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Interestingly, the MUMi distances between W1481 with any strain of known subspecies were all above the defined cut-off, supporting our view that W1481 might be a novel subspecies. W1481 was distantly separated from all known subspecies compared to the distances between any pair of known subspecies ( fig. 3A ). Supplementary table S1 , Supplementary Material online, shows the full sets of MUMi values generated by comparing all 36 strains of Fusobacterium species.

The MUMi results were further supported by the ANI and AAI analysis (using the same set of genomes with W1481 as the reference genome) which also showed that the closest strain to W1481, belonged to F. nucleatum subsp. animalis as shown in supplementary figure S1 , Supplementary Material online.

To further investigate the genomic structure and gene content of F. nucleatum W1481, we performed pan-genome analysis using the protein sequences of 21 F. nucleatum strains. All the genome sequences of Fusobacterium used in the study were also annotated using the RAST pipeline to ensure uniformity. The genome size of the strains analyzed ranged from 1.71 to 3.68 Mb with GC composition varying between 25.2% and 35.2%. An overview of all the 21 Fusobacterium genomes used in the present study is shown in table 1 . The results showed that F. nucleatum pan-genome contained 6,666 gene clusters which were composed of 880 (13%) core gene clusters, 2,850 (43%) dispensable gene clusters and 2,936 (44%) strain-specific gene clusters. The accessory genes (dispensable and strain-specific genes) which cover 87% of the total F. nucleatum pan-genome suggested that this species contains a very diverse genomic structure.

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INTENDED USE

Hardy Diagnostics CatScreen™ is a rapid test for the detection of the enzyme butyrate esterase in bacteria isolated on culture media for the presumptive identification of Moraxella ( Branhamella ) catarrhalis .

The CatScreen™ is used to confirm the identification of Moraxella ( Branhamella ) catarrhalis when used in conjunction with chromogenic substrate tests or carbohydrate fermentations, oxidase reaction, Gram stain and morphology. For presumptive identification of M. catarrhalis , the CatScreen™ may be used along with the oxidase reaction, gram stain and colony morphology. (5)

The value of tributyrin hydrolysis for differentiating M. catarrhalis from Neisseria spp. was first reported by Berger in 1962. Subsequently, several authors have reported the usefulness of butyrate esterase in differentiating the two organisms using various substrates. (5) Butyrate hydrolysis is listed in standard reference texts as a key test in differentiating M. catarrhalis from Neisseria spp. (1,4,6)

Disks impregnated with bromo-chloro-indolyl butyrate serve as the substrate for the detection of butyrate esterase. Hydrolysis of the substrate by the butyrate esterase yields a chromogenic compound which appears blue to blue-green in color.

The CatScreen™ Kit contains 25 disks impregnated with 0.35% bromo-chloro-indolyl butyrate in a volatile organic solvent, followed by desiccation.

Upon receipt store at 2-8ºC, in the dark. This product should not be used if there are any signs of deterioration, contamination, or if the expiration date has passed. Protect disks from light and moisture. Leave desiccant in vial.

The expiration dating on the product label applies to the product in its intact packaging when stored as directed. The product may be used and tested up to the expiration date on the product label and incubated for the recommended quality control incubation times.

Refer to the document " Storage " on the Hardy Diagnostics Technical Document website for more information.

This product may contain components of animal origin. Certified knowledge of the origin and/or sanitary state of the animals does not guarantee the absence of transmissible pathogenic agents. Therefore, it is recommended that these products be treated as potentially infectious, and handle observing the usual universal blood precautions. Do not ingest, inhale, or allow to come into contact with skin.

This product is for in vitro diagnostic use only. It is to be used only by adequately trained and qualified laboratory personnel. Observe approved biohazard precautions and aseptic techniques. All laboratory specimens should be considered infectious and handled according to "standard precautions." The "Guidelines for Isolation Precautions" is available from the Centers for Disease Control and Prevention at www.cdc.gov/ncidod/dhqp/gl_isolation.html .

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