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Molecular interactions that enable movement of the Lyme disease agent from the tick gut into the hemolymph.

Zhang L, Zhang Y, Adusumilli S, Liu L, Narasimhan S, Dai J, Zhao YO, Fikrig E - PLoS Pathog. (2011)

Bottom Line: The decreased hemolymph infection results in lower salivary glands infection, and consequently attenuates mouse infection by tick-transmitted B. burgdorferi.Silencing tre31 also decreased the B. burgdorferi burden in the tick hemolymph.Delineating the specific spirochete and arthropod ligands required for B. burgdorferi movement in the tick may lead to new strategies to interrupt the life cycle of the Lyme disease agent.

View Article: PubMed Central - PubMed

Affiliation: Section of Infectious Diseases, Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut, United States of America.

ABSTRACT
Borrelia burgdorferi, the causative agent of Lyme disease, is transmitted to humans by bite of Ixodes scapularis ticks. The mechanisms by which the bacterium is transmitted from vector to host are poorly understood. In this study, we show that the F(ab)(2) fragments of BBE31, a B.burgdorferi outer-surface lipoprotein, interfere with the migration of the spirochete from tick gut into the hemolymph during tick feeding. The decreased hemolymph infection results in lower salivary glands infection, and consequently attenuates mouse infection by tick-transmitted B. burgdorferi. Using a yeast surface display approach, a tick gut protein named TRE31 was identified to interact with BBE31. Silencing tre31 also decreased the B. burgdorferi burden in the tick hemolymph. Delineating the specific spirochete and arthropod ligands required for B. burgdorferi movement in the tick may lead to new strategies to interrupt the life cycle of the Lyme disease agent.

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Related in: MedlinePlus

F(ab)2 fragments of anti-BBE31 IgG interfere with B.burgdorferi migration from I.scapularis gut into hemolymph.(A) Borreliacidal activity of BBE31 F(ab)2 fragments against B. burgdorferi. B.burgdorferi N40 were incubated in BSK medium in the absence (Control) or presence of F(ab)2 fragments prepared from normal rabbit serum [NRS F(ab)2] or BBE31 antiserum [BBE31 F(ab)2]. A borreliacidal serum from a patient with diagnosed Lyme Disease (B.burgdorferi antisera) was used as a positive control. The number of spirochetes was assessed by dark-field microscopy after 24 and 48 hours of incubation. Data are presented relative to controls without treatment. Data represent the number of spirochetes remaining viable after treatment (mean ± SD, n = 4). Difference between control and NRS F(ab)2- as well as BBE31 F(ab)2-treated samples were not statistically significant. (B) B.burgdorferi burden in I.scapularis hemolymph assayed by q-PCR. N40-infected nymphs fed on C3H mice, which had been injected with F(ab)2 fragments of either normal rabbit IgG (NRS-F(ab)2) or anti-BBE31 IgG (BBE31-F(ab)2), for 66 hours. DNA was extracted from the hemolymph for q-PCR analysis. Each dot represents hemolymph from 5 ticks. Data were collected from 2 times of tick-feedings. *: p<0.05. (C) B.burgdorferi burden in I.scapularis hemolymph assayed by confocal microscope. Tick feeding is the same as that described in part (A). The spirochetes in tick hemolymph were probed with FITC-labeled goat anti-borrelia antibodies (shown in green), the hemocyte nucleolus were stained with TO-PRO-3 (shown in blue). The FITC and TO-PRO-3 images were examined at ×25 magnifications and are presented as merged images. Scale bar represents 20 µm. (D) Spirochetes lacking bbe31 can survive in tick hemolymph and invade the salivary glands. B.burgdorferi wild-type strain MSK5 and the lp25-lacking isolate ML23 were grown in BSK medium, resuspended in PBS and microinjected into partially-fed clean nymphal hemocoel (clean nymphs fed on C3H mice for 48 hours before microinjection). Six hours later, both hemolymph (HL) and salivary glands (SG) were collected for B.burgdorferi quantification by q-PCR. Mean and SD were calculated from 5 mRNA samples and 5 ticks were grouped for 1 mRNA sample.
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ppat-1002079-g005: F(ab)2 fragments of anti-BBE31 IgG interfere with B.burgdorferi migration from I.scapularis gut into hemolymph.(A) Borreliacidal activity of BBE31 F(ab)2 fragments against B. burgdorferi. B.burgdorferi N40 were incubated in BSK medium in the absence (Control) or presence of F(ab)2 fragments prepared from normal rabbit serum [NRS F(ab)2] or BBE31 antiserum [BBE31 F(ab)2]. A borreliacidal serum from a patient with diagnosed Lyme Disease (B.burgdorferi antisera) was used as a positive control. The number of spirochetes was assessed by dark-field microscopy after 24 and 48 hours of incubation. Data are presented relative to controls without treatment. Data represent the number of spirochetes remaining viable after treatment (mean ± SD, n = 4). Difference between control and NRS F(ab)2- as well as BBE31 F(ab)2-treated samples were not statistically significant. (B) B.burgdorferi burden in I.scapularis hemolymph assayed by q-PCR. N40-infected nymphs fed on C3H mice, which had been injected with F(ab)2 fragments of either normal rabbit IgG (NRS-F(ab)2) or anti-BBE31 IgG (BBE31-F(ab)2), for 66 hours. DNA was extracted from the hemolymph for q-PCR analysis. Each dot represents hemolymph from 5 ticks. Data were collected from 2 times of tick-feedings. *: p<0.05. (C) B.burgdorferi burden in I.scapularis hemolymph assayed by confocal microscope. Tick feeding is the same as that described in part (A). The spirochetes in tick hemolymph were probed with FITC-labeled goat anti-borrelia antibodies (shown in green), the hemocyte nucleolus were stained with TO-PRO-3 (shown in blue). The FITC and TO-PRO-3 images were examined at ×25 magnifications and are presented as merged images. Scale bar represents 20 µm. (D) Spirochetes lacking bbe31 can survive in tick hemolymph and invade the salivary glands. B.burgdorferi wild-type strain MSK5 and the lp25-lacking isolate ML23 were grown in BSK medium, resuspended in PBS and microinjected into partially-fed clean nymphal hemocoel (clean nymphs fed on C3H mice for 48 hours before microinjection). Six hours later, both hemolymph (HL) and salivary glands (SG) were collected for B.burgdorferi quantification by q-PCR. Mean and SD were calculated from 5 mRNA samples and 5 ticks were grouped for 1 mRNA sample.

Mentions: We first tested whether BBE31 IgG F(ab)2 fragments influence borrelial burden in the tick hemolymph during tick feeding. N40-infected nymphs were allowed to feed on selected F(ab)2 fragments-treated mice for 66 hours. Hemolymph was collected as described in experimental procedures and the spirochete burden was measured by both q-PCR and confocal microscopy. When compared to ticks fed on mice treated with the normal rabbit IgG F(ab)2 fragments, ticks fed on BBE31 F(ab)2 fragments-treated mice showed a significantly lower spirochete burden in the hemolymph (Figure 5B & 5C). When we analyzed individual ticks, we found that the low hemolymph burden always correlated with decreased infection of salivary glands, suggesting that BBE31 antibodies decrease spirochete burden in tick salivary glands by reducing the spirochetes burden in the hemolymph and not by interfering with the spirochetes invasion of tick salivary glands.


Molecular interactions that enable movement of the Lyme disease agent from the tick gut into the hemolymph.

Zhang L, Zhang Y, Adusumilli S, Liu L, Narasimhan S, Dai J, Zhao YO, Fikrig E - PLoS Pathog. (2011)

F(ab)2 fragments of anti-BBE31 IgG interfere with B.burgdorferi migration from I.scapularis gut into hemolymph.(A) Borreliacidal activity of BBE31 F(ab)2 fragments against B. burgdorferi. B.burgdorferi N40 were incubated in BSK medium in the absence (Control) or presence of F(ab)2 fragments prepared from normal rabbit serum [NRS F(ab)2] or BBE31 antiserum [BBE31 F(ab)2]. A borreliacidal serum from a patient with diagnosed Lyme Disease (B.burgdorferi antisera) was used as a positive control. The number of spirochetes was assessed by dark-field microscopy after 24 and 48 hours of incubation. Data are presented relative to controls without treatment. Data represent the number of spirochetes remaining viable after treatment (mean ± SD, n = 4). Difference between control and NRS F(ab)2- as well as BBE31 F(ab)2-treated samples were not statistically significant. (B) B.burgdorferi burden in I.scapularis hemolymph assayed by q-PCR. N40-infected nymphs fed on C3H mice, which had been injected with F(ab)2 fragments of either normal rabbit IgG (NRS-F(ab)2) or anti-BBE31 IgG (BBE31-F(ab)2), for 66 hours. DNA was extracted from the hemolymph for q-PCR analysis. Each dot represents hemolymph from 5 ticks. Data were collected from 2 times of tick-feedings. *: p<0.05. (C) B.burgdorferi burden in I.scapularis hemolymph assayed by confocal microscope. Tick feeding is the same as that described in part (A). The spirochetes in tick hemolymph were probed with FITC-labeled goat anti-borrelia antibodies (shown in green), the hemocyte nucleolus were stained with TO-PRO-3 (shown in blue). The FITC and TO-PRO-3 images were examined at ×25 magnifications and are presented as merged images. Scale bar represents 20 µm. (D) Spirochetes lacking bbe31 can survive in tick hemolymph and invade the salivary glands. B.burgdorferi wild-type strain MSK5 and the lp25-lacking isolate ML23 were grown in BSK medium, resuspended in PBS and microinjected into partially-fed clean nymphal hemocoel (clean nymphs fed on C3H mice for 48 hours before microinjection). Six hours later, both hemolymph (HL) and salivary glands (SG) were collected for B.burgdorferi quantification by q-PCR. Mean and SD were calculated from 5 mRNA samples and 5 ticks were grouped for 1 mRNA sample.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3111543&req=5

ppat-1002079-g005: F(ab)2 fragments of anti-BBE31 IgG interfere with B.burgdorferi migration from I.scapularis gut into hemolymph.(A) Borreliacidal activity of BBE31 F(ab)2 fragments against B. burgdorferi. B.burgdorferi N40 were incubated in BSK medium in the absence (Control) or presence of F(ab)2 fragments prepared from normal rabbit serum [NRS F(ab)2] or BBE31 antiserum [BBE31 F(ab)2]. A borreliacidal serum from a patient with diagnosed Lyme Disease (B.burgdorferi antisera) was used as a positive control. The number of spirochetes was assessed by dark-field microscopy after 24 and 48 hours of incubation. Data are presented relative to controls without treatment. Data represent the number of spirochetes remaining viable after treatment (mean ± SD, n = 4). Difference between control and NRS F(ab)2- as well as BBE31 F(ab)2-treated samples were not statistically significant. (B) B.burgdorferi burden in I.scapularis hemolymph assayed by q-PCR. N40-infected nymphs fed on C3H mice, which had been injected with F(ab)2 fragments of either normal rabbit IgG (NRS-F(ab)2) or anti-BBE31 IgG (BBE31-F(ab)2), for 66 hours. DNA was extracted from the hemolymph for q-PCR analysis. Each dot represents hemolymph from 5 ticks. Data were collected from 2 times of tick-feedings. *: p<0.05. (C) B.burgdorferi burden in I.scapularis hemolymph assayed by confocal microscope. Tick feeding is the same as that described in part (A). The spirochetes in tick hemolymph were probed with FITC-labeled goat anti-borrelia antibodies (shown in green), the hemocyte nucleolus were stained with TO-PRO-3 (shown in blue). The FITC and TO-PRO-3 images were examined at ×25 magnifications and are presented as merged images. Scale bar represents 20 µm. (D) Spirochetes lacking bbe31 can survive in tick hemolymph and invade the salivary glands. B.burgdorferi wild-type strain MSK5 and the lp25-lacking isolate ML23 were grown in BSK medium, resuspended in PBS and microinjected into partially-fed clean nymphal hemocoel (clean nymphs fed on C3H mice for 48 hours before microinjection). Six hours later, both hemolymph (HL) and salivary glands (SG) were collected for B.burgdorferi quantification by q-PCR. Mean and SD were calculated from 5 mRNA samples and 5 ticks were grouped for 1 mRNA sample.
Mentions: We first tested whether BBE31 IgG F(ab)2 fragments influence borrelial burden in the tick hemolymph during tick feeding. N40-infected nymphs were allowed to feed on selected F(ab)2 fragments-treated mice for 66 hours. Hemolymph was collected as described in experimental procedures and the spirochete burden was measured by both q-PCR and confocal microscopy. When compared to ticks fed on mice treated with the normal rabbit IgG F(ab)2 fragments, ticks fed on BBE31 F(ab)2 fragments-treated mice showed a significantly lower spirochete burden in the hemolymph (Figure 5B & 5C). When we analyzed individual ticks, we found that the low hemolymph burden always correlated with decreased infection of salivary glands, suggesting that BBE31 antibodies decrease spirochete burden in tick salivary glands by reducing the spirochetes burden in the hemolymph and not by interfering with the spirochetes invasion of tick salivary glands.

Bottom Line: The decreased hemolymph infection results in lower salivary glands infection, and consequently attenuates mouse infection by tick-transmitted B. burgdorferi.Silencing tre31 also decreased the B. burgdorferi burden in the tick hemolymph.Delineating the specific spirochete and arthropod ligands required for B. burgdorferi movement in the tick may lead to new strategies to interrupt the life cycle of the Lyme disease agent.

View Article: PubMed Central - PubMed

Affiliation: Section of Infectious Diseases, Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut, United States of America.

ABSTRACT
Borrelia burgdorferi, the causative agent of Lyme disease, is transmitted to humans by bite of Ixodes scapularis ticks. The mechanisms by which the bacterium is transmitted from vector to host are poorly understood. In this study, we show that the F(ab)(2) fragments of BBE31, a B.burgdorferi outer-surface lipoprotein, interfere with the migration of the spirochete from tick gut into the hemolymph during tick feeding. The decreased hemolymph infection results in lower salivary glands infection, and consequently attenuates mouse infection by tick-transmitted B. burgdorferi. Using a yeast surface display approach, a tick gut protein named TRE31 was identified to interact with BBE31. Silencing tre31 also decreased the B. burgdorferi burden in the tick hemolymph. Delineating the specific spirochete and arthropod ligands required for B. burgdorferi movement in the tick may lead to new strategies to interrupt the life cycle of the Lyme disease agent.

Show MeSH
Related in: MedlinePlus