Discovery and Contribution of Nontypeable Haemophilus influenzae NTHI1441 to Human Respiratory Epithelial Cell Invasion.

Nontypeable Haemophilus influenzae (NTHi) is the primary cause of bacterially induced acute exacerbations of chronic obstructive pulmonary disease (COPD). NTHi adheres to and invades host respiratory epithelial cells as a means to persist in the lower airways of adults with COPD. Therefore, we mined the genomes of NTHi strains isolated from the airways of adults with COPD to identify novel proteins to investigate their role in adherence and invasion of human respiratory epithelial cells. An isogenic knockout mutant of the open reading frame NTHI1441 showed a 76.6% ± 5.5% reduction in invasion of human bronchial and alveolar epithelial cells at 1, 3, and 6 h postinfection. Decreased invasion of the NTHI1441 mutant was independent of either intracellular survival or adherence to cells. NTHI1441 is conserved among NTHi genomes. Results of whole-bacterial-cell enzyme-linked immunosorbent assay (ELISA) and flow cytometry experiments identified that NTHI1441 has epitopes expressed on the bacterial cell surface. Adults with COPD develop increased serum IgG against NTHI1441 after experiencing an exacerbation with NTHi. This study reveals NTHI1441 as a novel NTHi virulence factor expressed during infection of the COPD lower airways that contributes to invasion of host respiratory epithelial cells. The role in host cell invasion, conservation among strains, and expression of surface-exposed epitopes suggest that NTHI1441 is a potential target for preventative and therapeutic interventions for disease caused by NTHi.

Awareness and Preventative Behaviours Regarding Toxoplasma, Listeria and Cytomegalovirus Among Pregnant Women

Aim Infection with toxoplasma, listeria and cytomegalovirus (CMV) infections can negatively affect pregnancy outcomes. Awareness of these infections, knowledge of preventive practices and pertinent behaviours was assessed. Methods A survey of patients at antenatal clinics in a Dublin maternity hospital was conducted over a six month period. Analysis was undertaken using SPSS. Results The response rate was 36% (287/800). One in two respondents were aware of toxoplasma, one in four of Listeria but only 1 in 10 of CMV. Overall, knowledge was highest among older, more educated pregnant women. Nativity had a significant effect on knowledge and behaviour, but increasing parity did not. However the majority practised key safe behaviours. Conclusion Information must be conveyed to pregnant women in a user friendly format and in a culturally sensitive way.

Ponderal index (PI) vs birth weight centiles in the low-risk primigravid population: which is the better predictor of fetal wellbeing?

Our objective was to compare Ponderal index (PI) with birth weight centiles as predictors of perinatal morbidity and to determine which best reflects the presence of placental disease. We prospectively recruited 1,011 low-risk primigravidas and calculated PI and birth weight centiles following delivery. Perinatal morbidity was defined as: pre-term birth (PTB); fetal acidosis; an Apgar score <7 at 5 min or neonatal resuscitation. Placental disease was defined as chronic uteroplacental insufficiency (CUPI); villous dysmaturity; infection or vascular pathology. Ponderal index was statistically reduced (25.33 vs 27.79 p =0.001) and the incidence of infant birth weight <9th centile was statistically higher (11.1% vs 5.1%; p =0.004) in cases with PTB and in CUPI (26.23 vs 27.84; p =0.001 and 28.2.1% vs 10.4%; p =0.002). Both PI and infant birth weight centile <9th centile for gestational age correlate with PTB, however overall, both are poor predictors of neonatal and placental disease.

Time to positivity (TTP) of neonatal blood cultures: a trend analysis over a decade from Ireland.

Background and aims: Blood culture (BC) remains gold standard for the evaluation and diagnosis of neonatal sepsis. Time when BC becomes positive and the type of microorganism isolated are crucial in deciding the antimicrobial management. Likely pathogenicity of organisms growing in BC could potentially be predicted based on the "time to positivity" (TTP). We aimed to estimate the predictive value of isolating a likely pathogenic organism depending on TTP; evaluate the aetiological trend and neonatal mortality rate due to culture-proven neonatal sepsis for over a decade and verify whether the application of a "36 hour rule" to discontinue empiric antibiotics in well newborn infants with negative BC would be safe.Methods: Retrospective review of BC results over a 14-year period from a regional neonatal unit in Ireland. Laboratory data were independently extracted in relation to BC results from the laboratory information management system (LIMS-iLAB). Neonatal mortality data were collected from multiple sources. Statistical analysis included logistic regression, chi-square, and Mann-Whitney U-test.Results: Over a 14-year period 11,432 neonatal BC specimens were incubated of which 605 (5.3%) turned positive. Overall, the commonest organism grown was coagulase-negative Staphylococcus (CoNS), 416 (68.8%). Main pathogenic organisms were Staphylococcus aureus 23 (3.8%), Enterococcus spp. 22 (3.6%), E.coli 21 (3.5%), group B Streptococcus (GBS) 18 (3.0%), and Klebsiella species 9 (1.5%). Gram-negative organisms had the shortest TTP, with Klebsiella spp. having a median TTP of 10 h and E. coli 11 h. For Gram-positives, GBS had a median TTP of 12 h, Enterococcus species 14 h, with S. aureus growing at a median time of 15 h. All of the Klebsiella spp. and other Coliforms were detected within 24 h, with, 95.2% of E.coli, 94.4% of GBS, 95.5% of Enterococci, and 95.7% of S. aureus, flagging positive in 24 h. Using logistic regression the omnibus test of the coefficients in the resulting model was significant (p < .001). Our observed coefficient (ß) for TTP was 0.144; shorter the TTP higher was the likelihood of isolating a pathogenic organism, with an odds ratio (OR) of 1.155. We also report a relatively low blood culture proven sepsis-specific neonatal mortality rate of 0.403/1000 live births and in all such instances observed TTP was less than 24 h.Conclusion: Duration of this study exceeds that of most of the neonatal blood culture TTP analysis published to-date. A shorter TTP is an important adjunct to suggest the growth of a pathogenic organism while managing suspected neonatal sepsis. TTP if < 24 h per se would not necessarily confirm the growth of a highly pathogenic organism; however, if a positive growth is likely to happen for a significant neonatal pathogen, in more than 98% the TTP would be within 24 h. This offers the clinician more of negative predictive value than a positive one; when there is no growth in BC. Our observation on TTP reiterate the National Institute of Health and Care Excellence (NICE) guideline of discontinuation of empiric antibiotics after 36 h in and clinically well and BC negative newborn infants.

The duration of gonadotropin stimulation does not alter the clinical pregnancy rate in IVF or ICSI cycles.

BACKGROUND: Ovarian stimulation is an essential part of assisted reproduction treatments. Research on whether the duration of stimulation alters the success in assisted reproduction has not been conclusive. AIM: The purpose of the study was to establish whether the duration of ovarian stimulation alters the success in assisted reproduction treatments. METHODS: All fresh (non-donor) stimulation cycles performed in an academic tertiary referral ART centre over a period of 18 years, between 1st January 1997 and 31st December 2014, were identified. Data were prospectively and electronically collected. IVF and ICSI cycles were analysed independently. Each category was then subdivided into assisted reproduction cycles where the antagonist, long (down regulation) and flare protocol were used. Clinical pregnancy was the main outcome measured. A total of 10,478 stimulation cycles (6011 fresh IVF and 4467 fresh ICSI) reaching egg collection were included. RESULTS: We showed no significant difference in CP rates in IVF cycles for the long (p = 0.082), antagonist (p = 0.217) or flare (p = 0.741) protocol cycles or in ICSI cycles with the long (p = 0.223), antagonist (p = 0.766) or the flare (p = 0.690) protocol with regards the duration of stimulation. CONCLUSION: The duration of stimulation does not alter the CP rate in ICSI or IVF cycles using the long, antagonist or flare stimulation protocol.

Coagulation indices in very preterm infants from cord blood and postnatal samples.

BACKGROUND: Very premature infants are at high risk of bleeding complications; however, few data exist on ranges for standard coagulation tests. OBJECTIVES: The primary objective of this study was to measure standard plasma coagulation tests and thrombin generation in very premature infants compared with term infants. The secondary objective was to evaluate whether an association existed between coagulation indices and intraventricular hemorrhage (IVH). PATIENTS/METHODS: Cord and peripheral blood of neonates < 30 weeks gestational age (GA) was drawn at birth, on days 1 and 3 and fortnightly until 30 weeks corrected gestational age. Prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen and coagulation factor levels were measured and tissue factor-stimulated thrombin generation was characterized. Control plasma was obtained from cord blood of term neonates. RESULTS: One hundred and sixteen infants were recruited. Median (range) GA was 27.7 (23.7-29.9) weeks and mean (SD) birth weight was 1020 (255) g. Median (5th-95th percentile) day 1 PT, APTT and fibrinogen were 17.5 (12.7-26.6) s, 78.7 (48.7-134.3) s and 1.4 (0.72-3.8) g L(-1) , respectively. No difference in endogenous thrombin potential between preterm and term plasma was observed, where samples were available. Levels of coagulation factors II, VII, IX and X, protein C, protein S and antithrombin were reduced in preterm compared with term plasma. Day 1 APTT and PT were not associated with IVH. CONCLUSION: In the largest cross-sectional study to date of very preterm infants, typical ranges for standard coagulation tests were determined. Despite long clotting times, thrombin generation was observed to be similar in very preterm and term infants.

An in vitro model of innate lymphoid cell function and differentiation.

Innate lymphoid cells (ILC) are RAG-independent lymphocytes with important roles in innate immunity, and include group-1 (natural killer (NK) cell, ILC1), group-2 (ILC2), and group-3 (lymphoid tissue inducer (LTi), NCR(+) ILC3) subsets. Group-3 ILC express Rorγt, produce interleukin (IL)-22, and are critically important in the normal function of mucosal tissues. Here, we describe a novel model cell line for the study of ILC function and differentiation. The parental MNK cell line, derived from NKR-P1B(+) fetal thymocytes, shows a capacity to differentiate in γc cytokines. One IL-7-responsive subline, designated MNK-3, expresses Rorγt and produces high levels of IL-22 in response to IL-23 and IL-1ß stimulation. MNK-3 cells display surface markers and transcript expression characteristic of group-3 ILC, including IL-7Rα (CD127), c-kit (CD117), CCR6, Thy1 (CD90), RANK, RANKL, and lymphotoxin (LTα1ß2). Using an in vitro assay of LTi cell activity, MNK-3 cells induce ICAM-1 and VCAM-1 expression on stromal cells in a manner dependent upon LTα1ß2 expression. A second IL-2-responsive subline, MNK-1, expresses several NK cell receptors, perforin and granzymes, and shows some cytotoxic activity. Thus, MNK-1 cells serve as a model of ILC1/NK development and differentiation, whereas MNK-3 cells provide an attractive in vitro system to study the function of ILC3/LTi cells.

A randomized, double-blind, placebo-controlled, cross-over trial to assess the side effects of medroxyprogesterone acetate in hormone replacement therapy.

Cyclic progestin therapy has been widely advocated as an adjunct to postmenopausal estrogen replacement therapy to reduce the risk of endometrial carcinoma. Acceptance of this approach, however, appears to have preceded detailed evaluation of possible adverse side effects of progestins that could result in patient noncompliance. We evaluated the nonmenstrual physical and psychological side effects of oral medroxyprogesterone acetate given in conjunction with transdermal estrogen in two groups of women with previous hysterectomy and oophorectomy. Twenty-four women with prospectively documented severe premenstrual syndrome (PMS) before surgery and 24 women with no such history of adverse premenstrual changes received transdermal estrogen 100 micrograms on days 1-25 and either oral medroxyprogesterone acetate 10 mg daily or an identical placebo (days 12-25) in a randomized, double-blind, cross-over design. Mood and physical symptoms were monitored prospectively, using daily self-ratings on the Daily Symptoms Checklist. The Beck Depression Inventory and Premenstrual Tension Self-Rating Scale were completed on day 24. At the study's completion, the patients were asked which treatment period they preferred. Paired comparisons did not reveal any significant differences, and preference for treatment was equally divided between medroxyprogesterone acetate and placebo. We conclude that addition of medroxyprogesterone acetate 10 mg/day for 14 days to cyclic transdermal estrogen therapy (days 1-25) produces no consistent adverse physical or psychological effects on women for one cycle of treatment, regardless of their PMS history.

Relative risks of oral contraceptive usage.

PIP: Women seeking safe, reliable contraception must weigh the risks associated with such methods with their benefits. To make this assessment process more meaningful, the authors have prepared a chart that sets forth the risks of oral contraceptive (OC) use by different user factors (e.g., age, cigarette smoking) in proportion to the risks inherent in a variety of sporting activities and types of accidents. Mortality rates reported for OC use include those due to the method and those due to pregnancies resulting from method failure. Scrutiny of this chart reveals that the risk of death associated with swimming, boating, and automobile use exceeds the risk of death for nonsmokers who use OCs up to 34 years of age (1.6 deaths/100,000 population). OC use by women 15-24 years of age who smoke carries a lower mortality risk (3.0/1000) than scuba diving or falling, while OC users 25-34 years old who smoke are more likely to die as a result of homicide, suicide, or automobile accidents than they are as a result of their use of OCs (mortality rate, 10.2/1000). On the other hand, OC use by smokers 35-44 years of age is associated with a mortality rate of 84.5/1000, which is a higher risk than that associated with the sport of hang-gliding.^ieng

Characteristics of Escherichia coli isolated from septic foals.

Fifteen Escherichia coli isolates from the blood and tissue of foals with septicemia were compared with 15 from the feces of clinically normal horses. Comparisons were made with respect to survival in normal equine serum, production of aerobactin, and production of hemolysin. Isolates from the blood and tissues of septic foals were more likely to be resistant to equine serum than were isolates from feces of clinically normal horses. There were minimal differences between the isolates with respect to aerobactin and hemolysin production, almost all being nonhemolytic and aerobactin negative. Serum resistance is probably a virulence determinant for invasive E. coli in horses.

Antimicrobial susceptibility patterns of the A and T types of Pasteurella haemolytica.

Minimal concentrations of penicillin, ampicillin, cephalothin, chloramphenicol, tetracycline, erythromycin, nitrofurantoin, kanamycin and gentamicin inhibitory to Pasteurella haemolytica biotypes A and T were determined. They were found to be significantly higher for T than for A type strains in the case of all the antimicrobial drugs tested except the two aminoglycosides kanamycin and gentamicin, for which no differences were observed. In the case of penicillin the differences were so marked that they may be useful as a basis for biotyping P haemolytica isolates. The emergence of antibiotic-resistant strains observed in recent years, however, may limit such application.

Linkage of serum resistance, aerobactin production, and resistance to antimicrobial agents on conjugal plasmids in some strains of Escherichia coli isolated from septic foals.

Fifteen isolates of Escherichia coli obtained from the blood and tissues of septic foals had plasmid DNA of size ranging from 2.5 to 93 megadaltons. These isolates grew in normal equine serum (serum resistant), a trait previously documented to be expressed by isolates obtained from blood and tissues of septic foals, but not by isolates obtained from the feces of clinically normal horses. Of these isolates, 3 contained conjugal plasmids that encoded resistance to multiple antimicrobial agents linked to serum resistance and, in 1 isolate, to production of aerobactin as well. Serum resistance and production of aerobactin are related to virulence of septicemic E coli from non-equine sources.

Myocardial assessment using tissue doppler imaging in preterm very low-birth weight infants before and after red blood cell transfusion.

OBJECTIVE: To investigate myocardial velocities in anemic very low-birth weight (VLBW) preterm infants, pre and post red blood cells transfusion using tissue Doppler imaging echocardiography. STUDY DESIGN: Forty-eight VLBW preterm infants34 weeks and>2 weeks old were prospectively divided: Transfused symptomatic infants (Hematocrit (Hct)<0.30 (n=32)) and non transfused asymptomatic controls (control 1, Hct >0.30 (n=9) and control 2, Hct <0.30 (n=7)). Echocardiography was performed before and 3-5 days after transfusion in the transfused, and the controls were studied at similar intervals. Non parametric tests were used for statistical analysis. RESULT: Left ventricular (LV) systolic velocity increased (transfused (4.6±0.70 vs 6.0±0.65, P<0.01)) as did LV diastolic velocities (P<0.01) without significant difference over time in each control. The percentage change in LV velocity following transfusion correlated negatively (ρ=0.36) with pre transfusion Hct. CONCLUSION: There is a significant increase in myocardial performance following transfusion, which is related to the severity of the anemia.

The British Columbia Positive Women's Survey: a detailed profile of 110 HIV-infected women.

OBJECTIVE: To describe the health, social environment, medical care received and satisfaction with medical care of HIV-infected women in British Columbia. DESIGN: Self-administered 75-item questionnaire distributed by mail or in person between March 1994 and February 1996 through community AIDS organizations and physicians' offices. SETTING: British Columbia. PARTICIPANTS: A total of 110 HIV-positive women. OUTCOME MEASURES: Sociodemographic data, risk factors for HIV infection, details about HIV testing, health status and medical treatment, use of health care services, degree of satisfaction with medical care and psychosocial stressors. RESULTS: Most of the women surveyed were aged 25 to 39 years (70.0%), were Canadian born (76.4%) and were white (80.9%). Over one-third did not complete high school, and half had an annual household income of less than $20,000. Of the 110 women 51.8% had children, who were HIV-positive in 12.3% of cases. The most frequently reported risk factor for HIV infection was sex with a man (49.1%); 19.1% reported both sex with a man and injection drug use, and 12.7% reported injection drug use only. Seventy-five women indicated that they had become infected through sex with a man, with or without injection drug use. Of these, 65 indicated whether or not this was the result of sexual assault or rape; 8 (12.3%) answered affirmatively. Of the 81 women who responded to the question regarding prior sexual assault or abuse, 43 (53.1%) reported being sexually assaulted as an adult, 35 (43.2%) reported being sexually abused as a child, and 22 (27.2%) reported being sexually abused or assaulted both as a child and as an adult. Women who were sexually abused as a child were more likely than those who were not abused as a child to have injection drug use as a risk factor (54.3% v. 7.5%). Menstrual cycle changes were reported by 70.1% of the respondents. Most women stated that they had not received adequate pre- or post-test counselling, and 47.0% were not satisfied with their doctor's care. Psychosocial concerns identified to be of greatest importance were financial problems, lack of intimacy or satisfying sexual relationship, and fear of rejection or discrimination. CONCLUSION: Several important concerns for HIV-positive women were identified, including dissatisfaction with medical care, fear of discrimination, violence and abuse, and poverty.

Maternity Care Guidelines checklist. To assist physicians in implementing CPGs.

PROBLEM BEING ADDRESSED: Implementing the recommended clinical practice guidelines for prenatal care can be difficult for busy practitioners because the guidelines are numerous and continually being revised. OBJECTIVE OF PROGRAM: To develop a checklist outlining the current recommended activities for prenatal care to assist practitioners in providing evidence-based interventions to pregnant women. MAIN COMPONENTS OF PROGRAM: We reviewed guidelines for prenatal care from the Canadian Task Force on the Periodic Health Examination (CTFPHE) and from the report of the US Preventive Services Task Force (USPSTF). We searched MEDLINE for interventions commonly performed in pregnancy, but not reviewed by either task force. Interventions graded A or B are listed in bold type on the checklist. Interventions graded C by either task force or recommended by organizations not necessarily using the same rigorous criteria are listed in plain type. Recommended interventions are displayed along a time line under three headings: clinical maneuvers, investigations, and issues for discussion. Pilot testing by 12 practising physicians and 12 family practice residents showed that most respondents thought the checklist very useful. CONCLUSIONS: Providing a one-page checklist summarizing recommended clinical maneuvers, investigations, and topics for discussion should help physicians with implementing the many clinical practice guidelines for prenatal care.

The major heat-modifiable outer membrane protein CD is highly conserved among strains of Branhamella catarrhalis.

The outer membrane of Branhamella catarrhalis contains a major, heat-modifiable outer membrane protein called CD which has epitopes on the surface of the intact bacterium. The gene encoding CD was cloned and expressed in Escherichia coli. The protein migrates in gels as a doublet, indicating that CD is encoded by single gene whose gene product has two stable conformations. The nucleotide sequence of the gene encoding CD was determined and shows homology with the OprF outer membrane protein of Pseudomonas species. The CD protein contains a proline-rich region, which appears to account for its aberrant migration in gels. Restriction fragment-length analysis of 30 isolates of B. catarrhalis with oligonucleotide probes corresponding to sequences in the CD gene produced identical patterns in Southern blot assays. The major heat-modifiable outer membrane protein CD shares homology with the OprF protein and is highly conserved among strains of B. catarrhalis.

Landing patterns in netball: analysis of an international game.

The passing and landing patterns among netball players were investigated by examining all the 595 passes in an international match between Australia and New Zealand. Intra-class correlations were used to test inter-observer reliability. The independence of two variables was tested using a Pearson chi 2 test and a contingency coefficient was used to assess the degree of association between variables. An inter-rater reliability in the identification of movement patterns was 0.99. Most of the landings observed in this match were on the forefoot and not the hindfoot as generally reported in controlled experiments. Only 14% of the players leapt to receive a pass compared with 76% reported earlier under controlled environments. The distribution of passing techniques unexpectedly showed that approximately 50% of all the players did not reach for the ball as it was thrown directly to them. However, over 70% of the players threw straight passes which were mainly received in the chest quadrant and other types of passes such as loop, bounce and rebound were used less frequently. Nearly 50% of all players used a right-handed pass to dispose of the ball, while the next most popular pass was to use both hands (40.2%). Landing and passing patterns differed between players in different positions. Footfall patterns on landing after a catch were associated with the trajectory of the ball.

Analysis of antigenic structure and human immune response to outer membrane protein CD of Moraxella catarrhalis.

Moraxella catarrhalis is an important cause of otitis media in children and lower respiratory tract infections in adults with chronic obstructive pulmonary disease (COPD). Outer membrane protein CD (OMP CD) is a 45-kDa protein which is a potential vaccine antigen to prevent infections caused by M. catarrhalis. Eight monoclonal antibodies were used to study the antigenic structure of the OMP CD molecule by assaying recombinant peptides corresponding to the sequence of the protein. This approach identified two surface-exposed epitopes, including one near the amino terminus (amino acids 25 to 44) and one in the central region of the molecule (amino acids 261 to 331). Assays with serum and sputum supernatants of adults with COPD revealed variable levels of antibodies to OMP CD among individuals. To determine which portions of the OMP CD molecule were recognized by human antibodies, three human serum samples were studied with six recombinant peptides which span the sequence of OMP CD. All three sera contained immunoglobulin G antibodies which recognized exclusively the peptide corresponding to amino acids 203 to 260 by immunoblot assay. Adsorption experiments with whole bacteria established that some of the human antibodies are directed at surface-exposed epitopes on OMP CD. We conclude that OMP CD is a highly conserved molecule which contains at least two separate epitopes which are exposed on the bacterial surface. While individual adults with COPD show variability in the immune response to OMP CD, a specific region of the OMP CD molecule (amino acids 203 to 260) is important as a target of the human immune response.