Prateek Bhatia

Visceral leishmaniasis associated hemophagocytic lymphohistiocytosis – Case report and systematic review

BACKGROUND The clinical features of leishmaniasis overlap with that of hemophagocytic lymphohistiocytosis (HLH) and the diagnosis of visceral leishmaniasis (VL) related HLH can be challenging. OBJECTIVES To review information available on disease course, treatment, adjunctive therapy used and the outcomes of VL related HLH. METHODS We describe an illustrative case and review all reported cases of VL associated HLH in the English literature till March 2007. RESULTS VL associated HLH is rare, with 56 cases reported in the English literature. Clinical features lack discriminating value to recognize VL as the inciting etiology. Bone marrow aspiration (BMA) establishes the diagnosis in 78% of cases but is often negative at onset of the syndrome due to the pauci-microbial nature of the disease and patchy involvement. Repeated marrow aspiration, liver biopsy, blood cultures and serology may be required to establish the diagnosis. Liposomal amphotericin is the drug of choice. IVIG may be considered when there is an inadequate response to anti-leishmanial therapy in severe and refractory disease. CONCLUSIONS VL related HLH is often under-recognized because of overlapping clinical features and negative marrow evaluation at onset, leading to high mortality rates.

The deferiprone and deferasirox combination is efficacious in iron overloaded patients with β-thalassemia major: A prospective, single center, open-label study

The high cost, coupled with the need for continuous infusion, renders Desferrioxamine (DFO), a non‐feasible option for iron‐chelation in a large majority of patients with β‐thalassemia major in developing countries. Monotherapy with deferiprone (DFP) or deferasirox (DFX) may not always attain optimal control, particularly in heavily iron‐loaded patients. Combination of DFP and DFX is a potential alternative.

Neonatal cytomegalovirus infection: Diagnostic modalities available for early disease detection

CMV is a ubiquitous virus. In India, there is high seroendemicity with almost 99% adults showing IgG antibodies. Infection or re-activation becomes important in immunocompromised host (Transplant recipients, Cancer therapy patients and patients with HIV/AIDS). Neonates form a distinctive high risk population for congenital CMV infection and suffer disastrous sequlae of the same. Neonatal infections may be congenital in nature or may be acquired after birth during first month of life via infected breast milk or due to exposure to high risk blood products. The risk for transmission of the virus to the fetus is higher in primary infected mothers than in mothers with reactivated disease. Primary CMV infections are reported in 1–4% of seronegative women during pregnancy and the risk for viral transmission to fetus is 30–40%. Reactivation of a CMV infection during pregnancy is reported in 10–30% of seropositive women and the risk of transmitting the virus is about 1–3%. The adverse outcome of congenital neonatal CMV infection includes-microcephaly (70%), intellectual impairment (60%), sensineural hearing loss (35%), choriorenitis (22%), hepatosplenomegaly (70%), jaundice (68%), thrombocytopenia (65%), low birth weight (65%), pneumonitis (2–5%) and congenital heart disease (<5%). About 5–10% of congenitally infected asymptomatic infants will have neurological problems later in life the most common of which is unilateral or bilateral sensory neural hearing loss.All immunocompromised hosts, including pre-term neonates, mount weak antibody responses (IgM), making serological detection of CMV infection in them, fallacious. Thus, it is imperative to use antigen detection methods such as quantitative PCR or PP65 Antigenaemia assays to detect CMV infection in immunocompromised host. Sakhuja et al and Minz et al have demonstrated that PP65 Antigenaemia assay is very good for diagnosing CMV disease in renal transplant recipients. The present review tends to highlight the role of newer diagnostic modalities in early CMV infection detection in neonatal population.

5-Year Review and Reappraisal of Ultrasound-Guided Percutaneous Transabdominal Fine Needle Aspiration of Pancreatic Lesions

OBJECTIVE To reevaluate the efficacy and safety offine needle aspiration cytology (FNAC) of pancreatic lesions performed by transabdominal approach. STUDY DESIGN Retrospective 5-year (2001-2006) audit of all pancreatic FNA samples. RESULTS This series includes 267 patients (88 men, 179 women). Seven cases (2.6%) yielded insufficient material for diagnosis; 260 cases were classified as benign (n=118) and malignant (n=142) lesions. Of the 118 benign aspirates, the cytodiagnosis was acute/chronic inflammation in 24, tuberculosis in 1, benign cyst in 10 and a benign aspirate, not otherwise specified, in the remaining 83 cases. Of the 142 malignant aspirates, the cytodiagnosis was adenocarcinoma in 126, neuroendocrine/carcinoid tumor in 7, papillary solid epithelial neoplasm in 2, mucinous cystadenocarcinoma in 2, acinar cell carcinoma in 1 and metastatic small cell carcinoma in lung in 4 cases. Cytohistologic correlation yielded a sensitivity of 81% and specificity of 100%. CONCLUSION A spectrum of pancreatic lesions can be accurately diagnosed by the technique. The false negative rate can be minimized by proper positioning of the needle under guidance and adequate sampling. No postprocedural complications were encountered, proving that this procedure is safe if carried out by an experienced team in a hospital setting.

Cell Blocks from Scraping of Cytology Smear

OBJECTIVE To compare cytomorphology preservation and immunohistochemistry results between conventional cell blocks (CCB) and cytoscrape cell blocks (SCB). STUDY DESIGN Fine needle aspiration (FNAC) was done in 17 consecutive cases. Air-dried smears for May-Grunwald-Giemsa stain and wet-fixed smear for hematoxylin-eosin (H-E) stain were prepared. Simultaneously another pass was made in each case for preparation of material for CCB. One of the H-E-stained smears was spared for SCB. SCB was compared with CCB for cell morphology. Immunostaining was performed both cell blocks, as well as on FNA smears in 8 cases. Results were evaluated for intensity of staining and percentage of cells showing positivity. RESULTS CCB and SCB sections showed adequate cellularity in all cases. Morphologic preservation was good in SCB sections. There was good architectural and nuclear preservation in all cases of SCB. Immunostaining results showed better and clear intensity of staining with little background in all cell block cases. CONCLUSION SCB is a valuable technique in cell blocks from stained FNA smears. The cytomorphologic details are equally good in SCB and CCB. Additional panels of immunostaining can be done on SCB for better diagnosis and classification, particularly in cases in which repeat FNA is not possible.

The 5-Year EFS of Multisystem LCH With Risk-Organ Involvement Is Suboptimal: A Single-center Experience From India.

The study describes an 8-year experience of a single center in managing patients with langerhans cell histiocytosis on the basis of the langerhans cell histiocytosis-III platform. A retrospective case-file review of children diagnosed during 2006 to 2013 was performed. Group 1 (multisystem with risk-organ involvement) patients received an initial treatment of 6 to 12 weeks, followed by continuation treatment to complete 12 months. Drugs included vinblastine, prednisolone, and 6-mercaptopurine. Group 2 (multisystem without risk-organ involvement) patients received a similar treatment, except for 6-mercaptopurine. Group 3 (single-system/multifocal bone disease) patients were treated for a duration of 6 months. Forty-nine patients were treated: 24 (49%), 14 (28.6%), and 11 (22.4%) in groups 1, 2, and 3, respectively. The mean age at diagnosis was 31.6±28.4 months (range, 4 to 120 mo). Five patients abandoned treatment. There were 7 deaths, all in group 1. All patients who died had either a partial response or progressive disease after induction (P=0.000). Among patients with liver involvement, those with sclerosing cholangitis had a greater mortality (P=0.007). A relapse was observed in 12 (24.5%) patients. The frequency of relapse was not different in the 3 groups (P=0.833). The 5-year event-free survival in groups 1, 2, and 3 was 29.3±10%, 58.9±14.6%, and 69.3±15%, respectively (P=0.019). The 5-year overall survival was 100% in groups 2/3 and 68.9±9.8% in group 1 (P=0.011).

Emerging role of NUDT15 polymorphisms in 6-mercaptopurine metabolism and dose related toxicity in acute lymphoblastic leukaemia

Despite more than 80% long term survival in ALL, morbidity due to drug related myelotoxicity remains high. Germline variants of thiopurine metabolizing enzymes (TPMT and ITPA) have been described which are associated with increased drug toxicity during maintenance phase, but their prevalence in different ethnic groups is variable to account for relatively high myelotoxicity incidence. NUDT15 variant (rs116855232) has been recently identified as a novel polymorphism related with thiopurine-induced leucopenia in inflammatory bowel disease and ALL. Current review highlights the scientific data on NUDT15 enzyme variant and its relation to 6-MP toxicity in various ethnic populations.

Systematic evaluation of paediatric cohort with iron refractory iron deficiency anaemia (IRIDA) phenotype reveals multiple TMPRSS6 gene variations

Systematic screening identified patients with an iron refractory iron deficiency anaemia (IRIDA) phenotype and genotype in iron‐deficient children in the Indian subcontinent. Cases of moderate to severe microcytosis and anaemia with no obvious cause and normal C‐reactive protein, HbA2 and tissue transglutaminase antibody levels (n = 550) were put on a trial of oral iron for 4 weeks. Sixty of these 550 cases (11%) were variably refractory to oral iron therapy (<10 g/l Hb rise) at 4–6 weeks and were subsequently evaluated for plasma iron, ferritin and hepcidin levels. The mean age of this cohort was 2.06 years. Low‐normal to normal ferritin and normal to high hepcidin levels were noted in 25/60 (41.6%) and 47/60 (78.3%), respectively. An IRIDA phenotype was noted in 38.3% (23/60) based on standard criteria. TMPRSS6 gene sequencing in 20 cases with IRIDA phenotype revealed 9 potentially deleterious intronic and two benign exonic variations in 12/20 cases (60%). Of these, 4 intronic and both exonic variations were noted in multiple cases and are likely to act synergistically leading to an IRIDA phenotype. However, given that only 38% (23/60 cases) of cases with iron refractoriness had IRIDA phenotype, a balanced approach is needed and other causes for refractoriness should be investigated before genetic studies for TMPRSS6 are undertaken.

DNA Ploidy and S-phase Fraction Analysis in Paediatric B-cell Acute Lymphoblastic Leukemia Cases: a Tertiary Care Centre Experience

DNA ploidy is an important prognostic parameter in paediatric B-ALL, but the significance of the S-phase fraction is unclear. In present study, DNA ploidy was assessed in 40 pediatric B-ALL cases by flow cytometry. The DI (DNA index) and percentage of cells in S-phase were calculated using Modfit software. Aneuploidy was noted in 26/40 (65%) cases. A DI of 1.10-1.6 (hyperdiploidy B) was noted in 20/40 (50%) and 6/40 (15%) had a DI>1.60 (triploid and tetraploid range). Some 14/40 (35%) cases had a diploid DI between 0.90-1.05. None of the cases had a DI <0.90 (hypodiploid) or in the 1.06-1.09 (hyperdiploid A) range. The mean S-phase fraction was 2.6%, with 24/40 (60%) having low and 16/40 (40%) high S-phase fractions. No correlation was noted with standard ALL risk and treatment response factors with DI values or S-phase data, except for a positive correlation of low S-phase with high NCI risk category (p=0.032). Overall frequency of hyperdiploidy in our cohort of B-ALL patients was very high (65%). No correlation between hyperdiploidy B and low TLC or common B-phenotype was observed in our study as 42% cases with DI 1.10-1.6 had TLC> 50 x 109 and 57.1% CD 10 negativity. The study also highlighted that S-phase fraction analysis does not add any prognostic information and is not a useful parameter for assessment in ALL cases. However, larger studies with long term outcome analysis are needed to derive definitive conclusions.

The Accelerated Phase of Chediak-Higashi Syndrome: The Importance of Hematological Evaluation

To the Editor Chediak-Higashi syndrome is a rare autosomal recessive disease that was first described in 1943 by Bequez-Cesar in 3 siblings that bore the primary clinical features. In 1952 Chediak (a Cuban hematologist) and in 1954 Higashi (a Japanese pediatrician) described a series of cases characterized by misdistribution of myeloperoxidase in the patients’ neutrophilic granules [1,2]. Mean age of onset is 5.85 years; however, most patients die before age 10 years. In patients that do survive beyond childhood the neurologic problems persist and/or increase in magnitude [3]. Most cases are diagnosed clinically based on partial albinism and recurrent pyogenic infections. Both of our patients had hypopigmentation of the skin with patchy grey hairs, mild coagulation defects identified via the presence of petechial rashes on the skin, and a history of recurrent infections since birth. The characteristic hematological finding is massive lysosomal inclusions in all white cells, formed via a combined process of fusion, cytoplasmic injury, and phagocytosis due to a microtubular defect [3]. These granules exhibit both azurophilic and specific granular markers (Figure 1), and are strongly myeloperoxidase positive. Approximately 85% of cases develop a fatal accelerated phase characterized by pancytopenia, hemophagocytosis, and marked infiltration of organs by lymphocytes, leading to multi-organ dysfunction [4]. Herein we present 2 cases in the accelerated phase of Chediak-Higashi syndrome. Both were born to apparently healthy non-consanguineous parents. Figure 1 Large intracytoplasmic granules in a granulocytic cell (Giemsa stained bone marrow aspirate film; 1000×). Case 1 was a 2.5-year-old female that presented with a 6-month history of abdominal distension and moderate pallor, and a 5-d history of cough. General physical examination showed that she was thin built, and had normal facies with patchy grey hair and hypopigmentation of the skin. Systemic examination showed hepatosplenomegaly; the liver was 6 cm below the right costal margin and the spleen was 13 cm below the left costal margin. The cardiovascular and respiratory systems were normal. She also had bilateral cervical lymphadenopathy. Case 2 was a 3-year-old female that presented with a 3-month history of fever, and a 1-month history of abdominal distension, itching over the body, and moderate pallor. Physical examination showed patchy gray hair and generalized albinism with petechial spots on the thighs and abdomen. The patient weighed 7.6 kg, was of average build with normal facies and significant hepatosplenomegaly; the liver was 4 cm below the right costal margin and the spleen was 7 cm below the left costal margin. Both patients underwent bone marrow aspiration due to peripheral blood pancytopenia and organomegaly. Hemogram findings in cases 1 and 2 were, respectively, as follows: Hb: 4.2 g/dL and 3.9 g/dL; WBC: 4 × 109/L and 3 × 109/L; platelet count: 8 × 109/L and 6 × 109/L. Bone marrow aspiration findings in both cases are shown in the Table 1. Both patients had hematological findings characteristic of the accelerated phase of Chediak-Higashi syndrome-peripheral blood pancytopenia and marked hemophagocytosis (Figure 2) on bone marrow aspirate, along with lymphocyte infiltration on trephine biopsy section. Both cases were negative for HIV, CMV, and EBV serology. Table 1 Bone marrow aspirate findings in cases 1 and 2. Figure 2 Hemophagocytosis in bone marrow smear (Giemsa stain; 1000×). The genetic hallmark of Chediak-Higashi syndrome is mutations in the CHS1/LYST gene. The gene product is known to regulate lysosomal organelle function and size. The mutations could not be characterized in the presented cases because the DNA material for analysis was insufficient in one case and the other died due to the fulminant accelerated phase before hematological diagnosis was established. It is well known that Chediak-Higashi syndrome patients with deletions in the LYST gene usually present with a fulminant accelerated phase early in life, whereas those with missense mutations have a relatively better prognosis characterized by the absence of an accelerated phase and no neurological involvement [5]. The hematological findings of pancytopenia and hemophagocytosis suggest a possible deletion in the LYST gene in both of the presented cases, but ethnicity data and complete DNA analysis are needed to further substantiate the findings; nonetheless, both cases presented with the characteristic clinical and hematological profiles diagnostic of Chediak-Higashi syndrome. Careful examination of peripheral blood film in suspected cases can facilitate early diagnosis and further evaluation. In addition, recognition of early cytopenias should alert clinicians and hematologists of an impending accelerated phase. Furthermore, bone marrow aspiration is a useful diagnostic procedure for identifying tissue hemophagocytosis in such patients. Conflict of Interest Statement The authors of this paper have no conflicts of interest, including specific financial interests, relationships, and/ or affiliations relevant to the subject matter or materials included.