Co-infection of human parvovirus B19 with Plasmodium falciparum contributes to malaria disease severity in Gabonese patients.

BACKGROUND: High seroprevalence of parvovirus B19 (B19V) coinfection with Plasmodium falciparum has been previously reported. However, the impact of B19V-infection on the clinical course of malaria is still elusive. In this study, we investigated the prevalence and clinical significance of B19V co-infection in Gabonese children with malaria. METHODS: B19V prevalence was analyzed in serum samples of 197 Gabonese children with P. falciparum malaria and 85 healthy controls using polymerase chain reaction (PCR), enzyme-linked immunosorbent assay (ELISA), and direct DNA-sequencing. RESULTS: B19V was detected in 29/282 (10.28%) of Gabonese children. B19V was observed more frequently in P. falciparum malaria patients (14.21%) in comparison to healthy individuals (1.17%) (P<0.001). Notably, the mild-malaria group revealed significantly lower hematocrit levels in B19V/P. falciparum co-infection than in P. falciparum mono-infection (P<0.05). Genetic analysis revealed a predominance of B19V genotype-1 (71.43%) in the studied population. However, B19V-genotype 2 was observed significantly more often in children with severe-malaria than in mild-malaria (P=0.04). CONCLUSION: Our findings reveal that B19V-infection is frequent in Gabonese children with P. falciparum malaria and signifies a possible contribution of B19V on the clinical course of malaria in a genotype-dependent manner. B19V co-infection should be considered as a additional diagnostic measure in malaria patients with life threatening anemia.

Detection, prevalence and analysis of emerging porcine parvovirus infections.

A number of newly identified porcine parvoviruses had been described during the last decade, but the presence and prevalence of these viruses are unknown in Hungary and only partly known for Europe. The present study was conducted to detect and measure the prevalence of these viruses, namely porcine parvovirus (PPV) 2, PPV3, PPV4, porcine bocavirus (PBoV) 1, PBoV2, PBo-likeV and the 6V and 7V parvoviruses. The prevalence of PPV1 and porcine circovirus type 2 (PCV2) was also investigated. Faecal samples, blood serum samples, organ tissues, foetuses and semen were collected from different swine herds in Hungary and tested by polymerase chain reaction methods specific for the different viruses. The results indicated that all of the examined parvoviruses were present in Hungary, hence in Europe. The prevalence was 18.1% for PCV2, 0.5 % for PPV1, 6.4% for PPV2, 9.7% for PPV3, 6.4% for PPV4, 1.5% for PBo-likeV, 4.8% for PBoV1 and PBoV2 and 1.8% for 6V and 7V. Based on the analysis of partial PPV4 and PBo-likeV sequences, these viruses showed a high degree of sequence conservation, whereas PPV3 and the majority of PPV2, PBoV1, PBoV2, 6V and 7V sequences showed higher variability. Possible sites of recombination were also identified between PBoV1 and PBoV2 genomes.

Persistent infection of American bison (Bison bison) with bovine viral diarrhea virus and bosavirus.

Bovine viral diarrhea viruses (BVDV) are significant pathogens of cattle, leading to losses associated with reproductive failure, respiratory disease and immune dysregulation. While cattle are the reservoir for BVDV, a wide range of domestic and wild ruminants are susceptible to infection and disease caused by BVDV. Samples from four American bison (Bison bison) from a captive herd were submitted for diagnostic testing due to their general unthriftiness. Metagenomic sequencing on pooled nasal swabs and serum identified co-infection with a BVDV and a bovine bosavirus. The BVDV genome was more similar to the vaccine strain Oregon C24 V than to other BVDV sequences in GenBank, with 92.7 % nucleotide identity in the open reading frame. The conserved 5'-untranslated region was 96.3 % identical to Oregon C24 V. Bosavirus has been previously identified in pooled fetal bovine serum but its clinical significance is unknown. Sequencing results were confirmed by virus isolation and PCR detection of both viruses in serum and nasal swab samples from two of the four bison. One animal was co-infected with both BVDV and bosavirus while separate individuals were positive solely for BVDV or bosavirus. Serum and nasal swabs from these same animals collected 51 days later remained positive for BVDV and bosavirus. These results suggest that both viruses can persistently infect bison. While the etiological significance of bosavirus infection is unknown, the ability of BVDV to persistently infect bison has implications for BVDV control and eradication programs. Possible synergy between BVDV and bosavirus persistent infection warrants further study.

Seroprevalence of viral and vector-borne bacterial pathogens in domestic dogs (Canis familiaris) in northern Botswana.

BACKGROUND: Domestic dogs (Canis familiaris) have the potential to act as disease reservoirs for wildlife and are important sentinels for common circulating pathogens. Therefore, the infectious disease seroprevalence among domestic dogs in northern Botswana may be indicative of pathogen exposure of various wildlife species. The objective of this study was to assess the seroprevalence of Ehrlichia spp., Borrelia burgdorferi, Anaplasma spp., Dirofilaria immitis, canine adenovirus, canine parvovirus, and canine distemper virus in domestic dogs as proxies of disease prevalence in the local wildlife in the Okavango Delta region of Botswana. Statistical analysis assessed crude and factor-specific seroprevalence proportions in relation to age, sex, and geographical location as predictors of seropositivity. Logistic regression was used to identify adjusted predictors of seropositivity for each of the pathogens of interest. RESULTS: Samples from 233 dogs in a total of seven locations in Maun, Botswana, and surrounding villages were collected and serologically analyzed. No dogs were seropositive for B. burgdorferi, while low seroprevalence proportions were observed for Anaplasma spp. (2.2%) and D. immitis (0.9%). Higher seroprevalence proportions were observed for the tick-borne pathogen Ehrlichia spp. (21.0%), and 19.7% were seropositive for canine adenovirus (hepatitis). The highest seroprevalence proportions were for canine parvovirus (70.0%) and canine distemper virus (44.8%). The predictors of seropositivity revealed that adults were more likely to be seropositive for canine adenovirus, canine distemper virus, and canine parvovirus than juveniles, and location was a risk factor for canine adenovirus, canine distemper virus, canine parvovirus, and Ehrlichia spp. CONCLUSIONS: Results indicate that increasing tick control and vaccination campaigns for domestic dogs may improve the health of domestic animals, and potentially wildlife and humans in the Okavango Delta since viral and vector-borne bacterial pathogens can be transmitted between them.

Intestinal Microbial Dysbiosis in Beagles Naturally Infected with Canine Parvovirus.

Canine parvoviral enteritis (PVE) is an important intestinal disease of the puppies; however, the potential impact of the canine parvovirus (CPV) on the gut microbiota has not been investigated. Therefore, the aim of this study was to evaluate the gut microbial shifts in puppies naturally infected with CPV. Fecal samples were collected from healthy dogs and those diagnosed with PVE at 4, 6, 8, and 12 weeks of age. The distal gut microbiota of dogs was characterized using Illumina MiSeq sequencing of the bacterial 16S rRNA genes. The sequence data were analyzed using QIIME with an Operational Taxonomic Unit definition at a similarity cutoff of 97%. Our results showed that the CPV was associated with significant microbial dysbiosis of the intestinal microbiota. Alpha diversity and species richness and evenness in dogs with PVE decreased compared to those of healthy dogs. At the phylum level, the proportion of Proteobacteria was significantly enriched in dogs with PVE while Bacteroidetes was significantly more abundant in healthy dogs (p < 0.05). In dogs with PVE, Enterobacteriaceae was the most abundant bacterial family accounting for 36.44% of the total bacterial population compared to only 0.21% in healthy puppies. The two most abundant genera in healthy dogs were Prevotella and Lactobacillus and their abundance was significantly higher compared to that of dogs with PVE (p < 0.05). These observations suggest that disturbances of gut microbial communities were associated with PVE in young dogs. Evaluation of the roles of these bacterial groups in the pathophysiology of PVE warrants further studies.

Fecal microbiota transplantation in puppies with canine parvovirus infection.

BACKGROUND: Diarrhea associated with parvovirus infection is common in dogs. Supportive care is the mainstay of treatment, but recovery may be prolonged and mortality rate can be high. Modification of the intestinal bacterial microbiota has been promising in human and veterinary medicine as an adjunctive treatment of various enteric diseases. OBJECTIVES: To investigate the safety and efficacy of fecal microbiota transplantation (FMT) on the clinical recovery of puppies with acute hemorrhagic diarrhea syndrome. ANIMALS: Sixty-six puppies with parvovirus infection were evaluated at 2 veterinary hospitals. METHODS: Randomized clinical trial. Puppies were randomly distributed into 2 groups: standard treatment (STD) and standard treatment + FMT (STD + FMT). The STD puppies (n = 33) received only treatment with IV fluids and antimicrobials and the STD + FMT puppies (n = 33) received FMT in addition to standard treatment. For FMT, 10 g of feces from a healthy dog diluted in 10 mL of saline were administered rectally 6-12 hours post-admission. RESULTS: Among survivors, treatment with FMT was associated with faster resolution of diarrhea (P < .001) and shorter hospitalization time (P = .001; median, 3 days in STD + FMT; median, 6 days in STD) compared to standard treatment. Mortality in STD was 36.4% (12/33) as compared to 21.2% (7/33) in puppies treated with FMT, but there was no statistical difference between groups (P = .174). Polymerase chain reaction indicated that all animals carried canine parvovirus, strain CPV-2b. CONCLUSIONS: Fecal microbiota transplantation in parvovirus-infected puppies was associated with faster resolution of diarrhea.

Characterization of a virus that causes transient aplastic crisis.

Transient aplastic crisis in children with congenital hemolytic anemias has been linked epidemiologically to infection with a serum parvovirus-like virus (SPLV). The virus is found in the blood in the early stages of the crisis, and serum containing SPLV inhibits erythroid colony formation in vitro. After sedimentation of virus-containing sera through a sucrose density gradient, colony inhibitory activity is present in the particulate fraction and separate from serum immunoglobulins. No inhibitory activity can be recovered from convalescent-phase sera after similar fractionation procedures. Inhibition of erythroid colony formation in vitro is not a feature of sera from other viral infections. The pattern of resistance of SPLV activity to chemicals and enzymes is compatible with it being a parvovirus. By using replating techniques, a target of SPLV has been identified as a late erythroid progenitor cell. Neither SPLV antigen nor anti-SPLV IgM was present in the sera of patients with other forms of bone marrow failure.

Serologic and molecular detection of human Parvovirus B19 infection.

Following its identification by Yvonne Cossart in 1975, human Parvovirus B19 has been recognized as the causative agent of a wide range of diseases. In childhood, the most common disease is a typical exanthema called "fifth disease". In adults, viral infection may be responsible for fetal loss and for aplastic anaemia in immuno-compromised patients. Because persistent viral infection may induce an autoimmune response, Parvovirus B19 is emerging as an environmental factor linked to the pathogenesis of autoimmunity. As a result of its expanding disease spectrum, Parvovirus B19 is the subject of intense efforts to clarify the pathogenesis of virus-related disorders as well as improve diagnostic laboratory testing including standardization of serological and nucleic acid-based detection assays. Enzymatic immunoassays based on conformational antigens have proven to be the most important tools for accurate diagnosis in the majority of cases. In other selected clinical cases, the detection of Parvovirus B19 infection can be complemented by PCR and, more recently, by the real-time PCR.

Human parvovirus B19 experimental infection in human fibroblasts and endothelial cells cultures.

With the aim to detect what kind of cells, in addition to erythroid progenitors, could be involved in the pathogenesis of B19 infection in some connective tissue diseases, primary cultures of human fibroblasts (HF) and endothelial cells (HUVEC) were exposed to a B19 positive serum (350 genome copies/cell). The presence of NS1 and VP1 mRNA, in both HF and HUVEC cultures 1, 2 and 6 days after the exposure, indicated infection by B19 virus. However, no significant increase of B19 DNA level in the infected HF and HUVEC cultures was detectable through the entire incubation period of 6 days. It is possible that HF and HUVEC are not permissive for B19 virus replication or, alternatively, that few cells only get infected by B19 virus. HF and HUVEC stimulation with different growth factors or cytokines could be required for a B19 productive infection to occur.

Clinical manifestations of human parvovirus B19 in adults.

Human parvovirus B19 has been associated with various clinical effects in a number of uncontrolled reports. To define the usual manifestations of B19 infection in adults and the factors that influence them we present a clinicoepidemiological study of an outbreak of B19 infection centered on a junior school. Four hundred fifty-three of 475 adults in this community were interviewed and blood was obtained for serological diagnosis. Fifty-four cases of recent infection were identified and were HLA typed. Fourteen of the cases were asymptomatic; 32 had an influenzalike illness; 23 a rash; and 26 an acute-onset polyarthropathy that was more common in women and lasted for up to 7 months. HLA-A, -B, and -C antigen frequencies were similar to a local control population and showed no association with symptoms except that HLA-DR1 was absent in those with persistent arthropathy.

Characterization of acute rat parvovirus infection by in situ hybridization.

In situ hybridization and virus titration were used to characterize early stages of rat virus (RV) infection of rat pups after oronasal inoculation. Results suggest that virus enters through the lung and that early viremia leads rapidly to pantropic infection. Cells derived from all three germ layers were infected with RV, but those of endodermal and mesodermal origin were the predominant targets. Infection of vascular endothelium was widespread and was associated with hemorrhage and infarction in the brain. Convalescence from acute infection was accompanied by mononuclear cell infiltrates at sites containing RV DNA. Viral DNA was also detected in endothelium, fibroblasts and smooth muscle myofibers four weeks after inoculation. Further examination of these cells as potential sites of persistent infection is warranted.

Detection of parvovirus DNA in human serum using biotinylated RNA hybridisation probes.

Methods were established for the detection of parvovirus DNA in human serum using single-stranded RNA probes. The sensitivity of detection of virus using 32P-radiolabelled RNA versus non-radiolabelled biotinylated RNA probes using a streptavidin-polyalkaline phosphatase detection system was compared. Virus was detected using 32P-labelled and biotinylated RNA probes at serum dilutions of 10(-3), equivalent to approx. 3 pg of viral DNA. Using biotinylated RNA probes and a dot-blot system, diagnosis of numerous serum samples could be performed within 8 h of receipt of samples, using an RNA probe which was synthesised and stored at -20 degrees C for up to 12 months without loss of sensitivity. Our work demonstrates the potential of biotinylated RNA probes in the routine analysis of viral sequences in serum.

Human parvovirus infections.

Discovered by chance in 1974, the human serum parvovirus B19 is at present the only recognized, autonomous, pathogenic human parvovirus. For some years following its discovery, B19 was not associated with any defined clinical syndrome; although a high titre viraemia was often noted in infected individuals they were largely asymptomatic. In 1980 the causal association between B19 infection and aplastic crisis in chronic haemolytic anaemia began to emerge with the discovery of B19 as the agent responsible for aplastic crisis in sickle cell anaemia. This fulfilled the expectation of a disease of tissue comprising a large proportion of dividing cells, namely the erythropoietic elements of the bone marrow, anticipated in autonomous parvovirus infection where viral replication is confined to dividing cells. More recently, erythema infectiosum, an illness sharing many of the clinical features of rubella, has been found to be the common result of B19 infection, although a spectrum of disease is now emerging. Much effort is currently directed toward the elucidation of the effects of maternal B19 infection on the developing fetus.

Detection of B19 parvovirus infections by a dot-blot hybridization assay using a digoxigenin-labelled probe.

A non-radioactive dot-blot hybridization assay for the detection of B19 parvovirus infections was developed using a digoxigenin-labelled probe both on nylon and nitrocellulose filters. A 700 bp BamHI HindIII fragment of B19 DNA was used to construct the probe. Probe labelling was carried out by incorporating deoxyuridine triphosphate labelled with digoxigenin. The dot-blot hybridization assay was visualized by an immunoenzymatic reaction using antidigoxigenin Fab fragments labelled with alkaline phosphatase. The specificity and sensitivity of digoxigenin-labelled B19 DNA probe was compared with the results obtained with 32P-labelled B19 DNA probe. Out of the 504 serum samples tested, 3 samples were positive in all the hybridization assays performed and 494 were negative, 7 serum samples gave a weak positive reaction when Dig-B19 probe was used on nitrocellulose filters. The 77 pharyngeal swabs tested were negative in all the hybridization assays performed. Our hybridization assay showed a high sensitivity and reproducibility and it appears to be a rapid, practical and reliable test for routine screening of B19 parvovirus DNA in large numbers of clinical specimens.

IgG and IgM antibodies to human parvovirus B19 in the serum of patients with a clinical diagnosis of infection with the virus and in the general population of Saudi Arabia.

A total of 56 samples of serum from 32 patients with a clinical diagnosis of human parvovirus B19 infection were tested for specific immunoglobulin G (IgG) and M (IgM) antibodies by means of the recently available indirect enzyme-linked immunosorbent assay (ELISA) (Parvoscan-B19, Ferring Diagnostica, Sweden). The assay was also used in order to determine the age-specific prevalence of antibodies to the virus in the general population of Saudi Arabia. Specific IgM antibodies were detected in 94% specimens collected 1 week after the onset of illness and could be detected for up to 2 months. On the other hand, specific IgG antibodies were detected in 85% patients from whom acute- and convalescent-phase serum samples were collected. Saudis begin to be exposed to human parvovirus B19 early in life and prevalence of exposure increases with age in both sexes (overall prevalence 19.0%). The availability of a commercial ELISA makes it possible to diagnose infection with the virus routinely and will help in establishing the extent of exposure to it in various communities.

Unusual characteristics of a parvovirus isolated from a clinically ill steer.

A parvovirus was isolated from the feces of an 8- and 9-month-old steer that died acutely with hemorrhagic diarrhea and microscopic evidence of a coccidial infection. The concurrent intestinal parasitism in this steer appeared to play a role in the development of clinical disease. The viral isolate was identified as a bovine parvovirus (BPV) on the basis of its size (22 nm) and icosahedral morphology, the neutralization of viral cytopathology by antiserum to BPV, a strong immunofluorescent reaction with fluorescein-labeled antiserum to BPV, and the inhibition of viral hemagglutination of guinea-pig erythrocytes by antiserum to BPV. Cell cultures infected with this isolate showed a slight nuclear fluorescent reaction with fluorescein-labeled antiserum to canine parvovirus, suggesting an antigenic relationship to canine parvovirus. Patterns of hemagglutination for this isolate with human erythrocytes from 20 donors of various blood types differed from those obtained with the reference Abinanti strain of BPV. These results indicate that blood from multiple donors of a species may be necessary to confirm the presence or absence of viral hemagglutinating activity with clinical isolates of BPV.

Characterization of biological and antigenic properties of raccoon dog and blue fox parvoviruses: a monoclonal antibody study.

Parvovirus isolates from blue foxes and raccoon dogs were characterized by studying their haemagglutination properties, host range in vitro and antigenic structure. In all 3 characters, raccoon dog parvovirus resembled canine parvovirus (CPV), while blue fox parvovirus was similar to mink enteritis virus (MEV). Monoclonal antibodies (MAbs) were prepared against both viruses. Raccoon dog parvovirus, while resembling CPV, had a unique antigenic site which could be specified by MAbs. The pattern of MAbs prepared against blue fox parvovirus indicated that it is a member of Type 2 MEV.

Comparison of the viral proteins of canine parvovirus-2, mink enteritis virus and feline panleukopenia virus.

Canine parvovirus-2 (CPV-2), Mink enteritis virus (MEV) and feline panleukopenia virus (FPV) were produced using identical cell culture and purification techniques. The distributions of the haemagglutinating activity of the three different parvoviruses in a CsCl gradient were similar with haemagglutinating peaks identified at 1.48-1.49, 1.42, 1.36 and 1.30-1.31 g cm-3. The number and distribution of the viral proteins and the equivalent protein molecular weights are similar for all three viruses in SDS-polyacrylamide gels (10%). Four viral proteins were identified and their molecular weights were determined: protein A (77 500-79 500), protein B (63 000-63 500), protein C (61 500-63 000) and protein D (50 000-55 000). The viral protein D although reported for some other parvoviruses has not previously been demonstrated in CPV-2, MEV or FPV.

Parvovirus infection in pigs with necrotic and vesicle-like lesions.

Porcine parvovirus was isolated from many visceral organs and also from the brain, serum and skin specimens of swine with vesicular-like conditions. Severe lesions were reported to have occurred in the mouth, on the tongue and snout, on the coronary band and in the interdigital spaces. Also, parvoviral antigens were demonstrated, by immunofluorescence, in the outer layers of hair follicles in skin adjacent to coronary band lesions.

Pathogenicity of a skin isolate of porcine parvovirus in swine fetuses.

The pathogenic properties of a skin isolate of porcine parvovirus (PPV), designated Kresse isolate, were compared with NADL-8 isolate, a prototype isolate of PPV, by in utero inoculation of mid-term and late-term gestation swine fetuses. Fetuses from pregnant sows of mid-gestation were inoculated with either NADL-8 or Kresse virus. Both isolates were highly pathogenic to mid-gestation fetuses. In contrast, dramatic differences in pathogenicity between these 2 isolates were observed in fetuses inoculated late in gestation. Such fetuses from each of 4 sows were inoculated with NADL-8 or Kresse virus isolate and sacrificed at 10, 18, 21, or 23 days postinoculation (PI). NADL-8-inoculated fetuses were grossly normal. The pathogenic effects of Kresse isolate were evident by gross pathology in fetuses collected at 18, 21, and 23 days PI, but not at 10 days PI. Hemagglutination (HA) and fluorescent antibody (FA) methods were used to identify virus in various tissues of late-gestation fetuses collected at 10 and 21 days PI. At 10 days PI, HA antigens were detected only in livers of NADL-8-inoculated fetuses, but in all tissues examined of Kresse-inoculated fetuses, including the brain. PPV specific fluorescence was demonstrated in tissues of fetuses inoculated with NADL-8 and Kresse virus. The major difference was that virus antigen was found in the brains of fetuses inoculated with Kresse virus, but not in NADL-8 infected fetuses. At 21 days PI, HA antigen was not detected in any of the tissues of fetuses inoculated with NADL-8 virus, with PPV specific fluorescence by FA being found only in the kidney. However, fetuses inoculated with Kresse virus displayed HA antigen in liver and PPV-specific fluorescence in all tissues tested including the brain. Both isolates induced similar antibody responses, 1:128 to 1:256 at 10 days and 1:512 to 1:1024 at 21 days PI. In addition, immunoglobulin G (IgG) deposits were demonstrated in kidneys and skin of fetuses inoculated with Kresse virus and IgM in brain, but not in tissues from fetuses inoculated with NADL-8 virus.