Performance of molecular methods for the detection of Salmonella in human stool specimens

Abstract

Background: The relationship between asymptomatic Salmonella exposure within the gastrointestinal tract and Salmonella bacteraemia is poorly understood, in part due to the low sensitivity of stool culture and the lack of validated molecular diagnostic tests for the detection of Salmonella in the stool. The study aimed to determine a reliable molecular diagnostic test for Salmonella in stool specimens. Methods: We optimised an in-house monoplex real-time polymerase chain reaction (PCR) for the detection of Salmonella ttr and InvA genes in stool by including a selenite broth pre-culture step for Salmonella before DNA extraction and validated their specificity against other local common pathogens. Then we assessed their performance against a well-validated multiplex PCR targeting the same ttr and InvA genes and against stool culture using clinical stool specimens collected from a cohort of 50 asymptomatic healthy Malawian children that were sampled at 1-month intervals over 12 months. We employed a latent Markov model to estimate the specificities and sensitivities of PCR methods. Results: Ttr and InvA primers were both able to detect all the different Salmonella serovars tested and had superior limits of detection when DNA was extracted after selenite pre-culture. Ttr sensitivity and specificity for monoplex-PCR were (99.53%, 95.46%) and for multiplex-PCR (90.30%, 99.30%) respectively. InvA specificity and specificity for using monoplex-PCR was (95.06%, 90.31%) and multiplex-PCRs (89.41%, 98.00%) respectively. Sensitivity and specificity for standard stool culture were 62.88% and 99.99%, respectively. Culture showed the highest PPV (99.73%), and monoplex-ttr had the highest NPV (99.67%). Conclusion: Test methods demonstrated high concordance, although stool culture and monoplexed ttr primers had superior specificity and sensitivity, respectively. The use of selenite pre-enrichment step increased Salmonella detection rate. Taken together, molecular detection methods used here could be used to reveal the true extent of both asymptomatic and symptomatic Salmonella exposure events.

Evaluation of Real Time Polymerase Chain Reaction for Salmonella Invasion Gene A and Salmonella Tetrathionate Respiration Gene as a Diagnostic Test for Typhoid Fever

“…The test is rapid than culture even with the use of preenrichment step and can be used for batch processing of the specimens. The used primers and the probes for the ttr and Inv A were previously evaluated for use in stool samples and demonstrated the ability to evaluate various Salmonella strains, including S. Enteritidis, S. Typhimurium and S. Typhi 16 .…”

Blood Clots Per Rectum in a Pediatric Patient With Salmonella Enteritis: A Case Report

“…Therefore, fecal leukocyte examination and stool lactoferrin detection should not be used to establish the cause of acute infectious diarrhea [1]. Another factor is that stool culture is currently considered to be the gold standard for the detection of enteric salmonella but despite having high specificity, it has low sensitivity (<50%) and is time-consuming [8]. On the other hand, stool PCR is highly sensitive, less dependent on the quality of the specimen, and a quicker method of detection but comes at the cost of lower specificity compared to stool culture [1,8].…”

“…Another factor is that stool culture is currently considered to be the gold standard for the detection of enteric salmonella but despite having high specificity, it has low sensitivity (<50%) and is time-consuming [8]. On the other hand, stool PCR is highly sensitive, less dependent on the quality of the specimen, and a quicker method of detection but comes at the cost of lower specificity compared to stool culture [1,8]. Our literature search has revealed several case reports of massive gastrointestinal bleed and hematochezia as a complication of invasive Salmonella typhi infections in adults and children [4][5][6].…”

“…Although the specificity of stool culture is almost 100%, all PCR tests also have high specificity and the results come back in a shorter period. However, PCR testing is more costly[8].…”

A Study on the Clinical Profile, Blood Profile and Culture Sensitivity Pattern of Salmonella Typhi in Paediatric Patients

“…Blood was collected in three designated bottles and sent to the hospital's laboratory, where it was processed and examined by a consultant microbiologist, and then culture and sensitivity pattern was determined. 13 Multi-drug-resistant typhoid was defined as if a culture-sensitivity pattern showed resistance to all first-line antibiotics as described in guidelines. 14 All relevant socio-demographic, clinical and laboratory information was entered in a proforma designed for this study.…”