Normal Human Microbiota Research Articles

Coagulase-negative staphylococci (CoNS) meningitis: a narrative review of the literature from 2000 to 2020.

Coagulase-negative staphylococci (CoNS) are recognized as comprising the main part of human normal microbiota and are rarely associated with severe and intensive infections. However, these organisms can cause a number of infections in humans, especially immunocompromised patients and neonates. Bacterial meningitis, as an important and acute infection in the central nervous system, is still a major global health challenge and a serious infectious disease, causing a high rate of mortality and morbidity. CoNS as causative agents of meningitis are generally related to trauma or direct implantation of foreign bodies and the presence of a cerebrospinal fluid shunt. Numerous epidemiologic and clinical studies have shown that different CoNS isolates such as Staphylococcus capitis, Staphylococcus lugdunensis, Staphylococcus hominis, Staphylococcus epidermidis, Staphylococcus schleiferi, Staphylococcus saprophyticus, Staphylococcus warneri and Staphylococcus haemolyticus are more frequently associated with meningitis. This study attempts to determine the role of CoNS in meningitis and reviews the reported cases of meningitis induced by CoNS from the year 2000 to 2020 in the literature.

The Pathogenic Role of Actinomyces spp. and Related Organisms in Genitourinary Infections: Discoveries in the New, Modern Diagnostic Era.

Actinomycosis is a chronic, suppurative, granulomatous infectious disease, caused by different species of Actinomyces bacteria. To date, 26 validly published Actinomyces species have been described as part of a normal human microbiota or from human clinical specimens. Due to the rapid spread of new, modern diagnostic procedures, 13 of 26 of these species have been described in this century and the Actinomycetaceae family has undergone several taxonomic revisions, including the introduction of many novel species termed Actinomyces-like organisms (ALOs). There is scarce data available on the role of these novel bacterial species in various infectious processes in human medicine. The aim of this review is to provide a comprehensive overview of Actinomyces and closely related organisms involved in human diseases—with a special focus on newly described species—in particular their role in genitourinary tract infections in females and males.

Yarrowia lipolytica causes sporadic cases and local outbreaks of infections and colonisation.

Yarrowia lipolytica belongs to the normal human microbiota but is also found in substrates with high contents in lipids and used in biotechnological processes. It is sometimes reported as human pathogen and especially in catheter-related candidaemia. Two apparently grouped cases of infections and/or contamination were reported involving 3 and 9 patients, respectively, in two hospitals. The goal of this study was to design a molecular tool to study the genetic diversity of Ylipolytica and confirm or not the common source of contamination during these grouped cases. Given that there is no genotyping method, we used genomic markers assessed on environmental isolates to determine intra-species relationship. We selected five highly polymorphic intergenic regions, totalling more than 3200bp and sequenced them for clinical (n=20) and environmental (n=14) isolates. Antifungal susceptibility was determined by EUCAST broth microdilution method. Multiple alignment of the five sequences revealed divergence of 0%-5.8% between isolates as compared to approximately 0.2%-0.25% after alignment of whole genomes, suggesting their potential usefulness to establish genetic relatedness. The analysis showed the multiple origins of the isolates. It uncovered two grouped case of fungaemia involving 3 and 2 patients, respectively. It also revealed several unrelated sporadic cases despite their temporal relationship and one probable laboratory contamination by a common yet uncovered source, explaining several consecutive positive cultures without infection. All isolates had high minimal inhibitory concentration (MIC) for flucytosine, the majority (14/34) was susceptible to fluconazole, and all to the other antifungal agents tested. This method could help elucidate cases related to the opportunistic pathogen Y lipolytica.

Staphylococcus epidermidis facilitates intracellular pathogen clearance through upregulation of antimicrobial protein perforin-2 (P-2) in the human skin gamma delta T cells

Abstract Staphylococcus epidermidis (SE), nonvirulent Gram-positive (G+) bacterium, is a member of the normal human skin microbiota with beneficial relationship with the host. Gamma delta (GD) T cells as the major T cell population in epithelial tissues have been implicated in maintaining tissue integrity, regulating inflammation and defending against pathogens. Perforin-2 (P-2) is a recently described antimicrobial protein responsible for clearance of intracellular G+ and G− bacteria. We examine the relationship between SE and P-2 expression in the human skin. Healthy human skin tissue was used for flow cytometric and bacterial infection analyses. We analyzed P-2 expression at a single cell resolution using an amplified fluorescence in situ hybridization (FISH) technique for detection of P-2 mRNA in combination with immune-phenotyping. Methicillin resistant Staphylococcus aureus (MRSA) intracellular killing assay was performed on the skin cells that were pretreated with S. epidermidis for 24 h. We found increase in the frequency of skin GDT cells and induction of P-2 transcripts in GDT cells after S. epidermidis infection. Incubating skin cells with S. epidermidis for 24h prior infection with MRSA resulted in rapid intracellular clearance of MRSA. Our findings reveal a novel P-2 mediated mechanism by which skin commensal bacteria may exert their beneficial role in modulating host innate immune response. In contrast, establishment of S. aureus biofilm resulted in suppression of P-2 expression, revealing a mechanism by which S. aureus escapes cutaneous immunity to cause persistent biofilm wound infections.

Mechanisms of interaction of Cetylpyridinium chloride with Staphylococcus aureus in the presence of β-cyclodextrin

Cetylpyridinium chloride (CPC) is a cationic surfactant, which has a biocidal activity against a broad spectrum of bacteria, including Staphylococcus aureus. This microorganism, although frequently found in the normal human microbiota, can become a pathogen that causes a wide variety of infections. Thus, the present work aims to investigate the effect of the β-cyclodextrin (βCD) on CPC antimicrobial activity against S. aureus, especially in the mechanism of interaction with S. aureus membrane. For these purposes, in vitro antimicrobial susceptibility of CPC and CPC/βCD compounds against S. aureus were determined by calculating the lower concentration capable of reducing cell viability by 50 and 100 percent, and the kinetics of bacterial death were evaluated by kill curves for the two systems. After that, the colloidal mechanisms of interaction of the CPC and CPC/βCD against S. aureus were investigated by Dynamic Light Scattering (DLS) and Zeta Potential (ZP) titrations. Finally, the thermodynamic parameters of drug-cell interaction were determined by Isothermal Titration Calorimetry (ITC), in order to obtain deeper understanding of the mechanism of drug interactions. The results of DLS, ZP and ITC showed that in presence of βCD occurs a different mechanism of interaction with S. aureus membrane, explaining therefore the higher antimicrobial activity of CPC/βCD system.

The aerobic respiratory chain of Pseudomonas aeruginosa cultured in artificial urine media: Role of NQR and terminal oxidases.

Pseudomonas aeruginosa is a Gram-negative γ-proteobacterium that forms part of the normal human microbiota and it is also an opportunistic pathogen, responsible for 30% of all nosocomial urinary tract infections. P. aeruginosa carries a highly branched respiratory chain that allows the colonization of many environments, such as the urinary tract, catheters and other medical devices. P. aeruginosa respiratory chain contains three different NADH dehydrogenases (complex I, NQR and NDH-2), whose physiologic roles have not been elucidated, and up to five terminal oxidases: three cytochrome c oxidases (COx), a cytochrome bo3 oxidase (CYO) and a cyanide-insensitive cytochrome bd-like oxidase (CIO). In this work, we studied the composition of the respiratory chain of P. aeruginosa cells cultured in Luria Broth (LB) and modified artificial urine media (mAUM), to understand the metabolic adaptations of this microorganism to the growth in urine. Our results show that the COx oxidases play major roles in mAUM, while P. aeruginosa relies on CYO when growing in LB medium. Moreover, our data demonstrate that the proton-pumping NQR complex is the main NADH dehydrogenase in both LB and mAUM. This enzyme is resistant to HQNO, an inhibitory molecule produced by P. aeruginosa, and may provide an advantage against the natural antibacterial agents produced by this organism. This work offers a clear picture of the composition of this pathogen’s aerobic respiratory chain and the main roles that NQR and terminal oxidases play in urine, which is essential to understand its physiology and could be used to develop new antibiotics against this notorious multidrug-resistant microorganism.

Small Molecule Inhibitors of the Response Regulator ArsR Exhibit Bactericidal Activity against Helicobacter pylori.

Helicobacter pylori is considered the most prevalent bacterial pathogen in humans. The increasing antibiotic resistance evolved by this microorganism has raised alarm bells worldwide due to the significant reduction in the eradication rates of traditional standard therapies. A major challenge in this antibiotic resistance crisis is the identification of novel microbial targets whose inhibitors can overcome the currently circulating resistome. In the present study, we have validated the use of the essential response regulator ArsR as a novel and promising therapeutic target against H. pylori infections. A high-throughput screening of a repurposing chemical library using a fluorescence-based thermal shift assay identified several ArsR binders. At least four of these low-molecular weight compounds noticeably inhibited the DNA binding activity of ArsR and showed bactericidal effects against antibiotic-resistant strains of H. pylori. Among the ArsR inhibitors, a human secondary bile acid, lithocholic acid, quickly destroyed H. pylori cells and exhibited partial synergistic action in combination with clarithromycin or levofloxacin, while the antimicrobial effect of this compound against representative members of the normal human microbiota such as Escherichia coli and Staphylococcus epidermidis appeared irrelevant. Our results enhance the battery of novel therapeutic tools against refractory infections caused by multidrug-resistant H. pylori strains.

The Role of Secretory Pathways in Candida albicans Pathogenesis.

Candida albicans is a fungus that is a commensal organism and a member of the normal human microbiota. It has the ability to transition into an opportunistic invasive pathogen. Attributes that support pathogenesis include secretion of virulence-associated proteins, hyphal formation, and biofilm formation. These processes are supported by secretion, as defined in the broad context of membrane trafficking. In this review, we examine the role of secretory pathways in Candida virulence, with a focus on the model opportunistic fungal pathogen, Candida albicans.

Safety Aspects of Genetically Modified Lactic Acid Bacteria.

Lactic acid bacteria (LAB) have a long history of use in the food industry. Some species are part of the normal human microbiota and have beneficial properties for human health. Their long-standing use and considerable biotechnological potential have led to the development of various systems for their engineering. Together with novel approaches such as CRISPR-Cas, the established systems for engineering now allow significant improvements to LAB strains. Nevertheless, genetically modified LAB (GM-LAB) still encounter disapproval and are under extensive regulatory requirements. This review presents data on the prospects for LAB to obtain ‘generally recognized as safe’ (GRAS) status. Genetic modification of LAB is discussed, together with problems that can arise from their engineering, including their dissemination into the environment and the spread of antibiotic resistance markers. Possible solutions that would allow the use of GM-LAB are described, such as biocontainment, alternative selection markers, and use of homologous DNA. The use of GM-LAB as cell factories in closed systems that prevent their environmental release is the least problematic aspect, and this is also discussed.

Myoviridae phage PDX kills enteroaggregative Escherichia coli without human microbiome dysbiosis.

Introduction Bacteriophage therapy can be developed to target emerging diarrhoeal pathogens, but doing so in the absence of microbiome disruption, which occurs with antibiotic treatment, has not been established.Aim Identify a therapeutic bacteriophage that kills diarrhoeagenic enteroaggregative Escherichia coli (EAEC) while leaving the human microbiome intact.Methodology Phages from wastewater in Portland, OR, USA were screened for bacteriolytic activity by overlay assay. One isolated phage, PDX, was classified by electron microscopy and genome sequencing. A mouse model of infection determined whether the phage was therapeutic against EAEC. 16S metagenomic analysis of anaerobic cultures determined whether a normal human microbiome was altered by treatment.Results Escherichia virus PDX, a member of the strictly lytic family Myoviridae, killed a case-associated EAEC isolate from a child in rural Tennessee in a dose-dependent manner, and killed EAEC isolates from Columbian children. A single dose of PDX (multiplicity of infection: 100) 1 day post-infection reduced EAEC recovered from mouse faeces. PDX also killed EAEC when cultured anaerobically in the presence of human faecal bacteria. While the addition of EAEC reduced the β-diversity of the human microbiota, that of the cultures with either faeces alone, faeces with EAEC and PDX, or with just PDX phage was not different statistically.Conclusion PDX killed EAEC isolate EN1E-0007 in vivo and in vitro, while not altering the diversity of normal human microbiota in anaerobic culture, and thus could be part of an effective therapy for children in developing countries and those suffering from EAEC-mediated traveller’s diarrhoea without causing dysbiosis.

Finegoldia magna, an Anaerobic Gram-Positive Bacterium of the Normal Human Microbiota, Induces Inflammation by Activating Neutrophils.

The Gram-positive anaerobic commensal Finegoldia magna colonizes the skin and other non-sterile body surfaces, and is an important opportunistic pathogen. Here we analyzed the effect of F. magna on human primary neutrophils. F. magna strains ALB8 (expressing protein FAF), 312 (expressing protein L) and 505 (naturally lacking both protein FAF and L) as well as their associated proteins activate neutrophils to release reactive oxygen species, an indication for neutrophil oxidative burst. Co-incubation of neutrophils with the bacteria leads to a strong increase of CD66b surface expression, another indicator for neutrophil activation. Furthermore, all tested stimuli triggered the release of NETs from the activated neutrophils, pointing to a host defense mechanism in response to the tested stimuli. This phenotype is dependent on actin rearrangement, NADPH oxidases and the ERK1/2 pathway. Proteins FAF and L also induced the secretion of several pro-inflammatory neutrophil proteins; HBP, IL-8 and INFγ. This study shows for the first time a direct interaction of F. magna with human neutrophils and suggests that the activation of neutrophils plays a role in F. magna pathogenesis.

<i>Lactobacillus rhamnosus GG:</i> experience in pediatric gastroenterology

Human organism is considered as a symbiotic microorganism according to the standpoint of modern science which number of commensal microbes is many times greater than the number of host cells. Normal human microbiota is involved in many physiological processes in the organism, and dysbiosis is associated with the development of a number of diseases. The accumulated knowledge about the composition of the normal microbiome allowed to suggest that its modification can change the course of the disease, and so the study of probiotics was initiated. One of the most studied probiotic strains is Lactobacillus rhamnosus GG (GG), ATCC 53103, isolated in 1985. Due to the strain’s resistance to acid and bile, good growth characteristics and the ability to adhere to the intestinal epithelial layer, Lactobacillus rhamnosus GG is most widely used in a variety of commercially available probiotic products. The article provides a review of the literature on the studies of this probiotic in the treatment and prevention of gastroenterological pathology in children, in particular, current evidence of the effectiveness of Lactobacillus rhamnosus GG in the treatment of acute gastroenteritis, antibiotic-associated diarrhea, as well as the results of studies in necrotizing enterocolitis, functional disorders of the intestine, in the eradication therapy for infection caused by Helicobacter pylori.

Oxadiazolylthiazoles as novel and selective antifungal agents

Studying the structure-activity relationships (SAR) of oxadiazolylthiazole antibiotics unexpectedly led us to identify ethylenediamine- and propylenediamine-analogs as potential antimycotic novel lead structures. Replacement of the ethylenediamine moiety for the lead compound 7 with cis-diaminocyclohexyl group (compound 18) significantly enhanced the antifungal activity. In addition to the high safety margin of 18 against mammalian cells, it showed highly selective broad-spectrum activity against fungal cells without inhibiting the human normal microbiota. The antifungal activity of 18 was investigated against 20 drug-resistant clinically important fungi, including Candida species, Cryptococcus, and Aspergillus fumigatus strains. In addition to the low MIC values that mostly ranged between 0.125 and 2.0 μg/mL, compound 18 outperformed fluconazole in disrupting mature Candida biofilm.

The Risk of Development of Antimicrobial Resistance with the Use of Coccidiostats in Poultry Diets

The Risk of Development of Antimicrobial Resistance with the Use of Coccidiostats in Poultry Diets

Complete hybrid genome assembly of clinical multidrug-resistant Bacteroides fragilis isolates enables comprehensive identification of antimicrobial-resistance genes and plasmids.

Bacteroides fragilis constitutes a significant part of the normal human gut microbiota and can also act as an opportunistic pathogen. Antimicrobial resistance (AMR) and the prevalence of AMR genes are increasing, and prediction of antimicrobial susceptibility based on sequence information could support targeted antimicrobial therapy in a clinical setting. Complete identification of insertion sequence (IS) elements carrying promoter sequences upstream of resistance genes is necessary for prediction of AMR. However, de novo assemblies from short reads alone are often fractured due to repeat regions and the presence of multiple copies of identical IS elements. Identification of plasmids in clinical isolates can aid in the surveillance of the dissemination of AMR, and comprehensive sequence databases support microbiome and metagenomic studies. We tested several short-read, hybrid and long-lead assembly pipelines by assembling the type strain B. fragilis CCUG4856T (=ATCC25285=NCTC9343) with Illumina short reads and long reads generated by Oxford Nanopore Technologies (ONT) MinION sequencing. Hybrid assembly with Unicycler, using quality filtered Illumina reads and Filtlong filtered and Canu-corrected ONT reads, produced the assembly of highest quality. This approach was then applied to six clinical multidrug-resistant B. fragilis isolates and, with minimal manual finishing of chromosomal assemblies of three isolates, complete, circular assemblies of all isolates were produced. Eleven circular, putative plasmids were identified in the six assemblies, of which only three corresponded to a known cultured Bacteroides plasmid. Complete IS elements could be identified upstream of AMR genes; however, there was not complete correlation between the absence of IS elements and antimicrobial susceptibility. As our knowledge on factors that increase expression of resistance genes in the absence of IS elements is limited, further research is needed prior to implementing AMR prediction for B. fragilis from whole-genome sequencing.

Whole-Genome Sequences of Five Strains of Kocuria rosea, NCTC2676, NCTC7514, NCTC7512, NCTC7528, and NCTC7511.

Kocuria rosea is a Gram-positive coccus found in the environment and within normal human skin microbiota, and more recently, it has been potentially implicated as an opportunistic pathogen. Here, we describe the genome sequences of five strains of K. rosea (NCTC2676, NCTC7514, NCTC7512, NCTC7528, and NCTC7511).

Glycobiology of Human Fungal Pathogens: New Avenues for Drug Development.

Invasive fungal infections (IFI) are an increasing threat to the developing world, with fungal spores being ubiquitous and inhaled every day. Some fungal species are commensal organisms that are part of the normal human microbiota, and, as such, do not pose a threat to the immune system. However, when the natural balance of this association is disturbed or the host’s immune system is compromised, these fungal pathogens overtake the organism, and cause IFI. To understand the invasiveness of these pathogens and to address the growing problem of IFI, it is essential to identify the cellular processes of the invading organism and their virulence. In this review, we will discuss the prevalence and current options available to treat IFI, including recent reports of drug resistance. Nevertheless, the main focus of this review is to describe the glycobiology of human fungal pathogens and how various components of the fungal cell wall, particularly cell wall polysaccharides and glycoconjugates, are involved in fungal pathogenicity, their biosynthesis and how they can be potentially exploited to develop novel antifungal treatment options. We will specifically describe the nucleotide sugar transporters (NSTs) that are important in fungal survival and suggest that the inhibition of fungal NSTs may potentially be useful to prevent the establishment of fungal infections.

The novel use of oral antibiotic monotherapy in prosthetic valve endocarditis caused by Finegoldia magna: a case study

BackgroundFinegoldia magna, a Gram-positive anaerobic coccus, is part of the human normal microbiota as a commensal of mucocutaneous surfaces. However, it remains an uncommon pathogen in infective endocarditis, with only eight clinical cases previously reported in the literature.Currently, infective endocarditis is routinely treated with prolonged intravenous antibiotic therapy. However, recent research has found that switching patients to oral antibiotics is non-inferior to prolonged parenteral antibiotic treatment, challenging the current guidelines for the treatment of infective endocarditis.Case presentationThis case report focuses on a 52-year-old gentleman, who presented with initially culture-negative infective endocarditis following bioprosthetic aortic valve replacement. Blood cultures later grew Finegoldia magna. Following initial intravenous antibiotic therapy and re-do surgical replacement of the prosthetic aortic valve, the patient was successfully switched to oral antibiotic monotherapy, an unusual strategy in the treatment of infective endocarditis inspired by the recent publication of the POET trial. He made excellent progress on an eight-week course of oral antibiotics and was successfully discharged from surgical follow-up.ConclusionsThis case is the 9th reported case of Finegoldia magna infective endocarditis in the literature. Our case also raises the possibility of a more patient-friendly and cost-effective means of providing long-term antibiotic therapy in suitable patients with prosthetic valve endocarditis and suggests that the principles highlighted in the POET trial can also be applicable to post-operative patients after cardiac surgery.

A Review of Mixed Strain Clostridium difficile Colonization and Infection

Given that Clostridium difficile is not part of the normal human microbiota, if multiple strains are to accumulate in the colon implies successive exposure events and/or persistent colonization must occur. Evidence of C. difficile infection (CDI) with more than one strain was first described in 1983. Despite the availability of increasingly discriminatory bacterial fingerprinting methods, the described rate of dual strain recovery in patients with CDI has remained stable at ∼5–10%. More data are needed to determine when dual strain infection may be harmful. Notably, one strain may block the establishment of and infection by another. In humans, patients colonized by non-toxigenic C. difficile strain are at a lower risk of developing CDI. Further studies to elucidate the interaction between co-infecting or colonizing and infecting C. difficile strains may help identify potential exploitable mechanisms to prevent CDI.

The environmental monitoring in hospital pharmacy cleanroom and microbiota catalogue preparation

Knowing the normal cleanroom microbiota is the basis for ensuring microbiological quality; assess changes and the introduction of new sampling methods. During our study, we prepared a catalogue of cleanroom microorganisms located in four different cleanrooms in University Clinical Centre Ljubljana Pharmacy. Catalogue is prepared as a basis for assessing the suitability of the new rapid microbiological method and subsequent correlation of the results of both methods. The results of our study showed that 78% of isolated bacteria are Gram-positive. However, in more than 70% isolated bacteria were the part of the normal human microbiota, 10–15% of the microorganisms originated from the air, mainly spore-forming bacteria of the genus Bacillus and fungi, and 5–10% of the Gram-negative microorganisms that originated from the water and represent the potential endotoxins (pyrogens).