Borrelia Bavarriensis 58

What distinguishes B. bavariensis from other Borrelia species within the B. burgdorferi sensu lato complex?

B. bavariensis is one of several pathogenic species within the B. burgdorferi sensu lato complex, alongside B. burgdorferi sensu stricto, B. afzelii, B. garinii, B. spielmanii, and B. mayonii . Previously considered a subtype of B. garinii (OspA serotype 4 or NT29-like), B. bavariensis was proposed as a separate genospecies in 2009 and validated in 2013 .

This species demonstrates several distinctive characteristics:

• It shows a high propensity to infect humans and is associated with severe manifestations of Lyme borreliosis, particularly neuroborreliosis

• It is primarily rodent-adapted

• It is distributed across Eurasia, utilizing either Ixodes ricinus (Europe) or Ixodes persulcatus (Asia) as vectors

• Despite causing significant human disease, it is rarely recovered from field-collected Ixodes ticks

What is the estimated evolutionary timeline for B. bavariensis divergence?

Molecular clock analyses estimate that:

• The split between B. garinii and B. bavariensis occurred approximately 137,000 generations ago

• The divergence between the two B. bavariensis populations (European and Asian) happened more recently, around 92,000 generations ago

These estimates are based on a mutation rate of 10^-7 substitutions per site per generation, though it's important to note this rate has not been definitively proven for B. burgdorferi sensu lato .

What are the key genomic features of B. bavariensis isolates?

B. bavariensis genomes combine a high degree of genetic conservation with significant plasticity:

• All isolates share the main chromosome and five plasmids

• Beyond these conserved elements, the repertoire of other plasmids is highly variable

• Genome modification mechanisms include plasmid losses, gains through horizontal transfer, and plasmid fusions

European and Asian populations show distinct patterns:

• European isolates exhibit limited diversity in genome content with some geographic structure

• Asian isolates each possess unique plasmid repertoires without obvious geographic differentiation between Japanese and Russian isolates

How can researchers identify and classify plasmids within B. bavariensis genomes?

Plasmid identification in B. burgdorferi sensu lato genomes can be facilitated by identifying plasmid partition genes on assembled contigs. Specifically:

• Five such genes have been described in B. burgdorferi sensu stricto

• Each replicon typically contains only one copy of these genes unless it represents a fusion of two plasmids

• The PFam32 protein family sequences are commonly used to name plasmids based on homology to B. burgdorferi sensu stricto

• Not all plasmids possess PFam32, but PFam50 and 57/62 also appear unique for each plasmid type and can aid identification in such cases

For accurate plasmid reconstruction:

• A combination of long-read (Pacific Bioscience) and short-read (Illumina) sequencing techniques allows proper genome reconstruction in most cases

• When only short-read sequencing data is available, a very close reference is necessary for accurate assembly

How do B. bavariensis strains evade host immune responses?

B. bavariensis, like other Lyme disease spirochetes, must overcome the first line of defense of the innate immune system after transmission to mammalian hosts. This is accomplished through two primary mechanisms:

• Binding complement regulators

• Terminating the formation of the membrane attack complex (MAC)

Specifically in B. bavariensis, the proteins BGA66 and BGA71 (members of the PFam54 family) inhibit complement activation by:

• Interacting with the late complement components C7, C8, and C9

• Binding to the formed membrane attack complex

What research opportunities are presented by naturally occurring PFam54-deficient strains?

Two B. bavariensis strains, PBN and PNi, naturally lack the entire PFam54 gene array while maintaining over 95% genomic identity to the reference strain PBi . This natural occurrence presents unique research opportunities:

• These strains serve as natural knockouts, circumventing the technical hurdles of simultaneously deleting all PFam54 proteins

• Comparative studies reveal that PBN and PNi show reduced survival in human serum compared to PBi

• Complementation experiments demonstrate that introducing recombinant BGA66 and BGA71 restores serum resistance

• Each recombinant protein individually confers serum resistance, while their combination does not enhance protection further

The properties of these strains are summarized in the following table:

What animal models are appropriate for studying B. bavariensis infections?

BALB/c mice have been successfully used to study B. bavariensis infection dynamics. Key methodological considerations include:

• Intradermal inoculation of spirochetes (with culture medium as a control)

• Assessment of spirochete burden at 21 days post-inoculation

• Tissue sampling should include the inoculation site, heart, bladder, ear, knee joints, and tibiotarsal joints

• Quantitative analysis of spirochete burden in these tissues provides insights into strain-specific tissue tropism

In comparative studies using this model, PFam54-containing (PBi) and PFam54-deficient strains (PBN and PNi) all established productive infections in mice, suggesting that:

• PFam54 is not essential for spirochete persistence in mice after intradermal infection

• Absence of PFam54 may lead to strain-specific differences in the efficiency of colonizing certain murine tissues

How can researchers validate the absence of PFam54 genes in Borrelia isolates?

To confirm the absence of PFam54 genes, researchers should employ multiple complementary approaches:

• Genomic sequencing using both long-read (PacBio) and short-read (Illumina) technologies

• Paralog-specific PCR using primers designed for individual PFam54 genes

• Verification of PCR products through Sanger sequencing to confirm identity

• Comparative analysis of lp54 plasmid length (typically shorter in PFam54-deficient strains)

When unexpected PCR products are observed, sequencing is essential to determine whether they represent PFam54 genes or unrelated genomic regions, as was noted when primers targeting bga68 amplified a chromosomal region in PFam54-deficient strains .

What structural insights have been gained regarding B. bavariensis immune evasion proteins?

The crystal structure of BGA71, a potent MAC inhibitor, has been determined at 2.9 Å resolution, revealing:

• A cysteine cross-linked homodimer structure

• Based on this structure and sequence alignment with CspA from B. burgdorferi, potential binding sites for C7 and C9 (MAC constituents) have been proposed

These structural insights:

• Illuminate the molecular mechanisms of immune evasion developed by pathogenic Borrelia species

• Aid understanding of Lyme disease pathogenesis

• May facilitate development of new strategies to prevent Lyme disease

What genetic diversity patterns are observed within B. bavariensis compared to other Borrelia species?

Comparative genetic diversity analysis shows specific patterns across Borrelia species:

These statistics suggest:

• B. bavariensis shows intermediate levels of diversity (π = 0.008) compared to other Borrelia species

• The negative Tajima's D value (-0.60) suggests potential population expansion or purifying selection

• Certain genes show evidence of balancing selection across Borrelia species, including the cytidine deaminase gene cdd (BB0618), which displays high diversity and elevated Tajima's D in both B. garinii and B. bavariensis

Understanding these patterns may help identify genes under selection that could be important during speciation events or population divergence.