Subtyping of Mycoplasma pneumoniae isolates based on extended genome sequencing and on expression profiles

Mycoplasma pneumoniae isolates from patients, collected over a period of 12 years in Germany, were characterized by various methods (parameters) including multilocus sequence typing, restriction fragment length polymorphisms, Western blotting with mono-specific antibodies directed against selected proteins or with polyspecific antibodies directed against the Triton X-114-soluble protein fraction, and two-dimensional gel electrophoresis. The results for 91 isolates from Germany, which were complemented with 14 isolates from the USA and 10 isolates from France, clearly showed that M. pneumoniae is a highly uniform species and that most of the isolates could be assigned to one of the two subtypes 1 and 2. The members of one subtype differ from the other with respect to the sequence of the P1 gene, the ORF6 gene, the P65 gene, and by a typical DNA restriction fragment pattern. We observed four isolates (variants), which seemed identical by the above mentioned criteria, but did not belong to either one of the two subtypes. They showed most of the subtype 2-specific features, but differed in the sequence of the P1 gene and showed a variation in the restriction fragment pattern. The appearance of subtype 1 or 2 over the last 12 years in Germany showed a dominance of subtype 1 between 1989 and 1996 and a dominance of subtype 2 between 1997 and 1998. The variant (neither subtype 1 nor subtype 2) was only detected in 1991 and 1995 but it had no epidemiological consequences.     

Peripheral blood chromosome aberrations in MDS.

The frequency of non-clonal structural and numerical chromosome aberrations in peripheral blood lymphocytes of 51 patients with MDS and 37 age-matched hematologically normal subjects is assessed. The frequency of aneuploid cells (p less than 0.001) and of structural aberrations (p less than 0.005) was significantly higher in MDS patients than in normal subjects, but showed no relationship with FAB type or with the presence of clonal karyotype abnormalities in the bone marrow. Exchange configurations were only observed in MDS patients (27.5%). The data also suggest that there may be an association between high peripheral blood aberration levels and rapidly progressive disease. This may indicate increased mutagen sensitivity and have implications for treatment.     

Comparative evaluation of the API 20S system and the automicrobic system gram-positive identification card for species identification of streptococci.

Two commercial methods, the API 20S system (API; Analytab Products, Inc., Plainview, N.Y.) and the Gram-Positive Identification Card (GPI; Vitek Systems, Inc., Hazelwood, Mo.), were evaluated without additional tests for the identification of 241 streptococcus strains. Organisms included 60 beta-hemolytic strains, 36 group D strains, 26 Streptococcus pneumoniae strains, and 119 viridans streptococcus strains. API correctly identified to species 68.3% of beta-hemolytic strains, 86.1% of group D strains, 53.9% of S. pneumoniae strains, and 12.6% of viridans streptococci. This method provided excellent identification of group A and B and S. faecalis strains. Overall, API correctly identified 41.9% of strains to species, with 41.1% good likelihood but low selectivity, 15.8% incorrect, and 1.2% not identified. GPI correctly identified to species 58.3% of beta-hemolytic strains, 97.2% of group D strains, 80.8% of S. pneumoniae strains, and 57.2% of viridans streptococci. Group A, B, and D strains were all accurately identified by this system. Overall, GPI correctly identified to species 66.0% of strains, with 8.7% correct preliminary identification, 20.8% incorrect, and 4.6% not identified. Both methods represent a worthwhile advance in streptococcal identification. Neither system, however, can be recommended for species identification of the viridans group at this time.     

DADOS-Survey: an open-source application for CHERRIES-compliant Web surveys

Background  

The Internet has been increasingly utilized in biomedical research. From online searching for literature to data sharing, the Internet has emerged as a primary means of research for many physicians and scientists. As a result, Web-based surveys have been employed as an alternative to traditional, paper-based surveys. We describe DADOS-Survey, an open-source Web-survey application developed at our institution that, to the best of our knowledge, is the first to be compliant with the Checklist for Reporting Results of Internet E-Surveys (CHERRIES). DADOS-Survey was designed with usability as a priority, allowing investigators to design and execute their own studies with minimal technical difficulties in doing so.     

Comparison of two automated multichannel haematology analysers in domestic animals

Blood samples from 33 dogs, 28 cats, 24 horses and 25 cattle were analysed in duplicate on the Coulter Counter S-Plus IV and the Baker 9000 multichannel haematology analysers. The precisions of each instrument and the correlations between the instruments were evaluated. The precisions of both systems were good but, the Baker 9000 system showed more variability between duplicate determinations for all parameters. The Baker 9000 had better precision for white blood cell count, haemoglobin concentration, and MCV. Correlations between the two instruments were excellent except for MCHC. Neither analyser reliably provided feline platelet counts and the Baker 9000 also failed to report error messages for 8 of 10 apparently false platelet counts. The Coulter instrument consistently reported higher haemoglobin concentrations and higher feline and lower canine white blood cell counts than the Baker 9000.     

Nutritionally variant Streptococcus pyogenes from a periorbital abscess.

A nutritionally variant Streptococcus pyogenes strain was isolated from a periorbital abscess. The organism was identified with the use of three rapid biochemical test kits, and the group A antigen was detected by conventional serology as well as direct antigen detection tests.     

A molecular and FISH analysis of structurally abnormal Y chromosomes in patients with Turner syndrome

Fourteen patients with Turner syndrome and a structurally abnormal Y chromosome were analysed by PCR amplification and fluorescence in situ hybridisation for the presence of sequences specific to defined regions of the Y chromosome. Thirteen patients had a mosaic karyotype including a 45,X cell line and one case was non-mosaic in cultured lymphocytes. Ten patients had a pseudodicentric Yp chromosome, two an isodicentric Yq, one a pseudodicentric Yq, and one a derived Y chromosome. Two of the patients with a psu dic(Yp) chromosome had complex karyotypes with more than two cell lines, one of which exhibited five morphologically distinct mar(Y) chromosomes, presumably derived from a progenitor psu dic(Yp). Nine of the ten psu dic(Yp) chromosomes were positive for all Yp and Yq probes used except DYZ1 which maps to Yq12, suggesting a common breakpoint near the Yq euchromatin/heterochromatin boundary. In the three patients with a dicentric Yq chromosome two different breakpoints were observed; in two it was between PABY and the subtelomeric repeat sequence and in one it was between DYZ5 and AMGY in proximal Yp. Our results suggest that the great majority of structurally abnormal Y chromosomes found in Turner syndrome mosaics contain two copies of virtually all of the functional Y chromosome euchromatin.     

The CA 125 tumour-associated antigen: a review of the literature

CA 125 is an antigenic determinant on a high-molecular-weight glycoprotein recognized by a monoclonal antibody which was raised using an ovarian cancer cell line as an immunogen. During the last 5 years the studies reviewed in this paper have provided information concerning the nature, distribution and clinical significance of CA 125. The CA 125 determinant is expressed by epithelial ovarian tumours and various other pathological and normal tissues of Müllerian origin. The function of the glycoprotein expressing CA 125 remains unclear but the distribution of the antigen suggests that it may have a physiological role. The highest serum levels of CA 125 are found in ovarian cancer patients, but elevation of serum CA 125 may also be associated with other malignancies and benign and physiological states, including pregnancy, endometriosis and menstruation. Despite limitations of sensitivity and specificity serum CA 125 estimation is of clinical value in the pre-operative diagnosis and monitoring of ovarian malignancy and may be a prognostic indicator for this disease. The role of CA 125 in screening for early-stage ovarian cancer is currently under investigation. Recent reports suggest that serum CA 125 measurement may also be of value as a prognostic indicator in endometrial cancer and as a reflection of disease status in advanced endometriosis.     

Drosophila Cyclin B3 is required for female fertility and is dispensable for mitosis like Cyclin B

Cyclin B3 has been conserved during higher eukaryote evolution as evidenced by its identification in chicken, nematodes, and insects. We demonstrate that Cyclin B3 is present in addition to Cyclins A and B in mitotically proliferating cells and not detectable in endoreduplicating tissues of Drosophila embryos. Cyclin B3 is coimmunoprecipitated with Cdk1(Cdc2) but not with Cdk2(Cdc2c). It is degraded abruptly during mitosis like Cyclins A and B. In contrast to these latter cyclins, which accumulate predominantly in the cytoplasm during interphase, Cyclin B3 is a nuclear protein. Genetic analyses indicate functional redundancies. Double and triple mutant analyses demonstrate that Cyclins A, B, and B3 cooperate to regulate mitosis, but surprisingly single mutants reveal that neither Cyclin B3 nor Cyclin B is required for mitosis. However, both are required for female fertility and Cyclin B also for male fertility.     

Matrix vesicle biogenesis in vitro by rachitic and normal rat chondrocytes.

Calcifying matrix vesicles (MVs) are released from chondrocytes and osteoblasts in monolayer culture. In the present studies, we tested the ability of rachitic versus normal rat growth plate chondrocytes in micromass or monolayer primary cultures to produce MVs. Unlike earlier reports of in vitro MV biogenesis by chicken chondrocytes in which most MVs were released into the medium, we found that most of the released rat matrix vesicles were entrapped in a newly formed cartilaginous matrix enveloping the cells. These matrix-associated MVs could be isolated by mild collagenase treatment and concentrated by differential centrifugation. Vesicle production slowed in the older 2- to 4-week-old cultures and, unlike vesicle release from cultured chicken chondrocytes, active vesicle production did not show a second burst of activity at 3 to 4 weeks. Alkaline phosphatase (ALP) activity diminished with time in culture in cells and matrix vesicles, suggesting a decrease in differentiative expression. Protein profiles on SDS polyacrylamide gels of native matrix vesicles and culture-derived MVs from rachitic and normal cells were quite similar and showed a typical simplified protein pattern as compared to chondrocyte plasma membrane proteins. There were distinctive proteins migrating at 130, 80 to 95, 66, 43, 20, and 14 kd. Culture-derived MVs showed vigorous in vitro calcifying activity that was ALP related. We conclude that 1) rachitic chondrocytes are essentially normal in their matrix vesicle production; 2) matrix entrapment of MVs is a characteristic of rat chondrocyte cultures; and 3) culture-produced MVs are similar to native MVs in protein profile and calcifiability, and thus can be studied as a model for normal MV composition and calcification.