Two Cases of Cutibacterium acnes (C acnes) Endophthalmitis Manifesting With Unusual Epiretinal Deposits.

This article describes two cases of delayed-onset Cutibacterium acnes (C acnes) endophthalmitis 1 month after cataract surgery manifesting with unusual epiretinal deposits. Both patients were referred for persistent inflammation after cataract surgery. After failing to respond to a vitreous tap with injection of antibiotics, the patients underwent pars plana vitrectomy and in one of the cases partial posterior capsulectomy for a posterior capsular plaque. Intraoperatively, both cases were found to have unusual multifocal epiretinal deposits. The clinical presentations described here represent a highly unique manifestation of C acnes endophthalmitis distinct from the classic anterior segment findings. [Ophthalmic Surg Lasers Imaging Retina. 2022;53:164-167.].

Bile acid biosynthesis in Smith-Lemli-Opitz syndrome bypassing cholesterol: Potential importance of pathway intermediates.

Bile acids are the end products of cholesterol metabolism secreted into bile. They are essential for the absorption of lipids and lipid soluble compounds from the intestine. Here we have identified a series of unusual Δ5-unsaturated bile acids in plasma and urine of patients with Smith-Lemli-Opitz syndrome (SLOS), a defect in cholesterol biosynthesis resulting in elevated levels of 7-dehydrocholesterol (7-DHC), an immediate precursor of cholesterol. Using liquid chromatography - mass spectrometry (LC-MS) we have uncovered a pathway of bile acid biosynthesis in SLOS avoiding cholesterol starting with 7-DHC and proceeding through 7-oxo and 7ß-hydroxy intermediates. This pathway also occurs to a minor extent in healthy humans, but elevated levels of pathway intermediates could be responsible for some of the features SLOS. The pathway is also active in SLOS affected pregnancies as revealed by analysis of amniotic fluid. Importantly, intermediates in the pathway, 25-hydroxy-7-oxocholesterol, (25R)26-hydroxy-7-oxocholesterol, 3ß-hydroxy-7-oxocholest-5-en-(25R)26-oic acid and the analogous 7ß-hydroxysterols are modulators of the activity of Smoothened (Smo), an oncoprotein that mediates Hedgehog (Hh) signalling across membranes during embryogenesis and in the regeneration of postembryonic tissue. Computational docking of the 7-oxo and 7ß-hydroxy compounds to the extracellular cysteine rich domain of Smo reveals that they bind in the same groove as both 20S-hydroxycholesterol and cholesterol, known activators of the Hh pathway.

Metabolism of Non-Enzymatically Derived Oxysterols: Clues from sterol metabolic disorders.

Cholestane-3ß,5α,6ß-triol (3ß,5α,6ß-triol) is formed from cholestan-5,6-epoxide (5,6-EC) in a reaction catalysed by cholesterol epoxide hydrolase, following formation of 5,6-EC through free radical oxidation of cholesterol. 7-Oxocholesterol (7-OC) and 7ß-hydroxycholesterol (7ß-HC) can also be formed by free radical oxidation of cholesterol. Here we investigate how 3ß,5α,6ß-triol, 7-OC and 7ß-HC are metabolised to bile acids. We show, by monitoring oxysterol metabolites in plasma samples rich in 3ß,5α,6ß-triol, 7-OC and 7ß-HC, that these three oxysterols fall into novel branches of the acidic pathway of bile acid biosynthesis becoming (25R)26-hydroxylated then carboxylated, 24-hydroxylated and side-chain shortened to give the final products 3ß,5α,6ß-trihydroxycholanoic, 3ß-hydroxy-7-oxochol-5-enoic and 3ß,7ß-dihydroxychol-5-enoic acids, respectively. The intermediates in these pathways may be causative of some phenotypical features of, and/or have diagnostic value for, the lysosomal storage diseases, Niemann Pick types C and B and lysosomal acid lipase deficiency. Free radical derived oxysterols are metabolised in human to unusual bile acids via novel branches of the acidic pathway, intermediates in these pathways are observed in plasma.

High Rate of Serotype V Streptococcus agalactiae Carriage in Pregnant Women in Botswana.

Maternal rectovaginal colonization is the major risk factor for early-onset neonatal sepsis due to Group B Streptococcus (GBS), a major cause of early life morbidity and mortality. Transmission generally occurs perinatally from colonized mothers to infants. Vaccines targeting a subset of GBS serotypes are under development, but GBS epidemiology remains poorly understood in many African nations. We performed a cross-sectional study of GBS colonization among pregnant women at two sites in Botswana, a country with minimal prior GBS carriage data. We found a rectovaginal colonization rate of 19%, comparable with studies in other regions; however, we also noted a striking predominance of serotype V (> 45% of strains). Although further studies are required to delineate the burden of invasive GBS disease in Botswana and the generalizability of type V epidemiology, these data provide a useful baseline for understanding the potential local impact of GBS prevention strategies, including vaccines.

Improving the Sensitivity of Real-time PCR Detection of Group B Streptococcus Using Consensus Sequence-Derived Oligonucleotides.

Group B Streptococcus (GBS) is a perinatal pathogen and an emerging cause of disease in adults. Culture-independent GBS detection relies on polymerase chain reaction (PCR) of conserved genes, including sip. We demonstrate suboptimal sensitivity of the existing sip PCR strategy and validate an improved method based on consensus sequences from >100 GBS genomes.

Identification of unusual oxysterols and bile acids with 7-oxo or 3ß,5α,6ß-trihydroxy functions in human plasma by charge-tagging mass spectrometry with multistage fragmentation.

7-Oxocholesterol (7-OC), 5,6-epoxycholesterol (5,6-EC), and its hydrolysis product cholestane-3ß,5α,6ß-triol (3ß,5α,6ß-triol) are normally minor oxysterols in human samples; however, in disease, their levels may be greatly elevated. This is the case in plasma from patients suffering from some lysosomal storage disorders, e.g., Niemann-Pick disease type C, or the inborn errors of sterol metabolism, e.g., Smith-Lemli-Opitz syndrome and cerebrotendinous xanthomatosis. A complication in the analysis of 7-OC and 5,6-EC is that they can also be formed ex vivo from cholesterol during sample handling in air, causing confusion with molecules formed in vivo. When formed endogenously, 7-OC, 5,6-EC, and 3ß,5α,6ß-triol can be converted to bile acids. Here, we describe methodology based on chemical derivatization and LC/MS with multistage fragmentation (MSn) to identify the necessary intermediates in the conversion of 7-OC to 3ß-hydroxy-7-oxochol-5-enoic acid and 5,6-EC and 3ß,5α,6ß-triol to 3ß,5α,6ß-trihydroxycholanoic acid. Identification of intermediate metabolites is facilitated by their unusual MSn fragmentation patterns. Semiquantitative measurements are possible, but absolute values await the synthesis of isotope-labeled standards.

Charge-tagging liquid chromatography-mass spectrometry methodology targeting oxysterol diastereoisomers.

The introduction of a hydroxy group to the cholesterol skeleton introduces not only the possibility for positional isomers but also diastereoisomers, where two or more isomers have different configurations at one or more of the stereocentres but are not mirror images. The differentiation of diastereoisomers is important as differing isomers can have differing biochemical properties and are formed via different biochemical pathways. Separation of diasterioisomers is not always easy by chromatographic methods Here we demonstrate, by application of charge-tagging and derivatisation with the Girard P reagent, the separation and detection of biologically relevant diastereoisomers using liquid chromatography - mass spectrometry with multistage fragmentation.

Cholesterolomics: An update.

Cholesterolomics can be regarded as the identification and quantification of cholesterol, its precursors post squalene, and metabolites of cholesterol and of its precursors, in a biological sample. These molecules include 1,25-dihydroxyvitamin D3, steroid hormones and bile acids and intermediates in their respective biosynthetic pathways. In this short article we will concentrate our attention on intermediates in bile acid biosynthesis pathways, in particular oxysterols and cholestenoic acids. These molecular classes are implicated in the aetiology of a diverse array of diseases including autoimmune disease, Parkinson's disease, motor neuron disease, breast cancer, the lysosomal storage disease Niemann-Pick type C and the autosomal recessive disorder Smith-Lemli-Opitz syndrome. Mass spectrometry (MS) is the dominant technology for sterol analysis including both gas-chromatography (GC)-MS and liquid chromatography (LC)-MS and more recently matrix-assisted laser desorption/ionisation (MALDI)-MS for tissue imaging studies. Here we will discuss exciting biological findings and recent analytical improvements.

Current trends in oxysterol research.

In this short review we provide a synopsis of recent developments in oxysterol research highlighting topics of current interest to the community. These include the involvement of oxysterols in neuronal development and survival, their participation in the immune system, particularly with respect to bacterial and viral infection and to Th17-cell development, and the role of oxysterols in breast cancer. We also discuss the value of oxysterol analysis in the diagnosis of disease.

OPTICAL COHERENCE TOMOGRAPHY ANGIOGRAPHY AND WIDEFIELD FUNDUS AUTOFLUORESCENCE IN PUNCTATE INNER CHOROIDOPATHY.

PURPOSE: To present the optical coherence tomography angiography and widefield fundus autofluorescence (FAF) imaging findings of a 34-year-old woman with punctate inner choroidopathy and Type 2 neovascularization. METHODS: In the retrospective case report, optical coherence tomography angiography imaging was performed with the RTVue XR Avanti (Optovue Inc, Fremont, CA) platform, which uses split spectrum amplitude decorrelation angiography to detect flow. PATIENTS: Single patient with the diagnosis of punctate inner choroidopathy. RESULTS: Spectral domain optical coherence tomography of the macula showed an inactive submacular scar but optical coherence tomography angiography demonstrated the fine detailed structure of a Type 2 neovascular membrane. Peripheral FAF abnormalities were detected despite the notable absence of these lesions with clinical examination and color fundus photography. These FAF findings were important in guiding clinical follow-up and response to therapy. The FAF lesions resolved with normal follow-up and treatment with oral naproxen. DISCUSSION: This is the first case report of optical coherence tomography angiography imaging of choroidal neovascularization associated with punctate inner choroidopathy and the first report of widefield FAF abnormalities in punctate inner choroidopathy. Optical coherence tomography angiography may provide a more noninvasive and sensitive imaging system for the evaluation of choroidal neovascularization than traditional fluorescein angiography and may provide a direct gauge of response to therapy. Widefield FAF may improve detection of inflammatory activity of retinal disease and monitoring of disease activity.

Development of the human pancreas from foregut to endocrine commitment.

Knowledge of human pancreas development underpins our interpretation and exploitation of human pluripotent stem cell (PSC) differentiation toward a ß-cell fate. However, almost no information exists on the early events of human pancreatic specification in the distal foregut, bud formation, and early development. Here, we have studied the expression profiles of key lineage-specific markers to understand differentiation and morphogenetic events during human pancreas development. The notochord was adjacent to the dorsal foregut endoderm during the fourth week of development before pancreatic duodenal homeobox-1 detection. In contrast to the published data from mouse embryos, during human pancreas development, we detected only a single-phase of Neurogenin 3 (NEUROG3) expression and endocrine differentiation from approximately 8 weeks, before which Nirenberg and Kim homeobox 2.2 (NKX2.2) was not observed in the pancreatic progenitor cell population. In addition to revealing a number of disparities in timing between human and mouse development, these data, directly assembled from human tissue, allow combinations of transcription factors to define sequential stages and differentiating pancreatic cell types. The data are anticipated to provide a useful reference point for stem cell researchers looking to differentiate human PSCs in vitro toward the pancreatic ß-cell so as to model human development or enable drug discovery and potential cell therapy.

Centriolar association of ALMS1 and likely centrosomal functions of the ALMS motif-containing proteins C10orf90 and KIAA1731.

Mutations in the human gene ALMS1 cause Alström syndrome, a rare progressive condition characterized by neurosensory degeneration and metabolic defects. ALMS1 protein localizes to the centrosome and has been implicated in the assembly and/or maintenance of primary cilia; however its precise function, distribution within the centrosome, and mechanism of centrosomal recruitment are unknown. The C-terminus of ALMS1 contains a region with similarity to the uncharacterized human protein C10orf90, termed the ALMS motif. Here, we show that a third human protein, the candidate centrosomal protein KIAA1731, contains an ALMS motif and that exogenously expressed KIAA1731 and C10orf90 localize to the centrosome. However, based on deletion analysis of ALMS1, the ALMS motif appears unlikely to be critical for centrosomal targeting. RNAi analyses suggest that C10orf90 and KIAA1731 have roles in primary cilium assembly and centriole formation/stability, respectively. We also show that ALMS1 localizes specifically to the proximal ends of centrioles and basal bodies, where it colocalizes with the centrosome cohesion protein C-Nap1. RNAi analysis reveals markedly diminished centrosomal levels of C-Nap1 and compromised cohesion of parental centrioles in ALMS1-depleted cells. In summary, these data suggest centrosomal functions for C10orf90 and KIAA1731 and new centriole-related functions for ALMS1.

Control of disseminated scedosporium prolificans infection and endophthalmitis.

PURPOSE: To report a case of endogenous endophthalmitis as the presenting sign of a disseminated Scedosporium prolificans infection. METHODS: A 38-year-old woman on chronic immunosuppressive treatment for a cardiac transplant was examined for blurry vision in her right eye. Dilated fundus examination in the right eye revealed a large serous retinal detachment with subretinal cream-colored masses. RESULTS: Complete pars plana vitrectomy with vitreal biopsies was performed and revealed S. prolificans on culture. Multiple intravitreal voriconazole injections as well as systemic voriconazole and terbinafine resulted in control and suppression of both the ocular and systemic disease. CONCLUSION: Scedosporium prolificans endogenous endophthalmitis represents a difficult to treat opportunistic infection in immunocompromised patients. However, newer-generation antifungals offer the potential for suppression of systemic disease and the possibility of avoiding enucleation in the setting of endophthalmitis.

Dark hypopyon in Streptococcus bovis endogenous endophthalmitis: clinicopathologic correlations.

PURPOSE: The aim of this report is to present a previously unreported causative organism associated with brown-pigmented hypopyon in a patient with endophthalmitis. METHODS: This is a retrospective case report which includes clinicopathologic correlations. RESULTS: Vitreous cultures demonstrated Streptococcus bovis infection resulting in a brown-pigmented hypopyon, with uveal pigment found intra- and extracellularly on pathologic examination of the pupillary membrane. CONCLUSIONS: S. bovis endophthalmitis may be a cause of dark hypopyon, especially in patients with a history of liver disease, and, when identified, warrants colonoscopy and cardiac workup.

Simultaneous presentation of Kimura disease and angiolymphoid hyperplasia with eosinophilia.

Kimura disease and angiolymphoid hyperplasia with eosinophilia are rare disease entities that can manifest as subcutaneous nodules, plaques, or papules of the head and neck, and less commonly the orbit and ocular adnexa. The merits of each entity have been debated with regard to whether they are truly individual or are separate points on a single spectrum. Current opinion favors the former. This clinicopathologic report of a patient who presented with a right lower eyelid mass with pathologic features consistent with Kimura disease and a left conjunctival mass with features consistent with angiolymphoid hyperplasia with eosinophilia poses a challenge to the notion that these are distinct conditions.

Amniotic membrane transplantation for intraoperative conjunctival repair during trabeculectomy with mitomycin C.

PURPOSE: To describe the use of amniotic membrane (AM) transplantation for the repair of conjunctival buttonholes during trabeculectomy with mitomycin C. METHODS: Four eyes of 3 patients with thin conjunctiva, precluding watertight conjunctival closure at the incision site with suturing, underwent intraoperative AM grafts over the leaking areas. RESULTS: In all eyes, a functional, nonleaking bleb was achieved. At the latest follow-up (8 to 30 mo), all eyes had intraocular pressures of 12 mm Hg or less without medications. CONCLUSIONS: AM transplantation was effective in the intraoperative closure of conjunctival buttonholes which developed at the incision line in 4 eyes. This intervention may be a useful addition to the glaucoma surgeon's repertoire of surgical techniques.

Electrical stimulation in normal and retinal degeneration (rd1) isolated mouse retina.

Stimulus threshold and response latencies were measured for electrically elicited retinal ganglion cell responses in retina isolated from the eyes of normal and retinal degenerate (rd1) mice. Stimulation of the ganglion cell-side in normal retina yielded a significantly lower mean threshold and shorter latency when compared with stimulation of the photoreceptor side in normal retina. The latency of the ganglion cell-side stimulation in normal retina also proved to be significantly shorter than the latency for stimulation of the ganglion cell side in rd1 retina. Thus both the electrode positioning as well as the health of the retinal tissue play a role in the stimulating current required to elicit a retinal response.

Implementation of a quality systems approach for laboratory practice in resource-constrained countries.

Under the direction of the US Global AIDS Coordinator's Office, Department of Health and Human Services, the CDC Global AIDS programme helps resource-constrained countries to address the global HIV/AIDS pandemic. Activities include laboratory capacity and laboratory infrastructure development in 25 resource-constrained countries. Medical practitioners and public health programme leaders in industrialized countries rely on the use of quality laboratory data for evidence-based medical decision-making to determine policy for the implementation of disease control measures, to monitor disease to determine the impact of control programmes, and to support surveillance activities. In these countries, laboratory data to support decision-making processes have a level of quality attributable to laws, regulations and guidelines developed over many years. However, resource-constrained countries have not had similar experiences. Few countries have developed laws, regulations or guidelines, nor is there a data-use culture (e.g. evidence-based medicine) for those in the decision-making environment in resource-constrained countries. The strengthening of laboratory capability and capacity in resource-constrained countries is an important goal to improve accurate and reliable data for the diagnosis, treatment and monitoring of disease.A process for the implementation of a quality systems approach for a laboratory is presented: (i) acknowledgement of the need to improve the laboratory programme in the country at the Ministry of Health and at all decision-making levels within the provinces/states of the country; (ii) assessment of capabilities, capacities, infrastructure, and training needs; (iii) implementation of a national meeting of laboratorians; (iv) designation of a national Quality Assurance Office and leadership within that office; (v) the development and provision of technical training.