The antibacterial activity of host defense peptides (HDP) is largely mediated by permeabilization of bacterial membranes. The lipid membrane of enveloped viruses might also be a target of antimicrobial peptides. Therefore, we screened a panel of naturally occurring HDPs representing different classes for inhibition of early, Env-independent steps in the HIV replication cycle. A lentiviral vector-based screening assay was used to determine the inhibitory effect of HDPs on early steps in the replication cycle and on cell metabolism.Human LL37 and porcine Protegrin-1 specifically reduced lentiviral vector infectivity, whereas the reduction of luciferase activities observed at high concentrations of the other HDPs is primarily due to modulation of cellular activity and/ or cytotoxicity rather than antiviral activity. A retroviral vector was inhibited by LL37 and Protegrin-1 to similar extent, while no specific inhibition of adenoviral vector mediated gene transfer was observed. Specific inhibitory effects of Protegrin-1 were confirmed for wild type HIV-1.Although Protegrin-1 apparently inhibits an early step in the HIV-replication cycle, cytotoxic effects might limit its use as an antiviral agent unless the specificity for the virus can be improved. The lowrn model ,22.Some of these peptides are induced at epithelial surfaces in response to invading organisms -25. ManyIn addition, several antiviral activities were reported. Recently it has been demonstrated that rabbit neutrophil peptide alpha-defensin NP1 protects cells from infection with HSV-1 and 2 . Other sThe antibacterial activity of HDPs is largely mediated by pore formation leading to permeablization of the bacterial membrane. Although some selectivity for bacterial membranes has been described, the lipid membrane of enveloped viruses might also be a target of antimicrobial peptides ,40. ThisGiven the potential cytotoxicity of HDPs, it was important to discriminate between modulation of cell metabolism and direct antiviral effects. We were concerned that HDPs could modulate host cell metabolism without affecting cell viability as assayed for example by standard MTT assays. We therefore decided to use immunodeficiency virus-based vectors transferring the luciferase gene to determine both, the HDP antiviral activity and modulation of cell metabolism. The luciferase activity of target cells, stably transduced with the lentiviral vector prior to HDP treatment should reveal any effect of HDPs on cellular transcriptional and translational efficacy. Treating cells with HDPs during infection with the lentiviral vector and comparison with results obtained with the stably transduced cells should then allow identifying HDPs that inhibit early steps in the viral replication cycle. To validate the luciferase-based assay for modulation of cell metabolism, 293 cells were stably transduced with a lentiviral vector transferring the luciferase gene. Incubation of transduced cells with increasing concentrations of Protegrin-1 and LL37 led to a dose-dependent inhibition of luciferase activity , fungal Plectasin and Novispirin G10, inhibited gram positive and gram negative bacteria revealing antimicrobial activity in the expected range (Table 50) of LL37 and PG-1, resulting in IC50s of approximately 30 μg/ml and 16.8 μg/ml, respectively . To evaluate whether LL37 and PG-1 directly target VSV-G or a lentiviral protein, the inhibitory effect of these HDP against the lentiviral vector was compared side by side to their effect on a retroviral vector containing the amphotropic MLV Env for entry into target cells. The dose dependent reduction in the luciferase activity in the target cells was very similar in cells transduced with the lentiviral or the MLV vector Fig. .50s of 25 μg/ml and 14 μg/ml were observed for LL37 and PG-1, respectively  [syn, 2 μg of pHIT-G and 5 μg of VLΔBH were transiently cotransfected by the CaPO4coprecipitation method into 293T cells as previously described [syn, pcTat [To generate lentiviral vector particles transferring the luciferase gene, a codon-optimized (Geneart GmbH, Regensburg, Germany) HIV-1 (Hgpsyn)  and a VS(Hgpsyn)  were use(Hgpsyn) .5 μg of escribed . An HIV escribed  was alson, pcTat , pcRev [n, pcTat  and pHITn, pcTat . The MLVn, pcTat . Two dayin vitro ligation [dl327 (kindly provided by T. Shenk, Princeton University, Princeton, NJ), utilizing the unique Bst1107 I restriction site. The vector was propagated in 293 cells and purified by two rounds of CsCl density centrifugation [2 and 10% glycerol four-times (1 hour each) at 4°C, and stored at -80°C until use. The concentration of the vector was determined by measuring absorbency at 260 nm [Beginning with a first generation E1- and E3-deleted adenoviral vector, we generated a replication-competent adenoviral vector Ad.OW126, which harbors in the E1 region the firefly luciferase cDNA (subcloned from pGEM-Luc (Promega, Madison, WI)), a IRES element ,51, and ligation  to H5dl3fugation , dialyzet 260 nm , and thet 260 nm . The ratsyn and pHIT-G. 293A cells were plated in 24 well plates at a density of 50.000 cells / well and transduced with 200 μl of VLΔBH-SIN vector for two hours. Two days after plating cells were transferred to one well of a six well plate and transduced again with 1 ml of VLΔBH-SIN vector. Cells were subsequently expanded resulting in 293-Luc cells.To stably transduce the luciferase gene into 293A cells, a self-inactivating version of VLΔBH, VLΔBH-SIN similar to VGΔBH-SIN  was packThe supernatant of infected 293A or 293-Luc cells, cultured in 96 well plates was removed and cells were lysed in 50 μl of cell lysis buffer (Promega, Pittsburgh, PA). 20 μl of the cell lysates were used in the firefly luciferase assay system of Promega as described by the manufacturer. Each single value of the triplicates was expressed as percent of the mean of triplicates of control cultures infected with the same vector in the absence of HDPs and the mean and the standard deviation of the percent values was calculated for each triplicate.in vitro and in vivo studies. Potential endotoxin contamination was monitored with the chromogenic Limulus amoebocyte lysate assay  using Escherichia coli endotoxin (supplied with the kit) as the standard. Endotoxin levels for the peptides were not detectable.The antimicrobial peptides  used in this study were prepared by solid phase synthesis and purified by RP-HPLC. Recombinant human β-Defensin-2 was produced by a molecular farming approach in transgenic potato tubers and purified by perfusion chromatography (data not shown). The peptides (≥ 98% pure) were dissolved in 0.01% acetic acid and used for all Acinetobacter baumannii (ATCC 19606), Escherichia coli (ATCC 25922) and Pseudomonas aeruginosa (ATCC 27853) Gram-positive strains: Staphylococcus aureus (ATCC 25923), Staphylococcus epidermidis (ATCC 12228) and Enterococcus faecalis (ATCC 29212).The following strains were used in this study: Gram-negative strains: 620 × 2.5 × 108.All bacterial strains were analyzed with API test strips (BioMerieux, Hazelwood, MO) to confirm identity and aliquots were stored frozen in 50 % skim milk at -80°C. Bacteria were grown overnight in trypticase soy broth (Becton Dickinson, Franklin Lakes, NJ) at 275 rpm and 37°C. An aliquot of the resulting stationary phase cultures was then transferred to 20 ml of trypticase soy broth and incubated at 37°C for 2.5 hours to reach log phase. This subculture was transferred to a 50 ml conical polystyrene tube and centrifuged for 10 min at 4°C at 880 g. The bacterial pellet was washed once with chilled phosphate buffered saline, pH 7.4, and resuspended in 5 ml of the same cold buffer. One milliliter was removed to measure its optical density at 620 nm. The bacterial concentration was calculated from the following formula: CFU/ml = ODin vitro a radial diffusion assay was performed as previously described [6 CFU) were mixed with 10 ml of underlay gel (43°C) and immediately poured into square 9 × 9 cm petri dishes. A series of 3 mm wells was punched after the agarose solidified. After appropriate serial dilutions were done, 5 μl of HDP, vancomycin (Abbott Labs, Chicago, IL), gentamicin, ciprofloxacin, or fluconazole (Sigma-Aldrich, St. Louis, MO) were added to the designated wells. Plates were incubated at 37°C for 3 hours. The bacteria-containing layer was covered with a 10 ml overlay of the nutrient rich agar. The overlay agar consisted of 6% (w/v) TSB and 1% agarose in PBS for all assays. After 18 h of incubation at 37°C, the plates were stained with 0.001% Coomassie blue for 10 h. The clear zones  around the punched wells indicated antibacterial activity. The diameters of the clear zones were converted into units by subtracting the well diameter and multiplying the difference by 10. Results were plotted using a semi log scale and correlation coefficients and X-intercepts obtained from linear regression analysis. The minimal effective concentration (MEC) corresponded to the X-intercept value. All assays were performed in triplicates and repeated at least once.To monitor bacterial growth inhibition escribed . Briefly3 cells / well. After 48 hours, 50 μl of MTT-solution (3 mg/ml) was added and incubated at 37° C under 5% CO2 for 1 hour. After this time medium was removed and 100 μl 0.04 N HCL + 10% SDS was used to dissolve the resulting blue formazan crystals in living cells. The optical density was determined at 550 nm. Each single value of the triplicates was expressed as percent of the mean of triplicates of control cultures infected with the same vector in the absence of HDPs and the mean and the standard deviation of the percent values was calculated for each triplicate. In addition the BrdU Cell proliferation ELISA with chemiluminescence detection (Roche Diagnostics GmbH, Mannheim, Germany) was performed. After 293-Luc cells or P4CCR5 cells were plated in 96 well plates at a density of 2 × 103 cells/well BrdU (5-bromo-2'-deoxyuridine) was added to the cells with a resulting concentration of 10 μM for the last 22 h of the incubation period. After removing the culture medium, the cells were fixed and DNA was denatured in one step with Fixdenat. Thereafter the cells were incubated with Anti-BrdU-POD for 1 h at room temperature. The chemiluminescence detection was measured after automatic injection of substrate solution with a microplate-luminometer (Orion, Berthold detection systems, Pforzheim, Germany).293-Luc cells were plated in 96 well plates at a density of 2 × 103 cells / well. After overnight incubation, the supernatant of the wells were removed and replaced by 25 μl vector preparation and 25 μl HDP at twice the final concentration indicated. 50 μl fresh medium with HDP at the final concentration indicated was added after two hours. One  or two  days after infection, the supernatant was removed and 50 μl of cell lysis buffer (Promega) was added. Lysates were stored at -80°C until determination of the luciferase activity of the extracts. The affect of HDPs on cell metabolism was determined in parallel by plating 293-Luc cells exactly the same way as 293A cells and the luciferase activity was determined after one  or two  days of incubation with HDPs. For GFP-expressing vectors, vector titers were calculated from the number of GFP positive cells per well as previously described [To determine the effect of HDPs on vector infectivity and cell metabolism, 293A target cells were plated in triplicates into 96-well plates at 2 × 10escribed  and the 3 cells / well of 96 well plate. 50 μl fresh medium with HDP at the final concentration indicated was added after two hours. Two days after infection, the supernatant was removed and cells were stained by X-Gal. The number of infected cells per well were counted in the microscope.Stocks of HIV-1 were generated by transient transfection of 293T cells with the molecular clone pNL4-3. 25 μl of the virus stock were incubated for 30 minutes at room temperature with 25 μl of LL37 or PG-1 adjusted to twice the final concentration indicated. The mixture was added to P4CCR5 cells plated the day before at a density of 2 × 10The author(s) declare that they have no competing interests.LS, BT and JM performed most of the experiments. LS, OW, ML, EL, HH, HS and KU participated in the experimental design, data interpretation and writing of the manuscript.


Elevated low-density lipoprotein (LDL)-cholesterol is associated with a significantly increased risk of coronary heart disease. Ezetimibe is the first member of a new class of selective cholesterol absorption inhibitors. It impairs the intestinal reabsorption of both dietary and hepatically excreted biliary cholesterol. Ezetimibe is an effective and safe agent for lowering LDL-C and non HDL-C. Short term clinical trials have established the role of ezetimibe monotherapy and its use in combination with statins. Furthermore, ezetimibe and statin combination therapy increased the percentage of patients who achieved their LDL-C treatment goal.Studies using surrogate markers of atherosclerosis have suggested a possible role of ezetimibe in combating atherosclerosis. Ezetimibe provides an effective therapeutic strategy for the management of homozygous familial hypercholesterolemia (HoFH) and sitosterolemia. The lack of outcomes and long term safety data is attributed to the relatively recent introduction of this medication. Over 60 million Americans suffer from cardiovascular disease (CHD). The incidence of CHD and stroke has been on the rise partly because of the increase in life expectancy and the explosive epidemic of diabetes and the metabolic syndrome . CHD is Animal and human studies have established the role of cholesterol in the development and progression of atherosclerosis. LDL-cholesterol (LDL-C) constitutes approximately 60–70 % of total serum cholesterol. Epidemiological studies directly implicated LDL-C to the development of atherosclerosis and CHD. Furthermore, LDL-C level appears to be directly related to the development and recurrence of CHD .Animal studies suggested a protective effect of low LDL-C against atherosclerosis . MultiplMost of the landmark CHD prevention trials involved the use of statin medications. LDL-C remains the primary target of treatment in most instances, and statins are the mainstay of LDL-C lowering treatment .The National Cholesterol Education Program/ Adult Treatment Panel III (NCEP/ATP III) updated guidelines table  for deteEzetimibe, a cholesterol absorption inhibitor, is the first agent of a new class of lipid-lowering compounds that selectively inhibits the intestinal absorption of cholesterol and related phytosterols. Ezetimibe undergoes extensive glucuronidation to an active metabolite in the intestinal mucosa . EzetimiAfter oral administration, ezetimibe is absorbed and extensively conjugated to a pharmacologically active phenolic glucuronide (ezetimibe-glucuronide) , the druPlasma concentrations for total ezetimibe were about 2-fold higher in older individuals (>65 years), levels were similar in adolescents to healthy adults and may be higher in women than in men . In patiAdverse experiences were reported in approximately 2% of patients treated with ezetimibe and included fatigue, arthralgia, diarrhea, abdominal pain and back pain. Angioedema and rash were reported after general clinical use of this medication . With co1. Primary Hypercholesterolemia , Ezetimibe is indicated in this case for use as both mono and combination therapy.2. The reduction of elevated total-C and LDL-C levels in patients with homozygous familial hypercholesterolemia (HoFH) either as primary or as an adjunct to other lipid-lowering treatments.3. In patients suffering from Homozygous Sitosterolemia, as adjunctive therapy to diet for the reduction of elevated sitosterol and campesterol levels in patients with homozygous familial sitosterolemia.Multiple studies conducted to examine the effects of ezetimibe monotherapy have concluded that this drug was effective in lowering LDL-C versus placebo.Analysis of multicenter, double-blind, placebo-controlled trials demonstrated that ezetimibe at the 10 mg once daily clinically approved dose significantly modified cholesterol and cholesterol subtypes in patients with hypercholesterolemia when compared to placebo. Ezetimibe significantly lowered total-Cholesterol (TC) (12 %), LDL-C (18 %), apolipoprotein B (Apo B) (15 %), and triglycerides (TG) (7%) and increased high density lipoprotein (HDL-C) (3.5%) -24. LipoIn a case series report, we analyzed the effects of Ezetimibe on cholesterol particle size and number using NMR technology . We found that Ezetimibe lowered cholesterol particle number by a mean 26 % and had no significant effect on cholesterol particle size [The effects of ezetimibe on cholesterol and its subtypes were not influenced by risk-factor status, gender, age, race, time of administration, or baseline lipid profile . The oveThe struggle to achieve the NCEP/ATP III guidelines LDL-C goals through primary utilization of statins is often frustrating for the clinician. In a study to examine the efficacy of statin titration on attainment of LDL-C goal, the authors concluded that for high risk patients, approximately half were able to achieve their LDL-C goal at the appropriate statin starting dose, and only one third of the titration group were able to achieve the NCEP/ATP III cholesterol goal . Now witHMG-CoA reductase inhibitors (statins) act on the rate-limiting step to inhibit HMG-CoA conversion to mevalonate, effectively decreasing LDL-C synthesis. They result in a decrease of LDL-C ranging between 30–60 %, depending on the individual statin and the dose administered. Statin induced LDL-C lowering appears to be effective in reducing CHD and CHD related mortality and morbidity. The extent of CHD and CHD related events reduction is proportionate to the extent of LDL-C reduction ,28. LDL-Reasons to initiate combination therapy to treat hyperlipidemia include: further LDL-C lowering, reducing side effects related to higher doses of statins, modifying other risk factors besides LDL-C such as HDL-C and TG.Increasing the dose of statins has a limited effect on reducing LDL-C, as it is well established that doubling the dose of a statins leads to a 5 % more reduction in total TC and 7 % more reduction in LDL-C with each doubling .Although statins have demonstrated similarity in CHD related events, they are heterogeneous not only in LDL-C lowering efficacy but also in their safety profiles. The bulk of the statins effect on LDL-C occurs at the initial recommended dose and they are safer when used at doses below the maximal recommended dose.Statins are the most effective drugs known to modify LDL-C, but in terms of HDL-C and TG modifying capacity, other classes of lipid lowering medications used alone or in combination with statins offer higher efficacy.The metabolism of cholesterol is an intricate process that involves both produced and ingested cholesterol. The mechanism of action of HMG-CoA reductase inhibitors affects the production of cholesterol, whereas that of cholesterol absorption inhibitors affects absorbed cholesterol, thereby offering potential synergism of action when the medication are used in combination. Trials examining the efficacy have demonstrated synergism and consistency in LDL-C lowering in the absence pharmacokinetic interaction between the statins and ezetimibe.In a relatively large, multicenter study, involving patients with primary hypercholesterolemia already receiving statin monotherapy , patients were randomized to receive either ezetimibe or placebo in addition to their current statin therapy. At the conclusion of this 8 week study, the ezetimibe and statin groups were found to have a significantly lower total-C, LDL-C, Apo B, and TG, and increased HDL-C when compared to the statin only and placebo groups. Furthermore, LDL-C reductions induced by ezetimibe were generally consistent across all statins groups .Another multicenter, double-blind, randomized trial examined the effects of ezetimibe on patients suffering from (HoFH). At the initiation of the trial patients were receiving either atorvastatin or simvastatin. The addition of ezetimibe reduced LDL-C by an additional 20.5 % in contrast to only 6.7 % reduction that resulted from doubling the statin dose . SimilarThe addition of ezetimibe to statins is superior to treatment with statins alone in lowering non-HDL-C, ezetimibe co-administered with simvastatin lowered non-HDL-C by 47.1% whereas, simvastatin monotherapy lowered non-HDL-C by 33.6% when results were pooled across different doses [In terms of modifying risk factors other than LDL-C, the co-administration of ezetimibe with statins had a more favorable effect on HDL-C and TG when compared to statins therapy alone . In concEzetimibe co-administration with fibric acid derivatives was examined in a randomized, evaluator-blind, placebo-controlled, parallel-group study of 32 healthy hypereholesterolemics. Ezetimibe co-administration with fenofibrate was found to produce clinically significant reductions in LDL-C (36.3%) compared to the fenofibrate group (22.3%) with a more favorable TG and HDL-C profile .Sitosterolemia is a rare inherited disorder caused by mutation in either the ABCG5 or ABCG8 genes located on chromosome (2p21) . First dClinical manifestations include: tendon and tuberous xanthomas, episodes of hemolysis, accelerated atherosclerosis, and premature coronary artery disease. It is important to note that close to 50 % of these patients have normal cholesterol levels -41.A recently reported trial demonstrated that treatment with ezetimibe reduces plant sterol levels in patients with sitosterolemia. The authors reported a decrease in sitosterol concentrations by 21% and campesterol by 24 % .It is generally accepted that atherosclerosis is an inflammatory disorder. It is believed that the atherosclerotic process begins with endothelial cell activation, which is triggered by multiple factors such as oxidized lipoproteins. Cholesterol lowering agents as a group have demonstrated great efficacy in prevention and cessation of the progression of atherosclerosis. The efficacy of ezetimibe in monotherapy or in combination on CHD morbidity and mortality has not been well established.One of the unique features of cholesterol absorption inhibitors is their ability to modify post-prandial hyperlipidemia. There is increasing evidence that post-prandial lipoproteins (particularly cholesterol-rich chylomicron remnant) are atherogenic. Ezetimibe has the potential to reduce the cholesterol content of chylomicrons by up to 60% , which mHigh-sensitivity C-reactive protein (hs-CRP) is an inflammatory mediator whose levels correlate with increased coronary risk. Ezetimibe co-administered with simvastatin resulted in significant incremental decreases in hs-CRP in patients with primary hypercholesterolemia. Changes in individual lipid parameters did not explain the observed decreases in hs-CRP and were possibly consistent with an additional anti-inflammatory effect compared with simvastatin monotherapy .In a prospective trial to study effects of ezetimibe co-administered with atorvastatin in patients with primary hypercholesterolemia, ezetimibe plus atorvastatin significantly provided an additional (10%) lowering of hs-CRP versus atorvastatin alone .Ezetimibe and its class of cholesterol absorption inhibitors are new, and there is a lack of outcomes data to explore whether its cholesterol modifying effects will translate to lower CHD mortality and morbidity. The safety of this medication has not yet been established with long term trials data as most of the studies conducted were short term. With the advent and increased utilization of combination therapy in the management of dyslipidemia, further trials are needed to explore the efficacy, indications and safety profile of ezetimibe use in combination with Peroxisome proliferator-activated receptors (PPARs), niacin and bile acid resins.The increased popularity of special weight loss diets such as the high protein diet, poses questions of whether such diets will alter the efficacy or safety of cholesterol absorption inhibitors.Finally, the efficacy of statins in reducing CHD related events has lead to the controversial hypothesis regarding whether or not statins poses a pleiotropic (non lipoprotein) effect. If a pleiotropic effect exists, one might argue that a statin at a higher dose might be more beneficial than combination therapy producing the same effect.Ezetimibe is the first clinically approved cholesterol absorption inhibitor. It is effective in lowering LDL-C as monotherapy or in combination with statins. The use of combination LDL-C lowering medication is expected to become a much more common modality of treatment, especially after the recent NCEP/ATP III update. Ezetimibe offers further lowering of LDL-C and non HDL-C that is consistent and probably safer than increasing the dose of the individual statin. It also provides another effective treatment option for HoFH and sitosterolemia patients. Because of its recent introduction, we still lack both outcomes and long term safety data.


After reading the Learning Forum by Fleming and Lynn (1) Morphology: the essential point of dermatological diagnosis is morphology, a low tech, but hard to master, skill. Dermatological diagnosis, as any other medical diagnosis, starts by collecting adequate information from the patient, and follows by its elaboration. Many doctors consider that dermatological diagnosis can be made on a quick recognition basis, but an ordered and syndromic approach is essential to get to an adequate diagnosis. I think that most dermatologists would agree that a good description of a patient by an experienced colleague is a better starting point for diagnosis than many pictures. I would describe the lesions seen in Figure 1 of [1] not simply as shallow ulcers, but as clearly polycyclic erosions (a finding highly suggestive of herpetic infection).PLoS Medicine, with many readers in less developed countries, this test should not be forgotten.(2) Indicated investigations: Tzanck test is the microscopic evaluation of cell morphology on a cutaneous smear. It can be done in about 15 minutes, requiring a microscope and a trained doctor. Access to this test is probably much easier than to viral cultures or polymerase chain reaction tests. In this setting, a positive Tzanck test would be enough to confirm the clinical diagnosis at a minimum cost. Considering the widespread audience of (3) This case, and the suspicion about systemic manifestations of skin disease, is a wonderful opportunity to disseminate an old concept, very frequently forgotten in medical literature: the skin is an organ, in fact, the biggest one in the body. Its main functions are to act as a barrier, to control temperature, to serve immunological and hormonal roles, and, physiologically less important but very important for patient well-being, to participate in personal relationships. When these functions are not adequately performed, skin failure appears, exactly as is the case with heart or renal failure. Skin failure can have many manifestations, including noninfectious fever, bacteremia, or sepsis. As is the case with renal or cardiac failure, it is easier and more practical to learn about this syndrome than to discuss the systemic manifestations of the many diseases that can cause it. I would highly recommend the following references for doctors interested in the subject: ,3.


Direct payments to local communities to conserve wildlife could prove effective but is biodiversity a commodity that can be bought and sold? The language of conservation is changing: protecting biodiversity is no longer just about ethics and aesthetics; the latest buzzwords are commodities and consumers. Traditionally, conservation initiatives have talked up the benefits they will bring to the global community—saving species, habitats, ecosystems, and ultimately the planet. But conservation also has its costs, and these are usually borne by local people prevented from exploiting the resources around them in other ways. It is unfair to expect a localised minority to pick up costs that ultimately benefit a dispersed majority, argue conservation biologists. There has to be more money made available by concerned individuals, non-governmental organisations, national governments, and international bodies, and there need to be better ways to spend this money if conservation is to be effective, they say. Biodiversity is a commodity that can be bought and sold. We are consumers and must pay.2 of the country , and is using direct payments to help reduce the slaughter of turtles by local fishermen. The Watamu Turtle Watch Program is currently paying fishermen just over $3 a turtle to release the animals from their nets rather than kill them. Before the scheme started in 2000, only around 50 turtles were being released from nets each year. By 2003, more than 500 a year were making it back into the sea. Elsewhere along the Kenyan coast, where fishermen do not get these payments, turtles continue to be killed, says Trott. However, the financial incentives are only part of a grander program of education and support to sensitise people to the conservation message, he says. Eventually, the plan is to stop payments altogether. ‘Payment will be reduced as education and awareness is increased to the point where it's phased out,’ he says.Marine biologist Steve Trott agrees. He is project coordinator for the Local Ocean Trust, a charity-based conservation organisation operating in the Watamu and Malindi Marine Parks and Reserve in Kenya  in Costa Rica . In the


Physicians often recommend to older adults that they should engage in mentally stimulating activity to reduce the risk of dementia. But is this recommendation based on sound evidence? Physicians now commonly advise older adults to engage in mentally stimulating activity as a way of reducing their risk of dementia. Indeed, the recommendation is often followed by the acknowledgment that evidence of benefit is still lacking, but “it can't hurt.” What could possibly be the problem with older adults spending their time doing crossword puzzles and anagrams, completing figural logic puzzles, or testing their reaction time on a computer? In certain respects, there is no problem. Patients will probably improve at the targeted skills, and may feel good—particularly if the activity is both challenging and successfully completed.But can it hurt? Possibly. There are two ways that encouraging mental activity programs might do more harm than good. First, they may offer false hope. Second, individuals who do develop dementia might be blamed for their condition. When heavy smokers get lung cancer, they are sometimes seen as having contributed to their own fates. People with Alzheimer disease might similarly be viewed as having brought it on themselves through failure to exercise their brains.Three types of evidence are cited to support the idea that mental exercise can improve one's chances of escaping Alzheimer disease.Having more years of education has been shown to be related to a lower prevalence of Alzheimer disease in cross-sectional, population-based studies In epidemiological studies, people cannot be randomly assigned to different levels of education, or to different kinds and levels of participation in leisure activities. Consequently, researchers must try to identify confounders and take them into account analytically. However, uncertainties remain. Both education and leisure activities are imperfect measures of mental exercise. For instance, leisure activities represent a combination of influences. Not only is there mental activation, but there may also be broader health effects, including stress reduction and improved vascular health—both of which may contribute to reducing dementia risk Another problem with these epidemiological studies is that reverse causation could be involved—in other words, that incipient dementia could be causing reduced engagement in leisure activities, although some prospective studies have been particularly attentive to controlling for this possibility Many studies support the possibility of enhancing memory and other cognitive performance, or of slowing cognitive decline in older adults without dementia These limitations are evident in one of the largest randomized controlled trials of cognitive training with older adults, a large, multisite study named ACTIVE  The third type of evidence suggesting that mental exercise may help to prevent Alzheimer disease comes from neurobiology studies that show greater brain complexity in those with higher levels of mental activity. Many such studies, done with animals, show greater neural complexity after having been exposed to an enriched environment that provides lots of stimulation, for example by including wheels, tunnels, toys, and gnawing sticks The concept of cognitive reserve is often used to explain why education and mental stimulation are beneficial. The term cognitive reserve is sometimes taken to refer directly to brain size or to synaptic density in the cortex. At other times, cognitive reserve is defined as the ability to compensate for acquired brain pathology. This definition encompasses coping skills as well as recruitment of other brain areas, with cognitive reserve thus accounting for individual differences in severity of cognitive dysfunction when there are pathological neural changes. People with a higher level of education have greater cognitive reserve. In some studies, education or occupation are even used as proxy measures of cognitive reserve, while others are beginning to measure neural substrates that correspond to reserve Taken together, the evidence is very suggestive that having greater cognitive reserve is related to a reduced risk of Alzheimer disease. But the evidence that mental exercise per se can increase cognitive reserve and stave off dementia is weaker. Epidemiological studies suggest that individual differences in cognitive reserve may actually be lifelong. In addition, people with greater cognitive reserve may choose mentally stimulating leisure activities and jobs, leading to a chicken-and-egg dilemma for the interpretation of the relationship between mentally stimulating activities in adulthood and dementia risk. Cognitive training has demonstrable effects on performance, on views of self, and on brain function—but the results are very specific to the skills that are trained, and it is as yet entirely unknown whether there is any effect on when or whether an individual develops Alzheimer disease. Further, the types of skills taught by practicing mental puzzles may be less helpful in everyday life than more prosaic “tricks,” such as concentrating, or taking notes, or putting objects in the same place each time so that they won't be lost.So far, we have little evidence that mental practice will help prevent the development of dementia. We have better evidence that good brain health is multiply determined, that brain development early in life matters, and that genetic influences are of great importance in accounting for individual differences in cognitive reserve and in explaining who develops Alzheimer disease and who does not. At least half of the explanation for individual differences in susceptibility to Alzheimer disease is genetic, although the genes involved have not yet been completely discovered For older adults, health practices that could influence the brain include sound nutrition, sufficient sleep, stress management, treatment of mood or anxiety disorders, good vascular health, physical exercise, and avoidance of head trauma. But there is no convincing evidence that memory practice and other cognitively stimulating activities are sufficient to prevent Alzheimer disease; it is not just a case of “use it or lose it.”


Genetically encoded fluorescent calcium indicator proteins (FCIPs) are promising tools to study calcium dynamics in many activity-dependent molecular and cellular processes. Great hopes—for the measurement of population activity, in particular—have therefore been placed on calcium indicators derived from the green fluorescent protein and their expression in (selected) neuronal populations. Calcium transients can rise within milliseconds, making them suitable as reporters of fast neuronal activity. We here report the production of stable transgenic mouse lines with two different functional calcium indicators, inverse pericam and camgaroo-2, under the control of the tetracycline-inducible promoter. Using a variety of in vitro and in vivo assays, we find that stimuli known to increase intracellular calcium concentration  and synaptic and sensory stimulation) can cause substantial and rapid changes in FCIP fluorescence of inverse pericam and camgaroo-2. Winfred Denk and colleagues succeed in generating transgenic mice that express one of two calcium indicators in their cells, creating a valuable tool to study neuronal activity Two classes of genetic Ca2+ indicators have been designed that use different mechanisms of action. The first class, called “cameleons”  regulation system (for a review see 2+]-depen-dent fluorescence changes, and inducibility. The FCIPs were flash pericam, inverse pericam (IP), G-CaMP, camgaroo-2 (Cg2), and the cameleons YC 2.12 and YC 3.12 (n = 5 cells) and IP (n = 2 cells), respectively  were used for analysis of expression patterns by immunohistochemistry . In high- and moderate-expression lines  mice, but there it had very different, much broader emission spectra  and one moderate-expression line MTH-Cg2-14), using confocal imaging spectrometry with excitation at 488 nm. The emission spectra of smooth and punctate fluorescence were similar to each other and to Cg2-expressing HEK cells . Imaging up to and occasionally beyond a depth of 500 μm was possible . Densely2+ response properties of FCIPs using a variety of different preparations and stimulation methods.Next we tested the CaMainly to test temporal response characteristics, we performed a series of somatic electrical recording and synaptic stimulation experiments on pyramidal cells in brain slices. Targeted whole-cell tight-seal recordings of FCIP-expressing layer-2/3 cortical cells (the cell identity was confirmed by the overlap of fluorescence from the FCIP and that from Alexa 568, which was contained in the pipette) showed that FCIP-expressing cells have normal electrophysiological properties A. SomatiFirst, cortical slices from a Cg2 animal (MTH-Cg2-14) were imaged using a CCD camera. Stimulation effectiveness was monitored in a distantly located soma (≈200 μm) by whole-cell tight-seal recording C. Short 2+ responses were seen (eight cells in two retinas tested), consistent with YC3.12 results in other tissues. In lines expressing Cg2 (MTH-Cg2-14), fluorescence became too weak and bleached too quickly for optophysiological measurements . In one of two IP-expressing mice tested , a subset of ganglion cells was strongly labeled see K but no endrites C and 7D.FCIPs are expressed in the olfactory bulb in afferent sensory axons and granule cells, with relative expression levels varying somewhat with FCIP type and across lines (data not shown). Substantial changes in fluorescence were observed for both IP and Cg2 (MTH-IP-12 and MTH-Cg2-19) in response to odor stimulation . We do n2+] dynamics in sensory afferents  rather than [Ca2+]. It is, however, unlikely that the changes we saw are dominated by pH effects for the following reasons. In the case of Cg2, stimulation-induced pH changes  certainly does not apply to IP and Cg2 since we found large responses even in mice aged 8–12 wks, which had been expressing indicators since the onset of αCamKII expression before birth. However, even in strongly expressing smooth-fluorescence lines, a substantial fraction of the indicator protein was found to be immobile and potentially nonfunctional at various ages. It is surprising that punctate fluorescence occurs predominantly in weak lines since precipitation typically occurs at high concentrations. It could, however, be that a limited number of binding and sequestration sites for FCIPs exist in the cell and that only after these sites are saturated does the accumulation of mobile see J, cytosog motifs  that lacromoters  but not romoters . This isromoters . SimilarOne of the remaining problems hampering imaging of population activity with FCIPs is that expression, while cell-type specific, is not complete, i.e., not in every cell of one type, even in lines that express FCIPs at high levels. One possible explanation is position effect variegation (PEV), which occurs when a transgene integrates adjacent to a heterochromatin domain in the genome. In such a situation, expression of the locus variegates, being active in some cells and silent in others . If this2+ indicators in mice, albeit for an unexpected reason. Unlike other promoter systems, such as the CMV promoter, which also appears to resist gene silencing, the TET system allows cell specificity via the expression of the transactivator, which appears to be controllable by cellular promoters without gene silencing.In any case, it appears that the use of the TET system allows the expression of genetically encoded Ca2+ indicators opens the way for the measurement of neural activity patterns in mammals in vivo and in vitro. While the sensitivity of genetic indicators does not (yet) quite reach those of synthetic compounds, it is sufficient for single-trial measurements at least in some applications  and FL were cloned into a Ptetbi vector . The resulting plasmids (Ptetbi-FL/FP, Ptetbi-FL/IP, Ptetbi-FL/CaMP, Pbi-FL/Cg2, Ptetbi-FL/YC2.12, and Ptetbi-FL/YC3.12) were sequenced and transfected into HeLa cells that stably express tTA   were fixed in 4% paraformaldehyde in PBS for 4 h and washed twice with PBS. Brain slices were cut to a thickness of approximately 70 μm using a vibratome . Distribution of Calontech)  and the http://rsb.info.nih.gov/ij/) and IgorPro (Wavemetrics, Lake Oswego, Oregon, United States).All two-photon measurements described in the following sections were done using custom-built two-photon microscopes. Fluorescence was two-photon-excited by a mode-locked Ti-sapphire laser (Coherent Mira F900, 930 nm, 100 fs, 78 MHz) coupled into a custom-built imaging system. The objective used was a 40X/0.8 NA water immersion lens (Nikon, Tokyo, Japan). A photomultiplier-based whole-field detector  detectedRegions rich in neurites were repeatedly scanned in the two-photon microscope. After a first bleaching run  from mice  were prepared according to published procedures for hippocampal experiments  and for Acute brain slices (see above) were maintained in ACSF. Whole-cell tight-seal recordings were made using pipettes made from borosilicate glass (5–10 MΩ) containing 135 mM K gluconate, 4 mM KCl, 10 mM HEPES, 10 mM Na2-phosphocreatine, 4 mM Mg-ATP, and 0.3 mM Na-GTP. Recording pipettes also contained 0.2% biocytin and 1–5 μM Alexa 568, which is spectrally distinct from the FCIPs and can be used to unambiguously identify the recorded cell. For synaptic stimulation we used saline-filled glass electrodes or tungsten microelectrodes (1 MΩ) (World Precision Instruments, Berlin, Germany). Synaptic transmission was blocked in some experiments by the addition of 40 μM 6-cyano-7-nitroquinoxaline-2,3-dione and 100 μM 2-amino-5-phosphovaleric acid.Wide-field images were taken with a MicroMax, 512 × 512 back-illuminated CCD camera (Roper Scientific, Tucson, Arizona, United States) binned 5 by 5. For experiments involving extracellular stimuli we calculated the change in fluorescence relative to the prestimulus period (ΔF/F) for each binned pixel frame by frame. The prestimulus period of regions of interest was fitted with an exponential curve, which was then subtracted from the entire fluorescence time course to correct for bleaching. No corrections were made for autofluorescence, so that the relative FCIP fluorescence changes are likely to be larger. When analyzing action-potential-induced signals from individual neurons, we calculated the fluorescence changes by subtracting the average of two nearby areas from the total fluorescence to account for background fluorescence  to record baseline fluorescence (100 pulses at 100 Hz or 20 pulses at 100 Hz). Image sequences were 6 or 12 s long. The background level (average intensity with the laser off) was subtracted from every frame in the image sequence. The average of the five prestimulus (rest) images was subtracted from the average of five “response” images (response minus rest). The built-in smoothing function of ImageJ was used to reduce the noise further. The brightness of the difference image was enhanced for better display. Localized small fluorescent structures or hot spots in neurites were identified visually. For IP, traces with high time resolution were acquired using 64-pixel line scans at 500 Hz. Fluorescence was averaged over the width of the soma. The fluorescence from a neighboring region was subtracted to account for background. Fluorescence changes (percent ΔF/F) were calculated relative to the resting fluorescence.Mice were dark-adapted for several hours before the experiments and all subsequent procedures were carried out under dim red illumination to minimize photobleaching. Animals were anesthetized with halothane and subsequently killed by cerebral dislocation or by intraperitoneal injection of pentobarbital. Immediately afterward, both eyes were removed and dissected free in Ames medium (Sigma-Aldrich). A piece was cut from a retina and placed photoreceptor side down into the recording chamber, and maintained at 35 °C in Ames medium continuously perfused with oxygen. The remaining retina was kept for further use.2+-mediated fluorescence (emission 520 BP 30 nm) changes in retinal ganglion cells using a custom-built two-photon microscope. The laser (Coherent Mira 900F) was tuned to 925 nm (for YC3.12, the laser was tuned to 870 nm, see The stimulation and imaging procedures were as described elsewhere . In brieMice were anesthetized and dissected as previously described . Odors wMice were anesthesized with urethane (1.5 mg/g) and body temperature was maintained at 37 °C. For two-photon imaging, a custom-built headplate with an imaging window (4 mm × 3 mm) was glued to the top of the skull using cyano-acrylate (UHU, Buhl-Baden, Germany) and attached to a fixed metal bar before thinning of the skull. The combination of rigid headplate and thinned-skull reduces respiration and cardiac-pulsation-induced brain motion. The microscope objective was positioned at an angle so that the optical axis was perpendicular to the surface of the cortex.


Accidental poisoning is one of the leading causes of injury in the United States, second only to motor vehicle accidents. According to the Centers for Disease Control and Prevention, the rates of accidental poisoning mortality have been increasing in the past fourteen years nationally. In Texas, mortality rates from accidental poisoning have mirrored national trends, increasing linearly from 1981 to 2001. The purpose of this study was to determine if there are spatiotemporal clusters of accidental poisoning mortality among Texas counties, and if so, whether there are variations in clustering and risk according to gender and race/ethnicity. The Spatial Scan Statistic in combination with GIS software was used to identify potential clusters between 1980 and 2001 among Texas counties, and Poisson regression was used to evaluate risk differences.Several significant (p < 0.05) accidental poisoning mortality clusters were identified in different regions of Texas. The geographic and temporal persistence of clusters was found to vary by racial group, gender, and race/gender combinations, and most of the clusters persisted into the present decade. Poisson regression revealed significant differences in risk according to race and gender. The Black population was found to be at greatest risk of accidental poisoning mortality relative to other race/ethnic groups (Relative Risk (RR) = 1.25, 95% Confidence Interval (CI) = 1.24 – 1.27), and the male population was found to be at elevated risk (RR = 2.47, 95% CI = 2.45 – 2.50) when the female population was used as a reference.The findings of the present study provide evidence for the existence of accidental poisoning mortality clusters in Texas, demonstrate the persistence of these clusters into the present decade, and show the spatiotemporal variations in risk and clustering of accidental poisoning deaths by gender and race/ethnicity. By quantifying disparities in accidental poisoning mortality by place, time and person, this study demonstrates the utility of the spatial scan statistic combined with GIS and regression methods in identifying priority areas for public health planning and resource allocation. The accidental poisoning mortality rate has been increasing in the United States. In the 21-year period spanning 1981 to 2001, mortality rates due to accidental poisoning more than doubled from 2.0 per 100,000 in 1981 to 4.9 per 100,000 in 2001 [Accidental poisoning refers to the physiologic damage caused by inhalation, ingestion, or any other mode of exposure to various licit and illicit drugs, or chemicals such as those found in pesticides, household cleaning products and gases/vapors . AccordiMany of the studies regarding poisoning mortality have examined exclusively excess mortality due to long-term exposure to specific agents and have not explicitly studied the possible persistence of mortality due to accidental poisoning across space and time. They often examined either spatial or temporal attributes of the burden of the disease. A statistic for examining spatial and temporal data concurrently is available through the use of SaTScan software. The objectives of this study are to detect the existence of spatial and temporal clusters of accidental poisoning mortality in Texas and to determine if there are variations with respect to race and gender among these clusters.From 1980 to 2001, there were 10,774 deaths attributable to accidental poisoning, resulting in an average annual age-adjusted mortality rate of 2.8 deaths per 100 000. From 1981 to 2001, a linear increase was observed in unintentional poisoning mortality rates among Texas counties  the purely spatial scan statistic and 2) the space-time scan statistic non-parametrically adjusting for time. The results of these additional analyses indicated that most clusters detected using the default method were in mostly identical regions detected in the earlier analysis, only with slightly lower relative risks (i.e., observed deaths/expected deaths). For instance for the "all population group", the detected primary cluster in West Texas with a relative risk of 2.4 remained a cluster, but reduced to relative risks of 2.2 and 2.1 using additional methods that adjusted for time trend. In addition, most of the detected clusters still persisted to the present decade after the adjustments of time trend. Similar results were observed for most clusters detected in 'Male", "Female", "Black", "Non-Hispanic Whites" and "Hispanic" subpopulations using additional analyses options  identify most clusters in almost identical regions but with slightly lower relative risks. This observation, along with the fact that both the results of the space-time scans with and without adjusting for time non-parametrically both demonstrate the temporal persistence of accidental poisoning mortality clusters into the present decade , has provided supporting evidence that the clusters detected using the spatial-temporal method are geographic in nature, rather than an artefact of temporal trend.The burden of excess accidental poisoning mortality was found to be highest in the Black population, followed by the non-Hispanic White population, and the least among the Hispanic population and populations of other race/ethnicity categories. Consistent with the literature, the male population also exhibited an elevated risk of accidental poisoning mortality when compared to the female population.The results of this research presented both agreement and disagreement when compared other the results in which conventional techniques were used to identify geographic disparities by race/ethnicity, and gender. For instance, the results of this study were consistent with the literature in that the Black population was found to be at greatest risk of death from accidental poisoning ,11. In aAlthough there are few studies in the literature that explicitly record cluster detection for accidental poisoning mortality, there are a variety of studies that have examined trends of accidental poisoning mortality in space and time -5,11,14.There is also evidence in the literature to support the existence of temporal clustering of accidental poisoning mortality. In a study on organophosphate poisoning, Sahin, Sahin and Arabaci reported that deaths from accidental poisoning due to organophosphates were most frequent during certain months . In a reThe results of this study should be interpreted with several considerations. One, the process of cluster detection is necessarily ecological. The objective was to determine if there was an association between certain areas of Texas with excess accidental poisoning mortality. These results cannot be extrapolated to the level of the individual, i.e., one cannot interpret a relative risk above one as increased risk of accidental poisoning mortality for residents living in a given county. However, the risk estimate does provide valuable information about geographic disparity of accidental poisoning mortality.Another consideration concerns the utilization of county-level data. The scan statistic may not have been sensitive to small areas of excess mortality that may have been detected at a higher geographic resolution. However, in Texas the county is the smallest geographic area for which there is routinely reported or available deaths and population estimate information. Also, the county is the political level at which health policy actions are instituted, since many of the local public health departments in Texas are county health departments.th revision, and there may be differences in mortality estimates based on the codes in the 9th and 10th revisions. This introduces the possibility of misclassification bias in the results. However, there is no evidence to indicate that the misclassification would be differential [A third consideration is that this study relied on county-level mortality data; and there may be variations in coding or reporting practices from county to county. Furthermore, in 1998 the International Classification of Diseases underwent its 10erential . Hence, This study has demonstrated the existence of spatiotemporal clusters of accidental poisoning mortality in Texas. Additionally, this study has also shown variations in risk and clustering by gender and race/ethnicity. The results of this study have numerous implications. By quantifying accidental poisoning mortality disparities by geographic area, race/ethnicity, and gender, the results provide an evidence-base for health planning. The clusters identified in this study, specifically those that persist into the present decade, represent areas where health services, e.g., poison control centers and emergency rooms, may be deficient. Knowing the specific areas of excess mortality from accidental poisoning would help health policymakers to focus the scope of prevention programs and health care delivery, thus providing for efficient allocation of public health resources.The findings from this study illustrate the use of the scan statistic and GIS in public health surveillance. Longitudinal data for accidental poisoning mortality was analyzed to identify hotspot clusters not only for the total population but also for subpopulations. Identifying geographic areas and specific populations, which are at an elevated risk of accidental poisoning mortality helps in the process of hypothesis generation for possible etiological mechanisms. For example, the most likely cluster for the total Texas population is in a heavily industrial area. The population in this area may have experienced excess mortality from other occupational exposure-related health outcomes (e.g. different types of cancers). One may further examine into the excess accidental poisoning mortality observed in this area in order to determine if it was due to occupational exposures.The clusters identified in this study warrant further attention. Additional studies should be conducted to determine the causes behind the observed clustering, especially in the Gulf Coast region and the Central North Texas region where clustering was observed for the total Texas population and the two populations at greatest risk of accidental poisoning mortality, the Black population and the White population. More research is needed to assess different types of toxic exposures and their contribution to accidental poisoning mortality. Furthermore, the availability, locality and accessibility of health services such as poison control centers and health care facilities  should be studied, giving special attention to those in non-urban counties.). The data was extracted as counts by race, gender and age-group for ICD-9 codes 850 – 858  and 860–869  for the years 1980 – 1998 and ICD-10 codes X40 – X49 . The ICD-10 classification codes include exposure to drugs, gases and vapors, and other unspecified chemicals. Second, population (and estimates) files by year were obtained from the Texas State Data Center. Both deaths and population data included stratified information by age groups (16, representing the ages of 0–4 to 75 and above), gender  and race (4, representing Blacks, Hispanics, Non-Hispanic Whites and Others). Third, county centroids information of latitude and longitude was downloaded from the 2000 U.S. Gazetteer portion of the US Census Bureau (Available online at ).Three types of files were downloaded for the analysis. First, death files including the unintentional poisoning mortality data was obtained from Expert Health Data Programming Incorporated's Vitalweb (URL ) to perform scan analysis, and extracted cluster information from the SaTScan output file. Following this, the program linked the cluster dbase files with the county name file, and created a new file for mapping purposes. Using ArcGIS software (ESRI, ), we spatially joined the dbase file to a Texas county layer and produced maps displaying the poison mortality clusters of all populations and those in the populations stratified by gender, race/ethnicity, and gender-race/ethnicity combinations.We performed the data processing by using an automated Microsoft Visual Basic Application (VBA) program to organize the downloaded county information of death counts, population at risk and geographic files (i.e. county centroids), before exporting them to SaTScan for analysis and for GIS mapping. The VBA program first imported, aligned, and then exported data tables to a SatScan compatible format. The program then invoked the SaTScan batch executable file . To control for overdispersion, the Poisson model was used with the scaled deviance option. Estimates of the annual percent increase, and rate ratios for race and gender were calculated from the results of the analysis.To detect potential spatiotemporal variations of poisoning mortality among Texas counties, this study utilized the Spatial Scan Statistic developed by Kulldorff ,17 to deMany studies conducted on rare events such as unintentional poisoning deaths have aggregated the events to a higher geographic resolution, and have thus been unable to detect the existence of true clusters. The spatial scan statistic, however, is ideal for cluster detection of rare events since it includes the option of examining Poisson distributed data.The space-time parameter of the spatial scan statistic places a cylindrical window on the coordinates grid for the locations studied and moves the center of the cylinder base over the grid so that the sets of geographic units covered by the window are constantly changing. The statistic assumes no predetermined cluster size and utilizes scan windows of various sizes (of the population at risk) with a maximum window size specified by the user. In the space-time scan, this spatial scan window serves as the base of a cylinder, with time acting as the height of the cylinder. The cylinder is continuously expanded and extended up to the maximum specifications set by the user. Whenever the cylindrical window finds a new death, SatScan calculates a likelihood function to test for elevated risk within the cylinder as compared with outside the cylinder. The likelihood function for any given cylindrical window (under the Poisson assumption) is proportional to:d/n)d([D - d] / [D - n])D - d)(I     (1) the purely spatial scan statistic and (2) the space-time scan statistic  adjusting for time.ETN was responsible for the design and implementation of the project and the final paper. ETN also contributed to data analysis. CEH contributed to database design, visual basic application programming and data analysis. CEH also assisted in the preparation and editing of the final paper. VIH contributed to the preparation and editing of the final paper, including reviewing the relevant literature. AMH was responsible for the layout and design of the illustrative maps and also assisted in preparing and editing the final paper. All authors read and approved the final manuscript.


Although a rare disease, uveal melanoma is the most common primary intraocular malignancy in adults, with an incidence rate of up to 1.0 per 100,000 persons per year in Europe. Only a few consistent risk factors have been identified for this disease. We present the study design of an ongoing incident case-control study on uveal melanoma (acronym: RIFA study) that focuses on radiofrequency radiation as transmitted by radio sets and wireless telephones, occupational risk factors, phenotypical characteristics, and UV radiation.We conduct a case-control study to identify the role of different exposures in the development of uveal melanoma. The cases of uveal melanoma were identified at the Division of Ophthalmology, University of Essen, a referral centre for tumours of the eye. We recruit three control groups: population controls, controls sampled from those ophthalmologists who referred cases to the Division of Ophthalmology, University of Duisburg-Essen, and sibling controls. For each case the controls are matched on sex and age (five year groups), except for sibling controls. The data are collected from the study participants by short self-administered questionnaire and by telephone interview. During and at the end of the field phase, the data are quality-checked.To estimate the effect of exposures on uveal melanoma risk, we will use conditional logistic regression that accounts for the matching factors and allows to control for potential confounding. Although a rare disease, uveal melanoma of the eye is the most common primary intraocular malignancy in adults, with an incidence rate of up to 1.0 per 100,000 person years (age-standardized, world standard) in Europe ,3. AnothSome uveal melanoma are associated with neurofibromatosis. However, the vast majority of familial cases reported are non-syndromic . Some reOccupation may be also relevant, and may include chemical work ,12, arc The population-wide introduction of analog and digital mobile phone techniques in the recent years, which has been coined as the mobile revolution , has resUntil now, the majority of epidemiological cancer studies focussed on brain cancer because the brain may be exposed to RFR -20. WithThe RIFA study is planned to answer several etiologic questions with a special focus on electromagnetic radiation especially radio-frequency radiation as emitted by mobile phones and radio sets. First, is the finding of an increased risk of uveal melanoma among subjects with frequent use of RFR devices reproducible? Second, if there is an association, is this association site-specific in terms of laterality of the uveal melanoma and major site of mobile phone use? Third, if there is an association, is there a dose-response relationship between RFR and uveal melanoma risk? Another set of etiologic question relates to pigmentation characteristics including iris colour, hair colour, tendency of the skin to burn and to tan, freckling, number of cutaneous nevi. A further study questions relates to exposure to work and leisure time related ambient ultraviolet radiation and uveal melanoma risk. In addition, our study focuses on several occupational exposures or jobs that are suspected to be associated with an increased risk of uveal melanoma ,15 incluth, 2002 to September 24th, 2004 are eligible, if they are referred to the Division of Ophthalmology, University of Duisburg-Essen during the recruitment period, are aged 20–74 years at diagnosis, are living in Germany, and are capable to complete the interview in German. The majority (about 70–80%) of uveal melanoma treated at the University Hospital Essen receive episcleral plaque therapy without histological verification. For this reason we did not include a reference pathologist who reviews the diagnostic certainty of the cases. Experiences from our previous case-control studies showed that there is a nearly perfect agreement between the local eye doctors in the reference centre and the international reference pathologist (Dr. Ian Cree, London) [The case recruitment is hospital-based and takes place in the Division of Ophthalmology, University of Essen which is a the referral centre for eye cancer in Germany, currently treating about 400–500 eye cancer patients per year. Eligible uveal melanoma cases have to fulfil several criteria. Patients with newly diagnosed first uveal melanoma located in the choroid, iris, and/or ciliary body  during t London) .We considered a diagnosis of uveal melanoma as definite if the results of the clinical examination of the eye  and ultrasound (sometimes supplemented by fluorescence angiography, computertomography or magnetic resonance imaging) were unambiguous. The inclusion and exclusion criteria of the cases are listed in table th, 2002 through March 31th, 2004. Assuming comparable incidences of uveal melanoma in the federal states of Germany, the crude rate (referred cases divided by population at risk aged 20–74 years) may be considered as an indicator of the referral effect.Interim analyses showed that the majority of cases comes from the territory of former West Germany. Figure Obviously, the referral effect varies by federal states. However, it is difficult to judge whether the case referral to the Division of Ophthalmology, University of Duisburg-Essen is a random sample of all newly diagnosed uveal melanoma cases in Germany. We therefore decided to recruit three different control groups. First, if we assume that cases treated in Essen are a random sample of all cases in Germany, a population-based control group would be the most appropriate control group. For this approach, we randomly select controls from mandatory lists of residence that cover the total population of the city or local district. These lists are regarded as the most complete sampling frame for population-based studies in Germany.Second, if the referred cases are not a random sample of all newly diagnosed cases of uveal melanoma in Germany, a control group sampled from those ophthalmologists who referred cases to the Division of Ophthalmology, University of Duisburg-Essen, would be most appropriate ,26. To iIn addition, we recruit sibling controls of cases (matching ratio 1:1) in order to assess whether genetic factors may confound the effect of exposure. The sibling controls are matched in genetic background. The inclusion criteria of the three control groups are presented in table Based on our former uveal melanoma case-control studies ,22 we esPopulation-based prevalence estimates of mobile phone use in the general population are scant. A recent telephone survey from 2001 showed that 82% of male and 74% of female participants aged 14–44 years use mobile phones; within the age group 45–59 years, 63% of male and 58% of female participants use mobile phones. The oldest age group (> = 60 years) shows considerable lower prevalences of mobile phone use (men: 49%, women: 25%) . To deteTable The questionnaire on mobile phone use is the same instrument which has been used by the international case-control study on brain cancer and mobile phone use sponsored by the International Agency for Research on Cancer, called Interphone study . In contCompared to other studies on the aetiology of uveal melanoma, the RIFA study uses a detailed assessment of pigmentary characteristics. The self-administered questionnaire contains a eye and hair colour card that allows the participants to choose the most appropriate colour of their eyes and hairs at age 20 years. During the telephone interview, the skin reaction to sun exposure (tanning ability, burning tendency) is asked according the concept of Fitzpatrick . FitzpatFor the assessment of nevi of the upper arms and the dorsum of the feet with a diameter of at least 3 mm, participants receive a template with a 3 mm hole that enables them to count all nevi of this minimum size. In addition, the CATÍ (computer assisted telephone interview) includes a detailed history of sunburns in the recent 15 years before interview and tendency to freckle as a child. The CATI contains a question on eye colour with the typical categorical answers as has been used by several others (blue, grey, green, hazel, brown, black,  that wilThe course of the exposure assessment is displayed in figure For subjects who reported selected work tasks (e.g. cooking and food processing, welding and others), we use 16 job-specific supplementary questionnaires to obtain details of the job tasks and materials used.The quality control program includes several procedures. The study is designed to fulfill the recommendations of the German Good Epidemiologic Practises .Eight month before the main study started, a manual of standard operating procedures (MOP) was written and a pilot study of four weeks was conducted to test the field work and exposure questionnaires. After some minor revisions of the MOP, report forms, and questionnaire instruments, the principal investigator and participating epidemiologists had to sign that they fully agree with the final version of the MOP.Interviewers of the study were introduced into the field work and were blinded against our study hypotheses. After an initial interviewer training course, interviewers are regularly monitored and receive regular training courses.The recruitment progress, given as number of registered cases and controls, distribution of inclusion and exclusion criteria, response proportions, is monitored monthly. The analysis of nonresponse reasons is supplemented by an short questionnaire for subjects not willing to participate. This questionnaire includes few demographic and exposure items that help us to assess potential selection effects due to nonresponse.A plausibility control of the interview data is done quarterly and is the basis for the regular training courses of the interviewers.The completeness of case registration is checked by regular comparison of the list of registered cases with lists of admissions to the referral centre. In addition we compare our list of cases with data of the hospital information system that includes information on diagnoses.The self-administered short questionnaires are visually edited by the study personnel before the telephone interview starts. The visual editing includes a completeness check and coding of the life-long job history. For each job period, the occupation and branch of industry is coded according to ISCO-68  and NACEThe CATI contains internal quality checks that prevent data entry errors. For example, interviewers are not able to fill in the detailed questions on mobile phones, if the entry question on ever having used mobile phone has been answered with no.At the end of the field phase, the data are quality-checked. To estimate the effect of exposures on uveal melanoma risk, we will use conditional logistic regression that accounts for the matching factors and allows to control for potential confounding. We will classify people exposed to an occupational category if they ever worked within this category for at least six months. The quantification of mobile phone use will be based on average number of phone calls and average duration of phone calls per time unit. The association between pigmentary characteristics and uveal melanoma risk will be assessed by detailed matrix containing information on hair colour at age 20 years, eye colour, freckling tendency, and skin colour. Final results of these analyses are scheduled to be published in summer 2005.None declared.The pre-publication history for this paper can be accessed here:


The MidSouth Computational Biology and Bioinformatics Society (MCBIOS) describes its efforts to provide local opportunities for researchers to learn and connect with colleagues The rich get richer while the poor get poorer. This is as true for research in the life sciences as it is for society in general. Why is this? To put it simply: because existing resources can be leveraged to generate new ones. In life science research, these resources are often expressed in terms of people, equipment, and funding. Those who have more in these areas—large research organizations, support staff, extensive facilities, and pipelines of funding—can be more productive, and thereby more successful in maintaining and even expanding their infrastructure. In the United States, this “scientific wealth” is concentrated in certain geographic areas, particularly along the east and west coasts. While there are many highly competent individuals in other, less wealthy, regions, they often struggle to attract funding because they lack this infrastructure.Fortunately, scientific progress is also made through collaboration and cooperation—activities fostered through the exchange of viewpoints and the exploration of ideas. Given a framework in which this networking can occur, some of the problems that result from a lack of infrastructure can be overcome. By definition, collaboration involves the sharing of ideas, capabilities, and resources. While it may not be possible to access all necessary resources locally, finding partners—who have their own local capabilities and resources—can enhance one's own research environment. Through these kinds of efforts, “virtual infrastructures” grow. And through collaborative efforts, those of us in some of the “poorer” regions of the United States have opportunities to successfully compete for national sources of funding by becoming better able to address the “existing research environment” consideration on most grant applications.www.MCBIOS.org), created to serve a geographical area that includes Arkansas, Louisiana, western Tennessee, Missouri, Mississippi, Oklahoma, and east Texas. By its very nature, bioinformatics involves people from many different backgrounds and is therefore ideal for collaborative efforts. Like many other regional societies, our primary goal is to provide a framework in which collaboration and cooperation can occur. By sponsoring regional activities  we hope to bring educators, researchers, and especially students together with others who have similar and/or complementary interests. Through the society, not only can medical scientists make contact with computational experts, but researchers in Arkansas can connect with researchers in Oklahoma, educators from Missouri can interact with educators from Louisiana, and students from Mississippi can find others from Tennessee who are working in the same area. Communication across specialties and areas of expertise is especially important for fostering interdisciplinary efforts and exposing students to a broader range of topics than might be available at their own institution. Nowhere is this truer than in the application of informatics to a variety of disciplines.Building a virtual infrastructure is the primary purpose behind the formation, in the United States, of the MidSouth Computational Biology and Bioinformatics Society , for example, has recognized MCBIOS under their “regional affiliate” program. Likewise, MCBIOS is encouraging its members to form “local chapters” (for example, the Oklahoma chapter of MCBIOS, www.OKBIOS.org). These local chapters are eligible to host the annual MCBIOS conference and are able to support even closer interactions among their local participants. The hierarchy of affiliations provides an abundance of opportunities for members to participate based upon their interests, financial resources, and tolerance for travel.Regional societies need not be in competition with national or international societies. In fact, many of these larger organizations are finding that it is to their advantage to encourage close relationships with their regional counterparts. The International Society of Computational Biology , we have deliberately chosen locations for the MCBIOS annual conference that would be within a day's drive for all members. At the inaugural 2003 MCBIOS conference, for example, with a theme of “Building Networks,” a number of computer scientists attended who were able to find out more about bioinformatics and their possible contribution to life science research; they would probably not have been able to make the investment to attend a national or international meeting on bioinformatics.While still maintaining high standards, regional events also increase the number of venues for the presentation of research results and creative efforts in the educational arena. While the ISCB program, which is already highly self-selecting, accepts less than 15% of their submissions, almost all submissions at the first annual MCBIOS conference could be accommodated, at least in poster form. These regional events can also attract well-respected keynote speakers and provide training opportunities that might not otherwise be available to the membership. For example, Dr. Alan Leshner, CEO of the American Association for the Advancement of Science, gave the keynote address of the 2004 MCBIOS conference. In 2003, we were also able to host a special training session by the National Center for Biotechnology Information on their GenBank and molecular biology tools.While MCBIOS is still new, we have plans to extend our activities—and thus our impact—beyond our annual conference. We are dedicated to supporting our local chapters, and have plans to develop a speakers' bureau to serve them. Plans to collaborate on regional technology efforts  and on multi-institutional educational programs are also in the works. Through the auspices of the society, we hope to increase regional credibility and attract national funding in support of these infrastructure improvements.In the long term—and in addition to supporting the intellectual efforts of our members through a vibrant organizational community—our goal is to increase extramural funding for our represented institutions. We hope to achieve this by fostering competitive research. And this competitive research will be possible on a larger scale through our development of a virtual infrastructure—one that comes about through regional collaboration and a pooling of resources. In conjunction with other infrastructure-building efforts , we hope to see externally funded research in the life sciences significantly increase within our region, the American “Midsouth.”http://BRIN.UAMS.edu] of the National Center for Research Resources through a grant to underwrite student participation and awards at the conferences. The 2004 MCBIOS conference was also supported in part by a grant from the National Center for Toxicological Research [www.FDA.gov/NCTR].)(The 2003 and 2004 MCBIOS conferences have been supported in part by National Institutes of Health grant P20 RR-16460 from the Arkansas Biomedical Research Infrastructure Network Program [


We evaluated the impact of ibpAB, which is innocuous under physiological conditions, impaired culture growth during α-glucosidase production. At higher temperatures, accumulation of stress proteins including disaggregation chaperones (DnaK and ClpB) and components of the RNA degradosome, enolase and PNP, was intensified. Overexpression of ibpAB, conversely, suppressed the heat-shock response under these conditions. Inclusion bodies of α-glucosidase started to disaggregate after arrest of protein synthesis in a ClpB and DnaK dependent manner, followed by degradation or reactivation. IbpA/IbpB decelerated disaggregation and degradation at higher temperatures, but did hardly influence the disaggregation kinetics at 15°C. Overexpression of ibpAB concomitant to production at 42°C increased the yield of α-glucosidase activity during reactivation.Deletion of ibpAB, inclusion body removal is faster, but cells encounter more intense stress and growth impairment. IbpA/IbpB thus exert a major function in cell protection during stressful situations.IbpA/IbpB attenuate the accumulation of stress proteins, and – at high temperatures – save disaggregated proteins from degradation, at the cost, however, of delayed removal of aggregates. Without  Escherichia coli can induce several stress responses [Escherichia coli [Production of recombinant proteins as inclusion bodies in esponses -3 and maesponses -6. The mesponses ,8 may reesponses ,9,10. Moesponses ,12. Whilhia coli , the highia coli ,15. The Analogously, incubation at extreme temperatures is characterised by near exclusive heat-shock protein synthesis as a consequence of extensive aggregation of cellular proteins ,19. To dE. coli sHsps, IbpA and IbpB, reversibly dissociate upon prolonged incubation at 50°C from multimers of 2 – 3 MDa to monomers in the case of IbpA and to oligomers of 650 – 700 kDa in the case of IbpB, exposing hydrophobic sites [in vitro and facilitates their subsequent refolding by DnaK [in vitro and in vivo is mediated by ClpB together with DnaK [ibpAB as well as in some cases in strains lacking ibpAB [The small heat-shock proteins (sHsps) are intrinsic holding chaperones of multimeric structure. The multimers dissociate at high temperatures into smaller entities for exposure of client binding sites. After binding of misfolded client proteins, the sHsps form large globular assemblies . Therebyic sites ,25. IbpAic sites , but sho by DnaK -25, a ro by DnaK ,27. IbpA by DnaK ,28. The ith DnaK , can be ng ibpAB ,32,33. Ing ibpAB ,35.ibpAB confers resistance to heat and other stresses [ibpAB operon does not affect growth even at high temperatures. Only recovery from prolonged exposure to extreme temperatures such as 50°C are impaired in an ibpAB mutant, and growth of ibpAB dnaK double mutants at high temperatures [Saccharomyces cerevisiae produced in recombinant E. coli is a suitable model system. It accumulates as inclusion bodies and induces the heat-shock response including prolonged expression of the ibpAB operon [E. coli if produced at low temperature [The role of IbpA/IbpB in cell protection is less clear. Overexpression of stresses , but deleratures ,36. In tB operon . It is, perature , enablinibpAB operon was induced already during α-glucosidase production at 30°C, with moderately higher levels at elevated temperatures  or concomitant to production at lower temperature.Post-induction growth of the res Fig. , althougThus, the sHsp are effective only under conditions that favour inclusion body formation.ibpAB deletion mutant at all temperatures, in addition to the induction at elevated temperatures . While the concentration of α-glucosidase was 10–25% higher in dnaK and clpB deletion mutants than in the wildtype strain, the specific activities obtained during the production phase were less than 0.1 Umg-1, compared to about 0.3 Umg-1 in the other strains (data not shown). Importantly, neither disaggregation, nor reactivation nor degradation took place in these mutants within two hours, and the concentration of α-glucosidase inclusion bodies remained unchanged.The impact of chaperones on the metabolism of α-glucosidase inclusion bodies was examined in wildtype proteins . In the ibpAB, were investigated as a function of temperature. After production of α-glucosidase at 37°C, the cultures were transferred to various temperatures. As protein synthesis was arrested before the transfer, the initial IbpA/IbpB level did not depend on the disaggregation temperature. The α-glucosidase concentration in the insoluble cell fractions of all cultures decreased with time, but with different kinetics. In the ibpAB deletion strain, the disaggregation kinetics were not influenced by the incubation temperature .Disaggregation of α-glucosidase inclusion bodies was accompanied by an increase of the specific α-glucosidase activities in the cell extracts. The reactivation showed a clear temperature optimum in the range of 24 to 30°C, most evident with wildtype levels of IbpA/IbpB Fig. . The spened Fig. . Both, iibpAB deletion mutant at all temperatures . About one third of the α-glucosidase produced concomitant to ins Fig. . Also thins Fig. , resemblins Fig. . Thus, tibpAB overexpression U169 rpsL150 relA1 flbB5301 deoC1 ptsF25 rbsR and JGT17 MC4100 Δ ibpAB [argU tRNA and carrying the lacI repressor gene [HindIII restriction sides, coding for the ibpA and ibpB genes including their native promoter, was cloned into the HindIII site of the plasmid pKK177-3/GLUCP1 [GLUCPI gene and the 5ST1T2 terminator, to give the new plasmid named pKK177-3/GLUCP1_ibpAB, in which the ibpAB operon is under the control of its native promoter and of the tac-promoter upstream of the α-glucosidase gene. Proper orientation of the insert was tested by EarI digestion. Δ ibpAB  were use Δ ibpAB . Plasmidsor gene  was cotrsor gene . The seqsor gene  between 3/GLUCP1 , located-1, chloramphenicol 50 μg mL-1), on a rotary shaker at 37°C to an OD600 of 0.5, induced with 1 mM IPTG and transferred to different temperatures as indicated in the results section. For reactivation experiments, tetracycline was added to a final concentration of 25 μgmL-1 two hours after induction to arrest protein synthesis, and the cultures were transferred to the indicated temperatures.The cultures were incubated on Luria-Bertani (LB) medium, supplemented with appropriate antibiotics .α-glucosidase activity was measured as described . For the600 mL were dissolved in 200 μL lysis buffer containing 8 M urea, 4% CHAPS, 60 mM DDT, 2% Pharmalyte 3–10 and 0.002% bromophenol blue and incubated for one hour at room temperature. The first dimension was carried out on an IPGphor using 24 cm pH gradient 3–10 strips (amersham pharmacia biotech), with the voltage profile recommended by the manufacturer. The second dimension was run in an Ettan Dalt six (amersham pharmacia biotech) on 12.5% polyacrylamide gels. Gels were stained with Coomassie brilliant blue for quantification or by silver staining for visualisation. Gels for cultures incubated at 42°C were performed in quadruplicate, for other temperatures once. Protein spots were identified by comparison with the E. coli 2D database . Spot identities were verified by N-terminal sequencing for ClpB (RLDRLTN) and enolase (SKIVKII). α-glucosidase was not dissolved from the inclusion bodies and is not visible in the gels.Cell pellets from a culture volume of 8/OD


How genomics and related health biotechnologies can improve the health of the poor and contribute towards meeting the Millenium Development Goals Making New Technologies Work for Human Development, identified technical progress as the largest factor in reducing mortality rates and improving life expectancy from 1960 to 1990 Development experts and policy makers agree that investment in science and technology is important for economic growth and development. The 2001 United Nations (UN) Development Programme report, Genomics and Global Health, presented recently to Task Force 10, we addressed how genomics and related health biotechnologies can improve global health and contribute towards meeting the MDGs This capacity is essential if the world is to achieve the UN Millennium Development Goals (MDGs), which were adopted by all UN members in 2000 in a commitment to promote sustainable development and eliminate poverty in the world. As part of the Millennium Project, the UN established task forces to come up with strategies to help developing countries achieve the MDGs. One of these is the Task Force on Science, Technology, and Innovation (Task Force 10), created in recognition of the fact that many of the goals, especially those related to health and the environment, cannot be realized without a strong contribution from science and technology Genomics refers to the powerful new wave of health-related life sciences energized by the human genome project and the knowledge and tools it is spawning. It is a relatively new science that has tremendous potential to address health problems in developing countries.http://www.malariatest.com/]). Recombinant vaccines, a result of genetic engineering, promise to be safer, cheaper, and possibly easier to store and transport than traditional vaccines The role of genomics in global health has been highlighted previously in the World Health Organization's 2002 report Genomics and Global HealthIn The GGI could also facilitate the sharing of good practices. For example, the Canadian Prime Minister, Paul Martin, in his February 2004 reply to the Speech from the Throne, set a long-term target for Canada to devote 5% of its research and development spending to the challenges of developing countries. If successfully implemented and replicated by other industrialized countries, this target could make a real difference to global health.Our report concludes that the key actors are developing countries themselves. We explore how to put genomics and related technologies to work in developing countries within the next Today there are examples of strategies that some developing countries, including China, Cuba, Brazil, India, and South Africa, have followed to institute learning processes that are helping them to build NSIs in biotechnology. China seized the opportunity to take part in the Human Genome Project and quickly set up major institutions in genomics, such as the Beijing Genomics Institute, equipped with state-of-the-art sequencing facilities and computers. It has also followed a strategy of private-sector development in line with the NSI framework. Because of a government policy encouraging their return, Chinese expatriates are increasingly active in setting up small health biotechnology firms, bringing to the country knowledge from the world's major centers for genomics and related technologies.The government in Cuba became interested in biotechnology in the early 1980s when the field was still in its infancy and created an interdisciplinary group, the Biological Front, to explore the possibilities of the technology for Cuba. It has continued to support biotechnology even during periods of economic hardship, set up institutions with research, development, and manufacturing facilities, and encouraged linkages between these institutions by setting up a biotechnology cluster, the West Havana Scientific Pole. Encouraging linkages has been a core policy of the government in Cuba, and its health biotechnology development has benefited from the ties with a strong public health system.Brazil has a relatively long history of supporting science and technology, and the country is increasingly focusing on genomics and related technologies. The lack of commercialization of its cutting edge science and technology has been a weakness of the system in Brazil, but the country is now trying to overcome this weakness by proposing an “Innovation Law” that encourages cooperation between universities and the private sector.Since its independence in 1947, India has followed a vision to improve the quality of life of its people by emphasizing science and technology. Limited resources and a patenting system that did not allow patenting of pharmaceutical products but only patenting of processes encouraged firms to come up with low-cost process innovation. This has resulted in health products such as the Shanvac-B hepatitis B vaccine, which is produced in India for a fraction of the cost in developed countries.The South African government's Biotechnology Advisory Committee recognizes that successful commercialization of public-sector-supported research and development requires strong linkages within the NSI. The committee has recommended the creation of several Regional Innovation Centres to act as nuclei for the development of biotechnology platforms that can effectively launch new products and services. These strategies will provide important lessons for many other developing nations as they begin participating in the genomics revolution.First, the development gap between developing countries and the industrialized world continues to grow. The international community is beginning to promote science and technology to reduce this gap. The genomics revolution holds tremendous potential to improve health in developing countries and, if harnessed appropriately, could help to reduce the development divide.Second, genomics and related biotechnologies can help to achieve the UN MDGs. Fast, accurate molecular diagnostic devices, safer recombinant vaccines, female-controlled vaginal microbicides, and low-cost bioremediation tools are some examples of biotechnologies that can have an impact.Third, genomics knowledge has the characteristics of a global public good. In order to harness the benefits of genomics for development, the developing world needs, above all, access to genomics knowledge. Fourth, the promotion of the science of genomics as a global public good and the encouragement of global knowledge flows could best be achieved through international partnerships. A GGI involving an international partnership of public and private entities from both developed and developing countries could catalyze genomics knowledge and learning worldwide.Fifth, countries that have genomics capacity are best positioned to take advantage of the genomics revolution to meet their health needs. For the transfer of technologies to be effective and sustainable, it must be accompanied by transfer of science and knowledge. As well, receiving countries must have the capacity to absorb and use the technology.And sixth, learning is important for building genomics capacity, and is central to the creation of NSIs in biotechnology in developing countries. These countries can strengthen the building blocks of the NSI framework by doing the following: re-energizing academic institutions and public-sector research to strengthen their science base; training people and building human capital to use, adapt, and innovate biotechnologies; encouraging regional and international cooperation to create new channels for knowledge exchange and trade; improving the policy environment  to encourage the building of capacity; and fostering the growth of the private sector, encouraging it to address local health needs, and strengthening linkages between public and private sectors to create new biotechnology goods and services.