Since plasmid-mediated metallo-β-lactamase (MBL)-producing Pseudimonas aeruginosa was reported in Japan, MBL-producing Gram-negative rods (GNRs) have emerged worldwide. We developed a way to detect MBL-producing GNRs in routine examination using broth microdilution with the MBL inhibitor sodium mercaptoacetate (SMA) in frozen plates. Between 1996 and 2005, we evaluated this and other methods, including broth microdilution with another MBL inhibitor dipicolinic acid (DPA) in dry plates, conventional PCR, and a combined simple DNA preparation and enzymatic PCR product detection. The combined method is suitable for detecting IMP-type MBL-producing GNRs from numerous isolates. Broth microdilution with SMA at a concentration of 400μg/mL had high performance and detected most PCR-positive MBL-producing GNRs in routine antimicrobial susceptibility testing. DPA in dry plates at 400μg/mL yielded false positive results in 11.4% of isolates but worked satisfactorily at 175μg/mL and 400μg/mL of SMA in frozen plates. Until 1996, MBL had been detected from only 6 bacterial species, i.e., Pseudomonas aeruginosa, Pseudomonas putida, Pseudomonas stutzeri, Achromobacter xylosoxidans, Serratia marcescens, and Citrobacter freundii, MBL-producing GNRs were later found in other glucose nonfermenting GNRs such as Acinetobacter baumanii, Acinetobacter lwoffii, and Burkholderia cepacia complex and Enterobacteriaceae. Most MBL-producing bacteria were multidrug resistant and no antimicrobial agents exist that are active against such isolates in monotherapy, making their rapid detection very important in controlling infection control.

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Marked Streptococcus pneumoniae tolerance is attributed to a penicillin and macrolide resistance genotypes, yet no report has, to our knowledge, compared the ability of these two genotypes to grow. Deviations in the number of clinical appearances of Streptococcus pneumoniae depend on the combination of these genotypes, thought primarily due to the individual strain growth. To test this, we compared ATCC6303 and a strain of each genotype with an absorption spectrophotometer. Time points-TO, T2, T4, T8, T10, T12, and T24-were compared to each of the following factors: initial value, dullness time, doubling time (D.B.T.), peak attainment, peak value, and stationary value. ATCC6303 and the clinical strain (PSSP8) showed no difference in time points or tested factors. Penicillin susceptible Streptcoccus pneumoniae (PSSP) 8 and Penicillin intermediately resistant Streptcoccus pneumoniae (PISP) (2b, mefA) strains showed significant differences in some time points-T8, T10, and T12-and in certain factors-dullness time, D.B.T., and peak attainment.
In penicillin-resistant genotypes, the sequential growth rate priority was PSSP>PISP>PRSP, while in macrolide-resistant genotypes, it was mefA>non-mefA, and non-ermB>ermB. PISP (2x) and PISP (2x+2b, mefA) begin doubling immediately, suggesting that they proliferate earlier than other strains.
Differences in the start of doubling time and doubling speed suggest that different strains are distributed among clinical resistant S. pneumoniae strains.

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After developing the Ec-LPS array for Escherichia coli O-serogroup serodiagnosis (J. Jpn. Infect. Dis., 2007; 81: 26-32), we tested the array's usefulness in sera bled over 8 years ago from 24 patients with pediatric diarrhea. IgM and IgA antibodies in 20 sera among sera from the 24 reacted with a single LPS spot, making itpossible to diagnose the O-serogroup. IgG antibodies in almost all patient sera reacted with many LPS among the 58 O-serogroup LPS of E. coli. O-serogroup strains of E. coli isolated from the 24 patients numbered 15. Among 11 patients in who O-serogroups were serodiagnosed by both methods, 7 were diagnosed with the same O-serogroups. Based on these results, this array appears useful in the O-serogroup serodiagnosis of E. coil.

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To elucidate the origin of infection, we conducted epidemiological and bacteriological studies to clarify the origin of five sporadic outbreaks of enterohemorrhagic Escherichia coli (EHEC) O157: H7 between May and July 2007 in Saitama City and its outskirts. Of the 20 subjects were reported; including 6 patients and 5 infected persons, none of the 9 symptomatic subjects developed hemolytic-uremic syndrome. No association was confirmed between infection and food materials, but 11 organisms showed almost the same chromosomal DNA restriction patterns produced by pulsed-field gel electrophoresis. Our results suggested that these 5 sporadic outbreaks were part of a diffuse outbreak induced by an EHEC O157: H7 strain having a single origin.

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Changes in gelatin particle agglutination (PA), hemagglutination inhibition (HI), and neutralization (NT) antibodies were compared using sera from 124 individuals collected between 3 weeks and 10 years after measles vaccination, and the relationship between these changes and IgG avidity was studied. PA, HI, and NT antibodies peaked 4-5 months after vaccination. The rate of increase in mean antibody titer from 0-1 months after vaccination to peak levels was 1.7-fold for NT, 1.5-fold for HI, and 7.4-fold for PA antibodies. Peak mean antibody titer was 2^11.8 for PA, 2^6.7 for NT and 2^6.7 for HI antibodies. After peaking PA antibodies changed in parallel with NT and HI antibody titers, and correlated strongly with both antibodies (r=0.801 and 0.840). In contrast, NT and HI antibodies were consistent throughout the period. IgG avidity increased for 4-5 months following vaccination, peaking at 45%, and remaining constant at 40-50% for the next 10 years.
PA antibody is strongly influenced by IgG avidity, unlike NT and HI antibodies. Due to the effects of IgG avidity, PA antibodies increase more significantly than NT and HI antibodies as IgG antibodies mature following vaccination, resulting in a weak correlation between PA and NT or HI antibodies. Following the increase in IgG avidity to maturation, PA antibodies correlated strongly with NT and HI antibodies. PA assay detected IgM antibodies against measles virus more efficiently than the NT test. The PA assay thus differs from conventional, commonly used NT and HI assays. PA assay is simple and rapid, making it very useful for detecting measles antibodies provided that its unique features are taken into accounts.

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The Gonogen II test for rapid identification of Neisseria gonorrhoeae (Gonococcus, GC) was evaluated. The test is based on a colorimetric reaction with monoclonal antibody to GC outer membrane protein 1. Of the 50 clinical isolates of GC, 49 isolates tested positive and only one strain tested negative. Other Neisseria. spp, H. influenzae, H. parainfluenzae, E. corrodens, M. catarrhalis, and A. baumannii showed negative test results. Non-Neisseriae. spp, such as S. aureus. P. aeruginosa, E. faecalis, and E. coli also showed negative test results. No cross-reactivity was found between GC and other Neisseriae. spp or non-Neisseriae. spp. In a mixed suspension of GC and all of non-Neisseriae. spp as mentioned above, the GonoGen II test was positive. The specificity and sensitivity of the test for the identification of GC were 98% and 100%. The minimum limit of detection of GC was ≥1×10⁵cfu/mL. Decision making based on the test result is possible within 10 minutes. These findings also suggest that the test does not require pure GC. The GonoGen II test appears to be a reliable, quick and easy-to-use assay, and also to not require viable GC. Thus GonoGen II is shown to be a very useful test for the identification of GC.

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We applied a previously reported method to clarify whether a multidrug-resistant Shigella colonizes in a south Asian country. At Kansai Airport Quarantine Station, stool samples were collected from overseas travelers who reported a history of diarrhea. Shigella strains were isolated, ranging from 53 to 106 (average, 82.4) isolates/year (2001-2005), and almost 80% of the isolates were Shigella sonnei. The most frequent country of origin was India. Strains from the country of the most frequent origin were studied by antimicrobial susceptibility testing. Resistance to tetracycline, sulfamethoxazole-trimethoprim and nalidixic acid was observed at the highest frequency: in 23 of the 25 strains isolated in 2001, 5 of the 13 strains isolated in 2002, and 16 of the 19 strains isolated in 2005. Strains showing the most prevalent multidrug-resistance pattern were analyzed by pulsed-field gel electrophoresis (PFGE). The PFGE profiles showed that 27 of the 44 strains isolated in 2001, 2002, and 2005 were identical in PFGE pattern, as determined using two restriction enzymes. We concluded that a multidrug-resistant Shigella sonnei colonizes in a south Asian country.

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Neutralizing antibody (NA), hemagglutination inhibition antibody (HI), and antibody assayed by IgGenzyme immunoassay (EIA) against measles were tested as a set on sera of 119 out of 120 patients institutionalized for severe motor and intellectual disability and correlations among the 3 types of antibody titers were examined.
1) NA, HI, and EIA titers correlated positively.
2) If NA, HI, and EIA antibody titers were 4 (2²) or more, NA, HI, and EIA serum titers were positive. If they were 8 (2³) or more, all sera were positive for NA. If light absorption was 4 or more, serum HI was also positive for EIA.
3) Nine cases were certified as positive for measles infection, and 3 of these were vaccinated 2-3 years after infection. All 9 had positive NA and EIA even 15 years or more after infection. Of 29 cases certified as negative for measles infection and injected with measles vaccine, 21 (72.4%) had positive NA and 16 (72.4%) had positive HI. In EIA, 28 of the 29 (96.6%) showed positive. The only EIA-negative case was also the only one negative for both NA and HI, i.e., an 18-year-old man suffering from chromosomal aberration, 21-ring trisomy, and suspected of being a low responder to measles antigens, including NA, HI and EIA antigens.
4) The above facts suggest that EIA is a more sensitive test for positive history of wild measles virus or measles vaccine virus contact, making it possible to detect measles or measles vaccine injection. To prevent nosocominal infection, it is important to know whether institutionalized individuals are immune to measles, but many have no clear history of measles or measles vaccine injection, especially those 40 years old or older.
5) Institutionalized individuals 40 years old or more numbered 45. Their antibody titers against measles were positive, 82.2% in NA, 48.9% in HI and 91.1 % in EIA. The high positive EIA rate suggests that most would sufferered from measles before institutionalization, because they had little chance of measles vaccination as children and little possibility of measles infection in the institution at nosocomial infection currence in 1983, which was limited in other ward, and no nosocomial infection of measles has been experienced in this institution during this more than 20 years.

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Six cases of scrub typhus (tsutsugamushi disease) were reported to the Shizuoka Prefecture Hamamatsu City public health center during the seven years from 2001 to 2007. The content of the clinical record of the five cases were investigated. High serum titers of antibody to Gilliam-type Orientia tsutsugamushi were detected by immunofluorescense assay in most of these patients. Fever, rash, headache and relative bradycardia seen at a high frequency. On peripheral blood smear examination, atypical lymphocytes were detected in 3 cases. Serum electrolyte examination revealed hyponatremia in 4 (80%) patients; SIADH was suspected in one of these cases. All the patients improved promptly following start of therapy with intravenous or oral minocycline.

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Highly active antiretroviral therapy (HAART) has dramatically decreased the incidence of HIV-1-associated morbidity and mortality. During the initial months of HAART, immune reconstitution inflammatorysyndrome (IRIS), an adverse consequence of restoration of the pathogen-specific immune response, often occurred in terminal-stage in patients, with MAC infection the most frequently implicated in IRIS.
In August 2004, a 26-year-old Japanese woman with fever and general lymphadenopathy was diagnosedwith AIDS (HIV-1 RNA 5.7×10⁵copies/mL, CD4⁺T cell count 10/μL) and disseminated Mycobacterium avium (M. avium) infection, for which antimycobacterial treatment was initiated. The M. avium infection respondedwell to two months of this treatment, and HAART was begun. Despite good virologic response to HAART (HIV-1 RNA<50 copies/mL), she contracted pulmonary disease with parenchymal lung changes, endobronchiallesions, and localized supraclavicular lymphadenitis, which are M. avium-associated IRIS. Good immunologicalresponse (CD4⁺ T cell count 136/μL) and a stronger antimycobacterial treatment helped her overcoming M. avium-associated IRIS without systemic corticosteroids or the discontinuation of HAART.
The possibility of IRIS should always be watched for when treating AIDS patients with HAART and an antimycobacterial treatment regimen formulated that considers potential drug interactions with HAART.

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