title: Two-stage PCR assay for detection of human brucellosis in endemic areas.
abstract: BACKGROUND
  Brucellosis is a common zoonosis that can cause a severe febrile illness in humans. It constitutes a persistent health problem in many developing countries around the world. It is one of the most frequently reported diseases in Saudi Arabia and incidence is particularly high in the Central region, and around the city of Riyadh. The aim of this study was to evaluate a two-stage PCR assay for detection of human brucellosis particularly in endemic areas.
  
  
  METHODS
  A total of 101 serum samples were collected from patients with acute febrile illness (AFI) of unknown cause from two different locations in the Western region of Saudi Arabia. The first location (Northern) is characterized by a nomadic rural population while the second (Central) is a modern urban city. All samples were subjected to DNA extraction and Brucella genus-specific PCR amplification using B4/B5 primers of the bcsp31 gene. Positive B4/B5 samples were subjected to multiplex species-specific Brucella PCR amplification.
  
  
  RESULTS
  In the Northern location, 81.9% of the AFI samples were confirmed Brucella positive, while all the samples collected from the Central region proved to be Brucella negative. Samples positive for Brucella were subjected to multiplex species-specific Brucella amplification. B. abortus was detected in 10% and B. melitensis in 8% of the samples, while the majority (82%) of samples showed both B. abortus and B. melitensis. As expected, B. suis was not detected in any of the samples.
  
  
  CONCLUSIONS
  This study concluded that a two-stage PCR assay could be useful as a rapid diagnostic tool to allow the consideration of brucellosis as a possible cause of AFI, particularly in non-urban locations. It also recommends the collection of epidemiological data for such patients to obtain further information that may help in rapid diagnosis.
authors.affiliation: Biochemistry Department, Faculty of Science, King Abdulaziz University, Jeddah, Saudi Arabia. imbc57@yahoo.com
authors.forename: Ibrahim Hassan
authors.initials: IH
authors.lastname: Kamal
authors.forename: Basim
authors.initials: B
authors.lastname: Al Gashgari
authors.forename: Said Salama
authors.initials: SS
authors.lastname: Moselhy
authors.forename: Taha Abdullah
authors.initials: TA
authors.lastname: Kumosani
authors.forename: Khalid Omar
authors.initials: KO
authors.lastname: Abulnaja
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001431:Bacteriological Techniques / Q000379:methods*; D002002:Brucella / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D002006:Brucellosis / Q000175:diagnosis*; D002648:Child; D002675:Child, Preschool; D005260:Female; D006801:Humans; D007223:Infant; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D060885:Multiplex Polymerase Chain Reaction / Q000379:methods*; D012529:Saudi Arabia; D055815:Young Adult