# The protein degradation system encoded by hslUV (ClpYQ) is dispensable for the virulence of Haemophilus ducreyi in human volunteers

## TL;DR

The hslUV system in Haemophilus ducreyi is not essential for causing disease in humans, suggesting other systems can compensate for its function.

## Contribution

The study shows functional redundancy in protein degradation systems in H. ducreyi, challenging its necessity for virulence.

## Key findings

- Deletion of hslUV did not reduce lesion formation in human volunteers.
- H. ducreyi may use Lon protease, ClpXP, and ClpB/DnaK to manage protein stress.
- The hslUV system is dispensable for virulence in human hosts.

## Abstract

Haemophilus ducreyi causes cutaneous ulcers in children who live in yaws-endemic countries and the genital ulcer disease chancroid. In the human host, H. ducreyi resides in an abscess and may need to resist both heat and oxidative stress, which result in aggregation and misfolding of bacterial proteins. In Escherichia coli, the hslUV (clpYQ) operon encodes a proteasome-like complex that degrades misfolded proteins and is upregulated during heat shock. In previous studies, we showed that hslUV transcripts are upregulated in experimental lesions caused by H. ducreyi in human volunteers, suggesting that HslUV may help H. ducreyi adapt to the abscess environment. Here, we constructed an unmarked hslUV operon deletion mutant, 35000HPΔhslUV, in H. ducreyi. Whole-genome sequencing showed that compared to its parent (35000HP), the mutant contained only the deletion of interest. Six volunteers were inoculated at three sites on skin overlying the deltoid on opposite arms with 35000HP and 35000HPΔhslUV. Within 24 h, papules formed at 88.9% (95% CI [69%, 100%]) at both parent and mutant-inoculated sites (P = 1.0). Pustules formed at 44.4% (95% CI [25.6%, 64.3%]) at parent-inoculated sites and 33.3% (95% CI [2.5%, 64.1%]) at mutant-inoculated sites (P = 0.17). Thus, the proteosome-like complex encoded by hslUV was dispensable for H. ducreyi virulence in humans. In the absence of hslUV, H. ducreyi likely utilizes other systems such as the Lon protease, ClpXP, and ClpB/DnaK to combat protein aggregation and misfolding, underscoring the importance of the functional redundancy of such systems in gram-negative pathogens.