Interfacial Stresses within Droplets and Channels Influence Bacterial Physiology: A Perspective.

Bacterial cells frequently experience fluid motion in their natural environments, like water bodies, aerosols, fomites, human capillaries, etc., a phenomenon that researchers have largely overlooked. Nevertheless, some reports have suggested that the interfacial stresses caused by fluid motion inside evaporating droplets or shear flows within capillaries may trigger physiological and morphological changes in the bacterial cells. Remarkably, the virulence of bacterial cells exhibits significant alterations in response to fluctuations in stress levels and external environmental factors. The dynamics of bacterial systems are analogous to colloidal systems but with the distinction that bacterial systems exhibit responsiveness, necessitating thorough exploration in dynamic environments. In this perspective, we discuss the important issue pertaining to bacterial survival, virulence, and disease transmission. Furthermore, we delineate a pathway and underscore emerging opportunities that demand exploration to unveil new avenues in the domains of bacterial pathogenicity, drug development, and strategies for disease mitigation.

Weather-related changes in the dehydration of respiratory droplets on surfaces bolster bacterial endurance.

HYPOTHESIS: The study shows for the first time a fivefold difference in the survivability of the bacterium Pseudomonas Aeruginosa (PA) in a realistic respiratory fluid droplet on fomites undergoing drying at different environmental conditions. For instance, in 2023, the annual average outdoor relative humidity (RH) and temperature in London (UK) is 71 % and 11 °C, whereas in New Delhi (India), it is 45 % and 26 °C, showing that disease spread from fomites could have a demographic dependence. Respiratory fluid droplet ejections containing pathogens on inanimate surfaces are crucial in disease spread, especially in nosocomial settings. However, the interplay between evaporation dynamics, internal fluid flow and precipitation and their collective influence on the distribution and survivability of pathogens at different environmental conditions are less known. EXPERIMENTS: Shadowgraphy imaging is employed to study evaporation, and optical microscopy imaging is used for precipitation dynamics. Micro-particle image velocimetry (MicroPIV) measurements reveal the internal flow dynamics. Confocal imaging of fluorescently labelled PA elucidates the bacterial distribution within the deposits. FINDINGS: The study finds that the evaporation rate is drastically impeded during drying at elevated solutal concentrations, particularly at high RH and low temperature conditions. MicroPIV shows reduced internal flow under high RH and low temperature (low evaporation rate) conditions. Evaporation rate influences crystal growth, with delayed efflorescence and extending crystallization times. PA forms denser peripheral arrangements under high evaporation rates and shows a fivefold increase in survivability under low evaporation rates. These findings highlight the critical impact of environmental conditions on pathogen persistence and disease spread from inanimate surfaces.

Future research perspective on the interfacial physics of non-invasive glaucoma testing in pathogen transmission from the eyes.

Non-contact tonometry (NCT) is a non-invasive ophthalmologic technique to measure intraocular pressure (IOP) using an air puff for routine glaucoma testing. Although IOP measurement using NCT has been perfected over many years, various phenomenological aspects of interfacial physics, fluid structure interaction, waves on corneal surface, and pathogen transmission routes to name a few are inherently unexplored. Research investigating the interdisciplinary physics of the ocular biointerface and of the NCT procedure is sparse and hence remains to be explored in sufficient depth. In this perspective piece, we introduce NCT and propose future research prospects that can be undertaken for a better understanding of the various hydrodynamic processes that occur during NCT from a pathogen transmission viewpoint. In particular, the research directions include the characterization and measurement of the incoming air puff, understanding the complex fluid-solid interactions occurring between the air puff and the human eye for measuring IOP, investigating the various waves that form and travel; tear film breakup and subsequent droplet formation mechanisms at various spatiotemporal length scales. Further, from an ocular disease transmission perspective, the disintegration of the tear film into droplets and aerosols poses a potential pathogen transmission route during NCT for pathogens residing in nasolacrimal and nasopharynx pathways. Adequate precautions by opthalmologist and medical practioners are therefore necessary to conduct the IOP measurements in a clinically safer way to prevent the risk associated with pathogen transmission from ocular diseases like conjunctivitis, keratitis, and COVID-19 during the NCT procedure.

Observations on phenomenological changes in Klebsiella Pneumoniae under fluidic stresses.

In the present work, experiments are conducted to understand the consequence of stresses generated by flowing fluid on the bacterial morphology and virulence in microfluidic channels. We consider Klebsiella pneumoniae (KP, a clinical isolate), an ESKAPE pathogen, to be the model bacteria responsible for blood stream infections, bacteremia, including pneumonia, urinary tract infections and more. Four different stress conditions are generated by changing the flow rate and channel geometry subsequently altering the shear rate and stressing time (τ). We observe significant changes in the structural aspects of the stressed bacteria. With an increase in stressing parameters, the viability of the bacterial sample deteriorated. Most importantly, these stressed samples proliferate much more than unstressed samples inside the RAW264.7 murine macrophages. The results shed light on the complex relationship between flow stresses and bacterial virulence. Furthermore, the bacterial samples are challenged with ciprofloxacin to see how they behave under different stress conditions. The observations presented in the present study can be extended to model deadly diseases including bacteremia using organ-on-a-chip technology and to understand bacterial pathogenicity under realistic environments.

phoP maintains the environmental persistence and virulence of pathogenic bacteria in mechanically stressed desiccated droplets.

Despite extensive studies on kinematic features of impacting drops, the effect of mechanical stress on desiccated bacteria-laden droplets remains unexplored. In the present study, we unveiled the consequences of the impaction of bacteria-laden droplets on solid surfaces and their subsequent desiccation on the virulence of an enteropathogen Salmonella typhimurium (STM). The methodology elucidated the deformation, cell-cell interactions, adhesion energy, and roughness in bacteria induced by impact velocity and low moisture because of evaporation. Salmonella retrieved from the dried droplets were used to understand fomite-mediated pathogenesis. The impact velocity-induced mechanical stress deteriorated the in vitro viability of Salmonella. Of interest, an uninterrupted bacterial proliferation was observed in macrophages at higher mechanical stress. Wild-type Salmonella under mechanical stress induced the expression of phoP whereas infecting macrophages. The inability of STM ΔphoP to grow in nutrient-rich dried droplets signifies the role of phoP in sensing the mechanical stress and maintaining the virulence of Salmonella.

On the specific heat capacity of HITEC-salt nanocomposites for concentrated solar power applications.

High specific heat capacity or C P of molten salt is crucial for concentrated solar power plants as it will enhance the energy density of thermal energy storage. It can be achieved by doping nanoparticles into molten salts. However, reported results show inconsistency in C P enhancement (positive and negative). Since the results are based on Differential Scanning Calorimeter (DSC) measurements of small batches (<10 mg), the average C P obtained from these results may not represent the bulk-C P of the nanocomposite, which is an important parameter from an application viewpoint. Moreover, the methods of salt-nanoparticle composite production lack industrial scalability. In this work, we examined a potentially scalable method based on mechanical shear mixing. The molten-salt of choice was HITEC due to its lower melting point, while inexpensive alumina and silica nanoparticles were used as dopants. To compare and contrast variability in C P enhancement, the sample selection was made by random sampling; DSC measurement was performed on small-sized batches (<10 mg), and the T-history method was applied on large-sized batches (20 g). While DSC tests indicated a mean decrease in C P for alumina (-43%) and an increase in C P for silica nanocomposite (+15%), T-history tests indicated a mean decrement in the bulk-C P for both alumina (-49%) and silica nanocomposites (-3%). This anomalous behavior in C P values was further compared using a nonparametric statistical test, the Mann-Whitney U test, which revealed that the C P of small-sized batches is statistically different from that of large-sized batches. Given their industrial scale of usage, the C P of the nanocomposite must be measured using both DSC and T-history methods to ascertain the effect of nanoparticles.

Physics of self-assembly and morpho-topological changes of Klebsiella pneumoniae in desiccating sessile droplets.

HYPOTHESIS: The bacteria suspended in pure water self-assemble into unique patterns depending on bacteria-bacteria, bacteria-substrate and bacteria-liquid interactions. The physical forces acting on bacteria vary based on their respective spatial location inside the droplet cause an assorted magnitude of physical stress. The shear and dehydration induced stress on pathogens(bacteria) in drying bio-fluid droplets alters the viability and infectivity. EXPERIMENTS: We have investigated the flow and desiccation-driven self-assembly of Klebsiella pneumoniae in the naturally evaporating sessile droplets. Klebsiella pneumoniae exhibits extensive changes in its morphology and forms unique patterns as the droplet dries, revealing hitherto unexplored rich physics governing its survival and infection strategies. Self-assembly of bacteria at the droplet contact line is characterized by order-to-disorder packing transitions with high packing densities and excessive deformations (analysed using scanning electron microscopy and atomic force microscopy). In contrast, thin-film instability-led hole formation at the center of the droplet engenders spatial packing of bacteria analogous to honeycomb weathering. FINDINGS: Self-assembly favors the bacteria at the rim of the droplet, leading to enhanced viability and pathogenesis on the famously known "coffee ring" of the droplet compared to the bacteria present at the center of the droplet residue. Mechanistic insights gained via our study can have far-reaching implications for bacterial infection through droplets, e.g., through open wounds.

Evaporation dynamics of a surrogate respiratory droplet in a vortical environment.

HYPOTHESIS: Vortex droplet interaction is crucial for understanding the route of disease transmission through expiratory jet where several such embedded droplets continuously interact with vortical structures of different strengths and sizes. EXPERIMENTS: A train of vortex rings with different vortex strength, quantified with vortex Reynolds number (Re'=0,53,221,297) are made to interact with an isolated levitated droplet, and the evolution dynamics is captured using shadowgraphy, particle image velocimetry (PIV), and backlight imaging technique. NaCl-DI water solution of 0, 1, 10 and 20 wt% concentrations are used as test fluids for the droplet. FINDINGS: The results show the dependence of evaporation characteristics on vortex strength, while the crystallization dynamics was found to be independent of it. A reduction of 12.23% and 14.6% in evaporation time was seen in case of de-ionized (DI) water and 1% wt NaCl solution respectively in presence of vortex ring train at Re'=221. In contrast to this, a minimal reduction in evaporation time (0.6% and 0.9% for DI water and 1% wt NaCl solution, respectively) is observed when Re' is increased from 221 to 297. The mechanisms for evaporation time reduction due to enhancement of convective heat and mass transfer from the droplet and shearing away of vapor layer by vortex ring interaction are discussed in this work.

Malleable Patterns from the Evaporation of a Colloidal Liquid Bridge: Coffee Ring to the Scallop Shell.

The present article highlights an approach to generating contrasting patterns from drying colloidal droplets in a liquid bridge configuration, different from well-known coffee rings. Reduction of the confinement distance (the gap between the solid surfaces) leads to systematized nanoparticle agglomeration yielding spoke-like patterns similar to those found on scallop shells instead of circumferential edge deposition. Alteration of the confinement distance modulates the curvature that entails variations in the evaporation flux across the liquid-vapor interface. Consequently, flow inside different liquid bridges (LBs) varies significantly for different confinement distance. Small confinement distance results in the stick-slip motion of squeezed liquid bridges. On the contrary, the stretched LBs exhibit pinned contact lines. The confinement distance determines the characteristic length scales of the thin film formed near the contact line, and its theoretical estimations are validated against the experimental observations using reflection interferometry, further exhibiting good agreement (in order of magnitude). We decipher a proposition that a drying liquid thin film (height ∼ O(10-7)m) present during dewetting near the three-phase contact line is responsible for the aligned deposition of particles. The coupled interplay of contact line dynamics, evaporation induced advection, and dewetting of the thin film at a three-phase interface contributes to the differences in deposition patterns.

Multiscale vapor-mediateddendritic pattern formation and bacterial aggregation in complex respiratory biofluid droplets.

HYPOTHESIS: Deposits of biofluid droplets on surfaces (such as respiratory droplets formed during an expiratory) are composed of water-based salt-protein solution that may also contain an infection (bacterial/viral). The final patterns of the deposit formed and bacterial aggregation on the deposits are dictated by the fluid composition and flow dynamics within the droplet. EXPERIMENTS: This work reports the spatio-temporal, topological regulation of deposits of respiratory fluid droplets and control of bacterial aggregation by tweaking flow inside droplets using non-contact vapor-mediated interactions. Desiccated respiratory droplets form deposits with haphazard multiscale dendritic, cruciform-shaped precipitates when evaporated on a glass substrate. However, we showcase that short and long-range vapor-mediated interaction between the droplets can be used as a tool to control these deposits at nano-micro-millimeter scales. We morphologically control hierarchial dendrite size, orientation and subsequently suppress cruciform-shaped crystals by placing a droplet of ethanol in the vicinity of the biofluid droplet. Active living matter in respiratory fluids like bacteria is preferentially segregated and agglomerated without its viability and pathogenesis attenuation. FINDINGS: The nucleation sites can be controlled via preferential transfer of solutes in the droplets; thus, achieving control over crystal occurrence, growth dynamics, and the final topology of the deposit. For the first time, we have experimentally presented a proof-of-concept to control the aggregation of live active matter like bacteria without any direct contact. The methodology can have ramifications in biomedical applications like disease detection and bacterial segregation.