Jayasimha Rao

Indwelling urinary catheters frequently develop intraluminal biofilms, increasing the risk of catheter-associated urinary tract infections and systemic infections. Current prophylactic strategies, including antibiotics and antimicrobial coatings, have not consistently reduced the risk of infection. Histotripsy, a focused ultrasound ablation method, offers a promising non-invasive approach to mechanically disrupt biofilms and has demonstrated efficacy against catheter-associated biofilms in vitro. This study evaluated the ability of histotripsy to disrupt clinically derived intraluminal biofilms. Explanted urinary catheters were segmented and treated ex vivo with histotripsy, with matched untreated segments serving as controls. Following treatment, catheter segments were assessed for microbial viability by culture and for biofilm biomass using a crystal violet assay. Three silicone catheters were collected, each with four treated and four matched control segments. Based on Gram stain morphology, the three catheter tip cultures were consistent with Staphylococcus aureus, mixed Gram-positive and Gram-negative organisms, and a budding yeast suggestive of a Candida species, respectively. A single histotripsy treatment with a pulse repetition frequency of 200 Hz and estimated peak negative pressure of 31.6 MPa resulted in an 81.7% reduction in biofilm biomass compared with untreated controls (p < 0.001). Viable organisms remained detectable after treatment under the tested parameters. These findings support further investigation of histotripsy as an approach for managing catheter-associated biofilm infections. Additional studies are needed to optimize treatment parameters and explore potential clinical applications.

Providencia stuartii (Ps) is a clinically significant opportunistic pathogen often associated with “difficult-to-treat resistance” (DTR) infections due to pan-resistance to first-line antimicrobials. We report the clinical diagnosis and rapid genomic characterization of strain Ps-CMC-4104, recovered from a human splenic abscess in a patient with infected necrotizing pancreatitis. To resolve the complex genetic architecture of this strain, we utilized hybrid sequencing combining Oxford Nanopore (long-read) and Illumina (short-read) technologies. Analysis revealed a 4,504,925 bp circular chromosome featuring a unique genomic resistance island (GRI) closely related to Salmonella SGI1. Notably, the PsGRI contains multiple copies of NDM-1 and PER-1 carbapenem-resistance and -inhibitor genes, a repetitive structure typically unresolvable by standard short-read methods. Additionally, a large 278,489 bp low-copy circular plasmid harbored single copies of these carbapenemase and extended-spectrum β-lactamase genes alongside other antimicrobial resistance determinants and ISCR1 insertion sequences. Nanopore technology allowed us to precisely identify the duplications, providing critical insights into the strain’s pan-resistant phenotype. This study serves as proof-of-concept for the importance of integrating long-read sequencing into clinical workflows to identify complex resistance mechanisms in DTR pathogens, facilitating targeted antimicrobial stewardship and infection control.

Urinary catheterization is a common procedure, affecting 15%–25% of hospitalized patients. This procedure, however, often results in bacterial infections, primarily caused by biofilm‐forming Gram‐negative bacteria that adhere to the catheter surface. To address this challenge, we developed a polymer‐based antimicrobial coating using polydopamine (PD) embedded with gentamicin (Gent). We evaluated the antibiofilm efficacy of this coating (PD‐Gent) using a biofilm‐forming isolate of Pseudomonas aeruginosa (PAO1) as P. aeruginosa is commonly implicated in catheter‐related infections. We observed that the PD‐Gent coating significantly reduced biofilm formed by PAO1 compared to the uncoated control. Importantly, the coating maintained its antibiofilm activity across diverse substrates, including polystyrene, poly(vinyl chloride), and silicone. The approach was further extended to incorporate other antibiotics (tobramycin, amikacin), demonstrating adaptability to multiple antimicrobial agents. Finally, artificial urine inoculated with PAO1 was deployed through PD‐Gent–coated silicone Foley catheters under continuous flow for 24 h. Despite this continuous introduction of PAO1, the coated catheters inhibited biofilm formation by three‐fold compared to the uncoated control catheters. These findings underscore the promise of PD‐Gent as a robust, versatile coating with strong potential to significantly reduce the incidence of catheter‐associated infections in clinical settings.

Biofilms develop in sequential steps resulting in the formation of three-dimensional communities of microorganisms that are encased in self-produced extracellular polymeric substances. Biofilms play a key role in device-associated infections, such as catheter-associated urinary tract infections (CAUTIs), because they protect microorganisms from standard antimicrobial therapies. Current strategies to prevent biofilm formation in catheter-related infections, including prophylactic antibiotics and antibiotic-coated catheters, have been unsuccessful. This finding highlights a need for novel approaches to address this clinical problem. In this study, biofilm-forming phenotypes of common Gram-negative bacteria associated with CAUTIs were treated with antisense peptide nucleic acids (PNAs), and biofilm biomass and bacterial viability were quantified after 24 h of treatment. A cocktail of PNAs targeting the global regulator genes rsmA, amrZ, and rpoS in Pseudomonas aeruginosa significantly reduced viability and thus appropriately eliminated biofilm biomass. Antisense-PNAs against these same gene targets and the motility regulator gene motA inhibited biofilm formation among isolates of Klebsiella pneumoniae, Enterobacter cloacae, and Escherichia coli but did not reduce bacterial viability. These results suggest that antisense-PNAs are a promising new technology in preventing biofilm formation in urinary catheters, especially as a potential complement to conventional antimicrobials.

The COVID-19 pandemic initiated public interest in wastewater-based epidemiology (WBE). Public and private entities responded to the need to produce timely and accurate data. LuminUltra and Hach partnered to provide a rapid, field-based quantitative polymerase chain reaction (qPCR) test for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in wastewater. This study evaluates the Hach GeneCount SARS-CoV-2 Wastewater RT-qPCR Assay Kit and LuminUltra GeneCount® Q-16 RT-PCR instrument. The Hach LuminUltra methods were compared to the Promega Wizard® Enviro Total Nucleic Acid kit and Bio-Rad CFX Opus 96 Real-time PCR Detection System. Over a 12-week period, wastewater samples were collected weekly from seven locations in the Roanoke/Salem, VA sewersheds. Concentration and extraction of the viral RNA were followed by qPCR analysis. The target gene for detection was the nucleocapsid gene (N1) of the SARS-CoV-2 virus. Costs, ease of use, time to produce results, sample preparation, and data comparisons were considered. The comparison determined that the Hach LuminUltra method and instrument were more affordable, consumed less time, and required less technical expertise. While the new method was specific, it had low sensitivity. This evaluation suggests the Hach LuminUltra method should be reserved for limited situations requiring onsite field analysis where data accuracy is not essential.

We report the complete genome sequence of a clinical isolate of Providencia stuartii strain CMC-4104, isolated from a splenic abscess. Oxford Nanopore Technologies (ONT) and Illumina sequencing reads were assembled using Geneious to generate a 4,504,925-bp circular chromosome containing multiple copies of the NDM-1 and PER-1 genes in a genomic resistance island.

Objective . This paper is an initial work towards developing particle-mediated histotripsy (PMH) as a novel method of treating catheter-based medical device (CBMD) intraluminal biofilms. Impact Statement . CBMDs commonly become infected with bacterial biofilms leading to medical device failure, infection, and adverse patient outcomes. Introduction . Histotripsy is a noninvasive focused ultrasound ablation method that was recently proposed as a novel method to remove intraluminal biofilms. Here, we explore the potential of combining histotripsy with acoustically active particles to develop a PMH approach that can noninvasively remove biofilms without the need for high acoustic pressures or real-time image guidance for targeting. Methods . Histotripsy cavitation thresholds in catheters containing either gas-filled microbubbles (MBs) or fluid-filled nanocones (NCs) were determined. The ability of these particles to sustain cavitation over multiple ultrasound pulses was tested after a series of histotripsy exposures. Next, the ability of PMH to generate selective intraluminal cavitation without generating extraluminal cavitation was tested. Finally, the biofilm ablation and bactericidal capabilities of PMH were tested using both MBs and NCs. Results . PMH significantly reduced the histotripsy cavitation threshold, allowing for selective luminal cavitation for both MBs and NCs. Results further showed PMH successfully removed intraluminal biofilms in Tygon catheters. Finally, results from bactericidal experiments showed minimal reduction in bacteria viability. Conclusion . The results of this study demonstrate the potential for PMH to provide a new modality for removing bacterial biofilms from CBMDs and suggest that additional work is warranted to develop histotripsy and PMH for treatment of CBMD intraluminal biofilms.

We report the complete genome of a clinical strain of Pseudomonas aeruginosa , CMC-097, which was isolated from a ventilator-associated pneumonia patient with a chronic infection. Illumina sequence reads were assembled using Geneious to yield a 7,044,064-bp circular chromosome containing a carbapenem resistance integron, In 2020 .

We report the complete genome of clinical strain Pseudomonas aeruginosa CMC-115, which was isolated from an acute ventilator-associated pneumonia patient. Illumina sequencing reads were assembled using Geneious to yield a 6,375,262-bp circular chromosome that exhibited an unusual ferrichrome receptor in the pyoverdine synthesis locus and the absence of type 3 secretion system genes.

Background: Holarrhena antidysenterica is a medicinal plant widely used in traditional medicine for treating gastrointestinal disorders. Plant-derived antimicrobial agents have gained increasing attention as potential alternatives to synthetic drugs owing to the rising antimicrobial resistance. Objective: This study aimed to evaluate the phytochemical profile and antibacterial and antifungal activities of the ethanolic extract of H. antidysenterica. Materials and Methods: Bark powder was macerated in 80% ethanol, and the resulting extract underwent qualitative phytochemical evaluation. Analysis revealed major groups such as alkaloids, phenolics, tannins, steroids, glycosides, and coumarins, whereas saponins, terpenoids, carbohydrates, proteins, and anthraquinones were not detected. Antibacterial efficacy was evaluated using the agar well diffusion assay against E. coli, and S. flexneri (MTCC strains) at extract concentrations between 50 and 200 µg/mL. Results: A clear dose-dependent inhibition was recorded, with the largest inhibition zone observed for E. coli (28 mm), followed by S. flexneri (24 mm) at 200 µg/mL, approaching the efficacy of standard azithromycin. The extract, however, did not display any measurable antifungal activity against C. albicans or A. niger. The antibacterial activity is due to the combined action of alkaloids, phenols, and tannins. Conclusion: These results support the traditional medicinal use of H. antidysenterica and indicate its potential as a natural antibacterial source for the development of plant-based antimicrobial agents.