Researchers at an Australian university have developed DNA tests that can identify the five most common Salmonella subtypes in the country.

The tests can detect as few as 10 copies of DNA in eight minutes and work at a constant temperature unlike other methods that need specialized equipment for temperature cycling. They were developed using pure cultures so they will need validation in clinically relevant conditions, but they could be used for culture-independent serotyping of common Salmonella serovars from clinical samples.

Scientists at the University of New South Wales (UNSW) said pending further research, they could help public health laboratories and industry stop the spread of Salmonella outbreaks. Findings were published in The Journal of Molecular Diagnostics.

Tracing future outbreaks
In 2017, more than 16,000 cases of Salmonella were reported in Australia — a 30 percent increase on the previous 10-year average.

The five most common serovars cause more than 85 percent of Salmonella infections in the country. However, two of these — Typhimurium and Enteritidis — are also top types across the world, so researchers believe results are applicable to other geographies.

Senior author Professor Ruiting Lan, of the UNSW School of Biotechnology and Biomolecular Sciences, said the tests could play a role in tracing the origin of future Salmonella infections.

“It is essential for public health investigators to have a fast, simple way of tracking down the source of Salmonella outbreaks — so, the ability to test for different types of Salmonella is important,” he said.

Using an isothermal amplification technique, called multiple cross-displacement amplification (MCDA), seven assays were developed and evaluated for rapid detection and differentiation of the five most common Salmonella serovars in Australia: Typhimurium, Enteritidis, Virchow, Saintpaul, and Infantis.

Salmonella in a clinical or food sample or in fecal matter may exist in tiny amounts and requires sensitive methods to detect, said Lan.

“Our enhanced Multiple Cross Displacement Amplification method can detect tiny amounts of DNA rapidly and at a constant temperature, which makes it an excellent fit for a simple, rapid and sensitive bacterial detection test,” Lan said. “It’s a clear improvement upon the existing MCDA test for Salmonella that does not distinguish between different subtypes of Salmonella.”

Trend for culture-independent diagnostic tests
Traditional methods to distinguish Salmonella serotypes involve growing the bacteria in culture from samples and then testing them to assign them to a serovar.

Lan said the DNA tests developed in the study were unique because the gene markers used were seleced from analyzing thousands of Salmonella genomes. It paves the way for rapid serotyping directly from specimens.

“It’s difficult to know when our tests would become available, but they are part of the global trend toward culture-independent diagnostic tests which can identify the bacteria causing a foodborne illness without the need to culture the bacteria in a lab,” he said.

Sensitivity and specificity of the seven MCDA assays were evaluated using 79 target and 32 non-target strains. The assays were all sensitive and specific to target serovars, with the sensitivity ranging from 92.9 to 100 percent and specificity from 93.3 to 100 percent.

First author Xiaomei Zhang, UNSW science doctoral  candidate, said the detection method will be crucial to help control the spread of infection during outbreaks.

“Thousands of Salmonella serotypes exist, but we developed and evaluated seven MCDA tests for the rapid detection and differentiation of the five most common Salmonella serotypes in Australia,” said Zhang.

“It’s important to be able to detect the different serotypes because some are more likely to be associated with local infections while others are more likely to be associated with imported cases. Another advantage is the fact that our tests do not require specialized detection equipment, thereby simplifying future application in clinical or industrial settings.”