MALDI-TOF mass spectrometry of saliva samples as a prognostic tool for COVID-19

The SARS-CoV-2 infections are still imposing a great public health challenge despite the recent developments in vaccines and therapy. Searching for diagnostic and prognostic methods that are fast, low-cost and accurate is essential for disease control and patient recovery. The MALDI-TOF mass spectrometry technique is rapid, low cost and accurate when compared to other MS methods, thus its use is already reported in the literature for various applications, including microorganism identification, diagnosis and prognosis of diseases. Here we developed a prognostic method for COVID-19 using the proteomic profile of saliva samples submitted to MALDI-TOF and machine learning algorithms to train models for COVID-19 severity assessment. We achieved an accuracy of 88.5%, specificity of 85% and sensitivity of 91.5% for classification between mild/moderate and severe conditions. Then, we tested the model performance in an independent dataset, we achieved an accuracy, sensitivity and specificity of 67.18, 52.17 and 75.60% respectively. Saliva is already reported to have high inter-sample variation; however, our results demonstrates that this approach has the potential to be a prognostic method for COVID-19. Additionally, the technology used is already available in several clinics, facilitating the implementation of the method. Further investigation using a bigger dataset is necessary to consolidate the technique.

Microbiological analysis of the Cirina forda (Lepidoptera: Saturniidae) commercialized in North Togo

Edible insects are presented as a source of protein, fat and micronutrients. Consequently, they are an attractive growing environment for microorganisms. Although, professionals in this sector in Sub-Saharan Africa use traditional harvesting, processing and marketing techniques, there is little scientific data on the microorganisms that infest edible insects. The aim of this study was to identify the microorganisms present in the Cirina forda supply chain, the most commercially marketed insect species in Togo. A total of 300 samples of fresh, processed and commercial C. forda caterpillars were collected and analyzed using standard microbiological microorganism identification techniques. The caterpillar was 90% contaminated with Staphylococcus spp, 60% by Escherichia coli, 40% by Enterobacter spp, 40% by Aspergillus niger, 30% by Klebsiella pneumoniae and 10% by Mucor spp, Klebsiella oxytoca, Proteus spp, Serratia spp and Aspergillus flavus. Pathogenic microorganisms are found in all samples at all stages of the supply chain, so caterpillars handled using traditional methods are detrimental to the health of the consumer. The presence of germs indicative of contamination of the caterpillar analyzed exposes consumers to the risk of food poisoning. Keywords: Cirina forda, marketing, microorganisms, Togo

Species Composition of Microbiota in City Buses

Introduction: Regulatory documents of the Russian sanitary legislation provide no standardization of microbiological indicators for urban public transportation. At the same time, extensive studies of public transport microbiome conducted in different countries demonstrated that the external environment (stations and platforms) and the interior of vehicles contain a large number of various microorganisms, the number of which is independent of air temperature, route duration, or the number of passengers. The objective of our work was to study the species composition of microbiota in urban buses and to assess its structure using methods of synecological and epidemiological analysis. Methods: We analyzed wipe samples from generally accessible bus interior surfaces. In total, 41 buses of 16 routes from three conditional groups (“Nagornaya” (Upland), “Zarechnaya” (Transverse) and “Interdistrict”) were examined. We applied methods of classical bacteriology, MALDI-TOF mass spectrometry for microorganism identification, and synecological analysis. Results and discussion: We identified 85 types of microorganisms, 15 of which were found in all groups of routes. The bus microbiota followed the same ecological laws as natural microbiocenoses. The greatest number of species was represented by genera Staphylococcus (16), Acinetobacter (11), Bacillus (11), Pseudomonas (8), and Pantoea (5). The majority of identified microorganisms belonged to genera Acinetobacter, Enterobacter, and Pantoea. Among 15 species of microorganisms “common” to three groups of buses, 60 % were representatives of human microbiota. Microorganisms were found in large quantities on fabric seat backs (up to 18,600 CFU/cm3), seat back handles (up to 76,500 CFU/cm3), leather loop-holders (up to 6,400 CFU/cm3), and visually dirty surfaces (11,200 CFU/cm3). Conclusions: Our findings indicate the need to standardize microbiological indicators and develop guidelines for monitoring the efficiency of disinfection of interiors of public passenger vehicles.

Indwelling Device-Associated Biofilms in Critically Ill Cancer Patients—Study Protocol

Health care-associated infections are a leading cause of inpatient complications. Rapid pathogen detection/identification is a major challenge in sepsis management that highly influences the successful outcome. The current standard of microorganism identification relies on bacterial growth in culture, which has several limitations. Gene sequencing research has developed culture-independent techniques for microorganism identification, with the aim to improve etiological diagnosis and, therefore, to change sepsis outcome. A prospective, observational, non-interventional, single-center study was designed that assesses biofilm-associated pathogens in a specific subpopulation of septic critically ill cancer patients. Indwelling device samples will be collected in septic patients at the moment of the removal of the arterial catheter, central venous catheter, endotracheal tube and urinary catheter. Concomitantly, clinical data regarding 4 sites (nasal, pharyngeal, rectal and skin) of pathogen colonization at the time of hospital/intensive care admission will be collected. The present study aims to offer new insights into biofilm-associated infections and to evaluate the infection caused by catheter-specific and patient-specific biofilm-associated pathogens in association with the extent of colonization. The analysis relies on the two following detection/identification techniques: standard microbiological method and next generation sequencing (NGS). Retrospectively, the study will estimate the clinical value of the NGS-based detection and its virtual potential in changing patient management and outcome, notably in the subjects with missing sepsis source or lack of response to anti-infective treatment.

Human Skin Microbiome: Impact of Intrinsic and Extrinsic Factors on Skin Microbiota

The skin is the largest organ of the human body and it protects the body from the external environment. It has become the topic of interest of researchers from various scientific fields. Microorganisms ensure the proper functioning of the skin. Of great importance, are the mutual relations between such microorganisms and their responses to environmental impacts, as dysbiosis may contribute to serious skin diseases. Molecular methods, used for microorganism identification, allow us to gain a better understanding of the skin microbiome. The presented article contains the latest reports on the skin microbiota in health and disease. The review discusses the relationship between a properly functioning microbiome and the body’s immune system, as well as the impact of internal and external factors on the human skin microbiome.

Development of next-generation sequencing-based sterility test

The sterility testing methods described in pharmacopoeias require an incubation period of 14 days to obtain analysis results. An alternative method that can significantly shorten the detection time and improve the accuracy is in urgent need to meet the sterility testing requirements of regenerative medicine products with a short shelf life. In this study, we developed the next-generation sequencing-based sterility test (NGSST) based on sequencing and multiple displacement amplification. The NGSST can be finished within 48 hours with five steps including whole genome amplification, sequencing, alignment, sterility testing report, and microorganism identification. We use RPKM ratio to minorize the influence of environmental bacteria and determine its cutoff based AUC curve. The NGSST showed high sensitivity in reporting contaminates at 0.1 CFU in supernatant of biological product or 1 CFU in cell suspension. Furthermore, we identified microorganisms in 5 primary umbilical cord mesenchymal stem cell samples that were tested positive by BacT/ALERTR 3D. Overall, the NGSST can serve as a promising alternative for sterility testing of biological products.

Elucidation of Practices of Mobile Phone Hygiene and Identification of the Microorganisms: A Perspective Study from Riyadh, Saudi Arabia

Mobile phones (MPs) have become a tool for the transmission of microorganisms due to lack of personal hygiene and maybe the sharing of the mobile phone by more than one person that which leads it to be a suitable carrier for microbes. This study aimed to draw a bead on the practices of hygiene of MPs among people living in Riyadh, Saudi Arabia, by performing a cross-sectional survey of 204 participants. The response rate of this study was above ~95%. Results showed that 19.6 % of responders clean their MPs once in a day, but the majority (33.8%) never cleaned their MPs. More than a quarter of the respondents (28.4%) use tissue paper to clean MPs. Among the users, ~60% use MPs while eating, 76% realized that their MPs might be resource of transmitting microorganisms. The study was also carried out using standard techniques to identify and count the bacterial contamination using the MPs. A combined number of 75 MPs of the participants in the shopping malls of Riyadh were screened for microorganism identification. From 75 public MPs, 109 bacteria were isolated. Coagulase-negative staphylococci were the predominant organisms isolate (76.1%) and with 1.8% by Staphylococcus aureus. Micrococcus sps. was also found (12.8%). A small number of Kocuria sps. were also isolated (4.6%). These results showed that common peoples’ mobile phones were contaminated with various types of microorganisms. The results provide an evidence base for the development and enhancement of hygienic MPs using practices.

Rapid microbiological testing method

Classical microbiological methods currently have unacceptably long cycle times. Rapid microbiological strategies are accessible within the marketplace for about 10 years and are mostly used in the clinical laboratory and in food industries. However, their reapplication in the pharma industry has wide range of advantage. A comparison with ancient strategies to be conjointly performed. During this review, data concerning the validation of RMM strategies described within the document was given in addition as proof of the issue of validation of those strategies. A comparison with ancient strategies is additionally mentioned. This data is beneficial to the industries and in the labs which will doubtless be adopted. These strategies for microorganism free products. This methodology for microorganism identification will be delicate, accurate and fast. How the laboratory should be maintained for carrying out different tests, there should be good hygienic condition maintained. This article also includes different methods for identifying bacteria which is present in drug products as well as the material which are used for doing test. Presence of bacteria may affect the activity of drug product and bio availability may decreases and potency the drug may loss. How the laboratory should be maintained for carrying out different tests, there should be good hygienic condition maintained.

Preliminary Evidence of a Molecular Detection Method to Analyze Bacterial DNA as a Quality Indicator in Cosmetics

Cosmetics are a category of widely consumed and distributed products, and their manufacture is always subject to specific guidelines. Quality Control (QC) tests provide information supporting the absence of injurious organisms and regarding the microbiological stability of cosmetics. The microbiological risk analysis is typically performed using the plate count method, which is a time-consuming and operator-dependent approach. Molecular technologies allow a deeper and more sensitive testing than traditional cultures. The demand for rapid and sensitive methods is recently increasing. The aim of our study was to compare different DNA extraction methods in order to detect and quantify bacterial load in cosmetics using a qPCR system. Known numbers of microorganisms were spiked into six different cosmetics to simulate contaminated samples. DNA was extracted with seven extraction kits and then quantified by real-time qPCR. Results revealed differences in terms of cell recovery, DNA yield, and quality. The bead-beating approaches were the most suitable in our molecular workflow and lead to good quality DNA for analysis by qPCR within four hours. Combined with mechanical extraction, qPCR may represent an efficient and easy method for microorganism identification in cosmetics, and can be automated. This approach also is also applicable for the detection of probiotics used as beneficial biological components in cosmetic products. The results of our molecular method provided preliminary evidences for the rapid identification of cells (10–100) and nucleic acids in complex preparations employed for human health, in compliance with regulatory limits. The suggested methodology is easy, fast, and sensitive. Its scalability allows serial microbiological evaluation at every manufacturing step.

Ultrasound guided minimally invasive autopsies: combining imaging and pathology to characterize the systemic involvement of COVID-19 infection

Post mortem tissue sampling is of paramount importance to conduct in-situ and molecular studies of COVID-19. Ultrasound-based minimally invasive autopsies (MIA/US) is an inexpensive procedure to obtain tissue samples of several organs and, at the same time, reduce the risks of the autopsy procedure in situations of high contagiousness. The images obtained by ultrasound are good enough to localize and orient the sampling, and to select the most affected areas within each organ. The amount of tissue sampled is adequate for histological and molecular studies and microorganism identification, and delivers information within a rapid time window. The combination of the aforementioned aspects may extend the possibility of conducting autopsies in different parts of the world, perhaps helping to understand local characteristics of COVID-19 infections, within an extended range of genetic, social and economic diversity.