Microbial sensor for drug susceptibility testing of Mycobacterium tuberculosis.

AIMS: Drug susceptibility testing (DST) of clinical isolates of Mycobacterium tuberculosis is critical in treating tuberculosis. We demonstrate the possibility of using a microbial sensor to perform DST of M. tuberculosis and shorten the time required for DST. METHODS AND RESULTS: The sensor is made of an oxygen electrode with M. tuberculosis cells attached to its surface. This sensor monitors the residual oxygen consumption of M. tuberculosis cells after treatment with anti-TB drugs with glycerine as a carbon source. In principle, after drug pretreatment for 4-5 days, the response differences between the sensors made of drug-sensitive isolates are distinguishable from the sensors made of drug-resistant isolates. The susceptibility of the M. tuberculosis H37Ra strain, its mutants and 35 clinical isolates to six common anti-TB drugs: rifampicin, isoniazid, streptomycin, ethambutol, levofloxacin and para-aminosalicylic acid were tested using the proposed method. The results agreed well with the gold standard method (LJ) and were determined in significantly less time. The whole procedure takes approximately 11 days and therefore has the potential to inform clinical decisions. CONCLUSIONS: To our knowledge, this is the first study that demonstrates the possible application of a dissolved oxygen electrode-based microbial sensor in M. tuberculosis drug resistance testing. This study used the microbial sensor to perform DST of M. tuberculosis and shorten the time required for DST. SIGNIFICANCE AND IMPACT OF THE STUDY: The overall detection result of the microbial sensor agreed well with that of the conventional LJ proportion method and takes less time than the existing phenotypic methods. In future studies, we will build an O2 electrode array microbial sensor reactor to enable a high-throughput drug resistance analysis.

[Characteristics and related factors of viral nucleic acid negative conversion in children infected with Omicron variant strain of SARS-CoV-2].

Objective: To understand the characteristics and associated factors of viral nucleic acid conversion in children infected with Omicron variant strain of SARS-CoV-2 in Shanghai. Methods: The clinical symptoms, laboratory results and other data of 177 children infected with SARS-CoV-2 who were hospitalized in Shanghai Children's Hospital, School of Medicine, Shanghai Jiao Tong University (designated hospital for SARS-CoV-2 infection in Shanghai) from April 25 to June 8, 2022 were retrospectively analyzed. According to the chest imaging findings, the children were divided into mild and common type groups. According to their age, the unvaccinated children were divided into<3 years old group and 3-<18 years old group. According to the vaccination status, the children aged 3-<18 year were divided into non-vaccination group, 1-dose vaccination group and 2-dose vaccination group. Comparison between groups was performed by independent sample t-test and analysis of variance, and multivariate linear regression analysis was used for multivariate analysis. Results: Among the 177 children infected with Omicron variant of SARS-CoV-2, 96 were males and 81 were females, aged 3 (1, 6) years. The time of viral nucleic acid negative conversion was (10.3±3.1) days. The 177 children were 138 cases of mild type and 39 cases of common type. Among the children aged 3-<18 years old, 55 cases were not vaccinated, 5 cases received 1-dose and 36 cases received 2-dose vaccination. Among the 36 children who received 2 doses of vaccination, the time of viral nucleic acid negative conversion was shorter in those vaccinated within 6 months than those over 6 months ((7.1±1.9) vs. (10.8±3.0) d, t=-3.23, P=0.004). Univariate analysis showed that the time of nucleic acid negative conversion of SARS-CoV-2 was associated with age, underlying diseases, gastrointestinal symptoms, white blood cell count, proportion of neutrophils, proportion of lymphocytes, and the number of doses of SARS-CoV-2 vaccine (t=3.87, 2.55, 2.04, 4.24, 3.51, 2.92, F=16.27, all P<0.05). Multiple linear regression analysis showed that older age (ß=-0.33, 95% CI -0.485--0.182, P<0.001) and more doses of vaccination (ß=-0.79, 95% CI -1.463--0.120, P=0.021) were associated with shortened nucleic acid negative conversion time in children, while lower lymphocyte proportion (ß=-0.02, 95% CI -0.044--0.002, P=0.031) and underlying diseases (ß=1.52, 95% CI 0.363-2.672, P=0.010) were associated with prolonged nucleic acid negative conversion time in children. Conclusion: The children infected with Omicron variant of SARS-CoV-2 with reduced lymphocyte proportion and underlying diseases may have longer time of viral nucleic acid negative conversion,while children with older age and more doses of vaccination may have shorter time of viral nucleic acid negative conversion.

Comparison of hepatic epithelioid angiomyolipoma and non-hepatitis B, non-hepatitis C hepatocellular carcinoma on contrast-enhanced ultrasound.

PURPOSE: The purpose of this study was to retrospectively compare the imaging features of hepatic epithelioid angiomyolipoma (HEAML) to those of hepatocellular carcinoma negative for hepatitis B surface antigen and hepatitis C antibody (NBNC-HCC) on contrast-enhanced ultrasound (CEUS) with sulphur hexafluoride microbubbles. MATERIAL AND METHODS: Twenty-two patients (4 men, 18 women) with a mean age of 42.6±10.2 (SD) years (range: 22-63 years) with histopathologically confirmed HEMAL were included in the study. Forty-four patients (30 men, 14 women) with a mean age of 57.3±15.9 years (range: 19-85 years) with histopathologically confirmed NBNC-HCC were randomly selected from our institution's database as a control group. The CEUS characteristics of the two groups were compared. RESULTS: On conventional ultrasound, significant differences in tumor diameter were found between HEAML (4.0±2.0 [SD] cm; range: 1.3-8.9cm) and NBNC-HCC (8.4±4.4 [SD] cm; range: 1.6-18cm) (P<0.001) as well as in degrees of enhancement during the portal (P=0.001) and late phases (P=0.003), contrast distribution (P<0.001) and absence of pseudocaspule (P<0.001). On CEUS, hyperenhancement during the arterial phase was observed in 21/22 (95.5%) HEAMLs and in 43/44 (97.7%) NBNC-HCCs (P>0.999). Homogeneous enhancement was more frequent in HEAMLs (20/22; 90.9%) than in NBNC-HCCs (13/44; 29.6%) (P<0.001). Pseudocapsule was observed in 0/22 HEAMLs (0.0%) and in 36/44 NBNC-HCCs (81.8%) (P=0.017). A prolonged enhancement was observed in 5/22 HEAMLs (22.7%) and in 0/44 NBNC-HCCs (0.0%) (P<0.001) during the late phase. CONCLUSION: CEUS with sulphur hexafluoride microbubbles is helpful in discriminating between HEAML and NBNC-HCC. Homogeneous enhancement and lack of pseudocapsule are suggestive features for the diagnosis of HEAML.

Simultaneous determination of maltose and glucose using a screen-printed electrode system.

A screen-printed sensor system consisting of a glucose oxidase (GOD) electrode and an amyloglucosidase/glucose oxidase (A/G) electrode was constructed to determine maltose and glucose simultaneously in a mixture. Sensor construction was optimised so that it contained 20 units of GOD/40 units of amyloglucosidase and 0.2 mM 1,1'-ferrocenedimethanol. These components were deposited onto a screen-printed carbon electrode and an outer membrane was printed from 3.5% hydroxyethyl cellulose (HEC) solution. The optimum pH was 4.8. The linear range of the system was up to 40 mM glucose or 20 mmol/L maltose with coefficients of variation (CVs) ranging from 3.5% to 5.29%. The results obtained by using the enzyme electrode system agreed well with those obtained by the Fehling titration method. When stored dry, especially at 4 degrees C, the enzyme electrodes showed good stability over four months.

DNA optical sensor: a rapid method for the detection of DNA hybridization.

A DNA optical sensor system is proposed based on the combination of sandwich solution hybridization, magnetic bead capture, flow injection and chemiluminescence for rapid detection of DNA hybridization. Bacterial alkaline phosphatase (phoA) gene and Hepatitis B virus (HBV) DNA were used as target DNA. A biotinylated DNA probe was used to capture the target gene onto the streptavidin-coated magnetic beads and a calf intestine alkaline phosphatase (CAP)-labelled DNA probe was used for subsequent enzymatic chemiluminescence detection. The detection cycle was less than 30 min, excluding the DNA hybridization time, which was about 100 min. Both the phoA gene and HBV DNA could be detected at picogramme or femtomole level. No response signal was obtained when target DNA did not exist in the sample. Successive sample detection could be made by removing the magnetic field and a washing step.

Calcium antagonists in prevention of hypertension and stroke in stroke-prone spontaneously hypertensive rats.

To investigate the effect of Calcium antagonists (Ca-An) with different tissue specificity in the development of hypertension and stroke in salt-loading SHRSP, three experiments were conducted. In experiment I (1), 50 8-week-old male SHRSP were divided into three groups and given nifedipine (NF, 32 mg/kg/day), menidipine (MN 32 mg/kg/day) and placebo (control group) respectively. In the control group 83.3% (15/18) died of stroke and 17 showed renal vascular sclerosis. Their average lifespan was 84 days. NF and MN significantly reduced systolic blood pressure (SBP), and no stroke of renal vascular sclerosis developed. In experiment I (2), 54 7-week-old male SHRSP were divided into three groups (18 in each group). They were treated with nimodipine (NM) 20 mg/kg/day, 2 mg/kg/day and placebo respectively. NM (20 mg/kg/day) markedly lowered SBP and postponed the onset of stroke. Only 11% died in 17 weeks. NM (2 mg/kg/day did not lower SBP but postpond the onset of stroke. In experiment II (1), 29 10-week-old female SHRSP were divided into three groups: Group A was given NF 32 mg/kg/day, group B was parathyroidectomized (PTX) and group C served as control. PTX group did not lower SBP but could postpone the onset of stroke. In experiment II (2), 33 male SHRSP were divided into three groups and ticated as described above in experiment II (1) (11 in each group). Seven weeks after the experiment, the brain blood flow of NF group was significantly greater (67.5%) than that of the control and PTX groups. In experiment III, 27 7-week-old male SHRSP were divided into three groups as described above in experiment I (2).(ABSTRACT TRUNCATED AT 250 WORDS)

[Acute and chronic hypotensive effect of m-nifedipine].

Three groups of adult stroke--prone spontaneously hypertensive rats (SHRsp) were treated with m-nifedipine (m-Nif) 2.5, 5 and 10 mg.kg-1 by intraperitoneal injection. The systolic blood pressure (SBp) decreased significantly. The hypotensive effect persisted for 2-4, 6-8, and 12 h respectively and the peak effect appeared in 1 h. The range of hypotension was markedly greater than that of nifedipine (Nif). In SHPsp, treatment with m-Nif 10 or 30 mg.kg-1.d-1 for 4 weeks reduced significantly the SBp, but the hypotensive action is about the same as Nif.

Morphologies of fibroblast cells cultured on surfaces of PHB films implanted by hydroxyl ions.

Polyhydroxybutyrate (PHB) films were implanted with 40 keV hydroxyl ions with fluences ranging from 1 x 10(12) to 1 x 10(15) ions/cm2, respectively. The as-implanted PHB films were characterized by scanning electron microscopy (SEM), electron spectroscopy for chemical analysis (ESCA) and water contact angle measurements. The surface structures and properties of the as-implanted PHB films were closely related with hydroxyl ion fluence. They were further investigated by inoculating 3T6 fibroblasts cells on their surfaces. Morphologies of the 3T6 fibroblast cells cultured on surfaces of the as-implanted PHB films were observed by SEM. Characterization of the cultural 3T6 cells was analyzed qualitatively. The preliminary experimental results reveal that the bioactivity of the PHB films modified by hydroxyl ion implantation was improved at different levels, and the fluence of 1 x 10(13) ions/cm2 is optimal for PHB film.

Orthognathic surgery correction of mandibular hypoplasia accompanying obstructive sleep apnea syndrome.

OBJECTIVE: To examine the surgical correction methods for treating cases of severe mandibular hypoplasia accompanying obstructive sleep apnea syndrome (OSAS). METHODS: Sixteen cases of severe mandibular hypoplasia were studied in which OSAS was documented by polysomnography (PSG) and cephalometric study. The obstructive site was at the base of the tongue. Surgical procedures such as temporomandibular joint (TMJ) reconstruction and bimaxillary, chin, and hyoid bone advancement were performed to improve each patient's profile, function, and occlusion, and to treat the OSAS. RESULTS: There were great improvements in patient's sleep and daytime quality of life. The pre- and postoperative changes of most PSG values and some cephalometric values (SNB, PAS) were statistically significant. CONCLUSIONS: Severe mandibular hypoplasia can cause not only abnormalities in profile and occlusion but also OSAS. The evaluation of OSAS and its treatment effects depend on PSG. It is also very important to confirm the obstructive site in the upper airway by cephalometric study and fiberoptic endoscopy. Orthognathic surgery procedures can advance the maxillary, chin, and hyoid bone, and expand the upper airway simultaneously. These procedures can treat OSAS. Cases of TMJ ankylosis with OSAS should be treated step by step.

Anchor-chain molecular system for orientation control in enzyme immobilization.

An anchor-chain molecular system was constructed for controlled orientation and high activity in enzyme immobilization. A streptavidin recognition peptide (streptag) coding sequence was fused to the 3' end of the phoA gene, which codes for E. coli alkaline phosphatase (EAP). Both the wild-type (WT) and the Asp-101 --> Ser (D1O1S) mutant were modified with the streptag sequence with or without the insertion of a flexible linker peptide [-(Gly-Ser)(5)-] coding sequence. The fused genes were cloned into the vector pASK75 and expressed in the periplasm of the host cell Escherichia coli SM547. The proteins were released by osmotic shock and purified by ion-exchange chromatography. Enzyme activities of all proteins were measured spectrophotometrically with rho-nitrophenyl phosphate as the substrate. Specific activities of D101S-streptag and D101S-linker-streptag enzymes were increased 25- or 34-fold over the WT, respectively. These fusion proteins were then immobilized on microtiter plates through streptag-streptavidin binding reaction. After immobilization, the D101S-linker-streptag enzyme displayed the highest residual activity and the ratio of enzyme activities of the linker to nonlinker enzymes was 8.4. These results show that the addition of a linker peptide provides a spacer so as to minimize steric hindrance between the enzyme and streptavidin. The method provides a solution for controlled enzyme immobilization with high recover activity, which is especially important in construction of biosensors, biochips, or other biodevices.

[Cloning and expression of Aspergillus niger glucose oxidase gene in methylotrophic yeast].

The DNA fragment encoding A. niger glucose oxidase was amplified by PCR using A. niger genomic DNA as template, and was cloned into vector of pPIC9 for expression in Pichia pastoris. When transformed into methylotrophic yeast Pichia pastoris GS115, The constructed plasmid pPICGOD1 directed the synthesis and secretion of functionally active GOD. After induction in MM medium for 4 days, the GOD activity in the medium reached 30-40 u/mL. SDS-PAGE revealed that recombinant yeast GOD was expressed up to 60%-70% of the total soluble protein, and the secreted GOD could be purified to electrophoretic homogeneity with one purification step using Q Sepharose Fast Flow ion exchange chromatography. The recombinant yeast GOD had very high catalytic activity, showed about 1.6-fold increase of specific activity over the commercial A. niger GOD. Kinetic analysis clearly demonstrated that recombinant yeast GOD showed similar substrate affinity for glucose to A. niger GOD, but the turnover number of the GOD from yeast was determined to be much higher than that of A. niger GOD. In addition, the linear range of glucose electrode made with recombinant yeast GOD was efficiently widened due to the high catalytic activity of yeast GOD.

Role integration and perceived health in Asian American women caregivers.

In this study, role involvement, role integration (including role stress and role satisfaction), and perceived health were examined in 50 Asian American women who were caregivers of aging parents in addition to being wives, mothers, and employees. Their mean age was 47.8 years. Twenty-nine of the participants were Chinese and 21 Filipino, with an average length of caregiving for each group of 11 years. All participants were born outside the United States. Instruments used in the study were translated and back-translated into Chinese and Tagalog and tested for validity and reliability. The association of role involvement, role integration, role stress, and role satisfaction with perceived physical and psychological health in the combined and separate groups was examined. Role involvement was not associated with health in the combined group of caregivers but was associated with overall health in the sample of Chinese women. Role integration was positively associated with all three perceived health measures in the Filipino group but not in the Chinese group. Role satisfaction was consistently high in both groups. Role satisfaction and psychological well-being were significantly correlated for the combined group and for the Filipino caregivers. Total role stress was significantly correlated with overall health and current health only in the combined group. Thus, support that helps to decrease role stress and to increase role satisfaction may be more effective than efforts to decrease the extent of role involvement.

An adaptation of Brislin's translation model for cross-cultural research.

BACKGROUND: The number of nurse scientists pursuing cross-cultural research using quantitative designs is increasing. Preparation of instruments that are conceptually and functionally appropriate in the language of the participants is a complex process that needs examination. OBJECTIVES: Brislin's classic model for translation and validation of instruments for cross-cultural research is critiqued. Adaptations and extensions of that model are recommended. METHODS: Brislin's model guided tool preparation in a cross-cultural investigation. The process is described and lessons learned are outlined and discussed. CONCLUSIONS: Steps toward a more efficient and valid approach to the preparation of instruments are suggested.

Construction of a fusion enzyme system by gene splicing as a new molecular recognition element for a sequence biosensor.

A bifunctional fusion enzyme system constructed by gene splicing is proposed as a new model to develop sequence biosensors, taking maltose biosensor as an example. The cDNA fragment of Aspergillus niger glucoamylase (E.C 3.2.1.3, GA) was fused to the 3' end of Aspergillus niger glucose oxidase (E.C 1.1.3.4, GOD) gene with the insertion of a flexible linker peptide [-(Ser-Gly)5-] coding sequence. The fusion gene was cloned into the vector pPIC9 and expressed in Pichia pastoris GS115 under the control of the AOX1 promoter. It was found that a bifunctional hybrid protein with a molecular weight of 430 kDa was secreted after induction with methanol. The fusion enzyme GOD-(Ser-Gly)5-GA (GLG) was purified using Q Sepharose Fast Flow ion-exchange chromatography. Kinetic analysis demonstrated that GLG retained the typical kinetic properties of both GA and GOD. After being immobilized on an aminosilanized glass slide through covalent bonding by glutaraldehyde, GLG showed much higher sequential catalytic efficiency than the mixture of separately expressed GA and GOD (GA/GOD). Maltose biosensors were fabricated with GLG and GA/GOD, respectively. The performance characteristics of the maltose biosensor with respect to reproducibility, signal level, and linearity were effectively improved by using the fusion enzyme. Our findings offer a basis for the development of other sequence biosensors.