Single Pathogen Research Articles

Ten-year longitudinal molecular epidemiology study of Escherichia coli and Klebsiella species bloodstream infections in Oxfordshire, UK

BackgroundThe incidence of Gram-negative bloodstream infections (BSIs), predominantly caused by Escherichia coli and Klebsiella species, continues to increase; however, the causes of this are unclear and effective interventions are therefore hard to design.MethodsIn this study, we sequenced 3468 unselected isolates over a decade in Oxfordshire (UK) and linked this data to routinely collected electronic healthcare records and mandatory surveillance reports. We annotated genomes for clinically relevant genes, contrasting the distribution of these within and between species, and compared incidence trends over time using stacked negative binomial regression.ResultsWe demonstrate that the observed increases in E. coli incidence were not driven by the success of one or more sequence types (STs); instead, four STs continue to dominate a stable population structure, with no evidence of adaptation to hospital/community settings. Conversely in Klebsiella spp., most infections are caused by sporadic STs with the exception of a local drug-resistant outbreak strain (ST490). Virulence elements are highly structured by ST in E. coli but not Klebsiella spp. where they occur in a diverse spectrum of STs and equally across healthcare and community settings. Most clinically hypervirulent (i.e. community-onset) Klebsiella BSIs have no known acquired virulence loci. Finally, we demonstrate a diverse but largely genus-restricted mobilome with close associations between antimicrobial resistance (AMR) genes and insertion sequences but not typically specific plasmid replicon types, consistent with the dissemination of AMR genes being highly contingent on smaller mobile genetic elements (MGEs).ConclusionsOur large genomic study highlights distinct differences in the molecular epidemiology of E. coli and Klebsiella BSIs and suggests that no single specific pathogen genetic factors (e.g. AMR/virulence genes/sequence type) are likely contributing to the increasing incidence of BSI overall, that association with AMR genes in E. coli is a contributor to the increasing number of E. coli BSIs, and that more attention should be given to AMR gene associations with non-plasmid MGEs to try and understand horizontal gene transfer networks.

Simultaneous identification of Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium, and Trichomonas vaginalis ‒ multicenter evaluation of the Alinity m STI assay

Abstract Objectives Accurate and rapid diagnosis of sexually transmitted infections (STIs) is essential for timely administration of appropriate treatment and reducing the spread of the disease. We examined the performance of the new Alinity m STI assay, a qualitative real-time multiplex PCR test for simultaneous identification of Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Mycoplasma genitalium (MG), and Trichomonas vaginalis (TV) run on the fully automated Alinity m platform. Methods This international, multicenter study evaluated the accuracy, reproducibility, and clinical performance of the Alinity m STI assay compared to commonly used STI assays in a large series of patient samples encountered in clinical practice. Results The Alinity m STI assay identified accurately and precisely single and mixed pathogens from an analytical panel of specimens. The Alinity m STI assay demonstrated high overall agreement rates with comparator STI assays (99.6% for CT [n=2,127], 99.2% for NG [n=2,160], 97.1% for MG [n=491], and 99.4% for TV [n=313]). Conclusions The newly developed Alinity m STI assay accurately detects the 4 sexually transmitted target pathogens in various collection devices across clinically relevant specimen types, regardless of single or mixed infection status.

Application of metagenomic next-generation sequencing technology in pathogen detection in patients with burns and patients with acute or chronic wounds

Objective: To explore the value of using metagenomic next-generation sequencing (mNGS) technology to detect pathogens in patients with burns and patients with acute or chronic wounds. Methods: A retrospective observational study was conducted. From March 2019 to June 2020, 11 patients with burns and patients with acute or chronic wounds (including 10 males and 1 female, aged 23 to 85 years) in the Fourth Medical Center of PLA General Hospital met the inclusion criteria and were recruited. A total of 23 specimens were collected, including 6 whole blood specimens, 1 skin tissue specimen, 1 drained pus specimen, and 15 wound secretion swab specimens. Each specimen was divided into two parts, which were subjected for pathogen detection using microbial culture method and mNGS method, respectively. The number and types of pathogens detected by the 2 methods and the relative abundance detected by the mNGS method were recorded, and the consistency of the two methods were compared. Data were statistically analyzed with paired Wilcoxon rank sum test. Results: With the microbial culture method, no pathogen was detected in 5 of the 23 specimens, while 35 pathogens were detected in the remaining 18 specimens, belonging to 9 species of bacteria and 2 species of fungi. Five specimens had one pathogen while 9 specimens had 2 pathogens and 4 specimens had 3 pathogens detected in each specimen. With the mNGS method, no pathogen was detected in one of the 23 specimens, while 75 pathogens were detected in the remaining 22 specimens, belonging to 28 species of bacteria, 3 species of fungi, and 3 species of viruses. Eight specimens had one pathogen, 5 specimens had 2 pathogens, 2 specimens had 3 pathogens, 3 specimens had 4 pathogens, 2 specimens had 6 pathogens, and 1 specimen had 7 pathogens, and 1 specimen had 20 pathogens detected in each specimen. The number of pathogens detected in each specimen by microbial culture method was 2 (1, 2) types, which was significantly less than 2 (1, 4) types by mNGS method (Z=3.359, P<0.01). In 5 specimens, no bacteria were detected by microbial culture method but mNGS method detected bacteria in 2 specimens and virus in 2 different specimens. The mNGS method detected two or more types of bacteria in 13 specimens, the relative abundance of bacteria with the 1st relative abundance ranking ranged from 28.8% to 95.9% in each specimen. Of the 23 specimens detected by two detection methods, 7 specimens (30.4%) showed identical detection results, 5 specimens (21.7%) showed totally different detection results, and 11 specimens (47.8%) had partially consistent detection results. Conclusions: Compared with the traditional microbial culture method, the mNGS method has higher detection sensitivity and stronger capacity to detect pathogens, and can determine the relative abundance of pathogens in mixed infections. As a supplement to the culture method, the mNGS method is expected to play an important role in the diagnosis of infectious pathogens in burns and acute or chronic wounds.

Decision tree analysis for pathogen identification based on circumstantial factors in outbreaks of bovine respiratory disease in calves

Respiratory tract infections continue to be a leading cause of economic loss, hampered animal welfare and intensive antimicrobial use in cattle operations, worldwide. To better target antimicrobial therapy, control and prevention towards the involved pathogens, there is a growing interest in microbiological tests on respiratory samples. However, these tests are time consuming, cost money and sampling might compromise animal welfare. Therefore, the objective of the present study was to develop immediately applicable decision trees for pathogen identification in outbreaks of bovine respiratory disease based on circumstantial factors. Data from a cross sectional study, involving 201 outbreaks of bovine respiratory disease in dairy and beef farms between 2016 and 2019 was used. Pathogens were identified by a semi-quantitative PCR (polymerase chain reaction) on a pooled non-endoscopic broncho-alveolar lavage sample from clinically affected animals. Potential risk factors of involved animals, environment, management and housing were obtained by enquiry. Classification and regression tree analysis was used for decision tree development with cross-validation. Different trees were constructed, involving a general 3-group classification tree (viruses, Mycoplasma bovis or Pasteurellaceae family) and a tree for each single pathogen. The general 3- group classification tree was 52.7 % accurate and had a sensitivity of 81.5 % and a specificity 52.2 % for viruses, respectively 51.7 % and 84.4 % for M. bovis and 28.9 % and 93.6 % for Pasteurellaceae. The single-pathogen trees were more specific than sensitive: Histophilus somni (Se = 25.8 %; Sp = 94.5 %), Mannheimia haemolytica (Se = 69.2 %; Sp = 70.6 %), bovine coronavirus (Se = 42.2 %; Sp = 89.6 %) and bovine respiratory syncytial virus (Se = 34.0 %; Sp = 96.6 %). For Pasteurella multocida, M. bovis and parainfluenzavirus type 3 no meaningful tree was obtained. The concept and trees are promising, but currently lack sensitivity and specificity in order to be a reliable tool for practice. For now, the obtained trees can already be informative for decision making to some extend depending on the end node in which an outbreak falls.

Pleurostoma Decline of Olive Trees Caused by Pleurostoma richardsiae in California.

A single fungal pathogen was consistently isolated from symptomatic wood of olive trees (Olea europaea) displaying branch and trunk cankers in superhigh-density orchards in the Sacramento and San Joaquin Valleys of California. Morphological characters of the pathogen included two distinct types of conidia (thick-walled, dark brown, and globose and thin-walled, hyaline, and oblong to ellipsoid) and three types of phialides, indicating a pleurostoma-like fungus. Phylogenetic results of four nuclear loci including the internal transcribed spacer (ITS) region (ITS1-5.8S-ITS2) and partial sequences of the actin, beta-tubulin, and translation elongation factor 1-α genes confirmed the isolates as Pleurostoma richardsiae. Pathogenicity trials conducted in the field involving 2- to 3-year-old branches of three widely planted oil olive cultivars (Arbequina, Arbosana, and Koroneiki) satisfied Koch's postulates and confirmed the pathogenic nature of this species to cause the decline of olive trees in California. All three cultivars were equally susceptible to Pl. richardsiae, indicating no detectable resistance to the pathogen. Additional isolations from symptomatic hosts including almond, peach, pistachio, and plum, also confirmed this species, suggesting that Pl. richardsiae is widespread in agricultural systems and should be considered an emerging pathogen of fruit and nut crops in California.

Utility of multiplex real-time PCR for diagnosing paediatric acute respiratory tract infection in a tertiary care hospital

BackgroundAcute respiratory infections (ARIs) are responsible for considerable morbidity and mortality among children all over the world. Many of the etiologic agents of the infections especially viral go undiagnosed for lack of requisite facility and the cost factors. We have used a commercially available platform for diagnosis of ARIs in children receiving inpatient and outpatient services in a tertiary care centre. MethodsThe framework of the study was prospective and observational. In this study, clinical samples of children suffering from ARIs were subjected to real-time multiplex PCR targeting both viral and bacterial pathogens. ResultsOf 94 samples received at our centre (49 male and 45 female), the positivity for respiratory pathogens was detected in 50 (53.19%) samples. Clinical symptoms of patients and age distribution have been elaborated in text. A single pathogen (n = 29/50), two pathogens (15/50) and three pathogens (n = 6/50) were detected by multiplex RT-PCR. Of 77 isolates detected, maximum numbers were of human rhinovirus (HRV) (n = 14) (18.18%) Streptococcus pneumoniae (n = 14) (18.18%) followed by Staphylococcus aureus (n = 10) (12.98%). ConclusionThe epidemiology of ARIs considering viral etiologies is poorly understood due to less number of studies especially in Indian subcontinent. The advent of latest advanced molecular methods has made it possible to identify common respiratory pathogens and has contributed to cover the gap in existing knowledge.

Co-occurrence of pathogen assemblages in a keystone species the common cockle Cerastoderma edule on the Irish coast.

Despite coinfections being recognized as the rule in animal populations, most studies focus on single pathogen systems. Pathogen interaction networks and the drivers of such associations are lacking in disease ecology studies. Common cockle Cerastoderma edule populations are exposed to a great diversity of pathogens, thus making them a good model system to investigate. This study examined the diversity and prevalence of pathogens from different taxonomic levels in wild and fished C. edule on the Irish coast. Potential interactions were tested focussing on abiotic (seawater temperature and salinity) and biotic (cockle size and age, and epiflora on shells) factors. No Microsporidia nor OsHV-1μVar were detected. Single infections with Haplosporidia (37.7%) or Vibrio (25.3%) were more common than two-pathogen coinfected individuals (9.5%), which may more easily succumb to infection. Fished C. edule populations with high cockle densities were more exposed to infections. Higher temperature and presence of epiflora on cockle shells promoted coinfection in warmer months. Low seawater salinity, host condition and proximity to other infected host species influenced coinfection distribution. A positive association between two Minchinia spp. was observed, most likely due to their different pathogenic effect. Findings highlight the major influence that ecological factors have on pathogen interactions and host–pathogen interplay.

Novel urine biomarkers to distinguish UTI from culture-negative pyuria.

Emergency departments (EDs) often rely on urinalysis (UA) to rapidly identify urinary tract infections (UTIs) in children. However, the suboptimal test characteristics of UA can lead to false-positive results. Novel urinary biomarkers may increase the diagnostic precision of UA. In this study, we compared the concentrations of 6 pre-selected proteins: BH3 interacting domain death agonist (BID), B-cell lymphoma 6 protein, ras GTPase-activating protein 1, cathepsin S (CTSS), 3-hydroxyanthranilate 3,4-dioxygenase, and transgelin-2. In a pediatric ED, we prospectively enrolled 167 children with UA and urine culture collected. Pyuria was defined as either ≥ 5 white blood cells per high-power field on microscopy or positive leukocyte esterase (LE). The urine culture was considered positive if it yielded ≥ 50,000 colony-forming units per milliliter of any single urinary pathogen. Urine protein levels were measured by enzyme-linked immunosorbent assay and normalized to urine creatinine. BID was significantly higher in the UTI group compared to the culture-negative pyuria group with a mean ratio of 1.42 (95% confidence interval (CI), 1.15, 1.76) when uncorrected for creatinine concentration. When corrected for creatinine concentration, CTSS was significantly elevated in the UTI group compared to the culture-negative pyuria group with a mean ratio of 2.11 (95% CI, 1.39, 3.21). BID and CTSS concentrations were elevated in the urine of children with UTI compared to those with culture-negative pyuria. These proteins deserve further research into their utility to serve as novel biomarkers for UTI.

The interplay of SARS-CoV-2 evolution and constraints imposed by the structure and functionality of its proteins.

The unprecedented pace of the sequencing of the SARS-CoV-2 virus genomes provides us with unique information about the genetic changes in a single pathogen during ongoing pandemic. By the analysis of close to 200,000 genomes we show that the patterns of the SARS-CoV-2 virus mutations along its genome are closely correlated with the structural and functional features of the encoded proteins. Requirements of foldability of proteins’ 3D structures and the conservation of their key functional regions, such as protein-protein interaction interfaces, are the dominant factors driving evolutionary selection in protein-coding genes. At the same time, avoidance of the host immunity leads to the abundance of mutations in other regions, resulting in high variability of the missense mutation rate along the genome. “Unexplained” peaks and valleys in the mutation rate provide hints on function for yet uncharacterized genomic regions and specific protein structural and functional features they code for. Some of these observations have immediate practical implications for the selection of target regions for PCR-based COVID-19 tests and for evaluating the risk of mutations in epitopes targeted by specific antibodies and vaccine design strategies.

Involvement of Mobile phones as a source for Nosocomial infections

Background: Mobile phones are communication devices for communication nowadays and have emerged as a need of our life. Mobile phones also act as a reservoir for microorganisms. Human skin, wallets, bags, environment and food items are major sources of contamination for mobile phones. Medical students, healthcare workers and other non-teaching staff nowadays use mobile phones for communication in the workplace. Mobile phone contamination spreads from surfaces to hands.&#x0D; Aims and Objective: Our study was aimed to investigate the contamination of mobile phones by pathogens among health care workers. The objective of this study was to document for the role of mobile phones as sources in nosocomial infections.&#x0D; Materials and Methods: The study was done in Department of Microbiology and Pathology, Era’s Lucknow Medical College and Hospital, including teaching staffs from hospital and college, health care workers in all laboratories and wards. A total of 200 swab samples from various mobile phones of medical and technical staff of various wards, Outpatient Department Treatment, laboratories was collected after taking informed written consent from them. Culture was done on Blood Agar, MacConkey Agar, and Sabouraud’s dextrose agar. After overnight incubation, colonies were identified by Gram’s stain and Biochemical tests.&#x0D; Results: Highest mobile phone users are medical students, residents followed by teaching faculty. Smart phones or touch screen mobiles were used by maximum ie.90.5%. Among the isolated bacteria, Bacillus (39.5%), CONS (19%), E.coli (15.5%), Klebsiella (10.5%), Pseudomonas (5.5%), Staphylococcus aureus (4%), Enterococcus (2.5%). No growth was seen in 3.5%mobile phone sample cultures. Not a single fungal pathogen was isolated.&#x0D; Conclusion: The findings is the present study suggests that the mobile phones used by medical staff and health care workers act as source of transmission vehicles for pathogenic microorganisms.

Isothermal amplification assay for visual on-site detection of Staphylococcus aureus in Chevon

ABSTRACT The ability to rapidly detect pathogens in food is important from public health and food safety reasons. Traditional culture-based detection methods tend to be laborious, time consuming, technically demanding and may be limited due to their low sensitivity. Rapid detection methods for foodborne pathogens should be specific and sensitive to detect pathogens in low numbers. Sensitivity is important because a single pathogen present in food has the potential risk to cause infection. PCR and Real-time PCR are rapid, sensitive, and specific but require specific instruments and laboratory set-up. In the present study, a simple loop-mediated isothermal amplification (LAMP) assay was developed for rapid detection of Staphylococcus aureus. The LAMP assay was found to be ten times more sensitive than traditional end-point PCR with analytical sensitivity of 0.56 pg/μl and 5.6 pg/μl of DNA, respectively. In spiked chevon samples inoculated with S. aureus, the detection limit of LAMP and PCR assay was 3.3 × 105 CFU/g and 3.3 × 106 CFU/g, respectively, without enrichment. After enriching the samples for 6 h, the detection limit improved to 3.3 × 102 CFU/g and 3.3 × 104 CFU/g, respectively, indicating LAMP to be 100-fold more sensitive than PCR. The detection limit further improved to 3.3 CFU/g of meat after enrichment of 12 h. The developed LAMP was also found to be suitable when evaluated on field samples. The present study reports a simple LAMP assay for rapid visual detection of S. aureus which has potential for use in resource limited settings.

Anti-infection mechanism of a novel dental implant made of titanium-copper (TiCu) alloy and its mechanism associated with oral microbiology

This work was focused on study of anti-infection ability and its underlying mechanism of a novel dental implant made of titanium-copper (TiCu) alloy. In general, most studies on antibacterial implants have used a single pathogen to test their anti-infection ability using infectious animal models. However, dental implant-associated infections are polymicrobial diseases. We innovatively combine the classic ligature model in dogs with sucrose-rich diets to induce oral infections via the canine native oral bacteria. The anti-infection ability, biocompatibility and underlying mechanism of TiCu implant were systematically investigated in comparison with pure Ti implant via general inspection, hematology, imageology (micro-CT), microbiology (16S rDNA and metagenome), histology, and Cu ion detections. Compared with Ti implant, TiCu implant demonstrated remarkable anti-infection potentials with excellent biocompatibility. Additionally, the underlying anti-infection mechanism of TiCu implant was considered to involve maintaining the oral microbiota homeostasis. It was found that the carbohydrates in the plaques formed on the surface of TiCu implant were metabolized through the tricarboxylic acid cycle (TCA) cycles, which prevented the formation of an acidic microenvironment and inhibited the accumulation of acidogens and pathogens, thereby maintaining the microflora balance between aerobic and anaerobic bacteria.

Key issues affecting the current status of infectious diseases in Chinese cattle farms and their control through vaccination

Infectious diseases can have a major impact on the profitability of the cattle industry. To determine the occurrence of bovine infectious diseases in China and the adoption of vaccination to control them, a national-wide questionnaire and focus group meeting were performed. The questionnaire was administered to 189 farmers including 93 dairy farmers, 80 beef cattle farmers and 16 yak farmers. Since it is compulsory to vaccinate cattle against foot and mouth disease, the coverage of vaccination to this disease was the highest (100% of dairy and yak farms and 92.5% of beef farms). However, the implementation of vaccination against other diseases was vastly different between cattle types with less than 50% of farms adopting vaccination (except brucellosis vaccine in yak farms). In a focus group meeting of 36 cattle experts on the key issues affecting the frequency of infectious diseases in cattle and the vaccination practices adopted on Chinese cattle farms, the lack of effective vaccines against single or multiple pathogens, a lack of tools for the early and correct diagnosis of disease, difficulties in licensing novel vaccines and diagnostic agents, low efficiency in disseminating knowledge on diseases and control products to producers were identified as key issues. In conclusion, except for FMD, the control of most infectious diseases of cattle in China requires improving. Development of improved control measures and diagnostic tests along with the development and implementation of educational material for producers on cattle diseases should be given priority.

Unique Pathogen Peptidomes Facilitate Pathogen-Specific Selection and Specialization of MHC Alleles.

A key component of pathogen-specific adaptive immunity in vertebrates is the presentation of pathogen-derived antigenic peptides by major histocompatibility complex (MHC) molecules. The excessive polymorphism observed at MHC genes is widely presumed to result from the need to recognize diverse pathogens, a process called pathogen-driven balancing selection. This process assumes that pathogens differ in their peptidomes—the pool of short peptides derived from the pathogen’s proteome—so that different pathogens select for different MHC variants with distinct peptide-binding properties. Here, we tested this assumption in a comprehensive data set of 51.9 Mio peptides, derived from the peptidomes of 36 representative human pathogens. Strikingly, we found that 39.7% of the 630 pairwise comparisons among pathogens yielded not a single shared peptide and only 1.8% of pathogen pairs shared more than 1% of their peptides. Indeed, 98.8% of all peptides were unique to a single pathogen species. Using computational binding prediction to characterize the binding specificities of 321 common human MHC class-I variants, we investigated quantitative differences among MHC variants with regard to binding peptides from distinct pathogens. Our analysis showed signatures of specialization toward specific pathogens especially by MHC variants with narrow peptide-binding repertoires. This supports the hypothesis that such fastidious MHC variants might be maintained in the population because they provide an advantage against particular pathogens. Overall, our results establish a key selection factor for the excessive allelic diversity at MHC genes observed in natural populations and illuminate the evolution of variable peptide-binding repertoires among MHC variants.

Key Macrophage Responses to Infection With Mycobacterium tuberculosis Are Co-Regulated by microRNAs and DNA Methylation.

Tuberculosis (TB) is the leading cause of death from infection with a single bacterial pathogen. Host macrophages are the primary cell type infected with Mycobacterium tuberculosis (Mtb), the organism that causes TB. Macrophage response pathways are regulated by various factors, including microRNAs (miRNAs) and epigenetic changes that can shape the outcome of infection. Although dysregulation of both miRNAs and DNA methylation have been studied in the context of Mtb infection, studies have not yet investigated how these two processes may jointly co-regulate critical anti-TB pathways in primary human macrophages. In the current study, we integrated genome-wide analyses of miRNA abundance and DNA methylation status with mRNA transcriptomics in Mtb-infected primary human macrophages to decipher which macrophage functions may be subject to control by these two types of regulation. Using in vitro macrophage infection models and next generation sequencing, we found that miRNAs and methylation changes co-regulate important macrophage response processes, including immune cell activation, macrophage metabolism, and AMPK pathway signaling.

Recurrent UTI in Ventriculoperitoneal Shunted Children's

Objective: To determine the rate and the type of recurrent UTI in ventriculoperitoneal shunted children's admitted to Azady Teaching Hospital, Kirkuk.. Methods: From mid 2015 to end of 2020; UTI in ventriculoperitoneal shunted children's . Once infection was suspected , CSF (from shunt reservoir or from ventricular tapping), blood, and urine samples, analysis, and culture taken and empirical antibiotics were recommended. Results: 25.9% of patients with VP shunts had infections which represents 29.3% of the procedures. 40% of infected patients had recurrent episodes. 59.1% of infections occurred throughout the first two months following insertion. Single pathogen was isolated in each episode. E.coli represented 50% of isolated pathogens compared with 18.2% with Staphylococcus epidermidis. Conclusions: There is a high incidence of recurrent UTI in ventriculoperitoneal shunted children's in Azady Teaching Hospital, Kirkuk. when compared withother international centres. Gram negative organisms are the most common cause of the infection. Keywords: Urinary tract infection= UTI ,Renal scar.,BBD = Bladder Bowel Dysfunction; DMSA = dimercaptosuccinic acid; IV

An inter-laboratory study to investigate the impact of the bioinformatics component on microbiome analysis using mock communities

Despite the advent of whole genome metagenomics, targeted approaches (such as 16S rRNA gene amplicon sequencing) continue to be valuable for determining the microbial composition of samples. Amplicon microbiome sequencing can be performed on clinical samples from a normally sterile site to determine the aetiology of an infection (usually single pathogen identification) or samples from more complex niches such as human mucosa or environmental samples where multiple microorganisms need to be identified. The methodologies are frequently applied to determine both presence of micro-organisms and their quantity or relative abundance. There are a number of technical steps required to perform microbial community profiling, many of which may have appreciable precision and bias that impacts final results. In order for these methods to be applied with the greatest accuracy, comparative studies across different laboratories are warranted. In this study we explored the impact of the bioinformatic approaches taken in different laboratories on microbiome assessment using 16S rRNA gene amplicon sequencing results. Data were generated from two mock microbial community samples which were amplified using primer sets spanning five different variable regions of 16S rRNA genes. The PCR-sequencing analysis included three technical repeats of the process to determine the repeatability of their methods. Thirteen laboratories participated in the study, and each analysed the same FASTQ files using their choice of pipeline. This study captured the methods used and the resulting sequence annotation and relative abundance output from bioinformatic analyses. Results were compared to digital PCR assessment of the absolute abundance of each target representing each organism in the mock microbial community samples and also to analyses of shotgun metagenome sequence data. This ring trial demonstrates that the choice of bioinformatic analysis pipeline alone can result in different estimations of the composition of the microbiome when using 16S rRNA gene amplicon sequencing data. The study observed differences in terms of both presence and abundance of organisms and provides a resource for ensuring reproducible pipeline development and application. The observed differences were especially prevalent when using custom databases and applying high stringency operational taxonomic unit (OTU) cut-off limits. In order to apply sequencing approaches with greater accuracy, the impact of different analytical steps needs to be clearly delineated and solutions devised to harmonise microbiome analysis results.

Growth-inhibitory effects of tris-(1,10-phenanthroline) iron (II) against Mycobacterium tuberculosis in vitro and in vivo

Mycobacterium tuberculosis is the major etiological agent for tuberculosis (TB), which is the leading cause of single pathogen infection-related deaths worldwide. The End TB Strategy of the World Health Organization aimed to decrease the incidence of TB by 20% between 2015 and 2020, which was not achieved. Here, the growth-inhibitory effects of tris-(1,10-phenanthroline) iron (II) complex ([Fe(phen)3]2+), a known commercially available cheap chemical substance, were examined. The best in vitro results showed great activity with MIC ranging from 0.77 to 3.06 μM against clinical strains and at low pH (mimicking the granuloma) with MIC of 0.21 μM. Preliminary safety analysis revealed that the complex did not exhibit cytotoxic activity against different cell lines or mutagenic activity in vitro. The complex was orally bioavailable after 2 h of administration in vivo. Additionally, the results of the acute toxicity test revealed that the complex did not exert toxic effects in female BALB/c mice. The mechanism of action was performed using D29 mycobacteriophages where the treatment with different concentrations of the complex inhibited viral protein synthesis, which indicated that the anti-TB mechanisms of the complex involve protein synthesis inhibition. These findings suggested that [Fe(phen)3]2+ is a potential novel therapeutic for TB.

A Systematic Review on the Efficacy of Vaporized Hydrogen Peroxide as a Non-Contact Decontamination System for Pathogens Associated with the Dental Environment.

Aerosol generation and a wide range of pathogens originating from the oral cavity of the patient contaminate various surfaces of the dental clinic. The aim was to determine the efficacy of vaporized hydrogen peroxide fogging on pathogens related to the dental environment and its possible application in dentistry. PICOS statement (Population, Intervention, Comparison/Control, Outcome and Study design statement) was used in the review. Six electronic databases were searched for articles published from 2010 to 2020. Articles written in English reporting vaporized hydrogen peroxide on pathogens deemed to be relevant to the dental environment were assessed. The quality of the studies was assessed using the risk-of-bias assessment tool designed for the investigation of vaporized hydrogen peroxide application in dentistry. A total of 17 studies were included in the qualitative synthesis. The most commonly reported single bacterial pathogen was Methicillin-resistant Staphylococcus aureus in five studies, and the viruses Feline calicivirus, Human norovirus, and Murine norovirus were featured in three studies. The results of the studies reporting the log kill were sufficient for all authors to conclude that vaporized hydrogen peroxide generation was effective for the assessed pathogens. The studies that assessed aerosolized hydrogen peroxide found a greater log kill with the use of vaporized hydrogen peroxide generators. The overarching conclusion was that hydrogen peroxide delivered as vaporized hydrogen peroxide was an effective method to achieve large levels of log kill on the assessed pathogens. The hydrogen peroxide vapor generators can play a role in dental bio-decontamination. The parameters must be standardized and the efficacy assessed to perform bio-decontamination for the whole clinic. For vaporized hydrogen peroxide generators to be included in the dental bio-decontamination regimen, certain criteria should be met. These include the standardization and efficacy assessment of the vaporized hydrogen peroxide generators in dental clinics.

Using Data Mining and Network Analysis to Infer Arboviral Dynamics: The Case of Mosquito-Borne Flaviviruses Reported in Mexico.

Simple SummaryGiven the significant impact on both human and animal health of mosquito-borne flaviviruses, a better understanding of their transmission cycles, viewed as a complex multi pathogen-vector-host system is urgently required. Here, we use a spatial datamining framework, based on co-occurrence data that includes biotic niche variables to create models for Dengue, Yellow fever, West Nile Virus and St. Louis encephalitis in Mexico that predict: (i) which mosquito species are likely to be the most important vectors for a given pathogen; (ii) which species are most likely to be important from a multi-pathogenic viewpoint; and (iii) which mosquito and/or mammal assemblages are most likely to play an important role in the transmission cycles. Our predictions are consistent with known information about the dynamics of these mosquito-borne flaviviruses and predict new potential vectors. Our approach can improve disease surveillance efforts and generate useful information regarding public health and biodiversity conservation.Given the significant impact of mosquito-borne flaviviruses (MBFVs) on both human and animal health, predicting their dynamics and understanding their transmission cycle is of the utmost importance. Usually, predictions about the distribution of priority pathogens, such as Dengue, Yellow fever, West Nile Virus and St. Louis encephalitis, relate abiotic elements to simple biotic components, such as a single causal agent. Furthermore, focusing on single pathogens neglects the possibility of interactions and the existence of common elements in the transmission cycles of multiple pathogens. A necessary, but not sufficient, condition that a mosquito be a vector of a MBFV is that it co-occurs with hosts of the pathogen. We therefore use a recently developed modeling framework, based on co-occurrence data, to infer potential biotic interactions between those mosquito and mammal species which have previously been identified as vectors or confirmed positives of at least one of the considered MBFVs. We thus create models for predicting the relative importance of mosquito species as potential vectors for each pathogen, and also for all pathogens together, using the known vectors to validate the models. We infer that various mosquito species are likely to be significant vectors, even though they have not currently been identified as such, and are likely to harbor multiple pathogens, again validating the predictions with known results. Besides the above “niche-based” viewpoint we also consider an assemblage-based analysis, wherein we use a community-identification algorithm to identify those mosquito and/or mammal species that form assemblages by dint of their significant degree of co-occurrence. The most cohesive assemblage includes important primary vectors, such as A. aegypti, A. albopictus, C. quinquefasciatus, C. pipiens and mammals with abundant populations that are well-adapted to human environments, such as the white-tailed deer (Odocoileus virginianus), peccary (Tayassu pecari), opossum (Didelphis marsupialis) and bats (Artibeus lituratus and Sturnira lilium). Our results suggest that this assemblage has an important role in the transmission dynamics of this viral group viewed as a complex multi-pathogen-vector-host system. By including biotic risk factors our approach also modifies the geographical risk profiles of the spatial distribution of MBFVs in Mexico relative to a consideration of only abiotic niche variables.