Cabozantinib and nivolumab with or without live bacterial supplementation in metastatic renal cell carcinoma: a randomized phase 1 trial.

Supplementation with CBM588, a bifidogenic live bacterial product, has been associated with improved clinical outcomes in persons with metastatic renal cell carcinoma (mRCC) receiving nivolumab and ipilimumab. However, its effect on those receiving tyrosine kinase inhibitor-based combinations is unknown. In this open-label, randomized, investigator-initiated, phase 1 study, 30 participants with locally advanced or mRCC with histological confirmation of clear cell, papillary or sarcomatoid component were randomized in a 2:1 fashion to receive cabozantinib (an inhibitor of vascular endothelial growth factor receptor, MET and AXL) and nivolumab (anti-programmed cell death protein 1) with or without CBM588 as first-line treatment. Metagenomic sequencing was performed on stool samples to characterize their gut microbiome at baseline and 13 weeks into treatment. The primary endpoint was a change in the relative abundance of Bifidobacterium spp.; secondary endpoints included objective response rate (ORR), progression-free survival (PFS) and toxicity profile. The primary endpoint of the study was not met and the addition of CBM588 to cabozantinib and nivolumab did not result in a difference in the relative abundance of Bifidobacterium spp. or alpha diversity (as measured by the Shannon index). However, ORR was significantly higher in participants treated with CBM588 compared to those in the control arm (14 of 19, 74% versus 2 of 10, 20%; P = 0.01). PFS at 6 months was 84% (16 of 19) and 60% (6 of 10) in the experimental and control arms, respectively. No significant difference in toxicity profile was seen between the study arms. Our results provide a preliminary signal of improved clinical activity with CBM588 in treatment-naive participants with mRCC receiving cabozantinib and nivolumab. Further investigation is needed to confirm these findings and better characterize the underlying mechanism driving this effect.ClinicalTrials.gov identifier: NCT05122546.

Functional evidence of distinct electrophile-induced activation states of the ion channel TRPA1.

Transient Receptor Potential Ankyrin 1 (TRPA1) is a tetrameric, nonselective cation channel expressed on nociceptive sensory nerves whose activation elicits nocifensive responses (e.g. pain). TRPA1 is activated by electrophiles found in foods and pollution, or produced during inflammation and oxidative stress, via covalent modification of reactive cysteines, but the mechanism underlying electrophilic activation of TRPA1 is poorly understood. Here we studied TRPA1 activation by the irreversible electrophiles iodoacetamide and N-ethylmaleimide (NEM) following transient expression in HEK293 cells. We found that in Ca2+ imaging studies C621 is critical for electrophile-induced TRPA1 activation, but the role of C665 in TRPA1 activation is dependent on the size of the electrophile. We identified slower TRPA1 activation in whole-cell recordings compared to studies with intact cells, which is rescued by pipette solution supplementation with the antioxidant glutathione. Single-channel recordings identified two distinct electrophilic-induced TRPA1 activation phases: a partial activation that, in some channels, switched to full activation with continued electrophile exposure. Full activation but not the initial activation was regulated by C665. Fitting of open time distributions suggests that full activation correlated with an additional (and long) exponential component, thus suggesting the phases are manifestations of distinct activation states. Our results suggest that distinct NEM-induced TRPA1 activation states are evoked by sequential modification of C621 then C665.

Connecting the Dots: Translating the Vaginal Microbiome Into a Drug.

A Lactobacillus-dominated vaginal microbiota (VMB) has been associated with health and considered an important host defense mechanism against urogenital infections. Conversely, depletion of lactobacilli and increased microbial diversity, amplifies the risk of adverse gynecologic and obstetric outcomes. A common clinical condition that exemplifies dysbiosis is bacterial vaginosis (BV). BV is currently treated with antibiotics, but frequently recurs, due in part to persistent dysbiosis and failure of lactobacilli to repopulate the vagina. New treatment options are needed to address BV. The VMB is relatively simple and optimally dominated by one or several species of Lactobacillus. Lactobacillus crispatus is strongly associated with vaginal health and depleted in dysbiosis. Replenishing the dysbiotic VMB with protective L. crispatus CTV-05 is a promising approach to prevent recurrent infections and improve women's health. Here we discuss confirmation of this approach with the microbiome-based biologic drug, LACTIN-V (L. crispatus CTV-05), focusing on prevention of BV recurrence.

Microscope-Cockpit: Python-based bespoke microscopy for bio-medical science.

We have developed "Microscope-Cockpit" (Cockpit), a highly adaptable open source user-friendly Python-based Graphical User Interface (GUI) environment for precision control of both simple and elaborate bespoke microscope systems. The user environment allows next-generation near instantaneous navigation of the entire slide landscape for efficient selection of specimens of interest and automated acquisition without the use of eyepieces. Cockpit uses "Python-Microscope" (Microscope) for high-performance coordinated control of a wide range of hardware devices using open source software. Microscope also controls complex hardware devices such as deformable mirrors for aberration correction and spatial light modulators for structured illumination via abstracted device models. We demonstrate the advantages of the Cockpit platform using several bespoke microscopes, including a simple widefield system and a complex system with adaptive optics and structured illumination. A key strength of Cockpit is its use of Python, which means that any microscope built with Cockpit is ready for future customisation by simply adding new libraries, for example machine learning algorithms to enable automated microscopy decision making while imaging.

Effect of clindamycin and a live biotherapeutic on the reproductive outcomes of IVF patients with abnormal vaginal microbiota: protocol for a double-blind, placebo-controlled multicentre trial.

INTRODUCTION: Recent studies in in vitro fertilisation (IVF) patients have associated abnormal vaginal microbiota (AVM) with poor clinical pregnancy rates of 6%-9% per embryo transfer. The biological plausibility for this finding is hypothesised to be ascending infection to the endometrium which in turn hampers embryo implantation. New molecular based diagnosis may offer advantages compared to microscopical diagnosis of AVM which has huge inter-study variability ranging from 4 to 38%; however, the important question is whether screening and treatment of AVM would improve reproductive outcomes in IVF patients. Herein, we describe a protocol for an ongoing double-blind, placebo-controlled multicentre trial of IVF patients diagnosed with AVM and randomised in three parallel groups 1:1:1. METHODS AND ANALYSIS: This is a drug intervention study where IVF patients will be screened for AVM, using a qPCR assay targeting Atopobium vaginae and Gardnerella vaginalis. If positive, patients will be randomised to one of the three study arms. The first arm consists of clindamycin 300 mg ×2 daily for 7 days followed by vaginal Lactobacillus crispatus CTV-05 until clinical pregnancy scan week 7-9. The second arm consists of clindamycin and placebo L. crispatus CTV-05, whereas patients in the third arm will be treated with placebo/placebo. We used a superiority design to estimate that active treatment in both arms will increase the primary outcome, clinical pregnancy rate per embryo transfer, from 20% to 40%. A potential difference between the two active arms was considered exploratory. With a power of 80% and an alpha at 5%, the sample size is estimated to be 333 patients randomised. A pre-planned interim analysis is scheduled at 167 patients randomised. ETHICS AND DISSEMINATION: All patients have to give informed consent. Dissemination of results is ensured in clinical trial agreements whether they be positive or not. Ethics committee, Central Denmark Region approved this protocol. TRIAL REGISTRATION NUMBER: ICH-GCP monitored trial, EudraCT 2016-002385-31; Pre-results.

Aldosterone up-regulates voltage-gated potassium currents and NKCC1 protein membrane fractions.

Na+-K+-2Cl- Cotransporter (NKCC1) is a protein that aids in the active transport of sodium, potassium, and chloride ions across cell membranes. It has been shown that long-term systemic treatment with aldosterone (ALD) can enhance NKCC1 protein expression and activity in the aging cochlea resulting in improved hearing. In the present work, we used a cell line with confirmed NKCC1 expression to demonstrate that in vitro application of ALD increased outward voltage-gated potassium currents significantly, and simultaneously upregulated whole lysate and membrane portion NKCC1 protein expression. These ALD-induced changes were blocked by applying the mineralocorticoid receptor antagonist eplerenone. However, application of the NKCC1 inhibitor bumetanide or the potassium channel antagonist Tetraethyl ammonium had no effect. In addition, NKKC1 mRNA levels remained stable, indicating that ALD modulates NKCC1 protein expression via the activation of mineralocorticoid receptors and post-transcriptional modifications. Further, in vitro electrophysiology experiments, with ALD in the presence of NKCC1, K+ channel and mineralocorticoid receptor inhibitors, revealed interactions between NKCC1 and outward K+ channels, mediated by a mineralocorticoid receptor-ALD complex. These results provide evidence of the therapeutic potential of ALD for the prevention/treatment of inner ear disorders such as age-related hearing loss.

Novel Selective Inhibition of Lactobacillus iners by Lactobacillus-Derived Bacteriocins.

Lactobacillus iners is often associated with vaginal dysbiosis and bacterial vaginosis (BV), which are risk factors for adverse gynecological and obstetric outcomes. To discover natural inhibitors of L. iners, cell-free culture supernatants (CFSs) from 77 vaginal human Lactobacillus strains and 1 human intestinal strain were screened for inhibitory activity. Three active strains were identified, and Lactobacillus paragasseri K7 (K7), a human intestinal strain, produced the most potent L. iners-inhibitory activity. The active material was purified from the K7 CFS and yielded three active peptides, identified as components of two different class IIb, two-peptide bacteriocins, gassericin K7A (GasK7A) and gassericin K7B (GasK7B). The peptides corresponded to the GasK7A α peptide and the GasK7B α and ß peptides. While all three peptides exhibited individual activity against L. iners, GasK7B α was the most potent, with an MIC of 23 ng/ml (4 nM). When combined in equal amounts, the GasK7B α and ß peptides showed synergistic inhibition, with an MIC of 2 ng/ml (each peptide at 0.4 nM). Among the four major vaginal Lactobacillus species, the K7 bacteriocins selectively inhibited L. iners All 21 strains of L. iners tested (100%) were inhibited by the K7 bacteriocins, whereas <20% of the vaginal Lactobacillus crispatus, L. jensenii, and L. gasseri strains were inhibited. The combination of the BV treatment metronidazole and K7 bacteriocins completely killed both L. iners and Gardnerella vaginalis in a coculture experiment to mimic BV conditions. In contrast, this treatment did not inhibit L. crispatus cultures.IMPORTANCELactobacillus iners is a prevalent species of the vaginal microbiome, but unlike other major vaginal Lactobacillus species, it is not considered protective against BV and can coexist with BV-associated bacteria. L. iners is generally the first Lactobacillus species to emerge following the treatment of BV with metronidazole, and mounting evidence suggests that it may contribute to the onset and maintenance of vaginal dysbiosis. The discovery of highly potent bacteriocins that selectively kill L. iners while sparing protective vaginal lactobacilli may provide novel pharmacological tools to better understand the roles of this enigmatic bacterium in vaginal ecology and potentially lead to new and improved therapies for dysbiosis-related conditions such as BV.

Randomized Trial of Lactin-V to Prevent Recurrence of Bacterial Vaginosis.

BACKGROUND: Bacterial vaginosis affects 15 to 50% of women of reproductive age, and recurrence is common after treatment with an antibiotic agent. The high incidence of recurrence suggests the need for new treatments to prevent recurrent bacterial vaginosis. METHODS: We conducted a randomized, double-blind, placebo-controlled, phase 2b trial to evaluate the ability of Lactobacillus crispatus CTV-05 (Lactin-V) to prevent the recurrence of bacterial vaginosis. Women 18 to 45 years of age who had received a diagnosis of bacterial vaginosis and who had completed a course of vaginal metronidazole gel as part of the eligibility requirements were randomly assigned, in a 2:1 ratio, to receive vaginally administered Lactin-V or placebo for 11 weeks; follow-up occurred through week 24. The primary outcome was the percentage of women who had a recurrence of bacterial vaginosis by week 12. RESULTS: A total of 228 women underwent randomization: 152 to the Lactin-V group and 76 to the placebo group; of these participants, 88% in the Lactin-V group and 84% in the placebo group could be evaluated for the primary outcome. In the intention-to-treat population, recurrence of bacterial vaginosis by week 12 occurred in 46 participants (30%) in the Lactin-V group and in 34 participants (45%) in the placebo group (risk ratio after multiple imputation for missing responses, 0.66; 95% confidence interval [CI], 0.44 to 0.87; P = 0.01). The risk ratio for recurrence by week 24 (also calculated with multiple imputation for missing responses) was 0.73 (95% CI, 0.54 to 0.92). At the 12-week visit, L. crispatus CTV-05 was detected in 79% of participants in the Lactin-V group. The percentage of participants who had at least one adverse event related to Lactin-V or placebo by week 24 did not differ significantly between the groups. The percentage of participants with local or systemic adverse events was similar in the two groups. CONCLUSIONS: The use of Lactin-V after treatment with vaginal metronidazole resulted in a significantly lower incidence of recurrence of bacterial vaginosis than placebo at 12 weeks. (Funded by the National Institutes of Health; ClinicalTrials.gov number, NCT02766023.).

The exceptionally high reactivity of Cys 621 is critical for electrophilic activation of the sensory nerve ion channel TRPA1.

Activation of the sensory nerve ion channel TRPA1 by electrophiles is the key mechanism that initiates nociceptive signaling, and leads to defensive reflexes and avoidance behaviors, during oxidative stress in mammals. TRPA1 is rapidly activated by subtoxic levels of electrophiles, but it is unclear how TRPA1 outcompetes cellular antioxidants that protect cytosolic proteins from electrophiles. Here, using physiologically relevant exposures, we demonstrate that electrophiles react with cysteine residues on mammalian TRPA1 at rates that exceed the reactivity of typical cysteines by 6,000-fold and that also exceed the reactivity of antioxidant enzymes. We show that TRPA1 possesses a complex reactive cysteine profile in which C621 is necessary for electrophile-induced binding and activation. Modeling of deprotonation energies suggests that K620 contributes to C621 reactivity and mutation of K620 alone greatly reduces the effect of electrophiles on TRPA1. Nevertheless, binding of electrophiles to C621 is not sufficient for activation, which also depends on the function of another reactive cysteine (C665). Together, our results demonstrate that TRPA1 acts as an effective electrophilic sensor because of the exceptionally high reactivity of C621.

Expression of Human Immunodeficiency Virus Type 1 Neutralizing Antibody Fragments Using Human Vaginal Lactobacillus.

Eradication of human immunodeficiency virus type 1 (HIV-1) by vaccination with epitopes that produce broadly neutralizing antibodies is the ultimate goal for HIV prevention. However, generating appropriate immune responses has proven difficult. Expression of broadly neutralizing antibodies by vaginal colonizing lactobacilli provides an approach to passively target these antibodies to the mucosa. We tested the feasibility of expressing single-chain and single-domain antibodies (dAbs) in Lactobacillus to be used as a topical microbicide/live biotherapeutic. Lactobacilli provide an excellent platform to express anti-HIV proteins. Broadly neutralizing antibodies have been identified against epitopes on the HIV-1 envelope and have been made into active antibody fragments. We tested single-chain variable fragment m9 and dAb-m36 and its derivative m36.4 as prototype antibodies. We cloned and expressed the antibody fragments m9, m36, and m36.4 in Lactobacillus jensenii-1153 and tested the expression levels and functionality. We made a recombinant L. jensenii 1153-1128 that expresses dAb-m36.4. All antibody fragments m9, m36, and m36.4 were expressed by lactobacilli. However, we noted the smaller m36/m36.4 were expressed to higher levels, ≥3 µg/ml. All L. jensenii-expressed antibody fragments bound to gp120/CD4 complex; Lactobacillus-produced m36.4 inhibited HIV-1BaL in a neutralization assay. Using a TZM-bl assay, we characterized the breadth of neutralization of the m36.4. Delivery of dAbs by Lactobacillus could provide passive transfer of these antibodies to the mucosa and longevity at the site of HIV-1 transmission.

Robust vaginal colonization of macaques with a novel vaginally disintegrating tablet containing a live biotherapeutic product to prevent HIV infection in women.

MucoCept is a biotherapeutic for prevention of HIV-1 infection in women and contains a human, vaginal Lactobacillus jensenii that has been genetically enhanced to express the HIV-1 entry inhibitor, modified cyanovirin-N (mCV-N). The objective of this study was to develop a solid vaginal dosage form that supports sustained vaginal colonization of the MucoCept Lactobacillus at levels previously shown, with freshly prepared cultures, to protect macaques from SHIV infection and to test this formulation in a macaque vaginal colonization model. Vaginally disintegrating tablets were prepared by lyophilizing the formulated bacteria in tablet-shaped molds, then packaging in foil pouches with desiccant. Disintegration time, potency and stability of the tablets were assessed. For colonization, non-synchronized macaques were dosed vaginally with either one tablet or five tablets delivered over five days. Vaginal samples were obtained at three, 14, and 21 days post-dosing and cultured to determine Lactobacillus colonization levels. To confirm identity of the MucoCept Lactobacillus strain, genomic DNA was extracted from samples on days 14 and 21 and a strain-specific PCR was performed. Supernatants from bacteria were tested for the presence of the mCV-N protein by Western blot. The tablets were easy to handle, disintegrated within two minutes, potent (5.7x1011 CFU/g), and stable at 4°C and 25°C. Vaginal administration of the tablets to macaques resulted in colonization of the MucoCept Lactobacillus in 66% of macaques at 14 days post-dosing and 83% after 21 days. There was no significant difference in colonization levels for the one or five tablet dosing regimens (p=0.88 Day 14, p=0.99 Day 21). Strain-specific PCR confirmed the presence of the bacteria even in culture-negative macaques. Finally, the presence of mCV-N protein was confirmed by Western blot analysis using a specific anti-mCV-N antibody.

The effect of a three-tier formulary on antidepressant utilization and expenditures.

BACKGROUND: Health plans in the United States are struggling to contain rapid growth in their spending on medications. They have responded by implementing multi-tiered formularies, which label certain brand medications 'non-preferred' and require higher patient copayments for those medications. This multi-tier policy relies on patients' willingness to switch medications in response to copayment differentials. The antidepressant class has certain characteristics that may pose problems for implementation of three-tier formularies, such as differences in which medication works for which patient, and high rates of medication discontinuation. AIMS OF THE STUDY: To measure the effect of a three-tier formulary on antidepressant utilization and spending, including decomposing spending allocations between patient and plan. METHODS: We use claims and eligibility files for a large, mature nonprofit managed care organization that started introducing its three-tier formulary on January 1, 2000, with a staggered implementation across employer groups. The sample includes 109,686 individuals who were continuously enrolled members during the study period. We use a pretest-posttest quasi-experimental design that includes a comparison group, comprising members whose employer had not adopted three-tier as of March 1, 2000. This permits some control for potentially confounding changes that could have coincided with three-tier implementation. RESULTS: For the antidepressants that became nonpreferred, prescriptions per enrollee decreased 11% in the three-tier group and increased 5% in the comparison group. The own-copay elasticity of demand for nonpreferred drugs can be approximated as -0.11. Difference-in-differences regression finds that the three-tier formulary slowed the growth in the probability of using antidepressants in the post-period, which was 0.3 percentage points lower than it would have been without three-tier. The three-tier formulary also increased out-of-pocket payments while reducing plan payments and total spending. DISCUSSION: The results indicate that the plan enrollees were somewhat responsive to the changed incentives, shifting away from the drugs that became nonpreferred. However, the intervention also resulted in cost-shifting from plan to enrollees, indicating some price-inelasticity. The reduction in the proportion of enrollees filling any prescriptions contrasts with results of prior studies for non-psychotropic drug classes. Limitations include the possibility of confounding changes coinciding with three-tier implementation (if they affected the two groups differentially); restriction to continuous enrollees; and lack of data on rebates the plan paid to drug manufacturers. IMPLICATIONS FOR HEALTH CARE PROVISION AND USE: The results of this study suggest that the impact of the three-tier formulary approach may be somewhat different for antidepressants than for some other classes. IMPLICATIONS FOR HEALTH POLICY: Policymakers should monitor the effects of three-tier programs on utilization in psychotropic medication classes. IMPLICATIONS FOR FURTHER RESEARCH: Future studies should seek to understand the reasons for patients' limited response to the change in incentives, perhaps using physician and/or patient surveys. Studies should also examine the effects of three-tier programs on patient adherence, quality of care, and clinical and economic outcomes.

Conservation of virally encoded microRNAs in Kaposi sarcoma--associated herpesvirus in primary effusion lymphoma cell lines and in patients with Kaposi sarcoma or multicentric Castleman disease.

BACKGROUND: MicroRNAs are small noncoding RNAs that posttranscriptionally regulate gene expression. Kaposi sarcoma (KS)-associated herpesvirus (KSHV) encodes 12 distinct microRNA genes, all of which are located within the latency-associated region that is highly expressed in all KSHV-associated malignancies. METHODS: We amplified, cloned, and sequenced a 2.8-kbp-long region containing a cluster of 10 microRNAs plus a 646-bp fragment of K12/T0.7 containing the remaining 2 microRNAs from 5 primary effusion lymphoma-derived cell lines and from 17 patient samples. The patients included 2 with classic KS, 12 with AIDS-KS (8 from the United States, 1 from Europe, 3 from Africa, and 4 from Central/South America), and 2 with multicentric Castleman disease (MCD). Additionally, we analyzed the K1, open reading frame 75, and K15 genes to determine KSHV subtypes, and we performed a phylogenetic analysis. RESULTS: Phylogenetic analysis of the 2.8-kbp microRNA region revealed 2 distinct clusters of sequences: a major (A/C) and a variant (B/Q) cluster. The variant cluster included sequences from 3 patients of African origin and both patients with MCD. Some microRNAs were highly conserved, whereas others had changes that could affect processing and, therefore, biological activity. CONCLUSIONS: These data demonstrate that KSHV microRNA genes are under tight selection in vivo and suggest that they contribute to the biological activity and possibly the pathogenesis of KSHV-associated malignancies.

Image denoising using total least squares.

In this paper, we present a method for removing noise from digital images corrupted with additive, multiplicative, and mixed noise. An image patch from an ideal image is modeled as a linear combination of image patches from the noisy image. We propose to fit this model to the real-world image data in the total least square (TLS) sense, because the TLS formulation allows us to take into account the uncertainties in the measured data. We develop a method to reduce the contribution from the irrelevant image patches, which will sharpen the edges and reduce edge artifacts at the same time. Although the proposed algorithm is computationally demanding, the image quality of the output image demonstrates the effectiveness of the TLS algorithms.

Engineered vaginal lactobacillus strain for mucosal delivery of the human immunodeficiency virus inhibitor cyanovirin-N.

Women are at significant risk of human immunodeficiency virus (HIV) infection, with the cervicovaginal mucosa serving as a major portal for virus entry. Female-initiated preventatives, including topical microbicides, are urgently needed to help curtail the HIV/AIDS pandemic. Here we report on the development of a novel, live microbicide that employs a natural vaginal strain of Lactobacillus jensenii engineered to deliver the potent HIV inhibitor cyanovirin-N (CV-N). To facilitate efficient expression of CV-N by this bacterium, the L. jensenii 1153 genome was sequenced, allowing identification of native regulatory elements and sites for the chromosomal integration of heterologous genes. A CV-N expression cassette was optimized and shown to produce high levels of structurally intact CV-N when expressed in L. jensenii. Lactobacillus-derived CV-N was capable of inhibiting CCR5-tropic HIV(BaL) infectivity in vitro with a 50% inhibitory concentration of 0.3 nM. The CV-N expression cassette was stably integrated as a single copy into the bacterial chromosome and resolved from extraneous plasmid DNA without adversely affecting the bacterial phenotype. This bacterial strain was capable of colonizing the vagina and producing full-length CV-N when administered intravaginally to mice during estrus phase. The CV-N-producing Lactobacillus was genetically stable when propagated in vitro and in vivo. This work represents a major step towards the development of an inexpensive yet durable protein-based microbicide to block the heterosexual transmission of HIV in women.

Real-time PCR assay for detection and quantification of hepatitis B virus genotypes A to G.

The detection and quantification of hepatitis B virus (HBV) DNA play an important role in diagnosing and monitoring HBV infection as well as assessing therapeutic response. The great variability among HBV genotypes and the enormous range of clinical HBV DNA levels present challenges for PCR-based amplification techniques. In this study, we describe the development, evaluation, and validation of a novel real-time PCR assay designed to provide accurate quantification of DNA from all eight HBV genotypes in patient plasma specimens. A computer algorithm was used to design degenerate real-time PCR primers and probes based upon a large number (n = 340) of full-length genomic sequences including HBV genotypes A to H from Europe, Africa, Asia, and North and South America. Genotype performance was tested and confirmed using 59 genotype A to G specimens from two commercially available worldwide genotype panels. This assay has a dynamic range of at least 8 log(10) without the need for specimen dilution, good clinical intra- and interassay precision, and excellent correlation with the Bayer Diagnostics VERSANT HBV DNA 3.0 (branched DNA) assay (r = 0.93). Probit analysis determined the 95% detection level was 56 IU/ml, corresponding to 11 copies per PCR well. The high sensitivity, wide linear range, good reproducibility, and genotype inclusivity, combined with a small sample volume requirement and low cost, make this novel quantitative HBV real-time PCR assay particularly well suited for application to large clinical and epidemiological studies.

Joint demosaicing and denoising.

The output image of a digital camera is subject to a severe degradation due to noise in the image sensor. This paper proposes a novel technique to combine demosaicing and denoising procedures systematically into a single operation by exploiting their obvious similarities. We first design a filter as if we are optimally estimating a pixel value from a noisy single-color (sensor) image. With additional constraints, we show that the same filter coefficients are appropriate for color filter array interpolation (demosaicing) given noisy sensor data. The proposed technique can combine many existing denoising algorithms with the demosaicing operation. In this paper, a total least squares denoising method is used to demonstrate the concept. The algorithm is tested on color images with pseudorandom noise and on raw sensor data from a real CMOS digital camera that we calibrated. The experimental results confirm that the proposed method suppresses noise (CMOS/CCD image sensor noise model) while effectively interpolating the missing pixel components, demonstrating a significant improvement in image quality when compared to treating demosaicing and denoising problems independently.

Adaptive homogeneity-directed demosaicing algorithm.

A cost-effective digital camera uses a single-image sensor, applying alternating patterns of red, green, and blue color filters to each pixel location. A way to reconstruct a full three-color representation of color images by estimating the missing pixel components in each color plane is called a demosaicing algorithm. This paper presents three inherent problems often associated with demosaicing algorithms that incorporate two-dimensional (2-D) directional interpolation: misguidance color artifacts, interpolation color artifacts, and aliasing. The level of misguidance color artifacts present in two images can be compared using metric neighborhood modeling. The proposed demosaicing algorithm estimates missing pixels by interpolating in the direction with fewer color artifacts. The aliasing problem is addressed by applying filterbank techniques to 2-D directional interpolation. The interpolation artifacts are reduced using a nonlinear iterative procedure. Experimental results using digital images confirm the effectiveness of this approach.

Demosaicing using optimal recovery.

Color images in single-chip digital cameras are obtained by interpolating mosaiced color samples. These samples are encoded in a single-chip charge-coupled device by sampling the light after it passes through a color filter array (CFA) that contains different color filters (i.e., red, green, and blue) placed in some pattern. The resulting sparsely sampled images of the three-color planes are interpolated to obtain the complete color image. Interpolation usually introduces color artifacts due to the phase-shifted, aliased signals introduced by the sparse sampling of the CFAs. This paper introduces a nonlinear interpolation scheme based on edge information that produces high-quality visual results. The new method is especially good at reconstructing the image around edges, a place where the visual human system is most sensitive.