Progress 10/01/16 to 09/30/17

Outputs
Target Audience:Target audiences are the seafood industry, regulatory agencies, risk assessors, risk managers, and the scientific community. The efforts to reach the target audiences were presentations of the research findings at professional meetings, publications in peer-reviewed journals, class room discussion, workshops, personal communication with the seafood industry, state, and regulatory agencies, extension and outreach. Changes/Problems:We conducted several trial and error experiments to standardize the methods for Pulsed-field gel electrophoresis (PFGE) of Campylobacter jejuni recovered from seafood.In addition, the electrophoresis chamber of the PFGE apparatus was broken so we had to repair it. What opportunities for training and professional development has the project provided?One technician, one graduate student, four undergraduate students, and one high school student had the opportunity to learn state-of-the-art microbiology and molecular biology techniques. One M.S. student graduated with experience in seafood safety, microbiology and molecular biology. One doctoral student will defend his doctoral dissertation in April and will graduate officially in May 2018. One new collaboration was established with scientists (Dr. L. Wei, and F. Sai) from Beijing Center for Disease Control, China. One technician, students and investigators were able to attend the regional, national and international professional meetings to share the project findings with the scientific community and update the knowledge in molecular biology, food microbiology and safety. How have the results been disseminated to communities of interest?Four presentations were made at regional and national professional meetings. One manuscript was published and one manuscript is under review by co-authors. Findings of this project were also shared with the students of independent studies in animal and poultry sciences, advanced food microbiology, food microbiology laboratory, and seminar in food science and technology courses at the University of Maryland Eastern Shore, the seafood industry, and regulatory agencies through classroom discussion, personal communication, workshops, outreach and extension activities. What do you plan to do during the next reporting period to accomplish the goals?Molecular analysis (PFGE and virulence factors) of Campylobacter isolates and PFGE of Vibrio isolates will be completed by first week of March 2018. One doctoral student will defend his dissertation in April 2018 to graduate officially in spring 2018.

Impacts
What was accomplished under these goals? One isolate from each positive sample (n=127) was tested for the presence of Salmonella virulence genes invA, pagC, spvC, spvR by PCR. Seventy-three percent, 76 and 71 percent of isolates recovered from catfish, shrimp, and tilapia, respectively, were positive for InvA. PagC was detected in 59 percent, 71 percent and 74 percent of catfish, shrimp, and tilapia, respectively. Sixty-eight percent of catfish, 88 percent of shrimp, and 74 percent of tilapia contained SpvC. SpvR was found in 59 percent of catfish, 51 percent of shrimp, and 48 percent of tilapia. One Vibrio parahaemolyticus isolate from each positive sample (n=14) was tested for two virulence genes (thermostable direct hemolysin [tdh] and tdh-related hemolysin [thr]). All isolates were negative for tdh and trh. One V. vulnificus isolate from each positive sample (n=6) was also analyzed for a virulence gene (virulence correlated gene [vcgC]). Fifty percent of V. vulnificus isolates recovered from shrimp were positive for vcgC gene. There were no significant differences in the presence of virulence genes in Salmonella and Vibrio among samples. Pulsed-field gel electrophoresis (PFGE) patterns for Salmonella isolates recovered from seafood were completed using a molecular method. Currently, analysis of PFGE patterns of Salmonella isolates are underway using bionumeric software. Preliminary data analysis indicates that Salmonella isolates were genetically diverse and no association was observed between PFGE patterns of Salmonella and the seafood samples. The results suggest that Salmonella and V. vulnificus isolates recovered from seafood can possess virulence genes and thus have the potential to cause salmonellosis and V. vulnificus infections. Potential food safety hazards associated with seafood warrant further large-scale studies. Training of students and a technician in seafood safety, microbiology and molecular biology as well as establishment of collaborative research among UMES, FDA and University of Maryland, College Park (UMD) will enhance the national and international standing of UMES research programs, thereby enhancing student recruiting and the recognition of its microbiological research and outreach programs. So far, the project has motivated minority graduate and undergraduate students to pursue higher education in molecular biology, food microbiology, and seafood safety as well as careers in these disciplines in academia, industry, and federal and state agencies. The outreach activities significantly strengthen ongoing extension programs.

Publications

• Type: Journal Articles Status: Under Review Year Published: 2018 Citation: Elbashir, S., Parveen, S., Min, B. R. and Jahncke, M. 2018. Antimicrobial resistance of foodborne pathogens and antimicrobial residues in seafood: A Review. Journal of Food Science (under review by co-authors)
• Type: Journal Articles Status: Published Year Published: 2018 Citation: Elbashir, S., Parveen, S., Schwarz, J., Rippen, T., Jahncke, M., and DePaola, A. 2018. Seafood Pathogens and Information on Antimicrobial Resistance: A Review. Food Micro. 70:85-90.
• Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Parveen, S. 2017. Current perspectives on microbial poultry & seafood safety. Workshop for Cochran Fellows from Ghana. Summer 2017. University of Maryland Eastern Shore, Princess Anne, MD (invited oral).
• Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Parveen, S. 2017. Current perspectives on microbial seafood safety. Global issues related to food safety and health. ARD 18th Biennial Research Symposium. Apr. 1-4, 2017. Atlanta, GA (invited oral).
• Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Parveen, S. 2017. Is it safe to eat seafood? Sea food Safety Workshop. Jan. 06, 2017. Delaware State University, Dover, DE (invited oral).
• Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Elbashir, Salah, 2016. Microbiological Quality of Frozen Seafood. Joint Institute For Food Safety And Applied Nutrition (JIFSAN). Summer 2016. University of Maryland, College Park, MD 20740 (invited oral).

Progress 10/01/15 to 09/30/16

Outputs
Target Audience:Target audiences arethe seafood industry, regulatory agencies, risk assessors, risk managers, and the scientific community. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Two technicians, one graduate student, and two undergraduate students had opportunities to learn state-of-the-art microbiology and molecular biology techniques.The project director and students attended several national and international professional meetings to exchange ideas and update their knowledge in seafood safety. How have the results been disseminated to communities of interest?PI and one graduate student madeoral presentations at professional meetings. One article was submitted for publication to the peer reviewed journal "Food Microbiology". Findings of this project were also shared with students; the seafood industry; and regulatory agencies through classroom discussion, personal communication, and workshops. What do you plan to do during the next reporting period to accomplish the goals?To accomplish our goals and objectives, we are planning to conduct the following analyses/experiments during the next reporting period: To characterize three isolates from each positive sample by Real Time PCR assay and pulsed field gel electrophoresis (PFGE; objective 5). To analyze the data of antibiotic resistance patterns of pathogens and antibiotic residues in seafood to determine whether the antibiotic resistance patterns of pathogenic bacteria are correlated with antibiotic residues in imported and domestic shrimp, tilapia, and catfish (Objective 4). To develop outreach and extension programs for education and the control of foodborne pathogens and antibiotic residues in shrimp, catfish, and tilapia (objective 6).

Impacts
What was accomplished under these goals? Extraction of antimicrobials from seafood were successfully conducted and experiments were conducted to identify multidrug residues (MDR) in imported and domestic seafood (shrimp, tilapia and catfish). Fourteen tested antimicrobials were chloramphenicol, tetracycline, oxytetracycline, doxycycline, enrofloxacin, difloxacin, sarafloxacin, sulfamerazine, sulfamethazine, sulfadimethoxine, sulfapyridine, sulfathiazole, sulfamethoxazole, and malachite green. First, 468 seafood samples were grouped into 54 groups' composites based on: the type of seafood, month of sample collection, and country of origin. The results revealed detectable amounts of three types of sulfonamides (sulfamethazine, sulfathiazole, and Sulfamethoxazole) in 10 of the seafood groups. The 10 seafood groups were 7 shrimp groups, and 3 tilapia groups; and no antimicrobial residue was detected in the catfish groups. Sulfamethazine was detected in two groups of shrimp and one of tilapia. Sulfathiazole was detected in 4 shrimp and 2 tilapia groups. Sulfamethoxazole was detected in two shrimp groups only. The second test was a validation test and in which the ten above groups were tested at the level of individual samples that formed them. No detectable amounts of any of the antimicrobials were detected on individual samples. The results emphasized that there is no correlation between antimicrobial resistance of the pathogens (Vibrio, Salmonella, and Campylobacter) isolated from the same seafood samples and the antimicrobial residues. Pulsed-field gel electrophoresis patterns and virulence factors for Vibrio, Salmonella, and Campylobacter species recovered from frozen seafood (catfish, shrimp, and tilapia) were partially completed using molecular methods. So far, the project has motivated minority graduate and undergraduate students to pursue higher education in molecular biology, food microbiology, and seafood safety as well as careers in these disciplines in academia, industry, and federal and State agencies. In addition, analysis of domestic and imported seafood samples for antibiotic residues will provide valuable information about the prevalence of antibiotic resistance phenotypes and antibiotic residues in samples for further risk assessment upon which seafood safety policy decisions can be made. Moreover, the analysis of the antibiotic resistance phenotypes of Salmonella, Vibrio and Campylobacter isolates recovered from domestic and imported seafood will provide useful information for the seafood industry and regulatory agency, as the multiple drug resistant foodborne pathogens isolated from seafood and aquatic environments represent a major concern in aquaculture and human health. Objective 1, 2, and 3 have been completed. Objectives 4, 5 and 6 have been partially completed.

Publications

• Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Elbashir, Salah, 2016. Microbiological Quality of Frozen Seafood. Joint Institute For Food Safety And Applied Nutrition (JIFSAN). University of Maryland, College Park, MD 20740
• Type: Conference Papers and Presentations Status: Published Year Published: 2016 Citation: Parveen, S. 2016. Environmental Microbiology, Food Microbiology and Food Safety Research at UMES (How to write a successful grant workshop). Jan. 05, 2016. North South University, Dhaka, Bangladesh.
• Type: Journal Articles Status: Submitted Year Published: 2016 Citation: Elbashir, S., Parveen, S, Schwarz, Rippen, T, Jahncke, M, and DePaola. A. 2016. Seafood Pathogens and Information on Antimicrobial Resistance: A Review. Food microbiology (submitted).
• Type: Journal Articles Status: Under Review Year Published: 2016 Citation: Elbashir, S., Parveen, S., Byungrok, M., Schwarz, J., Rippen, T., and Jahncke, M. Antimicrobial Resistance of Foodborne Pathogens and Antimicrobial Residues in Frozen Seafood: A Review. Journal of Food Science. (Under view by co-authors)

Progress 10/01/14 to 09/30/15

Outputs
Target Audience:Target audiences are students, the seafood industry, regulatory agencies, risk assessors, risk managers, and the scientific community. The efforts to reach the target audiences were presentations of the research findings at professional meetings, classroom discussion, and personal communication with the scientific community, students, seafood industry, and state and regulatory agencies. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Two technician, two graduate students, and three undergraduate students had opportunities to learn state-of-the-art microbiology and molecular biology techniques. Existing collaboration was extended with collaborators Jurgen Schwarz, John Bowers, Angelo DePaola, Michael Jahncke, Byungrok Min, and Tom Rippen. The project director and students attended several national and international professional meetings to exchange ideas and update her knowledge in seafood safety. How have the results been disseminated to communities of interest?Six oral presentations were made by students and investigators at regional, national and international professional meetings. One undergraduate student made a poster presentation at Research Experiences for Undergraduates (REU) meeting. Four abstracts were published in proceedings of regional, national and international professional meetings. Moreover, a review article is under view by co-authors. In addition, findings of this project were also shared with students of the independent studies in animal and poultry sciences, Advanced Food Microbiology, and seminar in food science and technology courses at the University of Maryland Eastern Shore; the seafood industry; and regulatory agencies through classroom discussion, personal communication, and workshops. What do you plan to do during the next reporting period to accomplish the goals?To accomplish our goals and objectives, we are planning to conduct the following analysis/experiments during the next reporting period: 1) to characterize three isolates from each positive sample by Real Time PCR assay and pulsed field gel electrophoresis (PFGE; objective 5); 2) to determine the concentrations of antibiotic residues (chloramphenicol, nitrofuran, and tetracycline) in imported and domestic shrimp, tilapia and catfish; objective 3) to determine whether the antibiotic resistance patterns of pathogenic bacteria are correlated with antibiotic residues in imported and domestic shrimp, tilapia, and catfish; objective 4) to develop outreach and extension programs for education and the control of foodborne pathogens and antibiotic residues in shrimp, catfish, and tilapia (objective 6).

Impacts
What was accomplished under these goals? Two graduate students, three undergraduate students and two technicians were trained in microbiology and molecular biology. One graduate student was trained in analysis of antibiotic residues in seafood. A total of 127 Salmonella isolates recovered from imported and domestic seafood samples (shrimp, tilapia and catfish) were analyzed by serotyping. The results revealed that all isolated were Salmonella Typhimurium var-5. No associated was observed among serovar, seafood type and the country of origin. A total of 127 Salmonella, 26 Vibrio cholerae, 14 Vibrio parahaemolyticus, 6 Vibrio vulnificus and 16 Campylobacter jejuni isolated from imported and domestic seafood samples were tested for antibiotic resistance phenotypes to 17 different antibiotics, chosen by clinical usage for these pathogens and aquaculture practices, using the Sensititre® micro-broth dilution method according to the guidelines of the Clinical and Laboratory Standards Institute (CLSI). All tested Salmonella isolates were uniformly susceptible to five (amikacin, chloramphenicol, florfenicol, doxycycline, and imipenem) of the 17 tested antibiotics. Thirty five percent of the Salmonella isolate were resistant to at least one antimicrobial among them 27 percent were multidrug resistant. All tested Vibrio species were susceptible to five antimicrobials (amikacin, amoxicillin /clavulanic acid, chloramphenicol, doxycycline and florfenicol). Two of them (ciprofloxacin, and doxycycline) were recommended by CDC for treating Vibrio infections. Fifty seven percent of the isolates were resistant to at least one antimicrobial and 30 percent were resistant to multiple antimicrobials. Twenty percent of the Salmonella isolates and 23 percent of the Vibrio isolates were found to be completely susceptible to all antimicrobials tested. All Campylobacter isolates were uniformly susceptible to clindamycin and florfenicol. Sixty two percent were resistant to at least one antimicrobial. Forty four percent of the isolates were resistant to more than one antimicrobials and 13 percent were susceptible to all nine antimicrobials. Statistical analysis is underway to determine the association between antibiotic resistance phenotypes and type of seafood. Several experiments were conducted to standardize the methods for extraction of antibiotic residues from seafood. So far, the project has motivated minority graduate and undergraduate students to pursue higher education in molecular biology, food microbiology, and seafood safety as well as careers in these disciplines in academia, industry, and federal and state agencies. In addition, analysis of domestic and imported seafood samples for antibiotic residues will provide valuable information about the prevalence of antibiotic resistance phenotypes and antibiotic residues in samples for further risk assessment upon which seafood safety policy decisions can be made. Moreover, the analysis of the antibiotic resistance phenotypes of Salmonella, Vibrio and Campylobacter isolates recovered from domestic and imported seafood will provide useful information for the seafood industry and regulatory agency, as the multiple drug resistant foodborne pathogens isolated from seafood and aquatic environments represent a major concern in aquaculture and human health. Objective 1 has been completed, objectives 2, 3, 4 and 6 have been partially completed.

Publications

• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Allen, L., Elbashir, S., and Parveen, S. 2015. Antibiotic susceptibility of Salmonella isolates from seafood. Research Experiences for Undergraduate (REU) Summer Program. August 2015. Paul S. Sarbanes Coastal Ecology Center, Ocean city. Maryland.
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Elbashir, S., Parveen, S., Schwarz, J., Rippen, T., Jahncke, M. and John Bowers. 2015. Detection and Identification of Salmonella and Campylobacter jejuni in Seafood. UMES Symposium. April 20-23, 2015. Princess Anne, MD.
• Type: Journal Articles Status: Under Review Year Published: 2015 Citation: Elbashir, S., Parveen, S., Schwarz, J., Rippen, T., Jahncke, M. and DePaola, A. 2015. Major pathogens and information on antimicrobial resistance: A review. Food Microbiology (under review by co-authors).
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Elbashir, S., Parveen, S., Bowers, J., Rippen, T., Schwarz, J.,Jahncke. J., and DePaola, A. 2015. Microbiological quality of imported and domestic seafood. International Association for Food Protection (IAFP) Annual meeting. July 24-28, 2015. Portland. OR.
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Parveen, S. and Huq, A. 2015. Emerging Techniques for Detecting and Characterizing Food- and Water-Borne Pathogens for Food Safety and Security. The third International conference on global warming: Food Security, Ras Al Khaimah, UAE. May 5-7, 2015. Ras Al Khaimah, UAE (Invited presentation).
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Elbashir, S and Parveen, S., 2015. Evaluation of foodborne pathogens and antibiotic residues in seafood. Interstate Shellfish Sanitation Conference (ISSC), Ocean City. MD. April 14-16, 2015 (Invited presentation).
• Type: Conference Papers and Presentations Status: Published Year Published: 2015 Citation: Brown, J., Elbashir, S., Parveen, S. 2015. Prevalence of Vibrio parahaemolyticus and Vibrio vulnificus in shrimp. University of Maryland Eastern Shore (UMES) Symposium. April 20-23, 2015. Princess Anne, MD.

Progress 10/01/13 to 09/30/14

Outputs
Target Audience: Target audiences are students, the seafood industry, regulatory agencies, risk assessors, risk managers, and the scientific community. The efforts to reach the target audiences were presentations of the research findings at professional meetings, classroom discussion, and personal communication with the students, scientific community, seafood industry, and state and regulatory agencies. Changes/Problems: Nothing significant to report this reporting period. What opportunities for training and professional development has the project provided? One technician, one graduate student, and two undergraduate students had opportunities to learn state-of-the-art microbiology and molecular biology techniques. Existing collaboration was extended with collaborators Jurgen Schwarz, John Bowers, Angelo DePaola, Michael Jahncke, Byungrok Min and Tom Rippen. The project director attended the American Society for Microbiology General Meeting and the International Association for Food Protection meeting to update her knowledge in seafood safety. How have the results been disseminated to communities of interest? One oral presentation was made at the Louis Strokes Alliances for Minority Participation (LSMAP) Program meeting in 2014 at UMES, three abstracts were submitted for oral presentation at regional and national professional meetings. Moreover, one article is underreview by co-authors. Findings of this project were also shared with students of the independent studies in animal and poultry sciences, and seminar in food science and technology courses at the University of Maryland Eastern Shore; the seafood industry; and regulatory agencies through classroom discussion, personal communication, and workshops. What do you plan to do during the next reporting period to accomplish the goals? To accomplish our goals and objectives, we are planning to conduct the following analysis/experiments during the next reporting period: 1) to characterize three isolates from each positive sample by Real Time PCR assay, serotyping, antibiotic susceptibility testing and pulsed field gel electrophoresis (PFGE); 2) to determine the concentrations of antibiotic residues (chloramphenicol, nitrofuran, and tetracycline) in imported and domestic shrimp, tilapia and catfish; 3) to determine whether the antibiotic resistance patterns of pathogenic bacteria are correlated with antibiotic residues in imported and domestic shrimp, tilapia, and catfish; 4) to develop outreach and extension programs for education and the control of foodborne pathogens and antibiotic residues in shrimp, catfish, and tilapia.

Impacts
What was accomplished under these goals? One graduate student and three undergraduate students were trained in microbiology and molecular biology. One graduate student was trained in analysis of antibiotic residues in seafood. Investigators, a technician, and graduate and undergraduate students met a couple of times to discuss the project plan and to design experiments for the molecular analysis and antibiotic susceptibility testing of confirmed bacterial isolates. A total of 468 frozen catfish, shrimp, and tilapia imported (60, 85, 84) and domestic (96, 71, 72) samples were collected from three retail stores on the Eastern Shore of Maryland and analyzed for aerobic plate count (APC), total coliforms, Escherichia coli and three major foodborne-pathogens (Campylobacter jejuni, Salmonella, and Vibrio) using standard methods. Two hundred and sixty four (54 Campylobacter, 127 Salmonella, and 83 Vibrio) presumptive positive isolates maintained at -80 degreeC in brain heart infusion broth (BBL, Difco, BD, Sparks, MD) that contained 15 percent (vol/vol) glycerol (Sigma, St. Louis, MO) were confirmed by BAX Real-time PCR assays for detecting Campylobacter jejuni, Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus and Standard PCR assays for detecting Salmonella. Measurement outcomes were evaluated by one-way ANOVA or t-test when quantitative and by Fisher's exact test when qualitative. All samples were positive for APC and the average log CFU per gram for APC ranged from 3.9 to 4.1 in the three types of seafood. The highest count of APC was observed in shrimp (log 8.4 CFU per gram). Total coliforms were found in 41 percent of the samples and 8.8 percent were positive for E. coli. The counts of total coliforms and E. coli ranged from 3.18 log CFU per gram to 4.10 log CFU per gram and from 2.18 log CFU per gram to 3.88 log CFU per gram in domestic and imported seafood. The highest counts of total coliforms and E. coli were observed in catfish (3.8 log CFU per gram) and tilapia (3.7 log CFU per gram), respectively. Approximately 3 percent, 27percent and 8 percent of the samples were positive for Campylobacter jejuni, Salmonella, and Vibrio, respectively. The counts of Campylobacter jejuni, Salmonella, and Vibrio ranged from 1.0-2.1 percent, 19.4-33.3 percent, and 5-3 percent in domestic and imported seafood, respectively. With respect to comparisons between imported versus domestic, statistically significant differences in rate of detection of total coliforms were observed in shrimp but there were no statistically significant differences between average log CFU per gram for APC or the rate of detection of E. coli. With respect to the major pathogens, there was a relatively large difference between Salmonella prevalence in imported versus domestic tilapia (33.3 percent of imported and 19.4 percent of domestic). These results indicate potential food safety hazards associated with domestic and imported seafood. So far, the project has motivated minority graduate and undergraduate students to pursue higher education in molecular biology, food microbiology, and seafood safety as well as careers in these disciplines in academia, industry, and federal and state agencies. In addition, analysis of domestic and imported seafood samples for antibiotic residues will provide valuable information about the prevalence of antibiotic residues in samples for further risk assessment upon which seafood safety policy decisions can be made. Objective 1 has been completed and objective 2 has been partially completed.

Publications

• Type: Conference Papers and Presentations Status: Accepted Year Published: 2014 Citation: Brown, J., Elbashir, S., Parveen, S. 2014. Prevalence of Vibrio parahaemolyticus in shrimp. Louis Strokes Alliances for Minority Participation (LSAMP) Program meeting, 2014. UMES. Princess Anne, MD.
• Type: Conference Papers and Presentations Status: Submitted Year Published: 2015 Citation: Brown, J., Elbashir, S., Parveen, S. 2015. Prevalence of Vibrio parahaemolyticus and Vibrio vulnificus in shrimp. University of Maryland Eastern Shore (UMES) Symposium. April 20-23, 2015. Princess Anne, MD (submitted).
• Type: Conference Papers and Presentations Status: Submitted Year Published: 2015 Citation: Elbashir, S., Parveen, S., Bowers, J., Rippen, T., Schwarz, J., Jahncke, J., DePaola, A. 2015. Microbiological quality of imported and domestic seafood. International Association for Food Protection (IAFP) Annual meeting. July 24-28, 2015. Portland. OR (submitted)
• Type: Conference Papers and Presentations Status: Submitted Year Published: 2015 Citation: Elbashir, S., Parveen, S., Schwarz, J., Rippen, T., Jahncke, M. and John Bowers. 2015. Detection and Identification of Salmonella and Campylobacter jejuni in Seafood. UMES Symposium. April 20-23, 2015. Princess Anne, MD (submitted).
• Type: Conference Papers and Presentations Status: Under Review Year Published: 2015 Citation: Elbashir, S., Parveen, S., Schwarz, J., Rippen, T., Jahncke, M. and DePaola, A. 2015. Major pathogens and their antimicrobial susceptibility in seafood: A review. Food Microbiology (under review by co-authors).

Progress 01/01/13 to 09/30/13

Outputs
Target Audience: Target audiences are the seafood industry, regulatory agencies, risk assessors, risk managers, and the scientific community. The efforts to reach the target audiences were presentations of the research findings at professional meetings, classroom discussion, and personal communication with the scientific community, seafood industry, and state and regulatory agencies. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? One technician, three graduate students, and two undergraduate students had opportunities to learn state-of-the-art microbiology and molecular biology techniques. A new collaboration was established with a scientist (Byungrok Min) of the University of Maryland Eastern Shore (UMES) and an existing collaboration was extended with collaborators Jurgen Schwarz, John Bowers, Angelo DePaola, Michael Janhcke and Tom Rippen. One graduate student was able to attend the workshop on Methods for Determination of Drug Residues in Aquaculture, Meat and Poultry presented at the Joint Institute for Food Safety and Applied Nutrition (JIFSAN) in College Park, Maryland, to learn the techniques for the detection of antibiotic residues in imported and domestic seafood. The project director attended the International Association for Food Protection meeting to update her knowledge in seafood safety. A graduate student also visited Dr. Debu Das’ laboratory at the University of Maryland, College Park to learn the techniques for the detection of Campylobacter in seafood. How have the results been disseminated to communities of interest? One poster and four oral presentations were made at the REU research symposium, the NIFA project director’s meeting, the Geoscience Bridge Program, and the food microbiology laboratory meeting at UMES. Findings of this project were also shared with students of the advanced food toxicology, independent studies in animal and poultry sciences, and seminar in food science and technology courses at the University of Maryland Eastern Shore; the seafood industry; and regulatory agencies through classroom discussion, personal communication, and workshops. What do you plan to do during the next reporting period to accomplish the goals? To accomplish our goals and objectives, we are planning to conduct the following analysis/experiments during the next reporting period: 1) to collect and analyze 216 seafood samples for microbiological examination; 2) to conduct confirmatory tests for all presumptive isolates using BAX Real-time and Salmonella assays; 3) to characterize three isolates from each positive sample by antibiotic susceptibility testing and pulsed field gel electrophoresis (PFGE); 4) to determine the concentrations of antibiotic residues (chloramphenicol, nitrofuran, and tetracycline) in imported and domestic shrimp, tilapia and catfish; 5) to determine whether the antibiotic resistance patterns of pathogenic bacteria are correlated with antibiotic residues in imported and domestic shrimp, tilapia, and catfish; 6) to develop outreach and extension programs for education and the control of foodborne pathogens and antibiotic residues in shrimp, catfish, and tilapia.

Impacts
What was accomplished under these goals? One graduate student was hired and trained in January 2013. Investigators, a technician, and graduate and undergraduate students met in February 2013 and organized several conference calls in April 2013 and June 2013 to discuss the project plan and to design experiments for this study. Investigators and students also visited several retail stores on the Eastern Shore to find and purchase domestic and imported seafood samples. Several experiments were conducted to standardize the methods for the detection of Campylobacter and Salmonella in seafood. A total of 216 seafood samples [72 domestic catfish, 72 shrimp (41 domestic and 31 imported), and 72 imported tilapia] were collected from three retail stores on the Eastern Shore of Maryland and analyzed for the detection of total aerobic plate counts (APC), total coliforms, Escherichia coli, Campylobacter, Salmonella, and Vibrio species during the reporting period. All samples were positive for APC. The counts of APC ranged from 3.16 log CFU/g to 6.28 log CFU/g in imported and domestic tilapia, shrimp, and catfish. The highest count of APC was observed in catfish (log 6.28 CFU/g). Thirty nine and 13percent of samples were positive for total coliforms and E. coli, respectively. The counts of total coliforms and E. coli ranged from 3.18 log CFU/g to 4.10 log CFU/g and from 2.18 log CFU/g to 3.88 log CFU/g in domestic and imported seafood. The highest counts of total coliforms and E. coli were observed in catfish (3.8 log CFU/g) and tilapia (3.7 log CFU/g), respectively. Eight, 35, 8, and 10 percent of domestic and imported samples were positive for presumptive Campylobacter, Salmonella, Vibrio parahaemolyticus, and V. vulnificus, respectively. These results indicate potential food safety hazards associated with domestic and imported seafood. Presumptive positive isolates were maintained at -80 degree C in brain heart infusion broth (BBL, Difco, BD, Sparks, MD) that contained 15 percent (vol/vol) glycerol (Sigma, St. Louis, MO) for confirmation by BAX Real-time PCR assays for detecting Campylobacter jejuni, Vibrio parahaemolyticus, and Vibrio vulnificus and Standard PCR assays for detecting Salmonella. So far, the project has motivated minority graduate and undergraduate students to pursue higher education in molecular biology, food microbiology, and seafood safety as well as careers in these disciplines in academia, industry, and federal and state agencies. In addition, analysis of domestic and imported seafood samples for antibiotic residues will provide valuable information about the prevalence of antibiotic residues in samples for further risk assessment upon which seafood safety policy decisions can be made. Objectives 1 and 6 have been partially completed.

Publications

• Type: Other Status: Published Year Published: 2013 Citation: Parveen. S. 2013. Seafood safety research at the University of Maryland Eastern Shore (UMES). USDA NIFA directors meeting. Jan. 30, 2013. UMES. Princess Anne, MD (oral). Parveen, S. 2013. Pathogenic bacteria and viruses in seafood. Geoscience Bridge Program. July 16, 2013. UMES. Princess Anne, MD (oral). Jones, Remy 2013. Prevalence and antimicrobial susceptibility of Salmonella isolated from fresh gulf shrimp. REU Research Symposium. August 09, 2013. UMES. Princess Anne, MD (oral and poster). Elbashir, Salah 2014. Methods for determination of drug residues in aquaculture, meat, and poultry. Food Microbiology Laboratory meeting. February 12, 2014 (oral).