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Lactococcus lactis (L. lactis), the first genetically modified Generally Recognized As Safe (GRAS) category Lactic Acid producing Bacteria (LAB), is best known for its generalized health-promoting benefits and ability to express heterologous proteins. However, achieving the optimal probiotic effects requires a selective approach that would allow us to study in vivo microbial biodistribution, fate, and immunological consequences. Although the chemical conjugation of fluorophores and chromophores represent the standard procedure to tag microbial cells for various downstream applications, it requires a high-throughput synthesis scheme, which is often time-consuming and expensive. On the contrary, the genetic manipulation of LAB vector, either chromosomally or extra-chromosomally, to express bioluminescent or fluorescent reporter proteins has greatly enhanced our ability to monitor bacterial transit through a complex gut environment. However, with faster passage and quick washing out from the gut due to rhythmic contractions of the digestive tract, real-time tracking of LAB vectors, particularly non-commensal ones, remains problematic. To get a deeper insight into the biodistribution of non-commensal probiotic bacteria in vivo, we bioengineered L. lactis to express fluorescence reporter proteins, mCherry (bright red monomeric fluorescent protein) and mEGFP (monomeric enhanced green fluorescent protein), followed by microencapsulation with a mucoadhesive and biodegradable polymer, chitosan. We show that coating of recombinant Lactococcus lactis (rL. lactis) with chitosan polymer, cross-linked with tripolyphosphate (TPP), retains their ability to express the reporter proteins stably without altering the specificity and sensitivity of fluorescence detection in vitro and in vivo. Further, we provide evidence of enhanced intragastric stability by chitosan-TPP (CS) coating of rL. lactis cells, allowing us to study the spatiotemporal distribution for an extended time in the gut of two unrelated hosts, avian and murine. The present scheme involving genetic modification and chitosan encapsulation of non-commensal LAB vector demonstrates great promise as a non-invasive and intensive tool for active live tracking of gut microbes.
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Lactococcus lactis , Proteínas Luminiscentes , Lactococcus lactis/genética , Lactococcus lactis/metabolismo , Animales , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Vectores Genéticos , Genes Reporteros , Ratones , Probióticos , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Proteína Fluorescente RojaRESUMEN
The targeted depletion of potential gut pathogens is often challenging because of their intrinsic ability to thrive in harsh gut environments. Earlier, we showed that Campylobacter jejuni (C. jejuni) exclusively uses the Type-VI Secretion System (T6SS) to target its prey such as Escherichia coli (E. coli), and phenotypic differences between T6SS-negative and T6SS-positive C. jejuni isolates toward bile salt sensitivity. However, it remains unclear how the target-driven T6SS functionality prevails in a polymicrobial gut environment. Here, we investigated the fate of microbial competition in an altered gut environment via bacterial T6SS using a T6SS-negative and -positive C. jejuni or its isogenic mutant of the hemolysin-coregulated protein (hcp). We showed that in the presence of bile salt and prey bacteria (E. coli), T6SS-positive C. jejuni experiences enhanced intracellular stress leading to cell death. Intracellular tracking of fluorophore-conjugated bile salts confirmed that T6SS-mediated bile salt influx into C. jejuni can enhance intracellular oxidative stress, affecting C. jejuni viability. We further investigated whether the T6SS activity in the presence of prey (E. coli) perturbs the in vivo colonization of C. jejuni. Using chickens as primary hosts of C. jejuni and non-pathogenic E. coli as prey, we showed a marked reduction of C. jejuni load in chickens cecum when bile salt solution was administered orally. Analysis of local antibody responses and pro-inflammatory gene expression showed a reduced risk of tissue damage, indicating that T6SS activity in the complex gut environment can be exploited as a possible measure to clear the persistent colonization of C. jejuni in chickens.
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The increasing concentration of CO2 and CH4 in the environment is a global concern. Tri-reforming of methane (TRM) is a promising route for the conversion of these two greenhouse gases to more valuable synthesis gas with an H2/CO ratio of 1.5-2. In this study, a series of Zr-MOF synthesized via the solvothermal method and impregnation technique was used to synthesize the nickel impregnated on MOF-derived ZrO2 catalyst. The catalyst was characterized by various methods, including N2-porosimetry, X-ray diffraction (XRD), temperature programmed reduction (TPR), CO2-temperature programmed desorption (CO2-TPD), thermo-gravimetric analysis (TGA), chemisorption, field-emission scanning electron microscopy (FE-SEM), and high-resolution transmission electron microscopy (HR-TEM). Characterization results confirmed the formation of the Zr-MOF and nickel metal dispersed on MOF-derived ZrO2. Further, the tri-reforming activity of the catalyst developed was evaluated in a downflow-packed bed reactor. The various catalysts were screened for TRM activity at different temperatures (600-850 °C). Results demonstrated that TRM was highly favorable over the NZ-1000 catalyst due to its desirable physicochemical properties, including nickel metal surface area (2.3 m2/gcat-1), metal dispersion (7.1%), and nickel metal reducibility (45%), respectively. Over the NZ-1000 catalyst, an optimum H2/CO ratio of ~ 1.6-2 was achieved at 750 °C, and it was stable for a longer period of time.
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Metano , Níquel , Circonio , Metano/química , Catálisis , Níquel/química , Circonio/química , Estructuras Metalorgánicas/química , Dióxido de Carbono/química , Difracción de Rayos XRESUMEN
The application of transition metal complexes for antimicrobial photodynamic therapy (PDT) has emerged as an attractive alternative in mitigating a broad range of bacterial pathogens, including multidrug-resistant pathogens. In view of their photostability, long excited-state lifetimes, and tunable emission properties, transition metal complexes also contribute as bioimaging agents. In the present work, we designed mono and trinuclear cyclometalated iridium (III) complexes to explore their imaging application and antibacterial potential. For this, we used Methicillin-resistant S. aureus (MRSA), the most prevalent of community-associated (CA) multidrug-resistant (MDR) bacteria (CA MDR) and Lactococcus lactis (L. lactis) as Gram-positive while Campylobacter jejuni (C. jejuni) and E. coli as Gram-negative bacteria. In addition to differential bioimaging of these bacteria, we assessed the antibacterial effects of both mono and trinuclear Ir(III) complexes under exposure to 427â nm LED light. The data presented herein strongly suggest better efficacy of trinuclear Ir(III) complex over the mononuclear complex in imparting photoinduced cell death of MRSA. Based on the safety profile of these complexes, we propose that trinuclear cyclometalated iridium(III) complex holds great promise for selective recognition and targeting MDR bacteria with minimal off-target effect.
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Antibacterianos , Complejos de Coordinación , Iridio , Staphylococcus aureus Resistente a Meticilina , Fotoquimioterapia , Fármacos Fotosensibilizantes , Iridio/química , Iridio/farmacología , Complejos de Coordinación/química , Complejos de Coordinación/farmacología , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Antibacterianos/farmacología , Antibacterianos/química , Fármacos Fotosensibilizantes/química , Fármacos Fotosensibilizantes/farmacología , Pruebas de Sensibilidad Microbiana , Luz , Escherichia coli/efectos de los fármacos , HumanosRESUMEN
Background Surgical site infections (SSIs), the third most common nosocomial infection, endanger hospitals and patients. SSIs must be monitored continuously. This present study examined SSI incidence, risk factors, pathogens, and antibiotic sensitivity in emergency and elective or planned abdominal surgeries. Methods The Dr. S.N. Medical College General Surgery Department in Jodhpur, India, operated on 100 patients. The sample was divided into two 50-person groups. Group A includes emergency surgery patients, while Group B includes elective surgery patients. The samples were aseptically collected and processed according to microbiological methods. Data were processed with IBM SPSS Statistics for Windows, version 20 (released 2011; IBM Corp., Armonk, New York, United States). Results Out of a sample size of 100 patients, 17 individuals experienced SSIs. SSI incidence was 16.66% in male patients and 18.18% in female patients. In addition, the rate of SSIs was 26% in the emergency group and 8% in the planned group. The association was stronger among elderly individuals, diabetics (33.33% in Group A and 12.5% in Group B), and anemics with a history of smoking. The association was higher in those who underwent surgery for more than 60 minutes (34.37% in Group A and 18.8% in Group B). The incidence of SSIs was higher in emergency cases compared to elective surgeries, with rates of 26% and 8%, respectively, but was statistically insignificant. The infection rate in clean cases during planned surgery was 3.70%, while clean contaminated cases during planned surgery had a wound infection rate of approximately 13.04%. In emergency surgery, no clean case was operated on, but the SSI rate in the emergency group was 9.09%, 22.22%, and 47.36% in the clean-contaminated, contaminated, and dirty cases, respectively. In Group A, Escherichia coli was the predominant organism found in SSI wounds, while in Group B, Staphylococcus aureus was the predominant organism, accounting for 46.15% and 50% of infections, respectively. Amikacin and metronidazole exhibited the highest efficacy against E. coli, with amikacin demonstrating the highest sensitivity. Conclusion SSIs are more common in emergencies than planned procedures. Age, gender, diabetes, hypertension, smoking, and prolonged surgery are risk factors for SSIs. Effective antibiotic policy and infection control can greatly prevent SSIs.
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INTRODUCTION: Staghorn calculi (SC) occupy the renal pelvis and calyces and are common in females linked to repeated urinary tract infections (UTIs). Judicious surgery planning reduces the chance of further damage to the kidney due to renal SCs. Open stone surgery (OSS) is one of the various operative techniques to remove such huge stones with one operative intervention and protect the kidney from ongoing functional damage. CASE REPORT: A 47-year-old male patient presented with right-sided renal colic pain, and on further investigations, he was diagnosed with a large renal stone responsible for substantial renal function impairment on the same side. The SC measured 8 × 4 cm with another stone in the lower calyx. thinner parenchyma, and only 16% relative function. Therefore, open surgery was selected over less invasive approaches because multiple lithotripsy (ESWL) sittings may have been required in less invasive options. DISCUSSION: SCs, which can be complete or partial, often result in renal impairment. Hence, it is crucial to implement a proactive therapeutic approach that includes a thorough evaluation of the stone's size and position, the patient's choice, and institutional capacity. Complete elimination of SCs is preferred to maintain maximal renal function. Based on clinical, technical, and socioeconomic considerations, open pyelolithotomy or OSS was chosen over percutaneous nephrolithotomy for SC removal in the discussed case. CONCLUSION: The ability to remove large stones in a single intervention with open pyelolithotomy has been very effective due to its distinctive clinical presentation and pathological abnormalities.
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In this work, the aggregation-induced photoluminescence emission (AIPE) of three water-soluble heterobimetallic Ir-Pt complexes was reported with insight into their photophysical and electrochemical properties and imaging of bacterial cells. An alkyne appended Schiff's base L, bridges bis-cyclometalated iridium(III) and platinum(II) terpyridine centre. The Schiff's base (N-N fragment) serves as the ancillary ligand to the iridium(III) centre, while the alkynyl end is coordinated to platinum(II). The pH and ionic strength influence the aggregation kinetics of the alkynylplatinum(II) fragment, leading to metal-metal and π-π interactions with the emergence of a triplet metal-metal-to-ligand charge transfer (3MMLCT) emission. The excellent reversibility and photostability of aggregation-induced emission (AIE) of these aqua-friendly complexes were tested for their ability to sense and selectively image E. coli cells at various pH values.
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Iridio , Platino (Metal) , Iridio/química , Ligandos , Escherichia coli , Concentración de Iones de HidrógenoRESUMEN
Quantum chemistry calculations have been performed to access the efficacy of Cu-based catalysts in various mechanistic steps of the glycerol hydrogenolysis reaction. Calculations are first performed for reactants in the gas phase (noncatalyzed system) and reactants in the gas phase with a 3-atom Cu cluster (catalyzed system). We demonstrate that the glycerol to ethylene glycol conversion is preferred in the noncatalyzed system but glycerol conversion to 1,2-propanediol via the 2-acetol intermediate is preferred in the catalyzed system. We next analyze the adsorption energies of the reactant and product species involved in the glycerol to 1,2-PDO reaction on an 8-atom Cu cluster and Cu cluster doped with a Zn atom or a ZnO molecule. Finally, we study the effects of Zn or ZnO doping on the activation barriers of the two steps of the glycerol to 1,2-PDO reaction.
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The bacterial Type VI Secretion System (T6SS) functions as a nanomachine used by many gut pathogens. In the present protocol, we outlined how such molecular activities during interspecies interaction can be demonstrated at a population level. To this end, we first present a comprehensive protocol for isolation, identification, and functional characterization of T6SS-positive Campylobacter jejuni. Further, we developed straightforward techniques for unraveling how the T6SS targets prey populations and host cells when growing with or without environmental stressors. For complete details on the use and execution of this protocol, please refer to Gupta et al. (2021).