Study of the possibility of differentiating species of the Bactrocera tryoni complex based on regions of the mitochondrial genes COI and 16S
Description
The Queensland fruit fly Bactrocera (Bactrocera) tryoni (Froggatt, 1897) is a member of the Tephritidae family and is a serious pest of a wide range of fruit and vegetable crops in Australia. The object of this study, Bactrocera tryoni, belongs to the complex of B. tryoni species, which, in addition to it, includes the species Bactrocera aquilonis (May, 1965), Bactrocera neohumeralis (Hardy, 1951), and Bactrocera melas (Perkins & May, 1949). Since these species are morphologically very similar and some may infect similar products, there is a need for molecular genetic diagnostic methods that will reliably distinguish these species from other species and among themselves. According to the obtained data on the COI gene generally in the sample matrix, there is a hiatus - a gap between intra- and interspecific distances. However, the species B. tryoni, B. aquilonis, and B. neohumeralis on the dendrogram form clades corresponding to the same species, therefore, it is impossible to distinguish them using the sequences of the COI gene region. However, based on this sequence, it is possible to narrow down the identification by molecular genetic methods to these three species, since they are well separated from other studied species. According to the graph of the distribution of pairwise genetic distances for the 16S gene, there is a lower variability than for the COI gene. On the dendrogram, the species B. tryoni, B. aquilonis, and B. neohumeralis form the one clade, but other species formed separate clades. It is necessary to increase the number of 16S gene sequences in the sample for each species. Thus, based on the COI gene sequence, it is possible to separate the species B. tryoni, B. aquilonis, and B. neohumeralis from other species of the genus Bactrocera studied by us. To study the relationships within the complex of B. tryoni species and search for a gene sequence by which it will be possible to reliably differentiate the B. tryoni species, another, possibly more rapidly evolving gene is required.
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To construct trees, we used sequences taken from the GenBank NCBI database. 80 sequences of the COI gene region of representatives of the genus Bactrocera were taken from the GenBank NCBI database with a view to construct trees. Seven sequences of the species Dacus ciliatus Loew, 1862 were taken as the data of the outgroup. 11 sequences of the 16S gene region of representatives of the genus Bactrocera and 9 sequences of the species Dacus ciliatus as an outgroup were used for analysis. The matrix of pairwise genetic distances of the studied species, the frequency of pairwise genetic distances were calculated using the MEGA-X program. To analyze the taxonomic status of the studied representatives of the genus Bactrocera, we used the method of detecting the hiatus between intra- and interspecific genetic distances (“barcoding gap”) implemented in the ABGD program. The model of nucleotide substitutions was chosen in the programs PAUP4 and MrModelTest; the optimal models for the samples (COI and 16S) are GTR (General Time Reversible) +I+G and GTR+I, respectively. The dendrograms were built using the MEGA-X program using the nearest neighbor method (Neighbour-Joining, NJ). As a result of the analysis, the best tree was chosen. Bootstrap support (1000 replications) for each branch is listed on the tree. The phylogenetic Bayesian tree was built taking into account the model of nucleotide substitutions in the program MrBayes 3.2.6, the Bayesian posterior probabilities of support for each node are indicated on the tree; three independent mcmc-launches of 10 million generations were carried out, one tree for every thousand generations was selected for final processing. The remaining trees were combined into a consensus tree using the method of maximum confidence in the hoard. The tree was visualized in the FigTree v.1.4.2 program.