[In vivo confocal microscopy study of corneal endotheliitis].

OBJECTIVE: To study the cellular morphological characteristics and changes of corneal endotheliitis by corneal microstructure in vivo. METHODS: Forty-eight clinical diagnosed patients of corneal endotheliitis were examined by in vivo confocal microscopy. Confocal images of different layers were collected and observed. Of all the patients, 39 were disciform and 9 were diffuse. The history of the 48 patients was from 7 days to 23 years and the duration was from 4 to 180 days. RESULTS: Epithelium revealed cellular edema, enlarged intercellular gaps, and bubble between the cells. The density of sub-basal nerve plexus was significantly lower than that of normal, even disappeared in 24 patients. A numerous dendritic cells (Langerhans cells, LCs) presented in the basal epithelium layer and gradually abated with disease regression, but the density of LCs was significantly higher than that of the contralateral eye. The keratocyte revealed edema and to be activated. Inflammatory cell was found in stroma of the patients with long history and duration disease. Endothelium cells were observed edema, with enlarged intercellular gaps. Inflammatory cells was found to infiltrate into the endothelial layer, most them gathered to be keratic precipitates (KP), which were rounded or elliptic and inserted between the endothelium by pushing the endothelium away, and induced decayed area between the endothelium. CONCLUSION: Corneal endotheliitis was not only the inflammation of endothelium, but also varied pathological changes of all layer of the cornea. Corneal endotheliitis patients with long history and duration presented the mixed type with stromal keratitis. The lower density of sub-basal nerve plexus and the higher density of dendritic LCs were the characteristics of endotheliitis. Impairment of intercellular junction by KP might be another important role of cornea edema. Sufficient and accurate treatment of endotheliitis was important.

[Etiological analysis of ocular non-viral microbial isolates in children and their antibiotics susceptibility in vitro].

OBJECTIVE: To analyze the types, distribution characteristics and in vitro susceptibility of antibiotic agents of ocular non-viral microbial isolates in children. METHODS: Experimental research.785 specimens obtained from children below the age of 14 in Beijing Tongren Hospital during 1999 to 2004 were cultured and identified for bacterium, fungus and Acanthamoeba respectively at Beijing Institute of Ophthalmology. Microbial culture-positive rate, susceptibility patterns and the organisms distribution of positive cultures were retrospectively analyzed. The results were statistical analyzed by chi(2). RESULTS: Of the 785 specimens, 211 were culture positive. The percentage of positive culture was 26.88%. Of those positive cultures, 176 (83.41%) had pure bacterial infections, 32 (15.17%) had pure fungal infections, and 3 (1.42%) had pure cultures of Acanthamoeba. Of the 176 bacterial positive cultures, polymicrobial infection was present in 38 cases (33 bacterial, 5 fungal and bacterial). Of 214 bacterial isolates, the most common isolate was Neisseria gonorrhoeae (39/214, 18.22%), followed by Staphylococcus epidermidis (37/214, 17.29%), and Corynebacterium spp. (36/214, 16.82%). The most common fungal pathogen isolated was Fusarium spp. representing 25.00% (8/32) of all positive fungal cultures, followed by Aspergillus spp. (15.63%, 5/32). The sensitivity of tobramycin, norfloxacin, gentamicin, amikacin, ciprofloxacin, ofloxacin, rifampicin and levofloxacin of bacteria was 62.57%, 64.94%, 70.06%, 70.87%, 71.19%, 73.89%, 85.80% and 87.50%. Gram-positive isolates were susceptible to levofloxacin. Gram-negative isolates showed high susceptible to ciprofloxacin, ofloxacin and levofloxacin. The fungal isolates were most susceptible to natamycin, followed by terbinafine, but resistant to fluconazole. CONCLUSIONS: Bacteria are the most common pathogens in non-viral microbial eye infection in children. Neisseria gonorrhoeae, Staphylococcus epidermidis, and Corynebacterium spp. are the most common bacterial pathogens, which are highly susceptible to levofloxacin and rifampicin. Fusarium spp. and Aspergillus spp. are the most common fungal pathogens, which are highly susceptible to natamycin but resistant to fluconazole.

[Mutant prevention concentrations of antibacterial agents to ocular pathogenic bacteria].

OBJECTIVE: To establish a method to measure mutant prevention concentration (MPC) in vitro, and to measure MPC of antibacterial agents for ocular bacteria caused keratitis. METHODS: It was an experimental study. Forty strains of ocular bacteria were separated from cornea in Beijing Institute of Ophthalmology, which included 8 strains of Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pneumoniae, Pseudomonas aeruginosa and Klebsiella pneumoniae respectively. The minimal inhibitory concentration (MIC) of the levofloxacin (LVF), ofloxacin (OFL), ciprofloxacin (CIP), norfloxacin (NFL), tobramycin (TOB) and chloromycetin (CHL) were determined by agar dilution method from National Committee of Clinical Laboratory Standard (NCCLS). The MPC were measured by accumulate-bacterial methods with bacterial population inoculated more than 1.2 x 10(10) colony forming units per milliliter with Mueller-Hinton broth and tryptic soy agar plate. With the software of SPSS 11.0, the datum such as the range of MIC, MPC, MIC90 and MPC90 were calculated, and the selection index (MPC90/ MI90) and mutant selection window (MSW) were obtained. RESULTS: The MI90 of LVF and TOB (4 mg/L) to Staphylococcus aureus strains were the lowest. CIP showed the lowest MIC90 (0.25 mg/L) to Pseudomonas aeruginosa among six kinds of antibacterial agents. The MIC90 of LVF to Staphylococcus epidermidis (256 mg/L), Streptococcus pneumoniae (1 mg/L) and Klebsiella pneumoniae (0.25 mg/L) were lower than other antibacterial agents. The MPC90, MSW and the MPC90/MIC90 of levofloxacin showed lower values compared with other antibacterial medicines. From all the datum, the MIC90 of CHL was the highest and the activity was the weakest. Although the activity of LVF was higher to every kind of bacteria, CIP had the highest activity antibacterial to Pseudomonas aeruginosa. The capacity of CHL and TOB was weaker than Quinolones for restricting resistant mutants on ocular bacteria. LVF had the strongest capacity for restricting resistant mutants among Quinolones. CONCLUSION: LVF has better antibacterial effects and stronger capacity for restricting the selection of resistant mutants on ocular bacteria than other antibacterial agents.

[Operation of cornea grafting micro-surgery robot system].

OBJECTIVE: To design the robotic mechanism of the cornea grafting micro-surgery system and evaluate its experimental feasibility and efficacy. METHODS: It was an experimental study. Based on computer assisted cornea grafting surgery (CACGS), the prototype about cornea grafting robot, which consists of six subsystems, was developed. The system was used for looping incision and suturing for 20 rabbit corneas. Experimental data were studied including the error of target incision position, the actual cutting depth and the cutting edge span in order to evaluate the system's clinical value. All above statistical descriptions were performed using SPSS software version 11.5. RESULTS: 20 corneas were looping incised and sutured by the robot-assisted cornea grafting micro-surgery system successfully. The error of target incision position was (0.356 + or - 0.040) mm. With CACGS, the cutting edge span (A) was (0.855 + or - 0.040) and the actual cutting depth error was less than 10 microm. Corneal structure and thickness was showed no change and cutting edge of cornea was displayed regularity in pathological examination. To suturing system, suturing span was (2.15 + or - 0.09) mm and the error of span (0.15 + or - 0.02) mm. CONCLUSIONS: The cornea grafting micro-surgery system developed is used for cornea grafting of some animals successfully. The experiment results demonstrate the stability and efficiency of the robot.

[Analysis on susceptibility of ocular bacterial pathogens to four kinds of fluoroquinolone].

OBJECTIVE: To assess the in vitro susceptibilities of ocular bacterial isolates to Gatifloxacin and other fluoroquinolones which are often used in ophthalmic treatment. METHODS: Non-randomized, non-comparative, retrospective study. Ocular bacterial isolates (n = 619) from Beijing Institute of Ophthalmology were tested for their susceptibilities to gatifloxacin, ciprofloxacin, ofloxacin and levofloxacin in vitro by the Kirby-Bauer disk diffusion method. RESULTS: The total number of ocular bacterial isolates was 619 from January, 2005 to December, 2006 including gram-positive cocci 372(60.1%), gram-negative cocci 7 (1.1%), gram-positive bacilli 60 (9.7%), gram-negative bacilli 177 (28.6%), other 3 (0.5%). The percentage of susceptibility in total ocular bacterial isolates to gatifloxacin, ciprofloxacin, ofloxacin and levofloxacin was 88.4%, 72.1%, 64.8% and 54.4%. The susceptibility of Staphylococcus species to gatifloxacin (89.9%) was significantly higher than ciprofloxacin (38.8%), ofloxacin (48.2%) and levofloxacin (61.6%). Streptococcus species showed significantly higher susceptibility to gatifloxacin (93.2%), levofloxacin (89.2%) and ofloxacin (87.8%) than ciprofloxacin (62.2%). The susceptibility of gram-positive bacilli to gatifloxacin (86.7%) was significantly higher than ciprofloxacin (58.3%); All above differences are significant (P < 0.0083). Gram-negative cocci, enteric bacilli family and Pseudomonas species have the equivalent susceptibility to gatifloxacin, ciprofloxacin, ofloxacin and levofloxacin (P > 0.05). CONCLUSIONS: Most of ocular bacterial are susceptible to gatifloxacin in vitro, especially gram-positive bacteria including Staphylococcus species, Streptococcus species and gram-positive bacilli.

[Movement capture of knot-tying manipulation in microsurgeries].

The surgical dexterous suturing and knotting movements are photographed with binocular stereo vision and the movements' locus is captured through identifying the RGB values in a series of key-frames. Then a normal movement path of knot-tying is obtained through curve-fitting and smoothing. And meanwhile, the workspace of knot-tying manipulation in microsurgeries is got. And thus, a good foundation is laid for the design of an automatic knotting mechanism and for the establishment of a virtual surgery training system.

[Experimental study of non-tuberculous mycobacterial keratitis in rabbits].

OBJECTIVE: To study clinical manifestation and pathology change of different time points of rabbit keratitis caused by non-tuberculous mycobacteria (NTM). METHODS: Forty-eight eyes from 48 rabbits were randomly divided into three groups:under-flap infection group (UFI group), under-flap infection group with corticosteroid (UFIC group) and surface infection group (SI group). The cornea infiltration area of all rabbits was observed. Bacterial quantitative culture and histopathological examination were performed. RESULTS: In the earliest stage (5 days), the cornea was cloudy and edematous. In the middle stage (7 - 14 days), cornea stromal infiltration became its major clinical sign. In the last stage (21 days), corneal neovascularization and fibrosis were developed. The mean stromal infiltration area in UFIC group was greater than those of UFI group and SI group (F = 9.362, 8.341, P < 0.05). The number of NTM in UFIC group was higher than those of UFI group and SI group (F = 411.272, P < 0.05). In histopathological examination, there were infiltrates with polymorphonuclear leukocytes (PMN) in the corneal stroma in the early stage and acid-fast bacilli was found. A plenty of lymphocytes in the cornea was found in the middle stage of NTM keratitis. In the latter stage, the fibroblast cells proliferation and corneal ulcer were cured. During the course of this infection, the number of CD4+, CD8+ T lymphocyte increased significantly, and then decreased gradually following the regress of the infection. In the stage (5 - 21 days), CD4+ T lymphocyte decreased significantly in UFIC group compared with other two groups (F = 21.907, 196.521, 12.552, 11.100, P < 0.01). CONCLUSIONS: The clinical characteristic of rabbit keratitis is the multifocal dense superficial stromal infiltrates. CD4+ T lymphocyte may play an important role in NTM keratitis.

[Etiological analysis on bacterial endophthalmitis].

OBJECTIVE: To investigate the spectrum of organisms causing bacterial endophthalmitis and their sensitivities to commonly used ocular antibiotics. METHODS: A retrospective analysis on microbiological records of 728 intraocular specimens of patients with suspected endophthalmitis from January 1999 to December 2006. Antibiotic susceptibility of these isolates was determined for commonly used ocular antibiotics using the Kirby-Bauer disk-diffusion test. RESULTS: In all, 168 organisms were isolated from 728 intraocular specimens during the study interval. Overall, 96 (57.1%) of 168 isolates were Gram-positive cocci, 52 (31.0%) were Gram-negative bacilli, 18 (10.7%) were Gram-positive bacilli, and 2 (1.2%) were Gram-negative cocci. The most common organisms identified were Staphylococcus epidermidis in 21.4% (36/168), Staphylococcus aureus in 11.3% (19/168), and Pseudomonas aeruginosa in 8.9% (15/168). For Gram-positive cocci, sensitivities were the following: vancomycin 93.4% (85/91), levofloxacin 84.4% (81/96), ofloxacin 68.8% (66/96), ciprofloxacin 59.4% (57/96), gentamicin 56.3% (54/96), and tobramycin 43.8% (42/96). For Gram-negative bacilli, sensitivities were the following: levofloxacin 88.2% (45/51), ofloxacin 76.9% (40/52), ciprofloxacin 75.0% (39/52), gentamicin 49.0% (25/51), and tobramycin 48.1% (25/52). Comparing the data from 2003 to 2006 with the data from 1999 to 2002, the susceptibility of organisms to frequently employed antibiotics was decreasing. CONCLUSIONS: The spectrum of organisms causing bacterial endophthalmitis is complex, and no single antibiotic provided coverage for all of the microbes isolated from intraocular specimens with bacterial endophthalmitis. Combination therapy is recommended as the initial empiric treatment of suspected bacterial endophthalmitis.

[Molecular characteristics of Acanthamoeba strains from keratitis patients].

OBJECTIVE: To study the molecular characteristics of Acanthamoeba strains isolated from keratitis patients. METHODS: Twenty six Acanthamoeba strains were isolated from corneal ulcers between 1994 and 2002. Partial genome DNA sequences of all strains of Acanthamoeba were amplified using PCR and then the PCR products were sequenced. The partial DNA sequences were analyzed by software Clustal X and MEGA2. The results of Acanthamoeba obtained from Beijing Tongren Hospital were compared with strains from other countries and districts. RESULTS: Of 26 Acanthamoeba strains isolated from the patients, 25 (96%) were 18S rDNA (Rns) genotype T4 and one (4%) was T3. These results were similar to those obtained from other countries and districts. CONCLUSIONS: According to the Rns genotype classification criterion, most of Acanthamoeba strains isolated from keratitis patients in Beijing are genotype T4 and only few of these strains are genotype T3. The Acanthamoeba infecting the cornea might belong to the specific 18S rDNA genotype.