SGP-C: A Broad Host Range Temperate Bacteriophage; Against Salmonella gallinarum

Salmonella gallinarum is a poultry restricted-pathogen causing fowl-typhoid disease in adult birds with mortality rates up-to 80% and exhibit resistance against commonly used antibiotics. In this current study, a temperate broad host range bacteriophage SGP-C was isolated against S. gallinarum from poultry digesta. It showed infection ability in all the 15 tested field strains of S. gallinarum. The SGP-C phage produced circular, turbid plaques with alternate rings. Its optimum activity was observed at pH 7.0 and 37–42°C, with a latent period of 45 min and burst size of 187 virions/bacterial cell. The SGP-C lysogens, SGPC-L5 and SGPC-L6 exhibited super-infection immunity against the same phage, an already reported feature of lysogens. A virulence index of 0.5 and 0.001 as MV50 of SGP-C suggests its moderate virulence. The genome of SGP-C found circular double stranded DNA of 42 Kbp with 50.04% GC content, which encodes 63 ORFs. The presence of repressor gene at ORF49, and absence of tRNA sequence in SGP-C genome indicates its lysogenic nature. Furthermore, from NGS analysis of lysogens we propose that SGP-C genome might exist either as an episome, or both as integrated and temporary episome in the host cell and warrants further studies. Phylogenetic analysis revealed its similarity with Salmonella temperate phages belonging to family Siphoviridae. The encoded proteins by SGP-C genome have not showed homology with any known toxin and virulence factor. Although plenty of lytic bacteriophages against this pathogen are already reported, to our knowledge SGP-C is the first lysogenic phage against S. gallinarum reported so far.

Intracranial Hemorrhage Associated with Therapeutic Anticoagulation in Three Critically Ill COVID-19 Patients: A Case Series

Introduction: The COVID-19 pandemic has challenged global health with novel pathogenesis that is both severe and poorly understood. Several mechanisms have been proposed to explain the severity and complexity of the clinical illness including cytokine storm release, thromboembolic microangiopathy, direct cytotoxicity, and post-viral bacterial super infection. Cases: The patients in this case series were all admitted to the intensive care unit with respiratory failure from COVID-19 requiring invasive mechanical ventilation. They were all started on anticoagulation. All three patients developed acute kidney injuries. The first patient had hypertensive emergency at the time of the bleed. The second and third patient both had supratherapeutic heparin levels at the time of the bleed. Methods: We followed patients aged 18 years and above who were admitted to the ICU for COVID-19 during April and May 2020. We then followed those who required therapeutic anticoagulation for any indication and evaluated the ones that developed ICH. Results: Out of the 79 patients admitted to the ICU for COVID-19 related illness during April and May 2020, 31 were placed on therapeutic anticoagulation (intermediate or full-dose) for indications that included hypercoagulable state, ACS, atrial fibrillation, and deep vein thrombosis. 25% of patients on anticoagulation developed bleeding for which the anticoagulation had to be stopped. Three out of these 31 patients developed ICH while on anticoagulation, accounting for 3.8% of our ICU population with COVID-19 Discussion: Activation of coagulation pathways during cytokine storms can result in systemic thromboembolism, in both venous and arterial circulations posing risk of ischemic infarctions to any organ. Supratherapeutic heparin levels and acute kidney injuries are common in COVID-19 patients. The ideal candidates for anticoagulation, the recommended agent and dose, and duration of treatment remain unclear. Conclusion: The benefits of anticoagulation should be weighed against the potential risk of bleeding.

308. Secondary Infections in Patients Requiring Extracorporeal Membrane Oxygenation (ECMO) for Severe Acute Respiratory Distress Syndrome (ARDS) due to COVID-19 Pneumonia (PNA)

Background Rescue ECMO has been used worldwide in patients (pts) with ARDS caused by COVID-19. Bacterial super-infections affect 3.5-14.3% of hospitalized pts with COVID-19. Pts requiring ECMO may be at an increased risk of infection due to their severity of illness, gut translocation and ECMO impact on host immunity. Methods This was a retrospective review of pts requiring ECMO for COVID-19 from April 2020-2021 at a single center. Strict definitions of infections (including ventilator-associated PNA, VAP) were in accordance with CDC criteria. Results 43 ECMO pts with 1065 ECMO days were evaluated. Median age was 53 yrs (range: 21-62) and median BMI was 36.2 (range: 19.4-75.8). 70% were men and 65% were white. 37 patients (86%) experienced a total of 40 infectious episodes with a median onset from ECMO cannulation to first infection of 10.5d (range: 4-50). Median SOFA and SAPSII scores at time of infection were 12 (6-20) and 63 (30-90), respectively. PNA was the most common infection (78%, with 19% of cases complicated by bacteremia and 3% by empyema) (Fig. 1). The most common organisms isolated were Enterobacterales (37%), S. aureus (25%) and P. aeruginosa (16%) (Fig. 2). Only 2% of all organisms were multi-drug resistant. 3 pts had fungal infections (1 candidemia, 2 aspergillus PNA). Duration of ECMO was significantly longer for infected pts (26d, range: 5-92d) vs (11d, range: 3-24d), p=.01. 95% of infected pts had received steroids vs. 67% of uninfected pts, p=0.09. Treatment success at 1 week was 50%, and 24% and 40% of pts had recurrent infections and persistent/recurrent organisms in clinical cultures, respectively. S. aureus (54%) and Enterobacterales (26%) were associated with persistent or recurrent clinical cultures, requiring prolonged antimicrobial therapy. Mortality rate at 30 days was 65% and was significantly higher for pts with infection than those without (67% vs 33%, p=.02). Conclusion Super-infection (most commonly PNA) occurred in almost all COVID-19 pts requiring ECMO for >4 days, and was a significant risk factor for death. Recurrent infections among survivors were common, especially when caused by Enterbacterales or S. aureus. Super-infection and mortality rates of ARDS pts on ECMO for COVID-19 were worse than for ARDS pts on ECMO for influenza at our center. Disclosures Ryan K. Shields, PharmD, MS, Shionogi (Consultant, Research Grant or Support) Fernanda P. Silveira, MD, MS, FIDSA, Ansun (Individual(s) Involved: Self): Grant/Research Support; Novartis (Individual(s) Involved: Self): Grant/Research Support; Qiagen (Individual(s) Involved: Self): Grant/Research Support; Shire (Individual(s) Involved: Self): Advisor or Review Panel member, Grant/Research Support; SlieaGen (Individual(s) Involved: Self): Grant/Research Support; Whiscon (Individual(s) Involved: Self): Grant/Research Support Cornelius J. Clancy, MD, Merck (Grant/Research Support)

Murine Type III interferons are functionally redundant and correlate with bacterial burden during influenza/bacterial super-infection

Background Type III interferon, or interferon lambda (IFNλ) is a crucial antiviral cytokine induced by influenza infection. While IFNλ is important for anti-viral host defense, published data demonstrate that IFNλ is pathogenic during influenza/bacterial super-infection. It is known that polymorphisms in specific IFNλ genes affect influenza responses, but the effect of IFNλ subtypes on bacterial super-infection is unknown. Methods Using an established model of influenza, Staphylococcus aureus super-infection, we studied IFNλ3-/- and control mice to model a physiologically relevant reduction in IFNλ and to address its role in super-infection. Results Surprisingly, IFNλ3-/- mice did not have significantly lower total IFNλ than co-housed controls, and displayed no change in viral or bacterial clearance. Importantly, both control and IFNλ3-/- mice displayed a positive correlation between viral burden and total IFNλ in the bronchoalveolar lavage during influenza/bacterial super-infection, suggesting that higher influenza viral burden drives a similar total IFNλ response regardless of IFNλ3 gene integrity. Interestingly, total IFNλ levels positively correlated with bacterial burden, while viral burden and bronchoalveolar lavage cellularity did not. Conclusions These data suggest IFNλ2 can compensate for IFNλ3 to mount an effective antiviral and defense, revealing a functional redundancy in these highly similar IFNλ subtypes. Further, the IFNλ response to influenza, as opposed to changes in cellular inflammation or viral load, significantly correlates with susceptibility to bacterial super-infection. Moreover, the IFNλ response is regulated and involves redundant subtypes, suggesting it is of high importance to pulmonary pathogen defense.

Mucormycosis in CAPA, a Possible Fungal Super-Infection

The occurrence of pulmonary fungal superinfection due to Aspergillus spp. in patients with COVID-19 is a well-described complication associated with significant morbidity and mortality. This can be related to a directed effect of the virus and to the immunosuppressive role of the therapies administered for the disease. Here, we describe the first case of pulmonary infection due to Mucorales occurring in a patient with a concomitant diagnosis of COVID-19-associated pulmonary aspergillosis.

Strong Replication Interference Between Hepatitis Delta Viruses in Human Liver Chimeric Mice

BackgroundHepatitis D Virus (HDV) is classified into eight genotypes with distinct clinical outcomes. Despite the maintenance of highly conserved functional motifs, it is unknown whether sequence divergence between genotypes, such as HDV-1 and HDV-3, or viral interference mechanisms may affect co-infection in the same host and cell, thus hindering the development of HDV inter-genotypic recombinants. We aimed to investigate virological differences of HDV-1 and HDV-3 and assessed their capacity to infect and replicate within the same liver and human hepatocyte in vivo.MethodsHuman liver chimeric mice were infected with hepatitis B virus (HBV) and with one of the two HDV genotypes or with HDV-1 and HDV-3 simultaneously. In a second set of experiments, HBV-infected mice were first infected with HDV-1 and after 9 weeks with HDV-3, or vice versa. Also two distinct HDV-1 strains were used to infect mice simultaneously and sequentially. Virological parameters were determined by strain-specific qRT-PCR, RNA in situ hybridization and immunofluorescence staining.ResultsHBV/HDV co-infection studies indicated faster spreading kinetics and higher intrahepatic levels of HDV-3 compared to HDV-1. In mice that simultaneously received both HDV strains, HDV-3 became the dominant genotype. Interestingly, antigenomic HDV-1 and HDV-3 RNA were detected within the same liver but hardly within the same cell. Surprisingly, sequential super-infection experiments revealed a clear dominance of the HDV strain that was inoculated first, indicating that HDV-infected cells may acquire resistance to super-infection.ConclusionInfection with two largely divergent HDV genotypes could be established in the same liver, but rarely within the same hepatocyte. Sequential super-infection with distinct HDV genotypes and even with two HDV-1 isolates was strongly impaired, suggesting that virus interference mechanisms hamper productive replication in the same cell and hence recombination events even in a system lacking adaptive immune responses.

In vitro and in vivo co-infection and super-infection dynamics of Mayaro and Zika viruses in mosquito and vertebrate backgrounds

Factors related to increasing globalization and climate change have contributed to the simultaneous increase and spread of arboviral diseases. Co-circulation of multiple arboviruses in the same geographic regions provides impetus to study the impacts of multiple arbovirus infections in a single vector. In the present study we describe co-infection and super-infection with Mayaro virus (Family Togaviridae, genus Alphavirus) and Zika virus (family Flaviviridae, genus Flavivirus) in vertebrate cells, mosquito cells, and Aedes aegypti mosquitoes to understand the interaction dynamics of these pathogens and effects on viral infection, dissemination and transmission. In Aedes aegypti mosquitoes, co-infection has a negative impact on infection and dissemination rates for Zika virus, but not Mayaro virus, when compared to single infection scenarios, and super-infection of Mayaro virus with a previous Zika virus infection resulted in increased Mayaro virus infection rates. We found that co-infection and super-infection negatively affected Zika viral replication in vertebrate cells (Vero and Huh), resulting in the complete blockage of Zika virus replication in some scenarios. At the cellular level, we demonstrate that single vertebrate and insect cells can be simultaneously infected with Zika and Mayaro viruses. This study highlights the dynamics of arboviral co- and super-infections and emphasizes the importance of considering these dynamics during risk assessment in epidemic areas.

HDV Pathogenesis: Unravelling Ariadne’s Thread

Hepatitis Delta virus (HDV) lies in between satellite viruses and viroids, as its unique molecular characteristics and life cycle cannot categorize it according to the standard taxonomy norms for viruses. Being a satellite virus of hepatitis B virus (HBV), HDV requires HBV envelope glycoproteins for its infection cycle and its transmission. HDV pathogenesis varies and depends on the mode of HDV and HBV infection; a simultaneous HDV and HBV infection will lead to an acute hepatitis that will resolve spontaneously in the majority of patients, whereas an HDV super-infection of a chronic HBV carrier will mainly result in the establishment of a chronic HDV infection that may progress towards cirrhosis, liver decompensation, and hepatocellular carcinoma (HCC). With this review, we aim to unravel Ariadne’s thread into the labyrinth of acute and chronic HDV infection pathogenesis and will provide insights into the complexity of this exciting topic by detailing the different players and mechanisms that shape the clinical outcome.