Targeted genomic sequencing of avian influenza viruses in wetland sediment from wild bird habitats.

Diverse influenza A viruses (IAVs) circulate in wild birds, including highly pathogenic strains that infect poultry and humans. Consequently, surveillance of IAVs in wild birds is a cornerstone of agricultural biosecurity and pandemic preparedness. Surveillance is traditionally done by testing wild birds directly, but obtaining these specimens is labor intensive, detection rates can be low, and sampling is often biased toward certain avian species. As a result, local incursions of dangerous IAVs are rarely detected before outbreaks begin. Testing environmental specimens from wild bird habitats has been proposed as an alternative surveillance strategy. These specimens are thought to contain diverse IAVs deposited by a broad range of avian hosts, including species that are not typically sampled by surveillance programs. To enable this surveillance strategy, we developed a targeted genomic sequencing method for characterizing IAVs in these challenging environmental specimens. It combines custom hybridization probes, unique molecular index-based library construction, and purpose-built bioinformatic tools, allowing IAV genomic material to be enriched and analyzed with single-fragment resolution. We demonstrated our method on 90 sediment specimens from wetlands around Vancouver, Canada. We recovered 2,312 IAV genome fragments originating from all eight IAV genome segments. Eleven hemagglutinin subtypes and nine neuraminidase subtypes were detected, including H5, the current global surveillance priority. Our results demonstrate that targeted genomic sequencing of environmental specimens from wild bird habitats could become a valuable complement to avian influenza surveillance programs.IMPORTANCEIn this study, we developed genome sequencing tools for characterizing avian influenza viruses in sediment from wild bird habitats. These tools enable an environment-based approach to avian influenza surveillance. This could improve early detection of dangerous strains in local wild birds, allowing poultry producers to better protect their flocks and prevent human exposures to potential pandemic threats. Furthermore, we purposefully developed these methods to contend with viral genomic material that is diluted, fragmented, incomplete, and derived from multiple strains and hosts. These challenges are common to many environmental specimens, making these methods broadly applicable for genomic pathogen surveillance in diverse contexts.

The multifaceted virulence of adherent-invasive Escherichia coli.

The surge in inflammatory bowel diseases, like Crohn's disease (CD), is alarming. While the role of the gut microbiome in CD development is unresolved, the frequent isolation of adherent-invasive Escherichia coli (AIEC) strains from patient biopsies, together with their propensity to trigger gut inflammation, underpin the potential role of these bacteria as disease modifiers. In this review, we explore the spectrum of AIEC pathogenesis, including their metabolic versatility in the gut. We describe how AIEC strains hijack the host defense mechanisms to evade immune attrition and promote inflammation. Furthermore, we highlight the key traits that differentiate AIEC from commensal E. coli. Deciphering the main components of AIEC virulence is cardinal to the discovery of the next generation of antimicrobials that can selectively eradicate CD-associated bacteria.

Medical Masks Versus N95 Respirators for Preventing COVID-19 Among Health Care Workers : A Randomized Trial.

BACKGROUND: It is uncertain if medical masks offer similar protection against COVID-19 compared with N95 respirators. OBJECTIVE: To determine whether medical masks are noninferior to N95 respirators to prevent COVID-19 in health care workers providing routine care. DESIGN: Multicenter, randomized, noninferiority trial. (ClinicalTrials.gov: NCT04296643). SETTING: 29 health care facilities in Canada, Israel, Pakistan, and Egypt from 4 May 2020 to 29 March 2022. PARTICIPANTS: 1009 health care workers who provided direct care to patients with suspected or confirmed COVID-19. INTERVENTION: Use of medical masks versus fit-tested N95 respirators for 10 weeks, plus universal masking, which was the policy implemented at each site. MEASUREMENTS: The primary outcome was confirmed COVID-19 on reverse transcriptase polymerase chain reaction (RT-PCR) test. RESULTS: In the intention-to-treat analysis, RT-PCR-confirmed COVID-19 occurred in 52 of 497 (10.46%) participants in the medical mask group versus 47 of 507 (9.27%) in the N95 respirator group (hazard ratio [HR], 1.14 [95% CI, 0.77 to 1.69]). An unplanned subgroup analysis by country found that in the medical mask group versus the N95 respirator group RT-PCR-confirmed COVID-19 occurred in 8 of 131 (6.11%) versus 3 of 135 (2.22%) in Canada (HR, 2.83 [CI, 0.75 to 10.72]), 6 of 17 (35.29%) versus 4 of 17 (23.53%) in Israel (HR, 1.54 [CI, 0.43 to 5.49]), 3 of 92 (3.26%) versus 2 of 94 (2.13%) in Pakistan (HR, 1.50 [CI, 0.25 to 8.98]), and 35 of 257 (13.62%) versus 38 of 261 (14.56%) in Egypt (HR, 0.95 [CI, 0.60 to 1.50]). There were 47 (10.8%) adverse events related to the intervention reported in the medical mask group and 59 (13.6%) in the N95 respirator group. LIMITATION: Potential acquisition of SARS-CoV-2 through household and community exposure, heterogeneity between countries, uncertainty in the estimates of effect, differences in self-reported adherence, differences in baseline antibodies, and between-country differences in circulating variants and vaccination. CONCLUSION: Among health care workers who provided routine care to patients with COVID-19, the overall estimates rule out a doubling in hazard of RT-PCR-confirmed COVID-19 for medical masks when compared with HRs of RT-PCR-confirmed COVID-19 for N95 respirators. The subgroup results varied by country, and the overall estimates may not be applicable to individual countries because of treatment effect heterogeneity. PRIMARY FUNDING SOURCE: Canadian Institutes of Health Research, World Health Organization, and Juravinski Research Institute.

Ceftobiprole for Treatment of MRSA Blood Stream Infection: A Case Series.

While methicillin-resistant Staphylococcus aureus (MRSA) bacteremia has poor outcomes, we describe our experience with Ceftobiprole mainly as a combination therapy for the treatment of MRSA bacteremia. All the cases of MRSA bacteremia in our center at the King Abdulaziz Medical City (KAMC), Riyadh, that had undergone Ceftobiprole treatment were included. We had six cases of MRSA bacteremia between 2018 and 2019, secondary to different infectious syndromes including endocarditis. There was a severe infection that required intensive care unit (ICU) admission in four cases. Ceftobiprole is used in combination with vancomycin in four cases. On day 14, all cases had a favorable outcome with microbiological and clinical improvement. However, three patients died after months of suffering from bacteremia from unrelated causes for the infection. The clinical outcome in our series of treatment of MRSA bacteremia using Ceftobiprole was favorable. Further studies for the evaluation of the use of Ceftobiprole in MRSA bacteremia should be encouraged.

E3 ligase RFWD3 is a novel modulator of stalled fork stability in BRCA2-deficient cells.

BRCA1/2 help maintain genomic integrity by stabilizing stalled forks. Here, we identify the E3 ligase RFWD3 as an essential modulator of stalled fork stability in BRCA2-deficient cells and show that codepletion of RFWD3 rescues fork degradation, collapse, and cell sensitivity upon replication stress. Stalled forks in BRCA2-deficient cells accumulate phosphorylated and ubiquitinated replication protein A (ubq-pRPA), the latter of which is mediated by RFWD3. Generation of this intermediate requires SMARCAL1, suggesting that it depends on stalled fork reversal. We show that in BRCA2-deficient cells, rescuing fork degradation might not be sufficient to ensure fork repair. Depleting MRE11 in BRCA2-deficient cells does block fork degradation, but it does not prevent fork collapse and cell sensitivity in the presence of replication stress. No such ubq-pRPA intermediate is formed in BRCA1-deficient cells, and our results suggest that BRCA1 may function upstream of BRCA2 in the stalled fork repair pathway. Collectively, our data uncover a novel mechanism by which RFWD3 destabilizes forks in BRCA2-deficient cells.

Emergence of a Bilaterally Symmetric Pattern from Chiral Components in the Planarian Epidermis.

Most animals exhibit mirror-symmetric body plans, yet the molecular constituents from which they are formed are often chiral. In planarian flatworms, centrioles are arranged in a bilaterally symmetric pattern across the ventral epidermis. Here, we found that this pattern is generated by a network of centrioles with prominent chiral asymmetric properties. We identify centriole components required for establishing asymmetric connections between centrioles and balancing their effects to align centrioles along polarity fields. SMED-ODF2, SMED-VFL1, and SMED-VFL3 affect the assembly of centriole appendages that tether cytoskeletal connectors to position the centrioles. We further show that the medio-lateral polarization of centrioles relies on mechanisms that are partly distinct on the left and right sides of the planarian body. Our findings shed light on how bilaterally symmetrical patterns can emerge from chiral cellular organizations.

Dynamic Polarization of the Multiciliated Planarian Epidermis between Body Plan Landmarks.

Polarity is a universal design principle of biological systems that manifests at all organizational scales, yet its coordination across scales remains poorly understood. Here, we make use of the extreme anatomical plasticity of planarian flatworms to probe the interplay between global body plan polarity and local cell polarity. Our quantitative analysis of ciliary rootlet orientation in the epidermis reveals a dynamic polarity field with head and tail as independent determinants of anteroposterior (A/P) polarization and the body margin as determinant of mediolateral (M/L) polarization. Mathematical modeling rationalizes the global polarity field and its response to experimental manipulations as superposition of separate A/P and M/L fields, and we identify the core PCP and Ft/Ds pathways as their molecular mediators. Overall, our study establishes a framework for the alignment of cellular polarity vectors relative to planarian body plan landmarks and establishes the core PCP and Ft/Ds pathways as evolutionarily conserved 2D-polarization module.

Liposomal Therapy Attenuates Dermonecrosis Induced by Community-Associated Methicillin-Resistant Staphylococcus aureus by Targeting α-Type Phenol-Soluble Modulins and α-Hemolysin.

Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA), typified by the pulse-field type USA300, is an emerging endemic pathogen that is spreading rapidly among healthy people. CA-MRSA causes skin and soft tissue infections, life-threatening necrotizing pneumonia and sepsis, and is remarkably resistant to many antibiotics. Here we show that engineered liposomes composed of naturally occurring sphingomyelin were able to sequester cytolytic toxins secreted by USA300 and prevent necrosis of human erythrocytes, peripheral blood mononuclear cells and bronchial epithelial cells. Mass spectrometric analysis revealed the capture by liposomes of phenol-soluble modulins, α-hemolysin and other toxins. Sphingomyelin liposomes prevented hemolysis induced by pure phenol-soluble modulin-α3, one of the main cytolytic components in the USA300 secretome. In contrast, sphingomyelin liposomes harboring a high cholesterol content (66 mol/%) were unable to protect human cells from phenol-soluble modulin-α3-induced lysis, however these liposomes efficiently sequestered the potent staphylococcal toxin α-hemolysin. In a murine cutaneous abscess model, a single dose of either type of liposomes was sufficient to significantly decrease tissue dermonecrosis. Our results provide further insights into the promising potential of tailored liposomal therapy in the battle against infectious diseases.

Synergy between conventional antibiotics and anti-biofilm peptides in a murine, sub-cutaneous abscess model caused by recalcitrant ESKAPE pathogens.

With the antibiotic development pipeline running dry, many fear that we might soon run out of treatment options. High-density infections are particularly difficult to treat due to their adaptive multidrug-resistance and currently there are no therapies that adequately address this important issue. Here, a large-scale in vivo study was performed to enhance the activity of antibiotics to treat high-density infections caused by multidrug-resistant Gram-positive and Gram-negative bacteria. It was shown that synthetic peptides can be used in conjunction with the antibiotics ciprofloxacin, meropenem, erythromycin, gentamicin, and vancomycin to improve the treatment outcome of murine cutaneous abscesses caused by clinical hard-to-treat pathogens including all ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Enterobacter cloacae) pathogens and Escherichia coli. Promisingly, combination treatment often showed synergistic effects that significantly reduced abscess sizes and/or improved clearance of bacterial isolates from the infection site, regardless of the antibiotic mode of action. In vitro data suggest that the mechanisms of peptide action in vivo include enhancement of antibiotic penetration and potential disruption of the stringent stress response.

Computer-aided Discovery of Peptides that Specifically Attack Bacterial Biofilms.

Biofilms represent a multicellular growth state of bacteria that are intrinsically resistant to conventional antibiotics. It was recently shown that a synthetic immunomodulatory cationic peptide, 1018 (VRLIVAVRIWRR-NH2), exhibits broad-spectrum antibiofilm activity but the sequence determinants of antibiofilm peptides have not been systematically studied. In the present work, a peptide library consisting of 96 single amino acid substituted variants of 1018 was SPOT-synthesized on cellulose arrays and evaluated against methicillin resistant Staphylococcus aureus (MRSA) biofilms. This dataset was used to establish quantitative structure-activity relationship (QSAR) models relating the antibiofilm activity of these peptides to hundreds of molecular descriptors derived from their sequences. The developed 3D QSAR models then predicted the probability that a peptide would possess antibiofilm activity from a library of 100,000 virtual peptide sequences in silico. A subset of these variants were SPOT-synthesized and their activity assessed, revealing that the QSAR models resulted in ~85% prediction accuracy. Notably, peptide 3002 (ILVRWIRWRIQW-NH2) was identified that exhibited an 8-fold increased antibiofilm potency in vitro compared to 1018 and proved effective in vivo, significantly reducing abscess size in a chronic MRSA mouse infection model. This study demonstrates that QSAR modeling can successfully be used to identify antibiofilm specific peptides with therapeutic potential.

New Perspectives in Biofilm Eradication.

Microbial biofilms, which are elaborate and highly resistant microbial aggregates formed on surfaces or medical devices, cause two-thirds of infections and constitute a serious threat to public health. Immunocompromised patients, individuals who require implanted devices, artificial limbs, organ transplants, or external life support and those with major injuries or burns, are particularly prone to become infected. Antibiotics, the mainstay treatments of bacterial infections, have often proven ineffective in the fight against microbes when growing as biofilms, and to date, no antibiotic has been developed for use against biofilm infections. Antibiotic resistance is rising, but biofilm-mediated multidrug resistance transcends this in being adaptive and broad spectrum and dependent on the biofilm growth state of organisms. Therefore, the treatment of biofilms requires drug developers to start thinking outside the constricted "antibiotics" box and to find alternative ways to target biofilm infections. Here, we highlight recent approaches for combating biofilms focusing on the eradication of preformed biofilms, including electrochemical methods, promising antibiofilm compounds and the recent progress in drug delivery strategies to enhance the bioavailability and potency of antibiofilm agents.

Remaining clinical issues in hepatitis C treatment.

Key advances in the evaluation and treatment of hepatitis C virus (HCV) infection have positively transformed the management and outcomes of those living with this chronic viral infection. Previously difficult-to-cure populations, including those coinfected with HIV infection, now enjoy similarly high success rates with interferon-free, orally administered direct-acting antiviral (DAA) therapies. Nonetheless, relevant unresolved clinical questions remain. The role and impact of viral resistance testing on treatment selection and outcome remain to be fully determined. The consequences of developing resistance while on DAA treatments that ultimately prove unsuccessful requires further evaluation. Optimal HCV management strategies in decompensated liver disease are unclear, and the role for ribavirin in DAA treatment-naïve and treatment-experienced patients is uncertain. A chief concern for those with cirrhosis relates to the risk for de novo and recurrent hepatocellular carcinoma among DAA recipients. In this article, we present and interpret current data and consider pragmatic, clinically useful options.

Ciprofloxacin-nitroxide hybrids with potential for biofilm control.

As bacterial biofilms display extreme tolerance to conventional antibiotic treatments, it has become imperative to develop new antibacterial strategies with alternative mechanisms of action. Herein, we report the synthesis of a series of ciprofloxacin-nitroxide conjugates and their corresponding methoxyamine derivatives in high yield. This was achieved by linking various nitroxides or methoxyamines to the secondary amine of the piperazine ring of ciprofloxacin using amide bond coupling. Biological evaluation of the prepared compounds on preformed P. aeruginosa biofilms in flow cells revealed substantial dispersal with ciprofloxacin-nitroxide hybrid 25, and virtually complete killing and removal (94%) of established biofilms in the presence of ciprofloxacin-nitroxide hybrid 27. Compounds 25-28 were shown to be non-toxic in both human embryonic kidney 293 (HEK 293) cells and human muscle rhabdomyosarcoma (RD) cells at concentrations up to 40 µM. Significantly, these hybrids demonstrate the potential of antimicrobial-nitroxide agents to overcome the resistance of biofilms to antimicrobials via stimulation of biofilm dispersal or through direct cell killing.

New Mouse Model for Chronic Infections by Gram-Negative Bacteria Enabling the Study of Anti-Infective Efficacy and Host-Microbe Interactions.

Only a few, relatively cumbersome animal models enable long-term Gram-negative bacterial infections that mimic human situations, where untreated infections can last for weeks. Here, we describe a simple murine cutaneous abscess model that enables chronic or progressive infections, depending on the subcutaneously injected bacterial strain. In this model, Pseudomonas aeruginosa cystic fibrosis epidemic isolate LESB58 caused localized high-density skin and soft tissue infections and necrotic skin lesions for up to 10 days but did not disseminate in either CD-1 or C57BL/6 mice. The model was adapted for use with four major Gram-negative nosocomial pathogens, Acinetobacter baumannii, Klebsiella pneumoniae, Enterobacter cloacae, and Escherichia coli This model enabled noninvasive imaging and tracking of lux-tagged bacteria, the influx of activated neutrophils, and production of reactive oxygen-nitrogen species at the infection site. Screening antimicrobials against high-density infections showed that local but not intravenous administration of gentamicin, ciprofloxacin, and meropenem significantly but incompletely reduced bacterial counts and superficial tissue dermonecrosis. Bacterial RNA isolated from the abscess tissue revealed that Pseudomonas genes involved in iron uptake, toxin production, surface lipopolysaccharide regulation, adherence, and lipase production were highly upregulated whereas phenazine production and expression of global activator gacA were downregulated. The model was validated for studying virulence using mutants of more-virulent P. aeruginosa strain PA14. Thus, mutants defective in flagella or motility, type III secretion, or siderophore biosynthesis were noninvasive and suppressed dermal necrosis in mice, while a strain with a mutation in the bfiS gene encoding a sensor kinase showed enhanced invasiveness and mortality in mice compared to controls infected with wild-type P. aeruginosa PA14.IMPORTANCE More than two-thirds of hospital infections are chronic or high-density biofilm infections and difficult to treat due to adaptive, multidrug resistance. Unfortunately, current models of chronic infection are technically challenging and difficult to track without sacrificing animals. Here we describe a model of chronic subcutaneous infection and abscess formation by medically important nosocomial Gram-negative pathogens that is simple and can be used for tracking infections by imaging, examining pathology and immune responses, testing antimicrobial treatments suitable for high-density bacterial infections, and studying virulence. We propose that this mouse model can be a game changer for modeling hard-to-treat Gram-negative bacterial chronic and skin infections.

Alternative strategies for the study and treatment of clinical bacterial biofilms.

Biofilms represent an adaptive lifestyle where microbes grow as structured aggregates in many different environments, e.g. on body surfaces and medical devices. They are a profound threat in medical (and industrial) settings and cause two-thirds of all infections. Biofilm bacteria are especially recalcitrant to common antibiotic treatments, demonstrating adaptive multidrug resistance. For this reason, novel methods to eradicate or prevent biofilm infections are greatly needed. Recent advances have been made in exploring alternative strategies that affect biofilm lifestyle, inhibit biofilm formation, degrade biofilm components and/or cause dispersal. As such, naturally derived compounds, molecules that interfere with bacterial signaling systems, anti-biofilm peptides and phages show great promise. Their implementation as either stand-alone drugs or complementary therapies has the potential to eradicate resilient biofilm infections. Additionally, altering the surface properties of indwelling medical devices through bioengineering approaches has been examined as a method for preventing biofilm formation. There is also a need for improving current biofilm detection methods since in vitro methods often do not accurately measure live bacteria in biofilms or mimic in vivo conditions. We propose that the design and development of novel compounds will be enabled by the improvement and use of appropriate in vitro and in vivo models.

Antimicrobial Peptides: An Introduction.

The "golden era" of antibiotic discovery has long passed, but the need for new antibiotics has never been greater due to the emerging threat of antibiotic resistance. This urgency to develop new antibiotics has motivated researchers to find new methods to combat pathogenic microorganisms resulting in a surge of research focused around antimicrobial peptides (AMPs; also termed host defense peptides) and their potential as therapeutics. During the past few decades, more than 2000 AMPs have been identified from a diverse range of organisms (animals, fungi, plants, and bacteria). While these AMPs share a number of common features and a limited number of structural motifs; their sequences, activities, and targets differ considerably. In addition to their antimicrobial effects, AMPs can also exhibit immunomodulatory, anti-biofilm, and anticancer activities. These diverse functions have spurred tremendous interest in research aimed at understanding the activity of AMPs, and various protocols have been described to assess different aspects of AMP function including screening and evaluating the activities of natural and synthetic AMPs, measuring interactions with membranes, optimizing peptide function, and scaling up peptide production. Here, we provide a general overview of AMPs and introduce some of the methodologies that have been used to advance AMP research.

Bacterial Abscess Formation Is Controlled by the Stringent Stress Response and Can Be Targeted Therapeutically.

Cutaneous abscess infections are difficult to treat with current therapies and alternatives to conventional antibiotics are needed. Understanding the regulatory mechanisms that govern abscess pathology should reveal therapeutic interventions for these recalcitrant infections. Here we demonstrated that the stringent stress response employed by bacteria to cope and adapt to environmental stressors was essential for the formation of lesions, but not bacterial growth, in a methicillin resistant Staphylococcus aureus (MRSA) cutaneous abscess mouse model. To pharmacologically confirm the role of the stringent response in abscess formation, a cationic peptide that causes rapid degradation of the stringent response mediator, guanosine tetraphosphate (ppGpp), was employed. The therapeutic application of this peptide strongly inhibited lesion formation in mice infected with Gram-positive MRSA and Gram-negative Pseudomonas aeruginosa. Overall, we provide insights into the mechanisms governing abscess formation and a paradigm for treating multidrug resistant cutaneous abscesses.

Synthesis and Evaluation of Ciprofloxacin-Nitroxide Conjugates as Anti-Biofilm Agents.

As bacterial biofilms are often refractory to conventional antimicrobials, the need for alternative and/or novel strategies for the treatment of biofilm related infections has become of paramount importance. Herein, we report the synthesis of novel hybrid molecules comprised of two different hindered nitroxides linked to the piperazinyl secondary amine of ciprofloxacin via a tertiary amine linker achieved utilising reductive amination. The corresponding methoxyamine derivatives were prepared alongside their radical-containing counterparts as controls. Subsequent biological evaluation of the hybrid compounds on preformed P. aeruginosa flow cell biofilms divulged significant dispersal and eradication abilities for ciprofloxacin-nitroxide hybrid compound 10 (up to 95% eradication of mature biofilms at 40 µM). Importantly, these hybrids represent the first dual-action antimicrobial-nitroxide agents, which harness the dispersal properties of the nitroxide moiety to circumvent the well-known resistance of biofilms to treatment with antimicrobial agents.

Cell Competition Modifies Adult Stem Cell and Tissue Population Dynamics in a JAK-STAT-Dependent Manner.

Throughout their lifetime, cells may suffer insults that reduce their fitness and disrupt their function, and it is unclear how these potentially harmful cells are managed in adult tissues. We address this question using the adult Drosophila posterior midgut as a model of homeostatic tissue and ribosomal Minute mutations to reduce fitness in groups of cells. We take a quantitative approach combining lineage tracing and biophysical modeling and address how cell competition affects stem cell and tissue population dynamics. We show that healthy cells induce clonal extinction in weak tissues, targeting both stem and differentiated cells for elimination. We also find that competition induces stem cell proliferation and self-renewal in healthy tissue, promoting selective advantage and tissue colonization. Finally, we show that winner cell proliferation is fueled by the JAK-STAT ligand Unpaired-3, produced by Minute(-/+) cells in response to chronic JNK stress signaling.

High throughput screening methods for assessing antibiofilm and immunomodulatory activities of synthetic peptides.

The recent observation that certain cationic peptides possess potent antibiofilm activity demonstrated that small peptides could be used to treat biofilm-associated infections. Other so-called innate defense regulator peptides possess potent immunomodulatory properties such as leukocyte recruitment and suppression of harmful inflammation. A peptide that directly targets biofilm cells while favorably modulating the immune response would be particularly advantageous for treating serious skin infections caused by Staphylococcus aureus. In the present work, using SPOT-synthesized peptide arrays on cellulose membranes, we outline a strategy for systematically assessing the antibiofilm activity of hundreds of IDR-1002 (VQRWLIVWRIRK-NH2) and IDR-HH2 (VQLRIRVAVIRA-NH2) peptide variants against MRSA biofilms. In addition, the ability of these peptides to stimulate production of a monocyte chemoattractant protein (MCP-1) and suppress LPS-induced interleukin (IL)-1ß production in human peripheral blood mononuclear cells (PBMCs) was evaluated. These results informed the synthesis of second-generation peptides resulting in a new peptide, IDR-2009 (KWRLLIRWRIQK-NH2), with enhanced MCP-1 stimulatory activity, favorable IL-1ß suppression characteristics and strong antibiofilm activity against MRSA and Pseudomonas aeruginosa biofilms. This work provides a proof-of-concept that multiple peptide activities can be optimized simultaneously to generate novel sequences that possess a variety of biological properties.