Amniotic Fluid Research Articles

Array Comparative Genomic Hybridization (aCGH) Results among Patients Referred to Invasive Prenatal Testing after First-Trimester Screening: A Comprehensive Cohort Study.

Introduction: Invasive prenatal testing with chromosomal microarray analysis after first-trimester screening is a relevant option but there is still debate regarding the indications. Therefore, we evaluated the prevalence of numerical chromosomal aberrations detected by classic karyotype and clinically relevant copy number variants (CNVs) in prenatal samples using array comparative genomic hybridization (aCGH) stratified to NT thickness: <the 95th percentile, the 95th percentile-2.9 mm, 3.0-3.4 mm, 3.5-3.9 mm, 4.0-4.5 mm, and >4.5 mm, and by the presence/absence of associated structural anomalies detected by ultrasonography. Materials and Methods: Retrospective cohort study carried out at two tertiary Polish centers for prenatal diagnosis (national healthcare system) in central and south regions from January 2018 to December 2021. A total of 1746 prenatal samples were received. Indications for invasive prenatal testing included high risk of Down syndrome in the first-trimester combined test (n = 1484) and advanced maternal age (n = 69), and, in 193 cases, other reasons, such as parental request, family history of congenital defects, and genetic mutation carrier, were given. DNA was extracted directly from amniotic fluid (n = 1582) cells and chorionic villus samples (n = 164), and examined with classic karyotype and aCGH. Results: Of the entire cohort of 1746 fetuses, classical karyotype revealed numerical chromosomal aberrations in 334 fetuses (19.1%), and aCGH detected CNV in 5% (n = 87). The frequency of numerical chromosomal aberrations increased with NT thickness from 5.9% for fetuses with NT < p95th to 43.3% for those with NT > 4.5 mm. The highest rate of numerical aberrations was observed in fetuses with NT > 4.5 mm having at least one structural anomaly (50.2%). CNVs stratified by NT thickness were detected in 2.9%, 2.9%, 3.5%, 4.3%, 12.2%, and 9.0% of fetuses with NT < 95th percentile, 95th percentile-2.9 mm, 3.0-3.4 mm, 3.5-3.9 mm, 4.0-4.5 mm, and >4.5 mm, respectively. After exclusion of fetuses with structural anomalies and numerical aberrations, aCGH revealed CNVs in 2.0% of fetuses with NT < 95th percentile, 1.5% with NTp95-2.9 mm, 1.3% with NT 3.0-3.4 mm, 5.4% with NT 3.5-3.9 mm, 19.0% with NT 4.0-4.5 mm, and 14.8% with NT > 4.5 mm. Conclusions: In conclusion, our study indicates that performing aCGH in samples referred to invasive prenatal testing after first-trimester screening provides additional clinically valuable information over conventional karyotyping, even in cases with normal NT and anatomy.

Clinicopathological Insights to the Nerve Growth Factor NGF Associated Stress Response in Pregnancy and Therapeutic Potential in Fetal Neurodevelopment

The nerve growth factor has a significant role in fetal neurogenesis and sustaining pregnancy. Objectives: To investigate the effects of nerve growth factor in stress response during pregnancy on developing fetus to bring clinicopathological correlation on the role of nerve growth factor in maternal stress markers (cortisol levels, glucocorticoids, depression, anxiety, and brain-derived neurotrophic factor levels) and fetal brain development. Methods: Following PRISMA guidelines, this study was extracted from PubMed, ScienceDirect, Nature, and Google Scholar articles from January 2014 to April 2024. The examination of pregnant women in published research gave a possibility to understand the application of nerve growth factor as a suitable biomarker for brain stress and fetal neuronal development. To exclude studies with lower ranks, each of the selected studies was assessed for adherence to evidence-based research methodology. The studies were taken from China, Europe, America and South Asia (including Pakistan). Results: Increased nerve growth factor levels were associated with maternal stress reactions which caused changes in cortisol levels and the amygdaloidal complex area. However, the increased nerve growth factor level was linked to changes in the fetal brain such as the weight of the fetal brain and stress biomarkers in the amniotic fluid sample inferring a critical role in the modulation of maternal stress on the fetal neurodevelopmental spheres. Conclusions: It was concluded that it is important to note how stress and nerve growth factors interact during pregnancy to create effective interventions to reduce stress dependence for the better health of both the mother and child.

Metabolic landscaping of extracellular vesicles from body fluids by phosphatidylserine imprinted polymer enrichment and mass spectrometry analysis

Extracellular vesicles (EVs) are emerging as new source of biomarkers discovery in liquid biopsy due to their stabilization in body fluids, protected by phospholipid bilayers. However, the metabolomics study of EVs is very little reported due to the lack of efficient and high-throughput isolation methods for clinical samples. In this study, phosphatidylserine imprinted polymers were employed for rapid and efficient EVs isolation from five human body fluids, including plasma, urine, amniotic fluid, cerebrospinal fluid, and saliva. The isolated EVs were subsequently analyzed for metabolomic studies by high-resolution mass spectrometry. Metabolic landscaping was conducted between the body fluids and their EVs, indicating EVs contain a large number of metabolites that are completely specific to the body fluid source. Finally, quantitative metabolomic analysis of EVs was carried out with plasma samples of hepatocellular carcinoma. Several differentially expressed exosomal metabolites were revealed including the upregulation of sphingosine (d18:1), taurochenodeoxycholic acid (TCDCA), pipecolic acid (PA), and 4-hydroxynonenal (4-HNE) and down-regulation of piperine, caffeine, and indole. We believe the proposed methodology will provide a deeper understanding of the molecular composition and functions of EVs as an alternative source for biomarker discovery.

Differential use of various types of pessaries in isthmic-cervical insufficiency for prevention of preterm birth: A randomized prospective trial

Background. Obstetric pessary comprises one of the methods for treatment of isthmic-cervical insufficiency. Despite the variety of pessaries produced, the common purpose of their use consists in preventing premature birth. Various types of pessaries correct different cervical parameters, which is not always taken into account by doctors when choosing a pessary and reduces their potential effectiveness. Objective. To substantiate a differentiated approach to the selection of pessary type for correcting isthmic-cervical insufficiency and preventing preterm birth based on the evaluation of cervical parameters. Methods. A randomized prospective study enrolled 90 pregnant women diagnosed with isthmic-cervical insufficiency (ICD-10 code — О.34.3) at 19–24 weeks of gestation. Of these, 41 women underwent correction of isthmic-cervical insufficiency with an obstetric unloading pessary and 49 women — with a perforated cervical pessary. Transvaginal ultrasound cervicometry evaluated the parameters of the cervix before correcting isthmic-cervical insufficiency and in dynamics (once every 4 weeks) after inserting various types of pessaries. Statistical data processing was carried out using Statistica 10.0 (StatSoft, Tulsa, USA) and MedCalc 10.2.0.0 (MedCalc, Mariakerke, Belgium). The differences were considered to be statistically significant at p &lt;0.05. Results. Inserting an obstetric unloading pessary in isthmic-cervical insufficiency decreased the uterocervical angle from 115 (110; 130)° to 100 (90; 115)° (p = 0.021). A decrease in the uterocervical angle was observed during 16-week-use of obstetric unloading pessary. After insertion of perforated cervical pessaries, the length of the closed part of the cervical region increased from 23 (21; 24) mm to 25 (21; 27) mm (p = 0.009) for a period of 4 weeks with a subsequent decrease in this parameter. The effectiveness of both types of pessaries in preventing preterm birth was found to be identical. Urgent delivery occurred in 61% of cases of using an obstetric unloading pessary and in 64.7% of cases of using a perforated cervical pessary (p = 0.993). The gestational age at preterm birth against the background of the use of obstetric unloading pessaries and perforated cervical pessaries was found comparable and amounted to 247 (230; 253) days and 245 (225; 254) days, respectively (p = 0.870). Conclusion. A differentiated approach to selecting a type of pessary for the prevention of premature birth in isthmic-cervical insufficiency is determined by the initial ultrasound parameters of the cervix. Thus, an increase in the uterocervical angle serves as an indication for an obstetric unloading pessary, while a shortened part of the cervical region without an increase in the utero-cervical angle determines the use of a perforated cervical pessary. Additional dynamic ultrasound control after inserting pessaries of any type allows such complications as pessary displacement, cervical edema, amniotic fluid sludge, prolapse of fetal membranes in the vagina, and increased myometrial tone to be timely diagnosed and corrected, thereby increasing the effectiveness of using pessaries.

Characterization of Amniotic Fluid Ureaplasma Species from Pregnancies Complicated by Preterm Prelabor Rupture of Membranes.

The main aim of this study was to determine expanded sequence types (eSTs) of Ureaplasma species (U. spp.). DNA isolated from the amniotic fluid of pregnancies complicated by preterm prelabor rupture of membranes (PPROM) using an expanded multilocus sequence typing scheme. Additionally, the study sought to examine whether phylogenetic subgroups of U. spp. DNA differ with respect to maternal demographic and clinical parameters and selected aspects of short-term neonatal morbidity. This retrospective cohort study was focused on singleton pregnancies complicated by PPROM occurring between the gestational ages of 24+0 and 36+6 weeks, where amniocentesis was conducted to assess the intra-amniotic environment and the presence of U. spp. DNA in the amniotic fluid samples was confirmed. The stored aliquots of U. spp. DNA were used to assess differences in nucleotide sequences in six U. spp. genes (ftsH, rpL22, valS, thrS,ureG, and mba-np1) using the eMLST scheme. The expanded multilocus sequence typing scheme was performed in 73 samples of U. spp. DNA isolated from pregnancies complicated by PPROM. In total, 33 different U. spp. DNA eSTs were revealed, 21 (#20, 233-244, 248-251, 253, 255, 259, and 262) of which were novel. The most frequently identified eST was #41, identified in 18% (13/73) of the aliquots. Based on their genetic relationships, the U. spp. DNA was divided into two clusters and four subgroups [cluster I (U. parvum): A, 43% (n = 31); B, 15% (n = 11); and C, 26% (n = 19); cluster II (U. urealyticum): 1; 16% (n = 12)]. Cluster II had a higher rate of polymicrobial findings than cluster I (58% vs 16%; p = 0.005), while subgroup A had the highest rate of concomitant Mycoplasma hominis in the amniotic fluid samples (66%; p = 0.04). In conclusion, Ureaplasma spp. DNA obtained from PPROM consisted of 33 different eSTs of U. spp. DNA. No differences in maternal and neonatal characteristics were found among the phylogenetical subgroups of U. spp. DNA, except for a higher rate of polymicrobial amniotic fluid findings in those with U. urealyticumand the concomitant presence of M. hominis in the amniotic fluid in those with the presence of U. parvum.

The prevalence of pulmonary artery hypertension in neonates with meconium aspiration syndrome: a prospective study

Background: Meconium aspiration syndrome (MAS) is one of the most common causes of respiratory distress in the term and post term newborns. Meconium staining of amniotic fluid occurs in 10-15% of all deliveries, amongst which 5% of babies develop MAS. Pulmonary artery hypertension (PAH) and pneumothorax are severe complications of MAS and these contribute significantly to mortality and morbidity in the affected babies. The objective was to study the prevalence, risk factors and outcome of PAH in neonates with meconium aspiration syndrome. Methods: This prospective observational study included 250 newborns with MAS who were screened for the development of PAH by echocardiography during hospital stay. The prevalence, risk factors and outcome of PAH were determined. Results: The prevalence of PAH and PPHN among meconium aspiration syndrome was 45% and 29% respectively. Low Apgar score at birth and presence of thick meconium were identified as the risk factors for progression to PAH. The outcome of newborns with PPHN was poor. Conclusion: There is a high prevalence of PAH in babies born with MAS and those progressing to PPHN have a poorer outcome.

A green and sensitive electromembrane extraction (EME) method for the determination of dialkylphosphates in maternal urine and amniotic fluid

Organophosphates compounds are extensively used pesticides, whose metabolites, dialkyl phosphates, are considered as bioindicators of human exposure. Recently, research has indicated that they can affect thyroid endocrine function. Pregnant women are a particularly vulnerable cohort to these effects due to concerns about prenatal exposure to environmental pollutants. For the first time an electromembrane microextraction method is proposed for the simultaneous determination of seven dialkylphosphate metabolites in maternal urine and amniotic fluid, which is compatible with mass spectrometry detection and achieves highly sensitive levels of quantitation. Dialkylphosphates were extracted from a 10 mL donor solution at pH 7 into a 50 µL acceptor solution at pH 12 using 1-octanol as the supported liquid membrane and applying 60 V for 10 min at 600 rpm. The obtained extracts were analysed by ion pair liquid chromatography coupled to a triple quadrupole mass spectrometer, using an Acquity UPLC® BEH C18 column (100 mm × 2.1 mm i.d., 1.7 µm particle size) at 30 °C. Under optimal extraction conditions, the preconcentration factors were achieved were up to 178-fold, allowing for high sensitivity levels, in the order of ng mL−1. The limits of quantitation were within 0.075 ng mL−1 (diethyl thiophosphate) and 1.67 ng mL−1 (dimethyl phosphate) in maternal urine samples and within 0.015 ng mL−1 (diethyl thiophosphate) and 0.5 ng mL−1 (dimethyl dithiophosphate) in amniotic fluid samples. The comparative study revealed a significant improvement in terms of analysis time, sensitivity, greenness, and practicality. Finally, the applicability of the method for the determination of selected dialkylphosphates was demonstrated in paired maternal urine and amniotic fluid samples. These results suggest potential placental transfer to the offspring.

Trial of labor following cesarean among patients with polyhydramnios: a multicenter retrospective study.

This study aimed to assess maternal and neonatal outcomes in patients with polyhydramnios attempting trial of labor after cesarean (TOLAC) compared to those undergoing planned repeat cesarean delivery (PRCD). A multi-center retrospective cohort study was conducted and included women with term singleton viable pregnancies following a single low-segment transverse cesarean delivery (CD) with a polyhydramnios diagnosis (maximal vertical pocket > 8cm and/or Amniotic Fluid Index > 24cm) within 14days before birth who delivered between the years 2017 and 2021. Maternal and neonatal outcomes were compared between those attempting TOLAC and those opting for PRCD. The primary outcome was composite adverse maternal. Univariate analysis was followed by multivariate analysis to control for potential confounders. Out of 358 included births with a previous CD, 208 (58.1%) attempted TOLAC, while 150 had PRCD (41.9%). The successful vaginal birth after cesarean (VBAC) rate was 82.2%, and no cases of uterine rupture, hysterectomy, or maternal intensive care unit admission occurred in either group. After controlling for potential confounders, no independent association between TOLAC and composite adverse maternal (adjusted odds ratio [aOR] 0.62, 95% confidence interval [CI] 0.32-1.20, p = 0.16) and neonatal (aOR 0.89, 95% CI 0.51-1.53, p = 0.67) adverse outcomes was demonstrated. In patients with a term diagnosed polyhydramnios, TOLAC appears to be a reasonable alternative associated with favorable outcomes. Larger prospective studies are needed to refine management strategies and enhance maternal and neonatal outcomes in this context.

Lipidomic profiling of amniotic fluid reveals aberrant fetal lung development and fetal growth disrupted by lipid disorders during gestational asthma

This study aimed to investigate how maternal asthma during pregnancy disrupts fetal lung development by altering lipid metabolism in the amniotic fluid, which is crucial for fetal development. A pregnancy-induced asthma model was established in female rats using house dust mite (HDM) as a common allergen. The fetuses were divided into four groups based on whether the mother and fetus were exposed to the allergen: PBS+PBS, PBS+HDM, HDM+PBS, and HDM+HDM. Ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was employed to analyze changes in the lipid profile of the amniotic fluid and bronchoalveolar lavage fluid (BALF). Principal component analysis (PCA) and ChemRICH methods were used to explore the potential relationship between lipid metabolism abnormalities and impaired fetal lung development. The results indicate that maternal asthma exacerbates asthma-related inflammatory markers in fetuses, leading to pathological changes in the lungs and elevated levels of cytokines IL-5, IL-13, and IgE. Additionally, 18 differential lipids, primarily oxygenated lipids, were identified in the amniotic fluid after modeling, suggesting an enhanced oxidative stress environment for the fetus. This environment causes metabolic disturbances in various lipid groups in fetal lungs, with the HDM+HDM group showing significant abnormalities in lipids critical for lung development, including phosphatidylethanolamine (PE), phosphatidylglycerol (PG), and fatty acids (FA). In conclusion, gestational asthma can reshape the lipid profile in the amniotic fluid and BALF, significantly disrupting fetal growth and lung development. Restoring normal lipid metabolism in the amniotic fluid and fetal lungs may offer a potential therapeutic approach to managing aberrant fetal lung development in asthmatic mothers.

The Implications of a "Flat" Oral Glucose Tolerance Test Curve in Pregnancy.

To determine whether pregnant women with "flat" oral glucose tolerance test (OGTT) curves are at increased risk of adverse perinatal outcomes. We conducted a retrospective cohort study comparing the perinatal outcomes of pregnant women whose 100g OGTT curve was "flat" (fasting serum glucose below 95 mg/dl and the remaining values below 100 mg/dl) to those of women whose OGTT curve was "normal". Primary outcomes compared were the prevalence of macrosomic and small for gestational age (SGA) neonates. Secondary outcomes included hypertensive disorders of pregnancy (HDP), intrauterine fetal demise, placental abruption, induction of labor, meconium-stained amniotic fluid (MSAF), mode of delivery, postpartum hemorrhage (PPH), blood product transfusion, postpartum readmission, umbilical artery pH <7.1, Apgar score <7 at 5 minutes, neonatal intensive care unit (NICU) admission, neonatal respiratory and infectious morbidity, and hypoglycemia. Composite adverse maternal and neonatal outcomes were also evaluated. There were 1,060 patients in the study group and 10,591 patients in the control group. Patients with a flat OGTT had a reduced risk of delivering a macrosomic neonate (11.4% vs. 15.1% OR 0.7 [0.58-0.89] p=0.001) and having an unplanned cesarean delivery (7.5% vs 10.2% OR 0.8 [0.58-0.96] p=0.002). There was no difference in the rate of composite adverse maternal (14.0% vs 15.4% OR 0.9 [0.7-1.0] p=0.1) or neonatal outcome (5.3% vs. 4.5% OR 1.2 [0.9-1.5] p=0.15). Neonates had a slightly lower mean birthweight (3474g vs. 3505g, p=0.04), but the rate of SGA was similar in the two groups (2.5% vs.1.8% OR 1.3[0.9-2.0] p=0.08). Conclusions Pregnant women whose OGTT curve is flat have a lower risk of delivering macrosomic neonates and undergoing unplanned cesarean delivery and are not at increased risk of adverse maternal or neonatal outcomes. More research is required to evaluate the relationship between different OGTT curves and th.

Quantitative cervicovaginal fluid fetal fibronectin: A liquid biopsy for intra-amniotic inflammation.

Intra-amniotic inflammation is causally linked to spontaneous preterm labor. The gold standard for the diagnosis of intra-amniotic inflammation is the determination of an amniotic fluid profile obtained from transabdominal amniocentesis, which is invasive. Cervicovaginal fluid fetal fibronectin (fFN) is a widely-used predictive biomarker for spontaneous preterm labor. The aims of this study are to determine (1) whether a quantitative cervicovaginal fluid fFN test can be used to identify the presence of intra-amniotic inflammation; and (2) an appropriate cut-off value of a cervicovaginal fluid fFN concentration for the identification of intra-amniotic inflammation. This prospective cohort study included 78 patients with preterm labor and intact membranes who had a sample collected for quantitative cervicovaginal fluid fFN measurement and underwent transabdominal amniocentesis. Intra-amniotic inflammation was defined as an amniotic fluid interleukin-6 concentration ≥2.6 ng/mL. Clinicians were masked from the results of cervicovaginal fluid fFN and amniotic fluid interleukin-6 concentrations. Logistic regression analysis was used to determine which factors were significant predictors of intra-amniotic inflammation. The diagnostic indices of the cervicovaginal fluid fFN test for the identification of intra-amniotic inflammation were calculated. (1) Frequency of intra-amniotic inflammation was 26.9% (21/78); (2) the higher the cervicovaginal fluid fFN concentration, the greater the risk of intra-amniotic inflammation (p < 0.001); (3) cervicovaginal fluid fFN concentration ≥125 ng/mL had an area under the curve of 0.91 (95% confidence interval: 0.83-0.96) for the identification of intra-amniotic inflammation with 100% sensitivity, 100% negative predictive value, 82.46% specificity and a positive likelihood ratio of 5.7; and (4) cervicovaginal fluid fFN cut-off of 125 ng/mL had a significant higher predictive performance than the traditional cut-off (50 ng/mL) for the identification of intra-amniotic inflammation. Quantitative cervicovaginal fluid fFN with a cut-off of 125 ng/mL had a high sensitivity and a negative predictive value as well as a positive likelihood ratio for the identification of intra-amniotic inflammation. Its high sensitivity and negative predictive value can be used to decrease an index of suspicion of intra-amniotic inflammation. This test may be useful as an initial assessment test to select appropriate patients for amniocentesis to determine intra-amniotic inflammation.

Changes of CD8+CD28- Tregs and Gamma-Delta-T-cells in a Neonate with Intrauterine Cytomegalovirus Infection: A Case Report

BackgroundCongenital cytomegalovirus (cCMV) infection can lead to a range of adverse outcomes. The majority of cCMV neonates with clinical symptoms are infected postnatally; however, established cases of intrauterine infection are uncommon, resulting in a paucity of reports on clinical findings and lymphocytes expression in CMV-infected neonates.Case presentationWe followed a neonate with cCMV infection from the onset of hospitalization to several months of follow-up. This infant was intrauterine CMV-positive in the amniotic fluid of the mother at 21 weeks’ gestation and received intravenous ganciclovir infusion and sequential oral valganciclovir after birth. The typical clinical signs manifested in the nervous system, liver, and peripheral blood and were documented during the hospitalizaion period and up to the follow-up visit. Flow cytometry was employed to examine the expression of T cells, their subsets, and the associated cytokines in peripheral blood samples at various time points. The flow data for the cCMV neonate were compared with those of the controls at each time point. Following treatment, clinical symptoms improved and the infant became CMV negative. However, developmental delays occurred later in life. The proportion of CD8+CD28− Tregs in the peripheral blood of the neonate with congenital CMV infection was higher than that in the controls at the three time points. The expression levels of perforin and granzyme B secreted by γδ T cells (Vδ1 and Vδ2 T cells), increased during the course of hospitalization until follow-up and were higher than those in the controls at the three time points.ConclusionsDespite the alleviation of clinical symptoms, developmental delay in later life remains inevitable in this intrauterine cCMV neonate. CD8+CD28− Tregs and Vδ1 and Vδ2 T cells secreting perforin and granzyme B may be involved in congenital CMV infection, although this hypothesis requires validation in a larger study. This report may contribute to our understanding of the effect of current treatment and the immune status of intrauterine cCMV-infected neonates.

Amnioinfusion by lactated Ringer’s solution via may reduce the toxicity on the exposed nerve tissue in spina bifida

Spina bifida is one of the most common developmental fetal malformations. It is a defect in dorsal spinal elements that exposes nerve tissue to the external environment – the amniotic fluid – leading to toxic exposure. By reviewing the development of fetal therapy, the physiopathology, and the mechanism of spina bifida, we hypothesize that the repetitive early replacement of the amniotic fluid with lactated Ringer’s solution via amnioinfusion may reduce toxicity to the exposed nerve tissue. Studies in fetal rat models suggest that amniotic fluid is toxic to the exposed spinal cord by an inflammatory response. Given the similarity between lactated Ringer’s solution and amniotic fluid, this replacement could mitigate toxicity without altering the intrauterine environment. Randomized controlled trials in animals and human fetuses are needed to test this hypothesis. If effective, this approach could benefit conservative couples and be applicable in low-income settings, improving outcomes after fetal surgery.

PSLBI-12 Screening and isolation of Fusobacterium necrophorum and Fusobacterium varium from seminal, and vagino-uterine microbiota of healthy cattle and sheep

Abstract Fusobacteria necrophorum is an important opportunistic bacterial pathogen associated with liver abscesses, foot rot, necrotic laryngitis and endometritis in both dairy and beef cattle. Recently, we identified F. necrophorum as one of the taxa in the uterine microbiota that are positively associated with pregnancy outcome in beef cows, and that Fusobacterium was identified as the most dominant genus in the bovine seminal microbiota. These findings indicate that Fusobacterium spp. may have a role in reproductive health and cattle fertility. Our objectives were to detect and isolate the F. necrophorum subsp necrophorum (FNN), F. necrophorum subsp. funduliforme (FNF) and F. varium (FV) in seminal, vaginal and uterine microbial ecosystems as well as fetal samples by culture and real-time PCR (qPCR) methods. The qPCR was performed on the DNA (n = 215) extracted from bull semen (n = 37), ram semen (n = 100), and vaginal (n = 25) and uterine swabs (n = 32), caruncles (n = 9), allantoic fluid (n = 6), and amniotic fluid (n = 6) from pregnant beef cows. The qPCR targeted the hgdA gene, and leukotoxin promoter regions (lktA-n and lktA-f) to detect and quantify FV, FNN and FNF, respectively. Cryopreserved samples (n = 174; 57 bull semen, 10 ram semen, 38 vaginal, and 47 uterine swabs) were cultured to detect and isolate FNN, FBF and FV by direct plating and after culturing in anaerobic peptone-yeast extract broth containing sodium lactate broth with josamycin, vancomycin and norfloxacin for selective enrichment of FN and FV. Also, 28 samples of 6-mo-old bovine fetuses caruncles, allantoic and amniotic fluids, and intestinal tissue) were screened for Fusobacterium species. By qPCR, FNF was detected in 75.7% of bull semen, and 8.0% of vaginal swabs as well as 33.3% of both allantoic fluid and caruncles, but was undetected in ram semen, uterine swabs, and amniotic fluids. Subspecies necrophorum was identified in 37.8% of bull semen, 3% of ram semen, 8.0% of vaginal and 15.6% of uterine 55.6 % of caruncles, 50% of allantoic fluid samples, but was not detected in amniotic fluid samples. Meanwhile FV was only detected in vaginal, uterine swabs and caruncles at 4%, 6.3%, and 11.1%, respectively. The culture method detected FNF in 63.2% of bull semen and 5.3% of vaginal samples, while FNN was identified in 1.8% of bull semen and 2.1% of uterine samples. F. varium was not isolated from any of the samples. Overall, FNF was the most prevalent subspecies present in the seminal microbiome of bulls. Identification of FNN subspecies and FV in seminal, vaginal, and uterine microbiota of healthy cattle, as well as in healthy calf fetus-associated samples suggests potential transfer from bulls to cows and then on to offspring, and potential involvement of the Fusobacterium spp. in cattle fertility.

Maternal Cytomegalovirus (CMV) Serology: The Diagnostic Limitations of CMV IgM and IgG Avidity in Detecting Congenital CMV Infection.

Congenital cytomegalovirus (cCMV) is a common congenital viral infection. Testing for cCMV usually begins with assessing maternal CMV serology, specifically IgM and IgG antibodies. A negative maternal CMV IgM suggests a low risk of recent maternal CMV infection, thereby suggesting a low risk of cCMV in the fetus. Consequently, cCMV is often ruled out when maternal CMV IgM is negative. In our perinatal autopsy and placental pathology database, we identified 5 cases of cCMV despite negative maternal CMV IgM results in the second trimester. In all 5 cases, fetal abnormalities were first detected by ultrasound in the second trimester, prompting maternal CMV testing. Since second trimester maternal CMV IgM was negative in all cases, cCMV was considered unlikely, thus precluding further prenatal CMV testing in 4 of these cases. The diagnosis of cCMV was subsequently made through placental and/or autopsy examinations. Following this diagnosis, retrospective CMV serology and IgG avidity testing was performed on stored frozen first-trimester maternal blood samples in 3 cases. Among these, the first-trimester samples in 2 cases were IgG+, IgM+, and exhibited low IgG avidity, suggesting a primary maternal CMV infection around the time of conception. In the third case, both first and second-trimester maternal blood samples were IgG+, IgM-, and showed high IgG avidity, suggesting a non-primary maternal CMV infection (i.e., reactivation or reinfection of CMV). A negative maternal CMV IgM in the second trimester cannot exclude cCMV infection. While CMV IgG avidity testing and analysis of stored frozen first-trimester maternal blood samples provide valuable insights, they have limitations. CMV PCR performed on amniotic fluid is a useful prenatal diagnostic tool. For cases of unexplained fetal abnormalities or death, autopsy and placental examination are recommended.

Clinical Characteristics and Outcomes of Intrauterine Blood Transfusion (IUT) for Infectious Etiologies.

Congenital viral infection may result in fetal anemia and thrombocytopenia. While intrauterine blood transfusions (IUTs) are more commonly performed for Rh alloimmunization, reports using IUT for infection have varying success. Our primary objective was to characterize the outcomes of patients undergoing IUT for infectious etiologies at our center compared with Rh disease. This was a case series of patients undergoing IUT from 2012-2023. Infectious etiologies were identified by maternal serologies and confirmed by amniotic fluid polymerase chain reactions (PCR). Clinical outcomes were obtained from electronic medical records. During the study period, 70 patients underwent IUT, 34% (24/70) for Rh alloimmunization and 17% (12/70) for infection. Those with infectious etiologies were more likely to be diagnosed at earlier gestational ages (22 vs. 25weeks, p=0.04), with hydrops (75 vs. 33%, p=0.03), and thrombocytopenia (27±33×103 vs. 163±112×103, p<0.01). Perinatal death was significantly greater in cases of CMV (4/5, 80%) compared to parvovirus (1/7, 14%) or Rh alloimmunization (5/24, 21%) (p=0.02). Anemias and thrombocytopenias related to fetal infection may be indications for IUT. Compared with Rh alloimmunization, IUT in fetal infections was performed significantly earlier, and hydrops were more common at the time of IUT. In the case of CMV, greater rates of IUFD (80%) were observed. Patients should be counseled on the various outcomes by indication.

Propagation of koi herpesvirus using embryonated chicken eggs: a potential substitute method for fish vaccine production?

Koi herpesvirus disease (KHVD) is caused by a large DNA virus that commonly infects carp, Cyprinus carpio (koi and common carp). KHVD has spread rapidly across the globe and caused high mortality in all ages of common carp and koi. Until now, no effective treatment has been applied to prevent KHV infection impacting the mass production of koi and common carp . An environmentally friendly alternative strategy for controlling fish disease is vaccination. One of the challenges facing conventional viral vaccine production is the requirement for koi or common carp cell cultures, which must be frequently maintained with expensive materials required for virus propagation. This study aims to obtain KHV that has been propagated in embryonated chicken eggs (ECE) to formulate an affordable KHV vaccine for the aquaculture industry. This research consisted of three stages; the first stage was virus inoculation into various parts of eggs (allantoic fluid, chorioallantoic membrane, amniotic fluid, and egg yolk). The second stage was observing viral growth and collecting ECE fluid and membranes. The third stage involved quantitatively determining the viral genomic copy numbers using the quantitative Polymerase Chain Reaction (qPCR) assay. KHV propagation in various parts of ECE resulted in varying viral genomic copy numbers with a high DNA copy number reported in allantoic fluid on the third day after inoculation. Further work is required to monitor virus titer in later passages and optimize methodology for using ECE as the potential alternative to cultured cells as the medium for virus propagation. In the future the system could be developed to produce promising vaccine candidates with more affordable vaccine prices for the Indonesian fish farming industry.

Transamniotic Fetal Immunotherapy with Secretory IgA: A Potential Novel Ancillary Strategy for the Prevention of Necrotizing Enterocolitis

Plain Language SummaryThis is an experimental study in an animal model showing that a special type of immunoglobulin called secretory immunoglobulin-A (SIgA), which can find its way to the fetal intestine after a simple injection into the amniotic fluid, is capable of attaching itself to intestinal bacteria after birth. The importance of that is the fact that SIgA has been previously shown to protect neonates, especially premature ones, from a dangerous disease called necrotizing enterocolitis (NEC). Since certain premature neonates cannot take SIgA by mouth, this new form of delivery of that immunoglobulin could become a novel minimally invasive alternative to give babies that molecule and hopefully help protect them from NEC. Further studies need to be performed before this can be used in humans, yet the original data presented in this study justify said studies while also suggesting that transamniotic fetal immunotherapy with SIgA could become a novel strategy for prevention of not only NEC but also possibly other perinatal infections.

Transamniotic Delivery of Surfactant Protein B mRNA in a Healthy Model

Introduction: We sought to determine whether exogenous surfactant protein B (SPB) mRNA could be incorporated and translated by the fetal lung after simple transamniotic administration. Methods: Fetuses (n = 149) of twelve time-dated dams underwent intra-amniotic injections of either human SPB (hSPB) mRNA encapsulated into lipopolyplex (mRNA, n = 99) or lipopolyplex without mRNA (control; n = 50) on gestational day 17 (E17, term = E21–22). Lungs were screened for hSPB by enzyme-linked immunosorbent assay daily until term. Phosphatidylcholine (PC) (a surrogate for surfactant production) was measured in the amniotic fluid by fluorometric assay. Statistical analysis included nonparametric Wilcoxon rank sum test. Results: Significantly improved survival in the mRNA group compared to controls was observed at E18 (100% vs. 85.7%) and E20 (100% vs. 83.3%) (both p < 0.001). When controlled by mRNA-free injections, hSPB protein was detected in the mRNA group’s lungs at E18, 19, and term (p = 0.002 to <0.001). Amniotic fluid PC levels were increased compared to control at term [285.9 (251.1, 363.9) μM versus 263.1 (222.8, 309.1) μM]; however, this did not reach significance (p = 0.33). Conclusions: Encapsulated exogenous SPB mRNA can be incorporated and translated by fetal lung cells following intra-amniotic injection in a healthy rat model. Transamniotic mRNA delivery could become a novel strategy for perinatal surfactant protein replacement.

Harnessing the power of child development records to detect early neurodevelopmental disorders using Bayesian analysis.

This study aims to analyse the developmental data from public health nurses (PHNs) to identify early indicators of neurodevelopmental disorders (NDDs) in young children using Bayesian network (BN) analysis to determine factor combinations that improve diagnosis accuracy. The study cohort was 501 children who underwent health checkups at 18 and 36-month. Data included demographics, pregnancy, delivery, neonatal factors, maternal interviews, and physical and neurological findings. Diagnoses were made by paediatricians and child psychiatrists using standardised tools. Predictive accuracy was assessed by the receiver operating characteristic (ROC) curve analysis. We identified several infant/toddler factors significantly associated with NDD diagnoses. Predictive factors included meconium-stained amniotic fluid, 1 min Apgar score, and early developmental milestones. ROC curve analysis showed varying predictive accuracies based on evaluation timing. The 10-month checkup was valid for screening but less reliable for excluding low-risk cases. The 18-month evaluation accurately identified children at NDD risk. The study demonstrates the potential of using developmental records for early NDD detection, emphasising early monitoring and intervention for at-risk children. These findings could guide future infant mental health initiatives in the community.