Rapid Diagnosis of Pandemic (H1N1) 2009 in Cuba

To the Editor: During 2005–2008, the Cuban National Influenza Center (NIC) at the Pedro Kouri Institute in Havana, Cuba, implemented a protocol for influenza surveillance proposed by the Pan American Health Organization and the US Centers for Disease Control and Prevention (1). One of the most essential features of this protocol was strengthening laboratory capacity for surveillance of seasonal influenza and timely detection of a new influenza virus with pandemic potential.

was designed to enable virus-specifi c identifi cation. Confi rmation was performed by subsequent sequencing of hemagglutinion, nucleoprotein, or neuraminidase genes by using the BigDye Terminator Cycle Sequencing Quick Start Kit (Beckman Coulter Inc., Krefeld, Germany).
The second phase of the algorithm was used during September 2009-August 2010. This phase included automated nucleic acid extraction (QIAcube; QIAGEN) and real-time RT-PCR kits (Pan American Health Organization, Washington, DC, USA, and Centers for Disease Control and Prevention, Atlanta, GA, USA) for detection and characterization of pandemic (H1N1) 2009 virus (10).
During January 3-July 31, 2009, a total of 2,156 specimens were submitted to the NIC for infl uenza surveillance. During January 2009-August 10, 2010, a total of 14,692 clinical samples were processed. In the next 6 months, the number of specimens submitted to NIC doubled. More specimens (7,978) were submitted during January 1-August 10, 2010, than during all of 2009. Most (5,601) clinical specimens processed were from patients with infl uenza-like illness. The highest percentage (45.9%) of infl uenzapositive samples was detected in specimens from these patients, followed by specimens from patients during outbreaks (18.0%).
Pandemic ( In this context, we believe that implementing national diagnostic algorithm enabled timely identifi cation of the novel virus and effective monitoring of its circulation, even before international diagnostic protocols and reagents were available in Cuba. This study shows the need for nucleic acid amplifi cation tools in laboratory diagnosis and surveillance of infl uenza viruses. As we prepare for future infl uenza pandemics, new and appropriate diagnostic methods and periodic assessment of infl uenza surveillance methods are needed as new information becomes available.

Acknowledgments
We thank the physicians and laboratory personnel for providing samples and clinical data; the technicians and researchers of the Virology Department and Microbiology  According to a Japanese Infectious Agents Surveillance Report, this virus is one of the major causes of herpangina, an acute febrile disease characterized by vesicles, ulcers, and redness around the uvula, which occurs mainly in young children and infants. (1).
In June 2011, a sudden increase in cases of hand, foot, and mouth disease (HFMD) at pediatric sentinel sites (≈3,000 pediatric hospitals and clinics) was reported to the National Epidemiologic Surveillance of Infectious Diseases System in Japan. Compared with past numbers of cases over 30 years of surveillance, the number of cases of HFMD per sentinel site peaked in week 28 (July) of 2011 (10.97 cases per sentinel), particularly in western Japan (2). According to the Infectious Agents Surveillance Report (as of September, 18, 2011), CVA6 was detected in 709 HFMD cases and 156 herpangina cases throughout Japan (1).
Clinical samples (throat swab specimens and feces) obtained from sentinel sites in Shimane, Hyogo, Hiroshima, and Shizuoka, Japan, were screened for enteroviruses by using an enterovirus-specifi c reverse transcription PCR and sequence analysis of the partial viral protein (VP)4/VP2 or VP1 region (3). Among 93 clinical samples from 108 HFMD case-patients, we identifi ed 74 casepatients as CVA6 positive by sequence analysis.
On the basis of sequence analysis of the entire VP1 region (GenBank accession nos. AB649286-AB649291), the consensus sequence