Publisher Correction: Whole-genome sequencing of a sporadic primary immunodeficiency cohort.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Whole-genome sequencing of a sporadic primary immunodeficiency cohort.

Primary immunodeficiency (PID) is characterized by recurrent and often life-threatening infections, autoimmunity and cancer, and it poses major diagnostic and therapeutic challenges. Although the most severe forms of PID are identified in early childhood, most patients present in adulthood, typically with no apparent family history and a variable clinical phenotype of widespread immune dysregulation: about 25% of patients have autoimmune disease, allergy is prevalent and up to 10% develop lymphoid malignancies1-3. Consequently, in sporadic (or non-familial) PID genetic diagnosis is difficult and the role of genetics is not well defined. Here we address these challenges by performing whole-genome sequencing in a large PID cohort of 1,318 participants. An analysis of the coding regions of the genome in 886 index cases of PID found that disease-causing mutations in known genes that are implicated in monogenic PID occurred in 10.3% of these patients, and a Bayesian approach (BeviMed4) identified multiple new candidate PID-associated genes, including IVNS1ABP. We also examined the noncoding genome, and found deletions in regulatory regions that contribute to disease causation. In addition, we used a genome-wide association study to identify loci that are associated with PID, and found evidence for the colocalization of-and interplay between-novel high-penetrance monogenic variants and common variants (at the PTPN2 and SOCS1 loci). This begins to explain the contribution of common variants to the variable penetrance and phenotypic complexity that are observed in PID. Thus, using a cohort-based whole-genome-sequencing approach in the diagnosis of PID can increase diagnostic yield and further our understanding of the key pathways that influence immune responsiveness in humans.

Fecal Immunochemical Testing in Patients With Low-Risk Symptoms of Colorectal Cancer: A Diagnostic Accuracy Study.

BACKGROUND: The fecal immunochemical test (FIT) is recommended for triaging primary care patients in England with low-risk symptoms of colorectal cancer (CRC). The evidence underpinning recommendations by the National Institute for Health and Care Excellence had limitations, with a paucity of primary care evidence. This study examines the diagnostic accuracy of FIT in a defined low-risk symptom primary care population. PATIENTS AND METHODS: Consecutive symptomatic adult patients referred for a FIT between October and December 2019 were included. Patients were derived from 225 primary care practices in England. Serious colorectal diseases (CRC, high-risk polyps, and inflammatory bowel disease [IBD]) were identified through patient follow-up over 18 months, using both primary and secondary healthcare records. Performance characteristics of FIT are reported according to differing thresholds, including the currently recommended threshold of ≥10 µg hemoglobin per gram of feces (µg/g). RESULTS: A total of 3,506 patients were included in the final analysis. Of these, 708 had a positive FIT result (≥10 µg/g). The prevalence of CRC was 1.3%. FIT positivity declined from 20.2% to 5.8% and 4.5% at cutoffs of 10, 80, and 120 µg/g, respectively. The sensitivity of FIT at ≥10 µg/g to detect CRC was 91.1% (95% CI, 77.9%-97.1%); its specificity was 80.7% (95% CI, 79.3%-82.0%); the positive predictive value (PPV) was 5.8% (95% CI, 4.2%-7.8%); and the negative predictive value (NPV) was 99.9% (95% CI, 99.6%-99.95%). The area under the receiver operating characteristic curve was 0.93 (0.91-0.96). PPV and specificity increased, whereas sensitivity and NPV decreased when serious colorectal diseases (CRC, high-risk polyps, and IBD) were combined. Age, sex, socioeconomic deprivation, and anemia did not significantly influence FIT sensitivity on subgroup analysis. CONCLUSIONS: Utilization of FIT at a threshold ≥10 µg/g can safely triage patients with low-risk symptoms in primary care, with negative results effectively ruling out CRC.

Production of trimeric SARS-CoV-2 spike protein by CHO cells for serological COVID-19 testing.

We describe scalable and cost-efficient production of full length, His-tagged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike glycoprotein trimer by Chinese hamster ovary (CHO) cells that can be used to detect SARS-CoV-2 antibodies in patient sera at high specificity and sensitivity. Transient production of spike in both human embryonic kidney (HEK) and CHO cells mediated by polyethyleneimine was increased significantly (up to 10.9-fold) by a reduction in culture temperature to 32°C to permit extended duration cultures. Based on these data GS-CHO pools stably producing spike trimer under the control of a strong synthetic promoter were cultured in hypothermic conditions with combinations of bioactive small molecules to increase yield of purified spike product 4.9-fold to 53 mg/L. Purification of recombinant spike by Ni-chelate affinity chromatography initially yielded a variety of co-eluting protein impurities identified as host cell derived by mass spectrometry, which were separated from spike trimer using a modified imidazole gradient elution. Purified CHO spike trimer antigen was used in enzyme-linked immunosorbent assay format to detect immunoglobulin G antibodies against SARS-CoV-2 in sera from patient cohorts previously tested for viral infection by polymerase chain reaction, including those who had displayed coronavirus disease 2019 (COVID-19) symptoms. The antibody assay, validated to ISO 15189 Medical Laboratories standards, exhibited a specificity of 100% and sensitivity of 92.3%. Our data show that CHO cells are a suitable host for the production of larger quantities of recombinant SARS-CoV-2 trimer which can be used as antigen for mass serological testing.

Phytosynthesis of gold nanoparticles using Mappia foetida leaves extract and their conjugation with folic acid for delivery of doxorubicin to cancer cells.

Mappia foetida leaves extract is used as bioreductant for the synthesis of gold nanoparticles and their application in the efficient delivery of doxorubicin to human cancer cells is reported here. The formation of gold nanoparticles is evident from their characteristic optical absorption at ~560 nm. X-ray diffraction pattern of gold nanoparticles confirmed their fcc structure. Fourier transform infrared spectroscopy shows the bioactive molecules from plant extract capped on the surface of gold nanoparticles and conjugation of doxorubicin along with activated folic acid as navigational molecules for targeted drug delivery. Such a conjugation of gold nanoparticles is characterized by their weight loss, ~35-40 %, due to thermal degradation of plant biomass and conjugated drug along with receptor, as observed in thermogravimetric analysis. The spherical shaped gold nanoparticles (Φ 10-20 nm) are observed by field emission scanning electron microscopy and transmission electron microscopy images and the expected elemental composition by energy dispersive X-ray spectroscopy. Gold nanoparticles conjugated with activated folic acid and doxorubicin complex is found to be toxic for human cancer cells viz., MDA-MB-231, HeLa, SiHa and Hep-G2. Furthermore, the amount of drug released was maximum at pH 5.3 (an ambient condition for intravenous cancer drugs) followed by pH 7.2 and pH 6.8.

Anti-arthritic Activity of Dashanga Ghana (An Ayurvedic Compound Formulation) Against Freund's Adjuvant Induced Arthritis in Charles Foster Albino Rats.

INTRODUCTION: Arthritis is the most common cause of disability, limiting the activities of adults throughout the world. Apart from the conventional treatment strategies using non-steroidal anti-inflammatory drugs, disease-modifying anti-rheumatic drugs, and glucocorticoids, newer and safer drugs are continuously being searched, as long-term usage of these drugs have resulted in adverse effects. Besides this, currently a number of medicinal plants are under scientific evaluation to develop a promising remedy in these cases. There is a need to investigate the complete therapeutic potential of these herbals for providing newer and safer treatment options with minimum side effects. Considering this, a polyherbal Ayurvedic compound formulation (Dashanga Ghana) has been studied in experimental animals to evaluate anti-arthritic activity. MATERIALS AND METHODS: Dashanga Ghana has been prepared in the laboratory by following standard guidelines. Charles Foster albino rats were used to evaluate the activity through Freund's adjuvant induced arthritis model. RESULTS AND CONCLUSIONS: Dashanga Ghana is found to possess significant anti-arthritic activity. Further studies are required to identify and characterize exact active phyto-constituents and to elucidate the exact mechanism of action, which is responsible for the observed pharmacological profile.

Serum proteomics of hepatitis C virus infection reveals retinol-binding protein 4 as a novel regulator.

Persistent infection of hepatitis C virus (HCV) can lead to liver cirrhosis and hepatocellular carcinoma, which are currently diagnosed by invasive liver biopsy. Approximately 15-20 % of cases of chronic liver diseases in India are caused by HCV infection. In North India, genotype 3 is predominant, whereas genotype 1 is predominant in southern parts of India. The aim of this study was to identify differentially regulated serum proteins in HCV-infected Indian patients (genotypes 1 and 3) using a two-dimensional electrophoresis approach. We identified eight differentially expressed proteins by MS. Expression levels of one of the highly upregulated proteins, retinol-binding protein 4 (RBP4), was validated by ELISA and Western blotting in two independent cohorts. We also confirmed our observation in the JFH1 infectious cell culture system. Interestingly, the HCV core protein enhanced RBP4 levels and partial knockdown of RBP4 had a positive impact on HCV replication, suggesting a possible role for this cellular protein in regulating HCV infection. Analysis of RBP4-interacting partners using a bioinformatic approach revealed novel insights into the possible involvement of RBP4 in HCV-induced pathogenesis. Taken together, this study provided information on the proteome profile of the HCV-infected Indian population, and revealed a link between HCV infection, RBP4 and insulin resistance.

Rejection of Fecal Immunochemical Tests Within the Lower Gastrointestinal Diagnostic Pathway: A Cohort Study.

INTRODUCTION/OBJECTIVES: The fecal immunochemical test (FIT) helps triage primary care patients at risk of colorectal cancer (CRC). Improving FIT returns has received recent attention, however uncertainty exists regarding the accurate completion of samples provided for laboratory analysis. This study aims to identify the rejection rate of returned FIT samples and determine rejection causes. METHODS: FIT samples from symptomatic patients within South Yorkshire, Bassetlaw, and North Derbyshire are processed at a central laboratory. Tests requests are made from 225 GP practices, which serve an estimated 2 million population. This study describes a retrospective review of FIT samples received in the central laboratory between 01/09/19 and 31/12/22. Locally held data was interrogated in March 2023 to determine the number of FIT samples received and rejected during the study period. Documented reasons for rejection were explored to identify common themes. RESULTS: Total FIT specimens received during the study period was 126 422. Of these, 5190 (4.1%) were rejected. Monthly rejection rates fell from 17.4% in September 2019 to 1.3% in December 2022 (P < .001). Sampling errors were the most frequent cause for FIT rejection (2151/5190), with other causes including: expired specimen; no sample collection date/ time, no request form, incomplete patient information and illegible handwriting. CONCLUSIONS: This is the first study exploring FIT rejection rates in symptomatic primary care patients, which shows improvements in rejection rates over time. Targeted interventions could improve rejection rates further, thereby reducing NHS resource use and costs and diagnostic delays.

Evaluation of Punarnavadi Mandura for haematinic activity against mercuric chloride-induced anemia in albino rats.

Background: Punarnavadi Mandura, a compound Ayurvedic formulation, is one of the most commonly used medicine in the treatment of anemia (Pandu) in Ayurveda. The safety profile of this formulation is well established; however, no pharmacological study has been reported to provide the scientific basis for its use in the treatment of anemia. Aim: To evaluate the hematinic effect of Punarnavadi Mandura against mercuric chloride-induced anemia in albino rats. Materials and methods: The test drug (Punarnavadi Mandura) was suspended in distilled water and administered orally in a dose of 450 mg/kg for 30 days in rats. Anemia was induced by simultaneous administration of mercuric chloride (9 mg/kg) for 30 consecutive days. Body weight was noted for each animals. At the end, haematological parameters, anaemia markers like serum iron, ferritin, and total iron binding capacity (TIBC), as well as relative weight of organs and histopathology investigation, were examined. Results: Exposure of mercuric chloride to rats for 30 days resulted in a significant decrease of body weight, an increase in the weight of the liver and kidney and a decrease in hemoglobin content. It also decreased serum ferritin to a significant extent and increased serum TIBC. Histopathology of the liver shows macro fatty changes, vacuolization, marked necrosis, and severe degenerative changes, while the kidney shows cell infiltration. All these changes were significantly attenuated by the administration of Punarnavadi Mandura. Conclusion: The present data indicate that Punarnavadi Mandura has possessing marked cytoprotective activity, significantly attenuated the HgCl2-induced adverse changes on red blood cell related parameters, and showing hematinic activity in albino rats.

Prevention of in vitro glucose-induced cataract by Vasanjana prepared by Yashtimadhu Kalka (paste of Glycyrrhyza glabra Linn).

BACKGROUND: Cataract is a major cause of blindness worldwide. Researchers received much attention from the traditional systems such as Ayurveda for the solutions of cataract through antioxidant activities apart from the surgical extraction. AIMS: A To study, the anti-cataract activity of Vasanjana (VK) prepared with Yashtimadhu (Glycyrrhyza glabra Linn) Kalka (paste) in Vasa (fat) of the domestic fowl (Gallus gallus) on glucose-induced cataract in ovine (sheep) lenses. MATERIALS AND METHODS: Artificial aqueous humor with 55 mM glucose was used to induce cataract in sheep eye lenses. Treatment was given with cow ghee (CG), plain fat, Vasanjana, and Vitamin E to the same media and lenses were incubated at the room temperature for 72 h. Biochemical parameters studied in the lens were total proteins, malondialdehyde (MDA), Na+ K+ ATPase activity and electrolytes (Na+ and K+). Photographic evaluation was also done. RESULTS: The complete opacification induced by the glucose in ovine lens was observed in 72 h. Cataractous lenses showed significant increase in Na+, MDA level and significant decrease in Na+ K+ ATPase activity and total protein content. Lenses treated with Vasanjana showed non-significant increase of total protein content and decreased MDA level and prevented formation and progress of cataract by glucose, as evidenced by photographic evaluation. Glucose-induced biochemical changes were found to be reversed in statistically significant manner in CG and Vitamin E treated lenses. CONCLUSION: The anti-cataract activity of Vasanjana and CG may be because of the antioxidant and free radical scavenging activity. Further in vitro and in vivo studies in various experimental models are required to validate their anti-cataract activity.

Toxicity and recovery studies of two ayurvedic preparations of iron.

Lauha Bhasma and Mandura Bhasma in 55 mg/kg dose (5 times the therapeutic effective dose) for 60 days exhibited no serious toxic effects in Charles Foster albino rats. Both the drugs showed significant recovery from chronic toxic effect after 45 days of recovery period.

In-vivo study of tissue reaction to Crotalaria pallida and Sansevieria roxburghiana fibers.

BACKGROUND: A suture material producing least tissue reaction is considered as ideal. Other characteristics like tensile strength, capacity to sustain sterilization process enhance its acceptability. In the present situation there is a need to reascertain the relevance and utility of these materials. Among the suture materials mentioned by Sushrutacharya, Moorva (Sansevieria roxburghiana) and Shana (Crotalaria pallida) have been showed insignificant tissue reaction in operated cases of inguinal hernia. An experimental study to confirm the extent of tissue reaction in deeper planes is needed before extending the use of materials in the deep tissues. OBJECTIVE: The objective of the study was to analyze deep tissue reaction and tensile strengths of plant fibres extracted from Crotalaria pallida and Sansevieria roxburghiana. MATERIALS AND METHODS: The study was conducted on 18 albino rats, 3 groups of 6 rats each for a period of 21 days inserting the suture materials in deeper tissue, studying histopathology changes of the deeper connective tissues, hydroxyproline content and blood parameters on 7th, 14th, 21st days of the study. The tensile strength of the two materials was also assessed on 7th day in three different conditions. Statistical analysis was carried out using paired and unpaired t tests. RESULTS: S. roxburghiana had least tissue reaction. C. pallida showed greater tensile strength in comparison to Moorva. CONCLUSION: C. pallida can be used for deep tissue approximation because of its moderate tissue reaction and tensile strength, successive increase in hydroxyproline content and its capacity to sustain sterilization.

British Lung Foundation/United Kingdom Primary Immunodeficiency Network Consensus Statement on the Definition, Diagnosis, and Management of Granulomatous-Lymphocytic Interstitial Lung Disease in Common Variable Immunodeficiency Disorders.

A proportion of people living with common variable immunodeficiency disorders develop granulomatous-lymphocytic interstitial lung disease (GLILD). We aimed to develop a consensus statement on the definition, diagnosis, and management of GLILD. All UK specialist centers were contacted and relevant physicians were invited to take part in a 3-round online Delphi process. Responses were graded as Strongly Agree, Tend to Agree, Neither Agree nor Disagree, Tend to Disagree, and Strongly Disagree, scored +1, +0.5, 0, -0.5, and -1, respectively. Agreement was defined as greater than or equal to 80% consensus. Scores are reported as mean ± SD. There was 100% agreement (score, 0.92 ± 0.19) for the following definition: "GLILD is a distinct clinico-radio-pathological ILD occurring in patients with [common variable immunodeficiency disorders], associated with a lymphocytic infiltrate and/or granuloma in the lung, and in whom other conditions have been considered and where possible excluded." There was consensus that the workup of suspected GLILD requires chest computed tomography (CT) (0.98 ± 0.01), lung function tests (eg, gas transfer, 0.94 ± 0.17), bronchoscopy to exclude infection (0.63 ± 0.50), and lung biopsy (0.58 ± 0.40). There was no consensus on whether expectant management following optimization of immunoglobulin therapy was acceptable: 67% agreed, 25% disagreed, score 0.38 ± 0.59; 90% agreed that when treatment was required, first-line treatment should be with corticosteroids alone (score, 0.55 ± 0.51).

Cerebral salt wasting syndrome in a patient with tuberculous meningitis.

We report a case of a 65 year male with meningitis who had polyuria, severe hyponatremia, volume depletion and very high urinary sodium excretion. He was diagnosed to have cerebral salt wasting syndrome based on clinical and laboratory parameters.

Development of quality standards of medicinal mistletoe - Helicanthes elastica (Desr.) Danser employing Pharmacopoeial procedures.

Helicanthes elastica (Desr.) Danser (Loranthaceae), commonly known as Indian mango mistletoe, is a parasitic shrub found widely growing on mango trees in southern India. Development of monographic quality standards is need of the hour for Pharmacopoeial/extra-Pharmacopoeial and folk medicinal plants. Systematic pharmacognostical evaluation of leaves of H. elastica has been carried out employing Pharmacopoeial procedures of testing herbal drugs. Macro-microscopic features of H. elastica leaf were recorded. Ethanolic extract was tested positive for alkaloids, steroids, carbohydrates, tannins, saponins and phenols. HPTLC fingerprint profile was developed for the identification of extracts using reference standard ß-sitosterol glucoside. Results of the present investigation would serve as a source of pharmacognostical information and a document to control the quality of H. elastica (Desr.) Danser.

Effect and Mechanism of Virechana Karma (Therapeutic Purgation) Over Fructose-Induced Metabolic Syndrome: An Experimental Study.

BACKGROUND: Panchakarma (biopurification methods) is one of the modes of ayurveda to treat disorders of the body. Virechana karma (therapeutic purgation), one among the Panchakarma, is a purification process that is commonly used to treat metabolic disorders like obesity and diabetes mellitus. Hence this study was planned to provide evidence through animal experiments. METHODS: Albino rats were subject to Virechana karma (therapeutic purgation) to evaluate the influence of therapy and its mechanism over fructose-induced metabolic syndrome. RESULTS: Results show that Virechana is effective in the management of the metabolic syndrome with decrease in the fecal fat content, fasting blood glucose, serum triglyceride, and reduced fatty changes in liver, heart, and kidney in comparison with the positive control group. CONCLUSION: Experimental evaluation showed decrease in fatty acid in the storage like liver, kidney, heart, and muscle adipose tissue can indirectly increase the insulin sensitivity in insulin receptor present at skeletal muscles.

Phytochemically Functionalized Cu and Ag Nanoparticles Embedded in MWCNTs for Enhanced Antimicrobial and Anticancer Properties.

Nanomedicine is an emerging field concerned with the use of precision engineered nanomaterials, which leads to the development of novel remedial and diagnostic modalities for human use. In this study, Cu(NO3)2 and AgNO3 precursors were reduced to copper nanoparticles (CuNPs) and silver nanoparticles (AgNPs) using Terminalia arjuna bark extracts under microwave irradiation in the presence of well-dispersed multi-walled carbon nanotubes (MWCNTs) in aqueous medium. The formation of CuNPs or AgNPs and their functionalization with MWCNTs via bioactive molecules of plant extract were evidenced from UV-Vis spectra, XRD, FTIR, FESEM, EDX, and TEM images. The phytochemically functionalized Cu-MWCNTs and Ag-MWCNTs nanomaterials showed enhanced biocide activity, and the inhibitory activity for bacteria was higher than that of fungus. Furthermore, these biohybrid nanomaterials are non-toxic to normal epithelial cells (Vero), whereas they are highly toxic for tested human cancer cells of MDA-MB-231, HeLa, SiHa, and Hep-G2. The cell viability was found to decrease with the increasing dose from 10 to 50 µg mL-1, as well as incubation time from 24 to 72 h. For instance, the cell viability was found to be ~91 % for normal Vero cells and ~76 % for cancer cells for lower dose of 10 µg mL-1.

High performance thin layer chromatography qualitative densitometry as a sensitive method to assess shelf life of polyherbal formulations: A study on Hutabhugadi Churna.

INTRODUCTION: Measuring chemical stability of polyherbal formulations is very challenging due to diversity in phytochemical composition. As there are no comprehensive guidelines for stability testing of herbal products, there is a need for a sensitive tool to detect how the quality of herbal products varies with time under the influence of environmental conditions. AIMS: To validate the employability of high-performance thin layer chromatography (HPTLC) for real-time stability of Hutabhugadi Churna (HC). MATERIALS AND METHODS: The chromatograms were developed using toluene/ethyl acetate/formic acid (10:5:1) and ethyl acetate/formic acid (10:1) as a mobile phase for chloroform and ethanolic extract, respectively. The plates were scanned under 254, 366, 540 (pre-derivatization) and 540 nm (post-derivatization). Samples were analyzed immediately after preparation and after 3(rd) and 6(th) months of storage. Alteration of fingerprint profiles from the initial pattern, in terms of number of peaks, was employed as diagnostic tools. Percentage variation in composition at given period was calculated. RESULTS: HC is found to be stable at room temperature up to 1.3 months using the method of calculation of 10% degradation period employing slope and intercept values for the initial, 3(rd) and 6(th) months' deviation in number of bands. The data obtained were subjected to regression analysis in context to number of bands obtained. The curve was found to be linear with R(2) value of 0.89-0.96 supported by their tolerance range of 0.04-0.11. CONCLUSION: The proposed model is a new logic with prospects to become working method for stability assessment of polyherbal formulations under controlled conditions.

Investigation of the anti-inflammatory effects of Paederia foetida.

A detailed investigation on the anti-inflammatory activity of the butanol fraction of a methanol extract (BMEL) of the defatted leaves of Paederia foetida was undertaken to find the pharmacological basis for the ethnomedical use of the plant. This fraction produced a significant inhibition of granulation tissue formation in cotton-pellet implanted rats. It decreased liver aspartate transaminase activity without affecting serum aspartate transaminase activity. It did not, however, affect adrenal weight and ascorbic acid content significantly, thus ruling out a stimulation of the adrenal-pituitary axis. BMEL antagonised hyposaline-induced haemolysis of human red blood cells and an elevation of rat serum acid phosphatase activity, indicating the presence of a membrane stabilising activity. It also inhibited the elevation of serum orosomucoid levels in rats, suggesting the possibility of the presence of disease-modifying antirheumatic activity. The results indicate that there is some rationale behind the ethnomedical use of the plant for treating inflammatory disorders.