OSELTAMIVIR RESISTANT 2009 INFLUENZA A (H1N1) VIRUS INFECTION IN AN INMUNOCOMPROMISED HIV POSITIVE PATIENT FROM ARGENTINA

BOUZAS, M.B.; MAMMANA, L.; ZAPIOLA, I.; FERNANDEZ GIULIANO, S.; TORRES, C.; CULASSO, A.; CAMPOS, R.H.; NENNA, A .; CUNTO, E.; NOGUERAS, M.; SAN JUAN, J .

Daytona Beach, Florida

Simposio; 26th Clinical Virology Symposium; 2010

Background: 2009 Influenza A (H1N1) virus infection cause illness and death among persons worldwide. Oseltamivir is one of the two anti- neuraminidase drugs, zanavivir the other one , that is currently widely used against influenza.The most common mutation associated with resistance has been the H274Y. Immunosupressed patients with influenza virus infection can shed virus for prolonged periods, increasing the chances for development of drug resistance. Selected specimens collected during the winter season from hospitalized patients were studied by sequencing analysis for oseltamivir resistance. Patients and Methods: Theirteen specimens (nasopharingeal swabs and/ or endothracheal aspirates) from 12 patients who were asisted at intensive care unit were selected for the present study. 2/12 patients were HIV positive, 2 were receiving outpatient chemotherapy, 1 was pregnant and 1 was diabetic. Patients were aged 23-66 years old and 8 of them died. Clinical data in all cases were recorded. Samples undergo nucleic acid extaction using the High Purthe Viral Nucleic Acid Kit (Roche), aliquots were prepared and kept at -70ºC. All specimens tested positive for 2009 H1N1 virus using Real Time ready Swine Inf A/H1N1 which detects 2 genes M2, swH1 (Light Cycler 2.0, Roche). External and internal primers were designed based on the segment six of the NA gene, strain A/California/04/2009(H1N1) access Nº FJ966084, targeting the nucleotide position: 334 to 1325 and 525 -1084.The target region was amplified by RT ?PCR and a fragment of 561 bp was sequenced. Philogenetic analysis was performed with Mega 4 program using Neighbor Joining and Maximum Composite likelihood model. Branch support was assesed by boostrap analysis. Results: Sequences were obtained from all 13 samples, and the H274Y mutation was detected in 2 , both of them belong to one of the HIV positive patients included in the study. All sequences clustered with 2009H1N1 pandemic sequences (boostrap 99). Our patient was hospitalized on june 19 with an acute pneumonia, HIV was diagnosed and he was treated as PCP. On june 24 he started with abdominal pain, diarrhea and Clostridium dificile was isolated . On july 3 the patient was febrile with dysnea a nasopharyngeal swab was collected and oseltamivir was installed. On july 11 the patient was moved to intensive care unit and additional swabs were collected on july 12 and 16 before the patient died. All 3 specimens collected from this patient were confirmed as 2009 H1N1 virus, sequences carrying the H274Y were those dated on july 12 and 16, sample collected on july 3, before the initiation of oseltamivir treatment, is under study. Conclusions: In this selected group of patients we found an oselltamivir ?resistant 2009 influenza A (H1N1) virus in an HIV positive patient. Analysis of the the first sample collected before the initiation of oseltamivir treatment would allow as to determine whether the patient was infected with /or the resistance emerged during the treatment. From Argentina only one sequence carrying the H274Y mutation has been reported in a patient of one year old not epidemiologically linked with our patient.