Bacterial Symbionts in Ceratitis capitata.

Ceratitis capitata (Diptera: Tephritidae) is responsible for extensive damage in agriculture with important economic losses. Several strategies have been proposed to control this insect pest including insecticides and the Sterile Insect Technique. Traditional control methods should be implemented by innovative tools, among which those based on insect symbionts seem very promising. Our study aimed to investigate, through the 16S Miseq analysis, the microbial communities associated with selected organs in three different medfly populations to identify possible candidates to develop symbiont-based control approaches. Our results confirm that Klebsiella and Providencia are the dominant bacteria in guts, while a more diversified microbial community has been detected in reproductive organs. Concertedly, we revealed for the first time the presence of Chroococcidiopsis and Propionibacterium as stable components of the medfly's microbiota. Additionally, in the reproductive organs, we detected Asaia, a bacterium already proposed as a tool in the Symbiotic Control of Vector-Borne Diseases. A strain of Asaia, genetically modified to produce a green fluorescent protein, was used to ascertain the ability of Asaia to colonize specific organs of C. capitata. Our study lays the foundation for the development of control methods for C. capitata based on the use of symbiont bacteria.

Draft genome of an extremely drug-resistant st551 Staphylococcus pseudintermedius from an Italian dog with otitis externa.

OBJECTIVES: To determine the draft genome sequence and analyse the genetic features of a Staphylococcus pseudintermedius clinical isolate according to the main typing schemes available, with a special focus on antibiotic resistance. METHODS: The strain was isolated from a case of otitis externa in a dog. Its identity and pattern of antibiotic susceptibility were determined using an automated system. The genome was sequenced using an Illumina platform. MLST, SCCmec typing, resistome, and mobile genetic elements were derived by comparative analysis using available specific databases. RESULTS: S. pseudintermedius CAM1 isolate has a chromosome size of 2 652 610 bp. It showed a wide pattern of phenotypic resistance, comprising beta-lactams, macrolides and lincosamides, aminoglycosides, fluoroquinolones, tetracyclines, and trimethoprim-sulfamethoxazole. The genetic determinants of the underlying mechanisms were all found by in silico analysis of the genome. The mecA gene for methicillin resistance was harboured by the Vc type of the SCCmec. MLST of the strain was st551. CONCLUSION: By comparison with the MLST database of S. pseudintermedius and data from published molecular epidemiology studies, CAM1 is the first st551 strain recorded in Italy and, in the context of an already extremely wide antibiotic resistance pattern, it harbours also the tetK gene, the prevalence of which is rare in MDR S. pseudintermedius.

Testing Surgical Face Masks in an Emergency Context: The Experience of Italian Laboratories during the COVID-19 Pandemic Crisis.

The first wave of the COVID-19 pandemic brought about a broader use of masks by both professionals and the general population. This resulted in a severe worldwide shortage of devices and the need to increase import and activate production of safe and effective surgical masks at the national level. In order to support the demand for testing surgical masks in the Italian context, Universities provided their contribution by setting up laboratories for testing mask performance before releasing products into the national market. This paper reports the effort of seven Italian university laboratories who set up facilities for testing face masks during the emergency period of the COVID-19 pandemic. Measurement set-ups were built, adapting the methods specified in the EN 14683:2019+AC. Data on differential pressure (DP) and bacterial filtration efficiency (BFE) of 120 masks, including different materials and designs, were collected over three months. More than 60% of the masks satisfied requirements for DP and BFE set by the standard. Masks made of nonwoven polypropylene with at least three layers (spunbonded-meltblown-spunbonded) showed the best results, ensuring both good breathability and high filtration efficiency. The majority of the masks created with alternative materials and designs did not comply with both standard requirements, resulting in suitability only as community masks. The effective partnering between universities and industries to meet a public need in an emergency context represented a fruitful example of the so-called university "third-mission".

Structure/activity virtual screening and in vitro testing of small molecule inhibitors of 8-hydroxy-5-deazaflavin:NADPH oxidoreductase from gut methanogenic bacteria.

Virtual screening techniques and in vitro binding/inhibitory assays were used to search within a set of more than 8,000 naturally occurring small ligands for candidate inhibitors of 8-hydroxy-5-deazaflavin:NADPH oxidoreductase (FNO) from Methanobrevibacter smithii, the enzyme that catalyses the bidirectional electron transfer between NADP+ and F420H2 during the intestinal production of CH4 from CO2. In silico screening using molecular docking classified the ligand-enzyme complexes in the range between - 4.9 and - 10.5 kcal/mol. Molecular flexibility, the number of H-bond acceptors and donors, the extent of hydrophobic interactions, and the exposure to the solvent were the major discriminants in determining the affinity of the ligands for FNO. In vitro studies on a group of these ligands selected from the most populated/representative clusters provided quantitative kinetic, equilibrium, and structural information on ligands' behaviour, in optimal agreement with the predictive computational results.

The volatile oils from the oleo-gum-resins of Ferula assa-foetida and Ferula gummosa: A comprehensive investigation of their insecticidal activity and eco-toxicological effects.

Since time immemorial, the oleo-gum-resins of Ferula assa-foetida and F. gummosa are used in the traditional medical systems as well as in foodstuffs, perfumery and cosmetics. In the present study, we explored the insecticidal efficacy of the essential oils obtained from these oleo-gum-resins to widen their fields of industrial applications. The two essential oils were mainly composed of sulfides [sec-butyl (Z)-propenyl disulfide, sec-butyl (E)-propenyl disulfide, sec-butyl (Z)-propenyl trisulfide and sec-butyl (E)-propenyl trisulfide)] and monoterpenes (α-pinene, ß-pinene and ß-phellandrene), respectively, as determined by GC-MS analysis. The two essential oils were assayed for toxicity on a panel of insects, represented by species of public health relevance (Culex quinquefasciatus and Musca domestica), agricultural (Spodoptera littoralis) and stored-product pests (Prostephanus truncatus and Trogoderma granarium). The ecotoxicological effects of the essential oils were assessed on the aquatic microcrustacean Daphnia magna and the earthworm Eisenia fetida, as well as on human cells. Overall, the two essential oils were effective against important insect pests and vectors. On the other hand, they resulted cytotoxic to fibroblasts and non-target aquatic microcrustaceans. Thus, further insights are needed to determine the full spectrum of their eco-toxicological effects.

Exploring new applications of tulip tree (Liriodendron tulipifera L.): leaf essential oil as apoptotic agent for human glioblastoma.

Liriodendron tulipifera L. (Magnoliaceae), also known as "tulip tree," is a hardwood plant native to North America, cultivated all over the world and used on an industrial level, especially for its fine wood and to make honey. It has also been traditionally exploited for its antimalarial properties. However, our knowledge about the bioactivity of its essential oil remains patchy. In this research, we focused on the biological evaluation of the volatile fractions obtained from different parts of the plant which are normally discharged by industry, including leaves, flowers, and fruits. For the purpose, the essential oils were obtained by hydrodistillation and analyzed by gas chromatography-mass spectrometry (GC-MS). Then, they were evaluated as radical scavenging, antioxidant, antimicrobial, and antiproliferative agents by using DPPH, ABTS, FRAP, disk diffusion, and MTT methods, respectively. The significant toxicity exhibited on human tumor cells, namely A375 malignant melanoma, HCT116 colon carcinoma, MDA-MB 231 breast adenocarcinoma, and T98G glioblastoma multiforme cell lines, prompted us to study the mechanism of action by acridine orange/ethidium bromide double staining and caspase 3 assays. Our findings shed light on the potential applications of tulip tree derivatives as anticancer drugs.

Quaternary Ammonium Leucine-Based Surfactants: The Effect of a Benzyl Group on Physicochemical Properties and Antimicrobial Activity.

Quaternary ammonium amphiphiles are a class of compounds with a wide range of commercial and industrial uses. In the pharmaceutical field, the most common quaternary ammonium surfactant is benzalkonium chloride (BAC), which is employed as a preservative in several topical formulations for ocular, skin, or nasal application. Despite the broad antimicrobial activity against Gram-positive and Gram-negative bacteria, as well as fungi and small enveloped viruses, safety concerns regarding its irritant and cytotoxic effect on epithelial cells still remain. In this work, quaternary ammonium derivatives of leucine esters (C10, C12 and C14) were synthesised as BAC analogues. These cationic surfactants were characterised in terms of critical micelle concentration (CMC, by tensiometry), cytotoxicity (MTS and LDH assays on the Caco-2 and Calu-3 cell lines) and antimicrobial activity on the bacterial species Staphylococcus aureus and Enterococcus faecalis among the Gram-positives, Escherichia coli and Pseudomonas aeruginosa among the Gram-negatives and the yeast Candida albicans. They showed satisfactory surface-active properties, and a cytotoxic effect that was dependent on the length of the hydrophobic chain. Lower minimum inhibiting concentration (MIC) values were calculated for C14-derivatives, which were comparable to those calculated for BAC toward Gram-positive bacteria and slightly higher for Gram-negative bacteria and C. albicans. Thus, the synthesised leucine-based quaternary ammonium cationic surfactants can potentially find application as promising surface-active compounds with antimicrobial activity.

High virulence gene diversity in Streptococcus pyogenes isolated in Central Italy.

Globally, Streptococcus pyogenes poses a continuous burden on human health, causing both self-limiting and life-threatening diseases. Therefore, studying the profile of virulence genes and their combinations is essential to monitor the epidemiology and pathogenic potential of this important species. Thus, the aim of this study was to analyze related genetic features of clinical strains collected in Italy in 2012 in order to obtain a valid picture of their virulence profile that could be compared to similar studies made in other countries approximately in the same period. We conducted emm typing and fibronectin-collagen-T antigen (FCT) region typing in 122 Streptococcus pyogenes strains. Furthermore, several additional virulence genes were screened by polymerase chain reaction. We found correlations between emm types and FCT region profiles. emm1 strains were mainly associated with FCT2 and FCT6, while emm89 and emm12 strains were associated with FCT4. FCT5 was mainly represented in emm4, emm6, and emm75 strains. Significantly, we defined subtypes for each FCT type based on the differences in single and double loci compared to the reference scheme used for the classification of the FCT region. In addition, new FCT-region variants with differences in multiple loci were also recorded. Cluster analysis based on virulence gene profiling showed a non-random distribution within each emm type. This study added new data to existing studies conducted worldwide and revealed new variability scores in circulating Streptococcus pyogenes strains and new assortments in well-established virulence gene signatures.

Bioactivity and Structural Properties of Novel Synthetic Analogues of the Protozoan Toxin Climacostol.

Climacostol (5-[(2Z)-non-2-en-1-yl]benzene-1,3-diol) is a resorcinol produced by the protozoan Climacostomum virens for defence against predators. It exerts a potent antimicrobial activity against bacterial and fungal pathogens, inhibits the growth of several human and rodent tumour cells, and is now available by chemical synthesis. In this study, we chemically synthesized two novel analogues of climacostol, namely, 2-methyl-5 [(2Z)-non-2-en-1-yl]benzene-1,3-diol (AN1) and 5-[(2Z)-non-2-en-1-yl]benzene-1,2,3-triol (AN2), with the aim to increase the activity of the native toxin, evaluating their effects on prokaryotic and free-living protists and on mammalian tumour cells. The results demonstrated that the analogue bearing a methyl group (AN1) in the aromatic ring exhibited appreciably higher toxicity against pathogen microbes and protists than climacostol. On the other hand, the analogue bearing an additional hydroxyl group (AN2) in the aromatic ring revealed its ability to induce programmed cell death in protistan cells. Overall, the data collected demonstrate that the introduction of a methyl or a hydroxyl moiety to the aromatic ring of climacostol can effectively modulate its potency and its mechanism of action.

A New Species of the γ-Proteobacterium Francisella, F. adeliensis Sp. Nov., Endocytobiont in an Antarctic Marine Ciliate and Potential Evolutionary Forerunner of Pathogenic Species.

The study of the draft genome of an Antarctic marine ciliate, Euplotes petzi, revealed foreign sequences of bacterial origin belonging to the γ-proteobacterium Francisella that includes pathogenic and environmental species. TEM and FISH analyses confirmed the presence of a Francisella endocytobiont in E. petzi. This endocytobiont was isolated and found to be a new species, named F. adeliensis sp. nov.. F. adeliensis grows well at wide ranges of temperature, salinity, and carbon dioxide concentrations implying that it may colonize new organisms living in deeply diversified habitats. The F. adeliensis genome includes the igl and pdp gene sets (pdpC and pdpE excepted) of the Francisella pathogenicity island needed for intracellular growth. Consistently with an F. adeliensis ancient symbiotic lifestyle, it also contains a single insertion-sequence element. Instead, it lacks genes for the biosynthesis of essential amino acids such as cysteine, lysine, methionine, and tyrosine. In a genome-based phylogenetic tree, F. adeliensis forms a new early branching clade, basal to the evolution of pathogenic species. The correlations of this clade with the other clades raise doubts about a genuine free-living nature of the environmental Francisella species isolated from natural and man-made environments, and suggest to look at F. adeliensis as a pioneer in the Francisella colonization of eukaryotic organisms.

Microbiological evaluation of ready-to-eat iceberg lettuce during shelf-life and effectiveness of household washing methods.

The aim of this study was to assess the microbiological quality of ready-to-eat (RTE) iceberg lettuce. Our investigation was based on the consumption tendency of university students considered a target market for this product. A total of 78 RTE samples were collected from chain supermarkets and analysed for the enumeration of aerobic mesophilic count (AMC), Escherichia coli and the detection of Salmonella spp. and Listeria monocytogenes. All samples were negative for the presence of pathogens. The mean value of AMC at the beginning, in the middle and after the expiration date was: 6.88, 8.51 and 8.72 log CFU g-1, respectively. The same investigation was performed on 12 samples of fresh iceberg lettuce samples. No pathogens were found and the mean value of AMC was lower than the RTE category (5.73 log CFU g-1; P<0.05). The effectiveness of 5 washing methods was determined on 15 samples of both fresh and RTE iceberg lettuce. Samples were washed for 15' and 30' in tap water (500 mL), tap water with NaCl (4 g/500 mL), tap water with bicarbonate (8 g/500 mL), tap water with vinegar (10 mL/500 mL) and tap water with chlorine-based disinfectant (10 mL/500 mL). A significant bacterial load reduction was recorded for vinegar and disinfectant after 30' and 15', respectively. Overall, these results showed that RTE iceberg lettuce is more contaminated than the fresh product. Also, the consumption in the first few days of packaging and after washing with disinfectants reduces the risk for health consumers.

Phylogenetic relationships, biofilm formation, motility, antibiotic resistance and extended virulence genotypes among Escherichia coli strains from women with community-onset primitive acute pyelonephritis.

The present work set out to search for a virulence repertoire distinctive for Escherichia coli causing primitive acute pyelonephritis (APN). To this end, the virulence potential of 18 E. coli APN strains was genotypically and phenotypically assessed, comparatively with 19 strains causing recurrent cystitis (RC), and 16 clinically not significant (control, CO) strains. Most of the strains belong to phylogenetic group B1 (69.8%; p<0.01), and APN strains showed unique features, which are the presence of phylogroup A, and the absence of phylogroup B2 and non-typeable strains. Overall, the most dominant virulence factor genes (VFGs) were ecpA and fyuA (92.4 and 86.7%, respectively; p<0.05), and the mean number of VFGs was significantly higher in uropathogenic strains. Particularly, papAH and malX were exclusive for uropathogenic strains. APN and RC strains showed a significantly higher prevalence of fyuA, usp, and malX than of CO strains. Compared to RC strains, APN ones showed a higher prevalence of iha, but a lower prevalence of iroN, cnf1, and kpsMT-II. Hierarchical cluster analysis showed a higher proportion of two gene clusters (malX and usp, and fyuA and ecpA) were detected in the APN and RC groups than in CO, whereas iutA and iha clusters were detected more frequently in APN strains. The motility level did not differ among the study-groups and phylogroups considered, although a higher proportion of swarming strains was observed in APN strains. Antibiotic-resistance rates were generally low except for ampicillin (37.7%), and were not associated with specific study- or phylogenetic groups. APN and RC strains produced more biofilm than CO strains. In APN strains, iha was associated with higher biofilm biomass formation, whereas iroN and KpSMT-K1 were associated with a lower amount of biofilm biomass. Further work is needed to grasp the virulence and fitness mechanisms adopted by E. coli causing APN, and hence develop new therapeutic and prophylactic approaches.

Antidiabetic "gliptins" affect biofilm formation by Streptococcus mutans.

Streptococcus mutans, a dental caries causing odontopathogen, produces X-prolyl dipeptidyl peptidase (Sm-XPDAP, encoded by pepX), a serine protease known to have a nutritional role. Considering the potential of proteases as therapeutic targets in pathogens, this study was primarily aimed at investigating the role of Sm-XPDAP in contributing to virulence-related traits. Dipeptidyl peptidase (DPP IV), an XPDAP analogous enzyme found in mammalian tissues,is a well known therapeutic target in Type II diabetes. Based on the hypothesis that gliptins, commonly used as anti-human-DPP IV drugs, may affect bacterial growth upon inhibition of Sm-XPDAP, we have determined their ex vivo antimicrobial and anti-biofilm activity towards S. mutans. All three DPP IV drugs tested reduced biofilm formation as determined by crystal violet staining. To link the observed biofilm inhibition to the human-DPP IV analogue present in S. mutans UA159, a pepX isogenic mutant was generated. In addition to reduced biofilm formation, CLSM studies of the biofilm formed by the pepX isogenic mutant showed these were comparable to those formed in the presence of saxagliptin, suggesting a probable role of this enzyme in biofilm formation by S. mutans UA159. The effects of both pepX deletion and DPP IV drugs on the proteome were studied using LC-MS/MS. Overall, this study highlights the potential of Sm-XPDAP as a novel anti-biofilm target and suggests a template molecule to synthesize lead compounds effective against this enzyme.

High prevalence of clonally diverse spa type t026 Staphylococcus aureus contaminating rural eggshells.

PURPOSE: The presence of Staphylococcus aureus in poultry and poultry products, including eggs, increases its potential to enter the food chain, resulting in foodborne diseases. In this context, eggshell colonization by staphylococci may represent a risk factor. This study aimed to investigate the contamination of rural eggshell by S. aureus and to characterize the key features of the isolated strains. METHODOLOGY: Antibiotic resistance was assessed by disc diffusion. Resistant isolates were analysed by PCR for the identification of associated genetic determinants of resistance. PCR was also used to screen for the presence of genes coding for toxins, namely, sea, sec, sei, sem, seo and tst. The genetic characterization was extended by means of agr locus typing and spa typing. RESULTS: 34 S. aureus were isolated. Macrolide- and tetracycline-resistant strains were prevalent. All strains were susceptible to oxacillin, cefoxitin and trimethoprim-sulfamethoxazole. PCR screening for genes encoding enterotoxins detected several virulence patterns, which, together with spa-typing and agr-locus typing, allowed cluster analysis and the description of novel clones. CONCLUSION: Continuous monitoring of staphylococci is needed also in rural or natural settings. Increasing the number of samples and expanding the geographical region will be needed to further extend the significance of the study.

Phenolic acids, antioxidant and antiproliferative activities of Naviglio® extracts from Schizogyne sericea (Asteraceae).

Schizogyne sericea, well-known as 'salado', is a halophytic shrub widespread on coastal rocks of Tenerife (Canary Islands). This plant is used traditionally as analgesic, astringent, anti-inflammatory and vulnerary agent. In the present work, we have analysed the aqueous and ethanolic extracts of S. sericea for the content of phenolic acids by HPLC-DAD. The dynamic solid-liquid Naviglio® extractor was used to extract the flowering aerial parts. Aqueous extracts showed higher levels of phenolics than ethanolic extracts. S. sericea extracts were rich in chlorogenic and isochlorogenic acids. The Naviglio® extracts obtained were assayed for in vitro biological activities, namely antioxidant, antimicrobial and cytotoxicity on tumour cells by DPPH, ABTS, FRAP, agar disc-diffusion and MTT methods, respectively. Results showed that aqueous extracts, being richer in phenolic acids, are endowed with relevant radical scavenging activity (TEAC values in the range 208-960 µmol TE/g) while ethanolic extracts exhibited noteworthy antiproliferative effects on tumour cells.

Antimicrobial and antioxidant activity of the essential oil from the Carpathian Thymus alternans Klokov.

The genus Thymus includes several species that are used as flavouring, food preservative as well as in cosmetics. Their secondary metabolites have been extensively studied for pharmacological effects. Nonetheless, some species are neglected and deserve to be explored for chemical composition and biological activities. This is the case with Thymus alternans, a Carpathian bush used as a food additive and for the preparation of a traditional herbal medicine. In this work, we have analysed the chemical composition of T. alternans essential oil by GC and GC-MS and evaluated its antimicrobial and antioxidant activity by disc diffusion, DPPH, ABTS and FRAP methods, respectively. Results showed that T. alternans belongs to the nerolidol chemotype, being rich of this sesquiterpene alcohol (15.8%) which might contribute to the antimicrobial (particularly effective on C. albicans growth) and antioxidant (weak inhibition on ABTS radical and reducing power) activities observed.

Genotypic and phenotypic heterogeneity in Streptococcus mutans isolated from diabetic patients in Rome, Italy.

Our study focuses on the antimicrobial susceptibility, genotypic and phenotypic heterogeneity, and serotype classification of the Streptococcus mutans isolated from type II diabetic patients (n = 25; age 42-68). Eighty-two percent of isolates were classified as serotype c. No serotype k was present. Macrorestriction analysis of genomic DNA of the isolates exhibited a clonal diversity that paralleled the phenotypic heterogeneity, which was also assessed in terms of biofilm forming ability. Isolates were susceptible to all the classes of antibiotics. In conclusion a great heterogeneity and no antimicrobial resistance were apparent in the considered S. mutans strains from diabetic patients.

Biological Activities of the Essential Oil from Erigeron floribundus.

Erigeron floribundus (Asteraceae) is an herbaceous plant widely used in Cameroonian traditional medicine to treat various diseases of microbial and non-microbial origin. In the present study, we evaluated the in vitro biological activities displayed by the essential oil obtained from the aerial parts of E. floribundus, namely the antioxidant, antimicrobial and antiproliferative activities. Moreover, we investigated the inhibitory effects of E. floribundus essential oil on nicotinate mononucleotide adenylyltransferase (NadD), a promising new target for developing novel antibiotics, and Trypanosoma brucei, the protozoan parasite responsible for Human African trypanosomiasis. The essential oil composition was dominated by spathulenol (12.2%), caryophyllene oxide (12.4%) and limonene (8.8%). The E. floribundus oil showed a good activity against Staphylococcus aureus (inhibition zone diameter, IZD of 14 mm, minimum inhibitory concentration, MIC of 512 µg/mL). Interestingly, it inhibited the NadD enzyme from S. aureus (IC50 of 98 µg/mL), with no effects on mammalian orthologue enzymes. In addition, T. brucei proliferation was inhibited with IC50 values of 33.5 µg/mL with the essential oil and 5.6 µg/mL with the active component limonene. The essential oil exhibited strong cytotoxicity on HCT 116 colon carcinoma cells with an IC50 value of 14.89 µg/mL, and remarkable ferric reducing antioxidant power (tocopherol-equivalent antioxidant capacity, TEAC = 411.9 µmol·TE/g).

Bioactive Secondary Metabolites from Schizogyne sericea (Asteraceae) Endemic to Canary Islands.

Schizogyne sericea (Asteraceae) is a halophytic shrub endemic to the Canary Islands and traditionally employed as analgesic, astringent, anti-inflammatory, and vulnerary. A comprehensive phytochemical investigation was conducted on the flowering aerial parts by analyzing both essential oil constituents and polar compounds. The essential oil was dominated by p-cymene, with the noteworthy occurrence of ß-pinene and thymol esters. From the EtOH extract, eight compounds were isolated and structurally elucidated. Essential oil, polar fractions, and isolates (2), (4), and (5) were separately in vitro assayed for antiproliferative activity on human tumor cell lines (A375, MDA-MB 231, and HCT116) by MTT assay, for antioxidant potential by DPPH, ABTS, and FRAP assays, and for antimicrobial activity by the agar disk diffusion method. Results revealed that essential oil and compounds 1 and 2 exert a strong inhibition on tumor cells, and in some cases, higher than that of cisplatin. Fractions containing thymol derivatives (1 and 2) and caffeoylquinic acid derivatives 4 and 5 displayed antioxidant activity comparable to that of Trolox, making S. sericea extract an interesting natural product with potential applications as preservative or in the treatment of diseases in which oxidative stress plays an important role.

Transcriptome Remodeling Contributes to Epidemic Disease Caused by the Human Pathogen Streptococcus pyogenes.

UNLABELLED: For over a century, a fundamental objective in infection biology research has been to understand the molecular processes contributing to the origin and perpetuation of epidemics. Divergent hypotheses have emerged concerning the extent to which environmental events or pathogen evolution dominates in these processes. Remarkably few studies bear on this important issue. Based on population pathogenomic analysis of 1,200 Streptococcus pyogenes type emm89 infection isolates, we report that a series of horizontal gene transfer events produced a new pathogenic genotype with increased ability to cause infection, leading to an epidemic wave of disease on at least two continents. In the aggregate, these and other genetic changes substantially remodeled the transcriptomes of the evolved progeny, causing extensive differential expression of virulence genes and altered pathogen-host interaction, including enhanced immune evasion. Our findings delineate the precise molecular genetic changes that occurred and enhance our understanding of the evolutionary processes that contribute to the emergence and persistence of epidemically successful pathogen clones. The data have significant implications for understanding bacterial epidemics and for translational research efforts to blunt their detrimental effects. IMPORTANCE: The confluence of studies of molecular events underlying pathogen strain emergence, evolutionary genetic processes mediating altered virulence, and epidemics is in its infancy. Although understanding these events is necessary to develop new or improved strategies to protect health, surprisingly few studies have addressed this issue, in particular, at the comprehensive population genomic level. Herein we establish that substantial remodeling of the transcriptome of the human-specific pathogen Streptococcus pyogenes by horizontal gene flow and other evolutionary genetic changes is a central factor in precipitating and perpetuating epidemic disease. The data unambiguously show that the key outcome of these molecular events is evolution of a new, more virulent pathogenic genotype. Our findings provide new understanding of epidemic disease.