Biology subjects

Lukomski, S.

Publications and source records attributed to Lukomski, S..

4 recordsLinked to original sources

bioRxiv2024

Group A Streptococcal Collagen-like Protein 1 Restricts Tumor Growth in Murine Pancreatic Adenocarcinoma and Inhibits Cancer-Promoting Neutrophil Extracellular Traps

Pancreatic ductal adenocarcinoma (PDAC) is a lethal cancer associated with an immunosuppressive environment. Neutrophil extracellular traps (NETs) were initially described in the context of infection but have more recently been implicated in contributing to the tolerogenic immune response in PDAC. Thus, NETs are an attractive target for new therapeutic strategies. Group A Streptococcus (GAS) has developed defensive strategies to inhibit NETs. In the present work, we propose utilizing intra-tumoral GAS injection to stimulate anti-tumor activity by inhibiting cancer-promoting NETs. Injection of three different M-type GAS strains reduced subcutaneous pancreatic tumor volume compared to control in two different murine PDAC models. Limitation of tumor growth was dependent on streptococcal collagen-like protein 1 (Scl1), as isogenic mutant strain devoid of Scl1 did not reduce tumor size. We further show that Scl1 plays a role in localizing GAS to the tumor site, thereby limiting the systemic spread of bacteria and off-target effects. While mice did elicit a humoral immune response to GAS antigens, tested sera were negative toward Scl1 antigen following intra-tumoral treatment with Scl1-expressing GAS. M1 GAS inhibited NET formation when co-cultured with neutrophils while Scl1-devoid mutant strain did not. Recombinant Scl1 protein inhibited NETs ex vivo in a dose-dependent manner by suppressing myeloperoxidase activity. Altogether, we demonstrate that intra-tumoral GAS injections reduce PDAC growth, which is facilitated by Scl1, in part through inhibition of cancer promoting NETs. This work offers a novel strategy by which NETs can be targeted through Scl1 protein and potentiates its use as a cancer therapeutic.

cancer biology↗

bioRxiv2023

Prevalence of Erythromycin Resistant emm92-type Invasive Group A Streptococcal Infections in West Virginia, United States, 2021-2023

BackgroundIncreasing incidence of invasive group A Streptococcus (iGAS) disease has been reported in Europe and United States over the past several years. Coupled with this are observations of higher rates of resistance to non-beta lactam antimicrobials. ObjectivesThe aim of this study was to characterize iGAS and pharyngitis isolates from West Virginia (WV), a region outside of the US national active bacteria core surveillance purview, where risk factors associated with iGAS infections are prevalent. MethodsSeventy-seven invasive group A Streptococcus isolates were collected from sixty-seven unique patients at the J.W. Ruby Memorial Hospital Clinical Microbiology Laboratory in WV from 2021-23. Invasive isolates and twenty unique pharyngitis isolates were tested for clindamycin and erythromycin susceptibilities in the clinical laboratory. Patient demographic and clinical information was retrieved from patient electronic health records. Isolates were further characterized based on emm-type and detection of MLSB resistance determinants. ResultsTwenty-six (39%) isolates were of a single emm-type, emm92. All emm92 isolates were uniformly erythromycin/clindamycin resistant with inducible or constitutive MLSB resistance imparted by the plasmid-borne erm(T) gene. The majority of emm92 infections were associated with adult patients who reported intravenous drug use, whereas no pharyngitis infections were caused by an emm92 strain. Overall, fifty-one (76%) of the sixty-seven iGAS isolates were determined to carry MLSB resistance. ConclusionsIsolates of emm-type 92 predominated in this collection, were uniformly erythromycin/clindamycin resistant, and were associated with adult intravenous drug use but not with pediatric pharyngitis.

microbiology↗

bioRxiv2022

Emergence of Erythromycin Resistant Invasive Group A Streptococcus in West Virginia, United States

Combination therapy with penicillin and clindamycin has been a mainstay for treatment of invasive group A Streptococcus (iGAS) infections yet increasing macrolide resistance may limit such treatment for strains displaying MLSB phenotypes. The CDC recently reported erythromycin and clindamycin resistance rates for iGAS exceeding 20% in 2017. Here, we investigated 76 iGAS isolates from 66 patients identified at J.W. Ruby Memorial Hospital in West Virginia from 2020-2021. emm typing was performed using the CDC protocol and database. Resistance genes were detected by PCR and sequencing, whereas antimicrobial susceptibility testing was performed in clinical and research laboratories with standard techniques. Median patient age was 42 years (23-86 range). 76% (n=50) of isolates were simultaneously resistant to erythromycin and clindamycin, which included both inducible (n=40) and constitutive (n=9) resistance. All emm92 (n=35) and emm11 (n=8) isolates were erythromycin resistant, while the remaining 11% (n=7) of resistant isolates comprised 5 emm types. Susceptible isolates primarily included emm89 (n=6) iGAS. Macrolide resistance was conferred by the plasmid-borne ermT gene in all emm92 isolates and by chromosomally-encoded ermA (n=7), ermB (n=7), and mefA (n=1) in other emm types. Macrolide-resistant iGAS were typically resistant to tetracycline and aminoglycoside antibiotics. Here, we characterized iGAS infections affecting non-pediatric residents across West Virginia. We showed a shift in emm-type distribution compared to historical and national reports, and dominance of macrolide-resistant isolates which raises concern for emerging resistance to commonly-prescribed antibiotics used in treatment of iGAS infections.

microbiology↗

bioRxiv2020

Burkholderia collagen-like protein 8, Bucl8, is a unique outer membrane component of a tetrapartite efflux pump in Burkholderia pseudomallei and Burkholderia mallei

Bacterial efflux pumps are an important pathogenicity trait because they extrude a variety of xenobiotics. Our laboratory previously identified in silico Burkholderia collagen-like protein 8 (Bucl8) in the Tier one select agents Burkholderia pseudomallei and Burkholderia mallei. We hypothesize that Bucl8, which contains two predicted tandem outer membrane efflux pump domains, is a component of a putative efflux pump. Unique to Bucl8, as compared to other outer membrane proteins, is the presence of an extended extracellular region containing a collagen-like (CL) domain and a non-collagenous C-terminus (Ct). Molecular modeling and circular dichroism spectroscopy with a recombinant protein, corresponding to this extracellular CL-Ct portion of Bucl8, demonstrated that it adopts a collagen triple helix, whereas functional assays screening for Bucl8 ligands identified binding to fibrinogen. Bioinformatic analysis of the bucl8 gene locus revealed it resembles a classical efflux-pump operon. The bucl8 gene is co-localized with downstream fusCDE genes encoding fusaric acid (FA) resistance, and with an upstream gene, designated as fusR, encoding a LysR-type transcriptional regulator. Using RT-qPCR, we defined the boundaries and transcriptional organization of the fusR-bucl8-fusCDE operon. We found exogenous FA induced bucl8 transcription over 80-fold in B. pseudomallei, while deletion of the entire bucl8 locus decreased the MIC of FA 4-fold in its isogenic mutant. We furthermore showed that the Bucl8 pump expressed in the heterologous Escherichia coli host confers FA resistance. On the contrary, the Bucl8 pump did not confer resistance to a panel of clinically-relevant antimicrobials in Burkholderia and E. coli. We finally demonstrated that deletion of the bucl8-locus drastically affects the growth of the mutant in L-broth. We determined that Bucl8 is a component of a novel tetrapartite efflux pump, which confers FA resistance, fibrinogen binding, and optimal growth. Author SummaryBurkholderia pseudomallei and Burkholderia mallei are highly infectious and multidrug resistant bacteria that are classified by the National Institute of Allergy and Infectious Diseases as Tier one select agents partly due to the intrinsic multidrug resistance associated with expression of the efflux pumps. To date, only few efflux pumps predicted in Burkholderia spp. have been studied in detail. In the current study we introduce Bucl8, an outer membrane component of an unreported putative efflux pump with a unique extended extracellular portion that forms a collagen triple helix and binds fibrinogen. We demonstrate Bucl8s role in fusaric acid resistance by defining its operon via bioinformatic and transcriptional analyses, as well as by employing loss-of-function and gain-of-function genetic approaches. Our studies also implicate the Bucl8-associated pump in metabolic and physiologic homeostasis. Understanding how Bucl8 efflux pump contributes to Burkholderia pathology will foster development of pump inhibitors targeting transport mechanism or identifying potential surface-exposed vaccine targets.

microbiology↗