Strain-dependent role of BrkA during Bordetella pertussis infection of the murine respiratory tract

Bordetella pertussis, the causative agent of whooping cough, expresses many virulence factors believed to be involved in infection and disease progression. While these factors as a group are required for infection, deletion of individual virulence factor genes generally has limited effects on the ability of B. pertussis to efficiently infect the respiratory tract of mice, suggesting they may perform noncritical or redundant functions. We have recently observed that a B. pertussis strain, putatively with a mutation of a single gene, brkA, results in a severe defect in vivo. Although BrkA has been shown to be required for B. pertussis to resist complement-mediated killing in vitro, the relevance of these findings to the in vivo role of BrkA during infection has not been examined. Transducing this mutation into multiple wild-type B. pertussis strains allowed us to confirm the in vitro phenotype of reduced resistance to serum complement. All DeltabrkA mutants were increased in their sensitivity to complement in vitro, both in the presence and absence of antibodies. However, these strains differed substantially in their phenotypes in vivo. DeltabrkA mutants of recent clinical isolates were indistinguishable from wild-type strains in their efficient infection of respiratory organs, suggesting that the function of BrkA in these strains is noncritical or redundant. In contrast, multiple DeltabrkA strains derived from Tohama I were severely defective during the first week postinoculation compared to their wild-type parent. This defect was present even in complement-deficient mice, revealing a complement-independent phenotype for the DeltabrkA mutant in respiratory tract infection.     

The Schizosaccharomyces pomberfc3 + gene encodes a homologue of the human hRFC36 and Saccharomyces cerevisiae Rfc3 subunits of replication factor C

In the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe replication factor C (RF-C) plays key roles both in chromosomal DNA replication and in DNA replication checkpoint function. At the replication fork, the five-subunit RF-C complex functions to load the trimeric polymerase accessory factor PCNA onto DNA. PCNA then acts as a sliding clamp, tethering Pol δ to the DNA to maximise its processivity. Here we describe the cloning of the S. pomberfc3 ⁺ gene, encoding a homologue of the S. cerevisiae Rfc3 and human hRFC36 proteins. The 1026 bp rfc3 ⁺ ORF is interrupted by five introns, ranging in size from 49 to 165 bp. The spliced ORF is predicted to encode a 342 amino-acid protein that is approximately 50% identical at the amino acid sequence level to the S. cerevisiae Rfc3 and human hRFC36 proteins. As expected, S. pomberfc3 ⁺ is an essential gene, with rfc3Δ cells being defective for DNA replication. Loss of rfc3 ⁺ function can be rescued by heterologous expression of either the S. cerevisiae Rfc3 or human hRFC36 proteins in S. pombe. Received: 15 October 1999     

Evaluation of dry and wet transported intravaginal swabs in detection of Chlamydia trachomatis and Neisseria gonorrhoeae infections in female soldiers by PCR

Screening women for sexually transmitted diseases (STD) in nonclinic settings is highly desirable because many infections are asymptomatic. This is especially true for military women, for whom logistical, social, and other job-related obstacles present barriers to accessing medical care. We assessed the accuracy of intravaginal swabs transported by mail in a wet versus a dry state for PCR (Amplicor CT/NG test) detection of chlamydia and gonorrhea infections in a cross-sectional study of 793 active-duty military women attending an STD clinic. PCR tests of vaginal swabs (wet and dry) were compared to local clinical methods used on cervical swabs. Standard wet vaginal swab PCR testing detected more chlamydia (11.6%) than cervical enzyme immunoassay (9.3%). For detection of chlamydia using wet swabs, the sensitivity and specificity compared with adjudicated true positives were 94.6% (87 of 92) and 99.3% (696 of 701), respectively. Comparing dry swabs to true-positives for chlamydia, the sensitivity was 91.3% (84 of 92) and the specificity was 99.3% (696 of 701). Standard wet vaginal swab PCR detected more gonorrhea (3.3%) than routine cervical culture (2.1%). The sensitivity and specificity of PCR testing of wet swabs compared to true-positives (infected patients) were 96.3% (26 of 27) and 98.2% (752 of 766) for gonorrhea, respectively. For gonorrhea, the sensitivity and specificity of dry swabs compared to true-positives (infected patients) were 88.9% (24 of 27) and 98.3% (753 of 766), respectively. PCR testing of wet and dry transported intravaginal swabs to detect chlamydia and gonorrhea infections was an accurate diagnostic method for military women.     

Dysmorphic features and learning disability in an adult male with pure partial trisomy 17q24-q25 due to a terminal duplication

We describe an adult male with severe learning disability, epilepsy, and dysmorphic features. Cytogenetic studies demonstrated a terminal duplication of the long arm of chromosome 17, resulting in partial trisomy 17q24-q25. Our patient shows some of the characteristic features of the distal 17q phenotype, but in addition has more unusual features such as epilepsy, sensorineural hearing loss, and long fingers and overlapping toes. We suggest that these features occur with terminal duplications of 17q.     

Maize NDC80 is a constitutive feature of the central kinetochore

In yeast and animals, Nuclear Division Cycle 80 (NDC80) is an important kinetochore protein that binds to microtubules and mediates chromosome movement. Its localization pattern is unusual, since it is generally not viewed as either an inner (centromeric chromatin) or outer (regulatory) component of the kinetochore. Here we report the characterization of NDC80 in a higher plant. By taking advantage of the large meiotic kinetochores of maize, we were able to show that NDC80 localizes outside of the constitutive kinetochore protein CENP-C. Further, a detailed analysis of mitosis indicates that NDC80 is stably present on kinetochores throughout the cell cycle. The quantity of NDC80 positively correlates with measured quantities of DNA and CENP-C, suggesting that NDC80 rapidly associates with DNA following replication and is stably maintained at centromeres during cell division. The data suggest that in plants NDC80 is on par with ‘foundation’ kinetochore proteins such as CENH3 and CENP-C.     

In vitro anti-anaerobic activity of the cephalosporin derivative RWJ 54428, compared to seven other compounds

Agar dilution MIC was used to test the activity of RWJ 54428, a new cephalosporin derivative, compared to imipenem, meropenem, ceftriaxone, piperacillin, piperacillin–tazobactam, clindamycin and metronidazole against 363 anaerobes isolated from clinical specimens. RWJ 54428 had low MICs against most β -lactamase-negative Gram-negative rods, and all Gram-positive strains except Clostridium difficile . Imipenem and meropenem had the lowest MICs (MIC₅₀s of 0.125 mg/L and MIC₉₀s of 1.0 mg/L). Piperacillin–tazobactam, clindamycin and metronidazole were active against most strains, and ceftriaxone was active mainly against β -lactamase-negative organisms.     

Effects of 2 ankle fatigue models on the duration of postural stability dysfunction

Context: Muscle fatigue is generally categorized in 2 ways: that caused by peripheral weakness (peripheral fatigue) and that caused by a progressive failure of voluntary neural drive (central fatigue). Numerous variables have been studied in conjunction with fatigue protocols, including postural stability, maximum voluntary contraction force, and reaction time. When torque recordings fall below 50% of a maximum voluntary contraction, the muscle is described as fatigued, but whether this value is a good indicator of fatigue has not been studied.Objective: To compare the effects of 2 ankle musculature fatigue protocols (30% and 50%) on the duration of postural stability dysfunction.Design: To assess differences between the 30% and 50% fatigue protocols, we calculated a 1 between-groups factor (subjects) and 2 within-groups factors (fatigue, test) analysis of variance.Setting: E.J. Nutter Athletic Training Facility.Patients or Other Participants: Twenty subjects (10 men, 10 women; age = 21.15 ± 2.23 years; height = 172.97 ± 9.86 cm; mass = 70.62 ± 14.60 kg) volunteered for this study. Subjects had no history of lower extremity injury, vestibular or balance disorders, functional ankle instability, or head injury in the past 6 months.Intervention(s): On separate days, subjects performed isokinetic fatiguing contractions of the plantar flexors and dorsiflexors in a 30% protocol (70% decrease in strength) and a 50% protocol (50% decrease in strength).Main Outcome Measure(s): Baseline and postfatigue postural stability scores were determined before and after the isokinetic fatiguing contractions. Plantar-flexion peak-torque measurements were obtained for the 2 fatiguing protocols. Three prefatigue and 12 postfatigue postural stability trials were recorded. Velocities for testing were 60°/s for plantar flexion and 120°/s for dorsiflexion.Results: Sway velocity was significantly greater when the ankle was fatigued to 30% (1.56°/s) than in the 50% condition (1.36°/s). For the 30% protocol, sway was significantly impaired when the pretest condition (1.19°/s) was compared with posttest trial 1 (2.34°/s), trial 2 (2.37°/s), and trial 3 (1.71°/s). For the 50% protocol, sway was significantly impaired when the pretest condition (1.27°/s) was compared with posttest trial 1 (2.02°/s).Conclusions: The 30% fatigue protocol resulted in significantly longer impairment of postural stability than the 50% protocol. Because the 30% protocol resulted in a greater effect but was relatively short-lived (approximately 75 to 90 s), it is more useful for research purposes.     

Organization of auditory cortex in the albino rat: sound frequency

1. Responses of neurons in the auditory cortex of the albino rat were examined using microelectrode mapping techniques. Characteristic frequencies were determined for numerous electrode penetrations across the cortical surface in individual animals. A primary auditory area was identified in the posterolateral neocortex that was characterized by short latency responses to tone bursts and tonotopic organization with high frequencies represented rostrally and low frequencies, caudally. Within this area cells with similar characteristic frequencies were aligned in a dorsoventral orientation to form isofrequency contours. 2. Tuning curves obtained from primary auditory cortex were characteristically "V" shaped with Q10's ranging from 0.97 to 28.4. Maximum Q10 values increased monotonically with characteristic frequency (CF). The lowest thresholds at CF closely approximated the behavioral audiogram for the albino rat. Many neurons, however, had CF thresholds well above the behavioral limit. 3. Areas were found dorsal and ventral to the primary auditory cortex in which CF's were clearly discontinuous with the neighboring isofrequency contours. These data suggest the presence of other auditory fields, the detailed characteristics of which have yet to be examined.     

Topographic organization of projections from the amygdala to the visual cortex in the macaque monkey

The topography of amygdaloid projections to the visual cortices in the macaque monkey was examined by injecting the fluorescent tracers Fast Blue and Diamidino Yellow at different locations in the occipital and temporal lobes and mapping the distribution of retrogradely labeled cells in the amygdala. Injections involving regions from rostral area TE to caudal area V1 all resulted in labeled cells within the basal nucleus of the amygdala. Relatively few double-labeled cells were observed even when the two injections were separated by less than 3 mm. The projections were rostrocaudally organized such that projections to caudal visual areas originated from dorsal and caudal portions of the magnocellular division of the basal nucleus while projections to more rostrally situated visual areas originated in more rostral and ventral portions of the basal nucleus. When injections involved rostral and medial portions of area TE, retrogradely labeled cells were observed in the accessory basal and lateral nuclei in addition to the basal nucleus. These data confirm that the amygdala gives rise to feedback projections to all levels of the "ventral stream" visual pathway. The projections do not appear to be diffusely distributed since few double-labeled cells were observed. The largest cells of the basal nucleus, those located in the magnocellular division, project the farthest in the visual system and innervate all occipital and temporal levels. The smaller cells, in the intermediate and parvicellular regions, project to more rostral and medial portions of the visual cortex. These results suggest that the amygdala may have substantial modulatory control over sensory processing at all stages of the ventral-stream visual cortical hierarchy.     

Immunosuppression by seminal prostaglandins.

In this paper we report studies undertaken to determine the contribution of seminal prostaglandins to some of the known immunosuppressive properties of human seminal plasma. Initial studies revealed that fractions of seminal plasma enriched in E series prostaglandins, obtained by reverse phase chromatography, had a pronounced inhibitory effect on the PHA-induced proliferation of peripheral blood lymphocytes and on the NK-cell-mediated lysis of K562 target cells. Additional investigations revealed that similar inhibitory effects could be achieved with purified PGE2 (10(-6) to 10(-9) M) and 19-OH PGE1 (10(-6) to 10(-7) M), both of which are present in uniquely high concentrations in human seminal plasma. In contrast, 19-OH PGF1 which is found in lower concentrations in semen was slightly stimulatory in proliferative assays and had no effect on NK-cell-mediated cytotoxicity. Removal of the seminal prostaglandins by absorption chromatography resulted in a dramatic decrease in immune suppressive activity. Further studies with fractions obtained by ion-exchange HPLC of desalted seminal plasma indicated that prostaglandins complexed with seminal proteins, and these too were immunosuppressive. The possible relevance of these results to sexually transmitted disease is discussed.