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Title:   Author  item
                                            Schultz, Carrieitem
                                            Edrington, Thomasitem
                                            Callaway, Todditem
                                            SCHROEDER, S - TX A&M UNIVERSITYitem
                                            HALLFORD, D - NM STATE UNIVERSITYitem
                                            Genovese, Kenneth

                                                         - *Ken*item
                                            Anderson, Robinitem
                                            Nisbet, David
  Submitted to: Letters in Applied Microbiology
                                    Publication Type: Peer Reviewed Journal
                                    Publication Acceptance Date: 12/20/2005
                                    Publication  Date: 6/5/2006
                                    Citation: Schultz, C.L., Edrington, T.S., Callaway, T.R., Schroeder, S.B., Hallford, D.M., Genovese, K.J., Anderson, R.C., Nisbet, D.J. 2006. The influence of melatonin on growth of E. coli O157:H7 in pure culture and exogenous melatonin on faecal shedding of E. coli O157:H7 in experimentally infected wethers. Letters in Applied Microbiology. 43:105-110.Interpretive Summary: Escherichia coli O157:H7 (E. coli O157:H7) is a pathogen responsible for a number of foodborne illnesses in the U.S. with an estimated 62,500 human cases reported per year.  Cattle and sheep are natural carriers of E. coli O157:H7.  Fecal shedding of E. coli O157:H7 by livestock is typically greater during the warmer months of the year, therefore resulting in seasonal differences in shedding.  The hormone, melatonin, is also affected by season with higher concentrations observed during the fall and winter.  To determine if this hormone plays a role in the amount of E. coli O157:H7 shed by livestock, sheep were orally administered melatonin.  Melatonin did not directly affect the growth of E. coli O157:H7 in a test tube or the fecal shedding of E. coli O157:H7 by sheep.  However, an interesting tendency for decreased E. coli O157:H7 shedding by sheep administered melatonin was observed towards the end of the experiment when their blood levels of melatonin were highest.  This decrease suggests melatonin may play a role in the seasonal shedding of E. coli O157:H7.
                                Technical Abstract: To determine if melatonin directly affects growth of E. coli O157:H7 in pure culture, an in vitro experiment was conducted.  Pure cultures of E. coli O157:H7 strains 933 and 6058 were individually added (0.5 mL each) to 9.0 mL tryptic soy broth (TSB).  Melatonin solutions were added to achieve final concentrations of 0.0, 0.43, 0.86, 1.72, 3.44, and 6.88 pmol mL**-1.  Culture tubes were sealed, vortexed, and incubated at 39 deg C.  Growth rates were estimated over time via measurement of optical density (O.D.).  To evaluate the effects of exogenous melatonin, wethers were orally administered gelatin capsules containing either ground alfalfa or 25 mg MEL plus ground alfalfa for CONT- and MEL-treated animals, respectively for 21 d. Seven d following initial MEL treatment, animals were experimentally infected with E. coli O157:H7 to monitor fecal shedding patterns. Sheep were euthanized on d 21 to evaluate gut populations of the experimental strain.  Blood was collected on d 7, 14, and 21 to determine total WBC and differential leukocyte counts.  Weekly blood samples were collected for analysis of MEL.  Growth rates (h**-1) of E. coli O157:H7 in pure culture containing increasing concentrations of added MEL were not affected (P > 0.1) by the addition of MEL, regardless of the concentration.  Daily faecal shedding patterns of the experimental strain were similar (P = 0.37) between the CON and treated group with shedding decreasing daily (P < 0.01) over the 14 d period.  By day 20, bacterial counts were 1.83 and 0.71 CFU g**-1 faeces (log10) for CON and MEL-treated animals, respectively.  Control sheep tended to, or shed more, E. coli O157:H7 on d 19 (P = 0.09) and d 20 (P = 0.05) than those receiving MEL.  However, bacterial counts from rectal samples taken on d 21 were similar (P = 0.68) between the CON and treated group.  No differences (P > 0.10) were observed in the populations of E. coli O157:H7 in ileal, caecal, or rectal contents.  Although neither treatment, nor treatment x day, effects were observed, total WBC counts decreased over the course of the study (P < 0.01) in all animals.  On d 7 (prior to inoculation with E. coli O157:H7), WBC counts were 1.5 x 10**4 and 1.0 x 10**4 for CON and MEL sheep, respectively, decreasing one log by d 21.  Differential leukocyte counts were also similar between the CON and treated group with no day or treatment x day effect observed.  Serum concentrations of MEL differed between groups (P < 0.01).  Initial serum MEL concentrations were 6.1 and 6.5 pg mL**-1 for CON and MEL sheep, respectively.  However, by the end of the experiment (d 21), average serum MEL concentrations were 12.1 pg mL**-1 for CON sheep compared to 71.3 pg mL**-1 for those receiving MEL.  Results indicate melatonin does not directly affect growth of E. coli O157:H7 in vitro, nor does exogenous melatonin affect fecal shedding patterns of E. coli O157:H7 in experimentally-infected wethers.  However, the tendency to observe lower fecal bacteria counts in MEL-treated sheep towards the end of the trial warrents further investigation.


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