Initial diet shapes resistance-gene composition and fecal microbiome dynamics in young ruminants during nursing.

This study was conducted to examine how colostrum pasteurization affects resistance genes and microbial communities in calf feces. Forty female Holstein calves were randomly assigned to either the control (CON) group, which received unheated colostrum, or the pasteurized colostrum (PAT) group. The calves body weight was measured weekly before morning feeding. Calf starter intake were measured and recorded daily before morning feeding. Samples of colostrum were collected before feeding. Blood was collected on d 1 and 70 before morning feeding. Ten calves were randomly selected from each group (n = 20 calves total) for fecal sampling on d 3, 28, 56 and 70 for subsequent DNA extraction and metagenomic sequencing. Total bacterial counts in the colostrum were markedly higher in the CON group than in the PAT group. Pasteurized colostrum administration substantially reduced the ARO diversity and diminishes the abundance of Enterobacteriaceae, thereby decreasing their contribution to resistance genes. Pasteurization also reduced glucoside hydrolase-66 activity in 3-day-old calves which led to an increase in the activity of aminoglycoside antibiotics, resulting in 52.63 % of PAT-enriched bacteria acquiring aminoglycoside resistance genes. However, from the perspective of overall microbial community, the proportion of aminoglycoside, beta-lactam and tetracycline resistance genes carried by microbial community in PAT group was lower than CON group (P < 0.05). Fecal samples from the PAT group contained greater abundances of Subdoligranulum (P < 0.05) and Lachnospiraceae_NK4A136_group (P < 0.05) on days 28 and 70 compared to CON. Network analysis and abundance variations of the different bacteria obtained by linear discriminant analysis effect size analysis showed that pasteurized colostrum feeding reduced the interactions among related bacteria and maintained stability of the hind-gut microbiome. In conclusion, these findings underscore the intricate interactions between early diet, calf resistance-gene transmission and microbial dynamics, which should be carefully considered in calf-rearing practices.

Systematic Review: Clinical Features, Antimicrobial Treatment, and Outcomes of Human Tularemia, 1993-2023.

BACKGROUND: Francisella tularensis, the causative agent of tularemia, is endemic throughout the Northern Hemisphere and requires as few as 10 organisms to cause disease, making this potential bioterrorism agent one of the most infectious bacterial pathogens known. Aminoglycosides, tetracyclines, and, more recently, fluoroquinolones are used for treatment of tularemia; however, data on the relative effectiveness of these and other antimicrobial classes are limited. METHODS: Nine databases, including Medline, Global Health, and Embase, were systematically searched for articles containing terms related to tularemia. Articles with case-level data on tularemia diagnosis, antimicrobial treatment, and patient outcome were included. Patient demographics, clinical findings, antimicrobial administration, and outcome (eg, intubation, fatality) were abstracted using a standardized form. RESULTS: Of the 8878 publications identified and screened, 410 articles describing 870 cases from 1993 to 2023 met inclusion criteria. Cases were reported from 35 countries; more than half were from the United States, Turkey, or Spain. The most common clinical forms were ulceroglandular, oropharyngeal, glandular, and pneumonic disease. Among patients treated with aminoglycosides (n = 452 [52%]), fluoroquinolones (n = 339 [39%]), or tetracyclines (n = 419 [48%]), the fatality rate was 0.7%, 0.9%, and 1.2%, respectively. Patients with pneumonic disease who received ciprofloxacin had no fatalities and the lowest rates of thoracentesis/pleural effusion drainage and intubation compared to those who received aminoglycosides and tetracyclines. CONCLUSIONS: Aminoglycosides, fluoroquinolones, and tetracyclines are effective antimicrobials for treatment of tularemia, regardless of clinical manifestation. For pneumonic disease specifically, ciprofloxacin may have slight advantages compared to other antimicrobials.

Improving the Safety of Clinical Management of COVID-19 Patients Receiving Aminoglycosides and Parenteral Nutrition: Y-site Compatibility Studies.

PURPOSE: Aminoglycosides (AMGs) are broad-spectrum bactericidal antibiotics that can resolve bacterial infections co-existing with COVID-19 or exploit their potential antiviral activities. Patients presenting the most severe forms of COVID-19 due to escalating catabolism and significant lean body mass loss often require the concomitant administration of parenteral nutrition (PN) and antibiotics. The Y-site administration is one of the approaches allowing the co-administration of two intravenous medications in patients with limited vascular access. Our study aimed to investigate the compatibility of AMGs and selected commercial PN admixtures enriched in omega-3 fatty acids. METHODS: Gentamycin (GM), amikacin (AM), and tobramycin (TM) solutions for infusion were combined with Nutriflex Omega Special (NOS) and Smofkabiven (SFK). Three different volume ratios were investigated: 1:2, 1:1, and 2:1, simulating Y-site administration. Samples underwent visual examination and determination of the lipid emulsion particle size, zeta potential, and pH immediately after preparation and after four hours of storage at room temperature (22 ± 2 °C) with sunlight exposure. RESULTS: GM and AM combined with NOS in all studied ratios met the set-up acceptance criteria. The addition of TM to NOS in a 2:1 volume ratio and all tested AMGs to SFK in all studied combinations significantly influenced the stability of the oil-water system leading to the appearance of globules larger than 5 µm exceeding the pharmacopeial limit of 0.05% immediately after preparation or after four hours of storage. CONCLUSION: In conclusion, our study showed that NOS was less prone to destabilization of oil-in-water systems by AMGs than SFK. In justified clinical cases, due to the lack of appearance of precipitate or enlarged lipid droplets, the combined administration of GM and AM with the NOS could be considered, provided tested volume ratios of the drug and MCB in the infusion line are maintained. However, it should be noted that such an infusion may be associated with the risk of changes in the pharmacokinetics of the drug.

A novel ruthenium-silver based antimicrobial potentiates aminoglycoside activity against Pseudomonas aeruginosa.

The rapid dissemination of antibiotic resistance combined with the decline in the discovery of novel antibiotics represents a major challenge for infectious disease control that can only be mitigated by investments in novel treatment strategies. Alternative antimicrobials, including silver, have regained interest due to their diverse mechanisms of inhibiting microbial growth. One such example is AGXX, a broad-spectrum antimicrobial that produces highly cytotoxic reactive oxygen species (ROS) to inflict extensive macromolecular damage. Due to the connections identified between ROS production and antibiotic lethality, we hypothesized that AGXX could potentially increase the activity of conventional antibiotics. Using the gram-negative pathogen Pseudomonas aeruginosa, we screened possible synergistic effects of AGXX on several antibiotic classes. We found that the combination of AGXX and aminoglycosides tested at sublethal concentrations led to a rapid exponential decrease in bacterial survival and restored the sensitivity of a kanamycin-resistant strain. ROS production contributes significantly to the bactericidal effects of AGXX/aminoglycoside treatments, which is dependent on oxygen availability and can be reduced by the addition of ROS scavengers. Additionally, P. aeruginosa strains deficient in ROS detoxifying/repair genes were more susceptible to AGXX/aminoglycoside treatment. We further demonstrate that this synergistic interaction was associated with a significant increase in outer and inner membrane permeability, resulting in increased antibiotic influx. Our study also revealed that AGXX/aminoglycoside-mediated killing requires an active proton motive force across the bacterial membrane. Overall, our findings provide an understanding of cellular targets that could be inhibited to increase the activity of conventional antimicrobials. IMPORTANCE The emergence of drug-resistant bacteria coupled with the decline in antibiotic development highlights the need for novel alternatives. Thus, new strategies aimed at repurposing conventional antibiotics have gained significant interest. The necessity of these interventions is evident especially in gram-negative pathogens as they are particularly difficult to treat due to their outer membrane. This study highlights the effectiveness of the antimicrobial AGXX in potentiating aminoglycoside activities against P. aeruginosa. The combination of AGXX and aminoglycosides not only reduces bacterial survival rapidly but also significantly re-sensitizes aminoglycoside-resistant P. aeruginosa strains. In combination with gentamicin, AGXX induces increased endogenous oxidative stress, membrane damage, and iron-sulfur cluster disruption. These findings emphasize AGXX's potential as a route of antibiotic adjuvant development and shed light on potential targets to enhance aminoglycoside activity.

Space Station-like Composite Nanoparticles for Co-Delivery of Multiple Natural Compounds from Chinese Medicine and Hydrogen in Combating Sensorineural Hearing Loss.

Ototoxic drugs such as aminoglycoside antibiotics and cisplatin (CDDP) can cause sensorineural hearing loss (SNHL), which is closely related to oxidative stress and the acidification of the inner ear microenvironment. Effective treatment of SNHL often requires multifaceted approach due to the complex pathology, and drug combination therapy is expected to be at the forefront of modern hearing loss treatment. Here, space-station-like composite nanoparticles (CCC@mPP NPs) with pH/oxidation dual responsiveness and multidrug simultaneous delivery capability were constructed and then loaded with various drugs including panax notoginseng saponins (PNS), tanshinone IIA (TSIIA), and ammonia borane (AB) to provide robust protection against SNHL. Molecular dynamics simulation revealed that carboxymethyl chitosan/calcium carbonate-chitosan (CCC) NPs and monomethoxy poly(ethylene glycol)-PLGA (mPP) NPs can rendezvous and dock primarily by hydrogen bonding, and electrostatic forces may be involved. Moreover, CCC@mPP NPs crossed the round window membrane (RWM) and entered the inner ear through endocytosis and paracellular pathway. The docking state was basically maintained during this process, which created favorable conditions for multidrug delivery. This nanosystem was highly sensitive to pH and reactive oxygen species (ROS) changes, as evidenced by the restricted release of payload at alkaline condition (pH 7.4) without ROS, while significantly promoting the release in acidic condition (pH 5.0 and 6.0) with ROS. TSIIA/PNS/AB-loaded CCC@mPP NPs almost completely preserved the hair cells and remained the hearing threshold shift within normal limits in aminoglycoside- or CDDP-treated guinea pigs. Further experiments demonstrated that the protective mechanisms of TSIIA/PNS/AB-loaded CCC@mPP NPs involved direct and indirect scavenging of excessive ROS, and reduced release of pro-inflammatory cytokines. Both in vitro and in vivo experiments showed the high biocompatibility of the composite NPs, even after long-term administration. Collectively, this work suggests that composite NPs is an ideal multi-drug-delivery vehicle and open new avenues for inner ear disease therapies.

Overcoming biological barriers to improve treatment of a Staphylococcus aureus wound infection.

Chronic wounds frequently become infected with bacterial biofilms which respond poorly to antibiotic therapy. Aminoglycoside antibiotics are ineffective at treating deep-seated wound infections due to poor drug penetration, poor drug uptake into persister cells, and widespread antibiotic resistance. In this study, we combat the two major barriers to successful aminoglycoside treatment against a biofilm-infected wound: limited antibiotic uptake and limited biofilm penetration. To combat the limited antibiotic uptake, we employ palmitoleic acid, a host-produced monounsaturated fatty acid that perturbs the membrane of gram-positive pathogens and induces gentamicin uptake. This novel drug combination overcomes gentamicin tolerance and resistance in multiple gram-positive wound pathogens. To combat biofilm penetration, we examined the ability of sonobactericide, a non-invasive ultrasound-mediated-drug delivery technology to improve antibiotic efficacy using an in vivo biofilm model. This dual approach dramatically improved antibiotic efficacy against a methicillin-resistant Staphylococcus aureus (MRSA) wound infection in diabetic mice.

Berberine chloride protects cochlear hair cells from aminoglycoside-induced ototoxicity by reducing the accumulation of mitochondrial reactive oxygen species.

Aminoglycoside, a medicinal category of antibiotics, are used in treatment of Gram-negative bacterial infections. Although they are the most widely-used antibiotics due to their high efficacy and low cost, several main adverse effects have been reported including nephrotoxicity and ototoxicity. Since drug-induced ototoxicity is one of the major etiological causes of acquired hearing loss, we examined cochlear hair cell damages caused by three aminoglycosides (amikacin, kanamycin, and gentamicin), and investigated protective property of an isoquinoline-type alkaloid, Berberine chloride (BC). Berberine, a well-known bioactive compound found from medicinal plants, has been known to have anti-inflammatory, antimicrobial effects. To determine protective effect of BC in aminoglycoside-induced ototoxicity, hair cell damages in aminoglycoside- and/or BC-treated hair cells using ex vivo organotypic culture system of mouse cochlea. Mitochondrial ROS levels and depolarization of mitochondrial membrane potential were analyzed, and TUNEL assay and immunostaining of cleaved caspase-3 were performed to detect apoptosis signals. As the results, it was found that BC significantly prevented aminoglycoside-induced hair cell loss and stereocilia degeneration by inhibiting excessive accumulation of mitochondrial ROS and subsequent loss of mitochondrial membrane potential. It eventually inhibited DNA fragmentation and caspase-3 activation, which were significant for all three aminoglycosides. This study is the first report suggested the preventative effect of BC against aminoglycoside-induced ototoxicity. Our data also suggests a possibility that BC has the potential to exert a protective effect against ototoxicity caused by various ototoxic drugs leading to cellular oxidative stress, not limited to aminoglycoside antibiotics.

Bioenergetic State of Escherichia coli Controls Aminoglycoside Susceptibility.

Aminoglycosides (AG) have been used against Gram-negative bacteria for decades. Yet, how bacterial metabolism and environmental conditions modify AG toxicity is poorly understood. Here, we show that the level of AG susceptibility varies depending on the nature of the respiratory chain that Escherichia coli uses for growth, i.e., oxygen, nitrate, or fumarate. We show that all components of the fumarate respiratory chain, namely, hydrogenases 2 and 3, the formate hydrogenlyase complex, menaquinone, and fumarate reductase are required for AG-mediated killing under fumarate respiratory conditions. In addition, we show that the AAA+ ATPase RavA and its Von Wildebrand domain-containing partner, ViaA, are essential for AG to act under fumarate respiratory conditions. This effect was true for all AG that were tested but not for antibiotics from other classes. In addition, we show that the sensitizing effect of RavA-ViaA is due to increased gentamicin uptake in a proton motive force-dependent manner. Interestingly, the sensitizing effect of RavA-ViaA was prominent in poor energy conservation conditions, i.e., with fumarate, but dispensable under high energy conservation conditions, i.e., in the presence of nitrate or oxygen. We propose that RavA-ViaA can facilitate uptake of AG across the membrane in low-energy cellular states. IMPORTANCE Antibiotic resistance is a major public health, social, and economic problem. Aminoglycosides (AG) are known to be highly effective against Gram-negative bacteria, but their use is limited to life-threatening infections because of their nephrotoxicity and ototoxicity at therapeutic dose. Elucidation of AG-sensitization mechanisms in bacteria would allow reduced effective doses of AG. Here, we have identified the molecular components involved in anaerobic fumarate respiration that are required for AG to kill. In addition to oxidoreductases and menaquinone, this includes new molecular players, RavA, an AAA+ ATPase, and ViaA, its partner that has the VWA motif. Remarkably, the influence of RavA-ViaA on AG susceptibility varies according to the type of bioenergetic metabolism used by E. coli. This is a significant advance because anaerobiosis is well known to reduce the antibacterial activity of AG. This study highlights the critical importance of the relationship between culture conditions, metabolism, and antibiotic susceptibility.

Chenodeoxycholic Acid-Amikacin Combination Enhances Eradication of Staphylococcus aureus.

The rise of antibiotic resistance and dearth of novel antibiotics have posed a serious health crisis worldwide. In this study, we screened a combination of antibiotics and nonantibiotics providing a viable strategy to solve this issue by broadening the antimicrobial spectrum. We found that chenodeoxycholic acid (CDCA), a cholic acid derivative of the traditional Chinese medicine (TCM) Tanreqing (TRQ), synergizes with amikacin against Staphylococcus aureus in vitro, and this synergistic killing was effective against diverse methicillin-resistant S. aureus (MRSA) variants, including small-colony variants (SCVs), biofilm strains, and persisters. The CDCA-amikacin combination protects a mouse model from S. aureus infections. Mechanistically, CDCA increases the uptake of aminoglycosides in a proton motive force-dependent manner by dissipating the chemical potential and potentiates reactive oxygen species (ROS) generation by inhibiting superoxide dismutase activity. This work highlights the potential use of TCM components in treating S. aureus-associated infections and extend the use of aminoglycosides in eradicating Gram-positive pathogens. IMPORTANCE Multidrug resistance (MDR) is spreading globally with increasing speed. The search for new antibiotics is one of the key strategies in the fight against MDR. Antibiotic resistance breakers that may or may not have direct antibacterial action and can either be coadministered or conjugated with other antibiotics are being studied. To better expand the antibacterial spectrum of certain antibiotics, we identified one component from a traditional Chinese medicine, Tanreqing (TRQ), that increased the activity of aminoglycosides. We found that this so-called agent, chenodeoxycholic acid (CDCA), sensitizes Staphylococcus aureus to aminoglycoside killing and protects a mouse model from S. aureus infections. CDCA increases the uptake of aminoglycosides in a proton motive force-dependent manner by dissipating the chemical potential and potentiates ROS generation by inhibiting superoxide dismutase activity in S. aureus. Our work highlights the potential use of TCM or its effective components, such as CDCA, in treating antibiotic resistance-associated infections.

The efficacy and safety of ceftolozane-tazobactam in the treatment of GNB infections: a systematic review and meta-analysis of clinical studies.

BACKGROUND: Ceftolozane-tazobactam is a novel cephalosporin/ß-lactamase inhibitor combination with activity against Gram-negative bacteria (GNB). We aimed to comprehensively evaluate the clinical efficacy and safety of ceftolozane-tazobactam in treating GNB infections in adult patients. RESEARCH DESIGN AND METHODS: PubMed, Embase, and Cochrane databases were retrieved until August 2022. Randomized trials and non-randomized controlled studies evaluating ceftolozane-tazobactam and its comparators in adult patients with GNB infections were included. RESULTS: A total of 13 studies were included. Overall, patients receiving ceftolozane-tazobactam had significant advantages in clinical cure (odds ratio [OR], 1.62; 95% CI, 1.05-2.51) and microbiological eradication (OR, 1.43; 95% CI, 1.19-1.71), especially in Pseudomonas aeruginosa-infected patients. Ceftolozane-tazobactam had a significant advantage in clinical success or microbial eradication compared with polymyxin/aminoglycosides (PL/AG) or levofloxacin. There were no significant differences in adverse events (AEs), Clostridium difficile infection (CDI), and mortality between ceftolozane-tazobactam and comparators. Notably, ceftolozane-tazobactam showed a significantly lower risk of acute kidney injury compared with PL/AG. CONCLUSIONS: Ceftolozane-tazobactam showed excellent clinical and microbiological efficacy in treating GNB, especially P. aeruginosa-induced infections. The overall safety profile of ceftolozane-tazobactam was comparable to other antimicrobials, with no increased risk of CDI and obvious advantage over antibacterial agents with high nephrotoxicity.

Design, synthesis, and bioassay of 5-epi-aminoglycosides.

For the purpose of seeking new antibiotics, researchers usually modify the already-existing ones. However, this strategy has been extensively used and is close to its limits, especially in the case of aminoglycosides, and it is difficult to find a proper aminoglycoside antibiotic for novel modification. In this paper, we reported the design, synthesis, and evaluation of a series of 5-epi-neamine derivatives based on the structural information of bacterial 16S RNA A-site binding with aminoglycosides. Bioassay results showed that our design strategy was feasible. Our study offers a new way to search for structurally novel aminoglycosides. Meanwhile, our study provides valuable structure-activity relationship information, which will lead to better understanding and exploitation of the drug target, and improved development of new aminoglycoside antibiotics.

Structural changes in the human stria vascularis induced by aminoglycosides and loop diuretics.

The human stria vascularis has been examined both by scanning and transmission electron microscopy in normal controls and from individuals who had received loop diuretics, aminoglycoside antibiotics or some combination of the two prior to their deaths. The tissues were preserved by perilymphatic perfusion of fixative within an hour of death and preservation was adequate. The normal ultrastructure is described and does not differ significantly from that found in experimental animals. The loop diuretics are associated with structural changes that cannot be distinguished from those found in animals treated with large doses of the same drugs. The aminoglycosides caused some changes, but the patients had been in renal failure and this probably contributed to the structural alterations. The combination of a loop diuretic and aminoglycoside was associated with a range of alterations from mild to severe. Overall, the three treatment groups had a series of ultrastructural changes resembling those found in animal models thereby justifying the use of experimental animals to predict human ototoxicity.

Combining microbial and chemical syntheses for the production of complex natural products.

Nature, a rich source of bioactive natural products, serves as a massive pool of drug candidates for the pharmaceutical industry. However, the supply of these structurally complex chemicals is costly as most of the natural products are scarce in nature, thus requiring de novo synthesis. The supply chain issue hinders the development of novel therapeutic agents from natural products. Microbial synthesis, based on the expression of biosynthetic genes in a suitable microbial host to produce certain chemicals, is a sustainable strategy to produce complex natural products. However, this strategy requires gaining insights into the biosynthesis of target molecules. Most natural products are biosynthetically unknown or not fully elucidated; thus, the sole application of microbial synthesis strategy to produce a given molecule is challenging. In this review, we highlight a strategy that combines microbial and chemical syntheses to tackle the supply chain issue in developing drugs from natural products. We believe this strategy can revive the drug development pipeline for natural products.

Aminoglycoside-mimicking carbonized polymer dots for bacteremia treatment.

Bacteremia and associated bacterial sepsis are potentially fatal and occur when the host response to microbial invasion is impaired or compromised. This motivated us to develop carbonized polymer dots (CPDsMan/AA) from a mixture of mannose (Man) and positively charged amino acids [AAs; lysine, arginine (Arg), or histidine] through a one-step mild pyrolysis procedure, which effectively inhibited drug-resistant bacterial strains isolated from septic patients. The as-prepared CPDsMan/AA showed broad-spectrum antibacterial activity, including multidrug-resistant bacteria, even in human plasma. The minimal inhibitory concentration of CPDsMan/Arg is ca. 1.0 µg mL-1, which is comparable to or lower than those of other tested antibiotics (e.g., ampicillin, gentamicin, and vancomycin). In addition to directly disrupting bacterial membranes, the CPDsMan/Arg feature a structure similar to aminoglycoside antibiotics that could bind to 16S rRNA, thereby blocking bacterial protein synthesis. In vitro cytotoxic and hemolytic assays demonstrated the high biocompatibility of the CPDsMan/AA. In addition, in vivo studies on methicillin-resistant Staphylococcus aureus-infected mice treated with the CPDsMan/Arg showed a significant decrease in mortality-even better than that of antibiotics. Overall, the synthesis of the CPDsMan/AA is cost-efficient, straightforward, and effective for treating bacteremia. The polymeric features of the CPDsMan/Arg, including cationic charges and specific groups, can be recognized as a safe and broad-spectrum biocide to lessen our reliance on antibiotics to treat systemic bacterial infections in the future.

Formulation of Chrysomycin A Cream for the Treatment of Skin Infections.

Chrysomycin A, a compound derived from marine microorganisms, proved to have a specific great in vitro inhibitory effect on methicillin-resistant Staphylococcus aureus (MRSA). It exhibits high safety for the skin, as well as a better therapeutic effect than the current clinical drug, vancomycin. Nevertheless, its poor water solubility highly limits the application and reduces the bioavailability. In view of this, we developed a cream of chrysomycin A (CA) to enhance the solubility for the treatment of skin infection, while avoiding the possible toxicity caused by systemic administration. A comprehensive orthogonal evaluation system composed of appearance, spreading ability, and stability was established to find the optimal formula under experimental conditions. The final product was odorless and easy to be spread, with a lustrous, smooth surface. The particle size of the product met Chinese Pharmacopoeia specifications and the entire cream showed long-term stability in destructive tests. The in vitro and in vivo studies indicated that CA cream had a similar anti-MRSA activity to commercially available mupirocin, showing its potential as an efficacious topical delivery system for skin infections treatment.

Potential risks of treating bacterial infections with a combination of ß-lactam and aminoglycoside antibiotics: A systematic quantification of antibiotic interactions in E. coli blood stream infection isolates.

BACKGROUND: Treatment of Blood Stream Infections (BSIs) with a combination of a ß-lactam and an aminoglycoside antibiotic is widely used in intensive care units (ICUs) around the world. However, no studies have systematically examined how these drugs interact and potentially influence the antimicrobial efficacy of the overall treatment. METHODS: We collected 500 E. coli isolates from the Uppsala University hospital that were isolated from blood of patients with suspicion of infection. Of those we tested the efficacy of combinations of 2 common ß-lactam antibiotics (Ampicillin and Cefotaxime) combined with 2 common aminoglycosides (Gentamicin and Tobramycin) on 254 isolates. The efficacy of all 4 pairwise combinations in inhibiting bacterial growth was then examined on all susceptible strains. That was done by quantifying the Fractional Inhibitory index (FICi), a robust metric for antibiotic combinatorial behaviour, of all possible treatments on every strain. When non additive interactions were identified, results of the original screen were verified with time kill assays. Finally, combination behaviours were analysed for potential cross correlations. FINDINGS: Out of the 4 antibiotic combinations screened none exhibited synergistic effects on any of the 254 strains. On the contrary all 4 exhibited important antagonistic effects on several isolates. Specifically, the combinations of AMP-GEN and CTX-GEN were antagonistic in 1.97% and 1.18% of strains respectively. Similarly, the combinations of AMP-TOB were antagonistic on 0.78% of all strains. PCA analysis revealed that an important factor on the responses to the combination treatments was the choice of a specific aminoglycoside over another. Subsequent cross correlation analysis revealed that the interaction profiles of combinations including the same aminoglycoside are significantly correlated (Spearman's cross correlation test p<0.001). INTERPRETATION: The findings of this study elucidate potential risks of the common combination treatment for blood stream infections. They also demonstrate, previously unquantified metrics on how antibiotics in combination therapies are not interchangeable with others of the same class. Finally, they reiterate the need for case-by-case testing of antibiotic interactions in a clinical setting. FUNDING: This work was funded by grants to DIA from the Swedish Research Council, the Wallenberg foundation and the Swedish Strategic Research Foundation.

Evaluating Mono- and Combination Therapy of Meropenem and Amikacin against Pseudomonas aeruginosa Bacteremia in the Hollow-Fiber Infection Model.

Debate continues as to the role of combination antibiotic therapy for the management of Pseudomonas aeruginosa infections. We studied the extent of bacterial killing by and the emergence of resistance to meropenem and amikacin as monotherapies and as a combination therapy against susceptible and resistant P. aeruginosa isolates from bacteremic patients using the dynamic in vitro hollow-fiber infection model. Three P. aeruginosa isolates (meropenem MICs of 0.125, 0.25, and 64 mg/L) were used, simulating bacteremia with an initial inoculum of ~1 × 105 CFU/mL and the expected pharmacokinetics of meropenem and amikacin in critically ill patients. For isolates susceptible to amikacin and meropenem (isolates 1 and 2), the extent of bacterial killing was increased with the combination regimen compared with the killing by monotherapy of either antibiotic. Both the combination and meropenem monotherapy were able to sustain bacterial killing throughout the 7-day treatment course, whereas regrowth of bacteria occurred with amikacin monotherapy after 12 h. For the meropenem-resistant P. aeruginosa isolate (isolate 3), only the combination regimen demonstrated bacterial killing. Given that tailored antibiotic regimens can maximize potential synergy against some isolates, future studies should explore the benefit of combination therapy against resistant P. aeruginosa. IMPORTANCE Current guidelines recommend that aminoglycosides should be used in combination with ß-lactam antibiotics as initial empirical therapy for serious infections, and otherwise, patients should receive ß-lactam antibiotic monotherapy. Given the challenges associated with studying the clinical effect of different antibiotic strategies on patient outcomes, useful data for subsequent informed clinical testing can be obtained from in vitro models like the hollow-fiber infection model (HFIM). Based on the findings of our HFIM, we propose that the initial use of combination therapy with meropenem and amikacin provides some bacterial killing against carbapenem-resistant P. aeruginosa isolates. For susceptible isolates, combination therapy may only be of benefit in specific patient populations, such as critically ill or immunocompromised patients. Therefore, clinicians may want to consider using the combination therapy for the initial management and ceasing the aminoglycosides once antibiotic susceptibility results have been obtained.

Metagenomic insights into the microbial community structure and resistomes of a tropical agricultural soil persistently inundated with pesticide and animal manure use.

Persistent use of pesticides and animal manure in agricultural soils inadvertently introduced heavy metals and antibiotic/antibiotic resistance genes (ARGs) into the soil with deleterious consequences. The microbiome and heavy metal and antibiotic resistome of a pesticide and animal manure inundated agricultural soil (SL6) obtained from a vegetable farm at Otte, Eiyenkorin, Kwara State, Nigeria, was deciphered via shotgun metagenomics and functional annotation of putative ORFs (open reading frames). Structural metagenomics of SL6 microbiome revealed 29 phyla, 49 classes, 94 orders, 183 families, 366 genera, 424 species, and 260 strains with the preponderance of the phyla Proteobacteria (40%) and Actinobacteria (36%), classes Actinobacteria (36%), Alphaproteobacteria (18%), and Gammaproteobacteria (17%), and genera Kocuria (16%), Sphingobacterium (11%), and Brevundimonas (10%), respectively. Heavy metal resistance genes annotation conducted using Biocide and Metal Resistance Gene Database (BacMet) revealed the detection of genes responsible for the uptake, transport, detoxification, efflux, and regulation of copper, cadmium, zinc, nickel, chromium, cobalt, selenium, tungsten, mercury, and several others. ARG annotation using the Antibiotic Resistance Gene-annotation (ARG-ANNOT) revealed ARGs for 11 antibiotic classes with the preponderance of ß-lactamases, mobilized colistin resistance determinant (mcr-1), macrolide-lincosamide-streptogramin (MLS), glycopeptide, and aminoglycoside resistance genes, among others. The persistent use of pesticide and animal manure is strongly believed to play a major role in the proliferation of heavy metal and antibiotic resistance genes in the soil. This study revealed that agricultural soils inundated with pesticide and animal manure use are potential hotspots for ARG spread and may accentuate the spread of multidrug resistant clinical pathogens.

Multi-Omics Analysis Reveals that the Antimicrobial Kasugamycin Potential Targets Nitrate Reductase in Didymella segeticola to Achieve Control of Tea Leaf Spot.

Because of the lack of effective disease management measures, tea leaf spot-caused by the fungal phytopathogen Didymella segeticola (syn. Phoma segeticola)-is an important foliar disease. The important and widely used agricultural antimicrobial kasugamycin (Ksg), produced by the Gram-positive bacterium Streptomyces kasugaensis, effects high levels of control against crop diseases. The results of this study indicated that Ksg could inhibit the growth of D. segeticola hyphae in vitro with a half-maximal effective concentration (EC50) of 141.18 µg ml-1. Meanwhile, the curative effect in vivo on the pathogen in detached tea leaves also demonstrated that Ksg induced some morphological changes in organelles, septa, and cell walls as observed by optical microscopy and by scanning and transmission electron microscopy. This may indicate that Ksg disturbs biosynthesis of key metabolites, inhibiting hyphal growth. Integrated transcriptomic, proteomic, and bioinformatic analyses revealed that differentially expressed genes or differentially expressed proteins in D. segeticola hyphae in response to Ksg exposure were involved with metabolic processes and biosynthesis of secondary metabolites. Molecular docking studies indicated that Ksg may target nitrate reductase (NR), and microscale thermophoresis assay showed greater affinity with NR, potentially disturbing nitrogen assimilation and subsequent metabolism. The results indicated that Ksg inhibits the pathogen of tea leaf spot, D. segeticola, possibly by binding to NR, disturbing fungal metabolism, and inducing subsequent changes in hyphal growth and development.

Clinical Pharmacogenetics Implementation Consortium Guideline for the Use of Aminoglycosides Based on MT-RNR1 Genotype.

Aminoglycosides are widely used antibiotics with notable side effects, such as nephrotoxicity, vestibulotoxicity, and sensorineural hearing loss (cochleotoxicity). MT-RNR1 is a gene that encodes the 12s rRNA subunit and is the mitochondrial homologue of the prokaryotic 16s rRNA. Some MT-RNR1 variants (i.e., m.1095T>C; m.1494C>T; m.1555A>G) more closely resemble the bacterial 16s rRNA subunit and result in increased risk of aminoglycoside-induced hearing loss. Use of aminoglycosides should be avoided in individuals with an MT-RNR1 variant associated with an increased risk of aminoglycoside-induced hearing loss unless the high risk of permanent hearing loss is outweighed by the severity of infection and safe or effective alternative therapies are not available. We summarize evidence from the literature supporting this association and provide therapeutic recommendations for the use of aminoglycosides based on MT-RNR1 genotype (updates at https://cpicpgx.org/guidelines/ and www.pharmgkb.org).