RESUMEN
Two Gram-stain positive, aerobic, short-rod-shaped, catalase-positive, oxidase-negative and non-motile strains, designated YIM 101343T and YIM 101645T, were isolated from faeces of Hylobates hoolock and Lemur catta, respectively. The results of 16S rRNA gene analysis indicated that both represented members of the genus Corynebacterium, and they shared a similarity of 98.0â% with each other. Corynebacterium marinum DSM 44953T showed the highest similarity with both strains YIM 101343T (99.0â%) and YIM 101645T (97.3â%). The results of phylogenetic analysis based on 16S rRNA gene indicated that strain YIM 101343T formed a cluster with C. marinum DSM 44953T and Corynebacterium comes 2019T, strain YIM 101645T formed a cluster with Corynebacterium halotolerans YIM 70093T, and the two clusters were neighbours. The genomic size of strain YIM 101343T was 3068751 bp and that of strain YIM 101645T was 3169714 bp. The dDDH, ANI and AAI values among strains YIM 101343T, YIM 101645T and the closely related species indicated that the two isolates represented two different novel species. Both strains contained meso-diaminopimelic acid and short-chain mycolic acids, and the major menaquinones were MK-9(H2) and MK-8(H2). The major polar lipids of the two strains were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and phosphatidylinositol mannoside. The major fatty acids (>10â%) of both strains were C16â:â0, summed feature 4 and C18â:â1ω9c, but C17â:â1 ω8c was only present as a major component in YIM 101645T . In addition, phenotypic and some chemotaxonomic characteristics of strains YIM 101343T, YIM 101645T and the closely related species were different. Thus, strains YIM 101343T and YIM 101645T should represent two novel species of the genus Corynebacterium, for which the names Corynebacterium hylobatis sp. nov. and Corynebacterium lemuris sp. nov. are proposed, respectively. The type strains are YIM 101343T (=DSM 45970T=CCTCC AB 2013221T) and YIM 101645T (=BCRC 16963T=CCTCC AB 2013281T=KCTC 39868T).
Asunto(s)
Ácidos Grasos , Fosfolípidos , Animales , Ácidos Grasos/química , Fosfolípidos/análisis , Filogenia , ARN Ribosómico 16S/genética , ADN Bacteriano/genética , Técnicas de Tipificación Bacteriana , Composición de Base , Análisis de Secuencia de ADN , Corynebacterium/genética , Primates , Heces/microbiologíaRESUMEN
Digestion and absorption of old cuticles during insect molting are necessary for new cuticle formation, during which complicated enzyme catalysis is essential. To date, a few carboxypeptidases, aminopeptidases and serine proteases (mostly trypsins) connected with cuticle digestion, zymogen activation and histological differentiation during the ecdysis of lepidopteran, dipteran and hymenopteran insects have been identified. However, little is known about these proteins in hemimetabolous insects. In this study, we identified 33 candidate trypsin and trypsin-like homologs, 14 metallocarboxypeptidase and 32 aminopeptidase genes in the brown planthopper Nilaparvata lugens, a hemipteran rice pest. Among the proteins encoded by these genes, 9 trypsin-like proteases, 3 metallocarboxypeptidases and 1 aminopeptidase were selected as potential procuticle hydrolases by bioinformatics analysis and in vivo validation. RNA interference targeting these genes demonstrated that 3 trypsin-like proteases (NlTrypsin-8, NlTrypsin-29 and NlTrypsin-32) genes and 1 metallocarboxypeptidase (NlCpB) gene were found to be essential for ecdysis in N. lugens; specifically, gene silencing led to incomplete cuticle degradation and arrested ecdysis, causing lethal morphological phenotype acquisition. Spatiotemporal expression profiling by quantitative PCR and western blotting revealed their specific expression in the integument and their periodic expression during each stadium, with a peak before ecdysis and eclosion. Transmission electron microscopy demonstrated corresponding ultrastructural defects after RNAi targeting, with NlCpB-silenced specimens having the most undigested old procuticles. Immunohistochemical staining revealed that NlTrypsin-8, NlTrypsin-29 and NlCpB were predominantly located in the exuvial space. This research further adds to our understanding of proteases and its potential role in insect ecdysis.
Asunto(s)
Hemípteros , Muda , Animales , Tripsina/metabolismo , Muda/genética , Hemípteros/metabolismo , Serina Proteasas/metabolismo , Interferencia de ARN , Aminopeptidasas/genética , Aminopeptidasas/metabolismo , Proteínas de Insectos/metabolismoRESUMEN
Heat shock 70kDa proteins (HSP70s) and their cochaperones DNAJs are ubiquitous molecular chaperones, which function as the "HSP70/DNAJ machinery" in a myriad of biological processes. At present, a number of HSP70s have been classified in many species, but studies on DNAJs, especially in insects, are lacking. Here, we first systematically identified and characterized the HSP70 and DNAJ family members in the brown planthopper (BPH), Nilaparvata lugens, a destructive rice pest in Asia. A total of nine HSP70 and 31 DNAJ genes were identified in the BPH genome. Sequence and phylogenetic analyses revealed the high diversity of the NlDNAJ family. Additionally, spatio-temporal expression analysis showed that most NlHSP70 and NlDNAJ genes were highly expressed in the adult stage and gonads. Furthermore, RNA interference (RNAi) revealed that seven NlHSP70s and 10 NlDNAJs play indispensable roles in the nymphal development, oogenesis, and female fertility of N. lugens under physiological growth conditions; in addition, one HSP70 (NlHSP68) was found to be important in the thermal tolerance of eggs. Together, our results in this study shed more light on the biological roles of HSP70/DNAJ in regulating life cycle, coping with environmental stresses, and mediating the interactions within, or between, the two gene families in insects.
Asunto(s)
Proteínas de Choque Térmico/genética , Hemípteros/genética , Animales , Asia , Fertilidad/genética , Chaperonas Moleculares/genética , Ninfa/genética , Oogénesis/genética , Oryza/parasitología , Filogenia , Interferencia de ARN/fisiología , Estrés Fisiológico/genéticaRESUMEN
Hsp90 (heat shock protein 90) chaperone machinery is considered to be a key regulator of proteostasis under both physiological and stress growth conditions in eukaryotic cells. The high conservation of both the sequence and function of Hsp90 allows for the utilization of various species to explore new phenotypes and mechanisms. In this study, three Hsp90 homologs were identified in the brown planthopper (BPH), Nilaparvata lugens: cytosolic NlHsp90, endoplasmic reticulum (ER) NlGRP94 and mitochondrial NlTRAP1. Sequence analysis and phylogenetic construction showed that these proteins belonged to distinct classes consistent with the predicted localization and suggested an evolutionary relationship between NlTRAP1 and bacterial HtpG (high-temperature protein G). Temporospatial expression analyses showed that NlHsp90 was inducible under heat stress throughout the developmental stage, while NlGRP94 was only induced at the egg stage. All three genes had a significantly high transcript level in the ovary. The RNA interference-mediated knockdown of NlHsp90 its essential role in nymph development and oogenesis under physiological conditions. NlGRP94 was also required during the early developmental stage and played a crucial role in oogenesis, fecundity and late embryogenesis. Notably, we first found that NlHsp90 and NlGRP94 were likely involved in the cuticle structure of female BPH. Together, our research revealed multifunctional roles of Hsp90s in the BPH.
Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Proteínas de Choque Térmico/metabolismo , Hemípteros/metabolismo , Calor , Ninfa/metabolismo , Oogénesis , Animales , Femenino , Fertilidad , Proteínas de Choque Térmico/antagonistas & inhibidores , Proteínas de Choque Térmico/genética , Hemípteros/genética , Hemípteros/crecimiento & desarrollo , Ninfa/genética , Ninfa/crecimiento & desarrollo , Filogenia , Interferencia de ARNRESUMEN
The rice water weevil (RWW) Lissorhoptrus oryzophilus Kuschel (Coleoptera: Curculionidae) is an invasive insect pest of rice Oryza sativa L. in China. Little is known about the interactions of this weevil with indigenous herbivores. In the present study, adult feeding and population density of the weevil, injury level of striped stem borer Chilo suppressalis (Walker) (Lepidoptera: Pyralidae) and pink stem borer Sesamia inferens (Walker) (Lepidoptera: Noctuidae) to rice, as well as growth status of their host plants were surveyed in a rice field located in Southeastern Zhejiang, China, in 2004 with the objective to discover interspecific interactions on the rice. At tillering stage, both adult feeding of the weevil and injury of the stem borers tended to occur on larger tillers (bearing 5 leaves) compared with small tillers (bearing 2~4 leaves), but the insects showed no evident competition with each other. At booting stage, the stem borers caused more withering/dead hearts and the weevil reached a higher density on the plants which had more productive tillers and larger root system; the number of weevils per tiller correlated negatively with the percentage of withering/dead hearts of plants in a hill. These observations indicate that interspecific interactions exist between the rice water weevil and the rice stem borers with negative relations occurring at booting or earlier developmental stages of rice.
Asunto(s)
Escarabajos/crecimiento & desarrollo , Oryza/parasitología , Gorgojos/crecimiento & desarrollo , Animales , Oryza/crecimiento & desarrollo , Densidad de PoblaciónRESUMEN
The rice water weevil, Lissorhoptrus oryzophilus Kuschel, has two generations in southern Zhejiang, China. To determine oogenesis in first-generation females (summer females) and its relations to temperature, females were collected from a rice field in early and mid-July and reared on young rice plants at 28, 31 and 34 degrees C in the laboratory. Percentage of females having oocytes, number of oocytes of different stages (stage-I, from early previtellogenesis to middle vitellogenesis; stage-II, late vitellogenesis; and mature-oocyte stage), and length of ovarioles were determined every 10 d of feeding. At each temperature, oogenesis took place in over 40% of females after 20~40 d of feeding, but only 0.0~3.3 stage-I, 0.0~0.8 stage-II and 0.0~1.1 mature oocytes were observed at each observation date. Temperature had significant effect on number of stage-I oocytes but not on number of stage-II and mature oocytes in early July females; temperature had no significant effect on number of oocytes of either stage in mid-July females. Conclusively, in southern Zhejiang, summer L. oryzophilus females have great potential to become reproductive on rice, but their oogenesis activity is very low, with the overall procedures little affected by temperature.
Asunto(s)
Gorgojos/genética , Animales , China , Femenino , Oogénesis , Oryza/parasitología , Estaciones del Año , Temperatura , Gorgojos/patogenicidadRESUMEN
AIM: To investigate the effect of benzyltetrahydropalmatine (BTHP) on the rapidly activating component of delayed rectifier K+ current (Ikr) in single guinea pig ventricular myocytes. METHODS: Whole-cell patch clamp technique was used to record Ikr. RESULTS: Ikr was blocked by 1-100 mumol.L-1 BTHP in concentration-, voltage-, and specifically frequency-dependent fashion, with IC50 of 13.5 mumol.L-1 (95% confidence range: 11.2-15.8 mumol.L-1). 30 mumol.L-1 BTHP reduced Ikr and Ikr.tail by (31 +/- 4)% and (36 +/- 5)% (n = 6, P < 0.01), respectively. The time constant for deactivation (tau') of the tail current was decreased by 30 mumol.L-1 BTHP from (238 +/- 16) ms to (196 +/- 14) ms, while drug had no any effect on the time constant for activation (tau) of Ikr,tail. CONCLUSION: BTHP inhibited Ikr in a frequency-dependent fashion.
Asunto(s)
Antiarrítmicos/farmacología , Alcaloides de Berberina/farmacología , Miocitos Cardíacos/efectos de los fármacos , Canales de Potasio con Entrada de Voltaje , Canales de Potasio/metabolismo , Animales , Separación Celular , Canales de Potasio de Tipo Rectificador Tardío , Femenino , Cobayas , Ventrículos Cardíacos/citología , Ventrículos Cardíacos/metabolismo , Masculino , Miocitos Cardíacos/metabolismo , Técnicas de Placa-Clamp , Canales de Potasio/efectos de los fármacosRESUMEN
AIM: To elucidate the possible mechanisms underlying antiarrhythmia of the non-selective Na+/H+ exchanger inhibitor--amiloride. METHODS: Single ventricular cells were isolated using a double-enzyme method. Effects of amiloride on voltage-dependent potassium and calcium currents in isolated guinea pig ventricular myocyte were recorded by using whole-cell patch clamp techniques. RESULTS: Exposure to amiloride (10 -100 micromol x L(-1)), the L-type and T-type calcium currents were depressed. Amiloride resulted in a concentration-dependent inhibition of peak (Ca,L), But amiloride did not change the shape of their I - V curves. It only decreased the amplitudes of the currents of the two types. When myocytes were incubated with 100 micromol x L(-1) amiloride, I(Kr) was slightly depressed and I(Ks) did not change. Amiloride (1 - 100 micromol x L(-10) depressed I(K1) in a concentration-dependent manner. CONCLUSION: Amiloride depressed potassium and calcium currents, which may give support to its uses in some diseases of the cardiovascular system.
Asunto(s)
Amilorida/farmacología , Canales de Calcio Tipo L/efectos de los fármacos , Canales de Calcio Tipo T/efectos de los fármacos , Miocitos Cardíacos/efectos de los fármacos , Canales de Potasio de Rectificación Interna/efectos de los fármacos , Animales , Antiarrítmicos/farmacología , Separación Celular , Femenino , Cobayas , Ventrículos Cardíacos/citología , Masculino , Técnicas de Placa-Clamp , Canales de Potasio con Entrada de Voltaje/efectos de los fármacos , Intercambiadores de Sodio-Hidrógeno/antagonistas & inhibidoresRESUMEN
OBJECTIVE: To evaluate the effects of Rbl on action potentials and force of contraction in guinea pig ventricular papillary muscles. METHOD: The ventricular papillary muscles of guinea pig were isolated regularly and immersed with Tyrode, s solution. The effects of Rbl (purified saponins of panaxnotoginseng) on the action potentials (AP), the slow action potentials and the force of contraction (FC) of the muscles were studied. The AP and FC were measured synchronously. RESULT: Rbl shortened the duration of AP, including APD2O and APD90, and reduced the FC(n = 5, P < 0.01), but didn't affect the rest potential (RP), the amplitude of action potential (APA), overshot (OS) and maximal upstrok velocity (Vmax). Rbl also decreased the APA of slow action potential, but quinidine had no such effects. CONCLUSION: Rbl may be a channel blocker.
Asunto(s)
Medicamentos Herbarios Chinos/farmacología , Ginsenósidos/farmacología , Contracción Miocárdica/efectos de los fármacos , Panax , Músculos Papilares/fisiología , Potenciales de Acción/efectos de los fármacos , Animales , Bloqueadores de los Canales de Calcio/farmacología , Medicamentos Herbarios Chinos/aislamiento & purificación , Femenino , Ginsenósidos/aislamiento & purificación , Cobayas , Masculino , Panax/químicaRESUMEN
Wolbachia spp. are obligate intracellular bacteria present in reproductive tissues of many arthropod species. Wolbachia infection status and roles in host reproduction were studied in the rice water weevil, Lissorhoptrus oryzophilus Kuschel (Coleoptera, Curculionidae), an introduced species in China. We examined Wolbachia infection status in five populations in China where it reproduces parthenogenetically, and one native population in Southeast Texas, where it reproduces bisexually. All populations were infected by Wolbachia, and all specimens in each population were infected by Wolbachia of a single strain. Phylogenetic analyses based on multilocus sequence typing system indicated that Wolbachia in non-native L. oryzophilus weevils diverges evidently from those in native weevils. After treatments with tetracycline, parthenogenetic weevils reduced the fecundity significantly and eggs were not viable. Our results suggest that Wolbachia are necessary for oocyte production in L oryzophilus.
Asunto(s)
Control Biológico de Vectores , Gorgojos/microbiología , Wolbachia/fisiología , Animales , Antibacterianos/farmacología , Proteínas Bacterianas/genética , China , Femenino , Datos de Secuencia Molecular , Tipificación de Secuencias Multilocus , Filogenia , Reacción en Cadena de la Polimerasa , Reproducción/efectos de los fármacos , Tetraciclina/farmacología , Texas , Gorgojos/efectos de los fármacos , Gorgojos/fisiología , Wolbachia/genéticaRESUMEN
The piggyBac transposon has been long used to integrate foreign DNA into insect genomes. However, undesirable transgene expression can result from random insertions into the genome. In this study, the efficiency of chimeric Gal4-piggyBac transposase in directing integration onto a DNA target plasmid was evaluated in cultured silkworm Bombyx mori Bm-12 and fruit fly Drosophila Schneider 2 (S2) cells. The Gal4-piggyBac transposase has a Gal4 DNA-binding domain (DBD), and the target plasmid has upstream activating sequences (UAS) to which the Gal4 DBD can bind with high affinity. The results indicate that, in the Bm-12 and S2 cells, transpositional activity of Gal4-piggyBac transposase was increased by 4.0 and 7.5 times, respectively, compared to controls, where Gal4-UAS interaction was absent. Moreover, the Gal4-piggyBac transposase had the ability of directing piggyBac element integration to certain sites of the target plasmid, although the target-directing specificity was not as high as expected. The chimeric piggyBac transposase has the potential for use in site-directed transgenesis and gene function research in B. mori.
Asunto(s)
Bombyx/genética , Drosophila/genética , Mutagénesis Insercional/métodos , Plásmidos/genética , Transposasas/genética , Animales , Línea CelularRESUMEN
Tetrandrine (Tet) is one of the best characterized Ca2+ channel blocker of plant origin. It can affect cardiovascular electrophysiologic properties in following field: inhibit the contractility, +/-dt/dpmax, and automaticity of myocardium, prolong the FRP, and exert concentration-dependent negative inotropic and chronotropic effects without altering cardiac excitability. Tet directly blocks both T-type and L-type calcium current in ventricular cells and vascular smooth muscle cells, but it does not shift the I-V relationship curve of ICa. All its effects would be beneficial in the treatment of angina, arrhythmias, and other cardiovascular disorders. Tet also directly inhibits the activity of BKCa channel in endothelial cell line and also inhibits Ca2+-release-activated channels in vessel endothelial cells, which might significantly contribute to the change of endothelial cell activity.
Asunto(s)
Alcaloides/farmacología , Bencilisoquinolinas/farmacología , Bloqueadores de los Canales de Calcio/farmacología , Corazón/efectos de los fármacos , Potenciales de Acción/efectos de los fármacos , Alcaloides/aislamiento & purificación , Animales , Bencilisoquinolinas/aislamiento & purificación , Canales de Calcio/efectos de los fármacos , Canales de Calcio/fisiología , Electrofisiología , Células Endoteliales/efectos de los fármacos , Corazón/fisiología , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Músculo Liso Vascular/citología , Músculo Liso Vascular/efectos de los fármacos , Contracción Miocárdica/efectos de los fármacos , Músculos Papilares/citología , Músculos Papilares/fisiologíaRESUMEN
AIM: To investigate the effects of benzyltetrahydropalmatine (BTHP) on rapidly activating component (I(Kr)), slowly activating component (I(Ks)) of delayed rectifier potassium current, inward rectifier potassium current (I(K1)), and transient outward potassium current (I(to)) in single ventricular myocytes. METHODS: Whole-cell patch clamp technique was used to record ionic currents. RESULTS: (1) BTHP 30 micromol/L reduced I(Kr) and I(Kr,tail) by 31 %+/-4 % and 36 %+/-5 % (n=6, P <0.01), respectively and inhibited I(Ks) and I(Ks,tail) by 40 %+/-6 % and 45 %+/-5 % (n=7, P <0.01), respectively. I(Kr) and I(Ks) were blocked by BTHP 1-100 micromol/L in a concentration-dependent fashion (IC50 value was 13.5 micromol/L and 95 % confidence limit: 11.2-15.8 micromol/L for I(Kr), 9.3 micromol/L and 95 % confidence limit: 7.8-11.8 micromol/L for I(Ks), respectively). (2) BTHP 5 micromol/L inhibited I(to) by 63 %+/-6 % (n=6, P <0.01). BTHP 1-100 micromol/L reduced I(to) in a concentration-dependent manner (IC50 value was 3.6 micromol/L and 95 % confidence limit: 2.9-4.3 micromol/L). (3) BTHP 200 micromol/L did not affect I(K1). CONCLUSION: BTHP inhibited I(Kr), I(Ks), and I(to), but not I(K1). The antiarrhythmic effects of BTHP may be mainly due to its blockade on potassium channels.
Asunto(s)
Antiarrítmicos/farmacología , Alcaloides de Berberina/farmacología , Miocitos Cardíacos/fisiología , Canales de Potasio con Entrada de Voltaje , Canales de Potasio/efectos de los fármacos , Animales , Separación Celular , Canales de Potasio de Tipo Rectificador Tardío , Femenino , Cobayas , Ventrículos Cardíacos/citología , Masculino , Miocitos Cardíacos/citología , Canales de Potasio de Rectificación Interna/efectos de los fármacos , Ratas , Ratas Wistar , Función VentricularRESUMEN
AIM: To study effects of anthopleurin-Q (AP-Q) on myocardial hypertrophy in rats and isolated atria in guinea pigs. METHODS: Two myocardial hypertrophy models in rats were established, one introduced by levothyroxine, the other by stenosis of abdominal aorta. Cardiac myocytes morphometry and functional experiments were employed to investigate effects of AP-Q. RESULTS: Low dose of AP-Q (1 microg/kg/d, ip) reduced morphologic changes of myocardial hypertrophy in both rat models. While high dose of AP-Q (10 microg/kg/d, ip) did not, and caused mild hydropic degeneration in cardiomyocytes. High concentration of AP-Q (30 nmol/L) enhanced the contractility, raised automaticity, and prolonged the functional refractory period (FRP) in isolated left atria of guinea pigs; higher concentration (100 nmol/L) triggered arrhythmia in right atria; low concentration of AP-Q (1 nmol/L)did not affect any myocardial properties above. CONCLUSION: Low dose of AP-Q without inotropic effect can hinder the experimental myocardial hypertrophy in rats; high dose with positive inotropic effect may be responsible for its toxic reaction.
Asunto(s)
Cardiomegalia/fisiopatología , Cardiotónicos/farmacología , Péptidos/farmacología , Animales , Estenosis de la Válvula Aórtica/complicaciones , Cardiomegalia/tratamiento farmacológico , Cardiomegalia/patología , Cobayas , Atrios Cardíacos/efectos de los fármacos , Atrios Cardíacos/fisiopatología , Técnicas In Vitro , Péptidos y Proteínas de Señalización Intercelular , Péptidos/efectos adversos , Péptidos/uso terapéutico , Distribución Aleatoria , Ratas , Ratas Sprague-Dawley , TiroxinaRESUMEN
AIM: To establish a perforated patch recording (PPR) mode with beta-escin and compare L-type calcium current (I(Ca,L)) recorded under PPR and normal whole-cell recording (WCR) condition in isolated guinea-pig ventricular myocytes. METHODS: Single myocytes were dissociated by enzymatic dissociation method. beta-escin was added to the pipette solution to perforate the cell membrane and obtain PPR mode. I(Ca,L) was recorded using PPR and WCR techniques. RESULTS: beta-Escin 20, 25, and 30 micromol/L could permeabilize the cell membrane and obtain PPR mode. With beta-escin 25 micromol/L, the success rate was highest (16/17, 94 %) and the time required for permibilization was 2-15 (8+/-4) min. Run-down of I(Ca,L) was considerably slower in PPR than in WCR condition. The amplitude of I(Ca,L) was decreased by 36 % at 20 min after the formation of WCR, while it was slowly decreased by 8 % at 30 min after the formation of PPR. The current-voltage relation (I-V) curves, activation and inactivation curves of I(Ca,L) were not significantly different between WCR and PPR. The inactivation rate of ICa,L was slower in PPR than in WCR, the faster inactivation time constant (tau(f)) was longer in PPR than in WCR at membrane potentials of -20 mV -- +10 mV (n=6, P<0.05), and the slower time constant (tau(s)) was also longer in PPR than in WCR at membrane potentials of -10 mV to +10 mV (n=6, P<0.05). There was no significant difference between the activation rate in WCR and PPR. CONCLUSION: Using beta-escin 25 micromol/L can easily obtain stable PPR in isolated guinea-pig ventricular myocytes, and this method is useful in dealing with channels, which show run-down under normal WCR such as L-type Ca channel.