Functional vulnerability of liver macrophages to capsules defines virulence of blood-borne bacteria.

Many encapsulated bacteria use capsules to cause invasive diseases. However, it remains largely unknown how the capsules enhance bacterial virulence under in vivo infection conditions. Here we show that the capsules primarily target the liver to enhance bacterial survival at the onset of blood-borne infections. In a mouse sepsis model, the capsules enabled human pathogens Streptococcus pneumoniae and Escherichia coli to circumvent the recognition of liver-resident macrophage Kupffer cells (KCs) in a capsular serotype-dependent manner. In contrast to effective capture of acapsular bacteria by KCs, the encapsulated bacteria are partially (low-virulence types) or completely (high-virulence types) "untouchable" for KCs. We finally identified the asialoglycoprotein receptor (ASGR) as the first known capsule receptor on KCs to recognize the low-virulence serotype-7F and -14 pneumococcal capsules. Our data identify the molecular interplay between the capsules and KCs as a master controller of the fate and virulence of encapsulated bacteria, and suggest that the interplay is targetable for therapeutic control of septic infections.

HtrA-mediated selective degradation of DNA uptake apparatus accelerates termination of pneumococcal transformation.

Natural transformation mediates horizontal gene transfer, and thereby promotes exchange of antibiotic resistance and virulence traits among bacteria. Streptococcus pneumoniae, the first known transformable bacterium, rapidly activates and then terminates the transformation state, but it is unclear how the bacterium accomplishes this rapid turn-around at the protein level. This work determined the transcriptomic and proteomic dynamics during the window of pneumococcal transformation. RNA sequencing revealed a nearly uniform temporal pattern of rapid transcriptional activation and subsequent shutdown for the genes encoding transformation proteins. In contrast, mass spectrometry analysis showed that the majority of transformation proteins were substantially preserved beyond the window of transformation. However, ComEA and ComEC, major components of the DNA uptake apparatus for transformation, were completely degraded at the end of transformation. Further mutagenesis screening revealed that the membrane-associated serine protease HtrA mediates selective degradation of ComEA and ComEC, strongly suggesting that breakdown of the DNA uptake apparatus by HtrA is an important mechanism for termination of pneumococcal transformation. Finally, our mutagenesis analysis showed that HtrA inhibits natural transformation of Streptococcus mitis and Streptococcus gordonii. Together, this work has revealed that HtrA regulates the level and duration of natural transformation in multiple streptococcal species.

A Cinchona Alkaloid Antibiotic That Appears To Target ATP Synthase in Streptococcus pneumoniae.

Optochin, a cinchona alkaloid derivative discovered over 100 years ago, possesses highly selective antibacterial activity toward Streptococcus pneumoniae. Pneumococcal disease remains the leading source of bacterial pneumonia and meningitis worldwide. The structure-activity relationships of optochin were examined through modification to both the quinoline and quinuclidine subunits, which led to the identification of analogue 48 with substantially improved activity. Resistance and molecular modeling studies indicate that 48 likely binds to the c-ring of ATP synthase near the conserved glutamate 52 ion-binding site, while mechanistic studies demonstrated that 48 causes cytoplasmic acidification. Initial pharmacokinetic and drug metabolism analyses of optochin and 48 revealed limitations of these quinine analogues, which were rapidly cleared, resulting in poor in vivo exposure through hydroxylation pendants to the quinuclidine and O-dealkylation of the quinoline. Collectively, the results provide a foundation to advance 48 and highlight ATP synthase as a promising target for antibiotic development.

Rifampicin can induce antibiotic tolerance in mycobacteria via paradoxical changes in rpoB transcription.

Metrics commonly used to describe antibiotic efficacy rely on measurements performed on bacterial populations. However, certain cells in a bacterial population can continue to grow and divide, even at antibiotic concentrations that kill the majority of cells, in a phenomenon known as antibiotic tolerance. Here, we describe a form of semi-heritable tolerance to the key anti-mycobacterial agent rifampicin, which is known to inhibit transcription by targeting the ß subunit of the RNA polymerase (RpoB). We show that rifampicin exposure results in rpoB upregulation in a sub-population of cells, followed by growth. More specifically, rifampicin preferentially inhibits one of the two rpoB promoters (promoter I), allowing increased rpoB expression from a second promoter (promoter II), and thus triggering growth. Disruption of promoter architecture leads to differences in rifampicin susceptibility of the population, confirming the contribution of rifampicin-induced rpoB expression to tolerance.