Levels Of Virulence Research Articles

Mechanisms and Implications of Phenotypic Switching in Bacterial Pathogens.

Bacteria encounter various stressful conditions within a variety of dynamic environments, which they must overcome for survival. One way they achieve this is by developing phenotypic heterogeneity to introduce diversity within their population. Such distinct subpopulations can arise through endogenous fluctuations in regulatory components, wherein bacteria can express diverse phenotypes and switch between them, sometimes in a heritable and reversible manner. This switching may also lead to antigenic variation, enabling pathogenic bacteria to evade the host immune response. Therefore, phenotypic heterogeneity plays a significant role in microbial pathogenesis, immune evasion, antibiotic resistance, host niche tissue establishment and environmental persistence. This heterogeneity can result from stochastic and responsive switches, as well as various genetic and epigenetic mechanisms. The development of phenotypic heterogeneity may create clonal populations that differ in their level of virulence, contribute to the formation of biofilms, and allow for antibiotic persistence within select morphological variants. This review delves into the current understanding of the molecular switching mechanisms underlying phenotypic heterogeneity, highlighting their roles in establishing infections caused by select bacterial pathogens.

Colletotrichum species associated with anthracnose disease on mango (Mangifera indica L.) cv. Azúcar in Colombia

The role of some Colletotrichum species is uncertain in crops of economic importance for Colombia such as mango (Mangifera indica L.) cv. Azúcar. Eight Colletotrichum isolates from asymptomatic plant tissues were characterized by their pathogenicity on fruits and identified by phylogenetic analysis of the sequences of the ITS, GADPH, Tub2, CHs-1 and Act genomic regions. Colletotrichum asianum, C. gloeosporioides and C. tropicale of the C. gloeosporioides species complex and C. karstii of the C. boninense species complex were identified. The highest virulence was observed for isolates of C. asianum, that was the most prevalent species, and for C. tropicale, that showed high and intermediate virulence levels, indicating diversity in pathogenicity within the same species. The isolate of C. karstii, which displayed intermediate virulence, is the first report on mango in Colombia. Colletotrichum gloeosporioides presented intermediate virulence and was isolated at a low frequency in the present research. The mycelial growth rate did not show differences between species and ranged between 6.1 and 8.3 mm per day. Colletotrichum asianum recorded higher values of conidia germination and appressoria formation than other species, at 77% and 79%, respectively (p < 0.001). The shape and size of conidia did not vary between species. Appressoria exhibited different shapes between isolates and the largest average diameter was for C. karstii with 7.6 µm (p < 0.001). There was a diversity of Colletotrichum species associated with mango cv. Azúcar in Colombia, with differential virulence among isolates of the same species.

Rapid phenotypic and genotypic change in a laboratory schistosome population.

Genomic analysis has revealed extensive contamination among laboratory-maintained microbes including malaria parasites, Mycobacterium tuberculosis and Salmonella spp. Here, we provide direct evidence for recent contamination of a laboratory schistosome parasite population, and we investigate its genomic consequences. The Brazilian Schistosoma mansoni population SmBRE has several distinctive phenotypes, showing poor infectivity, reduced sporocysts number, low levels of cercarial shedding and low virulence in the intermediate snail host, and low worm burden and low fecundity in the vertebrate rodent host. In 2021 we observed a rapid change in SmBRE parasite phenotypes, with a~10x increase in cercarial production and ~ 4x increase in worm burden. To determine the underlying genomic cause of these changes, we sequenced pools of SmBRE adults collected during parasite maintenance between 2015 and 2023. We also sequenced another parasite population (SmLE) maintained alongside SmBRE without phenotypic changes. While SmLE allele frequencies remained stable over the eight-year period, we observed sudden changes in allele frequency across the genome in SmBRE between July 2021 and February 2023, consistent with expectations of laboratory contamination. (i) SmLE-specific alleles rose in the SmBRE population from 0 to 41-46% across the genome between September and October 2021, documenting the timing and magnitude of the contamination event. (ii) After contamination, strong selection (s=~ 0.23) drove replacement of low fitness SmBRE with high fitness SmLE alleles. (iii) Allele frequency changed rapidly across the whole genome, except for a region on chromosome 4 where SmBRE alleles remained at high frequency. We were able to detect contamination in this case because SmBRE shows distinctive phenotypes. However, this would likely have been missed with phenotypically similar parasites. These results provide a cautionary tale about the importance of tracking the identity of parasite populations, but also showcase a simple approach to monitor changes within populations using molecular profiling of pooled population samples to characterize fixed single nucleotide polymorphisms. We also show that genetic drift results in continuous change even in the absence of contamination, causing parasites maintained in different labs (or sampled from the same lab at different times) to diverge.

Adaptations to cationic biocide exposure differentially influence virulence factors and pathogenicity in Pseudomonas aeruginosa

ABSTRACT Cationic biocides (CBs), which include quaternary ammonium compounds (QACs), are employed to mitigate the spread of infectious bacteria, but resistance to such surface disinfectants is rising. CB exposure can have profound phenotypic implications that extend beyond allowing microorganisms to persist on surfaces. Pseudomonas aeruginosa is a deadly bacterial pathogen that is intrinsically tolerant to a wide variety of antimicrobials and is commonly spread in healthcare settings. In this study, we pursued resistance selection assays to the QAC benzalkonium chloride and quaternary phosphonium compound P6P–10,10 to assess the phenotypic effects of CB exposure in P. aeruginosa PAO1 and four genetically diverse, drug-resistant clinical isolates. In particular, we sought to examine how CB exposure affects defensive strategies and the virulence-associated “offensive” strategies in P. aeruginosa. We demonstrated that development of resistance to BAC is associated with increased production of virulence-associated pigments and alginate as well as pellicle formation. In an in vivo infection model, CB-resistant PAO1 exhibited a decreased level of virulence compared to wild type, potentially due to an observed fitness cost in these strains. Taken together, these results illustrate the significant consequence CB resistance exerts on the virulence-associated phenotypes of P. aeruginosa.

Comparative Genomic Analyses of Colletotrichum lindemuthianum Pathotypes with Different Virulence Levels and Lifestyles.

Colletotrichum lindemuthianum is the most frequent pathogenic fungus of the common bean Phaseolus vulgaris. This filamentous fungus employs a hemibiotrophic nutrition/infection strategy, which is characteristic of many Colletotrichum species. Due to host-pathogen coevolution, C. lindemuthianum includes pathotypes with a diversity of virulence against differential common bean varieties. In this study, we performed comparative genomic analyses on three pathotypes with different virulence levels and a non-pathogenic pathotype, isolated from different geographical areas in Mexico. Our results revealed large genomes with high transposable element contents that have undergone expansions, generating intraspecific diversity. All the pathotypes exhibited a similar number of clusters of orthologous genes (COGs) and Gene Ontology (GO) terms. TFomes contain families that are typical in fungal genomes; however, they show different contents between pathotypes, mainly in transcription factors with the fungal-specific TF and Zn2Cys6 domains. Peptidase families mainly contain abundant serine peptidases, metallopeptidases, and cysteine peptidases. In the secretomes, the number of genes differed between the pathotypes, with a high percentage of candidate effectors. Both the virulence gene and CAZyme gene content for each pathotype was abundant and diverse, and the latter was enriched in hemicellulolytic enzymes. We provide new insights into the nature of intraspecific diversity among C. lindemuthianum pathotypes and the origin of their ability to rapidly adapt to genetic changes in its host and environmental conditions.

Transcriptomic Analyses to Unravel Cronobacter sakazakii Resistance Pathways.

The proliferation of antibiotic usage has precipitated the emergence of drug-resistant variants of bacteria, thereby augmenting their capacity to withstand pharmaceutical interventions. Among these variants, Cronobacter sakazakii (C. sakazakii), prevalent in powdered infant formula (PIF), poses a grave threat to the well-being of infants. Presently, global contamination by C. sakazakii is being observed. Consequently, research endeavors have been initiated to explore the strain's drug resistance capabilities, alterations in virulence levels, and resistance mechanisms. The primary objective of this study is to investigate the resistance mechanisms and virulence levels of C. sakazakii induced by five distinct antibiotics, while concurrently conducting transcriptomic analyses. Compared to the susceptible strains prior to induction, the drug-resistant strains exhibited differential gene expression, resulting in modifications in the activity of relevant enzymes and biofilm secretion. Transcriptomic studies have shown that the expression of glutathione S-transferase and other genes were significantly upregulated after induction, leading to a notable enhancement in biofilm formation ability, alongside the existence of antibiotic resistance mechanisms associated with efflux pumps, cationic antimicrobial peptides, and biofilm formation pathways. These alterations significantly influence the strain's resistance profile.

Replication Kinetics and Infectivity of African Swine Fever Virus (ASFV) Variants with Different Genotypes or Levels of Virulence in Cell Culture Models of Primary Porcine Macrophages

Replication Kinetics and Infectivity of African Swine Fever Virus (ASFV) Variants with Different Genotypes or Levels of Virulence in Cell Culture Models of Primary Porcine Macrophages

Genomic Interactions Between Mycobacterium tuberculosis and Humans.

Mycobacterium tuberculosis is considered by many to be the deadliest microbe, with the estimated annual cases numbering more than 10 million. The bacteria, including Mycobacterium africanum, are classified into nine major lineages and hundreds of sublineages, each with different geographical distributions and levels of virulence. The phylogeographic patterns can be a result of recent and early human migrations as well as coevolution between the bacteria and various human populations, which may explain why many studies on human genetic factors contributing to tuberculosis have not been replicable in different areas. Moreover, several studies have revealed the significance of interactions between human genetic variations and bacterial genotypes in determining the development of tuberculosis, suggesting coadaptation. The increased availability of whole-genome sequence data from both humans and bacteria has enabled a better understanding of these interactions, which can inform the development of vaccines and other control measures.

Host Range Characterization of Phytophthora sansomeana Across Corn, Soybean, Wheat, Winter Cereal Rye, Dry Bean, and Oats and an In Vitro Assessment of Seed Treatment Sensitivity.

Formally described in 2009, Phytophthora sansomeana is a pathogen of increasing interest in native, agricultural, and horticulturally important plant species. The objective of this study was to elucidate the symptomatic and asymptomatic host range of P. sansomeana on six agricultural crop species commonly used in field crop rotations in Michigan. In addition, sensitivity to oomicides commonly used in seed treatments, including oxathiapiprolin, mefenoxam, ethaboxam, and pyraclostrobin, was performed to aid in disease management recommendations. Plant biomass, quantity of P. sansomeana DNA in roots, and reisolations were used to assess pathogenicity and virulence of 18 isolates of P. sansomeana on each plant species using an inoculated seedling growth chamber assay. Isolates displayed varying levels of virulence to the hosts tested. Reisolations were completed for each plant species tested, and varying quantities of P. sansomeana DNA were found within all plant species root samples. Corn, wheat, soybean, dry bean, and winter cereal rye plants were symptomatic hosts with significant reduction observed in the total plant biomass. No significant reduction in total plant biomass was observed in oats, and oat roots harbored the least amount of P. sansomeana DNA. No P. sansomeana isolates were insensitive to the oomicide compounds tested with mean absolute inhibition (EC50) values of fungicide required for 50% growth inhibition values of 7.8 × 10-2 μg/ml for mefenoxam, 1.13 × 10-1 μg/ml for ethaboxam, 2.6 × 10-2 μg/ml for oxathiapiprolin, and 3.04 × 10-1 μg/ml for pyraclostrobin. These results suggest that common crop rotations in Michigan may not be a viable option to reduce soilborne inoculum accumulation and oomicide seed treatments could be considered for early-season management of P. sansomeana.

Intra-Phenotypic and -Genotypic Variations of Beauveria bassiana (Bals.) Vuill. Strains Infecting Aedes aegypti L. Adults.

Beauveria bassiana has potential for Aedes aegypti biological control. However, its efficacy depends on the strain's geographic location, host susceptibility, and virulence. The present study aimed to evaluate the effectiveness of B. bassiana strain BBPTG4 conidia in controlling Ae. aegypti adults and its detection via introns profile on exposed mosquito corpses. Morphologic characteristics among strains were highly similar. Comprehensive testing of these strains demonstrated that BBPT4 exhibited the ideal biological activity for Ae. aegypti control, with a median lethal time (TL50) of 7.5 d compared to ~3 d and ~10 d for BB01 and BB37 strains, respectively. Infected mosquitoes died after GHA and BBPTG4 exposure, and corpses were analyzed for infecting strains detection. Differences among the seven evaluated strains were determined, assessing five different insertion group I intron profiles in BBTG4, BB01, GHA, BB37, and BB02 strains. Mosquitoes infected by BBPTG4 and non-exposed (negative control) intron profiles were obtained. We detected the presence of introns in the BBPTG4 strain, which were not present in non-exposed mosquitoes. In conclusion, B. bassiana strains showed similarities in terms of their cultural and microscopic morphological characteristics and biologicals virulence level, but different intron profiles. BBPTG4 strain-infected Ae. aegypti adult corpses, showing specific amplicons, enabled us to identify B. bassiana at the strain level among infected mosquitoes. However, monitoring and detection of field-infected insects is essential for further verification.

Rapid phenotypic and genotypic change in a laboratory schistosome population.

Genomic analysis has revealed extensive contamination among laboratory-maintained microbes including malaria parasites, Mycobacterium tuberculosis and Salmonella spp. Here, we provide direct evidence for recent contamination of a laboratory schistosome parasite population, and we investigate its genomic consequences. The Brazilian Schistosoma mansoni population SmBRE has several distinctive phenotypes, showing poor infectivity, reduced sporocysts number, low levels of cercarial shedding and low virulence in the intermediate snail host, and low worm burden and low fecundity in the vertebrate rodent host. In 2021 we observed a rapid change in SmBRE parasite phenotypes, with a ~10x increase in cercarial production and ~4x increase in worm burden. To determine the underlying genomic cause of these changes, we sequenced pools of SmBRE adults collected during parasite maintenance between 2015 and 2023. We also sequenced another parasite population (SmLE) maintained alongside SmBRE without phenotypic changes. While SmLE allele frequencies remained stable over the eight-year period, we observed sudden changes in allele frequency across the genome in SmBRE between July 2021 and February 2023, consistent with expectations of laboratory contamination. (i) SmLE-specific alleles rose in the SmBRE population from 0 to 41-46% across the genome between September and October 2021, documenting the timing and magnitude of the contamination event. (ii) After contamination, strong selection (s = ~0.23) drove replacement of low fitness SmBRE with high fitness SmLE alleles. (iii) Allele frequency changed rapidly across the whole genome, except for a region on chromosome 4 where SmBRE alleles remained at high frequency. We were able to detect contamination in this case because SmBRE shows distinctive phenotypes. However, this would likely have been missed with phenotypically similar parasites. These results provide a cautionary tale about the importance of tracking the identity of parasite populations, but also showcase a simple approach to monitor changes within populations using molecular profiling of pooled population samples to characterize fixed single nucleotide polymorphisms. We also show that genetic drift results in continuous change even in the absence of contamination, causing parasites maintained in different labs (or sampled from the same lab at different times) to diverge.

Glucanases and Chitinases in Mangifera indica: Identification, Classification, Phylogeny, and Expression Analysis of Defense Genes against Colletotrichum spp.

Plant glucanases and chitinases are defense proteins that participate in pathogenesis; however, very little is known about the glucanase (GLUC) and chitinase (CHIT) gene families in mango. Some mango cultivars are of great economic importance and can be affected by anthracnose, a postharvest disease caused by fungi of the genus Colletotrichum spp. This study identified and characterized 23 putative glucanases and 16 chitinases in the mango genome cv. Tommy Atkins. We used phylogenetic analyses to classify the glucanases into three subclasses (A, B, and C) and the chitinases into four classes (I, II, IV, and V). Information on the salicylic, jasmonic acid, and ethylene pathways was obtained by analyzing the cis-elements of the GLUC and CHIT class I and IV gene promoters. The expression profile of GLUC, CHIT class I, and CHIT class IV genes in mango cv. Ataulfo inoculated with two Colletotrichum spp. revealed different profile expression related to these fungi's level of virulence. In general, this study provides the basis for the functional validation of these target genes with which the regulatory mechanisms used by glucanases and chitinases as defense proteins in mango can be elucidated.

Clubroot-Induced Changes in the Root and Rhizosphere Microbiome of Susceptible and Resistant Canola.

Clubroot is a soilborne disease of canola (Brassica napus) and other crucifers caused by the obligate parasite Plasmodiophora brassicae. In western Canada, clubroot is usually managed by planting-resistant cultivars, but the emergence of resistance-breaking pathotypes of P. brassicae represents a major threat to sustainable canola production. The rhizosphere and root contain beneficial microorganisms that can improve plant health. In this study, we evaluated the effect of two P. brassicae isolates (termed A and B) with different levels of virulence on the root and rhizosphere microbiomes of clubroot-resistant and clubroot-susceptible canola. Additionally, potential biocontrol microorganisms were identified based on taxa antagonistic to clubroot. Although both P. brassicae isolates were classified as pathotype 3A, isolate A caused a higher disease severity index in the resistant canola genotype compared with isolate B. Metabarcoding analysis indicated a shift in the bacterial and fungal communities in response to inoculation with either field isolate. Root endophytic bacterial and fungal communities responded to changes in inoculation, isolate type, sampling time, and canola genotype. In contrast, fungal communities associated with the rhizosphere exhibited significant differences between sampling times, while bacterial communities associated with the rhizosphere exhibited low variability.

EⅡB Mutation Reduces the Pathogenicity of Listeria monocytogenes by Negatively Regulating Biofilm Formation Ability, Infective Capacity, and Virulence Gene Expression.

To explore the role of the membrane permease ⅡB (EⅡB) gene of Listeria pathogenicity island 4 (LIPI-4) in the virulence of Listeria monocytogenes, both an EⅡB deletion strain (∆EⅡB) and a complemented strain were constructed. In vitro experiments demonstrated that EⅡB deletion affected the biofilm formation ability of the wild-type strain (Lm928). Moreover, this deletion decreased the intracellular proliferation abilities of L. monocytogenes. Mice infected with ∆EⅡB survived longer and experienced less weight loss on days 1, 2, and 3 post-infection. The bacterial load in the liver tissue of ∆EⅡB-infected mice was significantly reduced, and a considerable decrease in the blood levels of inflammatory cytokines IL-β, IL-6, IL-10, and TNF-α were observed. Following EⅡB deletion, 65% (13/20) of genes were downregulated, 25% (5/20) were upregulated, and 10% (2/20) showed no change. These findings suggest that EⅡB deletion may reduce both the in vivo and in vitro virulence levels as well as the biofilm formation ability of Lm928 by downregulating the transcription levels of genes associated with virulence and biofilm formation. These findings provide a foundation for further examining the pathogenic mechanisms of LIPI-4 and EⅡB in L. monocytogenes.

Aktivitas Enzimatik Cendawan dari Lalat Bactrocera dorsalis sebagai Kandidat Cendawan Entomopatogen

Extracellular enzymes produced by fungi can serve as a screening tool for entomopathogen candidates. Enzyme secretion ability is important in determining the virulence level of an entomopathogenic fungus. This study aims to see the potential of fungal isolates through chitinolytic, lipolytic, and proteolytic activities of five entomopathogenic fungal isolated from Bactrocera sp. imago from citrus plants. The research was carried out at the Microbiology Laboratory of the Department of Biology at the University of Lampung. Enzymatic activity is known through the method of observing the clear zone formed on agar media. Quantitative tests based on the calculation of enzymatic index values showed that Nigrospora sp. (IB1) had lipolytic activity, Penicillium sp. 2 (IB5) isolates had chitinolytic, lipolytic, and proteolytic activities. The isolates of Penicillium sp. 1 (IB2), Phytopthora sp. (IB3), and Gliocladium sp. (IB4) did not show the activity of the three enzymes. Keywords: Bactrocera sp., bioinsecticide, entomopathogenic fungi, enzyme

Chitosan: Biocontrol agent of Fusarium oxysporum in tomato fruit (Solanum lycopersicum L.)

Synthetic fungicides have experienced a significant increase in recent years, necessitating the search for more sustainable and environmentally friendly alternatives. In this regard, chitosan has emerged as an option to reduce reliance on these products. This study evaluated the effect of chitosan as a biocontrol agent against Fusarium oxysporum in tomato fruits. A fully randomized experimental design incorporating 6 treatments was employed, consisting of four chitosan treatments (0.5, 1, 2, and 3 g L-1), a negative control involving the application of a synthetic fungicide, and a positive control inoculated with F. oxysporum. Samples were taken from infected tomato fruits. The F4 isolate of Fusarium sp. was identified as F. oxysporum, and demonstrated the highest level of virulence. Among the four chitosan treatments, the 3 g L-1 treatment showed the highest a percentage of mycelial growth inhibition (PMGI) at 79.92% and the greatest reduction in biomass at 0.65 g, which did not differ significantly from the synthetic fungicide. Regarding disease severity and incidence, there were significant variations among each of the chitosan treatments, with the highest results obtained with the 2 and 3 g L-1 treatments. All chitosan treatments reduced disease severity in tomato fruits. Applying chitosan on fruits of the tomato plant presents an alternative for diminishing reliance on synthetic fungicides.

Differential Carbon Catabolite Repression and Hemicellulolytic Ability among Pathotypes of Colletotrichum lindemuthianum against Natural Plant Substrates.

Colletotrichum lindemuthianum is a phytopathogenic fungus that causes anthracnose in common beans (Phaseolus vulgaris) and presents a great diversity of pathotypes with different levels of virulence against bean varieties worldwide. The purpose of this study was to establish whether pathotypic diversity is associated with differences in the mycelial growth and secretion of plant-cell-wall-degrading enzymes (PCWDEs). We evaluated growth, hemicellulase and cellulase activity, and PCWDE secretion in four pathotypes of C. lindemuthianum in cultures with glucose, bean hypocotyls and green beans of P. vulgaris, and water hyacinth (Eichhornia crassipes). The results showed differences in the mycelial growth, hemicellulolytic activity, and PCWDE secretion among the pathotypes. Glucose was not the preferred carbon source for the best mycelial growth in all pathotypes, each of which showed a unique PCWDE secretion profile, indicating different levels of carbon catabolite regulation (CCR). The pathotypes showed a high differential hemicellulolytic capacity to degrade host and water hyacinth tissues, suggesting CCR by pentoses and that there are differences in the absorption and metabolism of different monosaccharides and/or disaccharides. We propose that different levels of CCR could optimize growth in different host tissues and could allow for consortium behavior in interactions with bean crops.

Variation in symptoms and morphology of Fusarium spp. on shallot associated with basal plate rot disease in Brebes District, Central Java Province, Indonesia

Abstract. Marianah L, Nawangsih AA, Munif A, Giyanto, Tondok ET. 2024. Variation in symptoms and morphology of Fusarium spp. on shallot associated with basal plate rot disease in Brebes District, Central Java Province, Indonesia. Biodiversitas 25: 2198-2208. Basal plate rot disease is an important disease of shallots that causes losses in the field and storage. The disease is caused by Fusarium complex species, with different levels of virulence and host susceptibility. The six species of Fusarium spp. reported to cause basal plate rot disease of onion were F. oxysporum f.sp. cepae, F. proliferatum, F. redolens, F. solani, F. acutatum and F. tricinctum. Information about the variation in symptoms and morphology of fungi that cause basal plate rot disease on shallots, especially in Indonesia, is still very limited. Besides, the species that cause basal plate rot disease on shallots has not been molecularly confirmed using specific primers. The objective of this study was to isolate, identify, and study the variation in symptoms of the fungus associated with basal plate rot disease. The fungi were isolated from shallot plants symptomatic of basal plate rot disease collected from Brebes, Central Java Province. Eight isolates of Fusarium with different morphologies were successfully isolated and identified. Five isolates were identified as F. solani (isolate BC1, BC2, BC3, BBS1, and BBS4), two isolates as F. oxysporum f.sp. cepae (isolate BC4 and BBS6) and one isolate F. proliferatum (isolate BBS5). All eight isolates were pathogenic on shallots with different levels of virulence and symptom variations. BC4 isolate had the highest level of virulence, resulting in plant death, and was identified as F. oxysporum f.sp. cepae.

Isolation and Identification of Entomopathogenic Fungi from the Soils of Northwest Iran and their Biological Control Efficacy on the Flour Moth (Ephestia kuehniella)

Entomopathogenic fungi have the potential to serve as alternatives to or supplements for chemical pesticides in insect biocontrol, with many of these fungi being utilized in commercial and integrated pest control practices globally. This study aimed to isolate and evaluate the efficacy of various entomopathogenic fungi for the biocontrol of the flour moth, Ephestia kuehniella (Lep: Pyralidae). Multiple isolates of entomopathogenic fungi were acquired from soils across different districts of Azerbaijan province in northwest Iran, and their virulence against E. kuehniella larvae was assessed. Seventeen isolates representing diverse entomopathogenic fungi were isolated and purified using the soil trap method from 20 distinct soil samples. Their pathogenicity against E. kuehniella was evaluated in three replicates through the immersion technique at concentrations of 104, 105, 106, 107, and 108 conidia/mL. Identification of selected fungal isolates was based on morphological features, mycological parameters, and sequencing of the internal transcribed spacer (ITS) region. The study identified seven fungal species, namely Lecanicillium lecanii, Beauveria bassiana, Metarhizium anisopliae, Purpureocillium lilacinum, Aspergillus oryzae, Penicillium citrinum, and Aspergillus lavus, among the 17 isolates collected. Bioassay assessments revealed the susceptibility of 4th-instar larvae of E. kuehniella to the selected isolates, with the larval mortality rate increasing with higher conidial concentrations. A. lavus, A. oryzae, P. citrinum, and P. lilacinum exhibited maximum virulence, causing nearly 90% lethality, while Metarhizium anisopliae demonstrated the lowest level of virulence, resulting in approximately 30% lethality in E. kuehniella. The virulence pattern of isolates across different concentrations was notably distinct. The findings of this research demonstrate that entomopathogenic fungi isolated in this investigation could be effectively incorporated into future pest management programs.

Transfer of micro-organisms from dry surface biofilms and the influence of long survival under conditions of poor nutrition and moisture on the virulence of Staphylococcusaureus

Biofilms on dry hospital surfaces can enhance the persistence of microorganisms on dry harsh clinical surfaces and can potentially act as reservoirs of infectious agents on contaminated surfaces. This study was conducted to quantify the transfer of viable Staphylococcus aureus cells from dry biofilms through touching and to investigate the impact of nutrient and moisture deprivation on virulence levels in S. aureus. Dry biofilms of S. aureus ATCC 25923 and a defective biofilm-forming ability mutant, S. aureus 1132 were formed in 24 well plates under optimised conditions mimicking dry biofilm formation on clinical surfaces. Microbial cell transfer was induced through the touching of the dry biofilms, which were quantified on nutrient agar. To investigate the impact of nutrient and moisture deprivation on virulence levels, dry and standard biofilms as well as planktonic cells of S. aureus ATCC 25923 were inoculated into Galleria mellonella and their kill rates compared. Results of this study showed that viable cells from dry biofilms of S. aureus ATCC 25923 were significantly more virulent and readily transferrable from dry biofilms through a touch test, therefore representing a greater risk of infection. The biofilm-forming capability of S. aureus strains had no significant impact on their transferability with more cells transferring when biofilm surfaces were wet. These findings indicate that dry biofilms on hospital surfaces may serve as a reservoir for the dissemination of pathogenic microorganisms in hospitals, thus highlighting the importance of regular cleaning and adequate disinfection of hospital surfaces.