Rinsho Biseibutsu Jinsoku Shindan Kenkyukai shi = JARMAM : Journal of the Association for Rapid Method and Automation in Microbiology最新文献

It was developed that an anti-norovirus single-chain Fv (scFv) antibody for immunochromathographic test kit by using the phage displayed technique and biopanning. To obtain the scFv having wide reactivity for several norovirus genotypes, a mixture of recombinant norovirus capsid proteins, including 11 norovirus genotypes, was used for biopanning. Then, one anti-norovirus scFv antibody that recognized both of norovirus genogroups GI and GII was identified. An immunochromatographic test strip was constructed by using the scFv and demonstrated it to detect norovirus in stool samples. The immunochromatographic strip showed similar sensitivity to a commercially available one on which several monoclonal antibodies are included.

Reports of community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) infections have recently increased in Japan. To determine the status of MRSA infections in our hospital, we investigated their Staphylococcal cassette chromosome mec (SCCmec) types and prevalence of Panton-Valentine leukocidin (PVL). In addition, we investigated the relation between their SCCmec and antimicrobial susceptibility. The 191 strains were isolated from January to July in 2011 and were classified as SCCmec type I (2, 1.0％), type II (136, 71.2％), type IV (36, 18.8％), type V (4, 2.1％) and type VIII (2, 1.0％). Eleven isolates (5.8％) were designated as nontypable. No isolates were PVL-positive in this study. The SCCmec type IV strains were more susceptible to imipenem (MIC90, 0.25 μg/ml) than SCCmec type II strains (MIC90, >16 μg/ml). This difference was also observed between SCCmec type IV and SCCmec type II in susceptibility levels to clarithromycin, clindamycin, minocycline, and levofloxacin, but not to gentamicin. In particular, SCCmec type IV strains were susceptible to imipenem and minocycline. The result indicates these susceptibility is useful to discriminate CA-MRSA from Hospital-associated MRSA (HA-MRSA).

Pathogenic bacteria of Mycoplasma pneumonia, Mycoplasma pneumoniae, do not respond to β-lactam antimicrobial agents. Therefore, it is clinically important to promptly identify infection with this type of bacteria. However, conventional examination methods such as culture and antibody tests are not useful for the rapid diagnosis of this bacterial type. In this study, we examined a Loopamp Mycoplasma P-detecting Reagent Kit developed based on the LAMP (Loop-mediated isothermal amplification) method, which may overcome the limitations of the conventional methods. The consistency rate for the conventional polymerase chain reaction (PCR) method was 98.3%. That for a rapid antibody test, immunocard Mycoplasma antibody, was 54.0%, suggesting the limitation of the rapid antibody test. We compared a pretreatment method using a Loopamp PURE DNA Extraction Kit, as a simple DNA extraction method, with that using a QIAamp DNA Mini Kit. The consistency rate was 91.4%, suggesting the usefulness of the simple DNA extraction method. The use of this kit may facilitate a more rapid diagnosis. The Loopamp Mycoplasma P-detecting Reagent Kit is useful for the accurate and rapid diagnosis of Mycoplasma pneumoniae infection.

Recently, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been challenged for the identification of Nocardia species. However, the standard ethanol-formic acid extraction alone is insufficient in allowing the membrane proteins of Nocardia species to be ionized by the matrix. We therefore aimed to establish our new extraction method for the MALDI-TOF MS-based identification of Nocardia species isolates. Our modified extraction procedure is through dissociation in 0.5% Tween-20 followed by bacterial heat-inactivation, mechanical breaking of the cell wall by acid-washed glass beads and protein extraction with formic acid and acetonitrile. As reference methods for species identification, full-length 16S rRNA gene sequencing and some phenotypical tests were used. In a first step, we made our own Nocardia database by analyzing 13 strains (13 different species including N. elegans, N. otitidiscaviarum, N. asiatica, N. abscessus, N. brasiliensis, N. thailandica, N. farcinica, N. nova, N. mikamii, N. cyriacigeorgica, N. asteroids, Nocardiopsis alba, and Micromonospora sp.) and registered to the MALDI BioTyper database. Then we established our database. The analysis of 12 challenge strains using the our database gave a 100% correct identification, including 8 strains identified to the species level and 4 strains to the genus level (N. elegans, N. nova, N. farcinica, Micromonospora sp.) according to the manufacture's log score specifications. In the estimation of reproducibility of our method intended for 4 strains, both within-run and between-run reproducibility were excellent. These data indicates that our method for rapid identification of Nocardia species is with reliability, reproducibility and cost effective.

Since we had experienced a case of misidentification for group B Streptococcus (GBS) (Streptococcus agalactiae) by automated identification system RAISUS, we have investigated the identification accuracy for GBS by automated identification system RAISUS system using our stock isolated of GBS. Among 31 strains including standard strain ATCC13813 of GBS, 30 strains were identified as GBS, while only 1 strain was misidentified as Streptococcus constellatus. We have needed 3 hours to identify 29 true GBS strains, while it has taken 6 hours for misidentified strain as S. constellatus. Since we have needed more than 4 hours for our misidentified strain of GBS, the tendency for identification time to become long has been recognized. Longer identification time might be associated with the misidentification. The improvement for galactose and glycerol tests of RAISUS by the manufacturer has resulted in better identification. As for the strains to be identified as S. constellatus by RAISUS, we might re-check it with other identification kits when there is higher clinical necessity.