ABSTRACT
Bovine tuberculosis (bTB) is a chronic inflammatory disease primarily caused by Mycobacterium bovis. The infection affects domestic animals and wildlife, posing a zoonotic risk to humans. To understand the dynamics of transmission and genetic diversity in Italy's M. bovis population, we conducted whole-genome sequencing (WGS) analysis on two prevalent genotypes, belonging to Spoligotype SB0120, identified in different geographical and temporal contexts. By comparing these genomes with international M. bovis isolates, we identified a distinct clade within the lineage La1.2, encompassing the Italian SB0120 isolates, indicating a genomic segregation of Italian M. bovis from other European isolates. Within Italy, a significant level of genetic variability emerged across regions, while isolates within epidemiologically linked outbreaks exhibited minimal genetic diversity. Additionally, isolates derived from cattle and wild boars within a tuberculosis hotspot in Central Italy and from cattle and black pigs in Sicily formed unified clonal clusters. This indicates the presence of persistent strains circulating in the examined regions. The genetic diversity within herds was limited, as specific clones endured over time within certain herds. This research enhances our comprehension of the epidemiology and transmission patterns of bTB in Italy, thereby aiding the development of precise control strategies and disease management. Using WGS and implementing standardized protocols and databases will be pivotal in combating bTB and promoting One-Health approaches to address this noteworthy public health concern.
ABSTRACT
Tularemia is a rare but potentially life threatening zoonotic disease caused by Francisella tularensis. F. novicida, previously considered a subspecies of F. tularensis, is currently considered a separate species. Human infections related to F. novicida are exceedingly rare but can cause morbidity and mortality in debilitated or immunocompromised individuals.A 42-year-old male presented at the hospital with vomiting, dehydration, constipation and pain in the right iliac fossa. He was first diagnosed with pancreatitis and admitted for further analysis. Chest computerized tomography scan showed the presence of parenchymal consolidation in the left upper and lower lobes of the lung with pleural effusion. Blood cultures isolated a Gram-negative coccobacillus, that was at first identified by MALDI-TOF as Francisella tularensis. Serological analysis for the detection of total antibodies against F. tularensis and Real-Time PCR targeting the gene coding for 23 kDa, resulting negative. Subsequently, PCR targeting helicases and tul4 genes, and the Regions of Difference RD1 and RD6 were performed allowing the identification of F. novicida. The isolate was further genetically characterized by whole genome sequencing (WGS).This is the first reported case of human infection caused by F. novicida in Italy.Given the rarity of human cases and the lack of specific symptoms, this pathogen is difficult to identify and the diagnosis can be extremely challenging. In this case report, despite the lack of amplification of the gene encoding for 23 kDa protein, the identification of Francisella species was achieved with the amplification of different genes and characterized by WGS.
ABSTRACT
Nanoplastics (NPs) affect fertility. We evaluated the effects of NPs treatment on luteal and endothelial cells. We examined crucial markers of growth and redox status. NPs treatment did not induce changes in ATP levels in luteal cells, while it increased (p< 0.05) their proliferation. In endothelial cells, no change in proliferation was detected, while an increase (p<0.05) in ATP levels was observed. The increase of reactive oxygen species, superoxide anion (p<0.05) and nitric oxide (p<0.001) was detected in both cell types, which also showed changes in superoxide dismutase enzyme activity as well as an increase of non-enzymatic antioxidant power (p<0.05). A decrease (p<0.05) in progesterone production as well as an increase of vascular endothelial growth factor A levels were detected (p<0.05). In addition, a dose-dependent accumulation of NPs in endothelial cells was shown, that likely occurred through adhesion and internalization. Results underline potential risk of NPs for corpus luteum functionality.
ABSTRACT
OBJECTIVES: The aim of the project was to develop and characterise powders containing a probiotic (Lactiplantibacillus plantarum [Lpb. plantarum], Lacticaseibacillus rhamnosus, or Lactobacillus acidophilus) to be administered to the lung for the containment of pathogen growth in patients with lung infections. METHODS: The optimised spray drying process for the powder manufacturing was able to preserve viability of the bacteria, which decreased of only one log unit and was maintained up to 30 days. RESULTS: Probiotic powders showed a high respirability (42%-50% of particles had a size < 5 µm) suitable for lung deposition and were proven safe on A549 and Calu-3 cells up to a concentration of 107 colony-forming units/mL. The Lpb. plantarum adhesion to both cell lines tested was at least 10%. Surprisingly, Lpb. plantarum powder was bactericidal at a concentration of 106 colony-forming units/mL on P. aeruginosa, whereas the other two strains were bacteriostatic. CONCLUSION: This work represents a promising starting point to consider a probiotic inhalation powder a value in keeping the growth of pathogenic microflora in check during the antibiotic inhalation therapy suspension in cystic fibrosis treatment regimen. This approach could also be advantageous for interfering competitively with pathogenic bacteria and promoting the restoration of the healthy microbiota.
Subject(s)
Lactobacillales , Probiotics , Pseudomonas Infections , Humans , Pseudomonas aeruginosa , Powders , Anti-Bacterial Agents/pharmacologyABSTRACT
Plastic is an important environmental issue and a more critical aspect concerns plastic fragments, mainly in term of nanoplastics (NPs). We demonstrated that NPs interfere with reproductive and adipose stromal cells. Since several research underlined an increased cardiovascular risk due to NPs, present study was undertaken to investigate their effect on aortic endothelial cells (AOC). We explored the specificity of their interaction with endothelial cells, quantifying their load in treated cells. Then, NPs effect was assessed on cell growth, generation of free radicals and antioxidant defence. Our data demonstrate that NPs colocalize with AOC. We found a significant (p < 0.01) increase both in metabolic activity and Vascular Endothelial Growth Factor (VEGF) production (p < 0.01). Redox status appeared to be disrupted (p < 0.05) by NPs. Taken together, the normal function of cultured AOC appeared negatively affected by AOC. Since NPs have been detected in blood, our present data appear of particular interest.
Subject(s)
Endothelial Cells , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth Factor A/metabolism , Microplastics , Oxidative Stress , AortaABSTRACT
IMPORTANCE: This study integrated population data with in vitro assessment of virulence phenotypes to unveil that a considerable part of the global population of Salmonella Derby is evolving to enhance its host adaptation to the swine host and that this evolution is simultaneously increasing its attenuation for humans. The study shows that the fixation of deleterious mutations in SPI-1 has a role in this process. This evidence indicates that SPI-1 has a key role for S. Derby virulence in humans but not for its circulation in swine. The results show that genes generally considered essential for Salmonella pathogenesis do not play the same key role for all Salmonella serovars or lineages and/or all hosts. The study helps in understanding the molecular mechanisms underlying the ecology and host adaptation of Salmonella showing that the adaptation process can vary for different types of Salmonella and hosts.
Subject(s)
Genomic Islands , Salmonella enterica , Humans , Animals , Swine , Salmonella enterica/genetics , Salmonella/genetics , Phenotype , Bacterial Proteins/geneticsABSTRACT
We report Listeria monocytogenes infection in a patient in Italy who was transfused with pooled platelet concentrate. Genomic analysis revealed that L. monocytogenes isolates from the donor blood unit, the transfused platelets, and the patient's blood culture were genetically closely related, confirming transfusion transmission. Additional surveillance and secondary bacterial screening could improve transfusion safety.
Subject(s)
Listeria monocytogenes , Listeriosis , Humans , Listeria monocytogenes/genetics , Blood Platelets , Platelet Transfusion/adverse effects , Listeriosis/microbiology , Italy/epidemiology , Food MicrobiologyABSTRACT
Rare cases of Pseudomonas aeruginosa community-acquired pneumonia (PA-CAP) were reported in non-immunocompromised patients. We describe a case of Pseudomonas aeruginosa (PA) necrotizing cavitary CAP with a fatal outcome in a 53-year-old man previously infected with SARS-CoV-2, who was admitted for dyspnea, fever, cough, hemoptysis, acute respiratory failure and a right upper lobe opacification. Six hours after admission, despite effective antibiotic therapy, he experienced multi-organ failure and died. Autopsy confirmed necrotizing pneumonia with alveolar hemorrhage. Blood and bronchoalveolar lavage cultures were positive for PA serotype O:9 belonging to ST1184. The strain shares the same virulence factor profile with reference genome PA01. With the aim to better investigate the clinical and molecular characteristics of PA-CAP, we considered the literature of the last 13 years concerning this topic. The prevalence of hospitalized PA-CAP is about 4% and has a mortality rate of 33-66%. Smoking, alcohol abuse and contaminated fluid exposure were the recognized risk factors; most cases presented the same symptoms described above and needed intensive care. Co-infection of PA-influenza A is described, which is possibly caused by influenza-inducing respiratory epithelial cell dysfunction: the same pathophysiological mechanism could be assumed with SARS-CoV-2 infection. Considering the high rate of fatal outcomes, additional studies are needed to identify sources of infections and new risk factors, along with genetic and immunological features. Current CAP guidelines should be revised in light of these results.
ABSTRACT
Listeria monocytogenes is an ubiquitous pathogen isolated from different host species including fish, crustaceans, and molluscs, but it is rarely a pathogenic microorganism to marine reptiles. In particular, only two cases of fatal disseminated listeriosis have been described in the loggerhead sea turtle (Caretta caretta). In this study, we describe a lethal case of L. monocytogenes infection in a loggerhead sea turtle. The turtle was found alive, stranded on a beach in North-eastern Italy, but perished soon after being rescued. The autoptic examination revealed that heart, lung, liver, spleen, and urinary bladder were disseminated with multiple, firm, 0.1-0.5 mm sized, nodular, white-green lesions. Microscopically, these lesions corresponded with heterophilic granulomas with Gram+ bacteria within the necrotic center. Furthermore, the Ziehl-Neelsen stain was negative for acid-fast organisms. Colonies isolated from heart and liver were tested through MALDI-TOF for species identification, revealing the presence of L. monocytogenes. Whole Genome Sequencing on L. monocytogenes isolates was performed and the subsequent in silico genotyping revealed the belonging to Sequence Type 6 (ST 6); the virulence profile was evaluated, showing the presence of pathogenicity islands commonly observed in ST 6. Our results further confirm that L. monocytogenes should be posed in differential diagnosis in case of nodular lesions of loggerhead sea turtles; thus, given the zoonotic potential of the microorganism, animals should be treated with particular caution. In addition, wildlife animals can play an active role as carriers of possibly pathogenetic and virulent strains and contribute to the distribution of L. monocytogenes in the environment.
ABSTRACT
Two Escherichia coli strains, respectively responsible for neonatal and adult meningitis, were isolated and their phenotypic antibiotic susceptibility and genomic features characterized by whole-genome sequencing (WGS). Multiplex real-time PCR targeting the principal microorganisms involved in meningitis etiology failed to identify either isolate. Afterwards, matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry was used to identify the isolates as E. coli strains. Genomic analysis showed that they belonged to sequence types 131 and 69 (ST131 and ST69). Neither of the isolates harbored the K1 capsular antigen or belonged to other capsular serotypes, but they shared different virulence factors, including ibe genes, responsible for invasion of brain endothelial cells. IMPORTANCE The extraintestinal pathogenic Escherichia coli group is characterized by the presence of uropathogenic E. coli (UPEC), sepsis-associated E. coli (SEPEC), and neonatal meningitis E. coli (NMEC). All of these members exhibit many virulence factors, such as lipopolysaccharides, toxins, iron acquisition factors, invasins, fimbriae, and capsules. Urinary infections are the most common infections caused by this group, followed by globally increasing numbers of both community- and nosocomially acquired bloodstream infections, associated with considerable patient morbidity and mortality. Some lineages tend to become dominant; in addition to enhanced fitness, this epidemiological success stems from increased virulence, antibiotic resistance, gut colonization, and greater host-to-host transmission. Our results underline the importance of continuous surveillance of these new emerging lineages and the need to develop new meningitis molecular assay panels able to identify them.
ABSTRACT
A clinical strain of Klebsiella pneumoniae typed as sequence type 307 carrying three different alleles of the flu gene encoding the Escherichia coli virulence factor antigen 43 associated with biofilm formation was detected and characterized. The flu alleles are located in the chromosome inside putative integrative conjugative elements. The strain displays the phenotypes associated with Ag43, i.e. bi-phasic colony morphology and enhanced biofilm production. Furthermore, the strain produces low amount of capsule known to affect Ag43 function. Analysis of 1431 worldwide deposited genomes revealed that 3.7% Klebsiella pneumoniae carry one or two flu alleles.
Subject(s)
Escherichia coli Proteins , Klebsiella pneumoniae , Alleles , Anti-Bacterial Agents , Antigens, Bacterial/genetics , Biofilms , Colistin , Escherichia coli/genetics , Escherichia coli Proteins/genetics , Klebsiella pneumoniae/genetics , Microbial Sensitivity Tests , Plasmids/geneticsABSTRACT
BACKGROUND: Klebsiella pneumoniae is a common species in the gut of mammals and is widely distributed in the environment. However, the environmental source of hvKp that precedes gut colonization is unclear, but once that it reaches the gut there is a possible generalized spread y fecal-oral transmission especially in endemic areas. Liver abscess might develop when the bacteria, using its virulence factors, cross the intestinal barrier and invade the liver by the portal circulation. This syndrome, prevalent mostly in Asian countries, is increasingly reported in Western Countries and leaves open questions about the source of infection. CASE: Here we describe for the first time in Italy, a case of pyogenic liver abscess caused by a hypervirulent Klebsiella pneumoniae (HvKp) complicated by endophthalmitis and other metastatic infections in lung and prostate in an immunocompetent Chinese healthy individual with no recent travel in Asia. CONCLUSION: This case underlines the need for increased awareness of hypervirulent K. pneumoniae, even in settings where it occurs infrequently and where there are not evident epidemiological links.
Subject(s)
Endophthalmitis , Klebsiella Infections , Liver Abscess , Male , Animals , Humans , Klebsiella pneumoniae , Virulence , Klebsiella Infections/complications , Liver Abscess/complications , Liver Abscess/microbiology , Endophthalmitis/diagnosis , MammalsABSTRACT
BACKGROUND: To date, whole genome sequencing has been performed mainly for isolates of Chlamydia trachomatis, C. pneumoniae, C. psittaci and C. abortus, but only a few isolates of C. pecorum have been entirely sequenced and this makes it difficult to understand its diversity and population structure. In this study the genome of two C. pecorum strains isolated from the lung of an Alpine chamois affected with pneumonia (isolate PV7855) and the brain of a water buffalo affected with meningoencephalomyelitis (isolate PV6959), were completely sequenced with MiSeq system (Illumina) and analyzed in their most polymorphic regions. RESULTS: The genome length and GC content of the two isolates were found to be consistent with other C. pecorum isolates and the gene content of polymorphic membrane proteins and plasticity zone was found to be very similar. Some differences were observed in the phospholipase genes for both isolates and in the number of genes in the plasticity zone, such as the presence of some hypothetical proteins in PV6959, not present in any other genomes analyzed in this study. Interestingly, PV6959 possesses an extra pmp and has an incomplete tryptophan biosynthesis operon. Plasmids were detected in both isolates. CONCLUSIONS: Genome sequencing of the two C. pecorum strains did not reveal differences in length and GC content despite the origin from different animal species with different clinical disease. In the plasticity zone, the differences in the genes pattern might be related to the onset of specific symptoms or infection of specific hosts. The absence of a tryptophan biosynthesis pathway in PV6959 may suggest a strict relationship between C. pecorum and its host.
Subject(s)
Rupicapra , Animals , Buffaloes , Chlamydia , Chlamydia trachomatis , Rupicapra/metabolism , Tryptophan/metabolismABSTRACT
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) emerge for their capability to better adapt to the human host aimed and enhance human-to-human transmission. Mutations in spike largely contributed to adaptation. Viral persistence is a prerequisite for intra-host virus evolution, and this likely occurred in immunocompromised patients who allow intra-host long-term viral replication. The underlying mechanism leading to the emergence of variants during viral persistence in the immunocompromised host is still unknown. Here, we show the existence of an ensemble of minor mutants in the early biological samples obtained from an immunocompromised patient and their dynamic interplay with the master mutant during a persistent and productive long-term infection. In particular, after 222 days of active viral replication, the original master mutant, named MB610, was replaced by a minor quasispecies (MB61222) expressing two critical mutations in spike, namely Q493K and N501T. Isolation of the two viruses allowed us to show that MB61222 entry into target cells occurred mainly by the fusion at the plasma membrane (PM), whereas endocytosis characterized the entry mechanism used by MB610. Interestingly, coinfection of two human cell lines of different origin with the SARS-CoV-2 isolates highlighted the early and dramatic predominance of MB61222 over MB610 replication. This finding may be explained by a faster replicative activity of MB61222 as compared to MB610 as well as by the capability of MB61222 to induce peculiar viral RNA-sensing mechanisms leading to an increased production of interferons (IFNs) and, in particular, of IFN-induced transmembrane protein 1 (IFITM1) and IFITM2. Indeed, it has been recently shown that IFITM2 is able to restrict SARS-CoV-2 entry occurring by endocytosis. In this regard, MB61222 may escape the antiviral activity of IFITMs by using the PM fusion pathway for entry into the target cell, whereas MB610 cannot escape this host antiviral response during MB61222 coinfection, since it has endocytosis as the main pathway of entry. Altogether, our data support the evidence of quasispecies fighting for host dominance by taking benefit from the cell machinery to restrict the productive infection of competitors in the viral ensemble. This finding may explain, at least in part, the extraordinary rapid worldwide turnover of VOCs that use the PM fusion pathway to enter into target cells over the original pandemic strain.
ABSTRACT
This case report describes for the first-time cases of severe gastroenteritis in water buffalo calves due to a new serovar of Salmonella enterica. The study was carried out on fecal matrix collected from live water buffalo calves that showed profuse diarrhea, severe dehydration and fever, exhibiting a systemic course. Culture and molecular investigations identified the pathogens isolated from intestinal contents as two Salmonella serovars, Salmonella enterica enterica O:35 and a new serovar of Salmonella enterica. The isolates showed multi-drug resistance. Timely diagnosis associated with a targeted antimicrobial treatment were found to be sufficient for the survival and recovery of the infected animals. Herd vaccines prepared from isolated pathogens were used to prevent further deaths of the calves.
ABSTRACT
This study aims to describe trends of mcr-positive Enterobacterales in humans based on laboratory surveillance with a defined catchment population. The data source is the Micro-RER surveillance system, established in Emilia-Romagna region (Italy), to monitor the trend of mcr resistance. Enterobacterales isolates from human clinical samples with minimum inhibitory concentration (MIC) ≥ 2 mg/L for colistin were sent to the study reference laboratory for the detection of mcr genes. Isolates prospectively collected in the period 2018-2020 were considered for the assessment of population rates and trends; further analyses were carried out for the evaluation of clonality and horizontal mcr gene transfer. Previous isolates from local laboratory collection were also described. In the period 2018-2020, 1164 isolates were sent to the reference laboratory, and 51 (4.4%) were confirmed as mcr-positive: 50 mcr-1 (42 Escherichia coli, 6 Klebsiella pneumoniae, 2 Salmonella enterica) and 1 mcr-4 (Enterobacter cloacae). The number of mcr-positive isolates dropped from 24 in the first half of 2018 to 3 in the whole of 2020 (trend p value < 0.001). Genomic analyses showed the predominant role of the horizontal transfer of mcr genes through plasmids or dissemination of transposable elements compared to clonal dissemination of mcr-positive microorganisms. The study results demonstrate a substantial decrease in the circulation of mcr-1 plasmid genes in Emilia-Romagna Region.
Subject(s)
Bacterial Proteins/metabolism , Enterobacteriaceae Infections/microbiology , Enterobacteriaceae/enzymology , Ethanolaminephosphotransferase/metabolism , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/genetics , Drug Resistance, Bacterial , Enterobacteriaceae/classification , Enterobacteriaceae/drug effects , Enterobacteriaceae/genetics , Enterobacteriaceae Infections/epidemiology , Ethanolaminephosphotransferase/genetics , Humans , Italy/epidemiology , Microbial Sensitivity Tests , Phylogeny , Retrospective StudiesABSTRACT
BACKGROUND: the co-production of carbapenemases and mcr-genes represents a worrisome event in the treatment of Enterobacteriaceae infections. The aim of the study was to characterize the genomic features of two clinical Enterobacter cloacae complex (ECC) isolates, co-producing VIM and MCR enzymes, in Italy. METHODS: species identification and antibiotic susceptibility profiling were performed using MALDI-TOF and broth microdilution methods, respectively. Transferability of the bla VIM- and mcr- type genes was verified through conjugation experiment. Extracted DNA was sequenced using long reads sequencing technology on the Sequel I platform (PacBio). RESULTS: the first isolate showed clinical resistance against ertapenem yet was colistin susceptible (EUCAST 2020 breakpoints). The mcr-9.2 gene was harbored on a conjugative IncHI2 plasmid, while the bla VIM-1 determinant was harbored on a conjugative IncN plasmid. The second isolate, resistant to both carbapenems and colistin, harbored: mcr-9 gene and its two component regulatory genes for increased expression on the chromosome, mcr-4.3 on non-conjugative (yet co-transferable) ColE plasmid, and bla VIM-1 on a non-conjugative IncA plasmid. CONCLUSIONS: to our knowledge, this is the first report of co-production of VIM and MCR in ECC isolates in Italy.
ABSTRACT
Background: The bacteria that compose the Mycobacterium tuberculosis complex (MTBC) cause tuberculosis (TB) in humans and in different animals, including livestock. Much progress has been made in understanding the population structure of the human-adapted members of the MTBC by combining phylogenetics with genomics. Accompanying the discovery of new genetic diversity, a body of operational nomenclature has evolved to assist comparative and molecular epidemiological studies of human TB. By contrast, for the livestock-associated MTBC members, Mycobacterium bovis, M. caprae and M. orygis, there has been a lack of comprehensive nomenclature to accommodate new genetic diversity uncovered by emerging phylogenomic studies. We propose to fill this gap by putting forward a new nomenclature covering the main phylogenetic groups within M. bovis, M. caprae and M. orygis. Methods: We gathered a total of 8,736 whole-genome sequences (WGS) from public sources and 39 newly sequenced strains, and selected a subset of 829 WGS, representative of the worldwide diversity of M. bovis, M. caprae and M. orygis. We used phylogenetics and genetic diversity patterns inferred from WGS to define groups. Results: We propose to divide M. bovis, M. caprae and M. orygis in three main phylogenetic lineages, which we named La1, La2 and La3, respectively. Within La1, we identified several monophyletic groups, which we propose to classify into eight sublineages (La1.1-La1.8). These sublineages differed in geographic distribution, with some being geographically restricted and others globally widespread, suggesting different expansion abilities. To ease molecular characterization of these MTBC groups by the community, we provide phylogenetically informed, single nucleotide polymorphisms that can be used as barcodes for genotyping. These markers were implemented in KvarQ and TB-Profiler, which are platform-independent, open-source tools. Conclusions: Our results contribute to an improved classification of the genetic diversity within the livestock-associated MTBC, which will benefit future molecular epidemiological and evolutionary studies.
ABSTRACT
Salmonella enterica variants exhibit diverse host adaptation, outcome of infection, and associated risk to food safety. Analysis of the distribution of Salmonella enterica serovar Derby (S. Derby) subtypes in human and swine identified isolates with a distinct PFGE profile that were significantly under-represented in human infections, consistent with further host adaptation to swine. Here we show that isolates with this PFGE profile form a distinct phylogenetic sub-clade within S. Derby and exhibit a profound reduction in invasion of human epithelial cells, and a relatively small reduction in swine epithelial cells. A single missense mutation in hilD, that encodes the master-regulator of the Salmonella Pathogenicity Island 1 (SPI-1), was present in the adapted lineage. The missense mutation resulted in a loss of function of HilD that accounted for reduced invasion in human epithelial cells. The relatively small impact of the mutation on interaction with swine cells was consistent with an alternative mechanism of invasion in this pathogen-host combination.