The heterogeneity of members of the Streptococcus anginosus group (SAG) has traditionally hampered their correct identification. Recently, the group was subdivided into 6 taxa whose prevalence among human infections is poorly described. We evaluated the accuracy of the Rapid ID32 Strep test, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS), and a PCR multiplex method to identify 212 SAG isolates recovered from human infections to the species and subspecies level by using multilocus sequence analysis (MLSA) as the gold standard. We also determined the antimicrobial susceptibilities of the isolates. Representatives of all SAG taxa were found among our collection. MALDI-TOF MS and the Rapid ID32 Strep test correctly identified 92% and 68% of the isolates to the species level, respectively, but showed poor performance at the subspecies level, and the latter was responsible for major identification errors. The multiplex PCR method results were in complete agreement with the MLSA identifications but failed to distinguish the subspecies Streptococcus constellatus subsp. pharyngis and S. constellatus subsp. viborgensis. A total of 145 MLSA sequence types were present in our collection, indicating that within each taxon a number of different lineages are capable of causing infection. Significant antibiotic resistance was observed only to tetracycline, erythromycin, and clindamycin and was present in most taxa. MALDI-TOF MS is a reliable method for routine SAG species identification, while the need for identification to the subspecies level is not clearly established. Initially included in the viridans group streptococci, the species now recognized as the Streptococcus anginosus group (SAG) were first described as Streptococcus milleri in 1956 (1), but their taxonomic classification has remained in flux. Currently, the group includes three species, Streptococcus anginosus, Streptococcus constellatus, and Streptococcus intermedius (2-5) with unique genome signature regions (5). The species S. constellatus was subdivided into the subspecies S. constellatus subsp. pharyngis and S. constellatus subsp. constellatus (6). Recently, Jensen et al., based on multilocus sequence analysis (MLSA), divided S. anginosus into two subspecies-S. anginosus subsp. anginosus (the isolates formerly classified as S. anginosus) and S. anginosus subsp. whileyi-and they described the novel subspecies S. constellatus subsp. viborgensis (7).Most SAG strains share common features, such as small colony size, low growth rate, and a distinctive caramel smell (8, 9). However, SAG also present diversity in their other characteristics. These organisms may be nonhemolytic or display beta-or alphahemolysis when grown on blood agar plates (4, 5, 10), and they may present any of the A, C, F, or G Lancefield group antigens or lack Lancefield group antigen altogether (4,5,8,10). SAG, as with other members of the viridans group streptococci, are part of the human microbiota, colonizing the oral cavity, nasopharynx, and g...

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Autographa californica nucleopolyhedrovirus is a very productive expression vector for recombinant proteins in insect cells. Most vectors are based on the polyhedrin gene promoter, which comprises a TAAG transcription initiation motif flanked by 20 base pairs upstream and 47 base pairs downstream before the native ATG. Many transfer vectors also include a short sequence downstream of the ATG, in which case this sequence is mutated to ATT to abolish translation. However, the ATT sequence, or AUU in the mRNA, is known to be leaky. If a target-coding region is placed in the frame with the AUU, then some products will comprise a chimeric molecule with part of the polyhedrin protein. In this study, we showed that if AUU is placed in the frame with a Strep tag and eGFP coding region, we could identify a protein product with both sequences present. Further work examined if alternative codons in lieu of AUG might reduce translation initiation further. We found that AUA was used slightly more efficiently than AUU, whereas AUC was the least efficient at initiating translation. The use of this latter codon suggested that there might also be a slight improvement of protein yield if this is incorporated into expression vectors.