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to the global rate of viral clearance (~25%). This is the first study on Egyptian viral strains that could help to elucidate some of the viral factors that may be responsible for viral clearance. This case report showed how a genotype variation in an important stem loop of the IRES can induce a direct phenotype change exemplified in the clearance of the virus from the patient. Such critical sequences may be used as therapeutic targets to abolish HCV replication using RNA silencing and/or antisense RNA. Further studies on patients who cleared the infection and analysis of the IRES region is required to assert the conclusion of this report, along with experimental verification of the changes in secondary structure of the IRES’s subdomains.
Gallego J, Varani G. The hepatitis C virus internal ribosome-entry site: A new target for antiviral research. Biochem Soc Trans 2002;30:140-5. Jubin R, Vantuno NE, Kieft JS, Murray MG, Doudna JA, Lau JY, et al. Hepatitis C virus internal ribosome entry site (IRES) stem loop IIId contains a phylogenetically conserved GGG triplet essential for translation and IRES folding. J Virol 2000;74:10430-7. Wang C, Sarnow P, Siddiqui A. Translation of human hepatitis C virus RNA in cultured cells is mediated by an internal ribosome-binding mechanism. J Virol 1993;67:3338-44. Thurner C, Witwer C, Hofacker IL, Stadler PF. Conserved RNA secondary structures in Flaviviridae genomes. J Gen Virol 2004;85:1113-24. Collier AJ, Gallego J, Klinck R, Cole PT, Harris SJ, Harrison GP, et al. A conserved RNA structure within the HCV IRES eIF3-binding site. Nat Struct Biol 2002;9:375-80. Odreman-Macchioli FE, Tisminetzky SG, Zotti M, Baralle FE, Buratti E. Influence of correct secondary and tertiary RNA folding on the binding of cellular factors to the HCV IRES. Nucleic Acids Res 2000;28:875-85. Gallego J, Varani G.The hepatitis C virus internal ribosome-entry site: A new target for antiviral research. Biochem Soc Trans 2002;30:140-5. El-Awady MK, Ismail SM, El-Sagheer M, Sabour YA, Amr KS, Zaki EA. Assay for hepatitis C virus in peripheral
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blood mononuclear cells enhances sensitivity of diagnosis and monitoring of HCV-associated hepatitis. Clin Chim Acta 1999;283:1-14. Sergi C, Arnold JC, Rau W, Otto HF, Hofmann WJ. Single nucleotide insertion in the 5’-untranslated region of hepatitis C virus with clearance of the viral RNA in a liver transplant recipient during acute hepatitis B virus superinfection. Liver 2002;22:79-82. Zhang J, Yamada O, Ito T, Akiyama M, Hashimoto Y, Yoshida H, et al. A single nucleotide insertion in the 5’-untranslated region of hepatitis C virus leads to enhanced cap-independent translation. Virology 1999;261:263-70. El Awady MK, Azzazy HM, Fahmy AM, Shawky SM, Badreldin NG, Yossef SS, et al. Positional effect of stem loop III mutations in HCV genotype 4a IRES on initial response of chronic patients to interferon 2a plus ribavirin combined therapy World J Gastroenterol 2009;15:1480-6. Lukavsky PJ, Otto GA, Lancaster AM, Sarnow P, Puglisi JD. Structures of two RNA domains essential for hepatitis C virus internal ribosome entry site function. Nat Struct Biol 2000;7:1105-10. Doshi KJ, Cannone JJ, Cobaugh CW, Gutell RR. Evaluation of the suitability of free-energy minimization using nearest-neighbor energy parameters for RNA secondary structure prediction. BMC Bioinformatics 2004;5:105.
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Website: www.ijmm.org PMID: *** DOI: 10.4103/0255-0857.148835
How to cite this article: Bader El Din NG, El Hefnawy MM, Omran MH, Dawood RM, El Abd Y, Ibrahim MK, El Awady MK. Spontaneous clearance of chronic hepatitis C infection is associated with an internal ribosomal entry site IIId stem loop structure variant. Indian J Med Microbiol 2015;33:S143-8. Source of Support: Nil, Conflict of Interest: None declared.
Fatal cryptococcosis involving multiple sites in an immunocompetent child H Kaur, K Zaman, BR Thapa, *SM Rudramurthy
Abstract Disseminated cryptococcosis is less common in immunocompetent individuals. Herein, we report a fatal case of cryptococcosis in apparently immunocompetent child with multiple site involvement. The yeast isolated from cerebrospinal fluid, blood, endotracheal, gastric and lymph node aspirate was identified by molecular method as Cryptococcus neoformans var. grubii. Key words: Cryptococcus neoformans var. grubii, disseminated cryptococcosis, immunocompetent child
*Corresponding author (email: ) Department of Medical Microbiology (HK, KZ, SMR), Department of Gastroenterology (BRT), Postgraduate Institute of Medical Education and Research, Chandigarh, India Received: 23-05-2014 Accepted: 08-07-2014
Introduction Cryptococcus is encapsulated yeast, which is present in environment, especially in soil contaminated with bird excreta. Predominantly, it affects the immunocompromised individuals, especially those infected by human immunodeficiency virus (HIV). The infections of pulmonary and central nervous system (CSI) are the most common among the infections caused by this agent. However, disseminated infection can occur in immunosuppressed patients. Such disseminated infection is rarely reported in immunocompetent children.[2-6] Herein, we present the first case from India, of an immunocompetent female child, who died due to disseminated infection by Cryptococcus neoformans var grubii. Case Report An 11-year-old female child, resident of Jammu and Kashmir presented with 2 months history of intermittent pain in epigastrium and right hypochondrium and intermittent fever documented up to 102°F associated with vomiting. She had jaundice since 1 month with dark coloured urine, loss of weight and appetite. She was admitted in local hospital for these symptoms but no improvement was seen. When she was presented to our hospital, she had headache in addition to recurrent vomiting since 2 days and had diminished response to stimuli for 1 day. She had raised intracranial pressure and had an episode of seizures with up rolling of eyes. She had no history of pruritis, bleeding from any site, focal neurological deficit or any change in behaviour or trauma, with no history of diabetes mellitus, tuberculosis, malignancy or chronic illness. Her family did not give any history of exposure to bird excreta. Clinical examination revealed icterus and pallor along with lymphadenopathy. Liver was enlarged with presence of distended veins over the abdomen. Kernig’s sign was positive and the fundus showed papilloedema. She had altered sensorium with Glasgow Coma Score (GCS) E2 M4 V2. Laboratory investigations showed haemoglobin of 7.7 gm, total leucocyte count (TLC) of 18,000 with predominant neutrophils. Other findings included hyperglycaemia, hypokalaemia, hypernatraemia and azotaemia. She was non reactive for HIV. Investigations for cell-mediated, humoural and phagocytic immunity did not show any evidence of immunodeficiency. Ultrasonography (USG) revealed multiple, enlarged lymph nodes at portal, peri-pancreatic regions and
mesentery. Lymph node fine needle aspiration cytology (FNAC) showed presence of yeast cells with capsule on histopathology. Magnetic resonance imaging (MRI) of brain showed multiple, small non-enhancing nodular lesions in bilateral cerebral hemispheres (parietal region, occipital region and putamen) suggestive of cryptococcomas. Contrast-enhanced computed tomography (CECT) head showed dilated third ventricles. Yeast cells with capsule were detected on direct microscopy of Indian ink preparation of cerebrospinal fluid (CSF). Cryptococcal antigen test by latex agglutination (CALAS®, Meridian Bioscience Inc.) was positive in CSF. Cryptococcus was isolated on Sabouraud dextrose agar and bird seed agar. The same fungus was detected from blood, gastric aspirate, and endotracheal aspirate culture. The isolate was urease positive and there was no colour change on canavanine glycine bromothymol blue (CGB) agar (25°C for 5 days). Antifungal susceptibility testing was performed as per Clinical Laboratories Standard Institute document for broth dilution technique (M 27 A3). The minimum inhibitory concentration (MIC) of isolate was: Amphotericin B-0.25 g/ml; fluconazole-1.0 g/ml; itraconazole-0.03g/ml;voriconazole-0.03g/ml;caspofungin-2.0 g/ml; anidulafungin-2.0 g/ml; micafungin-2.0 g/ml. The growth of Cryptococcus was used to extract the deoxyribose nucleic acid (DNA) and was amplified using ITS 1, 4 and 26S rRNA primers. It was then subjected to sequencing by Big Dye Terminator Cycle Sequencing Kit, Version 3.1 (Applied Biosystems, Foster city, CA, USA) and analysed by ABI 3130 Genetic Analyser (Applied Biosystems). The sequence obtained was compared with GenBank DNA database. The strain showed 100% similarity with standard strain of Cryptococcus neoformans var. grubii (CBS 8710, GenBank accession FJ534879). The isolate was deposited at the National Culture Collection of Pathogenic Fungi (NCCPF), Post Graduate Institute of Medical Education and Research (PGIMER), Chandigarh, India with accession NCCPF 250472. The nucleotide sequence is deposited in GenBank with accession KJ777814. Urine sample showed growth of Escherichia coli. Acid-fast stain for gastric aspirate, sputum and lymph node aspirate were negative. Initially, anti-tubercular therapy (ATT) was given for three days as the symptoms were quite suggestive of tuberculosis. However, after FNAC for lymph node demonstrated Cryptococcus, patient was started on liposomal amphotericin B (6 mg/kg/day) and 5 flucytosine (100 mg/kg/day in four divided doses). Two days later, liposomal formulation was replaced by conventional amphotericin B (1 mg/kg/day), which led to the development of hypokalaemia, for which she was given potassium supplements. She was given mannitol, 3% hypertonic saline, and glycerol for decreasing
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intracranial tension. She also developed hyperglycaemia (central diabetes insipidus) and was started on insulin and vasopressin therapy. Phenytoin (5 mg/kg/day) was given for epileptic seizure. She went into shock with persistent fever spikes and intermittent episodes of hyperventilation and was started on dopamine. She was also given antibiotics including vancomycin and meropenem. She progressed to ventricular tachycardia, multiorgan dysfunction syndrome, pulmonary haemorrhage and finally refractory septic shock, after which she could not be revived. Discussion Cryptococcus neoformans causes infection most commonly in immunocompromised patients including HIV positive, solid organ transplant and haematology malignancy. Disseminated infection is defined by a positive culture from at least two different sites or a positive blood culture. In the present case, positive cultures have been obtained from multiple sites (CSF, endotracheal aspirate, gastric aspirate, and lymph node aspirate) and from blood (cryptococcaemia) as well. Cirrhosis and liver disease have been found to be an important risk factors for disseminated infection.[8,9] In this case as well, liver involvement was present as shown by jaundice and dark-coloured urine. However, liver disease could not be investigated. The most commonly infected organ system in disseminated disease is central nervous system (CNS) followed by lung. This child had involvement of both CNS and lung. In this case, the presentation of the disease was insidious and chronic, as the child had complaints of vomiting and headache since two months. Though amphotericin B and 5 flucytosine were started immediately after diagnosis, it was too late as the infection had spread to multiple organs. Cryptococcosis in non-HIV patients has high mortality, possibly due to low suspicion of infection and delay in diagnosis. Cryptococcus neoformans var. neoformans and Cryptococcus neoformans var. grubii cause infection generally in immunocompromised individuals, whereas Cryptococcus gattii is responsible for disease in immunocompetent hosts. Disseminated infection in immunocompetent children in earlier reports was caused by Cryptococcus neoformans var. neoformans and Cryptococcus neoformans var. humicolus, both of which showed remission after treatment.[3,4] In the present case, the isolate was identified to be C. neoformans var. grubii, which has been rarely reported to cause disseminated infection in immunocompetent hosts, especially children. The present case highlights the importance of suspecting this aetiologic agent in immunocompetent children with liver failure
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presenting with complaints that are similar to this case. The major challenge in such cases is the differential diagnosis of tuberculosis, which produces similar symptoms. It is very important to initiate treatment after prompt diagnosis is made, so as to reduce mortality due to disseminated cryptococcosis in immunocompetent hosts, especially children. References 1.
Pappas PG, Perfect JR, Cloud GA, Larsen RA, Pankey GA, Lancaster DJ, et al. Cryptococcosis in human immunodeficiency virus-negative patients in the era of effective azole therapy. Clin Infect Dis 2001;33:690-9. 2. Suchitha S, Sheeladevi CS, Sunila R, Manjunath GV. Disseminated cryptococcosis in an immunocompetent patient: A case report. Case Rep Pathol 2012;2012:652351. 3. Chaudhary MW, Sardana K, Kumar P, Dewan V, Anand VK. Disseminated infection with Cryptococcus neoformans var neoformans in an 8 years immunocompetent girl. Indian J Pediatr 2005;72:85. 4. Godbole R, Nayar P, Pradhan A, Manchanda R. Disseminated cryptococcosis in an immunocompetent child. Indian J Hematol Blood Transfus 2007;23:122-4. 5. Tarai B, Gupta A, Ray P, Shivaprakash MR, Chakrabarti A. Polymerase chain reaction for early diagnosis of post-operative fungal endophthalmitis. Indian J Med Res 2006;123:671-8. 6. Jain BB, Bose D, Mondal R, Chattopadhyay S. Disseminated Cryptococcosis in an Immunocompetent Child. Turk Patoloji Derg 2014. 7. Yehia BR, Eberlein M, Sisson SD, Hager DN. Disseminated cryptococcosis with meningitis, peritonitis, and cryptococcemia in a HIV-negative patient with cirrhosis: A case report. Cases J 2009;2:170. 8. Chuang YM, Ho YC, Chang HT, Yu CJ, Yang PC, Hsueh PR. Disseminated cryptococcosis in HIV-uninfected patients. Eur J Clin Microbiol Infect Dis 2008;27:307-10. 9. Jean SS, Fang CT, Shau WY, Chen YC, Chang SC, Hsueh PR, et al. Cryptococcaemia: Clinical features and prognostic factors. QJM 2002;95:511-8. 10. Capoor MR, Nair D, Deb M, Gupta B, Aggarwal P. Clinical and mycological profile of cryptococcosis in a tertiary care hospital. Indian J Med Microbiol 2007;25:401-4. Access this article online Quick Response Code:
Website: www.ijmm.org PMID: *** DOI: 10.4103/0255-0857.150935
How to cite this article: Kaur H, Zaman K, Thapa BR, Rudramurthy SM. Fatal cryptococcosis involving multiple sites in an immunocompetent child. Indian J Med Microbiol 2015;33:S148-50. Source of Support: Nil, Conflict of Interest: None declared.
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