scholarly journals Motility Is Crucial for the Infectious Life Cycle of Borrelia burgdorferi

2013 ◽  
Vol 81 (6) ◽  
pp. 2012-2021 ◽  
Author(s):  
Syed Z. Sultan ◽  
Akarsh Manne ◽  
Philip E. Stewart ◽  
Aaron Bestor ◽  
Patricia A. Rosa ◽  
...  
Keyword(s):  
Life Cycle ◽  
Borrelia Burgdorferi ◽  
Mammalian Host ◽  
Major Protein ◽  
Content Type ◽  
Absolute Requirement ◽  
Wave Morphology ◽  
Minor Protein ◽  
Periplasmic Flagella ◽  
First Time

ABSTRACTThe Lyme disease spirochete,Borrelia burgdorferi, exists in a zoonotic cycle involving an arthropod tick and mammalian host. Dissemination of the organism within and between these hosts depends upon the spirochete's ability to traverse through complex tissues. Additionally, the spirochete outruns the host immune cells while migrating through the dermis, suggesting the importance ofB. burgdorferimotility in evading host clearance.B. burgdorferi's periplasmic flagellar filaments are composed primarily of a major protein, FlaB, and minor protein, FlaA. By constructing aflaBmutant that is nonmotile, we investigated for the first time the absolute requirement for motility in the mouse-tick life cycle ofB. burgdorferi. We found that whereas wild-type cells are motile and have a flat-wave morphology, mutant cells were nonmotile and rod shaped. These mutants were unable to establish infection in C3H/HeN mice via either needle injection or tick bite. In addition, these mutants had decreased viability in fed ticks. Our studies provide substantial evidence that the periplasmic flagella, and consequently motility, are critical not only for optimal survival in ticks but also for infection of the mammalian host by the arthropod tick vector.

scholarly journals Motor Rotation Is Essential for the Formation of the Periplasmic Flagellar Ribbon, Cellular Morphology, and Borrelia burgdorferi Persistence within Ixodes scapularis Tick and Murine Hosts

2015 ◽  
Vol 83 (5) ◽  
pp. 1765-1777 ◽  
Author(s):  
Syed Z. Sultan ◽  
Padmapriya Sekar ◽  
Xiaowei Zhao ◽  
Akarsh Manne ◽  
Jun Liu ◽  
...  
Keyword(s):  
Borrelia Burgdorferi ◽  
Ixodes Scapularis ◽  
Mutant Cell ◽  
Cellular Morphology ◽  
Flagellar Motor ◽  
Tick Feeding ◽  
Content Type ◽  
Wave Morphology ◽  
Periplasmic Flagella ◽  
Mutant Cells

Borrelia burgdorferimust migrate within and between its arthropod and mammalian hosts in order to complete its natural enzootic cycle. During tick feeding, the spirochete transmits from the tick to the host dermis, eventually colonizing and persisting within multiple, distant tissues. This dissemination modality suggests that flagellar motor rotation and, by extension, motility are crucial for infection. We recently reported that a nonmotileflaBmutant that lacks periplasmic flagella is rod shaped and unable to infect mice by needle or tick bite. However, those studies could not differentiate whether motor rotation or merely the possession of the periplasmic flagella was crucial for cellular morphology and host persistence. Here, we constructed and characterized amotBmutant that is nonmotile but retains its periplasmic flagella. Even though ΔmotBbacteria assembled flagella, part of the mutant cell is rod shaped. Cryoelectron tomography revealed that the flagellar ribbons are distorted in the mutant cells, indicating that motor rotation is essential for spirochetal flat-wave morphology. The ΔmotBcells are unable to infect mice, survive in the vector, or migrate out of the tick. Coinfection studies determined that the presence of these nonmotile ΔmotBcells has no effect on the clearance of wild-type spirochetes during murine infection and vice versa. Together, our data demonstrate that while flagellar motor rotation is necessary for spirochetal morphology and motility, the periplasmic flagella display no additional properties related to immune clearance and persistence within relevant hosts.

scholarly journals The Decrease in FlaA Observed in a flaB Mutant of Borrelia burgdorferi Occurs Posttranscriptionally

2004 ◽  
Vol 186 (12) ◽  
pp. 3703-3711 ◽  
Author(s):  
M. A. Motaleb ◽  
Melanie S. Sal ◽  
Nyles W. Charon
Keyword(s):  
Borrelia Burgdorferi ◽  
Translational Control ◽  
Pcr Analysis ◽  
Insertion Mutation ◽  
Brachyspira Hyodysenteriae ◽  
Wave Morphology ◽  
Minor Protein ◽  
A Minor ◽  
Filament Protein ◽  
Periplasmic Flagella

ABSTRACT The Lyme disease bacterium Borrelia burgdorferi is a motile spirochete with a flat-wave morphology. The periplasmic flagella, which are situated between the outer membrane sheath and cell cylinder, are essential for both the cell's wavy shape and motility. Here we focus on the structure and regulation of its periplasmic flagella. Previous studies have suggested that the periplasmic flagella consist of a polymer of the major filament protein FlaB and a minor protein, FlaA. We used immunoprecipitation methodology to present further evidence that FlaA is indeed a flagellar protein. In addition, in contrast to FlaA of the spirochete Brachyspira hyodysenteriae, B. burgdorferi FlaA did not impact the overall helical shape of the periplasmic flagella. We have previously shown that B. burgdorferi lacks the sigma factor-dependent cascade control of motility gene transcription found in other bacteria. To begin to understand motility gene regulation in B. burgdorferi, we examined the effects of an insertion mutation in flaB on the amounts of proteins encoded by motility genes. Of several motility gene-encoded proteins examined, only the amount of FlaA was decreased in the flaB mutant; it was 13% compared to the wild-type amount. Real-time reverse transcriptase PCR analysis indicated that this inhibition was not the result of a decrease in flaA mRNA. In addition, protein stability analysis suggested that FlaA was turned over in the flaB mutant. Our results indicate that the lack of FlaB negatively influences the amount of FlaA found in the cell and that this effect is at the level of either translational control or protein turnover.

scholarly journals Borrelia burgdorferi Lacking BBK32, a Fibronectin-Binding Protein, Retains Full Pathogenicity

2006 ◽  
Vol 74 (6) ◽  
pp. 3305-3313 ◽  
Author(s):  
Xin Li ◽  
Xianzhong Liu ◽  
Deborah S. Beck ◽  
Fred S. Kantor ◽  
Erol Fikrig
Keyword(s):  
Life Cycle ◽  
Borrelia Burgdorferi ◽  
Deletion Mutant ◽  
Binding Protein ◽  
Kanamycin Resistance ◽  
Binding Activity ◽  
Mammalian Host ◽  
Wild Type ◽  
Fibronectin Binding Protein ◽  
Fibronectin Binding

ABSTRACT BBK32, a fibronectin-binding protein of Borrelia burgdorferi, is one of many surface lipoproteins that are differentially expressed by the Lyme disease spirochete at various stages of its life cycle. The level of BBK32 expression in B. burgdorferi is highest during infection of the mammalian host and lowest in flat ticks. This temporal expression profile, along with its fibronectin-binding activity, strongly suggests that BBK32 may play an important role in Lyme pathogenesis in the host. To test this hypothesis, we constructed an isogenic BBK32 deletion mutant from wild-type B. burgdorferi B31 by replacing the BBK32 gene with a kanamycin resistance cassette through homologous recombination. We examined both the wild-type strain and the BBK32 deletion mutant extensively in the experimental mouse-tick model of the Borrelia life cycle. Our data indicated that B. burgdorferi lacking BBK32 retained full pathogenicity in mice, regardless of whether mice were infected artificially by syringe inoculation or naturally by tick bite. The loss of BBK32 expression in the mutant had no adverse effect on spirochete acquisition (mouse-to-tick) and transmission (tick-to-mouse) processes. These results suggest that additional B. burgdorferi proteins can complement the function of BBK32, fibronectin binding or otherwise, during the natural spirochete life cycle.

scholarly journals An Ixodes scapularis Protein Disulfide Isomerase Contributes to Borrelia burgdorferi Colonization of the Vector

2020 ◽  
Vol 88 (12) ◽  
Author(s):  
Yongguo Cao ◽  
Connor Rosen ◽  
Gunjan Arora ◽  
Akash Gupta ◽  
Carmen J. Booth ◽  
...  
Keyword(s):  
Life Cycle ◽  
Borrelia Burgdorferi ◽  
Protein Disulfide Isomerase ◽  
Inflammatory Responses ◽  
Ixodes Scapularis ◽  
Vertebrate Host ◽  
Content Type ◽  
Disulfide Isomerase ◽  
Complex Interactions ◽  
Protein Disulfide

ABSTRACT Borrelia burgdorferi causes Lyme disease, the most common tick-transmitted illness in North America. When Ixodes scapularis feed on an infected vertebrate host, spirochetes enter the tick gut along with the bloodmeal and colonize the vector. Here, we show that a secreted tick protein, I. scapularis protein disulfide isomerase A3 (IsPDIA3), enhances B. burgdorferi colonization of the tick gut. I. scapularis ticks in which ispdiA3 has been knocked down using RNA interference have decreased spirochete colonization of the tick gut after engorging on B. burgdorferi-infected mice. Moreover, administration of IsPDIA3 antiserum to B. burgdorferi-infected mice reduced the ability of spirochetes to colonize the tick when feeding on these animals. We show that IsPDIA3 modulates inflammatory responses at the tick bite site, potentially facilitating spirochete survival at the vector-host interface as it exits the vertebrate host to enter the tick gut. These data provide functional insights into the complex interactions between B. burgdorferi and its arthropod vector and suggest additional targets to interfere with the spirochete life cycle.

scholarly journals A Novel Tool for the Generation of Conditional Knockouts To Study Gene Function across the Plasmodium falciparum Life Cycle

mBio ◽  
2019 ◽  
Vol 10 (5) ◽  
Author(s):  
Marta Tibúrcio ◽  
Annie S. P. Yang ◽  
Kazuhide Yahata ◽  
Pablo Suárez-Cortés ◽  
Hugo Belda ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Life Cycle ◽  
Genetic Modification ◽  
Gene Deletion ◽  
Genetically Modify ◽  
Complex Life Cycle ◽  
Mosquito Vector ◽  
Parasite Line ◽  
Content Type ◽  
First Time

ABSTRACT Plasmodium falciparum has a complex life cycle that involves interaction with multiple tissues inside the human and mosquito hosts. Identification of essential genes at all different stages of the P. falciparum life cycle is urgently required for clinical development of tools for malaria control and eradication. However, the study of P. falciparum is limited by the inability to genetically modify the parasite throughout its life cycle with the currently available genetic tools. Here, we describe the detailed characterization of a new marker-free P. falciparum parasite line that expresses rapamycin-inducible Cre recombinase across the full life cycle. Using this parasite line, we were able to conditionally delete the essential invasion ligand AMA1 in three different developmental stages for the first time. We further confirm efficient gene deletion by targeting the nonessential kinase FIKK7.1. IMPORTANCE One of the major limitations in studying P. falciparum is that so far only asexual stages are amenable to rapid conditional genetic modification. The most promising drug targets and vaccine candidates, however, have been refractory to genetic modification because they are essential during the blood stage or for transmission in the mosquito vector. This leaves a major gap in our understanding of parasite proteins in most life cycle stages and hinders genetic validation of drug and vaccine targets. Here, we describe a method that supports conditional gene deletion across the P. falciparum life cycle for the first time. We demonstrate its potential by deleting essential and nonessential genes at different parasite stages, which opens up completely new avenues for the study of malaria and drug development. It may also allow the realization of novel vaccination strategies using attenuated parasites.

scholarly journals Analysis of the HD-GYP Domain Cyclic Dimeric GMP Phosphodiesterase Reveals a Role in Motility and the Enzootic Life Cycle of Borrelia burgdorferi

2011 ◽  
Vol 79 (8) ◽  
pp. 3273-3283 ◽  
Author(s):  
Syed Z. Sultan ◽  
Joshua E. Pitzer ◽  
Tristan Boquoi ◽  
Gerry Hobbs ◽  
Michael R. Miller ◽  
...  
Keyword(s):  
Life Cycle ◽  
Borrelia Burgdorferi ◽  
Double Mutant ◽  
Ixodes Scapularis ◽  
Metabolizing Enzymes ◽  
Content Type ◽  
Motility Regulation ◽  
Mutant Cells ◽  
Single Set

ABSTRACTHD-GYP domain cyclic dimeric GMP (c-di-GMP) phosphodiesterases are implicated in motility and virulence in bacteria.Borrelia burgdorferipossesses a single set of c-di-GMP-metabolizing enzymes, including a putative HD-GYP domain protein, BB0374. Recently, we characterized the EAL domain phosphodiesterase PdeA. A mutation inpdeAresulted in cells that were defective in motility and virulence. Here we demonstrate that BB0374/PdeB specifically hydrolyzed c-di-GMP with aKmof 2.9 nM, confirming that it is a functional phosphodiesterase. Furthermore, by measuring phosphodiesterase enzyme activity in extracts from cells containing thepdeA pdeBdouble mutant, we demonstrate that no additional phosphodiesterases are present inB. burgdorferi.pdeBsingle mutant cells exhibit significantly increased flexing, indicating a role for c-di-GMP in motility. Constructing and analyzing apilZpdeBdouble mutant suggests that PilZ likely interacts with chemotaxis signaling. While virulence in needle-inoculated C3H/HeN mice did not appear to be altered significantly inpdeBmutant cells, these cells exhibited a reduced ability to survive inIxodes scapularisticks. Consequently, those ticks were unable to transmit the infection to naïve mice. All of these phenotypes were restored when the mutant was complemented. Identification of this role ofpdeBincreases our understanding of the c-di-GMP signaling network in motility regulation and the life cycle ofB. burgdorferi.

scholarly journals The Borrelia burgdorferi Glycosaminoglycan Binding Protein Bgp in the B31 Strain Is Not Essential for Infectivity despite Facilitating Adherence and Tissue Colonization

2017 ◽  
Vol 86 (2) ◽  
Author(s):  
Samantha Schlachter ◽  
Janakiram Seshu ◽  
Tao Lin ◽  
Steven Norris ◽  
Nikhat Parveen
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Binding Protein ◽  
Mammalian Host ◽  
Infection Model ◽  
Binding Assays ◽  
Content Type ◽  
Mouse Infection Model ◽  
Successful Infection

ABSTRACTThe Lyme disease-causing organismBorrelia burgdorferiis transmitted into the mammalian host by an infected-tick bite. Successful infection relies on the ability of this extracellular pathogen to persist and colonize different tissues.B. burgdorferiencodes a large number of adhesins that are able to interact with host ligands to facilitate adherence and tissue colonization. Multiple glycosaminoglycan binding proteins present inB. burgdorferioffer a degree of redundancy of function during infection, and this highlights the importance of glycosaminoglycans as host cell receptors for spirochete adherence. Of particular interest in this study isBorreliaglycosaminoglycan binding protein (Bgp), which binds to heparin-related glycosaminoglycans. The properties of abgptransposon mutant and atrans-complemented derivative were compared to those of the wild-typeB. burgdorferiin thein vitrobinding assays and in infection studies using a C3H/HeJ mouse infection model. We determined that the loss of Bgp impairs spirochete adherence, infectivity, and tissue colonization, resulting in a reduction of inflammatory manifestations of Lyme disease. Although Bgp is not essential for infectivity, it is an important virulence factor ofB. burgdorferithat allows adherence and tissue colonization and contributes to disease severity.

scholarly journals The Lon-1 Protease Is Required by Borrelia burgdorferi To Infect the Mammalian Host

2020 ◽  
Vol 88 (6) ◽  
Author(s):  
Christina Thompson ◽  
Charlotte Mason ◽  
Shidoya Parrilla ◽  
Zhiming Ouyang
Keyword(s):  
Borrelia Burgdorferi ◽  
Bacterial Infection ◽  
Bacterial Resistance ◽  
Critical Role ◽  
Mammalian Host ◽  
Infection Model ◽  
Lon Protease ◽  
Content Type ◽  
Growth Defects ◽  
Functional Homolog

ABSTRACT Borrelia burgdorferi encodes a functional homolog of canonical Lon protease termed Lon-2. In addition, B. burgdorferi encodes a second Lon homolog called Lon-1. Recent studies suggest that Lon-1 may function differently from the prototypical Lon protease. However, the function of Lon-1 in B. burgdorferi biology remains virtually unknown. Particularly, the contribution of Lon-1 to B. burgdorferi fitness and infection remains hitherto unexplored. Herein, we show that Lon-1 plays a critical role for the infection of B. burgdorferi in a mammalian host. We found that lon-1 was highly expressed during animal infection, implying an important function of this protein in bacterial infection. We further generated a lon-1 deletion mutant and an isogenic complemented strain. Relative to that of the wild-type strain, the infectivity of the mutant was severely attenuated in a murine infection model. Our data also showed that the mutant displayed growth defects in regular BSK-II medium. Furthermore, bacterial resistance to osmotic stress was markedly reduced when lon-1 was inactivated. When exposed to tert-butyl hydroperoxide, survival of the lon-1 mutant was impaired. In addition, production of several virulence factors, such as BosR, RpoS, and OspC, was elevated in the mutant. These phenotypes were restored when the lon-1 mutation was complemented. Finally, we created a lon-1(S714A) mutant and found that this mutant failed to infect mice, suggesting that the proteolytic activity of Lon-1 is essential for bacterial infection. Taken together, these results demonstrate that Lon-1 is required by B. burgdorferi to infect animal hosts and to cope with environmental stresses.

scholarly journals BB0238, a Presumed Tetratricopeptide Repeat-Containing Protein, Is Required during Borrelia burgdorferi Mammalian Infection

2014 ◽  
Vol 82 (10) ◽  
pp. 4292-4306 ◽  
Author(s):  
Ashley M. Groshong ◽  
Danielle E. Fortune ◽  
Brendan P. Moore ◽  
Horace J. Spencer ◽  
Robert A. Skinner ◽  
...  
Keyword(s):  
Borrelia Burgdorferi ◽  
Protein Interactions ◽  
Point Mutations ◽  
Structural Motif ◽  
Tetratricopeptide Repeat ◽  
Mammalian Host ◽  
Content Type ◽  
Mammalian Infection ◽  
Tpr Domain

ABSTRACTThe Lyme disease spirochete,Borrelia burgdorferi, occupies both a tick vector and mammalian host in nature. Considering the unique enzootic life cycle ofB. burgdorferi, it is not surprising that a large proportion of its genome is composed of hypothetical proteins not found in other bacterial pathogens.bb0238encodes a conserved hypothetical protein of unknown function that is predicted to contain a tetratricopeptide repeat (TPR) domain, a structural motif responsible for mediating protein-protein interactions. To evaluate the role ofbb0238during mammalian infection, abb0238-deficient mutant was constructed. Thebb0238mutant was attenuated in mice infected via needle inoculation, and complementation ofbb0238expression restored infectivity to wild-type levels.bb0238expression does not change in response to varying culture conditions, and thus, it appears to be constitutively expressed underin vitroconditions.bb0238is expressed in murine tissues during infection, though there was no significant change in expression levels among different tissue types. Localization studies indicate that BB0238 is associated with the inner membrane of the spirochete and is therefore unlikely to promote interaction with host ligands during infection.B. burgdorfericlones containing point mutations in conserved residues of the putative TPR motif of BB0238 demonstrated attenuation in mice that was comparable to that in thebb0238deletion mutant, suggesting that BB0238 may contain a functional TPR domain.

scholarly journals Analysis of the Borrelia burgdorferi Cyclic-di-GMP-Binding Protein PlzA Reveals a Role in Motility and Virulence

2011 ◽  
Vol 79 (5) ◽  
pp. 1815-1825 ◽  
Author(s):  
Joshua E. Pitzer ◽  
Syed Z. Sultan ◽  
Yoshihiro Hayakawa ◽  
Gerry Hobbs ◽  
Michael R. Miller ◽  
...  
Keyword(s):  
Life Cycle ◽  
Borrelia Burgdorferi ◽  
Binding Protein ◽  
Survival Rates ◽  
Wild Type ◽  
Content Type ◽  
Infectious Dose ◽  
Swimming Pattern ◽  
Bacteria Inactivation ◽  
Mutant Cells

ABSTRACTThe cyclic-dimeric-GMP (c-di-GMP)-binding protein PilZ has been implicated in bacterial motility and pathogenesis. Although BB0733 (PlzA), the only PilZ domain-containing protein inBorrelia burgdorferi, was reported to bind c-di-GMP, neither its role in motility or virulence nor it's affinity for c-di-GMP has been reported. We determined that PlzA specifically binds c-di-GMP with high affinity (dissociation constant [Kd], 1.25 μM), consistent withKdvalues reported for c-di-GMP-binding proteins from other bacteria. Inactivation of the monocistronically transcribedplzAresulted in an opaque/solid colony morphology, whereas the wild-type colonies were translucent. While the swimming pattern of mutant cells appeared normal, on swarm plates, mutant cells exhibited a significantly reduced swarm diameter, demonstrating a role ofplzAin motility. Furthermore, theplzAmutant cells were significantly less infectious in experimental mice (as determined by 50% infectious dose [ID50]) relative to wild-type spirochetes. The mutant also had survival rates in fed ticks lower than those of the wild type. Consequently,plzAmutant cells failed to complete the mouse-tick-mouse infection cycle, indicatingplzAis essential for the enzootic life cycle ofB. burgdorferi. All of these defects were corrected when the mutant was complemented incis. We propose that failure ofplzAmutant cells to infect mice was due to altered motility; however, the possibility that an unidentified factor(s) contributed to interruption of theB. burgdorferienzootic life cycle cannot yet be excluded.

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