Clinical efficacy and imaging analysis of oblique lateral lumbar interbody fusion in the treatment of different types of lumbar intervertebral foramen stenosis.

PURPOSE: To analyze and study the clinical efficacy and imaging indexes of oblique lateral lumbar interbody fusion (OLIF) in the treatment of lumbar intervertebral foramen stenosis(LFS) caused by different causes. METHOD: 33 patients with LFS treated with OLIF from January 2018 to May 2022 were reviewed. Oswestry Dysfunction Index (ODI) and visual analogue scale (VAS) were calculated before and after operation. Segmental lordotic angle (SLA), lumbar lordotic angle (LLA) and segmental scoliosis angle (SSA), disc height (DH), posterior disc height (PDH), lateral disc height (LDH), foraminal height (FH), foramen width (FW) and foraminal cross-sectional area (FSCA) were measured before and after operation. RESULT: The VAS and ODI after operation were significantly improved as compared with those before operation. Compared with pre-operation, the DH, PHD increased by 67.6%, 94.6%, LDH increased by 107.4% (left), 101.7% (right), and FH increased by 30.2% (left), 34.5% (right). The FSCA increased by 93.1% (left), 89.0% (right), and the FW increased by 137.0% (left), 149.6% (right). The postoperative SSA was corrected by 74.5%, the postoperative SLA, LLA were corrected by 70.2%, 38.1%, respectively. All the imaging indexes were significantly improved (p < 0.01). CONCLUSION: The clinical efficacy and imaging data of OLIF in the treatment of LFS caused by low and moderate lumbar spondylolisthesis, intervertebral disc bulge and reduced intervertebral space height, degenerative lumbar scoliosis, articular process hyperplasia or dislocation have been well improved. OLIF may be one of the better surgical treatments for LFS caused by the above conditions.

How to Choose Surgical Corridor in Left Oblique Approach Lumbar Interbody Fusion at the L5-S1 Segment: A Prospective Cohort Study.

OBJECTIVE: There are 2 surgical corridors to L5-S1 lumbar interbody fusion via the left oblique approach: anterior to psoas-oblique lateral interbody fusion (ATP-OLIF) and oblique-anterior lumbar interbody fusion (O-ALIF). The aim of this study was to evaluate criteria to guide the selection of surgical corridors for L5-S1 lumbar interbody fusion via the left oblique approach. METHODS: According to the structure of L5-S1 segment left common iliac vein (LCIV) in axial magnetic resonance image, the LCIV was divided into 6 types. O-ALIF was performed for type I and type II. ATP-OLIF was performed for type A and type B. For sexually active men, ATP-OLIF was chosen. Between April 2020 and April 2022, 22 patients were assigned to ATP-OLIF or O-ALIF based on the type of LCIV. Clinical outcomes and radiographic outcomes were assessed. RESULTS: There were 11 cases in O-ALIF group (type I, n = 10; type II, n = 1) and 11 cases in ATP-OLIF group (type A, n = 8; type B, n = 3). No differences were observed in clinical outcomes (Oswestry Disability Index, VAS, and complication rate); radiographic outcomes (mean disk height and segmental lordosis angle); length of hospital stay; operation time; and blood loss. No vascular injury occurred in either group. CONCLUSIONS: This may be an appropriate criterion to guide the selection of surgical corridor for L5-S1 lumbar interbody fusion through the left oblique approach. O-ALIF was performed for type I and type II. ATP-OLIF was performed for type A and type B. For sexually active men, ATP-OLIF was chosen. According to this standard, the operation can be performed safely and with good clinical results.

Bridging the cervicothoracic junction during posterior cervical laminectomy and fusion for the treatment of multilevel cervical ossification of the posterior longitudinal ligament: a retrospective case series.

BACKGROUND: The purpose of this study was to investigate the surgical efficacy of crossing the cervicothoracic junction during posterior cervical laminectomy and fusion for the treatment of multilevel cervical ossification of the posterior longitudinal ligament (OPLL). METHODS: From October 2009 to October 2017, 46 consecutive patients with multilevel cervical OPLL underwent posterior cervical laminectomy and crossing the cervicothoracic junction fusion were obtained in the study. Their medical records were retrospectively collected. Cervical lordosis and cervical sagittal balance were used to assess radiographic outcomes. Japanese Orthopedic Association (JOA), axial symptom, C5 root palsy, blood loss, and operation time were used to assess clinical outcomes. The mean follow-up period was 20.7 ± 8.3 months. RESULTS: The operation time was 205.2 ± 39.8 min and the intraoperative blood loss was 352.2 ± 143.7 ml. Analysis of the final follow-up data showed significant differences in JOA score (P < 0.01), C2-C7 lordosis angle (P < 0.01), and C2-C7 SVA (P < 0.01). CT confirmed that grafted bone was completely fused in all patients and progression of OPLL was observed in two patients (4.3%) at final follow-up. No adjacent segment disease (ASD) or instrument failure occurred in any patients. CONCLUSIONS: Cervical laminectomy and crossing the cervicothoracic junction fusion are effective and safe methods to treat multilevel cervical OPLL. Randomized controlled studies compared constructs ending at cervical vertebrae or thoracic vertebrae are needed to confirm these results.

A clinical study on the factors associated with nasopharyngeal carcinoma among the Chinese population.

Nasopharyngeal carcinoma (NC) arises from the nasopharynx epithelium and the majority of NC cases globally are within China and Southeast Asia. Both short palate lung and nasal epithelium clone 1 (SPLUNC1) and myelodysplasia syndrome 1-ectopic viral integration site 1 (MDS1-EVI1) play an important role in carcinogenesis and have been found to be associated with nasopharyngeal carcinoma. In spite of their role in NC, the association between these genes and their polymorphisms in the development of NC has thus far not been studied. In the present study, the relationship between SPLUNC1 (rs2752903, T>C) and MDS1-EVI1 (rs6774494, G>A) polymorphisms and their role in the development of NC among the Chinese population were investigated. From a Chinese population of 1,059 patients with NC and 891 controls, genotype frequencies and the distribution of SPLUNC1 and MDS1-EVI1 polymorphisms were analyzed for possible susceptibility to NC. It was observed that those with MDS1-EVI1 CC (OR, 2.76; 95% CI, 1.96-3.81) and MDS1-EVI1 CT (OR, 1.51; 95% CI, 1.22-2.14) polymorphisms had an increased risk of developing NC. Those with SPLUNC1 AA genotypes also observed a higher risk for NC compared with SPLUNC1 GG genotypes (OR, 2.15; 95% CI, 1.62-3.15). When observing the gene-gene interaction between SPLUNC1 and MDS1-EVI1 polymorphisms, it was found that the presence of both SPLUNC1 CC and MDS1-EVI1 AA alleles was associated with a higher risk for NC compared with those who did not carry both alleles (OR, 6.75; 95% CI, 3.41-12.11). The present study suggested that the association between SPLUNC1 (rs2752903, T>C) and MDS1-EVI1 (rs6774494, G>A) polymorphisms may be a potent risk factor in the occurrence of NC.

Modified grade 4 osteotomy for the correction of post-traumatic thoracolumbar kyphosis: A retrospective study of 42 patients.

Many surgical procedures have been developed for the treatment of post-traumatic thoracolumbar kyphosis. But there is a significant controversy over the ideal management. The aim of this study was to illustrate the technique of modified grade 4 osteotomy for the treatment of post-traumatic thoracolumbar kyphosis and to evaluate clinical and radiographic results of patients treated with this technique.From May 2013 to May 2018, 42 consecutive patients experiencing post-traumatic thoracolumbar kyphosis underwent the technique of modified grade 4 osteotomy, and their medical records were retrospectively collected. Preoperative and postoperative sagittal Cobb angle, visual analog scale (VAS), Oswestry disability index (ODI), and American Spinal Injury Association (ASIA) were recorded. The average follow-up period was 29.7 ±â€Š14.2 months.The operation time was 185.5 ±â€Š26.8 minutes, the intraoperative blood loss was 545.2 ±â€Š150.1 mL. The Cobb angles decreased from 38.5 ±â€Š3.8 degree preoperatively to 4.2 ±â€Š2.6 degree 2 weeks after surgery (P < .001). The VAS reduced from 6.5 ±â€Š1.1 preoperatively to 1.5 ±â€Š0.9 at final follow-up (P < .001), and the ODI reduced from 59.5 ±â€Š15.7 preoperatively to 15.9 ±â€Š5.8 at final follow-up (P < .001). Kyphotic deformity was successfully corrected and bony fusion was achieved in all patients. Neurologic function of 7 cases was improved to various degrees.Modified grade 4 osteotomy, upper disc, and upper one-third to half of pedicle are resected, is an effective treatment option for post-traumatic thoracolumbar kyphosis. However, the long-term clinical effect still needs further studies.

Modified grade 4 osteotomy for kyphosis due to old osteoporotic vertebral compression fractures: Two case reports.

RATIONALE: The kyphosis caused by old osteoporotic vertebral compression fracture usually requires osteotomy to correct it. Various osteotomy techniques have been reported, but each has its own advantages and disadvantages. PATIENT CONCERNS: We reviewed 2 cases of old osteoporotic vertebral compression fractures with kyphosis in our hospital. One patient complained of persistent low-back pain, another patient complained of low-back pain and weakness of both lower extremities. DIAGNOSIS: Old osteoporotic vertebral compression fractures with kyphosis were diagnosed based on computer tomography and magnetic resonance imaging. INTERVENTIONS: We performed modified grade 4 osteotomy for 2 patients. OUTCOMES: Both patients said significant improvement in preoperative symptoms, and x-ray showed that the kyphosis was corrected. Both patients were satisfied with the treatment at the last follow-up, and the kyphosis was not aggravated. LESSONS: Modified grade 4 osteotomy is an effective option for the treatment of old osteoporotic fracture with kyphosis. It can restore the spine sequence and achieve better clinical result.

Identification modules of gastric cancer based on protein-protein interaction networks and gene expression data.

PURPOSE: The tumorigenesis of gastric cancer is an intricate process which contains genetic and epigenetic changes of proto-oncogenes and cancer-suppressor genes. The purpose of this study was to identify novel modules for gastric cancer based on protein-protein interaction networks and gene expression data. METHODS: Microarray data and corresponding annotated files of E-GEOD-15460 were downloaded from ArrayExpress database. All human protein-protein interactions were downloaded from STRING database. The fast depth-first assay was used to identify all maximal cliques of disease group and control group. Benjamini-Hochberg method was used to perform multiple corrections of p value. RESULTS: 248 modules for the control group and 30 modules for the disease group were determined in this research, and 734 pairs of similar or same modules of these two groups were detected through calculating module correlation density. Protein-protein interaction (PPI) network was identified, which comprised of 7899 genes and 48469 interrelationship pairs of genes. Finally, 6 modules with remarkable difference were found to be closely related with gastric cancer. CONCLUSIONS: Novel modules with significant difference and the related genes are useful biomarkers and therapeutic targets for gastric cancer.

Contralateral radiculopathy after transforaminal lumbar interbody fusion in the treatment of lumbar degenerative diseases: A Case Series.

RATIONALE: Transforaminal lumbar interbody fusion (TLIF) is an effective treatment for patients with degenerative lumbar disc disorder. Contralateral radiculopathy, as a complication of TLIF, has been recognized in this institution, but is rarely reported in the literature. PATIENT CONCERNS: In this article, we report 2 cases of contralateral radiculopathy after TLIF in our institution and its associated complications. DIAGNOSES: In the 2 cases, the postoperative computed tomography (CT) and magnetic resonance image (MRI) showed obvious upward movement of the superior articular process, leading to contralateral foraminal stenosis. INTERVENTIONS: Revision surgery was done at once to partially resect the opposite superior facet and to relieve nerve root compression. OUTCOMES: After revision surgery, the contralateral radiculopathy disappeared. LESSONS: Contralateral radiculopathy is an avoidable potential complication. It is very important to create careful preoperative plans and to conscientiously plan the use of intraoperative techniques. In case of postoperative contralateral leg pain, the patients should be examined by CT and MRI. If CT and MRI show that the superior articular process significantly migrated upwards, which leads to contralateral foraminal stenosis, revision surgery should be done at once to partially resect the contralateral superior facet so as to relieve nerve root compression and avoid possible long-term impairment.

Vertebral column decortication for the management of sharp angular spinal deformity in Pott disease: Case report.

RATIONALE: Extremely sharp angular spinal deformity of healed tuberculosis can be corrected by vertebral column resection (VCR). However, the VCR techniques have many limitations including spinal column instability, greater blood loss, and greater risk of neurologic deficit. PATIENT CONCERNS: We described a new spinal osteotomy technique to collect sharp angular spinal deformity in Pott disease. A 52-year-old woman presented with back pain and gait imbalance. DIAGNOSIS: The kyphosis of healed tuberculosis was diagnosed based on history and imaging examinations. INTERVENTION: A new posterior VCR was designed to treat this disease. OUTCOMES: The neurological function improved from Japanese Orthopedic Association scale 3 to 7. The back pain and neurological function were significantly improved. The Oswestry Disability Index decreased from 92 to 34. There was also a significant decrease in back pain visual analog scale from 9 to 2. LESSONS: For cases with extremely severe Pott kyphotic deformity, the technology of modified VCR offers excellent clinical and radiographic results.

Icariin exerts inhibitory effects on the growth and metastasis of KYSE70 human esophageal carcinoma cells via PI3K/AKT and STAT3 pathways.

Esophageal cancer is one of the leading causes of cancer related mortality across the globe. The current treatment options are insufficient and are associated with number of side effects. Therefore there is a pressing need to develop effective and more efficient strategies for the treatment of esophageal cancer. Consistently, natural products are considered potential candidates for develop of cancer chemotherapy. Icariin is a naturally occurring flavonol glucoside and has been reported to possess tremendous pharmacological potential ranging from neuroprotection to anticancer activity. However, the pharmacological role of icariin in esophageal cancer is still largely unclear. Here in the present study, icariin was evaluated for its anticancer activity against KYSE70 esophageal cancer cells and the possible underlying mechanism was determined. Icariin induced cytotoxicity with an IC50 of 40µM in esophageal cancer cells. These inhibitory effects were due to apoptosis through reactive oxygen species (ROS) mediated alterations in mitochondrial membrane potential (MMP). The results indicated that icariin enhanced the accretion of ROS upto 260% and reduced the MMP upto 48% at 100µM. Icariin also induced G2/M cell cycle arrest as evident from the significant increase in the G2 cell populations of KYSE70 esophageal cancer cells. Additionally, icariin inhibited esophageal cancer cell migration, invasion and metastasis by regulating the expression of epithelial to mesenchymal transition (EMT) markers. Results also indicated that icariin reduced cell viability and migration in part through suppression of the PI3K/AKT and STAT3 pathways. Taken together, our results indicate that icariin may prove a potential natural anticancer molecule against esophageal cancer.

HuR Stabilizes lnc-Sox5 mRNA to Promote Tongue Carcinogenesis.

Long noncoding RNAs (lncRNAs) have been recently regarded as systemic regulators in multiple biological processes including tumorigenesis. In this study, we report an ultra-highly expressed lncRNA, lnc-Sox5, in tongue tumor tissues. The results imply that lnc-Sox5 may play vital role in tongue carcinoma progression. We observed that the growth of Tca8113 cells was suppressed by lnc-Sox5 downregulation. Additionally, lnc-Sox5 knockdown simultaneously increased Tca8113 cell apoptosis, but the cell cycle was arrested. RNA immunoprecipitation suggested that HuR directly bound to and stabilized lnc-Sox5 RNA. Consistently, HuR knockdown reduced the level of lnc-Sox5 in Tca8113 cells. However, overexpression of HuR induced more lnc-Sox5 in Tca8113 cells. Both lnc-Sox5 knockdown and HuR knockdown suppressed Tca8113 cell tumorigenesis in xenograft models. These results suggest that lnc-Sox5, which was stabilized by HuR, could regulate carcinogenesis of tongue cancer and may serve as a predicted target for tongue carcinoma therapies.

Tigecycline targets nonsmall cell lung cancer through inhibition of mitochondrial function.

Nonsmall cell lung cancer (NSCLC) is the most common type of lung cancer with a high mortality rate and still remains therapeutically a challenge. A strategy to target NSCLC is to identify agents that are effective against NSCLC cells while sparing normal cells. We show that tigecycline, an FDA-approved antibiotic drug, preferentially targets NSCLC cells. Tigecycline is effective in inhibiting proliferation and inducing apoptosis of multiple cell lines derived from two common NSCLC subtypes: adenocarcinoma and squamous cell carcinoma. Tigecycline also dose-dependently inhibits colony formation of NSCLC subpopulation of cells with highly proliferative and invasive properties. Compared to NSCLC cells, tigecycline affects proliferation and survival of normal fibroblast cells significantly to a less extent. More importantly, tigecycline significantly inhibits NSCLC tumor growth through decreasing proliferation and increasing apoptosis of tumor cells in vivo. Tigecycline significantly inhibits mitochondrial respiration, mitochondrial membrane potential, and ATP levels and increases reactive oxygen species (ROS), suggesting that tigecycline impairs mitochondrial functions. Our study suggests that tigecycline may be a useful therapeutic agent, and inhibiting mitochondrial functions may represent a new targeted therapy for NSCLC.

Relationship between the laminoplasty opening size and the laminoplasty opening angle, increased sagittal canal diameter and the prediction of spinal canal expansion following open-door cervical laminoplasty.

OBJECTIVE: To clarify the relationship between the laminoplasty opening size (LOS), the laminoplasty opening angle (LOA) and the increase in sagittal canal diameter (SCD) and to predict the amount of canal enlargement during open-door cervical laminoplasty (ODCL). METHODS: Formula describing the relationship between LOS and LOA, the increase in SCD was deduced. The parameters of pre- and postoperative computed tomography scans of 36 patients who had undergone laminoplasty surgery were measured by picture archiving and communication system (PACS) software, and the amount of canal enlargement of these patients was predicted when the opening size of laminoplasty was 8, 10, 12, 14 and 16 mm according to the formula. RESULTS: For equivalent LOS, the amount of canal enlargement with each opening size differed throughout the cervical region. When the C3-C7 LOS was 10 mm the SCD increased >4.1 mm, and the canal area increased in C3-C6 >88 mm(2), and the canal area increased in C7 > 80 mm(2). When the C3-C7 LOS was 12 mm, the SCD increased >5.2 mm, and the canal area increased in C3-C6 >104 mm(2), and the canal area increased in C7 > 94 mm(2). CONCLUSION: Formula accurately showed the relationship between the LOS and the increase in SCD achieved by ODCL. The amount of canal enlargement following ODCL could be predicted by the formula. LOS of 10-12 mm at C3-C7 might be optimal during ODCL.

The relationship between laminoplasty opening angle and increased sagittal canal diameter and the prediction of spinal canal expansion following double-door cervical laminoplasty.

PURPOSE: To clarify the relationship between laminoplasty opening angle (LOA) and the increase in sagittal canal diameter (SCD) in double-door cervical laminoplasty (DDCL) and to predict the increase in SCD using the resulting formula. METHODS: We analyzed 20 patients with multilevel cervical spondylotic myelopathy who underwent DDCL between September 2010 and January 2013. The pre- and post-operative parameters of the cervical spinal canal were measured by computed tomography. We deduced a formula describing the relationship between LOA and the increase in SCD and used it to predict the increase in SCD of these patients as LOA increased. RESULTS: When the C3-C7 LOA was 25°-45°, the magnitude of the increase in SCD was notable (increases of 3.08-5.6 mm compared with the pre-operative SCD). When the C3-C7 LOA was more than 45°, the magnitude of the increase in SCD was relatively smaller; the increase in C3-C7 SCD with a 55° LOA was merely 0.4 mm more than with a 45° LOA. When LOA was 30° at C3-C6 or 40° at C7, the increase in SCD was more than 4 mm. When the C3-C6 LOA was 40°, SCD increased by more than 5 mm. CONCLUSIONS: The formula accurately showed the relationship between LOA and the increase in SCD in DDCL. Based on the LOA, increases in SCD following C3-C7 laminoplasty can be accurately predicted using this formula. This enables DDCL based on accurate individual LOAs, which prevents inadequate or excessive opening.

TMEM16A protein attenuates lipopolysaccharide-mediated inflammatory response of human lung epithelial cell line A549.

OBJECTIVE: To observe the expression of endogenous TMEM16A in rat alveolar type II epithelial cells (AT-II) and A549, and study the effect of TMEM16A on lipopolysaccharide (LPS)-induced proinflammatory cytokine secretion. METHODS: Rat AT-II cells were isolated and TMEM16A protein expression in rat AT-II cells was measured by Western blot. TMEM16A mRNA and protein expressions in A549 were measured by real-time quantitative polymerase chain reaction (PCR) and Western blot, respectively. TMEM16A gene was transfected into A549 using Lipofectamine 2000. Transfected cells were selected in the presence of G418 to create a stable TMEM16A overexpression A549 cell line. The expression of TMEM16A in A549 was knocked down by lentiviral vector-mediated RNA interference. TNF-α and IL-8 levels were determined by enzyme-linked immunosorbent assay (ELISA). A dual-luciferase reporter assay system was used to measure the transcriptional activity of NF-κB. RESULTS: (1) Endogenous TMEM16A was expressed in rat AT-II and A549. (2) TMEM16A expression in A549 significantly increased at 24 hours and 36 hours, and then decreased at 48 hours after LPS treatment. (3) TMEM16A mRNA and protein expressions were increased in the stable TMEM16A overexpression A549 cell line. (4) TMEM16A overexpression decreased the LPS-induced TNF-α and IL-8 secretions. (5) TMEM16A mRNA and protein expressions were knocked down in TMEM16A-siRNA lentivirus transfected A549. (6) TMEM16A knockdown increased the LPS-induced TNF-α and IL-8 secretions. (7) TMEM16A overexpression inhibited LPS-induced NF-κB activation. CONCLUSIONS: TMEM16A is expressed in AT-II. TMEM16A in A549 inhibits LPS-induced NF-κB activation and decreases proinflammatory cytokines release, protecting A549 from acute LPS-mediated damage.

Fuzhengpaidu granule regulates immune activation molecules CD38 and human leukocyte antigen-D related on CD4+ and CD8+ T cells in patients with acquired immunodeficiency syndrome/human immunodeficiency virus.

OBJECTIVE: To evaluate the effect of Fuzhengpaidu granule (FZPDG) on immune activation molecules CD38 and human leukocyte antigen-D related (HLA-DR) on CD4+ and CD8+ cells in HIV/AIDS patients, and to explore the underlying mechanism of this therapy. METHODS: Plasma changes in CD3+, CD4+, CD8+, CD3 + CD4 + CD38 +, CD3 + CD4 + HLA-DR+, CD3 + CD8+CD38+, and CD3+CD8+HLA-DR+ levels in HIV/ AIDS patients treated with FZPDG for six months were examined by flow cytometry and compared with levels in healthy controls. RESULTS: The clinical trial included 34 outpatients with HIV/AIDS. Before treatment, plasma levels of CD38+ and HLA-DR+ on CD4/CD8 cells were higher than those in 28 health controls (P < 0.05). There were no significant changes in serum levels of CD3+, CD4+, and CD8+ T cells between pretreatment baseline versus after treatment, which were 82.85% +/- 5.41%, 14.57% +/- 10.31% and 54.55% +/- 11.43% before treatment and 79.15% +/- 8.21%, 19.96% +/- 9.58% and 56.36% +/- 11.67% after treatment, respectively (P > 0.05). Plasma levels of CD3+ CD4+CD38+ and CD3+CD4+HLA-DR+ were 2.3% +/-2.2% and 7.8% +/- 5.5% before treatment and 1.2% +/-0.8% and 2.6% +/- 1.0% after treatment, respectively. Plasma levels of CD3+CD8+CD38+ and CD3+CD8+ HLA-DR+ were 41.4% +/- 13.4% and 17.8% +/- 11.3% before treatment, which changed to 27.1% +/- 10.2% and 3.8% +/- 2.4% after treatment, respectively (P < 0.05). CONCLUSION: HIV/AIDS patients exhibited an immune activation profile following FZPDG treatment. A potential mechanism of action for FZPDG appears to lie in its ability to up-regulate CD38 and HLA-DR levels on CD4+ T cells, and down-regulate them on CD8+ cells, thereby modulating immune activation of CD4+and CD8+T cells.

Development of a high-efficient transformation system of Bacillus pumilus strain DX01 to facilitate gene isolation via gfp-tagged insertional mutagenesis and visualize bacterial colonization of rice roots.

A Tn5 transposition vector, pMOD-tet-egfp, was constructed and used for the random insertional mutagenesis of Bacillus pumilus. Various parameters were investigated to increase the transformation efficiency B. pumilus DX01 via Tn5 transposition complexes (transposome): bacterial growth phase, type of electroporation buffer, electric field strength, and recovery medium. Transformation efficiency was up to 3 × 10(4) transformants/µg of DNA under the optimized electroporation conditions, and a total of 1,467 gfp-tagged transformants were obtained. Fluorescence-activated cell sorting analysis showed that all gfp-tagged bacterial cells expressed GFP, indicating that foreign DNA has been successfully integrated into the genome of B. pumilus and expressed. Finally, flanking DNA sequences were isolated from several transformants and colonization of rice roots by B. pumilus DX01 was also studied. The method developed here will be useful for creating an insertion mutant library of gram-positive bacteria, thus facilitating their molecular genetic and cytological studies.