Heterologous prime-boost-boost immunisation of Chinese cynomolgus macaques using DNA and recombinant poxvirus vectors expressing HIV-1 virus-like particles.

BACKGROUND: There is renewed interest in the development of poxvirus vector-based HIV vaccines due to the protective effect observed with repeated recombinant canarypox priming with gp120 boosting in the recent Thai placebo-controlled trial. This study sought to investigate whether a heterologous prime-boost-boost vaccine regimen in Chinese cynomolgus macaques with a DNA vaccine and recombinant poxviral vectors expressing HIV virus-like particles bearing envelopes derived from the most prevalent clades circulating in sub-Saharan Africa, focused the antibody response to shared neutralising epitopes. METHODS: Three Chinese cynomolgus macaques were immunised via intramuscular injections using a regimen composed of a prime with two DNA vaccines expressing clade A Env/clade B Gag followed by boosting with recombinant fowlpox virus expressing HIV-1 clade D Gag, Env and cholera toxin B subunit followed by the final boost with recombinant modified vaccinia virus Ankara expressing HIV-1 clade C Env, Gag and human complement protein C3d. We measured the macaque serum antibody responses by ELISA, enumerated T cell responses by IFN-γ ELISpot and assessed seroneutralisation of HIV-1 using the TZM-bl ß-galactosidase assay with primary isolates of HIV-1. RESULTS: This study shows that large and complex synthetic DNA sequences can be successfully cloned in a single step into two poxvirus vectors: MVA and FPV and the recombinant poxviruses could be grown to high titres. The vaccine candidates showed appropriate expression of recombinant proteins with the formation of authentic HIV virus-like particles seen on transmission electron microscopy. In addition the b12 epitope was shown to be held in common by the vaccine candidates using confocal immunofluorescent microscopy. The vaccine candidates were safely administered to Chinese cynomolgus macaques which elicited modest T cell responses at the end of the study but only one out of the three macaques elicited an HIV-specific antibody response. However, the antibodies did not neutralise primary isolates of HIV-1 or the V3-sensitive isolate SF162 using the TZM-bl ß-galactosidase assay. CONCLUSIONS: MVA and FP9 are ideal replication-deficient viral vectors for HIV-1 vaccines due to their excellent safety profile for use in humans. This study shows this novel prime-boost-boost regimen was poorly immunogenic in Chinese cynomolgus macaques.

Enhanced T-cell immunogenicity of plasmid DNA vaccines boosted by recombinant modified vaccinia virus Ankara in humans.

In animals, effective immune responses against malignancies and against several infectious pathogens, including malaria, are mediated by T cells. Here we show that a heterologous prime-boost vaccination regime of DNA either intramuscularly or epidermally, followed by intradermal recombinant modified vaccinia virus Ankara (MVA), induces high frequencies of interferon (IFN)-gamma-secreting, antigen-specific T-cell responses in humans to a pre-erythrocytic malaria antigen, thrombospondin-related adhesion protein (TRAP). These responses are five- to tenfold higher than the T-cell responses induced by the DNA vaccine or recombinant MVA vaccine alone, and produce partial protection manifest as delayed parasitemia after sporozoite challenge with a different strain of Plasmodium falciparum. Such heterologous prime-boost immunization approaches may provide a basis for preventative and therapeutic vaccination in humans.

Hereditary haemochromatosis, haemophagocytic lymphohistiocytosis and COVID-19.

BACKGROUND: Syndromes of iron overload have been shown to increase the risk of severe clinical disease in viral infections. Immune dysfunction is similarly described in hereditary haemochromatosis (HH). We present here the case of a 51-year-old man who developed severe coronavirus disease 2019 (COVID-19) complicated by suspected haemophagocytic lymphohistiocytosis (HLH). He was found to have HH post-mortem and we propose a link between his iron overload and the development of severe COVID-19. CASE REPORT: The initial clinical presentation consisted of cough, shortness of breath and fever. Pancytopenia, markedly elevated ferritin and d-dimer were present. Computed tomography (CT) showed bilateral ground glass changes consistent with COVID-19, widespread lymphadenopathy and splenomegaly. A subsequent combined nose and throat swab was positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). HLH was suspected based upon the H-score and Anakinra, an IL-1 receptor antagonist, was commenced. Liver function acutely worsened and magnetic resonance cholangiopancreatography (MRCP) revealed hepatic haemosiderosis. Intense splenic and cervical lymph node uptake were seen on a positron emission tomography (PET) scan and high doses of intravenous steroids were administered due to concerns over haematological malignancy. RESULTS: Day fourteen of admission heralded the start of progressive clinical deterioration with rapid increase in oxygen demands. Continuous positive airway pressure (CPAP) was trialled without success and the patient unfortunately died seventeen days into admission. Results returned after his death showed homozygous C282Y mutation of the HFE gene consistent with a diagnosis of HH. Post-mortem examination revealed widespread haemosiderin deposition in the liver along with lung pathology in keeping with severe COVID-19 and widespread splenic infarctions. CONCLUSION: An association between HH and COVID-19 is not currently described in the literature. What does exist, however, is an evidence base for the detrimental impacts iron overload has on viral infections in general and the negative effects of HH on the immune system. We therefore postulate that the underlying metabolic and immune disturbances seen in HH should be considered a potential risk factor for the development of severe COVID-19. This case also adds to the evidence that hyperinflammation appears to be a unique and interesting characteristic of this novel viral disease.

Pneumocystis pneumonia, a COVID-19 mimic, reminds us of the importance of HIV testing in COVID-19.

While clinical environments are highly focused on COVID-19, reports of missed or delayed treatment for conditions that imitate COVID-19, such as pneumonia caused by the fungus Pneumocystis jirovecii, are emerging. Given the uncertain spectrum of COVID-19 presentations and variable sensitivity of laboratory tests for SARS-CoV-2, there is a risk that, without a high index of suspicion, alternative aetiologies may be overlooked while pursuing a diagnosis of COVID-19. The British HIV Association has been calling for the inclusion of HIV testing in all patients admitted to hospital with suspected COVID-19. In this article we reflect on the importance of including HIV testing to prevent avoidable morbidity and mortality in our patients.

Hyperinflammation with COVID-19: The key to patient deterioration?

BACKGROUND: The potential risk of cytokine storm in patients with coronavirus disease 2019 (COVID-19) has been described [1]; we write to share our experience treating a 17-year-old male with haemophagocytic lymphohistiocytosis (HLH) secondary to COVID-19 infection. CASE REPORT: This patient presented with cough, sore throat, anorexia and pyrexia. On examination, he had gross cervical lymphadenopathy and palpable splenomegaly. Nose and throat swab for SARS-CoV-2 was positive and blood tests revealed pancytopaenia with very high ferritin, triglyceride and d-dimer levels. The patient's H-Score [2] was calculated at 220, suggesting probability of HLH of 93-96%. Considering Russell and colleagues' [3] comments about potential harm of corticosteroid use in patients with COVID-19 infection, the patient was commenced on treatment with the selective IL-1 receptor antagonist drug, Anakinra, and a two-day course of intravenous immunoglobulin. RESULTS: The patient responded rapidly to treatment, becoming apyrexial after 24 h. His lymph nodes and spleen began to normalise after the first 48 h, at which time point the ferritin also started to decrease. He was discharged after 11 days feeling fit and well. CONCLUSION: This case certainly illustrates the importance of hyperinflammation syndromes in COVID-19. It also raises the question - is the severe pneumonitis seen in patients with COVID-19 an immunological phenomenon? We know that the viral load of patients with COVID-19 seems to peak in the early stages of illness [4,5]; however, patients deteriorate later in the disease course, at around days 10-14. This patient, who had risk factors for deterioration (male, pancytopaenic), did not develop an oxygen requirement and clinically and biochemically improved rapidly on Anakinra with no adverse events. We might suggest Anakinra to the scientific community as a treatment option in COVID-19 infection.

Schistosomiasis.

Schistosomiasis is a common intravascular trematode infection. The snail/human lifecycle is illustrated. Travellers who acquire the infection are often asymptomatic, but nearly always give a history of fresh water exposure in endemic countries when asked. The various manifestations of symptomatic schistosomiasis are described, including the rare but important complication of neuroschistosomiasis. Guidelines for diagnostic tests, treatment with praziquantel and management of complications are given. Prospects for disease control in endemic countries by improvement in public health and mass treatment are discussed. Various vaccines are in development, but none are in clinical use yet.

Persistent Cryptococcal Brain Infection despite Prolonged Immunorecovery in an HIV-Positive Patient.

Background. HIV-positive people starting combined antiretroviral therapy may develop immune reconstitution to latent or treated opportunistic infections. Immune reconstitution to cerebral Cryptococcus is poorly understood and can be fatal. Case Presentation. A 33-year-old Zimbabwean female presented with cryptococcal meningitis and newly diagnosed HIV with a CD4 count of 51 cells/ µ L (4%). She was treated with amphotericin and flucytosine. Combined antiretroviral therapy was started four weeks later and she showed early improvement. However, over the ensuing 18 months, her clinical course was marked by periodic worsening with symptoms resembling cryptococcal meningitis despite having achieved CD4 counts ≥400 cells/ µ L. Although initially treated for relapsing cryptococcal immune reconstitution syndrome, a brain biopsy taken 17 months after initial presentation showed budding Cryptococci. Conclusion. This unusually protracted case highlights the difficulties in differentiating relapsing cryptococcal meningitis from immune reconstitution and raises questions concerning the optimum timing of initiation of combined antiretroviral therapy in such patients.