Eva Emmoth

A laboratory study of survival of selected microorganisms after heat treatment of biowaste used in biogas plants

The aim of the study was to assess the effect of pasteurisation, as set by the European regulation EC 1774/2002, on selected pathogens and indicator organisms. Unpasteurised substrate (biowaste), including animal by-products from a full-scale biogas plant was heat treated under laboratory conditions at 70 degrees C and 55 degrees C for 30 min and 60 min. Heat treatment at 55 degrees C for 60 min was not sufficient to achieve a hygienically acceptable product. Heat treatment at 70 degrees C for 30 min and 60 min was effective in reducing pathogenic bacteria, Ascaris suum eggs, Swine vesicular disease virus and indicator organisms. However, this level of pasteurisation will still not reduce the quantity of Clostridia spores, or completely inactivate heat-resistant viruses such as Porcine parvovirus or Salmonella phage 28B. The results still give cause for some concern regarding the use of digested residue from biogasplants in agriculture.

A Size‐Exclusion Nanocellulose Filter Paper for Virus Removal

This is the first time a 100% natural, unmodified nanofibrous polymer-based membrane is demonstrated capable of removing viruses solely based on the size-exclusion principle, with a log10 reduction value (LRV) ≥ 6.3 as limited by the assay lower detection limit and the feed virus titre, thereby matching the performance of industrial synthetic polymer virus removal filters.

Ammonia Disinfection of Hatchery Waste for Elimination of Single-Stranded RNA Viruses

ABSTRACT Hatchery waste, an animal by-product of the poultry industry, needs sanitation treatment before further use as fertilizer or as a substrate in biogas or composting plants, owing to the potential presence of opportunistic pathogens, including zoonotic viruses. Effective sanitation is also important in viral epizootic outbreaks and as a routine, ensuring high hygiene standards on farms. This study examined the use of ammonia at different concentrations and temperatures to disinfect hatchery waste. Inactivation kinetics of high-pathogenic avian influenza virus H7N1 and low-pathogenic avian influenza virus H5N3, as representatives of notifiable avian viral diseases, were determined in spiked hatchery waste. Bovine parainfluenza virus type 3, feline coronavirus, and feline calicivirus were used as models for other important avian pathogens, such as Newcastle disease virus, infectious bronchitis virus, and avian hepatitis E virus. Bacteriophage MS2 was also monitored as a stable indicator. Coronavirus was the most sensitive virus, with decimal reduction (D) values of 1.2 and 0.63 h after addition of 0.5% (wt/wt) ammonia at 14 and 25°C, respectively. Under similar conditions, high-pathogenic avian influenza H7N1 was the most resistant, with D values of 3.0 and 1.4 h. MS2 was more resistant than the viruses to all treatments and proved to be a suitable indicator of viral inactivation. The results indicate that ammonia treatment of hatchery waste is efficient in inactivating enveloped and naked single-stranded RNA viruses. Based on the D values and confidence intervals obtained, guidelines for treatment were proposed, and one was successfully validated at full scale at a hatchery, with MS2 added to hatchery waste.

Evolution of H3N8 equine influenza virus from 1963 to 1991.

The antigenic properties of H3N8 influenza viruses isolated from outbreaks of equine influenza in Sweden between 1979 and 1991 have been studied in hemagglutination inhibition tests with polyclonal and monoclonal antisera, and antigenic drift of the virus has been demonstrated. To clarify the basis of the antigenic drift, amino acid sequences of the globular head regions (HA1) of the hemagglutinin membrane glycoproteins of virus strains from 1979, 1984, 1988 and 1990 have been deduced from the nucleotide sequences of the hemagglutinin genes, and the sequence information has been used to construct a phylogenetic tree of H3N8 equine influenza strains. Several strains from previous studies have been included to give a clearer picture of viral evolution in an international context.

Composting for Avian Influenza Virus Elimination

ABSTRACT Effective sanitization is important in viral epizootic outbreaks to avoid further spread of the pathogen. This study examined thermal inactivation as a sanitizing treatment for manure inoculated with highly pathogenic avian influenza virus H7N1 and bacteriophages MS2 and ϕ6. Rapid inactivation of highly pathogenic avian influenza virus H7N1 was achieved at both mesophilic (35°C) and thermophilic (45 and 55°C) temperatures. Similar inactivation rates were observed for bacteriophage ϕ6, while bacteriophage MS2 proved too thermoresistant to be considered a valuable indicator organism for avian influenza virus during thermal treatments. Guidelines for treatment of litter in the event of emergency composting can be formulated based on the inactivation rates obtained in the study.

Cross-Protection of Chicken Immunoglobulin Y Antibodies against H5N1 and H1N1 Viruses Passively Administered in Mice

ABSTRACT Influenza viruses remain a major threat to global health due to their ability to undergo change through antigenic drift and antigenic shift. We postulated that avian IgY antibodies represent a low-cost, effective, and well-tolerated approach that can easily be scaled up to produce enormous quantities of protective antibodies. These IgY antibodies can be administered passively in humans (orally and intranasally) and can be used quickly and safely to help in the fight against an influenza pandemic. In this study, we raised IgY antibodies against H1N1, H3N2, and H5N1 influenza viruses. We demonstrated that, using whole inactivated viruses alone and in combination to immunize hens, we were able to induce a high level of anti-influenza virus IgY in the sera and eggs, which lasted for at least 2 months after two immunizations. Furthermore, we found that by use of in vitro assays to test for the ability of IgY to inhibit hemagglutination (HI test) and virus infectivity (serum neutralization test), IgYs inhibited the homologous as well as in some cases heterologous clades and strains of viruses. Using an in vivo mouse model system, we found that, when administered intranasally 1 h prior to infection, IgY to H5N1 protected 100% of the mice against lethal challenge with H5N1. Of particular interest was the finding that IgY to H5N1 cross-protected against A/Puerto Rico/8/34 (H1N1) both in vitro and in vivo. Based on our results, we conclude that anti-influenza virus IgY can be used to help prevent influenza virus infection.

Equine influenza virus from the 1991 Swedish epizootic shows major genetic and antigenic divergence from the prototype virus.

The antigenic properties of H3N8 equine influenza virus from the Swedish epizootic of 1991 differ from those of A/eq 2/Fontainebleau/79 (representative of the Swedish vaccine strain) in hemagglutination inhibition tests. The amino acid sequence of the hemagglutinin (HA) of an isolate from the 1991 outbreak was deduced from the nucleotide sequence and comparison was made to the A/eq 2/Fontainebleau/79 strain. Twenty-three amino acid substitutions were found, 10 mapping onto areas of the HA known to bind antibodies in human H3 influenza viruses. The amino acid changes together with the serological data suggest that a major antigenic drift has taken place in equine H3N8 viruses in Sweden and we conclude that recent strains of the virus must be incorporated into vaccines on a regular basis if epizootics of equine influenza are to be controlled in the future.

Comparison of two H1N2 swine influenza A viruses from disease outbreaks in pigs in Sweden during 2009 and 2010.

The influenza A virus subtypes H1N1, H1N2 and H3N2 are prevalent in pig populations worldwide. In the present study, two relatively uncommon swine influenza virus (SIV) H1N2 subtypes, isolated in Sweden in 2009 and 2010, were compared regarding their molecular composition and biological characteristics. The differences regarding markers purportedly related to pathogenicity, host adaptation or replication efficiency. They included a truncated PB1-F2 protein in the earlier isolate but a full length version in the more recent one; differences in the number of haemagglutinin glycosylation sites, including a characteristic human one; and a nuclear export protein with altered export signal. Of particular interest, the NS1 amino acid sequence of swine H1N2-2009 and 2010 has a ‘unique or very unusual’ PDZ binding domain (RPKV) at the C-terminal of the protein, a motif that has been implicated as a virulence marker. Concerning biological properties, these viruses reached lower titre and showed reduced cytopathogenicity in MDCK cells compared with an avian-like H1N1 isolate A/swine/Lidkoping/1193/2002 belonging to the same lineage as the 2009 and 2010 isolates. The findings should contribute to better understanding of factors related to the survival/extinction of this uncommon reassortant variant.

Biosecurity aspects and pathogen inactivation in acidified high risk animal by-products

The aim of the study was to investigate the effects of formic acid addition to ground high risk animal by-products (ABP 1) in terms of stabilization and pathogen inactivation and to evaluate the biosecurity risk connected with the ABP 1 based combustion fuel Biomal. Laboratory studies were performed on the persistence of Salmonella Typhimurium, Bacillus cereus spores, porcine herpes virus, avian influenza virus, bovine viral diarrhea virus, equine rhinitis A virus and porcine parvovirus in Biomal at different storage times. It was shown that Salmonella and enveloped viruses were inactivated within 1 day (24 h). Bacillus cereus spores were not reduced during 147 days and the non-enveloped virus porcine parvovirus was still detected after 168 days of storage. The conclusion that can be drawn from the study is that transmission of some highly contagious diseases such as foot-and-mouth-disease, swine vesicular disease and egg drop syndrome, caused by non-enveloped viruses, may follow accidental leakages of Biomal. In addition, there is a risk of transmission of the diseases anthrax and black leg, caused by sporulating bacteria.

Evaluation of Biological Indicator Spores as Tools for Assessment of Fumigation Decontamination Effectiveness

In high-containment laboratories and animal facilities common practice is to decontaminate the facilities prior to maintenance or in an emergency situation. Many laboratories use commercially available biological indicators (BIs) to validate the decontamination procedure. In this study the focus was to evaluate the reliability of four different commercial BIs in comparison to control microorganisms that are commonly used in laboratories. Two different fumigation decontamination procedures were chosen: formaldehyde (FA) and vaporized hydrogen peroxide (VHP). The control microorganisms were Salmonella typhimurium, Brucella melitensis, Bacillus thuringiensis spores, porcine parvovirus, equine rhinitis A virus, bovine viral diarrhea virus, and low pathogenic avian influenza virus. Exposure to formaldehyde caused a sufficient reduction of all the control microorganisms, including B. thuringiensis spores, whereas only one of the four commercial BIs was completely negative for growth. The VHP decontamination procedure was not able to reduce any of the control microorganisms more than 4 log10, except for the enveloped viruses, whereas the BIs with the lowest concentration (105 microorganisms per spore strip) indicated a satisfactory decontamination procedure. These results indicate that commercial BIs could be unreliable as general indicators of decontamination effectiveness. To ensure a reliable decontamination process, BIs have to be evaluated for each protocol in parallel with the microorganisms used in the laboratory.