Sea cucumber Holothuria leucospilota is one of the most widespread tropical holothurian species.In this study,eukaryotic organism composition in foregut and hindgut contents of H.leucospilota and surrounding sediments...Sea cucumber Holothuria leucospilota is one of the most widespread tropical holothurian species.In this study,eukaryotic organism composition in foregut and hindgut contents of H.leucospilota and surrounding sediments was assessed by 18S rRNA gene high-throughput sequencing.Eukaryon richness and diversity in the habitat sediments were significantly higher than those in foregut and hindgut contents of the sea cucumbers(P<0.05).The foregut content group,hindgut content group,and marine sediment group sequences were respectively assigned to 18.20±1.32,19.40±1.03,and 21.80±0.37 phyla.In the foregut contents,Nematoda(20.18%±9.59%),Mollusca(16.12%±10.49%),Chlorophyta(10.04%±4.85%),Annelida(8.72%±10.93%),Streptophyta(8.46%±4.65%),and Diatomea(5.99%±2.01%)were the predominant phyla,which showed the eukaryotic food sources of H.leucospilota were primarily belong to the above phyla.The predominant phyla in the hindgut contents were Streptophyta(45.55%±17.32%),Mollusca(4.93%±4.82%),Arthropoda(5.37%±3.08%),Diatomea(3.88%±2.34%),and Chlorophyta(3.79%±1.59%);and Annelida(37.80%±17.00%),Arthropoda(24.49%±12.53%),Platyhelminthes(7.14%±3.02%),Nematoda(4.14%±0.91%),and Diatomea(5.11%±1.35%)had large contents in the sediments.The comparatively high content of Paris genus in phylum Streptophyta in foregut contents indicated that land plants were one of the primary food sources of H.leucospilota,however the significantly higher contents of Streptophyta in hindgut contents than that in foregut contents might suggest a large part of the terrigenous detritus ingested might not be digested by H.leucospilota.UPGMA and PCoA analysis revealed that eukaryotic organism composition differed significantly between foregut contents of H.leucospilota and ambient sediments,indicating selective feeding feature of H.leucospilota.This study provided useful references for artificial feed of tropical sea cucumbers and enhanced understanding of the ecological roles of detritus-feeding macrobenthos.展开更多
Objective:To evaluate microscopy,OptiMAL<sup>?</sup> and multiplex PCR for the identification of Plasmodium falciparum（P.falciparum） and Plasmodium vivax（P.vivax） from the field isolates of Bikaner,Raj...Objective:To evaluate microscopy,OptiMAL<sup>?</sup> and multiplex PCR for the identification of Plasmodium falciparum（P.falciparum） and Plasmodium vivax（P.vivax） from the field isolates of Bikaner,Rajasthan（Northwest India）.Methods:In this study,a multiplex PCR（P.falciparum and P.vivax） was further developed with the incorporation of Plasmodium malariae（P.malariae） specific primer and also a positive control.The performance of microscopy,plasmodium lactate dehydrogenase（pLDH） based malaria rapid diagnostic test OptiMAL<sup>?</sup> and 18S rRNA gene based multiplex PCR for the diagnosis of P.falciparum and P.vivax was compared.Results:The three species multiplex PCR if.falciparum,P.vivax and P.malariae) with an inbuilt positive control was developed and evaluated.In comparison with multiplex PCR,which showed the sensitivity and specificity of 99.36%（95%CI,98.11%-100.00%） and 100.00%（95%CI,100.00%-100.00%）,the sensitivity and specificity of microscopy was 90.44%（95%CI,88.849-95.04%） and 99.22%（95% CI,97.71%-100.00%）,and OptiMAL<sup>?</sup> was 93.58%（95%CI,89.75%-97.42%） and 97.69%（95%CI, 95.10%-100.00%）.The efficiencies were 99.65%,95.10%and 95.45%for multiplex PCK.microscopy and OptiMAL<sup>?</sup>.respectively.Conclusions:Our results raise concerns over the overall sensitivities of microscopy and OptiMAL<sup>?</sup>,when compared to the multiplex PCR and thus stress the need for new molecular interventions in the accurate detection of the malarial parasites.This further highlights the fact that further developments are needed to improve the performance of rapid diagnostic tests at field level.展开更多
Objective:To identify the prevalence of Cryptosporidium from goats in three types of farm management systems in Terengganu,Malaysia and to determine the Cryptosporidium species infecting goats by using 18 S r RNA.Meth...Objective:To identify the prevalence of Cryptosporidium from goats in three types of farm management systems in Terengganu,Malaysia and to determine the Cryptosporidium species infecting goats by using 18 S r RNA.Methods:A total of 478 fecal samples were randomly collected from goats in three farms;199 samples were collected from intensive farm,179 samples from semi-intensive farm and 100 samples from extensive farm.The samples were processed by using formolether concentration technique and stained by using modified Ziehl–Neelsen.Positive samples were performed by using nested PCR analysis by using 18 S r RNA.Results:Out of 478 goats,207(43.3%) were found to be infected with Cryptosporidium.Goats reared under the intensive farm management system reported the highest prevalence of infection(49.7%),followed by intensive farm management system(41%) and the lowest prevalence was reported in the goats reared under semi-intensive management system(37.4%).Conclusions:The identified species found in goat was Cryptosporidium parvum.Future study on the zoonotic transmission of Cryptosporidium parvum in goats needs to be done in order to find the source of transmission of this parasite.展开更多
Ribosomal RNAs are important because they catalyze the synthesis of peptides and proteins.Comparative studies of the secondary structure of 18S rRNA have revealed the basic locations of its many length-conserved and l...Ribosomal RNAs are important because they catalyze the synthesis of peptides and proteins.Comparative studies of the secondary structure of 18S rRNA have revealed the basic locations of its many length-conserved and lengthvariable regions.In recent years,many more sequences of 18S rDNA with unusual lengths have been documented in GenBank.These data make it possible to recognize the diversity of the secondary and tertiary structures of 18S rRNAs and to identify the length-conserved parts of 18S rDNAs.The longest 18S rDNA sequences of almost every known eukaryotic phylum were included in this study.We illustrated the bioinformatics-based structure to show that,the regions that are more length-variable,regions that are less length-variable,the splicing sites for introns,and the sites of A-minor interactions are mostly distributed in different parts of the 18S rRNA.Additionally,this study revealed that some length-variable regions or insertion positions could be quite close to the functional part of the 18S rRNA of Foraminifera organisms.The tertiary structure as well as the secondary structure of 18S rRNA can be more diverse than what was previously supposed.Besides revealing how this interesting gene evolves,it can help to remove ambiguity from the alignment of eukaryotic 18S rDNAs and to improve the performance of 18S rDNA in phylogenetic reconstruction.Six nucleotides shared by Archaea and Eukaryota but rarely by Bacteria are also reported here for the first time,which might further support the supposed origin of eukaryote from archaeans.展开更多
目的基于线粒体细胞色素C氧化酶亚基Ⅰ(COⅠ)与核糖体18S小亚基(18S rRNA)基因序列,分析确定4种肉食螨合适的DNA条形码,从而进一步完善肉食螨DNA条形码数据库。方法2018年5月—2019年7月于安徽省阜阳、芜湖、铜陵等3市小型面粉作坊采集...目的基于线粒体细胞色素C氧化酶亚基Ⅰ(COⅠ)与核糖体18S小亚基(18S rRNA)基因序列,分析确定4种肉食螨合适的DNA条形码,从而进一步完善肉食螨DNA条形码数据库。方法2018年5月—2019年7月于安徽省阜阳、芜湖、铜陵等3市小型面粉作坊采集分离肉食螨样本,经形态学鉴定后,提取单个螨基因组DNA,经PCR扩增、克隆和测序获得COⅠ与18S rRNA基因序列,所获序列进行BLAST比对。结合已报道肉食螨序列,用Clustal X 1.83软件进行多序列比对,基于MEGA X软件进行序列分析并计算遗传距离,采用最大似然法构建系统发育树。结果马六甲肉食螨、转开肉食螨、鳞翅触足螨分子鉴定结果与形态学一致,而Gen Bank中缺少网真扇毛螨相关序列。4种肉食螨COI与18S r RNA基因序列(A+T)分别为69.6%和55.1%,碱基替换数分别为137对和46对。基于COⅠ与18S rRNA基因序列变异位点分析,发现4种肉食螨种间变异位点分别为154~321个和58~99个。4种肉食螨COⅠ与18S r RNA基因序列种内遗传距离均≤0.020,种间遗传距离分别为0.235~0.583和0.078~0.114。基于COⅠ与18S rRNA基因序列的系统发育树显示,4种肉食螨均能各自聚为一支,支持率均达100%,与形态学鉴定结果一致。结论线粒体COⅠ基因序列作为4种肉食螨DNA条形码优于18S rRNA基因序列,更适用于探索肉食螨的属、种低阶元亲缘关系。展开更多
Ribosome RNA(rRNA)accounts for more than 80%of the cell's total RNA,while the physiological functions of rRNA modifications are poorly understood.Mutations of 18S rRNA m6A methyltransferase METTL5 cause intellectu...Ribosome RNA(rRNA)accounts for more than 80%of the cell's total RNA,while the physiological functions of rRNA modifications are poorly understood.Mutations of 18S rRNA m6A methyltransferase METTL5 cause intellectual disability,microcephaly,and facial dysmorphisms in patients,however,little is known about the underlying mechanisms.In this study,we identified METTL5 protein complex and revealed that METTL5 mainly interacts with RNA binding proteins and ribosome proteins.Functionally,we found that Mettl5 knockout in mESCs leads to the abnormal craniofacial and nervous development.Moreover,using Mettl5 knockout mouse model,we further demonstrated that Mettl5 knockout mice exhibit intellectual disability,recapitulating the human phenotype.Mechanistically,we found that Mettl5 maintains brain function and intelligence by regulating the myelination process.Our study uncovered the causal correlation between mis-regulated 18S rRNA m6A modification and neural function defects,supporting the important physiological functions of rRNA modifications in human diseases.展开更多
We observed degradation ofβ‑actin mRNA and 18S rRNA in mouse spleen cells under constant temperature conditions in the different temperature group during postmortem intervals(PMIs)of 0-72 h.Thirty‑nine mice were sacr...We observed degradation ofβ‑actin mRNA and 18S rRNA in mouse spleen cells under constant temperature conditions in the different temperature group during postmortem intervals(PMIs)of 0-72 h.Thirty‑nine mice were sacrificed by cervical dislocation and kept at constant temperatures of 10℃,15℃,20℃,25℃,and 30℃.From 0 to 72 h after death,total RNA in spleen cells was extracted every 6 h.The cycle threshold(Ct)values ofβ‑actin mRNA and 18S rRNA were obtained by real‑time‑quantitative polymerase chain reaction.The results showed that,under the conditions of different and constant temperatures after mouse death at 72 h,the Ct values ofβ‑actin and 18S,Ct ratios ofβ‑actin to 18S,and relative ratios ofβ‑actin to 18S were significantly correlated with PMI.In addition,the relative degradation rates ofβ‑actin and 18S appeared to change from fast to slow with the increase of temperature.By interpolation and fitting analysis of the data,we obtained a ternary quintic equation of the relationship between the change in the relative ratios and PMI,which can be used to infer PMI within a certain temperature range(10℃-30℃).展开更多
Amoeboid protists,an assemblage of organisms belonging to different phylogenetic lineages,have drawn increasing attention due to their crucial ecological roles in various environments and their potential health risks....Amoeboid protists,an assemblage of organisms belonging to different phylogenetic lineages,have drawn increasing attention due to their crucial ecological roles in various environments and their potential health risks.Currently,18S rRNA gene sequencing is widely applied for the detection of amoebae.However,it is not clear which is the best primer pair for 18S rRNA gene amplification in amoebae.This study compared the four most commonly used primer pairs for revealing the diversity,composition,core species,and community assembly processes of amoebae in water and sediments.We found that the choice of primers artificially influences the detection of community composition of amoebae.We also found that short-read fragments may lead to mismatches in taxonomy and were not suitable for phylogenetic analyses.In contrast,full-length primers could detect the highest number of amoeba lineages and annotate 80%of reads belonging to amoebae to known species.However,full-length primers did not detect as many amoeba species as V4 primers.Moreover,we showed that beta diversity and community assembly determination were largely unaffected by primer choice,but different primers could influence our interpretations of the ecological process underlying stochasticity and determinism.This study indicates that full-length read sequencing and V4 region Illumina sequencing are suitable for profiling amoeba diversity in the environment.展开更多
基金Supported by the National Natural Science Foundation of China(Nos.42166005,42076097)the Hainan Provincial Key Research and Development Program(No.ZDYF2021XDNY130)+1 种基金the Natural Science Foundation of Hainan Province(No.321RC1023)the State Key Laboratory of Marine Resource Utilization in South China Sea Open Project(No.MRUKF2021008)。
文摘Sea cucumber Holothuria leucospilota is one of the most widespread tropical holothurian species.In this study,eukaryotic organism composition in foregut and hindgut contents of H.leucospilota and surrounding sediments was assessed by 18S rRNA gene high-throughput sequencing.Eukaryon richness and diversity in the habitat sediments were significantly higher than those in foregut and hindgut contents of the sea cucumbers(P<0.05).The foregut content group,hindgut content group,and marine sediment group sequences were respectively assigned to 18.20±1.32,19.40±1.03,and 21.80±0.37 phyla.In the foregut contents,Nematoda(20.18%±9.59%),Mollusca(16.12%±10.49%),Chlorophyta(10.04%±4.85%),Annelida(8.72%±10.93%),Streptophyta(8.46%±4.65%),and Diatomea(5.99%±2.01%)were the predominant phyla,which showed the eukaryotic food sources of H.leucospilota were primarily belong to the above phyla.The predominant phyla in the hindgut contents were Streptophyta(45.55%±17.32%),Mollusca(4.93%±4.82%),Arthropoda(5.37%±3.08%),Diatomea(3.88%±2.34%),and Chlorophyta(3.79%±1.59%);and Annelida(37.80%±17.00%),Arthropoda(24.49%±12.53%),Platyhelminthes(7.14%±3.02%),Nematoda(4.14%±0.91%),and Diatomea(5.11%±1.35%)had large contents in the sediments.The comparatively high content of Paris genus in phylum Streptophyta in foregut contents indicated that land plants were one of the primary food sources of H.leucospilota,however the significantly higher contents of Streptophyta in hindgut contents than that in foregut contents might suggest a large part of the terrigenous detritus ingested might not be digested by H.leucospilota.UPGMA and PCoA analysis revealed that eukaryotic organism composition differed significantly between foregut contents of H.leucospilota and ambient sediments,indicating selective feeding feature of H.leucospilota.This study provided useful references for artificial feed of tropical sea cucumbers and enhanced understanding of the ecological roles of detritus-feeding macrobenthos.
文摘Objective:To evaluate microscopy,OptiMAL<sup>?</sup> and multiplex PCR for the identification of Plasmodium falciparum（P.falciparum） and Plasmodium vivax（P.vivax） from the field isolates of Bikaner,Rajasthan（Northwest India）.Methods:In this study,a multiplex PCR（P.falciparum and P.vivax） was further developed with the incorporation of Plasmodium malariae（P.malariae） specific primer and also a positive control.The performance of microscopy,plasmodium lactate dehydrogenase（pLDH） based malaria rapid diagnostic test OptiMAL<sup>?</sup> and 18S rRNA gene based multiplex PCR for the diagnosis of P.falciparum and P.vivax was compared.Results:The three species multiplex PCR if.falciparum,P.vivax and P.malariae) with an inbuilt positive control was developed and evaluated.In comparison with multiplex PCR,which showed the sensitivity and specificity of 99.36%（95%CI,98.11%-100.00%） and 100.00%（95%CI,100.00%-100.00%）,the sensitivity and specificity of microscopy was 90.44%（95%CI,88.849-95.04%） and 99.22%（95% CI,97.71%-100.00%）,and OptiMAL<sup>?</sup> was 93.58%（95%CI,89.75%-97.42%） and 97.69%（95%CI, 95.10%-100.00%）.The efficiencies were 99.65%,95.10%and 95.45%for multiplex PCK.microscopy and OptiMAL<sup>?</sup>.respectively.Conclusions:Our results raise concerns over the overall sensitivities of microscopy and OptiMAL<sup>?</sup>,when compared to the multiplex PCR and thus stress the need for new molecular interventions in the accurate detection of the malarial parasites.This further highlights the fact that further developments are needed to improve the performance of rapid diagnostic tests at field level.
基金Supported by IIUM Research Initiative Grant,RIGS grant no.16-301-0465
文摘Objective:To identify the prevalence of Cryptosporidium from goats in three types of farm management systems in Terengganu,Malaysia and to determine the Cryptosporidium species infecting goats by using 18 S r RNA.Methods:A total of 478 fecal samples were randomly collected from goats in three farms;199 samples were collected from intensive farm,179 samples from semi-intensive farm and 100 samples from extensive farm.The samples were processed by using formolether concentration technique and stained by using modified Ziehl–Neelsen.Positive samples were performed by using nested PCR analysis by using 18 S r RNA.Results:Out of 478 goats,207(43.3%) were found to be infected with Cryptosporidium.Goats reared under the intensive farm management system reported the highest prevalence of infection(49.7%),followed by intensive farm management system(41%) and the lowest prevalence was reported in the goats reared under semi-intensive management system(37.4%).Conclusions:The identified species found in goat was Cryptosporidium parvum.Future study on the zoonotic transmission of Cryptosporidium parvum in goats needs to be done in order to find the source of transmission of this parasite.
基金the National Education Project in Basic Science for Special Subjects(Insect Systematics)[grant No.J0630963]the National Science Foundation Project[Grant No.30970350]the National Science Foundation Project for Distinguished Young Scholars[Grant No.30725005].
文摘Ribosomal RNAs are important because they catalyze the synthesis of peptides and proteins.Comparative studies of the secondary structure of 18S rRNA have revealed the basic locations of its many length-conserved and lengthvariable regions.In recent years,many more sequences of 18S rDNA with unusual lengths have been documented in GenBank.These data make it possible to recognize the diversity of the secondary and tertiary structures of 18S rRNAs and to identify the length-conserved parts of 18S rDNAs.The longest 18S rDNA sequences of almost every known eukaryotic phylum were included in this study.We illustrated the bioinformatics-based structure to show that,the regions that are more length-variable,regions that are less length-variable,the splicing sites for introns,and the sites of A-minor interactions are mostly distributed in different parts of the 18S rRNA.Additionally,this study revealed that some length-variable regions or insertion positions could be quite close to the functional part of the 18S rRNA of Foraminifera organisms.The tertiary structure as well as the secondary structure of 18S rRNA can be more diverse than what was previously supposed.Besides revealing how this interesting gene evolves,it can help to remove ambiguity from the alignment of eukaryotic 18S rDNAs and to improve the performance of 18S rDNA in phylogenetic reconstruction.Six nucleotides shared by Archaea and Eukaryota but rarely by Bacteria are also reported here for the first time,which might further support the supposed origin of eukaryote from archaeans.
文摘目的基于线粒体细胞色素C氧化酶亚基Ⅰ(COⅠ)与核糖体18S小亚基(18S rRNA)基因序列,分析确定4种肉食螨合适的DNA条形码,从而进一步完善肉食螨DNA条形码数据库。方法2018年5月—2019年7月于安徽省阜阳、芜湖、铜陵等3市小型面粉作坊采集分离肉食螨样本,经形态学鉴定后,提取单个螨基因组DNA,经PCR扩增、克隆和测序获得COⅠ与18S rRNA基因序列,所获序列进行BLAST比对。结合已报道肉食螨序列,用Clustal X 1.83软件进行多序列比对,基于MEGA X软件进行序列分析并计算遗传距离,采用最大似然法构建系统发育树。结果马六甲肉食螨、转开肉食螨、鳞翅触足螨分子鉴定结果与形态学一致,而Gen Bank中缺少网真扇毛螨相关序列。4种肉食螨COI与18S r RNA基因序列(A+T)分别为69.6%和55.1%,碱基替换数分别为137对和46对。基于COⅠ与18S rRNA基因序列变异位点分析,发现4种肉食螨种间变异位点分别为154~321个和58~99个。4种肉食螨COⅠ与18S r RNA基因序列种内遗传距离均≤0.020,种间遗传距离分别为0.235~0.583和0.078~0.114。基于COⅠ与18S rRNA基因序列的系统发育树显示,4种肉食螨均能各自聚为一支,支持率均达100%,与形态学鉴定结果一致。结论线粒体COⅠ基因序列作为4种肉食螨DNA条形码优于18S rRNA基因序列,更适用于探索肉食螨的属、种低阶元亲缘关系。
基金This work was supported by Excellent Youth Scholars grant from National Natural Science Foundation of China(No.81922052)Distinguished Young Scholars grant from Natural Science Foundation of Guangdong(No.2019B151502011)the Guangzhou People's Livelihood Science and Technology Project(No.201903010006).
文摘Ribosome RNA(rRNA)accounts for more than 80%of the cell's total RNA,while the physiological functions of rRNA modifications are poorly understood.Mutations of 18S rRNA m6A methyltransferase METTL5 cause intellectual disability,microcephaly,and facial dysmorphisms in patients,however,little is known about the underlying mechanisms.In this study,we identified METTL5 protein complex and revealed that METTL5 mainly interacts with RNA binding proteins and ribosome proteins.Functionally,we found that Mettl5 knockout in mESCs leads to the abnormal craniofacial and nervous development.Moreover,using Mettl5 knockout mouse model,we further demonstrated that Mettl5 knockout mice exhibit intellectual disability,recapitulating the human phenotype.Mechanistically,we found that Mettl5 maintains brain function and intelligence by regulating the myelination process.Our study uncovered the causal correlation between mis-regulated 18S rRNA m6A modification and neural function defects,supporting the important physiological functions of rRNA modifications in human diseases.
基金This work was supported by the Open Project of Key Laboratory of Forensic Genetics,Ministry of Public Security(No.2017FGKFKT05)Beijing Natural Science Foundation(No.7192121)Chinese Academy of Engineering(No.2019‑XZ‑31).
文摘We observed degradation ofβ‑actin mRNA and 18S rRNA in mouse spleen cells under constant temperature conditions in the different temperature group during postmortem intervals(PMIs)of 0-72 h.Thirty‑nine mice were sacrificed by cervical dislocation and kept at constant temperatures of 10℃,15℃,20℃,25℃,and 30℃.From 0 to 72 h after death,total RNA in spleen cells was extracted every 6 h.The cycle threshold(Ct)values ofβ‑actin mRNA and 18S rRNA were obtained by real‑time‑quantitative polymerase chain reaction.The results showed that,under the conditions of different and constant temperatures after mouse death at 72 h,the Ct values ofβ‑actin and 18S,Ct ratios ofβ‑actin to 18S,and relative ratios ofβ‑actin to 18S were significantly correlated with PMI.In addition,the relative degradation rates ofβ‑actin and 18S appeared to change from fast to slow with the increase of temperature.By interpolation and fitting analysis of the data,we obtained a ternary quintic equation of the relationship between the change in the relative ratios and PMI,which can be used to infer PMI within a certain temperature range(10℃-30℃).
基金supported by the National Natural Science Foundation of China(31970384,41907021,21806044,92051120,31802350)the Fundamental Research Funds for the Central Universities Sun Yat-sen University(22lgqb22,19lgzd28)+3 种基金the Innovation Group Project of Southern Marine Science and Engineering Guangdong Laboratory(Zhuhai)(311021006)the Southern Marine Science and Engineering Guangdong Laboratory(Zhuhai)(SML2021SP203)the Guangdong Natural Resources Department Contract(GDNRC[2021]62)Guangzhou Basic and Applied Basic Research Foundation(202102020257)。
文摘Amoeboid protists,an assemblage of organisms belonging to different phylogenetic lineages,have drawn increasing attention due to their crucial ecological roles in various environments and their potential health risks.Currently,18S rRNA gene sequencing is widely applied for the detection of amoebae.However,it is not clear which is the best primer pair for 18S rRNA gene amplification in amoebae.This study compared the four most commonly used primer pairs for revealing the diversity,composition,core species,and community assembly processes of amoebae in water and sediments.We found that the choice of primers artificially influences the detection of community composition of amoebae.We also found that short-read fragments may lead to mismatches in taxonomy and were not suitable for phylogenetic analyses.In contrast,full-length primers could detect the highest number of amoeba lineages and annotate 80%of reads belonging to amoebae to known species.However,full-length primers did not detect as many amoeba species as V4 primers.Moreover,we showed that beta diversity and community assembly determination were largely unaffected by primer choice,but different primers could influence our interpretations of the ecological process underlying stochasticity and determinism.This study indicates that full-length read sequencing and V4 region Illumina sequencing are suitable for profiling amoeba diversity in the environment.
