Fetal imaging and diagnosis services in developing countries – A call to action

Fetal congenital anomalies are among the leading causes of perinatal death or survival with disability worldwide. Their accurate antenatal detection  employing a range of fetal imaging techniques enables parental choices to be made and for postnatal care of affected babies to be planned. While such  prenatal care is well developed in developed countries of the world, it remains poor in many low‑ and middle‑income countries (LMICs). This review article  examines the scope of the problem and proffers strategies for service  organization and fetal imaging that will improve care in LMIC settings.Key words: Congenital abnormalities; fetus; imaging; pregnancy; ultrasound

Reproducibility and repeatability of measuring the electrical impedance of the pregnant human cervix-the effect of probe size and applied pressure

Background: The utility of cervical electrical impedance spectroscopy (EIS) as a diagnostic tool is being investigated in clinical trials. We sought to assess the reliability of two different sizes of tetrapolar probes used in measuring cervical impedance. Methods: Cervical transfer impedance was measured at 14 frequencies between 76 and 625 000 Hz from 11 pregnant subjects at term. Repeated measurements were taken with two probes (3 mm and 12 mm diameter) applied softly (approximately 0.7 Newton of force), and firmly (approximately 2.2 Newton) to the surface of the cervix by two observers. The intra-class correlation coefficient (ICC), coefficient of variation (CV) and repeatability standard deviations (SD) were derived from these measurements and compared. Results: Measurements taken by one observer were highly repeatable for both probes as demonstrated by high ICC and low CV values. Probe performance was improved further by firm application. Firm application of the 3 mm probe resulted in ICC values that ranged from 0.936 to 0.986 (p = 0.0001) and CV values between 1.0 and 3.4%. Firm pressure with the 12 mm probe resulted in ICC values that ranged between 0.914 and 0.988 (p = 0.0001) with CV values between 0.7 and 2.1%. In addition, the repeatability SD was low across all frequencies implying that there was low intra-observer variability. Measurements taken by 2 observers with firm application of the 12 mm probe demonstrated moderate reproducibility between 9.8 and 156 kHz, the frequency range in which previous clinical studies have shown predictive association between high cervical resistivity and vaginal delivery: ICC values ranged between 0.528 and 0.638 (p < 0.05), CV values were between 3.3 and 5.2% and reproducibility SD values were also low. In contrast the 3 mm probe demonstrated poor reproducibility at all study frequencies. Conclusion: Measuring cervical resistivity by a single observer with both the 3 and 12 mm probes is highly repeatable whilst inter-observer reproducibility is poor with the 3 mm probe but moderately good when the 12 mm probe is firmly applied to the cervix in the frequency range 9.8 to 156 kHz, consistent with our observations of probe performance in clinical trials

Female gut and genital tract microbiota-induced crosstalk and differential effects of short-chain fatty acids on immune sequelae

The gut and genital tract microbiota of females represent very complex biological ecosystems that are in continuous communication with each other. The crosstalk between these two ecosystems impacts host physiological, immunological and metabolic homeostasis and vice versa. The vaginal microbiota evolved through a continuous translocation of species from the gut to the vagina or through a mother-to-child transfer during delivery. Though the organisms retain their physio-biochemical characteristics while in the vagina, the immune responses elicited by their metabolic by-products appear to be at variance with those in the gut. This has critical implications for the gynecological, reproductive as well as overall wellbeing of the host and by extension her offspring. The homeostatic and immunomodulatory effects of the bacterial fermentation products (short chain fatty acids, SCFAs) in the gut are better understood compared to the genital tract. While gut SCFAs prevent a leakage of bacteria and bacterial products from the gut in to circulation (leaky gut) and consequent systemic inflammation (anti-inflammatory/protective role); they have been shown to exhibit dysbiotic and proinflammatory effects in the genital tract that can lead to unfavorable gynecological and reproductive outcomes. Therefore, this review was conceived to critically examine the correlation between the female gut and genital tract microbiota. Secondly, we explored the metabolic patterns of the respective microbiota niches; and thirdly, we described the diverse effects of products of bacterial fermentation on immunological responses in the vaginal and rectal ecosystems

Circulating levels of matrix proteases and their inhibitors in pregnant women with and without a history of recurrent pregnancy loss

Background: We have recently shown that serum relaxin-2 levels are attenuated in women with a history of recurrent pregnancy loss (RPL). We sought to determine whether a history of RPL is also associated with changes in serum matrix metalloproteases (MMPs) and tissue inhibitors of matrix metalloproteases (TIMP) -1 and -2. Methods: We obtained serum from 20 pregnant women with a history of RPL and 20 age-matched pregnant women with no history of RPL (NRPL) at 6-8, 10-12, 20, and 34 weeks gestation, and from cord blood. We quantified total serum concentrations of MMP-1, MMP-3, MMP-9 and TIMP-1 and TIMP-2 by ELISA. We determined whether these serum marker levels were associated with a history of RPL and delivery before 37 weeks gestation. Results: There was no difference in the rates of miscarriage, preterm birth or prelabour rupture of fetal membranes between RPL and NRPL. However babies born to RPL were lighter than those born to NRPL. Serum MMP-1, 9, and TIMP-1 did not differ between RPL and NRPL but MMP-3 was higher in RPL vs. NRPL at 6-8 weeks (P < 0.05). Serum TIMP-2 levels were higher in RPL women at all gestations (P < 0.01). The ratio of RLX-2 (reported previously) to TIMP-2 at 10-12 weeks gestation was more strongly associated with a history of RPL than either peptide separately - area under the ROC curves for RLX-2 0.79 (95% CI 0.57 to 0.92), TIMP-2 0.83 (95% CI 0.63 to 0.95), and for RLX-2: TIMP-2 ratio 0.92 (95% CI 0.74 to 0.99). Conclusions: Women with a history of RPL demonstrate increased serum TIMP-2 and reduced RLX-2 during a subsequent viable pregnancy. Determination of both markers in early pregnancy enhances the discrimination of women with a history of RPL. These observations suggest roles for these two peptides in early implantation and placental development. Whether these may prove to be reliable early predictive markers for subsequent pregnancy loss in the index pregnancy is unknown and will require further studies

The vaginal microenvironment: the physiologic role of lactobacilli

In addition to being a passage for sperm, menstruum, and the baby, the human vagina and its microbiota can influence conception, pregnancy, the mode and timing of delivery, and the risk of acquiring sexually transmitted infections. The physiological status of the vaginal milieu is important for the wellbeing of the host as well as for successful reproduction. High estrogen states, as seen during puberty and pregnancy, promote the preservation of a homeostatic (eubiotic) vaginal microenvironment by stimulating the maturation and proliferation of vaginal epithelial cells and the accumulation of glycogen. A glycogen-rich vaginal milieu is a haven for the proliferation of Lactobacilli facilitated by the production of lactic acid and decreased pH. Lactobacilli and their antimicrobial and anti-inflammatory products along with components of the epithelial mucosal barrier provide an effective first line defense against invading pathogens including bacterial vaginosis, aerobic vaginitis-associated bacteria, viruses, fungi and protozoa. An optimal host-microbial interaction is required for the maintenance of eubiosis and vaginal health. This review explores the composition, function and adaptive mechanisms of the vaginal microbiome in health and those disease states in which there is a breach in the host-microbial relationship. The potential impact of vaginal dysbiosis on reproduction is also outlined

Impaired autophagy with augmented apoptosis in a Th1/Th2-imbalanced placental micromilieu is associated with spontaneous preterm birth

Background: Despite decades of research, the pathogenesis of spontaneous preterm birth (PTB) remains largely unknown. Limited currently available data on PTB pathogenesis are based on rodent models, which do not accurately reflect the complexity of the human placenta across gestation. While much study has focused on placental infection and inflammation associated with PTB, two key potentially important cellular events in the placenta—apoptosis and autophagy—remained less explored. Understanding the role of these processes in the human placenta may unravel currently ill-understood processes in the pathomechanism of PTB. Methods: To address this necessity, we conducted qRT-PCR and ELISA assays on placental villous tissue from 20 spontaneous preterm and 20 term deliveries, to assess the inter-relationships between inflammation, apoptosis, and autophagy in villous tissue in order to clarify their roles in the pathogenesis of PTB. Results: We found disrupted balance between pro-apoptotic BAX and anti-apoptotic BCL2 gene/protein expression in preterm placenta, which was associated with significant reduction of BCL2 and increase of BAX proteins along with upregulation of active CASP3 and CASP8 suggesting augmented apoptosis in PTB. In addition, we detected impaired autophagy in the same samples, evidenced by significant accumulation of autophagosome cargo protein p62/SQSTM1 in the preterm villous placentas, which was associated with simultaneous downregulation of an essential autophagy gene ATG7 and upregulation of Ca2+-activated cysteine protease CAPN1. Placental aggregation of p62 was inversely correlated with newborn birth weight, suggesting a potential link between placental autophagy impairment and fetal development. These two aberrations were detected in a micromilieu where the genes of the Th2 cytokines IL10 and IL13 were downregulated, suggesting an alteration in the Th1/Th2 immune balance in the preterm placenta. Conclusion: Taken together, our observations suggest that impaired autophagy and augmented apoptosis in a Th1/Th2 imbalanced placental micro-environment may be associated with the pathogenesis of spontaneous PTB

Mid-gestational changes in cervicovaginal fluid cytokine levels in asymptomatic pregnant women are predictive markers of inflammation-associated spontaneous preterm birth

OBJECTIVES: Perturbation of the choriodecidual space before the onset of spontaneous preterm birth (sPTB) could lead to a concomitant rise in both cervicovaginal fluid (CVF) cytokine and fetal fibronectin (FFN), and assessing the concentrations of both markers could improve the prediction of sPTB (delivery before 37 completed weeks of gestation). Therefore, we prospectively determined mid-trimester changes in CVF cytokine and FFN concentrations, and their predictive capacity for sPTB in asymptomatic pregnant women. STUDY DESIGN: CVF collected at 20+0-22+6 weeks (n = 47: Preterm-delivered = 22, Term-delivered = 25) and 26+0-28+6 weeks (n = 50: Preterm-delivered = 17, Term-delivered = 33) from 63 asymptomatic pregnant women at risk of sPTB were examined. Cytokine and FFN concentrations were determined by multiplexed bead-based immunoassay and 10Q Rapid analysis (Hologic, MA, USA) respectively. The 20+0-22+6/26+0-28+6 weeks ratios of cytokines and FFN concentrations were compared between preterm- and term-delivered women using Receiver Operating Characteristics curves to predict sPTB. Also, bacterial 16S rDNA from 64 samples (20+0-22+6 weeks n = 36, 26+0-28+6 weeks n = 28) was amplified by polymerase chain reaction to determine associations between vaginal microflora, cytokine and FFN concentrations. RESULTS: Changes in RANTES and IL-1β concentrations between 20+0-22+6 and 26+0-28+6 weeks, expressed as a ratios, were predictive of sPTB, RANTES (AUC = 0.82, CI = 0.62-0.94) more so than IL-1β (AUC = 0.71, CI = 0.53-0.85) and FFN (not predictive). Combining these markers (AUC = 0.83, CI = 0.63-0.95) showed similar predictive capacity as RANTES alone. FFN concentrations at 26+0-28+6 weeks correlated with IL-1β (r = 0.4, P = 0.002) and RANTES (r = 0.3, P = 0.03). In addition, there was increased prevalence of vaginal anaerobes including Bacteroides, Fusobacterium and Mobiluncus between gestational time points in women who experienced sPTB compared to the term women (P = 0.0006). CONCLUSIONS: CVF RANTES and IL-1β in mid-trimester of pregnancy correlate with quantitative FFN. The levels of CVF RANTES and IL-1β decline significantly in women who deliver at term unlike women who deliver preterm. This observation suggests that sPTB may be characterised by sustained choriodecidual inflammation and may have clinical value in serial screening for sPTB if confirmed by larger studies

Transcriptomic analysis of the human placenta reveals trophoblast dysfunction and augmented Wnt signalling associated with spontaneous preterm birth

Preterm birth (PTB) is the leading cause of death in under-five children. Worldwide, annually, over 15 million babies are born preterm and 1 million of them die. The triggers and mechanisms of spontaneous PTB remain largely unknown. Most current therapies are ineffective and there is a paucity of reliable predictive biomarkers. Understanding the molecular mechanisms of spontaneous PTB is crucial for developing better diagnostics and therapeutics. To address this need, we conducted RNA-seq transcriptomic analysis, qRT-PCR and ELISA on fresh placental villous tissue from 20 spontaneous preterm and 20 spontaneous term deliveries, to identify genes and signalling pathways involved in the pathogenesis of PTB. Our differential gene expression, gene ontology and pathway analysis revealed several dysregulated genes (including OCLN, OPTN, KRT7, WNT7A, RSPO4, BAMBI, NFATC4, SLC6A13, SLC6A17, SLC26A8 and KLF8) associated with altered trophoblast functions. We identified dysregulated Wnt, oxytocin and cellular senescence signalling pathways in preterm placentas, where augmented Wnt signalling could play a pivotal role in the pathogenesis of PTB due to its diverse biological functions. We also reported two novel targets (ITPR2 and MYLK2) in the oxytocin signalling pathways for further study. Through bioinformatics analysis on DEGs, we identified four key miRNAs, - miR-524-5p, miR-520d-5p, miR-15a-5p and miR-424-5p - which were significantly downregulated in preterm placentas. These miRNAs may have regulatory roles in the aberrant gene expressions that we have observed in preterm placentas. We provide fresh molecular insight into the pathogenesis of spontaneous PTB which may drive further studies to develop new predictive biomarkers and therapeutics

Postnatal pelvic floor muscle stiffness measured by vaginal elastometry in women with obstetric anal sphincter injury : a pilot study

Introduction and hypothesis Vaginal childbirth is associated with pelvic floor muscle (PFM) damage in a third of women. The biomechanics prediction, detection and management of PFM damage remain poorly understood. We sought in this pilot study to determine whether quantifying PFM stiffness postnatally by vaginal elastometry, in women attending a perineal trauma clinic (PTC) within 6 months of obstetric anal sphincter injury, correlates with their antecedent labour characteristics, pelvic floor muscle damage, or urinary/bowel/sexual symptoms, to inform future definitive prospective studies. Methods In this pilot study, we measured postnatal PFM stiffness by vaginal elastometry in 54 women. A subset of participants (n = 14) underwent magnetic resonance imaging (MRI) to define any levator ani (LA) muscle defects from vaginal childbirth. We investigated the association of PFM stiffness with demographics, labour and delivery characteristics, clinical features and MRI evidence of LA damage. Results Raised maternal BMI was associated with reduced pelvic floor stiffness (r = −0.4; p < 0.01). Higher stiffness values were associated with forceps delivery for delayed second stage of labour (n = 14) vs non-forceps vaginal delivery (n = 40; 630 ± 40 N/m vs 500 ± 30 N/m; p < 0.05), and a non-significant trend towards longer duration of the second stage of labour. Women with urinary, bowel or sexual symptoms (n = 37) demonstrated higher pelvic floor stiffness values than those without (570 ± 30 N/m vs 450 ± 40 N/m; p < 0.05). Conclusions A history of delayed second stage of labour and forceps delivery was associated with higher PFM stiffness values in the postnatal period. Whether high pelvic muscle stiffness antenatally is a risk factor for instrumental vaginal delivery and LA avulsion is unknown

The value of cervical electrical impedance spectroscopy to predict spontaneous preterm delivery in asymptomatic women: the ECCLIPPx prospective cohort study

Objectives Preterm birth (PTB) accounts for two‐thirds of deaths of structurally normal babies and is associated with colossal lifetime health care costs. Prevention of PTB remains limited by prediction methods of modest accuracy– transvaginal ultrasound (TVUS) cervical length (CL) and quantitative cervicovaginal fetal fibronectin (FFN) estimation. We report the first substantive study detailing the predictive performance of a cervical probe device based on Electrical Impedance Spectroscopy (EIS) for PTB – the EleCtriCaL Impedance Prediction of Preterm birth by spectroscopy of the cervix (ECCLIPPx) studies. We aimed to compare the accuracy of cervical EIS‐based prediction of spontaneous PTB in asymptomatic women in mid‐trimester to TVUS CL and FFN. Methods We studied 449 pregnant women at 20‐22 weeks (V1) and 26‐28 weeks (V2). EIS was performed using the Sheffield Mark 5.0 device that makes measurements in the frequency range 76Hz to 625kHz using a small probe housing tetrapolar electrodes. TVUS CL and FFN were also measured. Associations between cervical EIS, TVUS CL, and FFN and spontaneous delivery < 37 weeks and < 32 weeks were determined by multivariate linear and non‐linear logistic regression analyses. The areas under the Receiver Operator Characteristic (AuROC) curve plots of sensitivity against specificity were used to compare the predictive performance of all parameters, singly and in combination. Results Of the 365 asymptomatic women studied at 20‐22 weeks who received no treatments, there were 29 spontaneous PTBs, 14 indicated PTBs, and 322 term births. At the higher frequencies assessed, cervical EIS predicted spontaneous delivery < 37 weeks (AuROC 0.76, 95% CI 0.71‐0.81) compared to TVUS CL (AuROC 0.72, 95% CI 0.66‐0.76) and FFN (AuROC 0.62, 95%CI 0.56‐0.67). Combining all three assessments improved prediction of spontaneous PTB <37 weeks (AuROC 0.80, 95% CI 0.74‐0.83) compared to TVUS CL and FFN alone. Incorporating previous history of PTB into the cervical EIS prediction model improved accuracy of prediction of spontaneous PTB < 37 weeks (AuROC 0.83, 95% CI 0.78‐0.87) and < 32 weeks (AuROC 0.86, 95% CI 0.82‐0.90). Conclusion Mid‐trimester cervical EIS assessment predicts spontaneous PTB. Larger confirmatory studies investigating its potential clinical utility and to inform effective preventive interventions are required