Development of the new microsatellite markers of Lucilia sericata (Diptera: Calliphoridae) from Korea.

BACKGROUND: Lucilia sericata is a medical and veterinary important insect species because its larvae feed on tissues of vertebrates including humans. Very few microsatellite makers have been reported from the species to illuminate its genetic variability and population genetic structure. METHODS AND RESULTS: In this study, L. sericata samples were collected from four different localities in Korea to develop the microsatellite markers to provide basic information on the genetic variability and population genetic structure in Korea of this species. In total, ten new microsatellite markers were sequenced and analyzed. Genetic diversity was performed using these microsatellite markers. The observed heterozygosity varied from 0.205 to 0.824, with an average of 0.546. The expected heterozygosity ranged from 0.579 to 0.886, with an average of 0.804. PIC value varied from 0.553 to 0.876. CONCLUSIONS: The markers developed in the present study are expected as informative for estimating genetic diversity of L. sericata.

Development of polymorphic microsatellite markers for the Trichoglossus haematodus and cross-species amplification in Trichoglossus moluccanus.

BACKGROUD: Trichoglossus haematodus is the most popular parrots globally and one of the most bred species in Korea's zoos. However, despite its popularity, there are limited studies on the population genetics of this species. METHODS AND RESULTS: In this study, 10 polymorphic microsatellite markers were developed for T. haematodus. The number of alleles ranged from 6 to 9 (mean 7.9). Null alleles were present in two loci (TH-07 and TH-08). The observed heterozygosity ranged from 0.4444 to 1.0000 (mean 0.7000). One locus (TH-08) indicated a significant deviation from the Hardy-Weinberg equilibrium after Bonferroni correction (p < 0.005). The mean inbreeding coefficient (FIS) of the 10 loci was positive, suggesting that there is inbreeding in the population. Since the polymorphism information content (PIC) values were more than 0.7 in all loci, all markers developed in this study were classified as informative. The parentage exclusion probabilities considering all loci were higher than 0.99 in all three cases (P1, P2, and P3). The cross-species amplification of the 10 markers was tested in T. moluccanus, a close relative species of T. haematodus. These markers were also informative for T. moluccanus with PIC values higher than 0.7 in all loci. Additionally, the parentage exclusion probabilities (P1, P2 and P3) for T. moluccanus were above 0.99. However, due to the small number of T. haematodus and T. moluccanus investigated in this study, the 10 microsatellite markers should be analyzed with more individuals of these two species in future studies. CONCLUSIONS: The markers developed in this study might be helpful for investigations of genetic diversity and parentage analysis of T. haematodus and T. moluccanus.

An Investigation of the Variations in Complete Mitochondrial Genomes of Lingula anatina in the Western Pacific Region.

Lingula anatina is a brachiopod widely distributed in the western Pacific region. Even though L. anatina has been targeted for a number of biological studies, there is still limited information on intraspecific genetic variations of L. anatina. In this study, L. anatina specimens were collected from Korea and Vietnam, and complete mitochondrial genome (mitogenome) sequences were analyzed and compared with previous records. The total mitogenomes of L. anatina were 24,875 bp and 25,305 bp in size for Korean and Vietnamese specimens, respectively. Those mitogenomes are extraordinarily longer than the typical mitogenome size for an animal but shorter than the previous record from Yanagawa (Japan) for this species. The gene orders and the sizes of the protein-coding genes are also different from those for the Japanese specimen. Furthermore, the nonsynonymous (Ka) and synonymous (Ks) substitution rates in protein-coding genes (PCGs) were calculated to test the idea of evolutionary rate differences in mitochondrial genomes. The analyses showed relatively low Ka and Ks for the complete mitogenomes from Buan (Korea), Doson (Vietnam) and Yanagawa (Japan). The Ka/Ks ratio was less than 1 in comparisons of three localities, indicating the existence of purifying selection in this species. The phylogenetic analyses showed that L. anatina diverged among localities in the western Pacific region.

De Novo Development of mtDNA Deletion Due to Decreased POLG and SSBP1 Expression in Humans.

Defects in the mitochondrial genome (mitochondrial DNA (mtDNA)) are associated with both congenital and acquired disorders in humans. Nuclear-encoded DNA polymerase subunit gamma (POLG) plays an important role in mtDNA replication, and proofreading and mutations in POLG have been linked with increased mtDNA deletions. SSBP1 is also a crucial gene for mtDNA replication. Here, we describe a patient diagnosed with Pearson syndrome with large mtDNA deletions that were not detected in the somatic cells of the mother. Exome sequencing was used to evaluate the nuclear factors associated with the patient and his family, which revealed a paternal POLG mutation (c.868C > T) and a maternal SSBP1 mutation (c.320G > A). The patient showed lower POLG and SSBP1 expression than his healthy brothers and the general population of a similar age. Notably, c.868C in the wild-type allele was highly methylated in the patient compared to the same site in both his healthy brothers. These results suggest that the co- deficient expression of POLG and SSBP1 genes could contribute to the development of mtDNA deletion.

Molecular Identification of Necrophagous Dermestes Species in South Korea Using Cytochrome c Oxidase Subunit I Nucleotide Sequences (Genus Dermestes).

Species identification of necrophagous insects found on a dead body is an essential key in applying medicolegal entomology to the estimation of postmortem interval (PMI). Due to limited morphological identification of insect evidence, several studies have identified species using molecular information such as DNA markers. While considerable cytochrome c oxidase subunit I (COI) gene sequence data of necrophagous fly species have been collected and annotated, those of necrophagous beetle species have not. Since necrophagous beetles such as Dermestes species have a larval period longer than that of flies, beetles are useful in even the late decomposition phase in estimating minimum PMI. To obtain the full-length COI gene sequences of six Dermestes species collected from South Korea, we designed primers for polymerase chain reaction amplification and sequencing. The obtained full COI nucleotide sequences were used for performing phylogenic analysis and comparison with previously reported sequences. The results demonstrated that the COI gene sequences could be used to identify forensically important Dermestes species in South Korea.

Comparative analyses of the three complete mitochondrial genomes from forensic important beetle genus Dermestes with phylogenetic relationships.

Necrophagous Dermestes species have high forensic importance in relation to the estimation of elapsed time since death or death season. To further supplement the genome-level features for related species, the complete mitochondrial genome (mitogenome) of Dermestes species D. essellatocollis, D. frischii and D. coarctatus are amplified, sequenced, annotated, analyzed, and compared with other twelve species of the infraorder Bostrichoidea. The mitochondrial genomes were typical circular molecules with 16,218, 15,873 and 15,873 bp in length, respectively. They included 13 protein coding genes, two rRNAs, and 22 tRNAs, as well as the putative control region. The gene orders and orientations are identical to those of other recorded bostrichiformian species and had the ancestral insect gene composition. Furthermore, phylogenetic analyses based on all the mitochondrial protein coding genes for 13 Bostrichoidea and 16 outgroup taxa were performed using Bayesian and Maximum Likelihood analyses. The inferred trees indicate that the genus Dermestes is monophyletic. The monophyly of infraorder Bostrichiformia is not supported. This study provides genomic data for mitochondrial genome library of the genus Dermestes to investigate evolutionary and systematic studies.

Characterization of the complete mitochondrial genome of Triceratopyga calliphoroides (Rohdendorf, 1931) (Insecta: Diptera: Calliphoridae).

Triceratopyga calliphoroides is a blowfly species which is the only member of the genus Triceratopyga. Because of their forensic importance, we sequenced the complete mitogenome of the T. calliphoroides and analyzed phylogenetic relationships. According to data, it has the longest mitogenome in the family with 16,529 bp in length. In the phylogenetic tree, T. calliphoroides were positioned in the subfamily Calliphorinae, and the closest species is Caliphora vomitoria. This is the first complete mitogenome record for the species.

The complete mitochondrial genome of a blowfly Calliphora nigribarbis (Vollenhoven, 1863) (Insecta: Diptera: Calliphoridae).

In the present study, the complete mitochondrial genome of a blowfly Calliphora nigribarbis has been sequenced and analyzed. The length of complete the mitochondrial genome is 16,279 bp, with 39.50% A, 13.20% C, 9.30% G, and 38.0% T nucleotide distribution. The complete mitochondrial genome consists of 13 protein-coding genes, 22 transfer RNAs, and 2 ribosomal RNAs likewise the most metazoan mitochondrial genomes. Furthermore, phylogenetic relationships of C. nigribarbis in the subfamily Calliphorinae investigated. The results suggested that C. vomitoria is the most related species to C. nigribarbis and the genus Calliphora is not monophyletic. This study provides the first complete mitochondrial genome sequence for the species.

Sequencing and analysis of the complete mitogenome of Doriprismatica atromarginata (Cuvier, 1804).

Doriprismatica atromarginata is a common nudibranch in the tropical and sub-tropical Indo-Pacific region. In this study, the complete mitogenome of D. atromarginata from Vietnam was recorded for the first time. The circular mitogenome had a size of 14,421 bp and consisted of 37 genes (13 protein-coding genes, two ribosomal RNA genes, and 22 tRNA genes). Phylogenetic tree based on amino acid sequences of coding genes demonstrated that D. atromarginata has sister group relationship with the genus Chromodoris.

Comparative Transcriptome Analysis for Understanding Predator-Induced Polyphenism in the Water Flea Daphnia pulex.

The crustacean Daphnia pulex is one of the best model organisms for studying inducible defense mechanisms due to their inducible morphology in response to the predator Chaoborus larvae. In this study, multiple developmental stages of D. pulex were exposed to C. flavicans larvae and transcriptome profiles of samples from late embryo to fifth instar were sequenced by the RNA-seq technique to investigate the genetic background underlying inducible defenses. In comparison, differentially expressed genes between defensive and normal morphs were identified, including 908 genes in late embryo, 1383 genes in the first-third (1⁻3) instar, and 1042 genes in fourth-fifth (4⁻5) instar. Gene ontology enrichment analysis showed that structural constituents of the cuticle and structural molecule activity genes were prominent up-regulated genes in late embryos. Down-regulated genes in late embryos and 1⁻3 instar comprised metabolic process, hydrolase activity, and peptidase activity gene classes. Pathway analysis indicated that small molecule neurotransmitter pathways were potentially involved in the development of inducible defenses. The characterization of genes and pathways in multiple developmental stages can improve our understanding of inducible defense responses of D. pulex to predation at the molecular level.

The complete mitogenome of ghost jellyfish Cyanea nozakii (Cnidaria, Semaeostomeae, Cyaneidae).

The cnidarian jellyfishes are impressive organisms to show animal mitochondrial genomic diversities. Their mitogenome structure is linear and tRNA content has one or two in numbers, which is highly different than other metazoans. In this study, a complete mitogenome of the ghost jellyfish Cyanea nozakii (Cnidaria, Semaeostomeae, Cyaneidae) was sequenced and analyzed. The mitgenome is 17,381 bp long with 38.5% A, 16.0% C, 13.9% G, and 31.6% T nucleotide distributions. In addition, phylogenetic relationship of C. nozakii in the class Scyphozoa was investigated by using mitochondrial protein coding genes. Due to results, C. nozakii was positioned in the paraphyletic order Semaeostomeae. This is the first complete mitogenome from the genus Cyanea.

Complete mitochondrial genome of xanthid crab Atergatis floridus (Linnaeus, 1767).

The complete mitochondrial genome sequenced from the floral egg crab Atergatis floridus (Linnaeus, 1767) and the determination of the position of the species in the reconstructed phylogenetic tree of the infraorder Brachyura using the protein coding mitochondrial genes are presented. Results show the mitochondrial genome length of A. floridus is 16,435 bp with nucleotide distribution as 33.4% A, 20.3% C, 10.5% G and 35.8% T. The structure of the complete mitochondrial genome of the species is the same as with the previous xanthid record. The result of the phylogenetic analysis suggests that A. floridus is the closest species to other Xanthidae species in the brachyuran records. This is the first complete mitochondrial genome record from the genus Atergatis.

Complete mitochondrial genome of Atergatis integerrimus (Decapoda, Xanthidae) from the Philippines.

In this study, a complete mitochondrial genome from the red egg crab, Atergatis integerrimus was sequenced and analyzed. The mitochondrial genome length is 16,333 bp and is composed of 13 protein-coding, two ribosomal RNA and 22 tRNA genes. The structure and gene orientation of the mitochondrial genome was found to be identical with the other brachyurans. The position of A. intergerrimus in the superfamily Xanthoidea was determined based on the mitochondrial protein-coding genes. The result of the phylogenetic relationship study showed that A. integerrimus was closest to Leptodius sanguineus.

The complete mitochondrial genome of Hemigrapsus penicillatus (De Haan, 1835) (Decapoda, Varunidae).

Hemigrapsus penicillatus is a small grapsoid crab which is found in Japan, Taiwan, Korea, and China coasts. In this study a complete mitogenome of Korean H. penicillatus was analyzed and phylogenetic relationships in the family Varunidae were investigated. The mitogenome size is 16,486 bp with 34.1% A, 18.1% C, 11.4% G, and 36.4% T nucleotide distributions. Genome structure and gene orientations are identical with previous records from the family and mitochondrial protein-coding gene based phylogenetic tree suggested that the closest species to H. penicillatus is H. sanguineus. This is the second complete mitogenome record from the genus Hemigrapsus and the first record for the species.

Analysis of complete mitochondrial genome of Etisus anaglyptus (Arthropoda, Decapoda, Xanthidae) with phylogenetic consideration.

The complete mitochondrial genome is sequenced and analyzed from a xanthid crab Etisus anaglyptus, which is the first complete mitochondrial genome for the genus. The mitochondrial genome length of E. anaglyptus is 16,435 bp and it is composed of 13 protein coding genes, two ribosomal RNA genes, and 22 tRNA genes. The structure and gene orientation of the mitochondrial genome is identical with the other brachyuran records. Furthermore, phylogenetic relationships of the infraorder Brachyura evaluated by mitochondrial protein coding genes. The phylogenetic study showed that E. anaglyptus is positioned in the superfamily Xanthoidea and the closest species to E. anaglyptus is Leptodius sanguineus.

Sequencing and analyzing complete mitochondrial DNA of Muscina angustifrons (Insecta, Diptera, Muscidae).

In this study, the complete mitochondrial genome sequenced and analyzed from a fly, Muscina angustifrons which collected from South Korea. The size of mitochondrial genome is 16,316 bp with 40.9% A, 12.3% C, 8.4% G and 38.4% T distribution. Furthermore, phylogenetic relationships of the superfamily Muscoidea evaluated due to mitochondrial protein coding genes. The results showed that the family Muscidae is a paraphyletic group and the closest species to M. angustifrons is M. levida. This is the third complete mitochondrial genome for the genus Muscina and the first genus record from South Korea.

Complete mitochondrial genome analysis of Phascolosoma sp. (Sipuncula, Phascolosomatida, Phascolosomatidea) from Micronesia.

In this study a Sipuncula species Phascolosoma sp. was collected from seagrass area from Chuuk lagoon Micronesia and its complete mitochondrial genome analyzed. This is the second complete mitochondrial genome record from the genus after Phascolosoma esculenta. The total length of mitochondrial genome of the species is 16,571 bp, which is longer than P. esculenta record. Also, locations of tRNA-Gly and putative control region are different between two records. Furthermore, phylogenetic relationship of Phascolosoma sp. are investigated due to protein-coding genes of mitochondrial genome. Due to the lack of recorded data, P. esculenta has been observed is the closest species to Phascolosoma sp. and they are belonging to the monophyletic group.

The complete mitogenome of a peanut worm Phascolosoma pacificum (Sipuncula, Phascolosomatida, Phascolosomatidea).

Sipuncula (peanut worms) is a traditional phylum which consists of unsegmented bilateral symmetric marine worms. Although it is accepted as a phylum, the phylogenetic position of Sipuncula has been questionable. There is lack of molecular record on Sipuncula species while morphological data is not enough to distinguish the closer relationships between Annelida and Sipuncula. In order to add more data to genomic library of sipunculan species, Phascolosoma pacificum (Keferstein, 1866) was collected from seagrass area of Chuuk lagoon/Micronesia and its complete mitochondrial genome sequenced. Furthermore, phylogenetic relationship of the phylum Sipuncula and the other Trochozoan phyla investigate due to mitochondrial protein coding genes. Although there is lack of recorded data, our results support the idea that Sipuncula are nested within Annelida according to phylogenetic analyses of mitochondrial protein coding genes.

Complete mitochondrial DNA analysis of Hydrotaea ignava (Insecta, Diptera, Muscidae).

The complete mitochondrial genome sequenced and analyzed from a black garbage fly, Hydrotaea ignava which is critically important species for forensic investigations. The size of mitochondrial genome is 17,026 bp with 40.8% A, 11.2% C, 8.2% G and 39.8% T distribution. This is the longest within complete mitochondrial genome records of the Muscidae species. The mitochondrial genome is composed of 13 protein coding, two ribosomal RNA and 22 tRNA genes and a putative control region. Furthermore, phylogenetic relationships of the superfamily Muscoidea evaluated due to mitochondrial protein coding genes. The results showed that the H. ignava placed in the paraphyletic Muscidae family and early diverged from a clade including Muscidae, Anthomyiidae and Scathophagidae species. This is the first complete mitochondrial genome for the genus Hydrotaea.

Complete mitogenome and phylogenetic analysis of hide beetle Dermestes maculatus (Insecta, Coleoptera, Dermestidae).

In this study, the complete mitochondrial genome of hide beetle Dermestes maculatus which was collected from Seoul, South Korea was sequenced by next-generation sequencing. The size of mitochondrial genome is 17,026 bp that composed of 13 protein coding, two ribosomal RNA and 22 tRNA genes which has the identical gene orientation with the other Bostrichiformia species. Additionally, the phylogenetic tree of the D. maculatus in the infraorder Bostrichiformia was reconstructed by using 13 protein-coding genes of complete mitochondrial genome. The results showed that the family Dermestidae is positioned in the infraorder Bostrichiformia early branched than family Bostrichidae. This study provides the first complete mitochondrial genome from the genus Dermestes.