Development of Next-Generation COVID-19 Vaccines: BARDA Supported Phase 2b Study Designs.

In response to the COVID-19 pandemic, vaccines were quickly and successfully developed and deployed, saving millions of lives globally. While first generation vaccines are safe and effective in preventing disease caused by SARSCoV-2, next-generation vaccines have the potential to improve efficacy and safety. Vaccines delivered by a mucosal route may elicit greater protective immunity at respiratory surfaces thereby reducing transmission. Inclusion of viral antigens in addition to the spike protein may enhance protection against emerging variants of concern. Next-generation vaccine platforms with a new mechanism of action may necessitate efficacy trials to fulfill regulatory requirements. The Biomedical Advanced Research and Development Authority (BARDA) will be supporting Phase 2b clinical trials of candidate next-generation vaccines. The primary endpoint will be improved efficacy in terms of symptomatic disease relative to a currently approved COVID-19 vaccine. In this paper, we discuss the planned endpoints and potential challenges to this complex program.

Outcomes of vitreoretinal surgery in eyes with retinitis pigmentosa without retinal detachment.

PURPOSE: Outcomes of retinal detachment (RD) have been discussed in detail in many reports of patients with retinitis pigmentosa (RP). This study tries to understand the outcomes of vitreoretinal (VR) surgery for indications other than RD in the eyes with RP. METHODS: This is a retrospective study that includes clinical data from January 2013 to December 2021. Patients with RP who were treated with a VR surgical intervention were included in the study. The primary outcome of the study was to assess the changes in best-corrected visual acuity. RESULTS: Forty-four eyes of 40 patients with RP were included in the study. Nearly half of the eyes (43%, 19/44) presented from 1 month to 1 year after the onset of diminished vision, with or without floaters. The mean ± standard deviation (SD) best-corrected visual acuity (BCVA) at presentation was 1.30 ± 0.79 logMAR (20/400 ± 20/125). The major surgical indications were vitreous opacities (43.2%, 19/44) and subluxated/dislocated cataractous lenses (25%, 11/44). The median follow-up duration was 8 months (interquartile range (IQR): 1.5-27). Approximately 77% (34/44) of the eyes had improvement in vision. The mean postoperative BCVA at the last follow-up was 0.95 ± 0.73 logMAR (p-value: 0.03). CONCLUSIONS: Most eyes with RP recovered well after VR surgical interventions, with short-term improvements in visual acuity. It may be crucial to address the vitreous opacities and membranes as they hinder the residual central island of vision in RP. However, appropriate counselling is required regarding the progressive nature of retinal neuronal degeneration.

Dual adhesive unipolar polysaccharides synthesized by overlapping biosynthetic pathways in Agrobacterium tumefaciens.

Agrobacterium tumefaciens is a member of the Alphaproteobacteria that pathogenises plants and associates with biotic and abiotic surfaces via a single cellular pole. A. tumefaciens produces the unipolar polysaccharide (UPP) at the site of surface contact. UPP production is normally surface-contact inducible, but elevated levels of the second messenger cyclic diguanylate monophosphate (cdGMP) bypass this requirement. Multiple lines of evidence suggest that the UPP has a central polysaccharide component. Using an A. tumefaciens derivative with elevated cdGMP and mutationally disabled for other dispensable polysaccharides, a series of related genetic screens have identified a large number of genes involved in UPP biosynthesis, most of which are Wzx-Wzy-type polysaccharide biosynthetic components. Extensive analyses of UPP production in these mutants have revealed that the UPP is composed of two genetically, chemically, and spatially discrete forms of polysaccharide, and that each requires a specific Wzy-type polymerase. Other important biosynthetic, processing, and regulatory functions for UPP production are also revealed, some of which are common to both polysaccharides, and a subset of which are specific to each type. Many of the UPP genes identified are conserved among diverse rhizobia, whereas others are more lineage specific.

Choroidal vascularity index in thyroid eye disease: comparison with controls and application in diagnosing non-inflammatory active disease.

PURPOSE: To report the differences in choroidal vascularity index (CVI) in thyroid eye disease (TED) and normals and its discriminatory value for differentiating various stages of TED. METHODS: Prospective, cross-sectional, non-interventional imaging study. Ninety-four eyes of 54 patients were included and divided into 5 groups - normal controls (C), inactive TED (I), active TED (A), non-inflammatory active TED (NIA) and systemic hyperthyroid disorder but no TED (SYS). Choroidal images were acquired using the swept-source optical coherence tomography and the choroid was binarized to calculate the CVI. RESULTS: Ninety-four eyes were included. Mean age was 44.52 ± 10.02 years (median 46 years, range 19-65 years). Mean IOP was 16.1 ± 3.37 mm Hg (median 16 mm Hg, range 16-24 mm Hg). Mean Spherical equivalent (SE) was -0.08 ± 1.86 diopters (median 0, range -2.5 to +2.25). Intra-rater agreement was 0.84 (p < 0.001). Inter-rater agreement was noted to be 0.85 (p < 0.001) for consistency and 0.77 (p < 0.001) for absolute agreement. CVI in the A group was 70.11 ± 3.38% and in the NIA group was 69.32 ± 3.5%. Both were comparable to each other and significantly higher than the C, I and SYS groups (p < 0.001). Multiple regression showed that the Clinical Activity Score (CAS) had a positive effect and spheroequivalent had a negative effect on the CVI. At CVI of 66.83%, active TED can be diagnosed with sensitivity of 91.67% and specificity of 82.14%                 . CONCLUSIONS: CVI is significantly higher in active TED and NIA TED compared to other groups. It has a good value in differentiating the non-inflammatory active TED eyes from the inactive eyes.

Clinical characteristics of comorbid retinal dystrophies and primary angle closure disease.

PURPOSE: To assess the clinical characteristics of comorbid retinal dystrophies and primary angle closure disease. DESIGN: Retrospective study from January 1992 to June 2020. METHODS: This descriptive study included 92 eyes of 46 patients with comorbid retinal dystrophies and primary angle closure disease (PACD) that included eyes with primary angle closure suspect, primary angle closure and primary angle closure glaucoma. Demographic profile, clinical characteristics of PACD and its association with retinal dystrophies are described. RESULTS: The study included 46 patients (92 eyes). Males were majority, 63%. Mean (± standard deviation) age when retinal dystrophy was diagnosed was 29.6 ± 9.4 years and PACD was diagnosed at 32.23 ± 7.92 years. Mean BCVA at presentation was 1.07 ± 0.87 log MAR [95% confidence interval (CI) 0.87, 1.26]. Mean Intraocular pressure at diagnosis of glaucoma was 27 ± 16 mmHg (95% CI 23.5, 31.5 mmHg). The most common retinal dystrophy associated with PACD was retinitis pigmentosa (RP) followed by RP with retinoschisis. The hospital-based prevalence of PACD among all patients with RP and retinoschisis was 0.19% and 0.15% respectively. Laser peripheral iridotomy was performed in 74 eyes (80.5%). Glaucoma was managed medically in majority of the eyes (58 eyes, 63.04%) and minority required surgical management with trabeculectomy (11, 11.9%). CONCLUSION: Retinitis pigmentosa is the most common retinal dystrophy associated with PACD. Comorbid PACD in eyes with retinal dystrophies was observed in second to third decade of life. This calls for screening for angle closure in eyes with retinal dystrophies from second decade onwards to identify the comorbid PACD and treat or refer them appropriately.

Spermidine Inversely Influences Surface Interactions and Planktonic Growth in Agrobacterium tumefaciens.

UNLABELLED: In bacteria, the functions of polyamines, small linear polycations, are poorly defined, but these metabolites can influence biofilm formation in several systems. Transposon insertions in an ornithine decarboxylase (odc) gene in Agrobacterium tumefaciens, predicted to direct synthesis of the polyamine putrescine from ornithine, resulted in elevated cellulose. Null mutants for odc grew somewhat slowly in a polyamine-free medium but exhibited increased biofilm formation that was dependent on cellulose production. Spermidine is an essential metabolite in A. tumefaciens and is synthesized from putrescine in A. tumefaciens via the stepwise actions of carboxyspermidine dehydrogenase (CASDH) and carboxyspermidine decarboxylase (CASDC). Exogenous addition of either putrescine or spermidine to the odc mutant returned biofilm formation to wild-type levels. Low levels of exogenous spermidine restored growth to CASDH and CASDC mutants, facilitating weak biofilm formation, but this was dampened with increasing concentrations. Norspermidine rescued growth for the odc, CASDH, and CASDC mutants but did not significantly affect their biofilm phenotypes, whereas in the wild type, it stimulated biofilm formation and depressed spermidine levels. The odc mutant produced elevated levels of cyclic diguanylate monophosphate (c-di-GMP), exogenous polyamines modulated these levels, and expression of a c-di-GMP phosphodiesterase reversed the enhanced biofilm formation. Prior work revealed accumulation of the precursors putrescine and carboxyspermidine in the CASDH and CASDC mutants, respectively, but unexpectedly, both mutants accumulated homospermidine; here, we show that this requires a homospermidine synthase (hss) homologue. IMPORTANCE: Polyamines are small, positively charged metabolites that are nearly ubiquitous in cellular life. They are often essential in eukaryotes and more variably in bacteria. Polyamines have been reported to influence the surface-attached biofilm formation of several bacteria. In Agrobacterium tumefaciens, mutants with diminished levels of the polyamine spermidine are stimulated for biofilm formation, and exogenous provision of spermidine decreases biofilm formation. Spermidine is also essential for A. tumefaciens growth, but the related polyamine norspermidine exogenously rescues growth and does not diminish biofilm formation, revealing that the growth requirement and biofilm control are separable. Polyamine control of biofilm formation appears to function via effects on the cellular second messenger cyclic diguanylate monophosphate, regulating the transition from a free-living to a surface-attached lifestyle.

Discrete Responses to Limitation for Iron and Manganese in Agrobacterium tumefaciens: Influence on Attachment and Biofilm Formation.

UNLABELLED: Transition metals such as iron and manganese are crucial trace nutrients for the growth of most bacteria, functioning as catalytic cofactors for many essential enzymes. Dedicated uptake and regulatory systems have evolved to ensure their acquisition for growth, while preventing toxicity. Transcriptomic analysis of the iron- and manganese-responsive regulons of Agrobacterium tumefaciens revealed that there are discrete regulatory networks that respond to changes in iron and manganese levels. Complementing earlier studies, the iron-responsive gene network is quite large and includes many aspects of iron-dependent metabolism and the iron-sparing response. In contrast, the manganese-responsive network is restricted to a limited number of genes, many of which can be linked to transport and utilization of the transition metal. Several of the target genes predicted to drive manganese uptake are required for growth under manganese-limited conditions, and an A. tumefaciens mutant with a manganese transport deficiency is attenuated for plant virulence. Iron and manganese limitation independently inhibit biofilm formation by A. tumefaciens, and several candidate genes that could impact biofilm formation were identified in each regulon. The biofilm-inhibitory effects of iron and manganese do not rely on recognized metal-responsive transcriptional regulators, suggesting alternate mechanisms influencing biofilm formation. However, under low-manganese conditions the dcpA operon is upregulated, encoding a system that controls levels of the cyclic di-GMP second messenger. Mutation of this regulatory pathway dampens the effect of manganese limitation. IMPORTANCE: Responses to changes in transition metal levels, such as those of manganese and iron, are important for normal metabolism and growth in bacteria. Our study used global gene expression profiling to understand the response of the plant pathogen Agrobacterium tumefaciens to changes of transition metal availability. Among the properties that are affected by both iron and manganese levels are those required for normal surface attachment and biofilm formation, but the requirement for each of these transition metals is mechanistically independent from the other.

Refractive outcomes following silicone oil tamponade in vitreoretinal surgery.

PURPOSE: To evaluate the factors influencing the refractive outcomes following silicone oil tamponade (SOT) and silicone oil removal (SOR) in different lens statuses post-vitreoretinal surgery. DESIGN: Retrospective analysis of three different lens statuses. MATERIALS AND METHODS: This was a descriptive study that included 150 eyes of 147 patients who had undergone pars plana vitrectomy with SOT and SOR between January 2017 and June 2021. Demographic profile, spherical equivalent refraction (SER), and its association with clinical features were evaluated with SOT and post-SOR. RESULTS: The mean (±standard deviation [SD]) age was 47 ± 17.8 years, including all three groups. SER was represented in diopters (D). The mean ± SD refraction with SOT in phakic, pseudophakic, and aphakic was 4.28 ± 2.59 D, 2.94 ± 2.58 D, and 3.98 ± 4.82 D. The mean SER post-SOR in phakic, pseudophakic, and aphakic was -2.72 ± 2.03 D, -1.12 ± 1.41 D, and 8.22 ± 3.70 D. The diagnosis of rhegmatogenous retinal detachment (RRD) among 96 eyes (64%) is the common indicator to perform vitreoretinal (VR) surgery. A minority of subjects were managed with retinal lasers before VR surgery (14%). The macula was attached in 100 eyes (67.6%), the belt buckle was done in 37 eyes (24.7%), and the silicone oil viscosity with 1000 centistoke was chosen in 129 eyes (86%). CONCLUSION: SOT was used as a tamponade in VR surgeries irrespective of lens status. The significant predictor for post-SOR refraction in phakic and aphakic is post-SOT refraction. In pseudophakic, gender and diagnosis of RRD are the predictors of SOR refraction.

Peripheral defocus of monofocal intraocular lenses.

PURPOSE: To quantify the angular dependence of monofocal intraocular lens (IOL) power. SETTING: Ophthalmic Biophysics Laboratory, Kallam Anji Reddy campus, L V Prasad Eye Institute, Hyderabad, India. DESIGN: Laboratory study. METHODS: Experiments were performed on IOLs from 2 different manufacturers (APPALENS 207, Appasamy Associates and SN60WF, Alcon Laboratories, Inc.). IOL powers ranged from 17 to 25 diopters (D). The IOLs were mounted in a fluid-filled chamber, and the on-axis and off-axis powers were measured using a laser ray tracing system over the central 3 mm zone with delivery angles ranging from -30 to +30 degrees in 5-degree increments. The position of the best focus was calculated for each IOL at each angle. The angular dependence of IOL power was compared with theoretical predictions. RESULTS: Peripheral defocus increased significantly with increasing incidence angle and power. The peripheral defocus at ±30 degrees increased from 5.8 to 8.5 D when the power increased from 17.5 to 24.5 D for APPALENS 207 and from 4.9 to 7.4 D when the power increased from 17 to 25 D for SN60WF. The mean difference between the measured and theoretical tangential power at ±30 degrees was 0.50 ± 0.16 D for the APPALENS 207 and -0.40 ± 0.10 D for the SN60WF, independent of IOL power. CONCLUSIONS: IOLs introduce a significant amount of peripheral defocus which varies significantly with IOL power and design. Given that peripheral defocus is related to lens power, replacement of the crystalline lens (approximately 24 D) with an IOL will produce a significant difference in peripheral defocus profile after surgery.

Prognostic factors for visual outcome after surgical repair of limbal corneal laceration at a tertiary eye care center in South India.

Purpose: To determine the factors affecting the visual outcome after surgical repair of limbal corneal lacerations at a tertiary eye care center in South India. Methods: A retrospective analysis of patients diagnosed with limbal tears between 2011 and 2021 was conducted. Demographic information such as age, gender, cause of injury, and size of the laceration was recorded. Comprehensive ocular examination was performed, including gentle B scan evaluation whenever not contraindicated for detailed posterior segment evaluation. Only those cases with a minimum follow-up of one year were included. Postoperative best-corrected visual acuity, intraocular pressure (IOP), cornea clarity, and integrity of the wound at last follow-up were noted. Results: Out of the 20 patients, 15 (75%) were males and 5 (25%) were females. The mean age was 42.6 ± 22.4 years. All 20 patients had a penetrating injury, with four (20%) injured by a stick, two (10%) by an iron rod, three (15%) due to road traffic accident (RTA), three (15%) by glass, and eight (40%) with other nonspecific objects [two (10%) with needle, two (10%) with elastic rope, two (10%) with bangle, and two (10%) with metal]. The average time between the injury and the surgery was 48 hours (2 days). Four (20%) patients underwent a second surgery within a week of repair. After limbal tear repair, at final follow-up at 3 years, 7 (35%) had VA worse than 20/800, 3 (15%) had VA between 20/100 and 20/800, and 10 (50%) achieved VA better than 20/80. Conclusion: Preoperative visual acuity (VA), mode of injury, and size of wound affect the final visual outcome after surgical repair of limbal corneal laceration. Preoperative VA and mode of injury were statistically significant even in the multivariate analysis.

Visual impairment and blindness due to retinitis pigmentosa in India: 15-year follow-up of the Andhra Pradesh Eye Disease Study cohort.

Purpose: To assess the incidence, visual impairment, and blindness due to retinitis pigmentosa (RP) in a rural southern Indian cohort. Methods: This is a population-based longitudinal cohort study of participants with RP from the Andhra Pradesh Eye Disease Study (APEDS) cohorts I and III, respectively. The study included participants with RP of APEDS I who were followed until APEDS III. Their demographic data along with ocular features, fundus photographs, and visual fields (Humphrey) were collected. Descriptive statistics using mean ± standard deviation with interquartile range (IQR) were calculated. The main outcome measures were RP incidence, visual impairment, and blindness as per the World Health Organization (WHO) definitions. Results: At baseline (APEDS I), 7771 participants residing in three rural areas were examined. There were nine participants with RP with a mean age at baseline of 47.33 ± 10.89 years (IQR: 39-55). There was a male preponderance (6:3), and the mean best-corrected visual acuity (BCVA) of 18 eyes from nine participants with RP was 1.2 ± 0.72 logarithm of minimum angle of resolution (logMAR; IQR: 0.7-1.6). Over a mean follow-up duration of 15 years, 5395/7771 (69.4%) were re-examined, which included seven RP participants from APEDS 1. Additionally, two new participants with RP were identified; so, the overall incidence was 370/ million in 15 years (24.7/million per year). The mean BCVA of 14 eyes of seven participants with RP who were re-examined in APEDS III was 2.17 ± 0.56 logMAR (IQR: 1.8-2.6), and five of these seven participants with RP developed incident blindness during the follow-up period. Conclusion: RP is a prevalent disease in southern India that warrants appropriate strategies to prevent this condition.

Retinitis pigmentosa in Usher syndrome in India: Electronic medical records driven big data analytics: Report III.

Purpose: To describe the clinical presentation and demographic distribution of retinitis pigmentosa (RP) in patients with Usher syndrome (USH). Methods: This is a cross-sectional observational hospital-based study including patients presenting between March 2012 and October 2020. In total, 401 patients with a clinical diagnosis of USH and RP in at least one eye were included as cases. The data were retrieved from the electronic medical record database. For better analysis, all 401 patients were reclassified into three subtypes (type 1, type 2, and type 3) based on the USH criteria. Results: In total, there were 401 patients with USH and RP, with a hospital-based prevalence rate of 0.02% or 2/10,000 population. Further, 353/401 patients were subclassified, with 121 patients in type 1, 146 patients in type 2, and 86 patients in the type 3 USH group. The median age at presentation was 27 years (IQR: 17.5-38) years. There were 246 (61.35%) males and 155 (38.65%) females. Males were more commonly affected in all three subtypes. Defective night vision was the predominant presenting feature in all types of USH (type 1: 43 (35.54%), type 2: 68 (46.58%), and type 3: 40 (46.51%) followed by defective peripheral vision. Patients with type 2 USH had more eyes with severe visual impairment. Conclusion: RP in USH is commonly bilateral and predominantly affects males in all subtypes. Patients with USH and RP will have more affection of peripheral vision than central vision. The key message of our study is early visual and hearing rehabilitation in USH patients with prompt referral to otolaryngologists from ophthalmologists and vice versa.

Clinical Presentation and Demographic Distribution of Retinitis Pigmentosa in India and Implications for Potential Treatments: Electronic Medical Records Driven Big Data Analytics: Report I.

AimTo describe the clinical presentation and demographic distribution of retinitis pigmentosa in patients presenting to a multi-tier ophthalmology hospital network in India and implications for potential treatments.DesignCross-sectional hospital-based study.MethodsThis study included 2,541,810 patients presenting between March 2012 and October 2020. Patients with a clinical diagnosis of retinitis pigmentosa in at least one eye were included as cases. The data were collected using an electronic medical record system.ResultsOverall, 15,062 (0.59%) new patients were diagnosed with retinitis pigmentosa and were included for analysis. The median age was 35 (IQR: 21-49) years and adult age (84.56%) presented more often. The most common presenting age group was between 21 and 30 years (20%). Majority of patients were male (61.68%) and had bilateral (96.39%) affliction. There was a family history in a tenth of the patients (10.46%) and a history of consanguinity in a minority (5.53%). The majority of the eyes had mild or no central visual acuity impairment of <20/70 (34.25%) followed by blindness <20/400 to 20/1200 (21.26%). The predominant retinal signs included disc pallor (76.43%), attenuated arterioles (82.61%) and bony spicule pigmentation (90.15%), and almost all had retinal pigmentary changes. A minority of the eyes underwent a surgical procedure (4.96%).ConclusionRetinitis pigmentosa is commonly bilateral and predominantly affects males. Most patients present in the third decade of life with severe visual acuity impairment. More than half of the eyes from our study are potential candidates for newer therapies. These results have implications for upcoming treatment choices and planning.

Choroidal Thickness in Thyroid Eye Disease: Comparison With Controls and Application in Diagnosing Non-Inflammatory Active Disease.

Introduction Choroidal thickness is known to vary in various systemic diseases. In the current study, we aim to report the differences in choroidal thickness in thyroid eye disease (TED) and normals and its discriminatory value for differentiating various stages of TED. Methods Prospective, cross-sectional, non-interventional imaging study. In an institutional practice, 102 eyes of 51 patients were included and divided into five groups: normal controls (C), inactive TED (I), active TED (A), non-inflammatory active TED (NIA) and systemic thyroid disorder but no TED (SYS). Choroidal images were acquired using the swept-source optical coherence tomography (Topcon DRI OCT Triton) with automatic layer segmentation which provided an automatic measurement of the subfoveal choroidal thickness and the mean in nine subfields based on the Early Treatment Diabetic Retinopathy Study (ETDRS) grid. One-way analysis of variance (ANOVA), Youden index and area under the receiver operating characteristic curves (AUROC) were reported. Results Central choroidal thickness in the A group was 279±37.52 microns and in the NIA group was 302.5±59.22 microns. Both were comparable to each other and significantly higher than the C, I and SYS groups (p<0.001). All ETDRS sub-fields showed significant AUROC to distinguish NIA from I. Most significant Youden index was for the inner nasal and central ETDRS subfields (0.55 and 0.61 respectively). Inner nasal sub-field showed 100% specificity while the central sub-field, showed 86.5% for predicting NIA. At a choroidal thickness of >266 microns, the central sub-field had the strongest discriminatory potential to predict NIA.  Conclusion Choroidal thickness is greater in active and non-inflammatory active TED. The inner nasal and central ETDRS sub-fields have value in differentiating the non-inflammatory active TED eyes from the inactive eyes.

Gastrointestinal mucormycosis in a child with acute lymphoblastic leukemia: An uncommon but ominous complication.

Invasive fungal infections constitute a major cause of morbidity and mortality in children undergoing therapy for hematological malignancies. We report a 1-year-old boy who was receiving chemotherapy for acute lymphoblastic leukemia. His clinical course was complicated by a clinical syndrome consistent with neutropenic enterocolitis to which he succumbed. Histopathology of the surgically resected bowel revealed evidence of mucormycosis. Gastrointestinal mucormycosis is an unusual presentation which requires high degree of clinical suspicion and aggressive management.

Using sound pulses to solve the crystal-harvesting bottleneck.

Crystal harvesting has proven to be difficult to automate and remains the rate-limiting step for many structure-determination and high-throughput screening projects. This has resulted in crystals being prepared more rapidly than they can be harvested for X-ray data collection. Fourth-generation synchrotrons will support extraordinarily rapid rates of data acquisition, putting further pressure on the crystal-harvesting bottleneck. Here, a simple solution is reported in which crystals can be acoustically harvested from slightly modified MiTeGen In Situ-1 crystallization plates. This technique uses an acoustic pulse to eject each crystal out of its crystallization well, through a short air column and onto a micro-mesh (improving on previous work, which required separately grown crystals to be transferred before harvesting). Crystals can be individually harvested or can be serially combined with a chemical library such as a fragment library.

Lipid releasing characteristics of microalgae species through continuous ultrasonication.

In this study, the lipid releasing characteristics of several microalgae species through continuous ultrasonication was examined. Two marine microalgae species, Tetraselmis suecica and Nannochloropsis sp., and one freshwater species, Chlorella sp. were ultrasonicated directly after cultivation. The cell disruption efficiency and lipids releasing pattern from microalgae cells were measured under various ultrasonication conditions. It was found that cell disruption efficiency correlates well with ultrasonication energy consumption despite the ultrasonication conditions. Lipids in Chlorella sp. that has rigid cell walls were released to the aqueous phase after cell disruption. T. suecica and Nannochloropsis sp. that have flexible cell membranes tend to coil up and retain the membrane lipids after disruption. Continuous ultrasonication can be a potential method to release the lipids in rigid walled microalgae species without expensive dewatering steps.

The malaria circumsporozoite protein has two functional domains, each with distinct roles as sporozoites journey from mosquito to mammalian host.

Plasmodium sporozoites make a remarkable journey from the mosquito midgut to the mammalian liver. The sporozoite's major surface protein, circumsporozoite protein (CSP), is a multifunctional protein required for sporozoite development and likely mediates several steps of this journey. In this study, we show that CSP has two conformational states, an adhesive conformation in which the C-terminal cell-adhesive domain is exposed and a nonadhesive conformation in which the N terminus masks this domain. We demonstrate that the cell-adhesive domain functions in sporozoite development and hepatocyte invasion. Between these two events, the sporozoite must travel from the mosquito midgut to the mammalian liver, and N-terminal masking of the cell-adhesive domain maintains the sporozoite in a migratory state. In the mammalian host, proteolytic cleavage of CSP regulates the switch to an adhesive conformation, and the highly conserved region I plays a critical role in this process. If the CSP domain architecture is altered such that the cell-adhesive domain is constitutively exposed, the majority of sporozoites do not reach their target organs, and in the mammalian host, they initiate a blood stage infection directly from the inoculation site. These data provide structure-function information relevant to malaria vaccine development.

Report of an NIAID workshop on dengue animal models.

Dengue is a mosquito-borne viral disease of humans that has re-emerged in many parts of the world and has become an important international public health threat. Dengue incidence and geographical spread has dramatically increased in the last few decades and is now affecting most tropical and sub-tropical regions of the world. Despite extensive research efforts for several decades, no vaccines or therapeutics are currently available to prevent and treat dengue infections. One of the main obstacles to the development of countermeasures has been the lack of good animal models that recapitulate dengue pathogenesis in humans and reliably predict the safety and efficacy of countermeasures against dengue. In September 2008, the National Institute of Allergy and Infectious Diseases (NIAID) held a workshop to consider the current state-of-the-art developments in animal models for dengue and discuss strategies to accelerate progress in this field. This report summarizes the main discussions and recommendations that resulted from the meeting.

A mosquito-specific protein family includes candidate receptors for malaria sporozoite invasion of salivary glands.

We describe a previously unrecognized protein family from Aedes and Anopheles mosquitoes, here named SGS proteins. There are no SGS homologues in Drosophila or other eukaryotes, but SGS presence in two mosquito genera suggests that the protein family is widespread among mosquitoes. Ae. aegypti aaSGS1 mRNA and protein are salivary gland specific, and protein is localized in the basal lamina covering the anatomical regions that are preferentially invaded by malaria sporozoites. Anti-aaSGS1 antibodies inhibited sporozoite invasion into the salivary glands in vivo, confirming aaSGS1 as a candidate sporozoite receptor. By homology to aaSGS1 we identified the complete complement of four SGS genes in An. gambiae, which were not recognized in the genome annotation. Two An. gambiae SGS genes display salivary gland specific expression like aaSGS1. Bioinformatic analysis predicts that SGS proteins possess heparin-binding domains, and have among the highest density of tyrosine sulphation sites of all An. gambiae proteins. The major sporozoite surface proteins (CS and TRAP) also bind heparin, and interact with sulphoconjugates during liver cell invasion. Thus, we speculate that sporozoite invasion of mosquito salivary glands and subsequently the vertebrate liver may share similar mechanisms based on sulphation. Phylogenomic analysis suggests that an SGS ancestor was involved in a lateral gene transfer.