Rapid Serotyping of Salmonella Isolates Based on Single Nucleotide Polymorphism-Like Sequence Profiles of a Salmonella-Specific Gene.

Although serotyping is the most important method of identification of taxonomy in Salmonella, conventional serotype determination with a complete set of antisera is time consuming and laborious. Recently, rapid serotyping procedures with polymerase chain reaction (PCR) have been developed. In this study, we established a novel PCR-based rapid serotyping method that employs a unique target gene. Alignment study of Salmonella-specific gene (Salmonella enterotoxin [stn]) revealed a correlation between the stn gene sequence and the serotype of the organism. In 750 bp of stn gene, 55 nucleotides indicated single nucleotide polymorphism (SNP)-like polymorphism, and the correlation between the SNP-like polymorphism and the serotype of the organism suggests that SNP-like sequences in stn gene can serve as an index for serotyping. To develop a rapid serotyping method based on the SNP-like polymorphism, we selected serotype-associated 12 SNP-like sites in the stn gene and established a method based on high-resolution melting (HRM) and PCR, which identifies nucleotides at SNP-like sites within 1.5 h. This newly established rapid serotyping procedure (stn-HRM) could identify nine serotypes, including the frequently isolated serovar Enteritidis. These nine serotypes cover 64.3% of cases of Salmonella, as reported by the World Health Organization/Global Foodborne Infection Network (WHO/GFN) Country Databank from 2001 to 2010. In this study, we employed a unique target gene, stn, which is completely independent of the genes that were targeted in previously reported rapid serotyping procedures. Therefore, the results obtained by our newly developed stn-HRM procedure are independent of the results obtained by other procedures. Besides, stn-HRM can ensure accurate identification of the bacterial species as stn is a Salmonella-specific gene. It is expected that the combination of newly constructed stn-HRM and previously reported procedures could further improve the credibility of Salmonella isolate serotyping.

Genomic Characterization of ß-Glucuronidase-Positive Escherichia coli O157:H7 Producing Stx2a.

Among Shiga toxin (Stx)-producing Escherichia coli (STEC) O157:H7 strains, those producing Stx2a cause more severe diseases. Atypical STEC O157:H7 strains showing a ß-glucuronidase-positive phenotype (GP STEC O157:H7) have rarely been isolated from humans, mostly from persons with asymptomatic or mild infections; Stx2a-producing strains have not been reported. We isolated, from a patient with bloody diarrhea, a GP STEC O157:H7 strain (PV15-279) that produces Stx2a in addition to Stx1a and Stx2c. Genomic comparison with other STEC O157 strains revealed that PV15-279 recently emerged from the stx1a/stx2c-positive GP STEC O157:H7 clone circulating in Japan. Major virulence genes are shared between typical (ß-glucuronidase-negative) and GP STEC O157:H7 strains, and the Stx2-producing ability of PV15-279 is comparable to that of typical STEC O157:H7 strains; therefore, PV15-279 presents a virulence potential similar to that of typical STEC O157:H7. This study reveals the importance of GP O157:H7 as a source of highly pathogenic STEC clones.

Diversity of Shiga toxin transducing phages in Escherichia coli O145:H28 and the different Shiga toxin 2 production levels associated with short- or long-tailed phages.

Shiga toxin (Stx)-producing Escherichia coli (STEC) causes serious gastrointestinal illness, including hemorrhagic colitis and hemolytic uremic syndrome. Two types of Stxs (Stx1 and Stx2) are known and both are encoded by bacteriophages (Stx phages), but the production of Stx2 is known to be a major risk factor for severe STEC infections. The production of Stx2, but not Stx1, is tightly coupled with the induction of Stx phages, and Stx2 production levels vary between STEC strains even within the same serotype. Here, we analyzed the genomic diversity of all Stx phages in 71 strains representing the entire O145:H28 lineage, one of the often highly pathogenic STECs, and the relationship between the variations in Stx phage genomes and the levels of Stx2 production by host strains. Our analysis reveals highly dynamic natures of Stx phages in O145:H28, including the independent acquisition of similar Stx phages by different sublineages, the recent transfer of Stx phage between different sublineages, and the frequent gain and loss of Stx phages in some sublineages. We also show the association of the Stx2 phage types with the Stx2 production levels of host strains: strains carrying short-tailed Stx2 phages exhibited significantly higher Stx2 production levels than those carrying long-tailed Stx2 phages. Detailed analyses of the Stx2 phage genomes revealed that both of short- and long-tailed phages exhibited sequence diversification and they were divided into two groups, respectively, based on the sequence similarity of the phage early region encoding genes responsible for phage induction, short-tailed phages contained early regions clearly different in genetic organization from those in long-tailed phages. Therefore, the variations in the early regions between short-and long-tailed Stx2 phages appeared to be linked to a striking difference in Stx2 production levels in their host strains. These results broaden our understanding of the diversification and dynamism of Stx phages in O145:H28 and the association of Stx2 phage types with the Stx2 production level in this STEC lineage.

Capuchin monkeys (Cebus apella) use conspecifics' emotional expressions to evaluate emotional valence of objects.

Emotional expressions provide important clues to other individuals' emotional states, as well as the environmental situations leading to such states. Although monkeys often modify their behavior in response to others' expressions, it is unclear whether this reflects understanding of emotional meanings of expressions, or simpler, non-cognitive processes. The present study investigated whether a New World monkey species, tufted capuchin monkeys, recognize objects as elicitors of others' expressions. Observer monkeys witnessed another individual (demonstrator) reacting either positively or negatively to the contents of one of two containers and were then allowed to choose one of the containers. The observer preferred the container that evoked positive expressions in the demonstrator and avoided the container that evoked negative expressions. Thus, the monkeys appropriately associated the emotional valence of others' expressions with the container. This finding supports the view that the ability to represent others' emotions is not limited to humans and apes.

Identification of Legionella rubrilucens isolated from a hot spring for foot-soaking in Niigata, Japan.

In May 2011, strain HYNE-20 (=JCM 17837) was isolated from a sample of hot spring water from a foot spa in Niigata, Japan, by a plating method using glycine vancomycin polymyxin B cycloheximide α-ketoglutarate (GVPCα) medium at 36°C for 7 d. The 16S rDNA sequences (1,469bp) of this strain (accession number: AB638719) had high (99.7%) similarity to Legionella rubrilucens, and we identified that this strain was indeed Legionella rubrilucens. When this strain was cultured on buffered charcoal yeast extract α-ketoglutarate (BCYEα) agar at 36°C for 7 d, it exhibited red autofluorescence under UV light (365 nm) . The dominant cellular fatty acids of the strain HYNE-20 were 16:1ω7c (29.9%) , and the guanine-plus-cytosine (G+C) content of DNA was 49.0 mol%. This is the first report that Legionella rubrilucens was isolated from a hot spring for foot soaking.

Surveillance of Staphylococcus aureus in cheese produced in Hokkaido.

Although the number of cheese manufacturing units in Hokkaido had increased every year and exceeds 60, many of these units are small-scale processors. We examined the cheese produced in Hokkaido for the presence of Staphylococcus aureus for 3 years after 2002. During the study period, S. aureus was isolated from 38 cheese samples: 3.6 to 9.2% of the total cheese samples examined and 13.0 to 20.0% of the total mozzarella-type cheese samples. The largest population of S. aureus was 2.0 x 10(4) CFU/g. The isolated S. aureus strains were subjected to PCR analysis to look for seven se genes. Of the 38 isolates, 20 did not possess the se gene, but the remaining 13 isolates had seg and sei genes. No enterotoxins were detected in the cheese samples analyzed with a commercial kit.

Capuchin monkeys (Cebus apella) modify their own behaviors according to a conspecific's emotional expressions.

Understanding conspecifics' emotional states is important for managing social interactions. We examined whether capuchin monkeys modify their own behaviors in response to a conspecific's emotional expressions. Six monkeys saw a demonstrator monkey responding emotionally to an object in a container; the subject monkeys could not see the object. The subjects reached for the container more frequently when the demonstrator showed an emotionally positive expression than when she showed an emotionally neutral or negative expression. This is the first report that New World monkeys are sensitive to the emotional valence of conspecifics' emotional expressions. The finding is consistent with the hypothesis that monkeys can recognize emotional meanings in others' expressions, an ability previously attributed only to humans and great apes.