Wolbachia are widespread maternally-inherited endosymbiotic bacteria that infect numerous arthropods.This study represents a thorough survey of the Wolbachia infection patterns in the pink stem borer,Sesamia inferens(...Wolbachia are widespread maternally-inherited endosymbiotic bacteria that infect numerous arthropods.This study represents a thorough survey of the Wolbachia infection patterns in the pink stem borer,Sesamia inferens(Walker),an important rice pest in China,based on nucleotide comparisons for the surface protein(wsp)and cell division protein(ftsZ)genes.The effects of Wolbachia on mtDNA variation and evolution of S.inferens were also investigated.Although we identified six genetically diverse strains,we found infections to be infrequent,with only 7.8%of hosts infected,and identified geographical differences in infection rates between southern and northern populations.Nucleotide indexes(haplotype diversity(Hd),nucleotide diversity(π)and number of variable sites(S)of mtDNA in infected populations were not significantly lower or higher than that in the uninfected populations.Furthermore,there was no association between Wolbachia infection status and phylogeny of mtDNA haplotypes.Analysis of molecular variance(AMOVA)showed that significant differentiation mainly existed within groups rather than among the groups.Additionally,using Tajima’s D and Fu’s F values,the mtDNA genes did not deviate significantly from neutral evolution.Taken together these results indicate that currently there were no effects of Wolbachia infection on host mtDNA variation and evolution in S.inferens.展开更多
The effects of rapid cold hardening （RCH） on the cold tolerance of the last instar larvae of Chilo suppressalis （Walker） were evaluated for the first time. The discriminating temperature, induction, detection, dur...The effects of rapid cold hardening （RCH） on the cold tolerance of the last instar larvae of Chilo suppressalis （Walker） were evaluated for the first time. The discriminating temperature, induction, detection, duration and extent of RCH of the larvae in the laboratory were tested, and the supercooling points （SCPs） and the contents of water and lipid of the larvae after RCH treatment were determined, respectively. The results showed that the discriminating temperature of the larvae was about -21℃. Mean survival rates of the larvae which exposed to either 0 or 5℃ for 2 and 4 h before exposure to the discriminating temperature for 2 h were significantly higher than those of the control groups （P 〈 0.05）. Moreover, the highest survival rate appeared in the larvae after 0℃ for 4 h treatment. The protection against low temperature gained by RCH at 0℃ for 4 h was rapidly lost on return to 28℃. Mean survival rates of RCH larvae were significantly higher than those of non-acclimated （NACC） larvae and acclimation （ACC） larvae when they were exposed to the discriminating temperature for 2 or 4 h （P〈 0.05）. Moreover, the rates of NACC, ACC, RCH and ACC ＋ RCH larvae from 2 to 6 h to the discriminating temperature resulted in a significant decline. The values of SCPs and the contents of lipid of the larvae which exposed to either 0 or 5℃ for 2 and 4 h showed no significant difference at 0.05 level compared to those of the control groups. But the contents of water in the larvae were obviously decreased. Therefore, it could be concluded that RCH could enhance cold tolerance and affect partly physiological and biochemical components of the larvae of C. suppressalis, but the underlying mechanisms needs to be further explored.展开更多
Small heat shock protei ns(sHSPs)are a very complex protei n superfamily that increase in sect temperature tolera nee.In order to deeply understand the function and role of sHSPs in Chilo suppressalis(Walker),this stu...Small heat shock protei ns(sHSPs)are a very complex protei n superfamily that increase in sect temperature tolera nee.In order to deeply understand the function and role of sHSPs in Chilo suppressalis(Walker),this study isolated and identified two CsHSP genes lacking introns from C.suppressalis,Cshsp23.9 and Cshsp27.3.The cDNA full-length of Cshsp23.9 and Cshsp27.3 were 909 and 1036 bp encoding 220 and 242 amino acids,respectively.Alignment with homologs and phylogenetic analysis indicated Cshsp23.9 and Cshsp27.3 were two new types of Cshsps in C.suppressalis.Real-time quantitative PCR(qPCR)revealed that Cshsp23.9 had the highest relative expression in hindgut compared with other tissues(head,epidermis,foregut,midgut,fat body,Malpighian tubules,and hemocytes),while Cs/?sp27.3 expressed the highest in fat body con tent.Whe n exposed to thermal stress from-11 to 43°C for 2 h,two genes showed differe nt expression patterns.Cshsp23.9 did not respond to low temperature,but could be up-regulated by high temperature and the highest expression temperature was at 36°C.Cshsp27.3 could only be induced by mild temperature,with the highest expression at 15 and 30°C.In conclusion,Cshsp23.9 and Cshsp27.3 existed in different tissues/organs of C.suppressalis,and played different important roles in C.suppressalis to resist temperature stress and regulate physiological activities.展开更多
Five genes encoding heat shock proteins（HSPs）, Cchsp40, Cchsp60, Cchsp70, Cchsc70 and Cchsp90, were cloned and sequenced from Cotesia chilonis using RT-PCR and RACE. The c DNA sequences of Cchsp40, Cchsp60, Cchsp70,...Five genes encoding heat shock proteins（HSPs）, Cchsp40, Cchsp60, Cchsp70, Cchsc70 and Cchsp90, were cloned and sequenced from Cotesia chilonis using RT-PCR and RACE. The c DNA sequences of Cchsp40, Cchsp60, Cchsp70, Cchsc70 and Cchsp90 were 1 265, 2 551, 2 094, 2 297 and 2 635 bp in length, respectively, with a molecular weight（MW） of 39.1, 60.6, 71.45, 70.19 and 82.92 k Da, respectively. The predicted amino acid sequences of these proteins showed high similarities with published HSPs of other insects in Hymenoptera. Analysis of genomic DNAs indicated that Cchsp40, Cchsp60, Cchsp70 and Cchsp90 lacked introns, but Cchsc70 contained an intron. The results also suggested that CcH SP40 in C. chilonis was the Type II HSP40, Cc HSP60 was a member of the mitochondrial HSP60 family, and Cc HSP90 was a part of cytoplasmic HSP90 A family. Expression patterns varied in the five Cchsps in response to temperature. Expression of Cchsp40 and Cchsp60 was induced significantly by cold but not heat stress. Cchsp70 and Cchsc70 showed similar response to the thermal stress and could be induced by both cold and heat, but their expression levels were consistently lower than that of Cchsp40 and Cchsp60. Cchsp90 could be induced by heat stress and mild cold, but not cold stress. In addition, the results demonstrated Cchsc70 might be constitutive and inducible protein that was expressed during normal cell functioning and also up-regulated in response to stressful stimuli while Cchsp70 was solely inducible protein induced by temperature changes. Overall, results generated from this study could significantly advance the understanding of Cchsps in response to temperature and provide important biological information for C. chilonis insects that reared under different temperatures.展开更多
Many proteins require assistance from molecular chaperones at various stages to attain correctly folded states and functional conformations during protein synthesis. In this study, the gene encoding T-complex polypept...Many proteins require assistance from molecular chaperones at various stages to attain correctly folded states and functional conformations during protein synthesis. In this study, the gene encoding T-complex polypeptide 1（TCP-1）, which belongs to the heat shock protein 60（HSP60） family, was isolated and characterized from the rice stem borer, Chilo suppressalis, by RACE and q PCR, respectively. The full-length c DNA of Tcp-1 was 2 144 bp and encoded a 1 635-bp ORF; the deduced translational product contained 545 amino acids with 5′-and 3′-UTRs and an isoelectric point of 5.29. Cluster analysis confirmed that the deduced amino acid sequence shared high identity（60–99%） with TCP-1 from other insects. To investigate Tcp-1 expression in response to abiotic stress, q PCR was used to analyze expression levels of Tcp-1 m RNA in C. suppressalis larvae exposed to temperatures ranging from –11 to 43°C. With respect to heat shock, Tcp-1 expression was higher than the control after a 2-h exposure to 30 and 36°C and declined at 39 and 43°C. Difference in Tcp-1 expression was observed at temperatures ranging from –11 to 27°C. q PCR analyses revealed that Tcp-1 expression was the highest in hindgut tissue as compared to heads, epidermis, fat body, foregut, midgut, and malpighian tubules. Our results indicated that Tcp-1 expression was differentially expressed in C. suppressalis tissues, and was impacted by temperature stress.展开更多
Heat shock protein 70（HSP70） is one of the most important members in the heat shock protein family, and plays important roles in the thermotolerance of insect. To explore the molecular mechanism of thermotolerance o...Heat shock protein 70（HSP70） is one of the most important members in the heat shock protein family, and plays important roles in the thermotolerance of insect. To explore the molecular mechanism of thermotolerance of Frankliniella occidentalis adults, the difference in the expression of HSP70s in F. occidentalis male or female adults under the thermal stress was studied under the laboratory conditions. Two full length c DNAs of HSP70s gene（Fohsc704 and Fohsc705） were cloned from F. occidentalis by using RT-PCR and RACE. The genomic sequence was demonstrated by genomic validation, and the position and size of the intron were analyzed by sequence analysis of c DNA. Real-time PCR was used to analyze the HSP70 expression patterns. The c DNA of Fohsc704 and Fohsc705 possessed 2 073 and 1 476 bp which encoded 690 and 491 amino acids（aa） with a calculated molecular weight of 75 and 54 k Da, respectively. Four introns in Fohsc704 and six introns in Fohsc705 protein were found. However, the HSP70 protein sequences in our study were ended with EKKN and GIFL, which were different from the reported Fo HSP70s. Various expression patterns of Fohsc704 and Fohsc705 were found in both genders of F. occidentalis under thermal stress. The expression of Fohsc704 and Fohsc705 reached to the highest level at –12 and –8°C in male adults, respectively, and Fohsc705 expressed the highest level at 33°C in female adults. In conclusion, HSP70s of F. occidentalis in our study are novel heat shock proteins. There were difference in expression patterns of the two hsc70s in genders of F. occidentalis, and the two HSP70s play important roles in the thermotolerance of F. occidentalis.展开更多
基金supported by the National Key R&D Program of China(2017YFD0200400)
文摘Wolbachia are widespread maternally-inherited endosymbiotic bacteria that infect numerous arthropods.This study represents a thorough survey of the Wolbachia infection patterns in the pink stem borer,Sesamia inferens(Walker),an important rice pest in China,based on nucleotide comparisons for the surface protein(wsp)and cell division protein(ftsZ)genes.The effects of Wolbachia on mtDNA variation and evolution of S.inferens were also investigated.Although we identified six genetically diverse strains,we found infections to be infrequent,with only 7.8%of hosts infected,and identified geographical differences in infection rates between southern and northern populations.Nucleotide indexes(haplotype diversity(Hd),nucleotide diversity(π)and number of variable sites(S)of mtDNA in infected populations were not significantly lower or higher than that in the uninfected populations.Furthermore,there was no association between Wolbachia infection status and phylogeny of mtDNA haplotypes.Analysis of molecular variance(AMOVA)showed that significant differentiation mainly existed within groups rather than among the groups.Additionally,using Tajima’s D and Fu’s F values,the mtDNA genes did not deviate significantly from neutral evolution.Taken together these results indicate that currently there were no effects of Wolbachia infection on host mtDNA variation and evolution in S.inferens.
文摘The effects of rapid cold hardening （RCH） on the cold tolerance of the last instar larvae of Chilo suppressalis （Walker） were evaluated for the first time. The discriminating temperature, induction, detection, duration and extent of RCH of the larvae in the laboratory were tested, and the supercooling points （SCPs） and the contents of water and lipid of the larvae after RCH treatment were determined, respectively. The results showed that the discriminating temperature of the larvae was about -21℃. Mean survival rates of the larvae which exposed to either 0 or 5℃ for 2 and 4 h before exposure to the discriminating temperature for 2 h were significantly higher than those of the control groups （P 〈 0.05）. Moreover, the highest survival rate appeared in the larvae after 0℃ for 4 h treatment. The protection against low temperature gained by RCH at 0℃ for 4 h was rapidly lost on return to 28℃. Mean survival rates of RCH larvae were significantly higher than those of non-acclimated （NACC） larvae and acclimation （ACC） larvae when they were exposed to the discriminating temperature for 2 or 4 h （P〈 0.05）. Moreover, the rates of NACC, ACC, RCH and ACC ＋ RCH larvae from 2 to 6 h to the discriminating temperature resulted in a significant decline. The values of SCPs and the contents of lipid of the larvae which exposed to either 0 or 5℃ for 2 and 4 h showed no significant difference at 0.05 level compared to those of the control groups. But the contents of water in the larvae were obviously decreased. Therefore, it could be concluded that RCH could enhance cold tolerance and affect partly physiological and biochemical components of the larvae of C. suppressalis, but the underlying mechanisms needs to be further explored.
基金This research was funded by the National Natural Science Foundation of China(31401733 and 31371937)and the National Basic Research and Development Program of China(2013CB127604).We sincerely thank Dr.Tom McAvoy from Virginia Tech,USA for editing on this manu script.
文摘Small heat shock protei ns(sHSPs)are a very complex protei n superfamily that increase in sect temperature tolera nee.In order to deeply understand the function and role of sHSPs in Chilo suppressalis(Walker),this study isolated and identified two CsHSP genes lacking introns from C.suppressalis,Cshsp23.9 and Cshsp27.3.The cDNA full-length of Cshsp23.9 and Cshsp27.3 were 909 and 1036 bp encoding 220 and 242 amino acids,respectively.Alignment with homologs and phylogenetic analysis indicated Cshsp23.9 and Cshsp27.3 were two new types of Cshsps in C.suppressalis.Real-time quantitative PCR(qPCR)revealed that Cshsp23.9 had the highest relative expression in hindgut compared with other tissues(head,epidermis,foregut,midgut,fat body,Malpighian tubules,and hemocytes),while Cs/?sp27.3 expressed the highest in fat body con tent.Whe n exposed to thermal stress from-11 to 43°C for 2 h,two genes showed differe nt expression patterns.Cshsp23.9 did not respond to low temperature,but could be up-regulated by high temperature and the highest expression temperature was at 36°C.Cshsp27.3 could only be induced by mild temperature,with the highest expression at 15 and 30°C.In conclusion,Cshsp23.9 and Cshsp27.3 existed in different tissues/organs of C.suppressalis,and played different important roles in C.suppressalis to resist temperature stress and regulate physiological activities.
基金funded by the National Key R&D Program of China (2017YFD0200400)the National Basic Research Program of China (973 Program, 2013CB127604)
文摘Five genes encoding heat shock proteins（HSPs）, Cchsp40, Cchsp60, Cchsp70, Cchsc70 and Cchsp90, were cloned and sequenced from Cotesia chilonis using RT-PCR and RACE. The c DNA sequences of Cchsp40, Cchsp60, Cchsp70, Cchsc70 and Cchsp90 were 1 265, 2 551, 2 094, 2 297 and 2 635 bp in length, respectively, with a molecular weight（MW） of 39.1, 60.6, 71.45, 70.19 and 82.92 k Da, respectively. The predicted amino acid sequences of these proteins showed high similarities with published HSPs of other insects in Hymenoptera. Analysis of genomic DNAs indicated that Cchsp40, Cchsp60, Cchsp70 and Cchsp90 lacked introns, but Cchsc70 contained an intron. The results also suggested that CcH SP40 in C. chilonis was the Type II HSP40, Cc HSP60 was a member of the mitochondrial HSP60 family, and Cc HSP90 was a part of cytoplasmic HSP90 A family. Expression patterns varied in the five Cchsps in response to temperature. Expression of Cchsp40 and Cchsp60 was induced significantly by cold but not heat stress. Cchsp70 and Cchsc70 showed similar response to the thermal stress and could be induced by both cold and heat, but their expression levels were consistently lower than that of Cchsp40 and Cchsp60. Cchsp90 could be induced by heat stress and mild cold, but not cold stress. In addition, the results demonstrated Cchsc70 might be constitutive and inducible protein that was expressed during normal cell functioning and also up-regulated in response to stressful stimuli while Cchsp70 was solely inducible protein induced by temperature changes. Overall, results generated from this study could significantly advance the understanding of Cchsps in response to temperature and provide important biological information for C. chilonis insects that reared under different temperatures.
基金funded by the National Natural Science Foundation of China (31401733)the Incubation Study Project of Science and Technology of Fuyang Normal University, China (2014KJFH02)
文摘Many proteins require assistance from molecular chaperones at various stages to attain correctly folded states and functional conformations during protein synthesis. In this study, the gene encoding T-complex polypeptide 1（TCP-1）, which belongs to the heat shock protein 60（HSP60） family, was isolated and characterized from the rice stem borer, Chilo suppressalis, by RACE and q PCR, respectively. The full-length c DNA of Tcp-1 was 2 144 bp and encoded a 1 635-bp ORF; the deduced translational product contained 545 amino acids with 5′-and 3′-UTRs and an isoelectric point of 5.29. Cluster analysis confirmed that the deduced amino acid sequence shared high identity（60–99%） with TCP-1 from other insects. To investigate Tcp-1 expression in response to abiotic stress, q PCR was used to analyze expression levels of Tcp-1 m RNA in C. suppressalis larvae exposed to temperatures ranging from –11 to 43°C. With respect to heat shock, Tcp-1 expression was higher than the control after a 2-h exposure to 30 and 36°C and declined at 39 and 43°C. Difference in Tcp-1 expression was observed at temperatures ranging from –11 to 27°C. q PCR analyses revealed that Tcp-1 expression was the highest in hindgut tissue as compared to heads, epidermis, fat body, foregut, midgut, and malpighian tubules. Our results indicated that Tcp-1 expression was differentially expressed in C. suppressalis tissues, and was impacted by temperature stress.
基金funded by the Special Fund for Agro-Scientific Research in the Public Interest of China (201103026, 200803025)the Science and Technology Innovation Project of Student in Yangzhou University,China (X20160637)
文摘Heat shock protein 70（HSP70） is one of the most important members in the heat shock protein family, and plays important roles in the thermotolerance of insect. To explore the molecular mechanism of thermotolerance of Frankliniella occidentalis adults, the difference in the expression of HSP70s in F. occidentalis male or female adults under the thermal stress was studied under the laboratory conditions. Two full length c DNAs of HSP70s gene（Fohsc704 and Fohsc705） were cloned from F. occidentalis by using RT-PCR and RACE. The genomic sequence was demonstrated by genomic validation, and the position and size of the intron were analyzed by sequence analysis of c DNA. Real-time PCR was used to analyze the HSP70 expression patterns. The c DNA of Fohsc704 and Fohsc705 possessed 2 073 and 1 476 bp which encoded 690 and 491 amino acids（aa） with a calculated molecular weight of 75 and 54 k Da, respectively. Four introns in Fohsc704 and six introns in Fohsc705 protein were found. However, the HSP70 protein sequences in our study were ended with EKKN and GIFL, which were different from the reported Fo HSP70s. Various expression patterns of Fohsc704 and Fohsc705 were found in both genders of F. occidentalis under thermal stress. The expression of Fohsc704 and Fohsc705 reached to the highest level at –12 and –8°C in male adults, respectively, and Fohsc705 expressed the highest level at 33°C in female adults. In conclusion, HSP70s of F. occidentalis in our study are novel heat shock proteins. There were difference in expression patterns of the two hsc70s in genders of F. occidentalis, and the two HSP70s play important roles in the thermotolerance of F. occidentalis.
