Salmonella has been implicated as a important pathogen associated with acute gastroenteritis or food borne diseases in humans. Rapid and accurate diagnostic tests for Salmonella are needed by both the food industry and clinical laboratories. A pair of oligonucleotide primers from the phoE gene was used to develop a polymerase chain reaction (PCR) procedure to detect Salmonella spp. When Chelex-extracted DNA from the SBG enrichiment culture were used as templates for the PCR amplification (E-Chx), the positive results were obtained with a few cells of Salmonella. This highly specific and sensitive technique gave an efficiency of 1.2 times (73 samples were positive) when tested against conventional culture method (62 samples were positive) using 125 fecal samples stored at 4°Cfor 1-16 months. Based on the results obtained in this study, it appears that the enrichement-PCR method (E-Chx) using Chelex-extracted DNA described here can be useful to screen a large number of samples such as foods or environmental specimens for contamination of Salmonella.
Knowing the importance of prevention of Staphylococcus aureus infections, especially that of Methicillin-resistant S. aureus (herein after abbreviated as MRSA) in the newborn room, we improved the nursing procedure in the room and examined subsequent changes in the onset condition of the above infection in newborns and infants less than 1-year-old. The infection was detected in 13 cases of stage 1 (stage according to the nursing procedure since the opening of our hospital): 6 cases (all MRSA) had their onset in the r000m. MRSA was detected in 11 cases, suspecting a hospital cross infection in relation to the period of birth and coagulase type. The infection was detected in 9 cases of stage 2 (stage of prevention of inter-newborn hospital cross infection through environmental purification within the newborn room, through hand and finger disinfection of medical workers, through improved bathing, etc.); 2 cases (1 of MRSA, 1 of Methicillin-sensitive S. aureus [hereinafter abbrevoated as MSSA]) had an onset in the room. MRSA was detected in 5 cases, but hospital cross infection was thought difficult to be responsible. The MRSA infection was detected in 4 pregnant cases (0.8%) of stage 3 (stage of effortful prevention of MRSA from carrying into the ward following examination of possible intra-nasal MRSA establishment in pregnant women in addition to similar newborn nursing to stage 2), indicating pregnant women's possible carrying MRSA into the newborn room. S. aureus infection was detected in 9 cases; 2 cases (MSSA) had an onset in the above room. MRSA was detected in 2 cases, suspecting intra-familial infections. The above results suggest that a decrease in the incidence of hospital MRSA infection can be achieved with due effort by preventing pregnant women from carrying MRSA into the newborn room and of hospital cross infection.
Chlarnydia pneumoniae was isolated from the throat of a 2-year-old girl with upper respiratory illness. The isolate, Shizuoka-37, was stained with C. pneumoniae specific monoclonal antibody (RR402), as well as the genus specific antibody (Cultureset®), but not with C. trachomatis specific monoclonal antibody (Micro-Trak®). C. pneumoniae genome was amplified by polymerase chain reaction in the isolate. Elementary bodies (EB) of the isolate was round shaped by electron micrograph.
In the present study, were examined the inhibitory effect of erythromycin on the biofilm formation by a temperature-sensitive mutant (Ts25) of Pseudomonas aeruginosa N-42 on the surface of Ishikawa cells which can produce a mucin-like glycoprotein physicochemically similar to mucoproteins produced in the human airway. Usually, 38-45 microcolonies (biofilms) were formed after 10 days of incubation in cultures of Ishikawa cells and P. aeruginosa Ts25. Erythromcin suppressed the adhension to Ishikawsa cells of P. aeruginosa and its subsequent biofilm formation at doses as low as 0.2μg/ml. Erythromycin also exhibited the suppressive effect on the production of glycoproteins by Ishikawa cells at doses higher than 1, ug/ml and the production of elastase and exoenxyme A fromP. aeruginosa at doses higher than 2μg/ml. These results suggest that erythromycin can inhibit the biofilm formation in culture of human epithelial cells and P. aeruginosa and the in-vivo bacterial biofilm formation may be attributed to host cell-derived factors rather than bacterial products.
Six hundred and seventy isolates from children with group A streptococcal infections from 1981 through 1990 were typed serologically and their antibiotic susceptibilities were determined. Productivity of streptococcal pyrogenic exotoxins was also investigated in some isolates. Four hundred and seventy-nine strains were isolated from patients with pharyngitis, 133 from those with scarlet fever, 35 from those with suppurative infection and 23 from those with nonsuppurative disease. With immediate treatment (antibiotics were started at the same day throat swabs were taken) for 10 days, 5.3% of the patients with pharyngitis including scarlet fever had relapses and 13.4% of those patients had recurrences. Of the episodes of recurrences 15.7% were due to the same M serotype strains. Six patients had two episodes of scarlet fever. M type of isolate was different in the first and the second episode of each patient. Pyrogenic exotoxin type was unprecedented in the second episode of 4 out of 6 patients. M-typable and T-typable rates of isolates were 90.7% and 97.3%, respectively. Coincidence between M and T types was 73.3%(83.0% if including strains with the same and mixed T-type). Prevalent M-serotypes were 12 and 4, but Ml, 3, or 28 was the most prevalent type of isolates in certain years. None of the 670 strains was resistant to penicillin G and cephalexin. Resistant rate of isolates to erythromycin and linecomycin was 26.5% in 1981 and 18.4% in 1982. But a marked decrease has noted since 1983 and only one has been resistant since 1986. Nineteen of the 21 erythromycin resistant strains were M-type 12 and the others were type 4 and 28. Chloramphenicol resistance was similar to erythromycin and lincomycin, and tetracycline resistance rate decreased gradually from 61.7% to less than 20% year by year.
In an attempt to know the applicability of non-cultural detection technique for N. gonorrhoeae and C. trachomatis on pharyngeal and rectal swabs, the swabs taken from N. gonorrhoeaeand C. trachomatis-free subjects were tested by Gonozyme, Chlamydiazyme, Ideia Chlamydia, Gen-Probe and Gen Probe Pace 2. As N gonorrhoeae-and C. trachomatis-free subjects, the castrated males older than age 70, with known prostate cancer treated at least for 6 months with anti-androgens who were seronegative for C. trachomatis and had a history of Ofloxacin treatment in the last 3 months and whose pharyngeal and rectal swabs were negative for N. gonorrhoeae, were subjected for study. The false negative rates were for N. gonorrhoeae from the pharynx by Gonozyme 57.7%, by Gen-Probe 0%, by Gen-Probe Pace 20%, from rectum by Gonozyme 0%, by Gen-Probe 0% by Gen-Probe Pace 20%, for C. trachomatis from pharynx by Chlamydiazyme 93.8%, by Ideia Chlamydia 8.3%, by Gen-Probe 0%, by Gen-Probe Pace 20%, from rectum by Chlamydiazyme 94.4%, by Ideia Chlamydia 70.9%, by Gen-Probe 20.0% and by Gen-Probe Pace 20%. To investigate the incidence of pharyngeal and rectal infection by N. gonorrhoeae among the patients with gonococcal genital infection, as well as pharyngeal and rectal infection by C. trachomatis among the patients with chlamydial genital infection, pharyngeal and recal swabs obtained from the patients with genital infection by N gonorrhoeae or C. trachomatis were tested by Gen-Probe Pace 2. Positive rate for N. gonorrhoeae of pharyngeal swab was in males 29.4%, in female 33.3% and of rectal swab in males 0%, in females 46.7%. Positive rate for C. trachomatis of pharyngeal swab was in males 3.9%, in females 10.5% and of rectal swab in males 0%, in females 53.3%. High infection rate of female rectum by both N. gonorrhoeae and C. trachomatis might suggest that rectal infection could be caused by direct contamination of cervical discharge. Difference of pharyngeal infection rates of both male and female between N. gonorrhoeae and C. trachomatis might suggest that pharyngeal infectibility of C. trachomatis would be lower than that of N. gonorrhoeae.
We experienced two outbreaks of influenza in the respiratory ward of our hospital in Feb. 1990 and Feb. 1993. Influenza-like symptoms were recognized in 42 of 67 cases (63%) in 1990, and in 22 of 56 cases (39%) in 1993. In the former outbreak, the prevalence of the serum CF titer for anti-influenza antibody was elevated in 25 of 42 cases (60%) (only the A2 antibody titer in 2 cases, B antibody titer in 4 cases in the latter one). Among asymptomatic inpatients, the serum titer was elevated in 3 of 25 cases (12%) in the former outbreak, but in 1 of 8 cases which the serum CF titer for anti-influenza antibody was measured in the latter one. A respiratory complications secondary to influenza were observed in 6 (2 cases pneumonia, 3 cases lower respiratory tract infection, 1 case asthma attack) of 67 cases during the former period, but only 1 of 56 cases during the latter period. In the two outbreaks of influenza in the respiratory ward we found the same results concerning the occurrence of influenza-like symptoms, an elevation of the serum antibody titer, but a significant difference was noted with regard to the phenotype of the prevalent influenza and the occurrence of complications.
An epidemic of Echo 11 virus infection occurred in Gifu Prefecture in children. Epidemiological and virological investigations were performed. The results are as follows: 1) Echo 11 viruses were isolated from patients from January to September. Most viruses were isolated in July. 2) Echo 11 viruses wsere isolated from all areas in Gifu Prefecture, especially from Seino and Hida areas. 3) Cases isolated Echovirus 11 ranged from 0 to 10 years of age, and 83.7% of them were in the 0 to 6 year of age group. 4) Five types of viruses were isolated from 51 of 86 cases (59.3%), and the Echovirus 11 was recovered from 43 cases (84.3%). 5) Echo 11 viruses were isolated from patients of various clinical conditions, but most patients in which isolated Echo 11 virus was isolated had aseptic meningitis, 17 cases (39.5%). Aseptic meningitis occurred in a higher frequency in those above 3 years of age. 6) By the cross neutralizing test between the prototype, isolated strain in 1982 and isolated strains in 1993 of Echovirus 11, a remarkable antigenic variation was not found in isolates in 1982 and 1993.
We had examined antibody titers against Legionella spp. of patients' sera which were mainly sent from other hospitals, performed with the indirect fluorescent antibody (IFA) method. The clinical status of the cases diagnosed as Legionella pneumonia serologically, were also studied. Out of 105 cases with clinically suspected Legionella pneumonia, 15 cases (14.3%) were seropositive. In 9 out of the 15 cases (60.0%) were caused by Legionella pneumophila serogroup 1. Clinical outline of these 15 cases did not contradict those reported in the literature, and erythromycin was effective in many cases. Significant rises (more than four times) of the titer were observed 3 to 4 weeks after onset in most of cases. We would like to emphasize that this should be performed serodiagnosis of Legionella pneumonia.
We performed transtracheal aspiration (TTA) in 1165 patients, who were suspected to have bronchopulmonary infection, from December 1978 to March 1993. We isolated pathogens from TTA in 806 patients (69.2%). We isolated H. influenzae (62 cases), S. pneumoniae (39 cases) andM. catarrhalis (24 cases) in patients with acute bronchitis, S. pneumoniae (65 cases), a-Streptococcus sp. (52 cases), H. influenzae (32 cases) and S. aureus (29 cases) in patients with pneumonia or lung abscess and H. influenzae (174 cases), S. pneumoniae (84 cases), P. aeruginosa (81 cases) and M. catarrhalis (42 cases) in patients with chronic lower respiratory tract infection. Anaerobic bacteria isolated from TTA included Peptostreptococcus sp. (19 cases), Bacteroides sp. (19 cases) and others. Mycoplasma pneumoniae was isolted from TTA in 8 patients with pneumo-nia without other organisms. Virus isolated from TTA included Rhinovirus (6 cases) and others. These results suggest that various pathogens affect the pathogenesis of bronchopulmonary infection. Therefore, we must diagnose the bronchopulmonary infection by the correct methods such as TTA.
Treponema pallidum hemagglutination (HA) is one of the most frequently used methods for the detection of Treponema pallidum (T. pallidum) antibodies. Recently, an innovative agglutination method using artificial carriers was newly developed, and is now available as a routine method. In order to compare the newly developed particle agglutination (PA) method (FUJIREBIO INC.) with the conventional HA method, T pallidum antibody titers of numerous sera were measured by respective methods. In the stability study, reconstituted reagent was stable for at least three weeks. Sample inactivation (56°C/30 min) demonstrated no effect on the test results. Among 800 sera, 132 (16.6%) positives (+), 633 (79.1%) negatives (-) and 34 (4.3%) indeterminates (+) were obtained by HA method. Meanwhile, 144 (18.0%) positives (+), 627 (78.4%) negatives (-) and 29 (3.6%) indeterminates (+) were obtained by PA method. The correlation between PA and HA method was 97.8%, and the antibody titers obtained by PA method showed good correlation with HA method. Those samples which showed discrepancy between PA and HA method in the above study were further examined with fluorescent treponemal antibody-absorption (FTA-ABS) method. The results obtained from FTA-ABS method were almost consistent with those obtained from PA method. For respective syphilis patients in stage I and II, antibody titer was monitored by HA, PA and RPR method. The results indicated that changes in antibody titer obtained from PA method was approximately the same as the titer changes obtained from RPR method. Namely, PA method detected the presence of IgM earlier than HA method. Further, changes in antibody level accompanied with the disease treatment were detected by PA method, and the changes in antibody titer correspond to the clinical progress of the disease.
Direct immunoperoxidase technique using human monoclonal antibody (C7) against a p65 antigen of cytomegalovirus (CMV) has been utilized to detect CMV antigen-positive leukocytes in the peripheral blood (CMV antigenemia). This technique was evaluated for enumeration of CMV antigen-positive leukocytes. The parameters included stability of the reagents, reproducibility of the results and quantitativity of the detection. The horseradish peroxidase (HRR)-labeled F (ab') 2 fractions of C7 antibody were found to be superior to the equivalent Fab' fractions, because the former was much more stable than the latter. Enumeration of the CMV antigen-positive leukocytes were very reproducible and quantitative. The detection limit was one CMV antigen-positive cell per 50, 000 leukocytes. Thus, this technique is reliable and practical to detect the CMV antigenemia, and it will contribute to early diagnosis of CMV diseases and initiation of antiviral therapy.