The Absence of Linear Plasmid 25 or 28-1 ofBorrelia burgdorferiDramatically Alters the Kinetics of Experimental Infection via Distinct Mechanisms

Abstract

The 25-kb linear plasmid lp25 and one of the 28-kb linear plasmids (lp28-1) are required for experimental infection in Borrelia burgdorferi, the etiologic agent of Lyme disease. The loss of these plasmids either eliminates infectivity (lp25) or significantly increases the 50% infective dose during a 2-week infection period (lp28-1). This study assessed the kinetics of bacterial dissemination in C3H/HeN mice infected with B. burgdorferi lacking either lp25 or lp28-1, as well as their wild-type parent, and tracked the development of specific borrelial antibodies over a 3-week period. The results indicated that the wild type and the lp28-1 ؊ strains were able to disseminate throughout the host, whereas the lp25 ؊ strain was cleared within 48 h of inoculation. While the wild-type B. burgdorferi persisted in tissues for the duration of the study, the lp28-1 ؊ mutant began clearing at day 8, with no detectable bacteria present by day 18. As expected, the wild-type strain persisted in C3H/HeN mice despite a strong humoral response; however, the lp28-1 ؊ mutant was cleared coincidently with the development of a modest immunoglobulin M response. The lp28-1 ؊ mutant was able to disseminate and persist in C3H-scid mice at a level indistinguishable from that of wild-type cells, confirming that acquired immunity was required for clearance in C3H/HeN mice. Thus, within an immunocompetent host, lp28-1-encoded proteins are not required for dissemination but are essential for persistence associated with Lyme borreliosis.Lyme disease or Lyme borreliosis is a multisystemic disorder transmitted by ticks of the genus Ixodes infected with the spirochetal bacterium Borrelia burgdorferi sensu lato, composed of B. burgdorferi sensu stricto, B. garinii, and B. afzelii (1,8,16). Lyme disease is the leading zoonotic infection in the United States, accounting for 95% of all reported vector-borne illness (10). The completed B. burgdorferi sensu stricto strain B31 annotated sequence revealed an unusual genome containing a linear chromosome of 910 kb in size as well as several extrachromosomal elements, including 12 linear plasmids of sizes ranging from 5 to 56 kb and 9 circular plasmids with sizes between 9 and 32 kb (9,14).Early studies had demonstrated a correlation between plasmid content and infectivity but were hindered by the plethora of plasmid species in B. burgdorferi and the inherent difficulty in resolving the many linear and circular species (24, 29-31). As such, prior to the information gained from the B. burgdorferi strain B31 genome sequence, it was difficult to definitively link a specific plasmid to a defect in infectivity. Exploiting the completed genome sequence of B. burgdorferi, we designed oligonucleotide primers that could be used in conjunction with PCR to specifically catalog the presence of plasmids from clonal isolates of B. burgdorferi (17). In our quest to link infectivity to any of the plasmids, we determined, along with others, that two linear plasmids, 25 and 28 kb in size (designated lp25 and lp28-1, respectiv...

Overexpression of CsrA (BB0184) Alters the Morphology and Antigen Profiles ofBorrelia burgdorferi

“…A clonal, noninfectious isolate of B. burgdorferi strain B31 (ML23 , Table 1), lacking linear plasmid 25 (lp25 negative), was used for the overexpression of CsrA Bb (44,45,65). All B. burgdorferi cultures used for transformations were grown in 1% CO 2 at 32°C in Barbour-StoennerKelly II (BSK-II) liquid medium (pH 7.6) supplemented with 6% normal rabbit serum (Pel-Freez Biologicals, Rogers, AR) (51,52).…”

Biology of Infection with Borrelia burgdorferi

“…Some are involved in early infection, such as OspC [112][113][114][115][116], which are presumably required for host colonization or resistance to innate immunity. Others are involved in resistance to acquired immunity, such as VlsE [13,109,117,118].…”

“…VlsE is a B. burgdorferi protein that is required for persistent mammalian infection [13,109,117,118] and whose synthesis begins around the time that OspC production ceases (Fig. 1) [103].…”

Biological Aspects of Lyme Disease Spirochetes: Unique Bacteria of the Borrelia Burgdorferi Species Group

“…However, some of those genes are necessary during natural infection course and for Borrelia persistence in the host as demonstrated by Labandeira-Rey et al 76,77 . They examined the infectivity of mutants lacking lp25 and lp28-1 in C3H mice and demonstrated that the loss of infectivity in lp25 -and elimination of lp28-1 -mutants from tissues coincident with the onset of specifi c IgM immune response.…”