Pin Yu

Intestinal inflammatory pseudotumour with regional lymph node involvement: identification of a new bacterium as the aetiological agent

Inflammatory pseudotumours are the morphological expression of diverse processes such as reactive/reparative, infective, and neoplastic. This paper reports an example of intestinal inflammatory pseudotumour, with identification of a newly characterized bacterium in the lesion. The patient presented with intestinal obstruction. Laparotomy revealed a tumour in the terminal ileum causing stricture, and multiple enlarged regional lymph nodes. Histologically, the tumour and lymph nodes were composed of plump spindle cells disposed in a vague storiform pattern, and associated with lymphocytes and plasma cells. Immunohistochemical studies showed that most of the spindle cells were histiocytes (CD68 positive), prompting a search for a bacterial aetiology, akin to mycobacterial spindle cell pseudotumour. All histochemical stains for micro‐organisms were unrewarding. Ultrastructural studies, however, revealed abundant bacteria within the spindle histiocytes. Polymerase chain reaction, using conserved oligonucleotide primers complementary to the 16S rRNA genes of eubacteria, was employed to amplify 16S rRNA gene fragments directly from the involved lymph node tissue. Phylogenetic analysis of the amplified DNA sequences revealed an organism with 99% sequence conformity to Pseudomonas veronii, a bacterium which has hitherto not been implicated in human infection. The importance of searching for an infective agent in inflammatory pseudotumour in the appropriate setting is re‐emphasized. Copyright

An Animal Model of MERS Produced by Infection of Rhesus Macaques With MERS Coronavirus

Abstract In 2012, a novel coronavirus (CoV) associated with severe respiratory disease, Middle East respiratory syndrome (MERS-CoV; previously known as human coronavirus–Erasmus Medical Center or hCoV-EMC), emerged in the Arabian Peninsula. To date, 114 human cases of MERS-CoV have been reported, with 54 fatalities. Animal models for MERS-CoV infection of humans are needed to elucidate MERS pathogenesis and to develop vaccines and antivirals. In this study, we developed rhesus macaques as a model for MERS-CoV using intratracheal inoculation. The infected monkeys showed clinical signs of disease, virus replication, histological lesions, and neutralizing antibody production, indicating that this monkey model is suitable for studies of MERS-CoV infection.

Novel Avian-Origin Human Influenza A(H7N9) Can Be Transmitted Between Ferrets via Respiratory Droplets

The outbreak of human infections caused by novel avian-origin influenza A(H7N9) in China since March 2013 underscores the need to better understand the pathogenicity and transmissibility of these viruses in mammals. In a ferret model, the pathogenicity of influenza A(H7N9) was found to be less than that of an influenza A(H5N1) strain but comparable to that of 2009 pandemic influenza A(H1N1), based on the clinical signs, mortality, virus dissemination, and results of histopathologic analyses. Influenza A(H7N9) could replicate in the upper and lower respiratory tract, the heart, the liver, and the olfactory bulb. It is worth noting that influenza A(H7N9) exhibited a low level of transmission between ferrets via respiratory droplets. There were 4 mutations in the virus isolated from the contact ferret: D678Y in the gene encoding PB2, R157K in the gene encoding hemagglutinin (H3 numbering), I109T in the gene encoding nucleoprotein, and T10I in the gene encoding neuraminidase. These data emphasized that avian-origin influenza A(H7N9) can be transmitted between mammals, highlighting its potential for human-to-human transmissibility.

Treatment With Lopinavir/Ritonavir or Interferon-β1b Improves Outcome of MERS-CoV Infection in a Nonhuman Primate Model of Common Marmoset

Abstract Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe disease in human with an overall case-fatality rate of >35%. Effective antivirals are crucial for improving the clinical outcome of MERS. Although a number of repurposed drugs, convalescent-phase plasma, antiviral peptides, and neutralizing antibodies exhibit anti-MERS-CoV activity in vitro, most are not readily available or have not been evaluated in nonhuman primates. We assessed 3 repurposed drugs with potent in vitro anti-MERS-CoV activity (mycophenolate mofetil [MMF], lopinavir/ritonavir, and interferon-β1b) in common marmosets with severe disease resembling MERS in humans. The lopinavir/ritonavir-treated and interferon-β1b-treated animals had better outcome than the untreated animals, with improved clinical (mean clinical scores ↓50.9%–95.0% and ↓weight loss than the untreated animals), radiological (minimal pulmonary infiltrates), and pathological (mild bronchointerstitial pneumonia) findings, and lower mean viral loads in necropsied lung (↓0.59–1.06 log10 copies/glyceraldehyde 3-phosphate dehydrogenase [GAPDH]; P < .050) and extrapulmonary (↓0.11–1.29 log10 copies/GAPDH; P < .050 in kidney) tissues. In contrast, all MMF-treated animals developed severe and/or fatal disease with higher mean viral loads (↑0.15–0.54 log10 copies/GAPDH) than the untreated animals. The mortality rate at 36 hours postinoculation was 67% (untreated and MMF-treated) versus 0–33% (lopinavir/ritonavir-treated and interferon-β1b-treated). Lopinavir/ritonavir and interferon-β1b alone or in combination should be evaluated in clinical trials. MMF alone may worsen MERS and should not be used.

Histopathological features and distribution of EV71 antigens and SCARB2 in human fatal cases and a mouse model of enterovirus 71 infection

Enterovirus 71 (EV71) is a neurotropic pathogen that causes hand, foot, and mouth disease. While infection is usually self-limiting, a minority of patients infected with EV71 develop severe neurological complications. In humans, EV71 has been reported to utilize the scavenger receptor class B, member 2 (SCARB2) as a receptor for infectious cellular entry. In this study, we define the pathological features of EV71-associated disease as well as the distribution of EV71 antigen and SCARB2 in human fatal cases and a mouse model. Histopathologically, human fatal cases showed severe central nervous system (CNS) changes, mainly in the brainstems, spinal cords, and thalamus. These patient further exhibited pulmonary edema and necrotic enteritis. Immunohistochemical analysis of human fatal cases demonstrated that EV71 antigen and SCARB2 were observed mainly in neurons, microglia cells and inflammatory cells in the CNS, and epithelial cells in the intestines. However, skeletal muscle tissue was negative for EV71 antigen. In a mouse model of EV71 infection, we observed massive necrotic myositis, different degrees of viral diseases in CNS, and extensive interstitial pneumonia. In mice, EV71 exhibits strong myotropism compared to the neurotropism seen in humans. EV71 antigen was detected in the spinal cord and brainstem of mice. However, there was no clear correlation between mouse SCARB2 and EV71 antigen distribution in the mouse model, consistent with previous results that SCARB2 functions as a receptor for EV71 in humans but not mice. The EV71-induced lesions seen in the mouse model resembled the pathological changes seen in human samples. These results increase our understanding of EV71 pathogenesis and will inform further work developing a mouse model for EV71 infection.

The mouse and ferret models for studying the novel avian-origin human influenza A (H7N9) virus

BackgroundThe current study was conducted to establish animal models (including mouse and ferret) for the novel avian-origin H7N9 influenza virus.FindingsA/Anhui/1/2013 (H7N9) virus was administered by intranasal instillation to groups of mice and ferrets, and animals developed typical clinical signs including body weight loss (mice and ferrets), ruffled fur (mice), sneezing (ferrets), and death (mice). Peak virus shedding from respiratory tract was observed on 2 days post inoculation (d.p.i.) for mice and 3–5 d.p.i. for ferrets. Virus could also be detected in brain, liver, spleen, kidney, and intestine from inoculated mice, and in heart, liver, and olfactory bulb from inoculated ferrets. The inoculation of H7N9 could elicit seroconversion titers up to 1280 in ferrets and 160 in mice. Leukopenia, significantly reduced lymphocytes but increased neutrophils were also observed in mouse and ferret models.ConclusionsThe mouse and ferret model enables detailed studies of the pathogenesis of this illness and lay the foundation for drug or vaccine evaluation.

Rapid adaptation of avian H7N9 virus in pigs

How the H7N9 avian influenza virus gained the distinct ability to infect humans is unclear. Pigs are an important host in influenza virus ecology because they are susceptible to infection with both avian and human influenza viruses and are often involved in interspecies transmission. Here, we passaged one avian isolate and one human isolate in pigs to examine the mammalian host adaptation of the H7N9 virus. The avian virus replicated to a high titer after one passage, whereas the human isolate replicated poorly after three passages in pig lungs. Sequence analysis found nine substitutions in the HA, NA, M and NS segments of the avian isolate, which enhanced the binding affinity for human-type receptors. These results indicate that avian H7N9 influenza viruses can be easily adapted to pigs and that pigs may act as an important intermediate host for the reassortment and transmission of such novel viruses.

Antigenicity and transmissibility of a novel clade 2.3.2.1 avian influenza H5N1 virus

A genetic variant of the H5N1 influenza virus, termed subclade 2.3.2.1, was first identified in Bulgaria in 2010 and has subsequently been found in Vietnam and Laos. Several cases of human infections with this virus have been identified. Thus, it is important to understand the antigenic properties and transmissibility of this variant. Our results showed that, although it is phylogenetically closely related to other previously characterized clade 2.3 viruses, this novel 2.3.2.1 variant exhibited distinct antigenic properties and showed little cross-reactivity to sera raised against other H5N1 viruses. Like other H5N1 viruses, this variant bound preferentially to avian-type receptors, but contained substitutions at positions 190 and 158 of the haemagglutinin (HA) protein that have been postulated to facilitate HA binding to human-type receptors and to enhance viral transmissibility among mammals, respectively. However, this virus did not appear to have acquired the capacity for airborne transmission between ferrets. These findings highlight the challenges in selecting vaccine candidates for H5N1 influenza because these viruses continue to evolve rapidly in the field. It is important to note that some variants have obtained mutations that may gain transmissibility between model animals, and close surveillance of H5N1 viruses in poultry is warranted.

Distribution of enterovirus 71 RNA in inflammatory cells infiltrating different tissues in fatal cases of hand, foot, and mouth disease

In previous studies of hand, foot, and mouth disease patients fatally infected with enterovirus 71 (EV71), the distribution of viral protein, but not the genome, was determined. To understand the pathogenesis of EV71, however, it is important to investigate the spread of the viral genome. There have been no pathological studies of in situ EV71 viral RNA in inflammatory cells infiltrating various tissues of fatal cases. We therefore first investigated the distribution and classification of inflammatory cells in various tissues and then performed in situ EV71 RNA hybridization in these tissues to better understand the pathogenesis of EV71 infection. EV71 RNA was found mainly in inflammatory cells infiltrating the central nervous system (CNS), intestines, lungs, and tonsils. Most EV71 RNA-positive inflammatory cells in the CNS were macrophages/microglia and neutrophils infiltrating the perivascular cuffing, microglial nodule, neuronophagia, and meninges. CD68+ macrophages and CD15+ neutrophils were diffusely distributed in tissues with severe pathological changes. This study demonstrates the presence of EV71 RNA in inflammatory cells infiltrating tissues in fatally infected patients. Our findings suggest that fatal EV71 infection with extensive infiltration of macrophages/microglia and neutrophils into the CNS results in severe neurological lesions.

Combinations of oseltamivir and fibrates prolong the mean survival time of mice infected with the lethal H7N9 influenza virus

The outbreak of human infections caused by the novel avian-origin H7N9 influenza viruses in China since March 2013 underscores the urgent need to find an effective treatment strategy against H7N9 infection in humans. In this study, we assessed the effectiveness of combinations of oseltamivir and two immunomodulators (simvastatin and fenofibrate) against H7N9 infection in a mouse model. Mice treated with oseltamivir plus fenofibrate exhibited the longest mean survival time, the largest reduction of viral titre in lung tissue, the highest levels of CD4(+) and CD8(+) T-lymphocytes, and the greatest decrease in pulmonary inflammation. Thus, the combination of oseltamivir plus fenofibrate improved the outcomes of mice infected with H7N9 virus by simultaneously reducing viral replication and normalizing the aberrant immune response. This drug combination should be considered in randomized controlled trials of treatments for H7N9 patients.