APHL 2022 POSTER ABSTRACTS
Food Safety
Rapid Sequencing : A Tale of iSeq and MinION
J Neal-McKinney 1 , O Kaluoch 2 , D Lau 2 , T Lee 2 , W-H Wu 1 , A Tran 2 and J Hu 1 ; 1 FDA Pacific Northwest Laboratory , 2 FDA San Francisco Laboratory
Whole genome sequencing of foodborne pathogens allows public health agencies to characterize bacterial isolates and identify similarities between strains . The bulk of bacterial genome sequencing done by the US Food and Drug Administration and GenomeTrakr network laboratories is performed using the Illumina MiSeq platform . In this study , we evaluate the utility of two more recent sequencing instruments , the iSeq from Illumina and the MinION from Oxford Nanopore . Both instruments are well-suited to the sporadic sequencing of real-time isolates as they rapidly produce sequence data on a smaller number of samples than the MiSeq , and less consumables are used per sequencing run . Reference strains of Escherichia coli , Salmonella enterica , Listeria monocytogenes , Campylobacter jejuni and Vibrio parahaemolyticus were sequenced on the iSeq using 2 x 150bp library preparation and the MinION using 1D library preparation . We found that the iSeq produces sequencing data in ~ 18 hours with quality metrics similar to MiSeq (> 80 % Q-Score 30 ). In contrast , the runtime of the MinION can be adjusted as desired , but we found that the bulk of data was produced within 12 hours . The MinION reads were much longer with an average length greater than 10kb , but the quality was greatly reduced ( average Q-score 10 ). Sequence data from both instruments was sufficient for bacterial species identification . Genome assemblies of the iSeq and MinION data were generated individually as well as combined into a hybrid genome assembly using Unicycler . The iSeq assemblies were very accurate yet fragmented into many contigs while the MinION assemblies were contiguous but contained regions of low-quality sequence . The hybrid genome assembly method resulted in complete and accurate genomes that provide greater detail than either method individually . Overall , the results of our studies show that both iSeq and MinION can be used to rapidly confirm and characterize the genetic identity of bacterial isolates .
Presenter : Jason Neal-McKinney , US Food and Drug Administration , jason . neal-mckinney @ fda . hhs . gov
Where in the World is Salmonella Sandiego ?
E Thomson , P O ’ Brien , K Kuwazaki , J Takiguchi , D Ornellas and M Ching-Lee , Hawaii State Department of Health
Introduction : Since 2017 , multiple cases of Salmonella Sandiego have been reported to the Hawaii State Department of Health ( HDOH ), primarily among Maui residents . No source has been identified . Hawaii has not previously had Salmonella Sandiego cases reported . An epidemiological investigation was launched in 2021 to investigate this cluster .
Methods : Case demographics and risk factors were assessed using a standardized Salmonella interview form . A case is defined as a person who experienced diarrhea and / or fever and had S . Sandiego confirmed by culture or culture-independent diagnostic test and serotyping from 2017 – present . Four cases with urine specimens were included . Specimens positive for Salmonella were sent to the State Laboratory for further analysis . Serotyping was conducted via conventional agglutination methods . From 2019 – 2021 , cluster isolates were characterized using whole genome sequencing , while isolates from 2017 – 2018 were characterized using pulsed-field gel electrophoresis . Environmental samples collected include four 2-L water samples from locations along the Kihei coastline .
Results and Discussion : Of 38 Salmonella Sandiego cases reported to HDOH from 2017 – present , ages ranged from 0 – 82 years with a median age of five years old . Fourteen ( 37 %) were ≤2 years old at time of illness . Twenty-two cases ( 58 %) were female and 16 ( 42 %) were male . Five cases ( 13 %) were hospitalized and no deaths reported . Sixty-eight percent of cases resided in a single coastal community ; 24 % who resided elsewhere reported spending time in that area . Three cases ( 8 %) were non-residents . The most recent case was in December 2021 . Of six ( 43 %) cases ≤2 years old in 2021 , 33 % were formula-fed and 67 % were breast-fed . Additional exposures assessed for cases included local produce ( 3 / 14 , 21 %), recreational water ( 4 / 14 , 29 %) and animals at home ( 4 / 14 , 29 %). Two out of three national isolates ( UT , WA ) reported exposures on west Maui . All Salmonella Sandiego isolates were within 0 – 14 alleles of each other . Cluster isolates received at NARMS indicate intermediate interpretation to colistin . After review of water quality reports on Maui , two consecutive samples had increased levels of indicator organisms at an ocean swimming area in September 2021 . Water samples taken in December 2021 from the Kihei coastline were filtered , enriched and isolated a Group B Salmonella , H-Factor I .
Conclusion : HDOH continues to investigate a Salmonella Sandiego cluster now comprising of 38 cases over a four-year period , with common exposure to a coastal Maui community . Standard investigative techniques have been unsuccessful in identifying the source . Next steps we are considering include open-ended interview techniques , applying the CDC hypothesis generating questionnaire , inquiring on food sampling from the cases ’ household and consultation from our foodborne illness center of excellence .
Presenter : Ellen Thomson , Hawaii State Department of Health , ellen . thomson @ doh . hawaii . gov
Development and Validation of a Multiplex Real-time Quantitative PCR Method for the Detection and Speciation of Vibrio Bacteria
M Eickhoff , F-M Wu , M Ellethy , R Siderits and T Kirm , New Jersey Public Health and Environmental Laboratories
Vibrios are marine bacteria that are highly abundant in coastal waters worldwide . Vibrio bacteria are found both free living in aquatic environments and associated with marine organisms including fish , crustaceans and algae . While most of the over 100 identified Vibrio species are not human pathogens , approximately 12 Vibrio species are the causative agents of human illness . In the United States , the CDC estimates that approximately 80,000 cases of Vibrio illness occur each year , of which approximately 52,000 cases are caused by eating foods contaminated with Vibrio bacteria , often raw or undercooked contaminated seafood . Most Vibrio infections result in mild or moderate gastroenteritis , while severe cases can lead to septicemia . One species , Vibrio vulnificus , can cause both foodborne illness and severe wound infections . Because Vibrios favor warm waters , most Vibrio infections occur during summer months , and several studies have correlated rising ocean temperatures with increased Vibrio concentrations and , likewise , infections . Here , we develop a multiplex real-time TaqMan PCR method for the rapid detection and speciation of Vibrio bacteria .