David V. Seal

Prophylaxis of postoperative endophthalmitis following cataract surgery: Results of the ESCRS multicenter study and identification of risk factors

PURPOSE: To identify risk factors and describe the effects of antibiotic prophylaxis on the incidence of postoperative endophthalmitis after cataract surgery based on analysis of the findings of the European Society of Cataract & Refractive Surgeons (ESCRS) multicenter study. SETTING: Twenty‐four ophthalmology units in Austria, Belgium, Germany, Italy, Poland, Portugal, Spain, Turkey, and the United Kingdom. METHODS: A prospective randomized partially masked multicenter cataract surgery study recruited 16 603 patients. The study was based on a 2 × 2 factorial design, with intracameral cefuroxime and topical perioperative levofloxacin factors resulting in 4 treatment groups. The comparison of case and non‐case data was performed using multivariable logistic regression analyses. Odds ratios (ORs) associated with treatment effects and other risk factors were estimated. RESULTS: Twenty‐nine patients presented with endophthalmitis, of whom 20 were classified as having proven infective endophthalmitis. The absence of an intracameral cefuroxime prophylactic regimen at 1 mg in 0.1 mL normal saline was associated with a 4.92‐fold increase (95% confidence interval [CI], 1.87‐12.9) in the risk for total postoperative endophthalmitis. In addition, the use of clear corneal incisions (CCIs) compared to scleral tunnels was associated with a 5.88‐fold increase (95% CI, 1.34‐25.9) in risk and the use of silicone intraocular lens (IOL) optic material compared to acrylic with a 3.13‐fold increase (95% CI, 1.47‐6.67). The presence of surgical complications increased the risk for total endophthalmitis 4.95‐fold (95% CI, 1.68‐14.6), and more experienced surgeons were more likely to be associated with endophthalmitis cases. When considering only proven infective endophthalmitis cases, the absence of cefuroxime and the use of silicone IOL optic material were significantly associated with an increased risk, and there was evidence that men were more predisposed to infection (OR, 2.70; 95% CI, 1.07‐6.8). CONCLUSIONS: Use of intracameral cefuroxime at the end of surgery reduced the occurrence of postoperative endophthalmitis. Additional risk factors associated with endophthalmitis after cataract surgery included CCIs and the use of silicone IOLs.

Use of Subgenic 18S Ribosomal DNA PCR and Sequencing for Genus and Genotype Identification of Acanthamoebae from Humans with Keratitis and from Sewage Sludge

ABSTRACT This study identified subgenic PCR amplimers from 18S rDNA that were (i) highly specific for the genus Acanthamoeba, (ii) obtainable from all known genotypes, and (iii) useful for identification of individual genotypes. A 423- to 551-bpAcanthamoeba-specific amplimer ASA.S1 obtained with primers JDP1 and JDP2 was the most reliable for purposes i and ii. A variable region within this amplimer also identified genotype clusters, but purpose iii was best achieved with sequencing of the genotype-specific amplimer GTSA.B1. Because this amplimer could be obtained from any eukaryote, axenic Acanthamoeba cultures were required for its study. GTSA.B1, produced with primers CRN5 and 1137, extended between reference bp 1 and 1475. Genotypic identification relied on three segments: bp 178 to 355, 705 to 926, and 1175 to 1379. ASA.S1 was obtained from single amoeba, from cultures of all known 18S rDNA genotypes, and from corneal scrapings of Scottish patients with suspected Acanthamoeba keratitis (AK). The AK PCR findings were consistent with culture results for 11 of 15 culture-positive specimens and detected Acanthamoeba in one of nine culture-negative specimens. ASA.S1 sequences were examined for 6 of the 11 culture-positive isolates and were most closely associated with genotypic cluster T3-T4-T11. A similar distance analysis using GTSA.B1 sequences identified nine South African AK-associated isolates as genotype T4 and three isolates from sewage sludge as genotype T5. Our results demonstrate the usefulness of 18S ribosomal DNA PCR amplimers ASA.S1 and GTSA.B1 for Acanthamoeba-specific detection and reliable genotyping, respectively, and provide further evidence that T4 is the predominant genotype in AK.

ESCRS study of prophylaxis of postoperative endophthalmitis after cataract surgery: Preliminary report of principal results from a European multicenter study.

PURPOSE: To report results in the European Society of Cataract & Refractive Surgeons (ESCRS) multicenter study of the prophylaxis of endophthalmitis after cataract surgery. SETTING: Twenty‐four ophthalmology units and eye clinics in Austria, Belgium, Germany, Italy, Poland, Portugal, Spain, Turkey, and the United Kingdom, with an administrative office in Ireland, coordinating center in England, and data management and statistical unit in Scotland. METHODS: This partially masked randomized placebo‐controlled multinational clinical study to evaluate prospectively the prophylactic effect of intracameral cefuroxime injection and/or perioperative levofloxacin eyedrops on the incidence of endophthalmitis after phacoemulsification cataract surgery began in September 2003 and was terminated early in January 2006. The study used random allocation of patients in a 2 × 2 factorial design. RESULTS: By the end of 2005, complete follow‐up records had been received for 13 698 study patients. Such a clear beneficial effect from the use of intracameral cefuroxime had been observed that it was agreed it would be unethical to continue the study and to wait for the completion of all follow‐up procedures before reporting this important result. If total reported cases of endophthalmitis are considered, the incidence rate observed in those treatment groups not receiving cefuroxime prophylaxis (23 cases in 6862 patients) was almost 5 times as high (odds ratio [OR], 4.59; 95% confidence interval [CI], 1.74‐12.08; P = .002) as that in the groups receiving this treatment (5 cases in 6836 patients). If only cases proved to be due to infection are considered, the rate was more than 5 times as high (OR, 5.32; 95% CI, 1.55‐18.26; P = .008) in the treatment groups not receiving cefuroxime. Although the use of perioperative levofloxacin eyedrops as prophylaxis was also associated with a reduction in the observed incidence rate of postoperative endophthalmitis, this effect was smaller and was not statistically significant, whether total reported cases or only cases proven to be due to infection are used in calculating the rates. As not all follow‐up procedures are complete, it is possible that further cases of endophthalmitis may be reported; however, it is not expected that this will alter the main conclusion. Nevertheless, it is anticipated that successful completion of follow‐up procedures in all patients will increase the total number in the study to approximately 16 000. CONCLUSION: Intracameral cefuroxime administered at the time of surgery significantly reduced the risk for developing endophthalmitis after cataract surgery.

ESCRS study of prophylaxis of postoperative endophthalmitis after cataract surgery: Case for a European multicenter study.

PURPOSE: To present the development and design of the European Society of Cataract & Refractive Surgeons multicenter study of the prevention of postsurgical infective endophthalmitis after phacoemulsification and to describe the process for its successful implementation and conduct. SETTING: Twenty‐four ophthalmology units and eye clinics in Austria, Belgium, Germany, Italy, Poland, Portugal, Spain, Turkey, and the United Kingdom, with an administrative office in Ireland, coordinating center in England, and data management and statistical unit in Scotland. METHODS: This partially masked randomized placebo‐controlled multinational clinical study was designed to evaluate prospectively the prophylactic effect of intracameral cefuroxime and/or perioperative topical levofloxacin on postoperative endophthalmitis after cataract surgery. Random allocation was based on a 2 × 2 factorial design that included participating centers as a class variable. Real‐time electronic data collection monitored study progress and provided weekly outcome tables, monthly recruitment summaries, and quarterly analytical reports for the studys Data Monitoring Committee, which evaluated the safety and efficacy by Internet‐based conferences. RESULTS: A 2‐year lead time was required to meet harmonized standards of clinical research in the European Union, obtain ministerial authorization in 3 countries, gain institutional approvals at 24 hospitals, and procure indemnity insurance for surgical centers. Informed consent instruments, designed to comply with national health policies, were translated into 8 languages. The use of information technology to collect study data enabled the organizers to evaluate individual eligibility at enrollment, adherence with study medications during and after surgery, and postoperative status during follow‐up. CONCLUSION: This international cooperative study provided the opportunity to estimate the current incidence of endophthalmitis after cataract surgery in Europe and determine whether 1 or both of 2 antimicrobial regimens reduces the risk for postsurgical intraocular infection.

Drug resistance and Acanthamoeba keratitis: the quest for alternative antiprotozoal chemotherapy.

Trophozoites and cysts of 20 isolates of Acanthamoeba from the cornea and five from related samples were tested in vitro for sensitivity to ten drugs (three aromatic dia-midines, two aminoglycosides, two macrolides, a polyene macrolide antibiotic, an organoarsenical and an antimetabolite) and two cationic antiseptics (chlorhexidine and polyhexamethylene biguanide, PHMB). Only chlorhexidine and PHMB showed uniform amoebacidal activity. Aromatic diamidines (pentamidine isethionate, propamidine isethionate and diminazene aceturate) generally proved effective against both forms of the amoeba; only pentamidine gave synergy with the biguanide while propamidine gave an additive effect. Other drugs tested proved erratic or ineffective against different isolates. Chlorhexidine alone, or together with propamidine, was subsequently used in two patients with proven Acanthamoeba keratitis; the causative isolates were sensitive to the individual compounds and to the combination in vitro. The treatment provided resolution of the clinical disease; amoebae were shown to be non-viable by histology and culture. The combination of chlorhexidine and propamidine is recommended for treatment of proven Acanthamoeba keratitis.

Amebic Keratitis in a Wearer of Disposable Contact Lenses due to a Mixed Vahlkampfia and Hartmannella Infection

PURPOSE To support the hypothesis that Acanthamoeba is not a unique cause of amebic keratitis, we report a case of amebic keratitis in which viable Acanthamoeba could not be isolated from corneal tissue. Vahlkampfia and Hartmannella, two other genera of free-living ameba, were isolated, however, using prolonged culture. METHODS A 24-year-old wearer of soft contact lenses had keratitis. Extensive histologic and microbiologic investigations were performed on corneal scrape, biopsy, and keratoplasty tissue. Contact lenses, storage case, and the home water supply, where contact lens hygiene was practiced, were examined for the presence of micro-organisms. RESULTS No viruses, pathogenic bacteria, or fungi were detected from corneal tissue samples. Amebae were observed using light and electron microscopy, but these could not be unequivocally classified using immunocytochemical staining. Viable Vahlkampfia and Hartmannella, but no Acanthamoeba, were isolated from the corneal biopsy sample. Indirect immunofluorescence with a range of polyclonal rabbit antisera raised against axenically cultivated stains of the three amebal genera was unhelpful because of cross-reactivity. A diverse range of micro-organisms was present within the storage case, including the three amebal species. Amebic cysts also were associated with the contact lens. CONCLUSION A mixed non-Acanthamoeba amebic keratitis has been identified in a wearer of soft contact lenses where lack of storage case hygiene provided the opportunity for the free-living protozoa Vahlkampfia and Hartmannella to be introduced to the ocular surface. When Acanthamoeba-like keratitis occurs, but where Acanthamoeba cannot be isolated using conventional laboratory culture methods, alternate means should be used to identify other amebae that may be present. Polyclonal immunofluorescent antibody staining was unreliable for generic identification of pathogenic free-living amebae in corneal tissue.

Determination of Amoebicidal Activities of Multipurpose Contact Lens Solutions by Using a Most Probable Number Enumeration Technique

ABSTRACT Six multipurpose contact lens solutions [All-in-One, All-in-One (Light), ReNu MultiPlus, Optifree Express, Complete, and Solo-care soft] were tested for their efficacies against Acanthamoeba castellanii trophozoites and cysts by using a most probable number (MPN) technique for amoebic enumeration. Against trophozoites, All-in-One, ReNu Multiplus, and Optifree Express achieved total kill (log reduction of >3) after the manufacturers minimum recommended disinfection time (MMRDT), with the remaining solutions failing to reach a log reduction of 1. After 24 h of exposure, all solutions proved trophozoiticidal, achieving, with the exception of Complete (log reduction of 3.13), total kill. Against cysts, All-in-One gave a log reduction of >3 within the MMRDT, with all other solutions failing to achieve a log reduction of 1. After 24 h of exposure, All-in-One achieved total kill of cysts (log reduction of 3.74), ReNu MultiPlus gave a log reduction of 3.15, and the remaining solutions reached log reductions of between 1.09 and 2.27. The MPN technique provides a simple, reliable, and reproducible method of amoebic enumeration that depends on simply establishing the presence or absence of growth on culture plates inoculated with a series of dilutions and determining the MPN of amoebae present from statistical tables. By use of this technique, two of the multipurpose solutions tested, ReNu MultiPlus and Optifree Express, demonstrated effective trophozoiticidal activities within the recommended disinfection times; however, only All-in-One proved effective against both trophozoites and cysts over the same time period. This MPN technique, which uses axenically produced trophozoites and mature, double-walled cysts, has the potential to form the basis of a national standard for amoebicidal efficacy testing of multipurpose contact lens disinfecting solutions.

Epidemiology of diarrhoea caused by enterotoxigenic Clostridium perfringens

Enterotoxigenic strains of Clostridium perfringens have recently been implicated in some cases of antibiotic-associated diarrhoea. We present here the results of an epidemiological study of this disease. Five cases of diarrhoea caused by C. perfringens serotype 41 occurred during a 9-week period, and then during a 6-week period there were three cases due to serotype 27 and two due to serotype 24; in all but one case two geriatric wards were involved. In total there were 16 cases in 22 months. All cases were identified by the detection of C. perfringens enterotoxin in the faeces. The mean number of C. perfringens in these cases was 10(8.8) cfu/g of faeces. Of 37 patients who had negative test results for C. perfringens enterotoxin, 18 had positive cultures for C. perfringens, with mean faecal counts of 10(5.3) cfu/g, and nine of these patients had diarrhoea. Thirteen different serotypes were isolated from these 18 patients, including type 41 from seven patients and type 27 from one. Hand carriage of the offending serotype was demonstrated in three of four infected patients, none of four controls and two of 14 ward staff. C. perfringens of serotypes causing disease was isolated from 59% of environmental areas where there was active disease, 27% of areas where there had been disease which had since resolved and 9% of areas where there was no history of disease. The findings imply that cross infection may occur.

Laboratory diagnosis of endophthalmitis: Comparison of microbiology and molecular methods in the European Society of Cataract & Refractive Surgeons multicenter study and susceptibility testing

PURPOSE: To investigate and compare the use of molecular biology with the use of traditional Gram stain and organism culture for the laboratory diagnosis of postoperative endophthalmitis. SETTING: Twenty‐four ophthalmology units together with 9 microbiology laboratories and 2 European reference molecular biology laboratories. METHODS: A prospective randomized partially masked multicenter cataract surgery study recruited 16 603 patients. This resulted in 29 cases of presumed postoperative endophthalmitis. Gram stain and culture were performed in the local laboratory according to agreed protocols. Samples of aqueous and/or vitreous were transported to the first referenced molecular biology laboratory (Regensburg, Germany) for polymerase chain reaction (PCR) testing, and an extracted aliquot of DNA was then referred to the second laboratory (Alicante, Spain) for PCR. RESULTS: Of the 29 who presented with presumed postoperative endophthalmitis, 20 were classified as proven infective endophthalmitis with positive Gram stain, culture, or PCR. Fourteen patients were culture‐positive; all but 1 of these was also positive by PCR. Six patients were positive by PCR but negative by Gram stain or culture. Nine patients were negative by both microbiology and PCR testing. CONCLUSIONS: Use of molecular biology technique increased the laboratory rate of identifying the pathogen by 20%, confirming the technique is very useful for the endophthalmitis specimen. Samples of both aqueous and vitreous should be collected and stored at −20°C for PCR at the time of the diagnostic taps.

Role of Staphylococcus aureus in Chronic Allergic Conjunctivitis

A study was undertaken to test the hypothesis that Staphylococcus aureus colonization of the lid margins could determine the expression of allergic eye disease in atopic patients. The authors compared lid isolates of S. aureus from 23 adults who had both atopic dermatitis and chronic conjunctivitis and isolates from 14 patients who had atopic dermatitis but who lacked ocular disease. No significant difference was found in either the staphylococcal protein A or hemolytic toxin production by isolates from the two disease groups, and there was no difference between groups in the quantity of serum IgG nor IgE antibodies to staphylococcal ribitol-teichoic acid. In seven patients with chronic allergic conjunctivitis who were challenged with staphylococcal protein A or heat-killed S. aureus, delayed-type hypersensitivity was not enhanced. These results suggest that although S. aureus colonization of the lids is common in atopic patients, neither the pattern of toxin production nor humoral or cell-mediated immunity to S. aureus play a role in the expression of chronic allergic conjunctivitis.