[The progression of applying transgenic mice as an animal model of high myopia].

Gene engineering technology provides new methods for medical research. The application of transgenic animals and knock out animals makes it possible to study the biological characteristics of genes associated with some human diseases in vivo. Transgenic mice were first achieved in the early 1980s, while they were used extensively in the field of ophthalmology doing some research a few years later after that. Therefore, it can possibly be a new path to investigate the pathogenesis by establishing transgenic mouse model of candidate gene of high myopia.

Comparison of total corneal power measurements obtained with different devices after myopic keratorefractive surgery.

AIM: To analyze the differences, agreements, and correlation among total corneal power parameters generated by different instruments after myopic keratorefractive surgery. METHODS: The prospective cross-sectional study included patients who underwent myopic keratorefractive surgery and received measurements of corneal power 3mo after surgery. Automated keratometer was used for the measurement of simulated keratometry (SimK), swept-source optical coherence tomography (SS-OCT) based biometer for total keratometry (TK), anterior segment-OCT for real keratometry (RK), and Scheimpflug keratometer for the true net power (TNP), the total corneal refractive power (TCRP) and equivalent K-readings (EKR). The differences among these parameters were analyzed, and the agreements and correlation between SimK and other total corneal power parameters were investigated. RESULTS: A total of 70 eyes of 70 patients after myopic keratorefractive surgery were included. The evaluated corneal power parameters were as follows: SimK 38.32±1.93 D, TK 37.54±2.12 D, RK 36.64±2.09 D, TNP 36.56±1.97 D, TCRP 36.70±2.01 D, and EKR 37.55±2.00 D. Pairwise comparison showed that there were significant differences (P<0.001) among all parameters except for between TK and EKR, RK and TNP, RK and TCRP (P=1.000, 1.000, 1.000, respectively). The limits of agreement between SimK and TK, RK, TNP, TCPR, and EKR were 1.08, 1.08, 1.43, 1.48, and 1.73 D, respectively. All parameters showed good correlation with SimK, and the correlation coefficients were 0.995, 0.994, 0.983, 0.982, and 0.975. CONCLUSION: Among the corneal power parameters after myopic keratorefractive surgery, the value of SimK is the largest, followed by TK and EKR, with TCRP, RK, and TNP being the smallest. The differences among the parameters may be attributable to the different calculation principles. Correct understanding and evaluation of corneal power parameters can provide a theoretical basis for taking advantage of the total corneal power to improve the accuracy of intraocular lens calculation after keratorefractive surgery.

Complete genome sequence of a novel clinical isolate, the nontuberculous Mycobacterium strain JDM601.

Mycobacteriosis is on the increase. Nontuberculous mycobacteria (NTM) are resistant to most antituberculosis drugs naturally. We determined the complete genome sequence of a novel NTM strain, JDM601, of the Mycobacterium terrae complex, which was isolated from a patient with tuberculosis-like disease and with various antibiotic resistances.

Evaluation of a novel fusion protein antigen for rapid serodiagnosis of tuberculosis.

The detection of Mycobacterium tuberculosis-specific antibodies in human sera has been a rapid and important diagnostic aid for tuberculosis (TB) control and prevention. However, any single antigen is not enough to be used to cover the antibody profiles of all TB patients. In this study, a novel fusion protein was constructed using gene splicing by overlap extension (SOEing), and then the antibody level against it in 171 TB patients and 86 controls was evaluated by enzyme-linked immunosorbent assay. Compared with the three individual antigen (16 kDa: sensitivity 19.9%, specificity 96.5%; MPT64: sensitivity 75.4%, specificity 34.9%; 38 kDa: sensitivity 33.3%, specificity 83.7%), the fusion protein antigen (sensitivity 42.1%, specificity 89.5%) gave the best diagnostic performance with the largest receiver operating characteristic curve area 0.656 (95% confidence interval [CI], 0.590-0.721; P<0.01). These results suggested that the novel fusion protein antigen successfully constructed by gene SOEing provided the improved diagnostic performance for TB, and other mycobacterial multiepitope fusion proteins may also be worthy of investigation for further enhancing the detection sensitivity.

Stability of the Barrett True-K formula for intraocular lens power calculation after SMILE in Chinese myopic eyes.

AIM: To compare the Barrett True-K formula with other formulas integrated in Lenstar 900 to predict intraocular lens (IOL) power after small-incision lenticule extraction (SMILE). METHODS: A theoretical prospective study was performed to predict the ratio of equivalent IOL power before and after SMILE using the SRK/T (Sanders, Retzlaff, Kraff/theoretical), Holladay 1, Haigis, and Barrett True-K formulas and compare the stability of their predictions. The study included 54 eyes (54 cases) with a manifest refraction spherical equivalent (MRSE) of -4.99±1.45 D. They were divided into two groups: 27 eyes with axial length of 24-26 mm in Group A, and 27 eyes with axial length >26 mm in Group B. All subjects enrolled in this study were examined with the Lenstar 900 before and 6mo after SMILE including measurements of axial length, corneal curvature, and anterior chamber depth (ACD). RESULTS: The prediction of equivalent IOL power of the two groups was more stable for the Barrett True-K formula, especially in long axial length eyes (Group B). The respective percentages for the SRK/T, Holladay 1, Haigis, and Barrett True-K formulas were 7.4%, 7.4%, 85.19%, and 88.89% for a margin of error within 0.5 D; 25.92%, 51.84%, 100%, and 100% for a margin of error within 1.0 D in Group A; 33.33%, 40.74%, 44.44%, and 81.48% for a margin of error within 0.5 D; and 44.44%, 59.26%, 66.66%, and 92.59% for a margin of error within 1.0 D in Group B. The respective percentages for Barrett True-K formulas were 100% for a margin of error within 2.0 D in Group B. CONCLUSION: Theoretically, the Barrett True-K formula provides more stable predictions than other formulas for cataract eyes after SMILE.

A Rapid and Accurate Detection Approach for Multidrug-Resistant Tuberculosis Based on PCR-ELISA Microplate Hybridization Assay.

Rapid and accurate diagnosis of multidrug-resistant tuberculosis (MDR-TB) is important for timely and appropriate therapy. In this study, a rapid and easy-to-perform molecular test that integrated polymerase chain reaction (PCR) amplification and a specific 96-well microplate hybridization assay, called PCR-ELISA (enzyme-linked immunosorbent assay), were developed for detection of mutations in rpoB, katG, and inhA genes responsible for rifampin (RIF) and isoniazid (INH) resistance and prediction of drug susceptibility in Mycobacterium tuberculosis clinical isolates. We evaluated the utility of this method by using 32 multidrug-resistent (MDR) isolates and 22 susceptible isolates; subsequently, we compared the results with data obtained by conventional drug susceptibility testing and DNA sequencing. The sensitivity and specificity of the PCR-ELISA test were 93.7% and 100% for detecting RIF resistance, and 87.5% and 100% for detecting INH resistance, respectively. These results were comparable to those yielded by commercially available molecular tests such as the GenoType MTBDRplus assay. Based on the aforementioned results, we conclude that the PCR-ELISA microplate hybridization assay is a rapid, inexpensive, convenient, and reliable test that will be useful for rapid diagnosis of MDR-TB, for improved clinical care.

Pharmacologic ascorbate as a pro-drug for hydrogen peroxide release to kill mycobacteria.

BACKGROUND AND PURPOSE: Tuberculosis is one of the most highly fatal diseases worldwide, and one-third of the world's population has been infected with Mycobacterium tuberculosis (M. tuberculosis). A previous study showed that M. tuberculosis was highly susceptible to being killed by ascorbate (i.e. vitamin C, VC), but the molecular mechanisms of the bactericidal activity of VC against M. tuberculosis are still not well understood. EXPERIMENTAL APPROACH: We assayed the effects of VC as an anti-tuberculosis drug against mycobacteria (i.e. M. bovis BCG or M. tuberculosis H37Rv) in macrophages (i.e. RAW 264.7 cells). Relative global protein expression changes in 5 mM VC-treated and control samples of H37Rv were investigated by Tandem mass tag (TMT)-based quantitative proteomic analysis. qRT-PCR was also used to measure the differential expression of six intracellular stress response mycobacteria genes. KEY RESULTS: Quantitative proteomic analysis showed that 11 peptide components including rip3, fdxA, Rv2028c, mtp, LH57_00670, hspX, pfkB, Rv1824, Rv1813c, LH57_08410 and Rv2030c were up-regulated and 17 peptide components were down-regulated in 5 mM VC-treated H37Rv compared with the control samples. qRT-PCR also verified that VC could induce the expression of six genes (hsp, fdxD, furA, devR, hspX, and dnaB) in BCG and H37Rv. We also found that exosomes from RAW 264.7 cells treated with pharmacologic VC could kill M. bovis BCG in vitro. CONCLUSION AND IMPLICATIONS: Our results demonstrated that the bactericidal activity of VC against mycobacteria, as a pro-drug for hydrogen peroxide formation (H2O2), was dependent on reactive oxygen species production and the activated oxidative stress pathway, which suggested that pharmaceutical VC and exosomes from macrophages treated with VC could be used as potential anti-tuberculosis drugs.

Consistent comparison of angle Kappa adjustment between Oculyzer and Topolyzer Vario topography guided LASIK for myopia by EX500 excimer laser.

AIM: To evaluate and compare the uniformity of angle Kappa adjustment between Oculyzer and Topolyzer Vario topography guided ablation of laser in situ keratomileusis (LASIK) by EX500 excimer laser for myopia. METHODS: Totally 145 cases (290 consecutive eyes )with myopia received LASIK with a target of emmetropia. The ablation for 86 cases (172 eyes) was guided manually based on Oculyzer topography (study group), while the ablation for 59 cases (118 eyes) was guided automatically by Topolyzer Vario topography (control group). Measurement of adjustment values included data respectively in horizontal and vertical direction of cornea. RESULTS: Horizontally, synclastic adjustment between manually actual values (dxmanu) and Oculyzer topography guided data (dxocu) accounts 35.5% in study group, with mean dxmanu/dxocu of 0.78±0.48; while in control group, synclastic adjustment between automatically actual values (dxauto) and Oculyzer topography data (dxocu) accounts 54.2%, with mean dxauto/dxocu of 0.79±0.66. Vertically, synclastic adjustment between dymanu and dyocu accounts 55.2% in study group, with mean dymanu/dyocu of 0.61±0.42; while in control group, synclastic adjustment between dyauto and dyocu accounts 66.1%, with mean dyauto/dyocu of 0.66±0.65. There was no statistically significant difference in ratio of actual values/Oculyzer topography guided data in horizontal and vertical direction between two groups (P=0.951, 0.621). CONCLUSION: There is high consistency in angle Kappa adjustment guided manually by Oculyzer and guided automatically by Topolyzer Vario topography during corneal refractive surgery by WaveLight EX500 excimer laser.

Identification and characterization of non-tuberculous mycobacteria isolated from tuberculosis suspects in Southern-central China.

The incidence of non-tuberculous mycobacteria (NTM)-related death has increased globally recently. To obtain information of the species and characterization of pathogens involved in NTM pulmonary infection in Southern-central China, we identified 160 non-tuberculous infection cases from 3995 acid-fast bacilli (AFB)-positive tuberculous suspects. We then randomly selected 101 non-tuberculous patients, isolated bacteria from their sputa and genotyped the pathogens using the 16S rRNA gene and 16S-23S rRNA internal transcribed spacer sequences. M. intracellulare (32.67%, 33/101), M. abscessus (32.67%, 33/101) and M. fortuitum (7.92%, 8/101) are identified in these isolates. Surprisingly, non-mycobacteria including Gordonia (8.91%, 9/101), Nocardia (5.94%, 6/101) and Tsukamurella (0.99%, 1/101) are also discovered, and the case of Tsukamurella pulmonis infection is first discovered in Southern-central China. Moreover, species of M. mucogenicum group, M. chubuense, M. kansasii, M. gastri, M. avium, M. porcinum and M. smegmatis are identified. In addition, nine immune compromised cases (8.91%, 9/101), including type two diabetes mellitus and HIV/AIDS are found to be infected with non-tuberculous bacteria. This study revealed the distribution and characteristics of non-tuberculous AFB pathogen infection occurred in Southern-central China, and suggested that physicians should be alert of the emerging of NTM and non-mycobacteria infection in AFB positive cases and take caution when choosing chemotherapy for tuberculosis-like pulmonary infections. Generally, this study may help with the development of new strategy for the diagnosis and treatment of mycobacterial infection.

Increased case finding of tuberculosis from sputum and sputum deposits after magnetic bead concentration of mycobacteria.

Concentration of mycobacteria from sputum by centrifugation prior to acid-fast microscopy increases case finding compared to direct microscopy of the sputum (direct smear). However, centrifugation has to be performed outside the safety cabinet and many laboratories do not have access to a centrifuge. Magnetic bead extraction of the mycobacteria is an alternative method that can be performed in a cabinet with just a magnet. Magnetic TB-Bead (Microsens Medtech Ltd) extraction of mycobacteria from sputum prior to microscopy was compared to direct smear on 78 sputum samples. Microscopy of the TB-Bead extracts identified all of 26 of the direct smear positive samples either with the same microscopy score or, in 19/27 of samples, with an increased microscopy score which aided microscopy detection. In addition, microscopy of the TB-Bead extracts identified 10 additional positive samples compared to direct smear; which represents a statistically significant increase in case finding of 38% (p = 0.002) compared to direct smear. In a separate study, TB-Beads enabled further 4 positive samples to be detected from 30 centrifuged pellets that were originally smear negative; two of these were subsequently found to be positive when the original deposits were reinvestigated by smear microscopy. By concentrating mycobacteria from sputum and sputum deposits, TB-Beads have been demonstrated to increase the number of positive sputum samples which could increase case-finding. The TB-Bead method is simple and rapid and compatible with use within a safety cabinet.