Salmonella Serovar Identification via LAMP

Whole-genome sequences of Salmonella serovars were obtained from the National Center for Biotechnology Information (NCBI) database GenBank (https://www.ncbi.nlm.nih.gov/, Bethesda, MD, USA) and compared using the Basic Local Alignment Search Tool (BLAST, https://blast.ncbi.nlm.nih.gov/Blast.cgi) to identify potential serovar-specific genome regions. Whole genome sequences (in GenBank format) of one representative of each serovar were compared in subsections of 100,000 base pairs in length using BLAST search to identify regions of lower sequence homology to other serovars. These regions were then used to calculate LAMP primer sets and the calculated amplicons were in turn analyzed for specificity by BLAST analysis once again. LAMP primer sets targeting the identified specific DNA sequences were designed via LAMP Designer software 1.16 (PREMIER Biosoft, San Francisco, CA, USA; Table 1).Table 1

Target Salmonella genes used for LAMP primers of the SalmoTyper assay

Assay parameter	Gene	Gene product	GenBank accession no	Primer	Sequence (5′-3′)	Amplicon size	Predicted cross-reactions^a
Salmonella spp.	invA	Invasion protein InvA	NC_003197.2	F3-SeinvA B3-SeinvA LF-SeinvA LB-SeinvA FIP-SeinvA BIP-SeinvA	ATCGCACTGAATATCGTACTG CCACGGTGACAATAGAGAAG GGTAATTAACAGTACCGCAGGA CTTGATTGAAGCCGATGCC AATGCCAGACGAAAGAGCGTTCGTTCTACATTGACAGAATCC TCGATCAGTACCAGCCGTCTTACGCCAATAACGAATTGCC	163 bp	None
S. Enteritidis	C0Z09_12860	Transcriptional regulator	CP025554.1	F3-Salend1 B3-Salend1 LF-Salend1 LB-Salend1 FIP-Salend1 BIP-Salend1	CTGGTACTTACGATGACAACTT GGGAGGGAGGAGCTTTAG TTCTTTCTCAGATTCAGGGAGT CTGCACAAAAGCGCCTAAA ACCTTTAAGCCGGTCAATGAGTCAGACAACAGGCTGATTTACTA AGAGGCTTTCCAGAACATGCTCATGTGGTTGGTTCGTCAC	165 bp	S. Javiana S. Newlands
S. Typhimurium	JYM79_16920	RHS repeat protein	CP074209.1	F3-Stymur1 B3-Stymur1 LF-Stymur1 LB-Stymur1 FIP-Stymur1 BIP-Stymur1	GGCGGGTTAAACCTTTATCA ATGTTTGCTCACCCACAAT CTACTGTCCCAGTATCAAGCAT TTCTATCGAAGGGCGATTAGC GGTAGGCAACATGAGTTGTCCCCTGCGGAATATCAGCAA CCTACAAATGAAGGATGGGCAATGTTGGCATTTCAGGAACA	255 bp	S. Abony S. Adjame S. Thompson S. Bredeney S. Give S. Quebec
S. Infantis	yecD_1	Putative isochorismatase hydrolase	LS483479.1	F3-Salinf2 B3-Salinf2 LF-Salinf2 LB-Salinf2 FIP-Salinf2 BIP-Salinf2	GGCTATTCATCCTGATGTCG TATGCCCAGGCTTCTGATA ACACCGGTAAGGATCAGGT CGTTGTGAGTGATTGCTGC AAGCACAACACCACTGGTAGTCAACTTGAGATGATCCTTCGTG TGACCTGGATTACCGCCTCAACATTGGTGTCATGATCTGG	163 bp	S. Sloterdijk
S. Derby	JF569_ 15,965	YadA-like family protein	CP066545.1	F3-Salder3 B3-Salder3 LF-Salder3 LB-Salder3 FIP-Salder3 BIP-Salder3	AATAGTCAGCCAGTGAATCAC ACCAGTATCACCAACCTGA CATCTACCAGTACCGACGC CATCCCATTTCAGTGCATCTTC CGTCACCGCTGGAACAATCTAAGAGTTGTCCACCGTTAAC CAGCGGTGAATACGCCTGATACGGTGACTAACCTTGGT	159 bp	S. Ouakam S. California
S. Choleraesuis	SCH_ 1198	Predicted antirepressor	CP012344.2	F3-Salchol B3-Salchol LF-Salchol LB-Salchol FIP-Salchol BIP-Salchol	TTGAATTATGGTGGCTTGGGTA GTTACATCGGTGTATTTGCTGAG ATGGTAGTCATAACGACAGCC TTATTTCCATAAGCCACATAAAGACG CATCAATAACGCAAAGTTCAGGTTGATAGCAGCTTGTGGTGATAAGT TTACTTCCTCACTGGCTGTTGCTTCGACAGAACAATCCAGGC	193 bp	None

^aAmplicon sequence homology > 95%

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Edel B., Glöckner S., Stoll S., Lindig N., Boden K., Wassill L., Simon S., Löffler B, & Rödel J. (2023). Development of a rapid diagnostic test based on loop-mediated isothermal amplification to identify the most frequent non-typhoidal Salmonella serovars from culture. European Journal of Clinical Microbiology & Infectious Diseases, 42(4), 461-470.

Publication 2023

Dna sequences Genes Genome Isochorismatase Primers Protein Salmonella

Corresponding Organization :

Other organizations : Jena University Hospital, Synlab (Germany), Robert Koch Institute

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Variable analysis

independent variables

Whole-genome sequences of Salmonella serovars

dependent variables

Identification of potential serovar-specific genome regions
Calculation of LAMP primer sets
Analysis of LAMP primer set specificity

control variables

Whole genome sequences were compared in subsections of 100,000 base pairs in length using BLAST search
LAMP primer sets were designed via LAMP Designer software 1.16

controls

Positive control: None mentioned
Negative control: None mentioned

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