Wastewater surveillance for bacterial targets: current challenges and future goals.

Wastewater-based epidemiology (WBE) expanded rapidly in response to the COVID-19 pandemic. As the public health emergency has ended, researchers and practitioners are looking to shift the focus of existing wastewater surveillance programs to other targets, including bacteria. Bacterial targets may pose some unique challenges for WBE applications. To explore the current state of the field, the National Science Foundation-funded Research Coordination Network (RCN) on Wastewater Based Epidemiology for SARS-CoV-2 and Emerging Public Health Threats held a workshop in April 2023 to discuss the challenges and needs for wastewater bacterial surveillance. The targets and methods used in existing programs were diverse, with twelve different targets and nine different methods listed. Discussions during the workshop highlighted the challenges in adapting existing programs and identified research gaps in four key areas: choosing new targets, relating bacterial wastewater data to human disease incidence and prevalence, developing methods, and normalizing results. To help with these challenges and research gaps, the authors identified steps the larger community can take to improve bacteria wastewater surveillance. This includes developing data reporting standards and method optimization and validation for bacterial programs. Additionally, more work is needed to understand shedding patterns for potential bacterial targets to better relate wastewater data to human infections. Wastewater surveillance for bacteria can help provide insight into the underlying prevalence in communities, but much work is needed to establish these methods.IMPORTANCEWastewater surveillance was a useful tool to elucidate the burden and spread of SARS-CoV-2 during the pandemic. Public health officials and researchers are interested in expanding these surveillance programs to include bacterial targets, but many questions remain. The NSF-funded Research Coordination Network for Wastewater Surveillance of SARS-CoV-2 and Emerging Public Health Threats held a workshop to identify barriers and research gaps to implementing bacterial wastewater surveillance programs.

Quaternary Ammonium Compounds: A Chemical Class of Emerging Concern.

Quaternary ammonium compounds (QACs), a large class of chemicals that includes high production volume substances, have been used for decades as antimicrobials, preservatives, and antistatic agents and for other functions in cleaning, disinfecting, personal care products, and durable consumer goods. QAC use has accelerated in response to the COVID-19 pandemic and the banning of 19 antimicrobials from several personal care products by the US Food and Drug Administration in 2016. Studies conducted before and after the onset of the pandemic indicate increased human exposure to QACs. Environmental releases of these chemicals have also increased. Emerging information on adverse environmental and human health impacts of QACs is motivating a reconsideration of the risks and benefits across the life cycle of their production, use, and disposal. This work presents a critical review of the literature and scientific perspective developed by a multidisciplinary, multi-institutional team of authors from academia, governmental, and nonprofit organizations. The review evaluates currently available information on the ecological and human health profile of QACs and identifies multiple areas of potential concern. Adverse ecological effects include acute and chronic toxicity to susceptible aquatic organisms, with concentrations of some QACs approaching levels of concern. Suspected or known adverse health outcomes include dermal and respiratory effects, developmental and reproductive toxicity, disruption of metabolic function such as lipid homeostasis, and impairment of mitochondrial function. QACs' role in antimicrobial resistance has also been demonstrated. In the US regulatory system, how a QAC is managed depends on how it is used, for example in pesticides or personal care products. This can result in the same QACs receiving different degrees of scrutiny depending on the use and the agency regulating it. Further, the US Environmental Protection Agency's current method of grouping QACs based on structure, first proposed in 1988, is insufficient to address the wide range of QAC chemistries, potential toxicities, and exposure scenarios. Consequently, exposures to common mixtures of QACs and from multiple sources remain largely unassessed. Some restrictions on the use of QACs have been implemented in the US and elsewhere, primarily focused on personal care products. Assessing the risks posed by QACs is hampered by their vast structural diversity and a lack of quantitative data on exposure and toxicity for the majority of these compounds. This review identifies important data gaps and provides research and policy recommendations for preserving the utility of QAC chemistries while also seeking to limit adverse environmental and human health effects.

Use of hemagglutinin and neuraminidase amplicon-based high-throughput sequencing with variant analysis to detect co-infection and resolve identical consensus sequences of seasonal influenza in a university setting.

BACKGROUND: Local transmission of seasonal influenza viruses (IVs) can be difficult to resolve. Here, we study if coupling high-throughput sequencing (HTS) of hemagglutinin (HA) and neuraminidase (NA) genes with variant analysis can resolve strains from local transmission that have identical consensus genome. We analyzed 24 samples collected over four days in January 2020 at a large university in the US. We amplified complete hemagglutinin (HA) and neuraminidase (NA) genomic segments followed by Illumina sequencing. We identified consensus complete HA and NA segments using BLASTn and performed variant analysis on strains whose HA and NA segments were 100% similar. RESULTS: Twelve of the 24 samples were PCR positive, and we detected complete HA and/or NA segments by de novo assembly in 83.33% (10/12) of them. Similarity and phylogenetic analysis showed that 70% (7/10) of the strains were distinct while the remaining 30% had identical consensus sequences. These three samples also had IAV and IBV co-infection. However, subsequent variant analysis showed that they had distinct variant profiles. While the IAV HA of one sample had no variant, another had a T663C mutation and another had both C1379T and C1589A. CONCLUSION: In this study, we showed that HTS coupled with variant analysis of only HA and NA genes can help resolve variants that are closely related. We also provide evidence that during a short time period in the 2019-2020 season, co-infection of IAV and IBV occurred on the university campus and both 2020/2021 and 2021/2022 WHO recommended H1N1 vaccine strains were co-circulating.

Decline and Pronounced Regional Disparities in Medical Cocaine Usage in the United States.

Background: Cocaine is a stimulant and Schedule II drug used as a local anesthetic and vasoconstrictor. Objective: This descriptive study characterized medical cocaine use in the United States. Methods: Retail drug distribution data from 2002 to 2017 were extracted for each state from the Drug Enforcement Administration, which reports on medical, research, and analytical chemistry use. The percentage of buyers (pharmacies, hospitals, and providers) was obtained. Use per state, corrected for population, was determined. Available cross-sectional data on cocaine use as reported by the Medicare and Medicaid programs for 2013-2017 and electronic medical records were examined. Results: Medical cocaine use decreased by -62.5% from 2002 to 2017. Hospitals accounted for 84.9% and practitioners for 9.9% of cocaine distribution in 2017. The number of pharmacies carrying cocaine dropped by -69.4%. The percentages of hospitals, practitioners, and pharmacies that carried cocaine in 2017 were 38.4%, 2.3%, and 0.3%, respectively. There was a 7-fold difference in 2002 (South Dakota, 76.1 mg/100 persons; Delaware, 10.1 mg/100 persons). Relative to the average state in 2017, those reporting the highest values (Montana, 20.1; North Dakota, 24.1 mg/100 persons) were significantly elevated. Cocaine use within the Medicare and Medicaid programs was negligible. Cocaine use within the Geisinger system was rare from 2002 to 2007 (<4 orders/100 000 patients per year) but increased to 48.7 in 2018. Conclusion and Relevance: If these pharmacoepidemiological patterns continue, licit cocaine may soon become a historical relic. The pharmacology and pharmacotherapeutics education of health care providers may need to be adjusted accordingly.

Nationwide Mass Inventory and Degradation Assessment of Plastic Contact Lenses in US Wastewater.

Plastics pose ecological and human health risks, with disposable contact lenses constituting a potential high-volume pollution source. Using sales data and an online survey of lens users (n = 416) alongside laboratory and field experiments at a conventional sewage treatment plant, we determined the environmental fate and mass inventories of contact lenses in the United States. The survey results revealed that 21 ± 0.8% of lens users flush their used lenses down the drain, a loading equivalent to 44 000 ± 1700 kg y-1 of lens dry mass discharged into US wastewater. Biological treatment of wastewater did not result in a measurable loss of plastic mass (p = 0.001) and caused only very limited changes in the polymer structure, as determined by µ-Raman spectroscopy. During sewage treatment, the lenses were found to accumulate as fragments in sewage sludge, resulting in an estimated accumulation of 24 000 ± 940 kg y-1 of microplastics destined for application on US agricultural soils contained in sewage sludge. Recycling of the contact lenses and their packaging amounted to only 0.04% of the total waste volume associated with contact lens use. This is the first study to identify contact lenses and more specifically silicone hydrogels, as a previously overlooked source of plastic and microplastic pollution.

Methyl mercury, but not inorganic mercury, associated with higher blood pressure during pregnancy.

Prior studies addressing associations between mercury and blood pressure have produced inconsistent findings; some of this may result from measuring total instead of speciated mercury. This cross-sectional study of 263 pregnant women assessed total mercury, speciated mercury, selenium, and n-3 polyunsaturated fatty acids in umbilical cord blood and blood pressure during labor and delivery. Models with a) total mercury or b) methyl and inorganic mercury were evaluated. Regression models adjusted for maternal age, race/ethnicity, prepregnancy body mass index, neighborhood income, parity, smoking, n-3 fatty acids and selenium. Geometric mean total, methyl, and inorganic mercury concentrations were 1.40µg/L (95% confidence interval: 1.29, 1.52); 0.95µg/L (0.84, 1.07); and 0.13µg/L (0.10, 0.17), respectively. Elevated systolic BP, diastolic BP, and pulse pressure were found, respectively, in 11.4%, 6.8%, and 19.8% of mothers. In adjusted multivariable models, a one-tertile increase of methyl mercury was associated with 2.83mmHg (0.17, 5.50) higher systolic blood pressure and 2.99mmHg (0.91, 5.08) higher pulse pressure. In the same models, an increase of one tertile of inorganic mercury was associated with -1.18mmHg (-3.72, 1.35) lower systolic blood pressure and -2.51mmHg (-4.49, -0.53) lower pulse pressure. No associations were observed with diastolic pressure. There was a non-significant trend of higher total mercury with higher systolic blood pressure. We observed a significant association of higher methyl mercury with higher systolic and pulse pressure, yet higher inorganic mercury was significantly associated with lower pulse pressure. These results should be confirmed with larger, longitudinal studies.

Prenatal exposure to tobacco smoke leads to increased mitochondrial DNA content in umbilical cord serum associated to reduced gestational age.

We investigated if prenatal exposures to tobacco smoke lead to changes in mitochondrial DNA content (mtDNA) in cord serum and adversely affect newborns' health. Umbilical cord serum cotinine levels were used to determine in utero exposure to smoking. Cord serum mtDNA was measured by quantitative polymerase chain reaction analysis of the genes coding for cytochrome c oxidase1 (MT-CO1) and cytochrome c oxidase2 (MT-CO2). Log transformed levels of mtDNA coding for MT-CO1 and MT-CO2 were significantly higher among infants of active smokers with higher serum level of cotinine (p < 0.05) and inversely associated with gestational age (p = 0.08; p = 0.02). Structural equation modeling results confirmed a positive association between cotinine and MT-CO1 and2 (p < 0.01) and inverse associations with gestational age (p = 0.02) and IGF-1 (p < 0.01). We identified a dose-dependent increase in the level of MT-CO1 and MT-CO2 associated to increased cord serum cotinine and decreased gestational age.

Mass Balance Assessment for Six Neonicotinoid Insecticides During Conventional Wastewater and Wetland Treatment: Nationwide Reconnaissance in United States Wastewater.

Occurrence and removal of six high-production high-volume neonicotinoids was investigated in 13 conventional wastewater treatment plants (WWTPs) and one engineered wetland. Flow-weighted daily composites were analyzed by isotope dilution liquid chromatography tandem mass spectrometry, revealing the occurrence of imidacloprid, acetamiprid, and clothianidin at ng/L concentrations in WWTP influent (60.5 ± 40.0; 2.9 ± 1.9; 149.7 ± 289.5, respectively) and effluent (58.5 ± 29.1; 2.3 ± 1.4; 70.2 ± 121.8, respectively). A mass balance showed insignificant removal of imidacloprid (p = 0.09, CI = 95%) and limited removal of the sum of acetamiprid and its degradate, acetamiprid-N-desmethyl (18 ± 4%, p = 0.01, CI = 95%). Clothianidin was found only intermittently, whereas thiamethoxam, thiacloprid, and dinotefuran were never detected. In the wetland, no removal of imidacloprid or acetamiprid was observed. Extrapolation of data from 13 WWTPs to the nation as a whole suggests annual discharges on the order of 1000-3400 kg/y of imidacloprid contained in treated effluent to surface waters nationwide. This first mass balance and first United States nationwide wastewater reconnaissance identified imidacloprid, acetamiprid, and clothianidin as recalcitrant sewage constituents that persist through wastewater treatment to enter water bodies at significant loadings, potentially harmful to sensitive aquatic invertebrates.

Mass Balance of Fipronil and Total Toxicity of Fipronil-Related Compounds in Process Streams during Conventional Wastewater and Wetland Treatment.

Attenuation of the pesticide fipronil and its major degradates was determined during conventional wastewater treatment and wetland treatment. Analysis of flow-weighted composite samples by liquid and gas chromatography-tandem mass spectrometry showed fipronil occurrence at 12-31 ng/L in raw sewage, primary effluent, secondary effluent, chlorinated effluent, and wetland effluent. Mean daily loads of total fipronil related compounds in raw sewage and in plant effluent after chlorination were statistically indistinguishable (p = 0.29; n = 10), whereas fipronil itself was partially removed (25 ± 3%; p = 0.00025; n = 10); the associated loss in toxicity was balanced by the formation of toxic fipronil degradates, showing conventional treatment to be unfit for reducing overall toxicity. In contrast to these findings at the municipal wastewater treatment, both parental fipronil and the sum of fipronil-related compounds were removed in the wetland with efficiencies of 44 ± 4% and 47 ± 13%, respectively. Total fipronil concentrations in plant effluent (28 ± 6 ng/L as fipronil) were within an order of magnitude of half-maximal effective concentrations (EC50) of nontarget invertebrates. This is the first systematic assessment of the fate of fipronil and its major degradates during full-scale conventional wastewater and constructed wetland treatment.

Organic Contaminants in Chinese Sewage Sludge: A Meta-Analysis of the Literature of the Past 30 Years.

The production of sewage sludge is increasing in China but with unsafe disposal practices, causing potential risk to human health and the environment. Using literature from the past 30 years (N = 159), we conducted a meta-analysis of organic contaminants (OCs) in Chinese sludge. Most data were available from developed and populated regions, and no data were found for Tibet. Since 1987, 35 classes of chemicals consisting of 749 individual compounds and 1 mixture have been analyzed, in which antibiotics and polycyclic aromatic hydrocarbons (PAHs) were the most targeted analytes. For 13 classes of principal OCs (defined as chemicals detected in over five studies) in sludge, the median (expressed in nanograms per gram dry weight) was the highest for phthalate esters (27 900), followed by alkylphenol polyethoxylates (12 000), synthetic musks (5800), antibiotics (4240), PAHs (3490), ultraviolet stabilizers (670), bisphenol analogs (160), organochlorine pesticides (110), polybrominated diphenyl ethers (100), pharmaceuticals (84), hormones (69), perfluorinated compounds (21), and polychlorinated biphenyls (15). Concentrations of PAHs in sludges collected between 1998 and 2012 showed a decreasing trend. Study findings suggest the need for a Chinese national sewage sludge survey to identify and regulate toxic OCs, ideally employing both targeted as well as nontargeted screening approaches.

Antimicrobial Chemicals Are Associated with Elevated Antibiotic Resistance Genes in the Indoor Dust Microbiome.

Antibiotic resistance is increasingly widespread, largely due to human influence. Here, we explore the relationship between antibiotic resistance genes and the antimicrobial chemicals triclosan, triclocarban, and methyl-, ethyl-, propyl-, and butylparaben in the dust microbiome. Dust samples from a mixed-use athletic and educational facility were subjected to microbial and chemical analyses using a combination of 16S rRNA amplicon sequencing, shotgun metagenome sequencing, and liquid chromatography tandem mass spectrometry. The dust resistome was characterized by identifying antibiotic resistance genes annotated in the Comprehensive Antibiotic Resistance Database (CARD) from the metagenomes of each sample using the Short, Better Representative Extract Data set (ShortBRED). The three most highly abundant antibiotic resistance genes were tet(W), blaSRT-1, and erm(B). The complete dust resistome was then compared against the measured concentrations of antimicrobial chemicals, which for triclosan ranged from 0.5 to 1970 ng/g dust. We observed six significant positive associations between the concentration of an antimicrobial chemical and the relative abundance of an antibiotic resistance gene, including one between the ubiquitous antimicrobial triclosan and erm(X), a 23S rRNA methyltransferase implicated in resistance to several antibiotics. This study is the first to look for an association between antibiotic resistance genes and antimicrobial chemicals in dust.

Mass Balance Model for Sustainable Phosphorus Recovery in a US Wastewater Treatment Plant.

In response to limited phosphorus (P) reserves worldwide, several countries have demonstrated the prospect of recovering significant amounts of P from wastewater treatment plants (WWTPs). This technique uses enhanced biological P removal (EBPR) to concentrate P in sludge followed by chemical precipitation of P as struvite, a usable phosphate mineral. The present study models the feasibility of this enhanced removal and recovery technique in a WWTP in Arizona with design parameters typical of infrastructure in the United States. A mass balance was performed for existing treatment processes and modifications proposed to estimate the quantity of P that could be recovered under current and future flow conditions. Modeling results show that about 71 to 96% of the P being lost potentially could be recovered as struvite. About 491 ± 64 t yr of struvite may be recovered after process modification, which corresponds to $150,000 ± $20,000 yr in P sales to fertilizer industries. The process was projected to be economically feasible, with a payback period of 45 ± 30 yr in the studied WWTP and a much shorter duration of 3 ± 1 yr for WWTPs already using an EBPR process. Furthermore, modeling results suggest that P recovery can improve the quality of biosolids by favorably reducing the P:N ratio. Implementation of this strategy at US WWTPs may increase national security by reducing dependence of limited P resources. Considering all aspects of the recovery process with respect to environmental, economic, and social implications, the examined technique is concluded to represent a cost-attractive and sustainable method for P management in US WWTPs.

Bioaccumulation of Legacy and Emerging Organochlorine Contaminants in Lumbriculus variegatus.

Freshwater sediment-dwelling Lumbriculus variegatus is known to serve as a vector for the transfer of contaminants from sediments to higher trophic level organisms, but limited data exist on the bioaccumulation of chemicals associated with sediments containing high total organic carbon (TOC). In the current study, sediments from the north shore area of Lake Apopka (Florida, USA), containing very high TOC [39 % (w/w)], were spiked with four chemicals-p,p'-dichlorordiphenyldichloroethylene (p,p'-DDE), dieldrin, fipronil, and triclosan-individually or in a mixture of the four and then used for bioaccumulation studies. Tissue concentrations of chemicals in L. variegatus were measured at 2, 7, 14, 21, and 28 days of exposure, and the bioaccumulation potential was evaluated using biosediment accumulation factors [BSAF (goc/glipid)]. Increase in total body burdens of all four chemicals in L. variegatus was rapid at day 2 and reached a steady-state level after 7 days in both single and mixture experiments. Tissue concentrations of fipronil peaked after 2 days and then decreased by 70 % in sediment experiments suggesting that in addition to the degradation of fipronil that occurred in the sediment, L. variegatus may also be able to metabolize fipronil. The calculated 28-day BSAF values varied among the chemicals and increased in the order fipronil (1.1) < triclosan (1.4) < dieldrin (21.8) < p,p'-DDE (49.8) in correspondence with the increasing degree of their hydrophobicity. The relatively high BSAF values for p,p'-DDE and dieldrin probably resulted from lower-than-expected sorption of chemicals to sediment organic matter either due to the nature of the plant-derived organic matter, as a result of the relatively short equilibration time among the various compartments, or due to ingestion of sediment particles by the worms.

Elucidating the molecular basis of adverse health effects from exposure to anthropogenic polyfluorinated compounds using toxicoproteomic approaches.

Linear, short-chain polyfluorinated and perfluorinated alkyl compounds, often referred to as PFCs, have been in worldwide use as surfactants and polymer precursors for decades, and environmental dispersal of these highly persistent compounds represents a public health threat. Whereas ubiquitous low-level exposure to these compounds has been demonstrated in human populations from around the world, the exact mechanisms of toxicity and their toxic potency remain subject to investigation and scientific dispute. As with other environmental exposures, a major hurdle for gaining a better understanding of their human health impacts is the limited utility of cell culture and animal models serving as convenient, yet imperfect proxies to human physiology and disease. The present communication provides a brief overview of the current understanding of potential health effects of PFC exposure and examines how new toxicoproteomic methodologies can provide insight into the molecular mechanism of PFC exposure. Furthermore, we showcase an exemplary data set to illustrate how toxicoproteomic, population-wide studies might overcome limitations of animal models to more fully understand the metabolism and effects of PFCs and other environmental stressors where it matters most, in human populations experiencing real-world, chronic, low-level exposures.

Occurrence of Bisphenol A Diglycidyl Ethers (BADGEs) and Novolac Glycidyl Ethers (NOGEs) in Archived Biosolids from the U.S. EPA's Targeted National Sewage Sludge Survey.

Epoxy resins incorporating bisphenol A diglycidyl ether (BADGE) and novolac glycidyl ether (NOGE) are used in a wide range of applications, including adhesives, structural and electrical laminates. However, little is known about the occurrence of BADGE, NOGE, and their derivatives in the environment. Using liquid chromatography-tandem mass spectrometry, BADGE, bisphenol F glycidyl ether (BFDGE), 3-ring NOGE, and eight of their derivatives (BADGE·2 H2O, BADGE·H2O, BADGE·HCl·H2O, BADGE·2 HCl, BADGE·HCl, BFDGE·2 H2O, and BFDGE·2 HCl) were determined in archived biosolid samples collected from 68 wastewater treatment plants (WWTPs) from the northeastern, midwestern, western, and southern regions of the USA. BADGE·2 H2O was the most frequently detected (DR = 99%) and the most abundant compound found (median: 93.6 ng/g dry weight [dw]) in this family. The highest total concentrations of target chemicals, ranging from 83.6 to 2490 ng/g dw, were found in biosolids collected from the northeastern United States. The sum of geometric mean (GM) concentration of BADGE, NOGE, and their derivatives in biosolids increased with the treatment capacity of WWTPs. Based on the measured concentrations in biosolids and predicted mass in wastewater, it was estimated that approximately 3.5% of the total production of BADGEs was emitted through WWTP discharges.

Characterization, Recovery Opportunities, and Valuation of Metals in Municipal Sludges from U.S. Wastewater Treatment Plants Nationwide.

U.S. sewage sludges were analyzed for 58 regulated and nonregulated elements by ICP-MS and electron microscopy to explore opportunities for removal and recovery. Sludge/water distribution coefficients (KD, L/kg dry weight) spanned 5 orders of magnitude, indicating significant metal accumulation in biosolids. Rare-earth elements and minor metals (Y, La, Ce, Pr, Nd, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb, Lu) detected in sludges showed enrichment factors (EFs) near unity, suggesting dust or soils as likely dominant sources. In contrast, most platinum group elements (i.e., Ru, Rh, Pd, Pt) showed high EF and KD values, indicating anthropogenic sources. Numerous metallic and metal oxide colloids (<100-500 nm diameter) were detected; the morphology of abundant aggregates of primary particles measuring <100 nm provided clues to their origin. For a community of 1 million people, metals in biosolids were valued at up to US$13 million annually. A model incorporating a parameter (KD × EF × $Value) to capture the relative potential for economic value from biosolids revealed the identity of the 13 most lucrative elements (Ag, Cu, Au, P, Fe, Pd, Mn, Zn, Ir, Al, Cd, Ti, Ga, and Cr) with a combined value of US $280/ton of sludge.

Use of amniotic fluid for determining pregnancies at risk of preterm birth and for studying diseases of potential environmental etiology.

Amniotic fluid (AF) is a biological medium uniquely suited for the study of early exposure of the human fetus to environmental contaminants acquired by the mother before and during pregnancy. Traditional diagnostic applications of AF have focused almost exclusively on the diagnosis of genetic aberrations such as Trisomy-21 and on heritable diseases in high-risk pregnancies. Since more than 50 anthropogenic compounds have been detected in AF, there is considerable potential in utilizing fetal protein biomarkers as indicators of health effects related to prenatal toxic exposure. Here, we focus on preterm birth (PTB) to illustrate opportunities and limitations of using AF as a diagnostic matrix. Representing a pervasive public health challenge worldwide, PTB cannot be managed simply by improving hygiene and broadening access to healthcare. This is illustrated by 15-year increases of PTB in the U.S. from 1989 to 2004. AF is uniquely suited as a matrix for early detection of the association between fetal exposures and PTB due to its fetal origin and the fact that it is sampled from women who are at higher risk of PTB. This critical review shows the occurrence in AF of a number of xenobiotics, including endocrine-disrupting compounds (EDCs), which are known or may reasonably be expected to shorten fetal gestation. It is not yet known whether EDCs, including bisphenol A, phytoestrogens, and polychlorinated biphenyls (PCBs), can affect the expression of proteins considered viable or potential biomarkers for the onset of PTB. As such, the diagnostic value of AF is broad and has not yet been fully explored for prenatal diagnosis of pregnancies at risk from toxic, environmental exposures and for the elucidation of mechanisms underlying important public health challenges including PTB.