The complete mitogenome of the horse stomach bot fly Gasterophilus pecorum (Fabricius) and a near-complete mitogenome of Wohlfahrt's wound myiasis fly Wohlfahrtia magnifica (Schiner) were sequenced. The mitogenomes contain the typical 37 mitogenes found in metazoans, organized in the same order and orientation as in other cyclorrhaphan Diptera. Phylogenetic analyses of mitogenomes from 38 calyptrate taxa with and without two non-calyptrate outgroups were performed using Bayesian Inference and Maximum Likelihood. Three sub-analyses were performed on the concatenated data: (1) not partitioned; (2) partitioned by gene; (3) 3rd codon positions of protein-coding genes omitted. We estimated the contribution of each of the mitochondrial genes for phylogenetic analysis, as well as the effect of some popular methodologies on calyptrate phylogeny reconstruction. In the favoured trees, the Oestroidea are nested within the muscoid grade. Relationships at the family level within Oestroidea are (remaining Calliphoridae (Sarcophagidae (Oestridae, Pollenia + Tachinidae))). Our mito-phylogenetic reconstruction of the Calyptratae presents the most extensive taxon coverage so far, and the risk of long-branch attraction is reduced by an appropriate selection of outgroups. We find that in the Calyptratae the ND2, ND5, ND1, COIII, and COI genes are more phylogenetically informative compared with other mitochondrial protein-coding genes. Our study provides evidence that data partitioning and the inclusion of conserved tRNA genes have little influence on calyptrate phylogeny reconstruction, and that the 3rd codon positions of protein-coding genes are not saturated and therefore should be included.

Stomach bot flies (Calyptratae: Oestridae, Gasterophilinae) are obligate endoparasitoids of Proboscidea (i.e. elephants), Rhinocerotidae (i.e. rhinos) and Equidae (i.e. horses and zebras, etc.), with their larvae developing in the digestive tract of hosts with very strong host specificity. They represent an extremely unusual diversity among dipteran, or even insect parasites in general, and therefore provide significant insights into the evolution of parasitism. The phylogeny of stomach bot flies was reconstructed based on extensive mitochondrial genomic data for Cobboldia , Gyrostigma and six of the eight known species of Gasterophilus . The phylogenetic tree, i.e. { Cobboldia , [ Gyrostigma , ( Gasterophilus pecorum , ( Gasterophilus intestinalis , ( Gasterophilus haemorrhoidalis , Gasterophilus inermis )), ( Gasterophilus nasalis , Gasterophilus nigricornis ))]}, provides a strong evolutionary reference to infer several biological patterns for the first time for this group: (i) host shifts of stomach bot flies from elephants to rhinoceroses and then from rhinoceroses to equids; (ii) dispersal with their hosts from the Afrotropical region into the Palaearctic and Oriental regions; (iii) oviposition site, originally on the host head, and egg production positively correlated with distance from the mouth; (iv) attachment of third‐instar larva originally in the stomach, with duodenal and large intestinal positions secondarily derived; and (v) guanine and cytosine enrichment of the mitogenome as an adaptation to larval life in the warm environment of the host digestive tract, combined with the need for a high evolutionary rate to cope with the fast evolution of their mammalian hosts.

The aim of this study was to detect the expression of proliferatng cell nuclear antigen (PCNA) and c-myc in Epstein‑Barr virus (EBV)-associated gastric carcinoma (EBVaGC) and EBV-negative gastric carcinoma (EBVnGC), as well as the expression of EBV-encoded proteins in EBVaGC and their effect on carcinogenesis and the development of gastric cancer. The PCNA and c-myc protein levels were assessed by immunohistochemistry in 13 EBVaGC and 45 EBVnGC specimens. The expression of related genes of EBV, including EB nuclear antigen (NA)-1 and EBNA2 genes, latent membrane protein 1 (LMP1) and early genes BARF1 and BHRF1 were tested by reverse transcription-polymerase chain reaction (RT-PCR) and southern blotting. The PCNA labeling index (LI) of EBVaGCs, EBVnGCs and the corresponding adjacent tissues of EBVaGCs were 49.3768±12.1832, 14.839±7.1847, 35.613±8.3831 and 24.2735±10.1332, respectively. The PCNA LI was significantly different between EBVaGC and EBVnGC of EBVaGC (t=4.686, P<0.01). The difference between EBVaGC and corresponding adjacent tissues of EBVaGC was also significant (t=8.805, P<0.01). The expression of c-myc protein was detected in 33 of 58 (55.39%) gastric carcinomas and in 21 of 58 (36.21%) adjacent tissues. There was a significant difference between the two groups (χ2=4.989, P0.05). EBNA1 mRNA was detected in all 13 EBVaGC cases, while EBNA2 and LMP1 mRNA was not detected in these cases. Of the 13 EBV-positive samples, 6 exhibited BARF1 transcripts and 2 exhibited BHRF1 transcripts. c-myc expression did not correlate with the presence of EBV in EBVaGC. EBV infection may induce PCNA expression. The lack of EBNA2 and LMP1 protein expression in EBVaGC suggests that EBNA2 and LMP1 do not correlate with cell apoptosis and c-myc expression. Early genes BARF1 and BHRF1 may play an important role in the development and progression of gastric carcinomas by immortalizing epithelial cells.