Progress 03/01/18 to 02/28/23
Outputs
Target Audience:Target audiences are the oyster harvesting and processing industries inMaryland and Florida, regulatory agencies, risk assessors, risk managers, students and the scientific community. The efforts to reach the target audiences include presentations of the research findings at professional meetings, publications in the peer reviewed journals, class room discussion, workshops, personal communication with the seafood industry, state, and regulatory agencies, extension and outreach. Changes/Problems:The delay in receiving the Iron Agar (only commercial supplier of Iron Agar is in Spain; it took several months to receive the product from the supplier due to back order and quality control issues).The technician was on sick leave for several weeks due to an injury and health issues. Due to COVID 19 pandemic restrictions, UMES were closed from March 2020-July 2020 and FAMU were closed from March 2020-Ocober 2020. No samples were collected during these periods. Moreover, we were not able to conduct any experiments during these times. What opportunities for training and professional development has the project provided?Three postdoctoral associate, three technicians, three graduate students, and 10 undergraduate students and two high school students had the opportunities to learn about the on-site water quality assessment, aquaculture, state-of-the-art microbiology and molecular biology techniques. One new collaboration was established with US Food and Drug Administration (USFDA) to investigate the Whole Genome Sequencing of Shewanella recovered from oysters and water samples. PI, Co-PI, postdoctoral associate and students were able to attend (virtual and in person) the regional, national and international professional meetings to share the project findings with the scientific community and update the knowledge in molecular biology, water quality, food microbiology and safety. The postdoctoral associate and one graduated student received permanent position in the academia and industry. How have the results been disseminated to communities of interest?Twelve abstracts were published and 15 presentations were made by PI, Co-PI, technicians, graduate and undergraduate students at regional, national and international professional meetings. Three manuscripts are under review by co-authors and two doctoral dissertations are under preparation. One graduate student received three awards for outstanding poster/oral presentations and 3MT presentation. 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?Three manuscript will be finalized for submission to peer-reviewed journals. Whole Genome Sequencing data will be analyzed for the preparation of a manuscript through the newcollaboration.
Impacts
What was accomplished under these goals?
Oyster and water samples were collected monthly from three site [Honga River (HR), Tangier Sound (TA), and Horn Point (HP)] in the Chesapeake Bay (CB) and one site in the Maryland Coastal Bay (MCB) from June 2019- October 2021. Additionally, physicochemical parameters such as water temperature, salinity, turbidity, dissolved oxygen, chlorophyll-a, and pH were recorded during the collection of samples. All samples were analyzed for Shewanella spp. using standardized methods. In brief, oyster homogenates and seawater samples were serially diluted and spread plated onto iron agar. After incubation plates were counted and at least 10% suspected Shewanella colonies (black) were picked and stored in glycerol at -80C. All isolates were confirmed using 16s rRNA sequencing. Identification was made by comparing the sequence to the GenBank database using the BLASTn program. The presumptive colonies were then multiplied by the percentage of isolates that were confirmed by 16S rRNA sequencing to be Shewanella and the data were converted to CFU/g or CFU/ml of sample. Pearson's correlation analysis was performed to assess correlation between CFU levels and physicochemical parameters. ANOVA tests were used to assess differences between sampling sites, sampling months, and sampling years. The Levene's test was used to assess variances among oyster and water samples. A new collaboration was established with US Food and Drug Administration (USFDA) to perform Whole Genome Sequencing. Seventy four percent of oyster (n=90) and 38% of water (n=93) samples were positive for total Shewanella, respectively.The highest counts of these bacterium were 1.0 x 106 CFU/g in oyster and 3.0 × 102 CFU/mL in water. In water, HP had higher Shewanella occurrences from June, July, August, and September with counts ranging from undetectable (0) to 2.7 × 102 CFU/mL undetectable to 7.0 × 102 CFU/mL at HR; undetectable to 3.1 × 101 CFU/mL at TA; and undetectable to 1.0 × 101 CFU/mL at MCB. In oyster, the highest detection for oysters were observed from June through October, oyster counts ranged from undetectable to 1.0 × 106 CFU/g at HP; undetectable to 5.7 × 104 CFU/g at HR; undetectable to 3.4 × 104 CFU/g at TA; and undetectable to 1.5 × 104 CFU/g at MCB. For both sample groups (water and oysters); sampling site, sampling month and sampling year showed significant differences between the abundance of Shewanella species found at the different sampling sites (p-value <0.05) and during the different sampling years (p-value = 0.03). The water temperature ranged from 17.8-20Cwith HP averaging the lowest temperature and MCB averaging the highest water temperature. Salinity ranged from 12-31 ppt. with MCB averaging the highest salinity levels, and HP averaging the lowest. Dissolved oxygen ranged from 7-9 mg/L with TAaveraging the highest levels and MCB. Turbidity ranged from 6-11 NTU; with HP and HR averaging the highest levels and TA averaging the lowest. The pH was for all sampling sites. Chlorophyll-a ranged from 6-8 µg/L with HP averaging the highest measurements and MCB with the lowest. Precipitation ranged at about 2.54-7.62 mm with HP receiving the least amount of average rainfall within 48 hours prior to sampling, with TA averaging the most. Total dissolved solids ranged from 1319-2976 mg/L with MCB averaging the highest and HP averaging the lowest. Atmospheric pressure consistently measured765 at all sampling sites. Shewanella were frequently isolated at higher concentrations in oysters than in seawater (p < 0.05). A weak correlation (defined as r = 0.40 to 0.49) was observed between seawater temperature and Shewanella counts (r = 0.45), while a negative correlation was observed between seawater dissolved oxygen levels and Shewanella abundance (r = -0.48). There was little to no correlation between the physicochemical parameters and Shewanella abundances in oysters. 16S rRNA sequencing was performed on 1,526 representative isolates of which 952 (777 oyster, 175 water) were confirmed as Shewanella within 18 different species. The top five species isolated from oyster samples were 345 S. khirikhana, 148 S. marisflavi, 98 S. loihica, 69 S. amazonensis, 48 S. algae and water samples the top five species were 86 S. khirikhana, 30 S. marisflavi, 23 S. algae, 12 S. loihica, 8 S. xiamenensis. Forty seven percent of identified Shewanella had beta hemolytic activity. Out of the 18 species identified, three species (S. khirikhana, S. marisflavi and S. loihica) observed more beta hemolytic activity than alpha for both oyster and water samples. The top 4 Shewanella species [S. algae (n=57), S. amazonensis (n=60), S. loihica (n=20), S. marisflavi (n=21)] were used for antibiotic susceptibility testing and three isolates from each samplewere selected. A total of 158 isolates were tested against 21 commonly used antibiotics. Since no existing resistance breakpoint data is available for Shewanella, the resistance breakpoints for Vibrio, enterobacteriaceae and FDA recommended standards were used for interpretation. Isolates that shared sequence relatedness to S. loihica and S. marisflavi were 85% susceptible to total antibiotics tested. Imipenem was the least effective antibiotic for preventive growth for isolates related to S. loihica (55%). Polymyxin B (84%) and Aztreonam (77%) were the least effective antibiotics to use for preventing S. algae related isolates. Almost 71% of the total isolates were resistant to multiple drugs. Overall, Polymyxin B had the highest resistance with 46% of the total isolates displaying growth at 4 µ/mL. Levofloxacin had the highest susceptibility with 0% of total isolates displaying growth at 8µ/mL. Cefepime, Cetazidime, Doxycycline, Minocycline, trimethoprim-sulfamethoxazole and Tigecycline with less than 2% of the isolates displaying resistance to the antibiotics used. In Florida, oyster and water samples were collected monthly from December 2020 until July 14, 2022, analyzed and characterized as described above. Similar trend was observed in the prevalence and counts of this bacterium in oyster and water samples in the Apalachicola Bay. A total of 13 species were identified using 16S rRNA sequencing. The four species were:S.algae - 52, S. loihica -41, S. khirikhana - 25, S. carassii - 21, S. xiamenensis - 8. The water temperature, salinity, pH, ranged from 10.3°C to 29.6C,15 to 28ppt; pH range 7.35 to 8.67. All of the isolates tested showed varied degrees of resistance to the antimicrobial agents used in the study. Interestingly, all the tested isolates were found resistant to the polymyxins. According to the updated classification of Multi Drug Resistant (MDR), our test results identified S. Khirikhana, S. oneidensis, S. carassii, S. seohaensis, and S. coralli as strains that are resistant to multiple drugs and considered as MDR. Moreover, we found that the species S. Khirikhana had the highest level of resistance, followed by the species S. oneidensis, S. carassii, S. seohaensis, S. corallii, S. xiamenensis. S. algae, S. aquimarina, S. Ioihica, and S. dokdonesis. Seventeen percent of isolates recovered from water samples were beta-hemolytic and 18.4 % recovered from oyster samples were tested as beta hemolytic. The results of this study indicate that S.amazonensis and S.algae have high numbers of multidrug resistance. This is concerning because S. algae is known to be pathogenic towards humans and pose a health threat through ingestion of contaminated seafood, or by exposing cuts in the skin to contaminated marine environments through recreational activities. While not much information is available on S. amazonensis, S. loihica or S. marisflavi, this experiment is aiding in bridging that data gap and better the understanding of Shewanella spp.
Publications
- Type:
Conference Papers and Presentations
Status:
Submitted
Year Published:
2018
Citation:
Hashem. F., S. Parveen, E. May. 2018. Water quality, fresh Produce and seafood safety research at UMES. Nov. 2018. University of Florida, Gainesville. FL.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2019
Citation:
Gleason, M. 2019. The prevalence of Vibrio and Shewanella spp. in oyster and seawater in Apalachicola Bay. Undergraduate thesis. Florida A and M University.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2019
Citation:
Hernandez, R. Meredith, J., and Parveen, S. 2019. Prevalence of Shewanella in oysters and seawater in the Chesapeake Bay and the Maryland Coastal Bays. REU presentation. August 2019. UMES, Paul S. Sarbanes Coastal Ecology Center, MD.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2020
Citation:
Hernandez, R. and Parveen, S. 2020. Prevalence of Shewanella in oysters and seawater in Maryland. 2020. Emerging Researchers National (ERN) Conference in STEM. Washington, D.C. February 6-8, 2020.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2020
Citation:
Parveen, S., 2020. Sea Food Safety and COVID-19. LSAMP Annual meeting, July 19, 2020, UMES. Virtual.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2021
Citation:
Parveen, S., 2021. Bangladesh Association for Food Protection (BAFP) in North America. The First Webinar on Bridging knowledge Gaps in Food and Water Safety. BAFP. February 20, 2021. Virtual.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2021
Citation:
Johnson, T., Parveen, S., Richards, G., Meredith, J., Almuhaideb, E., Rosales, D. 2021. Shewanella spp. as potential emerging pathogens in oyster and seawater from the Chesapeake Bay. USDA Congress Meeting, December 15, 2021. The Wye Research Center.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2021
Citation:
Parveen, S. 2021. A journey to food safety research and education. Mujib Centenary Seminar Series II, May 30, 2021. Dept. of Microbiology, University of Dhaka and DUMAA. Virtual.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2022
Citation:
Johnson, T., Parveen, S., Richards, G., Meredith, J., Almuhaideb, E., Rosales D., Chigbu, P., Dasilva, L. 2022. Incidence and pathogenic potential of Johnson, T., Almuhaideb, E., Meredith, J., Rosales, D., Chigbu, P., Dasilva, L., Parveen, S., Richards, G. 2022. Prevalence of Shewanella spp. in oyster and seawater. NOAA EPP/MSI 2022 Annual Forum. April 6-8, 2022. Tallahassee, FL. [Awarded 2nd place for Oral Presentation].
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2022
Citation:
Johnson, T., Almuhaideb, E., Meredith, J., Rosales, D., Chigbu, P., Dasilva, L., Parveen, S., Richards, G. 2022. Prevalence of Shewanella species in Chesapeake Bay oyster and seawater. ARD annual meeting. April 2-5, 2022. Atlanta, GA.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2022
Citation:
Johnson, T., Parveen, S., Richards, G., Meredith, J., Almuhaideb, E., Rosales D., Chigbu, P., Dasilva, L. 2022. Incidence and pathogenic potential of Shewanella species in oysters and seawater collected from the Chesapeake and Maryland Coastal Bays. International Association for Food Protection (IAFP) annual meeting, July 31-August 3, 2022. Pittsburg, PA.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2022
Citation:
Parveen, S., 2022. Food Safety and water quality: A journey into research and education at UMES. 1890 Research Directors Research (ARD) Research Symposium. April 02-05, 2022.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2023
Citation:
Johnson, T., Richards, G., Jacobs, J., Townsend, H., Almuhaideb, E., Meredith, J., Rosales, D., Chigbu, P., Dasilva, L., and Parveen, S. 2023. Prevalence of Shewanella species found in Oysters & Seawater from Mid-Atlantic Region. Living Marine Resources Cooperative Science Center (LMRCSC) Annual Meeting 2023. Virtual meeting. March 31, 2023.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2023
Citation:
Johnson, T., Richards, G., Jacobs, J., Townsend, H., Almuhaideb, E., Johnson, T., Richards, G., Jacobs, J., Townsend, H., Almuhaideb, E., Meredith, J., Rosales, D., Chigbu, P., Dasilva, L., and Parveen, S. 2023. Incidence and pathogenic potential of Shewanella species found in Chesapeake and Maryland Coastal Bays oysters and seawater. University of Maryland Eastern Shore Annual Research Symposium 2023. Princess Anne, MD. April 17-April 20, 2023. [2nd Place Award for Poster Presentation & 3MT].
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2023
Citation:
Johnson, T., Richards, G., Jacobs, J., Townsend, H., Almuhaideb, E., Meredith, J., Rosales, D., Chigbu, P., Dasilva, L., and Parveen, S. 2023.Occurrences and pathogenic potential of Shewanella species found in oysters and seawater from the Chesapeake and Maryland Coastal Bays. American Society of Microbiology (ASM) 2023 Conference. Houston TX. June 15-June 19, 2023.
- Type:
Journal Articles
Status:
Under Review
Year Published:
2023
Citation:
Johnson, T., Richards, G., Jacobs, J., Townsend, H., Almuhaideb, E., Meredith, J., Rosales, D., Chigbu, P., Dasilva, L., and Parveen, S. 2023. Prevalence and pathogenic potential of Shewanella species found in oysters and
seawater collected from the Chesapeake Bay and Maryland Coastal Bay. Applied and Environmental Microbiology (under review by co-authors)
|
Progress 03/01/21 to 02/28/22
Outputs
Target Audience:Target audiences are the oyster harvesting and processing industries in Florida and Maryland, regulatory agencies, risk assessors, risk managers, students, and the scientific community. The efforts to reach the target audiences will include presentations of the research findings at professional meetings, publications in peer-reviewed journals, classroom discussion, workshops, personal communication with the seafood industry, state, and regulatory agencies, extension and outreach. Changes/Problems:Our major problems are the aftermath of those reported in the previous report, namely COVID related effects and the Florida moratorium on harvesting oysters in Apalachicola Bay and its tributaries which is where our sample sites are located. Currently, we are collecting oysters from only one site that is exempted from this rule. What opportunities for training and professional development has the project provided?One technician, two graduate students and three undergraduate students had the opportunities to learn the state- of -the- art microbiology and molecular biology techniques. PIs and students attended the regional, national, and international virtual/in person meetings to share the project findings with the scientific community and update the knowledge in molecular biology, water quality, food microbiology and safety. How have the results been disseminated to communities of interest?One presentation was made at a regional professional meeting, two abstracts were accepted at regional and national/international professional meetings, and PI was invited to serve as speakers at national/international professional meetings. The objectives and findings of this projecthave been shared with middle and high school students through FAMU STEM Day and the Agri discovery program at the University of Maryland Eastern Shore (UMES). We expect to share results with other audiences later in the year. Findings of this project were also shared with the students of independent studies, seminar in food science and technology courses at UMES; 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?To accomplish the goal/objectives, we are planning to conduct the following analyses/experiments during the next reporting period: UMES team-- 1) to analyze data [objectives 1-2) and prepare manuscripts, 2) to characterize Shewanella recovered from oyster and seawater using phenotypic (antimicrobial susceptibility testing) and genotypic (Pulsed-Field Gel Electrophoresis) methods, 3) to share the research findings with the public, seafood industry and regulatory agencies, and 4) to increase capacity and enhance student educational and leadership opportunities. FAMU team--1) to complete collection of samples in fall 2022, 2) to characterize all new isolates and those stored in the freezer for antibiotic resistance and further genomic analysis, 3) to analyze the data, present results to communities of interest, prepare and submit at least one manuscript to a peer reviewed journal.
Impacts
What was accomplished under these goals?
The overall goal of this project is to investigate the prevalence of potentially pathogenic Shewanella spp. in oyster and seawater samples collected from the Apalachicola Bay (AB), Florida, the Chesapeake Bay (CB) and the Maryland Coastal Bays (MCBs), Maryland to enhance seafood safety and water quality research, outreach, and extension at two HBCUs. The specific objectives are:1) To investigate the prevalence of potentially pathogenic Shewanella species in AB, CB and MCBs; 2) To study the effect of environmental factors such as temperature, salinity, pH, and chlorophyll-a on the prevalence of Shewanella in oysters and seawater; 3) To characterize Shewanella recovered from oyster and seawater using phenotypic (antimicrobial susceptibility testing) and genotypic (Pulsed-Field Gel Electrophoresis) methods; 4) To share the research findings with the public, seafood industry and regulatory agencies; and 5) To increase capacity and enhance student educational and leadership opportunities. To accomplish these objectives, oyster and water samples were collected monthly from three sites [Honga River (HR), Tangier Sound (TA), and Horn Point (HP)] in the Chesapeake Bay and one site in the Maryland Coastal Bay (MCB) from March 2021 through October 2021. In addition, physiochemical parameters such as water temperature, salinity, turbidity, dissolved oxygen, chlorophyl-a, and pH were recorded during collection of samples. All samples were analyzed for Shewanella spp. using the standardized methods. In brief, all samples from this period were analyzed using Iron agar plats at 35ºC of 48 hours. Oyster homogenates and seawater samples were serially diluted using 10-fold dilutions (up to 10-4 for water and 10-6 for oyster) and spread plated onto Iron agar. After incubation, plates of 20 to 200 colonies (or fewer if no plate had 20 colonies) were counted and at least 10% (or up to 10) of suspect Shewanella (black) colonies were picked and grown in broth for freezing. Overall, 54% of water and 78% of oyster samples were positively identified as being Shewanella. Thirty-seven water and 34 oyster samples were collected in total. The counts ranged from <1 (January 2021) to 7.04 x 104 (July 2021) in oyster samples and <1 (January 2021) to 1.86 x 102 (September 2021) in water samples. The results indicates that the counts of presumptive Shewanella spp. were lower in water samples than in oyster samples. A total of 79 presumptive Shewanella isolates collected from water and 274 presumptive Shewanella isolates from oyster samples were collected and tested for beta hemolytic activity. In Maryland, 50% of oyster isolates and 31% of water isolates were positive for beta hemolysin. Horn point and Honga River, respectively had the highest percentage of beta hemolytic isolates in comparison to Tangier Sound and Maryland Coastal Bay. This is assumed to be correlated with the differences in water quality from site to site. Moreover, all isolates were confirmed using 16S rRNA sequencing. The top four species isolated from oyster and water samples were S. amazonensis, S. marisflavi, S. lohica and S. algae. There was a moderate significant correlation found for both oyster and water samples with temperature (r = .628) and dissolved oxygen (r = .662) to the prevalence of Shewanella species. Salinity (r= .446) and turbidity (r = .454) are also suspected to have a minor impact on the prevalence of Shewanella species being found in water samples. The pH ranged from 7.5-8.0. Salinity ranged from 8.3 ppt (January 2021) at HP to 34.4 ppt (September 2021) at MCB. Horn Point consistently had the lowest salinities and MCB consistently had the highest. Temperature ranged from 2.0ºC (February 2021) to 29.3ºC (August 2021). Turbidity ranged from 0 FNU (October 2021) to 28.8 FNU (March 2021). Dissolved oxygen ranged from 4.9 mg/L (August 2021) to 12.7 mg/L (February 2021). Chlorophyll-a ranged from 0.2 ug (August 2021) to 27.8 ug (February 2021). The antibiotic sensitivity testing of confirmed isolates has not been started; however, our team has already standardized the methods for this experiment. The result of this study indicates that Shewanella spp., especially beta hemolytic isolates are more prevalent in oyster and water samples during the warmer months. High numbers of Shewanella isolates in oysters and seawater during the summer months suggest the need to continue monitoring levels of pathogenic spp. in the Bays in Maryland and other geographical locations. Pathogenic Shewanella spp. could pose a health threat through the ingestion of contaminated seafood, by cuts or abrasions acquired in the marine environment, or by swimming and other recreational activities. Therefore, cautions should be taken during harvesting of oysters and recreation activities, especially, in the summer months. Our collaborator at the Florida A and M university (FAMU) also investigated the prevalence of Shewanella in Apalachicola Bay in Florida. Oyster and water samples were collected during the period March 2021 to Feb 2022 from one site, the Gulf Coast Marine Specimen Laboratory, because of a 5-year moratorium on oyster harvesting in the Apalachicola Bay. All samples were analyzed for Shewanella using standard methods. In addition, physicochemical parameters were recorded during collection of samples. There was a clear trend between temperature and the prevalence of Shewanella with lower numbers isolated in the winter months when the water temperature was lowest. The salinity was rather constant with little variation and does not appear to be a factor in the prevalence of Shewanella at this site. Moreover, pH had no effect on Shewanella. Seventeen percent (42 out of 242) of isolates recovered from water samples were beta-hemolytic and 18.4% (36 out of 195) recovered from oyster samples were tested as beta-hemolytic. Most of the isolates were confirmed by 16S rRNA gene analysis to be Shewanella algae. The investigators anticipate that the project will motivate minority graduate and undergraduate students to pursue higher education in molecular biology, food microbiology, and food safety as well as careers in these disciplines in academia, industry, and federal and state agencies. Successful completion of project objectives for enumeration of Shewanella species in oyster and seawater samples will provide a reliable tool for monitoring this emerging pathogen in shellfish that can be employed by commercial growers, and regulators. Furthermore, the projects objectives will aid in our understanding of the ecology, antibiotic resistance, and genetic diversity of Shewanella and provide valuable information pertaining to seafood safety and monitoring strategies as well as aquatic and human health.
Publications
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2022
Citation:
Johnson, T., Parveen, S., Richards, G., Meredith, J., Almuhaideb, E., Rosales D., Chigbu, P., Dasilva, L. 2022. Incidence and pathogenic potential of Shewanella species in oysters and seawater collected from the Chesapeake and Maryland Coastal Bays. International Association for Food Protection (IAFP) annual meeting, July 31-August 3, 2022. Pittsburg, PA (accepted).
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2022
Citation:
Parveen, S., 2022. Food Safety and water quality: A journey into research and education at UMES. 1890 Research Directors Research (ARD) Research Symposium. April 02-05, 2022 (invited, accepted).
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2022
Citation:
Johnson, T., Almuhaideb, E., Meredith, J., Rosales, D., Chigbu, P., Dasilva, L., Parveen, S., Richards, G. 2022. Prevalence of Shewanella species in Chesapeake Bay oyster and seawater. ARD annual meeting. April 2-5, 2022. Atlanta, GA (accepted).
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2021
Citation:
Johnson, T., Parveen, S., Richards, G., Meredith, J., Almuhaideb, E., Rosales, D. 2021. Shewanella spp. as potential emerging pathogens in oyster and seawater from the Chesapeake Bay. USDA Congress Meeting, December 15, 2021. The Wye Research Center.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2021
Citation:
Parveen, S. 2021. A journey to food safety research and education. Mujib Centenary Seminar Series II, May 30, 2021. Dept. of Microbiology, University of Dhaka and DUMAA. Virtual.
|
Progress 03/01/20 to 02/28/21
Outputs
Target Audience:Target audiences are the oyster harvesting and processing industries in Florida and Maryland, regulatory agencies, risk assessors, risk managers, students and the scientific community. The efforts to reach the target audiences will include 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:Challenges/problems that we have encountered so far that have caused some delays are: Due to COVID 19 pandemic restrictions, UMES were closed from March 2022-July 2020 and FAMU were closed from March 2021-Ocober 2020. No samples were collected during these periods. Moreover, we were not able to conduct any experiments during these times. The technician was on medical leave for a few week. In addition, FAMU had difficulty to get oysters from the Apalachicola Bay, Florida. What opportunities for training and professional development has the project provided?One postdoctoral associate and one undergraduate student had the opportunities to learn state-of-the-art microbiology and molecular biology techniques. PIs and students were to attend the regional, national and international virtual meetings to share the project findings with the scientific community and update the knowledge in molecular biology, water quality, food microbiology and safety. How have the results been disseminated to communities of interest?Three presentations were made at regional professional meetings. Findings of this project were also shared with the students of independent studies, 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?To accomplish the goal/objectives, we are planning to conduct the following analyses/experiments during the next reporting period: 1) analysis of 16 sRNA sequencing data of samples collected between 2019-2020; 2) collection and analysis of oyster and water samples in 2021 from all sites; 3) to study the effect of environmental factors such as temperature, salinity, pH, chlorophyll a etc. on the prevalence of Shewanella in oysters and seawater; 4) to characterize Shewanella recovered from oyster and seawater using phenotypic (antimicrobial susceptibility testing) and genotypic (Pulsed-Field Gel Electrophoresis) methods; 5) to share the research findings with the public, seafood industry and regulatory agencies; and 6) to increase capacity and enhance student educational and leadership opportunities.
Impacts
What was accomplished under these goals?
The overall goal of this project is to investigate the prevalence of potentially pathogenic Shewanella species in oyster and seawater samples collected from the Apalachicola Bay, Florida, the Chesapeake Bay and the Maryland Coastal Bays, Maryland to enhance seafood safety and water quality research, outreach, and extension at two HBCUs. To accomplish this goal, we made several conference calls with our collaborators to discuss the issues related to lockdown/limited access to the laboratories due to COVID 19 pandemic, non- availability of oysters in the Apalachicola Bay, methodology for collection and analysis of samples during this reporting period. Due to COVID-19, we were not able to collect samples from April through June. However, we collected samples monthly from July 2020 through February 2021 from three sites [Honga River (HR), Tangier Sounds (TA), and Horn Point (HP)] in the Chesapeake Bay and one site in the Maryland Coastal Bays (MCBs). All samples were analyzed for presumptive Shewanella species (spp.). In addition, physiochemical parameters, such as water temperature, salinity, turbidity, dissolved oxygen, chlorophyll a, and pH were recorded during collection of samples. All samples were analyzed for Shewanella spp. using the standardized methods. A total of 347 presumptive Shewanella colonies were collected. Horn point was the site to provide the most presumptive colonies with a total of 163, while MCBs displayed the least number of presumptive colonies with a total of 5. Honga River had 119 presumptive colonies and TA had 60. In the warmer months (July through September), we observed the highest level (46.3%) of beta-hemolytic presumptive Shewanella spp. in oyster and water samples. Data also showed that the most presumptive Shewanella colony growth on iron agar occurred during the months of July and August, specifically for the site HP. During these months, presumptive Shewanella count was averaged to be ~2 X 104 LogCFU/g for oyster samples and ≥ 9 X 101 LogCFU/mL for the water samples. These findings are consistent with the results of the last year. The pH ranged from 7.2 (September 2020) to 8.2 (January 2021). Salinity ranged from 7.1 psu. (July 2019) at Horn Point to 35.1 psu (September 2021) at MCBs. Horn Point and HR consistently had the lowest salinities and MCBs and TA consistently had the highest salinity. Temperature ranged from 2.0°C (February 2021) to 30.7°C (July 2021). Turbidity ranged from 0.5 Formazin Nephelometric Units (FNU) (January 2021) to 36.4 FNU (September 2020). Dissolved oxygen ranged from 3.7 mg/L (August 2020) to 12.7 mg/L (February 2021). Chlorophyll a ranged from 0.8 µg/L (January 2021) to 27.8 µg/L (February 2021). Interestingly, all chlorophyll a values ≥ to 10 µg/L were typically readings from the HP during both warm and cold months. These results suggest that beta hemolytic Shewanella spp. were more prevalent during the warmer months. We were also able to ship a total of 924 presumptive Shewanella spp. (collected 2019-2020) to Genewiz, a Biotech company, for 16S RNA sequencing. In addition, we developed the protocol for the antibiotic susceptibility testing of Shewanella spp. Our collaborators at Florida A and M University (FAMU) were not able to work in the laboratory due to operation restrictions imposed because of COVID-19 from March through October 2020. Samples collections were resumed in November 2020, but they found that the state of Florida had declared a five-year moratorium on oyster sampling in the Apalachicola Bay (AB) to allow for replenishment of the oyster population. Up to that point, oyster samples were routinely collected from two sites in the Apalachicola Bay. An oyster farm (OF) in Oyster Bay that they had gotten samples from raised their prices substantially, making it prohibitive for them to continue getting oysters from there. The last samples from that oyster farm were collected in November and December 2020. As a result of these challenges, FAMU had to find new locations outside of Apalachicola Bay from which to obtain oysters. One new site was the Gulf Specimen Marine Laboratory (GSML), from where samples were collected in December 2020. FAMU is currently in negotiation with another oyster farm in Apalachicola Bay to get oysters. During collection of samples, water temperature, pH and salinity were recorded. The highest level of presumptive Shewanella was observed in OF compared to that of AB and GSML. Sixty percent of presumptive Shewanella spp. were beta-hemolytic. The water temperature, salinity and pH ranged from 11.10-24.4?, 15.95-25.5 psu and 7.2-8.01, respectively. To identify at species level, presumptive Shewanella species were analyzed using the 16S rRNA sequencing. For this experiment, isolates were streaked onto TSA plates, and individual colonies were inoculated in the tryptic soy broth. Genomic DNA was isolated from these cultures and subjected to PCR for coding gene amplification. Further, PCR products were purified and subjected to sequencing. The results indicate that most of the isolates were Shewanella algae. The investigators anticipate that the project will motivate minority graduate and undergraduate students to pursue higher education in molecular biology, food microbiology, and food safety as well as careers in these disciplines in academia, industry, and federal and state agencies. In addition, successful development of project objectives for enumeration of Shewanella species in oyster and seawater samples will provide a simple, rapid and reliable tool for monitoring this emerging pathogen in shellfish that can be employed by commercial growers, state resource managers, regulators and educational institutions. In addition, the project's objectives will augment our understanding of the ecology, antibiotic resistance and genetic diversity of Shewanella and provide valuable information pertaining to seafood safety and monitoring strategies as well as aquatic and human health.
Publications
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2021
Citation:
Parveen, S., 2021. Bangladesh Association for Food Protection (BAFP) in North America. The First Webinar on Bridging knowledge Gaps in Food and Water Safety. BAFP. February 20, 2021. Virtual.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2020
Citation:
Parveen, S., 2020. Seafood Safety and global pandemic. Geosciences Summer Bridge Program, UMES. Aug. 06, 2020. Virtual.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2020
Citation:
Parveen, S., 2020. Sea Food Safety and COVID-19. LSAMP Annual meeting, July 19, 2020, UMES. Virtual.
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Progress 03/01/19 to 02/29/20
Outputs
Target Audience:Target audiences are the oyster harvesting and processing industries in Florida (FL) and Maryland (MD), regulatory agencies, risk assessors, risk managers, students and the scientific community. The efforts to reach the target audiences include presentations of the research findings at professional meetings, publications in peer-reviewed journals, classroom discussion, workshops, personal communication with the seafood industry, state, and regulatory agencies, extension and outreach. Changes/Problems:Due to nonavailablity of oysters, we collected samples from two sites in Florida instead of three sites. What opportunities for training and professional development has the project provided?One postdoctoral associate, one technician, two graduate students, and two undergraduate students had the opportunities to learn state-of-the-art microbiology and molecular biology techniques. One undergraduate student graduated with experience in food and environmental microbiology. PIs and students 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, water quality, food microbiology and safety. How have the results been disseminated to communities of interest?Two presentations were made and one undergraduate thesis was published. Findings of this project were also shared with the students of independent studies, 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?To accomplish the goal/objectives, we are planning to conduct the following analyses/experiments during the next reporting period: 1) analysis of samples collected in spring 2020; 2) sequencing and analysis of presumptive isolates for confirmation at species level; 3) investigating the prevalence of potentially pathogenic Shewanella species in AB, CB and MCBs; 4) determining the effect of environmental factors such as temperature, salinity, pH, dissolved oxygen, turbidity and chlorophyll a on the prevalence of Shewanella in oysters and seawater; 5) characterizing Shewanella recovered from oyster and seawater using phenotypic (antimicrobial susceptibility testing) and genotypic (Pulsed-Field Gel Electrophoresis) methods; 6) to share the research findings with the public, seafood industry and regulatory agencies; and 7) to increase capacity and enhance student educational and leadership opportunities.
Impacts
What was accomplished under these goals?
The overall goal of this project is to investigate the prevalence of potentially pathogenic Shewanella species (spp.) in oyster and seawater samples collected from the Apalachicola Bay (AB), Florida, the Chesapeake Bay and the Maryland Coastal Bays (MCBs), Maryland to enhance seafood safety and water quality research, outreach, and extension at two HBCUs. To accomplish this goal, we conducted several trial and error experiments to determine the optimum medium and incubation conditions for the growth of Shewanella from March through June 2019. We compared TSA with added salt and Iron agar. We also compared growth at 30 and 37 degree C and incubation times of 24 and 48 hrs. We determined the optimum medium is Iron agar and the best incubation conditions are 48 h at 37 degree C. In summer 2019, one graduate and one undergraduate students were hired and trained in microbiology and molecular biology to collect and analyze oyster and water samples. In fall 2019, one M.S. student was hired and trained in microbiology and molecular biology to conduct research for her Master's thesis in partial fulfillment of the requirements for her degree program. Florida A and M University also trained one undergraduate student in microbiology and molecular biology through direct involvement in this project to collect data for her undergraduate thesis. We also identified a biotech company to sequence presumptive isolates for confirmation using 16s RNA sequencing. During this period, the postdoctoral associate tried developing Shewanella-specific PCR and qPCR primers for easy detection and enumeration, targeting 16S rRNA genes, and found that targeting SSU ribosomal genes for this purpose will likely not be free of biases as this gene is highly conserved among the bacterial domain. She found a Shewanella-specific primer was previously designed and published, but it will amplify non-pathogenic strains as well, which is not our target. For this study, oyster and water samples were collected monthly from three sites [Honga River (HR), Tangier Sounds (TA), and Horns Point (HP)] in the Chesapeake Bay and one site in the MCBs from March 2019 through February 2020, and two sites [Cat point (CA)] and Oyster farm (OF)] in the AB. In addition, physiochemical parameters, such as water temperature, salinity, turbidity, dissolved oxygen, chlorophyll a, and pH were recorded during collection of samples. All samples were analyzed for Shewanella spp. using the standardized methods. As our preliminary experiments indicates that plating of samples on Iron agar plate and incubation of inoculated plates at 37 degree C for 48 hours provide optimum results, all samples were analyzed using Iron agar plates at 35 degree C for 48 hr. Oyster homogenates and seawater samples were serially diluted using 10-fold dilutions and spread plated onto Iron agar. Initially, we used both Iron agar and TSA, until we determined that Iron agar would be our medium of choice. After incubation, plates of 20 to 200 colonies (or fewer if no plate had 20 colonies) were counted and at least 10 percent of suspect Shewanella colonies (black colonies on Iron agar or orange colonies on TSA) were picked and grown in broth for freezing. During the months of June and July 2019, 53 suspect colonies were isolated from Honga River and Maryland Coastal Bay sites. These colonies were tested using the API 20E system for biochemical identification of Enterobacteriaceae. Of the samples that underwent biochemical analysis, 25% of the samples from MCBs and 83% from HR were confirmed to be S. putrefaciens. Overall, 92% and 88.8% of samples were positive for presumptive Shewanella spp. in Maryland and Florida, respectively. The counts ranged from <1 (March 2020) to 5x104 CFU/g (August 2019) in oysters in Maryland and <1 (December 2020) to 3.5x102 CFU/g (October) in oysters in Florida, respectively. In water samples, the counts ranged from <1 (November 2019) to 6.7 x 10 CFU/mL (October 2019) and <1 (December) to 4.7 x10 CFU/mL (October) except for the month of October (2.2x102 CFU/mL) in Maryland and Florida, respectively. The results indicate that the counts of presumptive Shewanella spp. were lower in water than in oyster samples in both regions. A total of 671 and 718 presumptive Shewanella isolates collected from Maryland and Florida were tested for beta hemolytic activity. In Maryland, 42% of isolates were positive for beta hemolysin. In the warmer months (June through August 2019), more than 25% isolate recovered from the Chesapeake Bay exhibited beta hemolytic activity. In contrast, only <1% isolate were positive for beta hemolytic activity in the MCBs. Approximately, 30% of isolates were beta-hemolytic in Florida. Currently, experiments are underway to confirm and characterize these isolates using genotypic and phenotypic methods. The pH ranged from 7.2 (July 2019) to 10.7 (November 2019). Salinity ranged from 6.26 ppt. (July 2019) at Horn Point to 31.82 ppt (December 2019 at MCBs). Horn Point and HR consistently had the lowest salinities and MCBs consistently had the highest salinity. Temperature ranged from 3.0°C (January 2020) to 31.3°C (July 2019). Turbidity ranged from 0.6 Formazin Nephelometric Units (FNU) (January 2020) to 48.1 FNU (July 2019), with the exception of one reading of 500.4 at HR in October. Dissolved oxygen ranged from 3.3 mg/L (July 2019) to 12.0 mg/L (December 2019). Chlorophyll ranged from 1.1 µg/L (December 2019) to 77.4 µg/L (August 2019). Interestingly, all chlorophyll values ≥ to 10 µg/L were from the HP or HR sites during both warm and cold months. In Florida, temperature, salinity, dissolved oxygen and pH were recorded during collection of samples only from CP. The water temperature ranged between 13.8 (February 2020) and 30.4? (August 2019). Salinity ranged between 0.31 (February 2020) and 32.89 ppt. (November 2019). pH ranged between 7.94 (November 2019) and 8.49 (August 2019), while dissolved oxygen ranged between 6.04 (October 2019) and 10.33 mg/L (February 2020). The results of this study indicate that Shewanella spp., especially beta hemolytic isolates were more prevalent in oyster and water samples during the warmer months in both geographical regions. High numbers of Shewanella isolates in oysters and seawater during the summer months suggest that monitoring the levels of pathogenic spp. should be continued in the Bays in Maryland, Florida and other geographical locations. Pathogenic Shewanella spp. could pose a health threat through the ingestion of contaminated seafood, by cuts or abrasions acquired in the marine environment, or by swimming and other recreational activities. Therefore, cautions should be taken during harvesting of oysters and recreational activities, especially, in the warmer months. The investigators anticipate that the project will motivate minority graduate and undergraduate students to pursue higher education in molecular biology, food microbiology, and food safety as well as careers in these disciplines in academia, industry, and federal and state agencies. In addition, successful development of project objectives for enumeration of Shewanella species in oyster and seawater samples will provide a simple, rapid and reliable tool for monitoring this emerging pathogen in shellfish that can be employed by commercial growers, state resource managers, regulators and educational institutions. In addition, the project's objectives will augment our understanding of the ecology, antibiotic resistance and genetic diversity of Shewanella and provide valuable information pertaining to seafood safety and monitoring strategies as well as aquatic and human health.
Publications
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2019
Citation:
Hernandez, R. Meredith, J., and Parveen, S. 2020. Prevalence of Shewanella in oysters and seawater in the Chesapeake Bay and the Maryland Coastal Bays. REU presentation. August 2019. UMES, Paul S. Sarbanes Coastal Ecology Center, MD.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2020
Citation:
Hernandez, R. and Parveen, S. 2020. Prevalence of Shewanella in oysters and seawater in Maryland. 2020. Emerging Researchers National (ERN) Conference in STEM. Washington, D.C. February 6-8, 2020.
- Type:
Theses/Dissertations
Status:
Accepted
Year Published:
2019
Citation:
Gleason, M. 2019. The prevalence of Vibrio and Shewanella spp. in oyster and seawater in Apalachicola Bay. Undergraduate thesis. Florida A and M University.
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Progress 03/01/18 to 02/28/19
Outputs
Target Audience:Target audiences are the oyster harvesting and processing industries in Florida and Maryland, regulatory agencies, risk assessors, risk managers, students and the scientific community. The efforts to reach the target audiences will include 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:Challenges/problems that we have encountered so far that have caused some delays are: the delay in receiving the Iron Agar (only commercial supplier of Iron Agar is in Spain; it took several months to receive the product from the supplier due to back order and quality control issues) and the technician was on sick leave for several weeks due to an injury. What opportunities for training and professional development has the project provided?One postdoctoral associate, one technician, two graduate students, and two undergraduate students had the opportunities to learn state-of-the-art microbiology and molecular biology techniques. How have the results been disseminated to communities of interest?The purpose, objectives and initial findings of the project have been shared through an invited presentation at a professional meeting. What do you plan to do during the next reporting period to accomplish the goals?To accomplish the goal/objectives, we are planning to conduct the following analyses/experiments during the next reporting period: 1) analysis of samples collected in spring 2019; 2) standardize the colony PCR method for the rapid detection and confirmation of this bacterium in oyster and water samples; 3) to investigate the prevalence of potentially pathogenic Shewanella species in AB, CB and MCBs; 4) to study the effect of environmental factors such as temperature, salinity, pH, and chlorophyll a on the prevalence of Shewanella in oysters and seawater; 5) to characterize Shewanella recovered from oyster and seawater using phenotypic (antimicrobial susceptibility testing) and genotypic (Pulsed-Field Gel Electrophoresis) methods; 6) to share the research findings with the public, seafood industry and regulatory agencies; and 7) to increase capacity and enhance student educational and leadership opportunities.
Impacts
What was accomplished under these goals?
The overall goal of this project is to investigate the prevalence of potentially pathogenic Shewanella species in oyster and seawater samples collected from the Apalachicola Bay, Florida, the Chesapeake Bay and the Maryland Coastal Bays, Maryland to enhance seafood safety and water quality research, outreach, and extension at two HBCUs. To accomplish this goal, we made several conference calls with our collaborators to discuss the hiring of students, technician, postdoctoral associate, and methodology for collection and analysis of samples in spring, summer and fall 2018. We established collaboration with three oyster farms in Maryland and Florida Fish and Wildlife Conservation Commission and one oyster farm in Florida. Florida Agricultural and Mechanical University (FAMU) initiated a search for a postdoctoral associate position and hired Dr. Kumari Richa in fall 2018. She has expertise in marine microbial ecology, cultivation and molecular techniques along with bioinformatics. University of Maryland Eastern Shore (UMES) trained one technician, two graduate and one undergraduate students to collect and analyze samples. In winter 2019, UMES identified one M.S. student to conduct research in partial fulfillment of the requirements for her degree program. FAMU also trained the postdoctoral associate and one undergraduate student in microbiology and molecular biology through direct involvement in this project. Moreover, they identified two summer interns (undergraduate students) to work on this project which will help them writing their undergraduate theses. This fall, FAMU also anticipates recruiting a graduate student to work on this project. Investigators, postdoctoral associates, technicians and students also visited the Chesapeake Bay and the Maryland Coastal Bays, Maryland and the Apalachicola Bay Florida to select sampling sites and to learn how to collect samples and physico-chemical parameters of water. Another accomplishment was setting up of our laboratories in terms of ordering the required supplies needed for this project, e.g. blenders, tubes, Iron Agar, other media and reagents. UMES developed a protocol and have had several conference calls with collaborators (FAMU and USDA ARS) to finalize procedures that we will be following and to discuss all aspects of the project from sample collection to laboratory testing. Several experiments were conducted to standardize the methods for enumeration of Shewanella species from oyster and water samples. Initial collection of samples was performed in February 2019 from one site in the Apalachicola Bay and one site in the Chesapeake Bay. In addition, physico-chemical parameters were recorded during the collection of samples. All samples were analyzed for Shewanella species using the standardized methods. Currently, experiments are underway to confirm and characterize these isolates using genotypic and phenotypic methods. In addition, FAMU is developing a colony PCR method using Shewanella-specific primers for rapid detection and confirmation of this bacterium recovered from oyster and water samples. The project is in its initial stage and results are inadequate to assess impact. The investigators anticipate that the project will motivate minority graduate and undergraduate students to pursue higher education in molecular biology, food microbiology, and food safety as well as careers in these disciplines in academia, industry, and federal and state agencies. In addition, successful development of project objectives for enumeration of Shewanella species in oyster and seawater samples will provide a simple, rapid and reliable tool for monitoring this emerging pathogen in shellfish that can be employed by commercial growers, state resource managers, regulators and educational institutions. In addition, the project's objectives will augment our understanding of the ecology, antibiotic resistance and genetic diversity of Shewanella and provide valuable information pertaining to seafood safety and monitoring strategies as well as aquatic and human health.
Publications
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2018
Citation:
Hashem. F., S. Parveen, E. May. 2018. Water quality, fresh Produce and seafood safety research at UMES. Nov. 2018. University of Florida, Gainesville. FL.
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