Effects of Bifidobacterium Pseudocatenulatum G4 on Clostridium Scindens and Clostridium Hiranonis

The purpose of this study was to investigate the deoxycholic acid (DCA) reduction by Bifidobacterium pseudocatenulatum G4 at in vitro conditions with the emphasis of its bile salt deconjugation ability and antagonistic activity against 7α-dehydroxylating bacteria. B. pseudocatenulatum G4 showed antagonistic activity against Clostridium scindens and C. hiranonis at colonic pH (5.7, 6.2 and 6.8), with highest activity (4mm inhibition zone) at pH 5.7 against C. hiranonis. Growth rates of B. pseudocatenulatum G4 (time required for the initial absorbance at zero time to increase by 0.3 units), decreased in the presence of 2.0% oxgall compared to absence of oxgall. Effect of B. pseudocatenulatum G4 on pH reduction was measured; B. pseudocatenulatum G4 reduced pH from 6.8 to 3.9 in TPY broth supplemented with 0.1% and pH 3.74 in TPY broth without oxgall (control), compared to pH 5.28 in TPY broth supplemented with 2.0% oxgall. This results showed oxgall exert inhibitory effect on pH reduction by B. pseudocatenulatum G4. Bile salt hydrolase (BSH) activity, which is the measurement of enzyme activity responsible for bile salt deconjugation, was quantified by high pressure liquid chromatography (HPLC) assay. B. pseudocatenulatum G4 gave deconjugation rate (disappearance of conjugated bile acid) in TPY broth supplemented with 0.25 mM (the highest concentration in colon) and 5.0 mM (the highest concentration in small intestine) of all 6 different conjugated bile acids (TCA, GCA, TCDCA, GCDCA, TDCA, and GDCA). Generally B. pseudocatenulatum G4 deconjugated glycoconjugated bile acids in higher amount compare to tauroconjugates. The percentage of deconjugation activity was higher in TPY medium supplemented with 0.25mM bile acids compared to TPY broth with 5.0mM bile acids. On other hand, pH 6.2 was observed as the optimum pH for BSH activity. Full factorial statistical design was used to evaluate the effects of three factors (pH, B. pseudocatenulatum G4 concentrations and Clostridiums) with different levels and interactions between them, on DCA production. Statistical analysis for results of the in vitro experiment showed that total DCA concentration produced by C. scindens and C. hiranonis in the presence of B. pseudocatenulatum G4 and low pH, were lower than the control group with highest pH and in the absence of B. pseudocatenulatum G4

Vedel-objektiiv abil salvestatud kaugseire piltide analüüs kasutades super-resolutsiooni meetodeid

Väitekirja elektrooniline versioon ei sisalda publikatsiooneKäesolevas doktoritöös uuriti nii riist- kui ka tarkvaralisi lahendusi piltide töötlemiseks. Riist¬varalise poole pealt pakuti lahenduseks uudset vedelläätse, milles on dielekt¬rilisest elastomeerist kihilise täituriga membraan otse optilisel teljel. Doktoritöö käigus arendati välja kaks prototüüpi kahe erineva dielektrilisest elastomeerist ki¬hilise täituriga, mille aktiivne ala oli ühel juhul 40 ja teisel 20 mm. Läätse töö vas¬tas elastomeeri deformatsiooni mehaanikale ja suhtelistele muutustele fookuskau¬guses. Muutuste demonstreerimiseks meniskis ja läätse fookuskauguse mõõtmiseks kasutati laserkiirt. Katseandmetest selgub, et muutuste tekitamiseks on vajalik pinge vahemikus 50 kuni 750 volti. Tarkvaralise poole pealt pakuti uut satelliitpiltide parandamise süsteemi. Paku¬tud süsteem jagas mürase sisendpildi DT-CWT laineteisenduse abil mitmeteks sagedusalamribadeks. Pärast müra eemaldamist LA-BSF funktsiooni abil suu¬rendati pildi resolutsiooni DWT-ga ja kõrgsagedusliku alamriba piltide interpo¬leerimisega. Interpoleerimise faktor algsele pildile oli pool sellest, mida kasutati kõrgsagedusliku alamriba piltide interpoleerimisel ning superresolutsiooniga pilt rekonst¬rueeriti IDWT abil. Käesolevas doktoritöös pakuti tarkvaraliseks lahenduseks uudset sõnastiku baasil töötavat super-resolutsiooni (SR) meetodit, milles luuakse paarid suure resolutsiooniga (HR) ja madala resolut-siooniga (LR) piltidest. Kõigepealt jagati vastava sõnastiku loomiseks HR ja LR paarid omakorda osadeks. Esialgse HR kujutise saamiseks LR sisendpildist kombineeriti HR osi. HR osad valiti sõnastikust nii, et neile vastavad LR osad oleksid võimalikult lähedased sisendiks olevale LR pil¬dile. Iga valitud HR osa heledust korrigeeriti, et vähendada kõrvuti asuvate osade heleduse erine¬vusi superresolutsiooniga pildil. Plokkide efekti vähendamiseks ar¬vutati saadud SR pildi keskmine ning bikuupinterpolatsiooni pilt. Lisaks pakuti käesolevas doktoritöös välja kernelid, mille tulemusel on võimalik saadud SR pilte teravamaks muuta. Pakutud kernelite tõhususe tõestamiseks kasutati [83] ja [50] poolt pakutud resolutsiooni parandamise meetodeid. Superreso¬lutsiooniga pilt saadi iga kerneli tehtud HR pildi kombineerimise teel alpha blen¬dingu meetodit kasutades. Pakutud meetodeid ja kerneleid võrreldi erinevate tavaliste ja kaasaegsete meetoditega. Kvantita-tiivsetest katseandmetest ja saadud piltide kvaliteedi visuaal¬sest hindamisest selgus, et pakutud meetodid on tavaliste kaasaegsete meetoditega võrreldes paremad.In this thesis, a study of both hardware and software solutions for image enhance¬ment has been done. On the hardware side, a new liquid lens design with a DESA membrane located directly in the optical path has been demonstrated. Two pro¬totypes with two different DESA, which have a 40 and 20 mm active area in diameter, were developed. The lens performance was consistent with the mechan¬ics of elastomer deformation and relative focal length changes. A laser beam was used to show the change in the meniscus and to measure the focal length of the lens. The experimental results demonstrate that voltage in the range of 50 to 750 V is required to create change in the meniscus. On the software side, a new satellite image enhancement system was proposed. The proposed technique decomposed the noisy input image into various frequency subbands by using DT-CWT. After removing the noise by applying the LA-BSF technique, its resolution was enhanced by employing DWT and interpolating the high-frequency subband images. An original image was interpolated with half of the interpolation factor used for interpolating the high-frequency subband images, and the super-resolved image was reconstructed by using IDWT. A novel single-image SR method based on a generating dictionary from pairs of HR and their corresponding LR images was proposed. Firstly, HR and LR pairs were divided into patches in order to make HR and LR dictionaries respectively. The initial HR representation of an input LR image was calculated by combining the HR patches. These HR patches are chosen from the HR dictionary corre-sponding to the LR patches that have the closest distance to the patches of the in¬put LR image. Each selected HR patch was processed further by passing through an illumination enhancement processing order to reduce the noticeable change of illumination between neighbor patches in the super-resolved image. In order to reduce the blocking effect, the average of the obtained SR image and the bicubic interpolated image was calculated. The new kernels for sampling have also been proposed. The kernels can improve the SR by resulting in a sharper image. In order to demonstrate the effectiveness of the proposed kernels, the techniques from [83] and [50] for resolution enhance¬ment were adopted. The super-resolved image was achieved by combining the HR images produced by each of the proposed kernels using the alpha blending tech-nique. The proposed techniques and kernels are compared with various conventional and state-of-the-art techniques, and the quantitative test results and visual results on the final image quality show the superiority of the proposed techniques and ker¬nels over conventional and state-of-art technique

Point-of-care diagnostics of childhood central nervous system infections, with a focus on usability in low-resource settings

Background: The inaccessibility of laboratory services sustains the high burden of paediatric infectious diseases, such as central nervous system (CNS) infections, in low-resource settings. New contextually fit and well-implemented point-of-care tests (POCTs) could relieve such a burden and narrow the diagnostic divide between rich and poor. Yet, current disengagement between product developers, end-users, and implementors of POCTs impedes their clinical use and utility in low-resource settings. Also, the lack of evidence gathered through field evaluations of many diagnostic instruments in low-resource settings raises questions of their clinical utility there. Objectives: The main aim of this thesis was to provide clinical and contextual guidance for developers of new POCTs for CNS infection diagnosis with high utility, especially in low-resource settings; and to implementors of POCT services towards their optimized clinical benefit. This was addressed through a multidisciplinary combination of qualitative, laboratory, and clinical studies. Methods: Qualitative focus group discussions were conducted with health care workers (HCW) in Mbarara, Uganda (Paper I), and in Stockholm, Sweden (Paper III). Discussions were audio recorded and transcribed verbatim. Qualitative content analysis with an inductive approach was pursued in for data analysis. Comparisons between the two settings were discussed. In Paper II, a vertical flow DNA microarray printed on paper was developed for the detection of Neisseria meningitidis – a major aetiology of paediatric bacterial CNS infection worldwide. The analytical performance of the microarray was laboratory evaluated on DNA extracted from the bacteria, through the detection of the ctrA gene sequence specific to N. meningitidis. In Paper IV, a commercially available polymerase chain reaction (PCR) instrument with the capability of multiplex single-sample cerebrospinal fluid (CSF) microbiology was prospectively field-evaluated for the diagnosis of paediatric CNS infection in Mbarara, Uganda. Clinical turnaround time (cTAT) was defined as time spent from lumbar puncture until reporting of microbiology analyses to clinicians. The PCR instrument’s influence on clinical and patient-centered outcomes (yield, cTAT, duration of hospitalization and antibiotic exposure, patient outcome) was compared to that of bacterial culture. Results: Fifty and 24 HCWs of different professions participated in the qualitative studies in Mbarara and Stockholm, respectively, expressing greater similarities than differences in perspectives of POCT use. POCTs were routinely used at both sites and credited for facilitating differential diagnostics and clinical decision-making. While the Ugandan setting with low laboratory accessibility was highly dependent on POCTs for sample analyses, the Swedish setting credited their use for having clinical and social value. Contrary to the described beneficial aspects, current POCTs were deemed contextually unfit in Mbarara, and their use to cause clinical distraction in Stockholm. Deficient implementation of POCT services was exposed in both places. Requests for ideal POCTs were aligned with those stipulated by the ‘ASSURED’ criteria of the World Health Organization. Specific POCTs for infectious diseases, including CNS infections, were requested. The laboratory study demonstrated an analytical sensitivity of 38 copies of ctrA per assay, with high specificity. The clinical study enrolled 212 children aged 0-12 years who were suspected of having CNS infection, with 193 of them being evaluated using the commercially available PCR instrument. A vast majority of children had been pre-administered antibiotics prior to lumbar puncture. Bacterial yield for the instrument was 12 % vs. 1.5 % for culture, with the addition of the instrument’s detection of viruses in 23 samples. Median cTAT for the instrument was 4.2 hours vs. 2 days for culture. Use of the instrument was associated with a statistically significant shorter antibiotic exposure of bacteria-negative vs. positive patients of five days, measured as from the time of reporting of laboratory results to the responsible clinicians. Similarly, its use was associated with a significantly shorter hospitalization for all-negative patients (five days) compared to those with any microorganism detected by it. No statistically significant differences in patient outcome were found due to its use, nor by its detection of any microorganisms. Conclusion: Point-of-care tests provide laboratory means to settings without laboratory capacity and to situations in need of timely results, and we could show how rapid molecular methods for CSF analysis could benefit paediatric children with suspected CNS infection. Yet, without any observed benefits in patient outcome, and at a cost not financially bearable in most lowresource settings. Contextually fit POCTs for paediatric CNS infections are needed in lowresource settings. Yet, there are design flaws in current POCTs and in implementations for their use, limiting their clinical benefits. Collaborative engagement of product developers, clinicians, laboratory professionals, and health policymakers would better serve low-resource settings with contextually fit POCTs and allow for their optimized implementation. The ‘POCTEST' framework for such an engagement is proposed in this thesis. Finally, as we provided proof of concept for a newly developed paper-printed molecular method, we will pursue its development towards contextual clinical utility in low-resource settings. Should we succeed, we hope to contribute to a decrease in preventable childhood mortality in such settings