Schistosoma mansoni were identified a decade ago, more flatworm genome data is available to identify their NR complement and to analyze the evolutionary relationship of Platyhelminth NRs. NRs are important transcriptional modulators that regulate development, differentiation and reproduction of animals. In this study, NRs are identified in genome databases of thirty-three species including in all Platyhelminth classes . Phylogenetic analysis shows that NRs in Platyhelminths follow two different evolutionary lineages: 1) NRs in a free-living freshwater flatworm (Schmidtea mediterranea) and all parasitic flatworms share the same evolutionary lineage with extensive gene loss. 2) NRs in a free-living intertidal zone flatworm (Macrostomum lignano) follow a different evolutionary lineage with a feature of multiple gene duplication and gene divergence. The DNA binding domain (DBD) is the most conserved region in NRs which contains two C4-type zinc finger motifs. A novel zinc finger motif is identified in parasitic flatworm NRs: the second zinc finger of parasitic Platyhelminth HR96b possesses a CHC2 motif which is not found in NRs of all other animals studied to date. In this study, novel NRs (members of NR subfamily 3 and 6) are identified in flatworms, this result demonstrates that members of all six classical NR subfamilies are present in the Platyhelminth phylum. NR gene duplication, loss and divergence in Platyhelminths are analyzed along with the evolutionary relationship of Platyhelminth NRs.Since the first complete set of Platyhelminth nuclear receptors (NRs) from  Platyhelminths (flatworms) are one of the largest animal phyla, which includes more than 20,000 species , 2. SomeDrosophila melanogaster) [Anopheles gambiae) [Caenorhabditis elegans and C. briggsae) [Ciona intestinalis) [Rattus norvegicus), mouse (Mus musculus) [Homo sapiens) [Nuclear receptors (NRs) are important transcriptional modulators that regulate development, differentiation and reproduction of animals. Most of NRs share a common tertiary structure: A/B-C-D-E domains. The A/B domain is highly variable, the C domain is the DNA-binding domain (DBD) which is the most conserved region containing two zinc finger motifs, the D domain a flexible hinge between the C and E domains and is poorly conserved, the E domain contains the ligand binding domain (LBD) which is involved in transcriptional activation. Atypical NRs are found in some animals, e.g. NRs with a DBD but no LBD are found in arthropods and nematodes, members without a DBD but with a LBD are present in some vertebrates, and NRs with two DBDs and a single LBD (2DBD-NRs) are identified in protostomes. A phylogenetic analysis of the NRs divides them into six classical subfamilies by alignment of the conserved DBD . The earogaster)  the mosqgambiae) , free-liriggsae) , 8, tunitinalis) , mammaliusculus)  and humasapiens) ) revealeSchistosoma mansoni a decade ago [Schistosoma mansoni [Echinococcus multilocularis [S. mansoni and E. multilocularis share the same evolutionary lineage as that of Deuterostomia and the arthropods in the Ecdysozoan clade of the Protostomia, but some divergent NRs in flatworms are not present in Deuterostomia or/and arthropods, for example 2DBD-NRs [Since we identified the first complete set of Platyhelminths nuclear receptors (NRs) from cade ago , more fl mansoni –14 and tocularis –15 shows2DBD-NRs . In this2DBD-NRs , 18. IdeA schematic that illustrates the different NR subfamilies and NRs in human, insects, Mollusca and Platyhelminths are shown in Nuclear receptors in Platyhelminths were mined from the genome databases in Wormbase ParaSite (version: WBPS15 (WS276)) , 18. AmiPhylogenetic trees were constructed from deduced amino acid sequences of the DBD and/or ligand binding domain (LBD), the sequences are aligned with ClustalW , phylogeSchistosoma mansoni and Echinococcus multilocularis) that have had their NR complement reported [NRs are identified in the genomes of 33 Platyhelminth species including 2 species . In this study, phylogenetic analysis of family 5 , but thefamily 5 –S17 Figs2+ ion. The first zinc finger (CI) contains a sequence element, the P-box [DBD is the most conserved region in NRs, it contains two C4-type zinc finger motifs. In each motif, four cysteine residues chelate one Znhe P-box , 63 whiche P-box .Schistosoma mansoni HR96b (SmNR96b) [Previously, we isolated a partial cDNA of a trematode SmNR96b) , unlike SmNR96b) , 65. In SmNR96b) , 65) is Macrostomum lignano (M. lignano NR3 (MlERRs) shows that all of them are clustered together as a separate group within NR3 subfamily with a high ML/Bayesian support value (Four members of NR subfamily 3 (MlERRs), one member of subfamily 6 (MlNR6) and an orthologue of fax-1 (Mlfax-1) are identified in Rhabditophora  lignano , this rert value . Though rt value , thus thCaenorhabditis elegans NHR236) was present in Cnidaria, Arthropoda, free-living Platyhelminths, Mollusca and Echinodermata [P. xenopodis  [Echinococcus granulosus HR3 (EgHR3) [mediterranea  , which i (EgHR3) , it is p   areDivergent NR refers to a NR which has a typical P-box sequence in the DBD but does not fall into any ‘typical’ NR groups, for example Platyhelminths NR1a and NR1b with a typical P-box of ‘CEGCKGFFRR’ belonging to NR subfamily 1 but they do not fit into any groups within the NR1 subfamily. It also refers to a NR which has an atypical P-box sequence in the DBD and does not fall into the present NR nomenclature .S. mediterranea and in Cestoda Mesocestoides corti. Most of the Platyhelminth divergent NRs possess an ‘atypical’ P-box sequence. For example, a NR with a P-box of ‘CEPCKVFFKR’ is identified in Monogenea G. salaris, a NR with a P-box of ‘CEACKAFFQQ’ is found in all analyzed species of Cestoda Hymenolepis family, a NR which has a P-box of ‘CDSCRAFFEM’ exists in Cestoda Taenia family, a NR with a P-box of ‘CEACKSFFKR’ is found in Cestoda Diphyllobothriidea order and seventeen NRs with various ‘atypical’ P-box sequences were identified in Rhabditophora M. lignano  which is the same as that of RXR/TR4/NR4A is found in Rhabditophora  lignano .M. lignano, Cestoda and Mollusca. The result suggests that these divergent NRs diverged and duplicated independently in different animal lineages.A phylogenetic tree of the divergent NRs including those of Platyhelminths and Mollusca was constructed . PhylogeM. lignano and three are identified in S. mediterranea; and two are identified in Cestoda and Trematoda, respectively. Our previous study showed that two TR homologues are present in Platyhelminths [M. lignano TRs are clustered in a group and three S. mediterranea TRs are clustered in another group. This result suggests that TRs duplicated independently in M. lignano, S. mediterranea and parasitic Platyhelminths. In parasitic Platyhelminth TR groups, trematode TRa  are clustered together with those of Monogenea and Cestoda, but trematode TRb  group only contain trematode TRs. This result suggested that one TR was present in a common ancestor of Platyhelminths and trematode TRa was an ancient TR gene. Phylogenetic analysis shows that the two Cestode M. corti TRs are clustered together in Cestoda TR group, this result suggests that TRs in Cestoda and Trematoda underwent duplication independently . Phylogenetic analysis shows HR96d is an ancient genewhich is supported by sequence analysis, with only members of Platyhelminth HR96d sharing the same DBD amino acid length and intron position with that of Mollusca HR96s  and RXR2 group (Schistosoma RXR2 orthologues). RXR2 group is clustered with RXRs of free-living flatworms, Drosophila, Mollusca and human, which suggests that parasitic Platyhelminth RXR2 is an orthologue of free-living flatworms, Drosophila, Mollusca and human RXRs. RXR1 group contains only parasitic Platyhelminth RXRs, which suggests that parasitic Platyhelminth RXR2 duplicated after the split of their common ancestor with free-living Platyhelminths. In free-living flatworms, two M. lignano RXRs are clustered in a group while two S. mediterranea RXRs are scattered in the phylogenetic tree, this result suggests that RXR duplicated independently in free-living flatworms M. lignano and S. mediterranea  and Coup-TFII group (Schistosoma Coup-TFII orthologues). Coup-TFI group contains members from all three parasitic classes and they are clustered with Coup-TFs of free-living flatworms, Drosophila, Mollusca and humans. This result suggests that parasitic Platyhelminth Coup-TFI is an ancient Coup-TF gene and it is the orthologue of free-living flatworms, Drosophila, Mollusca and human Coup-TFs. Parasitic Platyhelminth Coup-TFII group contains Monogenea and Trematoda members, which suggests that Coup-TFI duplicated and gave rise to Coup-TFII in a common ancestor of parasitic Platyhelminths. In free-living flatworms, two M. lignano Coup-TFs and two S. mediterranea Coup-TFs are scattered in the phylogenetic tree, it suggests M. lignano and S. mediterranea Coup-TFs duplicated independently (M. corti); Coup-TFII was missing in all analyzed species of Cestoda.One Coup-TF was identified in Cestoda; two are identified in Rhabditophora, Monogenea and Trematoda, respectively. Phylogenetic analysis shows that parasitic flatworms Coup-TFs are clustered in two different groups: Coup-TFI group , 2DBDb (S. mansoni 2DBDβ orthologues) and 2DBDg (S. mansoni 2DBDγ orthologues) groups. Parasitic Platyhelminth 2DBD-NRa and 2DBD-NRb groups contain members of three classes of parasitic flatworms, but 2DBD-NRg group contains 2DBD-NRs of all the four classes of Platyhelminths and members of the Mollusca. This result suggests that Parasitic Platyhelminth 2DBD-NRg is an ancient gene, and 2DBD-NRa and 2DBD-NRb were formed by a second round of duplication. In free living Platyhelminths, both Rhabditophora M. lignano 2DBD-NRs are clustered in 2DBD-NRg group, but the four S. mediterranea 2DBD-NRs are clustered with different parasitic helminth 2DBD-NR groups. This result suggests that M. lignano 2DBD-NR gene underwent duplication after a split of S. mediterranea and parasitic Platyhelminths. For the four S. mediterranea 2DBD-NRs, one is clustered in parasitic Platyhelminth 2DBD-NRg group, one is clustered in 2DBD-NRb group and two are clustered with 2DBD-NRa group. Since the two S. mediterranea 2DBD-NRs (2DBD-NRa1 and 2DBD-NRa2) in 2DBDa group form a polytomy, it suggests that 2DBD-NRa underwent another round of duplication and formed two 2DBD-NRs as a common ancestor of S. mediterranea and parasitic Platyhelminths and then one 2DBD-NRa was lost in a common ancestor of parasitic Platyhelminths . The number behind flatworm species/families in parentheses indicates the number of NRs identified.In this study, we identified NRs in different species of Platyhelminths and performed sequence and phylogenetic analysis of them. Phylogenetic analysis of the DBD of NRs using DBD sequences was carried out with Bayesian inference and Maximum Likelihood method, Comparison of the two methods, Bayesian inference highly support phylogenetic analysis of NRs using only DBD sequences. . This stS. mediterranea and parasitic Platyhelminths follow the first evolutionary lineage: gene duplication and lost, the events occurred in different flatworm lineages. For example, fax-1 was lost in a common ancestor of S. mediterranea and parasitic Platyhelminths, NHR236 was lost in Cestoda and Trematoda, and DSF was lost in Cestoda. In parasitic Platyhelminths, extensive NR gene loss occurred in Cestoda Hymenolepididae and Taeniidae families, e. g. RXR, Coup-TF and FTZ-F1 were retained in most species of Platyhelminths, but they were missing in the cestode families Hymenolepididae and Taeniidae.NRs in Rhabditophora Schistosoma and Trichobilharzia). NR complement in Monogenea and in Cestoda exhibit more differences among different families, in Monogenea, there are 18 NRs in Gyrodactylidae , but 23 NRs are present in Polyopisthocotylea (P. xenopodis). In Cestoda Cyclophyllidea, there are 15 NRs in Hymenolepididae family, 17 in Taeniidae family and 20 in Mesocestoididae family. NR gene duplication occurred in different flatworm lineages. For example, 2DBD-NR underwent a second round of duplication in a common ancestor of S. mediterranea and parasitic Platyhelminths, RXR gene duplicated separately in S. mediterranea and a common ancestor of parasitic Platyhelminths, and TR duplicated independently in Rhabditophora (S. mediterranea), Cestoda and Trematoda.Comparison of NR complement in different Platyhelminth families shows that NR complement is most conserved in Trematoda. All species of Trematoda share the same NR complement except that a HR96 member was lost in Strigeidida  and gene divergence. Multiple NR gene duplication was also found in other animals, for example, supplementary NRs (SupNRs) in nematodes and NR1H in Chordate Cephalochordate amphioxus [M. lignano may result from the adaption to its environment.NRs in Rhabditophora mphioxus , 69. DHRmphioxus . MlHR96 M. lignano. This is the first known occurrence of NRs in subfamily 3 and 6 in Platyhelminths, but it is still not clear whether NRs from subfamily 3 and 6 are present in other Rhabditophora since the genome data is unavailable. This study also shows that divergent NRs are present in different flatworm lineages suggesting that novel NRs were acquired in different flatworms to adapt to the different living environments.NRs in subfamily 3 and 6 were considered lost in Platyhelminths. In this study, we identified four ERRs and a NR6A in Rhabditophora E75/Rev, DHR3/ROR and EcR/LXR, which are present in Deuterostomia, Arthropods and Lophotrochozoa Mollusca, are missing in Platyhelminths. In insects, E75, HR3 and EcR are directly involved in the control of the ecdysone pathway. E75 acts as a repressor of DHR3 and may through direct interaction .Nucella lapillus RAR (NlRAR) showed that NlRAR binds to NR response elements as a heterodimer with RXR, but it does not bind all-trans retinoic acid or other retinoids [N. lapillus, PPAR-responsive pathways is related to tributyltin (TBT) induced imposex [RAR, PPAR, ER, which are missing in Arthropods but retained in Lophotrochozoa Mollusca, are also missing in Platyhelminths. In vertebrates, RARs are known to bind retinoic acid (RA). RA is a morphogen derived from vitamin A, it controls the patterning of the anteroposterior axis and the differentiation of various cell types . Study oetinoids . In vert imposex . Vertebr imposex .ERRs are orphan receptors in Chordata and Ecdysozoa, they play important roles in regulation of neurogenesis and metabolism in Chordata and are involved in control of larval growth in Ecdysozoa. The function of Lophotrochozoa ERRs is unknown . In thisApis mellifera and then in other insects, but it was missing in Drosophila. The missing of NR2E6 in the major model organisms delayed the identification and functional analysis of this protein. Sometimes NR2E6 is named as PNR-like or PNR, but the true insect homolog of vertebrate PNR is HR51 (NR2E3) [Caenorhabditis elegans HR236) was present in Cnidaria, Arthropoda, Platyhelminths, Mollusca and Echinodermata [The NR2E6 gene was first identified in the genome of the honeybee  (NR2E3) . We demoodermata . Insect S. mansoni HR96b (SmNR96b) [Previously, we isolated a partial cDNA of SmNR96b) , this meMacrostomum lignano NRs was published ) while this paper was under review. The main differences of identification of M. lignano NRs between Cheng et al. and this contribution are as follows . Cheng, Y., et al. identified 4 NR1Cs (PPARs) in M. lignano, we found no evidence for any PPARs in M. lignano. (3). Cheng, Y., et al. identified 2 NR1Fs (RORs). We identified the same sequences but our phylogenetical analysis showed that they belonged to NR1E (E78) group. (4). Cheng, Y., et al. identified 20 NR1Js (HR96s) in M. lignano including one sequence without a DBD sequence and 5 members each with an atypical P-box sequence which is different from the typical HR96 P-box sequence. We identified 16 HR96s in M. lignano, all of them possessed a typical HR96 P-box sequence. We also identified the same 5 members with an atypical P-box sequence that Cheng, Y., et al. identified, but put them into a divergent NR group. (5). Cheng, Y., et al. identified 10 NR3 members in M. lignano including four members that each possessed a typical ERR P-box sequences and 6 members that each possessed an atypical P-box sequence. We identified the same members but put the six members that possessed an atypical P-box sequence into a divergent NR group. (6). Cheng, Y., et al. identified a NR8 member, we identified the same member but our phylogenetical analysis showed it was a divergent member. (7). Cheng, Y., et al. identified 2 NR0 members. We identified the same members but our phylogenetic analysis showed one of them belong to subfamily 6 and the other one is a divergent member because it possessed an atypical P-box sequence.A paper on S1 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Ml: Macrostomum lignano, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show M. lignano NRs.The Bayesian tree was constructed with the deduced amino sequences of the DNA binding domain (DBD) with a mix amino acid replacement model + invgamma rates. The PPs values are shown above each branch, branches under the PPs 0.5 are shown as polytomies. The same data set is also tested by ML method using PHYML (v2.4.4) under LG+G+I substitution model , Number of substitution rate categories: 4, Gamma shape parameter: Estimated (0.956). Support values for the tree are obtained by bootstrapping a 1,000 replicates and bootstrap values above 500 and are indicated below each branch (or after MrBAYES BPPs separated by Slash). Star indicates the node obtained by Bayesian inference which is different from that obtained by ML method. Bg: (TIF)Click here for additional data file.S2 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni, Sme: Schmidtea mediterranea. Red highlighted NRs show S. mediterranea NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S3 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show P. xenopodis NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S4 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, Gs: Gyrodactylus salaris, h: Homo sapiens, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show G. salaris NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S5 FigH. microstoma NRs represents all analyzed Hymenolepis species because of the highly conserved DBD sequences in these species. Methods for construction of phylogenetic trees see Biomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Hm: Hymenolepis microstoma, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show H. microstoma NRs.The phylogenetic tree of (TIF)Click here for additional data file.S6 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Mc: Mesocestoides corti, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show M. corti NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S7 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Em: Echinococcus multilocularis, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show E. multilocularis NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S8 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Ht: Hydatigera taeniaeformis, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show H. taeniaeformis NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S9 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni, Ts: Taenia saginata. Red highlighted NRs show T. saginata NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S10 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Ss: Schitocephalus solidus, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show T. saginata NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S11 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Se: Spirometra erinaceieuropaei, Sm: Schistosoma mansoni. DBD of E78 sequence is partial, not used for construction of the tree. Red highlighted NRs show S. erinaceieuropaei NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S12 FigBiomphalaria glabrata, Cg: Crassostrea gigas, Cs: Clonorchis sinensis, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show C. sinensis NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S13 FigO viverrini NRs represents Opisthorchis viverrini and O. felineus because of the highly conserved DBD sequences in these two species. Methods for construction of phylogenetic trees see Biomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Ov: Opisthorchis viverrini, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show O. viverrini NRs.The phylogenetic tree of (TIF)Click here for additional data file.S14 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, Fh: Fasciola hepatica, h: Homo sapiens, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. Red highlighted NRs show F. hepatica NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S15 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, Eca: Echinostoma caproni, h: Homo sapiens, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni. EcaCoup-TFII sequence is partial and not used for construction of the tree. Red highlighted NRs show E. caproni NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S16 FigS haematobium NRs represents all analyzed Schistosoma species because of the highly conserved DBD sequences in these species. Methods for construction of phylogenetic trees see Biomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sh: Schistosoma haematobium, Sm: Schistosoma mansoni. Red highlighted NRs show S. haematobium NRs.The phylogenetic tree of (TIF)Click here for additional data file.S17 FigBiomphalaria glabrata, Cg: Crassostrea gigas, d: Drosophila melanogaster, h: Homo sapiens, Lg: Lottia gigantean, Px: Protopolystoma xenopodis, Sm: Schistosoma mansoni, Tr: Trichobilharzia regent. Red highlighted NRs show T. regent NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S18 FigBiomphalaria glabrata, d: Drosophila melanogaster, Dl: Dibothriocephalus latus, Eca: Echinostoma caproni, Eg: Echinococcus granulosus, Fh: Fasciola hepatica, h: Homo sapiens, Hd: Hymenolepis diminuta, Ht: Hydatigera taeniaeformis, Lg: Lottia gigantean, Mc: Mesocestoides corti, Ml: Macrostomum lignano, Ov: Opisthorchis viverrini, Px: Protopolystoma xenopodis, Se: Spirometra erinaceieuropaei, Sm: Schistosoma mansoni, Sme: Schmidtea mediterranea, Ss: Schitocephalus solidus, Tm: Taenia multiceps, Tr: Trichobilharzia regent. Red highlighted NRs show Platyhelminths NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S19 FigBiomphalaria glabrata, d: Drosophila melanogaster, Eg: Echinococcus granulosus, h: Homo sapiens, Lg: Lottia gigantean, Sme: Schmidtea mediterranea. Red highlighted NRs show Platyhelminths NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S20 FigBiomphalaria glabrata, d: Drosophila melanogaster, Eg: Echinococcus granulosus, h: Homo sapiens, Lg: Lottia gigantean, Sme: Schmidtea mediterranea. Red highlighted NRs show Platyhelminths NRs.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S21 FigBiomphalaria glabrata, d: Drosophila melanogaster, Dl: Dibothriocephalus latus, Ec: Echinococcus Canadensis, Eg: Echinococcus granulosus, Em: Echinococcus multilocularis, Gs: Gyrodactylus salaris, h: Homo sapiens, Hd: Hymenolepis diminuta, Hm: Of H. microstoma, Ht: Hydatigera taeniaeformis, Mc: Mesocestoides corti, Ml: Macrostomum lignano, Sme: Schmidtea mediterranea, Ta: Taenia asiatica, Tm: Taenia multiceps, Ts: Taenia saginata, Tr: Trichobilharzia regent, Tso: Taenia solium.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S22 FigBiomphalaria glabrata, Cs: Clonorchis sinensis, d: Drosophila melanogaster, Dl: Dibothriocephalus latus, Ec: Echinococcus Canadensis, Eca: Echinostoma caproni, Eg: Echinococcus granulosus, Em: Echinococcus multilocularis, Fh: Fasciola hepatica, Gs: Gyrodactylus salaris, h: Homo sapiens, Hd: Hymenolepis diminuta, Hm: Of H. microstoma, Hn: Hymenolepis nana, Ht: Hydatigera taeniaeformis, Lg: Lottia gigantean, Mc: Mesocestoides corti, Ml: Macrostomum lignano, Of: Opisthorchis felineus, Ov: Opisthorchis viverrini, Px: Protopolystoma xenopodis, Sb: Schistosoma bovis, Sc: Schistosoma curassoni, Se: Spirometra erinaceieuropaei, Sh: Schistosoma haematobium, Sj: Schistosoma japonicum, Sm: Schistosoma mansoni, Sma: Schistosoma margrebowiei, Smt: Schistosoma mattheei, Sme: Schmidtea mediterranea, Sr: Schistosoma rodhaini, Ss: Schitocephalus solidus, Ta: Taenia asiatica, Tm: Taenia multiceps, Ts: Taenia saginata, Tr: Trichobilharzia regent, Tso: Taenia solium.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S23 FigBiomphalaria glabrata, Cs: Clonorchis sinensis, d: Drosophila melanogaster, Dl: Dibothriocephalus latus, Ec: Echinococcus Canadensis, Eca: Echinostoma caproni, Eg: Echinococcus granulosus, Em: Echinococcus multilocularis, Fh: Fasciola hepatica, Gs: Gyrodactylus salaris, h: Homo sapiens, Lg: Lottia gigantean, Ml: Macrostomum lignano, Of: Opisthorchis felineus, Ov: Opisthorchis viverrini, Px: Protopolystoma xenopodis, Sb: Schistosoma bovis, Sc: Schistosoma curassoni, Se: Spirometra erinaceieuropaei, Sh: Schistosoma haematobium, Sj: Schistosoma japonicum, Sm: Schistosoma mansoni, Sma: Schistosoma margrebowiei, Smt: Schistosoma mattheei, Sme: Schmidtea mediterranea, Sr: Schistosoma rodhaini, Ss: Schitocephalus solidus, Tr: Trichobilharzia regent. B) Sequence alignment shows the intron position (red color >) in DBD of RXRs.A) Bayesian phylogenetic tree of Platyhelminth RXR. Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S24 FigBiomphalaria glabrata, Cs: Clonorchis sinensis, d: Drosophila melanogaster, Dl: Dibothriocephalus latus, Ec: Echinococcus Canadensis, Eca: Echinostoma caproni, Eg: Echinococcus granulosus, Em: Echinococcus multilocularis, Fh: Fasciola hepatica, Gs: Gyrodactylus salaris, h: Homo sapiens, Lg: Lottia gigantean, Ml: Macrostomum lignano, Of: Opisthorchis felineus, Ov: Opisthorchis viverrini, Px: Protopolystoma xenopodis, Sb: Schistosoma bovis, Sc: Schistosoma curassoni, Se: Spirometra erinaceieuropaei, Sh: Schistosoma haematobium, Sj: Schistosoma japonicum, Sm: Schistosoma mansoni, Sma: Schistosoma margrebowiei, Smt: Schistosoma mattheei, Sme: Schmidtea mediterranea, Sr: Schistosoma rodhaini, Ss: Schitocephalus solidus, Ta: Taenia asiatica, Tm: Taenia multiceps, Ts: Taenia saginata, Tr: Trichobilharzia regent, Tso: Taenia solium. B) Sequence alignment shows the intron position (red color >) in DBD of Coup-TFs.A) Bayesian phylogenetic tree of Platyhelminth Coup-TF. Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S25 FigBiomphalaria glabrata, d: Drosophila melanogaster, Gs: Gyrodactylus salaris, h: Homo sapiens, Hd: Hymenolepis diminuta, Lg: Lottia gigantean, Mc: Mesocestoides corti, Ml: Macrostomum lignano, Sm: Schistosoma mansoni, Sme: Schmidtea mediterranea.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S26 FigBiomphalaria glabrata, Cs: Clonorchis sinensis, d: Drosophila melanogaster, Dl: Dibothriocephalus latus, Em: Echinococcus multilocularis, Fh: Fasciola hepatica, Gs: Gyrodactylus salaris, h: Homo sapiens, Hd: Hymenolepis diminuta, Hm: H. microstoma, Lg: Lottia gigantean, Mc: Mesocestoides corti, Ml: Macrostomum lignano, Px: Protopolystoma xenopodis, Se: Spirometra erinaceieuropaei, Sm: Schistosoma mansoni, Sme: Schmidtea mediterranea, Ss: Schitocephalus solidus, Tr: Trichobilharzia regent.Methods for construction of phylogenetic trees see (TIF)Click here for additional data file.S1 File(DOCX)Click here for additional data file.S2 File(DOCX)Click here for additional data file.S3 File(DOCX)Click here for additional data file.S4 File(DOCX)Click here for additional data file.S5 File(XLSX)Click here for additional data file.


Acetylation at lysine residue in a protein mediates multiple cellular biological processes, including tumorigenesis. This study aimed to investigate the acetylated protein profile alterations and acetylation-mediated molecular pathway changes in human nonfunctional pituitary neuroendocrine tumors (NF-PitNETs). The anti-acetyl antibody-based label-free quantitative proteomics was used to analyze the acetylomes between NF-PitNETs (n = 4) and control pituitaries (n = 4). A total of 296 acetylated proteins with 517 acetylation sites was identified, and the majority of which were significantly down-acetylated in NF-PitNETs (p<0.05 or only be quantified in NF-PitNETs/controls). These acetylated proteins widely functioned in cellular biological processes and signaling pathways, including metabolism, translation, cell adhesion, and oxidative stress. The randomly selected acetylated phosphoglycerate kinase 1 (PGK1), which is involved in glycolysis and amino acid biosynthesis, was further confirmed with immunoprecipitation and western blot in NF-PitNETs and control pituitaries. Among these acetylated proteins, 15 lysine residues within 14 proteins were down-acetylated and simultaneously up-ubiquitinated in NF-PitNETs to demonstrate a direct competition relationship between acetylation and ubiquitination. Moreover, the potential effect of protein acetylation alterations on NF-PitNETs invasiveness was investigated. Overlapping analysis between acetylomics data in NF-PitNETs and transcriptomics data in invasive NF-PitNETs identified 26 overlapped molecules. These overlapped molecules were mainly involved in metabolism-associated pathways, which means that acetylation-mediated metabolic reprogramming might be the molecular mechanism to affect NF-PitNET invasiveness. This study provided the first acetylomic profiling and acetylation-mediated molecular pathways in human NF-PitNETs, and offered new clues to elucidate the biological functions of protein acetylation in NF-PitNETs and discover novel biomarkers for early diagnosis and targeted therapy of NF-PitNETs. Pituitary neuroendocrine tumors (PitNETs) are the second most common primary central nervous system tumors in adults , 2. BaseAcetylation is a reversible post-translational modification (PTM), and is co-regulated by lysine acetyltransferases (KATs) and lysine deacetylases (KDACs). KATs catalyze lysine residue to be acetylated with acetyl-coenzyme A (acetyl-CoA) as a cofactor, and KDACs reverse this process. Acetylation modulates biological functions of many proteins related to tumorigenesis. Histone acetylation facilitates chromatin decondensation to regulate transcriptional activation . When DNHowever, acetylomics analysis in NF-PitNETs has not been reported. Previous studies on the effect of acetylation on pituitary tumorigenesis only focused on some specific molecules –19. Thusvs. controls  were obtained between invasive NF-PitNETs and control tissues from Gene Expression Omnibus (GEO) database. The overlapping analysis between acetylated protein data and invasive DEG data was performed to identify acetylation-mediated molecular events for invasiveness of NF-PitNETs.This study will provide promising scientific data for insights into molecular mechanisms of NF-PitNETs, and discover potential biomarkers for early diagnosis and therapy of NF-PitNET patients.Quantitative acetylomics was performed between the mixed NF-PitNET samples (n =4) and mixed control samples (n =4) , and colEach tissue sample was dealt with 1 mL urea pyrolysis solution  and ice bath ultrasonic treatment. The solution was centrifuged , and the supernatant of each sample was equally divided into three parts. The protein content of each part was measured with a Bradford Protein Quantification Kit . An amount (20 µg) of each extracted protein sample  was mixed with 6X loading buffer  in a ratio of 6:1(v/v), boiled (5 min), and centrifuged . The supernatant was loaded onto 12.5% SDS-PAGE gel  for electrophoretic separation , followed by staining with Coommassie brilliant blue .Dithiothreitol  was added to each extracted protein sample , and achieved a final concentration of 10 mM DTT; and the mixture was incubated , and cooled to room temperature. Then indole-3-acetic acid  was added into each mixture, and achieved a final concentration of 50 mM IAA; and the mixture was kept . The water that was 5 times the volume of the mixture was added to make the concentration of urea to 1.5 M, followed by addition of trypsin into the mixture in a ratio of 1:50 to digest proteins for 18 h at 37 °C. The SPE C18 column  was used to desalt and lyophilize tryptic peptides.A volume (1.4 mL) of pre-cooled immunoaffinity purification (IAP) buffer was used to resuspend each lyophilized peptide sample (3x). The pre-processed anti-Ac-K antibody beads  were added in each tryptic peptide sample, and incubated . Afterwards, anti-Ac-K antibody beads with acetylated peptides were washed with 1 mL pre-cooled IAP (3x), and with 1 mL pre-cooled water (3x). A volume (40 μl) of 0.15% trifluoroacetic acid  was added to the washed anti-Ac-K antibody beads, and incubated , and then the same volume of TFA was added once again. The mixture was centrifuged . The supernatant was desalted with C18 STAGE Tips . The desm/z 350-1800. The top 20 intensive ions in MS scan (MS1) were selected for ion fragmentation with higher-energy collision dissociation (HCD) to generate MS/MS spectra (MS2). The MS1 resolution was 70,000 at m/z 200, and the MS2 resolution was 17,500 at m/z 200.LC-MS/MS was used to analyze the enriched acetylated peptides . Each enriched acetylated peptide sample was separated by high performance liquid chromatography (HPLC) system EASY-nLC1000 at nanoliter flow velocity. Chromatography column was balanced with 100% buffer A (0.1% acetonitrile formate aqueous solution that contained 2% acetonitrile). The enriched acetylated peptides were loaded onto the sample spindle, Thermo scientific EASY column (2 cm*100 μm 5 μm-C18), with an autosampler in buffer A, and then were separated when the sample flowed through analytical column (75 µm × 250 mm 3 µm-C18) at a flow rate of 250 nL/min in buffer B (0.1% acetonitrile formate aqueous solution that contained 84% acetonitrile). The liquid-phase gradient was buffer B linear gradient from 0 to 55% for 220 min, buffer B linear gradient from 55 to 100% for 8 min, and then maintained 100% buffer B for 12 min. The Q-Exactive mass spectrometer (Thermo Finnigan) was used to perform MS/MS analysis when the enriched acetylated peptides were separated with capillary HPLC. The parameter of mass spectrometer was set as time 240 min, positive ion detection mode, and scan range of precursor ion The Maxquant software (version 1.3.0.5) was used for database searching and data analysis of 6 original LC-MS/MS datasets . The database was uniprot_human_154578_20160815.fasta . Its primary parameters were set as main search ppm = 6, missed cleavage = 4, MS/MS tolerance ppm = 20, de-isotopic =TRUE, enzyme=trypsin, database = uniprot_human_154578_20160815.fasta, fixed modification = carbamidomethyl (C), variable modification = oxidation (M), acetyl (protein N-term), and acetyl (K), decoy database pattern = reverse, iBAQ = TRUE, match between runs = 2 min, peptide false discovery rate (FDR) = 0.01, and protein FDR = 0.01. The MS/MS data were used to determine the amino acid sequence and acetylation sites, label-free quantification was used to determine the acetylation level.Immunoprecipitation (IP) and western blot were used to semi-quantify PGK1 acetylation level in NF-PitNETs compared to controls. Three NF-PitNET tissue samples were equally mixed as the NF-PitNET sample, and five control protein samples were equally mixed as the control sample  analysis through David database. GO analysis included three categories - biological processes (BPs), cellular components (CCs), and molecular functions (MFs). The results of KEGG, BP, CC, and MF data were further clustered into different functional categories. Moreover, acetylation motif analysis was carried out by analysis of the sequences from −13 to +13 amino acid residues at those 517 acetylation sites within 296 acetylated proteins with Motif-X software to identify any motifs that were prone to be acetylated in NF-PitNETs.The acetylated protein data identified in this study were compared to the ubiquitinated protein data in our previous study , which fvs. controls were mined from the GEO database  derived from PGK1 (Swiss-Prot No.: P00558) . Its acetylation site was localized at residue K*361, which was only acetylated in controls (N) but not in NF-PitNETs (T)  . Its acetylation site was localized at residue K*225, and its acetylation level was significantly decreased in NF-PitNETs compared to controls   were identified  For BPs, a total of 97 BPs was identified, covering almost all cellular biological processes, including gene expression, metabolism, cell-cell (matrix) adhesion, apoptosis, and immune system regulation  in transaldolase (ID: P37837), K194 , and K117  in glyceraldehyde-3-phosphate dehydrogenase (ID: P04406), K81  in cytoplasmic isocitrate dehydrogenase [NADP] (ID: O75874), K314 , K185 (only acetylated in NF-PitNETs), K324 and K239 (only acetylated in controls) in mitochondrial malate dehydrogenase (ID: P40926), K298, and K236 (only acetylated in controls) in cytoplasmic malate dehydrogenase (ID: P40925), K7 (only acetylated in controls) in alcohol dehydrogenase class-3 (ID: P11766), K39  in phosphoglycerate mutase 1 (ID:P18669), K302  in mitochondrial aspartate aminotransferase (ID: P00505), K291, K146, K156 and K361 (only acetylated in controls) in phosphoglycerate kinase 1 (ID: P00558), K538 (only acetylated in NF-PitNETs) mitochondrial succinate dehydrogenase [ubiquinone] flavoprotein subunit (ID: P31040), K231 (only acetylated in controls), and K225  in triosephosphate isomerase (ID: P60174), K169 (only acetylated in controls) in catalase (ID: P04040), K330 and K200 (only acetylated in controls) in fructose-bisphosphate aldolase A (ID: P04075), K202 , K257 , and K124 (only acetylated in controls) in mitochondrial acetyl-CoA acetyltransferase (ID: P24752), and K267  in mitochondrial dihydrolipoylde hydrogenase (ID: P09622). The acetylation level of most lysine residues at these enzymes decreased in NF-PitNETs. (ii) Most microbe and mammalian cells depend on glycolysis to convert glucose into lactate, and produce energy in the absence of oxygen. However, most tumor cells uptake more glucose, and produce more lactate even in the presence of oxygen even though mitochondria function well, which is noted as aerobic glycolysis, or Warburg effect (p = 4.24E-03), and K117  in glyceraldehyde-3-phosphate dehydrogenase (ID: P04406), K7 (only acetylated in controls) in alcohol dehydrogenase class-3 (ID: P11766), K39  in phosphoglycerate mutase 1 (ID: P18669), K291, K146, K156, and K361 (only acetylated in controls) in phosphoglycerate kinase 1 (ID: P00558), K231 (only acetylated in controls), and K225  in triosephosphate isomerase (ID: P60174) and K330 and K200 (only acetylated in controls) in fructose-bisphosphate aldolase A (ID: P04075), and K267  in mitochondrial dihydrolipoylde hydrogenase (ID: P09622). The acetylation levels at more than 4/5 lysine residues in these enzymes enriched in glycolysis/gluconeogenesis pathways were decreased in NF-PitNETs, which might result in the convert of glycolysis to the aerobic glycolysis in NF-PitNETs, and further affect tumor progression. (iii) Pyruvate, the end product of glycolysis, is reduced to lactate in cytoplasm or transport into mitochondria to enter TCA cycle for full oxidation for ATP production, and sit at the switch point between these two important carbohydrate metabolism pathways (p=1.42E-04) and K77 (only acetylated in NF-PitNETs) were increased in NF-PitNETs in this study. In addition, this study found acetylation occurred at other enzymes enriched on pyruvate metabolism pathway, including acetylation at residues K30 (only acetylated in NF-PitNETs) in 4-trimethylaminobutyraldehyde dehydrogenase (ID: P49189), K314 , K185 (only acetylated in NF-PitNETs), K324 and K239 (only acetylated in controls) in mitochondrial malate dehydrogenase (ID: P40926), K298, and K236 (only acetylated in controls) in cytoplasmic malate dehydrogenase (ID: P40925), and K202 , K257  and K124 (only acetylated in controls) in mitochondrial acetyl-CoA acetyltransferase (ID: P24752). (iv) TCA cycle is the final metabolic pathway of carbohydrates, lipids, and amino acids, which is the central route for cellular oxidative phosphorylation and provides precursors for many anabolic pathways (p=2.20E-02), K185 (only acetylated in NF-PitNETs), K324 and K239 (only acetylated in controls) in mitochondrial malate dehydrogenase (ID: P40926), K267  in mitochondrial dihydrolipoylde hydrogenase (ID: P09622), and K538 (only acetylated in NF-PitNETs) in mitochondrial succinate dehydrogenase [ubiquinone] flavoprotein subunit (ID: P31040). The acetylation levels at most lysine residues in these enzymes were increased in NF-PitNETs. In addition, this study found acetylation occurred at other cytoplasmic enzymes enriched on TCA cycle, including acetylation at residues at K81  in cytoplasmic isocitrate dehydrogenase [NADP] (ID: O75874), and K298, and K236 (only acetylated in controls) in cytoplasmic malate dehydrogenase (ID: P40925). (v) Glyoxylate is a highly toxic substance because it is able to be rapidly oxidized to oxalate that forms insoluble crystals with calcium, which precipitates in various organs, especially the kidneys to cause renal failure. Therefore, glyoxylate metabolism in human mainly referred to glyoxylate detoxification (p=2.20E-02), K185 (only acetylated in NF-PitNETs), K324 and K239 (only acetylated in controls) in mitochondrial malate dehydrogenase (ID: P40926), K298, and K236 (only acetylated in controls) in cytoplasmic malate dehydrogenase (ID: P40925), K169 (only acetylated in controls) in catalase (ID: P04040), K202 , K257 , and K124 (only acetylated in controls) in mitochondrial acetyl-CoA acetyltransferase (ID: P24752), and K267  in mitochondrial dihydrolipoylde hydrogenase (ID: P09622). (vi) Valine, leucine and isoleucine, also known as branched-chain amino acids, are essential amino acids that have to be derived from diet, which can be oxidized in skeletal muscle for energy supply when exercise, and are also preferentially used by many tumor cells for protein synthesis and energy purposes. It is reported that many enzymes that catalyzed the degradation of valine, leucine and isoleucine were overexpressed in many cancers (p= 6.90E-03) in mitochondrial hydroxymethylglutaryl-CoA lyase (ID: P35914), K294 (only acetylated in NF-PitNETs) in mitochondrial 2-oxoisovalerate dehydrogenase subunit beta (ID: P21953), K30 (only acetylated in NF-PitNETs) in 4-trimethylaminobutyraldehyde dehydrogenase (ID: P49189), K204 (only acetylated in NF-PitNETs) in mitochondrial methylglutaconyl-CoA hydratase (ID: Q13825), K202 , K257 , and K124 (only acetylated in controls) in mitochondrial acetyl-CoA acetyltransferase (ID: P24752), and K267  in mitochondrial dihydrolipoylde hydrogenase (ID: P09622). The acetylation levels of most lysine residues at enzymes enriched in the valine, leucine and isoleucine degradation pathway increased in NF-PitNETs. (vii) Oxidative phosphorylation is the primary pathway for ATP synthesis and responsible for setting and maintaining metabolic homeostasis (p=7.61E-04) in mitochondrial cytochrome c oxidase subunit 5B (ID: P10606) and K21  in mitochondrial ATP synthase subunit epsilon (ID: P56381), the acetylation levels of all other lysine residues in enzymes enriched in oxidative phosphorylation pathway were increased in NF-PitNETs, including residues K538 (only acetylated in NF-PitNETs) in mitochondrial succinate dehydrogenase [ubiquinone] flavoprotein subunit (ID: P31040), K233, K154, and K244 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase F(0) complex subunit B1 (ID: P24539), K522 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit beta (ID: P06576), K60 , K199 and K162 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit O (ID: P48047), K136 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit delta (ID: P30049), K539 , and K427 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit alpha (ID: P25705), K63 , and K72  in mitochondrial ATP synthase subunit d (ID: O75947), and K37  in mitochondrial ATP synthase subunit epsilon (ID: P56381). (viii) Metabolic pathway involves many interconnected cellular pathways that ultimately provide cells with energy required to execute their function (p=1.52E-02) in transaldolase (ID: P37837), K194 , and K117  in glyceraldehyde-3-phosphate dehydrogenase (ID: P04406), K81  in cytoplasmic isocitrate dehydrogenase [NADP] (ID: O75874), K314 , K185 (only acetylated in NF-PitNETs), K324 and K239 (only acetylated in controls) in mitochondrial malate dehydrogenase (ID: P40926), K298, and K236 (only acetylated in controls) in cytoplasmic malate dehydrogenase (ID: P40925), K7 (only acetylated in controls) in alcohol dehydrogenase class-3 (ID: P11766), K39  in phosphoglycerate mutase 1 (ID: P18669), K302  in mitochondrial aspartate aminotransferase (ID: P00505), K291, K146, K156 and K361 (only acetylated in controls) in phosphoglycerate kinase 1 (ID: P00558), K538 (only acetylated in NF-PitNETs) in mitochondrial succinate dehydrogenase [ubiquinone] flavoprotein subunit (ID: P31040), K231(only acetylated in controls), and K225  in triosephosphate isomerase (ID: P60174), and K330 and K200 (only acetylated in controls) in fructose-bisphosphate aldolase A (ID: P04075), K202 , K257 , and K124 (only acetylated in controls) in mitochondrial acetyl-CoA acetyltransferase (ID: P24752), K267  in mitochondrial dihydrolipoylde hydrogenase (ID: P09622), K63 , and K72  in mitochondrial ATP synthase subunit d (ID: O75947), K30 (only acetylated in NF-PitNETs) in 4-trimethylaminobutyraldehyde dehydrogenase (ID: P49189), K539 , and K427 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit alpha (ID: P25705), K233, K154, and K244 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase F(0) complex subunit B1 (ID: P24539), K522 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit beta (ID: P06576), K60 , K199 and K162 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit O (ID: P48047), K86  in mitochondrial cytochrome c oxidase subunit 5B (ID: P10606), K37 , and K21  in mitochondrial ATP synthase subunit epsilon (ID: P56381), K294 (only acetylated in NF-PitNETs) in mitochondrial 2-oxoisovalerate dehydrogenase subunit beta (ID: P21953), K26 (only acetylated in controls) in nucleoside diphosphate kinase A (ID: P15531), K70  in mitochondrial NAD(P) transhydrogenase (ID: Q13423), K1704  in fatty acid synthase (ID: P49327), K189 (only acetylated in NF-PitNETs) in mitochondrial monofunctional C1-tetrahydrofolate synthase (ID: Q6UB35), K419  in retinal dehydrogenase 1 (ID: P00352), K48  in mitochondrial hydroxymethylglutaryl-CoA lyase (ID: P35914), K204 (only acetylated in NF-PitNETs) in mitochondrial methylglutaconyl-CoA hydratase (ID: Q13825), and K136 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit delta (ID: P30049). These acetylated proteins and acetylation sites involved in the metabolic pathway would expand the research data for the study of NF-PitNETs pathogenesis. (ix) Biosynthesis of amino acids is a crucial process constructing the precursor of proteins to participate in vital movement. The deregulated catabolism/anabolism of amino acids, especially serine, glutamine and glycine, were reported to function as metabolic regulators in promoting cancer cell growth (p=1.52E-02) in transaldolase (ID: P37837), K194 , and K117  in glyceraldehyde-3-phosphate dehydrogenase (ID: P04406), K81  in cytoplasmic isocitrate dehydrogenase [NADP] (ID: O75874), K39  in phosphoglycerate mutase 1 (ID: P18669), K302  in mitochondrial aspartate aminotransferase (ID: P00505), K291, K146, K156 and K361 (only acetylated in controls) in phosphoglycerate kinase 1 (ID: P00558), K231 (only acetylated in controls), and K225  in triosephosphate isomerase (ID: P60174), and K330 and K200 (only acetylated in controls) in fructose-bisphosphate aldolase A (ID: P04075).The pathways about metabolism and energy yield included carbon metabolism, glycolysis/gluconeogenesis, pyruvate metabolism, citrate cycle (TCA cycle), glyoxylate and dicarboxylate metabolism, metabolic pathways, biosynthesis of amino acids, oxidative phosphorylation, and valine, leucine and isoleucine degradation. (i) Carbon metabolism consisted of one-carbon metabolism and central carbon metabolism . Centralase ID: P837, K194e ID: P046, K81 , and K72  in mitochondrial ATP synthase subunit d (ID: O75947), K539 , and K427 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit alpha (ID: P25705), K233, K154, and K244 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase F(0) complex subunit B1 (ID: P24539), K522 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit beta (ID: P06576), K60 , K199 and K162 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit O (ID: P48047), K86  in mitochondrial cytochrome c oxidase subunit 5B (ID: P10606), K37 , and K21  in mitochondrial ATP synthase subunit epsilon (ID:P56381), and K136 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit delta (ID: P30049), K195 (only acetylated in controls) in ubiquitin carboxyl-terminal hydrolase isozyme L1 (ID: P09936), K72 (only acetylated in controls), and K427 (only acetylated in NF-PitNETs) in voltage-dependent anion-selective channel protein 2 (ID: P45880), K23 (only acetylated in NF-PitNETs) in ADP/ATP translocase 3 (ID: P12236), K23 (only acetylated in NF-PitNETs) in ADP/ATP translocase 1 (ID: P12235), K62 (only acetylated in controls) in protein deglycase DJ-1 (ID: Q99497), and K23 (only acetylated in NF-PitNETs) ADP/ATP translocase 2 (ID: P05141). (ii) This study found acetylation occurred at the Huntington’s disease pathway-related molecules (p=9.47E-05), and K72  in mitochondrial ATP synthase subunit d (ID: O75947), K539 , and K427 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit alpha (ID: P25705), K233, K154, and K244 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase F(0) complex subunit B1 (ID: P24539), K522 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit beta (ID: P06576), K60 , K199 and K162 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit O (ID: P48047), K86  in mitochondrial cytochrome c oxidase subunit 5B (ID: P10606), K37 , and K21  in mitochondrial ATP synthase subunit epsilon (ID: P56381), and K136 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit delta (ID: P30049), K72 (only acetylated in controls), and K427 (only acetylated in NF-PitNETs) in voltage-dependent anion-selective channel protein 2 (ID: P45880), K23 (only acetylated in NF-PitNETs) in ADP/ATP translocase 3 (ID: P12236), K23 (only acetylated in NF-PitNETs) in ADP/ATP translocase 1 (ID: P12235), K23 (only acetylated in NF-PitNETs) ADP/ATP translocase 2 (ID: P05141), K123  in superoxide dismutase [Cu-Zn] (ID: P00441), and K130 (only acetylated in NF-PitNETs) in mitochondrial superoxide dismutase [Mn] (ID: P04179). (iii) This study found acetylation occurred at the Alzheimer’s disease pathway-related molecules (p=9.47E-05), and K72  in mitochondrial ATP synthase subunit d (ID: O75947), K539 , and K427 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit alpha (ID: P25705), K233, K154, and K244 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase F(0) complex subunit B1 (ID: P24539), K522 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit beta (ID: P06576), K60 , K199 and K162 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit O (ID: P48047), K86  in mitochondrial cytochrome c oxidase subunit 5B (ID: P10606), K37 , and K21  in mitochondrial ATP synthase subunit epsilon (ID: P56381), and K136 (only acetylated in NF-PitNETs) in mitochondrial ATP synthase subunit delta (ID: P30049), K194 , and K117  in glyceraldehyde-3-phosphate dehydrogenase (ID: P04406), and K133 (only acetylated in controls) in calmodulin (Fragment) (ID: H0Y7A7).Three pathways associated with nervous system diseases were Parkinson’s disease pathway, Huntington’s disease pathway, and Alzheimer’s disease pathway. The acetylated proteins enriched in these three pathways were mainly enzymes involved in metabolism and energy yield, which indicated that the acetylation mainly occurred at enzymes, and their alterations might result in extensive influence in metabolism by various pathway. (i) This study found acetylation occurred at the Parkinson’s disease pathway-related molecules  The acetylated proteins enriched in pathogenic Escherichia Coli infection pathway extensively existed in cytoplasm and nucleus. Tubulin, actin, and ezrin constituted cytoskeleton, maintained cell morphology and motility, and regulated cell cycle or cell-cell adhesion. Nucleolin participated in the cleavage of rRNA precursors, DNA replication, and cell cycle regulation  in nucleolin (ID: P19338), K440 (only acetylated in controls) in tubulin alpha-1C chain (ID: F5H5D3), K61  and K326 (only acetylated in controls) in actin cytoplasmic 1 (ID: P60709), K394  and K336  in tubulin alpha-1B chain (ID: P68363), and K35 (only acetylated in controls) in ezrin (ID: P15311). (ii) One of proteins enriched in viral carcinogenesis pathway is 14-3-3 protein. 14-3-3 protein has many subtypes, including 14-3-3 protein gamma, theta, and epsilon, and many of them have carcinogenic potential (p=2.36E-02) in histone H4 (ID: P62805), K398  in alpha-actinin-1 (ID: P12814), K47 (only acetylated in controls) in histone H2B type 1-K (ID: O60814), K68 (only acetylated in controls) in 14-3-3 protein theta (ID: P27348), K150 (only acetylated in controls) in ran-specific GTPase-activating protein (ID: P43487), and K3 and K9 (only acetylated in controls) in 14-3-3 protein zeta/delta (ID: P63104). The acetylation level decreased in 14-3-3 protein in NF-PitNETs, the alteration of which might support NF-PitNETs tumorigenesis. (iii) Human African trypanosomiasis is a potentially fatal disease caused by the Trypanosoma Brucei sp (a kind of parasite) (p=4.29E-02) and K91  in hemoglobin subunit alpha (ID: P69905), K96 (only acetylated in controls) in mutant hemoglobin beta chain (Fragment) (ID: Q9BWU5), K57 (only acetylated in controls) in hemoglobin alpha-1 globin chain (Fragment) ID: E9M4D4), K12  in alpha globin chain (Fragment) (ID: U3PXP0), K96  in hemoglobin subunit beta (ID: P68871), K157 , K262, K230, and K206 (only acetylated in controls) in apolipoprotein A-I (ID: P02647), and K17  in alpha-2 globin chain (ID: D1MGQ2).Three pathways pathogenic gulation    (Supplep=1.52E-02) in transaldolase (ID: P37837), K194 , and K117  in glyceraldehyde-3-phosphate dehydrogenase (ID: P04406), K294 (only acetylated in NF-PitNETs) in mitochondrial 2-oxoisovalerate dehydrogenase subunit beta (ID: P21953), K81  in isocitrate dehydrogenase [NADP] cytoplasmic (ID: O75874), K314 , K185 (only acetylated in NF-PitNETs), K324 and K239 (only acetylated in controls) in mitochondrial malate dehydrogenase (ID: P40926), K298, and K236 (only acetylated in controls) in cytoplasmic malate dehydrogenase (ID: P40925), K7 (only acetylated in controls) in alcohol dehydrogenase class-3 (ID: P11766), K39  in phosphoglycerate mutase 1 (ID: P18669), K302  in mitochondrial aspartate aminotransferase (ID: P00505), K291, K146, K156 and K361 (only acetylated in controls) in phosphoglycerate kinase 1 (ID: P00558), K538 (only acetylated in NF-PitNETs) in mitochondrial succinate dehydrogenase [ubiquinone] flavoprotein subunit (ID: P31040), K231 (only acetylated in controls), and K225  in triosephosphate isomerase (ID: P60174), K330 and K200 (only acetylated in controls) in fructose-bisphosphate aldolase A (ID: P04075), K202 , K257 , and K124 (only acetylated in controls) in mitochondrial acetyl-CoA acetyltransferase (ID: P24752), and K267  in mitochondrial dihydrolipoylde hydrogenase (ID: P09622), K30 (only acetylated in NF-PitNETs) in 4-trimethylaminobutyraldehyde dehydrogenase (ID: P49189), and K169 (only acetylated in controls) in catalase (ID: P04040). The acetylation levels of most of these proteins were decreased in NF-PitNETs.In recent years, mammalian immune cells were found to synthesize antibiotics, itaconic acid, from citric acid cycle intermediate, to prevent bacteria from surviving in cells  (Supplep=6.90E-03) in mitochondrial hydroxymethylglutaryl-CoA lyase (ID: P35914), K81  in isocitrate dehydrogenase [NADP] cytoplasmic (ID: O75874), K130 , and K169 (only acetylated in controls) in peroxiredoxin-1 (ID: Q06830), K169 (only acetylated in controls) in catalase (ID: P04040), K123  in superoxide dismutase [Cu-Zn] (ID: P00441), and K130 (only acetylated in NF-PitNETs) in mitochondrial superoxide dismutase [Mn] (ID: P04179).Peroxisome is consisted of many kinds of oxidases, and contributes to cellular lipid metabolism and redox balance. Peroxisome has ability of detoxification, including removal of superoxide radicals originated from respiratory chain . The dysp=2.05E-03) and K153 (only acetylated in controls) in alpha-1-antitrypsin (ID: P01009), and K1176 (only acetylated in controls) in alpha-2-macroglobulin (ID: P01023).The last pathway was complement and coagulation cascade pathway, and all these DAPs enriched in this pathway were from blood. In the blood circulation, the coagulation system, platelets, complement system, and fibrinolysis system constructed a close and complex network. They activated and regulated each other, and mutually mediated tissue homeostasis and immune monitoring. The deregulation of each cascade system caused clinical manifestations and the progression of different diseases, such as C3 glomerulonephritis, sepsis, and systemic lupus erythematosus   proteins that were modified by both acetyl and ubiquitin in NF-PitNETs were involved in nucleosome or ribosome, hemoglobin, prolactin, ubiquitin hydrolase, membrane proteins, and proteins constituting cytoskeleton; and acetylation levels of these proteins were decreased, whereas their ubiquitination levels were increased in NF-PitNETs. (ii) The invasiveness-related acetylated proteins were mainly involved in biological processes and signaling pathways about metabolism and energy yield, which suggested that NF-PitNET invasive behaviors might be acetylation-mediated metabolic reprogramming process.Many tumor cells prefer to provide bioenergetics and growth requirements through glycolysis, rather than oxidative phosphorylation, during tumor growth progression, even with sufficient oxygen and normal mitochondria. Because glycolysis is able to provide sufficient cellular ATPs, and additional important metabolites to support the biosynthetic demands of consecutive cell proliferation . It is rThe invasive characteristics of NF-PitNETs have been a hot pot for a long time. Invasive PitNETs tend to suffer tumor postoperative residual and re-growth because of cavernous sinus invasion or the internal carotid artery encircling, which is able to cause poor prognosis. However, their underlying invasive mechanism remains unclear . This stThe co-regulation of acetylation and ubiquitination at specific lysine sites in some proteins might affect tumor development, such as Lys382 of p53 and Lys125 of SRSF5 , 25. OneMoreover, NF-PitNET and control tissue samples were very limited and precious, only very limited amount of proteins were obtained for subsequent quantitative acetylomics analysis. More acetylated sites and acetylated proteins are promised to be identified when the increased NF-PitNET protein samples available in future acetylomics analysis. This acetylome map of human NF-PitNETs described in this study is one component in the long-term program to find out NF-PitNET-specific acetylated proteins to clarify molecular mechanisms of NF-PitNETs. To achieve this goal, quantitative acetylomics needs to be further developed in the future.In summary, the current study provided the first human acetylomic data in NF-PitNETs, offered a valuable resource for further study in NF-PitNET tumorigenesis and progression, which contributed to the discovery of effective biomarkers for early diagnosis and therapy of NF-PitNETs.in vivo acetylated proteins and their acetylation sites, and to rationalize the functions of DAPs. A total of 296 acetylated proteins with 517 acetylated lysine sites provided a quantitative status of lysine acetylation in NF-PitNETs, and their bioinformatics analysis provided a new insight into the roles of protein lysine acetylation in formation and development of NF-PitNETs. The acetylation levels of more than half acetylated proteins were decreased in NF-PitNETs. Acetylation-mediated metabolic reprogramming might be considered as one of the underlying mechanisms in tumorigenesis and invasiveness of NF-PitNETs. Further investigation is needed to ascertain the biological significance of these lysine acetylation events and their relevance to NF-PitNET pathogenesis.This study provided a comprehensive approach that integrated anti-acetyl antibody-based enrichment, LC-MS/MS, and literature-based bioinformatics to discover The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/The studies involving human participants were reviewed and approved by the Xiangya Hospital Medical Ethics Committee of Central South University; the University of Tennessee Health Science Center Internal Review Board. The patients/participants provided their written informed consent to participate in this study.SW analyzed data, carried out western blot experiment and immunoprecipitation experiment, prepared figures and tables, designed and wrote the manuscript. JL, JY, BL, and NL participated in partial data analysis and experiments. XZ conceived the concept, designed experiments and manuscript, instructed experiments, analyzed data, obtained the acetylomics data, supervised results, coordinated, wrote and critically revised manuscript, and was responsible for its financial supports and the corresponding works. All authors contributed to the article and approved the submitted version.This work was supported by the Shandong First Medical University Talent Introduction Funds (to XZ), the Hunan Provincial Hundred Talent Plan (to XZ), Shandong Provincial Natural Science Foundation (ZR202103020356 to XZ), the National Natural Science Foundation of China (82172866), and the Academic Promotion Program of Shandong First Medical University (2019ZL002).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Correction to: BMC Infect Dis 21, 571 (2021)https://doi.org/10.1186/s12879-021-06287-1Following publication of the original article , the autFurthermore, a typo was found in an author’s affiliation.The incorrect affiliation name is:Center for Research in Epidemiology and Population Health ou (CESP), INSERM U1018, Paris-Saclay University, UVSQ, Montigny-Le-Bretonneux, FranceThe correct affiliation name is:Center for Research in Epidemiology and Population Health (CESP), INSERM U1018, Paris-Saclay University, UVSQ, Montigny-Le-Bretonneux, FranceThe original article has been corrected as well.Additional file 1: Supplement S1: Database description and patient selection. Table S1: ICD-10 codes used. Table S2: Selected bacteria included and associated resistance markers. Supplement S2: Algorithms for identification of risk factors. Table S3: Code list for definition of potential risk factors. Table S4: Conditional univariate logistic regression: Risk factors of having a community-acquired or healthcare-associated urinary tract infection caused by a resistant bacterium compared with a susceptible one, by gender. Table S5: Number of antibiotic dispensing during the previous 3 months. Table S6. Characteristics of patients excluded and included for analysis of association between antibiotic classes and resistant-bacterial acquisition.


First report of the fall armyworm, Spodoptera frugiperda (Lepidoptera: Noctuidae), an alien invasive pest on maize in India. Pest Manag Horticult Ecosyst. 24:23–29.The correct citation should be as follows; this has been updated:Sharanabasappa D, et al. 2018. First report of the Fall armyworm, Spodoptera frugiperda (JE Smith) (Lepidoptera: Noctuidae), an alien invasive pest onmaize in India. Pest Manag Horticult Ecosyst. 24:23–29.


Correction to: BMC Cancer 21, 378 (2021)https://doi.org/10.1186/s12885-021-08019-9Following publication of the original article , the autThe incorrect author name is: Fatima SkayaliThe correct author name is: Fatima KayaliThe Contributions section has therefore also been updated as follows:RJE and AH devised the study, AH, MEG, & FK extracted data, AH & BR analysed data, AC & EJC provided interpretation and assisted writing the manuscript. All authors have read and approved the manuscript.The author group has been updated above and the original article  has been


Mytilus coruscus) is widely distributed in the temperate seas of East Asia and is an important commercial bivalve in China. Chromosome-level genome information of this species will contribute not only to the development of hard-shelled mussel genetic breeding but also to studies on larval ecology, climate change biology, marine biology, aquaculture, biofouling, and antifouling.The hard-shelled mussel (M. coruscus is the sister taxon to the clade including Modiolus philippinarum and Bathymodiolus platifrons. Conserved chromosome synteny was observed between hard-shelled mussel and king scallop, suggesting that this is shared ancestrally. Transcriptomic profiling indicated that the pathways of catecholamine biosynthesis and adrenergic signaling in cardiomyocytes might be involved in metamorphosis.We applied a combination of Illumina sequencing, Oxford Nanopore Technologies sequencing, and high-throughput chromosome conformation capture technologies to construct a chromosome-level genome of the hard-shelled mussel, with a total length of 1.57 Gb and a median contig length of 1.49 Mb. Approximately 90.9% of the assemblies were anchored to 14 linkage groups. We assayed the genome completeness using BUSCO. In the metazoan dataset, the present assemblies have 89.4% complete, 1.9% incomplete, and 8.7% missing BUSCOs. Gene modeling enabled the annotation of 37,478 protein-coding genes and 26,917 non-coding RNA loci. Phylogenetic analysis showed that The chromosome-level assembly of the hard-shelled mussel genome will provide novel insights into mussel genome evolution and serve as a fundamental platform for studies regarding the planktonic-sessile transition, genetic diversity, and genomic breeding of this bivalve. Marine mussels, which belong to the phylum Mollusca, settle on most immersed surfaces of substrata and play a crucial role in marine ecosystems. As healthy and sustainable food items, these mussels are beneficial for humans owing to their high economic value for fishery and aquaculture, constituting >8% of mollusc aquaculture production . SimultaLike many other marine invertebrates, marine mussels also possess a free-swimming larval phase. After this stage, these minute larvae will settle on the substrata and finish metamorphosis transition, accompanied by dramatic remodeling of their anatomy , 15. MulMytilus coruscus Gould 1861, NCBI:txid42192, Fig. Mytilus has been assembled at the chromosome level, although a draft genome of M. coruscus ab initioon 2.2.1 , Swisspron 2.2.1 , 51 or aChlamys farreri (PRJNA185465), Pinctada fucata martensii (GCA_002216045.1), M. philippinarum (GCA_002080025.1), Crassostrea gigas (GCF_000297895.1), B. platifrons (GCA_002080005.1), Mizuhopecten yessoensis(GCA_002113 885.2), Penaeus vannamei (ASM378908v1), Pecten maximus (GCA_902652985.1), Scapharca (Anadara) broughtonii (PRJNA521075), Pomacea canaliculata (PRJNA427478), Haliotis discus hannai (PRJNA317403), and M. coruscus, using OrthoMCL  version 1.4 with a BLASTp cut-off value of 10−5 and an inflation value of 1.5  were identified in D-veliger, umbo, pediveliger, and juvenile larvae, respectively. Functional annotation indicated that these were mainly involved in “environmental information processing”  and “cellular processes” (“transport and catabolism”), in agreement with the key role of signal transduction and the endocrine system in larval development [To profile gene expression during development and metamorphosis in hard-shelled mussels, RNA-seq analysis was conducted at 5 developmental stages: trochophore, D-veliger, umbo, pediveliger, and juvenile (PRJNA689932). The quantification of gene expression enabled the detection of 33,743 transcripts with TPMs > 0 at all stages . The limelopment .Because the ability to effectuate metamorphosis develops during the pediveliger stage , we inveRRID:SCR_015008) v4.1.4 referencing metazoan and molluscan gene sets. In the metazoan dataset, the present assemblies have 89.4% complete (of which 1.0% were duplicated), 1.9% incomplete, and 8.7% missing BUSCOs, corresponding to a recovery of 91.3% of the entire BUSCO set. In the molluscan dataset, 85.5% complete (of which 1.3% were duplicated), 0.8% incomplete, and 13.7% missing BUSCOs were recorded, corresponding to 86.3% of the entire BUSCO set. Motifs with the characteristics of telomeric repeats were detected in 23 termini of the 13 chromosomes, suggesting the completeness of the assemblies  are available in Figshare [All of the raw Illumina and ONT read data underlying this article were deposited to NCBI SRA and the assembled genome was deposited to GenBank, under the accession No. Figshare  and GigaFigshare .Supplementary Table S1. Repetitive sequences in the hard-shelled mussel genomeSupplementary Table S2. Overview of the predicted non-coding RNAsSupplementary Table S3. Bidirectional BLASTp between the previously published gene models of the hard-shelled mussel and the predicted gene models in this studySupplementary Table S4. Gene expression profiles during 5 developmental stagesSupplementary Table S5. Genes involved in the pathways of catecholamine biosynthesis and adrenergic signaling in the cardiomyocytes were reported to affect metamorphosisSupplementary Table S6. Information on the motifs with the characteristic of telomeric repeatsSupplementary Figure S1. Circles showing genome-wide SNPs and indels from the farmed and wild populations. From the outer to the inner circle: first circle, marker distribution across 14 pseudochromosomes at a megabase scale; green circle, SNP density across the whole genome; red circle, indel density.α-ARA: alpha-1A adrenergic receptor-like; α-ARB: adrenergic receptor alpha-1B; β2AR: adrenergic receptor beta-2; AC1: adenylate cyclase 1; AC10: adenylate cyclase 10; Akt: RAC serine/threonine-protein kinase; BLAST: Basic Local Alignment Search Tool; bp: base pairs; BUSCO: Benchmarking Universal Single-Copy Orthologs; BWA: Burrows-Wheeler Aligner; CaM: calmodulin; CaMKII: calcium/calmodulin-dependent protein kinase (CaM kinase) II; CAV1: caveolin 1; CAV3: caveolin 3; CREB: cyclic AMP-responsive element-binding protein; DBH: dopamine beta-monooxygenase; DDC: aromatic-L-amino-acid decarboxylase; DHPR: voltage-dependent calcium channel gamma-1; Epac: Rap guanine nucleotide exchange factor; ERK: mitogen-activated protein kinase 1/3; FPS: farmed population specific; GATK: Genome Analysis Tool Kit; Gb: gigabase pairs; GC: guanine-cytosine; Gi: guanine nucleotide-binding protein G(i) subunit alpha; GO: Gene Ontology; Gq: guanine nucleotide-binding protein G(q) subunit alpha; Gs: guanine nucleotide-binding protein G(s) subunit alpha; Hi-C: high-throughput chromosome conformation capture; ICER: cAMP response element modulator; IKS: potassium voltage-gated channel KQT-like subfamily member 1; IMP2: mitochondrial inner membrane protease subunit 2; INaK: sodium/potassium-transporting ATPase subunit alpha; kb: kilobase pairs; KEGG: Kyoto Encyclopedia of Genes and Genomes; MAOA: monoamine oxidase A; MAOB: monoamine oxidase B; Mb: megabase pairs; MSK1: ribosomal protein S6 kinase alpha-5; NCBI: National Center for Biotechnology Information; NCX: solute carrier family 8 ; NF-κB: nuclear factor NF-kappa-B p105 subunit; NHE: solute carrier family 9 (sodium/hydrogen exchanger); ONT: Oxford Nanopore Technologies; p38MAPK: p38 MAP kinase; PASA: Program to Assemble Spliced Alignments; PE: paired end; PI3K: phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha/beta/delta; PKA: protein kinase A; PKCα: classical protein kinase C alpha type; PLC: phosphatidylinositol phospholipase C; PP1: serine/threonine-protein phosphatase PP1 catalytic subunit; RAxML: Randomized Axelerated Maximum Likelihood; RNA-Seq: RNA sequencing; SNP: single-nucleotide polymorphism; SRA: Sequence Read Archive; SSR: simple sequence repeat; TnI: Troponin I; TPMs: transcripts per million; TPM: tropomyosin; tRNA: transfer RNA; TYR: tyrosinase; WPS: wild population specific.The authors declare that they have no competing interests.This study was supported by the National Key Research and Development Program of China (2018YFD0900601), Key Program for International Science and Technology Cooperation Projects of Ministry of Science and Technology of China (No. 2018YFD0900101), Key Special Project for Introduced Talents Team of Southern Marine Science and Engineering Guangdong Laboratory (Guangzhou) (GML2019ZD0402), the National Natural Science Foundation of China , Program of Shanghai Academic Research Leader (20XD1421800), "Science and Technology Innovation Action Plan" The Belt and Road International Joint Laboratory Construction Projects (590750500), the Shanghai Sailing Program (18YF1410000), China Postdoctoral Science Foundation (2019M6614770), and Program for study on genetic resources, environment and strategy of mussel culture in coast of Gouqi Island offshore.J.L.Y., Y.L., and X.L. designed and supervised the study. K.C., J.K.X, Y.T.Z., and Y.F.L. collected the samples and extracted the genomic DNA and RNA. Y.L., J.L., and D.D.F. performed genome assembly and bioinformatics analysis. J.L.Y., D.D.F., X.L., J.L., and Y.L. wrote the original manuscript. All authors reviewed the manuscript.giab024_GIGA-D-20-00287_Original_SubmissionClick here for additional data file.giab024_GIGA-D-20-00287_Revision_1Click here for additional data file.giab024_GIGA-D-20-00287_Revision_2Click here for additional data file.giab024_Response_to_Reviewer_Comments_Original_SubmissionClick here for additional data file.giab024_Response_to_Reviewer_Comments_Revision_1Click here for additional data file.giab024_Reviewer_1_Report_Original_SubmissionMarco Gerdol -- 10/19/2020 ReviewedClick here for additional data file.giab024_Reviewer_1_Report_Revision_1Marco Gerdol -- 2/2/2021 ReviewedClick here for additional data file.giab024_Reviewer_2_Report_Original_SubmissionNathan Kenny -- 10/24/2020 ReviewedClick here for additional data file.giab024_Reviewer_2_Report_Revision_1Nathan Kenny -- 1/30/2021 ReviewedClick here for additional data file.giab024_Reviewer_3_Report_Original_SubmissionYi-Jyun Luo -- 10/26/2020 ReviewedClick here for additional data file.giab024_Reviewer_3_Report_Revision_1Yi-Jyun Luo -- 2/18/2021 ReviewedClick here for additional data file.giab024_Supplemental_FilesClick here for additional data file.


Differential and longitudinal immune gene patterns associated with reprogrammed microenvironment and viral mimicry in response to neoadjuvant radiotherapy in rectal cancer. J Immunother Cancer 2021;9:e001717. doi: 10.1136/jitc-2020-001717Wilkins A, Fontana E, Nyamundanda G, This paper has been updated since first published to amend author and ethics committee details.


A systemically deliverable Vaccinia virus with increased capacity for intertumoral and intratumoral spread effectively treats pancreatic cancer. J ImmunoThera Cancer 2021;9:e001624. doi:10.1136/jitc-2020-001624Marelli G, Chard Dunmall LS, Yuan M, This article has been corrected since it first published. The provenance and peer review statement has been added.


Phoneutria, mainly from P.nigriventer. In spite of the pharmaceutical potential demonstratedby P. nigriventer toxins, there is limited information onmolecules from venoms of the same genus, as their toxins remain poorlycharacterized. Understanding this diversity and clarifying the differencesin the mechanisms of action of spider toxins is of great importance forestablishing their true biotechnological potential. This prompted us tocompare three different venoms of the Phoneutria genus:P. nigriventer (Pn-V), P. eickstedtae(Pe-V) and P. pertyi (Pp-V).  Spider venoms induce different physio-pharmacological effects by bindingwith high affinity on molecular targets, therefore being of biotechnologicalinterest. Some of these toxins, acting on different types of ion channels,have been identified in the venom of spiders of the genus Biochemical and functional comparison of the venoms were carried out bySDS-PAGE, HPLC, mass spectrometry, enzymatic activities andelectrophysiological assays (whole-cell patch clamp).  The employed approach revealed that all three venoms had an overallsimilarity in their components, with only minor differences. The presence ofa high number of similar proteins was evident, particularly toxins in themass range of ~6.0 kDa. Hyaluronidase and proteolytic activities weredetected in all venoms, in addition to isoforms of the toxins Tx1 and Tx2-6.All Tx1 isoforms blocked Nav1.6 ion currents, with slight differences. Phoneutria genus, exposing their biotechnologicalpotential as a source for searching for new active molecules. Our findings showed that Pn-V, Pe-V and Pp-V are highly similar concerningprotein composition and enzymatic activities, containing isoforms of thesame toxins sharing high sequence homology, with minor modifications.However, these structural and functional variations are very important forvenom diversity. In addition, our findings will contribute to thecomprehension of the molecular diversity of the venoms of the other speciesfrom  Among the components of spider venoms, a wide variety of toxins evolved to bind tospecific targets of biological importance, such as ion channels, inducing biologicaleffects that allow spiders to immobilize their prey in a very efficient way, usingminor quantities of their toxins ,2. Thesethcentury and have advanced enormously up to the present time, suggesting theimmeasurable biotechnological potential of these molecules ethyl-(carboxymethyl)amino]acetic acid; EGTA: ethylene glycol-bis-(b-amino-ethyl ether)N,N,N´,N´-tetra-acetic acid; ESI: electrospray ionization; GMP: guanosinemonophosphate; HA: hyaluronic acid; HEPES:4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; HPLC: high-pressure liquidchromatography; IAA: iodoacetamide; KOH: potassium hydroxide; LC: liquidchromatography; MALDI-TOF: matrix associated laser desorption/ionization-time offlight; MgCl2: magnesium chloride; MS: mass spectrometry; MW: massweight; NaCl: sodium chloride; NaOH: sodium hydroxide; Nav: typevoltage-dependent sodium channel; NMDA N-methyl d-aspartate; NO: nitric oxide;Pe: Phoneutria eickstedtae; Pe-V: Phoneutriaeickstedtae venom; Ph: Phoneutria; Pn:Phoneutria nigriventer; Pn-V: Phoneutrianigriventer venom; Pp: Phoneutria pertyi; Pp-V:Phoneutria pertyi venom; RP: reverse-phase; SDS-PAGE:sodium dodecyl sulfate polyacrylamide gel electrophoresis; TEA-Cl:tetraethylammonium chloride; TFA: trifluoracetic acid; TNBS:2,4,6-trinitrobenzenesulfonic acid; Tx: toxin; UHPLC: ultra-high-pressure liquidchromatography. ACN: acetonitrile; ACU: acquired colorimetric unit; AMBIC: ammonium bicarbonate;CaCl


Andropromachus (Lonchodidae: Necrosciinae: Necrosciini) is a small genus of stick insects including four species with a distribution in northern Vietnam and south-western China.The genus Andropromachusgulinqingensissp. n. is described from Yunnan Province of China. Diagnostic characters of the new species are illustrated and an identification key to male and female Andropromachus is provided along with a checklist for all described species.A new species of stick insects  Andropromachus, including the two species A.scutatus Carl and A.bicolor Kirby, was erected within the tribe Necrosciini (Phasmatodea: Lonchodidae: Necrosciinae) by Lonchodinae by A.scutatus as the type-species. Spiniphasma Chen & He, 2000 is a junior synonym of the genus Andropromachus, and therefore transferred the species S.guangxiense to Andropromachus. Promachustonkinensis Brunner von Wattenwyl to Andropromachus as A.bicolor Kirby, which cannot be further examined because the type specimen of A.tonkinensis was lost and the original description was incomplete. Andropromachus back to Necrosciinae. A.scutatus was included in a recent phylogenetic study that also supported Andropromachus belonging to Necrosciinae .The genus tergites . At preshe world . Here, whe world  and two Adults, caught in the wild, were reared in boxes and some host plants (fern) were placed inside the boxes until female oviposition. After adults and eggs were killed by low temperatures (−20℃ to ca. −40℃), adults were pinned and eggs stored in small tubes. All materials studied were deposited in the Insect Collection of the Southwest Forestry University, Yunnan Province, China (SWFU).Morphological observations were made with a SOPTOP SZ stereomicroscope . Digital images were obtained using a Liyang Super Resolution System LY-WN-YH . Whole view images of the new specimens were taken with a Canon 5ds digital camera and LAOWA 100 mm F2.8 2X macro lens . Stacking was done using the software Zerene Stacker . Morphological terminology follows that of Bragg (2001)Xie & Qiansp. n.DFB7216B-DCFD-5416-AE84-268668CA5F8C303602BC-E9B7-4934-9CF7-7DAF8289E945Type status:Holotype. Occurrence: recordedBy: Xiang-Jin Liu; sex: female; lifeStage: adults; Taxon: scientificName: Andropromachusgulinqingensis; order: Phasmatodea; family: Lonchodidae; genus: Andropromachus; Location: country: China; stateProvince: Yunnan Province; county: Maguan County; municipality: Wenshan Zhuang and Miao Autonomous Prefecture; verbatimLocality: Maguan County, Gulinqing Provincial Nature Reserve; verbatimLatitude: N22.81339°; verbatimLongitude: E103.97092°; Event: year: 2020; month: 7; day: 15; Record Level: institutionCode: Southwest Forestry University, Yunnan Province, China (SWFU)Type status:Paratype. Occurrence: recordedBy: Xiang-Jin Liu; sex: 2 females, 2 males, 10 eggs; lifeStage: adults; Taxon: scientificName: Andropromachusgulinqingensis; order: Phasmatodea; family: Lonchodidae; genus: Andropromachus; Location: country: China; stateProvince: Yunnan Province; county: Maguan County; municipality: Wenshan Zhuang and Miao Autonomous Prefecture; verbatimLocality: Maguan County, Gulinqing Provincial Nature Reserve; verbatimLatitude: N22.81339°; verbatimLongitude: E103.97092°; Event: year: 2020; month: 7; day: 15; Record Level: institutionCode: Southwest Forestry University, Yunnan Province, China (SWFU)Medium size. Body robust. The general colouration of the body is green Fig. A-G.Head. Globose, longer than wide, vertex flat, sparsely covered with a few small granules and interspersed with a few acute small granules. Compound eyes rounded, occupying 1/4 of the genae. Antennae filiform, longer than forelegs; scapus rectangular, flattened and longer than pedicellus, pedicellus cylindrical and shorter than the third segment. Occiput prominently swollen and convex, with three pairs of spines on both sides of the median longitudinal sulci; anterior spines behind the compound eyes; median spines largest and with a few small branches; posterior spines close to median spines , but can be distinguished by: ♀♀, middle size (body length > 50 mm), without spines between the compound eyes (Fig. yes Fig. C, abdomiyes Fig. A and B, yes Fig. E, all feyes Fig. A and B; This specific epithet is derived from Gulinqing Provincial Nature Reserve where it was collected.


S. aureus bacteremia (SAB).Bacterial bloodstream infections (BSI) are one of the most described syndromes in infectious diseases, but the presence of racial disparities in BSI is unclear. The purpose of this project was to determine if racial disparities exist in patients with S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) infections. Statistical comparisons were performed for binary variables with Fisher’s Exact test and continuous variables with Kruskal-Wallis test.Data was used from a prospective cohort of patients with SAB at Duke University Medical Center from 1995-2015. Patients were categorized as African American (AA) or White. Characteristics of interest included discharge disposition, metastatic infection, persistence of SAB, and in-hospital mortality stratified by methicillin-susceptible Among the 2396 patients with SAB, 1496 (62.4%) were White and 900 (37.6%) were AA. 1241 patients (51.8%) had MSSA bacteremia overall. Whites comprised 63.6% of MSSA and 61.2% of MRSA infections. AA were younger  and more likely to be female . AA had higher rates of diabetes, hemodialysis, HIV infection for both MSSA and MRSA, but higher rates of injection drug use for MSSA only; Whites had higher rates of neoplasm, corticosteroid use, surgery for MSSA and MRSA, but higher rates of transplant for MRSA only . AA had higher Acute Physiology Scores . AA experienced increased rates of healthcare-associated infection . Although Whites were more likely to have in-hospital mortality for MRSA , discharge disposition, metastatic infection, and persistence did not vary significantly by race.Racial disparities exist in SAB, more so for patient characteristics than for outcomes. AA patients were younger, had a different set of comorbidities, and had more acute presentations. Although Whites had higher rates of in-hospital mortality, all other outcomes were similar.Vance G. Fowler, Jr., MD, MHS, Achaogen (Consultant)Advanced Liquid Logics (Grant/Research Support)Affinergy Affinium (Consultant)Akagera (Consultant)Allergan (Grant/Research Support)Amphliphi Biosciences (Consultant)Aridis (Consultant)Armata (Consultant)Basilea Bayer (Consultant)C3J (Consultant)Cerexa Contrafect Debiopharm Destiny (Consultant)Durata Genentech Green Cross Integrated Biotherapeutics (Consultant)Janssen Karius (Grant/Research Support)Locus (Grant/Research Support)Medical Biosurfaces (Grant/Research Support)Medicines Co. (Consultant)MedImmune Merck (Grant/Research Support)NIH (Grant/Research Support)Novadigm (Consultant)Novartis Pfizer (Grant/Research Support)Regeneron sepsis diagnostics Tetraphase (Consultant)Theravance Trius (Consultant)UpToDate Valanbio xBiotech (Consultant)


Incorrect name: Miriam Ly-Le MoalCorrect name: Myriam Ly‐Le MoalAn incorrect author nameIncorrect affiliation: Roche Pharma Research and Early Development, Roche Innovation Center Basel, F. Hofmann-La Roche Ltd., Grenzacherstrasse 124, 4070 Basel, Switzerland.Correct affiliation: Institut Jean Nicod, ENS, ParisThe wrong affiliation was listed for Tiziana ZallaThe author group has been updated above, and the original article [Following publication of the original article 1], the authors identified 2 errors:, the aut


Correction to: BMC Genomics 23, 20 (2022)https://doi.org/10.1186/s12864-021-08214-9Following publication of the original article , it was The incorrect sentence was:“Minimum inhibitory concentrations (MICs) were determined for the following 17 diferent antibiotics: cefotaxime (CTX), ceftazidime (CAZ), cefepime (FEP), colistin (CT), tigecycline (TGC), imipenem (IMP), ertapenem (ETP), meropenem (MEM), ciprofoxacin (CIP), fosfomycin (FOS), trimethoprim-sulfamethoxazole (SXT), piperacillin-tazobactam (PTZ), amikacin (AMK), gentamicin (GEN), chloramphenicol (CHL), tetracycline [5], and aztreonam (ATM) for EBSI041 using the agar microdilution method excepted for colistin using the broth microdilution method.”The correct sentence is:(TET), and aztreonam (ATM) for EBSI041 using the agar microdilution method excepted for colistin using the broth microdilution method.Minimum inhibitory concentrations (MICs) were determined for the following 17 diferent antibiotics: cefotaxime (CTX), ceftazidime (CAZ), cefepime (FEP), colistin (CT), tigecycline (TGC), imipenem (IMP), ertapenem (ETP), meropenem (MEM), ciprofoxacin (CIP), fosfomycin (FOS), trimethoprim-sulfamethoxazole (SXT), piperacillin-tazobactam (PTZ), amikacin (AMK), gentamicin (GEN), chloramphenicol (CHL), tetracycline The correct affiliations presentation is given in this Correction article and the original article  has been


Staphylococcus aureus (MRSA) present significant treatment challenges and can cause serious morbidity and mortality. Ceftobiprole, the active moiety of the prodrug ceftobiprole medocaril, is an advanced cephalosporin approved in many European and other countries for the treatment of adults with community- and hospital-acquired pneumonia, excluding ventilator-associated pneumonia. Ceftobiprole is currently in phase 3 clinical development to support a New Drug Application in the United States for acute bacterial skin and skin structure infections and S. aureus bacteremia. Here, the activity of ceftobiprole and comparators was evaluated against recent MDR S. aureus and MRSA clinical isolates.Multidrug-resistant (MDR) and methicillin-resistant S. aureus isolates were collected from patients with various infection types at 34 US medical centers from 2016–2020. Susceptibility to ceftobiprole and comparator agents was tested by CLSI methods. Current CLSI and EUCAST interpretive criteria were applied (Table). Isolates were categorized as MDR if they were non-susceptible  to ≥3 of the following antimicrobials: clindamycin (CM), daptomycin (DAP), erythromycin (ERY), gentamicin (GM), levofloxacin (LEV), linezolid (LZD), tetracycline (TET), tigecycline (TGC), trimethoprim-sulfamethoxazole (TMP-SMX), or vancomycin (VAN). Isolates displaying oxacillin MIC values ≥4 mg/L were categorized as MRSA.13,868 Ceftobiprole was more active than ceftaroline (CPT) against MRSA  (Table). Ceftobiprole maintained activity against 88.0% of the CPT-NS isolates, but CPT was only active against 6.5% of the ceftobiprole-NS isolates. Ceftobiprole was also highly active (97.7–100.0% S) against isolates NS to CM, DAP, ERY, GM, LEV, LZD, TET, TGC, or TMP-SMX. No VAN-NS isolates were detected. Importantly, ceftobiprole was more active (97.7% S) than CPT (83.0% S) against the subset of MDR-MRSA isolates.Conclusions: Ceftobiprole was highly active in vitro against MRSA and MDR S. aureus collected at US medical centers during 2016–2020. These results support the further development of ceftobiprole to treat S. aureus infections in the US.Leonard R. Duncan, PhD, AbbVie (formerly Allergan) (Research Grant or Support)Basilea Pharmaceutica International, Ltd. (Research Grant or Support)Cipla Therapeutics (Research Grant or Support)Cipla USA Inc. (Research Grant or Support)Department of Health and Human Services Shionogi (Research Grant or Support) Kamal Hamed, MD, MPH, Basilea Pharmaceutica International, Ltd (Employee)Department of Health and Human Services  Jennifer Smart, PhD, Basilea Pharmaceutica International, Ltd. (Employee)Department of Health and Human Services  Michael A Pfaller, MD, Basilea Pharmaceutica International, Ltd. (Research Grant or Support)Cidara Therapeutics, Inc. (Research Grant or Support)Department of Health and Human Services Pfizer, Inc. (Research Grant or Support) Helio S. Sader, MD, PhD, FIDSA, AbbVie (formerly Allergan) (Research Grant or Support)Basilea Pharmaceutica International, Ltd. (Research Grant or Support)Cipla Therapeutics (Research Grant or Support)Cipla USA Inc. (Research Grant or Support)Department of Health and Human Services Melinta Therapeutics, LLC (Research Grant or Support)Nabriva Therapeutics (Research Grant or Support)Pfizer, Inc. (Research Grant or Support)Shionogi (Research Grant or Support)Spero Therapeutics (Research Grant or Support)


Correction to: BMC Cancer 21, 676 (2021)https://doi.org/10.1186/s12885-021-08418-yBalearic Islands Health Research Institute (IdISBa), 07120 Palma de Mallorca, Illes Balears, SpainFollowing publication of the original article , the autThe affiliations have been correctly published in this correction and the original article  has been


ABSTRACT IMPACT: A short treatment of 8 obese postmenopausal women with conjugated estrogens and bazedoxifene does not alter insulin sensitivity or ectopic fat but increases serum markers of hepatic de novo lipogenesis and production of triacylglycerides. OBJECTIVES/GOALS: Combining conjugated estrogens (CE) with the selective estrogen receptor modulator bazedoxifene (BZA) is a novel, orally-administered menopausal therapy. We investigated the effect of CE/BZA on insulin sensitivity, energy metabolism, and serum metabolome in postmenopausal women with obesity. METHODS/STUDY POPULATION: We conducted a randomized, double-blind, crossover pilot trial, testing the effect of CE/BZA on cardiometabolic health in postmenopausal women. Eight postmenopausal women  were randomization to an 8-week CE/BZA or placebo treatment separated by an 8-week washout period [NCT02274571]. The primary outcome was insulin sensitivity (hyperinsulinemic-euglycemic clamp), while secondary outcomes included body composition (DXA); resting metabolic rate (RMR); substrate oxidation ; ectopic lipids (1H-MRS); fat cell size, adipose and skeletal muscle gene expression (biopsies); inflammatory markers; and serum metabolome (LC/MS). RESULTS/ANTICIPATED RESULTS: CE/BZA had no effect on insulin sensitivity, body composition, ectopic fat, or substrate oxidation, but resulted in a non-significant increase in RMR  compared to placebo. CE/BZA increased serum high-density lipoprotein cholesterol. CE/BZA also increased serum diacylglycerol (DAG) and triacylglycerol (TAG) species containing long-chain saturated, mono- and polyunsaturated fatty acids (FAs), and decreased long-chain acylcarnitines. These findings possibly reflect increased hepatic de novo FA synthesis and esterification into TAGs, and decreased FA oxidation, respectively (p<0.05). CE/BZA increased serum phosphatidylcholines, phosphatidylethanolamines, ceramides, and sphingomyelins, possibly reflecting the increase in lipoproteins (p<0.05). DISCUSSION/SIGNIFICANCE OF FINDINGS: A short treatment of postmenopausal women with CE/BZA did not alter insulin action or ectopic fat, but increased markers of hepatic de novo lipogenesis and TAG production. Study limitations include a small sample size and short treatment period. A larger, fully powered study is needed to validate the potential metabolic benefit of combining CE with BZA.


Following publication of the original article , the autThe sentence currently reads:Locked twins is a rare and hazardous obstetric complication, which occurs in approximately 1:100 twin pregnancies.The sentence should read:Locked twins is a rare and hazardous obstetric complication, which occurs in approximately 1:1000 twin pregnancies.The sentence currenty reads:Locked twins is a rare, hazardous obstetric complication, which occurs in approximately 1:100 twin pregnancies , 3.The sentence should read:Locked twins is a rare, hazardous obstetric complication, which occurs in approximately 1:1000 twin pregnancies , 3.The original article  has been


Nature Communications (2022) 13:1-7; 10.1038/s41467-022-28537-9, published online 18 February 2022.Correction to: In this article the affiliation details for Author L. Ruby Leung were incorrectly given as ‘Qingdao National Laboratory for Marine Science and Technology (QNLM), Qingdao, China’ but should have been ‘Atmospheric Sciences and Global Change Division, Pacific Northwest National Laboratory, Richland, WA, USA’. The original article has been corrected.