Chun-Yiu Law

Fatal viral infection-associated encephalopathy in two Chinese boys : a genetically determined risk factor of thermolabile carnitine palmitoyltransferase II variants

Influenza-associated encephalopathy (IAE) is a potentially fatal neurological complication of influenza infection usually in the presence of high and persistent fever. Thermolabile carnitine palmitoyltransferase II enzyme (CPT-II) predisposes IAE, so far only described in Japanese. As the genetic origins of Japanese and Chinese are alike, similar genetic risk factors in CPT-II are expected. We report the first two unrelated Chinese patients of thermolabile CPT-II variants that underlain the persistent high fever-triggered viral infection-associated encephalopathy, multi-organ failure and death. Elevated (C16:0+C18:1)/C2 acylcarnitines ratio and the CPT2 susceptibility variant allele [p.Phe352Cys; p.Val368Ile] were detected. The asymptomatic family members of one patient also had abnormal long-chain acylcarnitines. In our experience of biochemical genetics, the elevated (C16:0+C18:1)/C2 acylcarnitines ratio is unusual and specific for thermolabile CPT-II variants. Allele frequency of [p.Phe352Cys; p.Val368Ile] among Hong Kong Chinese was 0.104, similar to Japanese data, and [p.Phe352Cys] has not been reported in Caucasians. This may explain the Asian-specific phenomenon of thermolabile CPT-II-associated IAE. We successfully demonstrated the thermolabile CPT-II variants in patients with viral infection-associated encephalopathy in another Asian population outside Japanese. The condition is likely under-recognized. With our first cases, it is envisaged that more cases will be diagnosed in subsequent years. The exact pathogenic mechanism of how other factors interplay with thermolabile CPT-II variants and high fever leading to IAE, is yet to be elucidated. Fasting and decreased intake during illness may aggravate the disease. Further studies including high risk and neonatal screening are warranted to investigate its expressivity, penetrance and temperature-dependent behaviors in thermolabile CPT-II carriers. This may lead to discovery of the therapeutic golden window by aggressive antipyretics and L-carnitine administration in avoiding the high mortality and morbidity of IAE.

NMR-based metabolomic urinalysis: a rapid screening test for urinary tract infection.

BACKGROUNDnUrinary tract infection (UTI) is one of the most common bacterial infections in humans; however, there is no accurate and fast quantitative test to detect UTI. Dipstick urinalysis is semi-quantitative with a limited diagnostic accuracy, while urine culture is accurate but takes time. We described a quantitative biochemical method for the diagnosis of bacteriuria using a single marker.nnnMETHODSnWe compared the urine metabolomes from 88 patients with bacterial UTI and 61 controls using (1)H NMR spectroscopy followed by principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA). The biomarker identified was subsequently validated using independent samples.nnnRESULTSnThe urine acetic acid/creatinine (mmol/mmol) level was determined to be the most discriminatory marker for bacterial UTI with an area-under-receiver operating characteristic curve=0.97, sensitivity=91% and specificity=95% at the optimal cutoff 0.03 mmol/mmol. For validation, 60 samples were recruited prospectively. Using the optimal cutoff for acetic acid/creatinine, this method showed sensitivity=96%, specificity=94%, positive predictive value=92%, negative predictive value=97% and an overall accuracy=95%. The diagnostic performance was superior to dipstick urinalysis or microscopy. In addition, we also observed an increase of urinary trimethylamine (TMA) in patients with Escherichia coli-associated UTI. TMA is a mammalian-microbial co-metabolite and the high level of TMA generated is related to the bacterial enzyme, trimethylamine N-oxide (TMAO) reductase which reduces TMAO to TMA.nnnCONCLUSIONSnUrine acetic acid is a neglected metabolite that can be used for rapid diagnosis of UTI and TMA can be used for etiologic diagnosis of UTI. With the introduction of NMR-based clinical analyzers to clinical laboratories, NMR-based urinalysis can be translated for clinical use.

Quantitative metabolomics of urine for rapid etiological diagnosis of urinary tract infection: Evaluation of a microbial–mammalian co-metabolite as a diagnostic biomarker

BACKGROUNDnWe have previously reported a NMR-based urinalysis for the screening of urinary tract infection (UTI) with high accuracy and reproducibility. Urinary acetic acid per creatinine was found to be a diagnostic marker of bacterial UTI with an area-under-receiver operating characteristic (ROC) curve of 0.97. In addition, we identified trimethylamine (TMA) as a human-microbial marker of Escherichia coli (EC)-associated UTI. Here, we evaluate the clinical application of NMR-based urinalysis in aiding the etiological diagnosis of bacterial UTI.nnnMETHODSnProton NMR spectroscopy was acquired using a Bruker 600MHz spectroscopy for 88 urine samples from patients with bacterial UTI, confirmed by urine culture. The spectra were analyzed using orthogonal partial least squares-discriminant analysis (OPLS-DA). ROC curve analysis was performed after the quantitation of the urine metabolites.nnnRESULTSnThe TMA/creatinine (mmol/mmol) level was determined to be a specific marker for EC-associated UTI. It has an area-under-ROC=0.85 (95% confidence interval: 0.75-0.91). For the etiological diagnosis, the cutoff for 97.0% specificity was at 0.0117mmol/mmol creatinine for EC-associated UTI with a sensitivity of 66.7%. The mean of TMA/creatinine of EC is 21-fold that of non-EC.nnnCONCLUSIONSnThe co-metabolism of TMA by EC and human cells makes TMA an ideal urine biomarker for UTI. The presence of TMA in a freshly collected sample eliminates the possibility of contamination of urine by bacteria during the collection process resulting in a positive bacterial culture result. We envisage the NMR-based urinalysis of urinary TMA that can be a useful method for the etiological diagnosis of EC-associated UTI.

Untargeted mass spectrometry-based metabolomic profiling of pleural effusions: fatty acids as novel cancer biomarkers for malignant pleural effusions.

Untargeted mass spectrometry-based metabolomic profiling is a powerful analytical method used for broad-spectrum identification and quantification of metabolites in biofluids in human health and disease states. In this study, we exploit metabolomic profiling for cancer biomarker discovery for diagnosis of malignant pleural effusions. We envisage the result will be clinically useful since currently there are no cancer biomarkers that are accurate enough for the diagnosis of malignant pleural effusions. Metabolomes of 32 malignant pleural effusions from lung cancer patients and 18 benign effusions from patients with pulmonary tuberculosis were analyzed using reversed-phase liquid chromatography tandem mass spectrometry (LC-MS/MS) using AB SCIEX TripleTOF 5600. MS spectra were analyzed using XCMS, PeakView, and LipidView. Metabolome-Wide Association Study (MWAS) was performed by Receiver Operating Characteristic Curve Explorer and Tester (ROCCET). Insignificant markers were filtered out using a metabolome-wide significance level (MWSL) with p-value < 2 × 10(-5) for t test. Only compounds in Human Metabolome Database (HMDB) will be used as cancer biomarkers. ROCCET analysis of ESI positive and negative MS spectra revealed free fatty acid (FFA) 18:1 (oleic acid) had the largest area-under-ROC of 0.96 (95% CI = 0.87-1.00) in malignant pleural effusions. Using a ratio of FFA 18:1-to-ceramide (d18:1/16:0), the area-under-ROC was further increased to 0.99 (95% CI = 0.91-1.00) with sensitivity 93.8% and specificity 100.0%. Using untargeted metabolomic profiling, the diagnostic cancer biomarker with the largest area-under-ROC can be determined objectively. This lipogenic phenotype could be explained by overexpression of fatty acid synthase (FASN) in cancer cells. The diagnostic performance of FFA 18:1-to-ceramide (d18:1/16:0) ratio supports its use for diagnosis of malignant pleural effusions.

Clinical whole-exome sequencing reveals a novel missense pathogenic variant of GNAO1 in a patient with infantile-onset epilepsy

BACKGROUNDnThe cause of infantile-onset epilepsy is complex and is not easily recognized clinically, particularly in paediatric patients who present with non-specific neurological signs, no radiological abnormalities and no metabolic changes.nnnCASEnWe report a case of infantile-onset epilepsy in a 10-month-old Chinese girl who presented with non-specific neurological signs, no radiological abnormalities and no biochemical disturbances. She first presented at birth with twitching movements and convulsions of an unknown aetiology. Ambulatory EEG showed epileptic rhythmic activities, the presence of asynchrony and runs of sharp waves over the right parietal and central areas. Given the non-specific neurological features and negative structural and biochemical findings, we applied clinical whole-exome sequencing (WES) to determine the underlying aetiology. WES revealed a novel heterozygous missense pathogenic variant, GNAO1:NM_020988.2:c.118G>A; NP_066268.1:p.Gly40Arg. A genetic analysis of the family confirmed the variant identified is a de novo mutation.nnnCONCLUSIONSnClinical WES can streamline genetic analysis and sort out pathogenic genes in an unbiased approach. GNAO1 is a disease-causing gene for the autosomal dominant form of early infantile epileptic encephalopathy. The novel pathogenic variant identified in this case should contribute to our understanding of the expanding spectrum of infantile-onset epilepsy.

Lipid mediators of inflammation as novel plasma biomarkers to identify patients with bacteremia

OBJECTIVESnRapid diagnostic tests for bacteremia are important for early treatment to improve clinical outcome. We sought to identify plasma biomarkers that can identify patients with bacteremia using an untargeted global metabolomic analysis.nnnMETHODSnPlasma metabolomic profiles were analyzed for 145 adult patients with (cases) and without (controls) bacteremia using ultra-high-performance liquid chromatography/quadrupole-time-of-flight mass spectrometry (UHPLC-Q-TOF-MS). All metabolites were compared between cases and controls using a 2-tier filtering approach, and each metabolite underwent receiver operating characteristic (ROC) curve analysis. Individual metabolites that distinguish between cases and controls were characterized. Subgroup analysis was performed to identify metabolites with prognostic significance.nnnRESULTSnAfter 2-tier filtering, 128 molecular features were identified to be potential biomarkers that could distinguish cases from controls. Five metabolites had an area under the ROC curve (AUC) of >0.8 in ROC curve analysis, including a sphingolipid, an acylcarnitine, a fatty acid ester, and 2 glycerophosphocholines. These metabolites could distinguish cases from controls in the unsupervised hierarchical clustering analysis. Subgroup analysis of bacteremic patients showed that the level of trans-2,3,4-trimethoxycinnamate was lower in fatal than non-fatal cases.nnnCONCLUSIONSnPlasma lipid mediators of inflammation can distinguish bacteremia cases from non-bacteremia controls. These biomarkers may be used as targets for rapid test in clinical practice.

Novel nonsense CDC73 mutations in Chinese patients with parathyroid tumors

Hyperparathyroidism-jaw tumor syndrome (HPT-JT) is an autosomal dominant disease characterized by the occurrences of parathyroid tumors and ossifying fibroma of maxilla/mandible. It is caused by mutations in CDC73 gene and mutation carriers are at increased risk of parathyroid carcinoma. Hyperparathyroidism could be the sole manifestation. We reported two Chinese patients having parathyroid neoplasm with equivocal malignant potential and parathyroid carcinoma respectively with both germline and somatic CDC73 mutations detected. Both of them presented with severe hypercalcemia and primary hyperparathyroidism with no other HPT-JT associated tumors and negative family history. We identified one novel germline mutation CDC73 NM_024529.4: c.1475Gxa0>xa0A; NP_078805.3: p.Trp492X and one novel somatic mutation CDC73 NM_024529.4: c.142Gxa0>xa0T; NP_078805.3: p.Glu48X. The other germline mutation CDC73 NM_024529.4: c.226Cxa0>xa0T; NP_078805.3: p.Arg76X and somatic mutation CDC73 NM_024529.4: c.85delG; NP_078805.3: p.Glu29SerfsX8 were previously reported. This is the first report of CDC73 mutations in the Chinese population. Genetic analysis is reliable to confirm the underlying hereditary basis of hyperparathyroidism. By identification of mutations, the patient and the family members could benefit from regular surveillance for early detection of tumors.

Non-invasive screening of HLA-DPA1 and HLA-DPB1 alleles for persistent hepatitis B virus infection: Susceptibility for vertical transmission and toward a personalized approach for vaccination and treatment

BACKGROUNDnPolymorphisms in the major histocompatibility complex (MHC) and non-MHC genes were recently reported to be associated with persistent hepatitis B virus (HBV) infection and host response to hepatitis B vaccine in Asian populations. We aimed to confirm the associations in Chinese population and develop a non-invasive screening method for the risk loci.nnnMETHODSnWe genotyped 2 risk alleles on the MHC loci, HLA-DPA1 (rs3077) and HLA-DPB1 (rs9277535), and 1 risk allele near a non-MHC gene, FOXP1 (rs6789153) using high-resolution melting curve analysis. With minimal processing steps and time, salivary DNA was extracted with a modified protocol of a blood kit. We compared the genotyping fidelity between peripheral blood DNA and salivary DNA.nnnRESULTSnBoth rs3077 and rs9277535, but not rs6789153, are significantly associated with CHB in Chinese population (p-value<0.001). High genotype concordance between different sources of genomic DNA was obtained.nnnCONCLUSIONSnGenotyping salivary DNA using our modified methods provides a non-invasive fast screening for host susceptibility loci. The transmission mechanism of hepatitis B can now be modified by adding genetic susceptibility to the traditional vertical transmission model of hepatitis B.

Lipid metabolites as potential diagnostic and prognostic biomarkers for acute community acquired pneumonia

n Abstractn n Early diagnosis of acute community-acquired pneumonia (CAP) is important in patient triage and treatment decisions. To identify biomarkers that distinguish patients with CAP from non-CAP controls, we conducted an untargeted global metabolome analysis for plasma samples from 142 patients with CAP (CAP cases) and 97 without CAP (non-CAP controls). Thirteen lipid metabolites could discriminate between CAP cases and non-CAP controls with area-under-the-receiver-operating-characteristic curve of >0.8 (P ≤ 10−9). The levels of glycosphingolipids, sphingomyelins, lysophosphatidylcholines and L-palmitoylcarnitine were higher, while the levels of lysophosphatidylethanolamines were lower in the CAP cases than those in non-CAP controls. All 13 metabolites could distinguish CAP cases from the non-infection, extrapulmonary infection and non-CAP respiratory tract infection subgroups. The levels of trihexosylceramide (d18:1/16:0) were higher, while the levels of lysophosphatidylethanolamines were lower, in the fatal than those of non-fatal CAP cases. Our findings suggest that lipid metabolites are potential diagnostic and prognostic biomarkers for CAP.n n

A case of early-onset obesity, hypocortisolism, and skin pigmentation problem due to a novel homozygous mutation in the proopiomelanocortin (POMC) gene in an Indian boy.

Abstract Proopiomelanocortin (POMC) is the polypeptide precursor of several biologically active melanocortin peptides that have important roles in the regulation of food intake and energy homeostasis, adrenal steroidogenesis, melanocyte stimulation, and immune modulation. Mutation of the POMC gene has been associated with adrenal insufficiency, early-onset obesity, and red hair pigmentation. We describe an Indian boy with secondary hypocortisolism, hyperphagia, early-onset obesity, and skin pigmentation problem. Genetics analysis revealed a novel homozygous mutation in the POMC gene (p.Arg86Term). The boy also had central hypothyroidism in addition to the secondary hypocortisolism. Genetics analysis for the POMC gene should be considered in patients with secondary hypocortisolism, early-onset obesity, and pigmentary problems.