Reduction In Progeny Research Articles

Regulation ofAnopheles gambiaemale accessory gland genes influences postmating response in female

In Drosophila, the accessory gland proteins (Acps) secreted from the male accessory glands (MAGs) and transferred along with sperm into the female reproductive tract have been implicated in triggering postmating behavioral changes, including refractoriness to subsequent mating and propensity to egg laying. Recently, Acps have been found also in Anopheles, suggesting similar functions. Understanding the mechanisms underlying transcriptional regulation of Acps and their functional role in modulating Anopheles postmating behavior may lead to the identification of novel vector control strategies to reduce mosquito populations. We identified heat-shock factor (HSF) binding sites within the Acp promoters of male Anopheles gambiae and discovered three distinct Hsf isoforms; one being significantly up-regulated in the MAGs after mating. Through genome-wide transcription analysis of Hsf-silenced males, we observed significant down-regulation in 50% of the Acp genes if compared to control males treated with a construct directed against an unrelated bacterial sequence. Treated males retained normal life span and reproductive behavior compared to control males. However, mated wild-type females showed a ∼46% reduction of egg deposition rate and a ∼23% reduction of hatching rate (∼58% combined reduction of progeny). Our results highlight an unsuspected role of HSF in regulating Acp transcription in A. gambiae and provide evidence that Acp down-regulation in males leads a significant reduction of progeny, thus opening new avenues toward the development of novel vector control strategies.

Performance of pre-commercial release formulations of spinosad against five stored-product insect species on four stored commodities

Two liquid and one dry pre-commercial release spinosad formulations were evaluated at the labeled rate of 1 ppm against five stored-grain insect species on wheat, short-grain rice, long-grain rice, and maize. Except on maize, efficacy of spinosad was compared with a currently registered grain protectant, chlorpyrifos-methyl (3 ppm) plus deltamethrin (0.5 ppm). The 7- and 14-day mortalities of the lesser grain borer, Rhyzopertha dominica, were 99.0–100.0% on spinosad and chlorpyrifos-methyl plus deltamethrin-treated wheat, short-grain rice, and long-grain rice. Adult progeny of R. dominica after 42 days on these commodities decreased by 99.7–100.0% relative to progeny production on untreated wheat. Mortality and reduction in adult progeny of the rice weevil, Sitophilus oryzae, on the three commodities, and that of the maize weevil, Sitophilus zeamais, on maize and the red flour beetle, Tribolium castaneum, on wheat were 100.0% only with chlorpyrifos-methyl plus deltamethrin. The liquid spinosad formulations were most effective against the Indianmeal moth, Plodia interpunctella, on maize and wheat. Except for R. dominica, the effectiveness of spinosad on the other species varied with the formulation, exposure time, and commodity. Chlorpyrifos-methyl plus deltamethrin was effective against insect species on the commodities tested.

Molecular characterization of Drosophila cells persistently infected with Flock House virus

Little is known about the molecular determinants causing and sustaining viral persistent infections at the cellular level. We found that Drosophila cells persistently infected (PI) with Flock House virus (FHV) invariably harbor defective viral RNAs, which are replicated by the FHV RNA-dependent RNA polymerase. Some defective RNAs encoded a functional B2 protein, the FHV suppressor of RNA interference, which might contribute to maintenance of virus persistence. Viral small interfering RNAs (vsiRNAs) of both polarities were detected in PI cells and primarily mapped to regions of the viral genome that were preserved in the isolated defective RNAs. This indicated that defective RNAs could represent major sources of vsiRNAs. Immunofluorescence analysis revealed that mitochondria and viral proteins are differentially distributed in PI cells and lytically infected cells, which may partly explain the reduction in infectious viral progeny. Our results provide a basis for further investigations of the molecular mechanisms underlying persistent infections.

Toxicity and growth inhibitory activities of methanol extract and the β-carboline alkaloids of Peganum harmala L. against two coleopteran stored-grain pests

Methanol extract and the β-carboline alkaloids were extracted from the seeds of Peganum harmala (Zygophyllaceae). Their toxicity, growth inhibitory and effects on the progeny production of Tribolium castaneum and Rhyzopertha dominica was studied. To assess any additive effects among the extracted β-carbolines, they were tested as binary mixtures (1:1) or as a crude alkaloid fraction. All extracts exhibited a considerable adulticidal effect with increasing activities in response to increased exposure period. Using the contact toxicity bioassay, the crude β-carboline fraction was the most effective (LC 50’s were 20.1 and 36.7) μg/cm 2, 48 h post-treatment against R. dominica and T. castaneum, respectively. LC 50’s of (harmaline + harmine), (harmaline + harmane), and methanol extract were (31.2, 39.4), (33.7, 47.2), and (39.8, 65.2) μg/cm 2, 24 h post-treatment against R. dominica and T. castaneum, respectively. At 48 h post-treatment, LC 50 of (harmaline + harmine) reached 22.4 μg/cm 2 against R. dominica. When mixed with the insect’s diets, toxicity of all extracts were increased with the crude alkaloidal fraction the most toxic (LC 50’s were 7.8 and 14.7) mg/kg grains, 48 h post exposure against R. dominica and T. castaneum, respectively. When the 2nd instar larvae were fed sub-lethal doses-treated grains, development and F1 progeny of both insects were significantly affected ( P ≤ 0.001). At 3.5 mg/kg grains of the crude alkaloidal extract, percentages of malformed larvae and pupae of T. castaneum were 19.7 and 33.4%, respectively. In this case, a total life span of 81.3 days was recorded for the treated individuals compared to 44.2 for the control. A reduction in the adult progeny of 56.9, 44.0 and 43.6% was obtained with 3.5 mg/kg of the crude alkaloids, (harmaline + harmine) and methanol extract, respectively. Meanwhile, the reduction in adult progeny of R. dominica reached 79.2% with the same concentration of the crude alkaloid extract.

Chemical composition and the insecticidal activity of certain plants applied as powders and essential oils against two stored-products coleopteran beetles

Powders and essential oils were prepared from the aerial parts of Cinnamomum camphora, Ocimum basilicum, Chenopodium ambrosioides, and seeds of Pimpinella anisum. Their adulticidal activities and effects on the F1 progeny of Trogoderma granarium (Everts) and Tribolium castaneum (Herbst) were evaluated. The chemical composition of the plant oils were identified by gas chromatography (GC) and GC/mass spectrometry (MS). All of the tested botanicals showed insecticidal activities against the test insects in a dose-dependent manner with T. granarium was more susceptible to the tested plant products than T. castaneum. At a concentration of 5 g kg−1, many of the plant powders caused 100% mortality of both insects after 14 days of exposure. The powders of C. camphora, O. basilicum, and C. ambrosioides were effective against T. granarium, while that of C. ambrosioides caused 100% adult mortality of T. castaneum under the same assays conditions. A dose of 1.50 ml cm−2 of the oils of C. camphora and O. basilicum completely controlled T. granarium, while 100% mortality of T. castaneum adults was recorded with P. anisum oil. A significant and/or complete reduction (100% inhibition) of the F1 progeny of both insects was obtained as a result of parental exposure to the tested botanicals, especially at the highest doses applied. Botanicals under investigation showed a considerable grain protecting activity against the tested insect species and could be included in integrated pest management (IPM) strategies.

Efficacy of methoprene for control of five species of psocids (psocoptera) on wheat, rice, and maize.

The insect growth regulator methoprene was evaluated for control of Liposcelis bostrychophila Badonnel, Liposcelis decolor (Pearman), Liposcelis entomophila (Enderlein), Liposcelis paeta Pearman (Psocoptera: Liposcelididae), and Lepinotus reticulatus Enderlein (Trogiidae) at application rates of 1, 5, and 10 ppm on maize, wheat, and rice. Methoprene did not completely suppress progeny production during the 40-day test period, but did cause a significant reduction in adult progeny in all psocid species at the application rates of 5 and 10 ppm. At 1 ppm, numbers of adults were reduced for all species on wheat and maize, but only for L. paeta on rice. Nevertheless, the numbers of nymphs present after 40 days generally were not reduced, relative to the controls. Methoprene applied at rates of 1 to 10 ppm to stored grain would not provide adequate control of psocids.

<I>Beauveria bassiana</I> Infection Alters Colony Development and Defensive Secretions of the Beetles <I>Tribolium castaneum</I> and <I>Ulomoides dermestoides</I> (Coleoptera: Tenebrionidae)

We studied the effect of the entomopathogenic fungus Beauveria bassiana strain GHA on a) colony development of the beetles Tribolium castaneum (Herbst) and Ulomoides dermestoides (Fairmaire) (Coleoptera: Tenebrionidae) under laboratory conditions; and 2) the volatile blend released by both beetles, containing defensive pheromones, by using the solid phase microextraction technique. Colony development of both species was strongly altered 3 mo after treatment with B. bassiana, showing a significant reduction in progeny of 37.5% for T. castaneum and 50.0% for U. dermestoides. We also showed that the volatiles released by T. castaneum diminished close to 20% compared with those of healthy beetles, whereas in U. dermestoides secretions dramatically dropped to 5%, 7 d after immersion in 1 x 10(9) conidia per ml. These results suggest that after infection events take place, fungus-induced diminished secretion of the defensive pheromones may be a physiologic clue for behavioral changes in infected beetles.

Insecticidal activity of the root extract of Decalepis hamiltonii against stored-product insect pests and its application in grain protection

Root extracts of Decalepis hamiltonii were tested for insecticidal activity against the stored products pests, Rhyzopertha domonica, Sitophilus oryzae, Stigobium pancieum, Tribolium castaneum and Callosobruchus chinensis, in residual and contact toxicity bioassays. Methanolic extract showed LC50 value of 0.14 mg/cm(2) for all the test species in a filter paper residual bioassay. The extract was effective as a grain protectant for wheat and green gram. Reduction of F1 progeny was observed in treated grain stored for 3-4 months. The extract did not affect the germination of the treated grains. Our results indicate that methanolic extracts of D. hamiltonii has a potential to control stored product pests and could serve as a natural grain protectant.

Repelência e toxicidade de Cyperus iria L., em início de florescimento, ao gorgulho Sitophilus oryzae

Visa-se, com este trabalho, avaliar a repelência e a toxicidade de Cyperus iria L., no estádio de início de florescimento, na Sitophilus oryzae (L.). Realizaram-se dois bioensaios, avaliando-se pós de raízes, partes aéreas e inflorescências de C. iria misturados a grãos de trigo, nas concentrações de 0, 1, 2 e 5%. Em um terceiro bioensaio, avaliou-se a repelência de extratos aquosos produzidos a partir dos pós de C. iria impregnando-se papel sulfite nas concentrações de 0, 0,1, 0,5 e 1,0 mg de planta cm-2. Os pós produzidos com qualquer uma das estruturas de C. iria, repeliram a S. oryzae. A repelência e a toxicidade foram dependentes da concentração e da estrutura da planta empregada. A repelência foi maior com 5% de raízes e menor quando se usou qualquer uma das concentrações da inflorescência e se manteve, em raízes, por períodos superiores a 150 dias. A mortalidade e a redução na progênie foram maiores quando empregados pós de raízes de C. iria, exceto com 2% de raiz, todas as outras estruturas reduziram a progênie de S. oryzae até 60 dias. Houve repelência quando empregados extratos aquosos de raízes a 0,1 e 0,5 mg cm-2, de parte aérea 0,5 mg cm-2 e de inflorescência a 0,5 e 1,0 mg cm-2. O tratamento de grãos com pós de C. iria foi mais eficiente no controle de S. oryzae que os extratos aplicados em papel.

Characterization of a Newly Identified 35-Amino-Acid Component of the Vaccinia Virus Entry/Fusion Complex Conserved in All Chordopoxviruses

The original annotation of the vaccinia virus (VACV) genome was limited to open reading frames (ORFs) of at least 65 amino acids. Here, we characterized a 35-amino-acid ORF (O3L) located between ORFs O2L and I1L. ORFs similar in length to O3L were found at the same genetic locus in all vertebrate poxviruses. Although amino acid identities were low, the presence of a characteristic N-terminal hydrophobic domain strongly suggested that the other poxvirus genes were orthologs. Further studies demonstrated that the O3 protein was expressed at late times after infection and incorporated into the membrane of the mature virion. An O3L deletion mutant was barely viable, producing tiny plaques and a 3-log reduction in infectious progeny. A mutant VACV with a regulated O3L gene had a similar phenotype in the absence of inducer. There was no apparent defect in virus morphogenesis, though O3-deficient virus had low infectivity. The impairment was shown to be at the stage of virus entry, as cores were not detected in the cytoplasm after virus adsorption. Furthermore, O3-deficient virus did not induce fusion of infected cells when triggered by low pH. These characteristics are hallmarks of a group of proteins that form the entry/fusion complex (EFC). Affinity purification experiments demonstrated an association of O3 with EFC proteins. In addition, the assembly or stability of the EFC was impaired when expression of O3 was repressed. Thus, O3 is the newest recognized component of the EFC and the smallest VACV protein shown to have a function.

Effect of early oviposition experience on host acceptance in Trichogramma (Hymenoptera: Trichogrammatidae) and application of F1 sterility and T. principium to suppress the potato tuber moth (Lepidoptera: Gelechiidae)

Laboratory experiments with Trichogramma principium Sugonyaev and Sorokina females offered potato tuber moth Phthorimaea operculella (Zeller) eggs demonstrated that wasps’ rates of oviposition were highest the first day and decreased gradually thereafter. In addition, when T. principium females were sequentially offered eggs from 250 Gy irradiated parents or obtained from non-irradiated moths, the probability of host acceptance was not influenced by treatment of host eggs. In a concurrent laboratory study, a large cage test with combinant releases of T. principium and 250 Gy irradiated moths produced the greatest reduction in potato tuber moth F3-emerged progeny. Reductions obtained with irradiated moths alone, single release of irradiated moths with T. principium, and one or three releases of parasitoids were significantly higher than those in the control. From a pest management perspective, T. principium releases would synergistically complement the effects of F1 sterility against potato tuber moth infestation.

In vitro priming to direct neuronal fate in adult neural progenitor cells

We have previously reported methods that allow for the routine isolation, culture and transplantation of multipotent neural progenitor cells (NPCs) derived from the adult rat sub-ventricular zone (SVZ). In order to expand these techniques, the aim of the current study was to develop an in vitro strategy which can “prime” proliferating adult NPCs towards a neuronal fate during their subsequent differentiation under standard conditions. Our results show that transient exposure to lithium chloride during in vitro proliferation of adult SVZ-derived NPCs has the ability to alter differential fate in vitro and increase the proportion of cells expressing neuronal markers while at the same time causing a reduction in glial progeny. Treatment of adult NPCs with lithium chloride may therefore provide a novel mechanism by which adult NPCs can be primed towards a specific neuronal fate for cell replacement therapy.

Reduction in ovulation or male sex phenotype increases long-term anoxia survival in adaf-16-independent manner inCaenorhabditis elegans

Identifying genotypes and phenotypes that enhance an organism's ability to survive stress is of interest. We used Caenorhabditis elegans mutants, RNA interference (RNAi), and the chemical 5-fluorodeoxyuridine (FUDR) to test the hypothesis that a reduction in progeny would increase oxygen deprivation (anoxia) survival. In the hermaphrodite gonad, germ line processes such as spermatogenesis and oogenesis can be simultaneously as well as independently disrupted by genetic mutations. We analyzed genetic mutants [glp-1(q158), glp-4(bn2ts), plc-1(rx1), ksr-1(ku68), fog-2(q71), fem-3(q20), spe-9(hc52ts), fer-15(hc15ts)] with reduced progeny production due to various reproductive defects. Furthermore, we used RNAi to inhibit the function of gene products in the RTK/Ras/MAPK signaling pathway, which is known to be involved in a variety of developmental processes including gonad function. We determined that reduced progeny production or complete sterility enhanced anoxia survival except in the case of sterile hermaphrodites [spe-9(hc52ts), fer-15(hc15ts)] undergoing oocyte maturation and ovulation as exhibited by the presence of laid unfertilized oocytes. Furthermore, the fog-2(q71) long-term anoxia survival phenotype was suppressed when oocyte maturation and ovulation were induced by mating with males that have functional or nonfunctional sperm. The mutants with a reduced progeny production survive long-term anoxia in a daf-16- and hif-1-independent manner. Finally, we determined that wild-type males were able to survive long-term anoxia in a daf-16-independent manner. Together, these results suggest that the insulin signaling pathway is not the only mechanism to survive oxygen deprivation and that altering gonad function, in particular oocyte maturation and ovulation, leads to a physiological state conducive for oxygen deprivation survival.

Differential Response of a Virginia Common Lambsquarters (Chenopodium Album) Collection to Glyphosate

Control of common lambsquarters with POST applications of glyphosate in glyphosate-resistant crops generally has been effective. In 2002, common lambsquarters plants from Westmoreland County, VA, were identified after not being controlled with a POST glyphosate application in glyphosate-resistant soybean. Plants from this site that survived glyphosate were collected in both 2002 and 2004. The objective of this research was to evaluate the susceptibility of F1 and F2 progeny from these common lambsquarters plants, relative to the susceptibility of common lambsquarters collected in Montgomery County, VA. F1 progeny of the Westmoreland plants from both the 2002 and 2004 collections showed reduced response to glyphosate relative to the Montgomery collection. Vigor reduction of F1 progeny from three 2004 Westmoreland source plants with 0.84 kg ae ha−1 of glyphosate ranged from 66 to 85% at 28 d after treatment (DAT), compared with 89% for the Montgomery collection. Evaluation of four Westmoreland F2 common lambsquarters lines derived from 2002 collections indicated significant differences in glyphosate sensitivity. Fifteen F2 lines were generated from 2004 collections from each of three Westmoreland source plants and from the Montgomery source. For the least sensitive Westmoreland source, vigor reduction ranged from only 24 to 36% across F2 lines in response to 1.68 kg ha−1 of glyphosate at 28 DAT relative to 55 to 100% for the Montgomery source. I50 estimates for fresh weight reduction were 0.91 and 0.32 kg ha−1, for these sources, respectively. Sequential treatments of 0.42, 1.26, and 1.68 kg ha−1 applied at 3-wk intervals to the least susceptible 2004 Westmoreland F2 line resulted in only 37% vigor reduction and no mortality among 360 treated plants.

Grain-protective properties of herbal extracts against the bean weevil Acanthoscelides obtectus Say

Studies were conducted to evaluate the effect of the ethanol extracts from five aromatic medicinal plants against the bean weevil, Acanthoscelides obtectus (Coleoptera: Curculionidae). The extracts were tested for potential to protect stored legume seeds in terms of their repellency, toxicity and reduction of F1 progeny. Significant insecticidal activity was exhibited only by the 100% concentrated extracts from Urtica dioica L. and Taraxacum officinale L., whereas both 100 and 30% extracts from these plants were effective in repellency and reduction of F1 progeny. Although the extract form Achillea millefolium L. (100%) was ineffective in insecticidal activity, it provided a good level of repellency and reduction of F1 progeny. Extracts from Sambucus nigra L. and Juglans regia L. were ineffective in all conducted bioassays.

Varicella-zoster virus influences the activities of components and targets of the ERK signalling pathway

Varicella-zoster virus (VZV) is ultimately dependent upon its host cell for replication. To ensure its reproduction, VZV reorganizes various cellular functions by taking advantage of pre-existing signalling pathways. Recently, it was demonstrated that the activation of stress-related mitogen-activated protein kinase pathways following infection led to increased phosphorylation of cellular transcription factors involved in VZV gene expression. Here, it was shown that members of the extracellular signal-regulated kinase (ERK) pathway are also influenced following VZV infection: c-Raf remained inactive in infected MeWo cells, whereas MEK1/2 and ERK1/2 were phosphorylated transiently, reaching their highest level of phosphorylation at between 10 and 12 h post-infection. Inhibition of this pathway resulted in a severe reduction in viral progeny and in an increased apoptotic response, indicating that the functionality of this cascade is essential for successful high-rate replication. In addition, the activities of Bad, a cytoplasmic target of ERK via ribosomal S6 kinase, and the nuclear-localized target c-Myc were analysed. Bad is a member of the Bcl-2 family and has a key function in regulating apoptosis. Pro-apoptotic functions of Bad are repressed by phosphorylation. A 10-fold increase in Bad phosphorylation at Ser-112 was detected following infection, which was suppressed after inhibition of ERK. The transcription factor c-Myc is involved in the regulation of cell growth and apoptosis. By performing immunoblots and quantitative RT-PCR, suppression of c-Myc expression was demonstrated at both the transcriptional and translational levels in VZV-infected cells. These results suggest that VZV optimizes the conditions for its replication in different ways: upregulation of proviral-acting systems and suppression of potentially antiviral-acting systems.

The tick protective antigen, 4D8, is a conserved protein involved in modulation of tick blood ingestion and reproduction

The gene that encodes the tick protective antigen, 4D8, was cloned from 10 species belonging to 6 genera, and the nucleotide and amino acid sequences were analyzed. 4D8 nucleotide and protein sequences were conserved among these tick species with identity/similarity between 65–98 and 60–98%, respectively. The function of 4D8 was characterized by RNA interference (RNAi) in five tick species. After the ticks were allowed to feed, degeneration of gut, salivary glands and reproductive tissues was observed, and tick survival, weight and oviposition were significantly reduced. 4D8 RNAi effected >90% reduction in oviposition in all tick species tested. Because of the critical role that 4D8 plays during tick feeding and oviposition, which ultimately results in the reduction of tick progeny, we proposed the generic name “subolesin” (Latin, suboles: offspring, progeny) for tick 4D8 proteins and subA for the subolesin-encoding gene.

Effectiveness of spinosad as a grain protectant against resistant beetle and psocid pests of stored grain in Australia

Effectiveness of the bacterium-derived insecticide, spinosad, was determined against eight storage pests of Australia. Laboratory experiments were carried out on relevant resistant strains of four beetle and four psocid species, with the aim of determining the potential of spinosad as a new grain protectant. To explore the possibility that spinosad could have delayed effects, we exposed all insects for 14 d initially and then a further 14 and 28 d for psocids and beetles, respectively. Adult insects of each strain were exposed to untreated wheat (control) and wheat treated with spinosad at 0.1, 0.5 and 1 mg [a.i.]/kg of grain, and adult mortality and reduction of progeny were determined. Among beetles, spinosad was most effective against Rhyzopertha dominica (F.), with 100% adult mortality and progeny reduction after 14 d exposure at 1 mg [a.i.]/kg. Efficacy of spinosad was less with Sitophilus oryzae (L.), and least with Tribolium castaneum (Herbst) and Oryzaephilus surinamensis (L.). Against the psocids, spinosad was most effective against Liposcelis entomophila (Enderlein), with 100% adult mortality after 28 d exposure at 1 mg [a.i.]/kg and 92% progeny reduction after 14 d exposure and 100% subsequently. Spinosad was only moderately effective against Liposcelis bostrychophila Badonnel, L. decolor (Pearman) and L. paeta Pearman. Our findings suggest spinosad to be a potential protectant against R. dominica and L. entomophila in stored grain in Australia. This potential use would be in combination with another protectant capable of controlling other members of the pest complex.

Impact of neem and chinaberry fruit extracts on the pest/parasitoid (Pieris rapae/Hyposoter ebeninus) interactions

AbstractA new technique was performed to simulate exposure of the host Pieris rapae larvae to both botanical treatment and the parasitoid Hyposoter eheninus in different sequences. It was found that: host larvae subjected to both parasitoid and 1 % neem treatment showed significant or insignificant reductions both in pupal formation as well as adult emergence percentages when treatment preceded or followed parasitism, respectively. Both neem treatments revealed significant prolongation in the development of unparasitized and parasitized larvae (on average 4 to 5 days delay), whereas chinaberry caused significant prolongation only among parasitized larvae (on average 2.7 days delay in egg‐larval duration). Fate of parasitism among untreated hosts was found to be faster than among neem‐treated ones. Parasitism percentages among 1 and 0.5 % neem‐treated third instar host larvae held 7 days before parasitism reached 3 and 2 times that achieved among those reared on untreated diet for the same period, ­respectively. It was concluded that prolongation of the preferred target instars of the host, due to neem treatments, increased the chance for parasitism. Nevertheless, treatment with neem at the LC50 level exhibited a great reduction in parasitoid progeny. However, a lower concentration (LC25) could reasonably potentiate parasitism without drastic losses in parasitoid emergence.

Intracellular inhibition of the replication of hepatitis B virus by hammerhead ribozymes.

Chronic hepatitis B virus (HBV) infection is frequently associated with cirrhosis and hepatocellular carcinoma, and so has become a major worldwide health problem. Hammerhead ribozymes have recently gained some attention as potential tools to inhibit viral infection, for which there are no general effective therapies available. A hammerhead ribozyme, RzC, was designed to target the sequence encoding the tail region of the HBV core protein. The activities of the ribozyme were analyzed in vitro and in human hepatoma (HepG2) cells. In vitro, RzC cleaves HBV-RNA at its target site up to 30%, while the disabled ribozyme, dRzC, which has a one-base mutation in the catalytic site, did not cleave the target RNA at all. When the ribozymes were cotransfected into HepG2 cells with the HBV genome-containing plasmid, p3.6II, the inhibition of HBV replication by RzC was greater than that by dRzC, indicating that the active catalytic domain of the hammerhead ribozyme could increase the extent of antisense-mediated inhibition. In addition, there was a gradient of effectiveness in which the greater the amount of released ribozyme, the greater the reduction in HBV progeny DNA. These results suggest the possibility of hammerhead ribozyme-mediated gene therapy for the treatment of HBV infections.