Virus Excretion Research Articles

Beak and feather disease virus (BFDV) prevalence, load and excretion in seven species of wild caught common Australian parrots.

Pathogens pose a major risk to wild host populations, especially in the face of ongoing biodiversity declines. Beak and feather disease virus (BFDV) can affect most if not all members of one of the largest and most threatened bird orders world-wide, the Psittaciformes. Signs of disease can be severe and mortality rates high. Its broad host range makes it a risk to threatened species in particular, because infection can occur via spill-over from abundant hosts. Despite these risks, surveillance of BFDV in locally abundant wild host species has been lacking. We used qPCR and haemagglutination assays to investigate BFDV prevalence, load and shedding in seven abundant host species in the wild in south-east Australia: Crimson Rosellas (Platycercus elegans), Eastern Rosellas (Platycercus eximius), Galahs (Eolophus roseicapillus), Sulphur-crested Cockatoos (Cacatua galerita), Blue-winged Parrots (Neophema chrysostoma), Rainbow Lorikeets (Trichoglossus moluccanus) and Red-rumped Parrots (Psephotus haematonotus). We found BFDV infection in clinically normal birds in six of the seven species sampled. We focused our analysis on the four most commonly caught species, namely Crimson Rosellas (BFDV prevalence in blood samples: 41.8%), Sulphur-crested Cockatoos (20.0%), Blue-winged Parrots (11.8%) and Galahs (8.8%). Species, but not sex, was a significant predictor for BFDV prevalence and load. 56.1% of BFDV positive individuals were excreting BFDV antigen into their feathers, indicative of active viral replication with shedding. Being BFDV positive in blood samples predicted shedding in Crimson Rosellas. Our study confirms that BFDV is endemic in our study region, and can inform targeted disease management by providing comparative data on interspecies variation in virus prevalence, load and shedding.

Reflections on COVID-19 nasopharyngeal, fecal and peritoneal swabs in an infant with Wilms tumor: A case report

Proximal Sars-Cov-2 pandemic had radically changed the way surgeons work in many departments, forcing to reserve surgical treatment only for emergency and oncologic cases. We report a case of a ten months-old girl with right-sided Wilms tumor and a previous diagnosis of Sars-Cov-2 infection, who underwent open right nephrectomy. Surgery was planned after negativization of five nasopharyngeal tests, despite the simultaneous positivity of two rectal swabs. The procedure was performed safely with appropriate personal protective equipment (PPE). To better investigate viral excretion, the anesthetist repeated nasopharyngeal swab under general anesthesia, which resulted positive. At the same time, two peritoneal swabs were collected and showed the absence of the virus in the peritoneal fluid. This case highlights the importance of combining swabs from various sources to increase sensibility of the test. The value of nasopharyngeal swab under general anesthesia should be reinforced as it can result positive even after many negative tests. Very little is known about transmission of the virus through the peritoneum as both presence and absence of Sars-Cov-2 have been reported in the peritoneal fluid. Next literature will clarify which particular conditions determine viral penetration in this anatomical district.

SARS-CoV-2 infection (COVID-19): what can we expect?

- Case numbers in China are clearly declining, case numbers in many European regions are no longer increasing exponentially.- Data on mortality from SARS-CoV-2 infection are contradictory; mortality is certainly lower than for SARS and MERS, but probably higher than for most seasonal flu outbreaks in recent years- The main complication of SARS-CoV-2 infection is pneumonia with development of acute respiratory distress syndrome (ARDS)- Asymptomatic and oligosymptomatic courses with virus shedding are not uncommon; they may be more frequent in children than in adults. Virus excretion in asymptomatic people and in the pre-symptomatic phase of an infection is relevant for transmission- An effective antiviral therapy has not yet been established. Steroids for anti-inflammatory therapy are not recommended- It is very important to prepare all actors in the health care system for a longer-term burden of inpatients and complications and to create the necessary capacities. Low-threshold diagnostic testing and rapid detection of infection chains remain essential for better control of the pandemic. An effective vaccine is urgent.

Detection of Antibodies Against Canine Circovirus in Naturally and Experimentally Infected Canines by Recombinant Capsid Enzyme-Linked Immunosorbent Assay

Canine circovirus (CanineCV), a new pathogen, was found to be associated with canine hemorrhagic diarrhea, vasculitis, granulomatous lymphadenitis, and acute gastroenteritis. Although CanineCV was highly positive rate in diarrhea cases, its pathogenicity remains controversial. In this study, the seroprevalence and associated risk factors of CanineCV infection among domestic dogs in northeastern China was investigated by an indirect enzyme-linked immunosorbent assay (iELISA) based on recombinant capsid protein. Results revealed the proposed iELISA had no cross-reactivity with other related pathogens, and yielded good diagnostic values. Then, to evaluate the rCap iELISA, this study applied it to detect antibodies against CanineCV in 1,047 clinical serum samples obtained from northeastern China in 2016–2017. Results showed the positive rates in the five cities of Jilin, Liaoning, and Heilongjiang provinces ranged from 22.22 to 42.29%. Statistical analysis shows a significant difference in age between dogs <3 months old with respect to the >1-year-old dogs (p = 0.005), that is, the CanineCV infection was more frequently identified from older dogs. In the artificially infected experiment, the dogs developed seroconversion after 9 or 12 days and the main way of virus excretion was through feces. More interestingly, among the 32 ELISA-positive serum samples, 34.75% samples tested positive for the CanineCV DNA by qPCR, far higher than that in ELISA-negative serum samples (5.26%, 2/38). This report is the first to demonstrate that CanineCV infection is common in the dog population in northeastern China. The results showed obvious differences in the positive rate associated with diarrhea, age, but not with different cities. This study also provide basis for evaluating the pathogenic potential of CanineCV. But, the pathogenicity, the relationship between antibody level and immune protection, and the harmful effects of this virus remain to be established.

Foot-and-Mouth Disease Surveillance Using Pooled Milk on a Large-Scale Dairy Farm in an Endemic Setting.

Pooled milk is used for the surveillance of several diseases of livestock. Previous studies demonstrated the detection of foot-and-mouth disease virus (FMDV) in the milk of infected animals at high dilutions, and consequently, the collection of pooled milk samples could be used to enhance FMD surveillance. This study evaluated pooled milk for FMDV surveillance on a large-scale dairy farm that experienced two FMD outbreaks caused by the A/ASIA/G-VII and O/ME-SA/Ind-2001d lineages, despite regular vaccination and strict biosecurity practices. FMDV RNA was detected in 42 (5.7%) of the 732 pooled milk samples, and typing information was concordant with diagnostic reports of clinical disease. The FMDV positive milk samples were temporally clustered around reports of new clinical cases, but with a wider distribution. For further investigation, a model was established to predict real-time RT-PCR (rRT-PCR) CT values using individual cattle movement data, clinical disease records and virus excretion data from previous experimental studies. The model explained some of the instances where there were positive results by rRT-PCR, but no new clinical cases and suggested that subclinical infection occurred during the study period. Further studies are required to investigate the effect of vaccination on FMDV excretion in milk, and to evaluate more representative sampling methods. However, the results from this pilot study indicate that testing pooled milk by rRT-PCR may be valuable for FMD surveillance and has provided evidence of subclinical virus infection in vaccinated herds that could be important in the epidemiology of FMD in endemic countries where vaccination is used.

Sapovirus detection and quantification in fecal samples from allogeneic hematopoietic stem cell transplant recipients.

Sapovirus are important agents of acute gastroenteritis (AGE) and they are associated with outbreaks and sporadic cases worldwide. They infect people of all ages, but mainly children, the elderly and immunocompromised individuals are affected. The aim of this study was investigate sapovirus and to determine viral loads in fecal samples from patient undergoing allogeneic hematopoietic stem cell transplantation (allo-HSCT). Fecal samples were submitted to extraction of the genetic material using a commercial kit, and RT-qPCR TaqMan was used for sapovirus screening and determination of viral loads, using a standard curve with serial dilutions of a recombinant plasmid. Positive samples were sequence by Sanger method. Sapovirus was detected in one patient, 5.3% (1/19). Viral excretion lasted for 16days. Viral load varied from 1.73 × 106 to 8.97 × 106GC/g. One of the positive samples was characterized as GI.1 genotype. This is the first study to determine sapovirus loads in samples from allo-HSCT and to identify GI.1 genotype in immunocompromised patients.

Initial data from the COVID-19 Global Rheumatology Alliance provider registries.

Milena Gianfrancesco and colleagues1Gianfrancesco MA Hyrich KL Gossec L et al.Rheumatic disease and COVID-19: initial data from the COVID-19 Global Rheumatology Alliance provider registries.Lancet Rheumatol. 2020; (published online April 16.)https://doi.org/10.1016/S2665-9913(20)30095-3Summary Full Text Full Text PDF Scopus (162) Google Scholar underlined the potential use of disease-modifying drugs, such as hydroxychloroquine, for coronavirus disease 2019 (COVID-19) treatment, but I was struck by the absence of information about long-term hydroxychloroquine treatment among the reported patients.1Gianfrancesco MA Hyrich KL Gossec L et al.Rheumatic disease and COVID-19: initial data from the COVID-19 Global Rheumatology Alliance provider registries.Lancet Rheumatol. 2020; (published online April 16.)https://doi.org/10.1016/S2665-9913(20)30095-3Summary Full Text Full Text PDF Scopus (162) Google Scholar Antimalarial drugs have raised hopes and fierce debate among scientific and public communities worldwide, rapidly creating a political dimension.2Flinders M Coronavirus and the politics of crisis fatigue. The Conversation.http://theconversation.com/coronavirus-and-the-politics-of-crisis-fatigue-134702Date: March 25, 2020Date accessed: April 20, 2020Google Scholar Use of such drugs is a concern for both clinicians and the general population on different levels. It is unknown whether patients with inflammatory rheumatic disease under treatment are more at risk for infection or serious disease due to severe acute respiratory syndrome coronavirus 2 than the general population. The aim of the COVID-19 Global Rheumatology Alliance provider registries is to determine such features. Beyond this primary objective, description and understanding of the determinants of COVID-19 in patients taking immunomodulatory or immunosuppressive drugs (such as hydroxychloroquine and IL-6 receptor antagonists) might help with understanding the disease pathophysiology to prevent the appearance or decrease the severity of COVID-19 pneumonia or neurological events. It is critical to indicate in further reports the occurrence of clinically significant disease in patients treated with hydroxychloroquine, as this drug could be useful in preventing severe disease or minimising the viral excretion period. On April 6, a letter in the Annals of the Rheumatic Diseases stated that “hydroxychloroquine use might be an explanation for no report on systemic lupus erythematosus patients with COVID-19”,3Joob B Wiwanitkit V SLE, hydroxychloroquine and no SLE patients with covid-19: a comment.Ann Rheum Dis. 2020; (published online April 15.)DOI:10.1136/annrheumdis-2020-217506Crossref PubMed Scopus (20) Google Scholar showing the confidence clinicians might place in incomplete data. I declare no competing interests. Initial data from the COVID-19 Global Rheumatology Alliance provider registries – Authors' replyWe agree that information regarding long-term use of hydroxychloroquine and coronavirus disease 2019 (COVID-19) outcomes is needed. When reporting a case to the COVID-19 Global Rheumatology Alliance registry, providers are asked to report on various medications taken before the patient had COVID-19. Although specific details on individual medications might not have been included in our Comment,1 we report the percentage of cases on a number of immunomodulatory and immunosuppressive drugs, including antimalarials, on a weekly basis to our website , Twitter account , and mailing list, so these data are widely accessible and regularly updated. Full-Text PDF Rheumatic disease and COVID-19: initial data from the COVID-19 Global Rheumatology Alliance provider registriesIndividuals with inflammatory rheumatic disease require special consideration with regard to coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Many of these individuals are considered at-risk for serious infections due to their immunocompromised state resulting from their underlying immune conditions and use of targeted immune-modulating therapies such as biologics.1–4 However, some disease-modifying drugs commonly used to treat rheumatic diseases, such as hydroxychloroquine, are being investigated as potential therapies for COVID-19. Full-Text PDF

First viral replication of Covid-19 identified in the peritoneal dialysis fluid of a symptomatic patient

RETRACTED : the authors retracted see : https://bdd.rdplf.org/index.php/bdd/article/view/54713&#x0D; The COVID-19 pandemic is characterized by a disease with mainly respiratory tropism and varying severity.&#x0D; Viral excretion of COVID-19 has been described in both urine and stool with the risk of contamination by stool. No viral replication in the peritoneal dialysis fluid has been reported to date.&#x0D; We report an observation demonstrating the presence of the virus in the peritoneal dialysis drainage fluid of a COVID-19 patient. This underlines the importance in COVID-19 patients of considering dialysis fluid as a possible source of contamination.

Epidemiologic and clinical characteristics of 10 children with coronavirus disease 2019 in Changsha, China

BackgroundThe outbreak of a new coronavirus, first reported in Wuhan, China, is spreading around the world. Information on the characteristics of children with Coronavirus Disease 2019 (COVID-19) is limited. MethodsIn this retrospective study, we recruited 10 children infected with SARS-COV-2 from January 27 to March 10, 2020, in Changsha, China. We report the epidemiological, clinical, laboratory, and high-resolution CT findings for these children. Qualitative descriptive analysis was used to describe the key results. ResultsTen children were included. Three were male and seven were female. Three were from Wuhan, Hubei Province, and seven were from Changsha. All had a history of close contact with adults with COVID-19 before the onset of disease. Clinical manifestations included fever in four cases, respiratory symptoms in three cases, febrile convulsions in one case, vomiting in one case, abdominal pain in one case, and asymptomatic infection in two cases. All the children tested positive for nucleic acid in throat swabs at admission. Stool swabs of three cases were positive for nucleic acid after several days of fever. In nine children, blood routine results were normal, whereas in one case the white blood cell count was elevated. In four cases, CT findings of the lungs showed light ground-glass opacities, one case showed changes similar to bronchopneumonia, and the remaining cases were normal. All were treated with symptomatic support without complications. ConclusionOur findings indicate that intrafamily transmission may be the main form of transmission of COVID-19 in children, and persistent intestinal excretion of virus is another characteristic among children. The results of stool swab tests should be considered for discharge and release from isolation.

Molecular detection, histopathological analysis, and immunohistochemical characterization of equine infectious anemia virus in naturally infected equids.

Equine infectious anemia (EIA), a disease caused by equine infectious anemia virus (EIAV), is considered an obstacle to the development of the horse industry. There is no treatment or vaccine available for EIA, and its pathogenesis, as well as the immune response against the virus, is not fully understood. Therefore, an immunohistochemistry assay was developed for the detection of viral antigens in tissues of equids naturally infected with EIAV. Sections of organs of six equids from Apodi-RN, Brazil, that tested positive for EIA by serological tests (ELISA and AGID) were fixed in 10% formalin solution and embedded in paraffin. Immunohistochemistry was performed using a polyclonal anti-EIAV antibody. EIAV antigens were observed in red spleen pulp cells and hepatic sinusoids, as well as bronchiolar and alveolar epithelial cells of the lungs and proximal and distal tubules of the kidneys. The presence of EIAV in the spleen and liver was expected due to viral tropism by macrophages, which are abundantly present in these organs. However, EIAV was also found in lung and kidney epithelial cells, indicating that the virus infects cell types other than macrophages. In conclusion, the immunohistochemical assay standardized in this study was able to detect EIAV antigens in spleen, liver, kidney and lung cells from naturally infected EIAV equids. Immunostaining observed in the spleen confirms viral tropism by mononuclear phagocytes; however, the presence of EIAV in lung and kidney epithelial cells indicates that virus may be eliminated in urine and/or oronasal secretions, suggesting new routes for viral excretion.

Insertion of Basic Amino Acids in the Hemagglutinin Cleavage Site of H4N2 Avian Influenza Virus (AIV)-Reduced Virus Fitness in Chickens is Restored by Reassortment with Highly Pathogenic H5N1 AIV.

Highly pathogenic (HP) avian influenza viruses (AIVs) are naturally restricted to H5 and H7 subtypes with a polybasic cleavage site (CS) in hemagglutinin (HA) and any AIV with an intravenous pathogenicity index (IVPI) ≥ 1.2. Although only a few non-H5/H7 viruses fulfill the criteria of HPAIV; it remains unclear why these viruses did not spread in domestic birds. In 2012, a unique H4N2 virus with a polybasic CS 322PEKRRTR/G329 was isolated from quails in California which, however, was avirulent in chickens. This is the only known non-H5/H7 virus with four basic amino acids in the HACS. Here, we investigated the virulence of this virus in chickens after expansion of the polybasic CS by substitution of T327R (322PEKRRRR/G329) or T327K (322PEKRRKR/G329) with or without reassortment with HPAIV H5N1 and H7N7. The impact of single mutations or reassortment on virus fitness in vitro and in vivo was studied. Efficient cell culture replication of T327R/K carrying H4N2 viruses increased by treatment with trypsin, particularly in MDCK cells, and reassortment with HPAIV H5N1. Replication, virus excretion and bird-to-bird transmission of H4N2 was remarkably compromised by the CS mutations, but restored after reassortment with HPAIV H5N1, although not with HPAIV H7N7. Viruses carrying the H4-HA with or without R327 or K327 mutations and the other seven gene segments from HPAIV H5N1 exhibited high virulence and efficient transmission in chickens. Together, increasing the number of basic amino acids in the H4N2 HACS was detrimental for viral fitness particularly in vivo but compensated by reassortment with HPAIV H5N1. This may explain the absence of non-H5/H7 HPAIV in poultry.

Between roost contact is essential for maintenance of European bat lyssavirus type-2 in Myotis daubentonii bat reservoir: \u2018The Swarming Hypothesis\u2019

Many high-consequence human and animal pathogens persist in wildlife reservoirs. An understanding of the dynamics of these pathogens in their reservoir hosts is crucial to inform the risk of spill-over events, yet our understanding of these dynamics is frequently insufficient. Viral persistence in a wild bat population was investigated by combining empirical data and in-silico analyses to test hypotheses on mechanisms for viral persistence. A fatal zoonotic virus, European Bat lyssavirus type 2 (EBLV-2), in Daubenton’s bats (Myotis daubentonii) was used as a model system. A total of 1839 M. daubentonii were sampled for evidence of virus exposure and excretion during a prospective nine year serial cross-sectional survey. Multivariable statistical models demonstrated age-related differences in seroprevalence, with significant variation in seropositivity over time and among roosts. An Approximate Bayesian Computation approach was used to model the infection dynamics incorporating the known host ecology. The results demonstrate that EBLV-2 is endemic in the study population, and suggest that mixing between roosts during seasonal swarming events is necessary to maintain EBLV-2 in the population. These findings contribute to understanding how bat viruses can persist despite low prevalence of infection, and why infection is constrained to certain bat species in multispecies roosts and ecosystems.

SARS-CoV-2 infections with emphasis on pediatric patients: a narrative review.

ABSTRACTThis narrative review summarizes the main aspects underlying the new coronavirus SARS-CoV-2, its epidemiology, pathophysiology, pointing to differences of SARS-CoV-2 main receptors ACE2, in terms of expression and the amount of soluble ACE2 in the circulation of children, men and women, and also in those with risk factors such as the smokers and pregnant women or presenting with comorbidities (diabetes, obesity, hypertension and other cardiovascular diseases, renal and CNS pre-existing diseases). Clinical manifestations in adults and children were also described, emphasizing the particularities already seen in children, regarding signs, symptoms, viral excretion time and the involvement of all organs and systems. The COVID-19 in the pediatric population was divided into two sections: one dedicated to previously healthy children and adolescents with COVID-19, and the other to those who live with comorbidities and acquired COVID-19. A few paragraphs were reserved to the recently described severe multisystemic inflammatory syndrome associated with COVID-19 (MIS-C) that shares certain characteristics with Kawasaki disease. Some studies on the infection in pregnant and postpartum women, as well as neonates were shown. This review has also covered the laboratory diagnosis of COVID-19, passing through the imaging diagnosis made by the chest tomography revealing ground glass patching opacities, and results of non-specific exams such as the total blood with lymphopenia, the coagulation tests with increased prothrombin times, as well as marked increments of the D-dimer, troponin and proinflammatory cytokines. In the section devoted to the specific laboratory diagnosis of COVID-19, the most used RT-PCR protocols were described and some studies on the serological diagnosis with IgA, IgM and IgG detection were detailed, including the use of rapid immunochromatographic assays and discussing the ideal period after the onset of symptoms to perform each type of test. In the end, the management of pediatric patients with COVID-19 based mainly on supportive measures has been briefly commented.

Analysis of COVID-19 spreading and prevention strategy in schools based on continuous infection model

After the COVID-19 epidemic leveled off in China, many provinces have started to resume schooling. Long-term contact between students and teachers in such a closed environment in schooling can increase the possibility of the outbreak. Although the school closure can effectively alleviate the epidemic, large-scale students’ isolation not only causes social panic but also brings huge social and economic burden, so before the emergence of school epidemics, one should select and adopt more scientific prevention and control measures. In this study, according to the virus excretion of COVID-19 patients in the disease period, the infectious capacity of patients is redefined. After introducing it into the traditional suspected-exposed-infected-removed (SEIR) model, a continuous infection model that is more consistent with the actual transmission of COVID-19 patients is proposed. Secondly, the effective distance between students is calculated through real contact data. Based on the analysis of the effective distance, three types of isolation area prevention and control measures are proposed and compared with the recently proposed digital contact tracking prevention and control measures. Simulating the spread of COVID-19 in schools through real student contact data and continuous infection models, in order to compare the preventions and control effects of various prevention and control measures in the school epidemic situation, and evaluating the social influence of measures by accumulating the number of quarantines when prevention and control measures are adopted, we find that the COVID-19 can lead the cases to happen on a larger scale in the continuous infection model than in the traditional SEIR model, and the prevention and control measures verified in the continuous infection model are more convincing. Using digital contact tracking prevention and control measures in schools can achieve similar results to those in closed schools with the smallest number of quarantines. The research in this paper can help schools choose appropriate prevention and control measures, and the proposed continuous infection model can help researchers more accurately simulate the spread of COVID-19.

HIV+ Status and Cervical Cancer: Cytological Aspects of Cervical Smear in Cameroon Setting

Introduction: The early detection of precancerous lesions being very important for the preventive management of cervical cancer, we felt it was important to identify these lesions on potential backgrounds including HIV-positive (HIV+) women to suggest control strategies of cervical cancer in Cameroon. Objective: To determine the prevalence of precancerous lesions in women infected with the human immunodeficiency virus (HIV), to investigate the determinants of the HIV and cervical cancer association, and to make recommendations regarding cervical cancer screening in these patients of the city of Douala. Methodology: Our study was a case-control cross-sectional study from July 2017 to December 2017 (6 months) including 108 women among which 34 HIV+ matched with 74 HIV-. HIV serology was done using the complete HIV enzygnost test. Cervical smears for cytological lesions were fixed to the cyto-fixator and then stained by the Papanicolaou technique and read under an optical microscope. The cervical smear slides for viral excretion were fixed with a methanol-acetone mixture of equal volume; HPV testing was done by the indirect immune-peroxidase technique using P16 protein. The excretion of HSV type 1 & 2 was investigated by the indirect immunofluorescence technique using the Simplex Virus type 1 & 2 Rabbit anti-Herpes from DAKO (France). Results: The two groups of women were compared with the chi square test with a significance threshold of P 0.05. The average age was 40.07 with extremes of 21 and 71 years and a standard deviation of 9.99. Of the 34 HIV+ patients, 23 had an abnormal cervix compared to 36 cases of abnormal cervix among HIV- with a statistically significant difference (P = 0.006649). 12 cases of dysplasia were observed in the 34 HIV+ women and distributed as follows: 0% of mild dysplasia, 18.92% of moderate dysplasia and 13.51% of severe dysplasia. In HIV- women we detected 6 cases of dysplasia including 1.35% of mild dysplasia, 4.05% of moderate dysplasia and 2.70% of severe dysplasia. Regarding HPV infection, we observed 21 cases of HPV among 34 HIV+ women (61.76%) against 23 cases in HIV-lesions remains high in HIV+ women, hence the need to include routine screening for precancerous lesions in all HIV+ women at all ages, as well as the routine search for HPV excretion in all those with cellular dysplasia.

Productive Infection of Mouse Mammary Glands and Human Mammary Epithelial Cells by Zika Virus.

Zika virus (ZIKV) belongs to the large category of arboviruses. Surprisingly, several human-to-human transmissions of ZIKV have been notified, either following sexual intercourse or from the mother to fetus during pregnancy. Importantly, high viral loads have been detected in the human breast milk of infected mothers, and the existence of breastfeeding as a new mode of mother-to-child transmission of ZIKV was recently hypothesized. However, the maternal origin of infectious particles in breast milk is currently unknown. Here, we show that ZIKV disseminates to the mammary glands of infected mice after both systemic and local exposure with differential kinetics. Ex vivo, we demonstrate that primary human mammary epithelial cells were sensitive and permissive to ZIKV infection in this study. Moreover, by using in vitro models, we prove that mammary luminal- and myoepithelial-phenotype cell lines are both able to produce important virus progeny after ZIKV exposure. Our data suggest that the dissemination of ZIKV to the mammary glands and subsequent infection of the mammary epithelium could be one mechanism of viral excretion in human breast milk.

A Bivalent Live-Attenuated Vaccine for the Prevention of Equine Influenza Virus.

Vaccination remains the most effective approach for preventing and controlling equine influenza virus (EIV) in horses. However, the ongoing evolution of EIV has increased the genetic and antigenic differences between currently available vaccines and circulating strains, resulting in suboptimal vaccine efficacy. As recommended by the World Organization for Animal Health (OIE), the inclusion of representative strains from clade 1 and clade 2 Florida sublineages of EIV in vaccines may maximize the protection against presently circulating viral strains. In this study, we used reverse genetics technologies to generate a bivalent EIV live-attenuated influenza vaccine (LAIV). We combined our previously described clade 1 EIV LAIV A/equine/Ohio/2003 H3N8 (Ohio/03 LAIV) with a newly generated clade 2 EIV LAIV that contains the six internal genes of Ohio/03 LAIV and the HA and NA of A/equine/Richmond/1/2007 H3N8 (Rich/07 LAIV). The safety profile, immunogenicity, and protection efficacy of this bivalent EIV LAIV was tested in the natural host, horses. Vaccination of horses with the bivalent EIV LAIV, following a prime-boost regimen, was safe and able to confer protection against challenge with clade 1 (A/equine/Kentucky/2014 H3N8) and clade 2 (A/equine/Richmond/2007) wild-type (WT) EIVs, as evidenced by a reduction of clinical signs, fever, and virus excretion. This is the first description of a bivalent LAIV for the prevention of EIV in horses that follows OIE recommendations. In addition, since our bivalent EIV LAIV is based on the use of reverse genetics approaches, our results demonstrate the feasibility of using the backbone of clade 1 Ohio/03 LAIV as a master donor virus (MDV) for the production and rapid update of LAIVs for the control and protection against other EIV strains of epidemiological relevance to horses.

Dynamics and Differences in Systemic and Local Immune Responses After Vaccination With Inactivated and Live Commercial Vaccines and Subsequent Subclinical Infection With PRRS Virus.

The goals of our study were to compare the immune response to different killed and modified live vaccines against PRRS virus and to monitor the antibody production and the cell mediated immunity both at the systemic and local level. In the experiment, we immunized four groups of piglets with two commercial inactivated (A1—Progressis, A2—Suivac) and two modified live vaccines (B3—Amervac, B4—Porcilis). Twenty-one days after the final vaccination, all piglets, including the control non-immunized group (C5), were i.n., infected with the Lelystad strain of PRRS virus. The serum antibody response (IgM and IgG) was the strongest in group A1 followed by two MLV (B3 and B4) groups. Locally, we demonstrated the highest level of IgG antibodies in bronchoalveolar lavages (BALF), and saliva in group A1, whereas low IgA antibody responses in BALF and feces were detected in all groups. We have found virus neutralization antibody at DPV 21 (days post vaccination) and higher levels in all groups including the control at DPI 21 (days post infection). Positive antigen specific cell-mediated response in lymphocyte transformation test (LTT) was observed in groups B3 and B4 at DPV 7 and in group B4 at DPV 21 and in all intervals after infection. The IFN-γ producing lymphocytes after antigen stimulation were found in CD4−CD8+ and CD4+CD8+ subsets of all immunized groups 7 days after infection. After infection, there were obvious differences in virus excretion. The virus was detected in all groups of piglets in serum, saliva, and occasionally in feces at DPI 3. Significantly lower virus load was found in groups A1 and B3 at DPI 21. Negative samples appeared at DPI 21 in B3 group in saliva. It can be concluded that antibodies after immunization and infection, and the virus after infection can be detected in all the compartments monitored. Immunization with inactivated vaccine A1—Progressis induces high levels of antibodies produced both systemically and locally. Immunization with MLV-vaccines (Amervac and Porcilis) produces sufficient antibody levels and also cell-mediated immunity. After infection virus secretion gradually decreases in group B3, indicating tendency to induce sterile immunity.

Characterization of minimal lesions related to the presence of visna/maedi virus in the mammary gland and milk of dairy sheep

BackgroundIn order to characterize the complete range of lesions, especially minimal, affecting mammary gland and viral antigen distribution and target cells using immunohistochemistry in naturally Visna/maedi (VM) 84 infected sheep were studied, forty-four from flocks with clinical cases (A) and 35 randomly sampled from two abattoirs (B) together with five negative controls (C). An immunocytochemistry technique was developed and further milk samples (n = 39) were used to study viral excretion, carrier cells and the role of milk and colostrum in the transmission of the disease.ResultsAll sheep from group C and three sheep from group B were negative to VM in tissue sections by histopathology, immunohistochemistry and PCR, and also in serum using ELISA. Several degrees of CD3 + lymphocytic interstitial mastitis were observed in groups A and B: minimal (+) n = 26 sheep; moderate (++), n = 32 and severe (+++), n = 12. No differences in lesion distribution were observed between groups A and B. Viral presence was confirmed by immunohistochemistry using two different antibodies and/or PCR in every tissue with lesions while serology was negative in six sheep with lesions. Two milk samples taken from milk tanks from two flocks from group A and fourteen milk samples from 29 infected sheep from group B were positive to VM (most of them from animals with moderate and severe lesions). Positivity was only found in macrophages, even in focal and minimal lesions, while no positivity was observed in epithelial or any other cells in either tissue and milk samples.ConclusionsThis new observation of the minimal lesions described in this work increased the prevalence of VM lesions in mammary gland up to 90.9% and VM should be considered as a differential diagnosis when minimal interstitial lesions are detected. A high prevalence of VM was observed in intensive milk-producing sheep, ELISA serology did not detect as positivity all infected animals, while histology, IHC or PCR showed higher sensitivity. The cytological technique developed was very useful in milk-cell studies using hematoxylin and eosin and immunocytochemistry. Viral detection in milk samples (16/39) confirms a potential but limited role of milk/colostrum in viral transmission.

Development of a PPRV challenge model in goats and its use to assess the efficacy of a PPR vaccine

Peste des Petits Ruminants (PPR) is a severe disease of small ruminants and has high economic impacts in developing countries. Endemic in Africa, the Middle East and Asia, the disease is currently progressing with occurrences reported in North Africa, Turkey and in Georgia, and now threatens Europe. Much remains unknown about the infection dynamics, the virulence of the different strains and species/breed susceptibility. Robust experimental challenge models are needed to explore these fields and to confirm the efficacy of currently sold vaccines. We first assessed virulence of two PPR virus strains (CI89 and MA08) in Saanen goats. Whereas the MA08 strain led to classical severe clinical signs of PPR, the CI89 strain appeared to cause a mild disease in Saanen goats, highlighting the difference in virulence between strains in this animal model. We further demonstrated the importance of the inoculation route in the appearance of clinical signs and that ocular excretion is a better choice than blood for viral detection. After developing a robust challenge model, we assessed the efficacy of a vaccine (PPR-VAC®, BVI Botswana) against the MA08 strain and demonstrated that this vaccine blocked viral excretion and significantly reduced clinical signs. These results reinforce the paradigm that a strain from one lineage could protect against strains from other lineages.