Impact of Bilateral Subcostal Plus Lateral Transversus Abdominis Plane Block on Quality of Recovery After Laparoscopic Cholecystectomy: A Randomized Placebo-Controlled Trial.

BACKGROUND: Previous research has not evaluated the potential effect of transversus abdominis plane (TAP) block on quality of recovery following laparoscopic cholecystectomy. Therefore, we investigated whether addition of the bilateral subcostal and lateral TAP (bilateral dual TAP [BD-TAP]) blocks to multimodal analgesia would improve the quality of recovery as assessed with the Quality of Recovery-40 (QoR-40). METHODS: Patients age 18 to 60 years who were scheduled to undergo elective laparoscopic cholecystectomy were randomized to the BD-TAP or control group. The BD-TAP group received the BD-TAP block with multimodal analgesia under general anesthesia, using 0.25% ropivacaine, and the control group was treated with the same method, except that they received the sham block using 0.9% normal saline. Both groups had the same multimodal analgesia regimen, consisting of intravenous dexamethasone, propacetamol, ibuprofen, and oxycodone. The primary outcome was the QoR-40 score at 24 hours after surgery. Data were analyzed using the independent t test, Mann-Whitney U test, χ2 test, and Fisher exact test. RESULTS: Thirty-eight patients in each group were recruited. The mean QoR-40 score decreased by 13.6 (95% confidence interval [CI], 8.3-18.8) in the BD-TAP group and 15.6 (95% CI, 6.7-24.5) in the control group. The postoperative QoR-40 score at 24 hours after surgery did not differ between the 2 groups (BD-TAP group, median [interquartile range], 170.5 [152-178]; control group, 161 [148-175]; median difference, 3 [95% CI, -5 to 13]; P = .427). There were no differences between the 2 groups in the pain dimension of the QoR-40: 30.5 (95% CI, 27-33) in the BD-TAP group and 31 (95% CI, 26-32) in the control group; median difference was 0 (95% CI, -2 to 2); P = .77. CONCLUSIONS: Our results indicate that the BD-TAP block does not improve the quality of recovery or analgesic outcomes following laparoscopic cholecystectomy. Our results do not support the routine use of the BD-TAP block for this surgery.

Characterization of a novel cold-active esterase isolated from swamp sediment metagenome.

A functional screen of a metagenomic library from "Upo" swamp sediment in Korea identified a gene EstL28, the product of which displayed lipolytic properties on a tributyrin-supplemented medium. The EstL28 sequence encodes a 290 amino acid protein (designated as EstL28), with a predicted molecular weight of 31.3 kDa. The encoded EstL28 protein exhibited the highest sequence similarity (45 %) to a hydrolase found in Streptococcus sanguinis. Phylogenetic analysis indicated that EstL28 belongs to a currently uncharacterized family of esterases. Within the conserved α/ß-hydrolase 6 domain, the EstL28 retains the catalytic triad Ser103-Asp248-His268 that is typical of esterases. The Ser103 residue in the catalytic triad is located in the consensus pentapeptide motif GXSXG. The purified EstL28 enzyme worked optimally at 35 °C and pH 8.5 and remained stable at temperatures lower than 20 °C. The catalytic activity of EstL28 was maximal with p-nitrophenyl butyrate, indicating that it was an esterase. This enzyme also exhibited stable activity in the presence of methanol, ethanol, isopropanol, and dimethyl sulfoxide. Therefore, the level of stability in organic solvents and cold temperature suggests that EstL28 has potential for many biotechnological applications.

Effect of glycopyrrolate on the postoperative urine output of patients following kidney transplantation: a retrospective observational study.

OBJECTIVES: To compare the urine output and estimated glomerular filtration rate (eGFR) of patients postoperatively administered sugammadex or glycopyrrolate 7 days following kidney transplantation (KT). METHODS: We retrospectively enrolled 134 consecutive patients who underwent KT under general anesthesia. Their urine output and eGFR were recorded every 24 hours between postoperative day (POD) 1 and 7. We used regression analysis to evaluate the relationship between the reversal agent administered and the outcomes of the participants. RESULTS: The urine output and eGFR of the participants did not differ between the two groups. Multivariate analysis showed that body mass index (BMI) (odds ratio (OR) 1.21; 95% confidence interval (CI) 1.05-1.40), diabetes mellitus (OR 3.14; 95% CI 1.07-9.16), neurovascular disease (OR 7.00; 95% CI 1.61-30.42), and the duration of surgery (OR 1.01; 95% CI 1.00-1.01) were associated with lower urine output on POD 7. In addition, only BMI (OR 1.25; 95% CI 1.09-1.42) was associated with low eGFR on POD 7. CONCLUSIONS: The urine output and eGFR of patients administered sugammadex or glycopyrrolate following KT did not differ 7 days later. Moreover, glycopyrrolate does not affect urine output or eGFR on POD 7, according to multivariate regression analysis.

Overexpression of Brassica rapa SHI-RELATED SEQUENCE genes suppresses growth and development in Arabidopsis thaliana.

S HI-R ELATED SEQUENCE (SRS) genes are plant-specific transcription factors containing a zinc-binding RING finger motif, which play a critical role in plant growth and development. We have characterized six SRS genes in Brassica rapa. Overexpression of the SRSs BrSTY1, BrSRS7, and BrLRP1 induced dwarf and compact plants, and significantly decreased primary root elongation and lateral root formation. Additionally, the transgenic plants had upward-curled leaves of narrow widths and with short petioles, and had shorter siliques and low fertility. In stems, hypocotyls, and styles, epidermal cell lengths were also significantly reduced in transgenic plants. RT-PCR analysis of transgenic plants revealed that BrSTY1, BrSRS7, and BrLRP1 regulate expression of several gibberellin (GA)- and auxin-related genes involved in morphogenesis in shoot apical regions. We conclude that BrSTY1, BrSRS7, and BrLRP1 regulate plant growth and development by regulating expression of GA- and auxin-related genes.

Intraoperative use of noninvasive ventilation during spinal anaesthesia in patients with severe chronic obstructive pulmonary disease undergoing orthopaedic surgery: A case report.

Compared with invasive mechanical ventilation, noninvasive ventilation (NIV) improves patient comfort and neurocognitive function; and reduces the likelihood of nosocomial infections and the need for sedation. NIV can also be used perioperatively to prevent postoperative pulmonary complications. This current report describes a case of a 64-year-old female patient with chronic obstructive pulmonary disease and chronic respiratory failure that underwent spinal anaesthesia during surgery. She was sedated with propofol. She brought her home ventilator equipment to the operating room and it was used in biphasic-positive airway pressure mode for immediate treatment of respiratory depression.

Efficacy of preoxygenation with end-tidal oxygen when using different oxygen concentrations in patients undergoing general surgery: a single-center retrospective observational study.

BACKGROUND: Preoxygenation is a simple but very important procedure for preventing arterial desaturation. A higher fraction of inspired oxygen (FiO2) increases atelectasis and 80% oxygen results in significantly less atelectasis than 100% oxygen. We investigated whether there was a difference in the duration of adequate preoxygenation when using 100% and 80% oxygen. The proportion of patients for whom >3 min was required to achieve adequate preoxygenation was also investigated. METHODS: The VitalDB database of patients underwent general surgery between February 1, 2021 and November 12, 2021 was reviewed. The time between the start of preoxygenation and the point where a 10% difference between FiO2 and end-tidal oxygen (EtO2) was defined as the preoxygenation time. The patients were classified into 100% and 80% groups according to the oxygen concentration. Propensity score matching (PSM) was performed to control for potential confounding factors. RESULTS: Only 330 of the 1,377 patients had sufficient data for analysis: 179 in the 80% group and 151 in the 100% group. After PSM, 143 patients in each group were analyzed. The median preoxygenation time was 143 s [interquartile range (IQR): 120.5-181.5 s] and 144 s (IQR: 109.75-186.25 s) in the 80% and 100% groups, respectively [P=0.605; median difference =-1 s; 95% confidence interval (CI): -13 to 10]. Of the patients, 27% required >3 min for adequate preoxygenation. CONCLUSIONS: No difference in preoxygenation time was found between the 80% and 100% groups. For some patients, breathing for 3 min is not sufficient for adequate preoxygenation. EtO2 monitoring aids evaluation of whether preoxygenation was adequate.

Successful extracorporeal membrane oxygenation in a patient with central airway obstruction due to an endotracheal mass.

Extracorporeal membrane oxygenation (ECMO) assists blood circulation and gas exchange via a heart-lung machine. ECMO is used mainly in intensive care units as bridging therapy until heart and respiratory failure can be addressed or until transplantation can be performed. ECMO is sometimes used during surgery under general anaesthesia, depending on the patient's underlying diseases and the nature of the operation. If the oxygen supply and carbon dioxide removal capacity are limited, venovenous (VV)-ECMO can be helpful. Here, we describe the use of VV-ECMO for surgical resection of an endotracheal mass through rigid bronchoscopy in a patient who developed decompensating dyspnoea due to central airway obstruction (CAO).

Prediction of blood pressure changes associated with abdominal pressure changes during robotic laparoscopic low abdominal surgery using deep learning.

BACKGROUND: Intraoperative hypertension and blood pressure (BP) fluctuation are known to be associated with negative patient outcomes. During robotic lower abdominal surgery, the patient's abdominal cavity is filled with CO2, and the patient's head is steeply positioned toward the floor (Trendelenburg position). Pneumoperitoneum and the Trendelenburg position together with physiological alterations during anesthesia, interfere with predicting BP changes. Recently, deep learning using recurrent neural networks (RNN) was shown to be effective in predicting intraoperative BP. A model for predicting BP rise was designed using RNN under special scenarios during robotic laparoscopic surgery and its accuracy was tested. METHODS: Databases that included adult patients (over 19 years old) undergoing low abdominal da Vinci robotic surgery (ovarian cystectomy, hysterectomy, myomectomy, prostatectomy, and salpingo-oophorectomy) at Soonchunhyang University Bucheon Hospital from October 2018 to March 2021 were used. An RNN-based model was designed using Python3 language with the PyTorch packages. The model was trained to predict whether hypertension (20% increase in the mean BP from baseline) would develop within 10 minutes after pneumoperitoneum. RESULTS: Eight distinct datasets were generated and the predictive power was compared. The macro-average F1 scores of the datasets ranged from 68.18% to 72.33%. It took only 3.472 milliseconds to obtain 39 prediction outputs. CONCLUSIONS: A prediction model using the RNN may predict BP rises during robotic laparoscopic surgery.

Novel metagenome-derived carboxylesterase that hydrolyzes ß-lactam antibiotics.

It has been proposed that family VIII carboxylesterases and class C ß-lactamases are phylogenetically related; however, none of carboxylesterases has been reported to hydrolyze ß-lactam antibiotics except nitrocefin, a nonclinical chromogenic substrate. Here, we describe the first example of a novel carboxylesterase derived from a metagenome that is able to cleave the amide bond of various ß-lactam substrates and the ester bond of p-nitrophenyl esters. A clone with lipolytic activity was selected by functional screening of a metagenomic library using tributyrin agar plates. The sequence analysis of the clone revealed the presence of an open reading frame (estU1) encoding a polypeptide of 426 amino acids, retaining an S-X-X-K motif that is conserved in class C ß-lactamases and family VIII carboxylesterases. The gene was overexpressed in Escherichia coli, and the purified recombinant protein (EstU1) was further characterized. EstU1 showed esterase activity toward various chromogenic p-nitrophenyl esters. In addition, it exhibited hydrolytic activity toward nitrocefin, leading us to investigate whether EstU1 could hydrolyze ß-lactam antibiotics. EstU1 was able to hydrolyze first-generation ß-lactam antibiotics, such as cephalosporins, cephaloridine, cephalothin, and cefazolin. In a kinetic study, EstU1 showed a similar range of substrate affinities for both p-nitrophenyl butyrate and first-generation cephalosporins while the turnover efficiency for the latter was much lower. Furthermore, site-directed mutagenesis studies revealed that the catalytic triad of EstU1 plays a crucial role in hydrolyzing both ester bonds of p-nitrophenyl esters and amide bonds of the ß-lactam ring of antibiotics, implicating the predicted catalytic triad of EstU1 in both activities.

Ultrasound-guided pulsed radiofrequency treatment for distal suprascapular neuropathy: A case report.

RATIONALE: Suprascapular neuropathy is a rare cause of shoulder pain, and patients usually presents with posterosuperior shoulder pain and weakness on forward flexion and external rotation. Suprascapular neuropathy associated with rotator cuff pathology has received attention as an emerging cause of this condition. Suprascapular nerve (SSN) block can be used in these patients, and pulsed radio frequency (PRF) can be applied to achieve a long-term effect. Several studies have reported on PRF treatment of the SSN for shoulder pain, but most applied treatment to the nerve trunk under the transverse scapular ligament. This report describes a patient with suprascapular neuropathy treated with selective application of PRF to the distal SSN under ultrasound guidance. PATIENT CONCERNS: A 68-year-old woman suffered from right posterior shoulder pain after traumatic full thickness rotator cuff tear. Her pain was not diminished despite of 2 surgeries. DIAGNOSES: She was diagnosed with entrapment of the distal SSN in the spino-glenoid (SGN) notch and suprascapular neuropathy. INTERVENTIONS: She underwent surgery to decompress the entrapped SSN in the SGN. After that, we applied PRF on the distal SSN under ultrasound guidance for persistent pain. This treatment was repeated 3 times. OUTCOMES: PRF treatment resulted in a slight reduction in the visual analogue scale (VAS) pain score from 7-8/10 to 5-6/10 at the 2 weeks follow-up, and to 2-3/10 at the 1 month follow-up. The reduction in pain was maintained at the 1 year follow-up. LESSONS: PRF treatment of the SSN is typically approached from the main branch in the suprascapular notch. We selectively applied PRF to the distal SSN close to the SGN. This technique was safe and effective.

A case of one-lung ventilation using a single-lumen tube placed under fiberoptic bronchoscopic guidance in a 4-year-old child: A case report.

RATIONALE: One-lung ventilation (OLV) is essential for adequate visualization and exposure of the surgical site via a videoscopic approach. Although many instruments facilitating OLV are available, the choice is limited in pediatric patients. PATIENT CONCERNS: A 4-year-old female (weight: 18.6 kg, height: 100 cm) was admitted via our pediatric outpatient clinic because of recurrent hemoptysis, 2 weeks in duration. She had no medical or surgical history. DIAGNOSIS: Contrast-enhanced computed tomography (CT) revealed a 4.5-cm-diameter mass in the left, lower lung lobe. She was diagnosed with a congenital pulmonary airway malformation (CPAM). INTERVENTIONS: She was scheduled for emergency lobectomy via video-assisted thoracoscopic surgery (VATS). To ensure successful VATS, OLV was essential. As our hospital lacked a small-diameter fiberoptic bronchoscope and a proper bronchial blocker, we decided to use single-lumen tube (SLT) with adult fiberoptic bronchoscope. OUTCOMES: We performed successful bronchoscopic-guided OLV using a SLT. We aligned the tube to the right upper lobar bronchus and Murphy eye to prevent obstruction of the right upper lobe bronchus. At the end of surgery, the endotracheal tube lumen had been narrowed by blood clots, we decided to exchange the tracheal tube. The tube was immediately exchanged. After re-intubation, the pulse oximetry (SpO2) then gradually increased. LESSONS: Appropriate preparation and careful management should be considered to perform OLV in pediatric patients without significant complications.

Alpha- and beta-tubulin from Phytophthora capsici KACC 40483: molecular cloning, biochemical characterization, and antimicrotubule screening.

Internal fragments of alpha- and beta-tubulin genes were generated using reverse transcription polymerase chain reaction (RT-PCR), and the termini were isolated using 5'- and 3'-rapid amplification of cDNA ends. Phytophthora capsici alpha- and beta-tubulin specific primers were then used to generate full-length cDNA by RT-PCR. The recombinant alpha- and beta-tubulin genes were expressed in Escherichia coli BL21 (DE3), purified under denaturing conditions, and average yields were 3.38-4.5 mg of alpha-tubulin and 2.89-4.0 mg of beta-tubulin, each from 1-l culture. Optimum conditions were obtained for formation of microtubule-like structures. A value of 0.12 mg/ml was obtained as the critical concentration of polymerization of P. capsici tubulin. Benomyl inhibited polymerization with half-maximal inhibition (IC(50)) = 468 +/- 20 microM. Approximately 18.66 +/- 0.13 cysteine residues per tubulin dimer were accessible to 5,5'-dithiobis-(2-nitrobenzoic acid), a quantification reagent of sulfhydryl and 12.43 +/- 0.12 residues were accessible in the presence of 200 microM benomyl. The order of preference for accessibility to cysteines was benomyl > colchicine > GTP > taxol, and cysteine accessibility changes conformed that binding sites of these ligands in tubulin were folding correctly. Fluorescence resonance energy transfer technique was used for high throughput screening of chemical library in search of antimitotic agent. There was significant difference in relative fluorescence by 210-O-2 and 210-O-14 as compared to colchicine.

Identification of a novel 4-hydroxyphenylpyruvate dioxygenase from the soil metagenome.

4-Hydroxyphenylpyruvate dioxygenase (HPPD) is a Fe(II)-dependent, non-heme oxygenase that converts 4-hydroxyphenylpyruvate to homogentisate. Essential cofactors, such as plastoquinone and tocopherol, are produced by HPPD-dependent anabolic pathways in plants. To isolate a novel hppd using culture-independent method, a cosmid metagenomic library was constructed from soil in Korea. Screening of Escherichia coli metagenomic libraries led to the identification of a positive clone, YS103B, producing dark brown pigment in Luria-Bertani medium supplemented with l-tyrosine. In vitro transposon mutagenesis of YS103B showed that the 1.3kb insert was sufficient to produce the hemolytic brown pigment. Sequence analysis of YS103B disclosed one open reading frame encoding a 41.4kDa protein with the well-conserved prokaryotic oxygenase motif of the HPPD family of enzymes. The HPPD-specific beta-triketone herbicide, sulcotrione, inhibited YS103B pigmentation. The recombinant protein expressed in E. coli generated homogentisic acid. Thus, we present the successful heterologous expression of a previously uncharacterized hppd gene from an uncultured soil bacterium.

Characterization of a gene encoding cellulase from uncultured soil bacteria.

To detect cellulases encoded by uncultured microorganisms, we constructed metagenomic libraries from Korean soil DNAs. Screenings of the libraries revealed a clone pCM2 that uses carboxymethyl cellulose (CMC) as a sole carbon source. Further analysis of the insert showed two consecutive ORFs (celM2 and xynM2) encoding proteins of 226 and 662 amino acids, respectively. A multiple sequence analysis with the deduced amino acid sequences of celM2 showed 36% sequence identity with cellulase from the Synechococcus sp., while xynM2 had 59% identity to endo-1,4-beta-xylanase A from Cellulomonas pachnodae. The highest enzymatic CMC hydrolysis was observable at pH 4.0 and 45 degrees C with recombinant CelM2 protein. Although the enzyme CelM2 additionally hydrolyzed avicel and xylan, no substrate hydrolysis was observed on oligosaccharides such as cellobiose, pNP-beta-cellobioside, pNP-beta-glucoside, and pNP-beta-xyloside. These results showed that CelM2 is a novel endo-type cellulase.

Cloning, purification, and polymerization of Capsicum annuum recombinant alpha and beta tubulin.

Alpha and beta tubulin genes were cloned from the Capsicum annuum leaves using rapid amplification of cDNA ends (RACE)-PCR. Nucleotide sequence analysis revealed that 1,353 bp Capsicum annuum alpha/beta-tubulin (CAnm alpha/beta-TUB) encodes a protein of 450 amino acids (aa) each. The recombinant alpha/beta tubulin was overexpressed mainly as an inclusion body in Escherichia coli BL21 (DE3), upon induction with 0.2 mM isopropyl-beta-D-thiogalactopyranoside (IPTG), and its content was as high as 50% of the total protein content. Effective fusion protein purification and refolding are described. The average yields of alpha and beta tubulin were 2.0 and 1.3 mg/l of culture respectively. The apparent molecular weight of each tubulin was estimated to be 55 kDa by SDS-polyacrylamide gel electrophoresis (PAGE). The tubulin monomers were found to be assembly competent using a standard dimerization assay, and also retained antigenicity with anti-His/T7 antibodies. The purified tubulins were polymerized to microtubule-like structures in the presence of 2 mM guanosine 5'-triphosphate (GTP).

Analysis of caudal epidurogram in single center: A preliminary study of lumbar radiculopathy management.

A caudal epidural block involves placing a needle through the sacral hiatus and delivering medication into the epidural space. The procedure is safe and simple, but failure rates can be as high as 25%. The purpose of this study was to investigate the success rate of caudal epidural block by analyzing needle placement and dye flow pattern.We retrospectively analyzed the medical records of patients who underwent caudal epidural block under spinal stenosis. A case was defined as a failure if it met at least one of the following four criteria: the epidural needle was not placed correctly inside the caudal canal; blood regurgitation or aspiration in the needle was observed; the contrast dye was injected into a blood vessel; or a large amount of the dye leaked into the sacral foramen or did not reach the L5-S1 level.At least 1 failure criterion was observed in 14 cases (17.7%), while none of the failure criteria were satisfied in 65 successful cases (82.3%).No matter how experienced the anesthesiologist may be, delivery of adequate therapeutic agent is not achieved in approximately 20% of cases. Therefore, we recommend fluoroscopy-guided needle placement and confirmation by radio-contrast epidurograpy as the best choice.

Surfactin from Bacillus subtilis displays anti-proliferative effect via apoptosis induction, cell cycle arrest and survival signaling suppression.

The effect of surfactin on the proliferation of LoVo cells, a human colon carcinoma cell line, was examined. Surfactin strongly blocked the proliferation of LoVo cells by inducing pro-apoptotic activity and arresting the cell cycle, according to several lines of evidence on DNA fragmentation, Annexin V staining, and altered levels of poly (ADP-ribose) polymerase, caspase-3, p21(WAF1/Cip1), p53, CDK2 and cyclin E. The anti-proliferative activity of surfactin was mediated by inhibiting extracellular-related protein kinase and phosphoinositide 3-kinase/Akt activation, as assessed by phosphorylation levels. Therefore, our data suggest that surfactin may have anti-cancer properties as a result of its ability to downregulate the cell cycle and suppress its survival.

The genome sequence of Xanthomonas oryzae pathovar oryzae KACC10331, the bacterial blight pathogen of rice.

The nucleotide sequence was determined for the genome of Xanthomonas oryzae pathovar oryzae (Xoo) KACC10331, a bacterium that causes bacterial blight in rice (Oryza sativa L.). The genome is comprised of a single, 4 941 439 bp, circular chromosome that is G + C rich (63.7%). The genome includes 4637 open reading frames (ORFs) of which 3340 (72.0%) could be assigned putative function. Orthologs for 80% of the predicted Xoo genes were found in the previously reported X.axonopodis pv. citri (Xac) and X.campestris pv. campestris (Xcc) genomes, but 245 genes apparently specific to Xoo were identified. Xoo genes likely to be associated with pathogenesis include eight with similarity to Xanthomonas avirulence (avr) genes, a set of hypersensitive reaction and pathogenicity (hrp) genes, genes for exopolysaccharide production, and genes encoding extracellular plant cell wall-degrading enzymes. The presence of these genes provides insights into the interactions of this pathogen with its gramineous host.

Screening and characterization of an enzyme with beta-glucosidase activity from environmental DNA.

A novel beta-glucosidase gene, bglA, was isolated from uncultured soil bacteria and characterized. Using genomic libraries constructed from soil DNA, a gene encoding a protein that hydrolyzes a fluorogenic analog of cellulose, 4-methylumbelliferyl beta-D-cellobioside (MUC), was isolated using a microtiter plate assay. The gene, bglA, was sequenced using a shotgun approach, and expressed in E. coli. The deduced 55-kDa amino acid sequence for bglA showed a 56% identity with the family 1 glycosyl hydrolase Chloroflexus aurantiacus. Bg1A included two conserved family 1 glycosyl hydrolase regions. When using p-nitrophenyl-beta-D-glucoside (pNPG) as the substrate, the maximum activity of the purified beta-glucosidase exhibited at pH 6.5 and 55 degrees C, and was enhanced in the presence of Mn2+. The Km and Vmax values for the purified enzyme with pNPG were 0.16 mM and 19.10 micromol/min, respectively. The purified BglA enzyme hydrolyzed both pNPG and p-nitrophenyl-beta-D-fucoside. The enzyme also exhibited substantial glycosyl hydrolase activities with natural glycosyl substrates, such as sophorose, cellobiose, cellotriose, cellotetraose, and cellopentaose, yet low hydrolytic activities with gentiobiose, salicin, and arbutin. Moreover, Bg1A was able to convert the major ginsenoside Rb1 into the pharmaceutically active minor ginsenoside Rd within 24 h.

Generation of a specific marker to discriminate Gacillus anthracis from other bacteria of the Bacillus cereus group.

Bacillus anthracis is a soil pathogen capable of causing anthrax that is closely related to several environmental species, including B. cereus, B. mycoides, and B. thuringiensis. DNA homology studies showed that B. anthracis, B. cereus, B. mycoides, and B. thuringiensis are closely related, with a high sequence homology. To establish a method to specifically detect B. anthracis in situations such as environmental contamination, we initially performed RAPD-PCR with a 10-mer random primer and confirmed the presence of specific PCR bands only in B. anthracis species. One region specific for B. anthracis was cloned and sequenced, and an internal primer set was designed to amplify a 241-bp DNA fragment within the sequenced region. The PCR system involving these specific primer sets has practical applications. Using lyses methods to prepare the samples for PCR, it was possible to quickly amplify the 241-bp DNA segment from samples containing only a few bacteria. Thus, the PCR detection method developed in this study is expected to facilitate the monitoring of environmental B. anthracis contamination.