Surface-Related Features and Virulence Among Acinetobacter baumannii Clinical Isolates Belonging to International Clones I and II

Acinetobacter baumannii currently represents one of the most important nosocomial infection agent due to its multidrug-resistance and a propensity for the epidemic spread. The A. baumannii strains belonging to the international clonal lineages I (IC I) and II (IC II) are associated with the hospital outbreaks and a high virulence. However, the intra and inter lineage-specific features of strains belonging to these most worldwide spread A. baumannii clones are not thoroughly explored. In this study we have investigated a set of cell surface-related features of A. baumannii IC I (n = 20) and IC II (n = 16) lineage strains, representing 30 distinct pulsed-field gel electrophoresis types in the collection of clinical isolates obtained in Lithuanian tertiary care hospitals. We show that A. baumannii IC II strains are non-motile, do not form pellicle and display distinct capsular polysaccharide profile compared with the IC I strains. Moreover, in contrast to the overall highly hydrophobic IC I strains, IC II strains showed a greater variation in cell surface hydrophobicity. Within the IC II lineage, hydrophilic strains demonstrated reduced ability to form biofilm and adhere to the abiotic surfaces, also possessed twofold thicker cell wall and exhibited higher resistance to desiccation. Furthermore, these strains showed increased adherence to the lung epithelial cells and were more virulent in nematode and mouse infection model compared with the hydrophobic IC II strains. According to the polymerase chain reaction-based locus-typing, the reduction in hydrophobicity of IC II strains was not capsule or lipooligosaccharide locus type-dependent. Hence, this study shows that the most widespread A. baumannii clonal lineages I and II markedly differ in the series of cell surface-related phenotypes including the considerable phenotypic diversification of IC II strains at the intra-lineage level. These findings suggest that the genotypically related A. baumannii strains might evolve the features which could provide an advantage at the specific conditions outside or within the host

Microbial Diversity and Antimicrobial Resistance Profile in Microbiota From Soils of Conventional and Organic Farming Systems

Soil is one of the biggest reservoirs of microbial diversity, yet the processes that define the community dynamics are not fully understood. Apart from soil management being vital for agricultural purposes, it is also considered a favorable environment for the evolution and development of antimicrobial resistance, which is due to its high complexity and ongoing competition between the microorganisms. Different approaches to agricultural production might have specific outcomes for soil microbial community composition and antibiotic resistance phenotype. Therefore in this study we aimed to compare the soil microbiota and its resistome in conventional and organic farming systems that are continually influenced by the different treatment (inorganic fertilizers and pesticides vs. organic manure and no chemical pest management). The comparison of the soil microbial communities revealed no major differences among the main phyla of bacteria between the two farming styles with similar soil structure and pH. Only small differences between the lower taxa could be observed indicating that the soil community is stable, with minor shifts in composition being able to handle the different styles of treatment and fertilization. It is still unclear what level of intensity can change microbial composition but current conventional farming in Central Europe demonstrates acceptable level of intensity for soil bacterial communities. When the resistome of the soils was assessed by screening the total soil DNA for clinically relevant and soil-derived antibiotic resistance genes, a low variety of resistance determinants was detected (resistance to β-lactams, aminoglycosides, tetracycline, erythromycin, and rifampicin) with no clear preference for the soil farming type. The same soil samples were also used to isolate antibiotic resistant cultivable bacteria, which were predominated by highly resistant isolates of Pseudomonas, Stenotrophomonas, Sphingobacterium and Chryseobacterium genera. The resistance of these isolates was largely dependent on the efflux mechanisms, the soil Pseudomonas spp. relying mostly on RND, while Stenotrophomonas spp. and Chryseobacterium spp. on RND and ABC transporters

Inactivation of Opportunistic Pathogens <i>Acinetobacter baumannii</i> and <i>Stenotrophomonas maltophilia</i> by Antimicrobial Photodynamic Therapy

Acinetobacter baumannii and Stenotrophomonas maltophilia are opportunistic pathogens causing hospital infections with limited treatment options due to bacterial multidrug resistance. Here, we report that antimicrobial photodynamic therapy (aPDT) based on the natural photosensitizers riboflavin and chlorophyllin inactivates A. baumannii and S. maltophilia. The riboflavin and chlorophyllin photostability experiments assessed the photomodifications of photosensitizers under the conditions subsequently used to inactivate A. baumannii and S. maltophilia. A. baumannii planktonic cells were more sensitive to riboflavin-aPDT, while biofilm bacteria were more efficiently inactivated by chlorophyllin-aPDT. S. maltophilia planktonic and biofilm cells were more susceptible to chlorophyllin-aPDT compared to riboflavin-aPDT. The results suggest that riboflavin- and chlorophyllin-aPDT can be considered as a potential antimicrobial treatment for A. baumannii and S. maltophilia inactivation

The role of Acinetobacter baumannii response regulator BfmR in pellicle formation and competitiveness via contact-dependent inhibition system

Background: Acinetobacter baumannii is one of the most important opportunistic pathogens responsible for hospital acquired infections. It displays multi-drug resistance profile and has the ability to colonize surfaces and persist under harsh conditions. A. baumannii two-component signal transduction system BfmRS, consisting of response regulator BfmR and sensor kinase BfmS, has been implicated in the control of various virulence-related traits and has been suggested to act as a global modulator of A. baumannii physiology. Results: Here, we assessed the role of BfmR regulator in pellicle formation and bacterial competition, features important for the establishment of A. baumannii in clinical environment. We show that BfmR is required for the pellicle formation of A. baumannii, as ΔbfmRS mutant lacked this phenotype. The loss of bfmRS also greatly reduced the secretion of A. baumannii Hcp protein, which is a component of T6SS secretion system. However, the T6SS-mediated killing phenotype was not impaired in the ΔbfmRS mutant. On the contrary, the same mutation resulted in the transcriptional activation of contact-dependent inhibition (CDI) system, which A. baumannii used to inhibit the growth of another clinical A. baumannii strain and a closely related species Acinetobacter baylyi. Conclusions: The obtained results indicate that BfmR is not only required for the pellicle phenotype induction in A. baumannii, but also, due to the down-regulation of a CDI system, could allow the incorporation of other A. baumannii strains or related species, possibly increasing the likelihood of the pathogens’ survival

The toxin-antitoxin systems of the opportunistic pathogen Stenotrophomonas maltophilia of environmental and clinical origin

Stenotrophomonas maltophilia is a ubiquitous environmental bacterium that has recently emerged as a multidrug-resistant opportunistic pathogen causing bloodstream, respiratory, and urinary tract infections. The connection between the commensal environmental S. maltophilia and the opportunistic pathogen strains is still under investigation. Bacterial toxin-antitoxin (TA) systems have been previously associated with pathogenic traits, such as biofilm formation and resistance to antibiotics, which are important in clinical settings. The same species of the bacterium can possess various sets of TAs, possibly influencing their overall stress response. While the TA systems of other important opportunistic pathogens have been researched, nothing is known about the TA systems of S. maltophilia. Here, we report the identification and characterization of S. maltophilia type II TA systems and their prevalence in the isolates of clinical and environmental origins. We found 49 putative TA systems by bioinformatic analysis in S. maltophilia genomes. Despite their even spread in sequenced S. maltophilia genomes, we observed that relBE, hicAB, and previously undescribed COG3832-ArsR operons were present solely in clinical S. maltophilia isolates collected in Lithuania, while hipBA was more frequent in the environmental ones. The kill-rescue experiments in Escherichia coli proved higBA, hicAB, and relBE systems to be functional TA modules. Together with different TA profiles, the clinical S. maltophilia isolates exhibited stronger biofilm formation, increased antibiotic, and serum resistance compared to environmental isolates. Such tendencies suggest that certain TA systems could be used as indicators of virulence traits

Identification of colistin resistance gene origin in Escherichia coli isolated from migratory bird

Introduction: As antimicrobial resistance is becoming a major threat to global health, colistin remains as one of the very few antibiotics that can still be effectively used against multi-drug resistant bacteria. However, an outbreak of plasmid-mediated colistin resistance, conferred by mcr-1 gene, was recently reported. Since the mcr-1 plasmids can be transferred between different species of bacteria, it causes concern of the emergence of bacteria resistant to all known antibiotics. Therefore, to identify and understand the mechanisms of mcr-1 spread is crucial for reducing fatalities caused by untreatable bacterial infections. Aim: To identify the origin of mcr-1 from E. coli isolated from faeces of Larus argentatus in Kaunas city dump. To the best of our knowledge, it is the first known occurrence of mcr-1 in Lithuania. Materials and Methods: To ensure that the mcr-1 is located in a plasmid, it was transferred to another strain via conjugation. PCR was performed to confirm the presence of mcr-1 and to find other antibiotic resistance genes in the plasmid. NCBI GenBank Database was screened for occurrences of mcr-1 gene using BLAST (100% identity and coverage cut-off). Potential plasmids were evaluated by restriction analysis. Results: The mcr-1 was successfully transferred into E. coli K-12 laboratory strain, proving that it is located in a plasmid. Any other antibiotic resistance genes were not found in the plasmid. 143 complete plasmid sequences from NCBI GenBank database contained identical mcr-1 sequence. By data acquired from restriction analysis of the extracted target plasmid, we can conclude, that it belongs to a group of 23 highly similar plasmids. Conclusion: The mcr-1 gene in E. coli isolated from migratory bird was found to be located in a plasmid widely spread across the world and shared between both animal vectors and human hospital patients

OmpA Protein-Deficient Acinetobacter baumannii Outer Membrane Vesicles Trigger Reduced Inflammatory Response

Multidrug resistant Acinetobacter baumannii shows a growing number of nosocomial infections worldwide during the last decade. The outer membrane vesicles (OMVs) produced by this bacterium draw increasing attention as a possible treatment target. OMVs have been implicated in the reduction of antibiotic level in the surrounding environment, transfer of virulence factors into the host cells, and induction of inflammatory response. Although the evidence on the involvement of OMVs in A. baumannii pathogenesis is currently growing, their role during inflammation is insufficiently explored. It is likely that bacteria, by secreting OMVs, can expand the area of their exposure and prepare surrounding matrix for infection. Here, we investigated the impact of A. baumannii OMVs on activation of macrophages in vitro. We show that OmpA protein present in A. baumannii OMVs substantially contributes to the proinflammatory response in J774 murine macrophages and to the cell death in both lung epithelium cells and macrophages. The loss of OmpA protein in OMVs, obtained from A. baumannii ∆ompA mutant, resulted in the altered expression of genes coding for IL-6, NLRP3 and IL-1β proinflammatory molecules in macrophages in vitro. These results imply that OmpA protein in bacterial OMVs could trigger a more intense proinflammatory response