Diagnostic Procedures in Tularaemia with Special Focus on Molecular and Immunological Techniques

Abstract

Tularaemia is a severe bacterial zoonosis caused by the highly infectious agent Francisella tularensis. It is endemic in countries of the northern hemisphere ranging from North America to Europe, Asia and Japan. Very recently, Francisella-like strains causing disease in humans were described from tropical northern Australia. In the last decade, efforts have been made to develop sensitive and specific immunological and molecular techniques for the laboratory diagnosis of tularaemia and also for the definite identification of members of the species F. tularensis and its four subspecies. Screening for the keyword 'Francisella' a Medline search over the last decade was performed and articles describing diagnostic methods for tularaemia and its causative agent were selected. Besides classical microbiological techniques (cultivation, biochemical profiling, susceptibility testing) several new immunological and molecular approaches to identify F. tularensis have been introduced employing highly specific antibodies and various polymerase chain reaction (PCR)-based methods. Whereas direct antigen detection by enzyme-linked immunosorbent assay (ELISA) or immunofluorescence might allow early presumptive diagnosis of tularaemia, these methods--like all PCR techniques--still await further evaluation. Therefore, diagnosis of tularaemia still relies mainly on the demonstration of specific antibodies in the host. ELISA and immunoblot methods started to replace the standard tube or micro-agglutination assays. However, the diagnostic value of antibody detection in the very early clinical phase of tularaemia is limited. Francisella tularensis is regarded as a 'highest priority' biological agent (category 'A' according to the CDC, Atlanta, GA, USA), thus rapid and reliable diagnosis of tularaemia is required not only for a timely onset of therapy, the handling of outbreak investigations but also for the surveillance of endemic foci. Only very recently, evaluated test kits for serological diagnosis of human tularaemia became available, while the introduction of standardized molecular techniques for detection and typing is still missing.

Oropharyngeal tularemia cases admitted to a military hospital in Ankara, Turkey

“…The culture procedures should be conducted in level-3 biosafety laboratory conditions [25,26]. Due to the difficulty of reproduction of F. tularensis and the hazard it bears for laboratory personnel, serologic tests should be preferred in diagnosing the disease [3,10,16,24,26]. MA has been used as the diagnostic test for a long time.…”

Evaluation of an Immunochromatographic Test for Rapid and Reliable Serodiagnosis of Human Tularemia and Detection ofFrancisella tularensis-Specific Antibodies in Sera from Different Mammalian Species

“…Due to the highly infectious nature of F. tularensis, the difficulties caused by the special growth requirements, and the lack of standardized, well-evaluated PCR protocols, the clinical diagnosis of tularemia in humans is most commonly confirmed by serological proof (10,27,29). Definitive serological affirmation requires a 4-fold or greater rise in titers between acute and convalescent-phase sera (33).…”

“…In addition to its application in clinical microbiology, detection of antibodies against F. tularensis is useful to confirm successful vaccination after immunization with live or subunit vaccines and can also be applied for seroepidemiologic studies in endemic regions or populations at risk (24,27). In veterinary medicine, serology is used mainly for tularemia surveillance in rodents, hares, or surrogate animals such as boars or predators, including wolves or bears (1,26,34).…”

Tularemia: current diagnosis and treatment options

“…Another disadvantage is that the cost of the reagents may prohibit their routine usage in clinical microbiology laboratories, particularly in resource-limited environments. A final issue with PCR is that techniques are not standardized between laboratories [54]. …”