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Inhalation of small numbers of Francisella tularensis subspecies tularensis (Ftt) in the form of small particle aerosols causes severe morbidity and mortality in people and many animal species. For this reason, Ftt was developed into a bona fide biological weapon by the USA, by the former USSR, and their respective allies during the previous century. Although such weapons were never deployed, the 9/11 attack quickly followed by the Amerithrax attack led the U.S. government to seek novel countermeasures against a select group of pathogens, including Ftt. Between 2005-2009, we pursued a novel live vaccine against Ftt by deleting putative virulence genes from a fully virulent strain of the pathogen, SCHU S4. These mutants were screened in a mouse model, in which the vaccine candidates were first administered intradermally (ID) to determine their degree of attenuation. Subsequently, mice that survived a high dose ID inoculation were challenged by aerosol or intranasally (IN) with virulent strains of Ftt. We used the current unlicensed live vaccine strain (LVS), first discovered over 70 years ago, as a comparator in the same model. After screening 60 mutants, we found only one, SCHU S4 Delta clpB, that outperformed LVS in the mouse ID vaccination-respiratory-challenge model. Currently, SCHU S4 Delta clpB has been manufactured under current good manufacturing practice conditions, and tested for safety and efficacy in mice, rats, and macaques. The steps necessary for advancing SCHU S4 Delta clpB to this late stage of development are detailed herein. These include developing a body of data supporting the attenuation of SCHU S4 Delta clpB to a degree sufficient for removal from the U.S. Select Agent list and for human use; optimizing SCHU S4 Delta clpB vaccine production, scale up, and long-term storage; and developing appropriate quality control testing approaches.

Inhalation of small numbers of Francisella tularensis subspecies tularensis (Ftt) in the form of small particle aerosols causes severe morbidity and mortality in people and many animal species. For this reason, Ftt was developed into a bona fide biological weapon by the USA, by the former USSR, and their respective allies during the previous century. Although such weapons were never deployed, the 9/11 attack quickly followed by the Amerithrax attack led the U.S. government to seek novel countermeasures against a select group of pathogens, including Ftt. Between 2005-2009, we pursued a novel live vaccine against Ftt by deleting putative virulence genes from a fully virulent strain of the pathogen, SCHU S4. These mutants were screened in a mouse model, in which the vaccine candidates were first administered intradermally (ID) to determine their degree of attenuation. Subsequently, mice that survived a high dose ID inoculation were challenged by aerosol or intranasally (IN) with virulent strains of Ftt. We used the current unlicensed live vaccine strain (LVS), first discovered over 70 years ago, as a comparator in the same model. After screening 60 mutants, we found only one, SCHU S4 Delta clpB, that outperformed LVS in the mouse ID vaccination-respiratory-challenge model. Currently, SCHU S4 Delta clpB has been manufactured under current good manufacturing practice conditions, and tested for safety and efficacy in mice, rats, and macaques. The steps necessary for advancing SCHU S4 Delta clpB to this late stage of development are detailed herein. These include developing a body of data supporting the attenuation of SCHU S4 Delta clpB to a degree sufficient for removal from the U.S. Select Agent list and for human use; optimizing SCHU S4 Delta clpB vaccine production, scale up, and long-term storage; and developing appropriate quality control testing approaches.

Inhalation of small numbers of Francisella tularensis subspecies tularensis (Ftt) in the form of small particle aerosols causes severe morbidity and mortality in people and many animal species. For this reason, Ftt was developed into a bona fide biological weapon by the USA, by the former USSR, and their respective allies during the previous century. Although such weapons were never deployed, the 9/11 attack quickly followed by the Amerithrax attack led the U.S. government to seek novel countermeasures against a select group of pathogens, including Ftt. Between 2005-2009, we pursued a novel live vaccine against Ftt by deleting putative virulence genes from a fully virulent strain of the pathogen, SCHU S4. These mutants were screened in a mouse model, in which the vaccine candidates were first administered intradermally (ID) to determine their degree of attenuation. Subsequently, mice that survived a high dose ID inoculation were challenged by aerosol or intranasally (IN) with virulent strains of Ftt. We used the current unlicensed live vaccine strain (LVS), first discovered over 70 years ago, as a comparator in the same model. After screening 60 mutants, we found only one, SCHU S4 Delta clpB, that outperformed LVS in the mouse ID vaccination-respiratory-challenge model. Currently, SCHU S4 Delta clpB has been manufactured under current good manufacturing practice conditions, and tested for safety and efficacy in mice, rats, and macaques. The steps necessary for advancing SCHU S4 Delta clpB to this late stage of development are detailed herein. These include developing a body of data supporting the attenuation of SCHU S4 Delta clpB to a degree sufficient for removal from the U.S. Select Agent list and for human use; optimizing SCHU S4 Delta clpB vaccine production, scale up, and long-term storage; and developing appropriate quality control testing approaches.

Inhalation of small numbers of Francisella tularensis subspecies tularensis (Ftt) in the form of small particle aerosols causes severe morbidity and mortality in people and many animal species. For this reason, Ftt was developed into a bona fide biological weapon by the USA, by the former USSR, and their respective allies during the previous century. Although such weapons were never deployed, the 9/11 attack quickly followed by the Amerithrax attack led the U.S. government to seek novel countermeasures against a select group of pathogens, including Ftt. Between 2005-2009, we pursued a novel live vaccine against Ftt by deleting putative virulence genes from a fully virulent strain of the pathogen, SCHU S4. These mutants were screened in a mouse model, in which the vaccine candidates were first administered intradermally (ID) to determine their degree of attenuation. Subsequently, mice that survived a high dose ID inoculation were challenged by aerosol or intranasally (IN) with virulent strains of Ftt. We used the current unlicensed live vaccine strain (LVS), first discovered over 70 years ago, as a comparator in the same model. After screening 60 mutants, we found only one, SCHU S4 Delta clpB, that outperformed LVS in the mouse ID vaccination-respiratory-challenge model. Currently, SCHU S4 Delta clpB has been manufactured under current good manufacturing practice conditions, and tested for safety and efficacy in mice, rats, and macaques. The steps necessary for advancing SCHU S4 Delta clpB to this late stage of development are detailed herein. These include developing a body of data supporting the attenuation of SCHU S4 Delta clpB to a degree sufficient for removal from the U.S. Select Agent list and for human use; optimizing SCHU S4 Delta clpB vaccine production, scale up, and long-term storage; and developing appropriate quality control testing approaches.

Inhalation of small numbers of Francisella tularensis subspecies tularensis (Ftt) in the form of small particle aerosols causes severe morbidity and mortality in people and many animal species. For this reason, Ftt was developed into a bona fide biological weapon by the USA, by the former USSR, and their respective allies during the previous century. Although such weapons were never deployed, the 9/11 attack quickly followed by the Amerithrax attack led the U.S. government to seek novel countermeasures against a select group of pathogens, including Ftt. Between 2005-2009, we pursued a novel live vaccine against Ftt by deleting putative virulence genes from a fully virulent strain of the pathogen, SCHU S4. These mutants were screened in a mouse model, in which the vaccine candidates were first administered intradermally (ID) to determine their degree of attenuation. Subsequently, mice that survived a high dose ID inoculation were challenged by aerosol or intranasally (IN) with virulent strains of Ftt. We used the current unlicensed live vaccine strain (LVS), first discovered over 70 years ago, as a comparator in the same model. After screening 60 mutants, we found only one, SCHU S4 Delta clpB, that outperformed LVS in the mouse ID vaccination-respiratory-challenge model. Currently, SCHU S4 Delta clpB has been manufactured under current good manufacturing practice conditions, and tested for safety and efficacy in mice, rats, and macaques. The steps necessary for advancing SCHU S4 Delta clpB to this late stage of development are detailed herein. These include developing a body of data supporting the attenuation of SCHU S4 Delta clpB to a degree sufficient for removal from the U.S. Select Agent list and for human use; optimizing SCHU S4 Delta clpB vaccine production, scale up, and long-term storage; and developing appropriate quality control testing approaches.

INTRODUCTION: The CampySource project aims to identify risk factors for human Campylobacter infection in Australia. We will investigate locally relevant risk factors and those significant in international studies in a case-control study. Case isolates and contemporaneous isolates from food and animal sources will be sequenced to conduct source attribution modelling, and findings will be combined with the case-control study in a source-assigned analysis. METHODS AND ANALYSIS: The case-control study will include 1200 participants (600 cases and 600 controls) across three regions in Australia. Cases will be recruited from campylobacteriosis notifications to health departments. Only those with a pure and viable Campylobacter isolate will be eligible for selection to allow for whole genome sequencing of isolates. Controls will be recruited from notified cases of influenza, frequency matched by sex, age group and geographical area of residence. All participants will be interviewed by trained telephone interviewers using a piloted questionnaire.We will collect Campylobacter isolates from retail meats and companion animals (specifically dogs), and all food, animal and human isolates will undergo whole genome sequencing. We will use sequence data to estimate the proportion of human infections that can be attributed to animal and food reservoirs (source attribution modelling), and to identify spatial clusters and temporal trends. Source-assigned analysis of the case-control study data will also be conducted where cases are grouped according to attributed sources. ETHICS AND DISSEMINATION: Human and animal ethics have been approved. Genomic data will be published in online archives accompanied by basic metadata. We anticipate several publications to come from this study.

Using a strategy of gene deletion mutagenesis, we have examined the roles of genes putatively involved in lipopolysaccharide biosynthesis in the virulent facultative intracellular bacterial pathogen, Francisella tularensis subspecies tularensis, strain SCHU S4 in LPS biosynthesis, protein glycosylation, virulence and immunogenicity. One mutant, ∆wbtI, did not elaborate a long chain O-polysaccharide (OPS), was completely avirulent for mice, and failed to induce a protective immune response against challenge with wild type bacteria. Another mutant, ∆wbtC, produced a long chain OPS with altered chemical and electrophoretic characteristics. This mutant showed markedly reduced glycosylation of several known glycoproteins. Additionally this mutant was highly attenuated, and elicited a protective immune response against systemic, but not respiratory challenge with wild type SCHU S4. A third mutant, ∆kdtA, produced an unconjugated long chain OPS, lacking a detectable core structure, and which was not obviously expressed at the surface. It was avirulent and elicited partial protection against systemic challenge only.

Using a strategy of gene deletion mutagenesis, we have examined the roles of genes putatively involved in lipopolysaccharide biosynthesis in the virulent facultative intracellular bacterial pathogen, Francisella tularensis subspecies tularensis, strain SCHU S4 in LPS biosynthesis, protein glycosylation, virulence and immunogenicity. One mutant, ∆wbtI, did not elaborate a long chain O-polysaccharide (OPS), was completely avirulent for mice, and failed to induce a protective immune response against challenge with wild type bacteria. Another mutant, ∆wbtC, produced a long chain OPS with altered chemical and electrophoretic characteristics. This mutant showed markedly reduced glycosylation of several known glycoproteins. Additionally this mutant was highly attenuated, and elicited a protective immune response against systemic, but not respiratory challenge with wild type SCHU S4. A third mutant, ∆kdtA, produced an unconjugated long chain OPS, lacking a detectable core structure, and which was not obviously expressed at the surface. It was avirulent and elicited partial protection against systemic challenge only.

Using a strategy of gene deletion mutagenesis, we have examined the roles of genes putatively involved in lipopolysaccharide biosynthesis in the virulent facultative intracellular bacterial pathogen, Francisella tularensis subspecies tularensis, strain SCHU S4 in LPS biosynthesis, protein glycosylation, virulence and immunogenicity. One mutant, ∆wbtI, did not elaborate a long chain O-polysaccharide (OPS), was completely avirulent for mice, and failed to induce a protective immune response against challenge with wild type bacteria. Another mutant, ∆wbtC, produced a long chain OPS with altered chemical and electrophoretic characteristics. This mutant showed markedly reduced glycosylation of several known glycoproteins. Additionally this mutant was highly attenuated, and elicited a protective immune response against systemic, but not respiratory challenge with wild type SCHU S4. A third mutant, ∆kdtA, produced an unconjugated long chain OPS, lacking a detectable core structure, and which was not obviously expressed at the surface. It was avirulent and elicited partial protection against systemic challenge only.

Using a strategy of gene deletion mutagenesis, we have examined the roles of genes putatively involved in lipopolysaccharide biosynthesis in the virulent facultative intracellular bacterial pathogen, Francisella tularensis subspecies tularensis, strain SCHU S4 in LPS biosynthesis, protein glycosylation, virulence and immunogenicity. One mutant, ∆wbtI, did not elaborate a long chain O-polysaccharide (OPS), was completely avirulent for mice, and failed to induce a protective immune response against challenge with wild type bacteria. Another mutant, ∆wbtC, produced a long chain OPS with altered chemical and electrophoretic characteristics. This mutant showed markedly reduced glycosylation of several known glycoproteins. Additionally this mutant was highly attenuated, and elicited a protective immune response against systemic, but not respiratory challenge with wild type SCHU S4. A third mutant, ∆kdtA, produced an unconjugated long chain OPS, lacking a detectable core structure, and which was not obviously expressed at the surface. It was avirulent and elicited partial protection against systemic challenge only.

Using a strategy of gene deletion mutagenesis, we have examined the roles of genes putatively involved in lipopolysaccharide biosynthesis in the virulent facultative intracellular bacterial pathogen, Francisella tularensis subspecies tularensis, strain SCHU S4 in LPS biosynthesis, protein glycosylation, virulence and immunogenicity. One mutant, ∆wbtI, did not elaborate a long chain O-polysaccharide (OPS), was completely avirulent for mice, and failed to induce a protective immune response against challenge with wild type bacteria. Another mutant, ∆wbtC, produced a long chain OPS with altered chemical and electrophoretic characteristics. This mutant showed markedly reduced glycosylation of several known glycoproteins. Additionally this mutant was highly attenuated, and elicited a protective immune response against systemic, but not respiratory challenge with wild type SCHU S4. A third mutant, ∆kdtA, produced an unconjugated long chain OPS, lacking a detectable core structure, and which was not obviously expressed at the surface. It was avirulent and elicited partial protection against systemic challenge only.

The objective of this study was to determine the prevalence and genotypic range of Hepatitis E virus (HEV) in the pig population of northern Lao People's Democratic Republic (PDR). We collected 181 faecal samples from indigenous-breed pigs ≤6 months of age and the faeces was stored in RNA stabilisation buffer due to cold-chain and transport limitations. Twenty-one (11.6%) pigs had detectable HEV RNA and 43.5% of village pig herds were infected. Based on a 240 base pair-nucleotide sequence flanking the junction of open reading frames 1, 2 and 3 (ORF1, ORF2 and ORF3) the isolates were phylogenetically classified within genotype 4. Phylogenetic analyses revealed distinct genetic groupings of the Lao HEV isolates and two groups clustered with human and pig HEV isolates from China. This was the first study to demonstrate genotype 4 HEV in Lao PDR and indicates pigs are a potential reservoir for human HEV infection.

The objective of this study was to determine the prevalence and genotypic range of Hepatitis E virus (HEV) in the pig population of northern Lao People's Democratic Republic (PDR). We collected 181 faecal samples from indigenous-breed pigs ≤6 months of age and the faeces was stored in RNA stabilisation buffer due to cold-chain and transport limitations. Twenty-one (11.6%) pigs had detectable HEV RNA and 43.5% of village pig herds were infected. Based on a 240 base pair-nucleotide sequence flanking the junction of open reading frames 1, 2 and 3 (ORF1, ORF2 and ORF3) the isolates were phylogenetically classified within genotype 4. Phylogenetic analyses revealed distinct genetic groupings of the Lao HEV isolates and two groups clustered with human and pig HEV isolates from China. This was the first study to demonstrate genotype 4 HEV in Lao PDR and indicates pigs are a potential reservoir for human HEV infection.

Francisella tularensis subspecies tularensis is a highly virulent facultative intracellular pathogen of humans and a potential biological weapon. A live vaccine strain, F. tularensis LVS, was developed more than 50 years ago by pragmatic attenuation of a strain of the less virulent holarctica subspecies. LVS was demonstrated to be highly effective in human volunteers who were exposed to intradermal challenge with fully virulent subsp. tularensis, but was less effective against aerosol exposure. LVS faces regulatory hurdles that to date have prevented its licensure for general use. Therefore, a better defined and more effective vaccine is being sought. To this end we have created gene deletion mutants in the virulent subsp. tularensis strain and tested them for their ability to elicit a protective immune response against systemic or aerosol challenge with the highly virulent wild-type subsp. tularensis strain, SCHU S4. Both oral and intradermal (ID) primary vaccination routes were assessed in BALB/c and C3H/HeN mice as was oral boosting. One SCHU S4 mutant missing the heat shock gene, clpB, was significantly more attenuated than LVS whereas a double deletion mutant missing genes FTT0918 and capB was as attenuated as LVS. In general mice immunized with SCHU S4DeltaclpB were significantly better protected against aerosol challenge than mice immunized with LVS. A single ID immunization of BALB/c mice with SCHU S4DeltaclpB was at least as effective as any other regimen examined. Mice immunized with SCHU S4Delta0918DeltacapB were generally protected to a similar degree as mice immunized with LVS. A preliminary examination of immune responses to vaccination with LVS, SCHU S4DeltaclpB, or SCHU S4Delta0918DeltacapB provided no obvious correlate to their relative efficacies.

Cysticercosis caused by the zoonotic pork tapeworm Taenia solium is emerging as a constraint for the nutritional and economic wellbeing of small-holder farming communities in many underdeveloped areas of Southeast Asia. It occurs mainly in impoverished regions with inadequate sanitation, poor pig management practices and lack of meat inspection and control. Neurocysticercosis, the most serious form of the disease, is considered the most common parasitic infection of the human nervous system and the most frequent preventable cause of epilepsy in the developing world. Although theoretically easy to control and declared eradicable, T. solium taeniosis/cysticercosis remains a neglected disease. There is a lack of information and awareness of the burden and transmission of the disease at the regional and global level, partially explained by the unavailability of good quality diagnostic tools in field-applicable formats. These factors are further compounded by a lack of validated simple and sustainable intervention packages as part of integrated helminth control programmes. To date, T. solium taeniosis/cysticercosis has not been eliminated from any region by a specific programme in Southeast Asia, and no national control programmes are yet in place except in parts of the People's Republic of China. The presence, distribution, public health importance and economic relevance of cysticercosis need to be better documented in Southeast Asia in order to bring it to the attention of affected communities, decision-makers and funding bodies. A number of proven cost-effective intervention tools for combating cysticercosis appear to be available but need to be field validated. The Regional Network for Asian Schistosomiasis and Other Helminth Zoonoses (RNAS(+)) serves as an important regional 'driving force' for managing research, capacity building, knowledge and stakeholder engagement essential for controlling cysticercosis in the Southeast Asian region while ensuring that research

Cysticercosis caused by the zoonotic pork tapeworm Taenia solium is emerging as a constraint for the nutritional and economic wellbeing of small-holder farming communities in many underdeveloped areas of Southeast Asia. It occurs mainly in impoverished regions with inadequate sanitation, poor pig management practices and lack of meat inspection and control. Neurocysticercosis, the most serious form of the disease, is considered the most common parasitic infection of the human nervous system and the most frequent preventable cause of epilepsy in the developing world. Although theoretically easy to control and declared eradicable, T. solium taeniosis/cysticercosis remains a neglected disease. There is a lack of information and awareness of the burden and transmission of the disease at the regional and global level, partially explained by the unavailability of good quality diagnostic tools in field-applicable formats. These factors are further compounded by a lack of validated simple and sustainable intervention packages as part of integrated helminth control programmes. To date, T. solium taeniosis/cysticercosis has not been eliminated from any region by a specific programme in Southeast Asia, and no national control programmes are yet in place except in parts of the People's Republic of China. The presence, distribution, public health importance and economic relevance of cysticercosis need to be better documented in Southeast Asia in order to bring it to the attention of affected communities, decision-makers and funding bodies. A number of proven cost-effective intervention tools for combating cysticercosis appear to be available but need to be field validated. The Regional Network for Asian Schistosomiasis and Other Helminth Zoonoses (RNAS(+)) serves as an important regional 'driving force' for managing research, capacity building, knowledge and stakeholder engagement essential for controlling cysticercosis in the Southeast Asian region while ensuring that research

The type A subspecies of Francisella tularensis is a highly virulent facultative intracellular bacterial pathogen, and a potential biological weapon. Recently, there has been renewed interest in developing new vaccines and therapeutics against this bacterium. Natural cases of disease, tularemia, caused by the type A subspecies are very rare. Therefore, the United States Food and Drug Administration will rely on the so-called Animal Rule for efficacy testing of anti-Francisella medicines. This requires the human disease to be modeled in one or more animal species in which the pathogenicity of the agent is reasonably well understood. Mice are natural hosts for F. tularensis, and might be able to satisfy this requirement. Tularemia pathogenesis appears to be primarily due to the host inflammatory response which is poorly understood at the molecular level. Additionally, the extent to which this response varies depending on host and pathogen genetic background, or by pathogen challenge route or dose is unknown. Therefore, the present study examined sera and infected tissues from C57BL/6 and BALB/c mice challenged by natural intradermal (ID) and respiratory routes with one of two distinct type A strains of the pathogen for cytokine and chemokine responses that might help to explain the morbidity associated with tularemia. The results show that the molecular immune response was mostly similar regardless of the variables examined. For instance, mRNA for the proinflammatory cytokine IL-6, and chemokines KC, and IP-10 was consistently upregulated at all sites of infection. Upregulation of mRNA for several other cytokines and chemokines occurred in a more tissue restricted manner. For instance, IFN-gamma was highly upregulated in the skin of BALB/c, but not C57BL/6 mice after ID inoculation of the pathogen, whilst IL-10 mRNA upregulation was only consistently seen in the skin and lungs.

Taeniasis and cysticercosis are important but underreported parasitic zoonoses in the Lao People's Democratic Republic (Lao PDR). Reports of human and pig cysticercosis are rather limited and based largely on anecdotal evidence. To date, no structured surveys of disease prevalence or incidence have been reported. However, one unpublished pilot survey of pig cysticercosis in a slaughterhouse in northern Laos estimated prevalence to be 1.7%, without speciation of parasite cysts. Over the past 20 years, nine surveys of intestinal helminthic infection have been conducted; the prevalence of human taeniasis ranged from 0 to 14.0%. The study designs and sample sizes varied greatly, however a high degree of spatial and age variation in taeniasis prevalence was evident. These results are however inconclusive as the species of tapeworm infecting the people was not determined. To further our knowledge of taeniasis and cysticercosis in Lao PDR, structured community-based surveys in high-risk areas are required in combination with the use of sensitive and specific diagnostic tests capable of identifying the pork tapeworm, Taenia solium. This will enable the development and implementation of control measures that are both appropriate and sustainable if T. solium is shown to be a public health threat.

Approximately 75% of the population of Lao PDR is engaged in agriculture and the vast majority (approximately 90%) of these producers are in the smallholder sector. Livestock are an important contributor to national, agricultural and village economies and are relied on for food security. The pig population has increased over the past 5 years at an annual average increase of 4.7% at the national herd level and up to 20% in some provinces. Cattle and buffalo populations have grown at more modest rates of 1–2% (Figure 1). Disease, including foot-and-mouth disease (FMD) and classical swine fever (CSF), is a major constraint to efficient and sustainable livestock production. Up to 80–90% of pigs and 99% of cattle and buffalo are produced in the smallholder sector using low input practices; as such, there is limited private sector input. Disease reporting, diagnosis, control and prevention are addressed by the Lao Government through the National Department of Livestock and Fisheries (DLF) and local agriculture and forestry offices at provincial and district government levels. These activities are supported by international partners such as the Australian Centre for International Agricultural Research (ACIAR), Commonwealth Scientific and Investigation Research Organisation (CSIRO), Japanese International Cooperation Association (JICA), Food and Agriculture Organization (FAO), European Union (EU) and Office International des Epizooties (OIE). Disease reporting and communication are passive and reports are made from villages through government administrations at district and provincial levels and then to the national level—the DLF and the National Animal Health Centre (NAHC). Communication of FMD-related information at regional and international levels is coordinated by the OIE South-East Asian FMD regional coordination unit (SEAFMD RCU), where reports are submitted monthly. Disease reporting for CSF is less well coordinated and information is provided to the OIE.

At the close of the workshop, a facilitated session was held to explore future directions for the control and management of classical swine fever (CSF) and foot-and-mouth disease (FMD) in Lao PDR. The workshop was attended by experts in their fields from the People’s Republic of China, Thailand, Myanmar, Cambodia, Vietnam, Lao PDR and Australia, and other representatives from a range of nongovernment organisations and international development projects. The session was designed in such a way as to best capture the skills and experience of attending delegates to progress ideas in an environment where all participants were able to make a contribution to the discussion. The workshop participants were divided into five groups in a manner to ensure an even representation across countries. Two leaders were assigned to each group, one to record answers on a whiteboard and the other to facilitate discussion within the group. The leadership personnel comprised Lao and Australian project staff and the session was facilitated by the first author. The session method is described in Figure 1.

Classical swine fever (CSF) virus is endemic in Lao PDR, with major outbreaks each year resulting in significant production losses in all farming systems, including smallholder, semi-intensive and intensive farms. CSF is a vaccine-preventable disease and there are many vaccines, both live attenuated and subunit, commercially available. In Lao PDR a variety of livestock vaccines are produced at the National Vaccine Production Centre (NVPC), which is located at Nongteng village 15 km from Vientiane. The NVPC was established in 1980 and produces vaccines for CSF, haemorrhagic septicaemia, Newcastle disease, fowl cholera, infectious bronchitis, fowl pox and duck plague. CSF vaccine is produced from the live attenuated C-strain virus from homogenised rabbit spleen and mesenteric lymph nodes freeze dried in the presence of stabilisers in a rubber stoppered vial and stored at –20 °C.

Classical swine fever (CSF) virus is a highly contagious but vaccine-preventable disease of swine. A locally produced lapinised C-strain vaccine is used to control CSF in Lao PDR; however, vaccine failure has been reported. The CSF vaccine is produced at the National Vaccine Production Centre (NVPC) as a freeze-dried rabbit spleen homogenate in a rubber stoppered glass vial and stored at –20 °C with a recommended shelf life of 1 year. This paper describes two studies to (i) determine the stability of the locally produced vaccine when stored at 4 °C and –20 °C and (ii) determine if the vaccine elicits a protective immune response when delivered to village pigs under good transport conditions. The vaccine was found to be stable for only 4 months when stored at –20 °C and for less than 3 months when stored at 4 °C. Under field conditions, vaccine stored at –20 °C for 2 months and transported at temperatures less than 1 °C elicited an immune response in 89% of vaccinated pigs by day 35 and 100% of pigs by day 70 post vaccination.

Classical swine fever (CSF) and foot-and-mouth disease (FMD) are two major trans-boundary animal diseases (TADs) having economic impact on the South-East Asian region. This paper describes the various diagnostic tests available for CSF and FMD, the limitations of each and their potential application in a low technology setting. The need to have complementary field and laboratory operations including suitable samples and transport methods are discussed, and examples are given. The importance of a quality assurance system to assess the accuracy and precision of diagnostic results is highlighted.

Classical swine fever (CSF) is a highly contagious viral disease of swine that causes major losses in all pig production systems in many regions of the world. In Lao PDR, CSF is endemic and outbreaks have adverse effects on the predominantly smallholder farming sector. Laboratory testing is required to accurately identify CSF outbreaks because of the difficulty of making a diagnosis based solely on clinical signs. The National Animal Health Centre, located in the national capital, Vientiane, has the capacity to reliably detect CSF antigen in tissue samples using an antigen capture (AC)-ELISA, and antibodies to CSF virus from serum samples using the complex trapping blocking ELISA. This paper describes the use of immunomagnetic beads (IMB) as the solid phase for the portable detection of CSF antigen in spleen samples and for the reliable detection of antibodies to CSF in animals vaccinated with a lapinised C-strain vaccine. The portable IMB-ELISA for antigen detection was shown to be 100% sensitive and 91% specific in comparison to the AC-ELISA. The IMB-Antibody-ELISA was shown to be 97% sensitive and 95% specific in comparison to the gold standard—neutralising peroxidase linked assay. These new diagnostic tests have the potential to improve CSF management through portable and rapid identification of outbreaks and the reliable and inexpensive monitoring of vaccination programs.

The type A subspecies of Francisella tularensis is a highly virulent facultative intracellular bacterial pathogen, and a potential biological weapon. Recently, there has been renewed interest in developing new vaccines and therapeutics against this bacterium. Natural cases of disease, tularemia, caused by the type A subspecies are very rare. Therefore, the United States Food and Drug Administration will rely on the so-called Animal Rule for efficacy testing of anti-Francisella medicines. This requires the human disease to be modeled in one or more animal species in which the pathogenicity of the agent is reasonably well understood. Mice are natural hosts for F. tularensis, and might be able to satisfy this requirement. Tularemia pathogenesis appears to be primarily due to the host inflammatory response which is poorly understood at the molecular level. Additionally, the extent to which this response varies depending on host and pathogen genetic background, or by pathogen challenge route or dose is unknown. Therefore, the present study examined sera and infected tissues from C57BL/6 and BALB/c mice challenged by natural intradermal (ID) and respiratory routes with one of two distinct type A strains of the pathogen for cytokine and chemokine responses that might help to explain the morbidity associated with tularemia. The results show that the molecular immune response was mostly similar regardless of the variables examined. For instance, mRNA for the proinflammatory cytokine IL-6, and chemokines KC, and IP-10 was consistently upregulated at all sites of infection. Upregulation of mRNA for several other cytokines and chemokines occurred in a more tissue restricted manner. For instance, IFN-gamma was highly upregulated in the skin of BALB/c, but not C57BL/6 mice after ID inoculation of the pathogen, whilst IL-10 mRNA upregulation was only consistently seen in the skin and lungs.

Taeniasis and cysticercosis are important but underreported parasitic zoonoses in the Lao People's Democratic Republic (Lao PDR). Reports of human and pig cysticercosis are rather limited and based largely on anecdotal evidence. To date, no structured surveys of disease prevalence or incidence have been reported. However, one unpublished pilot survey of pig cysticercosis in a slaughterhouse in northern Laos estimated prevalence to be 1.7%, without speciation of parasite cysts. Over the past 20 years, nine surveys of intestinal helminthic infection have been conducted; the prevalence of human taeniasis ranged from 0 to 14.0%. The study designs and sample sizes varied greatly, however a high degree of spatial and age variation in taeniasis prevalence was evident. These results are however inconclusive as the species of tapeworm infecting the people was not determined. To further our knowledge of taeniasis and cysticercosis in Lao PDR, structured community-based surveys in high-risk areas are required in combination with the use of sensitive and specific diagnostic tests capable of identifying the pork tapeworm, Taenia solium. This will enable the development and implementation of control measures that are both appropriate and sustainable if T. solium is shown to be a public health threat.

Francisella tularensis subspecies tularensis (type A F. tularensis) is considered to be one of the most virulent of all bacterial pathogens. Mice are extremely susceptible to infection with this subspecies (LD100 via various inoculation routes is <10 cfu). However, it has not been established whether overt virulence differences exist amongst type A strains of F. tularensis. To this end, the present study compared the virulence of two distinct type A strains, FSC033 and SCHU S4, for naïve mice and mice immunized with the live vaccine strain of the pathogen, F. tularensis LVS. One nominal isolate of SCHU S4 was found to be completely avirulent. Another isolate was highly virulent, but all examined cases appeared somewhat less virulent than FSC033. The implication of these findings for future infection and immunity studies is discussed.

TGF alpha is a small protein which can bind to, and activate, epidermal growth factor receptors (EGF-R) on the surface of epithelial, mesenchymal and a range of cancer cells, causing the initiation of DNA synthesis and subsequent division. Despite intensive efforts using site-directed mutagenesis, synthetic peptide fragments, and determination of the secondary and tertiary folding of TGF alpha in solution, the role and importance of many structural features in mediating EGF-R activation remain ill-defined. We have assayed the ability of EGF-R to bind to peptide fragments of TGF alpha.

This report developes a technique for approximating the solution to a Cauchy problem for a class of second order elliptic partial differential equations in N independent variables. The method is based upon the determination of an a priori bound for an arbitrary u(alpha) at a point P in terms of the values of the u(alpha) and their gradients on the Cauchy surface, and of a functional of the elliptic operator applied to the u(alpha). (Author)