OBJECTIVES: To identify the common sarcosaprophagous flies in the Yangtze River Delta based on mitochondrial cytochrome c oxidase subunit Ⅰ(COⅠ) gene sequence and verify the reliability of this method. METHODS: Seven common genetically stable sarcosaprophagous flies in three families and six genera were collected from large domestic pig carcasses placed in the field and cultured in the laboratory for many generations. The whole genome DNA was extracted and the COⅠ gene fragment was amplified. The forward and reverse sequencing was followed by splicing. The base composition of the amplified fragment and the rate of interspecific evolutionary divergence were analyzed by software such as Mega 7.0.26. The phylogenetic tree of COⅠ gene sequence of common necrophagous flies in the Yangtze River Delta was established by neighbor joining (NJ) method and unweighted pair-group method with arithmetic means (UPGMA) method. RESULTS: The average base composition of different flies was A(30.14%), T(38.23%), C(15.98%), G(15.65%). The rate of interspecific evolutionary divergence ranged from 2.2% to 15.3%, the lowest rate was between Chrysomya megacephala and Chrysomya pinguis, the highest rate was between Muscina stabulans and Boettcherisca peregrina. CONCLUSIONS COⅠ gene can be used to identify the common necrophagous flies in the Yangtze River Delta.
Animals ; Cadaver ; Diptera/genetics* ; Phylogeny ; Reproducibility of Results ; Rivers

Objective To assess the feasibility of using 28S ribosomal RNA （28S rRNA） and mitochondrial cytochrome c oxidase subunit Ⅰ （COⅠ） gene sequences of nine necrophagous Calliphorid flies for the identification of common necrophagous Calliphorid flies, and to provide technical support for postmortem interval （PMI） estimation. Methods Twenty-three Calliphorid flies were collected and identified morphologically, and DNA were extracted from legs. The gene fragments of 28S rRNA and COⅠ were amplified and sequenced, then the sequence alignment was performed with BLAST. The composition of obtained sequences was analyzed and evolutionary divergence rate between species and intraspecies were established. The phylogeny tree was constructed with neighbor-joining method. Results The 23 necrophagous Calliphorid flies were identified to 9 species of 5 genera. The 715 bp from 28S rRNA and 637 bp from COⅠ gene were obtained and the online BLAST result showed more than 99% of similarity. The phylogeny tree showed that the necrophagous flies could cluster well into 9 groups, which was consistent with morphological identification results. The intraspecific difference in 28S rRNA was 0 and the interspecific difference was 0.001-0.033. The intraspecific difference in COⅠ was 0-0.008 and the interspecific difference was 0.006-0.101. Conclusion Combined use of 28S rRNA and COⅠ gene sequence fragments can effectively identify the nine Calliphorid flies in this study. However, for closely related blowfly species, more genetic markers should be explored and used in combination in future.
Animals ; DNA, Mitochondrial/genetics* ; Diptera/genetics* ; Phylogeny ; RNA, Ribosomal, 28S/genetics* ; Sequence Analysis, DNA ; Species Specificity

Objective To identify the species of common necrophagous flies in Fujian Province by gene fragment sequences of mitochondrial cytochrome c oxidase subunit Ⅰ （COⅠ） and 16S ribosomal deoxyribonucleic acid （16S rDNA）, and to explore the identification efficacy of these two molecular markers. Methods In total 22 common necrophagous flies were collected from the death scenes in 9 different regions in Fujian Province and DNA was extracted from the flies after morphological identification. The gene fragments of COⅠ and 16S rDNA were amplified and sequenced. All the sequences were uploaded to GeneBank and BLAST and MEGA 10.0 software were used to perform sequence alignment, homology analysis and intraspecific and interspecific genetic distance analysis. The phylogenetic trees of DNA fragment sequences of COⅠ and 16S rDNA of common necrophagous flies in Fujian Province were established by unweighted pair-group method with arithmetic means （UPGMA）, respectively. Results The flies were classified into 6 species, 5 genera and 3 families by morphological identification. The results of gene sequence analysis showed that the average number of interspecific and intraspecific genetic distance of 16S rDNA ranged from 1.8% to 8.9% and 0.0% to 2.4%, respectively. The average number of interspecific and intraspecific genetic distance of COⅠ ranged from 7.2% to 13.6% and 0.0% to 6.3%, respectively. Conclusion The gene sequences of COⅠ and 16S rDNA can accurately identify the species of different necrophagous flies, and 16S rDNA showed higher value in species identification of common calliphoridae necrophagous flies in Fujian Province.
Animals ; DNA, Ribosomal/genetics* ; Diptera/genetics* ; Humans ; Phylogeny ; RNA, Ribosomal, 16S/genetics* ; Sequence Analysis, DNA ; Species Specificity

Identifying sarcosaphagous flies specimens is an important first step in a forensic-entomological analysis. It is traditionally performed using morphological features of the Sarcosaphagous Flies. However, Morphological identification may be complicated by the numerical diversity of species and physical similarity between different species, particularly in immature stages. The sequences focused on some sections of the cytochrome oxidase I and II (CO I and CO II) encoding region of mtDNA could be as the prospective basis of a diagnostic technique. By Analysis of these sequences, forensic doctors can reveal abundant phylogenetic informative nucleotide substitutions that could effectively identify Sarcosaphagous flies to species group. It was not reported in our country before and was reviewed in this article now.
Animals ; DNA, Mitochondrial/genetics* ; Diptera/genetics* ; Electron Transport Complex IV/genetics* ; Forensic Medicine/methods* ; Humans ; Larva/genetics* ; Polymerase Chain Reaction/methods* ; Sequence Analysis, DNA ; Species Specificity

Species identification of sarcosaphagous insects is one of the important steps in forensic research based on the knowledge of entomology. Recent studies reveal that the application of molecular biology, especially the mtDNA sequences analysis, works well in the species identification of sarcosaphagous insects. The molecular biology characteristics, structures, polymorphism of mtDNA of sarcosaphagous insects, and the recent studies in species identification of sarcosaphagous insects are reviewed in this article.
Amino Acid Sequence ; Animals ; Base Sequence ; DNA, Mitochondrial/genetics* ; Diptera/genetics* ; Electron Transport Complex IV/genetics* ; Entomology ; Forensic Medicine/methods* ; Genes, Mitochondrial/genetics* ; Insecta/genetics* ; Polymerase Chain Reaction ; Polymorphism, Genetic ; RNA, Ribosomal/genetics* ; Sequence Analysis, DNA ; Species Specificity

OBJECTIVE: To solve the problems of identification of Sarcosaphagous flies and their larvae, pupas and eggs. METHODS: Sarcosaphagous Flies (Diptera) Samples were collected on the corpses of rabbits in the Huhhot district and a pig in the Dunhuang district. A 278bp region in the cytochrome oxidase subunit I (CO I) gene in mtDNA was analysed by DNA sequencing, A neighbour-joining tree using the Tamura and Nei model of nucleotide substitution was also constructed using the MEGA2.1 package. RESULTS: A 278 base pairs region of the gene for CO I encoding region of mtDNA of above all samples was showed less than 1% sequence divergence within species and about 3% divergence between species. CONCLUSION It is an effective, easy and accurate method to be used for identification of these Sarcosaphagous Flies (Diptera) to species group by sequencing the 278 base pairs region of the CO I encoding gene of mtDNA.
Animals ; Base Sequence ; DNA, Mitochondrial/genetics* ; Diptera/genetics* ; Electron Transport Complex IV/genetics* ; Forensic Medicine/methods* ; Larva/genetics* ; Phylogeny ; Polymerase Chain Reaction/methods* ; Pupa/genetics* ; Rabbits ; Sequence Analysis, DNA ; Species Specificity ; Swine

OBJECTIVE: To identify sarcosaphagous flies and their larvae, pupa. METHODS: Sarcosaphagous flies and their larvae, pupas were collected from human corpses and their surroundings in the Weifang city. A 304 bp region in COI gene was analyzed by mtDNA sequencing. RESULTS: The studied region showed no sequence divergence within same species and significant difference were found between different species in all samples. CONCLUSION It is a practical approach to identify these Sarcosaphagous flies and their larvae, pupas by sequence analysis of the 304bp region of the COI in mtDNA.
Animals ; Base Sequence ; China ; DNA Primers ; DNA, Mitochondrial/genetics* ; Diptera/genetics* ; Electron Transport Complex IV/genetics* ; Forensic Medicine ; Genes, Insect ; Humans ; Larva/genetics* ; Phylogeny ; Polymerase Chain Reaction/methods* ; Pupa/genetics* ; Sequence Analysis, DNA/methods* ; Species Specificity

Sarcosaphagous insects are very important to investigate some criminal cases. They are significant useful in estimating post-mortem interval (PMI) and corpse transfer post-mortem. Lucilia are very common sarcosaphagous insects. They like sunshine and are usually the earliest to touch the cadaver. These characteristics and others such as the stages of their larvae development can offer good evidences for criminal case investigation. This paper summarizes details of their application for estimating postmortem interval in recent years and reviews the methods to identify species and to determine the age of adult Lucilia with molecular biology and entomological morphology.
Animals ; DNA, Mitochondrial/genetics* ; Diptera/physiology* ; Entomology/methods* ; Feeding Behavior ; Forensic Medicine/methods* ; Larva/physiology* ; Polymerase Chain Reaction/methods* ; Postmortem Changes ; Seasons ; Sequence Analysis, DNA ; Species Specificity ; Weather

OBJECTIVE: To solve the difficulties of identification of Sarcosaphagous flies such as Lucilia sericata (Meigen) and Lucilia cuprina (Wiedemann) which could not be identified by analyzing the 278bp and 635 bp regions of the gene encoding for cytochrome oxidase subunit I and II (CO I and CO II) in mtDNA. METHODS: Specimens were collected from the corpses of rabbits on the grassland in Huhhot and Chengdu, the sequences of 551 bp region of 16S rDNA of their mtDNA were analyzed, the multiple-alignment program DNAMAN(version 4.0) and MEGA 2.1 sofeware were employed for sequence alignments neighbour-joining tree construction. RESULTS: Lucilia sericata (Meigen) and Lucilia cuprina (Wiedemann) were distinguished successfully by sequence analysis of The 551 bp region of the gene of 16S rDNA. CONCLUSION The 551 bp region of the gene of 16S rDNA of sarcosaphagous flies can be used for identifying them on species level effectively. It is likely to be a successful compliment to identify the sarcosaphagous flies by sequence analysis of CO I and CO II in mtDNA.
Animals ; DNA, Mitochondrial/genetics* ; DNA, Ribosomal/genetics* ; Diptera/genetics* ; Forensic Medicine/methods* ; Molecular Sequence Data ; Polymerase Chain Reaction/methods* ; RNA, Ribosomal, 16S/genetics* ; Rabbits ; Sequence Analysis, DNA ; Species Specificity

OBJECTIVE: Using CO II sequences to identify common species of carrion-breeding flies and larvae. METHODS: flies and larvae were collected on the corpses of rats in Zhengzhou district, DNA was extracted, CO II sequences were amplified and sequenced. Clustalx and MEGA 4.0 software were used to analyze the gene sequences and to construct the phylogenetic trees. RESULTS: There was no significant gene difference between adults and larvae. COII gene sequences could be used to identify Boettcherisca peregrina, Aldrichina grahami and Lucilia illustris but they could not distinguish Lucilia cuprina from the Lucilia sericata because of their close evolutionary distance and single nucleotide polymorphisms in aldrichina grahami and Lucilia illustris populations were found. CONCLUSION CO II sequence of mtDNA in Zhengzhou district can be used effectively to identify some common species of carrion-breeding fly. The method is simple and accurate.
Animals ; Base Sequence ; China ; DNA Primers ; DNA, Mitochondrial/genetics* ; Diptera/genetics* ; Electron Transport Complex IV/genetics* ; Entomology ; Forensic Medicine/methods* ; Genes, Insect ; Larva/genetics* ; Phylogeny ; Polymerase Chain Reaction/methods* ; Rats ; Sequence Analysis, DNA ; Species Specificity