An algorithmic approach to identifying the aetiology of acute encephalitis syndrome in India: results of a 4-year enhanced surveillance study.

BACKGROUND: Annual outbreaks of acute encephalitis syndrome pose a major health burden in India. Although Japanese encephalitis virus (JEV) accounts for around 15% of reported cases, the aetiology of most cases remains unknown. We aimed to establish an enhanced surveillance network and to use a standardised diagnostic algorithm to conduct a systematic evaluation of acute encephalitis syndrome in India. METHODS: In this large-scale, systematic surveillance study in India, patients presenting with acute encephalitis syndrome (ie, acute onset of fever with altered mental status, seizure, or both) to any of the 18 participating hospitals across Uttar Pradesh, West Bengal, and Assam were evaluated for JEV (serum and cerebrospinal fluid [CSF] IgM ELISA) per standard of care. In enhanced surveillance, JEV IgM-negative specimens were additionally evaluated for scrub typhus, dengue virus, and West Nile virus by serum IgM ELISA, and for Streptococcus pneumoniae, Haemophilus influenzae, Neisseria meningitidis, dengue virus, herpes simplex virus, and enterovirus by CSF PCR across five referral laboratories. In 2017, chikungunya and Leptospira serum IgM by ELISA and Zika virus serum and CSF by PCR were also tested. FINDINGS: Of 10 107 patients with acute encephalitis syndrome enrolled in enhanced surveillance between Jan 1, 2014, and Dec 31, 2017, 5734 (57·8%) of 9917 participants with available data were male and 6179 (62·7%) of 9856 were children aged 15 years and younger. Among patients who provided a sample of either CSF or serum in enhanced surveillance, an aetiology was identified in 1921 (33·2%) of 5786 patients enrolled between 2014 and 2016 and in 1484 (34·3%) of 4321 patients enrolled in 2017. The most commonly identified aetiologies were JEV (1023 [17·7%] of 5786 patients), scrub typhus (645 [18·5%] of 3489), and dengue virus (161 [5·2%] of 3124). Among participants who provided both CSF and serum specimens, an aetiology was identified in 1446 (38·3%) of 3774 patients enrolled between 2014 and 2016 and in 936 (40·3%) of 2324 enrolled in 2017, representing a 3·1-times increase in the number of patients with acute encephalitis syndrome with an identified aetiology compared with standard care alone (299 [12·9%]; p<0·0001). INTERPRETATION: Implementation of a systematic diagnostic algorithm in an enhanced surveillance platform resulted in a 3·1-times increase in identification of the aetiology of acute encephalitis syndrome, besides JEV alone, and highlighted the importance of scrub typhus and dengue virus as important infectious aetiologies in India. These findings have prompted revision of the national testing guidelines for this syndrome across India. FUNDING: US Centers for Disease Control and Prevention.

Dengue virus is an under-recognised causative agent of acute encephalitis syndrome (AES): Results from a four year AES surveillance study of Japanese encephalitis in selected states of India.

BACKGROUND: Acute encephalitis syndrome (AES) surveillance in India has indicated that Japanese encephalitis virus (JEV) accounts for 5-35% of AES cases annually; the etiology remains unknown in the remaining cases. We implemented comprehensive AES surveillance to identify other etiological agents of AES, with emphasis on dengue virus. METHODS: Serum and cerebrospinal fluid (CSF) specimens were collected from patients enrolled prospectively in AES surveillance from 2014-2017 at selected sites of three high burden states of India. All samples were initially tested for JEV IgM. Specimens negative for JEV by serology were tested for IgM to scrub typhus, dengue virus (DEN), and West Nile virus; all JEV IgM-negative CSF samples were tested by PCR for S. pneumoniae, N. meningitidis, H. influenzae, herpes simplex virus type 1, enteroviruses and DEN. RESULTS: Of 10,107 AES patients, an etiology could be established in 49.2% of patients including JEV (16%), scrub typhus (16%) and DEN (5.2%) as the top three agents. Amongst the DEN positive cases (359/6892), seven (2%) were positive only for dengue virus RNA: one in serum and six in CSF. CONCLUSION: Amongst the pathogens identified, dengue accounted for 5% of all AES cases and was one of the three common etiological agents. These results underscore the importance of including dengue virus in routine testing of AES cases.

Three-dimensional kinematic stress magnetic resonance image analysis shows promise for detecting altered anatomical relationships of tissues in the cervical spine associated with painful radiculopathy.

For some patients with radiculopathy a source of nerve root compression cannot be identified despite positive electromyography (EMG) evidence. This discrepancy hampers the effective clinical management for these individuals. Although it has been well-established that tissues in the cervical spine move in a three-dimensional (3D) manner, the 3D motions of the neural elements and their relationship to the bones surrounding them are largely unknown even for asymptomatic normal subjects. We hypothesize that abnormal mechanical loading of cervical nerve roots during pain-provoking head positioning may be responsible for radicular pain in those cases in which there is no evidence of nerve root compression on conventional cervical magnetic resonance imaging (MRI) with the neck in the neutral position. This biomechanical imaging proof-of-concept study focused on quantitatively defining the architectural relationships between the neural and bony structures in the cervical spine using measurements derived from 3D MR images acquired in neutral and pain-provoking neck positions for subjects: (1) with radicular symptoms and evidence of root compression by conventional MRI and positive EMG, (2) with radicular symptoms and no evidence of root compression by MRI but positive EMG, and (3) asymptomatic age-matched controls. Function and pain scores were measured, along with neck range of motion, for all subjects. MR imaging was performed in both a neutral position and a pain-provoking position. Anatomical architectural data derived from analysis of the 3D MR images were compared between symptomatic and asymptomatic groups, and the symptomatic groups with and without imaging evidence of root compression. Several differences in the architectural relationships between the bone and neural tissues were identified between the asymptomatic and symptomatic groups. In addition, changes in architectural relationships were also detected between the symptomatic groups with and without imaging evidence of nerve root compression. As demonstrated in the data and a case study the 3D stress MR imaging approach provides utility to identify biomechanical relationships between hard and soft tissues that are otherwise undetected by standard clinical imaging methods. This technique offers a promising approach to detect the source of radiculopathy to inform clinical management for this pathology.

Mycotic corneal ulcers in upper Assam.

PURPOSE: To study the association of various risk factors and epidemiological variables of mycotic keratitis treated at a tertiary referral hospital of upper Assam. MATERIALS AND METHODS: In this hospital-based prospective study a total of 310 consecutive corneal ulcer cases attending the ophthalmology outpatient department of Assam Medical College were enrolled between April 2007 and March 2009. After clinical and slit-lamp biomicroscopic examination in all suspected cases, smears and culture examination for fungus was done to establish the etiology. Demographic information and associated probable risk factors of individual cases were noted in a predesigned questionnaire. RESULTS: In 188 (60.6%) cases fungal etiology could be established. Out of them 67.6% were males. The most commonly affected age group was 41-50 years (25.5%). The maximum (23.4%) cases were reported during the paddy harvesting season in Assam (January and February). Fungal element could be demonstrated in 65.2% cases in direct potassium hydroxide (KOH) mount. The commonest predisposing factor was corneal injury (74.5%). While diabetes was a significant systemic predisposing factor in mixed bacterial and fungal infections in 11.1% cases, blocked naso-lacrimal duct was the local predisposing factor in 11.1% of cases. Fusarium solani (25%) was the commonest isolate followed by Aspergillus species (19%), Curvularia species (18.5%) and Penicillium species (15.2%). Yeasts were isolated in 2.7% (n=5) cases. CONCLUSIONS: Ocular trauma was the commonest cause of fungal corneal ulcer in Assam and Fusarium solani was the commonest species responsible for it. Most of the mycotic ulcer cases come from rural areas including the tea gardens.

Sodium MR imaging detection of mild Alzheimer disease: preliminary study.

BACKGROUND AND PURPOSE: There is significant interest in the development of novel noninvasive techniques for the diagnosis of Alzheimer disease (AD) and tracking its progression. Because MR imaging has detected alterations in sodium levels that correlate with cell death in stroke, we hypothesized that there would be alterations of sodium levels in the brains of patients with AD, related to AD cell death. MATERIALS AND METHODS: A total of 10 volunteers (5 with mild AD and 5 healthy control subjects) were scanned with a 20-minute sodium (23Na) MR imaging protocol on a 3T clinical scanner. RESULTS: After normalizing the signal intensity from the medial temporal lobes corresponding to the hippocampus with the ventricular signal intensity, we were able to detect a 7.5% signal intensity increase in the brains of patients with AD (AD group, 68.25% +/- 3.4% vs control group, 60.75% +/- 2.9%; P < .01). This signal intensity enhancement inversely correlated with hippocampal volume (AD group, 3.22 +/- 0.50 cm3 vs control group, 3.91 +/- 0.45 cm3; r2 = 0.50). CONCLUSIONS: This finding suggests that sodium imaging may be a clinically useful tool to detect the neuropathologic changes associated with AD.

Biomarkers for early detection of Alzheimer pathology.

The increasing prevalence of Alzheimer's disease and the devastating consequences of late-life dementia motivates the drive to develop diagnostic biomarkers to reliably identify the pathology associated with this disorder. Strategies to accomplish this include the detection of altered levels of tau and amyloid in cerebrospinal fluid, the use of structural MRI to identify disease-specific patterns of regional atrophy and MRI T(1)rho to detect disease-related macromolecular protein aggregation, and the direct imaging of amyloid deposits using positron emission tomography and single photon emission computerized tomography. Success will facilitate the ability to reliably diagnose Alzheimer's disease while the symptoms of brain failure are mild and may provide objective measures of disease-modifying treatment efficacy.

Mechanical ventilator for delivery of ¹7O2 in brief pulses.

The ¹7O nucleus has been used recently by several groups for magnetic resonance (MR) imaging of cerebral metabolism. Inhalational delivery of ¹7O(2) in very brief pulses could, in theory, have significant advantages for determination of the cerebral metabolic rate for oxygen (CMRO2) with MR imaging. Mechanical ventilators, however, are not typically capable of creating step changes in gas concentration at the airway. We designed a ventilator for large animal and human studies that provides mechanical ventilation to a subject inside an MR scanner through 25 feet of small-bore connecting tubing, and tested its capabilities using helium as a surrogate for ¹7O2. After switching the source gas from oxygen to helium, the 0-90% response time for helium concentration changes at the airway was 2.4 seconds. The capability for creating rapid step changes in gas concentration at the airway in large animal and human studies should facilitate the experimental testing of the delivery ¹7O2 in brief pulses, and its potential use in imaging CMRO2.

The Siva-1 putative amphipathic helical region (SAH) is sufficient to bind to BCL-XL and sensitize cells to UV radiation induced apoptosis.

The human Siva gene is localized to chromosome 14q32-33 and gives rise to the full-length predominant form, Siva-1 and a minor alternate form, Siva-2 that appears to lack the proapoptotic properties of Siva-1. Our recent work has shown that the missing region in Siva-2 encodes a unique twenty amino acid putative amphipathic helical region (SAH, residues 36-55 in Siva-1). Despite the fact that Siva-1 does not belong to the BCL-2 family, it specifically interacts with the anti-apoptotic protein BCL-XL and sensitizes MCF7 breast cancer cells expressing BCL-XL to UV radiation induced apoptosis. Deletion mutagenesis has mapped the necessary region to the SAH in Siva-1. In this paper we demonstrate that the SAH region in Siva-1 is sufficient to specifically interact with the anti-apoptotic members of the BCL2 family such as BCL-XL and BCL-2 but not its apoptotic member BAX. Using transient transfections and direct microinjection of synthetic SAH peptides, we also demonstrate that the SAH region is sufficient to inhibit the BCL-XL mediated cell survival and render MDA-MB-231 and MCF7 breast cancer cells expressing BCL-XL highly susceptible to UV radiation induced apoptosis. The underlying mechanism of action of SAH mediated inhibition of BCL-XL (and/or BCL2) cell survival appears to be due to loss of mitochondrial integrity as reflected in enhanced cytochrome c release leading to the activation of caspase 9 and finally caspase 3.

Proteoglycan-induced changes in T1rho-relaxation of articular cartilage at 4T.

Proteoglycan (PG) depletion-induced changes in T1rho (spin-lattice relaxation in rotating frame) relaxation and dispersion in articular cartilage were studied at 4T. Using a spin-lock cluster pre-encoded fast spin echo sequence, T1rho maps of healthy bovine specimens and specimens that were subjected to PG depletion were computed at varying spin-lock frequencies. Sequential PG depletion was induced by trypsinization of cartilage for varying amounts of time. Results demonstrated that over 50% depletion of PG from bovine articular cartilage resulted in average T1rho increases from 110-170 ms. Regression analysis of the data showed a strong correlation (R2 = 0.987) between changes in PG and T1rho. T1rho values were highest at the superficial zone and decreased gradually in the middle zone and again showed an increasing trend in the region near the subchondral bone. The potentials of this method in detecting early degenerative changes of cartilage are discussed. Also, T(1rho)-dispersion changes as a function of PG depletion are described.

Water distribution patterns inside bovine articular cartilage as visualized by 1H magnetic resonance imaging.

OBJECTIVE: To develop a magnetic resonance imaging (MRI) technique to non-invasively map water volume fraction (WVF) in articular cartilage. Special emphasis was placed on spatial resolution and temporal considerations, aimed at creating a procedure feasible for eventual human studies. DESIGN: Absolute proton density MR images of intact, ex vivo bovine patellae were calculated from fully T(1) relaxed, short echo time images. This was accomplished through compensation for T(2) decay with calculated T(2) maps. Calibration of the signal intensity in the image was accomplished with the use of H2O:D2O phantoms, where the WVF was varied from 0.95 to 0.75. Application of the calibration curve to the entire image yielded images that represent WVF on a pixel by pixel basis. Calculations of water content by weight were performed by considering the density of the solid content. RESULTS: Using four echo time points, experiments comparing MR images from single-echo and multi-echo spin echo sequences yielded similar results. T(2) decreased with depth through the cartilage, with a maximum at the articular surface of approx 100 ms, and a approximately 50 ms minimum at the bone/cartilage interface. The WVF through the depth of the cartilage showed a similar trend, decreasing from 0.9 at the surface, to 0.7 at the bone/cartilage interface. Translation to a weight percent yielded approximately 86% weight at the surface, trending down to approximately 63% at the bone/cartilage interface, with an average of 74.5% for five patellae. These MRI derived values were compared to the measured weight of water in excised cartilage plugs from the same patellae and showed remarkably close agreement. CONCLUSION: We have demonstrated that MRI can non-invasively map WVF in cartilage in a pixel by pixel manner. This was accomplished in a time span that was clinically feasible, allowing the routine use of this method in a clinical setting. Moreover, this procedure employed standard MRI equipment and pulse sequences, avoiding the need for hardware modifications and using simple post processing methods. However, baseline studies need to be performed prior to incorporation into a standard radiological evaluation. Implications in the diagnosis of osteoarthritis (OA) are discussed.

Sensitivity of MRI to proteoglycan depletion in cartilage: comparison of sodium and proton MRI.

OBJECTIVE: The purpose of this work was to evaluate the results from sodium and proton magnetic resonance imaging (MRI) in detecting small changes in proteoglycan (PG) content in bovine articular cartilage specimens. DESIGN: Articular cartilage from 15 specimens of bovine patellae were subjected to partial PG depletion with different concentrations of trypsin for 30 min. Sodium and proton MR images of the intact specimen were obtained on a 4T GE clinical MRI system. Two custom-built 7 cm-diameter solenoid coils tuned to proton and sodium frequencies were employed. Fast gradient echo and spin echo imaging sequences were used to determine sodium density, proton density and proton relaxation times (T(1)and T(2)) of the specimens. Spectrophotometric assay was performed after MRI to determine PG concentrations of the cartilage specimens. RESULTS: The sodium signal change correlated well with the observed PG loss (R(2)=0.85, P< 0.01) whereas the proton signal change was inconsistent (R(2)=0.10, P< 0.8). The change in proton T(1)and T(2)between the two regions did not correlate with PG loss (R(2)=0. 07 and R(2)=0.06, respectively). CONCLUSIONS: Results from these studies demonstrate that sodium MRI is both sensitive and specific in detecting small changes in PG concentration, whereas proton density and relaxation properties are not sensitive to small changes in PG content.

MR imaging of RF heating using a paramagnetic doped agarose phantom.

In this paper, we present the first description of a technique to visualize and quantitate radiofrequency (RF) heating of a tissue phantom during a magnetic resonance imaging (MRI) procedure. We evaluated the heating patterns of four 10 cm diameter transmit/receive surface coils with differing degrees of distributed capacitance. The tissue phantom was a 6% agarose gel doped with 40 mM Na(4)HTm[DOTP], and possesses a conductivity intermediate to human muscle and fat. Heating was discerned via phase difference mapping using the large temperature dependent chemical shift coefficient for 23Na in Na(4)HTm[DOTP]. This coefficient is -0.5 ppm/ degrees C. Heating was highest where the phantom was closest to the surface coils, dropping off towards the center of the coil. No significant difference was observed in the heating patterns between the different surface coils. For the experimental setups used in this study, electric field 'hot spots' at the areas corresponding to the placement of the capacitor gaps were not observed.

Temperature-dependent chemical shift and relaxation times of (23)Na in Na(4)HTm[DOTP].

We describe the characterization of a (23)Na temperature-dependent chemical shift and relaxation rates in the complex, Na(4)HTm[DOTP]. This is the first characterization of a (23)Na temperature-dependent chemical shift in a nonmetallic sample. The (23)Na temperature-dependent chemical shift coefficient is approximately -0. 5 PPM/ degrees C for both an aqueous solution and a 6% agarose gel of this compound. This is 50 times the magnitude of the temperature-dependent chemical shift coefficient of water protons. The relaxation times, T(1), T(2f), and T(2s) increased by 0.1, 0.01, and 0.05 ms/ degrees C, respectively. Applications of these unique properties for designing an MRI technique for monitoring heat deposition in tissue and tissue phantoms are discussed.

Sodium visibility and quantitation in intact bovine articular cartilage using high field (23)Na MRI and MRS.

Noninvasive methods of detecting cartilage degeneration can have an impact on identifying the early stages of osteoarthritis. Accurate measurement of sodium concentrations within the cartilage matrix provides a means for analyzing tissue integrity. Here a method is described for quantitating sodium concentration and visibility in cartilage, with general applications to all tissue types. The sodium concentration in bovine patellar cartilage plugs was determined by three different methods: NMR spectroscopy of whole cartilage plugs, NMR spectroscopy of liquefied cartilage in concentrated HCl, and inductively coupled plasma emission spectroscopy. Whole bovine patellae were imaged with relaxation normalized calibration phantoms to ascertain sodium concentrations inside the articular cartilage. Sodium concentrations in intact articular cartilage were found to range from approximately 200 mM on the edges to approximately 390 mM in the center, with an average of approximately 320 mM in five separate bovine patellae studied. In essence, we have created sodium distribution maps of the cartilage, showing for the first time, spatial variations of sodium concentration in intact cartilage. This average concentration measurement correlates very well with the values obtained from the spectroscopic methods. Furthermore, sodium was found to be 100% NMR visible in cartilage plugs. Applications of this method in diagnosing and monitoring treatment of osteoarthritis are discussed.

Time-domain quantification of multiple-quantum-filtered (23)Na signal using continuous wavelet transform analysis.

The application of continuous wavelet transform (CWT) analysis technique is presented to analyze multiple-quantum-filtered (MQF) (23)Na magnetic resonance spectroscopy (MRS) data. CWT acts on the free-induction-decay (FID) signal as a time-frequency variable filter. The signal-to-noise ratio (SNR) and frequency resolution of the output filter are locally increased. As a result, MQF equilibrium longitudinal magnetization and the apparent fast and slow transverse relaxation times are accurately estimated. A developed iterative algorithm based on frequency signal detection and components extraction, already proposed, was used to estimate the values of the signal parameters by analyzing simulated time-domain MQF signals and data from an agarose gel. The results obtained were compared to those obtained by measurement of signal height in frequency domain as a function of MQF preparation time and those obtained by a simple time-domain curve fitting. The comparison indicates that the CWT approach provides better results than the other tested methods that are generally used for MQF (23)Na MRS data analysis, especially when the SNR is low. The mean error on the estimated values of the amplitude signal and the apparent fast and slow transverse relaxation times for the simulated data were 2.19, 6. 63, and 16.17% for CWT, signal height in frequency domain, and time-domain curve fitting methods, respectively. Another major advantage of the proposed technique is that it allows quantification of MQF (23)Na signal from a single FID and, thus, reduces the experiment time dramatically.

In vivo triple quantum filtered twisted projection sodium MRI of human articular cartilage.

In this work, we present the first triple quantum filtered (TQF) sodium MR images of the human knee joint in vivo. A 3D TQF data set of 16 slices was obtained in 20 min using a TQF pulse sequence preencoded to a twisted projection imaging readout. Images clearly demarcate patellar cartilage and also demonstrate fluid signal suppressed by the triple quantum filter. Biexponential transverse relaxation times were calculated by fitting the TQF free induction decay to a theoretical signal expression. The average values from three healthy volunteers were T(2fall)(*) = 9.59 +/- 0.35 ms and T(2rise)(*) = 0.84 +/- 0.06 ms. Application of TQF imaging in biological tissues is discussed.

Specificity of starch synthase isoforms from potato.

In higher plants several isoforms of starch synthase contribute to the extension of glucan chains in the synthesis of starch. Different isoforms are responsible for the synthesis of essentially linear amylose chains and branched, amylopectin chains. The activity of granule-bound starch synthase I from potato has been compared with that of starch synthase II from potato following expression of both isoforms in Escherichia coli. Significant differences in their activities are apparent which may be important in determining their specificities in vivo. These differences include affinities for ADPglucose and glucan substrates, activation by amylopectin, response to citrate, thermosensitivity and the processivity of glucan chain extension. To define regions of the isoforms determining these characteristic traits, chimeric proteins have been produced by expression in E. coli. These experiments reveal that the C-terminal region of granule-bound starch synthase I confers most of the specific properties of this isoform, except its processive elongation of glucan chains. This region of granule-bound starch synthase I is distinct from the C-terminal region of other starch synthases. The specific properties it confers may be important in defining the specificity of granule-bound starch synthase I in producing amylose in vivo.