High Virulence Research Articles

Experimental infection of purebred Saanen goats high pathogenicity and virulence of Babesia aktasi.

Small ruminant babesiosis remains a neglected disease despite causing significant economic losses to sheep and goat herds in many regions around the world. The pathogenesis and clinical manifestations of ovine babesiosis are well-known, but there is a lack of information regarding caprine babesiosis. Since the discovery of the first Babesia spp. in 1888, several species/subspecies/genotypes, including Babesia aktasi, have been described. Our recent molecular survey revealed that the parasite is highly prevalent (22.5%) in indigenous goats from Mediterranean region of Türkiye. The aim of this experimental study was to determine the pathogenicity and virulence of B. aktasi in immunosuppressed (n = 5) and immunocompetent (n = 7) purebred Saanen goats. The goats were experimentally infected with fresh B. aktasi infected blood, and examined for clinical, parasitological, hematological, and serum biochemical findings throughout the infection. Following the parasite inoculation, intra-erythrocytic parasites were detected from the 1st day post-infection, followed by an increase in rectal temperature and parasitemia. The parasitemia was detected ranging from 4.3% to 33.5% in the immunosuppressed group, while it was 2.1% to 7.6% in the immunocompetent. Severe clinical symptoms characterized by anemia, jaundice, and hemoglobinuria developed in both groups. A statistically significant inverse correlation was observed between the increase in parasitemia and RBC, WBC, HCT, and Hb values in the goats compared to pre-infection levels. Values of AST, ALT, GGT, Total bilirubin, and Albumin showed a significant increase, with all the immunosuppressed goats dying on the 4th and 7th days post-infection, while four out of seven immunocompetent goats died on between 6-8th days. Severe edema in the lungs, frothy fluid in the trachea, jaundice in the subcutaneous and mesenteric fat, and dark red urine were detected in necropsy. The results obtained in this study demonstrated that B. aktasi was highly pathogenic to purebred Saanen goats. Current work assures valuable insights into the pathogenesis and virulence of B. aktasi and serves as a foundation for future studies to develop effective control strategies against caprine babesiosis.

Unraveling influenza sentinel surveillance in Pakistan 2008-2024: Epidemiological insights during the pre and post pandemic period of COVID-19.

The coronavirus pandemic has substantially influenced the transmission pattern of other respiratory viruses. However, screening and detecting other respiratory pathogens was unheeded during this time to combat the COVID-19 pandemic. High virulence and re assortment factors intensify the importance of influenza virus surveillance for effective disease management. Therefore, the present surveillance study was designed to determine the influenza positivity rate from 2008-24. This study will provide integral support in depicting a panoramic representation of two respiratory-pandemic periods, 2010-11 and 2019-2021, for influenza and COVID-19 pandemics, respectively. An inferential cross-sectional study was conducted from 2008 to 2024 by collecting influenza surveillance data from twelve sentinel sites in Pakistan. Clinical and demographic data was recorded at sample collection time. Specimens were collected through nasopharyngeal/throat swabs and stored in viral transport medium (VTM) at the sentinel site laboratory at 2-4°C. Viral RNA was isolated from the samples using KingFisher TM Flex Purification System and MagMAX™ Viral/Pathogen Nucleic Acid Extraction Kit. Within 16 years, 78118 samples were tested for influenza, of which 7999 (10.2%) appeared positive. The positivity rate appeared very low in recent years, with only a 3.5% positivity rate observed in 2020. Influenza A strain H1N1pdm09 seemed to be the prominent strain (n=3407, 42.6%), followed by influenza B (n=2125, 26.6%). The positivity of influenza samples was 10.2% and recorded in patients where typical clinical representation of influenza was absent. Fewer samples were reported during the coronavirus pandemic, which might be because influenza screening was hindered and overlooked to combat the SARS-CoV-2 virus, and the patient threshold was very high for COVID-19 virus screening.

Host-specific patterns of virulence and gene expression profiles of the broad-host-range entomopathogenic fungus Metarhizium anisopliae

Generalist pathogens with a broad host range encounter many different host environments. Such generalist pathogens are often highly versatile and adjust their expressed phenotype to the host being infected. Species in the fungal genus Metarhizium (Hypocreales: Clavicipitaceae) occupy various ecological niches, including plant rhizosphere symbionts, soil saprophytes, and insect pathogens with applications in biological control of pests. The species M. anisopliae is highly diverse combining the capability of association with plant roots and infection of a broad range of arachnid and insect hosts, from agricultural pests to vectors of human disease. It is among the most studied and applied biological control agents worldwide. Here, we investigate the phenotypic plasticity and differential gene expression of M. anisopliae blastospores during infection of different insect hosts. First, the virulence of M. anisopliae blastospores was evaluated against Tenebrio molitor (Coleoptera: Tenebrionidae), Spodoptera frugiperda (Lepidoptera: Noctuidae), Gryllus assimilis (Orthoptera: Gryllidae), and Apis mellifera (Hymenoptera: Apidae). Second, the percentage of appressorium formation on the membranous wings of the four hosts was determined, and third, the fungal transcriptome profile during penetration on the hosts was analyzed. Our findings reveal that M. anisopliae blastospores exhibit high virulence against Tenebrio molitor, with significantly higher appressorium formation on beetle wings compared to the other three tested insects. We also document distinct gene expression patterns in M. anisopliae blastospores during insect infection of T. molitor, S. frugiperda, and A. mellifera, with notable variations observed in G. assimilis. These differences are associated with the expression of enzymes involved in the degradation of specific compounds present in each insect wing, as well as hydrophobins, destruxins, and specialized metabolites related to virulence. The study emphasizes the differences in fungal gene expression during infection of the four insect orders and highlights the virulence-related genes specific to each infective process.

Comparative genomics and virulence potential of Campylobacter coli strains isolated from different sources over 25 years in Brazil

BackgroundCampylobacter spp. have been reported as a common cause of gastroenteritis in humans in many countries. However, in Brazil there is insufficient data to estimate the impact of Campylobacter in public health. In light of the importance of this foodborne pathogen, the aim of this study was to perform comparative analyses on 80 Brazilian Campylobacter coli genomes isolated from human feces, animals, the environment, and food. Methods include Average Nucleotide Identity (ANI), Gegenees, genomic plasticity, presence of pathogenicity, resistance, and metabolic islands. In addition, virulence analysis in Galleria mellonella were also performed for 18 selected C. coli strains.ResultsThe ANI values confirmed that all strains belonged to the C. coli species. Phylogenetic analyses demonstrated the evolutionary relationships among the studied strains, highlighting the genetic diversity among them. The differences in shared and deleted regions of the studied genomes were demonstrated, with 16 genomic islands identified, including 4 metabolic islands, 4 resistance islands, and 8 pathogenicity islands. We detected genes associated with chemotaxis, exotoxin production, antimicrobial resistance, stress response, defense mechanisms, and intracellular survival among these islands, highlighting the pathogenic potential of these strains. Two strains isolated from human and one from animal showed high virulence, killing 100% of Galleria mellonella larvae. Two strains isolated from the environment and two isolated from food killed 70–90% of the larvae and were classified as virulent. Three strains isolated from animal, two from human, two from the environment and one from food killed 30% to 60% of the larvae and were considered of intermediate virulence. Campylobacter jejuni ATCC 33291, one strain isolated from human and one from food killed 10 to 20% of the larvae and were considered of low virulence. One strain isolated from food did not kill any larvae and was considered avirulent.ConclusionsThe results obtained highlighted the genetic diversity, pathogenic and virulence potential of many of the C. coli strains studied, contributing for a more complete characterization of this important pathogen recognized as a cause of human gastroenteritis.

Virulence is not directly related to strain success in planta in Clavibacter nebraskensis.

Goss's wilt and leaf blight of maize is an economically important disease caused by the Gram-positive bacterium, Clavibacter nebraskensis (Cn). Little is known about the ecology and pathogenesis of this bacterium. Here, we used phenotypic assays and a high-throughput whole-genome sequencing approach to explore among-strain variation in virulence and multistrain reproductive success in planta. Our survey of 41 strains revealed that more recently sampled strains tended to have higher virulence than strains sampled before 2010 and tended to be more genetically divergent from the reference strain, isolated in 1971. More detailed assays with a representative sample of 13 of these strains revealed that host genotype (resistant or susceptible) did not strongly affect strain success and that strain success in planta in multi-strain communities was not closely associated with virulence in single-strain assays. Two weakly virulent strains, CIC354 and CIC370, had the greatest reproductive success, whereas the most highly virulent strains did not significantly change in frequency in any host genotype. A genomic analysis revealed candidate genes, including putative virulence factors (i.e., a secreted cellulase), responsible for among-strain variation in reproductive success.IMPORTANCENon-pathogenic strains of many bacterial pathogens are reported to coexist with pathogenic strains in symptomatic plants. To understand the ecology and pathogenesis of the pathogen population, it is essential to study strain dynamics in the context of the host. We created a community of 13 strains exhibiting diverse virulence phenotypes and used this community to infect the host plant. We compared the strain frequency of these strains before and after the host infection. Contrary to our hypothesis of highly virulent strains being selected by the susceptible host, we found that weakly virulent strains were selected by both resistant and susceptible host lines. We identified several genes associated with strain frequency shifts suggesting their role in strain colonization, virulence, and fitness.

Integrating Diagnostic Approaches in Infant Bacterial Meningitis Caused by a Non-K1 Escherichia coli: A Case Report

Background: Infant meningitis, particularly caused by Escherichia coli, remains a life-threatening condition, especially in premature and low-weight infants. Infections of the central nervous system can be fatal, necessitating prompt diagnosis and appropriate treatment. Acute infections caused by various pathogens, including E. coli, often present with similar clinical symptoms. The rapid identification of pathogens and their antimicrobial resistance mechanisms is critical for timely and effective treatment. We report the case of an 8-month-old patient who presented with fever, diarrhea, and convulsive seizures and was subsequently diagnosed with meningitis. Despite initial empirical treatment with ceftriaxone, the patient’s condition worsened. Methods: At Bambino Gesù Children’s Hospital, molecular diagnostic tools, including the FilmArray Meningitis/Encephalitis and Blood Culture Identification panels, were employed. Results: Although the Meningitis panel did not detect any pathogens due to the lack of the specific bacterial target, the off-label use of the Blood Culture Identification panel identified a non-K1 Escherichia coli strain carrying the CTX-M resistance gene, an extended-spectrum beta-lactamase (ESBL). Despite the rapid diagnostic approach and adjustment of antibiotic therapy, the patient succumbed to the infection due to the strain’s high virulence and multidrug resistance. Whole-genome sequencing further characterized the strain, revealing that it belonged to the ST131 group, a highly resistant and virulent strain associated with sepsis. Conclusions: This case highlights the importance of integrating advanced molecular diagnostics, such as whole-genome sequencing, with traditional methods to improve pathogen detection, especially in cases of emerging resistant strains that are not covered by standard diagnostic panels. It also emphasizes the need for the continuous adaptation of diagnostic tools to include non-K1 E. coli strains for more comprehensive and timely meningitis diagnosis.

A novel pigeon paramyxovirus type 1 isolated from a sick racing pigeon in the Qinghai-Tibet Plateau of China shows high virulence in chickens

A novel pigeon paramyxovirus type 1 isolated from a sick racing pigeon in the Qinghai-Tibet Plateau of China shows high virulence in chickens

Development of the duplex droplet digital PCR for quantitative monitoring of mixed infections of Meloidogyne incognita and M. enterolobii.

Root-knot nematodes (RKN, Meloidogyne spp.) are economically significant pests that cause immense damage to a wide range of crops. Among them, M. incognita and M. enterolobii are of particular concern, as their high virulence and broad host range. RKN are challenging for detection due to their subterranean lifestyle underground. Also, the mixed infection of nematodes in field crops complicates the need for more accurate diagnostic and quantification technologies. To address this challenge, we developed and optimized a novel duplex droplet digital PCR (ddPCR) method, using primer/probe sets targeting M. incognita and M. enterolobii, to simultaneously identify and quantify both species within a single assay. The innovative ddPCR diagnostic demonstrated excellent performance in terms of sensitivity, precision and reproductivity when quantifying the eggs and soil samples containing juveniles of both species. Moreover, the application of the duplex ddPCR method enables the monitoring of population dynamics of M. incognita and M. enterolobii under competitive environmental conditions. Our results indicated that the reproduction factor of M. incognita possibly inhibited when in mixed populations of M. enterolobii. In this study, we first applied duplex ddPCR technique for differentiating mixed infections of M. incognita and M. enterolobii, offering a valuable tool for species detection and quantification. It enables the monitoring of population dynamics for both species, which is crucial for providing theoretical guidance for the implementation of timely and effective control measures. © 2024 Society of Chemical Industry.

Identification of Pseudopestalotiopsis ampullacea as a new pathogen causing tea gray blight in India and its management strategies

Gray blight is a serious threat to the tea [Camellia sinensis (L.) O. Kuntze] production in major tea cultivating countries including India. The disease is caused by Pestalotiopsis-like species. In this study, five isolates of Pseudopestalotiopsis species isolated from symptomatic tea leaf samples in North Bengal, India were investigated. Based on the multi-locus phylogenetic analysis using concatenated sequences of three (ITS, tef-1 alpha, and tub-2) loci, cultural and micromorphological characters, and host association, the fungal isolates were identified as Pseudopestalotiopsis ampullacea F. Liu & L. Cai. The morphological analysis also revealed that the fungal isolates were evidently differentiated from other Pseudopestalotiopsis species. To date, P. ampullacea has not been reported on tea plants in India. Among the five isolates studied, isolate NKT0P03 was randomly selected for pathogenicity tests and its sensitivity to fungicides and microbial antagonists. In pathogenicity test, the isolate showed weak to high virulence reactions on 25 different tea cultivars. The pathogen showed an avirulent reaction on the cultivar TV11. In order to identify an effective management strategy against this new pathogen, synthetic fungicides and microbial biocontrol agents were evaluated in the laboratory. Results revealed that carbendazim + mancozeb, hexaconazole, propiconazole, and valextra were effective fungicides with an 85.1% to 89.8% range of inhibitory activity against P. ampullacea NKT0P03. Among microbial agents, Trichoderma harzianum, T. reesei, T. hamatum, Bacillus subtilis, and Microbacterium barkeri were efficient bioagents against P. ampullacea NKT0P03 with antagonistic activity ranging between 66.6% and 84.2%. Thus, these fungicides and microbial bioagents can be recommended as effective agents for the management of P. ampullacea causing tea gray blight after their field evaluations.

Emergence of polymyxin-resistant Yersinia enterocolitica strains in natural aquatic environments

Aquatic environments serve as ideal reservoirs for antibiotic-resistant bacteria and resistance genes. However, the presence of polymyxin-resistant Yersinia enterocolitica, the pathogen responsible for human yersiniosis, in aquatic environments remains poorly understood. Herein, we isolated polymyxin-resistant Y. enterocolitica strains from natural water for the first time. In addition to intrinsic resistance to ampicillin and cefazolin, the strains demonstrated high resistance to polymyxin B and polymyxin E. All isolates were capable of biofilm production and exerted high virulent effects in Galleria mellonella, with 90% mortality occurring within 48 h post-infection. Furthermore, whole genome sequencing identified 26 antibiotic resistance genes, including polymyxin resistance determinants (arnA and PmrF), beta-lactam resistance determinants (vatF and blaA), and 60 virulence genes such as yaxA and yaxB in Y. enterocolitica isolates. Notably, phylogenetic analysis revealed that Y. enterocolitica involved multilocus sequence types ST937 and ST631, which were clustered with strains isolated from a human in the United States or swine in China. The close relatedness to clinical isolates suggests that polymyxin-resistant Y. enterocolitica may pose considerable health risk to humans. Our findings provide evidence of the presence of polymyxin-resistant Y. enterocolitica in aquatic environments and raise concerns about health risks due to their potential high virulence.

Author Correction: Dominant HPAIV H5N1 genotypes of Germany 2021/2022 are linked to high virulence in Pekin ducklings

Author Correction: Dominant HPAIV H5N1 genotypes of Germany 2021/2022 are linked to high virulence in Pekin ducklings

Relationship Between the Presence of Red Complex Species and the Distribution of Other Oral Bacteria, Including Major Periodontal Pathogens in Older Japanese Individuals.

Red complex bacteria (Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia) have high virulence in periodontal disease. In the present study, we aimed to elucidate the detailed symbiotic relationships between the red complex and other oral bacteria in older Japanese individuals. Polymerase chain reaction was performed using dental plaque from 116 subjects and specific primers for ten periodontal pathogens. The detection rate of Prevotella intermedia and Capnocytophaga sputigena was significantly higher in P. gingivalis-positive subjects than in P. gingivalis-negative subjects (p < 0.05). The detection rate of Campylobacter rectus, Prevotella nigrescens, Capnocytophaga ochracea, and Eikenella corrodens was significantly higher in T. forsythia-positive subjects than in T. forsythia-negative subjects (p < 0.01). In a comprehensive analysis of oral microbiomes, three red complex species-positive subjects had significantly higher α-diversity than only P. gingivalis-positive subjects (p < 0.05) and had significantly lower β-diversity than only T. forsythia-positive subjects (p < 0.01). In the taxonomy analysis, Porphyromonas was significantly higher in three red complex species-positive subjects than in only P. gingivalis-positive and only T. forsythia-positive subjects (p < 0.01). These results suggest that each red complex species forms a unique oral microbiome and individuals positive for all red complex bacteria may harbor oral bacteria that confer a significant advantage in developing periodontal disease.

The overlooked shadow killer: Elizabethkingia anophelis as a potent pathogen in American bullfrogs (Aquarana catesbeiana)

Elizabethkingia anophelis has garnered significant attention due to its rising prevalence in clinical settings, yet its impact on animal infections remains underexplored. In this study, we isolated the strain AWYL-1 from infected American bullfrogs (Aquarana catesbeiana) and conducted comprehensive characterization of its biological properties. Morphologically and biochemically, AWYL-1 resembled the known bullfrog pathogen Elizabethkingia miricola, and exhibited multidrug resistance, being sensitive only to doxycycline. AWYL-1 demonstrated high virulence, with a 100 % mortality rate in bullfrogs at a concentration of 1 × 109 CFU/mL within 14 days, regardless of the infection route, including immersion and intramuscular injection. Gene expression analysis revealed significant upregulation of immune-related genes (e.g., P65, CXCL10, IFNγ) and cell death-related genes (e.g., BNIP3, Caspase-8) in the kidney and spleen, particularly at 72 h post-infection. Furthermore, experimental infection in mice resulted in a 50 % mortality rate, confirming the cross-species infection potential of AWYL-1. Histopathological examination revealed multi-organ hyperemia, edema, and inflammation, with significant pathological changes in the liver, spleen, kidneys, and lungs. These findings suggest that E. anophelis poses a zoonotic risk, capable of infecting both amphibians and mammals. This study provides critical data for the prevention and control of E. anophelis infections in aquaculture, highlighting the need for ongoing vigilance and further research into its potential impact on public health.

Machine Learning Approaches for Microorganism Identification, Virulence Assessment, and Antimicrobial Susceptibility Evaluation Using DNA Sequencing Methods: A Systematic Review.

Microbial infections pose a substantial global health challenge, particularly impacting immunocompromised individuals and exacerbating the issue of antimicrobial resistance (AMR). High virulence of pathogens can lead to severe infections and prolonged antimicrobial treatment, increasing the risk of developing resistant strains. Integrating machine-learning (ML) with DNA sequencing technologies offers potential solutions by enhancing microbial identification, virulence assessment, and antimicrobial susceptibility evaluation. This review explores recent advancements in these integrated approaches, addressing current limitations and identifying gaps in the literature. A comprehensive literature search was conducted across databases including PubMed, Scopus, Web of Science, and IEEE Xplore, covering publications from January 2014 to June 2024. Using a detailed Boolean search string, relevant studies focusing on ML applications in microorganism identification, antimicrobial susceptibility testing, and microbial virulence were included. The screening process involved a two-stage review of titles, abstracts, and full texts, with data extraction and critical appraisal performed using the QIAO tool. Data were analyzed through narrative synthesis to identify common themes and innovations. Out of 1,650 initially identified records, 19 studies met the inclusion criteria. These studies primarily focused on AMR, with additional research on microbial virulence and identification. Machine learning algorithms such as Random Forest, Support Vector Machines, and Convolutional Neural Networks, combined with DNA sequencing techniques like Whole Genome Sequencing and Metagenomic Sequencing, demonstrated significant advancements in predictive accuracy and efficiency. High-quality studies achieved impressive performance metrics, including F1-scores up to 0.88 and AUC scores up to 0.96. The integration of ML and DNA sequencing technologies has significantly enhanced microbial analysis, improving the identification of pathogens, assessment of virulence, and evaluation of antimicrobial susceptibility. Despite advancements, challenges such as data quality, high costs, and model interpretability persist. This review highlights the need for continued innovation and provides recommendations for future research to address these limitations and improve disease management and public health strategies. The systematic review is registered with PROSPERO (CRD42024571347).

Dominant HPAIV H5N1 genotypes of Germany 2021/2022 are linked to high virulence in Pekin ducklings

Highly pathogenic avian influenza viruses (HPAIV) of H5 clade 2.3.4.4b pose an ongoing threat worldwide. It remains unclear whether this panzootic situation would favor low virulent phenotypes expected by the ‘avirulence hypothesis’ of viral evolution. Assessing virulence in Pekin ducklings in an intramuscular infection model revealed that the two genotypes that dominated the epidemiological situation in Germany during the period 2021 and 2022 (EU-RL:CH and EU-RL:AB) were of high virulence. In contrast, rare genotypes were of intermediate virulence. The genetic constellation of these reassortants pointed to an important role of the viral polymerase complex (RdRP), particularly the PB1 genome segment, in shaping virulence in ducklings. Occulo-nasal infection of ducklings confirmed the phenotypes for two representative viruses and indicated a more efficient replication for the high virulence strain. These observations would be in line with the ‘virulence-transmission trade-off’ model for describing HPAIV epidemiology in wild birds in Germany.

NezhNPV, a new biocontrol agent for Nesodiprion zhejiangensis Zhou & Xiao (Hymenoptera: Diprionidae), an emerging forest pest.

Nesodiprion zhejiangensis, a multivoltine sawfly, is widely distributed in south China and has caused serious damage to forests. Historically, N. zhejiangensis management has relied heavily on synthetic chemicals. To reduce the reliance on chemical control, we previously isolated a nucleopolyhedrovirus, NezhNPV, from deceased N. zhejiangensis larvae. A subsequent pathogenicity assay confirmed its high virulence in a laboratory setting. In order to comprehensively examine the hypothesis that NezhNPV is an effective new biocontrol agent for N. zhejiangensis, we carried out a field test in Beijing, China, and characterized NezhNPV morphologically by electron microscopy and genetically by genome sequencing. Our field trials showed that NezhNPV was effective in controlling N. zhejiangensis in a naturally infested Himalayan blue pine forest. The occlusion bodies of NezhNPV consist of irregular polyhedra that occlude rod-shaped enveloped virions with a single nucleocapsid per virion. The NezhNPV genome is 80 637 bp in length, and contains 90 open reading frames, including 38 core, eight lepidopteran baculovirus, 34 hymenopteran baculovirus and 10 unique baculovirus genes, representing the smallest known genome among baculoviruses. The combined results based on phylogenetic analyses, Kimura-2-parameter distances and biological characteristics indicate that NezhNPV is a novel gammabaculovirus and candidate for species status with the provisional name Gammabaculovirus nezhejiangensis. NezhNPV is highly collinear with other gammabaculoviruses and contains nonsyntenic regions with an inversion and rearrangement between orf3 and orf35. The combined results from our field trials, coupled with morphological and genomic characterization clearly demonstrate the bioactivity of NezhNPV. This gammabaculovirus may be included in pest management practices against N. zhejiangensis as a novel biocontrol agent. © 2024 The Author(s). Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Whitefly‐Pathogenic Beauveria bassiana JEF‐507 as a Competitive Biopesticide, Chongchae‐Stop

ABSTRACTSilverleaf whitefly, Bemisia tabaci (Hemiptera: Aleyrodidae), is a destructive insect pest damaging to diverse crops by vectoring several plant pathogenic viruses, which consequently causes economic losses in crop production. As the resistance of whiteflies to chemical insecticides is increasing, this study aimed to investigate the potential of entomopathogenic fungi as an alternative. A total of 72 entomopathogenic fungal isolates, collected from soils using Tenebrio molitor larvae as an insect baiting method, were assessed for their virulence against 2nd nymphs of whitefly. Their virulence was assayed by dipping whitefly‐infested tomato leaves in fungal conidia suspensions at 1.0 × 107 conidia/mL. Among the tested isolates, two isolates of Beauveria bassiana JEF‐462 and JEF‐507 showed high virulence. In the assessment of virulence depending on conidia concentrations, the estimated LC50 values for JEF‐462 and JEF‐507 were similarly 8.7–14.0 × 107 conidia/mL. However, B. bassiana JEF‐507 showed higher conidial productivity and thermotolerance on most of tested 12 grain substrates than B. bassiana JEF‐462, and millet was the most suitable grain substrate. Additionally, siloxane as a surfactant was able to sufficiently exhibit the insecticidal activity of JEF‐507 against whitefly nymphs compared with other surfactants. In a pot‐based greenhouse trial, JEF‐507 showed higher control efficacy than chemical insecticides, dinotefuran and spinetoram. This work suggests that B. bassiana JEF‐507 could be competitively used as a biopesticide to control silverleaf whiteflies while overcoming current resistance issues. The JEF‐507 isolate has been registered in Korea, 2022 and successfully commercialised as the name of Chongchae‐Stop in this local market to control whitefly and thrips.

Characterization of a Highly Virulent Klebsiella michiganensis Strain Isolated from a Preterm Infant with Sepsis.

Klebsiella michiganensis is an opportunistic pathogen that causes an increasing number of serious infections. This study aimed to investigate the etiology of the severe clinical symptoms of sepsis in preterm infants and the characterization of K. michiganensis isolates. Whole-genome sequencing (WGS) was performed on three strains isolated from an infected preterm infant. Additionally, the genomic sequences of 534 K. michiganensis strains were obtained from the NCBI database. To gain deeper insights into these strains, we utilized the Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Clusters of Orthologous Groups (COG), and Pathogen Host Interactions (PHI) database annotation tools for comprehensive gene function analyses. Moreover, the multilocus sequence typing (MLST), EasyCGtree, and virulence factor database (VFDB) were employed to determine the sequence types (STs), construct phylogenetic trees, and identify potential virulence factors. Sequence analysis found that the three isolated strains had identical sequence characteristics and did not correspond to any of the known ST types. Virulence factor analysis revealed that the three strains harbored mrkABCDFHIJ, fimABCDEFGHIK, entABCDEFS, fepABCD, and capsule genes. These virulence factors are likely to play crucial roles in enhancing adhesion and metabolic capabilities, resisting phagocytosis (inducing immune cell damage), and ultimately contributing to prolonged bacteremia. The phylogenetic tree and comparative genomics of virulence factors showed the genetic and virulence factor diversity of the currently reported K. michiganensis strains. We identified a novel strain of K. michiganensis that exhibits high virulence and leads to severe septicemia phenotypes in preterm infants. Furthermore, comparative genomic analysis of previously reported K. michiganensis strains revealed the existence of three clades. This comprehensive analysis provides novel insights into the genetic relationships and virulence factor profiles of diverse strains of K. michiganensis. In future, it will be necessary to investigate the concept of the high virulence of K. michiganensis to determine the treatment method.

Genomic characterization of a clonal emergent Salmonella Minnesota lineage in Brazil reveals the presence of a novel megaplasmid of resistance and virulence.

Salmonella Minnesota has emerged in Brazil as the predominant serovar in poultry and poultry products, along with Salmonella Heidelberg. To understand the emergence of Salmonella Minnesota over the last few years in Brazil, we performed a comparative analysis between 69 selected S. Minnesota genomes from Pathogen Detection database and 65 clonal emergent genomes isolated from Brazil. We demonstrate the presence of multidrug resistance genes against tetracycline [tet(A)], sulfonamide (sul2), and AmpC beta-lactamase (blaCMY-2) in emergent genomes, along with the carriage of a megaplasmid of resistance and virulence (~210 kb), designated pESM (plasmid for emergent Salmonella Minnesota). pESM is an IncC/A2 plasmid predicted to increase S. Minnesota environmental tolerance to mercury (mer operon) and provide resistance to tetracycline and ampicillin due to the presence of tet(A) and blaCMY-2, respectively. Moreover, pESM carries the yersiniabactin siderophore (high-pathogenicity island of Yersinia) related to the iron uptake. The temporal inference demonstrated that the most recent common ancestor dated from ~1978 and that the clonal emergent genomes carrying the pESM belong to a completely different lineage of S. Minnesota. Our results indicate that the presence of pESM likely contributes to the emergence of S. Minnesota and is precisely related to the successful spread of this particular clonal lineage in Brazil.IMPORTANCESalmonella Minnesota has emerged in Brazil as one of the leading serovars related to human and animal infection, presenting high virulence and antibiotic resistance to drugs classified as the highest priority for clinical treatment in humans. This study performed whole-genome sequencing, temporal analysis, and phylogenetics to understand the genetic insights related to the emergence of Salmonella Minnesota in Brazil. Long-read sequencing has led to the identification and characterization of a unique megaplasmid carrying virulence, antibiotic resistance, and heavy-metal tolerance genes, which may play a central role in S. Minnesota's successful emergence in Brazil and possibly worldwide. The potentially high transmissibility of this plasmid between clones and serovars represents a risk to public health since its acquisition may increase Salmonella's fitness, virulence, resistance, and persistence. Understanding the genetic aspects related to the emergence of serovars can help devise measures to mitigate the spread of hazardous multidrug-resistant strains.

Isolation and characterization of a novel S-gene mutation porcine deltacoronavirus with high pathogenicity from diarrhea piglet in Zhejiang Province, China, 2022

Porcine deltacoronavirus (PDCoV) is a coronavirus that causes diarrhea in suckling piglets and has the potential for cross-species transmission. Monitoring PDCoV evolution and identifying potential vaccine candidates are crucial due to its high mutation rates in pig populations. In this study, a Chinese PDCoV strain named ZD2022 was successfully isolated from diarrhea piglets in Zhejiang province, followed by genetic evolutionary analysis, assessment of S proteins’ biological functions, in vitro cellular adaptation analysis and pathogenicity evaluation. Phylogenetic analyses placed the PDCoV ZD2022 strain within the Southeast Asia Lineage. Sequence analysis revealed 23 mutations in the S protein of ZD2022 compared to most of other Chinese PDCoV strains, including 8 unique mutations (T529I, L579F, Q614H, V709G, S959L, P1010S, V1016F, A1068V). In addition, bioinformatic predictions indicated these mutations impact the hydrophilicity/hydrophobicity, antigenic epitopes and N-glycosylation sites of the ZD2022 S protein. The virus growth curve of ZD2022 showed good cellular adaptation, with peak viral titers of 8.92 ± 0.31 Log10 TCID50/mL in ST cells. Furthermore, ZD2022 exhibited high virulence in suckling piglets, causing severe diarrhea in piglets at 60 h post-inoculation (hpi) and a mortality rate of 40 % (2/5) within 96 hpi. In summary, our findings indicate that the Chinese PDCoV strains continue to mutate, and the novel S gene mutation in strain ZD2022 offers strong cellular adaptation and high pathogenicity, making it a potential candidate strain for vaccine development.