Survey-based assessment of hematopoietic stem cell graft manipulation (minimal) by cell therapy labs supporting hematopoietic stem cell transplants in India.

BACKGROUND AIMS: Hematopoietic stem cell transplants (HSCTs) are increasingly being offered to patients in India for various conditions. The Indian Stem Cell Transplant Registry shows that a total of 2533 transplants were done in India in 2019. METHODS: An epidemiological descriptive cross-sectional survey (55 questions) of centers providing HSCT in India was planned to analyze variations in policies and practices regarding HSCT graft manipulation (i.e., plasma reduction, red blood cell [RBC] depletion and cryopreservation). A total of 63 of 102 centers responded to the survey (response rate, 61.7%), mostly from the northern part of India (27 of 63 [42.8%]). RESULTS: The majority of responding centers reported performing >50 HSCTs annually (n = 24 [38%]), and 92% (58 of 63) performed stem cell collections from a pediatric donor/patient (age <18 years). A total of 56 of 63 responding centers indicated that they did product manipulations involving cryopreservation (n = 45), plasma reduction (n = 42) and RBC depletion (n = 28). Cryopreservation was primarily done by blood centers (27 of 45 [60%]), with dimethyl sulfoxide (DMSO) being the primary constituent, used most commonly at a concentration of 5-10% (28 of 45 centers). Dump freezing was most commonly used (27 of 45) with a -80°C deep freezer. A 7-aminoactinomycin D based viability assessment was also most commonly used (30 of 45). Thawing of the product was done mainly at the bedside (30 of 45) using a wet-type thawer (36 of 45), and washing of DMSO was done by a few centers (seven of 45). Plasma reduction and RBC depletion were primarily done for ABO incompatibility at blood centers. CONCLUSIONS: This survey demonstrates the lack of standardization and uniformity in the minimal manipulation of hematopoietic stem cell grafts in centers supporting HSCT in India. This work also highlights the need for more studies and country-specific recommendations to establish best practices.

Production of MSTN Gene-Edited Embryos of Buffalo Using the CRISPR/Cas9 System and SCNT.

The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system and somatic cell nuclear transfer (SCNT) have been used to produce genome-edited farm animal species for improved production and health traits; however, these tools are rarely used in the buffalo and can play a pivotal role in milk and meat production in tropical and subtropical countries. In this study, we aimed to produce myostatin (MSTN) gene-edited embryos of the Murrah buffalo using the CRISPR/Cas9 system and SCNT. For this, fibroblast cells were electroporated with sgRNAs carrying all-in-one CRISPR/Cas9 plasmids targeting the first exon of the MSTN gene. Following puromycin selection, single-cell clonal populations were established and screened using the TA cloning and Sanger sequencing methods. Of eight single-cell clonal populations, one with a monoallelic and another with a biallelic heterozygous gene editing event were identified. These two gene-edited clonal cell populations were successfully used to produce blastocyst-stage embryos using the handmade cloning method. This work establishes the technical foundation for generation of genome-edited cloned embryos in the buffalo.

Neonatal and pediatric transfusion practices and policies in India: A survey-based cross-sectional assessment of blood centers.

BACKGROUND: Blood centers play a critical role in supporting neonatal and pediatric transfusions. We aim to study the variation in pediatric/neonatal transfusion policies and practices for blood centers in India. STUDY DESIGN AND METHODS: This is a survey-based (45 question) cross-sectional assessment of blood centers in India supporting pediatric/neonatal transfusions. RESULTS: One hundred three centers responded to the survey (response rate 51.2%; 103/201). As a part of pretransfusion testing, use of "microtainers" was reported by 58.4% (59/101) centers and only 57.4% (58/101) centers provide antibody screening. In case of absence of maternal sample, type O negative blood is most commonly used (48.5%; 49/101), and in case of ABO incompatibility, 68.3% (69/101) use units compatible with both mother and newborn. Leukoreduced RBCs are provided by 89% (90/101) centers and irradiated products are provided by 48.5% (49/101). 82% (83/101) of centers reported not receiving requests for CMV negative units considering the high incidence of CMV-seropositivity in donors. Fresh units are defined as <7 days old by 44.5% (45/101) and <5 days old by 31.6% (32/101) centers. 97% centers reported SAGM suspended RBCs in inventory but only 67.3% (68/101) use them for top-up transfusions to neonates. Overall >80% centers use the buffy-coat method for platelet concentrates preparation. Volume-based (ml/kg) aliquots preparation is done by 88% (89/101), mainly using the sterile connecting device (84.6%; 83/98). CONCLUSION: This survey shows heterogeneity and lack of standardization in practices across blood centers in India and highlights the need for more studies to establish best practices for this vulnerable age group.

Pluripotent Stem Cells for Livestock Health and Production.

Pluripotent stem cells (PSCs) have unlimited capacity for self-renewal and differentiation so that they can potentially produce any cell or tissue of animal's body. The PSCs derived from livestock represents a more appropriate model than a rodent for investigating human diseases due to their higher anatomical and physiological resemblance with human. Apart from that, livestock PSCs hold immense promises for innovative therapies, transgenic animal production and their biomedical interest. The realization of the full potential of PSCs, however, depends on the elucidation of the molecular mechanisms which play a critical role in the maintenance of pluripotency and reprogramming procedure remains poorly understood in livestock which in turn impedes the generation of true PSCs and their usage for clinical research. An in-depth understanding of pluripotency is extremely essential for improving health and welfare of livestock animals. Therefore, the present review focuses on the milestone achievements of PSCs in livestock animals and their potential application in health and production of livestock.

Autoimmune hemolytic anemia in children: Clinical presentation and treatment outcome.

BACKGROUND: Autoimmune hemolytic anemias (AIHA) are characterized by the destruction of red cells following the production of autoantibodies directed against them. Although AIHA in children is usually self-limiting, many still succumb to the illness due to delay in the diagnosis and treatment. AIHA in children may be secondary to autoimmune diseases, drugs, or immune deficiencies. Early diagnosis and appropriate immunohematological evaluation can aid in the diagnosis and treatment. OBJECTIVE: To analyze the evaluation, treatment, and outcome of AIHA in children. METHODS: Prospective data of patients aged 0-18 years diagnosed with AIHA between June 2017 and May 2019 were collected. INTERVENTION: Prednisolone was the first-line agent in all; second-line agents included cyclosporine and rituximab. Red cell transfusion was given in those with severe anemia with cardiac decompensation. RESULTS: Eleven patients were diagnosed during the study period. Hemoglobin ranged from 1.2 to 9 g/dl. The initial presentation was severe anemia in 8 children and moderate anemia with thrombocytopenia in 3. The trigger was infection in 5. Polyspecific direct coomb's test (DCT) was positive in 10 patients. 2/10 polyspecific DCT-positive cases on further evaluation had immunoglobulin G (IgG) and C3d positivity, whereas rest 8 had only IgG. One infant was diagnosed with DCT-negative immunoglobulin A-mediated AIHA. 4/11 attained remission following the short course of prednisolone. Cyclosporine was used as the second-line agent in 2 and rituximab was used in 3. Seven children are in sustained remission and off medication. One died within 12 h of diagnosis. CONCLUSION: AIHA is not an uncommon problem in children and can vary in its clinical severity. Early and correct diagnosis helps in deciding appropriate treatment.

Survey based cross-sectional study to analyse the variation of practices at blood centres during COVID-19 pandemic in India.

India has almost 3,000 blood centres collecting more than 11 million units annually. Maintaining blood supply during the COVID-19 pandemic is a huge challenge. We conducted a cross-sectional study by an online survey to analyse the variation of practices across blood centers of India during this pandemic. A total of 196 blood centers completely responded to the online survey. Most of the blood centres who responded were part of Government hospitals (60 %), part of an academic institutes (55.6 %) and were directly supporting a COVID hospital (67.5 %). Almost 95.4 % blood centers reported reduction of blood donation mainly due to lockdown (50 %) and inability to conduct camps (17.3 %). Scheduling blood donations was one of the most difficult to implement strategy for maintaining adequate blood donation (40.2 %). Blood center manpower management was also a challenge and upto 48 % blood centers operated in two batches to ensure social distancing in blood banks and reduce the risk of exposure. Hemato-oncology (36.8 %) and obstetrics (33.7 %) were major utilizer of blood during the pandemic. There were marked variations in use of PPE by blood banks staff as well as strategies adopted while conducting immunohematology tests on COVID-19 positive patients samples. This pandemic has highlighted some of the major limitations of the health services but blood services have risen to the challenge and strived to maintain the blood supply chain while ensuring blood donor and staff safety. The wide variations in the practices adopted highlights the need for uniform guidelines for blood services in future pandemics.

Convalescent plasma to aid in recovery of COVID-19 pneumonia in a child with acute lymphoblastic leukemia.

The natural history of COVID-19 infection in children is still evolving as the pandemic unfolds. Few cases of severe and often fatal COVID-19 have been reported although the infection is mild in the large majority. Children with cancers are recognised as a high risk group for all infections. Since there aren't any definite treatment guidelines established in children with severe COVID, treatment is guided by adult recommendations which too are often not evidence based. We report the case of a 4-year-old girl with severe COVID-19 associated pneumonia who presented to us as febrile neutropenia. The use of convalescent plasma along with steroids and IVIG showed dramatic results in this child and she recovered without the need for any specific treatment. This is highlighted as one of the earliest cases that is reporting the use of convalescent plasma in a child; the first ever in a child with underlying malignancy.

Cryobanking of primary somatic cells of elite farm animals - A pilot study in domesticated water buffalo (Bubalus bubalis).

Buffalo is an important farm animal species in South and South-east Asian countries. Cryopreservation allows long-term storage of somatic cells, which can be made available to research communities. This study aimed to 1) establish and cryopreserve somatic cells from elite buffaloes, and 2) share stored somatic cells and their associated data with researchers. To achieve these targets, somatic cells were established successfully from tail-skin biopsies of 17 buffaloes. The informative data such as buffalo details (breed, date of birth, sex, and age at the time of tissue biopsy collection, and production traits), the number of cryovials stored, and freezing dates were recorded in an electronic file and a printed inventory record. The established somatic cells were flat, spindle-shaped morphology, and expressed vimentin (a fibroblast-like cell type marker) and the negative expression of cytokeratin-18 (an epithelial cell type marker). Altogether, we cryopreserved 970 cryovials (0.1 million cells per vial) from two buffalo breeds, namely Murrah and Nili-Ravi (at least 45 cryovials per animal), for cryobanking. Somatic cell nuclear transfer (SCNT) experiments demonstrated the utility of cryopreserved cells to produce cloned buffaloes. Importantly, these cryopreserved somatic cells are made available to scientific communities. This study encourages the cryopreservation of somatic cells of elite farm animals for their utilization in cell-based research.

IgA-mediated autoimmune hemolytic anemia in an infant.

Autoimmune Hemolytic anemia (AIHA) a relatively uncommon form of hemolytic anemia in children, occurs due to the premature destruction of red blood cells caused by presence of autoantibodies directed against antigens on RBCs. Warm reactive AIHA is the most common form due to IgG isotype of immunoglobulin class binding to autologous RBCs at 370C and confirmed with a positive DAT screening. We present a case of DAT-negative primary warm AIHA in an infant due to IgA antibody. A 10 month old male infant presented with dark colored urine and irritability for past two months, with associated history of fever, diarrhea and vomiting. He had received one red cell transfusion 10 days prior. On physical examination he had pallor with tachycardia without splenomegaly. On investigation his hemoglobin was 5.8 g/dl, WBC 25.9 × 103/mm3 and normal platelets counts. Peripheral blood smear had spherocytes and biochemical values showed high bilirubin and LDH. Immunohematological work up revelaed polyspecific DAT was negative but monospecific DAT screening showed strong (4+) positivity for IgA and a weak IgG positivity. The patient was diagnosed as IgA-mediated Warm AIHA and was started on prednisolone at 2 mg/kg/day following which hemoglobin improved over the next 2 months. After 2 weeks, prednisolone was tapered and stopped by the end of 3 months. Patients with clinical and laboratory evidence of acute hemolysis, an additional screening for IgA antibody may be done even in cases where poly-specific DAT is negative. Early detection helps in avoiding further investigations and provide efficient management.

Cryptosporidiosis, whether it is more prevalent in Southern India.

Cryptosporidium species is identified as an important cause of morbidity in children and adults. Prevalence rate as reported from various studies in India is much lower when compared to developed word. Present study was designed to analyze the prevalence of cryptosporidiosis in West Godavari District, Andhra Pradesh. Stool samples from 306 patients suffering with gastrointestinal symptoms were examined for the presence of Cryptosporidium oocysts. A higher prevalence (25%) was found when compared to reports from north east and western India. Cryptosporidium was prevalent in 35-36% of adults, 17% of children and 20% of infants.

Macrodystrophia lipomatosa: a case report.

Macrodystrophia lipomatosa is a rare congenital form of localized gigantism, characterized by an increase in all mesenchymal elements, particularly fibroadipose tissue. The areas of predilection are segments supplied by the median or plantar nerves. We report such a rare case in a thirty four year old male patient, who presented with a swelling of the right upper limb and a marked increase in the size of the right thumb, index finger and radial half of the right hand, present since birth, with progressive increase to the present size. Amputation of the right thumb and debulking of the palm was done. The specimen measured 26 x 18 x 12 cms. and the thumb alone measured 13 x 7 x 7 cm. histology revealed hypertrophy of adipose tissue in the subcutaneous compartment and infiltration into the nerve sheaths and muscles.