Current status of diagnostic assays for emerging zoonotic viruses: Nipah and Hendra.

INTRODUCTION: Nipah and Hendra viruses belong to the Paramyxoviridae family, which pose a significant threat to human health, with sporadic outbreaks causing severe morbidity and mortality. Early symptoms include fever, cough, sore throat, and headache, which offer little in terms of differential diagnosis. There are no specific therapeutics and vaccines for these viruses. AREAS COVERED: This review comprehensively covers a spectrum of diagnostic techniques for Nipah and Hendra virus infections, discussed in conjunction with appropriate type of samples during the progression of infection. Serological assays, reverse transcriptase Real-Time PCR assays, and isothermal amplification assays are discussed in detail, along with a listing of few commercially available detection kits. Patents protecting inventions in Nipah and Hendra virus detection are also covered. EXPERT OPINION: Despite several outbreaks of Nipah and Hendra infections in the past decade, in-depth research into their pathogenesis, Point-of-Care diagnostics, specific therapies, and human vaccines is lacking. A prompt and accurate diagnosis is pivotal for efficient outbreak management, patient treatment, and the adoption of preventative measures. The emergence of rapid point-of-care tests holds promise in enhancing diagnostic capabilities in real-world settings. The patent landscape emphasizes the importance of innovation and collaboration within the legal and business realms.

An amphiphilic dansyl based multianalyte sensor for the detection of Hg2+, PPi, and TNP: A three-in-one chemical sensor.

A fluorescent dansyl-based amphiphilic probe, 5-(dimethylamino)-N-hexadecylnaphthalene-1-sulfonamide (DLC), was synthesized and characterized to detect multiple analytes at different sensing environments. In acetonitrile, DLC detects nitro explosives such as trinitrophenol (TNP) and 2,4-dinitrophenol (2,4-DNP) by an emission "on-off" response method, and the detection limits (LOD) were estimated to be as low as 4.3 µM and 17.4 µM, respectively. Amphiphilic long chains of the probe were embedded into lipid bilayers to form nanoscale vesicles DLC.Ves. Nanovesicular probe DLC.Ves was found to be highly selective for Hg2+ among other metal ions and for pyrophosphate (PPi) ions among various anions. DLC.Ves could detect Hg2+ with a turn "on-off" emission and PPi with ratiometric change in emission at 525 nm. It is proposed that DLC.Ves could detect Hg2+ via the Hg2+-induced aggregation quenching mechanism and PPi through the Hydrogen bonding. The LODs are estimated as 6.41 µM and 70.9 µM for Hg2+ and PPi, respectively. 1H NMR, SEM, and fluorescence lifetime measurements confirmed the binding mechanism. Thus, it is believed that the simple fluorescent probe DLC could be a prominent sensor to detect multiple analytes depending on the sensing medium.

Metarhabditis amsactae: A potential biopesticide isolated from Punjab (India) with potent insecticidal activity and immunomodulatory effects against Galleria mellonella (Lepidoptera: Pyralidae).

A survey was undertaken to isolate entomopathogenic nematodes from Amritsar district of Punjab, India. Out of 20 soil samples collected, two were found positive for the presence of nematodes. 18S and ITS rDNA gene sequencing revealed their identity as Metarhabditis amsactae. To assess its biocontrol potential, Galleria mellonella larvae were treated with concentrations of 20, 40, 80 and 160 IJs/L (infective juveniles/larva) and mortality was recorded from 24 h up to 96 h of nematode exposure. Distilled water without nematodes was used as an untreated control. M. amsactae showed potent larvicidal activity against G. mellonella that was found to be concentration and time dependent. Nematode infection caused 93.33 % larval mortality at 80 IJs/L after 72 h of treatment. 100 % mortality was observed after 96 h. No mortality was observed in control. To evaluate the immunomodulatory effects of M. amsactae, G. mellonella larvae were infected with 100 IJs/L and activities of antioxidant and detoxifying enzymes viz., superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APOX), phenol oxidase (PO), glutathione-S-transferase (GST) and acetylcholine esterase (AChE) were appraised after 12, 24, 36 and 48 h of nematode exposure. Malondialdehyde content was also determined. The results obtained demonstrated a significant elevation in all the enzyme activities at all time intervals in treated larvae when compared with untreated control. MDA levels were also enhanced in response to nematode infection. Thus, the present study revealed high insecticidal potential and immunomodulatory effects of M. amsactae on G. mellonella that should be further explored on other insect pests as well.

Liposome Based Near-Infrared Sensors for the Selective Detection of Hydrogen Sulfide.

Cyanine dye-based new amphiphilic compound NIR-Amp has been synthesised. NIR-Amp was embedded with phospholipids DOPC and DPPC to form liposomes based nanoscale chemical sensors NIR-Lip1 and NIR-Lip2. Here, two different phospholipids were used to demonstrate the influence of lipid structure, composition and fluidity on sensing of nanosensors. Both the probes show NIR absorption maximum at 790 nm and emission maximum at 815 nm. H2 S-triggered thiolation resulted a remarkable change in color from green to pale yellow. A decrease in UV-Vis absorption and emission in the NIR region was observed only with H2 S. NIR-Lip1 and NIR-Lip2 are highly selective for H2 S with a LOD of 0.57 µM and 1.24 µM, respectively. It was observed that in a solid-like gel state, NIR-Lip1 is slightly more sensitive towards H2 S than fluid-like NIR-Lip2. The H2 S sensing mechanism was confirmed by ESI-mass and infrared (IR) spectroscopic analysis. Based on the high sensitivity and selectivity, NIR-Lip1 was employed to detect H2 S in vegetable samples. Further, the probes are found to be non-toxic and established for H2 S fluorescence imaging in live cells.

Diverse presentations of ectopia lentis and lens coloboma in Marfan's syndrome.

Marfan's syndrome (MFS) is an autosomal dominant connective tissue disorder with defect in the fibrillin-1 gene. The most common ocular manifestation is subluxated lens in the superotemporal direction, accounting for 50%-85% of total cases. The association of lens coloboma with MFS has been described in literature, but the coexistence of lens coloboma with ectopia lentis is a rare feature. Here, we describe three cases of MFS including a case of bilateral lens coloboma with ectopia lentis: case 1 - a 39-year-old male with inferotemporal lens subluxation in the right eye and superotemporal lens subluxation in the left eye with open-angle glaucoma and high myopia, case 2 - a 15-year-old child with bilateral superonasal lens subluxation with lens coloboma, and case 3 - a 56-year-old female with bilateral lens coloboma. Case 1 and case 2 had clear lenses with good refractive correction; hence, they were optically rehabilitated with contact lenses, whereas case 3 was advised for cataract surgery. It is important to distinguish the lens coloboma from a more common entity, ectopia lentis as former usually remains stable while the latter might need a surgical intervention.

Klebsiella keratitis presenting as a ring infiltrate in an adolescent girl.

The purpose of the study was to report a unique case of Klebsiella keratitis presenting as a ring infiltrate in an adolescent girl. A 16-year-old girl presented with decreased vision in the right eye preceding an episode of fever with a rash associated with burning micturition. The patient was examined after taking appropriate consent. The slit-lamp examination revealed a ring-shaped corneal infiltrate with an epithelial defect in her right eye. Corneal scrapings were sent for microbiological evaluation which revealed Gram-negative rods and culture identified it as extended-spectrum beta-lactamase-producing Klebsiella pneumoniae colonies. The patient showed a good response to topical fortified amikacin and tobramycin. For her systemic complaints, the pediatrician did a thorough investigative workup out of which blood culture showed growth of K. pneumoniae. Hence, intravenous antibiotics were given based on the antibiogram report and the patient recovered. After 2 weeks, a paracentral infiltrate in her left eye was noted followed by anterior uveitis. The patient responded well to the topical course of steroids along with aminoglycosides. Four months later, she had a recurrence of anterior uveitis in the right eye preceded by fever. Blood investigations were negative. Hence, a diagnosis of recurrent uveitis secondary to endogenous infection was made and the patient was successfully treated with a short course of topical steroids. The patient is on follow-up for the past 6 months and maintaining the best-corrected visual acuity of 20/20 OU with normal intraocular pressure and quiet anterior chamber (AC). This is the first clinical report describing a ring infiltrate in endogenous Klebsiella keratitis and emphasizes thorough workup for prompt treatment.

Development of a PNA-LNA-LAMP Assay to Detect an SNP Associated with QoI Resistance in Erysiphe necator.

The repetitive use of quinone outside inhibitor fungicides (QoIs, strobilurins; Fungicide Resistance Action Committee [FRAC] 11) to manage grape powdery mildew has led to development of resistance in Erysiphe necator. While several point mutations in the mitochondrial cytochrome b gene are associated with resistance to QoI fungicides, the substitution of glycine to alanine at codon 143 (G143A) has been the only mutation observed in QoI-resistant field populations. Allele-specific detection methods such as digital droplet PCR and TaqMan probe-based assays can be used to detect the G143A mutation. In this study, a peptide nucleic acid-locked nucleic acid mediated loop-mediated isothermal amplification (PNA-LNA-LAMP) assay consisting of an A-143 reaction and a G-143 reaction, was designed for rapidly detecting QoI resistance in E. necator. The A-143 reaction amplifies the mutant A-143 allele faster than the wild-type G-143 allele, while the G-143 reaction amplifies the G-143 allele faster than the A-143 allele. Identification of resistant or sensitive E. necator samples was determined by which reaction had the shorter time to amplification. Sixteen single-spore QoI-resistant and -sensitive E. necator isolates were tested using both assays. Assay specificity in distinguishing the single nucleotide polymorphism (SNP) approached 100% when tested using purified DNA of QoI-sensitive and -resistant E. necator isolates. This diagnostic tool was sensitive to one-conidium equivalent of extracted DNA with an R2 value of 0.82 and 0.87 for the G-143 and A-143 reactions, respectively. This diagnostic approach was also evaluated against a TaqMan probe-based assay using 92 E. necator samples collected from vineyards. The PNA-LNA-LAMP assay detected QoI resistance in ≤30 min and showed 100% agreement with the TaqMan probe-based assay (≤1.5 h) for the QoI-sensitive and -resistant isolates. There was 73.3% agreement with the TaqMan probe-based assay when samples had mixed populations with both G-143 and A-143 alleles present. Validation of the PNA-LNA-LAMP assay was conducted in three different laboratories with different equipment. The results showed 94.4% accuracy in one laboratory and 100% accuracy in two other laboratories. The PNA-LNA-LAMP diagnostic tool was faster and required less expensive equipment relative to the previously developed TaqMan probe-based assay, making it accessible to a broader range of diagnostic laboratories for detection of QoI resistance in E. necator. This research demonstrates the utility of the PNA-LANA-LAMP for discriminating SNPs from field samples and its utility for point-of-care monitoring of plant pathogen genotypes.

A ruthenium nitrosyl complex-based highly selective colorimetric sensor for biological H2S and H2S-NO cross-talk regulated release of NO.

A ruthenium nitrosyl complex (1·NO) and 1·NO incorporated phospholipid-based liposomes (Lip-1·NO) were reported for highly selective colorimetric detection of H2S. The probe 1·NO "cross-talks" with H2S and releases nitric oxide (NO) in the process. The detection limit for H2S was found to be 0.31 µM and 0.45 µM in the cases of 1·NO and Lip-1·NO, respectively. The DAF-FM DA assay has been performed to confirm the H2S-induced NO release from 1·NO and Lip-1·NO. The sensing of H2S was also verified by ESI-MS and FT-IR spectroscopy. It was also observed that external stimuli, H2S and light worked in an almost similar way to release NO as observed by UV-Vis spectroscopy. A molecular logic gate operation "OR" was applied to the probe 1·NO in combination with inputs 'light' and 'H2S' to give the output 'NO release'. Hence, the probe 1·NO performs the dual work of sensing H2S with a colorimetric response, releasing NO upon cross-talk between NO and H2S.

Regulation of Nitric Oxide (NO) Release by Membrane Fluidity in Ruthenium Nitrosyl Complex-Embedded Phospholipid Vesicles.

Incorporating water-insoluble nitric oxide (NO)-releasing molecules into biocompatible vesicles may allow for the tunable control of NO release on a specific target site. In vesicles, membrane fluidity plays an important role and influences the final therapeutic efficiency of drugs loaded into the vesicles. Hence, we aimed to investigate the effect of lipid fluidity on the NO release behavior of the photo-controllable ruthenium nitrosyl (Ru-NO) complex. In this regard, a new photoactive ruthenium nitrosyl complex (L.Ru-NO) with amphiphilic terpyridine ligand was synthesized and characterized in detail. L.Ru-NO was incorporated with commercial phospholipids to form nanoscale vesicles L.Ru-NO@Lip. The photoactive {Ru-NO}6 type complex released NO in the organic solvent CH3CN and aqueous liposome solution by irradiating under low-intensity blue light (λ = 410 nm, 3 W). To demonstrate the effect of lipid structure and fluidity on NO release, four different liposome systems L.Ru-NO@Lip1-4 were prepared by using phospholipids such as DOPC, DSPC, DPPC, and DMPC having different chain lengths and saturation. The NO-releasing abilities of these liposomes in aqueous medium were studied by UV-vis spectrum, colorimetric Greiss, and fluorescent DAF assay. The results show that the rate of NO release could be easily tuned by varying the lipid fluidity. The effect of temperature and pH on NO release was also studied. Further, the complex L.Ru-NO and liposomes L.Ru-NO@Lip1 were assayed as an antibacterial agent against the strains of bacteria Escherichia coli and Staphylococcus aureus.

Multi analyte sensing of amphiphilic tridentate bis(benzimidazolyl)pyridine incorporated in liposomes and potential application in enzyme assay.

A liposome based nanosensor Lipo-1 for efficient detection of copper, cyanide (CN-) and ATP in a pure aqueous medium has been described. Lipo-1 shows a fluorescence ON-OFF response with copper. However, Lipo-1.Cu (Lipo-1 and copper ensemble) was used for the OFF-ON detection of ATP with nM and CN- with µM detection levels, lower than the WHO permissible level for safe drinking. Lipo-1 has better and enhanced binding properties over the counter organic amphiphilic compound Bzimpy-LC, which is not soluble in water. The significant changes in the emission spectra in the presence of Cu2+, CN- and ATP ions, as variable inputs, are used to construct INHIBIT and OR logic operations in a nano-scale environment. The fluorescent detection of CN- ions with Lipo-1.Cu was used to develop an enzyme assay for ß-glucosidase using amygdalin as the substrate. ß-Glucosidase enzymatic activity was monitored by the emission OFF-ON signal of the probe Lipo-1.Cu by CN- detection. This approach could be an efficient method for developing a fluorescence-based ß-glucosidase enzyme assay. A switch ON luminescence response, low detection limit, fast response, 100% aqueous solution, biocompatibility, multi-analyte detection, and improved sensitivity and selectivity of Bzimpy-LC in lipid bilayer membranes are the main features of the nanoprobe Lipo-1. These properties give it a clear advantage for analytical applications.

Flavan-based phytoconstituents inhibit Mpro, a SARS-COV-2 molecular target, in silico.

A well-validated in-silico approach can provide promising drug candidates for the treatment of the ongoing CoVID19 pandemic. In this study, we have screened 32 phytochemical constituents (PCCs) with Mpro binding site (PDB:6W63) based on which we identified three possible candidates that are likely to be effective against CoVID19-viz., licoleafol (binding energy: -8.1 kcal/mol), epicatechin gallate (-8.5 kcal/mol) and silibinin (-8.4 kcal/mol) that result in higher binding affinity than the known inhibitor, X77 (-7.7 kcal/mol). Molecular dynamics (MD) simulations of PCCs-Mpro complex confirmed molecular docking results with high structural and dynamical stability. The selected compounds were found to exhibit low mean squared displacements (licoleafol: 2.25 ± 0.43 Å, epicatechin gallate: 1.93 ± 0.35 Å, and silibinin: 1.39 ± 0.19 Å) and overall low fluctuations of the binding complexes (root mean squared fluctuations below 2 Å). Visualization of the MD trajectories and structural analyses revealed that they remain confined to the initial binding region, with mean fluctuations lower than 3 Å. To access the collective motion of the atoms, we performed principal component analysis demonstrating that the first 10 principal components are the major contributors (approximate contribution of 80%) and are responsible for the overall PCCs motion. Considering that the three selected PCCs share the same flavan backbone and exhibit antiviral activity against hepatitis C, we opine that licoleafol, epi-catechin gallate, and silibinin can be promising anti-CoVID19 drug candidates. Communicated by Ramaswamy H. Sarma.

Sensing and Bioimaging of the Gaseous Signaling Molecule Hydrogen Sulfide by Near-Infrared Fluorescent Probes.

A fluorescent probe for the monitoring of H2S levels in living cells and organisms is highly desirable. In this regard, near-infrared (NIR) fluorescent probes have emerged as a promising tool. NIR-I and NIR-II probes have many significant advantages; for instance, NIR light penetrates deeper into tissue than light at visible wavelengths, and it causes less photodamage during biosample analysis and less autofluorescence, enabling higher signal-to-background ratios. Therefore, it is expected that fluorescent probes having emission in the NIR region are more suitable for in vivo imaging. Consequently, a considerable increase in reports of new H2S-responsive NIR fluorescent probes appeared in the literature. This review highlights the advances made in developing new NIR fluorescent probes aimed at the sensitive and selective detection of H2S in biological samples. Their applications in real-time monitoring of H2S in cells and in vivo for bioimaging of living cells/animals are emphasized. The selection of suitable dyes for designing NIR fluorescent probes, along with the principles and mechanisms involved for the sensing of H2S in the NIR region, are described. The discussions are focused on small-molecule and nanomaterials-based NIR probes.

Nanoscale lipid vesicles functionalized with a nitro-aniline derivative for photoinduced nitric oxide (NO) delivery.

Nanoscale vesicles functionalized with a nitric oxide (NO) releasing molecule 4-nitro-3-(trifluoromethyl)aniline have been reported. The new NO-nano-vesicular donor material shows an effective photo-release of NO upon irradiation with blue light at 410 nm. The kinetics of NO release has been monitored by using simple spectroscopic techniques such as UV-Vis and fluorescence methods. Colorimetric Griess assay and fluorescence DAF assay have been used for the detection and quantification of NO released from vesicles. This new vesicular nanoscale NO donor has the advantages of facile preparation in water, capable of releasing NO in a pure aqueous medium, photo-controlled NO release, bio-compatibility and capacity to modulate the NO donor loading to achieve an essential amount of NO.

Fluorescent nanoprobes for the sensing of gasotransmitters hydrogen sulfide (H2S), nitric oxide (NO) and carbon monoxide (CO).

Fluorescent nanomaterials as sensing probes have experienced immense growth in recent years due to the intrinsic optical and physicochemical properties, high sensitivity, specificity, targeting ability, and suitability for medicinal applications. The fluorescent detection of gaseous signaling molecules, such as Hydrogen sulfide (H2S), nitric oxide (NO) and carbon monoxide (CO) are very important due to their potential therapeutic application. This review intends to provide the recent progress in the detection of H2S, CO and NO via fluorescent based nano probes. These probes work based on different mechanisms such as fluorescence enhancement and quenching, also defined as "turn-on" and "turn-off" responses respectively. It could be achieved through PET, FRET or ratiometric methods. In this article, we have discussed about a variety of fluorescent nanoprobes of QDS, CDs, AuNPs and UCNPS, working on the fluorescent sensing mechanisms and applicable for the detection of H2S, CO and NO in biological and environmental samples. Methods used for the detection, structural features of nanomaterials, type of fluorescence response observed, fluorescence sensing mechanism and their sensitivity are highlighted.

Development and characterization of embelin-loaded nanolipid carriers for brain targeting.

The objective of present work was to enquire the potential use of embelin-loaded nanolipid carriers for brain targeting. The average particle size and polydispersity index (PDI) of optimized formulation (F19) were found to be 152 ± 19.7 nm and 0.143 ± 0.023, respectively. Nanolipid carrier (NLC) was also significantly attenuated pentylenetetrazole (PTZ)-induced biochemical parameters in comparison to plain embelin that results in an increase in the level of malondialdehyde (MDA), nitrite, and reduction in the level of glutathione. From the results, it was concluded that embelin-NLCs developed as a beneficial carrier to achieve sustained release and brain targeting through nasal route.

Peptide Receptor Radionuclide Therapy Outcomes in a North American Cohort With Metastatic Well-Differentiated Neuroendocrine Tumors.

OBJECTIVES: The objective of this study was to describe the outcomes of patients in the University of Iowa Neuroendocrine Tumor (NET) Database treated with peptide receptor radionuclide therapy (PRRT). METHODS: One hundred thirty-five patients from the University of Iowa NET Database who received PRRT were analyzed, their characteristics were described, and survival was calculated. RESULTS: The median age at diagnosis was 51 years, and 64% were men. The primary tumor was located in the small bowel (SBNET) in 37.8%, in the pancreas (PNET) in 26.0%, in the lung in 13.3%, in unknown primary in 9.6%, and in other sites in 13.3%. A radiographic response of any magnitude was observed in 65.8%, 11.1% had a mixed response, and 15.4% showed progression. The overall survival (OS) from the first PRRT was 40 months, and the median time to progression was 23.9 months. Higher pretreatment chromogranin A and pancreastatin levels predicted inferior OS. CONCLUSIONS: Peptide receptor radionuclide therapy resulted in a relatively long OS and time to progression in heavily pretreated North American patients with advanced NETs. Elevated pretreatment chromogranin A and pancreastatin predicted shorter OS after therapy. Peptide receptor radionuclide therapy is a valuable treatment option in patients with advanced NETs, especially SBNETS.

Comparison of transarterial liver-directed therapies for low-grade metastatic neuroendocrine tumors in a single institution.

OBJECTIVE: We compared the clinical outcomes of patients with metastatic neuroendocrine tumors treated with hepatic artery embolization (HAE), chemoembolization (HACE), and selective internal radiation therapy (SIRT) at our institution over the last 10 years. METHODS: The medical records of 42 patients with metastatic neuroendocrine tumors with hepatic metastases treated with HAE, HACE, or SIRT at the University of Iowa from 2001 to 2011 were analyzed. RESULTS: A total of 13 patients had HAE, 17 patients had HACE, and 12 patients had SIRT as their initial procedure. Time to progression (TTP) was similar between SIRT (15.1 months) and HACE/HAE groups (19.6 months; P = 0.968). There was a trend toward increased TTP in patients receiving HACE (33.4 months) compared with HAE (12.1 months) or SIRT (15.1 months), although not statistically significant (P = 0.512). The overall survival for all patients from the first intervention was 41.9 months. There was no difference between HACE/HAE and SIRT in posttherapy change of chromogranin A (P = 0.233) and pancreastatin (P = 0.158) levels. Time to progression did not correlate with the change in the posttherapy chromogranin A (P = 0.299) or pancreastatin (P = 0.208) levels. CONCLUSIONS: There was no significant difference in TTP in patients treated with SIRT compared with patients treated with HAE or HACE. Baseline and posttherapy marker changes were not predictive of TTP.