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[Display omitted] • A new species known as R. qinlinganus sp. nov. within the subgenus Aeschyntelus , has been identified and established based on a comprehensive analysis of both morphological and molecular. • Discrepancies arising from nuclear and mitochondrial data suggest the presence of mito-nuclear discordance in the subgenus Aeschyntelus. • The subgenus Aeschyntelus likely originated in East Asia during the middle Miocene. • The development of Aeschyntelus biodiversity in the southwestern mountains of China occurred via an uplift-driven diversification process. The subgenus Aeschyntelus includes six species that show variations in body color and shape, thus making it difficult to identify them based on morphological identification alone. To date, no genetic study has evaluated species within this genus. Herein, we collected 171 individuals from 90 localities of Rhopalus and employed an integrative taxonomic approach that incorporated morphological data, mitochondrial genomic data (COI , whole mitochondrial data) and nuclear genomic data (18S + 28S rRNAs, nuclear genome-wide SNPs) to delineate species boundaries. Our analyses confirmed the status of nine described species of Rhopalus and proposed the recognition of one new species known as Rhopalus qinlinganus sp. nov., which is classified within the subgenus Aeschyntelus. Discrepancies arising from nuclear and mitochondrial data suggest the presence of mito-nuclear discordance. Specifically, mitochondrial data indicated admixture within Clade A, comprising R. kerzhneri and R. latus , whereas genome-wide SNPs unambiguously identified two separate species, aligning with morphological classification. Conversely, mitochondrial data clearly distinguished Clade B- consisting of R. sapporensis into two lineages, whereas genome-wide SNPs unequivocally identified a single species. Our study also provides insights into the evolutionary history of Aeschyntelus , thus indicating that it likely originated in East Asia during the middle Miocene. The development of Aeschyntelus biodiversity in the southwestern mountains of China occurred via an uplift-driven diversification process. Our findings highlight the necessity of integrating both morphological and multiple molecular datasets for precise species identification, particularly when delineating closely related species. Additionally, it reveals the important role of mountain orogenesis on speciation within the southwestern mountains of China. [ABSTRACT FROM AUTHOR]

Abstract: Marine propulsion unit (MPU) is one of the dominant vibration and noise sources onboard ship. Its vibration can be attenuated effectively by isolating MPU with low-frequency mounting system. But this is difficult to implement due to the stringent requirement of MPU alignment with the propulsion shafting. In this paper a novel air spring mounting system (ASMS) for propulsion system is proposed consisting of air spring subsystem, alignment control subsystem and safety protection subsystem. The load distribution optimization method and dynamic model of ASMS are presented. The factors that affect system stability and natural frequencies are analyzed, as well as the design measures to enhance system performance. A theoretical model is presented to estimate the isolation effect of ASMS. The monitoring model of alignment between MPU and propulsion shafting is established, followed by the alignment control algorithm and converge rule which assures the fast and uniform convergence of both air springs׳ load distribution and alignment control process. Safety protection mechanism is designed to ensure that the MPU can operate safely in case of ASMS failure or other extreme circumstances. A scaled ASMS prototype is manufactured and tested on a special experimental setup. Experimental results validate the effectiveness of theoretical models and show that the performance of ASMS satisfies the operation requirements of MPU. [Copyright &y& Elsevier]

[Display omitted] • Phylogeography of Riptortus pests in East Asia. • Similar phylogeographic structure within two bean bugs based on dd-rad data. • Phylogeographic patterns of two bean bugs are less influence by soybean cultivation. Comparative phylogeographic studies of closely related species sharing co-distribution areas can elucidate the role of shared historical factors and environmental changes in shaping their phylogeographic pattern. The bean bugs, Riptortus pedestris and Riptortus linearis, which both inhabit subtropical regions in East Asia, are recognized as highly destructive soybean pests. Many previous studies have investigated the biological characteristics, pheromones, chemicals and control mechanisms of these two pests, but few studies have explored their phylogeographic patterns and underlying factors. In this study, we generated a double-digest restriction site-associated DNA sequencing (ddRAD-seq) dataset to investigate phylogeographic patterns and construct ecological niche models (ENM) for both Riptortus species. Our findings revealed similar niche occupancies and population genetic structures between the two species, with each comprising two phylogeographic lineages (i.e., the mainland China and the Indochina Peninsula clades) that diverged approximately 0.1 and 0.3 million years ago, respectively. This divergence likely resulted from the combined effects of temperatures variation and geographical barriers in the mountainous regions of Southwest China. Further demographic history and ENM analyses suggested that both pests underwent rapid expansion prior to the Last Glacial Maximum (LGM). Furthermore, ENM predicts a northward shift of both pests into new soybean-producing regions due to global warming. Our study indicated that co-distribution soybean pests with overlapping ecological niches and similar life histories in subtropical regions of East Asia exhibit congruent phylogeographic and demographic patterns in response to shared historical biogeographic drivers. [ABSTRACT FROM AUTHOR]

[Display omitted] • Erthesina fullo with a strong east–west differentiation phylogeographical structure in East Asia. • The violent climate fluctuation might cause population divergence during the Middle and Upper Pleistocene. • The coastal populations of the east lineage were identified as the source of invasion in Europe. • The coastal populations of the east lineage were identified as the source of invasion in Europe. • The widest geographical distribution, a star-like haplotype network with the main haplotype as the core and rapid population expansion history, indicates the source lineage might have stronger diffusion ability and adaptability than the other lineages. The yellow spotted stink bug (YSSB), Erthesina fullo (Thunberg, 1783) is an important Asian pest that has recently successfully invaded Europe and an excellent material for research on the initial stage of biological invasion. Here, we reported the native evolutionary history, recent invasion history, and potential invasion threats of YSSB for the first time based on population genetic methods [using double digest restriction-site associated DNA (ddRAD) data and mitochondrial COI and CYTB] and ecological niche modelling. The results showed that four lineages (east, west, southwest, and Hainan Island) were established in the native range with a strong east–west differentiation phylogeographical structure, and the violent climate fluctuation might cause population divergence during the Middle and Upper Pleistocene. In addition, land bridges and monsoon promote dispersal and directional genetic exchanging between island populations and neighboring continental populations. The east lineage (EA) was identified as the source of invasion in Albania. EA had the widest geographical distribution among all other lineages, with a star-like haplotype network with the main haplotype as the core. It also had a rapid population expansion history, indicating that the source lineage might have stronger diffusion ability and adaptability. Our findings provided a significant biological basis for fine tracking of invasive source at the lineage or population level and promote early invasion warning of potential invasive species on a much subtler lineage level. [ABSTRACT FROM AUTHOR]

The Manila clam (Ruditapes philippinarum), one of the major marine aquaculture species in China, is susceptible to infection with the pathogen Vibrio , which results in massive mortality and economic losses. Toll-like receptors (TLRs) are significant pattern recognition receptors (PRRs) of innate immunity that are involved in immune regulation against pathogenic invasion. Molecular characterization of Manila clam TLRs and investigations of their immune functions are essential to prevent and control Vibrio infection. In the present research, eight cDNA sequences of R. philippinarum TLRs (RpTLRs) were identified from previous transcriptome libraries and then classified into four groups, namely, P-TLR (one sequence), V-TLR (one sequence), Ls-TLR (two sequences) and sP-TLR (four sequences), based on the corresponding LRR domain arrangement of their protein structures within the typical TLR motifs. A selective pressure test firstly suggested that the molluscan P-TLR, V-TLR, Ls-TLR and sP-TLR families underwent positive selection, and different numbers of positive selection sites (PSSs) were identified in different domains of the four types of RpTLRs, as determined by PAML and analysis of website data. These findings indicated that the evolution of RpTLRs may be associated with their immune recognition and function. Furthermore, tissue-specific expression analysis showed that all RpTLRs were ubiquitously expressed in all test tissues and were dominant in hemocytes. Quantitative real-time PCR revealed that the cDNA expression of all eight RpTLRs was upregulated after injection with Vibrio anguillarum (P < 0.01) in R. philippinarum hemocytes, revealing that these RpTLRs play important roles in responding to pathogenic stimulation. In summary, these findings provide a foundation for future investigations of the molecular classification and evolutionary patterns of Toll-like receptors in invertebrates, and the innate immune responses of TLR signaling pathways in Mollusca. [Display omitted] • P-TLR, V-TLR,Ls-TLR and sP-TLR genes were identified and characterized from Ruditapes philippinarum transcriptome. • The four families of molluscan TLRs underwent positive selection, and PSSs of them are located in different domain. • RpTLRs dominantly exist in hemocytes and significantly up-regulated by Vibrio anguillarum stimulation. [ABSTRACT FROM AUTHOR]

Toll-like receptors, the best known pattern recognition receptors, play important roles in recognizing non-self molecules and binding pathogen-associated molecular patterns in the innate immune system. In the present research, the cDNA and protein characterization of the TLR signalling pathway genes including IRAK4 , TRAK6 and IKKα (named CsIRAK4 , CsTRAF6 and CsIKKα , respectively) with the typical motifs from Cyclina sinensis showed significant similarity with their homologues from other shellfish. Furthermore, the mRNA transcripts of these three genes are ubiquitously expressed in all tissues tested and are dominantly expressed in C. sinensis haemocytes ( P < 0.05). Moreover, IRAK4 , TRAK6 and IKKα cDNA expression levels were all up-regulated after injection with Vibrio anguillarum , Micrococcus luteus and poly I:C ( P < 0.01) as shown by quantitative real-time PCR, indicating that they were involved in responding to pathogenic stimulation. We explored the function of the TLR13-MyD88-NF-κB signalling pathway in the innate immune responses of C. sinensis by RNA interference and immune challenges. The results suggested the mRNA expression patterns of CsMyD88 , CsIRAK4 , CsTRAF6 , CsIKKα , CsIκB , CsNF-κB , CsC-LYZ and CsAMP were all down-regulated ( P < 0.01) in normal and stimulated C. sinensis haemocytes, revealing the involvement of the TLR13-MyD88-NF-κB signalling pathway in innate immunity by positively adjusting internal signalling factors and immune-related genes. In summary, a TLR13-MyD88-NF-κB signalling pathway exists and plays vital roles in innate immune responses in C. sinensis . These findings collectively lay the foundation for studying the functional characterization of internal signalling factors and establishing a regulatory network for the TLR signalling pathway in molluscs. [ABSTRACT FROM AUTHOR]

[Display omitted] Accurate taxonomy and delimitation are of great importance for pest control strategies and management programs. Here, we focus on Cletus (Insecta: Hemiptera: Coreidae), which includes many crop pests. The species boundaries still conflict and only cytochrome c oxidase subunit I (COI) barcoding has been previously used for molecular studies. We generated new mitochondrial genome and nuclear genome-wide SNPs to explore the species boundaries of 46 Cletus samples from China using multiple species delimitation approaches. All results recovered a monophyly with high support, except for two closely related species in clade I - C. punctiger and C. graminis. Mitochondrial data demonstrated admixture in clade I, while genome-wide SNPs unambiguously identified two separate species, which were confirmed by morphological classification. Inconsistent nuclear and mitochondrial data indicated mito-nuclear discordance. Mitochondrial introgression is the most likely explanation, and more extensive sampling and more comprehensive data are needed to ascertain a pattern. Accurate species delimitation will shed light on species status; thus, an accurate taxonomy is of particular concern, as there is a pressing need to implement precise control of agricultural pests and to perform further research on diversification. [ABSTRACT FROM AUTHOR]

The Manila clam, Ruditapes philippinarum , is one of the most economically important aquatic clams that are harvested on a large scale by the mariculture industry in China. However, increasing reports of bacterial pathogenic diseases have had a negative effect on the aquaculture industry of R. philippinarum . In the present study, the two transcriptome libraries of untreated (termed H) and challenged Vibrio anguillarum (termed HV) hepatopancreas were constructed and sequenced from Manila clam using an Illumina-based paired-end sequencing platform. In total, 75,302,886 and 66,578,976 high-quality clean reads were assembled from 101,080,746 and 99,673,538 raw data points from the two transcriptome libraries described above, respectively. Furthermore, 156,116 unigenes were generated from 210,685 transcripts, with an N50 length of 1125 bp, and from the annotated SwissProt, NR, NT, KO, GO, KOG and KEGG databases. Moreover, a total of 4071 differentially expressed unigenes (HV vs H) were detected, including 903 up-regulated and 3168 down-regulated genes. Among these differentially expressed unigenes, 226 unigenes were annotated using KEGG annotation in 16 immune-related signaling pathways, including Toll-like receptor, NF-kappa B, MAPK, NOD-like receptor, RIG-I-like receptor, and the TNF and chemokine signaling pathways. Finally, 20,341 simple sequence repeats (SSRs) and 214,430 potential single nucleotide polymorphisms (SNPs) were detected from the H and HV transcriptome libraries. In conclusion, these studies identified many candidate immune-related genes and signaling pathways and conducted a comparative analysis of the differentially expressed unigenes from Manila clam hepatopancreas in response to V. anguillarum stimulation. These data laid the foundation for studying the innate immune systems and defense mechanisms in R. philippinarum. [ABSTRACT FROM AUTHOR]

[Display omitted] • Total-evidence phylogeny largely resolved corixid phylogenetic relationships. • Corixoidea began to diversify in the Middle Triassic (approximately 245 Ma). • Reconstruction of ancestral ranges suggest a Gondwanan origin for Corixoidea. • The breakup of Pangea promoted diversification of the Holarctic corixid lineage. The water boatmen of Corixoidea, a group of aquatic bugs with more than 600 extant species, is one of the largest superfamilies of Nepomorpha. Contrary to the other nepomorphan lineages, the Corixoidea are most diverse in the Laurasian remnant Holarctic region. To explicitly test whether the present-day Holarctic distribution of diverse corixids is associated with the arising of the Laurasian landmass that was separated from Gondwana, we investigated the phylogeny, divergence times and historical biogeography of Corixoidea based on morphological and molecular characters sampled from 122 taxa representing all families, subfamilies, tribes and approximately 54 % of the genera. Our results were largely congruent with the phylogenetic relationships within the established nepomorphan phylogenetic context. The fossil calibrated chronogram, diversification analysis and ancestral ranges reconstruction indicated that Corixoidea began to diversify in Gondwana in the late Triassic approximately at 224 Ma and the arising of the most diverse subfamily Corixinae in Corixidae in the Holarctic region was largely congruent with the time of separation of Laurasia from Gondwana. The large-scale expansion of the temperate and cold zones on the northward-moving Laurasian landmass after the breakup of the Pangea provided new aquatic niches and ecological opportunities for promoting rapid diversification for the Holarctic corixid lineage. [ABSTRACT FROM AUTHOR]

Heat shock protein 70 (HSP70) is an important member of the heat shock protein superfamily and is involved in protecting organisms against various stressors. In the present study, we used RACE to clone a full-length Cyclina sinensis HSP70 cDNA termed CsHSP70 . The full length of the CsHSP70 cDNA was 2308 bp, with a 5′ untranslated region (UTR) of 42 bp, a 3′ UTR of 268 bp, and an open reading frame (ORF) of 1998 bp encoding a polypeptide of 655 amino acids with an estimated molecular mass of 72.75 kDa and an estimated isoelectric point of 5.48. Quantitative real-time PCR was employed to analyze the tissue distribution and temporal expression of the CsHSP70 gene after bacterial challenge and cadmium (Cd) exposure. The CsHSP70 mRNA transcript was expressed ubiquitously in five examined tissues, with the highest expression in hemocytes ( P < 0.05) and with the lowest expression in the hepatopancreas. Furthermore, the expression level of CsHSP70 in hemocytes at 3 h after Vibrio anguillarum challenge was extremely significantly up-regulated ( P < 0.01). Moreover, the CsHSP70 transcript was up-regulated significantly following exposure to a safe Cd concentration (0.1 mg/L). Finally, after the CsHSP70 gene was silenced by RNA interference, the expression of the CsTLR13 and CsMyD88 genes were extremely significantly decreased ( P < 0.01). The results indicated that CsHSP70 could play an important role in mediating the environmental stress and immune responses, and regulating TLR signaling pathway in C. sinensis . [ABSTRACT FROM AUTHOR]

Toll-like receptors (TLRs) are an ancient family of pattern recognition receptors that play a critical role in initiating and activating the innate immune system. In this study, we identified two TLR genes ( CsTLR4 and CsTLR13 ) and the MyD88 ( CsMyD88 ) gene using a transcriptome library from Cyclina sinensis . The sequence features and mRNA expression profiles of the genes were characterized, and their functions in the immune response were investigated to validate the TLR signaling pathway and its potential role in immune defense. The expression patterns of CsTLR4 , CsTLR13 and CsMyD88 were detected in all the tissues examined from healthy clams and were primarily expressed in the hemocytes ( P < 0.05), as shown by real-time PCR. Upon challenge with Vibrio anguillarum and Micrococcus luteus , they were significantly increased in hemocytes ( P < 0.01), whereas only CsTLR13 and CsMyD88 were up-regulated ( P < 0.01) by poly (I:C) challenge. In addition, the mRNA expression level of CsC-LYZ and CsAMP was down-regulated at 72 h ( P < 0.01) after injection with CsMyD88 RNAi. These findings might be valuable for understanding the innate immune signaling pathways of C. sinensis and enabling future studies on host–pathogen interactions. [ABSTRACT FROM AUTHOR]

Abstract: The SSU nrDNA (18S), is one of the most frequently sequenced molecular markers in phylogenetic studies. However, the length-hyper-variation at multiple positions of this gene can affect the accuracy of alignment greatly and this length variation makes alignment across arthropod orders a serious problem. The analyses of Hexapoda phylogeny is such a case. A more clear recognition of the distribution of the length-variable-regions is needed. In this study, the secondary structure of some length-variable-regions in the SSU nrRNA of Arthropoda was adjusted by the principle of co-variation. It is found that the extent of plasticity of some length-variable-region can extraordinarily be higher than 600 bases in hexapods. And the numbers of hyper length-variable-regions are largest in Strepsiptera and Sternorrhyncha (Hemiptera). Our study shows that some length-variable-regions can serve as synapomorphies for some groups. The phylogenetic comparison also suggested that the expansion of a lateral bulge could be the origin of a helix. [Copyright &y& Elsevier]

Abstract: Cimicomorpha, which consists of 16 families representing more than 19,400 species, is the largest infraorder in Heteroptera, Insecta. We present the first molecular phylogenetic investigation of family relationships of Cimicomorpha, including 46 taxa from 12 of 16 Cimicomorphan families. Three genes, with a total of 3277bp of sequence data (nuclear 18S rDNA: 2022bp, 28S rDNA: 755bp, and mitochondrial 16S rDNA: 498bp) were analyzed. Data partitions were analyzed separately and in combination, by employing ML (maximum likelihood), MP (maximum parsimony), and Bayesian methods. As saturation was detected in substitutions of 16S rDNA, influence of data conflict in combined analyses was further explored by three methods: the incongruence length difference (ILD) test, the partitioned Bremer support (PBS), and the partition addition bootstrap alteration approach (PABA). PBS and PABA approaches suggested that 16S rDNA was not very suitable for addressing relationships at this level in Cimicomorpha. Our results also supported the nabid–cimicoid lineage for Cimicoidea proposed by Schuh and Štys [Schuh, R.T., Štys, P., 1991. Phylogenetic analysis of Cimicomorphan family relationships (Heteroptera). J. NY Entomol. Soc. 99 (3), 298–350]. Data incongruence and the utility of the three genes were briefly discussed. [Copyright &y& Elsevier]

Abstract: The small subunit of nuclear ribosomal RNA (SSU nrRNA), whose sedimentation is mostly 18S in eukaryotes, is considered a relatively conservative marker for resolving phylogenetic relationship at the order level or higher. Length variation in SSU nrDNA is common, and can be rather large in some groups. In studies of Hexapoda phylogeny, the SSU nrDNA has been repeatedly used as a marker. Sternorrhyncha has been rarely included. The lengths of SSU nrDNAs of sternorrhynchids, the basal group of Hemiptera identified in the previous study are 0.3–0.6kb longer than the usual ones in Hexapoda (1.8–1.9kb). To use the entire SSU nrDNA sequences or the length-variable parts could cause alignment trouble and therefore affect phylogenetic results, as shown in this study of Euhemiptera phylogeny. Two problems are particularly noticeable. One is that two hyper-variable regions flanking a short length-conservative region could become overlapped in the alignment. This will destroy the positional homology over a larger range. The other is that, when a base pair in a stem of the secondary structure is located near the length-variable regions (LVRs), the simultaneous positional homology of these two bases in the pair is always lost in the alignment results. In this study, the secondary structure model of Hexapoda SSU nrRNA was slightly adjusted and the LVR distributions in it were finely positioned. The noise caused by the hyper LVRs was eliminated and the simultaneous homology for the paired bases was recovered based on the secondary structure model. These corrections improved the quality of the data matrix and hence improved the resolving behavior of the algorithm used. This study provided more convincing evidence for resolving the Euhemiptera suborders phylogeny as (Archaeorrhyncha+(Clypeorrhyncha+(Coleorrhyncha+Heteroptera))). This result provided a more solid background for outgroup determination according to the phylogenetic studies inside each suborder. The problems caused by LVRs have seldom been well addressed. As phylogenetic reconstruction depends more on the data matrix itself than on the algorithm, and length variation of SSU/LSU rRNA exists more or less in any group, it is necessary to closely investigate the effect of rRNA length variation on alignment and phylogenetic reconstruction in more groups. [Copyright &y& Elsevier]