Mitogenome-based phylogeny of mosquitoes (Diptera: Culicidae).

Mosquitoes are of great medical significance as vectors of many deadly diseases. Mitogenomes have been widely used in phylogenetic studies, but mitogenome knowledge within the family Culicidae is limited, and Culicidae phylogeny is far from resolved. In this study, we surveyed the mitogenomes of 149 Culicidae species, including 7 newly sequenced species. Comparative analysis of 149 mosquito mitogenomes shows gene composition and order to be identical to that of an ancestral insect, and the AT bias, length variation, and codon usage are all consistent with that of other reported Dipteran mitogenomes. Phylogenetic analyses based on the DNA sequences of the 13 protein-coding genes from the 149 species robustly support the monophyly of the subfamily Anophelinae and the tribes Aedini, Culicini, Mansoniini, Sabethini, and Toxorhynchitini. To resolve ambiguous relationships between clades within the subfamily Culicinae, we performed topological tests and show that Aedini is a sister to Culicini and that Uranotaeniini is a sister to (Mansoniini + (Toxorhynchitini + Sabethini)). In addition, we estimated divergence times using a Bayesian relaxation clock based on the sequence data and 3 fossil calibration points. The results show mosquitoes diverged during the Early Jurassic with massive Culicinae radiations during the Cretaceous, coincident with the emergence of angiosperms and the burst of mammals and birds. Overall, this study, which uses the largest number of Culicidae mitogenomes sequenced to date, comprehensively reveals the mitogenome characteristics and mitogenome-based phylogeny and divergence times of Culicidae, providing information for further studies on the mitogenome, phylogeny, evolution, and taxonomic revision of Culicidae.

Complete mitogenomes of Anopheles peditaeniatus and Anopheles nitidus and phylogenetic relationships within the genus Anopheles inferred from mitogenomes.

BACKGROUND: Despite the medical importance of mosquitoes of the genus Anopheles in the transmission of malaria and other human diseases, its phylogenetic relationships are not settled, and the characteristics of mitochondrial genome (mitogenome) are not thoroughly understood. METHODS: The present study sequenced and analyzed the complete mitogenomes of An. peditaeniatus and An. nitidus, investigated genome characteristics, and inferred the phylogenetic relationships of 76 Anopheles spp. RESULTS: The complete mitogenomes of An. peditaeniatus and An. nitidus are 15,416 and 15,418 bp long, respectively, and both include 13 PCGs, 22 tRNAs, two tRNAs and one control region (CR). Mitogenomes of Anopheles spp. are similar to those of other insects in general characteristics; however, the trnR and trnA have been reversed to "trnR-trnA," as has been reported in other mosquito genera. Genome variations mainly occur in CR length (493-886 bp) with six repeat unit types identified for the first time that demonstrate an evolutionary signal. The subgenera Lophopodomyia, Stethomyia, Kerteszia, Nyssorhynchus, Anopheles and Cellia are inferred to be monophyletic, and the phylogenetic analyses support a new phylogenetic relationship among the six subgenera investigated, in that subgenus Lophopodomyia is the sister to all other five subgenera, and the remaining five subgenera are divided into two clades, one of which is a sister-taxon subgenera Stethomyia + Kerteszia, and the other consists of subgenus Nyssorhynchus as the sister to a sister-group subgenera Anopheles + Cellia. Four series (Neomyzomyia, Pyretophorus, Neocellia and Myzomyia) of the subgenus Cellia, and two series (Arribalzagia and Myzorhynchus) of the subgenus Anopheles were found to be monophyletic, whereas three sections (Myzorhynchella, Argyritarsis and Albimanus) and their subdivisions of the subgenus Nyssorhynchus were polyphyletic or paraphyletic. CONCLUSIONS: The study comprehensively uncovered the characteristics of mitogenome and the phylogenetics based on mitogenomes in the genus Anopheles, and provided information for further study on the mitogenomes, phylogenetics and taxonomic revision of the genus.

The impact of COVID-19 on short-term prognosis of ST-segment elevation myocardial infarction patients receiving primary percutaneous coronary intervention in Wuhan China.

ST-segment elevation myocardial infarction (STEMI) is a common cardiovascular emergency for which timely reperfusion therapies are needed to minimize myocardial necrosis. The aim of this study was to investigate the impact of the COVID-19 pandemic and reorganization of chest pain centers (CPC) on the practice of primary percutaneous coronary intervention (PPCI) and prognosis of STEMI patients. This single-center retrospective survey included all patients with STEMI admitted to our CPC from January 22, 2020 to April 30, 2020 (during COVID-19 pandemic in Wuhan), compared with those admitted during the analogous period in 2019, in respect of important time points of PPCI and clinical outcomes of STEMI patients. In the present article, we observed a descending trend in STEMI hospitalization and a longer time from symptom onset to first medical contact during the COVID-19 pandemic as compared to the control period (4.35 h versus 2.58 h). With a median delay of 17 minutes in the door to balloon time (D2B), the proportion of in-hospital cardiogenic shock was significantly higher in the COVID-19 era group (47.6% versus 19.5%), and major adverse cardiac events (MACE) tend to increase in the 6-month follow-up period (14.3% versus 2.4%). Although the reorganization of CPC may prolong the D2B time, immediate revascularization of the infarct-related artery could be offered to most patients within 90 minutes upon arrival. PPCI remained the preferred treatment for patients with STEMI during COVID-19 pandemic in the context of timely implementation and appropriate protective measures.

The complete mt genomes of Lutzia halifaxia, Lt. fuscanus and Culex pallidothorax (Diptera: Culicidae) and comparative analysis of 16 Culex and Lutzia mt genome sequences.

BACKGROUND: Despite the medical importance of the genus Culex, the mitochondrial genome (mt genome) characteristics of Culex spp. are not well understood. The phylogeny of the genus and particularly the generic status of the genus Lutzia and the subgenus Culiciomyia remain unclear. METHODS: The present study sequenced and analyzed the complete mt genomes of Lutzia halifaxia, Lutzia fuscanus and Cx. (Culiciomyia) pallidothorax and assessed the general characteristics and phylogenetics of all known 16 mt genome sequences for species in the genera Culex and Lutzia. RESULTS: The complete mt genomes of Lt. halifaxia, Lt. fuscanus and Cx. pallidothorax are 15,744, 15,803 and 15,578 bp long, respectively, including 13 PCGs, 22 tRNAs, two tRNAs and a control region (CR). Length variations in the Culex and Lutzia mt genomes involved mainly the CR, and gene arrangements are the same as in other mosquitoes. We identified four types of repeat units in the CR sequences, and the poly-T stretch exists in all of these mt genomes. The repeat units of CR are conserved to different extent and provide information on their evolution. Phylogenetic analyses demonstrated that the Coronator and Sitiens groups are each monophyletic, whereas the monophyletic status of the Pipiens Group was not supported; Cx. pallidothorax is more closely related to the Sitiens and Pipiens groups; and both phylogenetics analysis and repeat unit features in CR show that Lutzia is a characteristic monophyletic entity, which should be an independent genus. CONCLUSIONS: To our knowledge, this is the first comprehensive review of the mt genome sequences and taxonomic discussion based on the mt genomes of Culex spp. and Lutzia spp. The research provides general information on the mt genome of these two genera, and the phylogenetic and taxonomic status of Lutzia and Culiciomyia.

UDP-glycosyltransferase genes and their association and mutations associated with pyrethroid resistance in Anopheles sinensis (Diptera: Culicidae).

BACKGROUND: UDP-glycosyltransferase (UGT) is an important biotransformation superfamily of enzymes. They catalyze the transfer of glycosyl residues from activated nucleotide sugars to acceptor hydrophobic molecules, and function in several physiological processes, including detoxification, olfaction, cuticle formation, pigmentation. The diversity, classification, scaffold location, characteristics, phylogenetics, and evolution of the superfamily of genes at whole genome level, and their association and mutations associated with pyrethroid resistance are still little known. METHODS: The present study identified UGT genes in Anopheles sinensis genome, classified UGT genes in An. sinensis, Anopheles gambiae, Aedes aegypti and Drosophila melanogaster genomes, and analysed the scaffold location, characteristics, phylogenetics, and evolution of An. sinensis UGT genes using bioinformatics methods. The present study also identified the UGTs associated with pyrethroid resistance using three field pyrethroid-resistant populations with RNA-seq and RT-qPCR, and the mutations associated with pyrethroid resistance with genome re-sequencing in An. sinensis. RESULTS: There are 30 putative UGTs in An. sinensis genome, which are classified into 12 families (UGT301, UGT302, UGT306, UGT308, UGT309, UGT310, UGT313, UGT314, UGT315, UGT36, UGT49, UGT50) and further into 23 sub-families. The UGT308 is significantly expanded in gene number compared with other families. A total of 119 UGTs from An. sinensis, An. gambiae, Aedes aegypti and Drosophila melanogaster genomes are classified into 19 families, of which seven are specific for three mosquito species and seven are specific for Drosophila melanogaster. The UGT308 and UGT302 are proposed to main families involved in pyrethroid resistance. The AsUGT308D3 is proposed to be the essential UGT gene for the participation in biotransformation in pyrethroid detoxification process, which is possibly regulated by eight SNPs in its 3' flanking region. The UGT302A3 is also associated with pyrethroid resistance, and four amino acid mutations in its coding sequences might enhance its catalytic activity and further result in higher insecticide resistance. CONCLUSIONS: This study provides the diversity, phylogenetics and evolution of UGT genes, and potential UGT members and mutations involved in pyrethroid resistance in An. sinensis, and lays an important basis for the better understanding and further research on UGT function in defense against insecticide stress.

Comparative genomics of chemosensory protein genes (CSPs) in twenty-two mosquito species (Diptera: Culicidae): Identification, characterization, and evolution.

Chemosensory proteins (CSP) are soluble carrier proteins that may function in odorant reception in insects. CSPs have not been thoroughly studied at whole-genome level, despite the availability of insect genomes. Here, we identified/reidentified 283 CSP genes in the genomes of 22 mosquitoes. All 283 CSP genes possess a highly conserved OS-D domain. We comprehensively analyzed these CSP genes and determined their conserved domains, structure, genomic distribution, phylogeny, and evolutionary patterns. We found an average of seven CSP genes in each of 19 Anopheles genomes, 27 CSP genes in Cx. quinquefasciatus, 43 in Ae. aegypti, and 83 in Ae. albopictus. The Anopheles CSP genes had a simple genomic organization with a relatively consistent gene distribution, while most of the Culicinae CSP genes were distributed in clusters on the scaffolds. Our phylogenetic analysis clustered the CSPs into two major groups: CSP1-8 and CSE1-3. The CSP1-8 groups were all monophyletic with good bootstrap support. The CSE1-3 groups were an expansion of the CSP family of genes specific to the three Culicinae species. The Ka/Ks ratios indicated that the CSP genes had been subject to purifying selection with relatively slow evolution. Our results provide a comprehensive framework for the study of the CSP gene family in these 22 mosquito species, laying a foundation for future work on CSP function in the detection of chemical cues in the surrounding environment.

Complete mitochondrial genomes of Anopheles stephensi and An. dirus and comparative evolutionary mitochondriomics of 50 mosquitoes.

To better understand the phylogeny and evolution of mosquitoes, the complete mitochondrial genome (mitogenome) of Anopheles stephensi and An. dirus were sequenced and annotated, and a total of 50 mosquito mitogenomes were comparatively analyzed. The complete mitogenome of An. stephensi and An. dirus is 1,5371 bp and 1,5406 bp long, respectively. The main features of the 50 mosquito mitogenomes are conservative: 13 protein-coding genes (PCGs), two ribosomal RNA genes, 22 transfer RNA genes, positive AT-skew and negative GC-skew. The gene order trnA-trnR in ancestral insects is rearranged. All tRNA genes have the typical clover leaf secondary structure but tRNA Ser . The control regions are highly variable in size. PCGs show signals of purifying selection, but evidence for positive selection in ND2, ND4 and ND6 is found. Bayesian and Maximum Likelihood phylogenetic analyses based on all PCG nucleotides produce an identical tree topology and strongly support the monophyly of subgenera Cellia, Anopheles, Keterszia and Nyssorhynchus, the sister relationship of the subgenera Nyssorhynchus and Keterszia, and Cellia and Anopheles. The most recent ancestor of the genus Anopheles and Culicini + Aedini exited ~145 Mya ago. This is the first comprehensive study of mosquito mitogenomes, which are effective for mosquito phylogeny at various taxonomic levels.

Sequencing and analysis of the complete mitochondrial genome of Culex gelidus (Diptera: Culicidae).

In this study, we sequenced and analyzed the complete mitochondrial genome of Culex gelidus. The mitogenome is 15,600 bp long, and contains 13 protein-coding genes (PCGs), 22 tRNA genes, two rRNA genes, and a control region. The gene order and composition are identical with those mitogenomes reported in other mosquito species. The whole base composition is A (39.8%), T (39.2%), G (8.8%), and C (12.2%). The PCGs have the initiation codon ATN except for COI with a TCG, and possess the complete termination codon TAA or incomplete T. The phylogenetic analysis of known Culex mitogenome sequences was carried out based on the nucleotide sequences of 13 PCGs, and the result showed that Cx. P. pipiens, Cx. P. pallens, and Cx. p. quinquefasciatus were claded into the Pipiens Complex in the Pipiens Group, and Cx. gelidus might be inappropriate to be classified into the Pipiens Group. Whether the mosquito species group with Cx. gelidus as type should be regarded as a Gelidus Group or Subgroup and its taxonomic position need to be elucidated with more molecular data.

Sequencing and analysis of the complete mitochondrial genome in Anopheles culicifacies species B (Diptera: Culicidae).

The complete mitochondrial genome sequence of Anopheles culicifacial species B was sequenced in this study. The length of the mitochondrial genome is 15 330 bp, which contains 13 protein-coding genes (PCGs), 22 tRNA genes, two rRNA genes, and a non-coding control region. The gene order and the gene composition are consistent with those previously reported for other mosquito species. The initiation codon of the PCGs complies with the ATN rule except for COI using TCG and ND5 using GTG as a start codon, and the termination codon is TAA or imcomplete, an only T. The total base composition is 40.4% A, 38.1% T, 12.4% C, and 9.1% G. The phylogenetic tree based on the sequences of 13 protein-coding genes showed that these species were classified into two clades, corresponding to the subgenus Cellia and subgenus Nyssorhynchus. An. culicifacies species B of Myzomyia Series was clustered with An. gambiae of Pyretophorus Series with a high bootstrap value of 100%. The complete mitogenome data can provide a basis for molecular identification and phylogenetic studies of mosquito species.

Compaction dynamics of single DNA molecules under tension.

Using transverse magnetic tweezers, we studied the dynamics of DNA compaction induced by hexaammine cobalt chloride under constant forces. Discontinuous DNA compaction events were found for forces ranging from 0.5 to 1.7 pN, with approximately 270 nm DNA adsorbed in each compaction event. Forces larger than 6 pN were found able to unravel the toroid in a similar intermittent stepwise manner. The observations indicate that the folding/unfolding events are transitions between two metastable structural states which are separated by a tension-dependent energy barrier. Analysis of the waiting time revealed that the degree of the package ordering of DNA in a toroid depends on the compaction kinetics.