Tetrazolium reduction allows assessment of biofilm formation by Campylobacter jejuni in a food matrix model.

AIMS: To develop a staining method for specific detection of metabolically active (viable) cells in biofilms of the foodborne pathogen Campylobacter jejuni. METHODS AND RESULTS: Conversion of 2,3,5 triphenyltetrazolium chloride (TTC) to insoluble, red 1,3,5-triphenylformazan (TPF) was dependent on metabolic activity of Camp. jejuni. When used with chicken juice, TTC staining allowed quantification of Camp. jejuni biofilm levels, whereas the commonly used dye, crystal violet, gave high levels of nonspecific staining of food matrix components (chicken juice). The assay was optimized to allow for monitoring of biofilm levels and adapted to monitor levels of Camp. jejuni in broth media. CONCLUSIONS: Staining with TTC allows for the quantification of metabolically active Camp. jejuni and thus allows for quantification of viable cells in biofilms and food matrices. The TTC staining method can be adapted to quantify bacterial cell concentration in a food matrix model, where the accepted method of A600 measurement is not suitable due to interference by components of the food matrix. SIGNIFICANCE AND IMPACT OF THE STUDY: 2,3,5 Triphenyltetrazolium chloride (TTC) staining is a low-cost technique suitable for use in biofilm analysis, allowing rapid and simple imaging of metabolically active cells and increasing the methods available for biofilm assessment and quantification.

Cervicovaginal fistula presenting during miscarriage.

Cervicovaginal fistula is a recognized complication of induced mid-trimester termination of pregnancy, but more recently it has also been recognized as representing a complication of prior cervical cerclage. We report the ultrasound findings of prolapse of the amniotic sac through a cervicovaginal fistula in a woman with prior cervical cerclage. A woman with cervical incompetence and prior failed McDonald cerclage presented for prophylactic Shirodkar cerclage. Before the procedure, transvaginal ultrasonography revealed a live intrauterine pregnancy at 14 weeks' gestation. Upon further ultrasound examination, the amniotic sac appeared to protrude through the posterior wall of the cervix into the vaginal vault. Pelvic examination verified prolapse of the amniotic sac through a cervicovaginal fistula. The patient underwent an uncomplicated dilatation and evacuation. Women with a history of cervical cerclage are at risk for the development of cervicovaginal fistula, the detection of which is important to prevent potential morbidity.

Quantifying regeneration in patients following peripheral nerve injury.

Healthy nerve function provides humans with the control of movement; sensation (such as pain, touch and temperature) and the quality of skin, hair and nails. Injury to this complex system creates a deficit in function, which is slow to recover, and rarely, if ever, returns to what patients consider to be normal. Despite promising results in pre-clinical animal experimentation effective translation is challenged by a current inability to quantify nerve regeneration in human subjects and relate this to measurable and responsible clinical outcomes. In animal models, muscle and nerve tissue samples can be harvested following experimental intervention. This allows direct quantification of muscle mass and quality and quantity of regeneration of axons; such an approach is not applicable in human medicine as it would ensure a significant functional deficit. Nevertheless a greater understanding of this process would allow the relationship that exists between neural and neuromuscular regeneration and functional outcome to be more clearly understood. This article presents a combined commentary of current practice from a specialist clinical unit and research team with regard to laboratory and clinical quantification of nerve regeneration. We highlight how electrophysiological diagnostic methods (which are used with significant recognised limitations in the assessment of clinical medicine) can potentially be used with more validity to interpret and assess the processes of neural regeneration in the clinical context, thus throwing light on the factors at play in translating lab advances into the clinic.

Degradation of proteoglycan in articular cartilage.

Adult rabbit articular cartilage was labelled in vivo over 48 h with [35S]sulphate and was then incubated in organ culture at pH 7.2. Approx. 65% of the tissue content of [35S]proteoglycan was released into the culture medium during the first 48 h of incubation. The average molecular size of the released proteoglycans, as assessed by fractionation on Sepharose 2B/CL and 4B/Cl, was only slightly smaller than that of the proteoglycans extracted from non-cultured cartilage with 4 M guanidine HCl. The percentage of released proteoglycans and extracted proteoglycans which formed aggregates with hyaluronic acid was approx. 25% and 75%, respectively. The results indicate that proteoglycan degradation in adult articular cartilage is initiated by a limited proteolysis of subunit core protein, with the production of non-aggregating species which diffuse readily from the tissue.

Quantitation of the five forms of plasma high molecular weight angiotensinogen in women with pregnancy-induced hypertension.

In the human pregnant state a high molecular weight form of angiotensinogen (HMrA) is present in significant quantities in addition to the usual low molecular weight angiotensinogen (LMrA). In a previous study involving a small number of white women, it was found that women who had developed pregnancy-induced hypertension (PIH) had significantly higher levels of plasma HMrA. It has been determined that there are five isoforms of HMrA. The objectives of this study were to expand the previous study with the inclusion of black women and to determine which isoform(s) of plasma HMrA are elevated in PIH. Plasma LMrA and HMrA were quantitated in 24 normotensive pregnant women and 65 women with PIH. The PIH group had higher levels of HMrA and somewhat lower levels of LMrA than the normotensive group. The HMrA/LMrA ratio was elevated in 47% of the PIH group. The five isoforms of HMrA were quantitated in plasma from 10 white women with PIH, 10 black women with PIH, and 6 normotensive pregnant white women. Half of both the white and black women with PIH had an elevated HMrA/LMrA ratio. The relative proportion of the HMrA isomers was similar in all groups. These studies show that half the women with PIH have a distinct abnormality in their renin angiotensin system. Both white and black women show this abnormality. In those women who have an elevated total HMrA, all five isoforms of HMrA are equally elevated.

Ultrasonic recognition of the small-for-gestational-age fetus.

Sonar biparietal diameter, femur length, abdominal circumference, femur length/abdominal circumference ratio, ponderal index, and estimated fetal weight were obtained within ten days of delivery of small-for-gestational-age (SGA) (N = 102) and non-SGA (N = 204) newborns. The effectiveness of each ultrasound variable in the antenatal recognition of the SGA fetus was assessed. An abnormal abdominal circumference was the best predictor, confirming SGA in 98% of cases. Other variables (biparietal diameter, femur length, femur length/abdominal circumference ratio, ponderal index, or estimated fetal weight) were less accurate for predicting SGA. When we determined expected results based on a 10% prevalence of SGA, negative predictive value was greater than 92% for all variables studied; however, with the exception of estimated fetal weight and femur length, positive predictive values were disappointing, including abdominal circumference (21%). This report establishes the limits of ultrasound-derived growth variables in the antenatal identification of the SGA fetus.

Premature placental calcification in maternal cigarette smokers.

Ultrasonographic examinations, including placental grading, were done in 145 smoking and 100 nonsmoking low-risk obstetric patients at 37 weeks' gestation. Extensive calcification--grade III changes--occurred significantly more often in smokers than in nonsmokers (36 versus 14%; P less than .0001). Two groups of smokers, consuming five to 15 cigarettes per day or one or more packs per day, also had significant differences in grade III placenta when compared with nonsmokers. Smokers under age 20 years were more likely to have premature grade III changes; however, parity did not influence premature placental calcification in smokers and nonsmokers. The incidence of small for gestational age infants delivered was not significantly higher in smokers, and grade III placental changes appeared to occur no more often in SGA infants of smokers than of nonsmokers. Our findings suggest that the smoking gravida is at increased risk for premature placental calcification.

Clinical chorioamnionitis is not predicted by an ultrasonic biophysical profile in patients with premature rupture of membranes.

A modified biophysical profile was assessed serially in 47 patients with premature rupture of membranes who were not in labor. This profile included fetal movement, fetal tone, fetal breathing, amniotic fluid volume, and placental grade. The most recent study, obtained within 2 days of delivery, was compared with pregnancy outcome as reflected by the development of chorioamnionitis and/or neonatal sepsis. No study patient received antibiotics, steroids, or tocolytics before labor. Neither the composite biophysical profile nor any of its components were found to be different between patients with and without clinical chorioamnionitis. Neonatal sepsis was not observed. These data do not support the use of the biophysical profile as a predictor of maternal infection.

Umbilical cord blood gas assessment of twins.

Umbilical cord arterial and venous blood gas values were compared in 63 twin pairs, of which 57 pairs had birth weights of 1500 g or more each. Small differences between the first and second twins existed for PO2, PCO2, and pH. However, bicarbonate values did not differ significantly. These cord gas differences represent minor respiratory aberrations, as reflected by a tendency toward carbon dioxide retention by the second twin. Route of delivery, time interval between deliveries, and nonvertex presentations were not associated with significant deviations from these observed acid-base patterns.

Onchocercal dermatitis: ultrastructural studies of microfilariae and host tissues, before and after treatment with diethylcarbamazine (Hetrazan).

Specimens of skin from four Cameroon patients with severe onchocercal dermatitis, before and after treatment with diethylcarbamazine (DEC), were studied by light and electron microscopy. Microfilariae of Onchocerca volvulus have ultrastructural features resembling those of microfilariae of other genera. Between the surface layer of the cuticle and the trilaminate membrane, there is an electrolucent zone which is much wider in degenerating microfilariae than in intact microfilariae. Widening of the zone may result from DEC-induced release of component(s) of the cuticle, possibly collagen or mucopolysaccharide. Between the cuticle and dermal collagen there are granular deposits which might be immune complexes involving the collagenous component of cuticle. Others have shown that DEC does not kill microfilariae in vitro. Treatment with DEC presumably "unmasks" microfilariae in the skin so that they are recognized as foreign bodies and are destroyed by the host's defenses. Histiocytes and eosinophils are seen in close proximity to degenerating microfilariae. Enzymes from histiocytes and eosinophils might readily penetrate the cuticle altered by DEC treatment, and digest various components within the microfilariae. Alternatively, the widening of the electrolucent zone might result directly from the action of leucocytic or histiocytic enzymes, after the microfilaria has been killed by other mechanisms.

Cytomegalovirus infection.

Cytomegalovirus (CMV) infection is the most common perinatal infection and may result in severe injury to the fetus. Forty percent to 50% of infants delivered to mothers with primary CMV will have congenital infections. Of these, 5% to 18% will be overtly symptomatic at birth. The mortality rate in these children is almost 30%; approximately 80% of the survivors have severe neurological morbidity. The majority of congenitally infected infants will be asymptomatic at birth; 10% to 15% of these children subsequently have sequelae such as visual and auditory defects. If recurrent or reactivated CMV infection develops during pregnancy, the risk of serious fetal injury is very low. Similarly, neonatal infection acquired during delivery or from breast feeding also poses minimal risk to the child. Because antimicrobial therapy and immunoprophylaxis for CMV infection are unsatisfactory, pregnant women must be educated about preventive measures.

Effect of cocaine on intracellular calcium regulation in myometrium from pregnant women.

OBJECTIVE: To evaluate the effect of cocaine on intracellular free calcium ([Ca2+]i) regulation in human myometrial cells by determining the sources of Ca2+ it might mobilize, as well as assess the role cocaine might play in the catecholamine's effect on the cell's [Ca2+]i. METHODS: Primary culture of myometrial cells from pregnant women was used as an experimental model. [Ca2+]i relative changes in response to cocaine and norepinephrine were measured with fura-2 fluorometry and analyzed by means of one-way analysis of variance. RESULTS: Cocaine alone (10(-8) to 10(-3) mol/L) increased [Ca2+]i by up to 43 +/- 18% over basal level in a dose-dependent manner. Norepinephrine also elevated [Ca2+]i in a concentration-dependent manner (202 +/- 24% over basal level at 10(-4) mol/L). The norepinephrine-evoked increase was inhibited in Ca(2+)-free media by 48%, whereas the cocaine response was not affected. The Ca(2+)-channel antagonist nifedipine caused decrease in the [Ca2+]i response to 10(-5) mol/L of norepinephrine by 84%, whereas the [Ca2+]i rise to 10(-5) mol/L cocaine was not significantly changed. Inhibitor of the sarcoplasmic reticulum Ca2+ pump, thapsigargin, completely blocked cocaine-evoked increases in [Ca2+]i, whereas norepinephrine responses were greatly reduced. At the same time, cocaine (10(-8) to 10(-3) mol/L) did not potentiate norepinephrine-evoked Ca2+]i increases in the cells. CONCLUSION: These results indicate that cocaine increases [Ca2+]i in pregnant human myometrial cells, primarily by stimulating release of Ca2+ from intracellular stores rather than by direct stimulation of Ca2+ influx.

Anticardiolipin antibodies in unselected pregnant women. Relationship to fetal outcome.

Anticardiolipin antibodies (ACLA) have recently been associated with adverse fetal outcome. The prevalence of elevated ACLA has not been studied in unselected pregnant women, however. Twelve hundred unselected pregnant women were screened for IgG ACLA using an assay standardized by the first international workshop on ACLA. Fifteen (1.25%) were positive for IgG ACLA (greater than 3 SD above the mean) but only 0.5% had moderate to high levels of IgG ACLA (greater than 5 SD above the mean). Low levels of IgG ACLA were not associated with increased risk of fetal loss; however, 50% of women with moderate to high levels of antibody had fetal wastage. These findings further support the association of significantly elevated levels of IgG ACLA with fetal loss.

Umbilical cord thiocyanate and thyroid function in intrauterine growth-restricted infants of the smoking gravida.

OBJECTIVE: We evaluated maternal and umbilical cord thiocyanate and thyroid function in intrauterine growth-restricted infants of smoking mothers. STUDY DESIGN: One hundred six mother-infant pairs were studied and divided into four groups: smokers with appropriate for gestational age infants (n = 37), smokers with intrauterine growth-restricted infants (n = 19), nonsmokers with appropriate for gestational age infants (n = 33), and nonsmokers with intrauterine growth-restricted infants (n = 17). Sera from mothers were analyzed for thiocyanate levels. Serum from umbilical cord blood was analyzed for thiocyanate, thyroxine, resin triiodothyronine uptake, and thyroid-stimulating hormone. The free thyroxine index was then calculated. RESULTS: A significant correlation was found between maternal and umbilical cord thiocyanate levels (r = 0.90, p < 0.0001). Maternal and umbilical cord thiocyanate concentrations correlated with increased maternal cigarette use. For appropriate for gestational age and intrauterine growth-restricted infants, mean maternal and umbilical cord thiocyanate levels were not significantly different. The umbilical cord thiocyanate level did not correlate with thyroid function or birth weight. The umbilical cord thyroxine level was lower in nonsmokers than in smokers. CONCLUSION: Thiocyanate was not more concentrated in the intrauterine growth-restricted infant and did not correlate with thyroid function or birth weight. The thyroxine level was decreased significantly in intrauterine growth-restricted infants born to nonsmoking mothers. Further studies are necessary to evaluate the effects of smoking on thyroid function and on fetal growth and development.

The association of aneuploidy and unexplained elevated maternal serum alpha-fetoprotein.

OBJECTIVE: The purpose of this study was to examine fetal chromosomal abnormalities in pregnancies complicated by unexplained elevated maternal serum alpha-fetoprotein (MSAFP). STUDY DESIGN: We reviewed, using a computerized database, 58,162 obstetrical ultrasounds that were performed for various indications. Fetuses with MSAFP multiples of the median (MOM) > or = 2.5 and normal extensive ultrasounds were identified. Maternal demographic data and fetal karyotype were obtained. RESULTS: Seven hundred eighty-nine patients received ultrasounds for evaluation of elevated MSAFP. Of the 595 patients with normal scans, 195 (32.8%) underwent amniocentesis and cytogenetic evaluation. Two chromosomal abnormalities were detected (1.0%), including an inversion and a balanced translocation. CONCLUSION: The two karyotypic abnormalities identified in our study consisted of structural rearrangements. Patients undergoing karyotype analysis for unexplained elevated MSAFP should be counseled that the types of aneuploidy detected under this circumstance differ from those associated with advanced age and specific fetal anomalies (trisomy and triploidy).

Cervical prolapse complicating pregnancy.

Uterine cervical prolapse concurrent with pregnancy is rare. This article reports three cases of second-degree cervical prolapse during pregnancy. Two women developed prolapse in the late second trimester while one women had preexisting prolapse. Both women with prolapse developing during midpregnancy were treated unsuccessfully with a vaginal pessary to maintain cervical placement. Premature labor occurred in both of these women, resulting in one preterm birth. Although cervical prolapse is rarely encountered in pregnancy, the threat of preterm labor and delivery warrants close observation.

Cervical dilation as a predictor of pregnancy outcome following emergency cerclage.

OBJECTIVE: To determine whether cervical dilation at the time of physical examination indicated cerclage placement can predicts latency and gestational age at delivery. METHODS: A retrospective cohort study of all women who underwent physical examination indicated cerclage placement from 1996 to 2011 at Duke University Hospital (DUH) was performed. Physical examination indicated cerclage was defined as cerclage placement after 16 weeks in women with a cervical length of less than 2.5 cm and/or cervical dilation greater than or equal to 1 cm at time of procedure. Subjects were divided into two groups depending on cervical dilation at time of procedure (2 cm, <2 cm) for comparison. A multivariate linear regression model for the outcome gestational age of delivery was constructed, controlling for confounding variables. RESULTS: A total of 110 women with complete data were available for analysis. Median gestational age at cerclage placement was similar between the two groups (20.3 vs. 20.3 weeks, p = 0.8). Women with cervical dilatation ≥ 2 cm dilation delivered at an earlier median gestational age than women with cervical dilation <2 cm (27.0 vs. 35.6 weeks, p < 0.001). Cervical dilation at the time of cerclage placement independently predicted gestational age at delivery while controlling for use of intracervical Foley balloon catheter for membrane reduction, cerclage suture type, history of prior preterm birth, race, insurance status, and tobacco use. CONCLUSIONS: Women who receive a rescue cerclage are more likely to deliver at an earlier gestational age when cervical dilation is ≥ 2 cm at the time of procedure.

Climate change and air quality: the potential impact on health.

The objectives of the study were to: consider the potential health impacts in Australia and the region arising from changes in air quality occurring as a result of climate change, identify vulnerable groups and potential adaptation measures and discuss the implications for policy. The authors provide an overview of international and national information on the potential health impacts of air pollutants that would most likely be affected by climate change and a discussion of the policy implications. Climate change is likely to have an impact on levels of ozone and possibly particulates, both of which are associated with increased mortality and a range of respiratory and cardiovascular health effects. One of the implications is therefore a possible increase in adverse health effects due to air pollutants. Regional health impact assessments of climate change should address the issue of air quality, consider current coping capacity, and determine the need for adaptation, particularly for vulnerable groups. Implications for policy include the need for improved modeling and forecasting of air pollutant levels, increased efforts to reduce emissions of air pollutants, continued monitoring of air pollutant levels, and monitoring of the incidence of health effects associated with air pollutants in all countries in the region.