Invasive Bacterial Pathogens Research Articles

The arginine/ornithine binding protein ArgT plays an essential role in Brucella neotomae/Brucella melitensis to prevent intracellular killing and contribute to chronic persistence in the host

ABSTRACT Brucella species are facultative intracellular bacterial pathogens that cause the contagious zoonotic disease, brucellosis. Brucella spp. infect a wide range of animals, including livestock, wild animals, and marine mammals. Compared with other invasive bacterial pathogens, partial information is available on the virulence factors of Brucella that enable them to survive in the host. Here, we performed transposon-based random mutagenesis of B. neotomae and identified the arginine/ornithine binding protein, ArgT, as one of the crucial virulence determinants of Brucella. Deleting ArgT from B. neotomae or B. melitensis resulted in its attenuation in macrophages, which was restored upon complementation with an ArgT expression plasmid. We observed that macrophages infected with ΔArgT-B. neotomae produced elevated levels of NO due to the inability of these mutants to deplete the host intracellular arginine through their importer. Furthermore, defective survival of ΔArgT B. neotomae and B. melitensis was observed in the infected mice, which correlated with enhanced NO production in the mice. Our studies revealed that ArgT plays a vital role in preventing intracellular killing and contributes to the chronic persistence of B. neotomae/B. melitensis in the host. This study highlights the essential role of arginine in clearing intracellular infections and the subversion of this host defence mechanism by intracellular pathogens for their chronic persistence.

Coronatine orchestrates ABI1-mediated stomatal opening to facilitate bacterial pathogen infection through importin β protein SAD2.

Stomatal immunity plays an important role during bacterial pathogen invasion. Abscisic acid (ABA) induces plants to close their stomata and halt pathogen invasion, but many bacterial pathogens secrete phytotoxin coronatine (COR) to antagonize ABA signaling and reopen the stomata to promote infection at early stage of invasion. However, the underlining mechanism is not clear. SAD2 is an importin β family protein, and the sad2 mutant shows hypersensitivity to ABA. We discovered ABI1, which negatively regulated ABA signaling and reduced plant sensitivity to ABA, was accumulated in the plant nucleus after COR treatment. This event required SAD2 to import ABI1 to the plant nucleus. Abolition of SAD2 undermined ABI1 accumulation. Our study answers the long-standing question of how bacterial COR antagonizes ABA signaling and reopens plant stomata during pathogen invasion.

Butyrate affects bacterial virulence: a new perspective on preventing enteric bacterial pathogen invasion.

Enteric bacterial pathogens are a major threat to intestinal health. With the widespread use of antibiotics, bacterial resistance has become a problem, and there is an urgent need for a new treatment to reduce dependence on antibiotics. Butyrate can control enteric bacterial pathogens by regulating the expression of their virulence genes, promoting the posttranslational modification of their proteins, maintaining an anaerobic environment, regulating the host immune systemand strengthening the intestinal mucosal barrier. Here, thisreview describes the mechanisms by which butyrate regulates the pathogenicity of enteric bacterial pathogens from various perspectives and discusses the prospects and limitations of butyrate as a new option for the control of pathogenic bacteria.

Bacteriophages targeting protective commensals impair resistance against Salmonella Typhimurium infection in gnotobiotic mice.

Gut microbial communities protect the host against a variety of major human gastrointestinal pathogens. Bacteriophages (phages) are ubiquitous in nature and frequently ingested via food and drinking water. Moreover, they are an attractive tool for microbiome engineering due to the lack of known serious adverse effects on the host. However, the functional role of phages within the gastrointestinal microbiome remain poorly understood. Here, we investigated the effects of microbiota-directed phages on infection with the human enteric pathogen Salmonella enterica serovar Typhimurium (S. Tm), using a gnotobiotic mouse model (OMM14) for colonization resistance (CR). We show, that phage cocktails targeting Escherichia coli and Enterococcus faecalis acted in a strain-specific manner. They transiently reduced the population density of their respective target before establishing coexistence for up to 9 days. Infection susceptibility to S. Tm was markedly increased at an early time point after challenge with both phage cocktails. Surprisingly, OMM14 mice were also susceptible 7 days after a single phage inoculation, when the targeted bacterial populations were back to pre-phage administration density. Concluding, our work shows that phages that dynamically modulate the density of protective members of the gut microbiota can provide opportunities for invasion of bacterial pathogens, in particular at early time points after phage application. This suggests, that phages targeting protective members of the microbiota may increase the risk for Salmonella infection.

Characterization of pneumococcal serotype 7F in vaccine conjugation

Streptococcus pneumoniae is a highly invasive bacterial pathogen that can cause a range of illnesses. Pneumococcal capsular polysaccharides (CPS) are the main virulence factors that causes invasive pneumococcal disease (IPD). Pneumococcal CPS serotype 7F along with a few other serotypes is more invasive and likely to cause IPD. Therefore, 7F is a target for pneumococcal vaccine development, and is included in the two recently approved multi-valent pneumococcal conjugated vaccines, i.e. VAXNEUVANCE and PREVNAR 20.To support process and development of our 15-valent pneumococcal conjugated vaccine (PCV15), chromatographic methods have been developed for 7F polysaccharide and conjugate characterization. A size-exclusion chromatography (SEC) method with UV, light scattering and refractive index detections was employed for concentration, size and conformation analysis. A reversed-phase ultra-performance liquid chromatography (RP-UPLC) method was used for analysis of conjugate monosaccharide composition and degree of conjugation. The collective information obtained by these chromatographic analysis provided insights into the pneumococcal conjugate and conjugation process.Graphical

Bacteremia Caused by a Serotype Ob5 Vibrio cholerae Strain in a Cirrhotic Patient in China.

The increasing incidence of non-O1/non-O139 Vibrio cholerae (NOVC) has been observed worldwide. However, septicemia caused by NOVC remains a rare condition that has received limited attention. Currently, there are no established treatment guidelines for bloodstream infections caused by NOVC, and the understanding of this condition mainly relies on individual case reports. Although NOVC bacteremia can be fatal in a small percentage of cases, knowledge about its microbiological features remains limited. Here, we present a case of V. cholerae septicemia caused by NOVC in a 46-year-old man with chronic viral hepatitis and liver cirrhosis. The isolated strain, named V. cholerae VCH20210731 and classified as a new sequence type (ST), ST1553, was found to be susceptible to most of the antimicrobial agents tested. O-antigen serotyping of V. cholerae VCH20210731 revealed that it belonged to serotype Ob5. Interestingly, the ctxAB genes, which are typically associated with V. cholerae, were absent in VCH20210731. However, the strain possessed 25 other potential virulence genes, such as hlyA, luxS, hap, and rtxA. The resistome of V. cholerae VCH20210731 included several genes, including qnrVC4, crp, almG, and parE. Nevertheless, susceptibility testing demonstrated that the isolate was susceptible to most of the antimicrobial agents tested. Phylogenetic analysis indicated that the closest strain to VCH20210731 was strain 120 from Russia, differing by 630 single-nucleotide polymorphisms (SNPs). Our findings contribute to the understanding of the genomic epidemiological characteristics and antibiotic resistance mechanisms of this invasive bacterial pathogen. IMPORTANCE This study highlights the discovery of a novel ST1553 V. cholerae strain in China, providing valuable insights into the genomic epidemiology and global transmission dynamics of V. cholerae. It is important to note that clinical presentations of NOVC bacteremia can vary significantly, and the isolates demonstrate genetic diversity. Consequently, health care professionals and public health experts should remain vigilant about the potential for infection with this pathogen, particularly considering the elevated prevalence of liver disease in China.

High-force catch bonds between the Staphylococcus aureus surface protein SdrE and complement regulator factor H drive immune evasion

The invasive bacterial pathogen Staphylococcus aureus recruits the complement regulatory protein factor H (fH) to its surface to evade the human immune system. Here, we report the identification of an extremely high-force catch bond used by the S. aureus surface protein SdrE to efficiently capture fH under mechanical stress. We find that increasing the external force applied to the SdrE-fH complex prolongs the lifetime of the bond at an extraordinary high force, 1,400 pN, above which the bond lifetime decreases as an ordinary slip bond. This catch-bond behavior originates from a variation of the dock, lock and latch interaction, where the SdrE ligand binding domains undergo conformational changes under stress, enabling the formation of long-lived hydrogen bonds with fH. The binding mechanism dissected here represents a potential target for new therapeutics against multidrug-resistant S. aureus strains.

1319. Evolution of Invasive and Colonizing Pediatric Staphylococcus aureus Isolates

Abstract Background Staphylococcus aureus is the most common invasive bacterial pathogen in children. The constant evolution of S. aureus is a barrier to the development of successful interventions. A detailed understanding of the current molecular epidemiology of actively circulating, invasive S. aureus strains and a comparison to both colonization strains and historical invasive strains to determine which virulence factors and genetic lineages are preserved over time are important unmet needs in this field. Methods We compared four groups of S. aureus isolates: 45 invasive strains obtained during the epidemic methicillin-resistance S. aureus (MRSA) peak (2010-2012); 74 invasive contemporary strains (2015-2021); 61 colonizing strains from 2004; and 30 colonizing strains from 2021. Genetic relatedness (via clonal complex typing) and virulence factor presence were determined via whole genome sequencing. Results Among invasive isolates, the prevalence of clonal complex 8 (CC8) decreased significantly over time from 87% to 46% among typeable strains (p< 0.05, Fig.1). Similarly, the presence of Panton-Valentine leucocidin (PVL) decreased from 71% to 40% (p< 0.01). 56% of invasive isolates from 2010-2012 were MRSA, vs 37% from 2015-2021 (p= 0.05, Fig. 2). In contrast to PVL, the presence of leukocidins LukED and LukAB remained stably prevalent over time at 84% and 100% (Fig. 3). Colonization isolates overall remained diverse, with no dominant clonal complex (Fig. 1). Figure 1Figure 2Figure 3 Conclusion Invasive S. aureus molecular epidemiology has changed significantly over the past decade. The former dominance of CC8 and PVL-positive MRSA strains has shifted to a more diverse, predominantly MSSA group of contemporary strains. Other virulence factors (LukED, LukAB) have remained prevalent among invasive strains regardless of genetic lineage. Targets of intervention should be those that are preserved throughout the evolution of this major human pathogen. Disclosures All Authors: No reported disclosures.

Is eating slow or fast better for children?

Eating behaviors are an important component of risk for childhood obesity. These behaviors, which include the frequency of eating, the time of day that food is ingested, whether or not breakfast is a regular meal, and whether or not children are watching television while eating. One aspect of eating behavior that has not been well studied is eating speed. In some studies, it has been suggested that eating quickly may be associated with higher energy intake and increased BMI. In this volume of The Journal, Garcidueñas-Fimbres et al assessed the relationships among eating speed, adiposity, diet quality, and cardiometabolic risk factors in preschool children. They found that faster eating is associated with greater adiposity, lower adherence to a high-quality diet, as well as higher systolic blood pressure and higher fasting plasma glucose. These results suggest the potential for a behavioral intervention focused on slower eating and increased chewing as a possible approach to prevention of obesity or as a component of treatment of obesity. Article page 31 Following Birth Hospitalization: Invasive Bacterial Infections in Preterm Infants Aged 7-90 DaysThe Journal of PediatricsVol. 252PreviewTo assess the incidence rate of invasive bacterial infections in preterm infants and compare invasive bacterial infection rates and pathogens between preterm and full-term infants at age 7-90 days. Full-Text PDF Clostridioides difficile Infection in Hospitalized Pediatric Patients: Comparisons of Epidemiology, Testing, and Treatment from 2013 to 2019The Journal of PediatricsVol. 252PreviewTo compare the incidence, epidemiology, testing patterns, treatment, and outcomes of Clostridioides difficile infection (CDI) among hospitalized pediatric patients from 2013 to 2019. Full-Text PDF Extended Risk of Mortality in Children with Inborn Errors of Metabolism: A Longitudinal Cohort StudyThe Journal of PediatricsVol. 252PreviewTo determine the long-term risk of mortality among children with inborn errors of metabolism. Full-Text PDF Associations Between Eating Speed, Diet Quality, Adiposity, and Cardiometabolic Risk FactorsThe Journal of PediatricsVol. 252PreviewTo assess the associations between eating speed, adiposity, cardiometabolic risk factors, and diet quality in a cohort of Spanish preschool-children. Full-Text PDF Open AccessFebrile Preterm Infants: They are Not Just Small Febrile, Term InfantsThe Journal of PediatricsVol. 252PreviewAnnually, more than 200 000 febrile infants younger than 90 days are evaluated in US emergency departments and other outpatient settings.1 Although urinary tract infections are common in these infants, the main concerns involve invasive bacterial infections, defined by bacteremia or bacterial meningitis. The reported invasive bacterial infection rates range from approximately 1% to 5% in prospective studies, with higher rates in the youngest infants.2-4 To address the persistent variability in care of these febrile infants despite substantial research performed over several decades, the American Academy of Pediatrics published guidelines for the management of well-appearing febrile infants 8-60 days old. Full-Text PDF

Following Birth Hospitalization: Invasive Bacterial Infections in Preterm Infants Aged 7-90 Days

To assess the incidence rate of invasive bacterial infections in preterm infants and compare invasive bacterial infection rates and pathogens between preterm and full-term infants at age 7-90days. This is a retrospective cohort study of the incidence rate of invasive bacterial infections among all infants born at Kaiser Permanente Northern California (KPNC), with blood and cerebrospinal fluid cultures collected between 7 and 90days of chronological age from outpatient clinics, from emergency departments, and in the first 24hours of hospitalization presenting for care between January 1, 2005, and December 31, 2017. Incidence rates of invasive bacterial infection by chronological age and postmenstrual age (PMA) and pathogens were compared between preterm and full-term infants. Between January 1, 2005, and December 31, 2017, a total of 479 729 infants were born at KPNC, including 440 070 full-term infants and 39 659 preterm infants. There were 283 cases of bacteremia in 282 infants. The incidence rate of invasive bacterial infection was significantly higher for preterm infants compared with full-term infants. The highest incidence rates of invasive bacterial infection were in preterm infants at chronological age 7-28days and/or 37-39weeks PMA. There was a trend toward lower rates of invasive bacterial infection with increasing PMA in preterm infants aged 61-90days. Preterm infants aged 29-60days or at ≥40weeks PMA and those aged 61-90days or at ≥43weeks PMA had a rate of invasive bacterial infection equivalent to the overall rate seen in full-term infants of the same chronological age group. The distribution of pathogens causing bacteremia and meningitis did not differ between preterm and full-term infants. PMA and chronological age together were more useful than either alone in informing the incidence rate of invasive bacterial infection in preterm infants during the first 90days of life.

Iron Deficiency And Overload Attenuate Inflammatory Response and Modulate Metabolism in Alveolar Macrophage in Vitro

ObjectiveThe study aimed to characterize immunometabolic signatures of porcine alveolar macrophage (PAM) associated with cellular iron deficiency (ID) and iron overload (IE). We hypothesized that cellular iron imbalance modulates LPS‐induced inflammatory and metabolic responses in PAM cells.MethodsAlveolar macrophage cells were collected from 6‐week old donor piglets (N= 5‐6) and were cultured in complete media at 37 ˚C with 5% CO2 until attachment. Cells were treated with complete media (control, CON) or complete media supplemented with 500 µM deferiprone (DFP) or 200 µM ferric ammonium citrate (FAC) to induce iron deficiency (ID) or iron excess (IE), respectively. At the end of the 36‐hour treatment, cells were challenged with 100 ng/mL LPS (L) or sterile saline (S) for another 6 hours. Cells were tested for viability using XTT assay or harvested for analysis of gene, protein expression, and untargeted metabolome using RT‐qPCR, western blot, and GC time‐of‐flight MS, respectively. Cell culture media were collected for analysis of inflammatory cytokines (TNFα, IL‐6, and IL‐10) using ELISA kits. Data were analyzed for two‐way ANOVA using PROC MIXED of SAS (v.9.4). Statistical significance was declared when P‐value or FDR is less than 0.05 or 0.1, respectively.ResultsLPS challenge reduced cell viability by 10 – 20% regardless of the cellular iron status (P < 0.001). Ferritin H was markedly increased in IE cells compared to CON or ID cells (P < 0.001), suggesting cellular iron overload. The mRNA expression of transferrin receptor 1 was the highest in ID‐S cells and was significantly higher in ID and CON compared to IE cells, irrespective of the LPS challenge (P < 0.001). LPS significantly increased mRNA expression of divalent metal transporter 1 (DMT1)and zinc transporter (ZIP14) by 10‐300 fold (P< 0.001), while iron treatment had no significant effect. Unsurprisingly, LPS drastically induced (P < 0.001) genes encoding pro‐ (TNFA and IL1B) and anti‐(TGFB1 and IL10) inflammatory cytokines. Unexpectedly, both ID and IE diminished the induction of those genes caused by the LPS challenge (P < 0.05). The response of TNFA gene was mirrored by changes in TNFα concentration in culture media. Twenty‐nine, 14, and 5 metabolites were altered by cellular iron status, LPS challenge, and their interaction (FDR < 0.1), respectively. A total of 28 altered metabolites were mapped in the user database. Interestingly, metabolites involved in the glycolysis and TCA cycle (e.g. glucose‐6‐phosphate, citric acid, and aconitic acid) were most abundant in ID cells, whereas, in comparison with CON, both ID and IE decreased several amino acids in PAM (e.g. glycine, alanine, leucine, and lysine). LPS increased metabolites involved in glycolysis (fructose‐1‐phosphate) and pentose phosphate pathway (ribose‐5‐phosphate and fructose‐6‐phosphate). Cellular iron status modulated changes in itaconic acid, glucose, fumaric acid, and putrescine in response to LPS challenge; ID enhanced the LPS‐induced increase in itaconic acid, a metabolite marker of macrophage activation.ConclusionPresence of iron chelator and iron overexposure altered cellular iron metabolism of PAM cells and its inflammatory and metabolic responses to the invasion of bacterial pathogens.

Targeting the Gut Microbiota in Coronavirus Disease 2019: Hype or Hope?

Targeting the Gut Microbiota in Coronavirus Disease 2019: Hype or Hope?

The Global Landscape of Pediatric Bacterial Meningitis Data Reported to the World Health Organization-Coordinated Invasive Bacterial Vaccine-Preventable Disease Surveillance Network, 2014-2019.

BackgroundThe World Health Organization (WHO) coordinates the Global Invasive Bacterial Vaccine-Preventable Diseases (IB-VPD) Surveillance Network to support vaccine introduction decisions and use. The network was established to strengthen surveillance and laboratory confirmation of meningitis caused by Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis.MethodsSentinel hospitals report cases of children <5 years of age hospitalized for suspected meningitis. Laboratories report confirmatory testing results and strain characterization tested by polymerase chain reaction. In 2019, the network included 123 laboratories that follow validated, standardized testing and reporting strategies.ResultsFrom 2014 through 2019, >137 000 suspected meningitis cases were reported by 58 participating countries, with 44.6% (n = 61 386) reported from countries in the WHO African Region. More than half (56.6%, n = 77 873) were among children <1 year of age, and 4.0% (n = 4010) died among those with reported disease outcome. Among suspected meningitis cases, 8.6% (n = 11 798) were classified as probable bacterial meningitis. One of 3 bacterial pathogens was identified in 30.3% (n = 3576) of these cases, namely S. pneumoniae (n = 2177 [60.9%]), H. influenzae (n = 633 [17.7%]), and N. meningitidis (n = 766 [21.4%]). Among confirmed bacterial meningitis cases with outcome reported, 11.0% died; case fatality ratio varied by pathogen (S. pneumoniae, 12.2%; H. influenzae, 6.1%; N. meningitidis, 11.0%). Among the 277 children who died with confirmed bacterial meningitis, 189 (68.2%) had confirmed S. pneumoniae. The proportion of pneumococcal cases with pneumococcal conjugate vaccine (PCV) serotypes decreased as the number of countries implementing PCV increased, from 77.8% (n = 273) to 47.5% (n = 248). Of 397 H. influenzae specimens serotyped, 49.1% (n = 195) were type b. Predominant N. meningitidis serogroups varied by region.ConclusionsThis multitier, global surveillance network has supported countries in detecting and serotyping the 3 principal invasive bacterial pathogens that cause pediatric meningitis. Streptococcus pneumoniae was the most common bacterial pathogen detected globally despite the growing number of countries that have nationally introduced PCV. The large proportions of deaths due to S. pneumoniae reflect the high proportion of meningitis cases caused by this pathogen. This global network demonstrated a strong correlation between PCV introduction status and reduction in the proportion of pneumococcal meningitis infections caused by vaccine serotypes. Maintaining case-based, active surveillance with laboratory confirmation for prioritized vaccine-preventable diseases remains a critical component of the global agenda in public health.The World Health Organization (WHO)-coordinated Invasive Bacterial Vaccine-Preventable Disease (IB-VPD) Surveillance Network reported data from 2014 to 2019, contributing to the estimates of the disease burden and serotypes of pediatric meningitis caused by Streptococcus pneumoniae, Haemophilus influenzae and Neisseria meningitidis.

Staphylococcus aureus infections in children.

Staphylococcus aureus is the most common invasive bacterial pathogen infecting children in the U.S. and many parts of the world. This major human pathogen continues to evolve, and recognition of recent trends in epidemiology, therapeutics and future horizons is of high importance. Over the past decade, a relative rise of methicillin-susceptible S. aureus (MSSA) has occurred, such that methicillin-resistant S. aureus (MRSA) no longer dominates the landscape of invasive disease. Antimicrobial resistance continues to develop, however, and novel therapeutics or preventive modalities are urgently needed. Unfortunately, several recent vaccine attempts proved unsuccessful in humans. Recent scientific breakthroughs highlight the opportunity for novel interventions against S. aureus by interfering with virulence rather than by traditional antimicrobial mechanisms. A S. aureus vaccine remains elusive; the reasons for this are multifactorial, and lessons learned from prior unsuccessful attempts may create a path toward an effective preventive. Finally, new diagnostic modalities have the potential to greatly enhance clinical care for invasive S. aureus disease in children.

#31: Children with Invasive S. aureus Infection Produce Broadly Neutralizing Antibodies Against Distantly Related Variants of the Cytotoxin LukAB

Abstract Background Staphylococcus aureus is the most common invasive bacterial pathogen in children, and novel targets of intervention are urgently needed. The two-component leukotoxin, LukAB, is critical for S. aureus targeting and killing of human neutrophils and is abundantly produced in the setting of invasive human infection. LukAB is unique among S. aureus cytotoxins in that it exists in variant form across distantly related strains of clinically relevant S. aureus. The broad diversity of circulating clinical S. aureus strains must be taken into account to successfully develop new anti-staphylococcal preventives or therapeutics. This diversity and ongoing S. aureus evolution may explain previous unsuccessful attempts to intervene against S. aureus at the virulence factor level. We have previously shown that LukAB is ubiquitous among clinical isolates and that children with invasive disease mount a high-titer neutralizing response, but the breadth of function of this response has not previously been explored. Methods B-cells were isolated from children with invasive S. aureus infections (e.g. bacteremia or acute hematogenous osteomyelitis). Following EBV-transformation, cell supernatants were screened for LukAB binding and selected for generation of monoclonal hybridomas. MAbs were assessed for binding by ELISA and neutralizing function by in vitro cytotoxicity assays, where neutrophil-like HL-60 cells were cultured in the presence of human mAbs and diverse allelic variants of LukAB. The infecting S. aureus isolates were typed using multilocus sequence typing (MLST) to determine clonal complexes. Results 34 distinct human anti-LukAB mAbs were generated from 3 children with invasive S. aureus disease. Of these, 22% were isolated following infection with a strain belonging to clonal complex 8 (CC8), consistent with the epidemic USA300 clone, and 78% were generated against CC5 strains. Within this panel, all mAbs potently neutralized LukAB from the same clonal complex as the corresponding infecting isolate, but 23 also demonstrated neutralization against other allelic variants of LukAB. 7 mAbs were capable of broad, potent neutralization against LukAB variants from all clinically relevant clonal complexes tested (CC8, CC30, CC45, CC75, CC1, CC5, and CC398). Conclusions A subset of human mAbs isolated from children with invasive S. aureus disease were capable of broad neutralization against distantly related variants of the important toxin LukAB. This has two important implications: First, we found strong evidence of a conserved target (or targets) for antibody-mediated toxin neutralization across diverse strains of S. aureus. Second, this provides additional support for this toxin as a target of intervention, as some previous vaccine attempts were likely unsuccessful due to activity against a narrow subset of circulating S. aureus strains.

Inhibitory Concentrations of Ciprofloxacin Induce an Adaptive Response Promoting the Intracellular Survival of Salmonella enterica Serovar Typhimurium.

ABSTRACTAntimicrobial resistance (AMR) is a pressing global health crisis, which has been fueled by the sustained use of certain classes of antimicrobials, including fluoroquinolones. While the genetic mutations responsible for decreased fluoroquinolone (ciprofloxacin) susceptibility are known, the implications of ciprofloxacin exposure on bacterial growth, survival, and interactions with host cells are not well described. Aiming to understand the influence of inhibitory concentrations of ciprofloxacin in vitro, we subjected three clinical isolates of Salmonella enterica serovar Typhimurium to differing concentrations of ciprofloxacin, dependent on their MICs, and assessed the impact on bacterial growth, morphology, and transcription. We further investigated the differential morphology and transcription that occurred following ciprofloxacin exposure and measured the ability of ciprofloxacin-treated bacteria to invade and replicate in host cells. We found that ciprofloxacin-exposed S. Typhimurium is able to recover from inhibitory concentrations of ciprofloxacin and that the drug induces specific morphological and transcriptional signatures associated with the bacterial SOS response, DNA repair, and intracellular survival. In addition, ciprofloxacin-treated S. Typhimurium has increased capacity for intracellular replication in comparison to that of untreated organisms. These data suggest that S. Typhimurium undergoes an adaptive response under ciprofloxacin perturbation that promotes cellular survival, a consequence that may justify more measured use of ciprofloxacin for Salmonella infections. The combination of multiple experimental approaches provides new insights into the collateral effects that ciprofloxacin and other antimicrobials have on invasive bacterial pathogens.

Pattern and causes of the establishment of the invasive bacterial potato pathogen Dickeya solani and of the maintenance of the resident pathogen D.dianthicola.

Invasive pathogens can be a threat when they affect human health, food production or ecosystem services, by displacing resident species, and we need to understand the cause of their establishment. We studied the patterns and causes of the establishment of the pathogen Dickeya solani that recently invaded potato agrosystems in Europe by assessing its invasion dynamics and its competitive ability against the closely related resident D. dianthicola species. Epidemiological records over one decade in France revealed the establishment of D. solani and the maintenance of the resident D. dianthicola in potato fields exhibiting blackleg symptoms. Using experimentations, we showed that D. dianthicola caused a higher symptom incidence on aerial parts of potato plants than D. solani, while D. solani was more aggressive on tubers (i.e. with more severe symptoms). In co-infection assays, D. dianthicola outcompeted D. solani in aerial parts, while the two species co-existed in tubers. A comparison of 76 D. solani genomes (56 of which have been sequenced here) revealed balanced frequencies of two previously uncharacterized alleles, VfmBPro and VfmBSer , at the vfmB virulence gene. Experimental inoculations showed that the VfmBSer population was more aggressive on tubers, while the VfmBPro population outcompeted the VfmBSer population in stem lesions, suggesting an important role of the vfmB virulence gene in the ecology of the pathogens. This study thus brings novel insights allowing a better understanding of the pattern and causes of the D.solani invasion into potato production agrosystems, and the reasons why the endemic D. dianthicola nevertheless persisted.

Distribution of PDLIM1 at actin-rich structures generated by invasive and adherent bacterial pathogens.

The enteric bacterial pathogens Listeria monocytogenes (Listeria) and enteropathogenic Escherichia coli (EPEC) remodel the eukaryotic actin cytoskeleton during their disease processes. Listeria generate slender actin-rich comet/rocket tails to move intracellularly, and later, finger-like membrane protrusions to spread amongst host cells. EPEC remain extracellular, but generate similar actin-rich membranous protrusions (termed pedestals) to move atop the host epithelia. These structures are crucial for disease as diarrheal (and systemic) infections are significantly abrogated during infections with mutant strains that are unable to generate the structures. The current repertoire of host components enriched within these structures is vast and diverse. In this protein catalog, we and others have found that host actin crosslinkers, such as palladin and α-actinin-1, are routinely exploited. To expand on this list, we set out to investigate the distribution of PDLIM1, a scaffolding protein and binding partner of palladin and α-actinin-1, during bacterial infections. We show that PDLIM1 localizes to the site of initial Listeria entry into cells. Following this, PDLIM1 localizes to actin filament clouds surrounding immotile bacteria, and then colocalizes with actin once the comet/rocket tails are generated. Unlike palladin or α-actinin-1, PDLIM1 is maintained within the actin-rich core of membrane protrusions. Conversely, α-actinin-1, but not PDLIM1 (or palladin), is enriched at the membrane invagination that internalizes the Listeria-containing membrane protrusion. We also show that PDLIM1 is a component of the EPEC pedestal core and that its recruitment is dependent on the bacterial effector Tir. Our findings highlight PDLIM1 as another protein present within pathogen-induced actin-rich structures.

Perspective of a Pediatrician: Shared Pathogenesis of the Three Most Successful Pathogens of Children.

Highly successful invasive pathogens exploit host vulnerabilities by adapting tools to co-opt highly conserved host features. This is especially true when pathogens develop ligands to hijack trafficking routes or signaling patterns of host receptors. In this context, highly successful pathogens can be grouped together by the patterns of organs infected and diseases they cause. In the case of this perspective, the focus is on the historically most successful invasive bacterial pathogens of children that cause pneumonia, sepsis and meningitis: Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis. This triad shares a ligand to bind to PAF receptor to enter host cells despite early defenses by innate immunity. All three also target laminin receptor to cross endothelial barriers using a common set of molecular tools that may prove to be a design for a cross-protective vaccine.

Sublethal Levels of Antibiotics Promote Bacterial Persistence in Epithelial Cells.

Antibiotic therapy and host cells frequently fail to eliminate invasive bacterial pathogens due to the emergence of antibiotic resistance, resulting in the relapse and recurrence of infections. Bacteria evolve various strategies to persist and survive in epithelial cells, a front‐line barrier of host tissues counteracting invasion; however, it remains unclear how bacteria hijack cellular responses to promote cytoplasmic survival under antibiotic therapy. Here, it is demonstrated that extracellular bacteria show invasive behavior and survive in epithelial cells in both in vivo and in vitro models, to increase antibiotic tolerance. In turn, sublethal levels of antibiotics increase bacterial invasion through promoting the production of bacterial virulence factors. Furthermore, antibiotic treatments interrupt lysosomal acidification in autophagy due to the internalized bacteria, using Bacillus cereus and ciprofloxacin as a model. In addition, it is found that sublethal levels of ciprofloxacin cause mitochondrial dysfunction and reactive oxygen species (ROS) accumulation to impair lysosomal vascular tape ATPase (V‐ATPase) to further promote bacterial persistence. Collectively, these results highlight the potential of host cells mediated antibiotic tolerance, which markedly compromises antibiotic efficacy and worsens the outcomes of infection.