Detection of Chlamydia trachomatis and Neisseria gonorrhoeae by strand displacement amplification and relevance of the amplification control for use with vaginal swab specimens

Vaginal swab specimens may be preferable to cervical swab or urine specimens for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae because of the ease of specimen collection and transport. The purpose of this study was to evaluate whether vaginal swab specimens are equivalent to cervical swab specimens for the detection of N. gonorrhoeae and C. trachomatis by the Becton Dickinson strand displacement amplification assay (SDA) with the BDProbeTec ET instrument and then to evaluate the use of the amplification control in a clinical research setting. In the first phase, vaginal and cervical swab specimens were obtained from 455 symptomatic women aged 18 to 40 attending primary health care and sexually transmitted disease clinics. Thirty-nine specimens (8.6%) had true-positive results for N. gonorrhoeae and 37 specimens (8.1%) had true-positive results for C. trachomatis. The sensitivity of SDA was superior to that of culture for the detection of N. gonorrhoeae with vaginal swab specimens and equivalent to that of the Roche PCR for the detection of C. trachomatis with cervical swab specimens. In the second phase of the study, 1,411 consecutively collected vaginal swab specimens were evaluated, with 357 (25.3%) specimens giving indeterminate readings on the basis of the result for the amplification control. The prevalences of sexually transmitted pathogens in vaginal swab specimens with and without use of the amplification control were 6.0 and 5.8%, respectively, for C. trachomatis and 3.1 and 3.0%, respectively, for N. gonorrhoeae. Although, vaginal swab specimens were equivalent to cervical swab specimens for the detection of N. gonorrhoeae and C. trachomatis by SDA with respect to sensitivity, one in four vaginal swab specimens yielded an indeterminate result when the amplification control was used. The amplification control has limited value for use with vaginal swab specimens.     

Development of hatching blastocysts from immature human oocytes following in-vitro maturation and fertilization using a co-culture system

Recently, in-vitro maturation (IVM) of immature human oocytes recovered from non-stimulated follicles has been applied in the treatment of infertility. However, in previous reports, very few embryos cultured in conventional medium have reached the expanded blastocyst stage following in-vitro maturation and fertilization (IVM/IVF). The objective of this study was to investigate whether the developmental competence of human embryos following IVM/IVF could be enhanced by the use of a human ampullary cell co-culture system. Immature human oocytes were aspirated from small follicles at Caesarean section and then cultured in medium containing human menopausal gonadotrophin for 36 to 48 h, followed by insemination. Zygotes were randomly cultured either in conventional culture medium alone or in the co-culture system. Of 48 embryos cultured in conventional medium alone, all arrested at the 2-16- cell stage on day 3 after insemination. Of 46 embryos cultured in the co-culture system, 26 embryos (56.5%) arrested at the 2-16-cell stage. Six embryos (13%) developed to the morula stage. Fourteen embryos (30.4%) developed to expanded blastocysts and two blastocysts were hatching on day 7 after insemination. We conclude that co-culture significantly enhances the development of blastocysts in embryos resulting from IVM/IVF.      

Role of L3T4-bearing T-cell populations in experimental murine chlamydial salpingitis.

A role for both the cellular and humoral components of the immune response has been established for chlamydial infection. The significance of helper (L3T4) T cells was evaluated by using a Chlamydia trachomatis murine salpingitis model for upper genital tract chlamydial infection. Mouse oviducts were inoculated with C. trachomatis by using the mouse pneumonitis agent (MoPn) or control medium. Mice depleted of L3T4-bearing lymphocytes had significantly higher (P less than 0.05) numbers of organisms recovered at day 7 postinoculation. The rate of hydrosalpinx formation was significantly higher in the mice depleted of L3T4-bearing lymphocytes (27 of 31 [87%] ) than in the infected undepleted group (8 of 16 [50%] ) (P less than 0.01). The geometric mean antichlamydial immunoglobulin G titers at day 54 postinoculation were significantly higher in the L3T4-depleted mice (mean titer, 2,030) than in the undepleted group (mean titer, 776; P less than 0.05). The rate of fertility was lower in the L3T4-depleted group (2 of 31 [6%]) than in the infected, undepleted mice (2 of 16 [13%]), but this difference did not reach statistical significance. In conclusion, the greater persistence of organisms in the oviduct and higher rates of hydrosalpinx formation in mice depleted of L3T4-bearing cells suggests that these cells play a role in the clearing of organisms following infection and thus in reducing the degree of oviduct obstruction and damage.     

Electron and light microscopic observations on relationships between lymphocytes and intestinal epithelium

Correlated electron and light microscopic studies of the distribution, position, appearance, migration and fate of lymphocytes with relation to the intestinal epithelium of the young adult rat, mouse, hamster and man have been made. Under the conditions of this investigation, the lymphocytes which are normally present in the intestinal epithelium are intercellular in position, irrespective of their level within the epithelium. The great majority of these cells are located in the basal region of the epithelium, below the level of the epithelial cell nuclei. There was no evidence of lymphocytic degeneration. Lymphocytes migrated from the lamina propria into the epithelium and back again into the lamina propria, with no evidence of movement toward the free surface of the intestinal epithelium and extrusion into the lumen.     

The ratio of 2nd to 4th digit length: a predictor of sperm numbers and concentrations of testosterone, luteinizing hormone and oestrogen

The differentiation of the urinogenital system and the appendicular skeleton in vertebrates is under the control of Hox genes. The common control of digit and gonad differentiation raises the possibility that patterns of digit formation may relate to spermatogenesis and hormonal concentrations. This work was concerned with the ratio between the length of the 2nd and 4th digit (2D:4D) in humans. We showed that (i) 2D:4D in right and left hands has a sexually dimorphic pattern; in males mean 2D:4D = 0.98, i.e. the 4th digit tended to be longer than the 2nd and in females mean 2D:4D = 1.00, i.e. the 2nd and 4th digits tended to be of equal length. The dimorphism is present from at least age 2 years and 2D:4D is probably established in utero; (ii) high 2D:4D ratio in right hands was associated with germ cell failure in men (P = 0.04); (iii) sperm number was negatively related to 2D:4D in the right hand (P = 0.004); (iv) in men testosterone concentrations were negatively related to right hand 2D:4D and in women and men LH (right hand), oestrogen (right and left hands) and prolactin (right hand) concentrations were positively correlated with 2D:4D ratio and (v) 2D:4D ratio in right hands remained positively related to luteinizing hormone and oestrogen after controlling for sex, age, height and weight.      

Genetic linkage of the tricho-dento-osseous syndrome to chromosome 17q21

Tricho-dento-osseous syndrome (TDO), MIM# 190320, is transmitted as a highly penetrant autosomal dominant trait that is characterized by variable clinical expression. The principal clinical features include kinky/curly hair in infancy, enamel hypoplasia, taurodontism, as well as increased thickness and density of cranial bones. Possible genetic linkage has been reported for TDO with the ABO blood group locus, but the gene defect remains unknown. We have identified four multiplex families (n = 63, 39 affected, 24 unaffected) from North Carolina segregating TDO. We previously have excluded a major locus for TDO in the ABO region for these families. Utilizing a genome-wide search strategy, we obtained conclusive evidence for linkage of the TDO syndrome locus to markers on chromosome 17q21 (D17S791, Z max = 10.54, Theta = 0.00) with no indication of genetic heterogeneity. Multipoint analysis suggests the TDO locus is located in a 7 cM chromosomal segment flanked by D17S932 and D17S941. This finding represents the first step towards isolation and cloning of the TDO gene. Identification of this gene has important implications for understanding normal and abnormal craniofacial development of hair, teeth and bone.      

Cryopreservation of all prezygotes in patients at risk of severe hyperstimulation does not eliminate the syndrome, but the chances of pregnancy are excellent with subsequent frozen-thaw transfers

In-vitro fertilization patients (n = 15) at risk of ovarian hyperstimulation syndrome (OHSS) (oestradiol > or =4500 pg/ml on the day of human chorionic gonadotrophin administration and 25 or more follicles of intermediate or large size) underwent aspiration of all follicles and cryopreservation of all fertilized oocytes at the pronuclear stage. Patients were monitored for up to 2 weeks post- retrieval. Subsequent transfer of cryopreserved-thawed embryos was performed in programmed cycles using exogenous oestrogen and progesterone for endometrial preparation. Two patients (13%) developed OHSS necessitating hospitalization and vaginal aspiration of ascitic fluid. Two other patients (13%) developed moderate OHSS requiring ascitic fluid vaginal aspiration in the office setting, with dramatic improvement of the condition. Subsequent transfer of cryopreserved- thawed embryos yielded a clinical pregnancy rate of 58% per transfer and ongoing or delivery rates of 42 and 67% per transfer and per patient respectively. By eliminating pregnancy potential with cryopreservation of all prezygotes and examining the pregnancy potential with subsequent cryopreserved-thawed transfers, it is concluded that OHSS is reduced, but not eliminated for patients at risk. Subsequent transfer of cryopreserved-thawed prezygotes in a programmed cycle with exogenous steroids yields an excellent pregnancy rate.      

Charged cyclodextrin-mediated sample stacking in micellar capillary electrophoresis. A simple method for enhancing the detection sensitivity of hydrophobic compounds.

The development of on-line sample stacking techniques for enhancing limits of detection of neutral analytes in micellar capillary electrophoresis (MCE) has recently gained much attention. Utilizing high-conductivity sample matrices to invoke sample stacking is promising, but requires the limited use of sample solubilizing agents such as alcohols in the sample matrix. In this study, we show how simple replacement of the sample solvent (methanol) with a solution of sulfated beta-cyclodextrin (s beta-CD) allows a significant increase in the sensitivity of detection of model hydrophobic analytes. This increase in sensitivity is accompanied by significant peak sharpening. Sulfated CDs in the sample matrix allow for effective solubilization of hydrophobic analytes without the use of organic solvents such as methanol. The testing of various sample matrix s beta-CD concentrations for their effect on peak sharpening identified 3 to 5% as optimal for the estrogen standards. The use of a s beta-CD sample matrix allowed for hydrostatic injections (3.5 kPa) of 297 s, compared with 4 s when the analytes were dissolved in methanol. A mechanism explaining the s beta-CD-induced effect involves an analyte transfer mechanism where the s beta-CDs, despite providing anodic mobility to analytes in the sample zone, are able to transfer analytes to trailing separation buffer micelles for "recycling" back into the sample zone without compromising the stacking process. The overall improvement in sensitivity allows detection of estrogens in the parts-per-billion range and stands to improve the utility of MCE as a bioanalytical technique.