Activation and Autoinhibition Mechanisms of NLR Immune Receptor Pi36 in Rice.

Nucleotide-binding and leucine-rich repeat receptors (NLRs) are the most important and largest class of immune receptors in plants. The Pi36 gene encodes a canonical CC-NBS-LRR protein that confers resistance to rice blast fungal infections. Here, we show that the CC domain of Pi36 plays a role in cell death induction. Furthermore, self-association is required for the CC domain-mediated cell death, and the self-association ability is correlated with the cell death level. In addition, the NB-ARC domain may suppress the activity of the CC domain through intramolecular interaction. The mutations D440G next to the RNBS-D motif and D503V in the MHD motif autoactivated Pi36, but the mutation K212 in the P-loop motif inhibited this autoactivation, indicating that nucleotide binding of the NB-ARC domain is essential for Pi36 activation. We also found that the LRR domain is required for D503V- and D440G-mediated Pi36 autoactivation. Interestingly, several mutations in the CC domain compromised the CC domain-mediated cell death without affecting the D440G- or D503V-mediated Pi36 autoactivation. The autoactivate Pi36 variants exhibited stronger self-associations than the inactive variants. Taken together, we speculated that the CC domain of Pi36 executes cell death activities, whereas the NB-ARC domain suppressed CC-mediated cell death via intermolecular interaction. The NB-ARC domain releases its suppression of the CC domain and strengthens the self-association of Pi36 to support the CC domain, possibly through nucleotide exchange.

Blockade of CD300A enhances the ability of human NK cells to lyse hematologic malignancies.

OBJECTIVE: The human cluster of differentiation (CD)300A, a type-I transmembrane protein with immunoreceptor tyrosine-based inhibitory motifs, was investigated as a potential immune checkpoint for human natural killer (NK) cells targeting hematologic malignancies (HMs). METHODS: We implemented a stimulation system involving the CD300A ligand, phosphatidylserine (PS), exposed to the outer surface of malignant cells. Additionally, we utilized CD300A overexpression, a CD300A blocking system, and a xenotransplantation model to evaluate the impact of CD300A on NK cell efficacy against HMs in in vitro and in vivo settings. Furthermore, we explored the association between CD300A and HM progression in patients. RESULTS: Our findings indicated that PS hampers the function of NK cells. Increased CD300A expression inhibited HM lysis by NK cells. CD300A overexpression shortened the survival of HM-xenografted mice by impairing transplanted NK cells. Blocking PS-CD300A signals with antibodies significantly amplified the expression of lysis function-related proteins and effector cytokines in NK cells, thereby augmenting the ability to lyse HMs. Clinically, heightened CD300A expression correlated with shorter survival and an "exhausted" phenotype of intratumoral NK cells in patients with HMs or solid tumors. CONCLUSIONS: These results propose CD300A as a potential target for invigorating NK cell-based treatments against HMs.

Ambient PM2.5 components and prevalence of chronic kidney disease: a nationwide cross-sectional survey in China.

OBJECTIVES: Chronic kidney disease (CKD) is a global public health concern, and accumulating evidence has indicated that air pollution increases the odds of CKD. However, a limited number of studies have examined the long-term effects of ambient fine particulate matter (PM2.5) components on the risk of CKD among general population; thus, major knowledge gaps remain. METHODS: Using data from a nationwide representative cross-sectional survey in China and a validated PM2.5 composition dataset, we established generalized linear models to quantify the association between five major components of PM2.5 and CKD prevalence. RESULTS: There were significant associations between long-term exposure to three PM2.5 components [including black carbon (BC), sulfate (SO42-), organic matter (OM)] and increased odds of CKD prevalence. Along with an interquartile range (IQR) increment in BC (3.3 µg/m3), SO42- (9.7 µg/m3), and OM (16.2 µg/m3) at a 4-year moving average, the odds ratios (ORs) for CKD prevalence were 1.28 (95% CI 1.07, 1.54), 1.23 (95% CI 1.03, 1.45), and 1.23 (95% CI 1.02, 1.47), respectively. We did not detect any significant association of the other two PM2.5 components [nitrate (NO3-) or ammonium (NH4+)] with CKD prevalence. Stratified analyses revealed no differences (P ≥ 0.05) in the effect estimates of subgroups based on administrative region, sex, age, and other demographic characteristics. For instance, along with an IQR increment in BC at a 4-year moving average, the ORs of CKD prevalence among males and females were 1.30 (95% CI 0.98, 1.73) and 1.29 (95% CI 1.01, 1.65), respectively. The odds of CKD were generally higher with increasing PM2.5 composition concentration. CONCLUSIONS: Our study demonstrated that long-term exposure to specific PM2.5 components including BC, SO42-, and OM increased CKD risk in the general population. This study could provide new insights into source-directed PM2.5 control and CKD prevention.

[Early effectiveness of computer navigation system-assisted transiliac-transsacral screws placement for posterior pelvic ring injuries].

Objective: To investigate the early effectiveness of transiliac-transsacral screws internal fixation assisted by augmented reality navigation system HoloSight (hereinafter referred to as "computer navigation system") in the treatment of posterior pelvic ring injuries. Methods: A retrospective analysis was made in the 41 patients with posterior pelvic ring injuries who had been treated surgically with transiliac-transsacral screws between June 2022 and June 2023. The patients were divided into navigation group (18 cases, using computer navigation system to assist screw implantation) and freehand group (23 cases, using C-arm X-ray fluoroscopy to guide screw implantation) according to the different methods of transiliac-transsacral screws placement. There was no significant difference in gender, age, body mass index, causes of injuries, Tile classification of pelvic fracture, days from injury to operation, usage of unlocking closed reduction technique between the two groups ( P>0.05). The time of screw implantation, the fluoroscopy times, the guide wire adjustment times of each screw, and the incidence of complications were recorded and compared between the two groups. The position of the transiliac-transsacral screw was scanned by CT within 2 days after operation, and the position of the screw was classified according to Gras standard. Results: The operation was successfully completed in both groups. The time of screw implantation, the fluoroscopy times, and the guide wire adjustment times of each screw in the navigation group were significantly less than those in the freehand group ( P<0.05). There were 2 cases of incision infection in the freehand group, and the incision healed by first intention after active dressing change; there was no screw-related complication in the navigation group during operation and early period after operation; the difference in incidence of complications between the two groups (8.7% vs. 0) was not significant ( P=0.495). According to the Gras standard, the screw position of the navigation group was significantly better than that of the freehand group ( P<0.05). Conclusion: Compared with the traditional freehand method, the computer navigation system assisted transiliac-transsacral screws internal fixation in the treatment of posterior pelvic ring injuries has advantages of improving the accuracy of screw implantation and reducing radiation damage and the time of screw implantation.

Role of rhizobia in promoting non-enzymatic antioxidants to mitigate nitrogen-deficiency and nickel stresses in Pongamia pinnata.

The contribution of rhizobia in the mitigation of non-enzymatic antioxidants against nitrogen deficiency and heavy metal toxicity for legume plant is not clear. Therefore, it is hypothesized that the inoculation of rhizobia could mitigate nitrogen deficiency and nickel (Ni) stresses in P. pinnata tissues by enhancing the formation of certain non-enzymatic antioxidants. The effect of symbiotic nitrogen-fixing rhizobia on the mitigation of nitrogen-deficiency and Ni stresses in P. pinnata was evaluated by inoculating two different rhizobia, i.e., Rhizobium pisi PZHK2 and Ochrobacterium pseudogrignonense PZHK4, around the rhizosphere of P. pinnata grown in soil containing 40 mg kg-1 Ni2+ and without nitrogen addition. The inoculation with both rhizobial strains promoted the growth of P. pinnata under nickel stress or nitrogen-deficiency condition, increased nitrogen content in all plant tissues and nickel content in shoots and leaves, but reduced nickel accumulation in roots. The four non-enzymatic antioxidants including glutathione (GSH), proanthocyanidin (OPC), ascorbic acid (ASA) and flavonoids (FLA) distributed in roots, shoots and leaves were followed in descending order: GSH > OPC > ASA > FLA. The four non-enzymatic antioxidants showed different levels of change under the nitrogen-deficiency and nickel stresses and in the non-stress control. The inoculation of PZHK2 and PZHK4 significantly (p < 0.05) increased the four non-enzymatic antioxidants in P. pinnata tissues, especially in roots. Some non-enzymatic antioxidants showed correlations with nickel or nitrogen in P. pinnata tissues, and the four non-enzymatic antioxidants also had correlations among each other. Therefore, this research revealed an excellent role of rhizobia in promoting non-enzymatic antioxidants to mitigate nitrogen-deficiency or nickel stress for P. pinnata.

[Effectiveness analysis of disposable skin stretch closure in treatment of difficult to close skin and soft tissue defects].

Objective: To observe the effectiveness of disposable skin stretch closure in the treatment of wounds with skin and soft tissue defects that were difficult to close. Methods: The clinical data of 13 patients with skin and soft tissue defects that were difficult to close treated with disposable skin stretch closure and met the selection criteria between July 2021 and February 2022 were retrospectively reviewed. There were 9 males and 4 females, the age ranged from 15 to 71 years with a mean of 39.8 years. The causes of injury included falling injury in 5 patients, traffic accident injury in 5 patients, and falling from height injury in 3 patients. The causes of skin soft tissue defects included open fractures in 4 patients, wound infection in 4 patients, osteomyelitis in 3 patients, degloving injury in 1 patient, and necrosis of skin graft in 1 patient. The injury was located at calf in 8 patients, calcaneus in 3 patients, pelvis in 1 patient, and plantar in 1 patient. The skin and soft tissue defects ranged from 5.0 cm×2.0 cm to 10.5 cm×6.5 cm. Wound conditions (wound closure and wound healing) and the presence or absence of complications were recorded. Results: All 13 patients were followed up 32-225 days with a median of 164 days. The wound closure time ranged from 5 to 14 days, with a mean of 8.8 days. The wound closure speed ranged from 0.7 to 13.7 cm 2/day, with a mean of 3.6 cm 2/day. All wounds healed at grade A, and no complication such as skin edge injury, wound necrosis, infection, dehiscence, and edema occurred. No patient complained of pain or discomfort, and no obvious scarring was found during follow-up. The wound healing time ranged from 17 to 28 days, with a mean of 21.7 days. One of them was transferred to other department due to lung cancer condition changes after using disposable skin stretch closure, and the wound had directly healed without suturing at 17 days after operation. Conclusion: The effectiveness of disposable skin stretch closure in the treatment of wounds with skin and soft tissue defects that were difficult to close was exact, with short wound closure time, few complications, and easy operation.

Residential greenness and prevalence of chronic kidney disease: Findings from the China National Survey of Chronic Kidney Disease.

Green space is associated with many health benefits, but evidence concerning the effects on chronic kidney disease (CKD) has not been investigated. Using the nationwide cross-sectional study of 47,204 adults from the China National Survey of Chronic Kidney Disease dataset and residential greenness assessed by the normalized difference vegetation index (NDVI), this study evaluated the association between residential greenness and CKD prevalence. An interquartile range increase in NDVI1000m (0.26) was associated with decreased odds of CKD for all participants with an odds ratio (OR) of 0.79 (95% confidence interval [CI]: 0.73-0.86). Subgroup analyses demonstrated more apparent inverse associations in younger adults <65 years, male participants, people in higher socio-economic status, as well as people with smoking and alcohol drinking habit. In addition, more apparent inverse associations were found in regions with higher fine particulate matter (PM2.5) concentration levels, with OR of 0.56 (95% CI: 0.49, 0.65) for higher pollution regions, and OR of 0.95 (95% CI: 0.83, 1.09) for lower pollution regions (P for interaction <0.001). The exposure-response curves captured more apparent declines in OR of CKD when in lower NDVI1000m exposure ranges (<0.6), even controlling for the PM2.5 concentration. Our results indicated that residential greenness might be beneficial for the prevention and control of CKD at the population level, suggesting the positive significance of strengthening green space construction, particularly in regions with low greenness.

Synergistic anti-oxidative effects of Pongamia pinnata against nickel mediated by Rhizobium pisi and Ochrobacterium pseudogrignonense.

Nickel is widely spread by different anthropogenic activities and shows toxicity for plant growth and development. Whether rhizobia symbiotically fix nitrogen can eliminate or reduce nickel toxic effect on plant or not is still unknown. This study was aimed to investigate the effect of different rhizobia genus inoculation on growth, nitrogen fixing ability, metal accumulation and enzymatic antioxidative balance of Pongamia pinnnaa. Inoculation with Rhizobium pisi and Ochrobacterium pseudogrignonense increased the all the growth parameters both in 0 and 40 mg/kg nickel as comparison with control. Only shoot length increased in presence of nitrogen as compared with no supply of nitrogen. Nitrogen content also increased both in rhizobia inoculation as compared to no nitrogen supply and non-inoculation control, respectively. Nickel uptake was higher in shoots and leaves but lower in roots in case of inoculation as compared to non-inoculation control. Rhizobia inoculation improved the plant antioxidant capacity by increasing the activity of enzymatic scavengers catalase (CAT), superoxide dismutase (SOD), peroxidase (POD) and ascorbate (GR). However, 40 mg/kg of nickel adding showed mostly effect on the activity CAT, SOD, POD in leaves. All the enzymatic activity showed a significant increase in absence of nitrogen supply as compared nitrogen supply. Our results suggested that rhizobia inoculation effectively mediated nickel stress for legume plants by increasing nitrogen supplement and inducing antioxidant capacity.

Effects of Co-application of Cadmium-Immobilizing Bacteria and Organic Fertilizers on Houttuynia cordata and Microbial Communities in a Cadmium-Contaminated Field.

Cadmium pollution is a serious threat to the soil environment. The application of bio-based fertilizers in combination with beneficial microbial agents is a sustainable approach to solving Cd pollution in farm soil. The present study investigated the effects of co-application of a Cd-immobilizing bacterial agent and two fermented organic fertilizers (fermentative edible fungi residue; fermentative cow dung) on Houttuynia cordata and its microbial communities in a Cd-polluted field. It showed that both the application of the Cd-immobilizing bacterial agent alone and the combined application of bio-based soil amendments and the bacterial agent effectively reduced >20% of the uptake of Cd by the plant. Soil nitrogen level was significantly raised after the combined fertilization. The multivariate diversity analysis and co-occurrence network algorithm showed that a significant shift of microbial communities took place, in which the microbial populations tended to be homogeneous with reduced microbial richness and increased diversity after the co-application. The treatment of fermentative cow dung with the addition of the bacterial agent showed a significant increase in the microbial community dissimilarity (R = 0.996, p = 0.001) compared to that treated with cow dung alone. The co-application of the bacterial agent with both organic fertilizers significantly increased the abundance of Actinobacteria and Bacteroidetes. The FAPROTAX soil functional analysis revealed that the introduction of the microbial agent could potentially suppress human pathogenic microorganisms in the field fertilized with edible fungi residue. It also showed that the microbial agent can reduce the nitrite oxidation function in the soil when applied alone or with the organic fertilizers. Our study thus highlights the beneficial effects of the Cd-immobilizing bacterial inoculant on H. cordata and provides a better understanding of the microbial changes induced by the combined fertilization using the microbial agent and organic soil amendments in a Cd-contaminated field.

Comparative secretome analysis of different smut fungi and identification of plant cell death-inducing secreted proteins from Tilletia horrida.

BACKGROUND: Tilletia horrida is a basidiomycete fungus that causes rice kernel smut, one of the most important rice diseases in hybrid rice growing areas worldwide. However, little is known about its mechanisms of pathogenicity. We previously reported the genome of T. horrida, and 597 genes that encoded secreted proteins were annotated. Among these were some important effector genes related to pathogenicity. RESULTS: A secretome analysis suggested that five Tilletia fungi shared more gene families than were found in other smuts, and there was high conservation between them. Furthermore, we screened 597 secreted proteins from the T. horrida genome, some of which induced expression in host-pathogen interaction processes. Through transient expression, we demonstrated that two putative effectors could induce necrosis phenotypes in Nicotiana benthamiana. These two encoded genes were up-regulated during early infection, and the encoded proteins were confirmed to be secreted using a yeast secretion system. For the putative effector gene smut_5844, a signal peptide was required to induce non-host cell death, whereas ribonuclease catalytic active sites were required for smut_2965. Moreover, both putative effectors could induce an immune response in N. benthamiana leaves. Interestingly, one of the identified potential host interactors of smut_5844 was laccase-10 protein (OsLAC10), which has been predicted to be involved in plant lignification and iron metabolism. CONCLUSIONS: Overall, this study identified two secreted proteins in T. horrida that induce cell death or are involved in defense machinery in non-host plants. This research provides a useful foundation for understanding the interaction between rice and T. horrida.

The pathogenic mechanisms of Tilletia horrida as revealed by comparative and functional genomics.

Tilletia horrida is a soil-borne, mononucleate basidiomycete fungus with a biotrophic lifestyle that causes rice kernel smut, a disease that is distributed throughout hybrid rice growing areas worldwide. Here we report on the high-quality genome sequence of T. horrida; it is composed of 23.2 Mb that encode 7,729 predicted genes and 6,973 genes supported by RNA-seq. The genome contains few repetitive elements that account for 8.45% of the total. Evolutionarily, T. horrida lies close to the Ustilago fungi, suggesting grass species as potential hosts, but co-linearity was not observed between T. horrida and the barley smut Ustilago hordei. Genes and functions relevant to pathogenicity were presumed. T. horrida possesses a smaller set of carbohydrate-active enzymes and secondary metabolites, which probably reflect the specific characteristics of its infection and biotrophic lifestyle. Genes that encode secreted proteins and enzymes of secondary metabolism, and genes that are represented in the pathogen-host interaction gene database genes, are highly expressed during early infection; this is consistent with their potential roles in pathogenicity. Furthermore, among the 131 candidate pathogen effectors identified according to their expression patterns and functionality, we validated two that trigger leaf cell death in Nicotiana benthamiana. In summary, we have revealed new molecular mechanisms involved in the evolution, biotrophy, and pathogenesis of T. horrida.

Molecular hydrogen increases resilience to stress in mice.

The inability to successfully adapt to stress produces pathological changes that can lead to depression. Molecular hydrogen has anti-oxidative and anti-inflammatory activities and neuroprotective effects. However, the potential role of molecular hydrogen in stress-related disorders is still poorly understood. The present study aims to investigate the effects of hydrogen gas on resilience to stress in mice. The results showed that repeated inhalation of hydrogen-oxygen mixed gas [67%:33% (V/V)] significantly decreased both the acute and chronic stress-induced depressive- and anxiety-like behaviors of mice, assessed by tail suspension test (TST), forced swimming test (FST), novelty suppressed feeding (NSF) test, and open field test (OFT). ELISA analyses showed that inhalation of hydrogen-oxygen mixed gas blocked CMS-induced increase in the serum levels of corticosterone, adrenocorticotropic hormone, interleukin-6, and tumor necrosis factor-α in mice exposed to chronic mild stress. Finally, inhalation of hydrogen gas in adolescence significantly increased the resilience to acute stress in early adulthood, which illustrates the long-lasting effects of hydrogen on stress resilience in mice. This was likely mediated by inhibiting the hypothalamic-pituitary-adrenal axis and inflammatory responses to stress. These results warrant further exploration for developing molecular hydrogen as a novel strategy to prevent the occurrence of stress-related disorders.

Mitochondria-targeted peptide SS-31 attenuates renal injury via an antioxidant effect in diabetic nephropathy.

Oxidative stress is implicated in the pathogenesis of diabetic kidney injury. SS-31 is a mitochondria-targeted tetrapeptide that can scavenge reactive oxygen species (ROS). Here, we investigated the effect and molecular mechanism of mitochondria-targeted antioxidant peptide SS-31 on injuries in diabetic kidneys and mouse mesangial cells (MMCs) exposed to high-glucose (HG) ambience. CD-1 mice underwent uninephrectomy and streptozotocin treatment prior to receiving daily intraperitoneal injection of SS-31 for 8 wk. The diabetic mice treated with SS-31 had alleviated proteinuria, urinary 8-hydroxy-2-deoxyguanosine level, glomerular hypertrophy, and accumulation of renal fibronectin and collagen IV. SS-31 attenuated renal cell apoptosis and expression of Bax and reversed the expression of Bcl-2 in diabetic mice kidneys. Furthermore, SS-31 inhibited expression of transforming-growth factor (TGF)-ß1, Nox4, and thioredoxin-interacting protein (TXNIP), as well as activation of p38 MAPK and CREB and NADPH oxidase activity in diabetic kidneys. In vitro experiments using MMCs revealed that SS-31 inhibited HG-mediated ROS generation, apoptosis, expression of cleaved caspase-3, Bax/Bcl-2 ratio, and cytochrome c (cyt c) release from mitochondria. SS-31 normalized mitochondrial potential (ΔΨm) and ATP alterations, and inhibited the expression of TGF-ß1, Nox4, and TXNIP, as well as activation of p38 MAPK and CREB and NADPH oxidase activity in MMCs under HG conditions. SS-31 treatment also could reverse the reduction of thioredoxin (TRX) biologic activity and upregulate expression of thioredoxin 2 (TRX2) in MMCs under HG conditions. In conclusion, this study demonstrates a protective effect of SS-31 against HG-induced renal injury via an antioxidant mechanism in diabetic nephropathy.

Vitamin D receptor rs2228570 polymorphism and susceptibly to ovarian cancer: a meta-analysis.

The role of vitamin D receptor (VDR) rs2228570 polymorphism on the risk of ovarian cancer has been studied in many studies, but the relationship between VDR rs2228570 polymorphism and ovarian cancer is still unclear. We thus performed a meta-analysis of published studies to provide a comprehensive assessment of the association. Fourteen individual studies with a total of 10,964 subjects were finally included into the meta-analysis. We assessed the association by calculating the pooled odds ratio (OR) with 95 % confidence intervals (95 % CI). There was no heterogeneity among those included studies. Meta-analysis of 14 studies showed that the VDR rs2228570 polymorphism was associated with risk of ovarian cancer under three main comparison models (T versus C: OR = 1.09, 95 % CI 1.03 to 1.15, P = 0.004; TT versus CC: OR = 1.17, 95 % CI 1.04 to 1.32, P = 0.01; and TT/CT versus CC: OR = 1.12, 95 % CI 1.03 to 1.21, P = 0.007). Subgroup analysis in Caucasians further identified the obvious association. There was no evidence of publications bias. These data from the meta-analysis suggest that VDR rs2228570 polymorphism is associated with risk of ovarian cancer in Caucasians. More studies are warranted to assess the association between the VDR rs2228570 polymorphism and ovarian cancer in Asians and Africans.

Characterization of cry2-type genes of Bacillus thuringiensis strains from soil-isolated of Sichuan basin, China

Sichuan basin, situated in the west of China, is the fourth biggest basin in China. In order to describe a systematic study of the cry2-type genes resources from Bacillus thuringiensis strains of Sichuan basin, a total of 791 Bacillus thuringiensis strains have been screened from 2650 soil samples in different ecological regions. The method of PCR-restriction fragment length polymorphism (PCR-RFLP) was used to identify the type of cry2 genes. The results showed that 322 Bacillus thuringiensis strains harbored cry2-type genes and four different RFLP patterns were found. The combination of cry2Aa/cry2Ab genes was the most frequent (90.4 percent), followed by cry2Aa (6.8 percent) and cry2Ab alone (2.5 percent), and only one novel type of cry2 gene was cloned from one isolate (JF19-2). The full-length of this novel gene was obtained by the method of thermal asymmetric interlaced PCR (Tail-PCR), which was designated as cry2Ag1 (GenBank No. ACH91610) by the Bt Pesticide Crystal Protein Nomenclature Committee. In addition, the result of scanning electron microscopic (SEM) observation showed that these strains had erose, spherical, bipyramidal, and square crystal. And the results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that these strains harbored about one to three major proteins. These strains exhibited a wide range of insecticidal spectrum toxic to Aedes aegypti (Diptera) and Pieris rapae Linnaeus, 1758 (Lepidoptera). Particularly, JF19-2 contained cry2Ag gene had the highest insecticidal activity. All these researches mentioned above revealed the diversity and particularity of cry2-type gene resources from Bacillus thuringiensis strains in Sichuan basin.

Rapid cloning, identification, and application of one novel crystal protein gene cry30Fa1 from Bacillus thuringiensis.

In this study, a fast and efficient strategy has been developed for identifying and isolating novel cry genes from Bacillus thuringiensis by combining the PCR-restriction fragment length polymorphism and the single-oligonucleotide nested-PCR method. Using this method, one novel holotype cry gene, cry30Fa1, encoding a polypeptide of 687 amino acid residues with a molecular mass of 77.1 kDa, 74% identical to Cry30Aa1, was cloned from the B. thuringiensis strain BtMC28. Furthermore, the cry30Fa1 gene was successfully expressed in Escherichia coli BL21 (DE3). The Cry30Fa1 proteins, isolated from the cultures of recombinant E. coli, had remarkable insecticidal effects against Plutella xylostella and Aedes aegypti with LC50 at 6.477 and 15.359 mug mL(-1), respectively. Our results strongly suggest that this strategy is highly efficient and advantageous in terms of rapid cloning of holotype cry genes that have minimal identity to known genes. The cloning of the cry30Fa1 gene would be useful in the resources of the insecticidal crystal genes and may serve as an alternative choice of an insecticide for potential problems associated with insect resistance.