Towards Development of Improved Serodiagnostics for Tularemia by Use of Francisella tularensis Proteome Microarrays

Abstract

e Tularemia in humans is caused mainly by two subspecies of the Gram-negative facultative anaerobe Francisella tularensis: F. tularensis subsp. tularensis (type A) and F. tularensis subsp. holarctica (type B). The current serological test for tularemia is based on agglutination of whole organisms, and the reactive antigens are not well understood. Previously, we profiled the antibody responses in type A and B tularemia cases in the United States using a proteome microarray of 1,741 different proteins derived from the type A strain Schu S4. Fifteen dominant antigens able to detect antibodies to both types of infection were identified, although these were not validated in a different immunoassay format. Since type A and B subspecies are closely related, we hypothesized that Schu S4 antigens would also have utility for diagnosing type B tularemia caused by strains from other geographic locations. To test this, we probed the Schu S4 array with sera from 241 type B tularemia cases in Spain. Despite there being no type A strains in Spain, we confirmed the responses against some of the same potential serodiagnostic antigens reported previously, as well as determined the responses against additional potential serodiagnostic antigens. Five potential serodiagnostic antigens were evaluated on immunostrips, and two of these (FTT1696/GroEL and FTT0975/conserved hypothetical protein) discriminated between the Spanish tularemia cases and healthy controls. We conclude that antigens from the type A strain Schu S4 are suitable for detection of antibodies from patients with type B F. tularensis infections and that these can be used for the diagnosis of tularemia in a deployable format, such as the immunostrip.T ularemia is a zoonotic disease caused by the Gram-negative facultative anaerobe Francisella tularensis. The organism infects a multitude of different animals, including mammals, birds, fish, and insects. Transmission to humans can occur by inhalation, although the most common natural route is likely to be direct contact with infected animals or via ticks and other biting arthropods that feed on infected mammals. Important animal reservoirs appear to be rodents, hares, and rabbits. For example, the 1997 outbreak in Castilla y León, Spain (1, 2), was transmitted by the handling of infected hares. The following year, another outbreak which was attributed to crayfish handling occurred in Castilla-La Mancha, Spain (3). Human-tohuman transmission is rare.Four closely related subspecies have been defined: F. tularensis subsp. tularensis (type A), F. tularensis subsp. holarctica (type B), F. tularensis subsp. novicida, and F. tularensis subsp. mediasiatica. Each subspecies possesses a different pathogenic potential in humans, as well as a distinctive geographic distribution, host preference, and route of transmission. Type A is found mainly in North America and is associated with severe tularemia that may be fatal, whereas type B is found throughout the Northern Hemisphere and is less virulent in humans (4-6). F. tularensis subsp. ...

Transient Cannabinoid Receptor 2 Blockade during Immunization Heightens Intensity and Breadth of Antigen-specific Antibody Responses in Young and Aged mice

“…Immunogenic protein antigens FTT1269, FTT1329, FTT1483 ( Francisella tularensis ), WR148, WR101 ( Vaccinia virus ), BB_B19, BB_O34 ( Borrelia burgdorferi ), Rv3362 ( Mycobacterium tuberculosis ), and OTB1104 ( Orientia tsutsugamushi ) were purified as previously described 26 . Briefly, Protein-encoding T7-expression plasmids were used for protein expression in E. coli cells in vivo .…”

Francisella tularensis, Tularemia and Serological Diagnosis

“…The immunoblotting technique has also been used to determine the F. tularensis immunoreactive proteins using sera from infected or vaccinated persons (Havlasováet al, 2002;Eyles et al, 2007;Sundaresh et al, 2007;Nakajima et al, 2016). These studies have shown that the most immunoreactive F. tularensis included the chaperone proteins dnaK and groEL, two chaperonins (10 and 60 kDa), the elongation factor EF-Ts, membrane proteins (23 kDa hypothetical protein, 17 kDa TUL4 protein, OmpH and FopA), enzymes (pyruvate dehydrogenase, acetyl-CoA carboxylase, pyrrolidone-carboxylate peptidase, isocitrate dehydrogenase, glutamine synthetase, etc.)…”

“…This difference may reflect the higher specificity of the immunoblot technique but also low sensitivity of this method in patients with low antibody titers. Several studies have tried to dissect the immune response of patients infected with F. tularensis to develop more sensitive and specific serological tests (Havlasovaé t al., 2002;Eyles et al, 2007;Sundaresh et al, 2007;Nakajima et al, 2016). Although these studies have well defined the most immunodominant F. tularensis antigens, they have not given rise to the development of new serological tests with improved performances compared to those currently used for tularemia diagnosis.…”

Desarrollo y utilidad de las técnicas de ELISA y quimioluminiscencia para el diagnóstico de la tularemia humana

“…El estudio e identificación de los diferentes antígenos de membrana permitirá confirmar cuales son los antígenos inmunodominantes de Francisella tularensis y su importancia inmunopatogénica y protectora 3,24,25 .…”

“…En lo que respecta a las reacciones cruzadas con otras enfermedades, los estudios llevados a cabo en Castilla y León no encontraron relación con Yersinia enterocolitica ni con Proteus OX19 155,176 , aunque otros autores 25,177 han descrito casos de reacciones cruzadas con los antígenos de estas bacterias.…”