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High-throughput sequencing (HTS) technology has profoundly been involved in sequencing whole genomes of several organisms in a fast and cost-effective manner. Although HTS provides an alternative biomonitoring method to the time-consuming and taxonomy-expertise dependent morphological approach, still we cannot rule out the possibility of the impediment and misidentification biases. In this article we aim to retrieve whole mitochondrial genome (mitogenome) sequences from publicly available raw sequencing data for phylogenetic comparison of Ixodes persulcatus. For this comparison, we sequenced whole mitogenomes of four I. persulcatus ticks from Japan and constructed mitogenomes from raw sequencing data of 74 I. persulcatus ticks from China. Bayesian phylogenetic trees were inferred by the concatenated fifteen mitochondrial genes. We further tested our results by the phylogenetic analysis of cytochrome c oxidase subunit 1 (cox1) gene and internal transcribed spacer 2 (ITS2) sequences. Our findings showed that 70 constructed mitogenomes from China were clustered with the sequenced four mitogenomes of I. persulcatus from Japan. We also revealed that mitogenome sequences retrieved from two data sets CRR142297 and CRR142298 were clustered with Ixodes nipponensis. Moreover, other two mitogenome sequences from CRR142310 and CRR142311 formed a clade with Ixodes pavlovskyi. The phylogenetic analysis of cox1 gene and ITS2 sequences confirmed the identification errors of these four samples. The overall phylogenetics in our study concluded that accurate morphological identification is necessary before implementing HTS to avoid any misidentification biases.

Objective: This study sought to explore the mRNA and protein expression levels of eukaryotic translation elongation factor 1 alpha 2 (eEF1A2) and members of the PI3K-Akt signaling pathway in the context of cervical cancer. We sought to clarify the expression of eEF1A2, PI3K, Akt during cervical cancer tumorigenesis and development.Methods: Samples from 72 cases of cervical cancer were collected, as well as 46 cases of cervical intraepithelial neoplasia (CIN), which reflects the continuous process of cervical cancer development, divided into CIN I, CIN II, CIN III, and 40 cases of chronic cervicitis. qRT-PCR was to detect the mRNA levels of eEF1A2, PI3K, and Akt, with β-actin used as an internal control. eEF1A2, PI3K, p-Akt protein levels in cervical cancer, CIN, and chronic cervicitis tissues were detected by immunohistochemical. eEF1A2, PI3K and p-Akt protein expression levels in HeLa, SiHa, and human umbilical vein endothelial cells were detected by western blot.Results: qRT-PCR results showed that the level of mRNA expression of eEF1A2, PI3K, and Akt was higher in cervical cancer tissues than that in normal cervical tissues. Immunohistochemistry results showed that the eEF1A2, PI3K, and p-Akt protein levels were higher in cervical cancer tissues than in cervical chronic cervicitis tissues. The differences were statistically significant (P < 0.05). The expression of eEF1A2, PI3K, and p-Akt protein was higher in HeLa and SiHa cervical cell lines than that in normal epithelial cells.Conclusion: Together, these results suggest that the aberrant expression of eEF1A2, PI3K, and Akt may play a role in cervical cancer development and tumorigenesis.

Background The typical single-chromosome mitochondrial (mt) genome of animals has fragmented into multiple minichromosomes in the lineage Mitodivisia, which contains most of the parasitic lice of eutherian mammals. These parasitic lice differ from each other even among congeneric species in mt karyotype, i.e. the number of minichromosomes, and the gene content and gene order in each minichromosome, which is in stark contrast to the extremely conserved single-chromosome mt genomes across most animal lineages. How fragmented mt genomes evolved is still poorly understood. We use Polyplax sucking lice as a model to investigate how tRNA gene translocation shapes the dynamic mt karyotypes. Results We sequenced the full mt genome of the Asian grey shrew louse, Polyplax reclinata. We then inferred the ancestral mt karyotype for Polyplax lice and compared it with the mt karyotypes of the three Polyplax species sequenced to date. We found that tRNA genes were entirely responsible for mt karyotype variation among these three species of Polyplax lice. Furthermore, tRNA gene translocation observed in Polyplax lice was only between different types of minichromosomes and towards the boundaries with the control region. A similar pattern of tRNA gene translocation can also been seen in other sucking lice with fragmented mt genomes. Conclusions We conclude that inter-minichromosomal tRNA gene translocation orientated towards the boundaries with the control region is a major contributing factor to the highly dynamic mitochondrial genome organization in the parasitic lice of mammals.

International audience; The evolution and phylogenetic relationships of the ticks at both the family and genus levels are contested. The genus Amblyomma and its subgenera are in a state of flux; moreover, the relationships among the three tick families are controversial due to conflicting phylogenetic support for different arrangements of the three families of living ticks. With 18 newly sequenced mitochondrial (mt) genomes of ticks included, we executed the largest mt genome phylogenetic study of ticks so far. Phylogenetic trees were inferred from one sea spider mt genome, one horseshoe crab, five mite mt genomes and 146 tick mt genomes from 120 species: 153 mt genomes in total. Sixteen phylogenetic trees were inferred from 10 datasets using both maximum likelihood and Bayesian inference methods. We describe the first novel mt gene-arrangement for the metastriate Ixodidae in Amblyomma (Africaniella) transversale. Also, three unusual partial 16S rRNA gene inserts were found in the mt genome of Haemaphysalis (Alloceraea) kitaokai: we consider the possible role of past genome translocation events in the formation of these inserts. Our phylogenies revealed evidence that: (i) the genus Amblyomma is polyphyletic with respect to Amblyomma (Africaniella) transversale; (ii) the subgenus Aponomma is apparently embedded in the genus Amblyomma; (iii) Haemaphysalis (Segalia) parva and Haemaphysalis (Alloceraea) kitaokai form a clade to the exclusion of other Haemaphysalis species; and (iv) the phylogenetic position of the family Nuttalliellidae is unstable among phylogenies from different datasets.

Based on a comprehensive study of museum specimens, eight new species of sucking lice of the genus Hoplopleura Enderlein, 1904 (Psocodea: Phthiraptera: Hoplopleuridae), are described from six genera of Australian Old Endemic rodents: Conilurus Ogilby, 1838 (Rodentia: Muridae), Leggadina Thomas, 1910 (Rodentia: Muridae), Leporillus Thomas, 1906 (Rodentia: Muridae), Mesembriomys Palmer, 1906 (Rodentia: Muridae), Pogonomys Milne-Edwards, 1877 (Rodentia: Muridae), and Xeromys Thomas, 1889 (Rodentia: Muridae). The description of these new species increases the number of sucking louse species from endemic Australian rodents from 13 to 21 and extends the records of sucking lice to all of the 14 genera of endemic rodents in Australia. Our results show that sucking lice are much more diverse among rodents in Australia than previously known. Furthermore, the Australian Hoplopleura species are host specific—each Hoplopleura species, including the eight new species described in the present study, parasitizes only a single host species, except Hoplopleura irritans Kuhn and Ludwig, 1967 (Psocodea: Phthiraptera: Hoplopleuridae) and Hoplopleura melomydis Weaver, 2017 (Psocodea: Phthiraptera: Hoplopleuridae), each of which is found on two host species. An updated dichotomous key for identifying Australian Hoplopleura species is included.

We describe two new species of sucking lice in the genus Hoplopleura Enderlein, 1904 (Psocodea: Phthiraptera: Hoplopleuridae) from Australia: Hoplopleura gracilicaudatusa n. sp. from the eastern chestnut mouse Pseudomys gracilicaudatus (Gould) (Rodentia: Muridae), and Hoplopleura nanusa n. sp. from the western chestnut mouse Pseudomys nanus (Gould) (Rodentia: Muridae). Pseudomys Gray is the most speciose genus of rodents endemic to Australia with 24 species; however, only two Pseudomys species have been reported previously to be hosts of sucking lice. The description of the new species in the present study doubles the number of sucking louse species known to parasitize Pseudomys mice and increases the total number of sucking louse species known from endemic Australian rodents from 21 to 23. Pseudomys gracilicaudatus and P. nanus are closely related murines that diverged ~1 MYA with distinct and widely separated extant geographic distributions. The two new Hoplopleura species described in the present study share some morphological characters and likely co-evolved and co-speciated with their chestnut mouse hosts.

Chunhe Zhang,1,* Shaowei Fu,1,* Luyue Wang,1,* Fang Wang,1 Dan Wu,1 Xiangyi Zhe,1 Huizhen Xin,1 Hongtao Li,1 Dongmei Li,1 Fuyuan Jin,1 Renfu Shao,2 Zemin Pan1 1Department of Biochemistry and Molecular Biology, School of Medicine, Shihezi University, Xinjiang Endemic and Ethnic Disease and Education Ministry Key Laboratory, Shihezi, Xinjiang 832002, People’s Republic of China; 2School of Science, Technology and Engineering, Genecology Research Centre, University of the Sunshine Coast, Sippy Downs, Queensland 4556, Australia*These authors contributed equally to this workCorrespondence: Zemin Pan; Xiangyi ZheDepartment of Biochemistry and Molecular Biology, School of Medicine, Shihezi University, Xinjiang Endemic and Ethnic Disease and Education Ministry Key Laboratory, Shihezi, People’s Republic of ChinaEmail panteacher89@sina.com; 342480821@qq.comIntroduction: The aim of this study was to determine whether ZNF582 gene methylation and tissue protein expression can be used as a tool with high sensitivity and specificity for cervical cancer screening. We analyzed the correlation between promoter methylation of ZNF582 gene and cervical cancer and high risk HPV16/18 infection.Methods: Tissue samples of normal cervical or chronic cervicitis (n=51), CIN (cervical intraepithelial neoplasia) (n=35), and cervical carcinoma (n=68) were tested for HPV16/18 infection by polymerase chain reaction (PCR). We also detected the methylation status of the ZNF582 gene promoter in the same tissues by methylation-specific PCR (MSP), then analyzed the correlation between ZNF582 promoter methylation and HPV16/18 infection. Immunohistochemistry was used to analyze ZNF582 gene expression in 152 cervical tissues. We detected ZNF582 mRNA expression in cervical tissues (including cancer and non-cancer) by real-time fluorescence quantitative PCR (qPCR).Results: Among 93 high-grade cervical lesions (CINII and above) and cervical cancer samples, 57 cases were positive for HPV16/18 infection and 36 cases were negative. ZNF582 gene methylation occurred in 9 out of 51 cases in normal cervical tissues (17.6%), 16 of 35 cases in CIN tissues (45.7%), and 50 of 68 cases in cervical cancer (73.5%). The differences in methylation rate of the three groups were statistically significant (P< 0.05). The ZNF582 methylation rate in the positive HPV16/18 infection group was 73.7%, while the negative group was 63.9%. Compared with normal tissues, ZNF582 protein was highly expressed in cervical cancer tissues, but mRNA expression was low.Conclusion: While ZNF582 protein is highly expressed in cervical cancer tissues, it was not sufficient for use as a standard for cervical cancer staging. On the other hand, ZNF582 promoter methylation had high specificity and sensitivity in detecting CINII and highly diseased cervical lesions and could be used as a diagnostic marker for cervical cancer of women.Keywords: cervical cancer, ZNF582 gene, promoter methylation, HPV16/18, gene expression

We report for the first time the fragmented mitochondrial (mt) genomes of two Pedicinus species: Pedicinus obtusus and Pedicinus badii, and compared them with the lice of humans and chimpanzees. Despite being congeneric, the two monkey lice are distinct from each other in mt karyotype. The variation in mt karyotype between the two Pedicinus lice is the most pronounced among the congeneric species of sucking lice observed to date and is attributable to the opposite directions between them in mt karyotype evolution. Two of the inferred ancestral mt minichromosomes of the higher primate lice merged as one in the macaque louse whereas one of the ancestral minichromosomes split into two in the colobus louse after these two species diverged from their most recent common ancestor. Our results showed that mt genome fragmentation was a two-way process in the higher primate lice, and minichromosome merger was more common than previously thought.

Objective : This study aimed to: 1) investigate the status and genotype distribution of human papillomavirus (HPV) in infected Uygur and Han women in Xinjiang; 2) elucidate the variation of the HPV16 E6 gene sequence in the cervix of Uygur and Han women in Xinjiang; and 3) analyze the HPV16 E6 gene polymorphism site and relationship with the development of cervical cancer. Methods : A total of 2879 samples of cervical mucus from the exfoliated cells of Uygur and Han women were collected for an epidemiological analysis of HPV. Genomic DNA was extracted from the cervical HPV16-positive tissues of 110 Uygur and Han women, and E6 was amplified by PCR and sequenced. The HPV16 E6 sequence was analyzed using the European standard as the prototype, and an evolutionary tree analysis was performed. HPV16 E6-295T/350T-GV230, HPV16 E6-295G/350G-GV230, and HPV16 E6-295T/350G-GV230 were stably transfected into human cervical cancer C33A cells. HPV16 E6 protein expression was confirmed using a direct immunofluorescence assay. CCK8 and clonogenic assays were used to analyze C33A cell proliferation. Both a transwell and cell scratch assay were used to study C33A cell migration and invasion. C33A cell apoptosis was analyzed using FACS experiments. SPSS17.0 statistical software was used for statistical data processing. P < 0.05 was considered statistically significant. Results : The total HPV infection rate was 26.390% (760/2879), whereas the Uygur infection rate was 22.87% (196/857) and the Han infection rate was 27.89% (564/2022) (P < 0.05). HPV16, HPV 52, and HPV 53 were associated with higher detection rates in Uygur, whereas HPV16, HPV52, and HPV58 exhibited a higher detection rate in Han. HPV-infected women from Uygur and Han commonly exhibited a single infection. A total of 14 mutation sites were identified in the HPV16 E6 gene by sequencing 110 HPV16-positive samples, including eight missense and six synonymous mutations. Among these, 65 cases of E6 genes were mutated at nucleotide 350 (T350G) with the corresponding amino acids changing from leucine to valine (L83V) and a mutation rate of 59.09%. Moreover, there were seven cases of an E6 gene mutation at nucleotide 295 (T295G) with corresponding amino acid changes from aspartic to glutamic (D64E) and a mutation rate of 6.36%. It is important to note that these seven cases of HPV16 E6 T295G mutations were accompanied by the E6 T350G mutation. Phylogenetic analysis showed that there were HPV16 European (Ep), European variant (E), and three Asian (As) types in Uygur and Han women. No African (Af) and Asian American (AA) types were observed. When HPV16 E6 295T/350T, 295G/350G, and 295T/350G GV230 eukaryotic expression vector(s) were stably transfected into cervical cancer C33A cells, they were found to promote cellular proliferation, migration, invasion, and inhibit apoptosis. The 295T/350G-GV230 had the strongest effect on C33A cells and 295G/350G-GV230 was significantly stronger than 295T/350T-GV230 (P < 0.05). Conclusions : The positive HPV infection rates differed between the Uygur and Han groups in Xinjiang, and the genotype distribution of HPV infection was different. Between the Uygur and Han women in Xinjiang, the main types of HPV16 infection were European (E) and Asian (As). After stably transfecting C33A cells with a eukaryotic expression vector for different polymorphism sites (295T/350T, 295G/350G, and 295T/350G), the 295T/350G mutation site promoted the proliferation，migration, and invasion of C33A cells to a greater extent than 295G/350G; however, 295G/350G had a stronger effect than 295T/350T.

The booklouse, Liposcelis bostrychophila is an important storage pest worldwide. The mitochondrial (mt) genome of an asexual strain (Beibei, China) of the L. bostrychophila comprises two chromosomes; each chromosome contains approximate half of the 37 genes typically found in bilateral animals. The mt genomes of two sexual strains of L. bostrychophila, however, comprise five and seven chromosomes, respectively; each chromosome contains one to six genes. To understand mt genome evolution in L. bostrychophila, and whether L. bostrychophila is a cryptic species, we sequenced the mt genomes of six strains of asexual L. bostrychophila collected from different locations in China, Croatia, and the United States. The mt genomes of all six asexual strains of L. bostrychophila have two chromosomes. Phylogenetic analysis of mt genome sequences divided nine strains of L. bostrychophila into four groups. Each group has a distinct mt genome organization and substantial sequence divergence (48.7–87.4%) from other groups. Furthermore, the seven asexual strains of L. bostrychophila, including the published Beibei strain, are more closely related to two other species of booklice, L. paeta and L. sculptilimacula, than to the sexual strains of L. bostrychophila. Our results revealed highly divergent mt genomes in the booklouse, L. bostrychophila, and indicate that L. bostrychophila is a cryptic species.

Argasid systematics remains controversial with widespread adherence to the Hoogstraal (1985) classification scheme, even though it does not reflect evolutionary relationships and results in paraphyly for the main genera of soft ticks (Argasidae), namely Argas and Ornithodoros. The alternative classification scheme, proposed by Klompen and Oliver (1993), has problems of its own: most notably paraphyly of the subgenus Pavlovskyella and the controversial grouping together of the subgenera Alectorobius, Antricola, Carios, Chiropterargas, Nothoaspis, Parantricola, Reticulinasus and Subparmatus into the genus Carios. Recent phylogenetic analyses of 18S/28S rRNA sequences and mitochondrial genomes agree with the scheme of Klompen and Oliver (1993), with regard to the paraphyly of Pavlovskyella, placement of Alveonasus, Ogadenus, Proknekalia and Secretargas in the Argasinae and placement of Carios and Chiropterargas in the Ornithodorinae (Mans et al., 2019). The Carios clade and its constituent subgenera remain controversial, since the phylogenetic position of its type species Carios (Carios) vespertilionis Latreille, 1796 (formerly Argas vespertilionis) has not been determined with confidence. The current study aimed to resolve Carios sensu lato Klompen and Oliver, 1993, and Carios sensu stricto Hoogstraal, 1985, by determining and analysing phylogenetic nuclear and mitochondrial markers for C. (C.) vespertilionis. Both the nuclear and mitochondrial markers support placement of Carios s.s. within the subfamily Ornithodorinae, but to the exclusion of the clade that includes the 6 other subgenera that are part of Carios s.l. Klompen and Oliver (1993), namely Alectorobius, Antricola, Nothoaspis, Parantricola, Reticulinasus and Subparmatus. These 6 subgenera form a monophyletic clade that might be placed as new subgenera within the genus Alectorobius, or elevated to genera. Given the substantial differences in biology among these subgenera, we propose that these 6 subgenera be elevated to genera. Thus, we propose to modify the classification scheme of Mans et al. (2019) so that the subfamily Argasinae now has six genera, Alveonasus, Argas (subgenera Argas and Persicargas), Navis, Ogadenus, Proknekalia and Secretargas, and the subfamily Ornithodorinae has nine genera, Alectorobius, Antricola (subgenera Antricola and Parantricola), Carios, Chiropterargas, Nothoaspis, Ornithodoros (subgenera Microargas, Ornamentum, Ornithodoros, Pavlovskyella and Theriodoros), Otobius, Reticulinasus and Subparmatus (genera indicated in bold).

Background Debilitating skin infestations caused by the mite, Sarcoptes scabiei, have a profound impact on human and animal health globally. In Australia, this impact is evident across different segments of Australian society, with a growing recognition that it can contribute to rapid declines of native Australian marsupials. Cross-host transmission has been suggested to play a significant role in the epidemiology and origin of mite infestations in different species but a chronic lack of genetic resources has made further inferences difficult. To investigate the origins and molecular epidemiology of S. scabiei in Australian wildlife, we sequenced the mitochondrial genomes of S. scabiei from diseased wombats (Vombatus ursinus) and koalas (Phascolarctos cinereus) spanning New South Wales, Victoria and Tasmania, and compared them with the recently sequenced mitochondrial genome sequences of S. scabiei from humans. Results We found unique S. scabiei haplotypes among individual wombat and koala hosts with high sequence similarity (99.1% - 100%). Phylogenetic analysis of near full-length mitochondrial genomes revealed three clades of S. scabiei (one human and two marsupial), with no apparent geographic or host species pattern, suggestive of multiple introductions. The availability of additional mitochondrial gene sequences also enabled a re-evaluation of a range of putative molecular markers of S. scabiei, revealing that cox1 is the most informative gene for molecular epidemiological investigations. Utilising this gene target, we provide additional evidence to support cross-host transmission between different animal hosts. Conclusions Our results suggest a history of parasite invasion through colonisation of Australia from hosts across the globe and the potential for cross-host transmission being a common feature of the epidemiology of this neglected pathogen. If this is the case, comparable patterns may exist elsewhere in the ‘New World’. This work provides a basis for expanded molecular studies into mange epidemiology in humans and animals in Australia and other geographic regions. Electronic supplementary material The online version of this article (10.1186/s12862-017-1086-9) contains supplementary material, which is available to authorized users.

Eriophyoid mites (superfamily Eriophyoidea) comprise >4400 species worldwide. Despite over a century of study, the phylogenetic position of these mites within Acariformes is still poorly resolved. Currently, Eriophyoidea is placed in the order Trombidiformes. We inferred the high-level phylogeny of Acari with the mitochondrial (mt) genome sequences of 110 species including four eriophyoid species, and the nuclear small subunit (18S) rRNA gene sequences of 226 species including 25 eriophyoid species. Maximum likelihood (ML), Bayesian inference (BI) and Maximum parsimony (MP) methods were used to analyze the sequence data. Divergence times were estimated for major lineages of Acari using Bayesian approaches. Our analyses consistently recovered the monophyly of Eriophyoidea but rejected the monophyly of Trombidiformes. The eriophyoid mites were grouped with the sarcoptiform mites, or were the sister group of sarcoptiform mites+non-eriophyoid trombidiform mites, depending on data partition strategies. Eriophyoid mites diverged from other mites in the Devonian (384Mya, 95% HPD, 352-410Mya). The origin of eriophyoid mites was dated to the Permian (262Mya, 95% HPD 230-307Mya), mostly prior to the radiation of gymnosperms (Triassic-Jurassic) and angiosperms (early Cretaceous). We propose that the placement of Eriophyoidea in the order Trombidiformes under the current classification system should be reviewed.

Fragmented mitochondrial (mt) genomes have been reported in 11 species of sucking lice (suborder Anoplura) that infest humans, chimpanzees, pigs, horses, and rodents. There is substantial variation among these lice in mt karyotype: the number of minichromosomes of a species ranges from 9 to 20; the number of genes in a minichromosome ranges from 1 to 8; gene arrangement in a minichromosome differs between species, even in the same genus. We sequenced the mt genome of the guanaco louse, Microthoracius praelongiceps, to help establish the ancestral mt karyotype for sucking lice and understand how fragmented mt genomes evolved. The guanaco louse has 12 mt minichromosomes; each minichromosome has 2-5 genes and a non-coding region. The guanaco louse shares many features with rodent lice in mt karyotype, more than with other sucking lice. The guanaco louse, however, is more closely related phylogenetically to human lice, chimpanzee lice, pig lice, and horse lice than to rodent lice. By parsimony analysis of shared features in mt karyotype, we infer that the most recent common ancestor of sucking lice, which lived ∼75 Ma, had 11 minichromosomes; each minichromosome had 1-6 genes and a non-coding region. As sucking lice diverged, split of mt minichromosomes occurred many times in the lineages leading to the lice of humans, chimpanzees, and rodents whereas merger of minichromosomes occurred in the lineage leading to the lice of pigs and horses. Together, splits and mergers of minichromosomes created a very complex and dynamic mt genome organization in the sucking lice.

Extracellular vesicles (EVs) are small membrane vesicles secreted into the extracellular environment to play important roles in intercellular communications. Although there are some examples linking EVs to metastasis, there is a knowledge gaps about the common and unique roles for EVs in different cancer types. These systematic examination across different cancer types might provide the cancer specific metastatic mechanism. To this aim, we performed an integrative analysis on the literatures, thousands of cancer individuals, and reliable ligand-receptor information to present the first global analysis between EVs and metastasis. We identified 97 reliable genes present in EVs that contribute to metastasis with literature evidences. By exploring functional, network, and mutational features of these 97 genes, we found proteoglycans and microRNAs in cancer development might be the key links between EV and cancer metastasis. Most importantly, these 97 genes are significantly associated with cancer patient survival from a combination of 4975 cancer patients from various cancer types. The additional ligand-receptor network were reconstructed to reveal the cellular communication mechanisms for these key EV genes in cancer metastasis. Since the circulating EVs in body fluids could be a candidate for non-invasive liquid biopsy, our results provide a reliable resource of EV-related miRNAs and proteins important for cancer metastasis monitoring.

Background Xinjiang is one of the areas with the highest incidence of cervical cancer in China. Genetic variation in Human papillomavirus type 16 (HPV16) may increase the ability of the virus to mediate carcinogenesis and immune escape, which are risk factors for the progression of cervical cancer. We investigated polymorphism in HPV16 and the distribution of its sub-lineages in the region by analyzing the E6, E7 and long control region (LCR) gene sequences from women with HPV16-positive cervical samples in Xinjiang. Methods A total of 138 cases of cervical lesions and squamous cell carcinoma with infection of HPV16 virus were collected. The E6 and E7 genes and LCR of HPV16 virus were sequenced and compared with the HPV16 European prototype reference and other HPV16 mutants for single nucleotide polymorphisms. Neighbor-joining phylogenetic trees were constructed using E6, E7 and LCR sequences. Results Fourteen missense mutations were found in the E6 gene; the loci with the highest mutation frequency were T350G (36/75, 48%) and T178G (19/75, 25.3%). In the E7 gene, the locus with the highest mutation frequency was A647G (18/75, 24%). A total of 33 polymorphic sites were found in the LCR, of which T7447C (39/95, 40.1%) was the most frequent. Conclusion HPV16 in Xinjiang is mainly of the European variant, followed by the Asian variant type; no Africa 1, 2 or Asia–America variant types were found.

BACKGROUND:Colorectal cancer is an epigenetically heterogeneous disease, however the extent and spectrum of the CpG Island Methylator Phenotype (CIMP) is not clear.RESULTSAn unselected cohort of 216 colorectal cancers clustered into five clinically and molecularly distinct subgroups using Illumina 450K DNA methylation arrays. CIMP-High cancers were most frequent in the proximal colons of female patients. These dichotomised into CIMP-Hl and CIMP-H2 based on methylation profile which was supported by over representation of BRAF (74%, PKRAS (55%, P78). We identified oncogenes susceptible to gene body methylation and Wnt pathway antagonists resistant to gene body methylation. CIMP cluster specific mutations were observed for genes involved in chromatin remodelling, such as in the SWI/SNF and NuRD complexes, suggesting synthetic lethality.CONCLUSIONThere are five clinically and molecularly distinct subgroups of colorectal cancer based on genome wide epigenetic profiling. These analyses highlighted an unidentified role for gene body methylation in progression of serrated neoplasia. Subgroup-specific mutation of distinct epigenetic regulator genes revealed potentially druggable vulnerabilities for these cancers, which may provide novel precision medicine approaches.

Background The sucking louse fauna of endemic Australian rodents has been under-studied for decades. Sixty-five species of native rodents have been recorded in Australia. However, only 11 species of lice have been reported from 11 species of endemic Australian rodents. Results We describe a new species of sucking louse, Hoplopleura villosissima Wang (Psocodea: Phthiraptera: Hoplopleuridae), and report a new host record of the spiny rat louse, Polyplax spinulosa Burmeister, 1839 (Psocodea: Phthiraptera: Polyplacidae), from the long-haired rat, Rattus villosissimus Waite (Rodentia: Muridae), which is endemic to Australia. Conclusions This study is the first record of sucking louse from R. villosissimus and the first record of a species of Polyplax Enderlein, 1904 from an endemic Australian rodent. This study brings the total number of sucking louse species in endemic Australian rodents from 11 to 13. Previously, only the introduced brown rat, Rattus norvegicus Berkenhout and the black rat, Rattus rattus Linnaeus were recorded as the hosts of P. spinulosa in Australia. Because R. villosissimus overlaps with R. rattus in distribution but not with R. norvegicus, we propose that P. spinulosa transferred to R. villosissimus from R. rattus.

Organelle genome fragmentation has been found in a wide range of eukaryotic lineages; however, its use in phylogenetic reconstruction has not been demonstrated. We explored the use of mitochondrial (mt) genome fragmentation in resolving the controversial suborder-level phylogeny of parasitic lice (order Phthiraptera). There are approximately 5000 species of parasitic lice in four suborders (Amblycera, Ischnocera, Rhynchophthirina, and Anoplura), which infest mammals and birds. The phylogenetic relationships among these suborders are unresolved despite decades of studies. We sequenced the mt genomes of eight species of parasitic lice and compared them with 17 other species of parasitic lice sequenced previously. We found that the typical single-chromosome mt genome is retained in the lice of birds but fragmented into many minichromosomes in the lice of eutherian mammals. The shared derived feature of mt genome fragmentation unites the eutherian mammal lice of Ischnocera (family Trichodectidae) with Anoplura and Rhynchophthirina to the exclusion of the bird lice of Ischnocera (family Philopteridae). The novel clade, namely Mitodivisia, is also supported by phylogenetic analysis of mt genome and cox1 gene sequences. Our results demonstrate, for the first time, that organelle genome fragmentation is informative for resolving controversial high-level phylogenies.

This study aimed to explore: 1) DNA methylation in the promoter regions of Wilms tumor gene 1 (WT1), NK6 transcription factor related locus 1 gene (NKX6-1) and Deleted in bladder cancer 1 (DBC1) gene in cervical cancer tissues of Uygur women in Xinjiang, and 2) the correlation of gene methylation with the infection of HPV16/18 viruses. We detected HPV16/18 infection in 43 normal cervical tissues, 30 cervical intraepithelial neoplasia lesions (CIN) and 48 cervical cancer tissues with polymerase chain reaction (PCR) method. Methylation in the promoter regions of the WT1, NKX6-1 and DBC1 genes in the above-mentioned tissues was measured by methylation-specific PCR (MSP) and cloning sequencing. The expression level of these three genes was measured by real-time PCR (qPCR) in 10 methylation-positive cervical cancer tissues and 10 methylation-negative normal cervical tissues. We found that the infection of HPV16 in normal cervical tissues, CIN and cervical cancer tissues was 14.0, 36.7 and 66.7%, respectively. The infection of HPV18 was 0, 6.7 and 10.4%, respectively. The methylation rates of WT1, NKX6-1 and DBC1 genes were 7.0, 11.6 and 23.3% in normal cervical tissues, 36.7, 46.7 and 30.0% in CIN tissues, and 89.6, 77.1 and 85.4% in cervical cancer tissues. Furthermore, WT1, NKX6-1 and DBC1 genes were hypermethylated in the high-grade squamous intraepithelial lesion (CIN2, CIN3) and in the cervical cancer tissues with infection of HPV16/18 (both P< 0.05). The expression of WT1, NKX6-1 and DBC1 was significantly lower in the methylation-positive cervical cancer tissues than in methylation-negative normal cervical tissues. Our findings indicated that methylation in the promoter regions of WT1, NKX6-1 and DBC1 is correlated with cervical cancer tumorigenesis in Uygur women. The infection of HPV16/18 might be correlated with methylation in these genes. Gene inactivation caused by methylation might be related to the incidence and development of cervical cancer.

Table S2. Mitochondrial genome organization of Histiostoma feroniarum. (DOCX 16 kb)

Figure S2. The alignment of nucleotide sequences of nine mitochondrial putative tRNA genes (encoded by trnA, trnE, trnF, trnI, trnQ, trnR, trnS 1 , trnV and trnY) in the Sarcoptiformes mites. The conserved sequences in anticodon loops were marked. (PDF 4582 kb)

Table S1. Mitochondrial genome organization of Rhizoglyphus robini. (DOCX 13 kb)

Table S4. PCR primers used in this study. (DOCX 13 kb)

Table S5. Sarcoptiform mites included in this study. (DOCX 18 kb)

Table S3. Amino acid frequencies of the sarcoptiform mites. (DOCX 16 kb)

Figure S3. Relative synonymous codon usage (RSCU) and codon numbers for five amino acids (H, L2, P, S2 and W) in the mitochondrial genomes of sarcoptiform mites. The X-axis shows the sarcoptiform mites, and the Y-axis shows the RSCU or total number of codons. The blue column indicates the codons that match the anticodons of the corresponding mt tRNA genes. The red, green and purple column indicate the imperfect, synonymous codons to the anticodons of their corresponding mt tRNA genes. (PDF 635 kb)

Colon cancer is one of the most common cancers worldwide with high mortality. A major issue in colon cancer treatment is drug-resistance and metastasis that have been ascribed to the cancer stem cells. In this study, colon cancer stem cells were isolated through sphere culture and verified with the cancer stem cell markers CD133, CD44, and CD24. It was demonstrated that the PI3K/Akt/mTOR signalling pathway was highly activated in the colon cancer stem cells and that inhibition of the PI3K/Akt/mTOR pathway by the inhibitor BEZ235 suppressed the colon cancer stem cell proliferation with reduced stemness indicated by CD133 and Lgr5 expressions. Treatment with insulin as a known activator of the PI3K/Akt pathway increased CD133 expression and decreased the effects of BEZ235 on colon cancer proliferation and survival. The data presented here collectively suggest that the PI3K/Akt/mTOR pathway underpins the stemness of colon cancer stem cells and BEZ235 is potentially a good drug candidate for treatment of colon cancer drug resistance and metastasis.

Cervical cancer is a major disease with high mortality. All cervical cancers are caused by infection with human papillomaviruses (HPV). Although preventive vaccines for cervical cancer are successful, treatment of cervical cancer is far less satisfactory because of multidrug resistance and side effects. In this review, we summarize the recent application of nanotechnology to the diagnosis and treatment of cervical cancer as well as the development of HPV vaccines. Early detection of cervical cancer enables tumours to be efficiently removed by surgical procedures, leading to increased survival rate. The current method of detecting cervical cancer by Pap smear can only achieve 50% sensitivity, whereas nanotechnology has been used to detect HPVs with greatly improved sensitivity. In cervical cancer treatment, nanotechnology has been used for the delivery of anticancer drugs to increase treatment efficacy and decrease side effects. Nanodelivery of HPV preventive and therapeutic vaccines has also been investigated to increase vaccine efficacy. Overall, these developments suggest that nanoparticle-based vaccine may become the most effective way to prevent and treat cervical cancer, assisted or combined with some other nanotechnology-based therapy.

Ghrelin and its synthetic analog hexarelin are specific ligands of growth hormone secretagogue (GHS) receptor. GHS have strong growth hormone-releasing effect and other neuroendocrine activities such as stimulatory effects on prolactin and adrenocorticotropic hormone secretion. Recently, several studies have reported other beneficial functions of GHS that are independent of GH. Ghrelin and hexarelin, for examples, have been shown to exert GH-independent cardiovascular activity. Hexarelin has been reported to regulate peroxisome proliferator-activated receptor gamma (PPAR-γ) in macrophages and adipocytes. PPAR-γ is an important regulator of adipogenesis, lipid metabolism, and insulin sensitization. Ghrelin also shows protective effects on beta cells against lipotoxicity through activation of phosphatidylinositol-3 kinase/protein kinase B, c-Jun N-terminal kinase (JNK) inhibition, and nuclear exclusion of forkhead box protein O1. Acylated ghrelin (AG) and unacylated ghrelin (UAG) administration reduces glucose levels and increases insulin-producing beta cell number, and insulin secretion in pancreatectomized rats and in newborn rats treated with streptozotocin, suggesting a possible role of GHS in pancreatic regeneration. Therefore, the discovery of GHS has opened many new perspectives in endocrine, metabolic, and cardiovascular research areas, suggesting the possible therapeutic application in diabetes and diabetic complications especially diabetic cardiomyopathy. Here, we review the physiological roles of ghrelin and hexarelin in the protection and regeneration of beta cells and their roles in the regulation of insulin release, glucose, and fat metabolism and present their potential therapeutic effects in the treatment of diabetes and diabetic-associated heart diseases.

Rhipicephalus turanicus is an important tick species potentially carrying tick-borne pathogens. Several tick species have obvious subspecies divergence. However few studies aimed to examine the existence of divergence within R. turanicus. Therefore, a detailed morphological and molecular analysis was conducted for comparing R. turanicus from the Mediterranean Basin (represented by Albania) and Central Asia (Northwestern China). Altogether 315 adult ticks of R. turanicus (103 from Albania and 212 from China) were morphologically and molecularly analysed. DNA samples were used for mitochondrial 16S rRNA and cox1 gene sequences analysis. In addition, as potentially genetic markers, three fragments including partial nad1-16S rRNA, nad2-cox1, cox1-tRNA-Lys, were designed and then phylogenetically analyzed. Based on detailed morphological observations, only basis capituli length:width ratio (females), the length, the width and the length:width ratio of the scutum (males) had differences between R. turanicus from China and Albania. Gene divergences of 16S rRNA, cox1, partial nad1-16S rRNA, nad2-cox1 and cox1-tRNA-Lys from China and Albania ticks were 3.53–4.84, 3.57–4.92, 3.57–4.07, 3.57–4.39 and 3.18–4.69%, respectively. The evaluated five genetic markers revealed two phylogenetic branches in R. turanicus. Obvious differences exist within R. turanicus based on morphological and genetic analysis. Three newly designed genetic markers (partial nad1-16S rRNA, nad2-cox1 and cox1-tRNA-Lys) in this study may be suitable genetic tools for identification and analysis in R. turanicus. Subspecies analysis of R. turanicus from other regions of the world should be initiated in the future. Electronic supplementary material The online version of this article (10.1007/s10493-017-0189-8) contains supplementary material, which is available to authorized users.

Primer details for long range PCR. The four long range PCR fragments spaning across the mitochondrial genome are 4.4 kb, 4.0 kb, 3.8 kb and 1.7 kb long. These primers are positioned on four main genes; 12S rRNA gene, nd4, cob and cox1. (DOCX 13 kb)

Additional phylogenetic trees constructed for new NFL mitochondrial genomes. Five different phylogenetic trees were constructed; 1) Log-Det NeightbourNet, 2) Raw (Uncorrected) proportional distance and Neighbour-Net, 3) Tree-inference method with Jukes-Cantor (1969) distances, 4) Neighbour-joining tree using uncorrected distances and 5) Neighbour-joining tree with bootstrap values. (DOCX 343 kb)

Some people host lice on the clothing as well as the head. Whether body lice and head lice are distinct species or merely variants of the same species remains contentious. We sought to ascertain the extent to which lice from these different habitats might interbreed on doubly infected people by comparing their entire mitochondrial genome sequences. Toward this end, we analyzed two sets of published genetic data from double-infections of body lice and head lice: 1) entire mitochondrial coding regions (≈15.4 kb) from body lice and head lice from seven doubly infected people from Ethiopia, China, and France; and 2) part of the cox1 gene (≈486 bp) from body lice and head lice from a further nine doubly infected people from China, Nepal, and Iran. These mitochondrial data, from 65 lice, revealed extraordinary variation in the number of single nucleotide polymorphisms between the individual body lice and individual head lice of double-infections: from 1.096 kb of 15.4 kb (7.6%) to 2 bps of 15.4 kb (0.01%). We detected coinfections of lice of Clades A and C on the scalp hair of three of the eight people from Nepal: one person of the two people from Kathmandu and two of the six people from Pokhara. Lice of Clades A and B coinfected the scalp hair of one person from Atherton, Far North Queensland, Australia. These findings argue for additional large-scale studies of the body lice and head lice of double-infected people.

We assessed the suitability of HS3ST2 and CCNA1 genes as biomarkers for the early detection of cervical cancer in Uygur women in Xinjiang, China. Methylation-specific PCR (MSP) and HPV (HPV16 and HPV18)–specific PCR were performed on 110 cervical samples: 40 normal cervices, 10 cervical intraepithelial neoplasia 1 (CIN1), 10 CIN2, 10 CIN3 and 40 cervical cancer tissues. The expression of the 2 genes was measured by reverse transcription PCR (RT-PCR) in 10 methylation-positive and 10 methylation-negative cervical tissues. We found that both HS3ST2 and CCNA1 genes were methylated in 38 of the 40 cervical cancer tissues, 9 of the 10 CIN3, and 6 of the 10 CIN2. In contrast, methylation of these 2 genes was found in only 1 of the 40 normal tissues and none of 10 CIN1. Furthermore, hypermethylated HS3ST2 and CCNA1 genes were correlated with infection with HPV16 and HPV18 in high-grade squamous intraepithelial lesions (HSILs) and cervical cancer (both p

Endometrial cancer is the third most common cancer in women. Endometrial carcinomas (EC) are clinic histologically classified into two types. Type I tumours, which account for 80% of ECs, are estrogen-dependent and are low grade. Type II tumours are more aggressive with invasion into myometrium. Recently a new classification for endometrial cancer has been proposed based on molecular markers. Whether this classification is helpful for clinical management of endometrial cancer remains to be tested. At present, treatment outcomes of endometrial cancer are not satisfactory. Therefore, more effective approaches are sought. This review summarizes the recent studies about activation of PI3K/Akt pathway in EC and therapeutic implications of the inhibitors of the pathways with emphasis on dual inhibitors of PI3K and mTOR. Both genetic defects and environmental factors are involved in carcinogenesis and progression of EC via activation of multiple signal pathways including the PI3K/Akt pathway. Mutations of major components of the PI3K/Akt pathway are common in EC. Type I tumours usually have mutations in Ras, PTEN, PIK3CA, AKT1, beta-catenin and type II tumours have mutations in TP53. Environmental factor like obesity can also activate the PI3K/Akt pathway to increase the incidence of EC and to cause poorer prognosis. Therefore, inhibition of the PI3K/Akt pathway can be used for therapy of the disease. At present, mTOR inhibitors have been extensively studied and tested in clinical trials. A newly synthesised dual inhibitor of PI3K and mTOR BEZ235 has been shown to be more effective than mTOR inhibitor rapamycin. BEZ235 can inhibit feedback activation of PI3K/Akt pathway by rapamycin. It is promising to include effective PI3K/Akt inhibitors in current treatment regime of endometrial cancer to improve the therapeutic efficacy.

Over the past several years, there was a substantial increase in the number of cases of known and novel tick-borne infections in humans in China. To better understand the ticks associated with these infections, we collected hard ticks from animals or around livestock shelters in 29 localities in 5 provinces (Beijing, Henan, Hubei, Inner Mongolia, and Zhejiang) where cases of tick-borne illness were reported. We collected 2950 hard ticks representing 7 species of 4 genera (Dermacentor sinicus, Haemaphysalis flava, Haemaphysalis longicornis, Ixodes granulatus, Ixodes persulcatus, Rhipicephalus microplus, and Rhipicephalus sanguineus). These ticks were identified to species using morphological characters initially. We then sequenced the mitochondrial small subunit rRNA (12S rRNA) gene, cytochrome oxidase subunit 1 (COI) gene, and the second internal transcribed spacer (ITS2) gene of these ticks, and conducted phylogenetic analyses. Our analyses showed that the molecular and morphological data are consistent in the identification of the 7 tick species. Furthermore, all these 7 tick species from China were genetically closely related to the same species or related species found outside China. Rapid and accurate identification and long-term monitoring of these ticks will be of significance to the prevention and control of tick-borne diseases in China.

We sequenced the entire mitochondrial genomes of 3 species of metastriate ticks: Haemaphysalis formosensis, H. parva, and Amblyomma cajennense. We also sequenced two thirds (ca. 9500bp) of the mitochondrial genomes of H. humerosa and H. hystricis. We used these 5 mitochondrial genome sequences together with the 13 tick mitochondrial genomes we sequenced previously and the 2 tick mitochondrial genomes sequenced by Black and Roehrdanz (1998), as well as the nuclear rRNA genes from 84 ticks and mites, in phylogenetic analyses. Our analyses reveal deep phylogenetic structure within the genus Haemaphysalis, with at least 2 species, H. parva and H. inermis that are highly divergent from the rest of the genus Haemaphysalis. We identify a region of the 18S rRNA gene which correlates with this division of the genus Haemaphysalis as well as a novel insertion in the mitochondrial genome of H. parva. We reject the hypotheses of Hoogstraal and Aeschlimann (1982) and Barker and Murrell (2004) on the relationships among metastriate genera. Instead, our analysis provides further evidence for the division of the Metastriata into 2 major lineages: (i) Amblyomma s.s. plus Rhipicephalinae (i.e. Rhipicephalus, Hyalomma, Rhipicentor, and Dermacentor); and (ii) Haemaphysalis plus Bothriocroton plus Amblyomma sphenodonti. We also provide further evidence for the polyphyly of the genus Amblyomma with respect to A. sphenodonti and A. elaphense. The most likely position of A. elaphense is sister to the rest of the Metastriata; the most likely position of A. sphenodonti is sister to the genus Bothriocroton. These 2 species do not belong in the genus Amblyomma, and we propose that new genera are required for A. sphenodonti and A. elaphense.

Ascaridomorph nematodes threaten the health of humans and other animals worldwide. Despite their medical, veterinary and economic importance, the identification of species lineages and establishing their phylogenetic relationships have proved difficult in some cases. Many working hypotheses regarding the phylogeny of ascaridomorphs have been based on single-locus data, most typically nuclear ribosomal RNA. Such single-locus hypotheses lack independent corroboration, and for nuclear rRNA typically lack resolution for deep relationships. As an alternative approach, we analyzed the mitochondrial (mt) genomes of anisakids (~14 kb) from different fish hosts in multiple countries, in combination with those of other ascaridomorphs available in the GenBank database. The circular mt genomes range from 13,948-14,019 bp in size and encode 12 protein-coding genes, 2 ribosomal RNAs and 22 transfer RNA genes. Our analysis showed that the Pseudoterranova decipiens complex consists of at least six cryptic species. In contrast, the hypothesis that Contracaecum ogmorhini represents a complex of cryptic species is not supported by mt genome data. Our analysis recovered several fundamental and uncontroversial ascaridomorph clades, including the monophyly of superfamilies and families, except for Ascaridiidae, which was consistent with the results based on nuclear rRNA analysis. In conclusion, mt genome analysis provided new insights into the phylogeny and taxonomy of ascaridomorph nematodes.

Background Cervical cancer is a major cause of death in women worldwide. Interferon-induced transmembrane protein 1 (IFITM1) is involved in antivirus defense, cell adhesion, and carcinogenesis in different tissues. However, the role of IFITM1 gene in cervical squamous cell cancer is unclear. Methods To explore the role of IFITM1 in carcinogenesis of cervical cancer, we investigated the expression of IFITM1 gene in cervical squamous cell carcinoma. IFITM1 mRNA level was measured by real-time quantitative RT-PCR in cervical cancer tissues and their adjacent normal tissues. IFITM1 protein level was measured by immunohistochemistry. Methylation in the IFITM1 gene promoter was detected by methylation-specific PCR. We then transfected HeLa cells with IFITM1 expression vector or control vector. IFITM1 expression was examined; cell migration and invasion were analyzed by wound healing assay and matrigel-coated transwell migration assays, respectively. HeLa cell proliferation was measured by cell counting kit-8 assay and cell cycle analysis. Cell apoptosis was analyzed by Annexin V/propidium iodide double staining assay. Results The difference in IFITM1 protein expression between samples from chronic cervicitis and cervical carcinoma was statistically significant (P

Background Persistent infection of high-risk human papillomaviruses 16 (HPV16) has been considered as the leading cause of cervical cancer. In this study we assessed HPV16 sequence variation and genetic diversity of HPV16 variants in cervical cancer in Uigur women in Xinjiang, China. We analyzed the nucleotide sequences of the open reading frames of E6 and E7, and part of the open reading frames of L1 of HPV16 in Uigur women. Methods Biopsies of histologically confirmed HPV16 infections with cervical cancer were obtained from 43 Uigur women in Xinjiang, China. E6, E7 and L1 genes of HPV16 of all samples were amplified and sequenced; the sequences were used in phylogenetic analysis of HPV16 variants. Results Our analysis revealed nine nucleotide changes in E6 (five changes), E7 (one change) and L1 (three changes) gene. The most frequently observed variations were T350G (79.1 %). One variation T295G (D64E) at E6 were detected in 6 cases (KT959536, KT959542, KT959546, KT959550, KT959553, KT959558). Deletion (464Asp) along with insertion (448Ser) were observed in L1 (100 %). Most variants were European lineage (97.7 %); only one belongs to Asia variants with common T178G (D25E) in E6 and A647G (N29S) in E7. Conclusion The most prevalent HPV16 variants in the Uigur women we studied were of the European lineage. Our results indicate that HPV16 European lineage may serve as a harmful factor associated with the development and progression of cervical cancer. Electronic supplementary material The online version of this article (doi:10.1186/s13027-016-0089-2) contains supplementary material, which is available to authorized users.

The subclass Acari (mites and ticks) comprises two super-orders: Acariformes and Parasitiformes. Most species of the Parasitiformes known retained the ancestral pattern of mitochondrial (mt) gene arrangement of arthropods and their mt tRNAs have the typical cloverleaf structure. All of the species of the Acariformes known, however, have rearranged mt genomes and truncated mt tRNAs. We sequenced the mt genomes of two species of Eriophyoidea: Phyllocoptes taishanensis and Epitrimerus sabinae. The mt genomes of P. taishanensis and E. sabinae are 13,475 bp and 13,531 bp, respectively, are circular and contain the 37 genes typical of animals; most mt tRNAs are highly truncated in both mites. On the other hand, these two eriophyoid mites have the least rearranged mt genomes seen in the Acariformes. Comparison between eriophyoid mites and other Aacariformes mites showed that: 1) the most recent common ancestor of Acariformes mites retained the ancestral pattern of mt gene arrangement of arthropods with slight modifications; 2) truncation of tRNAs for cysteine, phenylalanine and histidine occurred once in the most recent common ancestor of Acariformes mites whereas truncation of other tRNAs occurred multiple times; and 3) the placement of eriophyoid mites in the order Trombidiformes needs to be reviewed.

Phylogenetic trees of HPV16 E6, E7 and L1 variants based on Neighbor Joining Molecular Phylogenetic analysis. Figure S1. Neighbor Joining Molecular Phylogenetic analysis using 43 nucleotide sequences of HPV16 E6 gene. Phylogenetic studies were performed on E6 nucleotide sequence alignment of 477 positions from each case, which was constructed by the neighbor joining method and the Kimura 2-Parameter model by MEGA 6 package. Bootstrap proportions were calculated with 1000 replicates. Study sequences are labeled in KT GenBank accession numbers, others are reference GenBank sequences. E, European variant; Ep, European prototype; As, Asia lineage; AA, Asian American lineage; Af, African lineage. Figure S2. Neighbor Joining Molecular Phylogenetic analysis using 43 nucleotide sequences of HPV16 E7 gene. Phylogenetic studies were performed on E7 nucleotide sequence alignment of 297 positions from each case, which was constructed by the neighbor joining method and the Kimura 2-Parameter model by MEGA 6 package. Bootstrap proportions were calculated with 1000 replicates. Study sequences are labeled in KT GenBank accession numbers, others are reference GenBank sequences. E, European variant; Ep, European prototype; As, Asia lineage; AA, Asian American lineage; Af, African lineage. Figure S3. Neighbor Joining Molecular Phylogenetic analysis using 39 nucleotide sequences of HPV16 L1 gene. Phylogenetic studies were performed on partial L1 nucleotide sequence alignment of 369 positions from each case, which was constructed by the neighbor joining method and the Kimura 2-Parameter model by MEGA 6 package. Bootstrap proportions were calculated with 1000 replicates. Study sequences are labeled in KT GenBank accession numbers, others are reference GenBank sequences. E, European variant; Ep, European prototype; As, Asia lineage; AA, Asian American lineage; Af, African lineage. (PDF 136 kb)

Ticks transmit more pathogens to humans and animals than any other arthropod. We describe the 2.1 Gbp nuclear genome of the tick, Ixodes scapularis (Say), which vectors pathogens that cause Lyme disease, human granulocytic anaplasmosis, babesiosis and other diseases. The large genome reflects accumulation of repetitive DNA, new lineages of retro-transposons, and gene architecture patterns resembling ancient metazoans rather than pancrustaceans. Annotation of scaffolds representing ∼57% of the genome, reveals 20,486 protein-coding genes and expansions of gene families associated with tick–host interactions. We report insights from genome analyses into parasitic processes unique to ticks, including host ‘questing’, prolonged feeding, cuticle synthesis, blood meal concentration, novel methods of haemoglobin digestion, haem detoxification, vitellogenesis and prolonged off-host survival. We identify proteins associated with the agent of human granulocytic anaplasmosis, an emerging disease, and the encephalitis-causing Langat virus, and a population structure correlated to life-history traits and transmission of the Lyme disease agent., This project has been funded in part with federal funds from the National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services (NIAID, NIH, DHHS) under contract numbers N01-AI30071, HHSN272200900007C, HHSN266200400001C and 5R01GM77117-5. Its contents are solely the responsibility of the authors and do not represent the official views of the NIH. Additional grants and contracts supporting work described in this manuscript were from the NIH-NIAID (HHSN266200400039C and HHSN272200900039C) to F.H.C., and a subcontract under HHSN272200900039C to C.A.H. and J.M.M., the Australian Research Council Discovery Project (DP120100240) to S.C.B. and R.S., the Ministerio de Ciencia e Innovación of Spain (BFU2007–6292; BFU2010–15484) to J.R., BIO2009–07990 and BIO2012–37926 to J.V. NIH-1R01AI090062 to Y.P., L.S., and J.K., NIH 1R21AI096268 and NSF IOS-0949194 to R.M.R., the Xunta de Galicia of Spain (10PXIB918057PR) to J.M.C.T. and M.T., BFU2011–23896 and EU FP7 ANTIGONE (278976) to J.F., the USDA-NRI/CREES (2008-35302-18820) and Texas AgriLife Research Vector Biology grant to P.V.P. and European Research Council Starting Independent Researcher Grant (205202) to R.B., J.M.R was supported by the intramural program of the NIAID, R.M.W. by a Marie Curie International Outgoing Fellowship PIOF-GA-2011–303312, E.M.Z. by Swiss National Science Foundation awards 31003A-125350 and 31003A-143936, J.M.G. by an NIH-NCATS award TL1 TR000162 and NSF Graduate Research Fellowship (DGE 1333468), V.C. by a Boehringer Ingelheim Ph.D. Fellowship, F.G.V. by a Fundação para a Ciência e a Tecnologia, Portugal fellowship (SFRH/BD/22360/2005), C.J.P.G. and F.H. by The Lundbeck Foundation (Denmark), and J.J.G. by NIH awards HHSN272200900040C, R01AI017828 and R01AI043006. Support from the Broad Genomics Platform is gratefully acknowledged.

Bilateral animals are featured by an extremely compact mitochondrial (mt) genome with 37 genes on a single circular chromosome. The human body louse, Pediculus humanus, however, has its mt genes on 20 minichromosomes. We sequenced the mt genomes of two other human lice: the head louse, P. capitis, and the pubic louse, Pthirus pubis. Comparison among the three human lice revealed the presence of fragmented mt genomes in their most recent common ancestor, which lived � 7 Ma. The head louse has exactly the same set of mt minichromosomes as the body louse, indicating that the number of minichromosomes, and the gene content and gene arrangement in each minichromosome have remained unchanged since the body louse evolved from the head louse � 107,000 years ago. The pubic louse has the same pattern of one protein-coding or rRNA gene per minichromosome (except one minichromosome with two protein-coding genes, atp6 and atp8) as the head louse and the body louse. This pattern is apparently ancestral to all human lice and has been stable for at least 7 Myr. Most tRNA genes of the pubic louse, however, are on different minichromosomes when compared with their counterparts in the head louse and the body louse. It is evident that rearrangement of four tRNA genes (for leucine, arginine and glycine) was due to gene-identity switch by point mutation at the third anticodon position or by homologous recombination, whereas rearrangement of other tRNA genes was by gene translocation between minichromosomes, likely caused by minichromosome split via gene degeneration and deletion.

Our understanding of the phylogenetic relationships among tick lineages has been limited by the lack of resolution provided by the most commonly used phylogenetic markers. Mitochondrial genomes are increasingly used to address controversial phylogenetic relationships. To date, the complete mitochondrial genomes of eleven tick species have been sequenced; however, only three of these species are metastriate ticks, the most speciose lineage of ticks. In this study, we present the nucleotide sequences of the complete mitochondrial genomes of five more species of metastriate ticks: Amblyomma elaphense, Amblyomma fimbriatum, Amblyomma sphenodonti, Bothriocroton concolor and Bothriocroton undatum. We use complete mitochondrial genome sequences to address the phylogenetic placement of two morphologically 'primitive' species -Am. elaphense and Am. sphenodonti - with respect to the genus Amblyomma. Our analysis of these five mitochondrial genomes with the other eleven tick mitochondrial genomes, as well as analysis of nuclear rRNA genes, provides strong evidence that the genus Amblyomma is polyphyletic with the inclusion of Am. sphenodonti and Am. elaphense. A new genus or two new genera may be required to describe Am. sphenodonti and Am. elaphense. It is also possible that these two species are sisters to two established genera, Bothriocroton in the case of Am. sphenodonti, and Haemaphysalis in the case of Am. elaphense. However, other arrangements of these taxa cannot be excluded with the current data. Thus, while Am. sphenodonti and Am. elaphense do not belong in the genus Amblyomma, the phylogenetic placement of these two species cannot be resolved without more data from metastriate ticks, either greater sampling of mitochondrial genomes, or a large data set of nuclear genes.

The mitochondrial (mt) genome of bilateral animals typically consists of a single chromosome, ~16 kb long, and contains 37 genes. Variation in mt gene sequence within an individual, i.e. heteroplasmy, is common in bilateral animals but usually occurs at very low levels. In the lineage that led to the human body louse, the typical mt chromosome has fragmented into 18 minichromosomes: each minichromosome is 3 to 4 kb long and contains 1 to 3 genes and a non- coding region. To understand the presence and the level of heteroplasmy in fragmented mt genomes, we cloned and sequenced seven copies of full-length cox1-minichromosome of a human body louse and a human head louse. We found 17 heteroplasmic sites in the coding region and 118 heteroplasmic sites in the non-coding region of the cox1- minichromosome. The level of heteroplasmy in the human lice appears to be much higher than that in other animals that have the typical mt genome. We propose that recombination between different minichromosomes of the fragmented mt genome may contribute to the high-level of heteroplasmy in the human lice.

The mitochondrial (mt) genome of the human body louse, Pediculus humanus, consists of 18 minichromosomes. Each minichromosome is 3 to 4 kb long and has 1 to 3 genes. There is unequivocal evidence for recombination between different mt minichromosomes in P. humanus. It is not known, however, how these minichromosomes recombine. Here, we report the discovery of eight chimeric mt minichromosomes in P. humanus. We classify these chimeric mt minichromosomes into two groups: Group I and Group II. Group I chimeric minichromosomes contain parts of two different protein-coding genes that are from different minichromosomes. The two parts of protein-coding genes in each Group I chimeric minichromosome are joined at a microhomologous nucleotide sequence; microhomologous nucleotide sequences are hallmarks of non-homologous recombination. Group II chimeric minichromosomes contain all of the genes and the non-coding regions of two different minichromosomes. The conserved sequence blocks in the non-coding regions of Group II chimeric minichromosomes resemble the “recombination repeats” in the non-coding regions of the mt genomes of higher plants. These repeats are essential to homologous recombination in higher plants. Our analyses of the nucleotide sequences of chimeric mt minichromosomes indicate both homologous and non-homologous recombination between minichromosomes in the mitochondria of the human body louse.

As an obligatory parasite of humans, the body louse ( Pediculus humanus humanus ) is an important vector for human diseases, including epidemic typhus, relapsing fever, and trench fever. Here, we present genome sequences of the body louse and its primary bacterial endosymbiont Candidatus Riesia pediculicola. The body louse has the smallest known insect genome, spanning 108 Mb. Despite its status as an obligate parasite, it retains a remarkably complete basal insect repertoire of 10,773 protein-coding genes and 57 microRNAs. Representing hemimetabolous insects, the genome of the body louse thus provides a reference for studies of holometabolous insects. Compared with other insect genomes, the body louse genome contains significantly fewer genes associated with environmental sensing and response, including odorant and gustatory receptors and detoxifying enzymes. The unique architecture of the 18 minicircular mitochondrial chromosomes of the body louse may be linked to the loss of the gene encoding the mitochondrial single-stranded DNA binding protein. The genome of the obligatory louse endosymbiont Candidatus Riesia pediculicola encodes less than 600 genes on a short, linear chromosome and a circular plasmid. The plasmid harbors a unique arrangement of genes required for the synthesis of pantothenate, an essential vitamin deficient in the louse diet. The human body louse, its primary endosymbiont, and the bacterial pathogens that it vectors all possess genomes reduced in size compared with their free-living close relatives. Thus, the body louse genome project offers unique information and tools to use in advancing understanding of coevolution among vectors, symbionts, and pathogens.