Detection Of Potential Pathogens Research Articles

A three-years survey of microbial contaminants in industrial hemp inflorescences from two Italian cultivation sites

BackgroundThe use of industrial Cannabis sativa L. for recreational, cosmeceutical, nutraceutical, and medicinal purposes has gained momentum due to its rich content of valuable phytochemicals, such as cannabidiol (CBD) and cannabigerol (CBG). However, there are concerns regarding the risk of microbial contamination in plants grown outside controlled environments. Microbes associated with hemp can be either epiphytes or endophytes and may pose a risk of infectious illness for humans.MethodsSeven Italian hemp genotypes, including Bernabeo, Carmagnola, Carmaleonte, Codimono, CS, Eletta Campana, and Fibranova, were cultivated in two distinct geographic locations, Catania and Rovigo, for three consecutive years from 2019 to 2021. Total aerobic microbes (TAMC), total combined yeasts/moulds (TYMC), the presence of bile-tolerant Gram-negative bacteria, and the absence of Escherichia coli and Salmonella spp. were evaluated and compared. The main phytocannabinoid content was measured and correlated with microbial contamination.ResultsMost samples analyzed in this study did not meet the European Pharmacopoeia microbiological limits. The detection of potential pathogens, such as E. coli and Salmonella spp., in the samples indicates that the use of inflorescences may represent a possible source of infection. Microbial contamination varied among harvesting seasons and production sites, with agroclimatic conditions influencing microbial load and composition. The presence of potentially pathogenic bacteria was less associated with seasonal climate variability and more likely affected by sporadic contamination from external sources. CBD concentration exhibited a negative correlation with bile-tolerant Gram-negative bacteria and total yeasts/moulds levels. Samples with lower CBD content were more contaminated than those with higher CBD levels, suggesting a potential protective effect of this phytochemical on the plant.ConclusionsThe threshing residues (inflorescences, floral bracts, and leaves) of industrial hemp varieties represent a valuable product and a source of beneficial phytochemicals that warrants further exploration. While post-harvest sterilization methods may reduce microbiological risks, they may also degrade heat- and light-sensitive bioactive phytochemicals. The most promising strategy involves implementing best agronomic practices to maintain healthy and uncontaminated cultures. Rigorous monitoring and quality certification protocols are essential to mitigate the microbiological risk associated with the consumption of hemp-derived products.

“Potential pathogens detection in Spodoptera frugiperda larvae maintained under laboratory conditions”

The microbiome of Spodoptera frugiperda plays a fundamental role during its biological cycle, since a great diversity of microorganisms interact through different mechanisms to maintain homeostasis. Their functions include, for example, defense against pathogens, degradation and the supply of functional molecules, among others. However, factors such as changes in diet, the use of insecticides, antibiotics, environmental factors, age or sex can interfere with the microbiome causing dysbiosis. Therefore, the objective of this work was to perform a metagenomic analysis of healthy and dead larvae with symptoms of infection to detect potential pathogenic agents. The results show that the genus Enterococcus presented an average relative abundance of 89.2% in healthy larvae and an average of 94.75% in dead larvae. The presence of Pseudomonas was also detected, with an average relative abundance of 10.8% in healthy larvae and 0.5% in dead larvae. While Acetobacter presented an average relative abundance in dead larvae of 4.5%. Interestingly, readings corresponding to viruses were 0.004% in healthy larvae and 1.48% in dead larvae, which were related to pathogenic viruses of the baculovirus family. The detection of potential pathogens that can affect S. frugiperda during rearing is of most importance to avoid their dispersal or affecting the colony.

Associations between indicators of new semi-quantitative express-test for procalcitonin and indicators of saturation in patients with community-acquired pneumonia

Objective. To evaluate the associations between the indicators of a new semi-quantitative express-test for procalcitonin and saturation levels in hospitalized patients with community-acquired pneumonia.
 Due to a high rate of growth of the incidence of severe community-acquired pneumonia, which leads to disability of the working population and claims thousands of lives, there is a need for early verification of the severity degree and a timely initiation of treatment. Traditional clinical, radiologic, laboratory and bacteriological criteria for inflammatory reactions (fever, increased pulmonary infiltration and leukocytosis, detection of potential pathogens in sputum or hemocultures) are not specific for pneumonia. Similar clinical and radiologic manifestations can also be noted in other pathological conditions, for example, in atelectasis of the lung, lung infarction, congestive heart failure, as well as acute respiratory distress syndrome. In this regard, the study of new diagnostic markers of community-acquired pneumonia is an actual direction in modern therapeutic diagnostics. Being a fast and highly sensitive method, the express test for procalcitonin can be a valuable predictive marker for the severe course of community-acquired pneumonia. Of interest is the relationship of procalcitonin with an indicator reflecting the presence of respiratory failure the level of saturation.
 Materials and methods. A single-stage comparative study involved 123 patients admitted to the hospital with a confirmed diagnosis of community-acquired pneumonia. On the first day after admission to the hospital, all patients underwent a general clinical examination, an overview X-ray of the chest organs, and determination of the plasma procalcitonin level using a semi-quantitative express-test as well as the saturation level. According to the results of the examination, the patients were divided into two groups: patients with saturation 94 % (n=43), and patients with saturation 94 % (n=80). The group of patients with saturation values 94 % consisted of 23 (53.5 %) women and 20 (46.5 %) men. The group of patients with saturation values 94 % consisted of 42 (52.5 %) women and 38 (47.5 %) men. The mean age of the patients was 56.719.1 and 45.620.7 (MSD) years, respectively, p=0.005. Statistical processing of the obtained data was carried out using the SPSS 13.0 software package.
 Results. Patients with low saturation values ( 94 %) had higher heart rate (p=0.001) and respiratory rate (p=0.001), and the levels of erythrocytes (p=0.001), hemoglobin (p=0.003) and albumin (p=0.001) were significantly lower than in patients with higher saturation levels 94 %. For patients with signs of respiratory failure and saturation indices 94 %, a more pronounced increase in the level of express-test for procalcitonin was characteristic (p=0.001). According to regression analysis, the express-test for procalcitonin 2 ng/ml by 13.8 % (CI 1.5131.3), age by 3 %, and heart rate by 11 % increase the risk of low saturation in patients with community-acquired pneumonia.
 Conclusion. Low values of blood oxygen saturation 94 % in hospitalized patients with community-acquired pneumonia are directly associated with high values of the new semi-quantitative express-test for procalcitonin (more than 2 ng/ml), that can be used for individual risk stratification.

Time-series prediction and detection of potential pathogens in bloodstream infection using mcfDNA sequencing.

Next-generation sequencing of microbial cell free DNA (mcfDNA-seq) has emerged as a promising diagnostic method for blood stream infection (BSI) and offers the potential to detect pathogens before blood culture. However, its application is limited by a lack of clinical validation. We conducted sequential mcfDNA-seq on blood samples from ICU participants at high risk of BSI due to pneumonia, or intravascular catheterization; and explored whether mcfDNA-seq could diagnose and detect pathogens in advance of blood culture positivity. Blood culture results were used as evaluation criteria. A total of 111 blood samples were collected during the seven days preceding and on the day of onset of 16 BSI episodes from 13 participants. The diagnostic and total predictive sensitivity of mcfDNA-seq were 90% and 87.5%, respectively. The proportion of pathogenic bacteria was relatively high in terms of both diagnosis and prediction. The reads per million of etiologic agents trended upwards in the days approaching the onset of BSI. Our work found that mcfDNA-seq has high diagnostic sensitivity and could be used to identify pathogens before the onset of BSI, which could help expand the clinical application of mcfDNA-seq.

Biological and chemical contamination of illegal, uncontrolled refuse storage areas in Poland

This study focused on assessing the microbiological and chemical contamination of air, soil and leachate in uncontrolled refuse storage areas in central Poland. The research included an analysis of the number of microorganisms (culture method), endotoxin concentration (gas chromatography-mass spectrometry), heavy metals level (atomic absorption spectrometry), elemental characteristics (elemental analyser), cytotoxicity assessment against A-549 (human lung) and Caco-2 (human colon adenocarcinoma) cell lines (PrestoBlue™ test) and toxic compound identification (ultra-high-performance liquid chromatography-quadrupole time-of-flight ultrahigh-resolution mass spectrometry).Microbial contamination differed depending on the dump and the group of tested microorganisms. The number of bacteria was: 4.3 × 102 – 1.8 × 103 CFU m−3 (air); 1.1 × 103 – 1.2 × 106 CFU mL−1 (leachate); 1.0 × 106 – 3.9 × 106 CFU g−1 (soil). Respectively, for air and soil the number of fungi was: 2.2 × 102 – 4.6 × 102 CFU m−3; 1.8 × 102 – 3.9 × 103 CFU g−1.Metal levels (Fe, Mn, Pb, Zn, Al, Hg, Cd, Cu, Cr) were higher than in the control sample; however, the average concentrations did not exceed the permissible standards.The cytotoxicity of soil and leachate samples depended on the dump, sample and cell line tested. The leachates were more cytotoxic than soil extracts. Compounds belonging to pesticides, surfactants and biocides, chemicals and/or polymer degradation products, medicinal drugs and insect repellents were found.The detection of potential pathogens in the air, soil and leachate, the presence of toxic compounds and the confirmation of the cytotoxic effect of leachate and soil on human cell lines justify the need for further research on the risks posed by illegal dumps. These studies should aim at developing a unified assessment method and a method to minimise the risk of contaminants spreading in the environment, including harmful biological agents.

The Arabidopsis E3 ubiquitin ligase PUB4 regulates BIK1 and is targeted by a bacterial type-III effector.

Plant immunity is tightly controlled by a complex and dynamic regulatory network, which ensures optimal activation upon detection of potential pathogens. Accordingly, each component of this network is a potential target for manipulation by pathogens. Here, we report that RipAC, a type III-secreted effector from the bacterial pathogen Ralstonia solanacearum, targets the plant E3 ubiquitin ligase PUB4 to inhibit pattern-triggered immunity (PTI). PUB4 plays a positive role in PTI by regulating the homeostasis of the central immune kinase BIK1. Before PAMP perception, PUB4 promotes the degradation of non-activated BIK1, while after PAMP perception, PUB4 contributes to the accumulation of activated BIK1. RipAC leads to BIK1 degradation, which correlates with its PTI-inhibitory activity. RipAC causes a reduction in pathogen-associated molecular pattern (PAMP)-induced PUB4 accumulation and phosphorylation. Our results shed light on the role played by PUB4 in immune regulation, and illustrate an indirect targeting of the immune signalling hub BIK1 by a bacterial effector.

Insights into the Bacterial Diversity and Detection of Opportunistic Pathogens in Mexican Chili Powder.

Chili powder is the most frequently consumed spice in Mexican diets. Thus, the dissemination of microorganisms associated with chili powder derived from Capsicum annuum L. is significant during microbial quality analysis, with special attention on detection of potential pathogens. The results presented here describe the initial characterization of bacterial community structure in commercial chili powder samples. Our results demonstrate that, within the domain Bacteria, the most abundant family was Bacillaceae, with a relative abundance of 99% in 71.4% of chili powder samples, while 28.6% of samples showed an average relative abundance of 60% for the Enterobacteriaceae family. Bacterial load for aerobic mesophilic bacteria (AMB) ranged from 104 to 106 cfu/g, while for sporulated mesophilic bacteria (SMB), the count ranged from 102 to 105 cfu/g. Bacillus cereus sensu lato (s.l.) was observed at ca. ˂600 cfu/g, while the count for Enterobacteriaceae ranged from 103 to 106 cfu/g, Escherichia coli and Salmonella were not detected. Fungal and yeast counts ranged from 102 to 105 cfu/g. Further analysis of the opportunistic pathogens isolated, such as B. cereus s.l. and Kosakonia cowanii, using antibiotic-resistance profiles and toxinogenic characteristics, revealed the presence of extended-spectrum β-lactamases (ESBLs) and Metallo-β-lactamases (MBLs) in these organisms. These results extend our knowledge of bacterial diversity and the presence of opportunistic pathogens associated with Mexican chili powder and highlight the potential health risks posed by its use through the spread of antibiotic-resistance and the production of various toxins. Our findings may be useful in developing procedures for microbial control during chili powder production.

The cellular community in the intestine of the shrimp <i>Penaeus penicillatus</i> and its culture environments

In the present study, bacterial community diversities were investigated in the shrimp intestines, outside seawater and culture-associated environments using clone library analysis. Comparative analyses revealed that the bacterial community in the intestinal contents was most similar to that of the sediment. Bacteria in the phyla Proteobacteria and Firmicutes constituted the dominant components of the intestinal bacteria in the shrimp. The potential pathogens and probiotics were dominant members of the intestine in healthy or diseased shrimp. Intestinal bacterial communities in the healthy shrimp were composed of Bacillales, Rhodobacterales, Vibrionales, and Alteromonadales, whereas fewer bacterial groups were observed in the intestines of diseased shrimp, which were dominated by Vibrionales and Bacillales. Phylogenetic analysis revealed that nearly half of the sequences in the intestines of diseased shrimp were closely related to the potential pathogen Photobacterium damselae and Vibrio harveyi. Obvious differences in microbial diversity were also observed between outside seawater and culture-associated environments. Eukaryotes in the outside seawater were dominated by Skeletonema, Cyclotella, Thalassiosira, Guinardia, and Chaetoceros. The pond water showed less diversity in the eukaryotic community and was dominated by Skeletonema, Thalassiosira, Chaetoceros, and Oithona. The implications of the present study may be significant in the management of shrimp cultivation, as well as the detection of potential pathogens and the application of probiotics.

An examination of the microbial community and occurrence of potential human pathogens in rainwater harvested from different roofing materials

While harvested rainwater can serve as an alternative water supply, microbial contaminants within the collection system can negatively affect water quality. Here, we investigated the impact of roofing material on the microbial quality of rainwater freshly harvested from pilot-scale roofs (concrete tile, cool, green, Galvalume® metal, and asphalt fiberglass shingle). The microbial quality of freshly harvested rainwater from six rain events over two years was analyzed by high-throughput sequencing and culture-dependent and -independent techniques. The concentrations of total coliform were significantly different among rainwaters harvested from the various roofing materials (p-value >0.05). However, the fecal coliform concentrations and the copy numbers of Enterococcus 23S rRNA genes and total Bacteria 16S rRNA genes did not vary by type of roofing material in a statistically significant way. Potential human pathogens such as Legionella, Escherichia coli O157:H7, Shiga-toxin-producing Escherichia coli, and adenovirus were detected at least once in rainwater harvested from the different roofing materials, even though the lowest occurrence of those potential human pathogens was noted from the metal roof. Also, substantial variation in the microbial communities from the different roofing materials was observed at the family and genus levels. These results demonstrate that the type of roofing material affects the microbial quality of freshly harvested rainwater, indicating that the choice of roofing material could shape the microbial community structure entering a rainwater storage tank. Given that detection of potential pathogens in the freshly harvested rainwater also differed between roofing materials, the type of roofing used to capture rainwater needs to be considered in rainwater harvesting system design, particularly if the water is intended for potable use.

The cellular community in the intestine of the shrimp Penaeus penicillatus and its culture environments

In the present study, bacterial community diversities were investigated in the shrimp intestines, outside seawater and culture-associated environments using clone library analysis. Comparative analyses revealed that the bacterial community in the intestinal contents was most similar to that of the sediment. Bacteria in the phyla Proteobacteria and Firmicutes constituted the dominant components of the intestinal bacteria in the shrimp. The potential pathogens and probiotics were dominant members of the intestine in healthy or diseased shrimp. Intestinal bacterial communities in the healthy shrimp were composed of Bacillales, Rhodobacterales, Vibrionales, and Alteromonadales, whereas fewer bacterial groups were observed in the intestines of diseased shrimp, which were dominated by Vibrionales and Bacillales. Phylogenetic analysis revealed that nearly half of the sequences in the intestines of diseased shrimp were closely related to the potential pathogen Photobacterium damselae and Vibrio harveyi. Obvious differences in microbial diversity were also observed between outside seawater and culture-associated environments. Eukaryotes in the outside seawater were dominated by Skeletonema, Cyclotella, Thalassiosira, Guinardia, and Chaetoceros. The pond water showed less diversity in the eukaryotic community and was dominated by Skeletonema, Thalassiosira, Chaetoceros, and Oithona. The implications of the present study may be significant in the management of shrimp cultivation, as well as the detection of potential pathogens and the application of probiotics.

Molecular Analysis of Bacterial Communities and Detection of Potential Pathogens in a Recirculating Aquaculture System for Scophthalmus maximus and Solea senegalensis

The present study combined a DGGE and barcoded 16S rRNA pyrosequencing approach to assess bacterial composition in the water of a recirculating aquaculture system (RAS) with a shallow raceway system (SRS) for turbot (Scophthalmus maximus) and sole (Solea senegalensis). Barcoded pyrosequencing results were also used to determine the potential pathogen load in the RAS studied. Samples were collected from the water supply pipeline (Sup), fish production tanks (Pro), sedimentation filter (Sed), biofilter tank (Bio), and protein skimmer (Ozo; also used as an ozone reaction chamber) of twin RAS operating in parallel (one for each fish species). Our results revealed pronounced differences in bacterial community composition between turbot and sole RAS, suggesting that in the systems studied there is a strong species-specific effect on water bacterial communities. Proteobacteria was the most abundant phylum in the water supply and all RAS compartments. Other important taxonomic groups included the phylum Bacteriodetes. The saltwater supplied displayed a markedly lower richness and appeared to have very little influence on bacterial composition. The following potentially pathogenic species were detected: Photobacterium damselae in turbot (all compartments), Tenacibaculum discolor in turbot and sole (all compartments), Tenacibaculum soleae in turbot (all compartments) and sole (Pro, Sed and Bio), and Serratia marcescens in turbot (Sup, Sed, Bio and Ozo) and sole (only Sed) RAS. Despite the presence of these pathogens, no symptomatic fish were observed. Although we were able to identify potential pathogens, this approach should be employed with caution when monitoring aquaculture systems, as the required phylogenetic resolution for reliable identification of pathogens may not always be possible to achieve when employing 16S rRNA gene fragments.

Effects of Differences in Lipid A Structure on TLR4 Pro-Inflammatory Signaling and Inflammasome Activation

The vertebrate immune system exists in equilibrium with the microbial world. The innate immune system recognizes pathogen-associated molecular patterns via a family of Toll-like receptors (TLR) that activate cells upon detection of potential pathogens. Because some microbes benefit their hosts, mobilizing the appropriate response, and then controlling that response is critical in the maintenance of health. TLR4 recognizes the various forms of lipid A produced by Gram-negative bacteria. Depending on the structural form of the eliciting lipid A molecule, TLR4 responses range from a highly inflammatory endotoxic response involving inflammasome and other pro-inflammatory mediators, to an inhibitory, protective response. Mounting the correct response against an offending microbe is key to maintaining health when exposed to various bacterial species. Further study of lipid A variants may pave the way to understanding how TLR4 responses are generally able to avoid chronic inflammatory damage.