Fate of Antibiotics and Antibiotic Resistance during Digestion and Composting: A Review.

Antibiotics and antibiotic-resistant bacteria (ARB) enter the environment through municipal and agricultural waste streams and pose a potential risk to human and livestock health through either direct exposure to antibiotic-resistant pathogens or selective pressure on the soil microbial community. This review summarizes current literature on the fate of antibiotics, ARB, and antibiotic resistance genes (ARGs) during anaerobic digestion and composting of manure and wastewater residuals. Studies have shown that removal of antibiotics varies widely during mesophilic anaerobic digestion, even within the same class of antibiotics. Research on ARB shows a wide range of removal under mesophilic conditions, with nearly complete removal under thermophilic conditions. Research on 16 antibiotics in 11 different studies using both bench-scale and farm-scale composting systems demonstrates that composting significantly reduces levels of extractable antibiotics in livestock manure in nearly all cases. Calculated half-lives ranged from 0.9 to 16 d for most antibiotics. There is more limited evidence that levels of ARB are also reduced by composting. Studies of the fate of ARGs show mixed evidence for removal during both mesophilic and thermophilic anaerobic digestion and during thermophilic composting. Antibiotic resistance genes are DNA structures, so they may persist until the DNA structure is degraded, yet the bacterium may have been rendered nonviable long before the DNA is completely degraded. Additional research would be of value to determine optimum anaerobic digestion and composting conditions for removal of ARB and to increase understanding of the fate of ARGs during anaerobic digestion and composting.

Not All Antibiotic Use Practices in Food-Animal Agriculture Afford the Same Risk.

The World Health Organization has identified quinolones, third- and fourth-generation cephalosporins, and macrolides as the most important antibiotics in human medicine. In the context of agricultural use of antibiotics, the principle zoonotic agents of concern are , spp., , and spp. Antibiotic exposure provides a selective advantage to resistant strains of these bacteria relative to their susceptible conspecifics. This is a dose-dependent process, and consequently antibiotic use practices that involve higher doses will exert greater and longer-lasting selective pressure in favor of resistant bacterial populations and will therefore increase the probability of transmission to people and other animals. Oral administration has a greater impact on enteric flora with the exception of fluoroquinolone treatments, which appear to affect the enteric flora equally if administered orally or parenterally. The use of quinolones in agriculture deserves heightened scrutiny because of the ease with which these broad-spectrum antibiotics favor spontaneously resistant bacteria in exposed populations. When present at sufficient concentrations, excreted antibiotics have the potential to selectively favor resistant bacteria in the environment and increase the probability of transmission to people and animals. The bioavailability of antibiotics varies greatly: some antibiotics remain active in soils (florfenicol, ß-lactams), whereas others may be rapidly sorbed and thus not bioavailable (tetracycline, macrolides, quinolones). When considering the risks of different antibiotic use practices in agriculture, it would be prudent to focus attention on practices that involve high doses, oral delivery, and residues of antibiotics that remain active in soils.

Hydrolysis of amphenicol and macrolide antibiotics: Chloramphenicol, florfenicol, spiramycin, and tylosin.

Antibiotics that enter the environment can present human and ecological health risks. An understanding of antibiotic hydrolysis rates is important for predicting their environmental persistence as biologically active contaminants. In this study, hydrolysis rates and Arrhenius constants were determined as a function of pH and temperature for two amphenicol (chloramphenicol and florfenicol) and two macrolide (spiramycin and tylosin) antibiotics. Antibiotic hydrolysis rates in pH 4-9 buffer solutions at 25°C, 50°C, and 60°C were quantified, and degradation products were characterized. All of the antibiotics tested remained stable and exhibited no observable hydrolysis under ambient conditions typical of aquatic ecosystems. Acid- and base-catalyzed hydrolysis occurred at elevated temperatures (50-60°C), and hydrolysis rates increased considerably below pH 5 and above pH 8. Hydrolysis rates also increased approximately 1.5- to 2.9-fold for each 10°C increase in temperature. Based on the degradation product masses found, the functional groups that underwent hydrolysis were alkyl fluoride, amide, and cyclic ester (lactone) moieties; some of the resultant degradation products may remain bioactive, but to a lesser extent than the parent compounds. The results of this research demonstrate that amphenicol and macrolide antibiotics persist in aquatic systems under ambient temperature and pH conditions typical of natural waters. Thus, these antibiotics may present a risk in aquatic ecosystems depending on the concentration present.

pH and temperature effects on the hydrolysis of three ß-lactam antibiotics: ampicillin, cefalotin and cefoxitin.

An understanding of antibiotic hydrolysis rates is important for predicting their environmental persistence. Hydrolysis rates and Arrhenius constants were determined as a function of pH and temperature for three common ß-lactam antibiotics, ampicillin, cefalotin, and cefoxitin. Antibiotic hydrolysis rates at pH4-9 at 25 °C, 50 °C, and 60 °C were quantified, and degradation products were identified. The three antibiotics hydrolyzed under ambient conditions (pH7 and 25 °C); half-lives ranged from 5.3 to 27 d. Base-catalyzed hydrolysis rates were significantly greater than acid-catalyzed and neutral pH hydrolysis rates. Hydrolysis rates increased 2.5- to 3.9-fold for a 10 °C increase in temperature. Based on the degradation product masses found, the likely functional groups that underwent hydrolysis were lactam, ester, carbamate, and amide moieties. Many of the proposed products resulting from the hydrolysis of ampicillin, cefalotin, and cefoxitin likely have reduced antimicrobial activity because many products contained a hydrated lactam ring. The results of this research demonstrate that ß-lactam antibiotics hydrolyze under ambient pH and temperature conditions. Degradation of ß-lactam antibiotics will likely occur over several weeks in most surface waters and over several days in more alkaline systems.

The effects of the antibiotics ampicillin, florfenicol, sulfamethazine, and tylosin on biogas production and their degradation efficiency during anaerobic digestion.

The impacts of four common animal husbandry antibiotics (ampicillin, florfenicol, sulfamethazine, and tylosin) on anaerobic digestion (AD) treatment efficiency and the potential for antibiotic degradation during digestion were evaluated. Sulfamethazine and ampicillin exhibited no impact on total biogas production up to 280 and 350 mg/L, respectively, although ampicillin inhibited biogas production rates during early stages of AD. Tylosin reduced biogas production by 10-38% between 130 and 913 mg/L. Florfenicol reduced biogas by ≈ 5%, 40% and 75% at 6.4, 36 and 210 mg/L, respectively. These antibiotic concentrations are higher than commonly seen for mixed feedlot manure, so impacts on full scale AD should be minimal. Antibiotic degradation products were found, confirming AD effectively degraded ampicillin, florfenicol, and tylosin, although some products were persistent throughout the process. Contamination of AD solid and liquid effluents with sulfamethazine and antibiotic transformation products from florfenicol and tylosin could present an environmental concern.

ß-lactams and florfenicol antibiotics remain bioactive in soils while ciprofloxacin, neomycin, and tetracycline are neutralized.

It is generally assumed that antibiotic residues in soils select for antibiotic-resistant bacteria. This assumption was tested by separately adding 10 different antibiotics (≥200 ppm) to three soil-water slurries (silt-loam, sand-loam, and sand; 20% soil [wt/vol]) and incubating mixtures for 24 h at room temperature. The antibiotic activity of the resultant supernatant was assessed by culturing a sensitive Escherichia coli strain in the filter-sterilized supernatant augmented with Luria-Bertani broth. We found striking differences in the abilities of supernatants to suppress growth of the indicator E. coli. Ampicillin, cephalothin, cefoxitin, ceftiofur, and florfenicol supernatants completely inhibited growth while bacterial growth was uninhibited in the presence of neomycin, tetracycline, and ciprofloxacin supernatants. High-performance liquid chromatography (HPLC) analysis demonstrated that cefoxitin and florfenicol were almost completely retained in the supernatants, whereas tetracycline and ciprofloxacin were mostly removed. Antibiotic dissipation in soil, presumably dominated by adsorption mechanisms, was sufficient to neutralize 200 ppm of tetracycline; this concentration is considerably higher than reported contamination levels. Soil pellets from the tetracycline slurries were resuspended in a minimal volume of medium to maximize the interaction between bacteria and soil particles, but sensitive bacteria were still unaffected by tetracycline (P = 0.6). Thus, residual antibiotics in soil do not necessarily exert a selective pressure, and the degree to which the pharmaceutical remains bioactive depends on the antibiotic. Efforts to control antibiotic contamination would be better directed toward compounds that retain biological activity in soils (e.g., cephalosporins and florfenicol) because these are the antibiotics that could exert a selective pressure in the environment.

High frequencies of elevated alkaline phosphatase activity and rickets exist in extremely low birth weight infants despite current nutritional support.

BACKGROUND: Osteopenia and rickets are common among extremely low birth weight infants (ELBW, <1000 g birth weight) despite current practices of vitamin and mineral supplementation. Few data are available evaluating the usual course of markers of mineral status in this population. Our objectives in this study were to determine the relationship between birth weight (BW) and peak serum alkaline phosphatase activity (P-APA) in ELBW infants and evaluate our experience with the diagnosis of rickets in these infants. METHODS: We evaluated all ELBW infants admitted to Texas Children's Hospital NICU in 2006 and 2007. Of 211 admissions, we excluded 98 patients who were admitted at >30 days of age or did not survive/stay for >6 weeks. Bone radiographs obtained in 32 infants were reviewed by a radiologist masked to laboratory values. RESULTS: In this cohort of 113 infants, P-APA was found to have a significant inverse relationship with BW, gestational age and serum phosphorus. In paired comparisons, P-APA of infants <600 g (957 +/- 346 IU/L, n = 20) and infants 600-800 g (808 +/- 323 IU/L, n = 43) were both significantly higher than P-APA of infants 800-1000 g (615 +/- 252 IU/L, n = 50), p < 0.01. Thirty-two patients had radiographic evaluation for evidence of rickets, based on P-APA greater than 800 IU/L, parenteral nutrition greater than 3 to 4 weeks, or clinical suspicion. Of these, 18 showed radiologic rickets and 14 showed osteopenia without rickets. Infants with BW <600 g were more likely to have radiologic rickets (10/20 infants) compared to those with BW 600-800 g (6/43 infants) and BW 800-1000 g (2/50 infants), p < 0.01 for each. P-APA was not significantly higher in infants with radiologic rickets (1078 +/- 356 IU/L) compared to those without radiologic evidence of rickets (943 +/- 346, p = 0.18). CONCLUSION: Elevation of P-APA >600 IU/L was very common in ELBW infants. BW was significantly inversely related to both P-APA and radiologic rickets. No single value of P-APA was related to radiological findings of rickets. Given the very high risk of osteopenia and rickets among ELBW infants, we recommend consideration of early screening and early mineral supplementation, especially among infants <600 g BW.

Dynamic changes in the cervical epithelial tight junction complex and differentiation occur during cervical ripening and parturition.

Cervical epithelia have numerous functions that include proliferation, differentiation, maintenance of fluid balance, protection from environmental hazards, and paracellular transport of solutes via tight junctions (TJs). Epithelial functions must be tightly regulated during pregnancy and parturition as the cervix undergoes extensive growth and remodeling. This study evaluated TJ proteins, as well as markers of epithelial cell differentiation in normal and cervical ripening defective mice to gain insights into how the permeability barrier is regulated during pregnancy and parturition. Although numerous TJ proteins are expressed in the nonpregnant cervix, claudins 1 and 2 are temporally regulated in pregnancy. Claudin 1 mRNA expression is increased, whereas claudin 2 expression declines. The cellular localization of claudin 1 shifts at the end of pregnancy (gestation d 18.75) to the plasma membrane in a lattice pattern, consistent with TJs in the apical cells. The timing of claudin 1-enriched TJs coincides with initiation of terminal differentiation of cervical squamous epithelia as evidenced by the increased expression of genes by differentiated epithelia late on gestation d 18. The cervical ripening defective steroid 5alpha-reductase type 1 deficient mouse, which has an elevated local progesterone concentration, also has aberrant claudin 1 and 2 expressions, fails to form claudin 1-enriched TJs, and lacks normal expression of genes involved in epithelial terminal differentiation. These data suggest that changes in permeability barrier properties during cervical ripening are, in part, negatively regulated by progesterone, and that dynamic changes in barrier properties of the cervix occur during pregnancy and parturition.