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Development of genetic system to inactivate a Borrelia turicatae surface protein selectively produced within the salivary glands of the arthropod vector.

Lopez JE, Wilder HK, Hargrove R, Brooks CP, Peterson KE, Beare PA, Sturdevant DE, Nagarajan V, Raffel SJ, Schwan TG - PLoS Negl Trop Dis (2013)

Bottom Line: We then applied a novel genetic system for B. turicatae to inactivate brpA and examined the role of the gene product for vector colonization and the ability to establish murine infection.These results demonstrate the complexity of protein production in a population of spirochetes within the tick.Additionally, the development of a genetic system is important for future studies to evaluate the requirement of specific B. turicatae genes for vector colonization and transmission.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Mississippi State University, Starkville, Mississippi, United States of America.

ABSTRACT

Background: Borrelia turicatae, an agent of tick-borne relapsing fever, is an example of a pathogen that can adapt to disparate conditions found when colonizing the mammalian host and arthropod vector. However, little is known about the genetic factors necessary during the tick-mammalian infectious cycle, therefore we developed a genetic system to transform this species of spirochete. We also identified a plasmid gene that was up-regulated in vitro when B. turicatae was grown in conditions mimicking the tick environment. This 40 kilodalton protein was predicted to be surface localized and designated the Borrelia repeat protein A (brpA) due to the redundancy of the amino acid motif Gln-Gly-Asn-Val-Glu.

Methodology/principal findings: Quantitative reverse-transcriptase polymerase chain reaction using RNA from B. turicatae infected ticks and mice indicated differential regulation of brpA during the tick-mammalian infectious cycle. The surface localization was determined, and production of the protein within the salivary glands of the tick was demonstrated. We then applied a novel genetic system for B. turicatae to inactivate brpA and examined the role of the gene product for vector colonization and the ability to establish murine infection.

Conclusions/significance: These results demonstrate the complexity of protein production in a population of spirochetes within the tick. Additionally, the development of a genetic system is important for future studies to evaluate the requirement of specific B. turicatae genes for vector colonization and transmission.

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Related in: MedlinePlus

IF-LSCM of spirochetes cultivated at 22°C.Chicken and rabbit serum generated against rFlaB (A) and rBrpA (B), respectively, were used to determine protein production within a population of spirochetes. Images were overlaid and spirochetes producing BrpA were counted (C). Chicken (D) and rabbit (E) preimmunization serum samples were used as negative controls. The secondary antibodies used were Alexa Fluor anti-chicken IgY 568 and anti-rabbit IgG 488. 5 µm bars are shown in each panel.
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pntd-0002514-g003: IF-LSCM of spirochetes cultivated at 22°C.Chicken and rabbit serum generated against rFlaB (A) and rBrpA (B), respectively, were used to determine protein production within a population of spirochetes. Images were overlaid and spirochetes producing BrpA were counted (C). Chicken (D) and rabbit (E) preimmunization serum samples were used as negative controls. The secondary antibodies used were Alexa Fluor anti-chicken IgY 568 and anti-rabbit IgG 488. 5 µm bars are shown in each panel.

Mentions: Performing IF-LSCM using rabbit and chicken serum generated against rBrpA and rFlaB, respectively, demonstrated that BrpA was present in a portion (24.6%) of the spirochete population during cultivation at 22°C (Figure 3 A–C). Rabbit and chicken pre-immunization serum samples were not reactive to BrpA and FlaB, respectively (Figure 3 D and E). Collectively, these results further suggested that BrpA is a tick-associated surface protein.


Development of genetic system to inactivate a Borrelia turicatae surface protein selectively produced within the salivary glands of the arthropod vector.

Lopez JE, Wilder HK, Hargrove R, Brooks CP, Peterson KE, Beare PA, Sturdevant DE, Nagarajan V, Raffel SJ, Schwan TG - PLoS Negl Trop Dis (2013)

IF-LSCM of spirochetes cultivated at 22°C.Chicken and rabbit serum generated against rFlaB (A) and rBrpA (B), respectively, were used to determine protein production within a population of spirochetes. Images were overlaid and spirochetes producing BrpA were counted (C). Chicken (D) and rabbit (E) preimmunization serum samples were used as negative controls. The secondary antibodies used were Alexa Fluor anti-chicken IgY 568 and anti-rabbit IgG 488. 5 µm bars are shown in each panel.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3814808&req=5

pntd-0002514-g003: IF-LSCM of spirochetes cultivated at 22°C.Chicken and rabbit serum generated against rFlaB (A) and rBrpA (B), respectively, were used to determine protein production within a population of spirochetes. Images were overlaid and spirochetes producing BrpA were counted (C). Chicken (D) and rabbit (E) preimmunization serum samples were used as negative controls. The secondary antibodies used were Alexa Fluor anti-chicken IgY 568 and anti-rabbit IgG 488. 5 µm bars are shown in each panel.
Mentions: Performing IF-LSCM using rabbit and chicken serum generated against rBrpA and rFlaB, respectively, demonstrated that BrpA was present in a portion (24.6%) of the spirochete population during cultivation at 22°C (Figure 3 A–C). Rabbit and chicken pre-immunization serum samples were not reactive to BrpA and FlaB, respectively (Figure 3 D and E). Collectively, these results further suggested that BrpA is a tick-associated surface protein.

Bottom Line: We then applied a novel genetic system for B. turicatae to inactivate brpA and examined the role of the gene product for vector colonization and the ability to establish murine infection.These results demonstrate the complexity of protein production in a population of spirochetes within the tick.Additionally, the development of a genetic system is important for future studies to evaluate the requirement of specific B. turicatae genes for vector colonization and transmission.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Mississippi State University, Starkville, Mississippi, United States of America.

ABSTRACT

Background: Borrelia turicatae, an agent of tick-borne relapsing fever, is an example of a pathogen that can adapt to disparate conditions found when colonizing the mammalian host and arthropod vector. However, little is known about the genetic factors necessary during the tick-mammalian infectious cycle, therefore we developed a genetic system to transform this species of spirochete. We also identified a plasmid gene that was up-regulated in vitro when B. turicatae was grown in conditions mimicking the tick environment. This 40 kilodalton protein was predicted to be surface localized and designated the Borrelia repeat protein A (brpA) due to the redundancy of the amino acid motif Gln-Gly-Asn-Val-Glu.

Methodology/principal findings: Quantitative reverse-transcriptase polymerase chain reaction using RNA from B. turicatae infected ticks and mice indicated differential regulation of brpA during the tick-mammalian infectious cycle. The surface localization was determined, and production of the protein within the salivary glands of the tick was demonstrated. We then applied a novel genetic system for B. turicatae to inactivate brpA and examined the role of the gene product for vector colonization and the ability to establish murine infection.

Conclusions/significance: These results demonstrate the complexity of protein production in a population of spirochetes within the tick. Additionally, the development of a genetic system is important for future studies to evaluate the requirement of specific B. turicatae genes for vector colonization and transmission.

Show MeSH
Related in: MedlinePlus