ResearchPad - superinfection https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[The influenza replication blocking inhibitor LASAG does not sensitize human epithelial cells for bacterial infections]]> https://www.researchpad.co/article/elastic_article_14740 Severe influenza virus (IV) infections still represent a major challenge to public health. To combat IV infections, vaccines and antiviral compounds are available. However, vaccine efficacies vary with very limited to no protection against newly emerging zoonotic IV introductions. In addition, the development of resistant virus variants against currently available antivirals can be rapidly detected, in consequence demanding the design of novel antiviral strategies. Virus supportive cellular signaling cascades, such as the NF-κB pathway, have been identified to be promising antiviral targets against IV in in vitro and in vivo studies and clinical trials. While administration of NF-κB pathway inhibiting agents, such as LASAG results in decreased IV replication, it remained unclear whether blocking of NF-κB might sensitize cells to secondary bacterial infections, which often come along with viral infections. Thus, we examined IV and Staphylococcus aureus growth during LASAG treatment. Interestingly, our data reveal that the presence of LASAG during superinfection still leads to reduced IV titers. Furthermore, the inhibition of the NF-κB pathway resulted in decreased intracellular Staphylococcus aureus loads within epithelial cells, indicating a dependency on the pathway for bacterial uptake. Unfortunately, so far it is not entirely clear if this phenomenon might be a drawback in bacterial clearance during infection.

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<![CDATA[Contrasting antibody responses to intrasubtype superinfection with CRF02_AG]]> https://www.researchpad.co/article/5989db50ab0ee8fa60bdbdea

HIV superinfection describes the sequential infection of an individual with two or more unrelated HIV strains. Intersubtype superinfection has been shown to cause a broader and more potent heterologous neutralizing antibody response when compared to singly infected controls, yet the effects of intrasubtype superinfection remain controversial. Longitudinal samples were analyzed phylogenetically for pol and env regions using Next-Generation Sequencing and envelope cloning. The impact of CRF02_AG intrasubtype superinfection was assessed for heterologous neutralization and antibody binding responses. We compared two cases of CRF02_AG intrasubtype superinfection that revealed complete replacement of the initial virus by superinfecting CRF02_AG variants with signs of recombination. NYU6564, who became superinfected at an early time point, exhibited greater changes in antibody binding profiles and generated a more potent neutralizing antibody response post-superinfection compared to NYU6501. In contrast, superinfection occurred at a later time point in NYU6501 with strains harboring significantly longer V1V2 regions with no observable changes in neutralization patterns. Here we show that CRF02_AG intrasubtype superinfection can induce a cross-subtype neutralizing antibody response, and our data suggest timing and/or superinfecting viral envelope characteristics as contributing factors. These results highlight differential outcomes in intrasubtype superinfection and provide the first insight into cases with CRF02_AG, the fourth most prevalent HIV-1 strain worldwide.

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<![CDATA[Classical Swine Fever Virus vs. Classical Swine Fever Virus: The Superinfection Exclusion Phenomenon in Experimentally Infected Wild Boar]]> https://www.researchpad.co/article/5989db43ab0ee8fa60bd78a6

Two groups with three wild boars each were used: Group A (animals 1 to 3) served as the control, and Group B (animals 4 to 6) was postnatally persistently infected with the Cat01 strain of CSFV (primary virus). The animals, six weeks old and clinically healthy, were inoculated with the virulent strain Margarita (secondary virus). For exclusive detection of the Margarita strain, a specific qRT-PCR assay was designed, which proved not to have cross-reactivity with the Cat01 strain. The wild boars persistently infected with CSFV were protected from superinfection by the virulent CSFV Margarita strain, as evidenced by the absence of clinical signs and the absence of Margarita RNA detection in serum, swabs and tissue samples. Additionally, in PBMCs, a well-known target for CSFV viral replication, only the primary infecting virus RNA (Cat01 strain) could be detected, even after the isolation in ST cells, demonstrating SIE at the tissue level in vivo. Furthermore, the data analysis of the Margarita qRT-PCR, by means of calculated ΔCt values, supported that PBMCs from persistently infected animals were substantially protected from superinfection after in vitro inoculation with the Margarita virus strain, while this virus was able to infect naive PBMCs efficiently. In parallel, IFN-α values were undetectable in the sera from animals in Group B after inoculation with the CSFV Margarita strain. Furthermore, these animals were unable to elicit adaptive humoral (no E2-specific or neutralising antibodies) or cellular immune responses (in terms of IFN-γ-producing cells) after inoculation with the second virus. Finally, a sequence analysis could not detect CSFV Margarita RNA in the samples tested from Group B. Our results suggested that the SIE phenomenon might be involved in the evolution and phylogeny of the virus, as well as in CSFV control by vaccination. To the best of our knowledge, this study was one of the first showing efficient suppression of superinfection in animals, especially in the absence of IFN-α, which might be associated with the lack of innate immune mechanisms.

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<![CDATA[Establishment of a Wolbachia Superinfection in Aedes aegypti Mosquitoes as a Potential Approach for Future Resistance Management]]> https://www.researchpad.co/article/5989db10ab0ee8fa60bcbece

Wolbachia pipientis is an endosymbiotic bacterium estimated to chronically infect between 40–75% of all arthropod species. Aedes aegypti, the principle mosquito vector of dengue virus (DENV), is not a natural host of Wolbachia. The transinfection of Wolbachia strains such as wAlbB, wMel and wMelPop-CLA into Ae. aegypti has been shown to significantly reduce the vector competence of this mosquito for a range of human pathogens in the laboratory. This has led to wMel-transinfected Ae. aegypti currently being released in five countries to evaluate its effectiveness to control dengue disease in human populations. Here we describe the generation of a superinfected Ae. aegypti mosquito line simultaneously infected with two avirulent Wolbachia strains, wMel and wAlbB. The line carries a high overall Wolbachia density and tissue localisation of the individual strains is very similar to each respective single infected parental line. The superinfected line induces unidirectional cytoplasmic incompatibility (CI) when crossed to each single infected parental line, suggesting that the superinfection would have the capacity to replace either of the single constituent infections already present in a mosquito population. No significant differences in fitness parameters were observed between the superinfected line and the parental lines under the experimental conditions tested. Finally, the superinfected line blocks DENV replication more efficiently than the single wMel strain when challenged with blood meals from viremic dengue patients. These results suggest that the deployment of superinfections could be used to replace single infections and may represent an effective strategy to help manage potential resistance by DENV to field deployments of single infected strains.

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<![CDATA[Bacteremia in Children Hospitalized with Respiratory Syncytial Virus Infection]]> https://www.researchpad.co/article/5989da7aab0ee8fa60b980c9

Background

The risk of bacteremia is considered low in children with acute bronchiolitis. However the rate of occult bacteremia in infants with RSV infection is not well established. The aim was to determine the actual rate and predictive factors of bacteremia in children admitted to hospital due to confirmed RSV acute respiratory illness (ARI), using both conventional culture and molecular techniques.

Methods

A prospective multicenter study (GENDRES-network) was conducted between 2011–2013 in children under the age of two admitted to hospital because of an ARI. Among those RSV-positive, bacterial presence in blood was assessed using PCR for Meningococcus, Streptococcus pneumoniae, Haemophilus influenzae, Streptococcus pyogenes, Klebsiella pneumoniae, Pseudomonas aeruginosa, Escherichia coli, and Staphylococcus aureus, in addition to conventional cultures.

Results

66 children with positive RSV respiratory illness were included. In 10.6% patients, bacterial presence was detected: H. influenzae (n = 4) and S. pneumoniae (n = 2). In those patients with bacteremia, there was a previous suspicion of bacterial superinfection and had received empirical antibiotic treatment 6 out of 7 (85.7%) patients. There were significant differences in terms of severity between children with positive bacterial PCR and those with negative results: PICU admission (100% vs. 50%, P-value = 0.015); respiratory support necessity (100% vs. 18.6%, P-value < 0.001); Wood-Downes score (mean = 8.7 vs. 4.8 points, P-value < 0.001); GENVIP scale (mean = 17 vs. 10.1, P-value < 0.001); and length of hospitalization (mean = 12.1 vs. 7.5 days, P-value = 0.007).

Conclusion

Bacteremia is not frequent in infants hospitalized with RSV respiratory infection, however, it should be considered in the most severe cases.

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<![CDATA[A Novel Controlled-Release System for Antibacterial Enzyme Lysostaphin Delivery Using Hydroxyapatite/Chitosan Composite Bone Cement]]> https://www.researchpad.co/article/5989da65ab0ee8fa60b91bab

In this work, a lysostaphin-loaded, control-released, self-setting and injectable porous bone cement with efficient protein delivery was prepared by a novel setting method using hydroxyapatite/chitosan (HA/CS) composite scaffold. The cement samples were made through cementitious reactions by mixing solid powder, a mixture of HA/CS composite particles, lysostaphin, Ca(OH)2, CaCO3 and NaHCO3, with setting liquid containing citric acid, acetic acid, NaH2PO4, CaCl2 and poloxamer. The setting parameters of the cement samples were determined. The results showed that the final setting time was 96.6±5.2 min and the pH value increased from approximately 6.2 to nearly 10 during the setting process and the porosity was 34% at the end. And the microstructure and composition were detected by scanning electron microscopy (SEM), x-ray diffraction and Fourier transform-infrared spectroscopy. For the release behavior of lysostaphin loaded in the cement sample, the in vitro cement extract experiment indicated that about 94.2±10.9% of the loaded protein was released before day 8 and the in vivo Qdot 625 fluorescence tracking experiment showed that the loaded protein released slower than the free one. Then the biocompatibility of the cement samples was evaluated using the methylthiazol tetrazolium assay, SEM and hematoxylin-eosin staining, which suggested good biocompatibility of cement samples with MC 3T3-E1 cells and subcutaneous tissues of mice. Finally the antibacterial activity assay indicated that the loaded lysostaphin had good release ability and strong antibacterial enzymatic activity against methicillin-resistant Staphylococcus aureus. Collectively, all the results suggested that the lysostaphin-loaded self-setting injectable porous bone cement released the protein in a controlled and effective way and the protein activity was well retained during the setting and releasing process. Thus this bone cement can be potentially applied as a combination of artificial bone substitute and controlled-release system for delivery of lysostaphin to treat bone defects and infections.

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<![CDATA[Uropathogenic Escherichia coli Superinfection Enhances the Severity of Mouse Bladder Infection]]> https://www.researchpad.co/article/5989da97ab0ee8fa60ba2691

Urinary tract infections (UTIs) afflict over 9 million women in America every year, often necessitating long-term prophylactic antibiotics. One risk factor for UTI is frequent sexual intercourse, which dramatically increases the risk of UTI. The mechanism behind this increased risk is unknown; however, bacteriuria increases immediately after sexual intercourse episodes, suggesting that physical manipulation introduces periurethral flora into the urinary tract. In this paper, we investigated whether superinfection (repeat introduction of bacteria) resulted in increased risk of severe UTI, manifesting as persistent bacteriuria, high titer bladder bacterial burdens and chronic inflammation, an outcome referred to as chronic cystitis. Chronic cystitis represents unchecked luminal bacterial replication and is defined histologically by urothelial hyperplasia and submucosal lymphoid aggregates, a histological pattern similar to that seen in humans suffering chronic UTI. C57BL/6J mice are resistant to chronic cystitis after a single infection; however, they developed persistent bacteriuria and chronic cystitis when superinfected 24 hours apart. Elevated levels of interleukin-6 (IL-6), keratinocyte cytokine (KC/CXCL1), and granulocyte colony-stimulating factor (G-CSF) in the serum of C57BL/6J mice prior to the second infection predicted the development of chronic cystitis. These same cytokines have been found to precede chronic cystitis in singly infected C3H/HeN mice. Furthermore, inoculating C3H/HeN mice twice within a six-hour period doubled the proportion of mice that developed chronic cystitis. Intracellular bacterial replication, regulated hemolysin (HlyA) expression, and caspase 1/11 activation were essential for this increase. Microarrays conducted at four weeks post inoculation in both mouse strains revealed upregulation of IL-1 and antimicrobial peptides during chronic cystitis. These data suggest a mechanism by which caspase-1/11 activation and IL-1 secretion could predispose certain women to recurrent UTI after frequent intercourse, a predisposition predictable by several serum biomarkers in two murine models.

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<![CDATA[Intestinal Translocation of Clinical Isolates of Vancomycin-Resistant Enterococcus faecalis and ESBL-Producing Escherichia coli in a Rat Model of Bacterial Colonization and Liver Ischemia/Reperfusion Injury]]> https://www.researchpad.co/article/5989daf5ab0ee8fa60bc2c20

The objectives of this study were to develop a rat model of gastrointestinal colonization with vancomycin-resistant Enterococcus faecalis (VRE) and extended-spectrum beta-lactamase (ESBL)-producing E. coli and to evaluate intestinal translocation to blood and tissues after total and partial hepatic ischemia. Methods - We developed a model of rat colonization with VRE and ESBL-E coli. Then we studied four groups of colonized rats: Group I (with hepatic pedicle occlusion causing complete liver ischemia and intestinal stasis); Group II (with partial liver ischemia without intestinal stasis); Group III (surgical manipulation without hepatic ischemia or intestinal stasis); Group IV (anesthetized without surgical manipulation). After sacrifice, portal and systemic blood, large intestine, small intestine, spleen, liver, lungs, and cervical and mesenteric lymph nodes were cultured. Endotoxin concentrations in portal and systemic blood were determined. Results – The best inocula were: VRE: 2.4×1010 cfu and ESBL-E. coli: 1.12×1010 cfu. The best results occurred 24 hours after inoculation and antibiotic doses of 750 µg/mL of water for vancomycin and 2.1 mg/mL for ceftriaxone. There was a significantly higher proportion of positive cultures for ESBL-E. coli in the lungs in Groups I, II and III when compared with Group IV (67%; 60%; 75% and 13%, respectively; p:0.04). VRE growth was more frequent in mesenteric lymph nodes for Groups I (67%) and III (38%) than for Groups II (13%) and IV (none) (p:0.002). LPS was significantly higher in systemic blood of Group I (9.761±13.804 EU/mL−p:0.01). No differences for endotoxin occurred in portal blood. Conclusion –We developed a model of rats colonized with resistant bacteria useful to study intestinal translocation. Translocation occurred in surgical procedures with and without hepatic ischemia-reperfusion and probably occurred via the bloodstream. Translocation was probably lymphatic in the ischemia-reperfusion groups. Systemic blood endotoxin levels were higher in the group with complete hepatic ischemia.

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<![CDATA[Does Viral Co-Infection Influence the Severity of Acute Respiratory Infection in Children?]]> https://www.researchpad.co/article/5989d9fcab0ee8fa60b72425

Background

Multiple viruses are often detected in children with respiratory infection but the significance of co-infection in pathogenesis, severity and outcome is unclear.

Objectives

To correlate the presence of viral co-infection with clinical phenotype in children admitted with acute respiratory infections (ARI).

Methods

We collected detailed clinical information on severity for children admitted with ARI as part of a Spanish prospective multicenter study (GENDRES network) between 2011–2013. A nested polymerase chain reaction (PCR) approach was used to detect respiratory viruses in respiratory secretions. Findings were compared to an independent cohort collected in the UK.

Results

204 children were recruited in the main cohort and 97 in the replication cohort. The number of detected viruses did not correlate with any markers of severity. However, bacterial superinfection was associated with increased severity (OR: 4.356; P-value = 0.005), PICU admission (OR: 3.342; P-value = 0.006), higher clinical score (1.988; P-value = 0.002) respiratory support requirement (OR: 7.484; P-value < 0.001) and longer hospital length of stay (OR: 1.468; P-value < 0.001). In addition, pneumococcal vaccination was found to be a protective factor in terms of degree of respiratory distress (OR: 2.917; P-value = 0.035), PICU admission (OR: 0.301; P-value = 0.011), lower clinical score (-1.499; P-value = 0.021) respiratory support requirement (OR: 0.324; P-value = 0.016) and oxygen necessity (OR: 0.328; P-value = 0.001). All these findings were replicated in the UK cohort.

Conclusion

The presence of more than one virus in hospitalized children with ARI is very frequent but it does not seem to have a major clinical impact in terms of severity. However bacterial superinfection increases the severity of the disease course. On the contrary, pneumococcal vaccination plays a protective role.

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<![CDATA[Role of Occult and Post-acute Phase Replication in Protective Immunity Induced with a Novel Live Attenuated SIV Vaccine]]> https://www.researchpad.co/article/5989dae4ab0ee8fa60bbccb4

In order to evaluate the role of persisting virus replication during occult phase immunisation in the live attenuated SIV vaccine model, a novel SIVmac239Δnef variant (SIVrtTA) genetically engineered to replicate in the presence of doxycycline was evaluated for its ability to protect against wild-type SIVmac239. Indian rhesus macaques were vaccinated either with SIVrtTA or with SIVmac239Δnef. Doxycycline was withdrawn from 4 of 8 SIVrtTA vaccinates before challenge with wild-type virus. Unvaccinated challenge controls exhibited ~107 peak plasma viral RNA copies/ml persisting beyond the acute phase. Six vaccinates, four SIVmac239Δnef and two SIVrtTA vaccinates exhibited complete protection, defined by lack of wild-type viraemia post-challenge and virus-specific PCR analysis of tissues recovered post-mortem, whereas six SIVrtTA vaccinates were protected from high levels of viraemia. Critically, the complete protection in two SIVrtTA vaccinates was associated with enhanced SIVrtTA replication in the immediate post-acute vaccination period but was independent of doxycycline status at the time of challenge. Mutations were identified in the LTR promoter region and rtTA gene that do not affect doxycycline-control but were associated with enhanced post-acute phase replication in protected vaccinates. High frequencies of total circulating CD8+T effector memory cells and a higher total frequency of SIV-specific CD8+ mono and polyfunctional T cells on the day of wild-type challenge were associated with complete protection but these parameters were not predictive of outcome when assessed 130 days after challenge. Moreover, challenge virus-specific Nef CD8+ polyfunctional T cell responses and antigen were detected in tissues post mortem in completely-protected macaques indicating post-challenge control of infection. Within the parameters of the study design, on-going occult-phase replication may not be absolutely required for protective immunity.

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