Pre-Transplant Screening for Latent Adenovirus in Donors and Recipients

Human adenoviruses are frequent cause of slight self-limiting infections in immune competent subjects, while causing life-threatening and disseminated diseases in immunocompromised patients, particularly in the subjects affected by acquired immunodeficiency syndrome and in bone marrow and organ transplant recipients. Here, infections interest lungs, liver, encephalon, heart, kidney and gastro enteric tract. To date, human adenoviruses comprise 51 serotypes grouped into seven species, among which species C especially possesses the capability to persist in infected tissues. From numerous works, it emerges that in the recipient, because of loss of immune-competence, both primary infection, via the graft or from the environment, and reactivated endogenous viruses can be responsible for transplantation related adenovirus disease. The transplants management should include the evaluation of anti-adenovirus pre-transplant screening similar to that concerning cytomegalovirus. The serological screening on cytomegalovirus immunity is currently performed to prevent viral reactivation from grafts and recipient, the viral spread and dissemination to different organs and apparatus, and potentially lethal outcome

Novel Human Astroviruses: Prevalence and Association with Common Enteric Viruses in Undiagnosed Gastroenteritis Cases in Spain

Adenovirus; Children; Classic astrovirusAdenovirus; Nens; Astrovirus clàssicAdenovirus; Niños; Astrovirus clásicoA remarkable percentage of acute gastroenteritis cases remain etiologically undiagnosed. The aim of the study was to determine the prevalence of common and emerging enteric viruses, such as novel human astroviruses, among undiagnosed samples from children with acute gastroenteritis. Epidemiological studies for novel human astroviruses are still scarce. Stool samples collected over two consecutive winter seasons (2016-2017) from children with gastroenteritis in Spain, which were negative for bacteria, rotavirus, and adenovirus by routine diagnostics were screened by real-time RT-PCR assays for the presence of classical and novel astrovirus, rotavirus, norovirus GI and GII, sapovirus, and adenovirus. Overall, 220/384 stool samples (57.3%) were positive for at least one virus. Co-infections were identified in 21% of cases. Among a total of 315 viruses identified, adenovirus was the most prevalent (n = 103), followed by rotavirus (n = 51), sapovirus (n = 50), classical astrovirus (n = 43), novel astroviruses (n = 42), and norovirus (n = 26). Novel astroviruses were present in 13.3% of virus-positive cases. Most novel astroviruses were found in children <2-year-old (30/39 children, 77%, p = 0.01) and were found in co-infection (66%). Only classical astroviruses demonstrated significant differences in the Cq values during mono-infections compared to co-infections. In conclusion, common enteric viruses may be frequently found in children with undiagnosed gastroenteritis, indicating the need to implement more sensitive diagnostic methods. Novel astroviruses circulate in the community and could be the cause of gastroenteritis among young children.Supported in part by the Biotechnology Reference Network (XRB) program of the Generalitat de Catalunya. This work was also supported in part by the Certest Biotec Company. Diem-Lan Vu was recipient of a fellowship from the Geneva University Hospital

Detection of Human Torovirus Like Particles and Adenovirus Type F in Children Attending to Babylon Maternity and Children Hospital

Toroviruses are enteric viruses belonging to the Nidovirales order that infect different animal species and human . Torovirus-like particales (TVLPs) that are immunologically related to BRV have been reported as etiological agents of gastroenteritis in humans. The lack of “in vitro” culture systems for toroviruses, except  for the prototype Berne virus or BEV, isolated originally from an infected horse, has hampered their study and the development of diagnostic assays. This  study  describes a real time RT-PCR method to detect  human  torovirus- like particles  (TVLPs) RNA in clinical  stool  samples using primers corresponding to the gene coding for  the nucleocapsid protein which are conserved in all (TVLPs) strains known to date. During this study, the CT value  measured  during real-time PCR analysis was used as an indication of the viral load found in the stool  sample . The assay was evaluated with 72 stool samples from children attending the Babylon maternity and children hospital. Fifty tow out of 72 (72.2%) children were shedding virus at  the time of sample collection, indicating a high incidence of TVLPs  infection  in Babylon Province. This  is the first study  attempted  for   estimating  the presence of TVLPs  in Iraq. The real time RT-PCR assay described in this study  provides a rapid, highly sensitive, specific and reliable detection and quantization  method enabling future TVLPs  epidemiological studies. In addition to that  the study included the development of real-time PCR assays for the detection of group F Adenovirus in 250 stool samples of pediatric subjects  exhibiting symptoms of diarrhea and/ or vomiting  which  were examined. PCR results of 10 positive Adenovirus group F diarrheic stool samples were confirmed by electron microscopy  examination which gave clear positive Adenovirus appearance . Till now there was no successful virus  culture growth for  isolation of diarrhegenic type 40 and 41 grow in routine cell culture . The result of this study by real time reverse transcription  – PCR  assay reflected in 72 .2 % and 58 % torovirus and adenovirus group F respectively. The genotyping results of adenoviruses(genotype 40 and 41)  highlight the significance of rapid molecular methods for the routine screening of stool samples in diagnostic laboratories to provide rapid and efficient methods . Keywords: Human Torovirus, Adenovirus, RT-PCR, Electron Microscopy

The development, implementation and evaluation of a real-time PCR-based diagnostic service for viral causes of infectious intestinal disease

Outbreaks and sporadic cases of viral Infectious Intestinal Disease (IID) are a major public health issue resulting in significant morbidity and sometimes mortality each year. The economic costs associated are substantial. Laboratory diagnosis of viral IID is important as the many infectious and non-infectious causes cannot be reliably differentiated using clinical or epidemiological characteristics alone. An accurate diagnosis can aid patient management, infection control procedures and reduce health care costs by preventing unnecessary treatments, testing for alternative causes and hospital stay. It also aids public health surveillance. At the start of the research described in this thesis the West of Scotland Specialist Virology Centre (WOSSVC) used Electron Microscopy (EM) as the frontline test for outbreaks and sporadic cases of IID. However, although rapid on a small number of samples, this technique has been shown to be insensitive, laborious and is not suited to testing large numbers of samples. The research presented in this thesis sought to examine whether molecular diagnostic techniques such as conventional gel-based or real-time Polymerase Chain Reaction (PCR) assays could be a viable replacement for EM as the frontline test(s) for viral IID in a routine laboratory service of this type, and whether their implementation could bring benefits to the laboratory service in terms of improved rapidity, sensitivity and throughput. The aim was to adapt published PCR methods for use in routine diagnostic work rather than for research purposes, an approach that distinguishes this research from previous work in this area. In order to achieve this aim, the appropriate PCR techniques were first selected from the literature, based on a combination of clinical and laboratory requirements, and were adapted for use in the laboratory service. A series of laboratory experiments was then carried out in order to compare the sensitivity of the adapted methods to existing techniques such as EM and antigen detection assays (EIAs) and to other methods that emerged during the period of study including alternative PCR assays. Where found to be suitable, the selected PCR tests were implemented in the routine diagnostic service for viral IID. The effects of these changes on the laboratory service were then examined. The results show that since the introduction of molecular tests at WOSSVC for the detection of viral pathogens in cases of gastroenteritis the number of samples tested has risen steadily, as have the detection rates for each of the main viral causes of IID. Furthermore, this has been achieved at the same time as a substantial reduction in sample turn-around-times. Such improvements will have a positive impact in several areas of public health relating to viral IID and are discussed fully, including patient management, infection control and national surveillance

PCR diagnostics and monitoring of adenoviral infections in hematopoietic stem cell transplantation recipients

After stem cell transplantation, human patients are prone to life-threatening opportunistic infections with a plethora of microorganisms. We report a retrospective study on 116 patients (98 children, 18 adults) who were transplanted in a pediatric bone marrow transplantation unit. Blood, urine and stool samples were collected and monitored for adenovirus (AdV) DNA using polymerase chain reaction (PCR) and real-time PCR (RT-PCR) on a regular basis. AdV DNA was detected in 52 (44.8%) patients, with mortality reaching 19% in this subgroup. Variables associated with adenovirus infection were transplantations from matched unrelated donors and older age of the recipient. An increased seasonal occurrence of adenoviral infections was observed in autumn and winter. Analysis of immune reconstitution showed a higher incidence of AdV infections during periods of low T-lymphocyte count. This study also showed a strong interaction between co-infections of AdV and BK polyomavirus in patients undergoing hematopoietic stem cell transplantations

Healthcare-associated Viral Gastroenteritis among Children in a Large Pediatric Hospital, United Kingdom

Enteric viruses introduced from the community are major causes of these illnesses

Novel Human Astroviruses: Prevalence and Association with Common Enteric Viruses in Undiagnosed Gastroenteritis Cases in Spain

A remarkable percentage of acute gastroenteritis cases remain etiologically undiagnosed. The aim of the study was to determine the prevalence of common and emerging enteric viruses, such as novel human astroviruses, among undiagnosed samples from children with acute gastroenteritis. Epidemiological studies for novel human astroviruses are still scarce. Stool samples collected over two consecutive winter seasons (2016-2017) from children with gastroenteritis in Spain, which were negative for bacteria, rotavirus, and adenovirus by routine diagnostics were screened by real-time RT-PCR assays for the presence of classical and novel astrovirus, rotavirus, norovirus GI and GII, sapovirus, and adenovirus. Overall, 220/384 stool samples (57.3%) were positive for at least one virus. Co-infections were identified in 21% of cases. Among a total of 315 viruses identified, adenovirus was the most prevalent (n = 103), followed by rotavirus (n = 51), sapovirus (n = 50), classical astrovirus (n = 43), novel astroviruses (n = 42), and norovirus (n = 26). Novel astroviruses were present in 13.3% of virus-positive cases. Most novel astroviruses were found in children <2-year-old (30/39 children, 77%, p = 0.01) and were found in co-infection (66%). Only classical astroviruses demonstrated significant differences in the Cq values during mono-infections compared to co-infections. In conclusion, common enteric viruses may be frequently found in children with undiagnosed gastroenteritis, indicating the need to implement more sensitive diagnostic methods. Novel astroviruses circulate in the community and could be the cause of gastroenteritis among young children

Linking digital surveillance and in-depth virology to study clinical patterns of viral respiratory infections in vulnerable patient populations

To improve the identification and management of viral respiratory infections, we established a clinical and virologic surveillance program for pediatric patients fulfilling pre-defined case criteria of influenza-like illness and viral respiratory infections. The program resulted in a cohort comprising 6,073 patients (56% male, median age 1.6 years, range 0–18.8 years), where every patient was assessed with a validated disease severity score at the point-of-care using the ViVI ScoreApp. We used machine learning and agnostic feature selection to identify characteristic clinical patterns. We tested all patients for human adenoviruses, 571 (9%) were positive. Adenovirus infections were particularly common and mild in children ≥1 month of age but rare and potentially severe in neonates: with lower airway involvement, disseminated disease, and a 50% mortality rate (n = 2/4). In one fatal case, we discovered a novel virus: HAdV-80. Standardized surveillance leveraging digital technology helps to identify characteristic clinical patterns, risk factors, and emerging pathogens.Peer Reviewe

Molecular Epidemiology of Acute Infectious Diarrhoea in Paediatric Cases at the Winnipeg Children’s Hospital Emergency Room

Acute infectious diarrhea (AcID) causes a significant health burden on the population of developed countries, and even a higher burden in the developing world. The only investigation into the causes of AcID in Winnipeg was conducted in the late 1970s. That study determined that 3%-5% of Children’s Hospital Emergency Room (CHER) visits were due to AcID, and about 50% of those were due to rotavirus infections. We conducted a prospective case-control study to reveal the current spectrum of viral pathogens associated with AcID and the distribution and frequency of their occurrence among the pediatric population of Winnipeg. In this study, nucleic acid detection (NAD) and genome sequence information confirmed the presence and identity of each pathogen, and established whether an etiological shift in the distribution of pathogens, both between families and strains of specific pathogens, occurred. Stool samples were collected from pediatric cases with AcID at the ER along with asymptomatic cases for control. A panel of viral nucleic acid detection (NAD) assays was established by the Viral Gastroenteritis Study Group for human astro, calici (Noro and Sapo), entro, polio, hepA, rota and reo viruses according to the published procedures. A new assay for Aichivirus was developed, and the VGSG has established a novel rotavirus assay which is capable of detecting rotavirus from at least 4 different host species (Human, bovine, porcine and simian). Amplified viral targets were sequenced and the information submitted to GeneBank to confirm the strain of each isolate. A total of 1128 patients visited WCHER and WC during the study period and among them 242 patients were enrolled. In 104 cases viruses were identified. A total number of 114 viruses were identified either by NAD or EM assay. Out of 114 viruses, prevalence of HAdV, NoV GI/II and HRV were 44%, 23% and 23% respectively. Mixed infections were found in 4% of cases.This knowledge of pathogen distribution will facilitate design of effective methods for prevention, treatment and intervention in the spread of AcID pathogens.February 201