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A 3½-year-old boy presented with generalized lymphadenopathy since 1 year of age. After 5 months of age, he had multiple infections and two episodes of pneumonia. At 2½ years, he developed chronic diarrhea. Colonoscopy showed ileal ulcers and inflammatory pan colitis with skip areas and histopathologic examination showed mild chronic ileitis, villous shortening, and diffused active colitis. Next-generation sequencing revealed a previously reported homozygous missense mutation Caspase-8. Unlike autoimmune lymphoproliferative syndrome, Caspase-8 deficiency states have immunodeficiency, autoimmunity, and early-onset inflammatory bowel disease in addition to lymphoproliferation. Early diagnosis aided by molecular confirmation is essential as haploidentical hematopoietic stem cell transplant is curative.

Objectives: Leucocyte adhesion defect 1 (LAD1) is an autosomal recessive defect in integrin expression. In this study we present a case series of 19 children with LAD1. We analysed their clinical and laboratory data and correlated it with integrin expression and underlying genetic defect. Material and Methods: Retrospective case record study of all children diagnosed with LAD1 between 2012 and 2021. Results: Ten females and 9 males were included. Eight cases were born of consanguineous marriage. 78% (15/19) had disease onset within one year of life and 8 of them were neonates. Neonates with LAD1 commonly presented with omphalitis, late-onset sepsis and respiratory infections (RTI). Most common organism isolated from blood was pseudomonas (4/8). CD11/CD18-expression ranged from severely reduced (6/8) to moderately reduced (2/8) but disease was uniformly fatal in the 7/8 patients who could not undergo hematopoietic stem cell transplant (HSCT). Seven children presented between 1month and 1year of age with RTI, bacterial sepsis and soft tissue abscess. Four expired, 1 survived after HSCT and 2 were alive at last follow up. CD11/CD18-expression ranged between 0-1.6% except in 2 cases. One child with 11%-expression is alive without transplant. Another child had 0%-CD11 but 17%-CD18-expression with very low MFI (mean florescence intensity). She succumbed at 20 months of age with perianal abscess, sepsis and RTI but prior to this fatal episode she developed autoimmune cutaneous lesions with lymphocytic infiltration. Four children presented beyond infancy. Median age at onset of symptom was 2 years with skin abscess, oral ulcers, severe periodontitis with tooth loss and pyogenic arthritis. Three of them survived without HSCT, 1 was alive at last follow up. CD11/CD18-expression ranged from 20-32% in these children. Details of cord separation was available in 17 cases and it was delayed (beyond 2 weeks of life) in 9 children. Of the 17 cases with genetic analysis of ITGB2 gene, 16 had a homozygous and 1 had compound heterozygous variant. (nonsense: 2; splice site variants:5; 8: missense variants; 2: deletion). Conclusion: Disease onset less than 1 year age was fatal, irrespective of CD11/CD18 expression. Integrin expression levels beyond 20% and splice site mutation were associated with 100% survival without HSCT but were not event free. Pseudomonas sepsis is the commonest infection in LAD1 neonates. Between 1month to 1year, RTI and gram-negative sepsis dominated and beyond 1st-year skin/perianal abscess was common. Delayed separation of cord was present only in 52% of cases.

Background : Primary immune regulatory disorders (PIRD) is a new term coined for a group of PID where autoimmune complications predominate. Herein we present a case series of 10 cases with lipopolysaccharide (LPS)-responsive and beige-like anchor protein (LRBA) deficiency who presented with multiple autoimmunities. Methods: 10 patients with molecular diagnosis of LRBA deficiency were included and their clinical data was evaluated. Results: The mean age of onset was 4.3 years. Male to female ratio was 7:3. 8/10 had 2 or more autoimmune disease with autoimmune cytopenia being the commonest. Other autoimmune diseases seen were autoimmune hepatitis, inflammatory bowel disease, enteropathy, diabetes melitis, thyroiditis, CNS vasculitis, dermatitis and alopecia areata. All patients had evidence of lymphoproliferation with generalized lymphadenopathy and/or hepatosplenomegaly. 5/10 had hypogammaglobinemia, 3/10 had low T cells, two of whom had CD4 lymphopenia. 4/10 had low B and NK cells. 2 had compound heterozygous and 8 had homozygous mutations in LRBA gene. The treatment was diverse and included corticosteroids, cyclosporine, azathioprine, cyclosporine, rituximab, sirolimus and abatacept. One child has undergone successful HSCT and currently doing well. Conclusion: Patients with LRBA deficiency present with a broad range of clinical manifestations. High index of suspicion for a monogenic cause for polyautoimmunity in early childhood can reduce time delay in diagnosis. With increasing availability of transplant facilities the outcome for these patients can be significantly better.

SWPs are the major virulence component of microsporidian spores. In microsporidia, SWPs can be found either in exospore or endospore to serve as a putative virulence factor for host cell invasion. SWP5 is a vital protein that involves in exospore localization and supports the structural integrity of the spore wall and this action potentially modulates the course of infection in N. bombycis. Here we report recombinant SWP5 purification using Ni-NTA IMAC and SEC. GFC analysis reveals SWP5 to be a monomer which correlates with the predicted theoretical weight and overlaps with ovalbumin peak in the chromatogram. The raised polyclonal anti-SWP5 antibodies was confirmed using blotting and enterokinase cleavage experiments. The resultant fusion SWP5 and SWP5 in infected silkworm samples positively reacts to anti-SWP5 antibodies is shown in ELISA. Immunoassays and Bioinformatic analysis reveal SWP5 is found to be localized on exospore and this action could indicate the probable role of SWP5 in host pathogen interactions during spore germination and its contribution to microsporidian pathogenesis. This study will support development of a field-based diagnostic kit for the detection N. bombycis NIK-1S infecting silkworms. The analysis will also be useful for the formulation of drugs against microsporidia and pebrine disease.

In 2014, a child with broad combined immunodeficiency (CID) who was homozygous for a private BCL10 allele was reported to have complete inherited human BCL10 deficiency. In the present study, we report a new BCL10 mutation in another child with CID who was homozygous for a BCL10 variant (R88X), previously reported as a rare allele in heterozygosis (minor allele frequency, 0.000003986). The mutant allele was a loss-of-expression and loss-of-function allele. As with the previously reported patient, this patient had complete BCL10 deficiency. The clinical phenotype shared features, such as respiratory infections, but differed from that of the previous patient that he did not develop significant gastroenteritis episodes or chronic colitis. Cellular and immunological phenotypes were similar to those of the previous patient. TLR4, TLR2/6, and Dectin-1 responses were found to depend on BCL10 in fibroblasts, and final maturation of T cell and B cell maturation into memory cells was affected. Autosomal-recessive BCL10 deficiency should therefore be considered in children with CID.

BackgroundWiskott Aldrich syndrome (WAS) is characterized by bleeding manifestations, recurrent infections, eczema, autoimmunity, and malignancy. Over the last decade, improved awareness and better in-house diagnostic facilities at several centers in India has resulted in increased recognition of WAS. This study reports collated data across major primary immunodeficiency diseases (PID) centers in India that are involved in care of children with WAS and highlights the varied clinical presentations, genetic profile, and outcomes of patients in India.MethodsRequest to share data was sent to multiple centers in India that are involved in care and management of patients with PID. Six centers provided requisite data that were compiled and analyzed.ResultsIn this multi-institutional cohort, clinical details of 108 patients who had a provisional diagnosis of WAS were received. Of these, 95 patients with ‘definite WAS’ were included Fourteen patients were classified as XLT and 81 patients as WAS. Median age at onset of symptoms of patients was 3 months (IQR 1.6, 6.0 months) and median age at diagnosis was 12 months (IQR 6,48 months). Clinical profile included bleeding episodes (92.6%), infections (84.2%), eczema (78.9%), various autoimmune manifestations (40%), and malignancy (2.1%). DNA analysis revealed 47 variants in 67 cases. Nonsense and missense variants were the most common (28.4% each), followed by small deletions (19.4%), and splice site defects (16.4%). We also report 24 novel variants, most of these being frameshift and nonsense mutations resulting in premature termination of protein synthesis. Prophylactic intravenous immunoglobulin (IVIg) was initiated in 52 patients (54.7%). Hematopoietic stem cell transplantation (HSCT) was carried out in 25 patients (26.3%). Of those transplanted, disease-free survival was seen in 15 patients (60%). Transplant related mortality was 36%. Outcome details were available for 89 patients. Of these, 37% had died till the time of this analysis. Median duration of follow-up was 36 months (range 2 weeks- 12 years; IQR 16.2 months- 70 months).ConclusionsWe report the first nationwide cohort of patients with WAS from India. Bleeding episodes and infections are common manifestations. Mortality continues to be high as curative therapy is not accessible to most of our patients.

Microsporidia are intracellular fungal parasites and they are the most common pathogens for sericulture. Microsporidian sp. can cause pebrine, a dreadful disease and lead to destructive disorder in Muga silkworm, Antheraea assamensis Helfer by vertical and horizontal transmission. This disease is the key factor obstructing the developmental progress of Muga culture in India. Nevertheless, molecular identification and characterization of pathogen associated with pebrine disease in A. assamensis is not yet established. Insect bioassay studies revealed that microsporidian infection in Muga silkworm, A. assamensis Helfer significantly reduced (P

Microsporidiosis (Pebrine) caused by the microsporidian parasite is one of the important devastating disease which affect the silk production leading to an unprofitable harvest. Till date ribosomal RNA (rRNA) gene was used as a target for detection of microsporidian species. In this study, we describe conventional and SYBR green based real-time PCR techniques alternatively targeting β-tubulin gene for quantitative detection of microsporidia infecting both the mulberry and non-mulberry silkworms. The modified DNA extraction method followed in our study was found to be easy, economical and could be used for both conventional and real time PCR as template. The real time qPCR revealed the expression of β-tubulin gene in different infected tissues of the silkworm Bombyx mori. The sensitivity of the SYBR green based real time PCR was found to be 100 times more than the conventional PCR and PCR was found more sensitive than the microscopic examination. The developed method did not produce any false positive results with the other silkworm pathogens and healthy silkworm. The data suggest that both the developed PCR methods targeting β-tubulin gene could be used effectively in quarantine process at seed centres for early detection of microsporidian infection in silkworms.

Hemophagocytic lymphohistiocytosis (HLH) is a syndrome of immune dysregulation characterized by hyperactivation of the immune system, excessive cytokine secretion and severe systemic inflammation. HLH is classified as familial (FHL) when associated with mutations in PRF1, UNC13D, STX11, and STXBP2 genes. There is limited information available about the clinical and mutational spectrum of FHL patients in Indian population. This study is a retrospective analysis of 101 molecularly characterized FHL patients over the last 10 years from 20 different referral centers in India. FHL2 and FHL3 together accounted for 84% of cases of FHL in our cohort. Patients belonging to different FHL subtypes were indistinguishable based on clinical and biochemical parameters. However, flow cytometry-based assays viz. perforin expression and degranulation assay were found to be specific and sensitive in diagnosis and classification of FHL patients. Molecular characterization of respective genes revealed 76 different disease-causing mutations including 39 (51%) novel mutations in PRF1, UNC13D, STX11, and STXBP2 genes. Overall, survival was poor (28%) irrespective of the age of onset or the type of mutation in our cohort. Altogether, this article sheds light on the current scenario of FHL in India. Our data reveal a wide genetic heterogeneity of FHL in the Indian population and confirms the poor prognosis of FHL. This study also emphasizes that though mutational analysis is important for diagnostic confirmation of FHL, flow cytometry based assays help significantly in rapid diagnosis and functional validation of novel variants identified.

BackgroundSevere Combined Immune Deficiency (SCID) is an inherited defect in lymphocyte development and function that results in life-threatening opportunistic infections in early infancy. Data on SCID from developing countries are scarce.ObjectiveTo describe clinical and laboratory features of SCID diagnosed at immunology centers across India.MethodsA detailed case proforma in an Excel format was prepared by one of the authors (PV) and was sent to centers in India that care for patients with primary immunodeficiency diseases. We collated clinical, laboratory, and molecular details of patients with clinical profile suggestive of SCID and their outcomes. Twelve (12) centers provided necessary details which were then compiled and analyzed. Diagnosis of SCID/combined immune deficiency (CID) was based on 2018 European Society for Immunodeficiencies working definition for SCID.ResultsWe obtained data on 277 children; 254 were categorized as SCID and 23 as CID. Male-female ratio was 196:81. Median (inter-quartile range) age of onset of clinical symptoms and diagnosis was 2.5 months (1, 5) and 5 months (3.5, 8), respectively. Molecular diagnosis was obtained in 162 patients - IL2RG (36), RAG1 (26), ADA (19), RAG2 (17), JAK3 (15), DCLRE1C (13), IL7RA (9), PNP (3), RFXAP (3), CIITA (2), RFXANK (2), NHEJ1 (2), CD3E (2), CD3D (2), RFX5 (2), ZAP70 (2), STK4 (1), CORO1A (1), STIM1 (1), PRKDC (1), AK2 (1), DOCK2 (1), and SP100 (1). Only 23 children (8.3%) received hematopoietic stem cell transplantation (HSCT). Of these, 11 are doing well post-HSCT. Mortality was recorded in 210 children (75.8%).ConclusionWe document an exponential rise in number of cases diagnosed to have SCID over the last 10 years, probably as a result of increasing awareness and improvement in diagnostic facilities at various centers in India. We suspect that these numbers are just the tip of the iceberg. Majority of patients with SCID in India are probably not being recognized and diagnosed at present. Newborn screening for SCID is the need of the hour. Easy access to pediatric HSCT services would ensure that these patients are offered HSCT at an early age.

BackgroundThere is paucity of literature on XLA from developing countries. Herein we report the clinical and molecular profile and outcome in a multicenter cohort of patients with XLA from India.MethodsData on XLA from all regional centers supported by the Foundation for Primary Immunodeficiency Diseases (FPID), USA and other institutions providing care to patients with PIDs were collated. Diagnosis of XLA was based on European Society for Immunodeficiencies (ESID) criteria.ResultsWe received clinical details of 195 patients with a provisional diagnosis of XLA from 12 centers. At final analysis, 145 patients were included (137 ‘definite XLA’ and eight ‘probable/possible XLA’). Median age at onset of symptoms was 12.0 (6.0, 36.0) months and median age at diagnosis was 60.0 (31.5, 108) months. Pneumonia was the commonest clinical manifestation (82.6%) followed by otitis media (50%) and diarrhea (42%). Arthritis was seen in 26% patients while 23% patients developed meningitis. Bronchiectasis was seen in 10% and encephalitis (likely viral) in 4.8% patients. Pseudomonas aeruginosa was the commonest bacterial pathogen identified followed by Streptococcus pneumoniae, Staphylococcus aureus and Klebsiella pneumoniae. Molecular analysis revealed 86 variants in 105 unrelated cases. Missense variants in BTK gene were the most common (36%) followed by frameshift (22%) and nonsense variants (21%). Most pathogenic gene variants (53%) were clustered in the distal part of gene encompassing exons 14–19 encoding for the tyrosine kinase domain. Follow-up details were available for 108 patients. Of these, 12% had died till the time of this analysis. The 5-year and 10-year survival was 89.9% and 86.9% respectively. Median duration of follow-up was 61 months and total duration of follow-up was 6083.2 patient-months. All patients received intravenous immunoglobulin (IVIg) replacement therapy. However, in many patients IVIg could not be given at recommended doses or intervals due to difficulties in accessing this therapy because of financial reasons and lack of universal health insurance in India. Hematopoietic stem cell transplant was carried out in four (2.8%) patients.ConclusionThere was a significant delay in the diagnosis and facilities for molecular diagnosis were not available at many centers. Optimal immunoglobulin replacement is still a challenge

Mendelian Susceptibility to Mycobacterial diseases (MSMD) are a group of innate immune defects with more than 17 genes and 32 clinical phenotypes identified. Defects in the IFN-γ mediated immunity lead to an increased susceptibility to intracellular pathogens like mycobacteria including attenuated Mycobacterium bovis-Bacillus Calmette-Guérin (BCG) vaccine strains and non-tuberculous environmental mycobacteria (NTM), Salmonella, fungi, parasites like Leishmania and some viruses, in otherwise healthy individuals. Mutations in the IL12RB1 gene are the commonest genetic defects identified. This retrospective study reports the clinical, immunological, and molecular characteristics of a cohort of 55 MSMD patients from 10 centers across India. Mycobacterial infection was confirmed by GeneXpert, Histopathology, and acid fast bacilli staining. Immunological workup included lymphocyte subset analysis, Nitro blue tetrazolium (NBT) test, immunoglobulin levels, and flow-cytometric evaluation of the IFN-γ mediated immunity. Genetic analysis was done by next generation sequencing (NGS). Disseminated BCG-osis was the commonest presenting manifestation (82%) with a median age of presentation of 6 months due to the practice of BCG vaccination at birth. This was followed by infection with Salmonella and non-typhi Salmonella (13%), Cytomegalovirus (CMV) (11%), Candida (7%), NTM (4%), and Histoplasma (2%). Thirty-six percent of patients in cohort were infected by more than one organism. This study is the largest cohort of MSMD patients reported from India to the best of our knowledge and we highlight the importance of work up for IL-12/IL-23/ISG15/IFN-γ circuit in all patients with BCG-osis and suspected MSMD irrespective of age.

Background: Systemic autoinflammatory diseases (SAID) are rare inherited disorders involving genes regulating innate immune signaling and are characterized by periodic or chronic multi-systemic inflammation.Objective: To describe spectrum of clinical, immunological, molecular features, and outcomes of patients with SAID in India.Methods: Request to share data was sent to multiple centers in India that are involved in care and management of patients with Inborn Errors of Immunity. Six centers provided requisite data that were compiled and analyzed.Results: Data on 107 patients with SAID were collated—of these, 29 patients were excluded due to unavailability of complete information. Twelve patients (15%) had type 1 interferonopathies, 21 (26%) had diseases affecting inflammasomes, 30 patients (41%) had non-inflammasome related conditions and 1five patients (19%) had Periodic Fever, Aphthous Stomatitis, Pharyngitis, Adenitis (PFAPA). Type1 interferonopathies identified in the cohort included patients with Deficiency of Adenosine Deaminase 2 (DADA2) (six patients; five families); STING-associated vasculopathy infantile-onset (SAVI) (three patients, one family); Spondyloenchondro-dysplasia with Immune Dysregulation (SPENCD) (two patients). Diseases affecting inflammasomes include Mevalonate Kinase Deficiency (eight patients); Cryopyrin-Associated Periodic Syndromes (CAPS) (seven patients); NLR Family, Pyrin domain-containing 12 (NLRP12) (two patients); Familial Mediterranean fever (FMF) (two patients); Autoinflammation and PLCG2-associated antibody deficiency and immune dysregulation (APLAID) (two patients). TNF receptor-associated periodic syndrome (TRAPS) (three patients); A20 haploinsufficiency (four patients); Deficiency of Interleukin 1 Receptor Antagonist (DIRA) (two patients) were categorized as non-inflammasome related conditions. There were significant delays in diagnosis Corticosteroids and other immunosuppressive agents were used for treatment as anti-IL-1 drugs and other biological agents were and still are not available in India. Eight (16.3%) patients had so far succumbed to their illness.Conclusions: This is the first nationwide cohort of patients with SAID from India. Clinical manifestations were diverse. Overlapping of clinical features with other relatively common rheumatological disorders often resulted in delays in diagnosis. More nationwide efforts are needed to enhance awareness of SAID among health care professionals and there is an urgent need to make targeted immunotherapies universally available.

Introduction:Hyper-IgE Syndrome (HIES) is a rare inborn error of immunity (IEI) characterized by a constellation of symptoms related to susceptibility toStaphylococcalskin and pulmonary infections, eczema, raised serum IgE (>2,000 IU/ml), craniofacial anomalies, and recurrent bone fractures. Data on HIES from the Indian subcontinent is scarce and restricted to small case series and case reports. This is the first compilation of national data on HIES.Materials and Methods:A total 103 cases clinically diagnosed and treated as HIES were analyzed from nine centers. Cases with clinical and/or molecular diagnosis of DOCK8 deficiency were not included. Patients were divided into two groups: group I for whom a heterozygous rare variant of STAT3 was identified, and group II, with clinical features similar to those of AD STAT3 deficiency, but without any genetic diagnosis.Results:Genetic diagnosis was available in 27 patients (26.2%) and all harbored rare variants in the STAT3 gene. Majority of these STAT3 HIES patients presented with recurrent skin abscesses (77.7%) or pneumonia (62.9%) or both (59.2%). Other features included eczema (37%), candidiasis (55.5%), facial dysmorphism (55.5%), recurrent fractures (11.1%), and retained primary teeth (7.4%).Mycobacterialinfections were seen in a significant 18.5%. Mortality was seen in three subjects (11.1%). A similar trend in the clinical presentation was observed when all the 103 patients were analyzed together. Twenty percent of patients without a rare variant in the STAT3 gene had an NIH score of ≥40, whereas, 51.9% of STAT3 HIES subjects had scores below the cut off of ≥40. TH17 cell numbers were low in 10/11 (90.9%) STAT3 HIES tested. Rare variants observed were 8 in exon 21; 8 in exon 13; 3 in exon 10; 2 in exon 15, and one each in exon 6, 16, 17, 19, 22, and splice site downstream of exon 12. Seven variants were novel and included F174S, N567D, L404Sfs*8, G419 =, M329K, T714I, R518X, and a splice site variant downstream of exon 12.Conclusions:The report includes seven novel STAT3 variants, including a rare linker domain nonsense variant and a CC domain variant.Mycobacterialdiseases were more frequent, compared to western literature.