Diagnostic utility of in‐house loop‐mediated isothermal amplification and real‐timePCRtargetingvirBgene for direct detection ofBrucella melitensisfrom clinical specimens

Abstract

Aims In this present study, the utility of a newly developed loop‐mediated isothermal amplification (LAMP) and real‐time PCR assays designed to amplify the virB gene region of Brucella melitensis was evaluated from human clinical specimens. Methods and Results Fifty‐four culture‐confirmed cases of brucellosis and 54 culture negative but clinically suspected cases of brucellosis were included in the study. Whole blood, serum and other nonblood specimens were collected and subjected to blood culture using automatic blood culture system, serological tests, LAMP assay and real‐time PCR. Overall sensitivities of LAMP and real‐time PCR assays were 67·5 and 68·3% respectively. For nonblood clinical specimens, we noticed a marked increase in the sensitivities of LAMP (88·9%) and real‐time PCR (100%) assays. Conclusions Performance of LAMP and real‐time PCR was not satisfactory for whole‐blood specimens because of the low abundance of bacteria or DNA. On the other hand, using nonblood specimens, both the assays showed higher sensitivity and specificity which makes them a good alternative for the rapid diagnosis of human brucellosis. Significance and Impact of the Study The developed LAMP and real‐time PCR assays are a specific and rapid diagnostic tool for direct and early detection of Brucella in clinical specimens.

Importance of brucellosis control programs of livestock on the improvement of one health

“…in milk and consumption of non-pasteurized dairy products pose an increasing risk to human consumers. Furthermore, it has been reported that the developed loop mediated isothermal amplification (LAMP) is a rapid and specific diagnostic tool for early and direct identification of Brucella in clinical specimens (Patra et al 2019). In addition, several methods for the optimization of DNA extraction from milk to improve subsequent PCR analysis have been set up leading to the reliable detection of Brucella spp.…”

Bovine brucellosis – a comprehensive review

“…Loop-mediated isothermal amplification (LAMP) of DNA as well as real-time PCR have been proved as significant, sensitive, quick and specific diagnostics for B. abortus and other Brucella spp. directly from clinical specimens (Karthik et al 2014b(Karthik et al , 2014c(Karthik et al , 2016Patra et al 2019). Real-time recombinase polymerase amplification (RPA) was developed targeting the bcsp31 gene and the sensitivity was found to be 94% (Qinet al 2019).…”

Development of a Smartphone-Based Nanozyme-Linked Immunosorbent Assay for Quantitative Detection of SARS-CoV-2 Nucleocapsid Phosphoprotein in Blood

“…If the diagnosis is based solely on viral nucleic acid detection, the workload will be too heavy as there are a large number of suspected cases for testing. Due to uncontrolled factors during sample collection or storage, a false negative or positive results may occur ( Tanner et al, 2015 ; Lim et al, 2018 ; Kim et al, 2019 ; Patra et al, 2019 ; Corman et al, 2020 ; Zhang et al, 2020 ).…”