BackgroundNeuraminidase (NA) inhibitors (NAIs) are currently the only antivirals effective against influenza infections due to widespread resistance to M2 inhibitors.MethodsInfluenza A and B viruses (n = 1079) collected worldwide between April 01, 2011, and September 30, 2011, were assessed for susceptibility to FDA‐approved NAIs, oseltamivir and zanamivir, and investigational peramivir, using the fluorescent‐based NA‐Fluor™ Influenza Neuraminidase Assay Kit. A subset of viruses (n = 98) were tested for susceptibility to the investigational NAI, laninamivir.ResultsInfluenza A(H1N1)pdm09 viruses (n = 326) were sensitive to all NAIs, except for two (0·6%) with H275Y (N1 numbering; H274Y in N2 numbering) substitution, which exhibited elevated IC 50s for oseltamivir and peramivir, and a third with previously unreported N325K substitution, exhibiting reduced susceptibility to oseltamivir. Influenza A(H3N2) viruses (n = 407) were sensitive to all NAIs. Influenza B viruses (n = 346) were sensitive to all NAIs, except two (0·6%) with H273Y (N1 numbering; H274Y in N2 numbering) substitution, exhibiting reduced susceptibility to oseltamivir and peramivir, and one with previously unreported G140R and N144K substitutions, exhibiting reduced susceptibility to oseltamivir, zanamivir, and peramivir. All influenza A and B viruses were sensitive to laninamivir. It is unknown whether substitutions N325K, G140R, and N144K were present in the virus prior to culturing because clinical specimens were unavailable for testing.ConclusionsThis study summarizes NAI susceptibility of influenza viruses circulating worldwide during the 2011 Southern Hemisphere (SH) season, assessed using the NA‐Fluor™ Kit. Despite low resistance to NAIs among tested influenza viruses, constant surveillance of influenza virus susceptibility to NAIs should be emphasized.

c Whole-genome sequencing of Cryptosporidium spp. is hampered by difficulties in obtaining sufficient, highly pure genomic DNA from clinical specimens. In this study, we developed procedures for the isolation and enrichment of Cryptosporidium genomic DNA from fecal specimens and verification of DNA purity for whole-genome sequencing. The isolation and enrichment of genomic DNA were achieved by a combination of three oocyst purification steps and whole-genome amplification (WGA) of DNA from purified oocysts. Quantitative PCR (qPCR) analysis of WGA products was used as an initial quality assessment of amplified genomic DNA. The purity of WGA products was assessed by Sanger sequencing of cloned products. Next-generation sequencing tools were used in final evaluations of genome coverage and of the extent of contamination. Altogether, 24 fecal specimens of Cryptosporidium parvum, C. hominis, C. andersoni, C. ubiquitum, C. tyzzeri, and Cryptosporidium chipmunk genotype I were processed with the procedures. As expected, WGA products with low (<16.0) threshold cycle (C T ) values yielded mostly Cryptosporidium sequences in Sanger sequencing. The cloning-sequencing analysis, however, showed significant contamination in 5 WGA products (proportion of positive colonies derived from Cryptosporidium genomic DNA, <25%). Following this strategy, 20 WGA products from six Cryptosporidium species or genotypes with low (mostly <14.0) C T values were submitted to whole-genome sequencing, generating sequence data covering 94.5% to 99.7% of Cryptosporidium genomes, with mostly minor contamination from bacterial, fungal, and host DNA. These results suggest that the described strategy can be used effectively for the isolation and enrichment of Cryptosporidium DNA from fecal specimens for whole-genome sequencing. C ryptosporidium spp. are an important cause of moderate to severe diarrhea in humans and various animals (1, 2). Over the past decade, great efforts have been made to understand the interaction between Cryptosporidium spp. and their hosts. Thus far, at least 26 species and more than 60 genotypes have been described (3), most with some host specificity. In contrast to the expanding knowledge of the taxonomic complexity of the genus Cryptosporidium, little progress has been made in understanding the molecular basis of phenotypic traits such as the host specificity. This is mainly due to the lack of whole-genome characterization of most Cryptosporidium species and genotypes.Thus far, the genomes of only four Cryptosporidium isolates have been sequenced. Sequence data of the whole genomes of C. parvum and C. hominis, the two species responsible for most human infections (4), were first published in 2004 (5, 6). Several years later, the genome of C. muris was sequenced, and data from the project are available on CryptoDB (http://cryptodb.org). More recently, the genome of anthroponotic subtype family IIc of C. parvum has been sequenced using the next-generation sequencing (NGS) tools (7). The availability of the whole-genome sequenc...