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Adenoviruses are double-stranded DNA viruses belonging to the family Adenoviridae. There are over 50 known serotypes of adenovirus, which are categorized into six subgenera (A to F). Adenoviruses are a common cause of acute febrile and respiratory infections in children and are generally self-limiting. Severe infections, including pneumonia, can occur in neonates and in adults with compromised immunity, such as those with hematopoietic stem cell transplants and in patients with human immunodeficiency virus (HIV) infection. Outbreaks of acute respiratory illness, including pneumonia, caused by adenovirus serotypes 3, 4, 7, 14 and 21 are common among military recruits, and fatal outcomes have occasionally been reported. Outbreaks of adenovirus infection in long-term nursing facilities and in hospital wards with associated cases of fatal pneumonia have also been described. In contrast, community-acquired adenovirus pneumonia in immunocompetent adult civilians has rarely been described. We report the case of a previously healthy and immunocompetent woman with severe adenovirus pneumonia who developed rapidly progressive respiratory failure requiring mechanical ventilation and who made a successful recovery after being treated with supportive measures. We also summarize the demographic, clinical, laboratory and radiological features of community-acquired adenovirus pneumonia cases in immunocompetent adult civilians that have previously been reported in the literature.
A 44-year-old Caucasian woman was admitted to our emergency department with a three-day history of a febrile illness associated with sore throat, dry cough, myalgia and diarrhea. One day prior to admission she had developed a widespread, non-pruritic, erythematous rash. Her medical history consisted of hypertension, for which she was taking atenolol, and several episodes of gout, for which she was taking allopurinol.
Her physical examination revealed that she was obese, had a body temperature of 39.0°C, a pulse rate of 112beats/minute and blood pressure of 145/90 mmHg. Her respiratory rate was 20 breaths/minute with oxygen saturation of 94% on room air. Her chest auscultation was unremarkable. She had a widespread, erythematous maculopapular rash with scattered petechiae on both legs. Examination of the oropharynx revealed erythema but no exudate.
Initial laboratory tests showed a white cell count of 9.2 × 109/L, a neutrophil count of 7.9 × 109/L, a lymphocyte count of 0.69 × 109/L, a platelet count of 254 × 109/L, a C-reactive protein concentration of 169 mg/L, an alanine aminotransferase level of 22 IU/mL, a creatinine phosphokinase (CPK) level of 950 IU/mL and a creatinine concentration of 73 μmol/L. Her HIV test was negative. Her anti-nuclear antibodies, rheumatoid factor and anti-neutrophil cytoplasmic antibodies were negative, and her complement components C3 and C4 and immunoglobulin levels were within the normal range. Her initial chest radiograph was unremarkable. She was commenced on intravenous ceftriaxone for presumed meningococcal disease.
Twenty-four hours following admission her condition rapidly deteriorated with acute respiratory failure and hypotension requiring admission to the intensive care unit for mechanical ventilation and vasopressor support. A repeat chest radiograph showed widespread interstitial infiltrates bilaterally (Figure 1). Her antibiotics were changed to imipenem and doxycycline to treat presumed bacterial pneumonia, and oseltamivir was empirically added to treat a possible 2009 pandemic influenza A (H1N1) infection.
Bacterial cultures of her blood and sputum, Legionella antigen testing of her urine, and a polymerase chain reaction (PCR) assay of her blood for Neisseria meningitidis and Streptococcus pneumoniae were all negative. Her nasopharyngeal and tracheal samples were negative for influenza A and B (including H1N1), respiratory syncytial virus (RSV) types A and B and parainfluenza virus (PIV) types 1 through 4, but they were positive for adenovirus DNA on the basis of PCR assay (using the hexon gene as the target for amplification), with a cycle threshold value of 18. Subsequent sequencing analysis performed at the respiratory Virus Reference Laboratory, London, revealed the isolate to belong to serotype 4.
The patient made an uncomplicated recovery without any specific antiviral therapy and was extubated on the fifth day of her admission. Antibiotics were stopped after a total of five days, and she was discharged to home on the ninth day of her admission. Further tests for immunodeficiency were negative.
We performed a literature search of MEDLINE for cases of community-acquired adenovirus pneumonia in immunocompetent adults. We used the search terms "adenovirus," "pneumonia," "immunocompetent," "adult" and "civilian." We excluded cases that involved military recruits, nosocomial cases and those cases in which bacterial pathogens were also implicated.
We identified 19 articles published between 1975 and 2008 describing 21 patients that matched our search terms. The demographic, laboratory, radiological and clinical details of these cases and our own are shown in Table 1.
Of the 21 cases retrieved in our literature search, 57% of the patients were men, and overall the patients' median age was 40 years (age range, 18 to 60 years). Where recorded, the commonest ethnic origin of patients was Caucasian (40%). Significant co-morbidity was uncommon among patients, but obesity was frequently noted as an examination finding.
The median duration of illness prior to admission to the hospital was five days. The following presenting symptoms were noted: fever (90%), cough (81%), dyspnea (70%), myalgia (57%), sore throat (29%), abdominal pain (14%) and diarrhea (10%). Common examination findings on presentation included abnormalities in chest auscultation (90%), pyrexia (89%) and hypoxia (66%). The presence of pharyngitis, conjunctivitis or rash was noted infrequently (19%, 19% and 5% respectively).
The median white cell count on admission to the hospital was 7.7 × 109 (range, 3.9 × 109 to 28 × 109), although neutrophilia was relatively common (33%). Lymphopenia and thrombocytopenia were noted in 52% and 19% of patients, respectively. Other frequently noted laboratory abnormalities were mildly elevated transaminases and elevated levels of CPK.
The chest radiograph at presentation was abnormal in 90% of patients. The most common pattern of abnormality was bilateral interstitial infiltrates (57%), although lobar consolidation was also noted reasonably frequently (24%).
Intubation and mechanical ventilation were required in 67% of patients and occurred at a median of one and half days following admission. Overall 24% of patients died. The median length of stay in the hospital was 21 days. Two patients received antiviral therapy with cidofovir, one of whom died.
Where recorded, the most common adenovirus serotypes identified were serotype 7 (24%), serotype 3 (19%), serotype 21 (14%) and serotype 4 (10%). The diagnosis was made most frequently on the basis of lower respiratory tract samples (principally bronchoscopic alveolar lavage fluid and lung biopsy tissue), and viral culture was the most common method of adenovirus detection (76%). There were no cases identified in the literature where molecular methods were used to diagnose adenovirus pneumonia.
In early 2009, the CNPRC housed 65 titi monkeys in one quadrant of an animal building. The index case, a healthy adult titi monkey, presented on May 14, 2009 with cough, lethargy, and decreased appetite (Fig. 1A, T1). Despite aggressive treatment with intravenous fluids and antibiotics, the animal developed severe respiratory distress and was humanely euthanized 5 days later. A second case presented 4 weeks later near the entrance to the building (Fig. 1A, T54). In the interim period, 3 healthy titi monkeys had been relocated from a separate building (Fig. 1A, T2, T3, and T19), with 2 of the 3 monkeys placed into the cage formerly occupied by the index case, reflecting a total at-risk population of 68. Over the ensuing 2 months, 21 additional monkeys, including one of the relocated monkeys, presented with clinical signs similar to those shown by the index case (attack rate = 23/68, or 34%) (Figs. 1A and 1B). Clinical signs in affected animals included cough, lethargy, poor appetite, tachypnea, and abdominal breathing. These symptoms progressed to overt respiratory distress and death or humane euthanasia within an average of 8 days. Chest radiographs typically revealed diffuse interstitial pulmonary changes and bronchoalveolar consolidation indicative of pneumonia, with right middle lobe predominance (Fig. 1C). Animals displaying clinical signs were quarantined and aggressively treated by veterinarians with supplemental oxygen, anti-inflammatory medications, bronchodilators (nebulized albuterol), broad-spectrum antibiotics, and antivirals (oseltamivir and/or ribavirin). In total, 19 animals died or were euthanized due to the illness during the outbreak (case fatality rate = 19/23, or 83%). Only 4 monkeys survived, even though the majority of sick animals (17/23, or 74%) consisted of apparently healthy adults and juveniles. Interestingly, none of the 133 rhesus macaques (Macaca mulatta) housed in the same building became sick during the outbreak, and neither did any of the Old World monkeys from surrounding outdoor colonies of rhesus and cynomolgus macaques (Macaca fascicularis).
Gross necropsy findings were similar in all titi monkeys and were characterized primarily by diffuse, consolidated pneumonias, with occasional evidence of fibrinous pleuritis, pericardial/pleural edema, and hemorrhage (Fig. 1D-1). Some livers, spleens, and lymph nodes were found to be abnormally enlarged. Hepatic necrosis and hemorrhage, along with ascites, were occasionally appreciated. On histologic examination, the normal cellular architecture of the lung and trachea was destroyed, and prominent intranuclear inclusion bodies were observed in the liver, lung, and trachea (Figs. 1D-2 and 1D-3).
A routine microbiological workup for infectious causes of the outbreak, including bacterial, mycoplasma, and fungal cultures, was negative. Respiratory viral testing failed to detect evidence of respiratory syncytial virus, adenovirus, influenza virus A and B, human metapneumovirus, and parainfluenza virus types 1, 2, and 3.
First discovered in 1953 by Rowe et al., human adenoviruses (HAdVs) are non-enveloped, double-stranded DNA viruses belonging to the Mastadenovirus genus (Adenoviridae family). They are common pathogens in children and cause a variety of diseases. HAdV accounts for at least 5 to 10% of pediatric and 1 to 7% of adult respiratory tract infections (RTIs). There are currently seven different HAdV species (A to G), some of which specifically attack the conjunctiva (species D), the upper and lower respiratory tracts (species B, C and E), and the gastrointestinal tract (species F and G). Of the 90 AdV serotypes, 55 are known to cause human diseases, with AdV3, 4, 7 and 14 being the most common types to cause respiratory disease outbreaks. HAdV has caused respiratory tract adenovirus outbreaks in Jiangsu and Taiwan provinces of China, as well as in Korea, Singapore and Malaysia [5–7]. Such infections have been estimated to cause 2–5% of RTIs overall and 4–10% of all pneumonias in City of Bethlehem.
Although HAdVs are associated with mild to moderate disease in most cases, life threatening disease can occur in some patients, particularly if they are immunocompromised. At present, China has not yet established a nationwide epidemiological surveillance program for adenovirus infections, and infections with it do not need to be legally reported, so the institutions for disease control and prevention cannot conduct early detection screening or issue early warnings. Neither are there any U.S. Food and Drug Administration approved antivirals for adenoviral infections. HAdVs play an important role in respiratory infections, particularly in children. Therefore, the aim of this study was to evaluate the epidemiological, clinical, and molecular characteristics of HAdV infections occurring among children with RTIs in a Chinese tertiary hospital from April 2017 to March 2018. Collectively, the findings from this study underscore the importance of monitoring the epidemiology of HAdV infections and protecting vulnerable patients as part of the suite of infection prevention strategies in hospitals.
The nasopharyngeal aspirate (NPA) samples (1276) used in this study were collected from hospitalized children (< 14 years) with RTIs at Beijing Friendship Hospital between April 2017 and March 2018. Informed consent from the parents or guardians of the children enrolled in the study was received. RTIs was defined as an illness that presented with at least two of the following clinical presentations: fever, cough, nasal obstruction, expectoration, sneeze and dyspnoea during the previous week. Patients, who were diagnosed with pneumonia by chest radiography, were also included in the study, even if they did not show the clinical features described above. All the specimens were stored at − 80 °C until further processing. Demographic and clinical data were obtained from the hospital’s database.
From January 2012 to December 2015 we collected specimens (nasal-pharyngeal and oral-pharyngeal swabs) and surveillance data for influenza and other viral respiratory pathogens from outpatients presenting with influenza-like-illness (ILI) at different sentinel sites in Senegal. Once collected, swabs are placed in 2-mL cryovials with viral transport medium (Universal Transport Medium; COPAN Diagnostics Inc., Murrieta, CA), and transported at a controlled temperature of 2°C—8°C to the laboratory. An ILI patient was defined as a person presenting with sudden onset of fever (>38°C) or history of sudden onset of fever in the recent past (≤ 3 days) and either cough or sore throat and/or rhinorrhea in the absence of other diagnosis, according to the CDC case definition. Each sample is accompanied by a case report form collecting demographic and clinical data. The questions included information on date of enrollment and symptom onset, sex, age, clinical symptoms, previous treatments, travelling history, vaccination status for influenza, and whether or not the patient was hospitalized. Upon arrival at the laboratory, the specimens were processed immediately for virus diagnosis. Aliquots of samples were also stored at −80°C for additional analysis (isolation and/or molecular characterization).
The data obtained daily were entered into an Epi Info database (Centers for Disease Control and Prevention, Atlanta, GA) and analyzed using Epi Info.
Adenoviruses, first isolated in the 1950s from explanted adenoid tissue, are double-stranded nonenveloped DNA viruses that naturally infect many vertebrates, including humans and nonhuman primates. The human adenoviruses in the Mastadenovirus genus, comprised of all mammalian adenoviruses, are classified into 7 species A-G, and at least 51 different serotypes (and 5 proposed types, HAdV-52 to HAdV-56) have been described to date,. Adenoviruses are the cause of an estimated 5–10% of febrile illnesses in children worldwide. Some serotypes, such as human adenovirus type 14 (HAdV-14), have been associated with severe and potentially fatal outbreaks of pneumonia in residential facilities and military bases. Adenoviruses have also been associated with other clinical syndromes including conjunctivitis, hepatitis, and diarrhea. In nonhuman primates, most epidemiologic studies of adenoviruses have focused on their identification in fecal samples from asymptomatic animals,,. Overt respiratory disease associated with simian adenoviruses has also been observed. Although adenoviruses are significant pathogens, genetically modified strains are being actively explored as potential vectors for vaccines and gene therapy.
Infection by adenoviruses has generally been thought to be species-specific. Human adenoviruses do not usually replicate in monkey cells in the absence of helper viruses, and do not productively infect rodents (and vice versa). Studies of sera from animal handlers and zoo workers exposed to chimpanzees in captivity fail to detect antibodies to chimpanzee adenoviruses,. However, recent serological surveys have found antibodies to New World and Old World monkey adenoviruses in donor human sera from regions where the monkeys are endemic,. In addition, phylogenetic analyses of adenoviruses from greater apes reveal that they fall precisely into “human” adenoviral species B, C, and E. The high degree of sequence relatedness within members of each species suggests that at least some adenoviral strains may be capable of infecting both nonhuman primates and humans.
Beginning in May of 2009, a deadly outbreak of fulminant pneumonia and hepatitis occurred in a closed colony of New World titi monkeys of the Callicebus genus at the California National Primate Research Center (CNPRC). Routine microbiological testing for an infectious etiology was negative. We previously developed the Virochip (University of California, San Francisco) as a broad-spectrum surveillance assay for identifying viral causes of unknown acute and chronic illnesses,,,,,,. The Virochip, a pan-viral microarray containing ∼19,000 probes derived from all viral species in GenBank (n∼2500),, has been previously successful in detection of novel outbreak viruses such as the SARS coronavirus, and the 2009 pandemic H1N1 influenza virus. Here we apply the Virochip to identify a novel and highly divergent adenovirus as the cause of the titi monkey outbreak. In addition, we present clinical and serological evidence that this virus may have infected a researcher at the CNPRC and a family member, thus demonstrating for the first time the potential for cross-species infection by adenoviruses.
Human adenoviruses (HAdVs) are highly contagious pathogens that are associated with a wide spectrum of human illnesses involving the respiratory, ocular, gastrointestinal, and genitourinary systems. They belong to the family Adenoviridae, genus Mastadenovirus with seven species (A-G), including each various types. Ubiquitous in the environment, HAdVs are non-enveloped, double stranded DNA viruses that vary in size from 70 to 100 nm. HAdVs are recognized as a common cause of respiratory infection in persons of all ages. The illnesses range from influenza-like fever and discomfort to pneumonia and death. Indeed, HAdVs infections are usely mild but some groups such as very young children, elderly, immunocompromised persons, or persons with underlying pulmonary or cardiac disease, might be at higher risk degree for severe disease. The most common HAdVs species that cause respiratory tract infections in children are B (HAdV-B3 and B7) and C (HAdV-C1, C2, and C5). Serotypes B3, B7, and B21 are the most frequent strains responsible for epidemics of acute febrile respiratory disease. Circulating HAdVs can vary temporally and geographically with possibility of emergent genomic variants which can be associated with more severe illness.
In the present study, we investigate the epidemiologic and viral molecular features of HAdVs circulating in Senegal after 4 consecutive years of sentinel surveillance of influenza-like Illness cases.
Human adenoviruses (HAdVs) belong to the genus Mastadenovirus within the family Adenoviridae. Adenoviruses are non-enveloped, icosahedral, double-stranded DNA viruses with genomes of 26–45 kb. The viral capsid is composed of two types of capsomeres: the hexon and the penton (which consists of the penton base and the fiber). Antigens at the surface of the virion are mainly type-specific. Hexons are involved in neutralization, and fibers in neutralization and haemagglutination-inhibition. A recombinant that has a unique combination of these three regions (penton base; hexon loops; fiber knob) derived from previously recognized genotypes will be assign a new genotype (http://hadvwg.gmu.edu).
Traditionally, the only basis for recognizing a new type of HAdV is by serology, and on the basis of their biological properties, HAdVs have been classified into 7 species (Human mastadenovirus A to G, HAdV-A to HAdV-G), including 52 human HAdV types, which are formally recognized by the International Committee on Taxonomy of Viruses (ICTV). In addition, novel HAdV genotypes (HAdV-53 to HAdV-68) were recently identified based on their bioinformatics and genomic analysis of the complete viral genome sequences (http://hadvwg.gmu.edu). Novel HAdV strains may arise from mutations or recombination among the different types of HAdVs.
HAdV can cause a variety of clinical diseases such as acute respiratory disease, gastroenteritis, and keratoconjunctivitis, which vary depending on the cell tropism of the viruses. Among the HAdV-associated respiratory diseases, viruses in species HAdV-B (HAdV-3, 7, 11, 14, 16, 21, 50, 55), species HAdV-C (HAdV-1, 2, 5, 6), and species HAdV-E (HAdV-4) [10–14] are recognized as the main pathogens responsible for the respiratory tract infection.
As the capital city of China, Beijing covers an area of 16,800 km2 with a large population of more than 19.72 million (Chinese Statistics Bureau, 2011). In order to elucidate the spectrum of the viral aetiology of acute respiratory infections and provide basic data to guide local disease prevention and control measures, a sentinel surveillance project on the viral aetiology of acute respiratory infections was initiated and sponsored by the Beijing Municipal Health Bureau in 2011. Adenovirus is one of the most common causes of viral acute respiratory infections. In this study, our primary aim was to identify the types of HAdV causing respiratory illness in Beijing since 2011, to avoid the overuse of antibiotics and to improve the level of diagnosis and treatment of respiratory viral disease especially HAdV associated disease in hospitals, and to provide scientific basis for prevention and control of HAdV causing respiratory illness.
Adenoviruses (AdVs) are non-enveloped icosahedral double-stranded DNA viruses that infect a number of vertebrate hosts, including humans and nonhuman primates. The genus Mastadenovirus within the Adenoviridae family includes 7 human adenoviral species A-G (HAdV-A through HAdV-G) and 1 simian adenoviral species A (SAdV-A). In humans, infections by adenoviruses cause conjunctivitis, gastroenteritis, hepatitis, myocarditis, and acute respiratory illness, ranging from the “common cold” syndrome to fatal outbreaks of pneumonia,. However, existing animal models of adenovirus infection to date have been primarily confined to rodents,, and no nonhuman primate (NHP) model has been established to study adenoviruses that infect humans and/or NHPs.
We previously identified a novel adenovirus, titi monkey adenovirus (TMAdV) in association with a fatal outbreak of pneumonia and hepatitis in a closed colony of captive New World titi monkeys (Callicebus cupreus), with evidence for potential cross-species transmission to a human researcher and family member with concurrent acute respiratory symptoms. The origin and natural host reservoir for TMAdV remained unknown, although neutralizing antibodies to TMAdV were found in 23 titi monkeys demonstrating clinical signs, 14 titi monkeys exposed to animals demonstrating clinical signs, 1 rhesus macaque (Macaca mulatta), and the 2 humans. Studies by other groups have also revealed the widespread presence of closely related adenoviruses in both human and nonhuman primates,,, and large-scale serological surveys have detected antibodies to monkey adenoviruses in humans living in endemic regions,. In addition, human adenovirus species E and G each contain only one member isolated from humans,, with the remaining members all isolated from monkeys or apes. Collectively, these data have raised concerns regarding the potential of adenoviruses as sources for emerging zoonotic disease in humans.
To further investigate the pathogenicity of the novel adenovirus TMAdV, we sought to develop an in vivo animal model of infection and disease for TMAdV. While TMAdV was originally discovered in a closed colony of captive titi monkeys (Callicebus cupreus) at the California National Primate Research Center (CNPRC), we recognized that in vivo testing in this monkey species was contraindicated due to the significant devastation to the colony caused by the virus. Since TMAdV was able to be successfully propagated in the marmoset lymphocyte cell line B95a in vitro (Table 1), we instead elected to pursue in vivo testing of TMAdV infection in the common marmoset (Callithrix jacchus).
The common marmoset is a New World primate that is small, easily handled, and highly susceptible to infectious agents, and thus a suitable nonhuman primate model for infectious disease,. Marmosets have been successfully used to characterize a number of emerging viral diseases, including filovirus- and arenavirus-induced hemorrhagic fevers, encephalitis, and severe acute respiratory syndrome (SARS). Since titi monkeys (Family Pithecidae, subfamily Callicebiniae) and marmoset monkeys (Family Cebidae, subfamily Callitrichinae) are classified into separate families, the use of the marmoset for in vivo testing of TMAdV infection also afforded the opportunity to directly test the capacity of TMAdV, and, by extension, adenoviruses in general, to cross the species barrier and cause productive infection in a related yet taxonomically distinct secondary host.
Diarrhea is the second most common cause of death worldwide and accounts for about 8 to 9% of the 5.9 million yearly deaths in children under the age of 5 (1, 2). Most of these deaths occur in Southeast Asia and sub-Saharan Africa (3, 4). The chances of infection with enteric viruses are higher in developing countries than developed countries, probably due to suboptimal sanitation and hygienic conditions and low quality of drinking water, especially in rural areas (5). In Cameroon, a limited number of studies have investigated the prevalence of enteric pathogens as the cause of gastroenteritis in humans. These studies mainly focused on the epidemiology of a limited number of pathogens such as rotavirus, norovirus, and enteroviruses, revealing significant differences in the prevalence of these viruses in different settings and time periods (4, 6, 7). In parts of Cameroon, a high prevalence of several enteric viruses such as enterovirus, norovirus, rotavirus, and adenovirus was found in children and adults (8). Generally in Africa, many episodes of gastroenteritis remain unexplained as no etiological agent is determined (9, 10). A proportion of the unexplained gastroenteritis cases are likely due to other known viruses, for which no tests were performed. However, a part of these gastroenteritis cases could also be caused by novel viral agents.
Transmission of these enteric viruses is predominantly fecal-oral, and humans are constantly exposed to these viruses through various routes (11). One of these routes is zoonosis from reservoirs in wild or domestic animals, either by insect vectors or by exposure to animal droppings or tissues. One rich but, until recently, underappreciated reservoir of emergent viruses is bats. Of the ∼5,500 known terrestrial species of mammals, about 20% are bats (12). Several viruses pathogenic to humans are believed to have originated in bats over the last several years, including severe acute respiratory syndrome (SARS)- and Middle East respiratory syndrome (MERS)-related coronaviruses, as well as filoviruses, such as Ebola and Marburg viruses, or henipaviruses, such as Nipah and Hendra viruses (13–18).
In the Southwest region of Cameroon, bats are hunted and eaten. Such close interactions provide ample opportunity for zoonotic events to occur (19).
Previously, we identified a plethora of known and novel eukaryotic viruses in Cameroonian fruit bats using a viral metagenomics approach, including viruses known to cause gastroenteritis in humans (sapovirus, sapelovirus, and rotaviruses A and H) and those not yet associated with gastroenteritis (bastrovirus and picobirna-like viruses) (20–23). In the current study, we metagenomically screened 221 human fecal samples collected in the same region (where bats are hunted and eaten), to assess (i) if any viruses of animal origin could be identified and (ii) which known human gastrointestinal viruses were present. These fecal samples were collected from children less than a year old to adults of more than 60 years who had gastroenteritis and/or were in contact with bats. Additionally, since the gut virome typically contains both eukaryotic and prokaryotic viruses (phages), of which the latter usually represents the largest fraction of the gut virome, we also analyzed the phageome of these samples.
Canine adenovirus (CAV) can be grouped into two distinct but related serotypes, CAV-1 and CAV-2, based on serological tests and molecular analyses [2–4]. Two types of Canine adenovirus (CAVs), Canine Adenovirus type 1 (CAV-1), the virus which causes infectious canine hepatitis, and Canine Adenovirus type 2 (CAV-2), which causes canine infectious laryngotracheitis, have been found in dogs. CAVs belong to the genus Mastadenovirus of the family Adenoviridae. Virus enters the host via direct contact with contaminated saliva, urine, and faeces. The incubation period is 4–7 days. CAV-1 replicates in vascular endothelial cells and causes a generalized infection characterized by hepatitis, whereas CAV-2 has an affinity for respiratory tract epithelium and is mainly associated with outbreaks of respiratory disease in kenneled dogs. CAV-1 causes fever, often above 40°C, apathy, anorexia, abdominal pain, blood in faeces, acute/chronic hepatitis and interstitial nephritis, tenderness, vomiting, and diarrhoea. Dogs may develop bronchopneumonia, conjunctivitis, photophobia, and a transient corneal opacity, “blue eye”, which may occur after clinical recovery as result of anterior uveitis and oedema [9, 10]. CAV-2 is characterized by respiratory disorders, with clinical signs that include tonsillitis, pharyngitis, tracheitis, and bronchitis [11–13]. Confirmation of diagnosis and identification of CAV-1 and CAV-2 infections are usually based on virus isolation, electron-microscopic observation and serological tests. There are distinct differences in structure, antigenicity, and pathogenicity between the two CAVs. Serological tests such as haemagglutination inhibition (HI), serum neutralization (SN), and enzyme-linked immunosorbent assay (ELISA) have been used detection of CAVs [15, 16]. The ELISA was found to be a highly efficient and rapid test to determine the immune status of dogs to infectious canine hepatitis virus and canine adenovirus type 2. The ELISA is a sensitive, reliable and fast method for the detection of anti-adenovirus antibody. When compared with SN test, the ELISA has several advantages. It does not require cell culture, the risk of contamination is less and the optimization is easier than other serological test methods. With the advent of molecular techniques, restriction endonuclease analysis (REA) of the viral genome has been said to differentiate between the two viruses [11, 14, 17].
The objective of this study was to determine the presence of antigen and the prevalence of CAV type 1 and 2 exposure in shelter-housed and household dogs in several regions of Turkey.
All the 21 patients who got HAdVs infections presented fever in between 38.2 to 40.0 degrees, seven (33.3%) of 21 patients had radiographic evidence of pneumonia, one patient (4.8%) had bronchitis, and others 13 patients (61.9%) had only upper respiratory tract infection symptoms such as cough and runny nose (Table 2). Among them, HAdV-55 infections (2 cases) and HAdV-7 infections (4 of 6 cases) seems led to patients of severe symptoms (pneumonia), while HAdV-3 and HAdV-4 infection caused minor symptoms (symptoms of upper respiratory tract infection or bronchitis), with only one HAdV-3 infection causing pneumonia. It is worth noting that the two patients infected with the undefined HAdV type appeared to have only mild symptoms such as fever and cough, and both patients affected by this recombinant virus are infants (below 1 years old), while patients infected with other HAdVs are all teenagers or adults.
No significant gross pathological lesions were observed. Only a few salient histologic findings were specific to TMAdV-inoculated animals (Table 2). A mild bronchitis (Fig. 3A–B) and focal area of atypical nodular hyperplasia of the liver (Fig. 3E–F) were observed in marmoset CJ29019 (Table 2). All 3 inoculated animals were found to have a mild enteritis (Fig. 3C–D) and/or colitis. Typical basophilic intranuclear inclusions consistent with active adenovirus infection were not observed in tissues from any of the TMAdV-infected marmosets. The paucity of histologic findings observed in marmosets experimentally infected with TMAdV is in sharp contrast with the striking lesions observed in moribund titi monkeys with TMAdV-associated pneumonia and hepatitis (Table 2).
Interest in the virome, or the entire population of viruses present in a biological sample, has increased recently due to improved availability of high throughput sequencing or next generation sequencing (NGS) technologies, and improved metagenomic analytical methods [1, 2]. The virome comprises all types of viruses, including those that infect prokaryotic and eukaryotic organisms, DNA or RNA viruses, and viruses that cause acute or chronic infections. Many of these viruses are difficult or impossible to propagate in cell culture, and molecular detection is difficult as no common gene such as the ribosomal 16S gene that is present in bacterial species exists in viruses. These limitations have hindered the identification and characterisation of uncultured viruses [3, 4]. Recently, due to the advent of molecular enrichment protocols, high throughput sequencing and new metagenomic analytical methods we are now able to explore, identify and characterise viruses from different biological and environmental samples with a greater capacity [2, 5–11]
In studies of human faeces, the virome has been shown to include viruses that infect eukaryotic organisms and viruses that infect prokaryotes (bacteriophages) [2, 5, 12–18]. Bacteriophages have been reported in many studies to be the most frequently detected viral constituent in the gut of humans [1, 2, 5, 8, 16, 19, 20]. The faecal virome has been characterised for several animal species including pigs, bats, cats, pigeons, horses and ferrets [2, 6, 7, 9–11, 21–31]. In dogs, the presence of enteric viral pathogens such as canine parvovirus, coronavirus, rotavirus and distemper virus (Paramyxoviridae) have been identified only through targeted studies [32–35]. To date, only one published study has used high throughput sequencing to investigate the faecal viral population in diarrhoeic dogs. These investigators analysed faeces from dogs with acute diarrhoea and detected two new virus species, canine sapovirus and canine kobuvirus; known canine enteric viruses such as canine coronavirus, canine parvovirus, canine rotavirus as well as plant and insect viruses were also reported.
The aim of this study was to describe the faecal virome of samples collected from healthy dogs, and compare these findings to the faecal virome of dogs with acute diarrhoea in Australia, using an Illumina MiSeq shotgun metagenomic sequencing approach.
The majority of emerging infectious diseases (EIDs) of humans are zoonoses, and the majority of these originate in wildlife (1–3). These diseases are largely viral (e.g., severe acute respiratory syndrome [SARS] and Nipah virus) and represent a significant global health threat. Analyses of trends in EIDs suggest that the rate of infectious disease emergence is increasing (3) and that the emergence of new viruses is not yet constrained by the richness (number of viruses) or diversity (genetic variability) of unknown viruses in wildlife, which is thought to be high. Systematically measuring viral richness, abundance, and diversity (here termed “virodiversity”) in wildlife is hindered by the large number of host species (e.g., around 5,500 mammals), their global distribution and often remote habitats (4), and the expense of collection, sampling, and viral identification or discovery (5), and it has not yet been achieved for even a single host species. In this study, we repeatedly sampled a mammalian host known to harbor emerging zoonotic pathogens (the Indian Flying Fox, Pteropus giganteus) and used PCR with degenerate primers targeting nine viral families to discover a large number and diversity of viruses. We then adapted the techniques normally used to estimate biodiversity in vertebrates and plants to estimate the total viral richness within these nine families in P. giganteus. Our analyses demonstrate proof-of-concept and provide the first statistically supported estimates of the unknown viral richness of a mammalian host and the sampling effort required to achieve it.
The virome is the community of viruses found in a particular ecosystem. Viromes characterized from animals and human are comprised of both prokaryotic and eukaryotic viruses. Commensal bacteriophages, which make up the major fraction of the fecal virome, can modulate the microbial community in the host body and influence host immunity. Although typically a smaller fraction of the enteric virome, mammalian viruses may cause diseases such as diarrhea resulting in malnutrition and dehydration. Deep sequencing of wild animal fecal viromes also unveiled many eukaryotic viruses whose pathogenicity, if any, remain mostly unknown. In the past, emergences of human infectious diseases have been initiated by zoonotic viruses originating from bats, rodents, and non-human primates. Ebola virus likely from bats, human immunodeficiency virus (HIV) from chimpanzees, and the Middle East respiratory syndrome coronavirus (MERS-CoV) from camels, have caused very large economic and public health disruptions. Therefore, it is important to identify the viruses within animals with the potential to spill over into human and result in pathogenic infections. Such zoonoses may take different routes including fecal-oral transmission. Outbreaks of zoonotic enteric viruses belonging to the families of Picornaviridae, Adenoviridae, Caliciviridae, and Reoviridae cause important enteric diseases in humans. Moreover, alteration of enteric virome in humans also affect bacterial microbiome stability and influence diseases such as inflammatory bowel disease and ulcerative colitis. Studies of intestinal and fecal bacterial communities have received much attention relative to that of the gut virome.
Cynomolgus macaque, a non-human primate species widely distributing across Southeast Asian countries have long been used for biological research including on influenza virus, Ebola virus, and simian/human immunodeficiency virus (SIV/HIV). The National Primate Research Center of Thailand–Chulalongkorn University (NPRCT-CU), maintains a colony of cynomolgus macaques captured from disturbed natural habitats. Although well-established biosecurity protocols are used to screen infectious viruses such as herpes B virus, simian retrovirus (SRV), simian immunodeficiency virus (SIV), simian-T-lymphotropic viruses (STLV) and foamy virus that might cause a sporadic outbreaks, the transmission of other viruses from wild-originating macaques remains possible. In addition, captivity may also influence gut microbiome and virome. A recent study illustrated that replacing the gut microbiome of inbred laboratory mice with that of wild mice restored their immune responses to better mimic those of wild animals. Here, we characterized and compared the fecal virome of wild and captive macaques and identified novel macaque viruses.
The median age of patients was 57.2 years (range, 23–88) and the male-to-female ratio was 1 to 2.27; 219 patients (92.02%) were farmers and 19 (7.98%) were workers or students. Among patients, 52 (21.85%) reported a tick bite within 2 weeks (5–14 days) before the onset of clinical manifestations; the remaining patients did not recall receiving a tick bite.
The main clinical features in confirmed patients included sudden onset of fever (>37.5°C −40°C) lasting up to 10 days, fatigue, anorexia, headache, myalgia, arthralgia, dizziness, enlarged lymph nodes, muscle aches, vomiting and diarrhea, upper abdominal pain, and relative bradycardia (Table 1). A small number of cases suffered more severe complications, including hypotension, mental status alterations, ecchymosis, gastrointestinal hemorrhage, pulmonary hemorrhage, respiratory failure, disseminated intravascular coagulation, multiple organ failure, and/or death. Most patients had a good outcome, but elderly patients and those with underlying diseases, neurological manifestations, coagulopathy, or hyponatremia tended to have a poorer outcome.
Laboratory tests showed that confirmed patients characteristically developed thrombocytopenia, leukopenia, proteinuria, and elevated serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels (Table 2). Biochemical tests revealed generally higher levels of lactate dehydrogenase, creatine kinase, AST and ALT enzymes, especially AST.
Acute respiratory infections (ARI) are a major cause of morbidity and mortality in children under five, the elderly, and vulnerable patients. Upper respiratory tract infection incidence is estimated at 7–8 per year in children under 4, and at 2–4 in adults.
Most respiratory viruses belong to five different viral families (Para- and Ortho-myxoviridae, Picornaviridae, Coronaviridae, Adenoviridae), and include 14 viral species, defining what is called the “respiratory panel”. Some of them have a high potential of emergence and can cause pandemics. Although some viruses have been associated with particular diseases (respiratory syncytial virus and bronchiolitis, parainfluenza virus 3 and laryngitis, rhinovirus and common cold, influenza virus and flu syndrome), there is no evidence for a clinical specificity, and only the virological diagnosis provides an accurate identification of the ARI [4, 5].
Detection of respiratory viruses is of little interest in general practice, in that the infection does not present a risk of severity for the patient. However, virological confirmation of ARI is needed in severe clinical presentations, requiring hospitalization in intensive care units and occurring in vulnerable subjects [4, 6]. The goal of early virological diagnosis would be an optimization of patient care, which could lead to reduction in length of hospital stay, a saving of antibiotics, and complementary examinations.
Virological tests allow for the establishment of accurate diagnosis of infection, assessment of evolving risks (bacterial infection, acute respiratory distress syndrome), and the establishment of measures to limit its spread (isolation, wearing gloves and masks).
Pandemics of Severe Acute Respiratory Syndrome (SARS, 2002-2003) and influenza A-H1N1 (2009) lead to the development of molecular biological techniques applied to virological diagnosis, mainly based on PCR (polymerase chain reaction). Performances of molecular methods in respiratory virology are so significant that they have replaced conventional techniques (culture, detection of viral antigens) as a reference method [8–11].
Multiplex PCR techniques are particularly suited to medical diagnosis because they can detect multiple viral targets in the same time, avoiding the virologist a selection of viral targets to search. There are now many commercial kits for the detection of a range of 12 to 15 respiratory viruses and some intracellular bacteria [6, 7, 12, 13]. Molecular techniques (real-time PCR) also make it possible to achieve a semi quantification of the viral molecular material present in the sample, giving additional information about the respiratory viral load (interest in therapeutic monitoring and infection transmission risk). A normalized viral load can be obtained by adding a cell quantification step.
The primary site for replication of respiratory viruses is the ciliated airway epithelium. The sample must be taken as soon as possible after the onset of symptoms. This is usually a nasal swab or nasopharyngeal aspiration (especially realized in children under 2). These samples are easily accessible and especially adapted to upper ARI. If a rich cell collection appears to be an important prerequisite for the quality of respiratory viral diagnosis, there is currently no information on a possible cellularity threshold that would validate the result of the viral molecular detection.
The main objective of this work is the study of cellularity in 800 respiratory specimens previously characterized virologically. The results should help to define the concept of “cellular richness” and determine the factors that influence it.
Acute respiratory tract infections (ARTIs) are the most common cause of childhood morbidity and mortality worldwide, accounting for about 30% of all childhood deaths in the developing world. The WHO estimates that ARTIs account for 1.9 to 2.2 million childhood deaths annually, with 42% occurring in Africa. The etiologic agents include viruses, bacteria, and fungi. Among the viruses responsible for ARTIs are members of the Orthomyxoviridae, Coronaviridae, Picornaviridae, Paramyxoviridae, Adenoviridae, and Parvoviridae. Bacteria, such as Streptococcus pneumoniae (S. pneumoniae), Haemophilus influenzae, Staphylococcus aureus, Moraxella catarrhalis, Mycoplasma pneumoniae (M. pneumoniae), and Chlamydia pneumoniae are the most common involved microbes. However, in general, viral infections are shown to be responsible for about 80% of ARTIs and are the cause of 90% of hospitalizations in children below 5 years of age.
According to the localization, ARTIs are divided into 2 categories: lower respiratory tract infections (LRTIs) and upper respiratory tract infections (URTIs). URTIs refer to pathogenic manifestations of rhinosinusitis, tonsillitis, pharyngitis, laryngitis/laryngotracheitis and otitis, while LRTIs include tracheitis, bronchitis, bronchiolitis, and bronchopneumonia. LRTIs are more harmful than URTIs and thus account for most of the serious disease burden. They are the major cause of hospital admissions in young children in the developed world.
The current epidemiological situation, patient’s age, clinical symptoms, radiographic and laboratory data and response to treatment may help to differentiate viral from bacterial infections. Even so, no clinical algorithm exists that will distinguish clearly between different causes of childhood respiratory infections. Accurate and rapid diagnostic tests that identify the cause of LRTIs in children can reduce the use of antibiotics, improve the targeted use of drugs and help to control nosocomial transmission. New technologies permit the detection of more than one pathogen in a single probe and are more relevant than non-multiplex approaches such as conventional PCR, viral cell culture and, immunofluorescence or immunochromatography assays. The recently developed commercial diagnostic assays based on multiplex real-time PCR (qPCR/qRT-PCR) allow sensitive and specific detection of a broad panel of conventional and emerging viruses in respiratory tract specimens. Thus, they have been established as a standard method within the past few years. These technological advances have changed the landscape of virus detection and provide the opportunity to improve our understanding of the epidemiology of respiratory viruses.
In the Arab Maghreb, few studies have been published describing the detection and the epidemiology of respiratory pathogens [10–12]. In Tunisia for example, studies were limited to few respiratory viruses and focused mostly on mono-detections such as influenza viruses (InfVs) [13, 14], MERS-Coronavirus, respiratory syncytial virus (RSV) [16–18] and, metapneumovirus (MPV). To our knowledge, a single Tunisian study describes the surveillance of InfVs including the detection of a panel of respiratory viruses by multiplex assay, but no information about patient’s clinical profile and its association with viral infection was provided.
The purpose of this study was to examine the distribution of viral pathogens, S. pneumoniae (most common cause for community-acquired bacterial LRTI), and M. pneumoniae in ARTIs from patients hospitalized between September 2013 and December 2014 in the Sousse area using real-time assays. In addition, we aimed to associate the manifestation of ARTIs with the occurrence of different respiratory pathogens. This study will be helpful for physicians to get a detailed profile about the distribution of respiratory pathogens in Sousse, Tunisia.
Over one-half of all known human pathogens originated from animals, and over 75% of emerging infectious diseases identified in the last three decades were zoonotic.1 The threat of veterinary pathogens to human health continues to grow because of increasing population density and urbanization, global movement of people and animals, and deforestation accompanied by increased proximity of human and wildlife habitats. Recent emerging infectious diseases have been concentrated in tropical Africa, Latin America, and Asia, with outbreaks usually occurring within populations living near wild animals.1 Identification of animal reservoirs from which zoonosis may emerge and detection and characterization of pathogens in these reservoirs will facilitate timely implementation of control strategies for new zoonotic infections.2 Therefore, pathogen discovery studies in animal reservoirs represent an integral part of public health surveillance.
Bats have long been known as natural hosts for lyssaviruses, and more recently, they have been recognized as potential reservoirs for emerging human pathogens, including henipaviruses, filoviruses, and severe acute respiratory syndrome (SARS) related coronaviruses.3,4 Novel viruses are documented in bats every year, which has drawn increasing attention to these mammalian reservoirs that are uniquely associated with a variety of known and potential zoonotic pathogens. In this study, we report the detection of nucleic acids of adenoviruses, rhabdoviruses, and paramyxoviruses in bats from Kenya.
A total of 221 human fecal samples (131 from Kumba and 90 from Lysoka) were collected from two hospitals in the Southwest region of Cameroon, for viral metagenomics screening. From these fecal samples, a total of 63 pools were constituted in categories based on age, bat contact status, and location (see Table S1 in the supplemental material). Illumina sequencing of all the 63 human pools generated in total approximately 708 million raw paired-end (PE) reads (between 4.3 and 53.4 million reads per pool). After trimming, 67% of the reads (471 million) were retained and 86% of these retained trimmed reads (405 million) were annotated using Diamond. Of these, 18% (74 million) could be attributed as viral.
Blood serum samples belonging to 188 dogs, which had either been admitted to the Internal Medicine Clinic of Selcuk University, Faculty of Veterinary Medicine, with clinical symptoms or had been sampled at the dog shelters they were cared after in Isparta and Burdur provinces, were examined using the ELISA method. Of these samples, 103 (54.7%) were found to be positive for antibodies against CAV infection (Table 3).
Of the 108 female animals sampled in the study, 55 (50.9%) were determined to be positive for CAV antibodies, while 48 (60%) of the sampled 80 male animals were confirmed to be positive (Table 3). Of the 7 animals below 1 year of age, only 1 (14.2%; 2-month-old female puppy) was positive, and the remaining ones were found to be negative for CAV antibodies. Of the 53 animals aged 1-2 years, 22 (41.5%); of the 58 animals aged 2 years, 31 (53.4%); of the 64 animals aged 3 years, 44 (68.7%); and of the 6 animals aged >4 years, 5 (83.33%) were found to be positive (Table 2).
Blood leukocyte samples from dogs were processed and inoculated onto confluent monolayers of MDCK cells using standard virological techniques. The inoculated cells were incubated at 37°C and observed daily for the appearance of cytopathic effect (CPE). After third passage, cells were examined by immunofluorescence test for virus isolation. No morphological changes were observed in cell cultures, and a positive result was not detected by immunofluorescence test.
Clinical Findings. Blood samples were taken from 111 dogs showing clinical symptoms which were brought to the Internal Medicine Clinic of Selcuk University, Faculty of Veterinary Medicine. Seventy-seven dogs were sampled from Isparta and Burdur dog shelters by random sampling, regardless of the clinical findings. Dogs showed a systemic disease, characterized by fever, diarrhea, vomiting, mucopurulent oculonasal discharge, mucopurulent conjunctivitis, severe moist cough, signs of pulmonary disease, and dehydration. Corneal opacity and photophobia were determined for two dogs.
Swimming pools have been implicated in the transmission of infections. The risk of infection has mainly been linked to fecal contamination of the water, generally due to feces released by bathers or to contaminated source water. Failure in disinfection has been recorded as the main cause of many of the outbreaks associated with swimming pools.
The majority of reported swimming pool-related outbreaks have been caused by enteric viruses. Sinclair and collaborators reported that 48% of viral outbreaks occur in swimming pools, 40% in lakes or ponds, and the remaining 12% in fountains, hot springs, and rivers (4% each).
Viruses cannot replicate outside their host’s tissues and cannot multiply in the environment. Therefore, the presence of viruses in a swimming pool is the result of direct contamination by bathers, who may shed viruses through unintentional fecal release, or through the release of body fluids such as saliva, mucus, or vomitus. Evidence suggests that skin may also be a potential source of pathogenic viruses.
All 285 patients with FTLS were from the Henan Province of China and were provisionally diagnosed as suspected HGA on the basis of similar clinical manifestations,. They represented four different epidemiologically linked sporadic cases and a few clusters of cases including 79 patients in 2007, seven patients in 2008, 47 patients in 2009, and 152 patients in 2010. The patients presented mainly between April and October, peaking in April-May during the tea-picking season in Henan. All patients resided in mountainous and hilly rural areas. In our study, 238 of 285 patients tested positive for novel bunyavirus infection by RT-PCR and/or IFA.