Molecular Diagnostic Methods of Brucellosis: a Note on Pitfalls

Document Type: Letters to the Editor

Authors

Dept. of Molecular Biology, Research Center of Health Reference Laboratory, Ministry of Health and Medical Education, Tehran, Iran

10.30699/ijp.13.2.294

Abstract

Clinical manifestations of the brucellosis may show extensive appearance because of its wide signs and symptoms. Patients with brucellosis are usually symptomatically treated by using different antibiotics at private clinics due to misdiagnosis of clinical laboratory findings specially those chronic forms. These subjects will have been hospitalized with a complicated form of signs after changing a few therapies (1).
At this time, classical methods such as culture based and serological procedures could not alone meet of clinicians’ demands. Many researchers have been being tried to introduce an alternative method better in accuracy, reliability and efficiency aspects. However, most reported comparison studies were not able to present desired results. There are not any significant points in existence of various and even contradictory results in laboratory interpretation reports. Those patients with recent brucellosis involvement or acute disease are better diagnosed than those hospitalized or chronic cases. Unfortunately, it is being observed the most researchers are not attending the patient clinical infectious form. Obviously, in-patients have more problematical conditions that results to have lower sensitivity rate in some test or even different from each other. Unfortunately, this concern has not been noticed in the majority of molecular epidemiology studies. Analyzed results of various tests are not fully in agreement with each other in these reports, confirming each test is better suited for some specific clinical conditions (2).
Another important problem is applying home brew protocols in all comparative studies. This type of  non-commercial diagnostic protocol are being surprisingly applied in most small Iranian clinical laboratories. These assays have been being followed from some reported papers without any proper optimization experiments or exact clinical trial specifically for Iranian populations, particularly when several of the effective parameters in the optimization may not be well reflected in these released reports. Some of them for proper optimization and standardization are mentioned in other studies (3). 
Proper selecting sampling is additional problems. Serum specimens would not be preferred samples since Brucella spp. agents are intracellular which is integrated in PMN cells. Serum specimens may give positive results, but in those has high antibody titer or acute cases. Although it is used as applied specimens instead of peripheral anti-coagulated blood (4). The chance of the recovery Brucella spp. DNA is obviously reduced in those chronic and complicated subjects. In addition, sampling from patients undergoing treatment may have low positive predictive value results (5,6). It would seem international approved extraction method accompanied with internal control as control the DNA isolation procedure and to check for possible PCR inhibition should be used in molecular diagnostic settings (4,7).
Finally the last point, various pathogenic Brucella species involve in many countries. At present, we have no significant documentary reports in frequency rate of Brucella spp.  in our community. Some reports represent considerable rate for B. abortus while some not. Therefore, applied protocol must be enough sensitive to detect and even differentiate at least B. abortus from  B. melitensis, although some reports illustrate the presence of B. canis as well as in Iran (8).

Molecular Diagnostic Methods of Brucellosis: a Note on Pitfalls

 

Clinical manifestations of the brucellosis may show extensive appearance because of its wide signs and symptoms. Patients with brucellosis are usually symptomatically treated by using different antibiotics at private clinics due to misdiagnosis of clinical laboratory findings specially those chronic forms. These subjects will have been hospitalized with a complicated form of signs after changing a few therapies (1).

At this time, classical methods such as culture based and serological procedures could not alone meet of clinicians’ demands. Many researchers have been being tried to introduce an alternative method better in accuracy, reliability and efficiency aspects. However, most reported comparison studies were not able to present desired results. There are not any significant points in existence of various and even contradictory results in laboratory interpretation reports. Those patients with recent brucellosis involvement or acute disease are better diagnosed than those hospitalized or chronic cases. Unfortunately, it is being observed the most researchers are not attending the patient clinical infectious form. Obviously, in-patients have more problematical conditions that results to have lower sensitivity rate in some test or even different from each other. Unfortunately, this concern has not been noticed in the majority of molecular epidemiology studies. Analyzed results of various tests are not fully in agreement with each other in these reports, confirming each test is better suited for some specific clinical conditions (2).

Another important problem is applying home brew protocols in all comparative studies. This type of  non-commercial diagnostic protocol are being surprisingly applied in most small Iranian clinical laboratories. These assays have been being followed from some reported papers without any proper optimization experiments or exact clinical trial specifically for Iranian populations, particularly when several of the effective parameters in the optimization may not be well reflected in these released reports. Some of them for proper optimization and standardization are mentioned in other studies (3). 

Proper selecting sampling is additional problems. Serum specimens would not be preferred samples since Brucella spp. agents are intracellular which is integrated in PMN cells. Serum specimens may give positive results, but in those has high antibody titer or acute cases. Although it is used as applied specimens instead of peripheral anti-coagulated blood (4). The chance of the recovery Brucella spp. DNA is obviously reduced in those chronic and complicated subjects. In addition, sampling from patients undergoing treatment may have low positive predictive value results (5,6). It would seem international approved extraction method accompanied with internal control as control the DNA isolation procedure and to check for possible PCR inhibition should be used in molecular diagnostic settings (4,7).

Finally the last point, various pathogenic Brucella species involve in many countries. At present, we have no significant documentary reports in frequency rate of Brucella spp.  in our community. Some reports represent considerable rate for B. abortus while some not. Therefore, applied protocol must be enough sensitive to detect and even differentiate at least B. abortus from  B. melitensis, although some reports illustrate the presence of B. canis as well as in Iran (8).

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