGATATATTAG-3') and the reverse NAD5R1 (5'-CGTGAATCTTGATTTTCCATTTTT-3') for amplification of a fragment of the NADH dehydrogenase subunit 5 (NADH5) gene (Janssen et al. 2016), and the forward TRANH (5'-TGAATTTTTTATTGTGATTAA-3') and the reverse MRH106 (5'-AATTTCTAAAGACTTTTCTTAGT-3') for amplification covering a portion of the cytochrome c oxidase subunit II (COII) and large subunit 16SrDNA (16S) gene (Stanton et al. 1997). DNA sequence of NADH5 gene fragment (accession no. MT795954) was 100% identical (532/532 bp) with a M. floridensis isolate from California and South Carolina (accession no. MH729181 and MN072363), while fragment of the COII and 16S genes (accession no. MT787563) was 99.76% identical (421/422 bp) with an isolate from Florida (accession no. DQ228697). The nematode females were also used for morphometric and perennial pattern analysis. Several micrographs with the inverted microscope (ZEISS Axio Vert.A1, Germany) and camera (ZEISS Axiocam 305 color, Germany) were taken from ten J2s for mean, standard deviation and range of body length: 362.7 ± 11.2 (340.4-379) µm, maximum body width: 15 ± 1.3 (12.4-16.4) µm, stylet length: 12.3 ± 1.3 (9.5-14) µm, hyaline tail terminus: 8.9 ± 1.1 (7.5-10.9) µm and tail length: 35.7 ± 4.4 (28.5-39.5) µm. Morphological measurements and configuration of perineal patterns (Fig. 2S) were comparable to previous reports of M. floridensis isolates from Florida (Handoo et al. 2004; Stanley et al. 2009). To the best of our knowledge, this is the first report of M. floridensis in Georgia as the fourth state in the USA after South Carolina, California and Florida. This nematode has been reported to parasitize several vegetable crops, including cucumber, eggplant, tomato, snap bean and squash. Furthermore, RKN resistant cultivars of tomato (harboring Mi-1 gene), pepper (harboring N gene), corn cv. Mp-710 and tobacco cv. NC 95 have been found susceptible to M. floridensis (Stanley et al. 2009), making it a serious threat.

First Report of Pythium sylvaticum Causing Corn Root Rot in Northeastern China.

As one of the most planted crops worldwide, corn has continuously increased in importance in China over the last decade. But in recent years, poor stands of corn seedlings have occurred frequently in northeastern China, causing significant economic loss. Mature plants were stunted, the roots were necrotic, and some plants collapsed. We collected soil samples from 5 fields with a history of poor stands of corn seedlings in the Heilongjiang province of China in October 2017. After being planted in the collected soil for 12 days, corn seedlings were uprooted. The pathogen was then isolated as described by Tang et al. (2019). Briefly, the rotted roots were washed in 0.5% NaOCl for 2 min, rinsed in sterile water, and then cut into 1-2 mm segments and placed on cornmeal agar amended with pimaricin (5 μg/ml), ampicillin (250 μg/ml), rifampicin (10 μg/ml), pentachloronitrobenzene (50 μg/ml), and benomyl (10 μg/ml) (PARP+B), which is selective for oomycetes (Jeffers and Martin 1986). After 3 days of incubation in the dark at 25℃, colonies were transferred to 10% V8 juice agar or potato dextrose agar (PDA) and grown for 7 days at 25℃. Based on morphological characteristics, one putative isolate (COPS) was identified as P. sylvaticum (Campbell and Hendrix 1967). On PDA, the culture (COPS) produced creamy white and floccus mycelium. P. sylvaticum (COPS) produced hyphal swellings, but no oogonia or zoospore. Hyphal swellings were globose, terminal, or intercalary, ranging from 12.22-18.55 μm diam. Sequence analysis was performed with the cytochrome c oxidase subunit Ⅱ (COⅡ) gene amplified with primers FM35/FM52 (Martin 2000) and the rDNA ITS amplified with primers DC6/ITS4 (Cooke et al. 2000). For COⅡ gene, BLAST analyses of the 773 bp segments showed 97.93% identity with P. sylvaticum isolate (GenBank Accession No. GU222164.1). For the ITS, BLAST analyses of the 880 bp segments showed 99.89% identity with P. sylvaticum isolate (GenBank Accession No. KY084736.1). Both sequences were submitted to GenBank with accession numbers MK648400 and MK606071 for COⅡ and ITS, respectively. For pathogenicity tests, similar to that described by Ling et al. (2018), four 9-cm petri plates containing 20 mL of 10% V8 juice agar were inoculated with an agar plug (5 mm diam) obtained from a 7-day-old P. sylvaticum culture (COPS) grown on 10% V8 juice agar and then incubated at 25℃ in the dark for 7 days. Nine corn seeds were placed on each plate, after which the plates were filled with 50 g sterilized organic peat substrate. For the controls, seeds were placed on non-inoculated plates of 10% V8 juice agar and filled with 50 g sterilized organic peat substrate. Four replications were inoculated. Plates were maintained in a greenhouse at 23℃. After 14 days, similar symptoms as to those observed in the field were present in the greenhouse, whereas control plants remained symptomless. P. sylvaticum (COPS) was re-isolated from diseased roots as described above, thus confirming Koch's postulates. To our knowledge, this is the first report of P. sylvaticum on corn in China. This pathogen may pose a risk to corn production. The identification of the pathogen will help to develop effective strategies to control the disease.

DIVERSITY OF Wasmannia auropunctata (HYMENOPTERA: FORMICIDAE) AND THE USE OF MITOCHONDRIAL INTERGENIC SPACER AND LEUCINE tRNA FOR ITS IDENTIFICATION

Formigas da espécie Wasmannia auropunctata (Roger, 1863) tem sido o foco de muitos estudos abordando o seu status de invasora, o seu sistema reprodutivo e o seu controle. Porém, poucos estudos têm analisado sua diversidade genética ou estudado novas técnicas para a identificação da espécie. No presente artigo foi estudada a diversidade de W. auropunctata em duas de suas regiões nativas, Brasil e Colômbia, testando novos métodos moleculares de identificação. Para tal, foi sequenciado o gene citocromo c oxidase subunidade I (COI), o espaçador intergênico (IGS), o RNA transportador (tRNA) e o gene citocromo c oxidase subunidade II (COII) de operárias de 31 localidades. Os resultados obtidos mostraram que 80% dos mitótipos identificados nunca haviam sido registrados anteriormente e que as amostras apresentaram uma alta diversidade haplotípica. A análise dos dados e comparação com diferentes estudos publicados indicam que o tRNA entre o IGS e o gene COII foi similar para todas as formigas dessa espécie, enquanto o IGS entre o gene COI e o tRNA variou em comprimento. Sendo assim, tais padrões poderiam contribuir para a identificação inter e intraespecífica, respectivamente. O presente trabalho demonstra a importância de mais estudos moleculares na área, uma vez que podem oferecer novas perspectivas sobre a diversidade da espécie, assim como métodos de identificá-la.

Mitochondrial DNA analyses reveal low genetic diversity in Culex quinquefasciatus from residential areas in Malaysia

The present study explored the intraspecific genetic diversity, dispersal patterns and phylogeographic relationships of Culex quinquefasciatus Say (Diptera: Culicidae) in Malaysia using reference data available in GenBank in order to reveal this species' phylogenetic relationships. A statistical parsimony network of 70 taxa aligned as 624 characters of the cytochrome c oxidase subunit I (COI) gene and 685 characters of the cytochrome c oxidase subunit II (COII) gene revealed three haplotypes (A1-A3) and four haplotypes (B1-B4), respectively. The concatenated sequences of both COI and COII genes with a total of 1309 characters revealed seven haplotypes (AB1-AB7). Analysis using tcs indicated that haplotype AB1 was the common ancestor and the most widespread haplotype in Malaysia. The genetic distance based on concatenated sequences of both COI and COII genes ranged from 0.00076 to 0.00229. Sequence alignment of Cx. quinquefasciatus from Malaysia and other countries revealed four haplotypes (AA1-AA4) by the COI gene and nine haplotypes (BB1-BB9) by the COII gene. Phylogenetic analyses demonstrated that Malaysian Cx. quinquefasciatus share the same genetic lineage as East African and Asian Cx. quinquefasciatus. This study has inferred the genetic lineages, dispersal patterns and hypothetical ancestral genotypes of Cx. quinquefasciatus.

Molecular systematics and phylogeography ofCebus capucinus(Cebidae, Primates) in Colombia and Costa Rica by means of the mitochondrial COII gene

We propose the first molecular systematic hypothesis for the origin and evolution of Cebus capucinus based on an analysis of 710 base pairs (bp) of the cytochrome c oxidase subunit II (COII) mitochondrial gene in 121 C. capucinus specimens sampled in the wild. The animals came from the borders of Guatemala and Belize, Costa Rica, and eight different departments of Colombia (Antioquia, Chocó, Sucre, Bolivar, Córdoba, Magdalena, Cauca, and Valle del Cauca). Three different and significant haplotype lineages were found in Colombia living sympatrically in the same departments. They all presented high levels of gene diversity but the third Colombian gene pool was determined likely to be the most ancestral lineage. The second Colombian mitochondrial (mt) haplogroup is likely the source of origin of the unique Central America mt haplogroup that was detected. Our molecular population genetics data do not agree with the existence of two well-defined subspecies in Central America (limitaneus and imitator). This Central America mt haplogroup showed significantly less genetic diversity than the Colombian mt haplogroups. All the C. capucinus analyzed showed evidence of historical population expansions. The temporal splits among these four C. capucinus lineages were related to the completion of the Panamanian land bridge as well as to climatic changes during the Quaternary Period.

Trypanosoma cruzi TcIII / Z3 genotype as agent of an outbreak of Chagas disease in the Brazilian Western Amazonia

Chagas' disease is an emerging and neglected disease in the Brazilian Amazon region, where T. cruzi I predominates among the acute cases of the disease; and T. cruzi III/Z3, a population cluster from sylvatic areas of the Amazon basin, is rarely associated with human infections. On 23rd April 2007, the Foundation for Health Surveillance of the State of Amazonas, Brazil reported an outbreak of acute Chagas disease in the municipality of Coari on the Solimões River banks. Fresh blood examination confirmed the infection in 25 patients. Parasite culture in LIT medium was successful for 18 isolates. Molecular characterization was performed by PCR of the non-transcribed spacer of the mini-exon and by sequencing of the mitochondrial cytochrome c oxidase subunit II (COII) gene. The T. cruzi isolates were all from genotype Z3, and sequencing revealed that all isolates had equal COII sequences compatible with TcIII type, suggesting a single source of infection. To our knowledge, this is the first outbreak of acute cases caused uniquely by the genotype TcIII/Z3. Wild vectors harbouring TcIII stocks contribute to transmission when the triatomine species reaches human food chain or when humans invade the forest environment, where sylvatic cycle constitutes a reservoir of parasites that might be associated with specific epidemiological and clinical traits of the emergent Chagas disease in the Amazon.

Molecular Phylogeny of the Subfamily Gerbillinae (Muridae, Rodentia) with Emphasis on Species Living in the Xinjiang-Uygur Autonomous Region of China and Based on the Mitochondrial Cytochromeband CytochromecOxidase Subunit II Genes

Rodents belonging to the subfamily Gerbillinae and living in the Xinjiang-Uygur autonomous region of China were collected in field surveys between 2001 and 2003. We found four Meriones species, including M. chengi M. liycus, M. meridianus, and M. tamariscinus, as well as related species from different genera, Rhombomys opimus and Brachiones przewaliskii For phylogenetic analyses of these gerbilline species, DNA sequences of parts of the mitochondrial cytochrome b (Cytb) and cytochrome c oxidase subunit II (COII) genes were examined with the neighbor Joining, maximum parsimony, maximum likelihood, and Bayesian inference methods. Our phylogenetic analyses suggest that the genus Meriones is not monophyletic and place M. tamaricinus as the sister taxon to a clade comprising Brachiones, Psammomys, Rhombomys, and the other Meriones species. The remaining Meriones species separate into three lineages: M. meridianus (including M. chengi), Meriones unguiculatus, and a clade that includes multiple Meriones species originating from Asia, the Middle East, and Africa. The phylogenetic relationships among the genera Brachines, Meriones, Psammomys, and Rhombomys remain ambiguous, probably due to the saturation of mutations that occurs in fast-evolving mitochondrial DNA. In addition, intraspecific variation was observed for M. meridianus, and this mostly correlated with collection localities, i.e., the northern and southern parts of the Xinjiang region. This variation corresponded to interspecific levels of divergence among other lineages of Meriones. Interestingly, no differences were observed in either the Cytb or COII gene sequences isolated from M. chengi collected from the Turfan Basin in the north and those from M. meridianus in the south, suggesting that M. chengi may be a synonym of M. meridianus.

Phylogenetic Relationships of South China Sea Snappers (Genus Lutjanus; Family Lutjanidae) Based on Mitochondrial DNA Sequences

Phylogenetic relationships of intra- and interspecies were elucidated based on complete cytochrome b (cyt b) and cytochrome c oxidase subunit II (COII) gene sequences from 12 recognized species of genus Lutjanus Bloch in the South China Sea (SCS). Using the combined data set of consensus cyt b and COII gene sequences, interspecific relationships for all 12 recognized species in SCS were consistent with Allen's morphology-based identifications, with strong correlation between the molecular and morphological characteristics. Monophyly of eight species (L. malabaricus, L. russellii, L. stellatus, L. bohar, L. johnii, L. sebae, L. fulvus, and L. fulviflamma) was strongly supported; however, the pairs L. vitta/L. ophuysenii and L. erythropterus/L. argentimaculatus were more similar than expected We inferred that L. malabaricus exists in SCS, and the introgression caused by hybridization is the reason for the unexpectedly high homogeneity.

Evolution of eutherian cytochrome c oxidase subunit II: heterogeneous rates of protein evolution and altered interaction with cytochrome c.

Cytochrome c oxidase subunit II (COII), encoded by the mitochondrial genome, exhibits one of the most heterogeneous rates of amino acid replacement among placental mammals. Moreover, it has been demonstrated that cytochrome c oxidase has undergone a structural change in higher primates which has altered its physical interaction with cytochrome c. We collected a large data set of COII sequences from several orders of mammals with emphasis on primates, rodents, and artiodactyls. Using phylogenetic hypotheses based on data independent of the COII gene, we demonstrated that an increased number of amino acid replacements are concentrated among higher primates. Incorporating approximate divergence dates derived from the fossil record, we find that most of the change occurred independently along the New World monkey lineage and in a rapid burst before apes and Old World monkeys diverged. There is some evidence that Old World monkeys have undergone a faster rate of nonsynonymous substitution than have apes. Rates of substitution at four-fold degenerate sites in primates are relatively homogeneous, indicating that the rate heterogeneity is restricted to nondegenerate sites. Excluding the rate acceleration mentioned above, primates, rodents, and artiodactyls have remarkably similar nonsynonymous replacement rates. A different pattern is observed for transversions at four-fold degenerate sites, for which rodents exhibit a higher rate of replacement than do primates and artiodactyls. Finally, we hypothesize specific amino acid replacements which may account for much of the structural difference in cytochrome c oxidase between higher primates and other mammals.

Mitochondrial Gene Sequences and the Molecular Systematics of the Artiodactyl Subfamily Bovinae

Nucleotide sequence evolution of the mitochondrial cytochrome c oxidase subunit II (COII) gene was used to examine the molecular phylogenetics and evolution of the Bovinae, a subfamily within the mammalian order Artiodactyla. The COII gene was sequenced in representatives of three bovine tribes (Bovini, Boselaphini, and Tragelaphini) and the outgroup taxonCapra(subfamily Caprinae). Although the phylogenetic analyses groupedBisonas sister toBos,the genusBisonwas paraphyletic, with the American bison being most closely related to species ofBosrather than to the European bison. COII data also supported a close relationship between African (Syncerus) and Asian (Bubalus) buffaloes, the monophyly of the tribe Bovini, and a sister-group relationship between the tribes Bovini and Boselaphini. Analysis of nucleotide substitutions in the COII gene prompted a system of differential weighting of nucleotide substitutions for inferring phylogenetic relationships across the range of divergence times examined here (2–20 million years). Rates of evolution in the COII gene are examined and compared to evolutionary rates in mtDNA tRNA/rRNA genes and the D-loop among other artiodactyl taxa.

Gene for cytochrome c oxidase subunit II in the mitochondrial DNA of Culex quinquefasciatus and Aedes aegypti (Diptera: Culicidae).

The cytochrome c oxidase subunit II (COII) gene between transfer RNA for Leu and Lys in the mitochondrial DNA of Culex quinquefasciatus Say and Aedes aegypti (L.) was amplified by the polymerase chain reaction (PCR) technique. Both the gene order and direction of transcription were identical to Anopheles and Drosophila. Nucleotide sequences of the PCR-amplified COII genes in these two mosquitoes exhibited 88% homology, and the frequency of transition was very close to that of transversion. The homology of deduced amino acid sequences of COII between these two mosquitoes was 95%. Two highly conserved segments of COII proteins were found in mosquitoes, fruit flies, locust, and honeybee. These segments contain the major amino acid residues of cytochrome c oxidase involved in electron transport and ligand binding. The amino acid residues are located at the positions similar to those of the mammalian enzymes. Two sets of the phylogenetic trees of a similar pattern were generated by comparing the divergences of nucleotide and amino acid sequences of COII. The branch lengths of the trees estimated by amino acid and nucleotide sequences showed different evolution rates of Aedes and Culex from their common ancestor.

Nucleotide sequence of a segment of Drosophila mitochondrial DNA that contains the genes for cytochrome c oxidase subunits II and III and ATPase subunit 6.

The nucleotide sequence of a segment of the mtDNA molecule of Drosophila yakuba has been determined, within which have been identified the genes for tRNAleuUUR, cytochrome c oxidase subunit II (COII), tRNAlys, tRNAasp, URFA6L, ATPase subunit 6 (ATPase6), cytochrome c oxidase subunit III (COIII) and tRNAgly. The genes are arranged in the order given and all are transcribed from the same strand of the molecule in a direction opposite to that in which replication proceeds around the molecule. The tRNAlys gene is unusual among mitochondrial tRNAlys genes in that it contains a CTT anticodon. The triplet AGA is used to specify an amino acid in all of the COII, COIII, ATPase6, and URFA6L genes. However, the AGA codons found in these four polypeptide genes correspond in position to codons which specify nine different amino acids, but never arginine, in the equivalent polypeptide gene which have been sequenced from mtDNAs of mouse, yeast and Zea mays.