Microorganisms, Volume 12, Issue 5 (May 2024) – 92 articles

Pyridoxine, also known as vitamin B₆, is an essential cofactor in numerous cellular processes. Its importance in various applications has led to a growing interest in optimizing its production through microbial biosynthesis. However, an imbalance in the net production of NADH disrupts intracellular cofactor levels, thereby limiting the efficient synthesis of pyridoxine. In our study, we focused on multiple cofactor engineering strategies, including the enzyme design involved in NAD⁺-dependent enzymes and NAD⁺ regeneration through the introduction of heterologous NADH oxidase (Nox) coupled with the reduction in NADH production during glycolysis. Finally, the engineered E. coli achieved a pyridoxine titer of 676 mg/L in a shake flask within 48 h by enhancing the driving force. Overall, the multiple cofactor engineering strategies utilized in this study serve as a reference for enhancing the efficient biosynthesis of other target products. Full article

African horse sickness is a severe and often fatal disease affecting all species of equids. The aetiological agent, African horse sickness virus (AHSV), can be differentiated into nine serotypes. The identification of AHSV serotypes is vital for disease management, as this can influence vaccine selection and help trace disease incursion routes. In this study, we report the development and optimisation of a novel, molecular-based assay that utilises multiplex PCR and microsphere-based technology to expedite detection and differentiation of multiple AHSV serotypes in one assay. We demonstrated the ability of this assay to identify all nine AHSV serotypes, with detection limits ranging from 1 to 277 genome copies/µL depending on the AHSV serotype. An evaluation of diagnostic sensitivity and specificity revealed a sensitivity of 88% and specificity of 100%. This method can serotype up to 42 samples per run and can be completed in approximately 4–6 h. It provides a powerful tool to enhance the rapidity and efficiency of AHSV serotype detection, thereby facilitating the generation of epidemiological data that can help understand and control the incidence of AHSV worldwide. Full article

The role of Bifidobacterium species and microbial metabolites such as short-chain fatty acids (SCFAs) and human milk oligosaccharides in controlling intestinal inflammation and the pathogenesis of obesity and type 1 diabetes (T1D) has been largely studied in recent years. This paper discusses the discovery of signature biomarkers for obesity and T1D based on data from a novel test for profiling several Bifidobacterium species, combined with metabolomic analysis. Through the NUTRISHIELD clinical study, a total of 98 children were recruited: 40 healthy controls, 40 type 1 diabetics, and 18 obese children. Bifidobacterium profiles were assessed in stool samples through an innovative test allowing high taxonomic resolution and precise quantification, while SCFAs and branched amino acids were measured in urine samples through gas chromatography–mass spectrometry (GC-MS). KIDMED questionnaires were used to evaluate the children’s dietary habits and correlate them with the Bifidobacterium and metabolomic profiles. We found that B. longum subs. infantis and B. breve were higher in individuals with obesity, while B. bifidum and B. longum subs. longum were lower compared to healthy individuals. In individuals with T1D, alterations were found at the metabolic level, with an overall increase in the level of the most measured metabolites. The high taxonomic resolution of the Bifidobacterium test used meant strong correlations between the concentrations of valine and isoleucine, and the relative abundance of some Bifidobacterium species such as B. longum subs. infantis, B. breve, and B. bifidum could be observed. Full article

The competitive colonization of bacteria on similar ecological niches has a significant impact during their establishment. The synthesis speeds of different chemical classes of molecules during early competitive colonization can reduce the number of competitors through metabolic effects. In this work, we demonstrate for the first time that Kosakonia cowanii Cp1 previously isolated from the seeds of Capsicum pubescens R. P. produced volatile organic compounds (VOCs) during competitive colonization against Pectobacterium aroidearum SM2, affecting soft rot symptoms in serrano chili (Capsicum annuum L.). The pathogen P. aroidearum SM2 was isolated from the fruits of C. annuum var. Serrano with soft rot symptoms. The genome of the SM2 strain carries a 5,037,920 bp chromosome with 51.46% G + C content and 4925 predicted protein-coding genes. It presents 12 genes encoding plant-cell-wall-degrading enzymes (PCDEWs), 139 genes involved in five types of secretion systems, and 16 genes related to invasion motility. Pathogenic essays showed soft rot symptoms in the fruits of C. annuum L., Solanum lycopersicum, and Physalis philadelphica and the tubers of Solanum tuberosum. During the growth phases of K. cowanii Cp1, a mix of VOCs was identified by means of HS-SPME-GC-MS. Of these compounds, 2,5-dimethyl-pyrazine showed bactericidal effects and synergy with acetoin during the competitive colonization of K. cowanii Cp1 to completely reduce soft rot symptoms. This work provides novel evidence grounding a better understanding of bacterial interactions during competitive colonization on plant tissue, where VOC synthesis is essential and has a high potential capacity to control pathogenic microorganisms in agricultural systems. Full article

Plasmids are mobile genetic elements known to carry secondary metabolic genes that affect the fitness and survival of microbes in the environment. Well-studied cases of plasmid-encoded secondary metabolic genes in marine habitats include toxin/antitoxin and antibiotic biosynthesis/resistance genes. Here, we examine metagenome-assembled genomes (MAGs) from the permanently-stratified water column of the Cariaco Basin for integrated plasmids that encode biosynthetic gene clusters of secondary metabolites (smBGCs). We identify 16 plasmid-borne smBGCs in MAGs associated primarily with Planctomycetota and Pseudomonadota that encode terpene-synthesizing genes, and genes for production of ribosomal and non-ribosomal peptides. These identified genes encode for secondary metabolites that are mainly antimicrobial agents, and hence, their uptake via plasmids may increase the competitive advantage of those host taxa that acquire them. The ecological and evolutionary significance of smBGCs carried by prokaryotes in oxygen-depleted water columns is yet to be fully elucidated. Full article

To measure associations between gardening with different compost amendments and the human gut microbiota composition, gardeners (n = 25) were provided with one of three types of compost: chicken manure (CM), dairy manure and plant material (DMP), or plant-based (P). Stool samples were collected before gardening (T1), after compost amendment (T2), and at peak garden harvest (T3). Compost and soil samples were collected. DNA was extracted, 16S rRNA libraries were established, and libraries were sequenced by Illumina MiSeq. Sequences were processed using mothur, and data were analyzed in R software version 4.2.2. Fast expectation-maximization microbial source tracking analysis was used to determine stool bacteria sources. At T2/T3, the gut microbiotas of P participants had the lowest Shannon alpha diversity, which was also the trend at T1. In stool from T2, Ruminococcus 1 were less abundant in the microbiotas of those using P compost as compared to those using CM or DMP. At T2, Prevotella 9 had the highest abundance in the microbiotas of those using CM compost. In participants who used CM compost to amend their gardening plots, a larger proportion of the human stool bacteria were sourced from CM compared to soil. Soil exposure through gardening was associated with a small but detectable change in the gardeners’ gut microbiota composition. These results suggest that human interactions with soil through gardening could potentially impact health through alterations to the gut microbiota. Full article

Candida auris has been identified by the World Health Organization (WHO) as a critical priority pathogen on its latest list of fungi. C. auris infections are reported in the bloodstream and less commonly in the cerebrospinal fluid and abdomen, with mortality rates that range between 30% and 72%. However, no large-scale epidemiology studies have been reported until now. The diagnosis of C. auris infections can be challenging, particularly when employing conventional techniques. This can impede the early detection of outbreaks and the implementation of appropriate control measures. The yeast can easily spread between patients and in healthcare settings through contaminated environments or equipment, where it can survive for extended periods. Therefore, it would be desirable to screen patients for C. auris colonisation. This would allow facilities to identify patients with the disease and take appropriate prevention and control measures. It is frequently unsusceptible to drugs, with varying patterns of resistance observed among clades and geographical regions. This review provides updates on C. auris, including epidemiology, clinical characteristics, genomic analysis, evolution, colonisation, infection, identification, resistance profiles, therapeutic options, prevention, and control. Full article

Because data on the fungal gut community structure of African children are scarce, we aimed to describe it by reanalysing rRNA ITS1 and ITS2 metabarcoding data from a study designed to assess the influence of microbiota in malaria susceptibility in Malian children from the Dogon country. More specifically, we aimed to establish the core gut mycobiome and compare the gut fungal community structure of breastfed children, aged 0–2 years, with other age groups. Briefly, DNA was extracted from 296 children’s stool samples. Both rRNA ITS1 and ITS2 genomic barcodes were amplified and subjected to Illumina MiSeq sequencing. The ITS2 barcode generated 1,975,320 reads and 532 operational taxonomic units (OTUs), while the ITS1 barcode generated 647,816 reads and 532 OTUs. The alpha diversity was significantly higher by using the ITS1 compared to the ITS2 barcode (p < 0.05); but, regardless of the ITS barcode, we found no significant difference between breastfed children, aged 0–2 years, compared to the other age groups. The core gut mycobiome of the Malian children included Saccharomyces cerevisiae, Candida albicans, Pichia kudriavzevii, Malassezia restricta, Candida tropicalis and Aspergillus section Aspergillus, which were present in at least 50% of the 296 children. Further studies in other African countries are warranted to reach a global view of African children’s core gut mycobiome. Full article

The integumentary system of the leatherback sea turtle (Dermochelys coriacea) is the most visible and defining difference of the species, with its smooth and waxy carapace and finely scaled skin, distinguishing it from the other six sea turtle species. The skin is the body’s largest organ and serves as a primary defense against the outside world and is thus essential to health. To date, we have begun to understand that the microorganisms located on the skin aid in these functions. However, many host–microbial interactions are not yet fully defined or understood. Prior to uncovering these crucial host–microbial interactions, we must first understand the communities of microorganisms present and how they differ through life-stage classes and across the body. Here, we present a comprehensive bacterial microbial profile on the skin of leatherbacks. Using next-generation sequencing (NGS), we identified the major groups of bacteria on the skin of neonates at emergence, neonates at 3–4 weeks of age (i.e., post-hatchlings), and nesting females. These data show that the predominant bacteria on the skin of the leatherback are different at each life-stage class sampled. This suggests that there is a shift in the microbial communities of the skin associated with life-stage class or even possibly age. We also found that different sample locations on the nesting female (i.e., carapace and front appendages = flipper) have significantly different communities of bacteria present. This is likely due to differences in the microhabitats of these anatomic locations and future studies should explore if this variation also holds true for neonates. These data define baseline skin microbiota on the leatherback and can serve as a foundation for additional work to broaden our understanding of the leatherbacks’ host–microbial interactions, the impacts of environmental changes or stressors over time, and even the pathogenicity of disease processes. Full article

Animal bites are a common reason for children to visit primary care and emergency departments. Dog bites are the most prevalent, followed by cat bites at 20–30%. Other animals such as bats, monkeys, snakes, and rats collectively contribute less than 1% of cases. Hospitalization is necessary in only 4% of animal bite incidents. The main aim of this narrative review is to summarize the main protocols currently followed in pediatrics in cases involving the most common bites from different animal species. Analysis of the literature showed that the management of common animal bites in children presents a multifaceted challenge requiring a comprehensive understanding of the epidemiology, clinical presentation, and treatment modalities associated with each specific species. Effective wound management is paramount in reducing the risk of infection and promoting optimal healing outcomes. Additionally, tetanus vaccination status should be assessed and updated as necessary, and prophylactic antibiotics may be indicated in certain cases to prevent secondary infections. Furthermore, the role of rabies prophylaxis cannot be overstated, particularly in regions where rabies is endemic or following bites from high-risk animals. In addition to medical management, psychosocial support for both the child and their caregivers is integral to the overall care continuum. Future studies exploring the efficacy of novel treatment modalities, such as topical antimicrobial agents or advanced wound dressings, may offer new insights into optimizing wound healing and reducing the risk of complications. Full article

Beneficial/probiotic strains protect the host from pathogens by competitive displacement and production of antibacterial substances, i.e., bacteriocins. The antiparasitic potential of bacteriocins/enterocins and their producing strains in experimental murine trichinellosis were tested as a new therapeutic strategy. Enterocin M and Durancin-like and their producers Enterococcus faecium CCM8558 and Enterococcus durans ED26E/7 were administered daily to mice that were challenged with Trichinella spiralis. Our study confirmed the antiparasitic effect of enterocins/enterococci, which reduced the number of adults in the intestine (Enterocin M—43.8%, E. faecium CCM8558—54.5%, Durancin-like—16.4%, E. durans ED26E/7—35.7%), suppressed the Trichinella reproductive capacity ex vivo (Enterocin M—61%, E. faecium CCM8558—74%, Durancin-like—38%, E. durans ED26E/7—66%), and reduced the number of muscle larvae (Enterocin M—39.6%, E. faecium CCM8558—55.7%, Durancin-like—15%, E. durans ED26E/7—36.3%). The direct effect of enterocins on Trichinella fecundity was documented by an in vitro test in which Durancin-like showed a comparable reducing effect to Enterocin M (40–60%) in contrast to the ex vivo test. The reducing activity of T.spiralis infection induced by Enterocin M was comparable to its strain E. faecium CCM8558; Durancin-like showed lower antiparasitic activity than its producer E. durans ED26E/7. Full article

This study aims to report the efficacy of a combined intrastromal injection in optimizing the outcome of severe mycotic keratitis. Herein, we report a case series of 20 consecutive patients with positive fungal cultures not responding to topical antifungal treatment. Patients received cycles of intrastromal injections of voriconazole (50 µg/0.1 mL) and amphotericin B (2.5 µg/0.1 mL); all patients continued their topical antifungal therapy. The organisms isolated were Fusarium (n = 5), Aspergillus (n = 4), Candida (n = 4), Rhodotorula (n = 2), Penicillium (n = 2), Alternaria (n = 1), Bipolaris (n = 1), and Curvularia (n = 1). The size of the infiltrate varied from 6.5 to 1.5 mm. At presentation, the best corrected visual acuity (BCVA, namely, the best visual acuity achieved with glasses, if needed) was less than 20/400 in all patients, improving to better than 20/400 in eleven patients. Seven patients required surgical intervention; four of them underwent penetrating keratoplasty (PK) à chaud one month after the first intrastromal injection. Patients who underwent surgery achieved a BCVA of 20/40 or better. Combined intrastromal injections before therapeutic penetrating keratoplasty (TPK) effectively reduced ulcer size and graft diameter, preventing infection recurrence. Our results highlight the efficacy of combined intrastromal injections in optimizing outcomes for severe mycotic keratitis undergoing TPK. Full article

Bacillus velezensis is well known as a plant growth-promoting rhizobacteria (PGPR) and biocontrol agent. Nevertheless, there are very few reports on the study of B. velezensis on tomato early blight, especially the biocontrol effects among different inoculation concentrations. In this study, an IAA-producing strain, Bacillus velezensis YXDHD1-7 was isolated from the tomato rhizosphere soil, which had the strongest inhibitory effect against Alternaria solani. Inoculation with bacterial suspensions of this strain promoted the growth of tomato seedlings effectively. Furthermore, inoculations at 10⁶, 10⁷, and 10⁸ cfu/mL resulted in control efficacies of 100%, 83.15%, and 69.90%, respectively. Genome sequencing showed that it possesses 22 gene clusters associated with the synthesis of antimicrobial metabolites and genes that are involved in the production of IAA. Furthermore, it may be able to produce spermidine and volatile compounds that also enhance plant growth and defense responses. Our results suggest that strain YXDHD1-7 prevents early blight disease by promoting growth and enhancing the defense enzyme activities in tomato plants. This strain is a promising candidate for an excellent microbial inoculant that can be used to enhance tomato production. Full article

Enterovirus (EV) infections are widespread and associated with a range of clinical conditions, from encephalitis to meningitis, gastroenteritis, and acute flaccid paralysis. Knowledge about the circulation of EVs in neonatal age and early infancy is scarce, especially in Africa. This study aimed to unveil the frequency and diversity of EVs circulating in apparently healthy newborns from the Free State Province, South Africa (SA). For this purpose, longitudinally collected faecal specimens (May 2021–February 2022) from a cohort of 17 asymptomatic infants were analysed using metagenomic next-generation sequencing. Overall, seven different non-polio EV (NPEV) subtypes belonging to EV-B and EV-C species were identified, while viruses classified under EV-A and EV-D species could not be characterised at the sub-species level. Additionally, under EV-C species, two vaccine-related poliovirus subtypes (PV1 and PV3) were identified. The most prevalent NPEV species was EV-B (16/17, 94.1%), followed by EV-A (3/17, 17.6%), and EV-D (4/17, 23.5%). Within EV-B, the commonly identified NPEV types included echoviruses 6, 13, 15, and 19 (E6, E13, E15, and E19), and coxsackievirus B2 (CVB2), whereas enterovirus C99 (EV-C99) and coxsackievirus A19 (CVA19) were the only two NPEVs identified under EV-C species. Sabin PV1 and PV3 strains were predominantly detected during the first week of birth and 6–8 week time points, respectively, corresponding with the OPV vaccination schedule in South Africa. A total of 11 complete/near-complete genomes were identified from seven NPEV subtypes, and phylogenetic analysis of the three EV-C99 identified revealed that our strains were closely related to other strains from Cameroon and Brazil, suggesting global distribution of these strains. This study provides an insight into the frequency and diversity of EVs circulating in asymptomatic infants from the Free State Province, with the predominance of subtypes from EV-B and EV-C species. This data will be helpful to researchers looking into strategies for the control and treatment of EV infection. Full article

The principal objective of this study was to isolate and identify the microorganisms present in commercial kefir grains, a novel kefir-fermented coconut water (CWK) and a novel coconut water kefir-fermented sourdough using phenotypic identification and Sanger sequencing and examine the microbial diversity of CWK and CWK-fermented sourdough throughout the fermentation process using the MiSeq Illumina sequencing method. The phenotypic characterisation based on morphology identified ten isolates of LAB, five AAB and seven yeasts from kefir (K), CWK and CWK-fermented sourdough (CWKS). The results confirm the presence of the LAB species Limosilactobacillus fermentum, Lactobacillus. plantarum, L. fusant, L. reuteri and L. kunkeei; the AAB species Acetobacter aceti, A. lovaniensis and A. pasteurianus; and the yeast species Candida kefyr, Rhodotorula mucilaginosa, Saccharomyces cerevisiae, C. guilliermondii and C. colliculosa. To the best of our knowledge, the identification of Rhodotorula from kefir is being reported for the first time. This study provides important insights into the relative abundances of the microorganisms in CWKS. A decrease in pH and an increase in the titratable acidity for CWK- and CWK-fermented sourdough corresponded to the increase in D- and L-lactic acid production after 96 h of fermentation. Significant reductions in the pHs of CWK and CWKS were observed between 48 and 96 h of fermentation, indicating that the kefir microorganisms were able to sustain highly acidic environments. There was also increased production of L-lactic acid with fermentation, which was almost twice that of D-lactic acid in CWK. Full article

Cryptosporidium spp. and Microsporidia are opportunistic microorganisms with remarkable zoonotic transmission potential due to their capacity to infect humans and animals. The aim of this study was to evaluate the prevalence of these microorganisms in stool samples of animal and human origin. In total, 369 stool samples (205 from human patients with diarrhea and 164 of animal origin) were included in the study. Cryptosporidium spp. and Microsporidia presence were determined by using multiplex nested PCR. Positive results were analyzed by using Sanger sequencing of the amplicon, utilizing BLASTN and ClustalX software to confirm identification. Cryptosporidium spp. were found in 0.97% and 4.26% of human and animal samples, respectively. Enterocytozoon bieneusi was detected in human and animal stools in 6.82% and 3.05% of the samples, respectively. No associations were found when analyzing the presence of Cryptosporidium spp. and E. bieneusi and the demographic and clinical variables of patients and animals. This study demonstrates the presence of these microorganisms in human and animal samples from different species, and the most interesting findings are the detection of Cryptosporidium spp. in pets (e.g., rodents) that are not usually included in this type of study, and the identification of E. bieneusi in patients with diarrhea without underlying disease. Full article

Streptococcus suis (S. suis) is a zoonotic pathogen capable of causing severe diseases in humans and pigs, including meningitis, sepsis, polyserositis, arthritis, and endocarditis. This study aimed to investigate the biological characteristics of 19 strains of S. suis isolated from diseased pigs in Hubei Province between 2021 and 2023. Through bioinformatics analysis, we investigated the serotype, MLST, pan-genome characteristics, SNP, AMR, and ICE of the 19 S. suis isolates. Among the 19 S. suis strains, ten serotypes were identified, and serotype 9 was the most prevalent (21.05%). Ten new alleles and nine new sequence types (STs) were discovered, with ST28 and ST243 emerging as the predominant STs. The results of the pan-genomic analysis of S. suis indicate that there are 943 core genes, 2259 shell genes, and 5663 cloud genes. Through SNP evolutionary analysis, we identified a strong genetic similarity between SS31 and the reference genome P1/7. The analysis of antibiotic resistance genes revealed widespread presence of erm(B) and tet(O) genes among 19 strains of S. suis. This association may be linked to the high resistance of S. suis to lincosamides, macrolides, and tetracyclines. Integrative and conjugative elements (ICEs) and integrative and mobilizable elements (IMEs) were identified in 16 strains, with a carriage rate of 84.21%, and resistance genes were identified within the ICE/IME elements of 8 strains. Antimicrobial susceptibility testing revealed that all strains showed sensitivity to vancomycin and lincomycin but resistance to tilmicosin, tiamulin, amoxicillin, and doxycycline. This study contributes to our understanding of the genomic diversity of S. suis in Hubei Province of China, providing essential data for the comprehensive prevention and control of S. suis infections in China. Full article

In contrast to “frank” pathogens, like Salmonella entrocolitica, Shigella dysenteriae, and Vibrio cholerae, that always have a probability of disease, “opportunistic” pathogens are organisms that cause an infectious disease in a host with a weakened immune system and rarely in a healthy host. Historically, drinking water treatment has focused on control of frank pathogens, particularly those from human or animal sources (like Giardia lamblia, Cryptosporidium parvum, or Hepatitis A virus), but in recent years outbreaks from drinking water have increasingly been due to opportunistic pathogens. Characteristics of opportunistic pathogens that make them problematic for water treatment include: (1) they are normally present in aquatic environments, (2) they grow in biofilms that protect the bacteria from disinfectants, and (3) under appropriate conditions in drinking water systems (e.g., warm water, stagnation, low disinfectant levels, etc.), these bacteria can amplify to levels that can pose a public health risk. The three most common opportunistic pathogens in drinking water systems are Legionella pneumophila, Mycobacterium avium, and Pseudomonas aeruginosa. This report focuses on these organisms to provide information on their public health risk, occurrence in drinking water systems, susceptibility to various disinfectants, and other operational practices (like flushing and cleaning of pipes and storage tanks). In addition, information is provided on a group of nine other opportunistic pathogens that are less commonly found in drinking water systems, including Aeromonas hydrophila, Klebsiella pneumoniae, Serratia marcescens, Burkholderia pseudomallei, Acinetobacter baumannii, Stenotrophomonas maltophilia, Arcobacter butzleri, and several free-living amoebae including Naegleria fowleri and species of Acanthamoeba. The public health risk for these microbes in drinking water is still unclear, but in most cases, efforts to manage Legionella, mycobacteria, and Pseudomonas risks will also be effective for these other opportunistic pathogens. The approach to managing opportunistic pathogens in drinking water supplies focuses on controlling the growth of these organisms. Many of these microbes are normal inhabitants in biofilms in water, so the attention is less on eliminating these organisms from entering the system and more on managing their occurrence and concentrations in the pipe network. With anticipated warming trends associated with climate change, the factors that drive the growth of opportunistic pathogens in drinking water systems will likely increase. It is important, therefore, to evaluate treatment barriers and management activities for control of opportunistic pathogen risks. Controls for primary treatment, particularly for turbidity management and disinfection, should be reviewed to ensure adequacy for opportunistic pathogen control. However, the major focus for the utility’s opportunistic pathogen risk reduction plan is the management of biological activity and biofilms in the distribution system. Factors that influence the growth of microbes (primarily in biofilms) in the distribution system include, temperature, disinfectant type and concentration, nutrient levels (measured as AOC or BDOC), stagnation, flushing of pipes and cleaning of storage tank sediments, and corrosion control. Pressure management and distribution system integrity are also important to the microbial quality of water but are related more to the intrusion of contaminants into the distribution system rather than directly related to microbial growth. Summarizing the identified risk from drinking water, the availability and quality of disinfection data for treatment, and guidelines or standards for control showed that adequate information is best available for management of L. pneumophila. For L. pneumophila, the risk for this organism has been clearly established from drinking water, cases have increased worldwide, and it is one of the most identified causes of drinking water outbreaks. Water management best practices (e.g., maintenance of a disinfectant residual throughout the distribution system, flushing and cleaning of sediments in pipelines and storage tanks, among others) have been shown to be effective for control of L. pneumophila in water supplies. In addition, there are well documented management guidelines available for the control of the organism in drinking water distribution systems. By comparison, management of risks for Mycobacteria from water are less clear than for L. pneumophila. Treatment of M. avium is difficult due to its resistance to disinfection, the tendency to form clumps, and attachment to surfaces in biofilms. Additionally, there are no guidelines for management of M. avium in drinking water, and one risk assessment study suggested a low risk of infection. The role of tap water in the transmission of the other opportunistic pathogens is less clear and, in many cases, actions to manage L. pneumophila (e.g., maintenance of a disinfectant residual, flushing, cleaning of storage tanks, etc.) will also be beneficial in helping to manage these organisms as well. Full article

This research explores the association between ABO blood groups and susceptibility to SARS-CoV-2 infection, analyzing Convalescent COVID-19 plasma (CCP) donors (n = 500) and healthy whole blood donors (BDs) (n = 9678) during the pandemic (1 May 2020 to 30 April 2021). A comparison is made with pre-pandemic BDs (n = 11,892) from 1 May 2018 to 30 April 2019. Significant differences in blood group distribution are observed, with blood group A individuals being three times more likely to be CCP donors. Conversely, blood groups B, O, and AB are less associated with CCP donation. Notably, blood group O is more prevalent among regular BDs, suggesting potential resistance to SARS-CoV-2 infection. This study underscores variations in blood group distribution during the pandemic compared to pre-pandemic periods. The findings support previous research indicating a link between blood group antigens and viral susceptibility, including SARS-CoV-2. Understanding these associations has implications for public health strategies, with potential for predicting COVID-19 outcomes and transmission patterns. Further research is crucial to explore molecular and immunological mechanisms, providing valuable insights for targeted preventive strategies and personalized healthcare in managing the impact of COVID-19. Full article

The search for natural food additives makes propolis an exciting alternative due to its known antimicrobial activity. This work aims to investigate propolis’ behavior as a nitrite substitute ingredient in cooked ham (a ready-to-eat product) when confronted with pathogenic microorganisms of food interest. The microbial evolution of Listeria monocytogenes, Staphylococcus aureus, Bacillus cereus, and Clostridium sporogenes inoculated at known doses was examined in different batches of cooked ham. The design of a challenge test according to their shelf life (45 days), pH values, and water activity allowed the determination of the mesophilic aerobic flora, psychotropic, and acid lactic bacteria viability. The test was completed with an organoleptic analysis of the samples, considering possible alterations in color and texture. The cooked ham formulation containing propolis instead of nitrites limited the potential growth (δ < 0.5 log₁₀) of all the inoculated microorganisms until day 45, except for L. monocytogenes, which in turn exhibited a bacteriostatic effect between day 7 and 30 of the storage time. The sensory analysis revealed the consumer’s acceptance of cooked ham batches including propolis as a natural additive. These findings suggest the functionality of propolis as a promising alternative to artificial preservatives for ensuring food safety and reducing the proliferation risk of foodborne pathogens in ready-to-eat products. Full article

Pathogenic microorganisms released onto the soil from point or diffuse sources represent a public health concern. They can be transported by rainwater that infiltrates into subsoil and reach the groundwater where they can survive for a long time and contaminate drinking water sources. As part of the SCA.Re.S. (Evaluation of Health Risk Related to the Discharge of Wastewater on the Soil) project, we reviewed a selection of field-scale studies that investigated the factors that influenced the fate of microorganisms that were transported from the ground surface to the groundwater. A total of 24 studies published between 2003 and 2022 were included in the review. These studies were selected from the PubMed and Web of Science databases. Microbial contamination of groundwater depends on complex interactions between human activities responsible for the release of contaminants onto the soil, and a range of environmental and biological factors, including the geological, hydraulic, and moisture characteristics of the media traversed by the water, and the characteristics and the viability of the microorganisms, which in turn depend on the environmental conditions and presence of predatory species. Enterococci appeared to be more resistant in the underground environment than thermotolerant coliforms and were suggested as a better indicator for detecting microbial contamination of groundwater. Full article

Rickettsial pathogens are among the emerging and re-emerging vector-borne zoonoses of public health importance. Reports indicate human exposure to Rickettsial pathogens in Namibia through serological surveys, but there is a lack of data on infection rates in tick vectors, hindering the assessment of the relative risk to humans. Our study sought to screen Ixodid ticks collected from livestock for the presence of Rickettsia species in order to determine infection rates in ticks and to determine the Rickettsia species circulating in the country. We collected and pooled Hyalomma and Rhipicephalus ticks from two adjacent regions of Namibia (Khomas and Otjozondjupa) and observed an overall minimum Rickettsia infection rate of 8.6% (26/304), with an estimated overall pooled prevalence of 9.94% (95% CI: 6.5–14.3). There were no statistically significant differences in the estimated pooled prevalence between the two regions or tick genera. Based on the nucleotide sequence similarity and phylogenetic analysis of the outer membrane protein A (n = 9) and citrate synthase (n = 12) genes, BLAST analysis revealed similarity between Rickettsia africae (n = 2) and Rickettsia aeschlimannii (n = 11), with sequence identities ranging from 98.46 to 100%. Our initial study in Namibia indicates that both zoonotic R. africae and R. aeschlimannii are in circulation in the country, with R. aeschlimannii being the predominant species. Full article

Microbial communities inhabiting sedimentary environments in river source regions serve as pivotal indicators of pristine river ecosystems. While the correlation between antibiotic resistome and pathogenicity with core gut bacteria in humans is well established, there exists a significant knowledge gap concerning the interaction of antibiotic resistance genes (ARGs) and human pathogenic bacteria (HPB) with specific microbes in river source basins, often referred to as “terrestrial gut”. Understanding the microbial composition, including bacteria and resident genetic elements such as ARGs, HPB, Mobile Genetic Elements (MGEs), and Virulence Factors (VFs), within natural habitats against the backdrop of global change, is imperative. To address this gap, an enrichment-based culturomics complementary along with metagenomics was conducted in this study to characterize the microbial biobank and provide preliminary ecological insights into profiling the dissemination of ARGs in the Lancang River Source Basin. Based on our findings, in the main stream of the Lancang River Source Basin, 674 strains of bacteria, comprising 540 strains under anaerobic conditions and 124 under aerobic conditions, were successfully isolated. Among these, 98 species were identified as known species, while 4 were potential novel species. Of these 98 species, 30 were HPB relevant to human health. Additionally, bacA and bacitracin emerged as the most abundant ARGs and antibiotics in this river, respectively. Furthermore, the risk assessment of ARGs predominantly indicated the lowest risk rank (Rank Ⅳ) in terms of endangering human health. In summary, enrichment-based culturomics proved effective in isolating rare and unknown bacteria, particularly under anaerobic conditions. The emergence of ARGs showed limited correlation with MGEs, indicating minimal threats to human health within the main stream of the Lancang River Source Basin. Full article

Background: Infective endocarditis (IE) is a pathological condition caused by various microbial agents that can lead to severe complications affecting the heart. Accurate diagnosis is crucial for the effective management of patients with IE. Blood culture is the gold standard for identifying the primary infectious agents, which is a key factor in diagnosing IE using the modified Duke criteria. Objective: The main objective of this study was to investigate the distribution of the etiological agents of IE and the most common secondary diagnoses associated with it. Method: A total of 152 patients aged 23–95 years with a diagnosis of IE and proven etiology (through blood cultures or serological tests) were included in this study. Results: The most common etiological agent identified through blood tests was Enterococcus faecalis, which was detected in 39 patients (23.5%). Staphylococcus aureus was the second most common agent and was identified in 33 patients (19.9%), followed by Staphylococcus epidermidis, which was identified in 12 patients (13.1%). Nine patients (5.8%) had high levels of anti-Coxiella burnetti IgG phase I and II antibodies. Conclusions: IE is a leading cause of death in the Department of Infectious Diseases. Early and accurate diagnosis, along with interdisciplinary treatment, can significantly increase the chances of patient survival. Currently, Enterococcus faecalis and Staphylococcus aureus are the dominant etiological agents of IE, highlighting the need to revise protocols for prophylaxis, diagnosis, and initial treatment of this condition. Full article

Three patients with relapsing and remitting borreliosis, babesiosis, and bartonellosis, despite extended anti-infective therapy, were prescribed double-dose dapsone combination therapy (DDDCT) for 8 weeks, followed by one or several two-week courses of pulsed high-dose dapsone combination therapy (HDDCT). We discuss these patients’ cases to illustrate three important variables required for long-term remission. First, diagnosing and treating active co-infections, including Babesia and Bartonella were important. Babesia required rotations of multiple anti-malarial drug combinations and herbal therapies, and Bartonella required one or several 6-day HDDCT pulses to achieve clinical remission. Second, all prior oral, intramuscular (IM), and/or intravenous (IV) antibiotics used for chronic Lyme disease (CLD)/post-treatment Lyme disease syndrome (PTLDS), irrespective of the length of administration, were inferior in efficacy to short-term pulsed biofilm/persister drug combination therapy i.e., dapsone, rifampin, methylene blue, and pyrazinamide, which improved resistant fatigue, pain, headaches, insomnia, and neuropsychiatric symptoms. Lastly, addressing multiple factors on the 16-point multiple systemic infectious disease syndrome (MSIDS) model was important in achieving remission. In conclusion, DDDCT with one or several 6–7-day pulses of HDDCT, while addressing abnormalities on the 16-point MSIDS map, could represent a novel effective clinical and anti-infective strategy in CLD/PTLDS and associated co-infections including Bartonella. Full article

In this work, we studied the selective pressure and evolutionary analysis on the SARS-CoV-2 BF.7 and BQ.1.1 lineages circulating in Italy from July to December 2022. Two different datasets were constructed: the first comprised 694 SARS-CoV-2 BF.7 lineage sequences and the second comprised 734 BQ.1.1 sequences, available in the Italian COVID-19 Genomic (I-Co-Gen) platform and GISAID (last access date 15 December 2022). Alignments were performed with MAFFT v.7 under the Galaxy platform. The HYPHY software was used to study the selective pressure. Four positively selected sites (two in nsp3 and two in the spike) were identified in the BF.7 dataset, and two (one in ORF8 and one in the spike gene) were identified in the BQ.1.1 dataset. Mutation analysis revealed that R408S and N440K are very common in the spike of the BF.7 genomes, as well as L452R among BQ.1.1. N1329D and Q180H in nsp3 were found, respectively, at low and rare frequencies in BF.7, while I121L and I121T were found to be rare in ORF8 for BQ.1.1. The positively selected sites may have been driven by the selection for increased viral fitness, under circumstances of defined selective pressure, as well by host genetic factors. Full article

Chagas disease is caused by the single-flagellated protozoan Trypanosoma cruzi, which affects several million people worldwide. Understanding the signal transduction pathways involved in this parasite’s growth, adaptation, and differentiation is crucial. Understanding the basic mechanisms of signal transduction in T. cruzi could help to develop new drugs to treat the disease caused by these protozoa. In the present work, we have demonstrated that Fetal Calf Serum (FCS) can quickly increase the levels of both phosphorylated and unphosphorylated forms of T. cruzi DNA polymerase beta (TcPolβ) in tissue-cultured trypomastigotes. The in vitro phosphorylation sites on TcPolβ by protein kinases TcCK1, TcCK2, TcAUK1, and TcPKC1 have been identified by Mass Spectrometry (MS) analysis and with antibodies against phosphor Ser-Thr-Tyr. MS analysis indicated that these protein kinases can phosphorylate Ser and Thr residues on several sites on TcPolβ. Unexpectedly, it was found that TcCK1 and TcPKC1 can phosphorylate a different Tyr residue on TcPolβ. By using a specific anti-phosphor Tyr monoclonal antibody, it was determined that TcCK1 can be in vitro autophosphorylated on Tyr residues. In vitro and in vivo studies showed that phorbol 12-myristate 13-acetate (PMA) can activate the PKC to stimulate the TcPolβ phosphorylation and enzymatic activity in T. cruzi epimastigotes. Full article

Diagnosing canine visceral leishmaniasis (CVL) in Brazil faces challenges due to the limitations regarding the sensitivity and specificity of the current diagnostic protocol. Therefore, it is urgent to map new antigens or enhance the existing ones for future diagnostic techniques. Immunoinformatic tools are promising in the identification of new potential epitopes or antigen candidates. In this study, we evaluated peptides selected by epitope prediction for CVL serodiagnosis in ELISA assays. Ten B-cell epitopes were immunogenic in silico, but two peptides (peptides No. 45 and No. 48) showed the best performance in vitro. The selected peptides, both individually and in combination, were highly diagnostically accurate, with sensitivities ranging from 86.4% to 100% and with a specificity of approximately 90%. We observed that the combination of peptides showed better performance when compared to peptide alone, by detecting all asymptomatic dogs, showing lower cross-reactivity in sera from dogs with other canine infections, and did not detect vaccinated animals. Moreover, our data indicate the potential use of immunoinformatic tools associated with ELISA assays for the selection and evaluation of potential new targets, such as peptides, applied to the diagnosis of CVL. Full article

The vast array of interconnected microorganisms across Earth’s ecosystems and within holobionts has been called the “Internet of Microbes.” Bacteria and archaea are masters of energy and information collection, storage, transformation, and dissemination using both “wired” and wireless (at a distance) functions. Specific tools affecting microbial energy and information functions offer effective strategies for managing microbial populations within, between, and beyond holobionts. This narrative review focuses on microbial management using a subset of physical modifiers of microbes: sound and light (as well as related vibrations). These are examined as follows: (1) as tools for managing microbial populations, (2) as tools to support new technologies, (3) as tools for healing humans and other holobionts, and (4) as potential safety dangers for microbial populations and their holobionts. Given microbial sensitivity to sound, light, and vibrations, it is critical that we assign a higher priority to the effects of these physical factors on microbial populations and microbe-laden holobionts. We conclude that specific sound, light, and/or vibrational conditions are significant therapeutic tools that can help support useful microbial populations and help to address the ongoing challenges of holobiont disease. We also caution that inappropriate sound, light, and/or vibration exposure can represent significant hazards that require greater recognition. Full article

Vibrio species are naturally found in estuarine and marine ecosystems, but are also recognized as significant human enteropathogens, often linked to seafood-related illnesses. In aquaculture settings, Vibrio poses a substantial risk of infectious diseases, resulting in considerable stock losses and prompting the use of antimicrobials. However, this practice contributes to the proliferation of antimicrobial-resistant (AMR) bacteria and resistance genes. Our investigation aimed to explore the potential of biological agents such as bacteriophage CH20 and endolysin LysVPp1 in reducing Vibrio bacterial loads in both rotifer and fish larvae. LysVPp1’s lytic activity was assessed by measuring absorbance reduction against various pathogenic Vibrio strains. Phage CH20 exhibited a limited host range, affecting only Vibrio alginolyticus GV09, a highly pathogenic strain. Both CH20 and LysVPp1 were evaluated for their effectiveness in reducing Vibrio load in rotifers or fish larvae through short-setting bioassays. Our results demonstrated the significant lytic effect of endolysin LysVPp1 on strains of Vibrio alginolyticus, Vibrio parahaemolyticus, and Vibrio splendidus. Furthermore, we have showcased the feasibility of reducing the load of pathogenic Vibrio in live feed and fish larvae by using a non-antibiotic-based approach, such as lytic phage and endolysin LysVPp1, thus contributing to the progress of a sustainable aquaculture from a One Health perspective. Full article