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Endocrine disrupting chemicals (EDCs) are compounds that can interfere with hormone signaling pathways and are now recognized as pervasive in estuarine and marine waters. One prevalent EDC in California’s coastal waters is the xenoestrogen 4-nonylphenol (4-NP), which has been shown to impair reproduction, development, growth, and in some cases immune function of marine invertebrates. To further investigate effects of 4-NP on marine invertebrate immune function we measured total hemocyte counts (THC), relative transcript abundance of immune-relevant genes, and lysozyme activity in Pacific oysters (Crassostrea gigas) following bacterial infection. To quantify these effects we exposed oysters to dissolved phase 4-NP at high (100 μg l-1), low (2 μg l-1), or control (100 μl ethanol) concentrations for 7 days, and then experimentally infected (via injection into the adductor muscle) the oysters with the marine bacterium Vibrio campbellii. 4-NP significantly altered the effects of bacterial infection had on THC. Oysters exposed to both high and low 4-NP did not experience a bacteria-induced increase in THC, as seen in control oysters. We also determined that V. campbellii infection induced differential expression of a subset of immune-related genes tested (Cg-bigdef2, Cg-bpi1, Cg-lys1, Cg-timp) in some, but not all, tissues; 4-NP exposure altered expression patterns in two of these genes (Cg-bpi1 and Cg-tgase). Exposure to 4-NP alone also caused differential expression in some genes (Cg-bpi1, Cg-galectin1, Cg-clec2). Lastly, low levels of 4-NP significantly increased lysozyme activity 24 h post-infection. These results suggest that exposure to 4-NP can alter both cellular and humoral immune responses to bacterial infection in C. gigas.

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The iron chelating molecule, ethylenediaminetetraacetic acid (EDTA) is used in food applications for the preservation of oxidation prone ingredients. Research has suggested that EDTA is also capable of enhancing the antimicrobial effectiveness of various compounds including naturally-derived antimicrobials. With consumer demand for cleaner food labels, there remains an opportunity to introduce new chelating technology to replace synthetically-derived EDTA. Through photographting and chemical conversion, hydroxamic acid ligands were covalently bound to polypropylene films resulting in polypropylene-graft-poly(hydroxamic acid) (PP-g-PHA). The resulting films demonstrated an ability to chelate 64 nmol/cm2 from an iron saturated environment or 163 nmol/cm2 of magnesium and 139 nmol/cm2 of calcium from bacterial growth media. A surface pKa of 8.97 suggested that film ligands should remain protonated under acidic and neutral pH conditions. When combined with lysozyme, PP-g-PHA films were able to reduce inhibitory concentration of lysozyme for Listeria monocytogenes by half. When tested against Bacillus cereus, Pseudomonas fluorescens, and E. coli O157:H7; PP-g-PHA films were unable to inhibit growth and showed little enhancement of lysozyme. EDTA controls revealed that similar levels of soluble chelator were more effective than immobilized chelators. EDTA results also suggested that a chelating film with a higher affinity for iron (through coordination or ligand stability) may be able to control B. cereus growth. Both EDTA and PP-g-PHA caused P. fluorescens to produce siderophores (pyoerdines), suggesting that each treatment resulted in a low-iron growth environment. These findings suggest that surface bound chelating technology can affect the growth of L. monocytogenes and enhance the effectiveness of lysozyme. With improved surface chemistry (a higher binding constant with iron), this technology has the potential to influence the growth of other pathogens and spoilage microorganisms.