[Performance and cost calculation for a university ophthalmological outpatient clinic]. / Leistungs- und Kostenkalkulation für eine universitäre, augenheilkundliche Hochschulambulanz.

BACKGROUND: Outpatient procedures at a university hospital are generally considered to be unprofitable. In the present publication we evaluate the turnover and costs of the university eye outpatient department of the Hannover Medical School (MHH) in terms of a cost unit accounting as well as providing a summary of the workload. MATERIAL AND METHOD: Given the data of the hospital information system (IS-H/i.s.h.med from SAP) and a proprietary software (TimeElement), all patient contacts in the year 2019 were evaluated. The latter software is applied in a standardized manner to record the patient flow of our outpatient service in real time electronically. The total costs consist of personnel, material and room costs including infrastructure of the MHH and are compared to the flat-rate revenues according to the university outpatient contract (HSA) as well as further revenues from internal referral services, self-pay patients, outpatient surgery and cooperation contracts for intravitreal injections (IVOM). RESULTS: With an average full-time equivalent (FTE) headcount of 10.63 assistant physicians, 3.6 specialist physicians, and 21 nonphysicians (plus 4 Federal Volunteer Service, BUFDI) in our policlinic, we have determined €â€¯2,927,022 in personnel costs, including overheads, for the entire year. Including infrastructure (€â€¯524,942), material and equipment costs with overheads and internal cost allocation of €â€¯258.657, the total costs in 2019 resulted in €â€¯3,710,621. In contrast, the total income in 2019 was €â€¯3,524,737 generated through the abovementioned patient segments, resulting in a deficit of €â€¯-185,884 (5%). Our data provide evidence that regular outpatient revenues are insufficient and are mainly balanced by outpatient surgery, IVOMs and self-pay patients. In total, there were 19,453 patient contacts during regular office hours (with 17,305 billable cases). At n = 9943, the majority of the contacts were HSA visits; however, only 82% of the cases could effectively be charged due to multiple visits per quarter. The median total patient attendance was 3.21 h (mean 3.38 h). On average, 78 patient contacts were counted per working day. The analysis with TimeElement unveiled a median of n = 2 physician contacts per patient (mean n = 1.91). The median duration per interaction with a physician was 17.98 min (mean 23.23 min). For diagnostics, we counted a median of n = 2 interactions per patient (mean n = 2.31), with an entire interaction lasting a median of 18.30 min (mean 22.60 min). In total n = 37,363 individual diagnostic procedures were recorded in 2019, with SD-OCT being the primary procedure at n = 10,888. CONCLUSION: The cost/turnover calculation showed a marginal financial loss through our ophthalmological outpatient department. Thus, the costs of a university eye outpatient department in Lower Saxony do not seem to be sufficiently covered by direct outpatient revenues. Maintaining quarterly flat rates for all cases of the outpatient department would require a fee of about €â€¯214 in our setting to remain cost neutral. Currently, the lower flat rates in the HSA area are compensated by other areas. Obviously, the high content-related workload in our setting requires a high personnel expenditure with a considerable personnel cost contribution of nearly 80%.

[Influence of COVID-19 shutdown on the workload of a university eye clinic]. / Einfluss des COVID-19-Shutdowns auf die Arbeitsleistung einer Universitäts-Augenpoliklinik.

BACKGROUND: The outbreak of the novel coronavirus disease 2019 (COVID-19) has led to a significant reduction in the number of patients treated in elective medicine in hospitals in spring 2020. Up until now, there are no corresponding data for university policlinics. MATERIAL AND METHODS: With the help of the data of the clinic's electronic medical record system (IS-H/i.s.h.med from SAP, Cerner Corporation, North Kansas City, MO, United States of America and SAP SE, Walldorf, Germany) and the time recording and management software TimeElement (Hannover Medical School, Hannover, Germany), which was codeveloped at our clinic, all patient contacts of the COVID-19 shutdown were evaluated over a period of approximately 7 weeks from 18 March 2020 to 8 May 2020 and compared with the same period in 2019. In addition, quarterly revenues for the first and second quarters of 2019 and 2020 were put into perspective via SAP. RESULTS: During the COVID-19 period the total number of patient contacts was reduced by 59.5% compared to the previous year. The number of lockdown period cases decreased by 74.8% compared to the same period in 2019. In particular, the university outpatient/self-paying sector recorded the largest number of lost patient contacts with a reduction to 17.2% of the 2019 baseline. The reduced patient contacts and case numbers resulted in a significant financial loss of at least €â€¯218,000. Via TimeElement there was a 69.4% decrease in all special diagnostic services, with visual field examinations in particular also reduced by 75.3%. Optical coherence tomography (OCT) measurements recorded a decrease of 60.3%. Patient tracking revealed a reduction of average patient attendance times by approximately 23% (COVID 19: 145.8 ± 88.8 min versus 189.6 ± 97.2 min in 2019). DISCUSSION: The COVID-19 shutdown caused the work performance of our policlinic to drop to only about 40% of patient contacts and that of functional diagnostic examinations to only about 30%, compared to the performance in 2019; however, the reduction in the number of patients also led to a significant reduction in the time patients were present at the clinic compared to the regular workload. The associated financial losses are considerable and obviously not compensated by legally regulated compensation payments as in the inpatient sector.

[Bruch's membrane opening minimum rim width : Correlation and diagnostic accuracy in comparison to peripapillary retinal nerve fiber layer thickness]. / Die auf der Bruchschen Membranöffnung basierende Minimale Randsaumweite : Korrelation und diagnostische Genauigkeit im Vergleich zur peripapillären retinalen Nervenfaserschichtdicke.

BACKGROUND: Glaucoma is one of the main causes of blindness in the Western hemisphere. Because the disease often painlessly progresses it remains unnoticed until major optic nerve head damage occurs in many cases. That is why new, more sensitive diagnostic methods are needed. Bruch's membrane opening minimum rim width (BMO-MRW), measured with the new glaucoma module premium edition (GMPE) was recently introduced as a more accurate tool to detect glaucomatous changes. The purpose of this study was to assess the correlation of SPECTRALIS® spectral domain optical coherence tomography (SD-OCT) and the GMPE anatomic positioning module (APS module) for retinal nerve fiber layer thickness (RNFLT) measurements. The second aim was to assess the diagnostic accuracy of BMO-MRW. METHODS: Prospective study of 41 eyes (41 patients) with glaucoma and 26 eyes from 26 healthy controls. Scans were obtained using SPECTRALIS® SD-OCT and RNFLT was measured with both modules and compared using Spearman's rank test. The BMO-MRW was assessed by GMPE. Sensitivity, specificity and area under receiver operating characteristics curves (AUROC) of each sector of the optic nerve were calculated and compared using the method of Delong et al. RESULTS: We found a positive correlation (0.694-0.955, p < 0.0001) between RNFLT measurements by standard SD-OCT and all diameters of RNFLT of APS module within all sectors. The AUROC of RNFLT in standard SD-OCT was 0.693 for the inferior nasal sector (NI) and BMO-MRW was 0.85 in NI. The difference in AUROC was statistically significant (p = 0.0049). No other sector showed statistically significant differences. CONCLUSION: The RNFLT measurements of both modules showed a positive correlation and appear to be comparable. The BMO-MRW in one sector (NI) showed a significantly higher accuracy of measurement than standard RNFLT. All other sectors showed a comparable accuracy of measurement.

[Software-based visualization of patient flow at a university eye clinic]. / Elektronische Visualisierung des ambulanten Patientenflows in einer Universitäts-Augenklinik.

OBJECTIVE: This article presents a method for visualization and navigation of patient flow in outpatient eye clinics with a high level of complexity. MATERIAL AND METHODS: A network-based software solution was developed targeting long-term process optimization by structural analysis and temporal coordination of process navigation. RESULTS: Each examination unit receives a separate waiting list of patients in which the patient flow for every patient is recorded in a timeline. Time periods and points in time can be executed by mouse clicks and the desired diagnostic procedure can be entered. Recent progress in any of these diagnostic requests, as well as a variety of information on patient progress are collated and drawn into the corresponding timeline which can be viewed by any of the personnel involved. The software called TimeElement has been successfully tested in the practical implemenation for several months. As an example the patient flow regarding time stamps of defined events for intravitreous injections on 250 patients was recorded and an average attendance time of 169.71 min was found, whereby the time was also automatically recorded for each individual stage. CONCLUSION: Recording of patient flow data is a fundamental component of patient flow management, waiting time reduction, patient flow navigation with time and coordination in particular regarding timeline-based visualization for each individual patient. Long-term changes in process management can be planned and evaluated by comparing patient flow data. As using the software itself causes structural changes within the organization, a questionnaire is being planned for appraisal by the personnel involved.

[Intraoperative optical coherence tomography in retinal detachment]. / Intraoperative optische Kohärenztomographie bei Ablatio retinae.

BACKGROUND: Using intraoperative optical coherence tomography (iOCT) can be advantageous during macular surgery and lamellar keratoplasty. It is yet unknown if there is a distinct benefit in its application in retinal detachment surgery. OBJECTIVE: What can be shown using iOCT during retinal detachment surgery? Can therapeutically relevant decisions be made and do they have a prognostic implication on postoperative results? METHODS: Based on already published (11 patients/eyes) and our own new data (23 patients/eyes),findings by iOCT during retinal detachment surgery are presented. RESULTS: Outer retinal corrugations are a frequent feature in iOCT in retinal detachment. These corrugations persist during the application of heavy liquids. Even when the retina seems clinically reattached under the use of perfluoroctane, there is significant subfoveal fluid. Using perfluordecaline, there seems to be less subfoveal fluid. In patients with retinal detachment and macula off situation, subclinical full thickness macular holes seem to be more common than assumed. It is unclear if their incidence is influenced by the use of heavy liquids. They appear to have a negative predictive value regarding postoperative visual acuity. CONCLUSION: Even if there are no obvious benefits in using iOCT in retinal detachment surgery, this new technique offers deeper insights into the microarchitecture of the detached retina. Further investigations in more patients will show if the use of the iOCT will result in a better prognosis for our patients.

[Intraoperative real-time OCT in macular surgery]. / Intraoperative Echtzeit-OCT in der Makulachirurgie.

BACKGROUND: Spectral domain optical coherence tomography (SD-OCT) has become a standard diagnostic tool in the surgical management of vitreomacular interface disorders. The high-resolution cross-sectional information obtained from SD-OCT is a perfect complement to vitreoretinal surgery. It provides detailed intraoperative anatomical views that are not possible with a microscope. OBJECTIVE: To investigate the value of intraoperative real-time OCT with respect to improvement of surgical techniques in the management of vitreomacular disorders. METHODS: A review of the current literature was conducted and an analysis of own systematically evaluated data was included to provide a comprehensive overview of potential applications for the clinical use of intraoperative real-time OCT in macular surgery. RESULTS: Intraoperative real-time OCT can provide detailed visualization of epiretinal membranes and help to identify whether complete membrane removal has been achieved following surgery. In addition, it can provide qualitative and quantitative information that has previously not been available and assist in surgical decision-making. Intraoperative real-time OCT allows membrane peeling to be performed in selected cases without using retinal dyes, whereas it is not ideal for accurately guiding the surgeon while performing maneuvers. CONCLUSION: Intraoperative real-time OCT provides high-resolution visualization of the effects of surgical maneuvers on the microarchitecture of the retina and surrounding tissues and will fill a gap in the understanding of the pathophysiology and prognostic factors of vitreomacular disorders; however, with currently available systems, accurate intraoperative real-time guidance of surgical maneuvers is hindered by several limiting factors.

Switch of anti-VEGF agents is an option for nonresponders in the treatment of AMD.

BACKGROUND: Although anti-VEGF therapy of exudative AMD with bevacizumab and ranibizumab proved efficacious in the majority of patients, CNV activity does not respond to continued treatment after repeated injections in a considerable amount of patients. These are referred to as nonresponders. A change of the drug to bevacizumab or ranibizumab could possibly offer an alternative option for the treatment of nonresponding exudative AMD. METHODS AND MATERIALS: A total of 138 nonresponders who switched therapy from bevacizumab to ranibizumab (n=114) or vice versa (n=24) were included in a retrospective study. Visual acuity (VA) and foveal thickness before and after the switch of therapy were compared. By means of linear regression analysis, we analyzed possible prognostic factors associated with a favorable outcome for visual acuity. RESULTS: Linear regression analysis revealed a statistically significant benefit for nonresponders when treatment was changed to a different anti-VEGF drug (bevacizumab or ranibizumab). VA at the time of the switch was positively correlated with a beneficial development of VA after changing the drug. There was no significant correlation with age, macular thickness, number of injections before the switch, or the development of VA under treatment before the switch. Both patients switching to Avastin and Lucentis benefitted without statistically significant differences. CONCLUSIONS: An exchange of bevacizumab with ranibizumab or vice versa should be considered in nonresponders in the treatment of exudative AMD. Further prognostic factors may help to identify patients who might benefit from a switch. These factors should be investigated in further studies.

[Spontaneous macula hemorrhage. Subhyaloid/sub-inner limiting membrane (ILM)]. / Spontane Makulablutung. Subhyaloidal/Sub-Membrana limitans interna (MLI).

This case study describes the clinical findings of two patients with spontaneous subhyaloid and sub-inner limiting membrane preretinal macular hemorrhage including ophthalmoscopy, optical coherence tomography (OCT) and fluorescein angiography (FAG), which are typical for a Valsalva retinopathy. The improvement of the decreased visual acuity was achieved in one case by spontaneous resorption and in the other case by pars plana vitrectomy. The etiology, diagnostic criteria and therapeutic options are discussed.

[Sudden appearance and rapide progression of bilateral visual deterioration]. / Plötzlich aufgetretene und rasch progrediente beidseitige Visusverschlechterung.

This article describes the case of a 49-year-old patient with incomplete Vogt-Koyanagi-Harada syndrome. The anamnesis showed that intermittent alternating visual problems had begun 2 weeks before. The best corrected vision was 0.1 bilateral. In addition to a right-sided anterior uveitis multiple blister-like retinal alterations of both fundi were seen in fundoscopy. The fundoscopic findings could be confirmed by spectral domain optical coherence tomography (SD-OCT) as multiple intraretinal cysts and areas with neurosensory detachment. Topical and systemic steroid therapy resulted in a rapid reduction of these symptoms. The reversal in SD-OCT corresponded with a visual improvement.

[Ocular ischemic syndrome]. / Okuläres Ischämiesyndrom.

Ocular ischemic syndrome (OIS) is a group of ocular diseases caused by chronic artery occlusion usually involving the internal carotid artery. Patients suffer from visual loss and pain. OIS is a rare disease which can be confounded with diabetic retinopathy or an older central retinal vein occlusion. The only therapy is to treat the neovascular complications. Due to the high mortality of OIS patients, medical and neurological examinations are mandatory. We discuss the clinical findings and diagnostic and therapeutic options of OIS patients in this paper.

[Functional results with two multifocal intraocular lenses with different near addition]. / Funktionelle Ergebnisse zweier Multifokallinsen mit unterschiedlicher Nahaddition.

BACKGROUND: The reduction of the near addition of multifocal intraocular lenses from +4 D to +3 D should improve the visual function in the intermediate distance. The ReSTOR +3 D is a multifocal lens with a near addition of 3 D. PATIENTS AND METHODS: In a prospective study the new ReSTOR +3 D was compared to the former ReSTOR platform with 4 D near addition. A total of 40 patients received 1 of these 2 multifocal lenses bilaterally in both eyes after uneventful phacoemulsification in otherwise sound cataract eyes (20 patients ReSTOR +3 D, 20 patients ReSTOR + 4D). After 3 months the following parameters were assessed: uncorrected and corrected distance visual acuity, near vision at different distances, contrast sensitivity, defocus curve and anterior and posterior segments. The functional tests were performed monocularly and binocularly. RESULTS: Uncorrected binocular distance visual acuity (decimal) was 0.93 (logMAR 0.03 ± 0.12) in the ReSTOR +3 D group and 1.05 (-0.02 ± 0.1) in the ReSTOR +4 D group. In the intermediate distance the ReSTOR +3 D performed slightly better at 40 cm (0.68 versus 0.56) (logMAR 0.17 ± 0.14 versus 0.24 ± 0.18). The difference was significant (p<0.01) at 70 cm (0.69 versus 0.43) (logMAR 0.16 ± 0.27 vs 0.37 ± 0.2). The superior performance of the ReSTOR +3 D in the intermediate distance was also clearly visible in the defocus curves. All functional tests revealed significantly better results when performed binocularly. CONCLUSION: The ReSTOR +3 D multifocal intraocular lens performed significantly better in the intermediate distance compared to the ReSTOR +4 D. The other functional tests showed excellent and equal results with both lenses. The reduction of near addition does not increase dysphotopsia, particularly halos. Complaints of unsatisfactory vision in the intermediate distance were reported only by patients with the ReSTOR +4 D. The new ReSTOR +3 D is likely to improve the acceptance of multifocal intraocular lenses.

[Clinical diagnostics and therapy for non-arteritic central retinal artery occlusion]. / Diagnostik und Therapie beim nicht arteriitischen Zentralarterienverschluss.

Central retinal artery occlusion (CRAO) is an ophthalmological emergency situation. Known risk factors are arterial hypertension, cardial arrhythmia, arteriosclerosis, hypercholesterolemia and diabetes. Elderly patients should be examined for an arteritic genesis. Young patients (< 45 years) without typical risk factors may suffer from thrombophilia. There is no uniform recommendation on how to treat non-arteritic CRAO. Many different interventions have been suggested in the literature, i. e., massaging the eye, systemic or local reduction of intraocular pressure, anticoagulation, either systemically administered venous thrombolysis or supraselective intra-arterial thrombolysis. In this review we present the causes of CRAO and diagnostic means to detect causes; we also critically discuss previously described therapeutic options. It is our aim to provide a guide through the necessary interdisciplinary diagnostics in co-operation with internal medicine and neurology and to recommend a multimodal therapy in patients with non-arteritic CRAO.

[Coats disease]. / Morbus Coats.

Coats disease is characterized by idiopathic congenital retinal telangiectasis and exudative retinopathy. Mostly young males are affected. The disease is usually unilateral. Visual prognosis varies from full visual acuity to blindness according to the extent of retinal exudation. Retinoblastoma is a major differential diagnosis particularly in small children but other causes of exudative retinopathy must also be considered. Treatment aims include obliteration of the telangiectasis in order to resolve retinal exudation and achieve reattachment of the retina. Therapeutic options comprise laser coagulation or cryotherapy, drainage of subretinal fluid, buckling surgery and pars plana vitrectomy. In more advanced cases with therapy-resistant secondary glaucoma enucleation may be required. More recently, intravitreal administration of VEGF inhibitors has become an option.

Resolving the clinical acuity categories "hand motion" and "counting fingers" using the Freiburg Visual Acuity Test (FrACT).

PURPOSE: The Freiburg Visual Acuity Test (FrACT) has been suggested as a promising test for quantifying the visual acuity (VA) of patients with very low vision, a condition often classified using the semi-quantitative clinical scale "counting fingers" (CF), "hand motion" (HM), "light perception" (LP) and "no light perception". The present study was designed to assess FrACT performance in a sizable number of CF, HM, and LP patients in order to generate a setting for future clinical studies in the low vision range. METHODS: We examined a total of 41 patients (LP, n = 11; CF, n = 15; HM, n = 15) with various eye diseases (e.g., diabetic retinopathy, ARMD), covering the clinical VA scale from LP to CF. The FrACT optotypes were presented at a distance of 50 cm on a 17-inch LCD monitor with four random orientations. After training, two FrACT measurements (test and retest) were taken, each comprising 30 trials. RESULTS: FrACT measures reproducibly the VA of CF and HM patients. In CF patients, FrACT resulted in a mean logMAR = 1.98 +/- 0.24 (corresponding to a decimal VA of 0.010), for HM in a mean logMAR = 2.28 +/- 0.15 (corresponding to a decimal VA of 0.0052). In all LP patients the FrACT values were close to what would be obtained by random guessing. The mean test-retest 95% confidence interval was 0.21 logMAR for CF patients and 0.31 logMAR for HM respectively. Test-retest variability declined from 24 to 30 trials, showing that at least 30 trials are necessary. CONCLUSION: FrACT can reproducibly quantify VA in the CF and HM range. We observed a floor effect for LP, and it was not quantifiable further. Quantitative VA measures are thus obtainable in the very low-vision range using FrACT.

Studies on MIP-2 and CXCR2 expression in a mouse model of extrahepatic colorectal metastasis.

AIMS: The CXC chemokine macrophage inflammatory protein (MIP)-2 has been shown to promote outgrowth of colorectal liver metastasis by enhancing angiogenesis and tumor cell migration. However, the effect of MIP-2 on extrahepatic metastasis is not known yet. With a use of a murine model, we therefore studied cell proliferation and microvascularization of extrahepatic CT26.WT-GFP colorectal tumors after exposure to MIP-2. METHODS: Green fluorescent protein (GFP)-transfected CT26.WT colorectal cancer cells were implanted in dorsal skinfold chambers of syngeneic BALB/c mice. After 5 days, the tumors were locally exposed to 100 nM MIP-2. Cell proliferation as well as tumor microvascularization and growth were studied during a further 9-day period using intravital fluorescence microscopy, histology and immunohistochemistry. Tumors exposed to PBS served as controls. RESULTS: MIP-2 induced a marked CXCR2 expression and promoted a distinct tumor cell proliferation. This was associated with a significant increase of tumor size compared to PBS-treated controls. Of interest, MIP-2 did not affect dilation and permeability of the tumor microvessels, which would be indicators for an enhanced VEGF action. Accordingly, the angiogenic response, e.g. the outgrowth of new microvessels, was not affected, and the density of the established tumor microvascular network was even found decreased after MIP-2 exposure when compared to PBS controls. CONCLUSION: With the use of a murine tumor model, we demonstrate that MIP-2 accelerates growth of experimentally established extrahepatic colorectal metastases by inducing tumor cell proliferation rather than promoting vascularization.

[Von Hippel-Lindau disease. Interdisciplinary patient care]. / Von-Hippel-Lindau-Erkrankung. Interdisziplinäre Patientenversorgung.

Von Hippel-Lindau disease is an important hereditary tumor syndrome with a clear option for effective treatment if diagnosed in time. Interdisciplinary cooperation is the key to successful management. Major components of the disease are retinal capillary hemangioblastomas, hemangioblastomas of cerebellum, brain stem and spine, renal clear cell carcinomas, pheochromocytomas, multiple pancreatic cysts and islet cell carcinomas, tumors of the endolymphatic sac of the inner ear, and cystadenomas of the epididymis and broad ligament. A well structured screening program should be performed at yearly intervals.

[Retinal angiomatosis. Ocular manifestation of von Hippel-Lindau disease]. / Angiomatosis retinae. Okuläre Manifestation des Von-Hippel-Lindau-Syndroms.

Von Hippel-Lindau disease (VHL disease) is a rare multisystem disorder of autosomal dominant inheritance with high penetrance. Inactivation of the VHL-protein leads to an increased expression of hypoxia induced growth factors. Predilection sites for tumor growth are the retina, the central nervous system and various visceral organs. Retinal capillary hemangioblastoma is one of the earliest manifestations of VHL disease. The lifetime risk of permanent visual loss defined as a visual acuity of 0.5 or less is about 35% in gene carriers. It increases to 60% if there is already retinal capillary hemangioblastoma. If VHL disease is suspected, a careful ophthalmological examination should be included in the clinical screening program. Having confirmed the diagnosis, regular ophthalmoscopic monitoring is essential in order to detect developing tumors at an early stage. Therapeutic options for small to medium sized peripheral tumors are laser or cryocoagulation; larger- hemangioblastomas can be treated by brachytherapy using ruthenium plaques, while asymptomatic juxtapapillary tumors can be observed at regular intervals.

Development of a highly productive and scalable plasmid DNA production platform.

With the applications of DNA vaccines extending from infectious diseases to cancer, achieving the most efficient, reproducible, robust, scalable, and economical production of clinical grade plasmid DNA is paramount to the medical and commercial success of this novel vaccination paradigm. A first generation production process based on the cultivation of Escherichia coli in a chemically defined medium, employing a fed-batch strategy, delivered reasonable volumetric productivities (500-750 mg/L) and proved to perform very well across a wide range of E. coli constructs upon scale-up at industrial scale. However, the presence of monosodium glutamate (MSG) in the formulation of the cultivation and feed solution was found to be a potential cause of process variability. The development of a second generation process, based on a defined cultivation medium and feed solution excluding MSG, was undertaken. Optimization studies, employing a plasmid coding for the HIV gag protein, resulted in cultivation conditions that supported volumetric plasmid titers in excess of 1.2 g/L, while achieving specific yields ranging from 25 to 32 microg plasmid DNA/mg of dry cell weight. When used for the production of clinical supplies, this novel process demonstrated applicability to two other constructs upon scale-up in 2,000-L bioreactors. This second generation process proved to be scalable, robust, and highly productive.

Production of indole diterpenes by Aspergillus alliaceus.

Production of two related indole diterpenes (differing by a dimethyl leucine side chain) by Aspergillus alliaceus was improved through several pilot scale fermentations. Media were optimized through focus primarily on initial increases, as well as mid-cycle additions, of carbon and nitrogen sources. Fermentation conditions were improved by varying ventilation conditions using various combinations of air flowrate and back-pressure set points. Production improvements were quantified based on total indole diterpene concentration as well as the ratio of the major-to-minor by-product components. Those changes with a positive substantial impact primarily on total indole diterpene concentration included early cycle glycerol shots and enhanced ventilation conditions (high air flowrate, low back-pressure). Those changes with a significant impact primarily on ratio included higher initial cerelose, soybean oil, monosodium glutamate, tryptophan, or ammonium sulfate concentrations, higher broth pH, and enhanced ventilation conditions. A few changes (higher initial glycerol and monosodium glutamate concentrations) resulted in less notable and desirable titer or ratio changes when implemented individually, but they were adopted to more fully realize the impact of other improvements or to simplify processing. Overall, total indole diterpene titers were improved at the 600 L pilot scale from 125-175 mg/L with a ratio of about 2.1 to 200-260 mg/L with a ratio of about 3.3-4.5. Thus, the ability to optimize total indole diterpene titer and/or ratio readily exists for secondary metabolite production using Aspergillus cultures.

Bioprocess monitoring and computer control: key roots of the current PAT initiative.

This review article has been written for the journal, Biotechnology and Bioengineering, to commemorate the 70th birthday of Daniel I.C. Wang, who served as doctoral thesis advisor to each of the co-authors, but a decade apart. Key roots of the current PAT initiative in bioprocess monitoring and control are described, focusing on the impact of Danny Wang's research as a professor at MIT. The history of computer control and monitoring in biochemical processing has been used to identify the areas that have already benefited and those that are most likely to benefit in the future from PAT applications. Past applications have included the use of indirect estimation methods for cell density, expansion of on-line/at-line and on-line/in situ measurement techniques, and development of models and expert systems for control and optimization. Future applications are likely to encompass additional novel measurement technologies, measurements for multi-scale and disposable bioreactors, real time batch release, and more efficient data utilization to achieve process validation and continuous improvement goals. Dan Wang's substantial contributions in this arena have been one key factor in steering the PAT initiative towards realistic and attainable industrial applications.