Biology subjects

Paranjape, K.

Publications and source records attributed to Paranjape, K..

5 recordsLinked to original sources

bioRxiv2024

Host-specific adaptation of Legionella pneumophila to single and multiple hosts

Legionella pneumophila is an endosymbiotic bacterial species able to infect and reproduce in various protist and human hosts. Upon entry in human lungs, they may infect lung macrophages, causing Legionnaires disease (LD), an atypical pneumonia, using similar mechanisms as in their protozoan hosts, despite the two hosts being separated by a billion years of evolution. In this study, we used experimental evolution to identify genes conferring host-specificity to L. pneumophila. To this end, we passaged L. pneumophila in two different hosts - Acanthamoeba castellanii and the human macrophage-like cells U937 - separately and by switching between the hosts twice a week for a year. In total, we identified 1518 mutations present in at least 5% of the population at the time of sampling. Half of these were localized in five groups of repeated sequences, likely to be recombination hotspots. Forty-nine mutations were fixed in the 18 populations at the end of the experiment, representing four different groups. The first two groups involve adaptation to the specific selection conditions, including (i) two specific mutations in the 30S ribosomal protein S12 (RpsL) that conferred resistance to streptomycin, to which the bacteria were unexpectedly exposed to during serial passage and (ii) two mutations, one in the 30S ribosomal protein S4 (RpsD), and one in the chaperonin GroES, both of which are likely to be fitness-restoring compensatory mutations to the original RpsL mutations. Two more interesting groups of mutations included (iii) mutations in 4 different strain-specific genes involved in LPS synthesis, found only in the lineages passaged with A. castellanii and (iv) mutations in the gene coding for LerC, a key regulator of protein effector expression, which was independently mutated in 6 lineages grown in presence of the U937 cells. We propose that the mutations degrading the function of the regulator LerC improve the fitness of L. pneumophila in human-derived cells, and that modifications in the LPS are beneficial for growth in A. castellanii. This study is a first step in further investigating determinants of host specificity in L. pneumophila.

evolutionary biology↗

bioRxiv2022

Toxoflavin secreted by Pseudomonas alcaliphila inhibits growth of Legionella pneumophila and its host Vermamoeba vermiformis

Legionella pneumophila is a natural inhabitant of water systems. From there, it can be transmitted to humans by aerosolization resulting in severe pneumonia. Most large outbreaks are caused by cooling towers contaminated with L. pneumophila. The resident microbiota of the cooling tower is a key determinant for the colonization and growth of L. pneumophila. The genus Pseudomonas correlates negatively with the presence of L. pneumophila, but it is not clear which species is responsible. Therefore, we identified the Pseudomonas species inhabiting 14 cooling towers using a Pseudomonas-specific 16S rRNA amplicon sequencing strategy. Cooling towers free of L. pneumophila contained a high relative abundance of members from the Pseudomonas alcaliphila/oleovorans phylogenetic cluster. In vitro, P. alcaliphila JCM 10630 inhibited the growth of L. pneumophila on agar plates. Analysis of the P. alcaliphila genome revealed the presence of a genes cluster predicted to produce toxoflavin. L. pneumophila growth was inhibited by pure toxoflavin and by extract from P. alcaliphila culture found to contain toxoflavin by LC-ESI-MS. In addition, toxoflavin inhibits growth of Vermameoba vermiformis, a host cell of L. pneumophila. Our study indicates that P. alcaliphila may be important to restrict growth of L. pneumophila in water systems through the production of toxoflavin. A sufficiently high concentration is likely not achieved in the bulk water but might have a local inhibitory effect such as in biofilm.

microbiology↗

bioRxiv2020

Local adaptation of Legionella pneumophila within a hospital hot water system increases tolerance to copper

In large-building water systems, Legionella pneumophila is exposed to common environmental stressors such as copper. The aim of this study was to evaluate the susceptibility to copper of L. pneumophila isolates recovered from various sites: two clinical and seven environmental from hot water systems biofilm & water, and from cooling tower water. After one-week acclimation in simulated drinking water, strains were exposed to various copper concentrations (0.8 to 5 mg/L) for over 672 hours. Complete loss of culturability was observed for three isolates, following copper exposure to 5 mg/L for 672h. Two ST1427-like isolates were highly sensitive to copper, while the other two, isolated from biofilm samples, were resistant. The expression of the copper resistance gene copA evaluated by RT-qPCR was significantly higher for the biofilm isolates. All four ST1427-like isolates were recovered from the same water system during an outbreak. Whole genome sequencing results confirmed that the four isolates are very close phylogenetically, differing by only 29 single nucleotide polymorphisms, suggesting in situ adaptation to microenvironmental conditions, possibly due to epigenetic regulation. These results indicate that the immediate environment within a building water distribution system influences the tolerance of L. pneumophila to copper. Increased contact of L. pneumophila biofilm strains with copper piping or copper alloys in the heat exchanger might lead to local adaptation. The phenotypic differences observed between water and biofilm isolates from the hot water system of a healthcare facility warrants further investigation to assess the relevance of evaluating disinfection performances based on water sampling alone. ImportanceLegionella pneumophila is a pathogen indigenous to natural and large building water systems in the bulk and the biofilm phases. The immediate environment within a system can impact the tolerance of L. pneumophila to environmental stressors, including copper. In healthcare facilities, copper levels in water can vary, depending on water quality, plumbing materials and age. This study evaluated the impact of the isolation site (water vs biofilm, hot water system vs cooling tower) within building water systems. Closely related strains isolated from a healthcare facility hot water system exhibited variable tolerance to copper stress shown by differential expression of copA, with biofilm isolates displaying highest expression and tolerance. Relying on the detection of L. pneumophila in water samples following exposure to environmental stressor such as copper may underestimate the prevalence of L. pneumophila, leading to inappropriate risk management strategies and increasing the risk of exposure for vulnerable patients.

microbiology↗

bioRxiv2019

Impact of temperature on Legionella pneumophila, its protozoan host cells, and the microbial diversity of the biofilm community of a pilot cooling tower

Legionella pneumophila (Lp) is a waterborne bacterium known for causing Legionnaires Disease, a severe pneumonia. Cooling towers are a major source of outbreaks, since they provide ideal conditions for Lp growth and produce aerosols. In such systems, Lp typically grow inside protozoan hosts. Several abiotic factors such as water temperature, pipe material and disinfection regime affect the colonization of cooling towers by Lp. The local physical and biological factors promoting the growth of Lp in water systems and its spatial distribution are not well understood. Therefore, we built a lab-scale cooling tower to study the dynamics of Lp colonization in relationship to the resident microbiota and spatial distribution. The pilot was filled with water from an operating cooling tower harboring low levels of Lp. It was seeded with Vermamoeba vermiformis, a natural host of Lp, and then inoculated with Lp. After 92 days of operation, the pilot was disassembled, the water was collected, and biofilm was extracted from the pipes. The microbiome was studied using 16S rRNA and 18S rRNA genes amplicon sequencing. The communities of the water and of the biofilm were highly dissimilar. The relative abundance of Legionella in water samples reached up to 11% whereas abundance in the biofilm was extremely low ([≤]0.5 %). In contrast, the host cells were mainly present in the biofilm. This suggest that Lp grows in host cells associated with biofilm and is then released back into the water following host cell lysis. In addition, water temperature shaped the bacterial and eukaryotic community of the biofilm, indicating that different parts of the systems may have different effects on Legionella growth.

microbiology↗

bioRxiv2019

Presence of Legionella spp. in cooling towers: the role of microbial diversity, Pseudomonas, and continuous chlorine application.

Legionnaires Disease (LD) is a severe pneumonia caused by Legionella pneumophila. Cooling towers are the main source of L. pneumophila during large outbreaks. Colonization, survival, and proliferation of L. pneumophila in cooling towers are necessary for outbreaks to occur. These steps are affected by chemical and physical parameters of the cooling tower environment. We hypothesize that the bacterial community residing in the cooling tower could also affect the presence of L. pneumophila. A 16S rRNA targeted amplicon sequencing approach was used to study the bacterial community of cooling towers and its relationship with the Legionella spp. and L. pneumophila communities. The results indicated that the water source shaped the bacterial community of cooling towers. Several taxa were enriched and positively correlated with Legionella spp. and L. pneumophila. In contrast, Pseudomonas showed a strong negative correlation with Legionella spp. and several other genera. Most importantly, continuous chlorine application reduced microbial diversity and promoted the presence of Pseudomonas creating a non-permissive environment for Legionella spp. This suggests that disinfection strategies as well as the resident microbial population influences the ability of Legionella spp. to colonize cooling towers.

microbiology↗