Science.gov

Sample records for highly pathogenic h5ni

  1. Highly pathogenic avian influenza.

    PubMed

    Swayne, D E; Suarez, D L

    2000-08-01

    Highly pathogenic (HP) avian influenza (AI) (HPAI) is an extremely contagious, multi-organ systemic disease of poultry leading to high mortality, and caused by some H5 and H7 subtypes of type A influenza virus, family Orthomyxoviridae. However, most AI virus strains are mildly pathogenic (MP) and produce either subclinical infections or respiratory and/or reproductive diseases in a variety of domestic and wild bird species. Highly pathogenic avian influenza is a List A disease of the Office International des Epizooties, while MPAI is neither a List A nor List B disease. Eighteen outbreaks of HPAI have been documented since the identification of AI virus as the cause of fowl plague in 1955. Mildly pathogenic avian influenza viruses are maintained in wild aquatic bird reservoirs, occasionally crossing over to domestic poultry and causing outbreaks of mild disease. Highly pathogenic avian influenza viruses do not have a recognised wild bird reservoir, but can occasionally be isolated from wild birds during outbreaks in domestic poultry. Highly pathogenic avian influenza viruses have been documented to arise from MPAI viruses through mutations in the haemagglutinin surface protein. Prevention of exposure to the virus and eradication are the accepted methods for dealing with HPAI. Control programmes, which imply allowing a low incidence of infection, are not an acceptable method for managing HPAI, but have been used during some outbreaks of MPAI. The components of a strategy to deal with MPAI or HPAI include surveillance and diagnosis, biosecurity, education, quarantine and depopulation. Vaccination has been used in some control and eradication programmes for AI.

  2. 76 FR 24793 - Highly Pathogenic Avian Influenza

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-05-03

    ... (76 FR 4046-4056, Docket No. APHIS-2006-0074) an interim rule that amended the regulations governing... Inspection Service 9 CFR Parts 93, 94, and 95 RIN 0579-AC36 Highly Pathogenic Avian Influenza AGENCY: Animal... products from regions where any subtype of highly pathogenic avian influenza is considered to exist....

  3. Current situation on highly pathogenic avian influenza

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Avian influenza is one of the most important diseases affecting the poultry industry worldwide. Avian influenza viruses can cause a range of clinical disease in poultry. Viruses that cause severe disease and mortality are referred to as highly pathogenic avian influenza (HPAI) viruses. The Asian ...

  4. Determinants of pathogenicity of H5N1 highly pathogenic avian influenza viruses in ducks

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ducks have been implicated in the dissemination and evolution of the H5N1 highly pathogenic avian influenza (HPAI) viruses. The pathogenicity of H5N1 HPAI viruses in domestic ducks has increased over time with some viruses producing 100% mortality in very short time. The determinants of pathogenic...

  5. Shigella: A Highly Virulent and Elusive Pathogen

    PubMed Central

    Estrada-García, Teresa

    2014-01-01

    Despite a significant decrease in Shigella-related mortality, shigellosis continues to carry a significant burden of disease worldwide, particularly in Asia and Africa. Shigella is a highly virulent pathogen comprised of four major species with numerous subtypes. Shigella dysenteriae and Shigella flexneri infections are predominant in resource-limited settings. Clinical presentations range from mild watery diarrhea to severe dysentery with systemic complications such as electrolyte imbalance, seizures and hemolytic uremic syndrome. S. dysenteriae subtype 1, the producer of Shiga toxin, causes the most severe illness and highest mortality. Susceptible strains of Shigella may be effectively treated with inexpensive oral antibiotics such as ampicillin or trimethoprim-sulfamethoxazole. Unfortunately, multidrug resistant strains have emerged that have rendered most antibiotics, including fluoroquinolones and extended-spectrum cephalosporins, ineffective. Management and prevention of shigellosis represents a major public health challenge. The development of an effective vaccine is urgently needed to decrease its global impact. PMID:25110633

  6. Shigella: A Highly Virulent and Elusive Pathogen.

    PubMed

    Zaidi, Mussaret Bano; Estrada-García, Teresa

    2014-06-01

    Despite a significant decrease in Shigella-related mortality, shigellosis continues to carry a significant burden of disease worldwide, particularly in Asia and Africa. Shigella is a highly virulent pathogen comprised of four major species with numerous subtypes. Shigella dysenteriae and Shigella flexneri infections are predominant in resource-limited settings. Clinical presentations range from mild watery diarrhea to severe dysentery with systemic complications such as electrolyte imbalance, seizures and hemolytic uremic syndrome. S. dysenteriae subtype 1, the producer of Shiga toxin, causes the most severe illness and highest mortality. Susceptible strains of Shigella may be effectively treated with inexpensive oral antibiotics such as ampicillin or trimethoprim-sulfamethoxazole. Unfortunately, multidrug resistant strains have emerged that have rendered most antibiotics, including fluoroquinolones and extended-spectrum cephalosporins, ineffective. Management and prevention of shigellosis represents a major public health challenge. The development of an effective vaccine is urgently needed to decrease its global impact.

  7. Pathogenicity of reassortant H5N1 highly pathogenic avian influenza viruses in domestic ducks

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The pathogenicity of H5N1 highly pathogenic avian influenza (HPAI) viruses in domestic ducks has increased over time. These changes in virulence have been reported with viruses from countries with high population of domestic ducks, including Egypt. In order to understand which viral genes are contri...

  8. USGS highly pathogenic avian influenza research strategy

    USGS Publications Warehouse

    Harris, M. Camille; Miles, A. Keith; Pearce, John M.; Prosser, Diann J.; Sleeman, Jonathan M.; Whalen, Mary E.

    2015-09-09

    Avian influenza viruses are naturally occurring in wild birds such as ducks, geese, swans, and gulls. These viruses generally do not cause illness in wild birds, however, when spread to poultry they can be highly pathogenic and cause illness and death in backyard and commercial farms. Outbreaks may cause devastating agricultural economic losses and some viral strains have the potential to infect people directly. Furthermore, the combination of avian influenza viruses with mammalian viruses can result in strains with the ability to transmit from person to person, possibly leading to viruses with pandemic potential. All known pandemic influenza viruses have had some genetic material of avian origin. Since 1996, a strain of highly pathogenic avian influenza (HPAI) virus, H5N1, has caused infection in wild birds, losses to poultry farms in Eurasia and North Africa, and led to the deaths of several hundred people. Spread of the H5N1 virus and other influenza strains from China was likely facilitated by migratory birds. In December 2014, HPAI was detected in poultry in Canada and migratory birds in the United States. Since then, HPAI viruses have spread to large parts of the United States and will likely continue to spread through migratory bird flyways and other mechanisms throughout North America. In the United States, HPAI viruses have severely affected the poultry industry with millions of domestic birds dead or culled. These strains of HPAI are not known to cause disease in humans; however, the Centers for Disease Control and Prevention (CDC) advise caution when in close contact with infected birds. Experts agree that HPAI strains currently circulating in wild birds of North America will likely persist for the next few years. This unprecedented situation presents risks to the poultry industry, natural resource management, and potentially human health. Scientific knowledge and decision support tools are urgently needed to understand factors affecting the persistence

  9. USGS highly pathogenic avian influenza research strategy

    USGS Publications Warehouse

    Harris, M. Camille; Miles, A. Keith; Pearce, John M.; Prosser, Diann J.; Sleeman, Jonathan M.; Whalen, Mary E.

    2015-01-01

    Avian influenza viruses are naturally occurring in wild birds such as ducks, geese, swans, and gulls. These viruses generally do not cause illness in wild birds, however, when spread to poultry they can be highly pathogenic and cause illness and death in backyard and commercial farms. Outbreaks may cause devastating agricultural economic losses and some viral strains have the potential to infect people directly. Furthermore, the combination of avian influenza viruses with mammalian viruses can result in strains with the ability to transmit from person to person, possibly leading to viruses with pandemic potential. All known pandemic influenza viruses have had some genetic material of avian origin. Since 1996, a strain of highly pathogenic avian influenza (HPAI) virus, H5N1, has caused infection in wild birds, losses to poultry farms in Eurasia and North Africa, and led to the deaths of several hundred people. Spread of the H5N1 virus and other influenza strains from China was likely facilitated by migratory birds. In December 2014, HPAI was detected in poultry in Canada and migratory birds in the United States. Since then, HPAI viruses have spread to large parts of the United States and will likely continue to spread through migratory bird flyways and other mechanisms throughout North America. In the United States, HPAI viruses have severely affected the poultry industry with millions of domestic birds dead or culled. These strains of HPAI are not known to cause disease in humans; however, the Centers for Disease Control and Prevention (CDC) advise caution when in close contact with infected birds. Experts agree that HPAI strains currently circulating in wild birds of North America will likely persist for the next few years. This unprecedented situation presents risks to the poultry industry, natural resource management, and potentially human health. Scientific knowledge and decision support tools are urgently needed to understand factors affecting the persistence

  10. Pathogenicity of two Egyptian H5N1 highly pathogenic avian influenza viruses in domestic ducks

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Domestic ducks have been implicated in the dissemination and evolution of H5N1 highly pathogenic avian influenza (HPAI) viruses. Interestingly, the pathogenicity of H5N1 HPAI viruses in domestic ducks has increased over time with some viruses producing 100% mortality in ducks. These changes in vir...

  11. Rapidly Expanding Range of Highly Pathogenic Avian Influenza Viruses.

    PubMed

    Hall, Jeffrey S; Dusek, Robert J; Spackman, Erica

    2015-07-01

    The movement of highly pathogenic avian influenza (H5N8) virus across Eurasia and into North America and the virus' propensity to reassort with co-circulating low pathogenicity viruses raise concerns among poultry producers, wildlife biologists, aviculturists, and public health personnel worldwide. Surveillance, modeling, and experimental research will provide the knowledge required for intelligent policy and management decisions.

  12. Rapidly Expanding Range of Highly Pathogenic Avian Influenza Viruses

    PubMed Central

    Dusek, Robert J.; Spackman, Erica

    2015-01-01

    The movement of highly pathogenic avian influenza (H5N8) virus across Eurasia and into North America and the virus’ propensity to reassort with co-circulating low pathogenicity viruses raise concerns among poultry producers, wildlife biologists, aviculturists, and public health personnel worldwide. Surveillance, modeling, and experimental research will provide the knowledge required for intelligent policy and management decisions. PMID:26079209

  13. Rapidly expanding range of highly pathogenic avian influenza viruses

    USGS Publications Warehouse

    Hall, Jeffrey S.; Dusek, Robert J.; Spackman, Erica

    2015-01-01

    The movement of highly pathogenic avian influenza (H5N8) virus across Eurasia and into North America and the virus’ propensity to reassort with co-circulating low pathogenicity viruses raise concerns among poultry producers, wildlife biologists, aviculturists, and public health personnel worldwide. Surveillance, modeling, and experimental research will provide the knowledge required for intelligent policy and management decisions.

  14. Detection of highly pathogenic and low pathogenic avian influenza subtype H5 (Eurasian lineage) using NASBA.

    PubMed

    Collins, Richard A; Ko, Lung Sang; So, Ka Lun; Ellis, Trevor; Lau, Lok Ting; Yu, Albert Cheung Hoi

    2002-05-16

    Nucleic acid sequence-based amplification (NASBA) is a technique that allows the rapid amplification of specific regions of nucleic acid obtained from a diverse range of sources. It is especially suitable for amplifying RNA sequences. A NASBA technique has been developed that allows the detection of avian influenza A subtype H5 from allantoic fluid harvested from inoculated chick embryos. The amplified viral RNA is detected by electrochemiluminescence. The NASBA technique described below is rapid and specific for the identification of influenza A subtype H5 viruses of the Eurasian lineage. More importantly, it can be used to distinguish highly pathogenic and low pathogenic strains of the H5 subtype.

  15. Pathogenicity of two Egyptian H5N1 highly pathogenic avian influenza viruses in domestic ducks.

    PubMed

    Wasilenko, J L; Arafa, A M; Selim, A A; Hassan, M K; Aly, M M; Ali, A; Nassif, S; Elebiary, E; Balish, A; Klimov, A; Suarez, D L; Swayne, D E; Pantin-Jackwood, M J

    2011-01-01

    Domestic ducks have been implicated in the dissemination and evolution of H5N1 highly pathogenic avian influenza (HPAI) viruses. In this study, two H5N1 HPAI viruses belonging to clade 2.2.1 isolated in Egypt in 2007 and 2008 were analyzed for their pathogenicity in domestic Pekin ducks. Both viruses produced clinical signs and mortality, but the 2008 virus was more virulent, inducing early onset of neurological signs and killing all ducks with a mean death time (MDT) of 4.1 days. The 2007 virus killed 3/8 ducks with a MDT of 7 days. Full-genome sequencing and phylogenetic analysis were used to examine differences in the virus genes that might explain the differences observed in pathogenicity. The genomes differed in 49 amino acids, with most of the differences found in the hemagglutinin protein. This increase in pathogenicity in ducks observed with certain H5N1 HPAI viruses has implications for the control of the disease, since vaccinated ducks infected with highly virulent strains shed viruses for longer periods of time, perpetuating the virus in the environment and increasing the possibility of transmission to susceptible birds.

  16. Persistence of Highly Pathogenic Avian Influenza Viruses in Natural Ecosystems

    PubMed Central

    Feare, Chris J.; Renaud, François; Thomas, Frédéric; Gauthier-Clerc, Michel

    2010-01-01

    Understanding of ecologic factors favoring emergence and maintenance of highly pathogenic avian influenza (HPAI) viruses is limited. Although low pathogenic avian influenza viruses persist and evolve in wild populations, HPAI viruses evolve in domestic birds and cause economically serious epizootics that only occasionally infect wild populations. We propose that evolutionary ecology considerations can explain this apparent paradox. Host structure and transmission possibilities differ considerably between wild and domestic birds and are likely to be major determinants of virulence. Because viral fitness is highly dependent on host survival and dispersal in nature, virulent forms are unlikely to persist in wild populations if they kill hosts quickly or affect predation risk or migratory performance. Interhost transmission in water has evolved in low pathogenic influenza viruses in wild waterfowl populations. However, oropharyngeal shedding and transmission by aerosols appear more efficient for HPAI viruses among domestic birds. PMID:20587174

  17. Highly pathogenic avian influenza virus among wild birds in Mongolia

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The central Asian country of Mongolia supports large populations of migratory water birds that migrate across much of Asia where highly pathogenic avian influenza (HPAI) virus subtype H5N1 is endemic. This, together with the near absence of domestic poultry, makes Mongolia an ideal location to unde...

  18. Rapidly expanding range of highly pathogenic avian influenza viruses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The recent introduction of highly pathogenic avian influenza virus (HPAIV) H5N8 into Europe and North America poses significant risks to poultry industries and wildlife populations and warrants continued and heightened vigilance. First discovered in South Korean poultry and wild birds in early 2014...

  19. Imperfect Vaccination Can Enhance the Transmission of Highly Virulent Pathogens

    PubMed Central

    Read, Andrew F.; Baigent, Susan J.; Powers, Claire; Kgosana, Lydia B.; Blackwell, Luke; Smith, Lorraine P.; Kennedy, David A.; Walkden-Brown, Stephen W.; Nair, Venugopal K.

    2015-01-01

    Could some vaccines drive the evolution of more virulent pathogens? Conventional wisdom is that natural selection will remove highly lethal pathogens if host death greatly reduces transmission. Vaccines that keep hosts alive but still allow transmission could thus allow very virulent strains to circulate in a population. Here we show experimentally that immunization of chickens against Marek's disease virus enhances the fitness of more virulent strains, making it possible for hyperpathogenic strains to transmit. Immunity elicited by direct vaccination or by maternal vaccination prolongs host survival but does not prevent infection, viral replication or transmission, thus extending the infectious periods of strains otherwise too lethal to persist. Our data show that anti-disease vaccines that do not prevent transmission can create conditions that promote the emergence of pathogen strains that cause more severe disease in unvaccinated hosts. PMID:26214839

  20. Imperfect Vaccination Can Enhance the Transmission of Highly Virulent Pathogens.

    PubMed

    Read, Andrew F; Baigent, Susan J; Powers, Claire; Kgosana, Lydia B; Blackwell, Luke; Smith, Lorraine P; Kennedy, David A; Walkden-Brown, Stephen W; Nair, Venugopal K

    2015-07-01

    Could some vaccines drive the evolution of more virulent pathogens? Conventional wisdom is that natural selection will remove highly lethal pathogens if host death greatly reduces transmission. Vaccines that keep hosts alive but still allow transmission could thus allow very virulent strains to circulate in a population. Here we show experimentally that immunization of chickens against Marek's disease virus enhances the fitness of more virulent strains, making it possible for hyperpathogenic strains to transmit. Immunity elicited by direct vaccination or by maternal vaccination prolongs host survival but does not prevent infection, viral replication or transmission, thus extending the infectious periods of strains otherwise too lethal to persist. Our data show that anti-disease vaccines that do not prevent transmission can create conditions that promote the emergence of pathogen strains that cause more severe disease in unvaccinated hosts.

  1. Pathogenicity, Transmission and Antigenic Variation of H5N1 Highly Pathogenic Avian Influenza Viruses

    PubMed Central

    Jiao, Peirong; Song, Hui; Liu, Xiaoke; Song, Yafen; Cui, Jin; Wu, Siyu; Ye, Jiaqi; Qu, Nanan; Zhang, Tiemin; Liao, Ming

    2016-01-01

    H5N1 highly pathogenic avian influenza (HPAI) was one of the most important avian diseases in poultry production of China, especially in Guangdong province. In recent years, new H5N1 highly pathogenic avian influenza viruses (HPAIV) still emerged constantly, although all poultry in China were immunized with H5N1 vaccinations compulsorily. To better understand the pathogenicity and transmission of dominant clades of the H5N1 HPAIVs in chicken from Guangdong in 2012, we chose a clade 7.2 avian influenza virus named A/Chicken/China/G2/2012(H5N1) (G2) and a clade 2.3.2.1 avian influenza virus named A/Duck/China/G3/2012(H5N1) (G3) in our study. Our results showed that the chickens inoculated with 103 EID50 of G2 or G3 viruses all died, and the titers of virus replication detected in several visceral organs were high but different. In the naive contact groups, virus shedding was not detected in G2 group and all chickens survived, but virus shedding was detected in G3 group and all chickens died. These results showed that the two clades of H5N1 HPAIVs had high pathogenicity in chickens and the contact transmission of them was different in chickens. The results of cross reactive HI assay showed that antigens of G2 and G3 were very different from those of current commercial vaccines isolates (Re-4, Re-6, and D7). And to evaluate the protective efficacy of three vaccines against most isolates form Guangdong belonging to clade 2.3.2.1 in 2012, G3 was chosen to challenge the three vaccines such as Re-4, Re-6, and D7. First, chickens were immunized with 0.3 ml Re-4, Re-6, and D7 inactivated vaccines by intramuscular injection, respectively, and then challenged with 106 EID50 of G3 on day 28 post-vaccination. The D7 vaccine had 100% protection against G3 for chickens, the Re-6 vaccine had 88.9%, and the Re-4 vaccine only had 66.7%. Our results suggested that the D7 vaccine could prevent and control H5N1 virus outbreaks more effectively in Guangdong. From the above, it was

  2. Emergence of a Highly Pathogenic Avian Influenza Virus from a Low-Pathogenic Progenitor

    PubMed Central

    Fusaro, Alice; Nelson, Martha I.; Bonfanti, Lebana; Mulatti, Paolo; Hughes, Joseph; Murcia, Pablo R.; Schivo, Alessia; Valastro, Viviana; Moreno, Ana; Holmes, Edward C.; Cattoli, Giovanni

    2014-01-01

    ABSTRACT Avian influenza (AI) viruses of the H7 subtype have the potential to evolve into highly pathogenic (HP) viruses that represent a major economic problem for the poultry industry and a threat to global health. However, the emergence of HPAI viruses from low-pathogenic (LPAI) progenitor viruses currently is poorly understood. To investigate the origin and evolution of one of the most important avian influenza epidemics described in Europe, we investigated the evolutionary and spatial dynamics of the entire genome of 109 H7N1 (46 LPAI and 63 HPAI) viruses collected during Italian H7N1 outbreaks between March 1999 and February 2001. Phylogenetic analysis revealed that the LPAI and HPAI epidemics shared a single ancestor, that the HPAI strains evolved from the LPAI viruses in the absence of reassortment, and that there was a parallel emergence of mutations among HPAI and later LPAI lineages. Notably, an ultradeep-sequencing analysis demonstrated that some of the amino acid changes characterizing the HPAI virus cluster were already present with low frequency within several individual viral populations from the beginning of the LPAI H7N1 epidemic. A Bayesian phylogeographic analysis revealed stronger spatial structure during the LPAI outbreak, reflecting the more rapid spread of the virus following the emergence of HPAI. The data generated in this study provide the most complete evolutionary and phylogeographic analysis of epidemiologically intertwined high- and low-pathogenicity viruses undertaken to date and highlight the importance of implementing prompt eradication measures against LPAI to prevent the appearance of viruses with fitness advantages and unpredictable pathogenic properties. IMPORTANCE The Italian H7 AI epidemic of 1999 to 2001 was one of the most important AI outbreaks described in Europe. H7 viruses have the ability to evolve into HP forms from LP precursors, although the mechanisms underlying this evolutionary transition are only poorly

  3. Decontamination of High-risk Animal and Zoonotic Pathogens

    PubMed Central

    Menrath, Andrea; Tomuzia, Katharina; Braeunig, Juliane; Appel, Bernd

    2013-01-01

    Preparedness for the decontamination of affected environments, premises, facilities, and products is one prerequisite for an immediate response to an animal disease outbreak. Various information sources provide recommendations on how to proceed in an outbreak situation to eliminate biological contaminants and to stop the spread of the disease. In order to facilitate the identification of the right decontamination strategy, we present an overview of relevant references for a collection of pathogenic agents. The choice of pathogens is based on a survey of lists containing highly pathogenic agents and/or biological agents considered to be potential vehicles for deliberate contamination of food, feed, or farm animals. European legislation and guidelines from national and international institutions were screened to find decontamination protocols for each of the agents. Identified recommendations were evaluated with regard to their area of application, which could be facilities and equipment, wastes, food, and other animal products. The requirements of a disinfectant for large-scale incidents were gathered, and important characteristics (eg, inactivating spectrum, temperature range, toxicity to environment) of the main recommended disinfectants were summarized to assist in the choice of a suitable and efficient approach in a crisis situation induced by a specific high-risk animal or zoonotic pathogen. The literature search revealed numerous relevant recommendations but also legal gaps for certain diseases, such as Q fever or brucellosis, and legal difficulties for the use of recommended disinfectants. A lack of information about effective disinfectants was identified for some agents. PMID:23971795

  4. Identification of peptidases in highly pathogenic vs. weakly pathogenic Naegleria fowleri amebae.

    PubMed

    Vyas, Ishan K; Jamerson, Melissa; Cabral, Guy A; Marciano-Cabral, Francine

    2015-01-01

    Naegleria fowleri, a free-living ameba, is the causative agent of Primary Amebic Meningoencephalitis. Highly pathogenic mouse-passaged amebae (Mp) and weakly pathogenic axenically grown (Ax) N. fowleri were examined for peptidase activity. Zymography and azocasein peptidase activity assays demonstrated that Mp and Ax N. fowleri exhibited a similar peptidase pattern. Prominent for whole cell lysates, membranes and conditioned medium (CM) from Mp and Ax amebae was the presence of an activity band of approximately 58 kDa that was sensitive to E64, a cysteine peptidase inhibitor. However, axenically grown N. fowleri demonstrated a high level of this peptidase activity in membrane preparations. The inhibitor E64 also reduced peptidase activity in ameba-CM consistent with the presence of secreted cysteine peptidases. Exposure of Mp amebae to E64 reduced their migration through matrigel that was used as an extracellular matrix, suggesting a role for cysteine peptidases in invasion of the central nervous system (CNS). The collective results suggest that the profile of peptidases is not a discriminative marker for distinguishing Mp from Ax N. fowleri. However, the presence of a prominent level of activity for cysteine peptidases in N. fowleri membranes and CM, suggests that these enzymes may serve to facilitate passage of the amebae into the CNS. PMID:25066578

  5. Identification of peptidases in highly pathogenic vs. weakly pathogenic Naegleria fowleri amebae.

    PubMed

    Vyas, Ishan K; Jamerson, Melissa; Cabral, Guy A; Marciano-Cabral, Francine

    2015-01-01

    Naegleria fowleri, a free-living ameba, is the causative agent of Primary Amebic Meningoencephalitis. Highly pathogenic mouse-passaged amebae (Mp) and weakly pathogenic axenically grown (Ax) N. fowleri were examined for peptidase activity. Zymography and azocasein peptidase activity assays demonstrated that Mp and Ax N. fowleri exhibited a similar peptidase pattern. Prominent for whole cell lysates, membranes and conditioned medium (CM) from Mp and Ax amebae was the presence of an activity band of approximately 58 kDa that was sensitive to E64, a cysteine peptidase inhibitor. However, axenically grown N. fowleri demonstrated a high level of this peptidase activity in membrane preparations. The inhibitor E64 also reduced peptidase activity in ameba-CM consistent with the presence of secreted cysteine peptidases. Exposure of Mp amebae to E64 reduced their migration through matrigel that was used as an extracellular matrix, suggesting a role for cysteine peptidases in invasion of the central nervous system (CNS). The collective results suggest that the profile of peptidases is not a discriminative marker for distinguishing Mp from Ax N. fowleri. However, the presence of a prominent level of activity for cysteine peptidases in N. fowleri membranes and CM, suggests that these enzymes may serve to facilitate passage of the amebae into the CNS.

  6. Mobility of the Yersinia High-Pathogenicity Island (HPI): transfer mechanisms of pathogenicity islands (PAIS) revisited (a review).

    PubMed

    Benedek, Orsolya; Schubert, S

    2007-06-01

    Horizontal gene transfer (HGT) plays a key role in the evolution of bacterial pathogens. The exchange of genetic material supplies prokaryotes with several fitness traits enhancing their adaptive response to environmental changes. Pathogenicity islands (PAIs) represent an important and in most cases already immobilized subset of the different vehicles for HGT. Encoding several virulence factors PAls represent a major contribution to bacterial pathogenicity. Nonetheless, the transfer mechanisms of PAIs still remain elusive. We summarise the currently available data regarding the major ways of genetic mobilisation with a focus on the transfer of the Yersinia High-Pathogenicity Island (HPI).

  7. Highly pathogenic avian influenza outbreak mitigated by seasonal low pathogenic strains: insights from dynamic modeling.

    PubMed

    Bourouiba, L; Teslya, A; Wu, J

    2011-02-21

    The spread of highly pathogenic avian influenza (HPAI) H5N1 remains a threat for both wild and domestic bird populations, while low pathogenic avian influenza (LPAI) strains have been reported to induce partial immunity to HPAI in poultry and some wild birds inoculated with both HPAI and LPAI strains. Here, based on the reported data and experiments, we develop a two-strain avian influenza model to examine the extent to which this partial immunity observed at the individual level can affect the outcome of the outbreaks among migratory birds in the wild at the population level during different seasons. We find a distinct mitigating effect of LPAI on the death toll induced by HPAI strain, and this effect is particularly important for populations previously exposed to and recovered from LPAI. We further investigate the effect of the dominant mode of transmission of an HPAI strain on the outcome of the epidemic. Four combinations of contact based direct transmission and indirect fecal-to-oral (or environmental) routes are examined. For a given infection peak of HPAI, indirect fecal-to-oral transmission of HPAI can lead to a higher death toll than that associated with direct transmission. The mitigating effect of LPAI can, in turn, be dependent on the route of infection of HPAI. PMID:21146544

  8. High Recombinant Frequency in Extraintestinal Pathogenic Escherichia coli Strains.

    PubMed

    Rodríguez-Beltrán, Jerónimo; Tourret, Jérôme; Tenaillon, Olivier; López, Elena; Bourdelier, Emmanuelle; Costas, Coloma; Matic, Ivan; Denamur, Erick; Blázquez, Jesús

    2015-07-01

    Homologous recombination promotes genetic diversity by facilitating the integration of foreign DNA and intrachromosomal gene shuffling. It has been hypothesized that if recombination is variable among strains, selection should favor higher recombination rates among pathogens, as they face additional selection pressures from host defenses. To test this hypothesis we have developed a plasmid-based method for estimating the rate of recombination independently of other factors such as DNA transfer, selective processes, and mutational interference. Our results with 160 human commensal and extraintestinal pathogenic Escherichia coli (ExPEC) isolates show that the recombinant frequencies are extremely diverse (ranging 9 orders of magnitude) and plastic (they are profoundly affected by growth in urine, a condition commonly encountered by ExPEC). We find that the frequency of recombination is biased by strain lifestyle, as ExPEC isolates display strikingly higher recombination rates than their commensal counterparts. Furthermore, the presence of virulence factors is positively associated with higher recombination frequencies. These results suggest selection for high homologous recombination capacity, which may result in a higher evolvability for pathogens compared with commensals.

  9. Quantifying Transmission of Highly Pathogenic and Low Pathogenicity H7N1 Avian Influenza in Turkeys

    PubMed Central

    Saenz, Roberto A.; Essen, Steve C.; Brookes, Sharon M.; Iqbal, Munir; Wood, James L. N.; Grenfell, Bryan T.; McCauley, John W.; Brown, Ian H.; Gog, Julia R.

    2012-01-01

    Outbreaks of avian influenza in poultry can be devastating, yet many of the basic epidemiological parameters have not been accurately characterised. In 1999–2000 in Northern Italy, outbreaks of H7N1 low pathogenicity avian influenza virus (LPAI) were followed by the emergence of H7N1 highly pathogenic avian influenza virus (HPAI). This study investigates the transmission dynamics in turkeys of representative HPAI and LPAI H7N1 virus strains from this outbreak in an experimental setting, allowing direct comparison of the two strains. The fitted transmission rates for the two strains are similar: 2.04 (1.5–2.7) per day for HPAI, 2.01 (1.6–2.5) per day for LPAI. However, the mean infectious period is far shorter for HPAI (1.47 (1.3–1.7) days) than for LPAI (7.65 (7.0–8.3) days), due to the rapid death of infected turkeys. Hence the basic reproductive ratio, is significantly lower for HPAI (3.01 (2.2–4.0)) than for LPAI (15.3 (11.8–19.7)). The comparison of transmission rates and are critically important in relation to understanding how HPAI might emerge from LPAI. Two competing hypotheses for how transmission rates vary with population size are tested by fitting competing models to experiments with differing numbers of turkeys. A model with frequency-dependent transmission gives a significantly better fit to experimental data than density-dependent transmission. This has important implications for extrapolating experimental results from relatively small numbers of birds to the commercial poultry flock size, and for how control, including vaccination, might scale with flock size. PMID:23028760

  10. High-level fluorescence labeling of gram-positive pathogens.

    PubMed

    Aymanns, Simone; Mauerer, Stefanie; van Zandbergen, Ger; Wolz, Christiane; Spellerberg, Barbara

    2011-01-01

    Fluorescence labeling of bacterial pathogens has a broad range of interesting applications including the observation of living bacteria within host cells. We constructed a novel vector based on the E. coli streptococcal shuttle plasmid pAT28 that can propagate in numerous bacterial species from different genera. The plasmid harbors a promoterless copy of the green fluorescent variant gene egfp under the control of the CAMP-factor gene (cfb) promoter of Streptococcus agalactiae and was designated pBSU101. Upon transfer of the plasmid into streptococci, the bacteria show a distinct and easily detectable fluorescence using a standard fluorescence microscope and quantification by FACS-analysis demonstrated values that were 10-50 times increased over the respective controls. To assess the suitability of the construct for high efficiency fluorescence labeling in different gram-positive pathogens, numerous species were transformed. We successfully labeled Streptococcus pyogenes, Streptococcus agalactiae, Streptococcus dysgalactiae subsp. equisimilis, Enterococcus faecalis, Enterococcus faecium, Streptococcus mutans, Streptococcus anginosus and Staphylococcus aureus strains utilizing the EGFP reporter plasmid pBSU101. In all of these species the presence of the cfb promoter construct resulted in high-level EGFP expression that could be further increased by growing the streptococcal and enterococcal cultures under high oxygen conditions through continuous aeration.

  11. Evidence for two evolutionary lineages of highly pathogenic Yersinia species.

    PubMed Central

    Rakin, A; Urbitsch, P; Heesemann, J

    1995-01-01

    Sensitivity to Yersinia pestis bacteriocin pesticin correlates with the existence of two groups of human pathogenic yersiniae, mouse lethal and mouse nonlethal. The presence of the outer membrane pesticin receptor (FyuA) in mouse-lethal yersiniae is a prerequisite for pesticin sensitivity. Genes that code for FyuA (fyuA) were identified and sequenced from pesticin-sensitive bacteria, including Y. enterocolitica biotype 1B (serotypes O8; O13, O20, and O21), Y. pseudotuberculosis serotype O1, Y. pestis, two known pesticin-sensitive Escherichia coli isolates (E. coli Phi and E. coli CA42), and two newly discovered pesticin-sensitive isolates, E. coli K49 and K235. A 2,318-bp fyuA sequence was shown to be highly conserved in all pesticin-sensitive bacteria, including E. coli strains (DNA sequence homology was 98.5 to 99.9%). The same degree of DNA homology (97.8 to 100%) was established for the sequenced 276-bp fragment of the irp2 gene that encodes high-molecular-weight protein 2, which is also thought to be involved in the expression of virulence by Yersinia species. Highly conserved irp2 was also found in all pesticin-sensitive E. coli strains. On the basis of the fyuA and irp2 sequence homologies, two evolutionary groups of highly pathogenic Yersinia species can be established. One group includes Y. enterocolitica biotype 1B strains, while the second includes Y. pestis, Y. pseudotuberculosis serotype O1, and irp2-positive Y. pseudotuberculosis serotype O3 strains. E. coli Phi, CA42, K49, and K235 belong to the second group. The possible proximity of these two iron-regulated genes (fyuA and irp2), as well as their high levels of sequence conservation and similar G+C contents (56.2 and 59.8 mol%), leads to the assumption that these two genes may represent part of an unstable pathogenicity island that has been acquired by pesticin-sensitive bacteria as a result of a horizontal transfer. PMID:7730256

  12. Highly Pathogenic Avian Influenza Virus among Wild Birds in Mongolia

    PubMed Central

    Gilbert, Martin; Jambal, Losolmaa; Karesh, William B.; Fine, Amanda; Shiilegdamba, Enkhtuvshin; Dulam, Purevtseren; Sodnomdarjaa, Ruuragchaa; Ganzorig, Khuukhenbaatar; Batchuluun, Damdinjav; Tseveenmyadag, Natsagdorj; Bolortuya, Purevsuren; Cardona, Carol J.; Leung, Connie Y. H.; Peiris, J. S. Malik; Spackman, Erica; Swayne, David E.; Joly, Damien O.

    2012-01-01

    Mongolia combines a near absence of domestic poultry, with an abundance of migratory waterbirds, to create an ideal location to study the epidemiology of highly pathogenic avian influenza virus (HPAIV) in a purely wild bird system. Here we present the findings of active and passive surveillance for HPAIV subtype H5N1 in Mongolia from 2005–2011, together with the results of five outbreak investigations. In total eight HPAIV outbreaks were confirmed in Mongolia during this period. Of these, one was detected during active surveillance employed by this project, three by active surveillance performed by Mongolian government agencies, and four through passive surveillance. A further three outbreaks were recorded in the neighbouring Tyva Republic of Russia on a lake that bisects the international border. No HPAIV was isolated (cultured) from 7,855 environmental fecal samples (primarily from ducks), or from 2,765 live, clinically healthy birds captured during active surveillance (primarily shelducks, geese and swans), while four HPAIVs were isolated from 141 clinically ill or dead birds located through active surveillance. Two low pathogenic avian influenza viruses (LPAIV) were cultured from ill or dead birds during active surveillance, while environmental feces and live healthy birds yielded 56 and 1 LPAIV respectively. All Mongolian outbreaks occurred in 2005 and 2006 (clade 2.2), or 2009 and 2010 (clade 2.3.2.1); all years in which spring HPAIV outbreaks were reported in Tibet and/or Qinghai provinces in China. The occurrence of outbreaks in areas deficient in domestic poultry is strong evidence that wild birds can carry HPAIV over at least moderate distances. However, failure to detect further outbreaks of clade 2.2 after June 2006, and clade 2.3.2.1 after June 2010 suggests that wild birds migrating to and from Mongolia may not be competent as indefinite reservoirs of HPAIV, or that HPAIV did not reach susceptible populations during our study. PMID:22984464

  13. Biological characteristics and pathogenicity of a highly pathogenic Shewanella marisflavi infected sea cucumber (Apostichopus uaponicus)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Shewanella marisflavi isolate AP629 was characterized as a novel pathogen of sea cucumber. The LD50 values (14 days) in sea cucumber and swordtail fish were 3.89 × 106 and 4.85 × 104 CFU g-1 body weight, respectively. Studies on S. marisflavi had been conducted, including morphology, physiological a...

  14. USGS role and response to highly pathogenic avian influenza

    USGS Publications Warehouse

    Harris, M. Camille; Miles, A. Keith; Pearce, John M.; Prosser, Diann J.; Sleeman, Jonathan M.; Whalen, Mary E.

    2015-09-09

    Avian influenza viruses are naturally occurring in wild birds such as ducks, geese, swans, and gulls. These viruses generally do not cause illness in wild birds, however, when spread to poultry they can be highly pathogenic and cause illness and death in backyard and commercial farms. Outbreaks may cause devastating agricultural economic losses and some viral strains have the potential to infect people directly. Furthermore, the combination of avian influenza viruses with mammalian viruses can result in strains with the ability to transmit from person to person, possibly leading to viruses with pandemic potential. All known pandemic influenza viruses have had some genetic material of avian origin. Since 1996, a strain of highly pathogenic avian influenza (HPAI) virus, H5N1, has caused infection in wild birds, losses to poultry farms in Eurasia and North Africa, and led to the deaths of several hundred people. Spread of the H5N1 virus and other influenza strains from China was likely facilitated by migratory birds. In December 2014, HPAI was detected in poultry in Canada and migratory birds in the United States. Since then, HPAI viruses have spread to large parts of the United States and will likely continue to spread through migratory bird flyways and other mechanisms throughout North America. In the United States, HPAI viruses have severely affected the poultry industry with millions of domestic birds dead or culled. These strains of HPAI are not known to cause disease in humans; however, the Centers for Disease Control and Prevention (CDC) advise caution when in close contact with infected birds. Experts agree that HPAI strains currently circulating in wild birds of North America will likely persist for the next few years. This unprecedented situation presents risks to the poultry industry, natural resource management, and potentially human health. Scientific knowledge and decision support tools are urgently needed to understand factors affecting the persistence

  15. USGS role and response to highly pathogenic avian influenza

    USGS Publications Warehouse

    Harris, M. Camille; Miles, A. Keith; Pearce, John M.; Prosser, Diann J.; Sleeman, Jonathan M.; Whalen, Mary E.

    2015-01-01

    Avian influenza viruses are naturally occurring in wild birds such as ducks, geese, swans, and gulls. These viruses generally do not cause illness in wild birds, however, when spread to poultry they can be highly pathogenic and cause illness and death in backyard and commercial farms. Outbreaks may cause devastating agricultural economic losses and some viral strains have the potential to infect people directly. Furthermore, the combination of avian influenza viruses with mammalian viruses can result in strains with the ability to transmit from person to person, possibly leading to viruses with pandemic potential. All known pandemic influenza viruses have had some genetic material of avian origin. Since 1996, a strain of highly pathogenic avian influenza (HPAI) virus, H5N1, has caused infection in wild birds, losses to poultry farms in Eurasia and North Africa, and led to the deaths of several hundred people. Spread of the H5N1 virus and other influenza strains from China was likely facilitated by migratory birds. In December 2014, HPAI was detected in poultry in Canada and migratory birds in the United States. Since then, HPAI viruses have spread to large parts of the United States and will likely continue to spread through migratory bird flyways and other mechanisms throughout North America. In the United States, HPAI viruses have severely affected the poultry industry with millions of domestic birds dead or culled. These strains of HPAI are not known to cause disease in humans; however, the Centers for Disease Control and Prevention (CDC) advise caution when in close contact with infected birds. Experts agree that HPAI strains currently circulating in wild birds of North America will likely persist for the next few years. This unprecedented situation presents risks to the poultry industry, natural resource management, and potentially human health. Scientific knowledge and decision support tools are urgently needed to understand factors affecting the persistence

  16. Emergence of highly pathogenic virus during selective chicken passage of the prototype mildly pathogenic chicken/Pennsylvania/83 (H5N2) influenza virus.

    PubMed

    Brugh, M; Perdue, M L

    1991-01-01

    The prototype mildly pathogenic A/chicken/Pennsylvania/21525/83 (H5N2) avian influenza virus, which was isolated more than 5 months before the emergence of highly pathogenic virus in the major 1983 Pennsylvania outbreak, was examined for the presence of minority subpopulations of highly pathogenic virus. Selective serial passage of the parental mildly pathogenic virus in leghorn hens did not lead to recovery of highly pathogenic virus. However, several highly pathogenic reisolates were recovered from hens inoculated with either of two mildly pathogenic virus clones selected for their ability to efficiently produce plaques in trypsin-free chicken embryo fibroblasts. Unlike the parental virus, these reisolates caused high mortality in chickens and produced postmortem lesions typical of highly pathogenic avian influenza. Electrophoretic mobilities of the hemagglutinin glycoproteins of the highly pathogenic derivatives resembled those of the prototype highly pathogenic A/chicken/Pennsylvania/1370/83 (H5N2) virus isolated in October 1983. These results suggest that unrecognized subpopulations of highly pathogenic virus may have infected Pennsylvania chickens for several months before emerging as the clinically manifest component of the virus population.

  17. High prevalence of pathogenic Yersinia enterocolitica in pig cheeks.

    PubMed

    Laukkanen-Ninios, Riikka; Fredriksson-Ahomaa, Maria; Maijala, Riitta; Korkeala, Hannu

    2014-10-01

    Samples from pork cuts for minced meat and cheeks from processing plants and a slaughterhouse, and modified atmosphere (MA) packaged pork from retail were studied to estimate the prevalence of pathogenic, i.e. virulence plasmid bearing, Yersinia enterocolitica and Yersinia pseudotuberculosis in pork, as well as to quantify pathogenic Y. enterocolitica in pork cuts. Pathogenic (virF-positive) Y. enterocolitica was isolated from 17 pig cheeks (23%) but not from any of the MA-packaged 54 retail pork samples and only from one of the 155 pork cut (0.6%). Most (16/17) of the cheek samples were contaminated with pathogenic Y. enterocolitica 4/O:3 and one with bioserotype 2/O:9. No Y. pseudotuberculosis was isolated. The prevalence of pathogenic Y. enterocolitica was clearly higher (39%) in 155 pork cuts when studied with nested PCR targeting yadA on the virulence plasmid pYV although the contamination level was low varying between 0.1 and 1.6 MPN/g. Raw pork cuts and especially pig cheeks may serve as possible sources for yersiniosis caused by pathogenic Y. enterocolitica.

  18. Control strategies for highly pathogenic avian influenza: a global perspective.

    PubMed

    Lubroth, J

    2007-01-01

    Comprehensive programmes for the prevention, detection and control of highly pathogenic avian influenza (HPAI) require a national dimension and relevant national legislation in which veterinary services can conduct surveillance, competent diagnosis and rapid response. Avian influenza was controlled and prevented by vaccination long before the current H5N1 crisis. The use of vaccine cannot be separated from other essential elements of a vaccination campaign, which include education in poultry production practices, such as hygiene, all in-all out production concepts, separation of species, biosecurity (bio-exclusion to keep the disease out and biocontainment to keep the disease from spreading once suspected or detected), competence in giving the vaccine and the role of vaccination teams, post-vaccination monitoring to ensure efficacy and to detect the circulation of wild-type virus, surveillance and buffer zones in outbreak areas, and performance indicators to determine when vaccination can cease. Reporting of disease can be improved through well-structured, adequately financed veterinary services and also by fair compensation for producers who suffer financial loss. A rapid response to suspected cases of HPAI should be ensured in simulation exercises involving various sectors of the food production and marketing chain, policy-makers, official veterinary structures and other government personnel. As for other transboundary animal diseases, national approaches must be part of a regional strategy and regional networks for cooperation and information sharing, which in turn reflect global policies and international standards, such as the quality of vaccines, reporting obligations, humane interventions, cleaning and disinfection methods, restocking times, monitoring and safe trade.

  19. Rapid and high-throughput detection of highly pathogenic bacteria by Ibis PLEX-ID technology.

    PubMed

    Jacob, Daniela; Sauer, Uschi; Housley, Roberta; Washington, Cicely; Sannes-Lowery, Kristin; Ecker, David J; Sampath, Rangarajan; Grunow, Roland

    2012-01-01

    In this manuscript, we describe the identification of highly pathogenic bacteria using an assay coupling biothreat group-specific PCR with electrospray ionization mass spectrometry (PCR/ESI-MS) run on an Ibis PLEX-ID high-throughput platform. The biothreat cluster assay identifies most of the potential bioterrorism-relevant microorganisms including Bacillus anthracis, Francisella tularensis, Yersinia pestis, Burkholderia mallei and pseudomallei, Brucella species, and Coxiella burnetii. DNA from 45 different reference materials with different formulations and different concentrations were chosen and sent to a service screening laboratory that uses the PCR/ESI-MS platform to provide a microbial identification service. The standard reference materials were produced out of a repository built up in the framework of the EU funded project "Establishment of Quality Assurances for Detection of Highly Pathogenic Bacteria of Potential Bioterrorism Risk" (EQADeBa). All samples were correctly identified at least to the genus level.

  20. Increased virulence in ducks of H5N1 highly pathogenic avian influenza viruses from Egypt

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The pathogenicity of H5N1 highly pathogenic avian influenza (HPAI) viruses in domestic ducks has increased over time. These changes in virulence have been reported with viruses from countries with high population of domestic ducks. Since 2006, H5N1 HPAI outbreaks in Egypt have been occurring in po...

  1. Detecting human bacterial pathogens in wastewater treatment plants by a high-throughput shotgun sequencing technique.

    PubMed

    Cai, Lin; Zhang, Tong

    2013-05-21

    Human pathogens are one of the major threats to global public health. Wastewater treatment plants (WWTPs) serve as city guts to receive and digest various human pathogens. Several techniques have been developed to detect human pathogens in WWTPs and to assess potential environmental risks. In this study, we employed 24 metagenomic DNA data sets derived from a high-throughput shotgun sequencing technique to more accurately and efficiently detect human bacterial pathogens in influent, activated sludge, and effluent of two Hong Kong WWTPs. Each data set was quality-filtered and normalized to 12,500,000 DNA sequences with a length of 150-190 bp. Then, a BLASTN search against Greengenes general 16S rRNA gene database and human pathogenic bacteria 16S rRNA gene database, a BLASTX search against human pathogenic bacteria virulence factor database, as well as MetaPhlAn analysis were conducted to survey the distribution, diversity, and abundance of human bacterial pathogens. The results revealed that (i) nine bacterial pathogens were detected; (ii) the overall pathogenic bacteria abundance was estimated as 0.06-3.20% in the total bacteria population using 16S rRNA gene fingerprinting; (iii) pathogenic bacteria detected in activated sludge and effluent shared similar profiles but were different from influent based on both 16S rRNA gene and virulence factor fingerprintings; (iv) Mycobacterium tuberculosis -like species may present potential pathogenic risks because it was detected with high abundance in both activated sludge and effluent. These findings provided a comprehensive profile of commonly concerned human pathogens in two Hong Kong WWTPs and demonstrated that the high-throughput shotgun sequencing technique is a feasible and effectual approach for environmental detection of human bacterial pathogens.

  2. Highly pathogenic avian influenza challenge studies in waterfowl

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Waterfowl are the natural hosts of avian influenza (AI) virus. The majority of AI viruses are classified as low pathogenicity (LP) based on their virulence in chickens, which are the reference species for pathotype testing and can be any of the 16 hemagglutinin subtypes (H1-16). Circulation of H5 ...

  3. High-Throughput Microfluidic Method To Study Biofilm Formation and Host-Pathogen Interactions in Pathogenic Escherichia coli

    PubMed Central

    Tremblay, Yannick D. N.; Vogeleer, Philippe; Jacques, Mario

    2015-01-01

    Biofilm formation and host-pathogen interactions are frequently studied using multiwell plates; however, these closed systems lack shear force, which is present at several sites in the host, such as the intestinal and urinary tracts. Recently, microfluidic systems that incorporate shear force and very small volumes have been developed to provide cell biology models that resemble in vivo conditions. Therefore, the objective of this study was to determine if the BioFlux 200 microfluidic system could be used to study host-pathogen interactions and biofilm formation by pathogenic Escherichia coli. Strains of various pathotypes were selected to establish the growth conditions for the formation of biofilms in the BioFlux 200 system on abiotic (glass) or biotic (eukaryotic-cell) surfaces. Biofilm formation on glass was observed for the majority of strains when they were grown in M9 medium at 30°C but not in RPMI medium at 37°C. In contrast, HRT-18 cell monolayers enhanced binding and, in most cases, biofilm formation by pathogenic E. coli in RPMI medium at 37°C. As a proof of principle, the biofilm-forming ability of a diffusely adherent E. coli mutant strain lacking AIDA-I, a known mediator of attachment, was assessed in our models. In contrast to the parental strain, which formed a strong biofilm, the mutant formed a thin biofilm on glass or isolated clusters on HRT-18 monolayers. In conclusion, we describe a microfluidic method for high-throughput screening that could be used to identify novel factors involved in E. coli biofilm formation and host-pathogen interactions under shear force. PMID:25681176

  4. Engineered nanoconstructs for the multiplexed and sensitive detection of high-risk pathogens

    NASA Astrophysics Data System (ADS)

    Seo, Youngmin; Kim, Ji-Eun; Jeong, Yoon; Lee, Kwan Hong; Hwang, Jangsun; Hong, Jongwook; Park, Hansoo; Choi, Jonghoon

    2016-01-01

    Many countries categorize the causative agents of severe infectious diseases as high-risk pathogens. Given their extreme infectivity and potential to be used as biological weapons, a rapid and sensitive method for detection of high-risk pathogens (e.g., Bacillus anthracis, Francisella tularensis, Yersinia pestis, and Vaccinia virus) is highly desirable. Here, we report the construction of a novel detection platform comprising two units: (1) magnetic beads separately conjugated with multiple capturing antibodies against four different high-risk pathogens for simple and rapid isolation, and (2) genetically engineered apoferritin nanoparticles conjugated with multiple quantum dots and detection antibodies against four different high-risk pathogens for signal amplification. For each high-risk pathogen, we demonstrated at least 10-fold increase in sensitivity compared to traditional lateral flow devices that utilize enzyme-based detection methods. Multiplexed detection of high-risk pathogens in a sample was also successful by using the nanoconstructs harboring the dye molecules with fluorescence at different wavelengths. We ultimately envision the use of this novel nanoprobe detection platform in future applications that require highly sensitive on-site detection of high-risk pathogens.

  5. Highly Pathogenic New World and Old World Human Arenaviruses Induce Distinct Interferon Responses in Human Cells

    PubMed Central

    Huang, Cheng; Kolokoltsova, Olga A.; Yun, Nadezhda E.; Seregin, Alexey V.; Ronca, Shannon; Koma, Takaaki

    2015-01-01

    ABSTRACT The arenavirus family includes several important pathogens that cause severe and sometimes fatal diseases in humans. The highly pathogenic Old World (OW) arenavirus Lassa fever virus (LASV) is the causative agent of Lassa fever (LF) disease in humans. LASV infections in severe cases are generally immunosuppressive without stimulating interferon (IFN) induction, a proinflammatory response, or T cell activation. However, the host innate immune responses to highly pathogenic New World (NW) arenaviruses are not well understood. We have previously shown that the highly pathogenic NW arenavirus, Junin virus (JUNV), induced an IFN response in human A549 cells. Here, we report that Machupo virus (MACV), another highly pathogenic NW arenavirus, also induces an IFN response. Importantly, both pathogenic NW arenaviruses, in contrast to the OW highly pathogenic arenavirus LASV, readily elicited an IFN response in human primary dendritic cells and A549 cells. Coinfection experiments revealed that LASV could potently inhibit MACV-activated IFN responses even at 6 h after MACV infection, while the replication levels of MACV and LASV were not affected by virus coinfection. Our results clearly demonstrated that although all viruses studied herein are highly pathogenic to humans, the host IFN responses toward infections with the NW arenaviruses JUNV and MACV are quite different from responses to infections with the OW arenavirus LASV, a discovery that needs to be further investigated in relevant animal models. This finding might help us better understand various interplays between the host immune system and highly pathogenic arenaviruses as well as distinct mechanisms underlying viral pathogenesis. IMPORTANCE Infections of humans with the highly pathogenic OW LASV are accompanied by potent suppression of interferon or proinflammatory cytokine production. In contrast, infections with the highly pathogenic NW arenavirus JUNV are associated with high levels of IFNs and

  6. Reduced Set of Virulence Genes Allows High Accuracy Prediction of Bacterial Pathogenicity in Humans

    PubMed Central

    Iraola, Gregorio; Vazquez, Gustavo; Spangenberg, Lucía; Naya, Hugo

    2012-01-01

    Although there have been great advances in understanding bacterial pathogenesis, there is still a lack of integrative information about what makes a bacterium a human pathogen. The advent of high-throughput sequencing technologies has dramatically increased the amount of completed bacterial genomes, for both known human pathogenic and non-pathogenic strains; this information is now available to investigate genetic features that determine pathogenic phenotypes in bacteria. In this work we determined presence/absence patterns of different virulence-related genes among more than finished bacterial genomes from both human pathogenic and non-pathogenic strains, belonging to different taxonomic groups (i.e: Actinobacteria, Gammaproteobacteria, Firmicutes, etc.). An accuracy of 95% using a cross-fold validation scheme with in-fold feature selection is obtained when classifying human pathogens and non-pathogens. A reduced subset of highly informative genes () is presented and applied to an external validation set. The statistical model was implemented in the BacFier v1.0 software (freely available at ), that displays not only the prediction (pathogen/non-pathogen) and an associated probability for pathogenicity, but also the presence/absence vector for the analyzed genes, so it is possible to decipher the subset of virulence genes responsible for the classification on the analyzed genome. Furthermore, we discuss the biological relevance for bacterial pathogenesis of the core set of genes, corresponding to eight functional categories, all with evident and documented association with the phenotypes of interest. Also, we analyze which functional categories of virulence genes were more distinctive for pathogenicity in each taxonomic group, which seems to be a completely new kind of information and could lead to important evolutionary conclusions. PMID:22916122

  7. Pathogenicity of the Korean H5N8 highly pathogenic avian influenza virus in commercial domestic poultry species.

    PubMed

    Lee, Dong-Hun; Kwon, Jung-Hoon; Noh, Jin-Yong; Park, Jae-Keun; Yuk, Seong-Su; Erdene-Ochir, Tseren-Ochir; Lee, Joong-Bok; Park, Seung-Yong; Choi, In-Soo; Lee, Sang-Won; Song, Chang-Seon

    2016-01-01

    In 2014, the highly pathogenic avian influenza (HPAI) virus H5N8 triggered outbreaks in wild birds and poultry farms in South Korea. In the present study, we investigated the pathogenicity of the H5N8 HPAI virus, belonging to the clade 2.3.4.4, in different species of poultry. For this, we examined clinical signs and viral shedding levels following intranasal inoculation of the virus in 3-week-old commercial layer chickens and quails, 10-week-old Korean native chickens, and 8-week-old Muscovy ducks. Intranasal inoculation with 10(6.0) viruses at 50% egg-infective dose resulted in 100% mortality in the layer chickens (8/8) and quails (4/4), but 60% and 0% deaths in the Korean native chickens (3/5) and Muscovy ducks (0/4), respectively. In addition, transmission of the inoculated virus to contact-exposed birds was evident in all the species used in this study. Based on our results, we conclude that the H5N8 HPAI virus has lower pathogenicity and transmissibility in poultry species compared with previously reported H5N1 HPAI viruses. PMID:26814367

  8. High doses of highly pathogenic avian influenza virus in chicken meat are required to infect ferrets

    PubMed Central

    2014-01-01

    High pathogenicity avian influenza viruses (HPAIV) have caused fatal infections in mammals through consumption of infected bird carcasses or meat, but scarce information exists on the dose of virus required and the diversity of HPAIV subtypes involved. Ferrets were exposed to different HPAIV (H5 and H7 subtypes) through consumption of infected chicken meat. The dose of virus needed to infect ferrets through consumption was much higher than via respiratory exposure and varied with the virus strain. In addition, H5N1 HPAIV produced higher titers in the meat of infected chickens and more easily infected ferrets than the H7N3 or H7N7 HPAIV. PMID:24894438

  9. Differential immune response of mallard duck peripheral blood mononuclear cells to two highly pathogenic avian influenza H5N1 viruses with distinct pathogenicity in mallard ducks.

    PubMed

    Cui, Zhu; Hu, Jiao; He, Liang; Li, Qunhui; Gu, Min; Wang, Xiaoquan; Hu, Shunlin; Liu, Huimou; Liu, Wenbo; Liu, Xiaowen; Liu, Xiufan

    2014-02-01

    CK10 and GS10 are two H5N1 highly pathogenic influenza viruses of similar genetic background but differ in their pathogenicity in mallard ducks. CK10 is highly pathogenic whereas GS10 is low pathogenic. In this study, strong inflammatory response in terms of the expression level of several cytokines was observed in mallard duck peripheral blood mononuclear cells (PBMC) infected with CK10 while mild response was triggered in those by GS10 infection. Two remarkable and intense peaks of immune response were induced by CK10 infection within 24 hours (at 8 and 24 hours post infection, respectively) without reducing the virus replication. Our observations indicated that sustained and intense innate immune responses may be central to the high pathogenicity caused by CK10 in ducks.

  10. The multigenic nature of the differences in pathogenicity of H5N1 highly pathogenic avian influenza viruses in domestic ducks

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Eurasian H5N1 highly pathogenic avian influenza (HPAI) viruses have evolved into many genetic lineages. The divergent strains that have arisen express distinct pathobiological features and increased virulence for many bird species including domestic waterfowl. The pathogenicity of H5N1 HPAI vi...

  11. Highly Pathogenic Avian Influenza Virus Infection of Mallards with Homo- and Heterosubtypic Immunity Induced by Low Pathogenic Avian Influenza Viruses

    PubMed Central

    Fereidouni, Sasan R.; Starick, Elke; Beer, Martin; Wilking, Hendrik; Kalthoff, Donata; Grund, Christian; Häuslaigner, Rafaela; Breithaupt, Angele; Lange, Elke; Harder, Timm C.

    2009-01-01

    The potential role of wild birds as carriers of highly pathogenic avian influenza virus (HPAIV) subtype H5N1 is still a matter of debate. Consecutive or simultaneous infections with different subtypes of influenza viruses of low pathogenicity (LPAIV) are very common in wild duck populations. To better understand the epidemiology and pathogenesis of HPAIV H5N1 infections in natural ecosystems, we investigated the influence of prior infection of mallards with homo- (H5N2) and heterosubtypic (H4N6) LPAIV on exposure to HPAIV H5N1. In mallards with homosubtypic immunity induced by LPAIV infection, clinical disease was absent and shedding of HPAIV from respiratory and intestinal tracts was grossly reduced compared to the heterosubtypic and control groups (mean GEC/100 µl at 3 dpi: 3.0×102 vs. 2.3×104 vs. 8.7×104; p<0.05). Heterosubtypic immunity induced by an H4N6 infection mediated a similar but less pronounced effect. We conclude that the epidemiology of HPAIV H5N1 in mallards and probably other aquatic wild bird species is massively influenced by interfering immunity induced by prior homo- and heterosubtypic LPAIV infections. PMID:19693268

  12. Pathogenesis of highly-pathogenic Asian PRRSV in pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In 2006, Chinese investigators reported a unique syndrome in growing swine that was highlighted by clinical signs of high fever, anorexia, listlessness, red discoloration of skin, respiratory distress and very high morbidity and mortality rates. Originally known as porcine high fever disease (PHFD),...

  13. Highly pathogenic avian influenza viruses and generation of novel reassortants,United States, 2014–2015

    USGS Publications Warehouse

    Dong-Hun Lee,; Justin Bahl,; Mia Kim Torchetti,; Mary Lea Killian,; Ip, Hon S.; David E Swayne,

    2016-01-01

    Asian highly pathogenic avian influenza A(H5N8) viruses spread into North America in 2014 during autumn bird migration. Complete genome sequencing and phylogenetic analysis of 32 H5 viruses identified novel H5N1, H5N2, and H5N8 viruses that emerged in late 2014 through reassortment with North American low-pathogenicity avian influenza viruses.

  14. Highly Pathogenic Avian Influenza Viruses and Generation of Novel Reassortants, United States, 2014–2015

    PubMed Central

    Lee, Dong-Hun; Bahl, Justin; Torchetti, Mia Kim; Killian, Mary Lea; Ip, Hon S.; DeLiberto, Thomas J.

    2016-01-01

    Asian highly pathogenic avian influenza A(H5N8) viruses spread into North America in 2014 during autumn bird migration. Complete genome sequencing and phylogenetic analysis of 32 H5 viruses identified novel H5N1, H5N2, and H5N8 viruses that emerged in late 2014 through reassortment with North American low-pathogenicity avian influenza viruses. PMID:27314845

  15. Highly Pathogenic Avian Influenza Viruses and Generation of Novel Reassortants, United States, 2014-2015.

    PubMed

    Lee, Dong-Hun; Bahl, Justin; Torchetti, Mia Kim; Killian, Mary Lea; Ip, Hon S; DeLiberto, Thomas J; Swayne, David E

    2016-07-01

    Asian highly pathogenic avian influenza A(H5N8) viruses spread into North America in 2014 during autumn bird migration. Complete genome sequencing and phylogenetic analysis of 32 H5 viruses identified novel H5N1, H5N2, and H5N8 viruses that emerged in late 2014 through reassortment with North American low-pathogenicity avian influenza viruses. PMID:27314845

  16. Pathogenicity of Highly Pathogenic Avian Influenza Virus H5N1 in Naturally Infected Poultry in Egypt.

    PubMed

    Hagag, Ibrahim Thabet; Mansour, Shimaa M G; Zhang, Zerui; Ali, Ahmed A H; Ismaiel, El-Bakry M; Salama, Ali A; Cardona, Carol J; Collins, James; Xing, Zheng

    2015-01-01

    Highly pathogenic avian influenza virus (HPAIV) H5N1 has been endemic in Egypt since 2006, and there is increasing concern for its potential to become highly transmissible among humans. Infection by HPAIV H5N1 has been described in experimentally challenged birds. However, the pathogenicity of the H5N1 isolated in Egypt has never been reported in naturally infected chickens and ducks. Here we report a 2013 outbreak of HPAIV H5N1 in commercial poultry farms and backyards in Sharkia Province, Egypt. The main symptoms were ecchymosis on the shanks and feet, cyanosis of the comb and wattles, subcutaneous edema of the head and neck for chickens, and nervous signs (torticollis) for ducks. Within 48-72 hrs of the onset of illness, the average mortality rates were 22.8-30% and 28.5-40% in vaccinated chickens and non-vaccinated ducks, respectively. Tissue samples of chickens and ducks were collected for analyses with cross-section immunohistochemistry and real-time RT-PCR for specific viral RNA transcripts. While viral RNA was detected in nearly all tissues and sera collected, viral nucleoprotein was detected almost ubiquitously in all tissues, including testis. Interestingly, viral antigen was also observed in endothelial cells of most organs in chickens, and clearly detected in the trachea and brain in particular. Viral nucleoprotein was also detected in mononuclear cells of various organs, especially pulmonary tissue. We performed phylogenetic analyses and compared the genomic sequences of the hemagglutinin (HA) and nonstructural proteins (NS) among the isolated viruses, the HPAIV circulated in Egypt in the past and currently, and some available vaccine strains. Further analysis of deduced amino acids of both HA and NS1 revealed that our isolates carried molecular determinants of HPAIV, including the multibasic amino acids (PQGERRRK/KR*GLF) in the cleavage site in HA and glutamate at position 92 (D92E) in NS1. This is the first report of the pathogenicity of the HPAIVH5N

  17. Pathogenicity of Highly Pathogenic Avian Influenza Virus H5N1 in Naturally Infected Poultry in Egypt

    PubMed Central

    Hagag, Ibrahim Thabet; Mansour, Shimaa M. G.; Zhang, Zerui; Ali, Ahmed A. H.; Ismaiel, El-Bakry M.; Salama, Ali A.; Cardona, Carol J.; Collins, James; Xing, Zheng

    2015-01-01

    Highly pathogenic avian influenza virus (HPAIV) H5N1 has been endemic in Egypt since 2006, and there is increasing concern for its potential to become highly transmissible among humans. Infection by HPAIV H5N1 has been described in experimentally challenged birds. However, the pathogenicity of the H5N1 isolated in Egypt has never been reported in naturally infected chickens and ducks. Here we report a 2013 outbreak of HPAIV H5N1 in commercial poultry farms and backyards in Sharkia Province, Egypt. The main symptoms were ecchymosis on the shanks and feet, cyanosis of the comb and wattles, subcutaneous edema of the head and neck for chickens, and nervous signs (torticollis) for ducks. Within 48-72 hrs of the onset of illness, the average mortality rates were 22.8-30% and 28.5-40% in vaccinated chickens and non-vaccinated ducks, respectively. Tissue samples of chickens and ducks were collected for analyses with cross-section immunohistochemistry and real-time RT-PCR for specific viral RNA transcripts. While viral RNA was detected in nearly all tissues and sera collected, viral nucleoprotein was detected almost ubiquitously in all tissues, including testis. Interestingly, viral antigen was also observed in endothelial cells of most organs in chickens, and clearly detected in the trachea and brain in particular. Viral nucleoprotein was also detected in mononuclear cells of various organs, especially pulmonary tissue. We performed phylogenetic analyses and compared the genomic sequences of the hemagglutinin (HA) and nonstructural proteins (NS) among the isolated viruses, the HPAIV circulated in Egypt in the past and currently, and some available vaccine strains. Further analysis of deduced amino acids of both HA and NS1 revealed that our isolates carried molecular determinants of HPAIV, including the multibasic amino acids (PQGERRRK/KR*GLF) in the cleavage site in HA and glutamate at position 92 (D92E) in NS1. This is the first report of the pathogenicity of the HPAIVH5N

  18. High temperature and bacteriophages can indirectly select for bacterial pathogenicity in environmental reservoirs.

    PubMed

    Friman, Ville-Petri; Hiltunen, Teppo; Jalasvuori, Matti; Lindstedt, Carita; Laanto, Elina; Örmälä, Anni-Maria; Laakso, Jouni; Mappes, Johanna; Bamford, Jaana K H

    2011-03-15

    The coincidental evolution hypothesis predicts that traits connected to bacterial pathogenicity could be indirectly selected outside the host as a correlated response to abiotic environmental conditions or different biotic species interactions. To investigate this, an opportunistic bacterial pathogen, Serratia marcescens, was cultured in the absence and presence of the lytic bacteriophage PPV (Podoviridae) at 25°C and 37°C for four weeks (N = 5). At the end, we measured changes in bacterial phage-resistance and potential virulence traits, and determined the pathogenicity of all bacterial selection lines in the Parasemia plantaginis insect model in vivo. Selection at 37°C increased bacterial motility and pathogenicity but only in the absence of phages. Exposure to phages increased the phage-resistance of bacteria, and this was costly in terms of decreased maximum population size in the absence of phages. However, this small-magnitude growth cost was not greater with bacteria that had evolved in high temperature regime, and no trade-off was found between phage-resistance and growth rate. As a result, phages constrained the evolution of a temperature-mediated increase in bacterial pathogenicity presumably by preferably infecting the highly motile and virulent bacteria. In more general perspective, our results suggest that the traits connected to bacterial pathogenicity could be indirectly selected as a correlated response by abiotic and biotic factors in environmental reservoirs.

  19. Making pathogens sociable: The emergence of high relatedness through limited host invasibility

    PubMed Central

    van Leeuwen, Edwin; O'Neill, Sarah; Matthews, Andrew; Raymond, Ben

    2015-01-01

    Cooperation depends upon high relatedness, the high genetic similarity of interacting partners relative to the wider population. For pathogenic bacteria, which show diverse cooperative traits, the population processes that determine relatedness are poorly understood. Here, we explore whether within-host dynamics can produce high relatedness in the insect pathogen Bacillus thuringiensis. We study the effects of host/pathogen interactions on relatedness via a model of host invasion and fit parameters to competition experiments with marked strains. We show that invasibility is a key parameter for determining relatedness and experimentally demonstrate the emergence of high relatedness from well-mixed inocula. We find that a single infection cycle results in a bottleneck with a similar level of relatedness to those previously reported in the field. The bottlenecks that are a product of widespread barriers to infection can therefore produce the population structure required for the evolution of cooperative virulence. PMID:26125685

  20. Local poultry biosecurity risks to highly pathogenic avian influenza in Kaduna State, Nigeria.

    PubMed

    Paul, Abdu A; Assam, Assam; Ndang, Tabe-Ntui L

    2013-01-01

    The study appraised local poultry biosecurity risks to highly pathogenic avian influenza by assessing farmers' knowledge, beliefs and poultry practices using a standard questionnaire. Farmers' knowledge on transmission and prevention was high but low on disease recognition. Radio was ineffective at informing Islamic educated farmers. Extensive knowledge on transmission and protection did not result in behavioural change as farmers engaged in risky practices of selling, eating or medicating infected poultry and not reporting poultry death. Islamic educated farmers do not believe highly pathogenic avian influenza is a serious and preventable disease. Women are more likely to self medicate when experiencing influenza-like illness. Audio-visual aids would improve avian influenza recognition while involvement of community leaders would enhance disease reporting. Outbreak of highly pathogenic avian influenza in local poultry in Nigeria would follow a similar pattern in Southeast Asia if the risk perception among farmers is not urgently articulated.

  1. Broad-Range Survey of Tick-Borne Pathogens in Southern Germany Reveals a High Prevalence of Babesia microti and a Diversity of Other Tick-Borne Pathogens

    PubMed Central

    Crowder, Chris D.; Carolan, Heather E.; Rounds, Megan A.; Ecker, David J.; Haag, Heike; Mothes, Benedikt; Nolte, Oliver

    2014-01-01

    Abstract Ticks harbor numerous pathogens of significance to human and animal health. A better understanding of the pathogens carried by ticks in a given geographic area can alert health care providers of specific health risks leading to better diagnosis and treatments. In this study, we tested 226 Ixodes ricinis ticks from Southern Germany using a broad-range PCR and electrospray ionization mass spectrometry assay (PCR/ESI-MS) designed to identify tick-borne bacterial and protozoan pathogens in a single test. We found 21.2% of the ticks tested carried Borrelia burgdorferi sensu lato consisting of diverse genospecies; a surprisingly high percentage of ticks were infected with Babesia microti (3.5%). Other organisms found included Borrelia miyamotoi, Rickettsia helvetica, Rickettsia monacensis, and Anaplasma phagocytophilum. Of further significance was our finding that more than 7% of ticks were infected with more than one pathogen or putative pathogen. PMID:25072989

  2. Host immune responses of ducks infected with H5N1 highly pathogenic avian influenza viruses of different pathogenicities.

    PubMed

    Wei, Liangmeng; Jiao, Peirong; Song, Yafen; Cao, Lan; Yuan, Runyu; Gong, Lang; Cui, Jin; Zhang, Shuo; Qi, Wenbao; Yang, Su; Liao, Ming

    2013-10-25

    Our previous studies have illustrated three strains of duck-origin H5N1 highly pathogenic avian influenza viruses (HPAIVs) had varying levels of pathogenicity in ducks (Sun et al., 2011). However, the host immune response of ducks infected with those of H5N1 HPAIVs was unclear. Here, we compared viral distribution and mRNA expression of immune-related genes in ducks following infection with the two HPAIV (A/Duck/Guangdong/212/2004, DK212 and A/Duck/Guangdong/383/2008, DK383). DK383 could replicate in the tested tissue of ducks (brain, spleen, lungs, cloacal bursa, kidney, and pancreas) more rapid and efficiently than DK212 at 1 and 2 days post-inoculation. Quantitative real-time PCR analysis showed that the expression levels of TLR3, IL-6, IL-8, and MHC class II in brains were higher than those of respective genes in lungs during the early stage of post infection. Furthermore, the expression levels of IL-6 and IL-8 in the brain of ducks following infection with DK383 were remarkably higher than those of ducks infected with DK212, respectively. Our results suggest that the shift in the H5N1 HPAIVs to increased virulence in ducks may be associated with efficient and rapid replication of the virus, accompanied by early destruction of host immune responses. These data are helpful to understand the underlying mechanism of the different outcome of H5N1 HPAIVs infection in ducks.

  3. Bacillus cereus from the environment is genetically related to the highly pathogenic B. cereus in Zambia.

    PubMed

    Ogawa, Hirohito; Ohnuma, Miyuki; Squarre, David; Mweene, Aaron Simanyengwe; Ezaki, Takayuki; Fujikura, Daisuke; Ohnishi, Naomi; Thomas, Yuka; Hang'ombe, Bernard Mudenda; Higashi, Hideaki

    2015-08-01

    To follow-up anthrax in Zambia since the outbreak in 2011, we have collected samples from the environment and the carcasses of anthrax-suspected animals, and have tried to isolate Bacillus anthracis. In the process of identification of B. anthracis, we collected two isolates, of which colonies were similar to B. anthracis; however, from the results of identification using the molecular-based methods, two isolates were genetically related to the highly pathogenic B. cereus, of which clinical manifestation is severe and fatal (e.g., pneumonia). In this study, we showed the existence of bacteria suspected to be highly pathogenic B. cereus in Zambia, indicating the possibility of an outbreak caused by highly pathogenic B. cereus.

  4. High prevalence of the amphibian chytrid pathogen in Gabon.

    PubMed

    Bell, Rayna C; Gata Garcia, Adriana V; Stuart, Bryan L; Zamudio, Kelly R

    2011-03-01

    Amphibian chytridiomycosis is an infectious disease caused by the fungus Batrachochytrium dendrobatidis (Bd) that is implicated in the worldwide decline and extinction of amphibians. Africa has been proposed as a potential source for the global expansion of Bd, yet the distribution of Bd across the continent remains largely unexplored. Using quantitative polymerase chain reaction (qPCR), we screened for the presence of Bd in 166 adult anurans from two national parks in Gabon (Monts de Cristal and Ivindo). Bd was detected in 20 of the 42 species and was present at all three sites surveyed (two in Monts de Cristal, and one in Ivindo) with high prevalence (19.6%-36.0%). Both national parks were Bd-positive at all elevations and across habitat types, though no dead or dying frogs were encountered. To our knowledge, this study presents the first evidence of Bd in Gabon and the first record of infection for 19 of the 20 species that were Bd-positive. Documenting the distribution and virulence of Bd across Africa will be essential for understanding the dynamics of amphibian chytridiomycosis across the globe.

  5. False positives complicate ancient pathogen identifications using high-throughput shotgun sequencing

    PubMed Central

    2014-01-01

    Background Identification of historic pathogens is challenging since false positives and negatives are a serious risk. Environmental non-pathogenic contaminants are ubiquitous. Furthermore, public genetic databases contain limited information regarding these species. High-throughput sequencing may help reliably detect and identify historic pathogens. Results We shotgun-sequenced 8 16th-century Mixtec individuals from the site of Teposcolula Yucundaa (Oaxaca, Mexico) who are reported to have died from the huey cocoliztli (‘Great Pestilence’ in Nahautl), an unknown disease that decimated native Mexican populations during the Spanish colonial period, in order to identify the pathogen. Comparison of these sequences with those deriving from the surrounding soil and from 4 precontact individuals from the site found a wide variety of contaminant organisms that confounded analyses. Without the comparative sequence data from the precontact individuals and soil, false positives for Yersinia pestis and rickettsiosis could have been reported. Conclusions False positives and negatives remain problematic in ancient DNA analyses despite the application of high-throughput sequencing. Our results suggest that several studies claiming the discovery of ancient pathogens may need further verification. Additionally, true single molecule sequencing’s short read lengths, inability to sequence through DNA lesions, and limited ancient-DNA-specific technical development hinder its application to palaeopathology. PMID:24568097

  6. A Single Amino Acid in the M1 Protein Responsible for the Different Pathogenic Potentials of H5N1 Highly Pathogenic Avian Influenza Virus Strains

    PubMed Central

    Nao, Naganori; Kajihara, Masahiro; Manzoor, Rashid; Maruyama, Junki; Yoshida, Reiko; Muramatsu, Mieko; Miyamoto, Hiroko; Igarashi, Manabu; Eguchi, Nao; Sato, Masahiro; Kondoh, Tatsunari; Okamatsu, Masatoshi; Sakoda, Yoshihiro; Kida, Hiroshi; Takada, Ayato

    2015-01-01

    Two highly pathogenic avian influenza virus strains, A/duck/Hokkaido/WZ83/2010 (H5N1) (WZ83) and A/duck/Hokkaido/WZ101/2010 (H5N1) (WZ101), which were isolated from wild ducks in Japan, were found to be genetically similar, with only two amino acid differences in their M1 and PB1 proteins at positions 43 and 317, respectively. We found that both WZ83 and WZ101 caused lethal infection in chickens but WZ101 killed them more rapidly than WZ83. Interestingly, ducks experimentally infected with WZ83 showed no or only mild clinical symptoms, whereas WZ101 was highly lethal. We then generated reassortants between these viruses and found that exchange of the M gene segment completely switched the pathogenic phenotype in both chickens and ducks, indicating that the difference in the pathogenicity for these avian species between WZ83 and WZ101 was determined by only a single amino acid in the M1 protein. It was also found that WZ101 showed higher pathogenicity than WZ83 in mice and that WZ83, whose M gene was replaced with that of WZ101, showed higher pathogenicity than wild-type WZ83, although this reassortant virus was not fully pathogenic compared to wild-type WZ101. These results suggest that the amino acid at position 43 of the M1 protein is one of the factors contributing to the pathogenicity of H5N1 highly pathogenic avian influenza viruses in both avian and mammalian hosts. PMID:26368015

  7. Highly Pathogenic Avian Influenza Virus (H5N1) Isolated from Whooper Swans, Japan

    PubMed Central

    Uchida, Yuko; Mase, Masaji; Yoneda, Kumiko; Kimura, Atsumu; Obara, Tsuyoshi; Kumagai, Seikou; Yamamoto, Yu; Nakamura, Kikuyasu; Tsukamoto, Kenji; Yamaguchi, Shigeo

    2008-01-01

    On April 21, 2008, four whooper swans were found dead at Lake Towada, Akita prefecture, Japan. Highly pathogenic avian influenza virus of the H5N1 subtype was isolated from specimens of the affected birds. The hemagglutinin (HA) gene of the isolate belongs to clade 2.3.2 in the HA phylogenetic tree. PMID:18760011

  8. Inhibition of enteric pathogens using integrated high intensity 405 nm LED on the surface of almonds

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The disinfecting properties of 405 nm light were investigated against Escherichia coli O157:H7, Salmonella, and their non-pathogenic surrogates inoculated onto the surface of almonds. High intensity monochromatic light was generated from an array of narrow-band 405 nm light emitting diodes (LED). Al...

  9. Pathogenesis and transmission of highly pathogenic avian influenza H5Nx in swine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Introduction Influenza A viruses (IAV) periodically transmit between pigs, people, and birds. If two IAV strains infect the same host, genes can reassort to generate progeny virus with potential to be more infectious or avoid immunity. Pigs pose a risk for such reassortment. Highly pathogenic avian ...

  10. Human Infection with Highly Pathogenic A(H7N7) Avian Influenza Virus, Italy, 2013

    PubMed Central

    Rossini, Giada; Facchini, Marzia; Vaccari, Gabriele; Di Trani, Livia; Di Martino, Angela; Gaibani, Paolo; Vocale, Caterina; Cattoli, Giovanni; Bennett, Michael; McCauley, John W.; Rezza, Giovanni; Moro, Maria Luisa; Rangoni, Roberto; Finarelli, Alba Carola; Landini, Maria Paola; Castrucci, Maria Rita; Donatelli, Isabella

    2014-01-01

    During an influenza A(H7N7) virus outbreak among poultry in Italy during August–September 2013, infection with a highly pathogenic A(H7N7) avian influenza virus was diagnosed for 3 poultry workers with conjunctivitis. Genetic analyses revealed that the viruses from the humans were closely related to those from chickens on affected farms. PMID:25271444

  11. Highly pathogenic avian influenza (H5N1) outbreaks in wild birds and poultry, South Korea.

    PubMed

    Kim, Hye-Ryoung; Lee, Youn-Jeong; Park, Choi-Kyu; Oem, Jae-Ku; Lee, O-Soo; Kang, Hyun-Mi; Choi, Jun-Gu; Bae, You-Chan

    2012-03-01

    Highly pathogenic avian influenza (H5N1) among wild birds emerged simultaneously with outbreaks in domestic poultry in South Korea during November 2010-May 2011. Phylogenetic analysis showed that these viruses belonged to clade 2.3.2, as did viruses found in Mongolia, the People's Republic of China, and Russia in 2009 and 2010.

  12. Highly pathogenic avian influenza virus and generation of novel reassortants, United States, 2014-2015

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Asian highly pathogenic avian influenza A(H5N8) viruses spread into North America in 2014 during autumn bird migration. Complete genome sequencing and phylogenetic analysis of 32 H5 viruses identified novel H5N1, H5N2, and H5N8 viruses that emerged in late 2014 through reassortment with North Americ...

  13. Novel Eurasian highly pathogenic avian influenza A H5 viruses in wild birds, Washington, USA, 2014.

    PubMed

    Ip, Hon S; Torchetti, Mia Kim; Crespo, Rocio; Kohrs, Paul; DeBruyn, Paul; Mansfield, Kristin G; Baszler, Timothy; Badcoe, Lyndon; Bodenstein, Barbara; Shearn-Bochsler, Valerie; Killian, Mary Lea; Pedersen, Janice C; Hines, Nichole; Gidlewski, Thomas; DeLiberto, Thomas; Sleeman, Jonathan M

    2015-05-01

    Novel Eurasian lineage avian influenza A(H5N8) virus has spread rapidly and globally since January 2014. In December 2014, H5N8 and reassortant H5N2 viruses were detected in wild birds in Washington, USA, and subsequently in backyard birds. When they infect commercial poultry, these highly pathogenic viruses pose substantial trade issues.

  14. Novel Eurasian highly pathogenic influenza A H5 viruses in wild birds, Washington, USA

    USGS Publications Warehouse

    Ip, Hon S.; Torchetti, Mia Kim; Crespo, Rocio; Kohrs, Paul; DeBruyn, Paul; Mansfield, Kristin G.; Baszler, Timothy; Badcoe, Lyndon; Bodenstein, Barbara L.; Shearn-Bochsler, Valerie I.; Killian, Mary Lea; Pederson, Janice C.; Hines, Nichole; Gidlewski, Thomas; DeLiberto, Thomas; Sleeman, Jonathan M.

    2015-01-01

    Novel Eurasian lineage avian influenza A(H5N8) virus has spread rapidly and globally since January 2014. In December 2014, H5N8 and reassortant H5N2 viruses were detected in wild birds in Washington, USA, and subsequently in backyard birds. When they infect commercial poultry, these highly pathogenic viruses pose substantial trade issues.

  15. Two Asian highly pathogenic strains of Type 2 PRRSV in United States swine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Highly pathogenic PRRSV (HP-PRRSV) has been circulating in Asia for 7 years. rJXwn06 and rSRV07 were rescued from infectious clones of two HP-PRRSV for investigation at the National Animal Disease Center. The clinical disease and viral replication kinetics of HP-PRRSV were compared to prototype stra...

  16. Global expansion of high pathogenicity avian influenza: implications on prevention and control programs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The H5N1 high pathogenicity avian influenza (HPAI) virus emerged in China during 1996 and has spread to infect poultry and/or wild birds in 63 countries during the past 18 years. The majority of the recent outbreaks of H5N1 HPAI have occurred in Indonesia, Egypt, Vietnam, and Bangladesh, in decreas...

  17. Global expansion of high pathogenicity avian influenza: implications on prevention and control programs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The H5N1 high pathogenicity avian influenza (HPAI) virus emerged in China during 1996 and has spread to infect poultry and/or wild birds in 63 countries during the past 18 years. The majority of the recent outbreaks of H5N1 HPAI have occurred in Indonesia, Egypt, Vietnam, and Bangladesh, in decreasi...

  18. Airborne transmission of H5N1 high pathogenicity avian influenza viruses during simulated home slaughter

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Most H5N1 human infections have occurred following exposure to H5N1 high pathogenicity avian influenza (HPAI) virus-infected poultry, especially when poultry are home slaughtered or slaughtered in live poultry markets. Previous studies have demonstrated that slaughter of clade 1 isolate A/Vietnam/1...

  19. Novel Eurasian Highly Pathogenic Avian Influenza A H5 Viruses in Wild Birds, Washington, USA, 2014

    PubMed Central

    Ip, Hon S.; Crespo, Rocio; Kohrs, Paul; DeBruyn, Paul; Mansfield, Kristin G.; Baszler, Timothy; Badcoe, Lyndon; Bodenstein, Barbara; Shearn-Bochsler, Valerie; Killian, Mary Lea; Pedersen, Janice C.; Hines, Nichole; Gidlewski, Thomas; DeLiberto, Thomas; Sleeman, Jonathan M.

    2015-01-01

    Novel Eurasian lineage avian influenza A(H5N8) virus has spread rapidly and globally since January 2014. In December 2014, H5N8 and reassortant H5N2 viruses were detected in wild birds in Washington, USA, and subsequently in backyard birds. When they infect commercial poultry, these highly pathogenic viruses pose substantial trade issues. PMID:25898265

  20. Chlorine inactivation of H5N1 highly pathogenic avian influenza virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Two Asian strains of H5N1 highly pathogenic avian influenza virus were studied to determine their resistance to chlorination. Experiments were conducted at two pH levels (pH 7 and 8) at 5 C. CT (chlorine concentration x exposure time) values were calculated for different levels of inactivation. R...

  1. Novel reassortant highly pathogenic avian influenza (H5N5) viruses in domestic ducks, China.

    PubMed

    Gu, Min; Liu, Wenbo; Cao, Yongzhong; Peng, Daxin; Wang, Xiaobo; Wan, Hongquan; Zhao, Guo; Xu, Quangang; Zhang, Wei; Song, Qingqing; Li, Yanfang; Liu, Xiufan

    2011-06-01

    In China, domestic ducks and wild birds often share the same water, in which influenza viruses replicate preferentially. Isolation of 2 novel reassortant highly pathogenic avian influenza (H5N5) viruses from apparently healthy domestic ducks highlights the role of these ducks as reassortment vessels. Such new subtypes of influenza viruses may pose a pandemic threat.

  2. Photoluminescent lateral-flow immunoassay revealed by graphene oxide: highly sensitive paper-based pathogen detection.

    PubMed

    Morales-Narváez, Eden; Naghdi, Tina; Zor, Erhan; Merkoçi, Arben

    2015-08-18

    A paper-based lateral flow immunoassay for pathogen detection that avoids the use of secondary antibodies and is revealed by the photoluminescence quenching ability of graphene oxide is reported. Escherichia coli has been selected as a model pathogen. The proposed device is able to display a highly specific and sensitive performance with a limit of detection of 10 CFU mL(-1) in standard buffer and 100 CFU mL(-1) in bottled water and milk. This low-cost disposable and easy-to-use device will prove valuable for portable and automated diagnostics applications. PMID:26205473

  3. Genomic reconnaissance of clinical isolates of emerging human pathogen Mycobacterium abscessus reveals high evolutionary potential.

    PubMed

    Choo, Siew Woh; Wee, Wei Yee; Ngeow, Yun Fong; Mitchell, Wayne; Tan, Joon Liang; Wong, Guat Jah; Zhao, Yongbing; Xiao, Jingfa

    2014-02-11

    Mycobacterium abscessus (Ma) is an emerging human pathogen that causes both soft tissue infections and systemic disease. We present the first comparative whole-genome study of Ma strains isolated from patients of wide geographical origin. We found a high proportion of accessory strain-specific genes indicating an open, non-conservative pan-genome structure, and clear evidence of rapid phage-mediated evolution. Although we found fewer virulence factors in Ma compared to M. tuberculosis, our data indicated that Ma evolves rapidly and therefore should be monitored closely for the acquisition of more pathogenic traits. This comparative study provides a better understanding of Ma and forms the basis for future functional work on this important pathogen.

  4. High-Throughput Biosensors for Multiplexed Food-Borne Pathogen Detection

    NASA Astrophysics Data System (ADS)

    Gehring, Andrew G.; Tu, Shu-I.

    2011-07-01

    Incidental contamination of foods by pathogenic bacteria and/or their toxins is a serious threat to public health and the global economy. The presence of food-borne pathogens and toxins must be rapidly determined at various stages of food production, processing, and distribution. Producers, processors, regulators, retailers, and public health professionals need simple and cost-effective methods to detect different species or serotypes of bacteria and associated toxins in large numbers of food samples. This review addresses the desire to replace traditional microbiological plate culture with more timely and less cumbersome rapid, biosensor-based methods. Emphasis focuses on high-throughput, multiplexed techniques that allow for simultaneous testing of numerous samples, in rapid succession, for multiple food-borne analytes (primarily pathogenic bacteria and/or toxins).

  5. Dissemination of a highly virulent pathogen: tracking the early events that define infection.

    PubMed

    Gonzalez, Rodrigo J; Lane, M Chelsea; Wagner, Nikki J; Weening, Eric H; Miller, Virginia L

    2015-01-01

    The series of events that occurs immediately after pathogen entrance into the body is largely speculative. Key aspects of these events are pathogen dissemination and pathogen interactions with the immune response as the invader moves into deeper tissues. We sought to define major events that occur early during infection of a highly virulent pathogen. To this end, we tracked early dissemination of Yersinia pestis, a highly pathogenic bacterium that causes bubonic plague in mammals. Specifically, we addressed two fundamental questions: (1) do the bacteria encounter barriers in disseminating to draining lymph nodes (LN), and (2) what mechanism does this nonmotile bacterium use to reach the LN compartment, as the prevailing model predicts trafficking in association with host cells. Infection was followed through microscopy imaging in addition to assessing bacterial population dynamics during dissemination from the skin. We found and characterized an unexpected bottleneck that severely restricts bacterial dissemination to LNs. The bacteria that do not pass through this bottleneck are confined to the skin, where large numbers of neutrophils arrive and efficiently control bacterial proliferation. Notably, bottleneck formation is route dependent, as it is abrogated after subcutaneous inoculation. Using a combination of approaches, including microscopy imaging, we tested the prevailing model of bacterial dissemination from the skin into LNs and found no evidence of involvement of migrating phagocytes in dissemination. Thus, early stages of infection are defined by a bottleneck that restricts bacterial dissemination and by neutrophil-dependent control of bacterial proliferation in the skin. Furthermore, and as opposed to current models, our data indicate an intracellular stage is not required by Y. pestis to disseminate from the skin to draining LNs. Because our findings address events that occur during early encounters of pathogen with the immune response, this work can

  6. In Vitro Emergence of High Persistence upon Periodic Aminoglycoside Challenge in the ESKAPE Pathogens.

    PubMed

    Michiels, Joran Elie; Van den Bergh, Bram; Verstraeten, Natalie; Fauvart, Maarten; Michiels, Jan

    2016-08-01

    Health care-associated infections present a major threat to modern medical care. Six worrisome nosocomial pathogens-Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.-are collectively referred to as the "ESKAPE bugs." They are notorious for extensive multidrug resistance, yet persistence, or the phenotypic tolerance displayed by a variant subpopulation, remains underappreciated in these pathogens. Importantly, persistence can prevent eradication of antibiotic-sensitive bacterial populations and is thought to act as a catalyst for the development of genetic resistance. Concentration- and time-dependent aminoglycoside killing experiments were used to investigate persistence in the ESKAPE pathogens. Additionally, a recently developed method for the experimental evolution of persistence was employed to investigate adaptation to high-dose, extended-interval aminoglycoside therapy in vitro We show that ESKAPE pathogens exhibit biphasic killing kinetics, indicative of persister formation. In vitro cycling between aminoglycoside killing and persister cell regrowth, evocative of clinical high-dose extended-interval therapy, caused a 37- to 213-fold increase in persistence without the emergence of resistance. Increased persistence also manifested in biofilms and provided cross-tolerance to different clinically important antibiotics. Together, our results highlight a possible drawback of intermittent, high-dose antibiotic therapy and suggest that clinical diagnostics might benefit from taking into account persistence. PMID:27185802

  7. Profile and Fate of Bacterial Pathogens in Sewage Treatment Plants Revealed by High-Throughput Metagenomic Approach.

    PubMed

    Li, Bing; Ju, Feng; Cai, Lin; Zhang, Tong

    2015-09-01

    The broad-spectrum profile of bacterial pathogens and their fate in sewage treatment plants (STPs) were investigated using high-throughput sequencing based metagenomic approach. This novel approach could provide a united platform to standardize bacterial pathogen detection and realize direct comparison among different samples. Totally, 113 bacterial pathogen species were detected in eight samples including influent, effluent, activated sludge (AS), biofilm, and anaerobic digestion sludge with the abundances ranging from 0.000095% to 4.89%. Among these 113 bacterial pathogens, 79 species were reported in STPs for the first time. Specially, compared to AS in bulk mixed liquor, more pathogen species and higher total abundance were detected in upper foaming layer of AS. This suggests that the foaming layer of AS might impose more threat to onsite workers and citizens in the surrounding areas of STPs because pathogens in foaming layer are easily transferred into air and cause possible infections. The high removal efficiency (98.0%) of total bacterial pathogens suggests that AS treatment process is effective to remove most bacterial pathogens. Remarkable similarities of bacterial pathogen compositions between influent and human gut indicated that bacterial pathogen profiles in influents could well reflect the average bacterial pathogen communities of urban resident guts within the STP catchment area.

  8. Multiple introductions of highly pathogenic avian influenza H5N1 viruses into Bangladesh

    PubMed Central

    Marinova-Petkova, Atanaska; Feeroz, Mohammed M; Rabiul Alam, SM; Kamrul Hasan, M; Akhtar, Sharmin; Jones-Engel, Lisa; Walker, David; McClenaghan, Laura; Rubrum, Adam; Franks, John; Seiler, Patrick; Jeevan, Trushar; McKenzie, Pamela; Krauss, Scott; Webby, Richard J; Webster, Robert G

    2014-01-01

    Highly pathogenic H5N1 and low pathogenic H9N2 influenza viruses are endemic to poultry markets in Bangladesh and have cocirculated since 2008. H9N2 influenza viruses circulated constantly in the poultry markets, whereas highly pathogenic H5N1 viruses occurred sporadically, with peaks of activity in cooler months. Thirty highly pathogenic H5N1 influenza viruses isolated from poultry were characterized by antigenic, molecular, and phylogenetic analyses. Highly pathogenic H5N1 influenza viruses from clades 2.2.2 and 2.3.2.1 were isolated from live bird markets only. Phylogenetic analysis of the 30 H5N1 isolates revealed multiple introductions of H5N1 influenza viruses in Bangladesh. There was no reassortment between the local H9N2 influenza viruses and H5N1 genotype, despite their prolonged cocirculation. However, we detected two reassortant H5N1 viruses, carrying the M gene from the Chinese H9N2 lineage, which briefly circulated in the Bangladesh poultry markets and then disappeared. On the other hand, interclade reassortment occurred within H5N1 lineages and played a role in the genesis of the currently dominant H5N1 viruses in Bangladesh. Few ‘human-like' mutations in H5N1 may account for the limited number of human cases. Antigenically, clade 2.3.2.1 H5N1 viruses in Bangladesh have evolved since their introduction and are currently mainly homogenous, and show evidence of recent antigenic drift. Although reassortants containing H9N2 genes were detected in live poultry markets in Bangladesh, these reassortants failed to supplant the dominant H5N1 lineage. PMID:26038508

  9. The irp2 and fyuA genes in High Pathogenicity Islands are involved in the pathogenesis of infections caused by avian pathogenic Escherichia coli (APEC).

    PubMed

    Tu, Jian; Xue, Ting; Qi, Kezong; Shao, Ying; Huang, Boyan; Wang, Xueyan; Zhou, Xiuhong

    2016-01-01

    Avian pathogenic Escherichia coli (APEC) is a major bacterial infectious disease that may lead to local or systemic infections in chickens with clinical manifestations. The irp2-fyuA gene cluster has been confirmed to be the main genes involved in the synthesis of HPI. The objective of this study was to determine the influence of the irp2 and fyuA genes in the high pathogenicity island (HPI) of avian pathogenic Escherichia coli (APEC) on its pathogenicity by knocking out these genes. The ΔAE17 (lacking irp2) and ΔΔAE17 (lacking irp2 and fyuA) strains of APEC were constructed. The ΔAE17 and ΔΔAE17 strains showed significantly impaired capacity to adhere onto DF-1 cells. The LD50 results indicated that the virulence of the ΔAE17 and ΔΔAE17 strains was decreased in comparison with that of the AE17 strain. We concluded that the knock-out of the core HPI genes weakened APEC adhesion onto DF-1 cells, inhibited transcription of virulence genes, and reduced pathogenicity in chicks. The effects of genetic deletion of irp2 and fyuA on APEC were more severe than those produced by deletion of irp2 only, indicating that irp2 and fyuA co-regulate APEC pathogenicity.

  10. Draft Genome Sequence of a Highly Virulent Strain of the Plant Pathogen Dickeya solani, IFB0099

    PubMed Central

    Golanowska, M.; Galardini, M.; Bazzicalupo, M.; Hugouvieux-Cotte-Pattat, N.; Mengoni, A.; Potrykus, M.; Slawiak, M.

    2015-01-01

    Dickeya solani is an important bacterial pathogen of potato cultivars in Europe. Here, we present the draft genome of D. solani strain IFB0099 isolated from potato in Poland that shows a high level of pectinolytic activity and a high virulence. This genome sequence is 5,094,121 bp and contains 4,365 protein-coding sequences. PMID:25792047

  11. Development of High Hydrostatic Pressure Applied in Pathogen Inactivation for Plasma.

    PubMed

    Yang, Chunhui; Bian, Guohui; Yang, Hong; Zhang, Xinmin; Chen, Limin; Wang, Jingxing

    2016-01-01

    High hydrostatic pressure has been used to inactivate pathogens in foods for decades. There is a great potential to adapt this technology to inactivate pathogens in plasma and derivatives. To better evaluate the potential of this method, pathogen inoculated plasma samples were pressurized under different pressure application modes and temperatures. The inactivation efficacy of pathogens and activities of plasma proteins were monitored after treatment. The CFUs of E.coli was examined as the indicator of the inactivation efficiency. The factor V and VIII were chosen as the indicator of the plasma function. Preliminary experiments identified optimized treatment conditions: 200-250MPa, with 5×1 minute multi-pulsed high pressure at near 0°C (ice-water bath). Under this conditions, the inactivation efficacy of EMCV was >8.5log. The CFUs of E. coli were reduced by 7.5log, B. cereus were 8log. However, PPV and S. aureus cannot be inactivated efficiently. The activities of factor II, VII, IX, X, XI, XII, fibrinogen, IgG, IgM stayed over 95% compared to untreated. Factor V and VIII activity was maintained at 46-63% and 77-82%, respectively. PMID:27561010

  12. Development of High Hydrostatic Pressure Applied in Pathogen Inactivation for Plasma

    PubMed Central

    Yang, Hong; Zhang, Xinmin; Chen, Limin; Wang, Jingxing

    2016-01-01

    High hydrostatic pressure has been used to inactivate pathogens in foods for decades. There is a great potential to adapt this technology to inactivate pathogens in plasma and derivatives. To better evaluate the potential of this method, pathogen inoculated plasma samples were pressurized under different pressure application modes and temperatures. The inactivation efficacy of pathogens and activities of plasma proteins were monitored after treatment. The CFUs of E.coli was examined as the indicator of the inactivation efficiency. The factor V and VIII were chosen as the indicator of the plasma function. Preliminary experiments identified optimized treatment conditions: 200-250MPa, with 5×1 minute multi-pulsed high pressure at near 0°C (ice-water bath). Under this conditions, the inactivation efficacy of EMCV was >8.5log. The CFUs of E. coli were reduced by 7.5log, B. cereus were 8log. However, PPV and S. aureus cannot be inactivated efficiently. The activities of factor II, VII, IX, X, XI, XII, fibrinogen, IgG, IgM stayed over 95% compared to untreated. Factor V and VIII activity was maintained at 46–63% and 77–82%, respectively. PMID:27561010

  13. Resistance of highly pathogenic Naegleria fowleri amoebae to complement-mediated lysis.

    PubMed

    Whiteman, L Y; Marciano-Cabral, F

    1989-12-01

    Weakly pathogenic and nonpathogenic Naegleria spp. are readily lysed by human and guinea pig complement. Highly pathogenic Naegleria fowleri are resistant to complement-mediated lysis. Electrophoretic analysis of normal human serum (NHS) incubated with pathogenic or nonpathogenic Naegleria spp. demonstrates that amoebae activate the complement cascade, resulting in the production of C3 and C5 complement cleavage products. To determine whether surface constituents play a role in resistance to complement lysis, trophozoites of Naegleria spp. were subjected to enzymatic treatments prior to incubation in NHS. Treatment of trophozoites with papain or trypsin for 1 h, but not with neuraminidase, increased susceptibility of highly pathogenic Naegleria fowleri to complement lysis. Treatment of trophozoites with actinomycin D or cycloheximide during incubation with NHS or pretreatment with various protease inhibitors for 4 h did not increase the susceptibility of N. fowleri amoebae to lysis. Neither a repair process involving de novo protein synthesis nor a complement-inactivating protease appears to account for the increased resistance of N. fowleri amoebae to complement-mediated lysis.

  14. Chinese and Vietnamese strains of HP-PRRSV cause different pathogenic outcomes in United States high health swine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An infectious clone of a highly pathogenic PRRSV strain from Vietnam (rSRV07) was prepared, analyzed and compared to Chinese highly pathogenic PRRSV rJXwn06 and US Type 2 prototype VR-2332 in order to examine the effects of virus phenotype and genotype on growth in MARC-145 cells, as well as the imp...

  15. Widespread detection of highly pathogenic H5 influenza viruses in wild birds from the Pacific Flyway of the United States

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A novel highly pathogenic avian influenza virus belonging to the H5 clade 2.3.4.4 variant viruses was detected in North America in late 2014. Motivated by the identification of these viruses in domestic poultry in Canada, an intensive study was initiated to conduct highly pathogenic avian influenza ...

  16. Multiplex cytokine profiling with highly pathogenic material: use of formalin solution in luminex analysis.

    PubMed

    Dowall, Stuart D; Graham, Victoria A; Tipton, Thomas R W; Hewson, Roger

    2009-08-31

    Work with highly pathogenic material mandates the use of biological containment facilities, involving microbiological safety cabinets and specialist laboratory engineering structures typified by containment level 3 (CL3) and CL4 laboratories. Consequences of working in high containment are the practical difficulties associated with containing specialist assays and equipment often essential for experimental analyses. In an era of increased interest in biodefence pathogens and emerging diseases, immunological analysis has developed rapidly alongside traditional techniques in virology and molecular biology. For example, in order to maximise the use of small sample volumes, multiplexing has become a more popular and widespread approach to quantify multiple analytes simultaneously, such as cytokines and chemokines. The luminex microsphere system allows for the detection of many cytokines and chemokines in a single sample, but the detection method of using aligned lasers and fluidics means that samples often have to be analysed in low containment facilities. In order to perform cytokine analysis in materials from high containment (CL3 and CL4 laboratories), we have developed an appropriate inactivation methodology after staining steps, which although results in a reduction of median fluorescent intensity, produces statistically comparable outcomes when judged against non-inactivated samples. This methodology thus extends the use of luminex technology for material that contains highly pathogenic biological agents.

  17. 2.1 Natural History of Highly Pathogenic Avian Influenza H5N1

    PubMed Central

    Sonnberg, Stephanie; Webby, Richard J.; Webster, Robert G.

    2013-01-01

    The ecology of highly pathogenic avian influenza (HPAI) H5N1 has significantly changed from sporadic outbreaks in terrestrial poultry to persistent circulation in terrestrial and aquatic poultry and potentially in wild waterfowl. A novel genotype of HPAI H5N1 arose in 1996 in southern China and through ongoing mutation, reassortment, and natural selection, has diverged into distinct lineages and expanded into multiple reservoir hosts. The evolution of Goose/Guangdong-lineage highly pathogenic H5N1 viruses is ongoing: while stable interactions exist with some reservoir hosts, these viruses are continuing to evolve and adapt to others, and pose an un-calculable risk to sporadic hosts, including humans. PMID:23735535

  18. Wild bird surveillance for highly pathogenic avian influenza H5 in North America

    USGS Publications Warehouse

    Flint, Paul L.; Pearce, John M.; Franson, J. Christian; Derksen, Dirk V.

    2015-01-01

    It is unknown how the current Asian origin highly pathogenic avian influenza H5 viruses arrived, but these viruses are now poised to become endemic in North America. Wild birds harbor these viruses and have dispersed them at regional scales. What is unclear is how the viruses may be moving from the wild bird reservoir into poultry holdings. Active surveillance of live wild birds is likely the best way to determine the true distribution of these viruses. We also suggest that sampling be focused on regions with the greatest risk for poultry losses and attempt to define the mechanisms of transfer to enhance biosecurity. Responding to the recent outbreaks of highly pathogenic avian influenza in North America requires an efficient plan with clear objectives and potential management outcomes.

  19. Potential strategies and biosafety protocols used for dual-use research on highly pathogenic influenza viruses

    PubMed Central

    Du, Lanying; Li, Ye; Gao, Jimin; Zhou, Yusen; Jiang, Shibo

    2013-01-01

    Summary Influenza A viruses (IAVs), particularly the highly pathogenic avian influenza (HPAI) H5N1, have posed a substantial threat to public health worldwide. Although the laboratory generation of the mutant influenza virus H5N1 with airborne transmissibility among mammals, which has been considered as a dual-use research, may benefit the development of effective vaccines and therapeutics against the emerging infectious agents, it may also pose threats to national biosecurity, laboratory biosafety, and/or public health. This review introduces the classification and characterization of IAVs, pinpoints historic pandemics and epidemics caused by IAVs, emphasizes the significance and necessity of biosafety, summarizes currently established biosafety-related protocols for IAV research, and provides potential strategies to improve biosafety protocols for dual-use research on the highly pathogenic avian influenza viruses and other emerging infectious agents. PMID:22987727

  20. Research efforts to control highly pathogenic arenaviruses: a summary of the progress and gaps.

    PubMed

    Kerber, R; Reindl, S; Romanowski, V; Gómez, R M; Ogbaini-Emovon, E; Günther, S; ter Meulen, J

    2015-03-01

    Significant progress has been made in the past 10 years in unraveling the molecular biology of highly pathogenic arenaviruses that are endemic in several West African countries (Lassa fever virus) and in some regions of South America (Argentine and Bolivian hemorrhagic fever viruses). While this has resulted in proof-of-concept studies of novel vaccine candidates in non-human primates and in the discovery of several novel antiviral small molecule drug candidates, none of them has been tested in the clinic to date. The recent Ebola outbreak in West Africa has demonstrated very clearly that there is an urgent need to develop the prophylactic and therapeutic armamentarium against viral hemorrhagic fever viruses as part of a global preparedness for future epidemics. As it pertains to this goal, the present article summarizes the current knowledge of highly pathogenic arenaviruses and identifies opportunities for translational research. PMID:25549822

  1. Wild bird surveillance for highly pathogenic avian influenza H5 in North America.

    PubMed

    Flint, Paul L; Pearce, John M; Franson, J Christian; Derksen, Dirk V

    2015-09-28

    It is unknown how the current Asian origin highly pathogenic avian influenza H5 viruses arrived, but these viruses are now poised to become endemic in North America. Wild birds harbor these viruses and have dispersed them at regional scales. What is unclear is how the viruses may be moving from the wild bird reservoir into poultry holdings. Active surveillance of live wild birds is likely the best way to determine the true distribution of these viruses. We also suggest that sampling be focused on regions with the greatest risk for poultry losses and attempt to define the mechanisms of transfer to enhance biosecurity. Responding to the recent outbreaks of highly pathogenic avian influenza in North America requires an efficient plan with clear objectives and potential management outcomes.

  2. Genetic characterization of highly pathogenic H5 influenza viruses from poultry in Taiwan, 2015.

    PubMed

    Huang, Pei-Yu; Lee, Chang-Chun David; Yip, Chun-Hung; Cheung, Chung-Lam; Yu, Guangchuang; Lam, Tommy Tsan-Yuk; Smith, David K; Zhu, Huachen; Guan, Yi

    2016-03-01

    Phylogenetic analysis of the highly pathogenic avian influenza (HPAI) H5 viruses causing recent outbreaks in Taiwan showed that they belonged to the Asian HPAI H5 lineage, clade 2.3.4.4 viruses, and were apparently introduced by migratory birds. These viruses reassorted with Eurasian influenza gene pool viruses and formed five genotypic variants. As Taiwan has a similar influenza ecosystem to southern China, the HPAI H5 lineage could become established and enzootic in the island.

  3. Effect of species, breed and route of virus inoculation on the pathogenicity of H5N1 highly pathogenic influenza (HPAI) viruses in domestic ducks

    Technology Transfer Automated Retrieval System (TEKTRAN)

    H5N1 highly pathogenic avian influenza (HPAI) viruses continue to be a threat to poultry in many regions of the world. Domestic ducks have been recognized as one of the primary factors in the spread of H5N1 HPAI. To improve the control of this disease it’s necessary to better understand the pathog...

  4. Growth and Pathogenic Potential of Naturally Selected Reassortants after Coinfection with Pandemic H1N1 and Highly Pathogenic Avian Influenza H5N1 Viruses

    PubMed Central

    Song, Min-Suk; Baek, Yun Hee; Pascua, Philippe Noriel Q.; Kwon, Hyeok-il; Kim, Eun-Ha; Park, Su-Jin; Kim, Se Mi; Kim, Young-Il; Choi, Won-Suk; Kim, Eung-Gook; Kim, Chul-Joong

    2015-01-01

    Coinfection of ferrets with H5N1 and pH1N1 viruses resulted in two predominate genotypes in the lungs containing surface genes of highly pathogenic avian influenza H5N1 virus in the backbone of pandemic H1N1 2009 (pH1N1). Compared to parental strains, these reassortants exhibited increased growth and virulence in vitro and in mice but failed to be transmitted indirectly to naive contact ferrets. Thus, this demonstrates a possible natural reassortment following coinfection as well as the pathogenicity of the potential reassortants. PMID:26491154

  5. Growth and Pathogenic Potential of Naturally Selected Reassortants after Coinfection with Pandemic H1N1 and Highly Pathogenic Avian Influenza H5N1 Viruses.

    PubMed

    Song, Min-Suk; Baek, Yun Hee; Pascua, Philippe Noriel Q; Kwon, Hyeok-Il; Kim, Eun-Ha; Park, Su-Jin; Kim, Se Mi; Kim, Young-Il; Choi, Won-Suk; Kim, Eung-Gook; Kim, Chul-Joong; Choi, Young Ki

    2016-01-01

    Coinfection of ferrets with H5N1 and pH1N1 viruses resulted in two predominate genotypes in the lungs containing surface genes of highly pathogenic avian influenza H5N1 virus in the backbone of pandemic H1N1 2009 (pH1N1). Compared to parental strains, these reassortants exhibited increased growth and virulence in vitro and in mice but failed to be transmitted indirectly to naive contact ferrets. Thus, this demonstrates a possible natural reassortment following coinfection as well as the pathogenicity of the potential reassortants. PMID:26491154

  6. Within-Host Models of High and Low Pathogenic Influenza Virus Infections: The Role of Macrophages

    PubMed Central

    Pawelek, Kasia A.; Dor, Daniel; Salmeron, Cristian; Handel, Andreas

    2016-01-01

    The World Health Organization identifies influenza as a major public health problem. While the strains commonly circulating in humans usually do not cause severe pathogenicity in healthy adults, some strains that have infected humans, such as H5N1, can cause high morbidity and mortality. Based on the severity of the disease, influenza viruses are sometimes categorized as either being highly pathogenic (HP) or having low pathogenicity (LP). The reasons why some strains are LP and others HP are not fully understood. While there are likely multiple mechanisms of interaction between the virus and the immune response that determine LP versus HP outcomes, we focus here on one component, namely macrophages (MP). There is some evidence that MP may both help fight the infection and become productively infected with HP influenza viruses. We developed mathematical models for influenza infections which explicitly included the dynamics and action of MP. We fit these models to viral load and macrophage count data from experimental infections of mice with LP and HP strains. Our results suggest that MP may not only help fight an influenza infection but may contribute to virus production in infections with HP viruses. We also explored the impact of combination therapies with antivirals and anti-inflammatory drugs on HP infections. Our study suggests a possible mechanism of MP in determining HP versus LP outcomes, and how different interventions might affect infection dynamics. PMID:26918620

  7. High-Throughput Screening of a Diversity Collection Using Biodefense Category A and B Priority Pathogens

    PubMed Central

    Barrow, Esther W.; Clinkenbeard, Patricia A.; Duncan-Decocq, Rebecca A.; Perteet, Rachel F.; Hill, Kimberly D.; Bourne, Philip C.; Valderas, Michelle W.; Bourne, Christina R.; Clarkson, Nicole L.; Clinkenbeard, Kenneth D.; Barrow, William W.

    2013-01-01

    One of the objectives of the National Institutes of Allergy and Infectious Diseases (NIAID) Biodefense Program is to identify or develop broad-spectrum antimicrobials for use against bioterrorism pathogens and emerging infectious agents. As a part of that program, our institution has screened the 10 000-compound MyriaScreen Diversity Collection of high-purity druglike compounds against three NIAID category A and one category B priority pathogens in an effort to identify potential compound classes for further drug development. The effective use of a Clinical and Laboratory Standards Institute–based high-throughput screening (HTS) 96-well–based format allowed for the identification of 49 compounds that had in vitro activity against all four pathogens with minimum inhibitory concentration values of ≤16 μg/mL. Adaptation of the HTS process was necessary to conduct the work in higher-level containment, in this case, biosafety level 3. Examination of chemical scaffolds shared by some of the 49 compounds and assessment of available chemical databases indicates that several may represent broad-spectrum antimicrobials whose activity is based on novel mechanisms of action. PMID:22653912

  8. Surfaces presenting α-phenyl mannoside derivatives enable formation of stable, high coverage, non-pathogenic Escherichia coli biofilms against pathogen colonization.

    PubMed

    Zhu, Zhiling; Wang, Jun; Lopez, Analette I; Yu, Fei; Huang, Yongkai; Kumar, Amit; Li, Siheng; Zhang, Lijuan; Cai, Chengzhi

    2015-06-01

    Prevention of pathogenic colonization on medical devices over a long period of time remains a great challenge, especially in a high-nutrient environment that accelerates the production of biomass leading to biofouling of the device. Since biofouling and the subsequent pathogen colonization is eventually inevitable, a new strategy using non-pathogenic bacteria as living guards against pathogenic colonization on medical devices has attracted increasing interest. Crucial to the success of this strategy is to pre-establish a high coverage and stable biofilm of benign bacteria on the surface. Silicone elastomers are one of the most widely used materials in biomedical devices. In this work, we modified silicone surfaces to promote formation of high coverage and stable biofilms by a non-pathogenic Escherichia coli strain 83972 with type 1 fimbriae (fim+) to interfere with the colonization of an aggressive biofilm-forming, uropathogenic Enterococcus faecalis. Although it is well known that mannoside surfaces promote the initial adherence of fim+ E. coli through binding to the FimH receptor at the tip of the type 1 fimbriae, it is not clear whether the fast initial adherence could lead to a high coverage and stable protective biofilm. To explore the role of mannoside ligands, we synthesized a series of alkyl and aryl mannosides varied in the structure and immobilized them on silicone surfaces pre-coated with a poly(amidoamine) (PAMAM) dendrimer. We found that stable and densely packed benign E. coli biofilms were formed on the surfaces presenting biphenyl mannoside with the highest initial adherence of fim+ E. coli. These non-pathogenic biofilms prevented the colonization of E. faecalis for 11 days at a high concentration (10(8) CFU mL(-1), 100,000 times above the diagnostic threshold for urinary tract infection) in the nutrient-rich Lysogeny Broth (LB) media. The result shows a correlation among the initial adherence of fim+ E. coli 83972, the coverage and long

  9. Comparative Genomic Analysis Reveals a Possible Novel Non-Tuberculous Mycobacterium Species with High Pathogenic Potential.

    PubMed

    Choo, Siew Woh; Dutta, Avirup; Wong, Guat Jah; Wee, Wei Yee; Ang, Mia Yang; Siow, Cheuk Chuen

    2016-01-01

    Mycobacteria have been reported to cause a wide range of human diseases. We present the first whole-genome study of a Non-Tuberculous Mycobacterium, Mycobacterium sp. UM_CSW (referred to hereafter as UM_CSW), isolated from a patient diagnosed with bronchiectasis. Our data suggest that this clinical isolate is likely a novel mycobacterial species, supported by clear evidence from molecular phylogenetic, comparative genomic, ANI and AAI analyses. UM_CSW is closely related to the Mycobacterium avium complex. While it has characteristic features of an environmental bacterium, it also shows a high pathogenic potential with the presence of a wide variety of putative genes related to bacterial virulence and shares very similar pathogenomic profiles with the known pathogenic mycobacterial species. Thus, we conclude that this possible novel Mycobacterium species should be tightly monitored for its possible causative role in human infections. PMID:27035710

  10. International biosecurity symposium : securing high consequence pathogens and toxins : symposium summary.

    SciTech Connect

    Not Available

    2004-06-01

    The National Nuclear Security Administration (NNSA) Office of Nonproliferation Policy sponsored an international biosecurity symposium at Sandia National Laboratories (SNL). The event, entitled 'Securing High Consequence Pathogens and Toxins', took place from February 1 to February 6, 2004 and was hosted by Dr. Reynolds M. Salerno, Principal Member of the Technical Staff and Program Manager of the Biosecurity program at Sandia. Over 60 bioscience and policy experts from 14 countries gathered to discuss biosecurity, a strategy aimed at preventing the theft and sabotage of dangerous pathogens and toxins from bioscience facilities. Presentations delivered during the symposium were interspersed with targeted discussions that elucidated, among other things, the need for subsequent regional workshops on biosecurity, and a desire for additional work toward developing international biosecurity guidelines.

  11. Comparative Genomic Analysis Reveals a Possible Novel Non-Tuberculous Mycobacterium Species with High Pathogenic Potential

    PubMed Central

    Choo, Siew Woh; Dutta, Avirup; Wong, Guat Jah; Wee, Wei Yee; Ang, Mia Yang; Siow, Cheuk Chuen

    2016-01-01

    Mycobacteria have been reported to cause a wide range of human diseases. We present the first whole-genome study of a Non-Tuberculous Mycobacterium, Mycobacterium sp. UM_CSW (referred to hereafter as UM_CSW), isolated from a patient diagnosed with bronchiectasis. Our data suggest that this clinical isolate is likely a novel mycobacterial species, supported by clear evidence from molecular phylogenetic, comparative genomic, ANI and AAI analyses. UM_CSW is closely related to the Mycobacterium avium complex. While it has characteristic features of an environmental bacterium, it also shows a high pathogenic potential with the presence of a wide variety of putative genes related to bacterial virulence and shares very similar pathogenomic profiles with the known pathogenic mycobacterial species. Thus, we conclude that this possible novel Mycobacterium species should be tightly monitored for its possible causative role in human infections. PMID:27035710

  12. Vaccination of gallinaceous poultry for H5N1 highly pathogenic avian influenza: Current questions and new technology

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Historically, vaccination for avian influenza virus (AIV) in poultry has not been routine for either high pathogenicity (HP) AIV or low pathogenicity (LP) AIV although it has been used in some locations in recent years where AIV is present (i.e. vaccine is not used unless there is a known challenge)...

  13. Characaterization of H5N1 highly pathogenic avian influenza viruses isolated from poultry in Pakistan 2006-2008

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Nine avian influenza viruses (AIV), H5N1 subtype, were isolated from dead poultry in the Karachi region of Pakistan from 2006-2008. The intravenous pathogenicity indices and HA protein cleavage sites of all nine viruses were consistent with highly pathogenic AIV. Based on phylogenetic analysis of ...

  14. Experimental infection of mallard ducks with different subtype H5 and H7 highly pathogenic avian influenza viruses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Highly pathogenic avian influenza viruses (HPAIV’s) remain a threat to poultry worldwide. Avian influenza viruses, including HPAIV, are usually non-pathogenic for ducks and other wild aquatic birds, with the exception of some Asian lineage H5N1 HPAIVs which can cause severe disease in ducks. With ...

  15. Pathobiological features of a novel, highly pathogenic avian influenza A(H5N8) virus.

    PubMed

    Kim, Young-Il; Pascua, Philippe Noriel Q; Kwon, Hyeok-Il; Lim, Gyo-Jin; Kim, Eun-Ha; Yoon, Sun-Woo; Park, Su-Jin; Kim, Se Mi; Choi, Eun-Ji; Si, Young-Jae; Lee, Ok-Jun; Shim, Woo-Sub; Kim, Si-Wook; Mo, In-Pil; Bae, Yeonji; Lim, Yong Taik; Sung, Moon Hee; Kim, Chul-Joong; Webby, Richard J; Webster, Robert G; Choi, Young Ki

    2014-10-01

    The endemicity of highly pathogenic avian influenza (HPAI) A(H5N1) viruses in Asia has led to the generation of reassortant H5 strains with novel gene constellations. A newly emerged HPAI A(H5N8) virus caused poultry outbreaks in the Republic of Korea in 2014. Because newly emerging high-pathogenicity H5 viruses continue to pose public health risks, it is imperative that their pathobiological properties be examined. Here, we characterized A/mallard duck/Korea/W452/2014 (MDk/W452(H5N8)), a representative virus, and evaluated its pathogenic and pandemic potential in various animal models. We found that MDk/W452(H5N8), which originated from the reassortment of wild bird viruses harbored by migratory waterfowl in eastern China, replicated systemically and was lethal in chickens, but appeared to be attenuated, albeit efficiently transmitted, in ducks. Despite predominant attachment to avian-like virus receptors, MDk/W452(H5N8) also exhibited detectable human virus-like receptor binding and replicated in human respiratory tract tissues. In mice, MDk/W452(H5N8) was moderately pathogenic and had limited tissue tropism relative to previous HPAI A(H5N1) viruses. It also induced moderate nasal wash titers in inoculated ferrets; additionally, it was recovered in extrapulmonary tissues and one of three direct-contact ferrets seroconverted without shedding. Moreover, domesticated cats appeared to be more susceptible than dogs to virus infection. With their potential to become established in ducks, continued circulation of A(H5N8) viruses could alter the genetic evolution of pre-existing avian poultry strains. Overall, detailed virological investigation remains a necessity given the capacity of H5 viruses to evolve to cause human illness with few changes in the viral genome. PMID:26038499

  16. Pathobiological features of a novel, highly pathogenic avian influenza A(H5N8) virus

    PubMed Central

    Kim, Young-Il; Pascua, Philippe Noriel Q; Kwon, Hyeok-Il; Lim, Gyo-Jin; Kim, Eun-Ha; Yoon, Sun-Woo; Park, Su-Jin; Kim, Se Mi; Choi, Eun-Ji; Si, Young-Jae; Lee, Ok-Jun; Shim, Woo-Sub; Kim, Si-Wook; Mo, In-Pil; Bae, Yeonji; Lim, Yong Taik; Sung, Moon Hee; Kim, Chul-Joong; Webby, Richard J; Webster, Robert G; Choi, Young Ki

    2014-01-01

    The endemicity of highly pathogenic avian influenza (HPAI) A(H5N1) viruses in Asia has led to the generation of reassortant H5 strains with novel gene constellations. A newly emerged HPAI A(H5N8) virus caused poultry outbreaks in the Republic of Korea in 2014. Because newly emerging high-pathogenicity H5 viruses continue to pose public health risks, it is imperative that their pathobiological properties be examined. Here, we characterized A/mallard duck/Korea/W452/2014 (MDk/W452(H5N8)), a representative virus, and evaluated its pathogenic and pandemic potential in various animal models. We found that MDk/W452(H5N8), which originated from the reassortment of wild bird viruses harbored by migratory waterfowl in eastern China, replicated systemically and was lethal in chickens, but appeared to be attenuated, albeit efficiently transmitted, in ducks. Despite predominant attachment to avian-like virus receptors, MDk/W452(H5N8) also exhibited detectable human virus-like receptor binding and replicated in human respiratory tract tissues. In mice, MDk/W452(H5N8) was moderately pathogenic and had limited tissue tropism relative to previous HPAI A(H5N1) viruses. It also induced moderate nasal wash titers in inoculated ferrets; additionally, it was recovered in extrapulmonary tissues and one of three direct-contact ferrets seroconverted without shedding. Moreover, domesticated cats appeared to be more susceptible than dogs to virus infection. With their potential to become established in ducks, continued circulation of A(H5N8) viruses could alter the genetic evolution of pre-existing avian poultry strains. Overall, detailed virological investigation remains a necessity given the capacity of H5 viruses to evolve to cause human illness with few changes in the viral genome. PMID:26038499

  17. Dynamics of low and high pathogenic avian influenza in wild and domestic bird populations.

    PubMed

    Tuncer, Necibe; Torres, Juan; Martcheva, Maia; Barfield, Michael; Holt, Robert D

    2016-01-01

    This paper introduces a time-since-recovery structured, multi-strain, multi-population model of avian influenza. Influenza A viruses infect many species of wild and domestic birds and are classified into two groups based on their ability to cause disease: low pathogenic avian influenza (LPAI) and high pathogenic avian influenza (HPAI). Prior infection with LPAI provides partial immunity towards HPAI. The model introduced in this paper structures LPAI-recovered birds (wild and domestic) with time-since-recovery and includes cross-immunity towards HPAI that can fade with time. The model has a unique disease-free equilibrium (DFE), unique LPAI-only and HPAI-only equilibria and at least one coexistence equilibrium. We compute the reproduction numbers of LPAI ([Formula: see text]) and HPAI ([Formula: see text]) and show that the DFE is locally asymptotically stable when [Formula: see text] and [Formula: see text]. A unique LPAI-only (HPAI-only) equilibrium exists when [Formula: see text] ([Formula: see text]) and it is locally asymptotically stable if HPAI (LPAI) cannot invade the equilibrium, that is, if the invasion number [Formula: see text] ([Formula: see text]). We show using numerical simulations that the ODE version of the model, which is obtained by discarding the time-since-recovery structures (making cross-immunity constant), can exhibit oscillations, and also that the pathogens LPAI and HPAI can coexist with sustained oscillations in both populations. Through simulations, we show that even if both populations (wild and domestic) are sinks when alone, LPAI and HPAI can persist in both populations combined. Thus, reducing the reproduction numbers of LPAI and HPAI in each population to below unity is not enough to eradicate the disease. The pathogens can continue to coexist in both populations unless transmission between the populations is reduced. PMID:26667351

  18. Inactivation of bacterial pathogens in human milk by high-pressure processing.

    PubMed

    Viazis, S; Farkas, B E; Jaykus, L A

    2008-01-01

    Low-temperature, long-time (LTLT) pasteurization assures the safety of banked human milk; however, heat can destroy important nutritional biomolecules. High-pressure processing (HPP) shows promise as an alternative for pasteurization of breast milk. The purpose of this study was to investigate the efficacy of HPP for inactivation of selected bacterial pathogens in human milk. Human milk was inoculated with one of five pathogens (10(8) to 10(9) CFU/ml), while 0.1% peptone solution solutions with the same levels of each organism were used as controls. The samples were subjected to 400 MPa at 21 to 31 degrees C for 0 to 50 min or to 62.5 degrees C for 0 to 30 min (capillary tube method) to simulate LTLT pasteurization. Tryptic soy agar and selective media were used for enumeration. Traditional thermal pasteurization resulted in inactivation (> 7 log) of all pathogens within 10 min. In human milk and in peptone solution, a 6-log reduction was achieved after 30 min of HPP for Staphylococcus aureus ATCC 6538. After 30 min, S. aureus ATCC 25923 was reduced by 8 log and 6 log in human milk and peptone solution, respectively. Treatments of 4 and 7 min resulted in an 8-log inactivation of Streptococcus agalactiae ATCC 12927 in human milk and peptone solution, respectively, while Listeria monocytogenes ATCC 19115 required 2 min for an 8-log inactivation in human milk. Escherichia coli ATCC 25922 was inactivated by 8 log after 10 min in peptone solution and by 6 log after 30 min in human milk. These data suggest that HPP may be a promising alternative for pasteurization of human milk. Further research should evaluate the efficacy of HPP in the inactivation of relevant viral pathogens.

  19. Pathogenicity of H5N8 highly pathogenic avian influenza viruses isolated from a wild bird fecal specimen and a chicken in Japan in 2014.

    PubMed

    Tanikawa, Taichiro; Kanehira, Katsushi; Tsunekuni, Ryota; Uchida, Yuko; Takemae, Nobuhiro; Saito, Takehiko

    2016-04-01

    Poultry outbreaks caused by H5N8 highly pathogenic avian influenza viruses (HPAIVs) occurred in Japan between December 2014 and January 2015. During the same period; H5N8 HPAIVs were isolated from wild birds and the environment in Japan. The hemagglutinin (HA) genes of these isolates were found to belong to clade 2.3.4.4 and three sub-groups were distinguishable within this clade. All of the Japanese isolates from poultry outbreaks belonged to the same sub-group; whereas wild bird isolates belonged to the other sub-groups. To examine whether the difference in pathogenicity to chickens between isolates of different HA sub-groups of clade 2.3.4.4 could explain why the Japanese poultry outbreaks were only caused by a particular sub-group; pathogenicities of A/chicken/Miyazaki/7/2014 (Miyazaki2014; sub-group C) and A/duck/Chiba/26-372-48/2014 (Chiba2014; sub-group A) to chickens were compared and it was found that the lethality of Miyazaki2014 in chickens was lower than that of Chiba2014; according to the 50% chicken lethal dose. This indicated that differences in pathogenicity may not explain why the Japanese poultry outbreaks only involved group C isolates. PMID:26916882

  20. Newly Emergent Highly Pathogenic H5N9 Subtype Avian Influenza A Virus

    PubMed Central

    Yu, Yang; Wang, Xingbo; Jin, Tao; Wang, Hailong; Si, Weiying; Yang, Hui; Wu, Jiusheng; Yan, Yan; Liu, Guang; Sang, Xiaoyu; Wu, Xiaopeng; Gao, Yuwei; Xia, Xianzhu; Yu, Xinfen; Pan, Jingcao; Gao, George F.

    2015-01-01

    ABSTRACT The novel H7N9 avian influenza virus (AIV) was demonstrated to cause severe human respiratory infections in China. Here, we examined poultry specimens from live bird markets linked to human H7N9 infection in Hangzhou, China. Metagenomic sequencing revealed mixed subtypes (H5, H7, H9, N1, N2, and N9). Subsequently, AIV subtypes H5N9, H7N9, and H9N2 were isolated. Evolutionary analysis showed that the hemagglutinin gene of the novel H5N9 virus originated from A/Muscovy duck/Vietnam/LBM227/2012 (H5N1), which belongs to clade 2.3.2.1. The neuraminidase gene of the novel H5N9 virus originated from human-infective A/Hangzhou/1/2013 (H7N9). The six internal genes were similar to those of other H5N1, H7N9, and H9N2 virus strains. The virus harbored the PQRERRRKR/GL motif characteristic of highly pathogenic AIVs at the HA cleavage site. Receptor-binding experiments demonstrated that the virus binds α-2,3 sialic acid but not α-2,6 sialic acid. Identically, pathogenicity experiments also showed that the virus caused low mortality rates in mice. This newly isolated H5N9 virus is a highly pathogenic reassortant virus originating from H5N1, H7N9, and H9N2 subtypes. Live bird markets represent a potential transmission risk to public health and the poultry industry. IMPORTANCE This investigation confirms that the novel H5N9 subtype avian influenza A virus is a reassortant strain originating from H5N1, H7N9, and H9N2 subtypes and is totally different from the H5N9 viruses reported before. The novel H5N9 virus acquired a highly pathogenic H5 gene and an N9 gene from human-infecting subtype H7N9 but caused low mortality rates in mice. Whether this novel H5N9 virus will cause human infections from its avian host and become a pandemic subtype is not known yet. It is therefore imperative to assess the risk of emergence of this novel reassortant virus with potential transmissibility to public health. PMID:26085150

  1. Development of a high- versus low-pathogenicity model of the free-living amoeba Naegleria fowleri.

    PubMed

    Burri, Denise C; Gottstein, Bruno; Zumkehr, Béatrice; Hemphill, Andrew; Schürch, Nadia; Wittwer, Matthias; Müller, Norbert

    2012-10-01

    Species in the genus Naegleria are free-living amoebae of the soil and warm fresh water. Although around 30 species have been recognized, Naegleria fowleri is the only one that causes primary amoebic meningoencephalitis (PAM) in humans. PAM is an acute and fast progressing disease affecting the central nervous system. Most of the patients die within 1-2 weeks of exposure to the infectious water source. The fact that N. fowleri causes such fast progressing and highly lethal infections has opened many questions regarding the relevant pathogenicity factors of the amoeba. In order to investigate the pathogenesis of N. fowleri under defined experimental conditions, we developed a novel high- versus low-pathogenicity model for this pathogen. We showed that the composition of the axenic growth media influenced growth behaviour and morphology, as well as in vitro cytotoxicity and in vivo pathogenicity of N. fowleri. Trophozoites maintained in Nelson's medium were highly pathogenic for mice, demonstrated rapid in vitro proliferation, characteristic expression of surface membrane vesicles and a small cell diameter, and killed target mouse fibroblasts by both contact-dependent and -independent destruction. In contrast, N. fowleri cultured in PYNFH medium exhibited a low pathogenicity, slower growth, increased cell size and contact-dependent target cell destruction. However, cultivation of the amoeba in PYNFH medium supplemented with liver hydrolysate (LH) resulted in trophozoites that were highly pathogenic in mice, and demonstrated an intermediate proliferation rate in vitro, diminished cell diameter and contact-dependent target cell destruction. Thus, in this model, the presence of LH resulted in increased proliferation of trophozoites in vitro and enhanced pathogenicity of N. fowleri in mice. However, neither in vitro cytotoxicity mechanisms nor the presence of membrane vesicles on the surface correlated with the pathologic potential of the amoeba. This indicated that the

  2. The hidden face of academic researches on classified highly pathogenic microorganisms.

    PubMed

    Devaux, Christian A

    2015-01-01

    Highly pathogenic microorganisms and toxins are manipulated in academic laboratories for fundamental research purposes, diagnostics, drugs and vaccines development. Obviously, these infectious pathogens represent a potential risk for human and/or animal health and their accidental or intentional release (biosafety and biosecurity, respectively) is a major concern of governments. In the past decade, several incidents have occurred in laboratories and reported by media causing fear and raising a sense of suspicion against biologists. Some scientists have been ordered by US government to leave their laboratory for long periods of time following the occurrence of an incident involving infectious pathogens; in other cases laboratories have been shut down and universities have been forced to pay fines and incur a long-term ban on funding after gross negligence of biosafety/biosecurity procedures. Measures of criminal sanctions have also been taken to minimize the risk that such incidents can reoccur. As United States and many other countries, France has recently strengthened its legal measures for laboratories' protection. During the past two decades, France has adopted a series of specific restriction measures to better protect scientific discoveries with a potential economic/social impact and prevent their misuse by ill-intentioned people without affecting the progress of science through fundamental research. French legal regulations concerning scientific discoveries have progressively strengthened since 2001, until the publication in November 2011 of a decree concerning the "PPST" (for "Protection du Potentiel Scientifique et Technique de la nation", the protection of sensitive scientific data). Following the same logic of protection of sensitive scientific researches, regulations were also adopted in an order published in April 2012 concerning the biology and health field. The aim was to define the legal framework that precise the conditions for authorizing

  3. The hidden face of academic researches on classified highly pathogenic microorganisms.

    PubMed

    Devaux, Christian A

    2015-01-01

    Highly pathogenic microorganisms and toxins are manipulated in academic laboratories for fundamental research purposes, diagnostics, drugs and vaccines development. Obviously, these infectious pathogens represent a potential risk for human and/or animal health and their accidental or intentional release (biosafety and biosecurity, respectively) is a major concern of governments. In the past decade, several incidents have occurred in laboratories and reported by media causing fear and raising a sense of suspicion against biologists. Some scientists have been ordered by US government to leave their laboratory for long periods of time following the occurrence of an incident involving infectious pathogens; in other cases laboratories have been shut down and universities have been forced to pay fines and incur a long-term ban on funding after gross negligence of biosafety/biosecurity procedures. Measures of criminal sanctions have also been taken to minimize the risk that such incidents can reoccur. As United States and many other countries, France has recently strengthened its legal measures for laboratories' protection. During the past two decades, France has adopted a series of specific restriction measures to better protect scientific discoveries with a potential economic/social impact and prevent their misuse by ill-intentioned people without affecting the progress of science through fundamental research. French legal regulations concerning scientific discoveries have progressively strengthened since 2001, until the publication in November 2011 of a decree concerning the "PPST" (for "Protection du Potentiel Scientifique et Technique de la nation", the protection of sensitive scientific data). Following the same logic of protection of sensitive scientific researches, regulations were also adopted in an order published in April 2012 concerning the biology and health field. The aim was to define the legal framework that precise the conditions for authorizing

  4. The development of high hydrostatic pressure processes as an alternative to other pathogen reduction methods.

    PubMed

    Demazeau, G; Rivalain, N

    2011-06-01

    In biology, scientist's interest for high hydrostatic pressure (HHP) has increased over the last 20 years, for both research and industrial developments, mainly because of the low energy associated with its application in liquid phase and its capacity to inactivate pathogens. It is now considered as an interesting alternative to heat treatments for the inactivation of contaminants in many products, from foods to pharmaceutical preparations. This last statement implies different objectives according to the type of product. The therapeutic properties of pharmaceutical preparations or other biological media of physiological importance are in general associated with specific and well-defined molecules such as proteins. Their activity mainly depends on their spatial conformation, maintained by weak chemical bonds that are often pressure sensitive. In this case, the optimization of a HHP process can be more complex than for foods, for which the organoleptic molecules are less pressure sensitive, and the evaluation of their preservation is more subjective and highly dependent on the consumers acceptance. The objective of this review is therefore to underline how, even if the basic concept for the optimization of a pathogen reduction process using HHP is the same whatever the product, major differences arise from the product itself and its final use.

  5. Use of high-throughput mass spectrometry to elucidate host pathogen interactions in Salmonella

    SciTech Connect

    Rodland, Karin D.; Adkins, Joshua N.; Ansong, Charles; Chowdhury, Saiful M.; Manes, Nathan P.; Shi, Liang; Yoon, Hyunjin; Smith, Richard D.; Heffron, Fred

    2008-12-01

    Capabilities in mass spectrometry are evolving rapidly, with recent improvements in sensitivity, data analysis, and most important, from the standpoint of this review, much higher throughput allowing analysis of many samples in a single day. This short review describes how these improvements in mass spectrometry can be used to dissect host-pathogen interactions using Salmonella as a model system. This approach enabled direct identification of the majority of annotated Salmonella proteins, quantitation of expression changes under various in vitro growth conditions, and new insights into virulence and expression of Salmonella proteins within host cell cells. One of the most significant findings is that a very high percentage of the all annotated genes (>20%) in Salmonella are regulated post-transcriptionally. In addition, new and unexpected interactions have been identified for several Salmonella virulence regulators that involve protein-protein interactions, suggesting additional functions of these regulators in coordinating virulence expression. Overall high throughput mass spectrometry provides a new view of pathogen-host interactions emphasizing the protein products and defining how protein interactions determine the outcome of infection.

  6. Use of high-throughput mass spectrometry to elucidate host-pathogen interactions in Salmonella

    SciTech Connect

    Rodland, Karin D.; Adkins, Joshua N.; Ansong, Charles; Chowdhury, Saiful M.; Manes, Nathan P.; Shi, Liang; Yoon, Hyunjin; Smith, Richard D.; Heffron, Fred

    2008-12-01

    New improvements to mass spectrometry include increased sensitivity, improvements in analyzing the collected data, and most important, from the standpoint of this review, a much higher throughput allowing analysis of many samples in a single day. This short review describes how host-pathogen interactions can be dissected by mass spectrometry using Salmonella as a model system. The approach allowed direct identification of the majority of annotate Salmonella proteins, how expression changed under various in vitro growth conditions, and how this relates to virulence and expression within host cell cells. One of the most significant findings is that a very high percentage of the all annotated genes (>20%) are regulated post-transcriptionally. In addition, new and unexpected interactions have been identified for several Salmonella virulence regulators that involve protein-protein interactions suggesting additional functions of the regulator in coordinating virulence expression. Overall high throughput mass spectrometer provides a new view of pathogen-host interaction emphasizing the protein products and defining how protein interactions determine the outcome of infection.

  7. Incorporating risk communication into highly pathogenic avian influenza preparedness and response efforts.

    PubMed

    Voss, Shauna J; Malladi, Sasidhar; Sampedro, Fernando; Snider, Tim; Goldsmith, Timothy; Hueston, William D; Lauer, Dale C; Halvorson, David A

    2012-12-01

    A highly pathogenic avian influenza (HPAI) outbreak in the United States will initiate a federal emergency response effort that will consist of disease control and eradication efforts, including quarantine and movement control measures. These movement control measures will not only apply to live animals but also to animal products. However, with current egg industry "just-in-time" production practices, limited storage is available to hold eggs. As a result, stop movement orders can have significant unintended negative consequences, including severe disruptions to the food supply chain. Because stakeholders' perceptions of risk vary, waiting to initiate communication efforts until an HPAI event occurs can hinder disease control efforts, including the willingness of producers to comply with the response, and also can affect consumers' demand for the product. A public-private-academic partnership was formed to assess actual risks involved in the movement of egg industry products during an HPAI event through product specific, proactive risk assessments. The risk analysis process engaged a broad representation of stakeholders and promoted effective risk management and communication strategies before an HPAI outbreak event. This multidisciplinary team used the risk assessments in the development of the United States Department of Agriculture, Highly Pathogenic Avian Influenza Secure Egg Supply Plan, a comprehensive response plan that strives to maintain continuity of business. The collaborative approach that was used demonstrates how a proactive risk communication strategy that involves many different stakeholders can be valuable in the development of a foreign animal disease response plan and build working relationships, trust, and understanding. PMID:23402134

  8. Characterization of a highly pathogenic avian influenza H5N1 virus isolated from an ostrich.

    PubMed

    Yang, Penghui; Dongmei; Wang, Cheng; Tang, Chong; Xing, Li; Luo, Deyan; Zhan, Zhongpeng; Duan, Yueqiang; Jia, Weihong; Peng, Daxin; Liu, Xiufan; Wang, Xiliang

    2010-06-11

    The continued spread of a highly pathogenic avian influenza (HPAI) H5N1 virus among poultry and wild birds has posed a potential threat to human public health. An influenza pandemic happens, when a new subtype that has not previously circulated in humans emerges. Almost all of the influenza pandemics in history have originated from avian influenza viruses (AIV). Birds are significant reservoirs of influenza viruses. In the present study, we performed a survey of avian influenza virus in ostriches and H5N1 virus (A/Ostrich/SuZhou/097/03, China097) was isolated. This H5N1 virus is highly pathogenic to both chickens and mice. It is also able to replicate in the lungs of, and to cause death in, BALB/c mice following intranasal administration. It forms plaques in chicken embryo fibroblast (CEF) cells in the absence of trypsin. The hemagglutinin (HA) gene of the virus is genetically similar to A/Goose/Guangdong/1/96(H5N1) and belongs to clade 0. The HA sequence contains multiple basic amino acids adjacent to the cleavage site, a motif associated with HPAI viruses. More importantly, the existence of H5N1 isolates in ostriches highlights the potential threat of wild bird infections to veterinary and public health. PMID:20497905

  9. Bacterial pathogens and community composition in advanced sewage treatment systems revealed by metagenomics analysis based on high-throughput sequencing.

    PubMed

    Lu, Xin; Zhang, Xu-Xiang; Wang, Zhu; Huang, Kailong; Wang, Yuan; Liang, Weigang; Tan, Yunfei; Liu, Bo; Tang, Junying

    2015-01-01

    This study used 454 pyrosequencing, Illumina high-throughput sequencing and metagenomic analysis to investigate bacterial pathogens and their potential virulence in a sewage treatment plant (STP) applying both conventional and advanced treatment processes. Pyrosequencing and Illumina sequencing consistently demonstrated that Arcobacter genus occupied over 43.42% of total abundance of potential pathogens in the STP. At species level, potential pathogens Arcobacter butzleri, Aeromonas hydrophila and Klebsiella pneumonia dominated in raw sewage, which was also confirmed by quantitative real time PCR. Illumina sequencing also revealed prevalence of various types of pathogenicity islands and virulence proteins in the STP. Most of the potential pathogens and virulence factors were eliminated in the STP, and the removal efficiency mainly depended on oxidation ditch. Compared with sand filtration, magnetic resin seemed to have higher removals in most of the potential pathogens and virulence factors. However, presence of the residual A. butzleri in the final effluent still deserves more concerns. The findings indicate that sewage acts as an important source of environmental pathogens, but STPs can effectively control their spread in the environment. Joint use of the high-throughput sequencing technologies is considered a reliable method for deep and comprehensive overview of environmental bacterial virulence.

  10. Bacterial Pathogens and Community Composition in Advanced Sewage Treatment Systems Revealed by Metagenomics Analysis Based on High-Throughput Sequencing

    PubMed Central

    Lu, Xin; Zhang, Xu-Xiang; Wang, Zhu; Huang, Kailong; Wang, Yuan; Liang, Weigang; Tan, Yunfei; Liu, Bo; Tang, Junying

    2015-01-01

    This study used 454 pyrosequencing, Illumina high-throughput sequencing and metagenomic analysis to investigate bacterial pathogens and their potential virulence in a sewage treatment plant (STP) applying both conventional and advanced treatment processes. Pyrosequencing and Illumina sequencing consistently demonstrated that Arcobacter genus occupied over 43.42% of total abundance of potential pathogens in the STP. At species level, potential pathogens Arcobacter butzleri, Aeromonas hydrophila and Klebsiella pneumonia dominated in raw sewage, which was also confirmed by quantitative real time PCR. Illumina sequencing also revealed prevalence of various types of pathogenicity islands and virulence proteins in the STP. Most of the potential pathogens and virulence factors were eliminated in the STP, and the removal efficiency mainly depended on oxidation ditch. Compared with sand filtration, magnetic resin seemed to have higher removals in most of the potential pathogens and virulence factors. However, presence of the residual A. butzleri in the final effluent still deserves more concerns. The findings indicate that sewage acts as an important source of environmental pathogens, but STPs can effectively control their spread in the environment. Joint use of the high-throughput sequencing technologies is considered a reliable method for deep and comprehensive overview of environmental bacterial virulence. PMID:25938416

  11. Pathogenicity in domestic ducks and mice of highly pathogenic H5N1 clade 2.3.2.1 influenza viruses recently circulating in Eastern Asia.

    PubMed

    Kang, Hyun-Mi; Choi, Jun-Gu; Kim, Kwang-Il; Kim, Bang-Sil; Batchuluun, Damdinjav; Erdene-Ochir, Tseren-Ochir; Kim, Min-Chul; Kwon, Jun-Hun; Park, Choi-Kyu; Lee, Youn-Jeong

    2013-12-27

    Influenza virus A (H5N1) clade 2.3.2.1 has recently caused widespread outbreaks of disease in domestic poultry and wild birds in Eastern Asia. In the current study, the antigenicity and pathogenicity of three clade 2.3.2.1 viruses (Ck/Kr/Gimje/08, Ws/Mongolia/1/09, and Ws/Mongolia/7/10) were investigated in domestic ducks and mice. The H5N1 influenza viruses in this study were antigenically similar to each other (r-values of 0.35-1.4). The three viruses replicated systemically in all tissues tested in domestic ducks, indicating high pathogenicity. However, the viruses produced different clinical signs and mortality rates: Ck/Kr/Gimje/08 and Ws/Mongolia/1/09 resulted in 100% mortality with severe neurological signs, whereas Ws/Mongolia/7/10 resulted in 50% mortality with relatively mild neurological signs. In mice, infection with Ck/Kr/Gimje/08 and Ws/Mongolia/7/10 resulted in weight loss that peaked at 4 days post-infection (22.3% and 20.8%, respectively), same MLD50 (2.2 Log10 EID50) and systemic replication. The three viruses had K deletion at the -2 position of the HA1-connecting peptide (PQRERRRK-R), which is associated with increased virulence in domestic ducks and harbored NA stalk deletion, NS1 deletion and mutation of P42S in NS1, and full length (90aa) in PB1-F2, which confer increased virulence in mice. Our study shows that clade 2.3.2.1 viruses from Korea and Mongolia are antigenically similar and highly pathogenic in both domestic ducks and mice. Moreover, we provide molecular determinants of the clade 2.3.2.1 viruses associated with the pathogenicity in domestic ducks and mice, respectively.

  12. Model-based evaluation of highly and low pathogenic avian influenza dynamics in wild birds

    USGS Publications Warehouse

    Hénaux, Viviane; Samuel, Michael D.; Bunck, Christine M.

    2010-01-01

    There is growing interest in avian influenza (AI) epidemiology to predict disease risk in wild and domestic birds, and prevent transmission to humans. However, understanding the epidemic dynamics of highly pathogenic (HPAI) viruses remains challenging because they have rarely been detected in wild birds. We used modeling to integrate available scientific information from laboratory and field studies, evaluate AI dynamics in individual hosts and waterfowl populations, and identify key areas for future research. We developed a Susceptible-Exposed-Infectious-Recovered (SEIR) model and used published laboratory challenge studies to estimate epidemiological parameters (rate of infection, latency period, recovery and mortality rates), considering the importance of age classes, and virus pathogenicity. Infectious contact leads to infection and virus shedding within 1–2 days, followed by relatively slower period for recovery or mortality. We found a shorter infectious period for HPAI than low pathogenic (LP) AI, which may explain that HPAI has been much harder to detect than LPAI during surveillance programs. Our model predicted a rapid LPAI epidemic curve, with a median duration of infection of 50–60 days and no fatalities. In contrast, HPAI dynamics had lower prevalence and higher mortality, especially in young birds. Based on field data from LPAI studies, our model suggests to increase surveillance for HPAI in post-breeding areas, because the presence of immunologically naïve young birds is predicted to cause higher HPAI prevalence and bird losses during this season. Our results indicate a better understanding of the transmission, infection, and immunity-related processes is required to refine predictions of AI risk and spread, improve surveillance for HPAI in wild birds, and develop disease control strategies to reduce potential transmission to domestic birds and/or humans.

  13. A highly sensitive and flexible magnetic nanoprobe labeled immunochromatographic assay platform for pathogen Vibrio parahaemolyticus.

    PubMed

    Liu, Yingying; Zhang, Zhaohuan; Wang, Yilong; Zhao, Yong; Lu, Ying; Xu, Xiaowei; Yan, Jun; Pan, Yingjie

    2015-10-15

    A magnetic nanoprobe labeled immunochromatographic test strip (MNP/ICTS) was developed to detect food-borne pathogen Vibrio parahaemolyticus. Specific antibody against V. parahaemolyticus was used as test line by coating onto the nitrocellulose membrane. Magnetic nanoprobe was prepared by immobilizing the specific antibody onto the surface of superparamagnetic nanoparticles. Specificity and sensitivity of the MNP/ICTS system were verified by artificially contaminated shrimp homogenate samples. Reliability and application feasibility of the MNP/ICTS system were demonstrated by using seafood samples (n=36). Comparing with polymerase chain reaction (PCR) and traditional culture methods, the MNP/ICTS system is found to be not only a rapid qualitative analysis (~10 min), but also an accurately quantitative detection platform. Through its rapid magnetic separation property, the MNP/ICTS system is capable to flexibly combine with a sample enrichment and pre-incubation process. This combination makes the qualitative sensitivity for the food samples surged more than 100-fold. A naked-eye observation of 1.58×10(2) CFU/g V. parahaemolyticus was realized. This sensitivity could meet the V. parahaemolyticus test threshold value in many countries. Also, the total sample pre-treatment plus MNP/ICTS assay only needs about 4.5h. Namely, we can get test results in a day. Hence, the developed MNP/ICTS assay platform is simple, rapid and highly sensitive. It is a flexible test platform for pathogen detection. The favorable comparison with PCR and culture methods further proves that the developed MNP/ICTS is applicable into food-borne pathogen or other areas where a simple, rapid, sensitive and point-of-care analysis is desirable. PMID:26188497

  14. A highly sensitive and flexible magnetic nanoprobe labeled immunochromatographic assay platform for pathogen Vibrio parahaemolyticus.

    PubMed

    Liu, Yingying; Zhang, Zhaohuan; Wang, Yilong; Zhao, Yong; Lu, Ying; Xu, Xiaowei; Yan, Jun; Pan, Yingjie

    2015-10-15

    A magnetic nanoprobe labeled immunochromatographic test strip (MNP/ICTS) was developed to detect food-borne pathogen Vibrio parahaemolyticus. Specific antibody against V. parahaemolyticus was used as test line by coating onto the nitrocellulose membrane. Magnetic nanoprobe was prepared by immobilizing the specific antibody onto the surface of superparamagnetic nanoparticles. Specificity and sensitivity of the MNP/ICTS system were verified by artificially contaminated shrimp homogenate samples. Reliability and application feasibility of the MNP/ICTS system were demonstrated by using seafood samples (n=36). Comparing with polymerase chain reaction (PCR) and traditional culture methods, the MNP/ICTS system is found to be not only a rapid qualitative analysis (~10 min), but also an accurately quantitative detection platform. Through its rapid magnetic separation property, the MNP/ICTS system is capable to flexibly combine with a sample enrichment and pre-incubation process. This combination makes the qualitative sensitivity for the food samples surged more than 100-fold. A naked-eye observation of 1.58×10(2) CFU/g V. parahaemolyticus was realized. This sensitivity could meet the V. parahaemolyticus test threshold value in many countries. Also, the total sample pre-treatment plus MNP/ICTS assay only needs about 4.5h. Namely, we can get test results in a day. Hence, the developed MNP/ICTS assay platform is simple, rapid and highly sensitive. It is a flexible test platform for pathogen detection. The favorable comparison with PCR and culture methods further proves that the developed MNP/ICTS is applicable into food-borne pathogen or other areas where a simple, rapid, sensitive and point-of-care analysis is desirable.

  15. Pseudotyping of vesicular stomatitis virus with the envelope glycoproteins of highly pathogenic avian influenza viruses.

    PubMed

    Zimmer, Gert; Locher, Samira; Berger Rentsch, Marianne; Halbherr, Stefan J

    2014-08-01

    Pseudotype viruses are useful for studying the envelope proteins of harmful viruses. This work describes the pseudotyping of vesicular stomatitis virus (VSV) with the envelope glycoproteins of highly pathogenic avian influenza viruses. VSV lacking the homotypic glycoprotein (G) gene (VSVΔG) was used to express haemagglutinin (HA), neuraminidase (NA) or the combination of both. Propagation-competent pseudotype viruses were only obtained when HA and NA were expressed from the same vector genome. Pseudotype viruses containing HA from different H5 clades were neutralized specifically by immune sera directed against the corresponding clade. Fast and sensitive reading of test results was achieved by vector-mediated expression of GFP. Pseudotype viruses expressing a mutant VSV matrix protein showed restricted spread in IFN-competent cells. This pseudotype system will facilitate the detection of neutralizing antibodies against virulent influenza viruses, circumventing the need for high-level biosafety containment.

  16. High Throughput, Multiplexed Pathogen Detection Authenticates Plague Waves in Medieval Venice, Italy

    PubMed Central

    Tran, Thi-Nguyen-Ny; Signoli, Michel; Fozzati, Luigi; Aboudharam, Gérard; Raoult, Didier; Drancourt, Michel

    2011-01-01

    Background Historical records suggest that multiple burial sites from the 14th–16th centuries in Venice, Italy, were used during the Black Death and subsequent plague epidemics. Methodology/Principal Findings High throughput, multiplexed real-time PCR detected DNA of seven highly transmissible pathogens in 173 dental pulp specimens collected from 46 graves. Bartonella quintana DNA was identified in five (2.9%) samples, including three from the 16th century and two from the 15th century, and Yersinia pestis DNA was detected in three (1.7%) samples, including two from the 14th century and one from the 16th century. Partial glpD gene sequencing indicated that the detected Y. pestis was the Orientalis biotype. Conclusions These data document for the first time successive plague epidemics in the medieval European city where quarantine was first instituted in the 14th century. PMID:21423736

  17. Isolation of recombinant phage antibodies targeting the hemagglutinin cleavage site of highly pathogenic avian influenza virus.

    PubMed

    Dong, Jinhua; Sakurai, Akira; Nomura, Namiko; Park, Enoch Y; Shibasaki, Futoshi; Ueda, Hiroshi

    2013-01-01

    Highly pathogenic avian influenza (HPAI) H5N1 viruses, which have emerged in poultry and other wildlife worldwide, contain a characteristic multi-basic cleavage site (CS) in the hemagglutinin protein (HA). Because this arginine-rich CS is unique among influenza virus subtypes, antibodies against this site have the potential to specifically diagnose pathogenic H5N1. By immunizing mice with the CS peptide and screening a phage display library, we isolated four antibody Fab fragment clones that specifically bind the antigen peptide and several HPAI H5N1 HA proteins in different clades. The soluble Fab fragments expressed in Escherichia coli bound the CS peptide and the H5N1 HA protein with nanomolar affinity. In an immunofluorescence assay, these Fab fragments stained cells infected with HPAI H5N1 but not those infected with a less virulent strain. Lastly, all the Fab clones could detect the CS peptide and H5N1 HA protein by open sandwich ELISA. Thus, these recombinant Fab fragments will be useful novel reagents for the rapid and specific detection of HPAI H5N1 virus.

  18. [Highly pathogenic avian influenza in poultry (fowl plague); implications for human health].

    PubMed

    Brugere-Picoux, Jeanne

    2005-11-01

    Since 1997, high-pathogenicity avian influenza (HPAI) virus infection in poultry "avian plague" has emerged as a potential threat to human health, with some fatal cases of bird-to-human transmission. These sporadic infections are caused by H7N7 and H5N1 viruses in Europe and Asia, respectively. The persistence of H5N1 viruses in poultry in several Asian countries, and their appearance in Europe, has raised concerns that the virus might mutate or recombine to create a human pandemic influenza A virus. Wild waterfowl are the natural reservoir of all influenza A viruses, and rarely develop the disease. Since 2002, some H5N1 HPAI viruses have become lethal for waterfowl, cats and humans, indicating an expanding host range. Transmission of H5N1 HPAI viruses from domestic poultry back to resistant domestic and wild ducks and to terrestrial birds (sparrows, pigeons, falcons, etc.) has increased the risk of geographic spread in Asia. These viruses spread through fecal contamination of the environment (particularly groundwater). Low-pathogenicity avian influenza (LPAI) viruses cause localized respiratory and gastrointestinal tract infection and, unlike HPAI viruses, are not detected in blood, muscle or eggs. Detection of HPAI viruses in meat, blood and internal organs of chickens and ducks raises public health concerns and underlines the need to thoroughly cook poultry and eggs consumed in Asia. The last case of HPA1 virus infection in France was notified in 1955.

  19. High reliability transformation of the wheat pathogen Bipolaris sorokiniana using Agrobacterium tumefaciens.

    PubMed

    Nizam, Shadab; Verma, Sandhya; Singh, Kunal; Aggarwal, Rashmi; Srivastava, Krishna Dutt; Verma, Praveen K

    2012-03-01

    Bipolaris sorokiniana, the causal agent of spot blotch of wheat, significantly reduces grain yield worldwide. In order to study pathogenic mechanisms of the fungus, conditions for efficient transformation using Agrobacterium-mediated transformation were investigated. To study different stages of hyphal fusion and pathogenic mechanisms of the fungus, two fluorescence markers viz. the red fluorescent protein (DsRed-Express) and the green fluorescent protein (EGFP1) were constitutively expressed. Southern hybridizations confirmed the presence of T-DNA in all hygromycin B or geneticin resistant transformants, and also showed random and single copy integration. Fluorescence microscopy suggested the high level expression of both DsRed and EGFP fluorescent proteins in spores and mycelia. The results signify that DsRed and EGFP can be used as efficient reporter gene for monitoring B. sorokiniana hyphal fusion as well as colonization in the host tissues. This work will be useful to develop methodologies for understanding the mechanisms of Bipolaris-wheat interaction and functional genomics of B. sorokiniana for various applications including insertional mutagenesis, targeted disruption of specific genes, ectopic complementation of loss-of-function strains and over-expression.

  20. Low-molecular-mass thiol compounds from a free-living highly pathogenic amoeba, Naegleria fowleri.

    PubMed

    Ondarza, Raúl N; Iturbe, Angélica; Hernández, Eva; Hurtado, Gerardo

    2003-04-01

    Acid extracts labelled with the fluorescent reagent monobromobimane and separated by HPLC have enabled the detection of low-molecular-mass thiol compounds in Naegleria fowleri for the first time. The amounts detected are expressed in nmol/1 x 10(6) trophozoites cultivated at various stages of growth in the appropriate culture medium. N. fowleri is a highly pathogenic free-living amoeba, in which we found important thiol compounds, some of them in their reduced and oxidized forms. Unlike cysteine and glutathione, a number of these are not represented in normal human lymphocytes. Some of these thiol compounds from Naegleria must have their respective disulphide reductases, although the presence of thiol-disulphide exchange reactions must be considered. Ovothiol A, with antioxidant properties, is an example of a compound that is kept reduced by trypanothione in trypanosomatids, although no disulphide reductase for ovothiol A has yet been discovered. In our case we were unable to detect this biothiol in Naegleria. The presence of thiol compounds that seem to be particular to this pathogen and which are not present in human lymphocytes opens the possibility of searching for disulphide-reducing enzymes that can serve as drug targets.

  1. Pathogenicity of Genetically Similar, H5N1 Highly Pathogenic Avian Influenza Virus Strains in Chicken and the Differences in Sensitivity among Different Chicken Breeds

    PubMed Central

    Matsuu, Aya; Kobayashi, Tomoko; Patchimasiri, Tuangthong; Shiina, Takashi; Suzuki, Shingo; Chaichoune, Kridsada; Ratanakorn, Parntep; Hiromoto, Yasuaki; Abe, Haruka; Parchariyanon, Sujira; Saito, Takehiko

    2016-01-01

    Differences in the pathogenicity of genetically closely related H5N1 highly pathogenic avian influenza viruses (HPAIVs) were evaluated in White Leghorn chickens. These viruses varied in the clinical symptoms they induced, including lethality, virus shedding, and replication in host tissues. A comparison of the host responses in the lung, brain, and spleen suggested that the differences in viral replication efficiency were related to the host cytokine response at the early phase of infection, especially variations in the proinflammatory cytokine IL-6. Based on these findings, we inoculated the virus that showed the mildest pathogenicity among the five tested, A/pigeon/Thailand/VSMU-7-NPT/2004, into four breeds of Thai indigenous chicken, Phadu-Hung-Dang (PHD), Chee, Dang, and Luang-Hung-Khao (LHK), to explore effects of genetic background on host response. Among these breeds, Chee, Dang, and LHK showed significantly longer survival times than White Leghorns. Virus shedding from dead Thai indigenous chickens was significantly lower than that from White Leghorns. Although polymorphisms were observed in the Mx and MHC class I genes, there was no significant association between the polymorphisms in these loci and resistance to HPAIV. PMID:27078641

  2. Pathogenicity of Genetically Similar, H5N1 Highly Pathogenic Avian Influenza Virus Strains in Chicken and the Differences in Sensitivity among Different Chicken Breeds.

    PubMed

    Matsuu, Aya; Kobayashi, Tomoko; Patchimasiri, Tuangthong; Shiina, Takashi; Suzuki, Shingo; Chaichoune, Kridsada; Ratanakorn, Parntep; Hiromoto, Yasuaki; Abe, Haruka; Parchariyanon, Sujira; Saito, Takehiko

    2016-01-01

    Differences in the pathogenicity of genetically closely related H5N1 highly pathogenic avian influenza viruses (HPAIVs) were evaluated in White Leghorn chickens. These viruses varied in the clinical symptoms they induced, including lethality, virus shedding, and replication in host tissues. A comparison of the host responses in the lung, brain, and spleen suggested that the differences in viral replication efficiency were related to the host cytokine response at the early phase of infection, especially variations in the proinflammatory cytokine IL-6. Based on these findings, we inoculated the virus that showed the mildest pathogenicity among the five tested, A/pigeon/Thailand/VSMU-7-NPT/2004, into four breeds of Thai indigenous chicken, Phadu-Hung-Dang (PHD), Chee, Dang, and Luang-Hung-Khao (LHK), to explore effects of genetic background on host response. Among these breeds, Chee, Dang, and LHK showed significantly longer survival times than White Leghorns. Virus shedding from dead Thai indigenous chickens was significantly lower than that from White Leghorns. Although polymorphisms were observed in the Mx and MHC class I genes, there was no significant association between the polymorphisms in these loci and resistance to HPAIV.

  3. Airborne Transmission of Highly Pathogenic H7N1 Influenza Virus in Ferrets

    PubMed Central

    Finch, Courtney; Shao, Hongxia; Angel, Matthew; Chen, Hongjun; Capua, Ilaria; Cattoli, Giovanni; Monne, Isabella

    2014-01-01

    ABSTRACT Avian H7 influenza viruses are recognized as potential pandemic viruses, as personnel often become infected during poultry outbreaks. H7 infections in humans typically cause mild conjunctivitis; however, the H7N9 outbreak in the spring of 2013 has resulted in severe respiratory disease. To date, no H7 viruses have acquired the ability for sustained transmission among humans. Airborne transmission is considered a requirement for the emergence of pandemic influenza, and advanced knowledge of the molecular changes or signature required for transmission would allow early identification of pandemic vaccine seed stocks, screening and stockpiling of antiviral compounds, and eradication efforts focused on flocks harboring threatening viruses. Thus, we sought to determine if a highly pathogenic influenza A H7N1 (A/H7N1) virus with no history of human infection could become capable of airborne transmission among ferrets. We show that after 10 serial passages, A/H7N1 developed the ability to be transmitted to cohoused and airborne contact ferrets. Four amino acid mutations (PB2 T81I, NP V284M, and M1 R95K and Q211K) in the internal genes and a minimal amino acid mutation (K/R313R) in the stalk region of the hemagglutinin protein were associated with airborne transmission. Furthermore, transmission was not associated with loss of virulence. These findings highlight the importance of the internal genes in host adaptation and suggest that natural isolates carrying these mutations be further evaluated. Our results demonstrate that a highly pathogenic avian H7 virus can become capable of airborne transmission in a mammalian host, and they support ongoing surveillance and pandemic H7 vaccine development. IMPORTANCE The major findings of this report are that a highly pathogenic strain of H7N1 avian influenza virus can be adapted to become capable of airborne transmission in mammals without mutations altering receptor specificity. Changes in receptor specificity have been

  4. Controlling highly pathogenic avian influenza outbreaks: An epidemiological and economic model analysis.

    PubMed

    Backer, J A; van Roermund, H J W; Fischer, E A J; van Asseldonk, M A P M; Bergevoet, R H M

    2015-09-01

    Outbreaks of highly pathogenic avian influenza (HPAI) can cause large losses for the poultry sector and for animal disease controlling authorities, as well as risks for animal and human welfare. In the current simulation approach epidemiological and economic models are combined to compare different strategies to control highly pathogenic avian influenza in Dutch poultry flocks. Evaluated control strategies are the minimum EU strategy (i.e., culling of infected flocks, transport regulations, tracing and screening of contact flocks, establishment of protection and surveillance zones), and additional control strategies comprising pre-emptive culling of all susceptible poultry flocks in an area around infected flocks (1 km, 3 km and 10 km) and emergency vaccination of all flocks except broilers around infected flocks (3 km). Simulation results indicate that the EU strategy is not sufficient to eradicate an epidemic in high density poultry areas. From an epidemiological point of view, this strategy is the least effective, while pre-emptive culling in 10 km radius is the most effective of the studied strategies. But these two strategies incur the highest costs due to long duration (EU strategy) and large-scale culling (pre-emptive culling in 10 km radius). Other analysed pre-emptive culling strategies (i.e., in 1 km and 3 km radius) are more effective than the analysed emergency vaccination strategy (in 3 km radius) in terms of duration and size of the epidemics, despite the assumed optimistic vaccination capacity of 20 farms per day. However, the total costs of these strategies differ only marginally. Extending the capacity for culling substantially reduces the duration, size and costs of the epidemic. This study demonstrates the strength of combining epidemiological and economic model analysis to gain insight in a range of consequences and thus to serve as a decision support tool in the control of HPAI epidemics.

  5. High antiviral effects of hibiscus tea extract on the H5 subtypes of low and highly pathogenic avian influenza viruses

    PubMed Central

    BAATARTSOGT, Tugsbaatar; BUI, Vuong N.; TRINH, Dai Q.; YAMAGUCHI, Emi; GRONSANG, Dulyatad; THAMPAISARN, Rapeewan; OGAWA, Haruko; IMAI, Kunitoshi

    2016-01-01

    Viral neuraminidase inhibitors are widely used as synthetic anti-influenza drugs for the prevention and treatment of influenza. However, drug-resistant influenza A virus variants, including H5N1 highly pathogenic avian influenza viruses (HPAIVs), have been reported. Therefore, the discovery of novel and effective antiviral agents is warranted. We screened the antiviral effects of 11 herbal tea extracts (hibiscus, black tea, tencha, rosehip tea, burdock tea, green tea, jasmine tea, ginger tea, lavender tea, rose tea and oak tea) against the H5N1 HPAIV in vitro. Among the tested extracts, only the hibiscus extract and its fractionated extract (frHibis) highly and rapidly reduced the titers of all H5 HPAIVs and low pathogenic AIVs (LPAIVs) used in the pre-treatment tests of Madin–Darby canine kidney (MDCK) cells that were inoculated with a mixture of the virus and the extract. Immunogold electron microscopy showed that anti-H5 monoclonal antibodies could not bind to the deformed H5 virus particles pretreated with frHibis. In post-treatment tests of MDCK cells cultured in the presence of frHibis after infection with H5N1 HPAIV, the frHibis inhibited viral replication and the expression of viral antigens and genes. Among the plants tested, hibiscus showed the most prominent antiviral effects against both H5 HPAIV and LPAIV. PMID:27193820

  6. Immune responses of locusts to challenge with the pathogenic fungus Metarhizium or high doses of laminarin.

    PubMed

    Mullen, Lisa M; Goldsworthy, Graham J

    2006-04-01

    Two isolates of Metarhizium anisopliae var acridum were tested for their effects on the locust immune system and for comparison with the effects of challenge by injection with laminarin. Isolate IMI 330189 (referred to hereafter as Met 189) is highly pathogenic whether applied topically as conidia or injected as blastospores. However, isolate ARSEF 728 (referred to hereafter as Met 728) is pathogenic only when injected as blastospores, suggesting that the lack of pathogenicity of topically applied conidia from this isolate is due to a failure to penetrate the insect cuticle and gain access to the haemocoel. After topical application of conidia from Met 189, no activation of prophenoloxidase is detected, but injection of blastospores from Met 189 brings about a transient increase in phenoloxidase activity in the haemolymph in both adult locusts and 5th instar nymphs, although this does not prevent fungal-induced mortality. Co-injection of adipokinetic hormone-I (AKH-I) with blastospores prolongs the activation of prophenoloxidase in the haemolymph of adult locusts, and enhances it in nymphs. It is argued that the lack of activation of prophenoloxidase in nymphs shown previously (Mullen, L., Goldsworthy, G., 2003. Changes in lipophorins are related to the activation of phenoloxidase in the haemolymph of Locusta migratoria in response to injection of immunogens. Insect Biochemistry and Molecular Biology 33, 661-670), reflects differences in the sensitivity of the immune system between adults and nymphs rather than distinct qualitative differences, and this is confirmed in this study by the demonstration that doses of laminarin higher than those used previously (>or=100 microg) do activate the prophenoloxidase cascade in 5th instar nymphs. Nodules are formed in locusts of all ages in response to fungal infection or injection of laminarin, although there is wide variation in the number, size and distribution of nodules formed. During the examination of 5th instar nymphs

  7. Economic issues in vaccination against highly pathogenic avian influenza in developing countries.

    PubMed

    McLeod, A; Rushton, J; Riviere-Cinnamond, A; Brandenburg, B; Hinrichs, J; Loth, L

    2007-01-01

    We consider the use of vaccination against highly pathogenic avian influenza (HPAI) in three contexts: as part of a stamping-out programme, as a government-led action for disease prevention and as private insurance by farmers. Poultry systems in developing countries cover all four of the poultry sectors defined by FAO and the OIE, each with particular economic aspects that might motivate farmers to take part in vaccination programmes or to initiate and finance them. Outbreaks in flocks of different types have different potential impacts in terms of disease spread and economic effects, which influence the potential benefits of vaccination as a means to prevent or control outbreaks. We use data from three countries to illustrate the costs of vaccination and discuss measures of cost-effectiveness and ways to improve it. We also consider the question of funding sources and their impact on the sustainability of vaccination programmes.

  8. Fluorescent Protein Nanowire-Mediated Protein Microarrays for Multiplexed and Highly Sensitive Pathogen Detection.

    PubMed

    Men, Dong; Zhou, Juan; Li, Wei; Leng, Yan; Chen, Xinwen; Tao, Shengce; Zhang, Xian-En

    2016-07-13

    Protein microarrays are powerful tools for high-throughput and simultaneous detection of different target molecules in complex biological samples. However, the sensitivity of conventional fluorescence-labeling protein detection methods is limited by the availability of signal molecules for binding to the target molecule. Here, we built a multifunctional fluorescent protein nanowire (FNw) by harnessing self-assembly of yeast amyloid protein. The FNw integrated a large number of fluorescent molecules, thereby enhancing the fluorescent signal output in target detection. The FNw was then combined with protein microarray technology to detect proteins derived from two pathogens, including influenza virus (hemagglutinin 1, HA1) and human immunodeficiency virus (p24 and gp120). The resulting detection sensitivity achieved a 100-fold improvement over a commercially available detection reagent. PMID:27315221

  9. Innate immune response to arenaviral infection: a focus on the highly pathogenic New World hemorrhagic arenaviruses

    PubMed Central

    Koma, Takaaki; Huang, Cheng; Kolokoltsova, Olga A; Brasier, Allan R; Paessler, Slobodan

    2013-01-01

    Arenaviruses are enveloped, negative-stranded RNA viruses that belong to the family Arenaviridae. This diverse family can be further classified into OW (Old World) and NW (New World) arenaviruses based on their antigenicity, phylogeny, and geographical distribution. Many of the NW arenaviruses are highly pathogenic viruses that cause systemic human infections characterized by hemorrhagic fever and/or neurological manifestations, constituting public health problems in their endemic regions. NW arenavirus infection induces a variety of host innate immune responses, which could contribute to the viral pathogenesis and/or influence the final outcome of virus infection in vitro as well as in vivo. On the other hand, NW arenaviruses have also developed several strategies to counteract the host innate immune response. We will review current knowledge regarding the interplay between the host innate immune response and NW arenavirus infection in vitro and in vivo, with emphasis on viral-encoded proteins and their effect on the type I interferon response. PMID:24075870

  10. Modeling highly pathogenic avian influenza transmission in wild birds and poultry in West Bengal, India

    PubMed Central

    Pandit, Pranav S.; Bunn, David A.; Pande, Satish A.; Aly, Sharif S.

    2013-01-01

    Wild birds are suspected to have played a role in highly pathogenic avian influenza (HPAI) H5N1 outbreaks in West Bengal. Cluster analysis showed that H5N1 was introduced in West Bengal at least 3 times between 2008 and 2010. We simulated the introduction of H5N1 by wild birds and their contact with poultry through a stochastic continuous-time mathematical model. Results showed that reducing contact between wild birds and domestic poultry, and increasing the culling rate of infected domestic poultry communities will reduce the probability of outbreaks. Poultry communities that shared habitat with wild birds or those indistricts with previous outbreaks were more likely to suffer an outbreak. These results indicate that wild birds can introduce HPAI to domestic poultry and that limiting their contact at shared habitats together with swift culling of infected domestic poultry can greatly reduce the likelihood of HPAI outbreaks. PMID:23846233

  11. High-antibacterial activity of Urtica spp. seed extracts on food and plant pathogenic bacteria.

    PubMed

    Körpe, Didem Aksoy; İşerı, Özlem Darcansoy; Sahin, Feride Iffet; Cabi, Evren; Haberal, Mehmet

    2013-05-01

    The aim of this study was to comparatively evaluate antibacterial activities of methanol (MetOH) and aqueous (dw) leaf (L), root (R) and seed (S) extracts of Urtica dioica L. (Ud; stinging nettle) and Urtica pilulifera L. (Up; Roman nettle) on both food- and plant-borne pathogens, with total phenolic contents and DPPH radical scavenging activities (DRSA). MetOH extracts of leaves and roots of U. dioica had the highest DRSA. Extracts with high antibacterial activity were in the order Up-LMetOH (13/16) > Ud-SMetOH (11/16) > Up-SMetOH (9/16). Results obtained with Up-SMetOH against food spoiling Bacillus pumilus, Shigella spp. and Enterococcus gallinarum with minimum inhibitory concentrations (MICs) in 128-1024 μg/ml range seem to be promising. Up-SMetOH also exerted strong inhibition against Clavibacter michiganensis with a considerably low MIC (32 μg/ml). Ud-SMetOH and Up-LMetOH were also effective against C. michiganensis (MIC = 256 and 1024 μg/ml, respectively). Ud-SMetOH and Ud-RMetOH had also antimicrobial activity against Xanthomonas vesicatoria (MIC = 512 and 1024 μg/ml, respectively). Results presented here demonstrate high-antibacterial activity of U. pilulifera extracts and U. dioica seed extract against phytopathogens for the first time, and provide the most comprehensive data on the antibacterial activity screening of U. pilulifera against food-borne pathogens. Considering limitations in plant disease control, antibacterial activities of these extracts would be of agricultural importance.

  12. Continued evolution of highly pathogenic avian influenza A (H5N1): updated nomenclature

    PubMed Central

    2011-01-01

    Please cite this paper as: WHO/OIE/FAO. (2012) Continued evolution of highly pathogenic avian influenza A(H5N1): Updated nomenclature. Influenza and Other Respiratory Viruses 6(1), 1–5. Background  Continued evolution of highly pathogenic avian influenza A (H5N1) throughout many regions of the eastern hemisphere has led to the emergence of new phylogenetic groups. A total of 1637 new H5N1 hemagglutinin (HA) sequences have become available since the previous nomenclature recommendations described in 2009 by the WHO/OIE/FAO H5N1 Evolution Working Group. A comprehensive analysis including all the new data is needed to update HA clade nomenclature. Methods  Phylogenetic trees were constructed from data sets of all available H5N1 HA sequences. New clades were designated on the basis of phylogeny and p‐distance using the pre‐established nomenclature system (Emerg Infec Dis 2008; 14:e1). Each circulating H5N1 clade was subjected to further phylogenetic analysis and nucleotide sequence divergence calculations. Results  All recently circulating clades (clade 1 in the Mekong River Delta, 2.1.3 in Indonesia, 2.2 in India/Bangladesh, 2.2.1 in Egypt, 2.3.2, 2.3.4 and 7 in Asia) required assignment of divergent HA genes to new second‐, third‐, and/or fourth‐order clades. At the same time, clades 0, 3, 4, 5, 6, 8, 9, and several second‐ and third‐order groups from clade 2 have not been detected since 2008 or earlier. Conclusions  New designations are recommended for 12 HA clades, named according to previously defined criteria. In addition, viruses from 13 clades have not been detected since 2008 or earlier. The periodic updating of this dynamic classification system allows continued use of a unified nomenclature in all H5N1 studies. PMID:22035148

  13. High-antibacterial activity of Urtica spp. seed extracts on food and plant pathogenic bacteria.

    PubMed

    Körpe, Didem Aksoy; İşerı, Özlem Darcansoy; Sahin, Feride Iffet; Cabi, Evren; Haberal, Mehmet

    2013-05-01

    The aim of this study was to comparatively evaluate antibacterial activities of methanol (MetOH) and aqueous (dw) leaf (L), root (R) and seed (S) extracts of Urtica dioica L. (Ud; stinging nettle) and Urtica pilulifera L. (Up; Roman nettle) on both food- and plant-borne pathogens, with total phenolic contents and DPPH radical scavenging activities (DRSA). MetOH extracts of leaves and roots of U. dioica had the highest DRSA. Extracts with high antibacterial activity were in the order Up-LMetOH (13/16) > Ud-SMetOH (11/16) > Up-SMetOH (9/16). Results obtained with Up-SMetOH against food spoiling Bacillus pumilus, Shigella spp. and Enterococcus gallinarum with minimum inhibitory concentrations (MICs) in 128-1024 μg/ml range seem to be promising. Up-SMetOH also exerted strong inhibition against Clavibacter michiganensis with a considerably low MIC (32 μg/ml). Ud-SMetOH and Up-LMetOH were also effective against C. michiganensis (MIC = 256 and 1024 μg/ml, respectively). Ud-SMetOH and Ud-RMetOH had also antimicrobial activity against Xanthomonas vesicatoria (MIC = 512 and 1024 μg/ml, respectively). Results presented here demonstrate high-antibacterial activity of U. pilulifera extracts and U. dioica seed extract against phytopathogens for the first time, and provide the most comprehensive data on the antibacterial activity screening of U. pilulifera against food-borne pathogens. Considering limitations in plant disease control, antibacterial activities of these extracts would be of agricultural importance. PMID:23067263

  14. Novel Reassortant H5N6 Influenza A Virus from the Lao People's Democratic Republic Is Highly Pathogenic in Chickens.

    PubMed

    Butler, Jeffrey; Stewart, Cameron R; Layton, Daniel S; Phommachanh, Phouvong; Harper, Jennifer; Payne, Jean; Evans, Ryan M; Valdeter, Stacey; Walker, Som; Harvey, Gemma; Shan, Songhua; Bruce, Matthew P; Rootes, Christina L; Gough, Tamara J; Rohringer, Andreas; Peck, Grantley R; Fardy, Sarah J; Karpala, Adam J; Johnson, Dayna; Wang, Jianning; Douangngeun, Bounlom; Morrissy, Christopher; Wong, Frank Y K; Bean, Andrew G D; Bingham, John; Williams, David T

    2016-01-01

    Avian influenza viruses of H5 subtype can cause highly pathogenic disease in poultry. In March 2014, a new reassortant H5N6 subtype highly pathogenic avian influenza virus emerged in Lao People's Democratic Republic. We have assessed the pathogenicity, pathobiology and immunological responses associated with this virus in chickens. Infection caused moderate to advanced disease in 6 of 6 chickens within 48 h of mucosal inoculation. High virus titers were observed in blood and tissues (kidney, spleen, liver, duodenum, heart, brain and lung) taken at euthanasia. Viral antigen was detected in endothelium, neurons, myocardium, lymphoid tissues and other cell types. Pro-inflammatory cytokines were elevated compared to non-infected birds. Our study confirmed that this new H5N6 reassortant is highly pathogenic, causing disease in chickens similar to that of Asian H5N1 viruses, and demonstrated the ability of such clade 2.3.4-origin H5 viruses to reassort with non-N1 subtype viruses while maintaining a fit and infectious phenotype. Recent detection of influenza H5N6 poultry infections in Lao PDR, China and Viet Nam, as well as six fatal human infections in China, demonstrate that these emergent highly pathogenic H5N6 viruses may be widely established in several countries and represent an emerging threat to poultry and human populations.

  15. Novel Reassortant H5N6 Influenza A Virus from the Lao People's Democratic Republic Is Highly Pathogenic in Chickens.

    PubMed

    Butler, Jeffrey; Stewart, Cameron R; Layton, Daniel S; Phommachanh, Phouvong; Harper, Jennifer; Payne, Jean; Evans, Ryan M; Valdeter, Stacey; Walker, Som; Harvey, Gemma; Shan, Songhua; Bruce, Matthew P; Rootes, Christina L; Gough, Tamara J; Rohringer, Andreas; Peck, Grantley R; Fardy, Sarah J; Karpala, Adam J; Johnson, Dayna; Wang, Jianning; Douangngeun, Bounlom; Morrissy, Christopher; Wong, Frank Y K; Bean, Andrew G D; Bingham, John; Williams, David T

    2016-01-01

    Avian influenza viruses of H5 subtype can cause highly pathogenic disease in poultry. In March 2014, a new reassortant H5N6 subtype highly pathogenic avian influenza virus emerged in Lao People's Democratic Republic. We have assessed the pathogenicity, pathobiology and immunological responses associated with this virus in chickens. Infection caused moderate to advanced disease in 6 of 6 chickens within 48 h of mucosal inoculation. High virus titers were observed in blood and tissues (kidney, spleen, liver, duodenum, heart, brain and lung) taken at euthanasia. Viral antigen was detected in endothelium, neurons, myocardium, lymphoid tissues and other cell types. Pro-inflammatory cytokines were elevated compared to non-infected birds. Our study confirmed that this new H5N6 reassortant is highly pathogenic, causing disease in chickens similar to that of Asian H5N1 viruses, and demonstrated the ability of such clade 2.3.4-origin H5 viruses to reassort with non-N1 subtype viruses while maintaining a fit and infectious phenotype. Recent detection of influenza H5N6 poultry infections in Lao PDR, China and Viet Nam, as well as six fatal human infections in China, demonstrate that these emergent highly pathogenic H5N6 viruses may be widely established in several countries and represent an emerging threat to poultry and human populations. PMID:27631618

  16. Inactivation of foodborne pathogens in ground beef by cooking with highly controlled radio frequency energy.

    PubMed

    Schlisselberg, Dov B; Kler, Edna; Kalily, Emmanuel; Kisluk, Guy; Karniel, Ohad; Yaron, Sima

    2013-01-01

    The consumer demand for fresh tasting, high quality, low salt, preservative-free meals which require minimal preparation time magnifies the safety concern and emphasizes the need to use innovative technologies for food processing. A modern technique to uniformly heat and cook foods is based on a combination of convection and controlled radio frequency (RF) energy. However any advantage conferred on meat cooked by this method would be lost if application of the technology results in decreased safety. Our main goal was to study the inactivation efficacy of this method of cooking against pathogens in ground meat in comparison to standard convection cooking. Meat balls were artificially inoculated with GFP expressing Escherichia coli, Salmonella Typhimurium and Listeria monocytogenes as well as spores of Bacillus cereus and Bacillus thuringiensis and cooked by convection heating (220°C, 40 min), by using energy generated from frequencies in the RF bandwidth (RF cooking, 7.5 min) or by combined heating (5.5 min), until the center temperature of each sample reached 73°C. The mean reductions in total indigenous bacteria obtained by RF and convection were 2.8 and 2.5 log CFU/g, respectively. Cooking of meat balls with convection reduced the E. coli population (8 log CFU/g) by 5.5 log CFU/g, whilst treatment with RF reduced E. coli population to undetectable levels. The mean reductions of S. Typhimurium obtained by RF and convection were 5.7 and 6.5 log CFU/g, respectively. The combined treatment reduced the Salmonella population to undetectable levels. In contrast, L. monocytogenes was poorly affected by RF cooking. The mean reduction of L. monocytogenes obtained by RF energy was 0.4 log CFU/g, while convection cooking resulted in undetectable levels. Interestingly, the combined treatment also resulted with undetectable levels of Listeria although time of cooking was reduced by 86%. One-step cooking had negligible effects on the Bacillus spores and therefore a 2-step

  17. Benefits of a European Project on Diagnostics of Highly Pathogenic Agents and Assessment of Potential “Dual Use” Issues

    PubMed Central

    Grunow, Roland; Ippolito, G.; Jacob, D.; Sauer, U.; Rohleder, A.; Di Caro, A.; Iacovino, R.

    2014-01-01

    Quality assurance exercises and networking on the detection of highly infectious pathogens (QUANDHIP) is a joint action initiative set up in 2011 that has successfully unified the primary objectives of the European Network on Highly Pathogenic Bacteria (ENHPB) and of P4-laboratories (ENP4-Lab) both of which aimed to improve the efficiency, effectiveness, and response capabilities of laboratories directed at protecting the health of European citizens against high consequence bacteria and viruses of significant public health concern. Both networks have established a common collaborative consortium of 37 nationally and internationally recognized institutions with laboratory facilities from 22 European countries. The specific objectives and achievements include the initiation and establishment of a recognized and acceptable quality assurance scheme, including practical external quality assurance exercises, comprising living agents, that aims to improve laboratory performance, accuracy, and detection capabilities in support of patient management and public health responses; recognized training schemes for diagnostics and handling of highly pathogenic agents; international repositories comprising highly pathogenic bacteria and viruses for the development of standardized reference material; a standardized and transparent Biosafety and Biosecurity strategy protecting healthcare personnel and the community in dealing with high consequence pathogens; the design and organization of response capabilities dealing with cross-border events with highly infectious pathogens including the consideration of diagnostic capabilities of individual European laboratories. The project tackled several sensitive issues regarding Biosafety, Biosecurity and “dual use” concerns. The article will give an overview of the project outcomes and discuss the assessment of potential “dual use” issues. PMID:25426479

  18. Genetic Characterization of Highly Pathogenic Avian Influenza (H5N8) Virus from Domestic Ducks, England, November 2014.

    PubMed

    Hanna, Amanda; Banks, Jill; Marston, Denise A; Ellis, Richard J; Brookes, Sharon M; Brown, Ian H

    2015-05-01

    Genetic sequences of a highly pathogenic avian influenza (H5N8) virus in England have high homology to those detected in mainland Europe and Asia during 2014. Genetic characterization suggests this virus is an avian-adapted virus without specific affinity for zoonoses. Spatio-temporal detections of H5N8 imply a role for wild birds in virus spread.

  19. High Pathogenicity of Wild-Type Measles Virus Infection in CD150 (SLAM) Transgenic Mice

    PubMed Central

    Sellin, Caroline I.; Davoust, Nathalie; Guillaume, Vanessa; Baas, Dominique; Belin, Marie-Françoise; Buckland, Robin; Wild, T. Fabian; Horvat, Branka

    2006-01-01

    Measles virus (MV) infection causes an acute childhood disease, associated in certain cases with infection of the central nervous system and development of a severe neurological disease. We have generated transgenic mice ubiquitously expressing the human protein SLAM (signaling lymphocytic activation molecule), or CD150, recently identified as an MV receptor. In contrast to all other MV receptor transgenic models described so far, in these mice infection with wild-type MV strains is highly pathogenic. Intranasal infection of SLAM transgenic suckling mice leads to MV spread to different organs and the development of an acute neurological syndrome, characterized by lethargy, seizures, ataxia, weight loss, and death within 3 weeks. In addition, in this model, vaccine and wild-type MV strains can be distinguished by virulence. Furthermore, intracranial MV infection of adult transgenic mice generates a subclinical infection associated with a high titer of MV-specific antibodies in the serum. Finally, to analyze new antimeasles therapeutic approaches, we created a recombinant soluble form of SLAM and demonstrated its important antiviral activity both in vitro and in vivo. Taken together, our results show the high susceptibility of SLAM transgenic mice to MV-induced neurological disease and open new perspectives for the analysis of the implication of SLAM in the neuropathogenicity of other morbilliviruses, which also use this molecule as a receptor. Moreover, this transgenic model, in allowing a simple readout of the efficacy of an antiviral treatment, provides unique experimental means to test novel anti-MV preventive and therapeutic strategies. PMID:16775330

  20. Mastitis Pathogens with High Virulence in a Mouse Model Produce a Distinct Cytokine Profile In Vivo

    PubMed Central

    Johnzon, Carl-Fredrik; Artursson, Karin; Söderlund, Robert; Guss, Bengt; Rönnberg, Elin; Pejler, Gunnar

    2016-01-01

    Mastitis is a serious medical condition of dairy cattle. Here, we evaluated whether the degree of virulence of mastitis pathogens in a mouse model can be linked to the inflammatory response that they provoke. Clinical isolates of Staphylococcus aureus (S. aureus) (strain 556 and 392) and Escherichia coli (E. coli) (676 and 127), and laboratory control strains [8325-4 (S. aureus) and MG1655 (E. coli)], were injected i.p. into mice, followed by the assessment of clinical scores and inflammatory parameters. As judged by clinical scoring, E. coli 127 exhibited the largest degree of virulence among the strains. All bacterial strains induced neutrophil recruitment. However, whereas E. coli 127 induced high peritoneal levels of CXCL1, G-CSF, and CCL2, strikingly lower levels of these were induced by the less virulent bacterial strains. High concentrations of these compounds were also seen in blood samples taken from animals infected with E. coli 127, suggesting systemic inflammation. Moreover, the levels of CXCL1 and G-CSF, both in the peritoneal fluid and in plasma, correlated with clinical score. Together, these findings suggest that highly virulent clinical mastitis isolates produce a distinct cytokine profile that shows a close correlation with the severity of the bacterial infection. PMID:27713743

  1. Characterizing ncRNAs in Human Pathogenic Protists Using High-Throughput Sequencing Technology

    PubMed Central

    Collins, Lesley Joan

    2011-01-01

    ncRNAs are key genes in many human diseases including cancer and viral infection, as well as providing critical functions in pathogenic organisms such as fungi, bacteria, viruses, and protists. Until now the identification and characterization of ncRNAs associated with disease has been slow or inaccurate requiring many years of testing to understand complicated RNA and protein gene relationships. High-throughput sequencing now offers the opportunity to characterize miRNAs, siRNAs, small nucleolar RNAs (snoRNAs), and long ncRNAs on a genomic scale, making it faster and easier to clarify how these ncRNAs contribute to the disease state. However, this technology is still relatively new, and ncRNA discovery is not an application of high priority for streamlined bioinformatics. Here we summarize background concepts and practical approaches for ncRNA analysis using high-throughput sequencing, and how it relates to understanding human disease. As a case study, we focus on the parasitic protists Giardia lamblia and Trichomonas vaginalis, where large evolutionary distance has meant difficulties in comparing ncRNAs with those from model eukaryotes. A combination of biological, computational, and sequencing approaches has enabled easier classification of ncRNA classes such as snoRNAs, but has also aided the identification of novel classes. It is hoped that a higher level of understanding of ncRNA expression and interaction may aid in the development of less harsh treatment for protist-based diseases. PMID:22303390

  2. Characterizing ncRNAs in Human Pathogenic Protists Using High-Throughput Sequencing Technology.

    PubMed

    Collins, Lesley Joan

    2011-01-01

    ncRNAs are key genes in many human diseases including cancer and viral infection, as well as providing critical functions in pathogenic organisms such as fungi, bacteria, viruses, and protists. Until now the identification and characterization of ncRNAs associated with disease has been slow or inaccurate requiring many years of testing to understand complicated RNA and protein gene relationships. High-throughput sequencing now offers the opportunity to characterize miRNAs, siRNAs, small nucleolar RNAs (snoRNAs), and long ncRNAs on a genomic scale, making it faster and easier to clarify how these ncRNAs contribute to the disease state. However, this technology is still relatively new, and ncRNA discovery is not an application of high priority for streamlined bioinformatics. Here we summarize background concepts and practical approaches for ncRNA analysis using high-throughput sequencing, and how it relates to understanding human disease. As a case study, we focus on the parasitic protists Giardia lamblia and Trichomonas vaginalis, where large evolutionary distance has meant difficulties in comparing ncRNAs with those from model eukaryotes. A combination of biological, computational, and sequencing approaches has enabled easier classification of ncRNA classes such as snoRNAs, but has also aided the identification of novel classes. It is hoped that a higher level of understanding of ncRNA expression and interaction may aid in the development of less harsh treatment for protist-based diseases.

  3. High-throughput real-time electrochemical monitoring of LAMP for pathogenic bacteria detection.

    PubMed

    Safavieh, Mohammadali; Ahmed, Minhaz Uddin; Ng, Andy; Zourob, Mohammed

    2014-08-15

    One of the significant challenges in healthcare is the development of point-of-care (POC) diagnostics. POC diagnostics require low-cost devices that offer portability, simplicity in operation and the ability for high-throughput and quantitative analysis. Here, we present a novel roll-to-roll ribbon fluid-handling device for electrochemical real-time monitoring of nucleic acid (NA) amplification and bacteria detection. The device rendered loop-mediated isothermal amplification (LAMP) and real-time electrochemical detection based on the interaction between LAMP amplicon and the redox-reactive osmium complex. We have shown the detection of 30CFU/ml of Escherichia coli (in the range between 30 and 3×10(7)CFU/ml) and 200CFU/ml of Staphylococcus aureus (in the range of 200-2×10(5)CFU/ml) cultured samples in both real-time and end point detection. This device can be used for the detection of various Gram-negative and a number of Gram-positive bacterial pathogens with high sensitivity and specificity in a high-throughput format. Using a roll-to-roll cassette approach, we could detect 12 samples in one assay. Since the LAMP and electrochemical analysis are implemented within sealed flexible biochips, time-consuming processing steps are not required and the risk of contamination is significantly reduced.

  4. Effect of species, breed and route of virus inoculation on the pathogenicity of H5N1 highly pathogenic influenza (HPAI) viruses in domestic ducks

    PubMed Central

    2013-01-01

    H5N1 highly pathogenic avian influenza (HPAI) viruses continue to be a threat to poultry in many regions of the world. Domestic ducks have been recognized as one of the primary factors in the spread of H5N1 HPAI. In this study we examined the pathogenicity of H5N1 HPAI viruses in different species and breeds of domestic ducks and the effect of route of virus inoculation on the outcome of infection. We determined that the pathogenicity of H5N1 HPAI viruses varies between the two common farmed duck species, with Muscovy ducks (Cairina moschata) presenting more severe disease than various breeds of Anas platyrhynchos var. domestica ducks including Pekin, Mallard-type, Black Runners, Rouen, and Khaki Campbell ducks. We also found that Pekin and Muscovy ducks inoculated with two H5N1 HPAI viruses of different virulence, given by any one of three routes (intranasal, intracloacal, or intraocular), became infected with the viruses. Regardless of the route of inoculation, the outcome of infection was similar for each species but depended on the virulence of the virus used. Muscovy ducks showed more severe clinical signs and higher mortality than the Pekin ducks. In conclusion, domestic ducks are susceptible to H5N1 HPAI virus infection by different routes of exposure, but the presentation of the disease varied by virus strain and duck species. This information helps support the planning and implementation of H5N1 HPAI surveillance and control measures in countries with large domestic duck populations. PMID:23876184

  5. Effect of species, breed and route of virus inoculation on the pathogenicity of H5N1 highly pathogenic influenza (HPAI) viruses in domestic ducks.

    PubMed

    Pantin-Jackwood, Mary; Swayne, David E; Smith, Diane; Shepherd, Eric

    2013-07-22

    H5N1 highly pathogenic avian influenza (HPAI) viruses continue to be a threat to poultry in many regions of the world. Domestic ducks have been recognized as one of the primary factors in the spread of H5N1 HPAI. In this study we examined the pathogenicity of H5N1 HPAI viruses in different species and breeds of domestic ducks and the effect of route of virus inoculation on the outcome of infection. We determined that the pathogenicity of H5N1 HPAI viruses varies between the two common farmed duck species, with Muscovy ducks (Cairina moschata) presenting more severe disease than various breeds of Anas platyrhynchos var. domestica ducks including Pekin, Mallard-type, Black Runners, Rouen, and Khaki Campbell ducks. We also found that Pekin and Muscovy ducks inoculated with two H5N1 HPAI viruses of different virulence, given by any one of three routes (intranasal, intracloacal, or intraocular), became infected with the viruses. Regardless of the route of inoculation, the outcome of infection was similar for each species but depended on the virulence of the virus used. Muscovy ducks showed more severe clinical signs and higher mortality than the Pekin ducks. In conclusion, domestic ducks are susceptible to H5N1 HPAI virus infection by different routes of exposure, but the presentation of the disease varied by virus strain and duck species. This information helps support the planning and implementation of H5N1 HPAI surveillance and control measures in countries with large domestic duck populations.

  6. Importation and Recombination Are Responsible for the Latest Emergence of Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus in China

    PubMed Central

    Zhao, Kuan; Ye, Chao; Chang, Xiao-Bo; Jiang, Cheng-Gang; Wang, Shu-Jie; Cai, Xue-Hui; Tong, Guang-Zhi; Tian, Zhi-Jun

    2015-01-01

    In China, a majority of the highly pathogenic porcine reproductive and respiratory syndrome (HP-PRRSV) strains were seeded by the 2006 outbreak. However, the most recently emerged (2013-2014) HP-PRRSV strain has a very different genetic background. It is a NADC30-like PRRSV strain recently introduced from North America that has undergone genetic exchange with the classic HP-PRRSV strains in China. Subsequent isolation and characterization of this variant suggest high pathogenicity, so it merits special attention in control and vaccine strategies. PMID:26246582

  7. Development of a consensus microarray method for identification of some highly pathogenic viruses.

    PubMed

    Xiao-Ping, Kang; Yong-Qiang, Li; Qing-Ge, Sun; Hong, Liu; Qing-Yu, Zhu; Yin-Hui, Yang

    2009-11-01

    Some highly pathogenic viruses, such as Chikungunya virus, Japanese encephalitis virus, Yellow fever virus, Dengue virus, Hanta virus, SARS-CoV, and H5N1 avian influenza virus can cause severe infectious diseases. However, the consensus method for detecting these viruses has not been well established. A rapid and sensitive microarray approach for detection of these viruses and a panel of specific probes covering nine genera and 16 virus species were designed. 70-mer oligonucleotides were used at the genus level and 50-mer oligonucleotides were at the species level, respectively. To decrease the interference of the host genome in hybridization, the consensus genus primers were designed and used to reverse transcribe only virus genome. The synthesis of the second strand was carried out with a random primer sequence (5'-GTTTCCCAGTAGGTCTCNNNNNNNN-3'). The amplified products were labeled and processed for microarray analyses. This microarray-based method used the highly conserved consensus primers to synthesize specifically the virus cDNA and could identify effectively Chikungunya virus, Japanese encephalitis virus, Yellow fever virus, Dengue virus, Tick borne encephalitis virus, and H5N1 avian influenza virus. Using this method, one unknown virus isolated from pig brain in Shanxi Province, China was identified. This method may have an important potential application for the diagnosis of virus infection.

  8. Greater virulence of highly pathogenic H5N1 influenza virus in cats than in dogs.

    PubMed

    Kim, Heui Man; Park, Eun Hye; Yum, Jung; Kim, Hyun Soo; Seo, Sang Heui

    2015-01-01

    Highly pathogenic H5N1 influenza virus continues to infect animals and humans. We compared the infectivity and pathogenesis of H5N1 virus in domestic cats and dogs to find out which animal is more susceptible to H5N1 influenza virus. When cats and dogs were infected with the H5N1 virus, cats suffered from severe outcomes including death, whereas dogs did not show any mortality. Viruses were shed in the nose and rectum of cats and in the nose of dogs. Viruses were detected in brain, lung, kidney, intestine, liver, and serum in the infected cats, but only in the lung in the infected dogs. Genes encoding inflammatory cytokines and chemokines, Toll-like receptors, and apoptotic factors were more highly expressed in the lungs of cats than in those of dogs. Our results suggest that the intensive monitoring of dogs is necessary to prevent human infection by H5N1 influenza virus, since infected dogs may not show clear clinical signs, in contrast to infected cats.

  9. Distribution and dynamics of risk factors associated with highly pathogenic avian influenza H5N1

    PubMed Central

    Zhang, L.; Guo, Z. W.; Bridge, E. S.; Li, Y. M.; Xiao, X. M.

    2016-01-01

    SUMMARY Within China’s Poyang Lake region, close interactions between wild migratory birds and domestic poultry are common and provide an opportunity for the transmission and subsequent outbreaks of highly pathogenic avian influenza (HPAI) virus. We overlaid a series of ecological factors associated with HPAI to map the risk of HPAI in relation to natural and anthropogenic variables, and we identified two hotspots for potential HPAI outbreaks in the Poyang Lake region as well as three corridors connecting the two hotspot areas. In hotspot I, there is potential for migratory birds to bring new avian influenza (AI) strains that can reassort with existing strains to form new AI viruses. Hotspot II features high-density poultry production where outbreaks of endemic AI viruses are likely. The three communication corridors that link the two hotspots further promote HPAI H5N1 transmission and outbreaks and lead to the persistence of AI viruses in the Poyang Lake region. We speculate that the region’s unevenly distributed poultry supply-and-demand system might be a key factor inducing HPAI H5N1 transmission and outbreaks in the Poyang Lake region. PMID:23398949

  10. Multiple reassortment events among highly pathogenic avian influenza A(H5N1) viruses detected in Bangladesh.

    PubMed

    Gerloff, Nancy A; Khan, Salah Uddin; Balish, Amanda; Shanta, Ireen S; Simpson, Natosha; Berman, Lashondra; Haider, Najmul; Poh, Mee Kian; Islam, Ausraful; Gurley, Emily; Hasnat, Md Abdul; Dey, T; Shu, Bo; Emery, Shannon; Lindstrom, Stephen; Haque, Ainul; Klimov, Alexander; Villanueva, Julie; Rahman, Mahmudur; Azziz-Baumgartner, Eduardo; Ziaur Rahman, Md; Luby, Stephen P; Zeidner, Nord; Donis, Ruben O; Sturm-Ramirez, Katharine; Davis, C Todd

    2014-02-01

    In Bangladesh, little is known about the genomic composition and antigenicity of highly pathogenic avian influenza A(H5N1) viruses, their geographic distribution, temporal patterns, or gene flow within the avian host population. Forty highly pathogenic avian influenza A(H5N1) viruses isolated from humans and poultry in Bangladesh between 2008 and 2012 were analyzed by full genome sequencing and antigenic characterization. The analysis included viruses collected from avian hosts and environmental sampling in live bird markets, backyard poultry flocks, outbreak investigations in wild birds or poultry and from three human cases. Phylogenetic analysis indicated that the ancestors of these viruses reassorted (1) with other gene lineages of the same clade, (2) between different clades and (3) with low pathogenicity avian influenza A virus subtypes. Bayesian estimates of the time of most recent common ancestry, combined with geographic information, provided evidence of probable routes and timelines of virus spread into and out of Bangladesh.

  11. Multiplex and quantitative pathogen detection with high-resolution capillary electrophoresis-based single-strand conformation polymorphism.

    PubMed

    Hwang, Hee Sung; Shin, Gi Won; Chung, Boram; Na, Jeongkyeong; Jung, Gyoo Yeol

    2013-01-01

    Among the molecular diagnostic methods for bacteria-induced diseases, capillary electrophoresis-based single-strand conformation polymorphism (CE-SSCP) combined with 16S rRNA gene-specific PCR has enormous potential because it can separate sequence variants using a simple procedure. However, conventional CE-SSCP systems have limited resolution and cannot separate most 16S rRNA gene-specific markers into separate peaks. A high-resolution CE-SSCP system that uses a poly(ethyleneoxide)-poly(propyleneoxide)-poly(ethyleneoxide) triblock copolymer matrix was recently developed and shown to effectively separate highly similar PCR products. In this report, a protocol for the detection of 12 pathogenic bacteria is provided. Pathogen markers were amplified by PCR using universal primers and separated by CE-SSCP; each marker peak was well separated at baseline and showed a characteristic mobility, allowing the easy identification of the pathogens.

  12. The Use of High Pressure Freezing and Freeze Substitution to Study Host-Pathogen Interactions in Fungal Diseases of Plants

    NASA Astrophysics Data System (ADS)

    Mims, C. W.; Celio, Gail J.; Richardson, Elizabeth A.

    2003-12-01

    This article reports on the use of high pressure freezing followed by freeze substitution (HPF/FS) to study ultrastructural details of host pathogen interactions in fungal diseases of plants. The specific host pathogen systems discussed here include a powdery mildew infection of poinsettia and rust infections of daylily and Indian strawberry. The three pathogens considered here all attack the leaves of their hosts and produce specialized hyphal branches known as haustoria that invade individual host cells without killing them. We found that HPF/FS provided excellent preservation of both haustoria and host cells for all three host pathogen systems. Preservation of fungal and host cell membranes was particularly good and greatly facilitated the detailed study of host pathogen interfaces. In some instances, HPF/FS provided information that was not available in samples prepared for study using conventional chemical fixation. On the other hand, we did encounter various problems associated with the use of HPF/FS. Examples included freeze damage of samples, inconsistency of fixation in different samples, separation of plant cell cytoplasm from cell walls, breakage of cell walls and membranes, and splitting of thin sections. However, we believe that the outstanding preservation of ultrastructural details afforded by HPF/FS significantly outweighs these problems and we highly recommend the use of this fixation protocol for future studies of fungal host-plant interactions.

  13. The use of high pressure freezing and freeze substitution to study host-pathogen interactions in fungal diseases of plants.

    PubMed

    Mims, C W; Celio, Gail J; Richardson, Elizabeth A

    2003-12-01

    This article reports on the use of high pressure freezing followed by freeze substitution (HPF/FS) to study ultrastructural details of host-pathogen interactions in fungal diseases of plants. The specific host-pathogen systems discussed here include a powdery mildew infection of poinsettia and rust infections of daylily and Indian strawberry. The three pathogens considered here all attack the leaves of their hosts and produce specialized hyphal branches known as haustoria that invade individual host cells without killing them. We found that HPF/FS provided excellent preservation of both haustoria and host cells for all three host-pathogen systems. Preservation of fungal and host cell membranes was particularly good and greatly facilitated the detailed study of host-pathogen interfaces. In some instances, HPF/FS provided information that was not available in samples prepared for study using conventional chemical fixation. On the other hand, we did encounter various problems associated with the use of HPF/FS. Examples included freeze damage of samples, inconsistency of fixation in different samples, separation of plant cell cytoplasm from cell walls, breakage of cell walls and membranes, and splitting of thin sections. However, we believe that the outstanding preservation of ultrastructural details afforded by HPF/FS significantly outweighs these problems and we highly recommend the use of this fixation protocol for future studies of fungal host-plant interactions. PMID:14750987

  14. Unraveling the Architecture and Structural Dynamics of Pathogens by High-Resolution in vitro Atomic Force Microscopy

    SciTech Connect

    Malkin, A J; Plomp, M; Leighton, T J; McPherson, A; Wheeler, K E

    2005-04-12

    Progress in structural biology very much depends upon the development of new high-resolution techniques and tools. Despite decades of study of viruses, bacteria and bacterial spores and their pressing importance in human medicine and biodefense, many of their structural properties are poorly understood. Thus, characterization and understanding of the architecture of protein surface and internal structures of pathogens is critical to elucidating mechanisms of disease, immune response, physicochemical properties, environmental resistance and development of countermeasures against bioterrorist agents. Furthermore, even though complete genome sequences are available for various pathogens, the structure-function relationships are not understood. Because of their lack of symmetry and heterogeneity, large human pathogens are often refractory to X-ray crystallographic analysis or reconstruction by cryo-electron microscopy (cryo-EM). An alternative high-resolution method to examine native structure of pathogens is atomic force microscopy (AFM), which allows direct visualization of macromolecular assemblies at near-molecular resolution. The capability to image single pathogen surfaces at nanometer scale in vitro would profoundly impact mechanistic and structural studies of pathogenesis, immunobiology, specific cellular processes, environmental dynamics and biotransformation.

  15. Experimental infection of SPF and Korean native chickens with highly pathogenic avian influenza virus (H5N8).

    PubMed

    Lee, Eun-Kyoung; Song, Byung-Min; Kang, Hyun-Mi; Woo, Sang-Hee; Heo, Gyeong-Beom; Jung, Suk Chan; Park, Yong Ho; Lee, Youn-Jeong; Kim, Jae-Hong

    2016-05-01

    In 2014, an H5N8 outbreak of highly pathogenic avian influenza (HPAI) occurred in South Korea. The H5N8 strain produced mild to moderate clinical signs and mortality rates in commercial chicken farms, especially Korean native chicken farms. To understand the differences between their pathogenicity in SPF chicken and Korean native chicken., we evaluated the mean bird lethal doses (BLD50) of the Korean representative H5N8 virus (A/broiler duck/Korea/Buan2/2014) The BLD50values of the H5N8 virus were 10(5.3)EID50 and 10(6.7)EID50 in SPF and Korean native chickens, respectively. In addition, the mean death time was much longer, and the viral titers in tissues of H5N8-infected chickens were significantly lower, in the Korean group than in the SPF group. These features of the H5N8 virus likely account for its mild-to-moderate pathogenicity in commercial chicken farms, especially Korean native chicken flocks, despite the fact that it is a highly pathogenic virus according to the OIE criteria. To improve current understanding and management of HPAI, pathogenic characterization of novel emerging viruses should be performed by natural route in major poultry species in each country.

  16. Smartphone-Based Fluorescent Diagnostic System for Highly Pathogenic H5N1 Viruses.

    PubMed

    Yeo, Seon-Ju; Choi, Kyunghan; Cuc, Bui Thi; Hong, Nguyen Ngoc; Bao, Duong Tuan; Ngoc, Nguyen Minh; Le, Mai Quynh; Hang, Nguyen Le Khanh; Thach, Nguyen Co; Mallik, Shyam Kumar; Kim, Hak Sung; Chong, Chom-Kyu; Choi, Hak Soo; Sung, Haan Woo; Yu, Kyoungsik; Park, Hyun

    2016-01-01

    Field diagnostic tools for avian influenza (AI) are indispensable for the prevention and controlled management of highly pathogenic AI-related diseases. More accurate, faster and networked on-site monitoring is demanded to detect such AI viruses with high sensitivity as well as to maintain up-to-date information about their geographical transmission. In this work, we assessed the clinical and field-level performance of a smartphone-based fluorescent diagnostic device with an efficient reflective light collection module using a coumarin-derived dendrimer-based fluorescent lateral flow immunoassay. By application of an optimized bioconjugate, a smartphone-based diagnostic device had a two-fold higher detectability as compared to that of the table-top fluorescence strip reader for three different AI subtypes (H5N3, H7N1, and H9N2). Additionally, in a clinical study of H5N1-confirmed patients, the smartphone-based diagnostic device showed a sensitivity of 96.55% (28/29) [95% confidence interval (CI): 82.24 to 99.91] and a specificity of 98.55% (68/69) (95% CI: 92.19 to 99.96). The measurement results from the distributed individual smartphones were wirelessly transmitted via short messaging service and collected by a centralized database system for further information processing and data mining. Smartphone-based diagnosis provided highly sensitive measurement results for H5N1 detection within 15 minutes. Because of its high sensitivity, portability and automatic reporting feature, the proposed device will enable agile identification of patients and efficient control of AI dissemination. PMID:26877781

  17. Smartphone-Based Fluorescent Diagnostic System for Highly Pathogenic H5N1 Viruses

    PubMed Central

    Yeo, Seon-Ju; Choi, Kyunghan; Cuc, Bui Thi; Hong, Nguyen Ngoc; Bao, Duong Tuan; Ngoc, Nguyen Minh; Le, Mai Quynh; Hang, Nguyen Le Khanh; Thach, Nguyen Co; Mallik, Shyam Kumar; Kim, Hak Sung; Chong, Chom-Kyu; Choi, Hak Soo; Sung, Haan Woo; Yu, Kyoungsik; Park, Hyun

    2016-01-01

    Field diagnostic tools for avian influenza (AI) are indispensable for the prevention and controlled management of highly pathogenic AI-related diseases. More accurate, faster and networked on-site monitoring is demanded to detect such AI viruses with high sensitivity as well as to maintain up-to-date information about their geographical transmission. In this work, we assessed the clinical and field-level performance of a smartphone-based fluorescent diagnostic device with an efficient reflective light collection module using a coumarin-derived dendrimer-based fluorescent lateral flow immunoassay. By application of an optimized bioconjugate, a smartphone-based diagnostic device had a two-fold higher detectability as compared to that of the table-top fluorescence strip reader for three different AI subtypes (H5N3, H7N1, and H9N2). Additionally, in a clinical study of H5N1-confirmed patients, the smartphone-based diagnostic device showed a sensitivity of 96.55% (28/29) [95% confidence interval (CI): 82.24 to 99.91] and a specificity of 98.55% (68/69) (95% CI: 92.19 to 99.96). The measurement results from the distributed individual smartphones were wirelessly transmitted via short messaging service and collected by a centralized database system for further information processing and data mining. Smartphone-based diagnosis provided highly sensitive measurement results for H5N1 detection within 15 minutes. Because of its high sensitivity, portability and automatic reporting feature, the proposed device will enable agile identification of patients and efficient control of AI dissemination. PMID:26877781

  18. Smartphone-Based Fluorescent Diagnostic System for Highly Pathogenic H5N1 Viruses.

    PubMed

    Yeo, Seon-Ju; Choi, Kyunghan; Cuc, Bui Thi; Hong, Nguyen Ngoc; Bao, Duong Tuan; Ngoc, Nguyen Minh; Le, Mai Quynh; Hang, Nguyen Le Khanh; Thach, Nguyen Co; Mallik, Shyam Kumar; Kim, Hak Sung; Chong, Chom-Kyu; Choi, Hak Soo; Sung, Haan Woo; Yu, Kyoungsik; Park, Hyun

    2016-01-01

    Field diagnostic tools for avian influenza (AI) are indispensable for the prevention and controlled management of highly pathogenic AI-related diseases. More accurate, faster and networked on-site monitoring is demanded to detect such AI viruses with high sensitivity as well as to maintain up-to-date information about their geographical transmission. In this work, we assessed the clinical and field-level performance of a smartphone-based fluorescent diagnostic device with an efficient reflective light collection module using a coumarin-derived dendrimer-based fluorescent lateral flow immunoassay. By application of an optimized bioconjugate, a smartphone-based diagnostic device had a two-fold higher detectability as compared to that of the table-top fluorescence strip reader for three different AI subtypes (H5N3, H7N1, and H9N2). Additionally, in a clinical study of H5N1-confirmed patients, the smartphone-based diagnostic device showed a sensitivity of 96.55% (28/29) [95% confidence interval (CI): 82.24 to 99.91] and a specificity of 98.55% (68/69) (95% CI: 92.19 to 99.96). The measurement results from the distributed individual smartphones were wirelessly transmitted via short messaging service and collected by a centralized database system for further information processing and data mining. Smartphone-based diagnosis provided highly sensitive measurement results for H5N1 detection within 15 minutes. Because of its high sensitivity, portability and automatic reporting feature, the proposed device will enable agile identification of patients and efficient control of AI dissemination.

  19. Identification of Highly Variable Supernumerary Chromosome Segments in an Asexual Pathogen.

    PubMed

    Huang, Xiaoqiu; Das, Anindya; Sahu, Binod B; Srivastava, Subodh K; Leandro, Leonor F; O'Donnell, Kerry; Bhattacharyya, Madan K

    2016-01-01

    Supernumerary chromosome segments are known to harbor different transposons from their essential counterparts. The aim of this study was to investigate the role of transposons in the origin and evolution of supernumerary segments in the asexual fungal pathogen Fusarium virguliforme. We compared the genomes of 11 isolates comprising six Fusarium species that cause soybean sudden death syndrome (SDS) or bean root rot (BRR), and identified significant levels of genetic variation in A+T-rich repeat blocks of the essential chromosomes and in A+T-neutral regions of the supernumerary segments. The A+T-rich repeat blocks in the essential chromosomes were highly variable between F. virguliforme and non-F. virguliforme isolates, but were scarcely variable between F. virguliforme isolates. The A+T-neutral regions in the supernumerary segments, however, were highly variable between F. virguliforme isolates, with a statistically significant number (21 standard deviations above the mean) of single nucleotide polymorphisms (SNPs). And supernumerary sequence types and rearrangement patterns of some F. virguliforme isolates were present in an isolate of F. cuneirostrum but not in the other F. virguliforme isolates. The most variable and highly expressed region in the supernumerary segments contained an active DNA transposon that was a most conserved match between F. virguliforme and the unrelated fungus Tolypocladium inflatum. This transposon was absent from two of the F. virguliforme isolates. Furthermore, transposons in the supernumerary segments of some F. virguliforme isolates were present in non-F. virguliforme isolates, but were absent from the other F. virguliforme isolates. Two supernumerary P450 enzymes were 43% and 57% identical to their essential counterparts. This study has raised the possibility that transposons generate genetic variation in supernumerary chromosome segments by frequent horizontal transfer within and between closely related species. PMID:27341103

  20. Identification of Highly Variable Supernumerary Chromosome Segments in an Asexual Pathogen

    PubMed Central

    Huang, Xiaoqiu; Das, Anindya; Sahu, Binod B.; Srivastava, Subodh K.; Leandro, Leonor F.; O’Donnell, Kerry; Bhattacharyya, Madan K.

    2016-01-01

    Supernumerary chromosome segments are known to harbor different transposons from their essential counterparts. The aim of this study was to investigate the role of transposons in the origin and evolution of supernumerary segments in the asexual fungal pathogen Fusarium virguliforme. We compared the genomes of 11 isolates comprising six Fusarium species that cause soybean sudden death syndrome (SDS) or bean root rot (BRR), and identified significant levels of genetic variation in A+T-rich repeat blocks of the essential chromosomes and in A+T-neutral regions of the supernumerary segments. The A+T-rich repeat blocks in the essential chromosomes were highly variable between F. virguliforme and non-F. virguliforme isolates, but were scarcely variable between F. virguliforme isolates. The A+T-neutral regions in the supernumerary segments, however, were highly variable between F. virguliforme isolates, with a statistically significant number (21 standard deviations above the mean) of single nucleotide polymorphisms (SNPs). And supernumerary sequence types and rearrangement patterns of some F. virguliforme isolates were present in an isolate of F. cuneirostrum but not in the other F. virguliforme isolates. The most variable and highly expressed region in the supernumerary segments contained an active DNA transposon that was a most conserved match between F. virguliforme and the unrelated fungus Tolypocladium inflatum. This transposon was absent from two of the F. virguliforme isolates. Furthermore, transposons in the supernumerary segments of some F. virguliforme isolates were present in non-F. virguliforme isolates, but were absent from the other F. virguliforme isolates. Two supernumerary P450 enzymes were 43% and 57% identical to their essential counterparts. This study has raised the possibility that transposons generate genetic variation in supernumerary chromosome segments by frequent horizontal transfer within and between closely related species. PMID:27341103

  1. Pathogenesis of highly pathogenic porcine reproductive and respiratory syndrome virus in Chinese Tibetan swine.

    PubMed

    Fan, Baochao; Zhang, Hongjian; Bai, Juan; Liu, Xing; Li, Yufeng; Wang, Xianwei; Jiang, Ping

    2016-10-01

    Highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) was first characterized in 2006 in China, and it causes great economic losses to the Chinese swine production industry. A China Landrace pig, the Tibetan pig, which has striking phenotypic and physiological differences from lowland pigs, is mainly distributed in the Tibetan highlands of China. The susceptibility of the Tibetan pig to HP-PRRSV has not been reported. In this study, 15 4-week-old Tibetan piglets were divided into three groups, and their susceptibility to HP-PRRSV was examined in the highland region. Five pigs in group 1 were inoculated intranasally with HP-PRRSV strain BB0907. At 2days post-inoculation, five other pigs were introduced into this group and then removed to a separated room to serve as contact group 2. Meanwhile, five pigs in group 3 were mock infected and used as controls. The results showed that the pigs in the inoculated and contact groups showed high fevers and clear clinical signs, including depression, anorexia, lethargy, sticky eye secretions, and hind limb paralysis, with high mortality. The main symptom was interstitial pneumonia. Viremia appeared on days 4 to 14 post-infection. HP-PRRSV infection resulted in inflammatory responses within the first week of infection, as evidenced by the expression of tumor necrosis factor alpha, interleukin (IL)-1β, IL-6, and IL-10. All the data indicate that the Tibetan pig is susceptible to HP-PRRSV infection. Thus, it is necessary to investigate and prevent PRRSV infections in the highland region in China. PMID:27663367

  2. New Reassortant H5N6 Highly Pathogenic Avian Influenza Viruses in Southern China, 2014

    PubMed Central

    Jiao, Peirong; Cui, Jin; Song, Yafen; Song, Hui; Zhao, Zhishan; Wu, Siyu; Qu, Nannan; Wang, Nianchen; Ouyang, Guowen; Liao, Ming

    2016-01-01

    New reassortant H5N6 highly pathogenic avian influenza viruses (AIVs) were isolated from apparently healthy domestic ducks in Southern China in 2014. Our results show that the viruses grew efficiently in eggs and replicated systemically in chickens. They were completely lethal in chicken (100% mortality), and the mean death time was 6 to 7 days post-inoculation. The viruses could transmit in chickens by naïve contact. BLAST analysis revealed that their HA gene was most closely related to A/wild duck/Shangdong/628/2011 (H5N1), and their NA genes were most closely related to A/swine/Guangdong/K6/2010 (H6N6). The other genes had the highest identity with A/wild duck/Fujian/1/2011(H5N1). The results of phylogenetic analysis showed that their HA genes clustered into clade 2.3.4.4 of the H5N1 viruses and all genes derived from H5 were Mix-like or H6-like viruses. Thus, the new H5N6 viruses were reassortmented of H5N1 and H6N6 virus. Therefore, the circulation of the new H5N6 AIVs may become a threat to poultry and human health. PMID:27242767

  3. Genotypic diversity of H5N1 highly pathogenic avian influenza viruses.

    PubMed

    Zhao, Zi-Ming; Shortridge, Kennedy F; Garcia, Maricarmen; Guan, Yi; Wan, Xiu-Feng

    2008-09-01

    Besides enormous economic losses to the poultry industry, recent H5N1 highly pathogenic avian influenza viruses (HPAIVs) originating in eastern Asia have posed serious threats to public health. Up to April 17, 2008, 381 human cases had been confirmed with a mortality of more than 60 %. Here, we attempt to identify potential progenitor genes for H5N1 HPAIVs since their first recognition in 1996; most were detected in the Eurasian landmass before 1996. Combinations among these progenitor genes generated at least 21 reassortants (named H5N1 progenitor reassortant, H5N1-PR1-21). H5N1-PR1 includes A/Goose/Guangdong/1/1996(H5N1). Only reassortants H5N1-PR2 and H5N1-PR7 were associated with confirmed human cases: H5N1-PR2 in the Hong Kong H5N1 outbreak in 1997 and H5N1-PR7 in laboratory confirmed human cases since 2003. H5N1-PR7 also contains a majority of the H5N1 viruses causing avian influenza outbreaks in birds, including the first wave of genotype Z, Qinghai-like and Fujian-like virus lineages. Among the 21 reassortants identified, 13 are first reported here. This study illustrates evolutionary patterns of H5N1 HPAIVs, which may be useful toward pandemic preparedness as well as avian influenza prevention and control.

  4. New Reassortant H5N6 Highly Pathogenic Avian Influenza Viruses in Southern China, 2014.

    PubMed

    Jiao, Peirong; Cui, Jin; Song, Yafen; Song, Hui; Zhao, Zhishan; Wu, Siyu; Qu, Nannan; Wang, Nianchen; Ouyang, Guowen; Liao, Ming

    2016-01-01

    New reassortant H5N6 highly pathogenic avian influenza viruses (AIVs) were isolated from apparently healthy domestic ducks in Southern China in 2014. Our results show that the viruses grew efficiently in eggs and replicated systemically in chickens. They were completely lethal in chicken (100% mortality), and the mean death time was 6 to 7 days post-inoculation. The viruses could transmit in chickens by naïve contact. BLAST analysis revealed that their HA gene was most closely related to A/wild duck/Shangdong/628/2011 (H5N1), and their NA genes were most closely related to A/swine/Guangdong/K6/2010 (H6N6). The other genes had the highest identity with A/wild duck/Fujian/1/2011(H5N1). The results of phylogenetic analysis showed that their HA genes clustered into clade 2.3.4.4 of the H5N1 viruses and all genes derived from H5 were Mix-like or H6-like viruses. Thus, the new H5N6 viruses were reassortmented of H5N1 and H6N6 virus. Therefore, the circulation of the new H5N6 AIVs may become a threat to poultry and human health. PMID:27242767

  5. The avian and mammalian host range of highly pathogenic avian H5N1 influenza

    PubMed Central

    Kaplan, Bryan S.; Webby, Richard J.

    2013-01-01

    Highly pathogenic H5N1 influenza viruses have been isolated from a number of avian and mammalian species. Despite intensive control measures the number of human and animal cases continues to increase. A more complete understanding of susceptible species and of contributing environmental and molecular factors is crucial if we are to slow the rate of new cases. H5N1 is currently endemic in domestic poultry in only a handful of countries with sporadic and unpredictable spread to other countries. Close contact of terrestrial bird or mammalian species with infected poultry/waterfowl or their biological products is the major route for interspecies transmission. Intra-species transmission of H5N1 in mammals, including humans, has taken place on a limited scale though it remains to be seen if this will change; recent laboratory studies suggest that it is indeed possible. Here we review the avian and mammalian species that are naturally susceptible to H5N1 infection and the molecular factors associated with its expanded host range. PMID:24025480

  6. Identifying risk factors of highly pathogenic avian influenza (H5N1 subtype) in Indonesia

    PubMed Central

    Leo, Loth; Marius, Gilbert; Jianmei, Wu; Christina, Czarnecki; Muhammad, Hidayat; Xiangming, Xiao

    2016-01-01

    Highly pathogenic avian influenza (HPAI), subtype H5N1, was first officially reported in Indonesia in 2004. Since then the disease has spread and is now endemic in large parts of the country. This study investigated the statistical relationship between a set of risk factors and the presence or absence of HPAI in Indonesia during 2006 and 2007. HPAI was evaluated through participatory disease surveillance (PDS) in backyard village chickens (the study population), and risk factors included descriptors of people and poultry distribution (separating chickens, ducks and production sectors), poultry movement patterns and agro-ecological conditions. The study showed that the risk factors “elevation”, “human population density” and “rice cropping” were significant in accounting for the spatial variation of the PDS-defined HPAI cases. These findings were consistent with earlier studies in Thailand and Vietnam. In addition “commercial poultry population”, and two indicators of market locations and transport; “human settlements” and “road length”, were identified as significant risk factors in the models. In contrast to several previous studies carried out in Southeast Asia, domestic backyard ducks were not found to be a significant risk factor in Indonesia. The study used surrogate estimates of market locations and marketing chains and further work should focus on the actual location of the live bird markets, and on the flow of live poultry and poultry products between them, so that patterns of possible transmission, and regions of particular risk could be better inferred. PMID:21813198

  7. Cockroaches as a Source of High Bacterial Pathogens with Multidrug Resistant Strains in Gondar Town, Ethiopia.

    PubMed

    Moges, Feleke; Eshetie, Setegn; Endris, Mengistu; Huruy, Kahsay; Muluye, Dagnachew; Feleke, Tigist; G/Silassie, Fisha; Ayalew, Getenet; Nagappan, Raja

    2016-01-01

    Background. Cockroaches are source of bacterial infections and this study was aimed to assess bacterial isolates and their antimicrobial profiles from cockroaches in Gondar town, Ethiopia. Methods. A total of 60 cockroaches were collected from March 1 to May 30, 2014, in Gondar town. Bacterial species were isolated from external and internal parts of cockroaches. Disk diffusion method was used to determine antibiotic susceptibility patterns. Data were entered and analyzed by using SPSS version 20; P values <0.005 were considered as statistically significant. Results. Of 181 identified bacteria species, 110 (60.8%) and 71 (39.2%) were identified from external and internal parts of cockroaches, respectively. Klebsiella pneumoniae 32 (17.7%), Escherichia coli 29 (16%), and Citrobacter spp. 27 (15%) were the predominant isolates. High resistance rate was observed to cotrimoxazole, 60 (33.1%), and least resistance rate was noted to ciprofloxacin, 2 (1.1%). Additionally, 116 (64.1%) of the isolates were MDR strains; Salmonella spp. were the leading MDR isolates (100%) followed by Enterobacter (90.5%) and Shigella spp. (76.9%). Conclusion. Cockroaches are the potential source of bacteria pathogens with multidrug resistant strains and hence effective preventive and control measures are required to minimize cockroach related infections.

  8. Cockroaches as a Source of High Bacterial Pathogens with Multidrug Resistant Strains in Gondar Town, Ethiopia

    PubMed Central

    Huruy, Kahsay; Muluye, Dagnachew; Feleke, Tigist; G/Silassie, Fisha; Ayalew, Getenet; Nagappan, Raja

    2016-01-01

    Background. Cockroaches are source of bacterial infections and this study was aimed to assess bacterial isolates and their antimicrobial profiles from cockroaches in Gondar town, Ethiopia. Methods. A total of 60 cockroaches were collected from March 1 to May 30, 2014, in Gondar town. Bacterial species were isolated from external and internal parts of cockroaches. Disk diffusion method was used to determine antibiotic susceptibility patterns. Data were entered and analyzed by using SPSS version 20; P values <0.005 were considered as statistically significant. Results. Of 181 identified bacteria species, 110 (60.8%) and 71 (39.2%) were identified from external and internal parts of cockroaches, respectively. Klebsiella pneumoniae 32 (17.7%), Escherichia coli 29 (16%), and Citrobacter spp. 27 (15%) were the predominant isolates. High resistance rate was observed to cotrimoxazole, 60 (33.1%), and least resistance rate was noted to ciprofloxacin, 2 (1.1%). Additionally, 116 (64.1%) of the isolates were MDR strains; Salmonella spp. were the leading MDR isolates (100%) followed by Enterobacter (90.5%) and Shigella spp. (76.9%). Conclusion. Cockroaches are the potential source of bacteria pathogens with multidrug resistant strains and hence effective preventive and control measures are required to minimize cockroach related infections. PMID:27340653

  9. Cockroaches as a Source of High Bacterial Pathogens with Multidrug Resistant Strains in Gondar Town, Ethiopia.

    PubMed

    Moges, Feleke; Eshetie, Setegn; Endris, Mengistu; Huruy, Kahsay; Muluye, Dagnachew; Feleke, Tigist; G/Silassie, Fisha; Ayalew, Getenet; Nagappan, Raja

    2016-01-01

    Background. Cockroaches are source of bacterial infections and this study was aimed to assess bacterial isolates and their antimicrobial profiles from cockroaches in Gondar town, Ethiopia. Methods. A total of 60 cockroaches were collected from March 1 to May 30, 2014, in Gondar town. Bacterial species were isolated from external and internal parts of cockroaches. Disk diffusion method was used to determine antibiotic susceptibility patterns. Data were entered and analyzed by using SPSS version 20; P values <0.005 were considered as statistically significant. Results. Of 181 identified bacteria species, 110 (60.8%) and 71 (39.2%) were identified from external and internal parts of cockroaches, respectively. Klebsiella pneumoniae 32 (17.7%), Escherichia coli 29 (16%), and Citrobacter spp. 27 (15%) were the predominant isolates. High resistance rate was observed to cotrimoxazole, 60 (33.1%), and least resistance rate was noted to ciprofloxacin, 2 (1.1%). Additionally, 116 (64.1%) of the isolates were MDR strains; Salmonella spp. were the leading MDR isolates (100%) followed by Enterobacter (90.5%) and Shigella spp. (76.9%). Conclusion. Cockroaches are the potential source of bacteria pathogens with multidrug resistant strains and hence effective preventive and control measures are required to minimize cockroach related infections. PMID:27340653

  10. Identifying risk factors of highly pathogenic avian influenza (H5N1 subtype) in Indonesia.

    PubMed

    Loth, Leo; Gilbert, Marius; Wu, Jianmei; Czarnecki, Christina; Hidayat, Muhammad; Xiao, Xiangming

    2011-10-01

    Highly pathogenic avian influenza (HPAI), subtype H5N1, was first officially reported in Indonesia in 2004. Since then the disease has spread and is now endemic in large parts of the country. This study investigated the statistical relationship between a set of risk factors and the presence or absence of HPAI in Indonesia during 2006 and 2007. HPAI was evaluated through participatory disease surveillance (PDS) in backyard village chickens (the study population), and risk factors included descriptors of people and poultry distribution (separating chickens, ducks and production sectors), poultry movement patterns and agro-ecological conditions. The study showed that the risk factors "elevation", "human population density" and "rice cropping" were significant in accounting for the spatial variation of the PDS-defined HPAI cases. These findings were consistent with earlier studies in Thailand and Vietnam. In addition "commercial poultry population", and two indicators of market locations and transport; "human settlements" and "road length", were identified as significant risk factors in the models. In contrast to several previous studies carried out in Southeast Asia, domestic backyard ducks were not found to be a significant risk factor in Indonesia. The study used surrogate estimates of market locations and marketing chains and further work should focus on the actual location of the live bird markets, and on the flow of live poultry and poultry products between them, so that patterns of possible transmission, and regions of particular risk could be better inferred.

  11. New reassortant H5N8 highly pathogenic avian influenza virus from waterfowl in Southern China

    PubMed Central

    Song, Yafen; Cui, Jin; Song, Hui; Ye, Jiaqi; Zhao, Zhishan; Wu, Siyu; Xu, Chenggang; Jiao, Peirong; Liao, Ming

    2015-01-01

    New reassortant H5N8 highly pathogenic avian influenza viruses were isolated from waterfowl in Southern China. Blast analysis demonstrated that the PB2 gene in these viruses were most closely related to A/wild duck/Shangdong/628/2011 (H5N1), while their NP genes were both more closely related to A/wild duck/Shandong/1/2011 (H5N1) and A/duck/Jiangsu/k1203/2010 (H5N8). However, the HA, NA, PB1, PA, M, and NS genes had the highest identity with A/duck/Jiangsu/k1203/2010 (H5N8). Phylogenetic analysis revealed that their HA genes belonged to the same GsGd H5 clade 2.3.4.4 detected in China in 2010. Therefore, we supposed that these H5N8 viruses might be novel reassortant viruses that have a H5N8 backbone while acquiring PB2 and NP genes from H5N1 viruses. This study is useful for better understanding the genetic and antigenic evolution of H5 avian influenza viruses in Southern China. PMID:26557113

  12. Susceptibility of Canada Geese (Branta canadensis) to highly pathogenic avian influenza virus (H5N1).

    PubMed

    Pasick, John; Berhane, Yohannes; Embury-Hyatt, Carissa; Copps, John; Kehler, Helen; Handel, Katherine; Babiuk, Shawn; Hooper-McGrevy, Kathleen; Li, Yan; Mai Le, Quynh; Lien Phuong, Song

    2007-12-01

    Migratory birds have been implicated in the long-range spread of highly pathogenic avian influenza (HPAI) A virus (H5N1) from Asia to Europe and Africa. Although sampling of healthy wild birds representing a large number of species has not identified possible carriers of influenza virus (H5N1) into Europe, surveillance of dead and sick birds has demonstrated mute (Cygnus olor) and whooper (C. cygnus) swans as potential sentinels. Because of concerns that migratory birds could spread H5N1 subtype to the Western Hemisphere and lead to its establishment within free-living avian populations, experimental studies have addressed the susceptibility of several indigenous North American duck and gull species. We examined the susceptibility of Canada geese (Branta canadensis) to HPAI virus (H5N1). Large populations of this species can be found in periagricultural and periurban settings and thus may be of potential epidemiologic importance if H5N1 subtype were to establish itself in North American wild bird populations.

  13. Risk factors for highly pathogenic avian influenza in commercial layer chicken farms in bangladesh during 2011.

    PubMed

    Osmani, M G; Thornton, R N; Dhand, N K; Hoque, M A; Milon, Sk M A; Kalam, M A; Hossain, M; Yamage, M

    2014-12-01

    A case-control study conducted during 2011 involved 90 randomly selected commercial layer farms infected with highly pathogenic avian influenza type A subtype H5N1 (HPAI) and 175 control farms randomly selected from within 5 km of infected farms. A questionnaire was designed to obtain information about potential risk factors for contracting HPAI and was administered to farm owners or managers. Logistic regression analyses were conducted to identify significant risk factors. A total of 20 of 43 risk factors for contracting HPAI were identified after univariable logistic regression analysis. A multivariable logistic regression model was derived by forward stepwise selection. Both unmatched and matched analyses were performed. The key risk factors identified were numbers of staff, frequency of veterinary visits, presence of village chickens roaming on the farm and staff trading birds. Aggregating these findings with those from other studies resulted in a list of 16 key risk factors identified in Bangladesh. Most of these related to biosecurity. It is considered feasible for Bangladesh to achieve a very low incidence of HPAI. Using the cumulative list of risk factors to enhance biosecurity pertaining to commercial farms would facilitate this objective.

  14. H5N1 highly pathogenic avian influenza in Indonesia: retrospective considerations.

    PubMed

    Daniels, Peter; Wiyono, Agus; Sawitri, Elly; Poermadjaja, Bagoes; Sims, L D

    2013-01-01

    Indonesia is one of the five countries where highly pathogenic avian influenza viruses of the H5N1 subtype (H5N1 HPAI) remain endemic in poultry. Importantly, it is one of the countries where the virus causes human infections. WHO data indicate that as of 2 May 2012, 189 human cases of Influenza A (H5N1) had been reported in Indonesia, with 157 human deaths. These human cases included a small number in which limited human-to-human transmission could have occurred. Hence, there remains a critical need in Indonesia for a more effective One Health approach to the control and prevention of this disease in people and in poultry. This chapter explores a number of aspects of the evolution of this disease in Indonesia, the virus that causes it and the control and preventive measures introduced, focusing on the successes and shortcomings of veterinary and One Health approaches. Indonesia provides many examples of situations where this latter approach has been successful, and others where further work is needed to maximize the benefits from coordinated responses to this disease leading to effective management of the risk to human health.

  15. Financial effects of the highly pathogenic avian influenza outbreaks on the Turkish broiler producers.

    PubMed

    Aral, Y; Yalcin, C; Cevger, Y; Sipahi, C; Sariozkan, S

    2010-05-01

    This research aimed at assessing the financial effects of the 2005 to 2006 highly pathogenic avian influenza outbreaks on Turkish broiler enterprises. The data were obtained from an interview survey carried out in 499 enterprises randomly selected from 14 provinces that accounted for 79% of the national broiler production. The research revealed that the contracted broiler producers lost on average 1.38 cycles of production and their management fee reduced by 14.7% in 8 mo after the outbreaks. As a result, the broiler production and the enterprise income declined by 34.8 and 44.3%, respectively. The bank loan of the producers rose by 161%. A total of 93% of the producers did not do any other supplementary work during the idle production period in spite of the fact that broiler production was the only business of 36% of them. Furthermore, more than half of the producers (56%) stated that they were considering expanding their business, but suspended this idea due to the outbreak. Approximately 87% of the producers increased the biosecurity measures after the outbreaks. The nationwide effects of the avian influenza outbreaks on the contracted broilers farms were estimated to be US$100.8 million (US$7,967/broiler house). The futures of the contracted broiler producers are fully dependent upon those of the integrated firms. Any negative effects on the latter appeared to be transferred directly to the former. However, the government neglected the integrated firms in the avian influenza compensation programs.

  16. Assessment of the Potential Distance of Dispersal of High Pathogenicity Avian Influenza Virus by Wild Mallards.

    PubMed

    Śmietanka, Krzysztof; Bocian, Łukasz; Meissner, Włodzimierz; Ziętek-Barszcz, Anna; Żółkoś, Katarzyna

    2016-05-01

    This work presents the results of studies aimed at assessing the median and maximum distances covered by wild mallards (Anas platyrhynchos; n = 38), hypothetically infected with the high pathogenicity avian influenza virus (HPAIV) during spring migrations, using GPS-GSM tracking and published data on the susceptibility to HPAIV infection and duration of shedding. The model was based on the assumptions that the birds shed virus in the absence of clinical signs during infectious periods (IP) that were assumed to last 1 day (IP1), 4 days (IP4), and 8 days (IP8) and that each day of migration is a hypothetical day of the onset of IP. Using the haversine formula over a sliding timeframe corresponding to each IP, distances were estimated for each duck that undertook migration and then the maximum distance (Dmax) was selected. Ten mallards undertook spring migrations but, due to the loss of signal in the GPS-GSM devices, only three ducks were observed during autumn migrations. The following ranges of Dmax values were calculated for spring migrations: 124-382 km for IP1 (median 210 km), 208-632 km for IP4 (median 342 km), and 213-687 km for IP8 (median 370 km). The present study provides information that can be used as a data source to perform risk assessment related to the contribution of wild mallards in the dispersal of HPAIV over considerable distances. PMID:27309073

  17. Scavenging ducks and transmission of highly pathogenic avian influenza, Java, Indonesia.

    PubMed

    Henning, Joerg; Wibawa, Hendra; Morton, John; Usman, Tri Bhakti; Junaidi, Akhmad; Meers, Joanne

    2010-08-01

    In Java, Indonesia, during March 2007-March 2008, 96 farms with scavenging ducks that were not vaccinated against highly pathogenic avian influenza (HPAI) were monitored bimonthly. Bird-level (prevalence among individual birds) H5 seroprevalence was 2.6% for ducks and 0.5% for chickens in contact with ducks. At least 1 seropositive bird was detected during 19.5% and 2.0% of duck- and chicken-flock visits, respectively. Duck flocks were 12.4x more likely than chicken flocks to have seropositive birds. During 21.4% of farm visits,

  18. Influenza A(H5N8) Virus Similar to Strain in Korea Causing Highly Pathogenic Avian Influenza in Germany.

    PubMed

    Harder, Timm; Maurer-Stroh, Sebastian; Pohlmann, Anne; Starick, Elke; Höreth-Böntgen, Detlef; Albrecht, Karin; Pannwitz, Gunter; Teifke, Jens; Gunalan, Vithiagaran; Lee, Raphael T C; Sauter-Louis, Carola; Homeier, Timo; Staubach, Christoph; Wolf, Carola; Strebelow, Günter; Höper, Dirk; Grund, Christian; Conraths, Franz J; Mettenleiter, Thomas C; Beer, Martin

    2015-05-01

    Highly pathogenic avian influenza (H5N8) virus, like the recently described H5N8 strain from Korea, was detected in November 2014 in farmed turkeys and in a healthy common teal (Anas crecca) in northeastern Germany. Infected wild birds possibly introduced this virus. PMID:25897703

  19. Inhibition of enteric pathogens and surrogates using integrated, high intensity 405nm led light on the surface of almonds

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The disinfecting properties of 405 nm light were investigated against Escherichia coli O157:H7, Salmonella, and their non-pathogenic surrogate bacteria on the surface of almonds. High intensity monochromatic blue light (MBL) was generated from an array of narrow-band 405 nm light-emitting diodes (LE...

  20. Widespread detection of highly pathogenic H5 influenza viruses in wild birds from the Pacific Flyway of the United States.

    PubMed

    Bevins, S N; Dusek, R J; White, C L; Gidlewski, T; Bodenstein, B; Mansfield, K G; DeBruyn, P; Kraege, D; Rowan, E; Gillin, C; Thomas, B; Chandler, S; Baroch, J; Schmit, B; Grady, M J; Miller, R S; Drew, M L; Stopak, S; Zscheile, B; Bennett, J; Sengl, J; Brady, Caroline; Ip, H S; Spackman, E; Killian, M L; Torchetti, M K; Sleeman, J M; Deliberto, T J

    2016-01-01

    A novel highly pathogenic avian influenza virus belonging to the H5 clade 2.3.4.4 variant viruses was detected in North America in late 2014. Motivated by the identification of these viruses in domestic poultry in Canada, an intensive study was initiated to conduct highly pathogenic avian influenza surveillance in wild birds in the Pacific Flyway of the United States. A total of 4,729 hunter-harvested wild birds were sampled and highly pathogenic avian influenza virus was detected in 1.3% (n = 63). Three H5 clade 2.3.4.4 subtypes were isolated from wild birds, H5N2, H5N8, and H5N1, representing the wholly Eurasian lineage H5N8 and two novel reassortant viruses. Testing of 150 additional wild birds during avian morbidity and mortality investigations in Washington yielded 10 (6.7%) additional highly pathogenic avian influenza isolates (H5N8 = 3 and H5N2 = 7). The geographically widespread detection of these viruses in apparently healthy wild waterfowl suggest that the H5 clade 2.3.4.4 variant viruses may behave similarly in this taxonomic group whereby many waterfowl species are susceptible to infection but do not demonstrate obvious clinical disease. Despite these findings in wild waterfowl, mortality has been documented for some wild bird species and losses in US domestic poultry during the first half of 2015 were unprecedented. PMID:27381241

  1. The pathobiology of highly pathogenic H5N2 avian influenza virus in Ruddy ducks and Lesser Scaup

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The susceptibility and pathogenesis of avian influenza virus (AIV) has not been characterized in numerous duck species, especially diving ducks, some of which migrate across the continental U.S. The pathobiology of highly pathogenic (HP) H5N2 AIV was characterized in two diving duck species, Ruddy ...

  2. Widespread detection of highly pathogenic H5 influenza viruses in wild birds from the Pacific Flyway of the United States

    USGS Publications Warehouse

    Bevins, S.N.; Dusek, Robert J.; White, C. LeAnn; Gidlewski, Thomas; Bodenstein, B.; Mansfield, Kristin G.; DeBruyn, Paul; Kraege, Donald K.; Rowan, E.L.; Gillin, Colin; Thomas, B.; Chandler, S.; Baroch, J.; Schmit, B.; Grady, M. J.; Miller, R. S.; Drew, M.L.; Stopak, S.; Zscheile, B.; Bennett, J.; Sengl, J.; Brady, Caroline; Ip, Hon S.; Spackman, Erica; Killian, M. L.; Torchetti, Mia Kim; Sleeman, Jonathan M.; DeLiberto, T.J.

    2016-01-01

    A novel highly pathogenic avian influenza virus belonging to the H5 clade 2.3.4.4 variant viruses was detected in North America in late 2014. Motivated by the identification of these viruses in domestic poultry in Canada, an intensive study was initiated to conduct highly pathogenic avian influenza surveillance in wild birds in the Pacific Flyway of the United States. A total of 4,729 hunter-harvested wild birds were sampled and highly pathogenic avian influenza virus was detected in 1.3% (n = 63). Three H5 clade 2.3.4.4 subtypes were isolated from wild birds, H5N2, H5N8, and H5N1, representing the wholly Eurasian lineage H5N8 and two novel reassortant viruses. Testing of 150 additional wild birds during avian morbidity and mortality investigations in Washington yielded 10 (6.7%) additional highly pathogenic avian influenza isolates (H5N8 = 3 and H5N2 = 7). The geographically widespread detection of these viruses in apparently healthy wild waterfowl suggest that the H5 clade 2.3.4.4 variant viruses may behave similarly in this taxonomic group whereby many waterfowl species are susceptible to infection but do not demonstrate obvious clinical disease. Despite these findings in wild waterfowl, mortality has been documented for some wild bird species and losses in US domestic poultry during the first half of 2015 were unprecedented.

  3. Emergence of H5N1 high pathogenicity avian influenza strains in Indonesia that are resistant to vaccines

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Vaccines have been used to protect poultry in Asia against H5N1 high pathogenicity avian influenza (HPAI) since 2002. Reports of vaccine “failures” began to emerge in 2006 in Indonesia, with identification of clinical disease consistent with HPAI or isolation of H5N1 HPAIV in vaccinated flocks or in...

  4. Mechanisms of transmission and spread of H5N1 high pathogenicity avian influenza virus in birds and mammals

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Eurasian-African H5N1 high pathogenicity avian influenza (HPAI) virus has crossed multiple species barriers to infect poultry, captive and wild birds, carnivorous mammals and humans. The specific transmission mechanisms are unclear in most cases, but experimental studies and field data sug...

  5. Surveillance for highly pathogenic avian influenza virus in wild birds during outbreaks in domestic poultry, Minnesota, 2015

    USGS Publications Warehouse

    Jennelle, Christopher S.; Carstensen, Michelle; Hildebrand, Erik C.; Cornicelli, Louis; Wolf, Paul C.; Grear, Daniel; Ip, Hon S.; VanDalen, Kaci K.; Minicucci, Larissa A.

    2016-01-01

    In 2015, a major outbreak of highly pathogenic avian influenza virus (HPAIV) infection devastated poultry facilities in Minnesota, USA. To clarify the role of wild birds, we tested 3,139 waterfowl fecal samples and 104 sick and dead birds during March 9–June 4, 2015. HPAIV was isolated from a Cooper’s hawk but not from waterfowl.

  6. Widespread detection of highly pathogenic H5 influenza viruses in wild birds from the Pacific Flyway of the United States

    PubMed Central

    Bevins, S. N.; Dusek, R. J.; White, C. L.; Gidlewski, T.; Bodenstein, B.; Mansfield, K. G.; DeBruyn, P.; Kraege, D.; Rowan, E.; Gillin, C.; Thomas, B.; Chandler, S.; Baroch, J.; Schmit, B.; Grady, M. J.; Miller, R. S.; Drew, M. L.; Stopak, S.; Zscheile, B.; Bennett, J.; Sengl, J.; Brady, Caroline; Ip, H. S.; Spackman, E.; Killian, M. L.; Torchetti, M. K.; Sleeman, J. M.; Deliberto, T. J.

    2016-01-01

    A novel highly pathogenic avian influenza virus belonging to the H5 clade 2.3.4.4 variant viruses was detected in North America in late 2014. Motivated by the identification of these viruses in domestic poultry in Canada, an intensive study was initiated to conduct highly pathogenic avian influenza surveillance in wild birds in the Pacific Flyway of the United States. A total of 4,729 hunter-harvested wild birds were sampled and highly pathogenic avian influenza virus was detected in 1.3% (n = 63). Three H5 clade 2.3.4.4 subtypes were isolated from wild birds, H5N2, H5N8, and H5N1, representing the wholly Eurasian lineage H5N8 and two novel reassortant viruses. Testing of 150 additional wild birds during avian morbidity and mortality investigations in Washington yielded 10 (6.7%) additional highly pathogenic avian influenza isolates (H5N8 = 3 and H5N2 = 7). The geographically widespread detection of these viruses in apparently healthy wild waterfowl suggest that the H5 clade 2.3.4.4 variant viruses may behave similarly in this taxonomic group whereby many waterfowl species are susceptible to infection but do not demonstrate obvious clinical disease. Despite these findings in wild waterfowl, mortality has been documented for some wild bird species and losses in US domestic poultry during the first half of 2015 were unprecedented. PMID:27381241

  7. Outbreaks of highly pathogenic Eurasian H5N8 avian influenza in two commercial poultry flocks in California

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In January 2015, a highly pathogenic Eurasian lineage H5N8 avian influenza (AI) virus was detected in a commercial meat turkey flock in Stanislaus County, California. Approximately 3 weeks later, a similar case was diagnosed in commercial chickens from a different company located in Kings County, C...

  8. High pathogenicity avian influenza outbreaks since 2008 except multi-continental panzootic of H5 Goose/Guangdong-lineage viruses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Since 2008, seven countries from five continents have experienced highly pathogenic avian influenza (HPAI) outbreaks in poultry due to viruses unrelated to H5 Goose/Guangdong lineage viruses. These have covered a range of virus subtypes and affected different production species from chickens to ost...

  9. Bronchointerstitial pneumonia in guinea pigs following inoculation with H5N1 high pathogenicity avian influenza virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The H5N1 high pathogenicity avian influenza (HPAI) viruses have caused widespread disease of poultry in Asia, Africa and the Middle East, and sporadic human infections. The guinea pig model has been used to study human H3N2 and H1N1 influenza viruses, but knowledge is lacking on H5N1 HPAI virus inf...

  10. Validation of high-throughput real time polymerase chain reaction assays for simultaneous detection of invasive citrus pathogens.

    PubMed

    Saponari, Maria; Loconsole, Giuliana; Liao, Hui-Hong; Jiang, Bo; Savino, Vito; Yokomi, Raymond K

    2013-11-01

    A number of important citrus pathogens are spread by graft propagation, arthropod vector transmission and inadvertent import and dissemination of infected plants. For these reasons, citrus disease management and clean stock programs require pathogen detection systems which are economical and sensitive to maintain a healthy industry. To this end, multiplex quantitative real-time PCR (qPCR) assays were developed allowing high-throughput and simultaneous detection of some major invasive citrus pathogens. Automated high-throughput extraction comparing several bead-based commercial extraction kits were tested and compared with tissue print and manual extraction to obtain nucleic acids from healthy and pathogen-infected citrus trees from greenhouse in planta collections and field. Total nucleic acids were used as templates for pathogen detection. Multiplex reverse transcription-qPCR (RT-qPCR) assays were developed for simultaneous detection of six targets including a virus, two viroids, a bacterium associated with huanglongbing and a citrus RNA internal control. Specifically, two one-step TaqMan-based multiplex RT-qPCR assays were developed and tested with target templates to determine sensitivity and detection efficiency. The first assay included primers and probes for 'Candidatus Liberibacter asiaticus' (CLas) and Citrus tristeza virus (CTV) broad spectrum detection and genotype differentiation (VT- and T3-like genotypes). The second assay contained primers and probes for Hop stunt viroid (HSVd), Citrus exocortis viroid (CEVd) and the mitochondrial NADH dehydrogenase (nad5) mRNA as an internal citrus host control. Primers and TaqMan probes for the viroids were designed in this work; whereas those for the other pathogens were from reports of others. Based on quantitation cycle values, automated high-throughput extraction of samples proved to be as suitable as manual extraction. The multiplex RT-qPCR assays detected both RNA and DNA pathogens in the same dilution series

  11. Using Extreme Value Theory Approaches to Forecast the Probability of Outbreak of Highly Pathogenic Influenza in Zhejiang, China

    PubMed Central

    Chen, Jiangpeng; Lei, Xun; Zhang, Li; Peng, Bin

    2015-01-01

    Background Influenza is a contagious disease with high transmissibility to spread around the world with considerable morbidity and mortality and presents an enormous burden on worldwide public health. Few mathematical models can be used because influenza incidence data are generally not normally distributed. We developed a mathematical model using Extreme Value Theory (EVT) to forecast the probability of outbreak of highly pathogenic influenza. Methods The incidence data of highly pathogenic influenza in Zhejiang province from April 2009 to November 2013 were retrieved from the website of Health and Family Planning Commission of Zhejiang Province. MATLAB “VIEM” toolbox was used to analyze data and modelling. In the present work, we used the Peak Over Threshold (POT) model, assuming the frequency as a Poisson process and the intensity to be Pareto distributed, to characterize the temporal variability of the long-term extreme incidence of highly pathogenic influenza in Zhejiang, China. Results The skewness and kurtosis of the incidence of highly pathogenic influenza in Zhejiang between April 2009 and November 2013 were 4.49 and 21.12, which indicated a “fat tail” distribution. A QQ plot and a mean excess plot were used to further validate the features of the distribution. After determining the threshold, we modeled the extremes and estimated the shape parameter and scale parameter by the maximum likelihood method. The results showed that months in which the incidence of highly pathogenic influenza is about 4462/2286/1311/487 are predicted to occur once every five/three/two/one year, respectively. Conclusions Despite the simplicity, the present study successfully offers the sound modeling strategy and a methodological avenue to implement forecasting of an epidemic in the midst of its course. PMID:25710503

  12. Lab-on-a-chip modules for detection of highly pathogenic bacteria: from sample preparation to detection

    NASA Astrophysics Data System (ADS)

    Julich, S.; Kopinč, R.; Hlawatsch, N.; Moche, C.; Lapanje, A.; Gärtner, C.; Tomaso, H.

    2014-05-01

    Lab-on-a-chip systems are innovative tools for the detection and identification of microbial pathogens in human and veterinary medicine. The major advantages are small sample volume and a compact design. Several fluidic modules have been developed to transform analytical procedures into miniaturized scale including sampling, sample preparation, target enrichment, and detection procedures. We present evaluation data for single modules that will be integrated in a chip system for the detection of pathogens. A microfluidic chip for purification of nucleic acids was established for cell lysis using magnetic beads. This assay was evaluated with spiked environmental aerosol and swab samples. Bacillus thuringiensis was used as simulant for Bacillus anthracis, which is closely related but non-pathogenic for humans. Stationary PCR and a flow-through PCR chip module were investigated for specific detection of six highly pathogenic bacteria. The conventional PCR assays could be transferred into miniaturized scale using the same temperature/time profile. We could demonstrate that the microfluidic chip modules are suitable for the respective purposes and are promising tools for the detection of bacterial pathogens. Future developments will focus on the integration of these separate modules to an entire lab-on-a-chip system.

  13. Genetic evolution of H5 highly pathogenic avian influenza virus in domestic poultry in Vietnam between 2011 and 2013.

    PubMed

    Lee, Eun-Kyoung; Kang, Hyun-Mi; Kim, Kwang-Il; Choi, Jun-Gu; To, Thanh Long; Nguyen, Tho Dang; Song, Byung-Min; Jeong, Jipseol; Choi, Kang-Seuk; Kim, Ji-Ye; Lee, Hee-Soo; Lee, Youn-Jeong; Kim, Jae-Hong

    2015-04-01

    In spite of highly pathogenic avian influenza H5N1 vaccination campaigns for domestic poultry, H5N1 viruses continue to circulate in Vietnam. To estimate the prevalence of avian influenza virus in Vietnam, surveillance was conducted between November 2011 and February 2013. Genetic analysis of 312 highly pathogenic avian influenza H5 viruses isolated from poultry in Vietnam was conducted and possible genetic relationships with strains from neighboring countries were investigated. As previously reported, phylogenetic analysis of the avian influenza virus revealed two H5N1 HPAI clades that were circulating in Vietnam. Clade 1.1, related to Cambodian strains, was predominant in the southern provinces, while clade 2.3.2.1 viruses were predominant in the northern and central provinces. Sequence analysis revealed evidence of active genetic evolution. In the gene constellation of clade 2.3.2.1, genotypes A, B, and B(II) existed during the 2011/2012 winter season. In June 2012, new genotype C emerged by reassortment between genotype A and genotype B(II), and this genotype was predominant in 2013 in the northern and central provinces. Interestingly, enzootic Vietnamese clade 2.3.2.1C H5 virus subsequently reassorted with N2, which originated from wild birds, to generate H5N2 highly pathogenic avian influenza, which was isolated from duck in the northeast region. This investigation indicated that H5N1 outbreaks persist in Vietnam and cause genetic reassortment with circulating viruses. It is necessary to strengthen active influenza surveillance to eradicate highly pathogenic avian influenza viruses and sever the link between highly pathogenic avian influenza and other circulating influenza viruses.

  14. Intersubtype Reassortments of H5N1 Highly Pathogenic Avian Influenza Viruses Isolated from Quail.

    PubMed

    Nguyen, Tinh Huu; Than, Van Thai; Thanh, Hien Dang; Hung, Vu-Khac; Nguyen, Duc Tan; Kim, Wonyong

    2016-01-01

    H5N1 highly pathogenic avian influenza (HPAI) viruses are considered a threat to national animal industries, causing production losses and high mortality in domestic poultry. In recent years, quail has become a popular terrestrial poultry species raised for production of meat and eggs in Asia. In this study, to better understand the roles of quail in H5N1 viral evolution, two H5N1-positive samples, designated A/quail/Vietnam/CVVI-49/2010 (CVVI-49/2010) and A/quail/Vietnam/CVVI-50/2014 (CVVI-50/2014), were isolated from quail during H5N1 outbreaks in Vietnam, and their whole genome were analyzed. The phylogenetic analysis reveals new evolutionary variation in the worldwide H5N1 viruses. The quail HA genes were clustered into clades 1.1.1 (CVVI-49/2010) and clade 2.3.2.1c (CVVI-50/2014), which may have evolved from viruses circulating from chickens and/or ducks in Cambodia, mainland of China, Taiwan, Indonesia, and South Korea in recent years. Interestingly, the M2 gene of the CVVI-49/2010 strain contained amino acid substitutions at position 26L-I and 31S-N that are related to amantadine-resistance. In particular, the CVVI-50/2014 strain revealed evidence of multiple intersubtype reassortment events between virus clades 2.3.2.1c, 2.3.2.1b, and 2.3.2.1a. Data from this study supports the possible role of quail as an important intermediate host in avian influenza virus evolution. Therefore, additional surveillance is needed to monitor these HPAI viruses both serologically and virologically in quail. PMID:26900963

  15. Highly Pathogenic Influenza A(H5N1) Virus Survival in Complex Artificial Aquatic Biotopes

    PubMed Central

    Horm, Viseth Srey; Gutiérrez, Ramona A.; Nicholls, John M.; Buchy, Philippe

    2012-01-01

    Background Very little is known regarding the persistence of Highly Pathogenic Avian Influenza (HPAI) H5N1 viruses in aquatic environments in tropical countries, although environmental materials have been suggested to play a role as reservoirs and sources of transmission for H5N1 viruses. Methodology/Principal Findings The survival of HPAI H5N1 viruses in experimental aquatic biotopes (water, mud, aquatic flora and fauna) relevant to field conditions in Cambodia was investigated. Artificial aquatic biotopes, including simple ones containing only mud and water, and complex biotopes involving the presence of aquatic flora and fauna, were set up. They were experimentally contaminated with H5N1 virus. The persistence of HPAI H5N1 virus (local avian and human isolates) was determined by virus isolation in embryonated chicken eggs and by real-time reverse-polymerase chain reaction. Persistence of infectious virus did not exceed 4 days, and was only identified in rain water. No infectious virus particles were detected in pond and lake water or mud even when high inoculum doses were used. However, viral RNA persisted up to 20 days in rain water and 7 days in pond or lake water. Viral RNA was also detected in mud samples, up to 14 days post-contamination in several cases. Infectious virus and viral RNA was detected in few cases in the aquatic fauna and flora, especially in bivalves and labyrinth fish, although these organisms seemed to be mostly passive carriers of the virus rather than host allowing virus replication. Conclusions/Significance Although several factors for the survival and persistence of HPAI viruses in the environment are still to be elucidated, and are particularly hard to control in laboratory conditions, our results, along with previous data, support the idea that environmental surveillance is of major relevance for avian influenza control programs. PMID:22514622

  16. Intersubtype Reassortments of H5N1 Highly Pathogenic Avian Influenza Viruses Isolated from Quail.

    PubMed

    Nguyen, Tinh Huu; Than, Van Thai; Thanh, Hien Dang; Hung, Vu-Khac; Nguyen, Duc Tan; Kim, Wonyong

    2016-01-01

    H5N1 highly pathogenic avian influenza (HPAI) viruses are considered a threat to national animal industries, causing production losses and high mortality in domestic poultry. In recent years, quail has become a popular terrestrial poultry species raised for production of meat and eggs in Asia. In this study, to better understand the roles of quail in H5N1 viral evolution, two H5N1-positive samples, designated A/quail/Vietnam/CVVI-49/2010 (CVVI-49/2010) and A/quail/Vietnam/CVVI-50/2014 (CVVI-50/2014), were isolated from quail during H5N1 outbreaks in Vietnam, and their whole genome were analyzed. The phylogenetic analysis reveals new evolutionary variation in the worldwide H5N1 viruses. The quail HA genes were clustered into clades 1.1.1 (CVVI-49/2010) and clade 2.3.2.1c (CVVI-50/2014), which may have evolved from viruses circulating from chickens and/or ducks in Cambodia, mainland of China, Taiwan, Indonesia, and South Korea in recent years. Interestingly, the M2 gene of the CVVI-49/2010 strain contained amino acid substitutions at position 26L-I and 31S-N that are related to amantadine-resistance. In particular, the CVVI-50/2014 strain revealed evidence of multiple intersubtype reassortment events between virus clades 2.3.2.1c, 2.3.2.1b, and 2.3.2.1a. Data from this study supports the possible role of quail as an important intermediate host in avian influenza virus evolution. Therefore, additional surveillance is needed to monitor these HPAI viruses both serologically and virologically in quail.

  17. Intersubtype Reassortments of H5N1 Highly Pathogenic Avian Influenza Viruses Isolated from Quail

    PubMed Central

    Nguyen, Tinh Huu; Than, Van Thai; Thanh, Hien Dang; Hung, Vu-Khac; Nguyen, Duc Tan; Kim, Wonyong

    2016-01-01

    H5N1 highly pathogenic avian influenza (HPAI) viruses are considered a threat to national animal industries, causing production losses and high mortality in domestic poultry. In recent years, quail has become a popular terrestrial poultry species raised for production of meat and eggs in Asia. In this study, to better understand the roles of quail in H5N1 viral evolution, two H5N1-positive samples, designated A/quail/Vietnam/CVVI-49/2010 (CVVI-49/2010) and A/quail/Vietnam/CVVI-50/2014 (CVVI-50/2014), were isolated from quail during H5N1 outbreaks in Vietnam, and their whole genome were analyzed. The phylogenetic analysis reveals new evolutionary variation in the worldwide H5N1 viruses. The quail HA genes were clustered into clades 1.1.1 (CVVI-49/2010) and clade 2.3.2.1c (CVVI-50/2014), which may have evolved from viruses circulating from chickens and/or ducks in Cambodia, mainland of China, Taiwan, Indonesia, and South Korea in recent years. Interestingly, the M2 gene of the CVVI-49/2010 strain contained amino acid substitutions at position 26L-I and 31S-N that are related to amantadine-resistance. In particular, the CVVI-50/2014 strain revealed evidence of multiple intersubtype reassortment events between virus clades 2.3.2.1c, 2.3.2.1b, and 2.3.2.1a. Data from this study supports the possible role of quail as an important intermediate host in avian influenza virus evolution. Therefore, additional surveillance is needed to monitor these HPAI viruses both serologically and virologically in quail. PMID:26900963

  18. Evolution of highly pathogenic avian H5N1 influenza viruses

    SciTech Connect

    Macken, Catherine A; Green, Margaret A

    2009-01-01

    Highly pathogenic avian H5N1 viruses have circulated in Southeast Asia for more than a decade, are now endemic in parts of this region, and have also spread to more than 60 countries on three continents. The evolution of these viruses is characterized by frequent reassortment events that have created a significant number of different genotypes, both transient and longer lasting. However, fundamental questions remain about the generation and perpetuation of this substantial genetic diversity. These gaps in understanding may, in part, be due to the difficulties of genotyping closely related viruses, and limitations in the size of the data sets used in analysis. Using our recently published novel genotyping procedure ('two-time test'), which is amenable to high throughput analysis and provides an increased level of resolution relative to previous analyses, we propose a detailed model for the evolution and diversification of avian H5N1 viruses. Our analysis suggests that (i) all current H5N1 genotypes are derived from a single, clearly defined sequence of initial reassortment events; (ii) reassortment of the polymerase and NP genes may have played an important role in avian H5N1 virus evolution; (iii) the current genotype Z viruses have diverged into three distinguishable sub-genotypes in the absence of reassortment; (iv) some potentially significant molecular changes appear to be correlated with particular genotypes (for example, reassortment of the internal genes is often paralleled by a change in the HA clade); and (v) as noted in earlier studies of avian influenza A virus evolution, novel segments are typically derived from different donors (i.e., there is no obvious pattern of gene linkage in reassortment). The model of avian H5N1 viral evolution by reassortment and mutation that emerges from our study provides a context within which significant amino acid changes may be revealed; it also may help in predicting the 'success' of newly emerging avian H5N1 viruses.

  19. Evidence of infection by H5N2 highly pathogenic avian influenza viruses in healthy wild waterfowl

    USGS Publications Warehouse

    Gaidet, N.; Cattoli, G.; Hammoumi, S.; Newman, S.H.; Hagemeijer, W.; Takekawa, J.Y.; Cappelle, J.; Dodman, T.; Joannis, T.; Gil, P.; Monne, I.; Fusaro, A.; Capua, I.; Manu, S.; Micheloni, P.; Ottosson, U.; Mshelbwala, J.H.; Lubroth, J.; Domenech, J.; Monicat, F.

    2008-01-01

    The potential existence of a wild bird reservoir for highly pathogenic avian influenza (HPAI) has been recently questioned by the spread and the persisting circulation of H5N1 HPAI viruses, responsible for concurrent outbreaks in migratory and domestic birds over Asia, Europe, and Africa. During a large-scale surveillance programme over Eastern Europe, the Middle East, and Africa, we detected avian influenza viruses of H5N2 subtype with a highly pathogenic (HP) viral genotype in healthy birds of two wild waterfowl species sampled in Nigeria. We monitored the survival and regional movements of one of the infected birds through satellite telemetry, providing a rare evidence of a non-lethal natural infection by an HP viral genotype in wild birds. Phylogenetic analysis of the H5N2 viruses revealed close genetic relationships with H5 viruses of low pathogenicity circulating in Eurasian wild and domestic ducks. In addition, genetic analysis did not reveal known gallinaceous poultry adaptive mutations, suggesting that the emergence of HP strains could have taken place in either wild or domestic ducks or in non-gallinaceous species. The presence of coexisting but genetically distinguishable avian influenza viruses with an HP viral genotype in two cohabiting species of wild waterfowl, with evidence of non-lethal infection at least in one species and without evidence of prior extensive circulation of the virus in domestic poultry, suggest that some strains with a potential high pathogenicity for poultry could be maintained in a community of wild waterfowl.

  20. Characterization of low-pathogenic H5 subtype influenza viruses from Eurasia: implications for the origin of highly pathogenic H5N1 viruses.

    PubMed

    Duan, L; Campitelli, L; Fan, X H; Leung, Y H C; Vijaykrishna, D; Zhang, J X; Donatelli, I; Delogu, M; Li, K S; Foni, E; Chiapponi, C; Wu, W L; Kai, H; Webster, R G; Shortridge, K F; Peiris, J S M; Smith, Gavin J D; Chen, H; Guan, Y

    2007-07-01

    Highly pathogenic avian influenza (HPAI) H5N1 viruses are now endemic in many Asian countries, resulting in repeated outbreaks in poultry and increased cases of human infection. The immediate precursor of these HPAI viruses is believed to be A/goose/Guangdong/1/96 (Gs/GD)-like H5N1 HPAI viruses first detected in Guangdong, China, in 1996. From 2000 onwards, many novel reassortant H5N1 influenza viruses or genotypes have emerged in southern China. However, precursors of the Gs/GD-like viruses and their subsequent reassortants have not been fully determined. Here we characterize low-pathogenic avian influenza (LPAI) H5 subtype viruses isolated from poultry and migratory birds in southern China and Europe from the 1970s to the 2000s. Phylogenetic analyses revealed that Gs/GD-like virus was likely derived from an LPAI H5 virus in migratory birds. However, its variants arose from multiple reassortments between Gs/GD-like virus and viruses from migratory birds or with those Eurasian viruses isolated in the 1970s. It is of note that unlike HPAI H5N1 viruses, those recent LPAI H5 viruses have not become established in aquatic or terrestrial poultry. Phylogenetic analyses revealed the dynamic nature of the influenza virus gene pool in Eurasia with repeated transmissions between the eastern and western extremities of the continent. The data also show reassortment between influenza viruses from domestic and migratory birds in this region that has contributed to the expanded diversity of the influenza virus gene pool among poultry in Eurasia.

  1. Efficacy of direct detection of pathogens in naturally infected mice by using a high-density PCR array.

    PubMed

    Henderson, Kenneth S; Perkins, Cheryl L; Havens, Richard B; Kelly, Mee-Jin E; Francis, Brian C; Dole, Vandana S; Shek, William R

    2013-11-01

    We used a high-density array of real-time PCR assays for commonly reported rodent infectious agents (PRIA) to test naturally infected index mice and sentinel mice exposed by contact and soiled-bedding transfer. PRIA detected 14 pathogens--including viruses, bacteria, fur mites, pinworms, and enteric protozoa--in 97.2% of 28 pooled fecal samples, fur-perianal swabs, and oral swabs from 4 cages containing a total of 10 index mice. Among these pathogens, PRIA (like conventional health monitoring methods) failed to detect Mycoplasma pulmonis, Pasteurella pneumotropica, and Giardia spp. in all of the 9 contact and 9 soiled-bedding sentinels. PRIA demonstrated murine adenovirus and Cryptosporidium and Spironucleus spp. in contact but not soiled-bedding sentinels and detected Helicobacter and pinworms in fewer than half of the soiled-bedding sentinels. Of the 4 species of Helicobacter that species-specific PCR assays identified in index mice, only H. ganmani was found in soiled-bedding and contact sentinels. PRIA detected all of the pathogens in sentinels that were identified by conventional methods. Myobia musculi was detected by PCR in index and sentinel mice but missed by conventional parasitologic examinations. In summary, PRIA reproducibly detected diverse pathogens in heavily pooled specimens collected noninvasively from infected index mice antemortem. The inability of PRIA and conventional health monitoring methods (that is, parasitology, micro-biology, and serology) to demonstrate transmission of some pathogens to contact sentinels and the inefficient transmission of others to soiled-bedding sentinels underscores the importance of direct PCR testing to determine the pathogen status of rodents in quarantine and during routine colony surveillance. PMID:24351765

  2. Molecular pathogenesis of H5 highly pathogenic avian influenza: the role of the haemagglutinin cleavage site motif.

    PubMed

    Luczo, Jasmina M; Stambas, John; Durr, Peter A; Michalski, Wojtek P; Bingham, John

    2015-11-01

    The emergence of H5N1 highly pathogenic avian influenza has caused a heavy socio-economic burden through culling of poultry to minimise human and livestock infection. Although human infections with H5N1 have to date been limited, concerns for the pandemic potential of this zoonotic virus have been greatly intensified following experimental evidence of aerosol transmission of H5N1 viruses in a mammalian infection model. In this review, we discuss the dominance of the haemagglutinin cleavage site motif as a pathogenicity determinant, the host-pathogen molecular interactions driving cleavage activation, reverse genetics manipulations and identification of residues key to haemagglutinin cleavage site functionality and the mechanisms of cell and tissue damage during H5N1 infection. We specifically focus on the disease in chickens, as it is in this species that high pathogenicity frequently evolves and from which transmission to the human population occurs. With >75% of emerging infectious diseases being of zoonotic origin, it is necessary to understand pathogenesis in the primary host to explain spillover events into the human population. PMID:26467906

  3. Molecular pathogenesis of H5 highly pathogenic avian influenza: the role of the haemagglutinin cleavage site motif

    PubMed Central

    Luczo, Jasmina M.; Stambas, John; Durr, Peter A.; Michalski, Wojtek P.

    2015-01-01

    Summary The emergence of H5N1 highly pathogenic avian influenza has caused a heavy socio‐economic burden through culling of poultry to minimise human and livestock infection. Although human infections with H5N1 have to date been limited, concerns for the pandemic potential of this zoonotic virus have been greatly intensified following experimental evidence of aerosol transmission of H5N1 viruses in a mammalian infection model. In this review, we discuss the dominance of the haemagglutinin cleavage site motif as a pathogenicity determinant, the host‐pathogen molecular interactions driving cleavage activation, reverse genetics manipulations and identification of residues key to haemagglutinin cleavage site functionality and the mechanisms of cell and tissue damage during H5N1 infection. We specifically focus on the disease in chickens, as it is in this species that high pathogenicity frequently evolves and from which transmission to the human population occurs. With >75% of emerging infectious diseases being of zoonotic origin, it is necessary to understand pathogenesis in the primary host to explain spillover events into the human population. © 2015 The Authors. Reviews in Medical Virology published by John Wiley & Sons Ltd. PMID:26467906

  4. Continuing Reassortant of H5N6 Subtype Highly Pathogenic Avian Influenza Virus in Guangdong.

    PubMed

    Yuan, Runyu; Wang, Zheng; Kang, Yinfeng; Wu, Jie; Zou, Lirong; Liang, Lijun; Song, Yingchao; Zhang, Xin; Ni, Hanzhong; Lin, Jinyan; Ke, Changwen

    2016-01-01

    First identified in May 2014 in China's Sichuan Province, initial cases of H5N6 avian influenza virus (AIV) infection in humans raised great concerns about the virus's prevalence, origin, and development. To evaluate both AIV contamination in live poultry markets (LPMs) and the risk of AIV infection in humans, we have conducted surveillance of LPMs in Guangdong Province since 2013 as part of environmental sampling programs. With environmental samples associated with these LPMs, we performed genetic and phylogenetic analyses of 10 H5N6 AIVs isolated from different cities of Guangdong Province from different years. Results revealed that the H5N6 viruses were reassortants with hemagglutinin (HA) genes derived from clade 2.3.4.4 of H5-subtype AIV, yet neuraminidase (NA) genes derived from H6N6 AIV. Unlike the other seven H5N6 viruses isolated in first 7 months of 2014, all of which shared remarkable sequence similarity with the H5N1 AIV in all internal genes, the PB2 genes of GZ693, GZ670, and ZS558 more closely related to H6N6 AIV and the PB1 gene of GZ693 to the H3-subtype AIV. Phylogenetic analyses revealed that the environmental H5N6 AIV related closely to human H5N6 AIVs isolated in Guangdong. These results thus suggest that continued reassortment has enabled the emergence of a novel H5N6 virus in Guangdong, as well as highlight the potential risk of highly pathogenic H5N6 AIVs in the province.

  5. Evolution of highly pathogenic avian influenza H5N1 viruses in Egypt indicating progressive adaptation.

    PubMed

    Arafa, A; Suarez, D; Kholosy, S G; Hassan, M K; Nasef, S; Selim, A; Dauphin, G; Kim, M; Yilma, J; Swayne, D; Aly, M M

    2012-10-01

    Highly pathogenic avian influenza (HPAI) virus of the H5N1 subtype was first diagnosed in poultry in Egypt in 2006, and since then the disease became enzootic in poultry throughout the country, affecting the poultry industry and village poultry as well as infecting humans. Vaccination has been used as a part of the control strategy to help to control the disease. Epidemiological data with sequence analysis of H5N1 viruses is important to link the mechanism of virus evolution in Egypt. This study describes the evolutionary pattern of Egyptian H5N1 viruses based on molecular characterization for the isolates collected from commercial poultry farms and village poultry from 2006 to 2011. Genetic analysis of the hemagglutinin (HA) gene was done by sequencing of the full-length H5 gene. The epidemiological pattern of disease outbreaks in Egyptian poultry farms seems to be seasonal with no specific geographic distribution across the country. The molecular epidemiological data revealed that there are two major groups of viruses: the classic group of subclade 2.2.1 and a variant group of 2.2.1.1. The classic group is prevailing mainly in village poultry and had fewer mutations compared to the originally introduced virus in 2006. Since 2009, this group has started to be transmitted back to commercial sectors. The variant group emerged by late 2007, was prevalent mainly in vaccinated commercial poultry, mutated continuously at a higher rate until 2010, and started to decline in 2011. Genetic analysis of the neuraminidase (NA) gene and the other six internal genes indicates a grouping of the Egyptian viruses similar to that obtained using the HA gene, with no obvious reassortments. The results of this study indicate that HPAI-H5N1 viruses are progressively evolving and adapting in Egypt and continue to acquire new mutations every season. PMID:22760662

  6. Depression as sickness behavior? A test of the host defense hypothesis in a high pathogen population.

    PubMed

    Stieglitz, Jonathan; Trumble, Benjamin C; Thompson, Melissa Emery; Blackwell, Aaron D; Kaplan, Hillard; Gurven, Michael

    2015-10-01

    Sadness is an emotion universally recognized across cultures, suggesting it plays an important functional role in regulating human behavior. Numerous adaptive explanations of persistent sadness interfering with daily functioning (hereafter "depression") have been proposed, but most do not explain frequent bidirectional associations between depression and greater immune activation. Here we test several predictions of the host defense hypothesis, which posits that depression is part of a broader coordinated evolved response to infection or tissue injury (i.e. "sickness behavior") that promotes energy conservation and reallocation to facilitate immune activation. In a high pathogen population of lean and relatively egalitarian Bolivian forager-horticulturalists, we test whether depression and its symptoms are associated with greater baseline concentration of immune biomarkers reliably associated with depression in Western populations (i.e. tumor necrosis factor alpha [TNF-α], interleukin-1 beta [IL-1β], interleukin-6 [IL-6], and C-reactive protein [CRP]). We also test whether greater pro-inflammatory cytokine responses to ex vivo antigen stimulation are associated with depression and its symptoms, which is expected if depression facilitates immune activation. These predictions are largely supported in a sample of older adult Tsimane (mean±SD age=53.2±11.0, range=34-85, n=649) after adjusting for potential confounders. Emotional, cognitive and somatic symptoms of depression are each associated with greater immune activation, both at baseline and in response to ex vivo stimulation. The association between depression and greater immune activation is therefore not unique to Western populations. While our findings are not predicted by other adaptive hypotheses of depression, they are not incompatible with those hypotheses and future research is necessary to isolate and test competing predictions. PMID:26044086

  7. Continuing Reassortant of H5N6 Subtype Highly Pathogenic Avian Influenza Virus in Guangdong

    PubMed Central

    Yuan, Runyu; Wang, Zheng; Kang, Yinfeng; Wu, Jie; Zou, Lirong; Liang, Lijun; Song, Yingchao; Zhang, Xin; Ni, Hanzhong; Lin, Jinyan; Ke, Changwen

    2016-01-01

    First identified in May 2014 in China's Sichuan Province, initial cases of H5N6 avian influenza virus (AIV) infection in humans raised great concerns about the virus's prevalence, origin, and development. To evaluate both AIV contamination in live poultry markets (LPMs) and the risk of AIV infection in humans, we have conducted surveillance of LPMs in Guangdong Province since 2013 as part of environmental sampling programs. With environmental samples associated with these LPMs, we performed genetic and phylogenetic analyses of 10 H5N6 AIVs isolated from different cities of Guangdong Province from different years. Results revealed that the H5N6 viruses were reassortants with hemagglutinin (HA) genes derived from clade 2.3.4.4 of H5-subtype AIV, yet neuraminidase (NA) genes derived from H6N6 AIV. Unlike the other seven H5N6 viruses isolated in first 7 months of 2014, all of which shared remarkable sequence similarity with the H5N1 AIV in all internal genes, the PB2 genes of GZ693, GZ670, and ZS558 more closely related to H6N6 AIV and the PB1 gene of GZ693 to the H3-subtype AIV. Phylogenetic analyses revealed that the environmental H5N6 AIV related closely to human H5N6 AIVs isolated in Guangdong. These results thus suggest that continued reassortment has enabled the emergence of a novel H5N6 virus in Guangdong, as well as highlight the potential risk of highly pathogenic H5N6 AIVs in the province. PMID:27148209

  8. MALDI-TOF mass spectrometry compatible inactivation method for highly pathogenic microbial cells and spores.

    PubMed

    Lasch, Peter; Nattermann, Herbert; Erhard, Marcel; Stämmler, Maren; Grunow, Roland; Bannert, Norbert; Appel, Bernd; Naumann, Dieter

    2008-03-15

    Identification of microorganisms, specifically of vegetative cells and spores, by intact cell mass spectrometry (ICMS) is an emerging new technology. The technique provides specific biomarker profiles which can be employed for bacterial identification at the genus, species, or even at the subspecies level holding the potential to serve as a rapid and sensitive identification technique in clinical or food microbiology and also for sensitive detection of biosafety level (BSL) 3 microorganisms. However, the development of ICMS as an identification technique for BSL-3 level microorganisms is hampered by the fact that no MALDI-TOF (matrix-assisted laser desorption/ionization time-of-flight) compatible inactivation procedure for microorganisms, and particularly for bacterial endospores, has been evaluated so far. In this report we describe a new methodology for effective inactivation of microorganisms which is compatible with the analysis of microbial protein patterns by MALDI-TOF mass spectrometry. The main challenge of this work was to define the conditions that ensure microbial inactivation and permit at the same time comprehensive analysis of microbial protein patterns. Among several physical, chemical, and mechanical inactivation procedures, inactivation by trifluoroacetic acid (TFA) proved to be the best method in terms of bactericidal capacity and information content of the mass spectra. Treatment of vegetative cells by 80% TFA alone for 30 min assured complete inactivation of microbial cells under all conditions tested. For spore inactivation, the "TFA inactivation protocol" was developed which is a combination of TFA treatment with basic laboratory routines such as centrifugation and filtering. This MALDI-TOF/ICMS compatible sample preparation protocol is simple and rapid (30 min) and assures reliable inactivation of vegetative cells and spores of highly pathogenic (BSL-3) microorganisms.

  9. DEPRESSION AS SICKNESS BEHAVIOR? A TEST OF THE HOST DEFENSE HYPOTHESIS IN A HIGH PATHOGEN POPULATION

    PubMed Central

    Stieglitz, Jonathan; Trumble, Benjamin C.; Thompson, Melissa Emery; Blackwell, Aaron D.; Kaplan, Hillard; Gurven, Michael

    2015-01-01

    Sadness is an emotion universally recognized across cultures, suggesting it plays an important functional role in regulating human behavior. Numerous adaptive explanations of persistent sadness interfering with daily functioning (hereafter “depression”) have been proposed, but most do not explain frequent bidirectional associations between depression and greater immune activation. Here we test several predictions of the host defense hypothesis, which posits that depression is part of a broader coordinated evolved response to infection or tissue injury (i.e. “sickness behavior”) that promotes energy conservation and reallocation to facilitate immune activation. In a high pathogen population of lean and relatively egalitarian Bolivian foragerhorticulturalists, we test whether depression and its symptoms are associated with greater baseline concentration of immune biomarkers reliably associated with depression in Western populations (i.e. tumor necrosis factor alpha [TNF-α], interleukin-1 beta [IL-1β], interleukin-6 [IL-6], and C-reactive protein [CRP]). We also test whether greater pro-inflammatory cytokine responses to ex vivo antigen stimulation are associated with depression and its symptoms, which is expected if depression facilitates immune activation. These predictions are largely supported in a sample of older adult Tsimane (mean±SD age=53.2±11.0, range=34-85, n=649) after adjusting for potential confounders. Emotional, cognitive and somatic symptoms of depression are each associated with greater immune activation, both at baseline and in response to ex vivo stimulation. The association between depression and greater immune activation is therefore not unique to Western populations. While our findings are not predicted by other adaptive hypotheses of depression, they are not incompatible with those hypotheses and future research is necessary to isolate and test competing predictions. PMID:26044086

  10. Highly Pathogenic Avian Influenza H5N1 in Mainland China.

    PubMed

    Li, Xin-Lou; Liu, Kun; Yao, Hong-Wu; Sun, Ye; Chen, Wan-Jun; Sun, Ruo-Xi; de Vlas, Sake J; Fang, Li-Qun; Cao, Wu-Chun

    2015-05-08

    Highly pathogenic avian influenza (HPAI) H5N1 has posed a significant threat to both humans and birds, and it has spanned large geographic areas and various ecological systems throughout Asia, Europe and Africa, but especially in mainland China. Great efforts in control and prevention of the disease, including universal vaccination campaigns in poultry and active serological and virological surveillance, have been undertaken in mainland China since the beginning of 2006. In this study, we aim to characterize the spatial and temporal patterns of HPAI H5N1, and identify influencing factors favoring the occurrence of HPAI H5N1 outbreaks in poultry in mainland China. Our study shows that HPAI H5N1 outbreaks took place sporadically after vaccination campaigns in poultry, and mostly occurred in the cold season. The positive tests in routine virological surveillance of HPAI H5N1 virus in chicken, duck, goose as well as environmental samples were mapped to display the potential risk distribution of the virus. Southern China had a higher positive rate than northern China, and positive samples were mostly detected from chickens in the north, while the majority were from duck in the south, and a negative correlation with monthly vaccination rates in domestic poultry was found (R = -0.19, p value = 0.005). Multivariate panel logistic regression identified vaccination rate, interaction between distance to the nearest city and national highway, interaction between distance to the nearest lake and wetland, and density of human population, as well as the autoregressive term in space and time as independent risk factors in the occurrence of HPAI H5N1 outbreaks, based on which a predicted risk map of the disease was derived. Our findings could provide new understanding of the distribution and transmission of HPAI H5N1 in mainland China and could be used to inform targeted surveillance and control efforts in both human and poultry populations to reduce the risk of future infections.

  11. Rapid detection of fungal pathogens in bronchoalveolar lavage samples using panfungal PCR combined with high resolution melting analysis.

    PubMed

    Bezdicek, Matej; Lengerova, Martina; Ricna, Dita; Weinbergerova, Barbora; Kocmanova, Iva; Volfova, Pavlina; Drgona, Lubos; Poczova, Miroslava; Mayer, Jiri; Racil, Zdenek

    2016-10-01

    Despite advances in the treatment of invasive fungal diseases (IFD), mortality rates remain high. Moreover, due to the expanding spectrum of causative agents, fast and accurate pathogen identification is necessary. We designed a panfungal polymerase chain reaction (PCR), which targets the highly variable ITS2 region of rDNA genes and uses high resolution melting analysis (HRM) for subsequent species identification. The sensitivity and specificity of this method was tested on a broad spectrum of the most clinically important fungal pathogens including Aspergillus spp., Candida spp. and mucormycetes. Despite the fact that fluid from bronchoalveolar lavage (BAL) is one of the most frequently tested materials there is a lack of literature sources aimed at panfungal PCR as an IFD diagnostic tool from BAL samples. The applicability of this method in routine practice was evaluated on 104 BAL samples from immunocompromised patients. Due to high ITS region variability, we obtained divergent melting peaks for different fungal species. Thirteen out of 18 patients with proven or probable IFD were positive. Therefore, the sensitivity, specificity, positive predictive value and negative predictive value of our method were 67%, 100%, 100%, and 94%, respectively. In our assay, fungal pathogens identification is based on HRM, therefore omitting the expensive and time consuming sequencing step. With the high specificity, positive and negative predictive values, short time needed to obtain a result, and low price, the presented assay is intended to be used as a quick screening method for patients at risk of IFD. PMID:27161789

  12. Identification of highly variable supernumerary chromosome segments in an asexual pathogen

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Supernumerary chromosome segments are known to harbor different transposons from their essential counterparts. The aim of this study was to investigate the role of transposons in the origin and evolution of supernumerary segments in the asexual fungal pathogen Fusariumvirguliforme. We compared the g...

  13. Major Histocompatibility Complex and Background Genes in Chickens Influence Susceptibility to High Pathogenicity Avian Influenza Virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The chicken’s major histocompatibility complex (MHC) haplotype has profound influence on the resistance or susceptibility to certain pathogens such as B21 MHC haplotype confers resistance to Marek’s disease (MD). However, non-MHC genes are also important in disease resistance. For example, both line...

  14. Expansion of Variant Diversity Associated with a High Prevalence of Pathogen Strain Superinfection under Conditions of Natural Transmission

    PubMed Central

    Ueti, Massaro W.; Tan, Yunbing; Broschat, Shira L.; Castañeda Ortiz, Elizabeth J.; Camacho-Nuez, Minerva; Mosqueda, Juan J.; Scoles, Glen A.; Grimes, Matthew; Brayton, Kelly A.

    2012-01-01

    Superinfection occurs when a second, genetically distinct pathogen strain infects a host that has already mounted an immune response to a primary strain. For antigenically variant pathogens, the primary strain itself expresses a broad diversity of variants over time. Thus, successful superinfection would require that the secondary strain express a unique set of variants. We tested this hypothesis under conditions of natural transmission in both temperate and tropical regions where, respectively, single-strain infections and strain superinfections of the tick-borne pathogen Anaplasma marginale predominate. Our conclusion that strain superinfection is associated with a significant increase in variant diversity is supported by progressive analysis of variant composition: (i) animals with naturally acquired superinfection had a statistically significantly greater number of unique variant sequences than animals either experimentally infected with single strains or infected with a single strain naturally, (ii) the greater number of unique sequences reflected a statistically significant increase in primary structural diversity in the superinfected animals, and (iii) the increase in primary structural diversity reflected increased combinations of the newly identified hypervariable microdomains. The role of population immunity in establishing temporal and spatial patterns of infection and disease has been well established. The results of the present study, which examined strain structure under conditions of natural transmission and population immunity, support that high levels of endemicity also drive pathogen divergence toward greater strain diversity. PMID:22585962

  15. Glycan:glycan interactions: High affinity biomolecular interactions that can mediate binding of pathogenic bacteria to host cells

    PubMed Central

    Day, Christopher J.; Tran, Elizabeth N.; Semchenko, Evgeny A.; Tram, Greg; Hartley-Tassell, Lauren E.; Ng, Preston S. K.; King, Rebecca M.; Ulanovsky, Rachel; McAtamney, Sarah; Apicella, Michael A.; Tiralongo, Joe; Morona, Renato; Korolik, Victoria; Jennings, Michael P.

    2015-01-01

    Cells from all domains of life express glycan structures attached to lipids and proteins on their surface, called glycoconjugates. Cell-to-cell contact mediated by glycan:glycan interactions have been considered to be low-affinity interactions that precede high-affinity protein–glycan or protein–protein interactions. In several pathogenic bacteria, truncation of surface glycans, lipooligosaccharide (LOS), or lipopolysaccharide (LPS) have been reported to significantly reduce bacterial adherence to host cells. Here, we show that the saccharide component of LOS/LPS have direct, high-affinity interactions with host glycans. Glycan microarrays reveal that LOS/LPS of four distinct bacterial pathogens bind to numerous host glycan structures. Surface plasmon resonance was used to determine the affinity of these interactions and revealed 66 high-affinity host–glycan:bacterial–glycan pairs with equilibrium dissociation constants (KD) ranging between 100 nM and 50 µM. These glycan:glycan affinity values are similar to those reported for lectins or antibodies with glycans. Cell assays demonstrated that glycan:glycan interaction-mediated bacterial adherence could be competitively inhibited by either host cell or bacterial glycans. This is the first report to our knowledge of high affinity glycan:glycan interactions between bacterial pathogens and the host. The discovery of large numbers of glycan:glycan interactions between a diverse range of structures suggests that these interactions may be important in all biological systems. PMID:26676578

  16. Glycan:glycan interactions: High affinity biomolecular interactions that can mediate binding of pathogenic bacteria to host cells.

    PubMed

    Day, Christopher J; Tran, Elizabeth N; Semchenko, Evgeny A; Tram, Greg; Hartley-Tassell, Lauren E; Ng, Preston S K; King, Rebecca M; Ulanovsky, Rachel; McAtamney, Sarah; Apicella, Michael A; Tiralongo, Joe; Morona, Renato; Korolik, Victoria; Jennings, Michael P

    2015-12-29

    Cells from all domains of life express glycan structures attached to lipids and proteins on their surface, called glycoconjugates. Cell-to-cell contact mediated by glycan:glycan interactions have been considered to be low-affinity interactions that precede high-affinity protein-glycan or protein-protein interactions. In several pathogenic bacteria, truncation of surface glycans, lipooligosaccharide (LOS), or lipopolysaccharide (LPS) have been reported to significantly reduce bacterial adherence to host cells. Here, we show that the saccharide component of LOS/LPS have direct, high-affinity interactions with host glycans. Glycan microarrays reveal that LOS/LPS of four distinct bacterial pathogens bind to numerous host glycan structures. Surface plasmon resonance was used to determine the affinity of these interactions and revealed 66 high-affinity host-glycan:bacterial-glycan pairs with equilibrium dissociation constants (K(D)) ranging between 100 nM and 50 µM. These glycan:glycan affinity values are similar to those reported for lectins or antibodies with glycans. Cell assays demonstrated that glycan:glycan interaction-mediated bacterial adherence could be competitively inhibited by either host cell or bacterial glycans. This is the first report to our knowledge of high affinity glycan:glycan interactions between bacterial pathogens and the host. The discovery of large numbers of glycan:glycan interactions between a diverse range of structures suggests that these interactions may be important in all biological systems. PMID:26676578

  17. Highly pathogenic avian influenza virus nucleoprotein interacts with TREX complex adaptor protein Aly/REF.

    PubMed

    Balasubramaniam, Vinod R M T; Hong Wai, Tham; Ario Tejo, Bimo; Omar, Abdul Rahman; Syed Hassan, Sharifah

    2013-01-01

    We constructed a novel chicken (Gallus gallus) lung cDNA library fused inside yeast acting domain vector (pGADT7). Using yeast two-hybrid screening with highly pathogenic avian influenza (HPAI) nucleoprotein (NP) from the strain (A/chicken/Malaysia/5858/2004(H5N1)) as bait, and the Gallus gallus lung cDNA library as prey, a novel interaction between the Gallus gallus cellular RNA export adaptor protein Aly/REF and the viral NP was identified. This interaction was confirmed and validated with mammalian two hybrid studies and co-immunoprecipitation assay. Cellular localization studies using confocal microscopy showed that NP and Aly/REF co-localize primarily in the nucleus. Further investigations by mammalian two hybrid studies into the binding of NP of other subtypes of influenza virus such as the swine A/New Jersey/1976/H1N1 and pandemic A/Malaysia/854/2009(H1N1) to human Aly/REF, also showed that the NP of these viruses interacts with human Aly/REF. Our findings are also supported by docking studies which showed tight and favorable binding between H5N1 NP and human Aly/REF, using crystal structures from Protein Data Bank. siRNA knockdown of Aly/REF had little effect on the export of HPAI NP and other viral RNA as it showed no significant reduction in virus titer. However, UAP56, another component of the TREX complex, which recruits Aly/REF to mRNA was found to interact even better with H5N1 NP through molecular docking studies. Both these proteins also co-localizes in the nucleus at early infection similar to Aly/REF. Intriguingly, knockdown of UAP56 in A549 infected cells shows significant reduction in viral titer (close to 10 fold reduction). Conclusively, our study have opened new avenues for research of other cellular RNA export adaptors crucial in aiding viral RNA export such as the SRSF3, 9G8 and ASF/SF2 that may play role in influenza virus RNA nucleocytoplasmic transport.

  18. Surveillance for Highly Pathogenic Avian Influenza Virus in Wild Birds during Outbreaks in Domestic Poultry, Minnesota, 2015

    PubMed Central

    Carstensen, Michelle; Hildebrand, Erik C.; Cornicelli, Louis; Wolf, Paul; Grear, Daniel A.; Ip, Hon S.; Vandalen, Kaci K.; Minicucci, Larissa A.

    2016-01-01

    In 2015, a major outbreak of highly pathogenic avian influenza virus (HPAIV) infection devastated poultry facilities in Minnesota, USA. To understand the potential role of wild birds, we tested 3,139 waterfowl fecal samples and 104 sick and dead birds during March 9–June 4, 2015. HPAIV was isolated from a Cooper’s hawk but not from waterfowl fecal samples. PMID:27064759

  19. Novel Highly Pathogenic Avian H5 Influenza A Viruses in Live Poultry Markets, Wuxi City, China, 2013−2014

    PubMed Central

    Ma, Mai-Juan; Chen, Shan-Hui; Wang, Guo-Lin; Zhao, Teng; Qian, Yan-Hua; Wu, Meng-Na; Liu, Ying; Gray, Gregory C.; Lu, Bing; Cao, Wu-Chun

    2016-01-01

    During 12 recent months of periodic influenza virus surveillance at 9 live poultry markets in Wuxi City China, we identified multiple highly pathogenic H5N6, H5N8, H5N2, and H5N1 avian influenza viruses. The variety of potentially pandemic viruses in this low-risk area is disconcerting and portends an increased pandemic threat. PMID:27186580

  20. High-quality complete genome sequence of Microbacterium sp. SUBG005, a plant pathogen.

    PubMed

    Rakhashiya, Purvi M; Patel, Pooja P; Thaker, Vrinda S

    2015-09-01

    Microbacterium sp. SUBG005 is a Gram positive bacterium, isolated from infected leaf of Mangifera indica L. in Rajkot (22.30°N, 70.78°E), Gujarat, India. The genome sequencing of Microbacterium sp. SUBG005 is having type I secretion system genes of pathogenicity as well as heavy metal resistance unique genes. The genome size is 7.01 Mb with G + C content of 64.80% and contains rRNA sequences. Genome sequencing analysis provides information about the microbe role in host-pathogen interaction. The whole genome sequencing has been deposited in DDBJ/EMBL/GenBank under the accession number JNNT00000000. PMID:26484276

  1. Transmutation of Personal Glucose Meters into Portable and Highly Sensitive Microbial Pathogen Detection Platform.

    PubMed

    Wang, Zhenzhen; Chen, Zhaowei; Gao, Nan; Ren, Jinsong; Qu, Xiaogang

    2015-10-01

    Herein, for the first time, we presented a simple and general approach by using personal glucose meters (PGM) for portable and ultrasensitive detection of microbial pathogens. Upon addition of pathogenic bacteria, glucoamylase-quaternized magnetic nanoparticles (GA-QMNPS) conjugates were disrupted by the competitive multivalent interactions between bacteria and QMNPS, resulting in the release of GA. After magnetic separation, the free GA could catalyze the hydrolysis of amylose into glucose for quantitative readout by PGM. In such way, PGM was transmuted into a bacterial detection device and extremely low detection limits down to 20 cells mL(-1) was achieved. More importantly, QMNPS could inhibit the growth of the bacteria and destroy its cellular structure, which enabled bacteria detection and inhibition simultaneously. The simplicity, portability, sensitivity and low cost of presented work make it attractive for clinical applications. PMID:26153225

  2. Proteogenomic analysis of pathogenic yeast Cryptococcus neoformans using high resolution mass spectrometry

    PubMed Central

    2014-01-01

    Background Cryptococcus neoformans, a basidiomycetous fungus of universal occurrence, is a significant opportunistic human pathogen causing meningitis. Owing to an increase in the number of immunosuppressed individuals along with emergence of drug-resistant strains, C. neoformans is gaining importance as a pathogen. Although, whole genome sequencing of three varieties of C. neoformans has been completed recently, no global proteomic studies have yet been reported. Results We performed a comprehensive proteomic analysis of C. neoformans var. grubii (Serotype A), which is the most virulent variety, in order to provide protein-level evidence for computationally predicted gene models and to refine the existing annotations. We confirmed the protein-coding potential of 3,674 genes from a total of 6,980 predicted protein-coding genes. We also identified 4 novel genes and corrected 104 predicted gene models. In addition, our studies led to the correction of translational start site, splice junctions and reading frame used for translation in a number of proteins. Finally, we validated a subset of our novel findings by RT-PCR and sequencing. Conclusions Proteogenomic investigation described here facilitated the validation and refinement of computationally derived gene models in the intron-rich genome of C. neoformans, an important fungal pathogen in humans. PMID:24484775

  3. Unusually High Mortality in Waterfowl Caused by Highly Pathogenic Avian Influenza A(H5N1) in Bangladesh

    PubMed Central

    Haider, N.; Sturm-Ramirez, K.; Khan, S. U.; Rahman, M. Z.; Sarkar, S.; Poh, M. K.; Shivaprasad, H. L.; Kalam, M. A.; Paul, S. K.; Karmakar, P. C.; Balish, A.; Chakraborty, A.; Mamun, A. A.; Mikolon, A. B.; Davis, C. T.; Rahman, M.; Donis, R. O.; Heffelfinger, J. D.; Luby, S. P.; Zeidner, N.

    2015-01-01

    Summary Mortality in ducks and geese caused by highly pathogenic avian influenza A (H5N1) infection had not been previously identified in Bangladesh. In June–July 2011, we investigated mortality in ducks, geese and chickens with suspected H5N1 infection in a north-eastern district of the country to identify the aetiologic agent and extent of the outbreak and identify possible associated human infections. We surveyed households and farms with affected poultry flocks in six villages in Netrokona district and collected cloacal and oropharyngeal swabs from sick birds and tissue samples from dead poultry. We conducted a survey in three of these villages to identify suspected human influenza-like illness cases and collected nasopharyngeal and throat swabs. We tested all swabs by real-time RT-PCR, sequenced cultured viruses, and examined tissue samples by histopathology and immunohistochemistry to detect and characterize influenza virus infection. In the six villages, among the 240 surveyed households and 11 small-scale farms, 61% (1789/2930) of chickens, 47% (4816/10 184) of ducks and 73% (358/493) of geese died within 14 days preceding the investigation. Of 70 sick poultry swabbed, 80% (56/70) had detectable RNA for influenza A/H5, including 89% (49/55) of ducks, 40% (2/5) of geese and 50% (5/10) of chickens. We isolated virus from six of 25 samples; sequence analysis of the hemagglutinin and neuraminidase gene of these six isolates indicated clade 2.3.2.1a of H5N1 virus. Histopathological changes and immunohistochemistry staining of avian influenza viral antigens were recognized in the brain, pancreas and intestines of ducks and chickens. We identified ten human cases showing signs compatible with influenza-like illness; four were positive for influenza A/H3; however, none were positive for influenza A/H5. The recently introduced H5N1 clade 2.3.2.1a virus caused unusually high mortality in ducks and geese. Heightened surveillance in poultry is warranted to guide

  4. Pathobiological Characterization of a Novel Reassortant Highly Pathogenic H5N1 Virus Isolated in British Columbia, Canada, 2015

    PubMed Central

    Berhane, Yohannes; Kobasa, Darwyn; Embury-Hyatt, Carissa; Pickering, Brad; Babiuk, Shawn; Joseph, Tomy; Bowes, Victoria; Suderman, Mathew; Leung, Anders; Cottam-Birt, Colleen; Hisanaga, Tamiko; Pasick, John

    2016-01-01

    In the current study, we describe the pathobiologic characteristics of a novel reassortant virus - A/chicken/BC/FAV-002/2015 (H5N1) belonging to clade 2.3.4.4 that was isolated from backyard chickens in British Columbia, Canada. Sequence analyses demonstrate PB1, PA, NA and NS gene segments were of North American lineage while PB2, HA, NP and M were derived from a Eurasian lineage H5N8 virus. This novel virus had a 19 amino acid deletion in the neuraminidase stalk. We evaluated the pathogenic potential of this isolate in various animal models. The virus was highly pathogenic to mice with a LD50 of 10 plaque forming units (PFU), but had limited tissue tropism. It caused only subclinical infection in pigs which did result in seroconversion. This virus was highly pathogenic to chickens, turkeys, juvenile Muscovy ducks (Cairnia moschata foma domestica) and adult Chinese geese (Anser cynoides domesticus) causing a systemic infection in all species. The virus was also efficiently transmitted and resulted in mortality in naïve contact ducks, geese and chickens. Our findings indicate that this novel H5N1 virus has a wide host range and enhanced surveillance of migratory waterfowl may be necessary in order to determine its potential to establish itself in the wild bird reservoir. PMID:26988892

  5. Pathobiological Characterization of a Novel Reassortant Highly Pathogenic H5N1 Virus Isolated in British Columbia, Canada, 2015.

    PubMed

    Berhane, Yohannes; Kobasa, Darwyn; Embury-Hyatt, Carissa; Pickering, Brad; Babiuk, Shawn; Joseph, Tomy; Bowes, Victoria; Suderman, Mathew; Leung, Anders; Cottam-Birt, Colleen; Hisanaga, Tamiko; Pasick, John

    2016-01-01

    In the current study, we describe the pathobiologic characteristics of a novel reassortant virus - A/chicken/BC/FAV-002/2015 (H5N1) belonging to clade 2.3.4.4 that was isolated from backyard chickens in British Columbia, Canada. Sequence analyses demonstrate PB1, PA, NA and NS gene segments were of North American lineage while PB2, HA, NP and M were derived from a Eurasian lineage H5N8 virus. This novel virus had a 19 amino acid deletion in the neuraminidase stalk. We evaluated the pathogenic potential of this isolate in various animal models. The virus was highly pathogenic to mice with a LD50 of 10 plaque forming units (PFU), but had limited tissue tropism. It caused only subclinical infection in pigs which did result in seroconversion. This virus was highly pathogenic to chickens, turkeys, juvenile Muscovy ducks (Cairnia moschata foma domestica) and adult Chinese geese (Anser cynoides domesticus) causing a systemic infection in all species. The virus was also efficiently transmitted and resulted in mortality in naïve contact ducks, geese and chickens. Our findings indicate that this novel H5N1 virus has a wide host range and enhanced surveillance of migratory waterfowl may be necessary in order to determine its potential to establish itself in the wild bird reservoir. PMID:26988892

  6. Pathobiological Characterization of a Novel Reassortant Highly Pathogenic H5N1 Virus Isolated in British Columbia, Canada, 2015.

    PubMed

    Berhane, Yohannes; Kobasa, Darwyn; Embury-Hyatt, Carissa; Pickering, Brad; Babiuk, Shawn; Joseph, Tomy; Bowes, Victoria; Suderman, Mathew; Leung, Anders; Cottam-Birt, Colleen; Hisanaga, Tamiko; Pasick, John

    2016-03-18

    In the current study, we describe the pathobiologic characteristics of a novel reassortant virus - A/chicken/BC/FAV-002/2015 (H5N1) belonging to clade 2.3.4.4 that was isolated from backyard chickens in British Columbia, Canada. Sequence analyses demonstrate PB1, PA, NA and NS gene segments were of North American lineage while PB2, HA, NP and M were derived from a Eurasian lineage H5N8 virus. This novel virus had a 19 amino acid deletion in the neuraminidase stalk. We evaluated the pathogenic potential of this isolate in various animal models. The virus was highly pathogenic to mice with a LD50 of 10 plaque forming units (PFU), but had limited tissue tropism. It caused only subclinical infection in pigs which did result in seroconversion. This virus was highly pathogenic to chickens, turkeys, juvenile Muscovy ducks (Cairnia moschata foma domestica) and adult Chinese geese (Anser cynoides domesticus) causing a systemic infection in all species. The virus was also efficiently transmitted and resulted in mortality in naïve contact ducks, geese and chickens. Our findings indicate that this novel H5N1 virus has a wide host range and enhanced surveillance of migratory waterfowl may be necessary in order to determine its potential to establish itself in the wild bird reservoir.

  7. Highly active modulators of indole signaling alter pathogenic behaviors in Gram-negative and Gram-positive bacteria.

    PubMed

    Minvielle, Marine J; Eguren, Kristen; Melander, Christian

    2013-12-16

    Indole is a universal signal that regulates various bacterial behaviors, such as biofilm formation and antibiotic resistance. To generate mechanistic probes of indole signaling and control indole-mediated pathogenic phenotypes in both Gram-positive and Gram-negative bacteria, we have investigated the use of desformylflustrabromine (dFBr) derivatives to generate highly active indole mimetics. We have developed non-microbicidal dFBr derivatives that are 27-2000 times more active than indole in modulating biofilm formation, motility, acid resistance, and antibiotic resistance. The activity of these analogues parallels indole, because they are dependent on temperature, the enzyme tryptophanase TnaA, and the transcriptional regulator SdiA. This investigation demonstrates that molecules based on the dFBr scaffold can alter pathogenic behaviors by mimicking indole-signaling pathways.

  8. Isolation of highly pathogenic avian influenza H5N1 virus from Saker falcons (Falco cherrug) in the Middle East.

    PubMed

    Marjuki, Henju; Wernery, Ulrich; Yen, Hui-Ling; Franks, John; Seiler, Patrick; Walker, David; Krauss, Scott; Webster, Robert G

    2009-01-01

    There is accumulating evidence that birds of prey are susceptible to fatal infection with highly pathogenic avian influenza (HPAI) virus. We studied the antigenic, molecular, phylogenetic, and pathogenic properties of 2 HPAI H5N1 viruses isolated from dead falcons in Saudi Arabia and Kuwait in 2005 and 2007, respectively. Phylogenetic and antigenic analyses grouped both isolates in clade 2.2 (Qinghai-like viruses). However, the viruses appeared to have spread westward via different flyways. It remains unknown how these viruses spread so rapidly from Qinghai after the 2005 outbreak and how they were introduced into falcons in these two countries. The H5N1 outbreaks in the Middle East are believed by some to be mediated by wild migratory birds. However, sporting falcons may be at additional risk from the illegal import of live quail to feed them.

  9. Longitudinal 2 years field study of conventional vaccination against highly pathogenic avian influenza H5N1 in layer hens.

    PubMed

    Rudolf, Miriam; Pöppel, Manfred; Fröhlich, Andreas; Breithaupt, Angele; Teifke, Jens; Blohm, Ulrike; Mettenleiter, Thomas; Beer, Martin; Harder, Timm

    2010-10-01

    A licensed, inactivated vaccine based on a low pathogenic avian influenza virus strain (H5N2) was evaluated in layer hens kept under field conditions during a 2-year period. Vaccine efficacy was investigated by specific antibodies and by challenge-contact experiments using highly pathogenic avian influenza viruses (HPAIV) H5N1. Basic immunization with two applications induced clinical protection. Virus excretion by vaccinated hens was significantly reduced compared to non-vaccinated controls; transmission to non-vaccinated and vaccinated contact birds was not fully interrupted. Vaccination efficacy is influenced by several factors including antigenic relatedness between vaccine and field strains, but also by species, age and type of commercial uses of the host. Limitations and risks of HPAIV vaccination as silent spread of HPAIV and emergence of escape mutants must be considered a priori and appropriate corrective measures have to be installed. PMID:20727963

  10. Expression of H5 hemagglutinin vaccine antigen in common duckweed (Lemna minor) protects against H5N1 high pathogenicity avian influenza virus challenge in immunized chickens

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A synthetic hemagglutinin (HA) gene from the highly pathogenic avian influenza (HPAI) virus A/chicken/Indonesia/7/2003 (H5N1) (Indo/03) was expressed in aquatic plant Lemna minor (rLemna-HA). In Experiment 1, efficacy of rLemna-HA was tested on specific pathogen free (SPF) birds immunized with 0.2 ...

  11. Variation in protection by seven inactivated H5 vaccine strains against eight H5N1 high pathogenicity avian influenza viruses in chickens

    Technology Transfer Automated Retrieval System (TEKTRAN)

    H5N1 high pathogenicity avian influenza virus (HPAIV) is an important pathogen for poultry. Vaccines have assisted in control for poultry, and for human pandemic preparedness. However the genetic diversity and rapid antigenic drifting of the field viruses have led to inadequate protection. This s...

  12. High-throughput detection of food-borne pathogenic bacteria using oligonucleotide microarray with quantum dots as fluorescent labels.

    PubMed

    Huang, Aihua; Qiu, Zhigang; Jin, Min; Shen, Zhiqiang; Chen, Zhaoli; Wang, Xinwei; Li, Jun-Wen

    2014-08-18

    Bacterial pathogens are mostly responsible for food-borne diseases, and there is still substantial room for improvement in the effective detection of these organisms. In the present study, we explored a new method to detect target pathogens easily and rapidly with high sensitivity and specificity. This method uses an oligonucleotide microarray combined with quantum dots as fluorescent labels. Oligonucleotide probes targeting the 16SrRNA gene were synthesized to create an oligonucleotide microarray. The PCR products labeled with biotin were subsequently hybridized using an oligonucleotide microarray. Following incubation with CdSe/ZnS quantum dots coated with streptavidin, fluorescent signals were detected with a PerkinElmer Gx Microarray Scanner. The results clearly showed specific hybridization profiles corresponding to the bacterial species assessed. Two hundred and sixteen strains of food-borne bacterial pathogens, including standard strains and isolated strains from food samples, were used to test the specificity, stability, and sensitivity of the microarray system. We found that the oligonucleotide microarray combined with quantum dots used as fluorescent labels can successfully discriminate the bacterial organisms at the genera or species level, with high specificity and stability as well as a sensitivity of 10 colony forming units (CFU)/mL of pure culture. We further tested 105 mock-contaminated food samples and achieved consistent results as those obtained from traditional biochemical methods. Together, these results indicate that the quantum dot-based oligonucleotide microarray has the potential to be a powerful tool in the detection and identification of pathogenic bacteria in foods.

  13. Rapid identification of bovine mastitis pathogens by high-resolution melt analysis of 16S rDNA sequences.

    PubMed

    Ajitkumar, Praseeda; Barkema, Herman W; De Buck, Jeroen

    2012-03-23

    Accurate identification of mastitis pathogens is often compromised when using conventional culture-based methods. Here, we report a novel, rapid assay tested for speciation of bacterial mastitis pathogens using high-resolution melt analysis (HRMA) of 16S rDNA sequences. Real-time PCR amplification of 16S rRNA gene fragment, spanning the variable region V5 and V6 was performed with a resulting amplicon of 290bp. First, a library was generated of melt curves of 9 common pathogens that are implicated in bovine mastitis. Six of the isolates, Escherichia coli, Streptococcus agalactiae, Klebsiella pneumoniae, Streptococcus uberis, Staphylococcus aureus and Mycoplasma bovis, were type strains while the other 3, Arcanobacterium pyogenes, Corynebacterium bovis and Streptococcus dysgalactiae, were bovine mastitis field isolates. Four of the type strains, E. coli, S. agalactiae, K. pneumoniae and S. aureus, were found to be of human origin, while the other 3 type strains were isolated from bovine infections. Secondly, the melt curves and corresponding amplicon sequences of A. pyogenes, E. coli, S. agalactiae, S. dysgalactiae, K. pneumoniae, S. uberis and S. aureus were compared with 10 bovine mastitis field isolates of each pathogen. Based on the distinct differences in melt curves and sequences between human and bovine isolates of E. coli and K. pneumoniae, it was deemed necessary to select a set of bovine strains for these pathogens to be used as reference strains in the HRMA. Next, the HRMA was validated by three interpreters analyzing the differential clustering pattern of melt curves of 60 bacterial cultures obtained from mastitis milk samples. The three test interpreters were blinded to the culture and sequencing results of the isolates. Overall accuracy of the validation assay was 95% as there was difficulty in identifying the streptococci due to heterogeneity observed in the PCR amplicons of S. uberis. The present study revealed that broad-range real-time PCR with

  14. Determining the Phylogenetic and Phylogeographic Origin of Highly Pathogenic Avian Influenza (H7N3) in Mexico

    PubMed Central

    Lu, Lu; Lycett, Samantha J.; Leigh Brown, Andrew J.

    2014-01-01

    Highly pathogenic (HP) avian influenza virus (AIV) H7N3 outbreaks occurred 3 times in the Americas in the past 10 years and caused severe economic loss in the affected regions. In June/July 2012, new HP H7N3 outbreaks occurred at commercial farms in Jalisco, Mexico. Outbreaks continued to be identified in neighbouring states in Mexico till August 2013. To explore the origin of this outbreak, time resolved phylogenetic trees were generated from the eight segments of full-length AIV sequences in North America using BEAST. Location, subtype, avian host species and pathogenicity were modelled as discrete traits upon the trees using continuous time Markov chains. A further joint analysis among segments was performed using a hierarchical phylogenetic model (HPM) which allowed trait rates (location, subtype, host species) to be jointly inferred across different segments. The complete spatial diffusion process was visualised through virtual globe software. Our result indicated the Mexico HP H7N3 originated from the large North America low pathogenicity AIV pool through complicated reassortment events. Different segments were contributed by wild waterfowl from different N. American flyways. Five of the eight segments (HA, NA, NP, M, NS) were introduced from wild birds migrating along the central North American flyway, and PB2, PB1 and PA were introduced via the western North American flyway. These results highlight a potential role for Mexico as a hotspot of virus reassortment as it is where wild birds from different migration routes mix during the winter. PMID:25226523

  15. Tracking the Emerging Human Pathogen Pseudallescheria boydii by Using Highly Specific Monoclonal Antibodies ▿

    PubMed Central

    Thornton, Christopher R.

    2009-01-01

    Pseudallescheria boydii has long been known to cause white grain mycetoma in immunocompetent humans, but it has recently emerged as an opportunistic pathogen of humans, causing potentially fatal invasive infections in immunocompromised individuals and evacuees of natural disasters, such as tsunamis and hurricanes. The diagnosis of P. boydii is problematic since it exhibits morphological characteristics similar to those of other hyaline fungi that cause infectious diseases, such as Aspergillus fumigatus and Scedosporium prolificans. This paper describes the development of immunoglobulin M (IgM) and IgG1 κ-light chain monoclonal antibodies (MAbs) specific to P. boydii and certain closely related fungi. The MAbs bind to an immunodominant carbohydrate epitope on an extracellular 120-kDa antigen present in the spore and hyphal cell walls of P. boydii and Scedosporium apiospermum. The MAbs do not react with S. prolificans, Scedosporium dehoogii, or a large number of clinically relevant fungi, including A. fumigatus, Candida albicans, Cryptococcus neoformans, Fusarium solani, and Rhizopus oryzae. The MAbs were used in immunofluorescence and double-antibody sandwich enzyme-linked immunosorbent assays (DAS-ELISAs) to accurately differentiate P. boydii from other infectious fungi and to track the pathogen in environmental samples. Specificity of the DAS-ELISA was confirmed by sequencing of the internally transcribed spacer 1 (ITS1)-5.8S-ITS2 rRNA-encoding regions of environmental isolates. PMID:19321690

  16. Tracking the emerging human pathogen Pseudallescheria boydii by using highly specific monoclonal antibodies.

    PubMed

    Thornton, Christopher R

    2009-05-01

    Pseudallescheria boydii has long been known to cause white grain mycetoma in immunocompetent humans, but it has recently emerged as an opportunistic pathogen of humans, causing potentially fatal invasive infections in immunocompromised individuals and evacuees of natural disasters, such as tsunamis and hurricanes. The diagnosis of P. boydii is problematic since it exhibits morphological characteristics similar to those of other hyaline fungi that cause infectious diseases, such as Aspergillus fumigatus and Scedosporium prolificans. This paper describes the development of immunoglobulin M (IgM) and IgG1 kappa-light chain monoclonal antibodies (MAbs) specific to P. boydii and certain closely related fungi. The MAbs bind to an immunodominant carbohydrate epitope on an extracellular 120-kDa antigen present in the spore and hyphal cell walls of P. boydii and Scedosporium apiospermum. The MAbs do not react with S. prolificans, Scedosporium dehoogii, or a large number of clinically relevant fungi, including A. fumigatus, Candida albicans, Cryptococcus neoformans, Fusarium solani, and Rhizopus oryzae. The MAbs were used in immunofluorescence and double-antibody sandwich enzyme-linked immunosorbent assays (DAS-ELISAs) to accurately differentiate P. boydii from other infectious fungi and to track the pathogen in environmental samples. Specificity of the DAS-ELISA was confirmed by sequencing of the internally transcribed spacer 1 (ITS1)-5.8S-ITS2 rRNA-encoding regions of environmental isolates.

  17. Lactobacillus crispatus L1: high cell density cultivation and exopolysaccharide structure characterization to highlight potentially beneficial effects against vaginal pathogens

    PubMed Central

    2014-01-01

    Background Vaginal lactic acid bacteria defend the host against pathogens through a combination of competitive exclusion, competition for nutrients, production of antimicrobial substances and through the activation of the immune system. A new human isolate named Lactobacillus crispatus L1 was characterized in this work, and a preliminary evaluation of its probiotic potential is described together with a process to obtain a high productivity of viable biomass. Results In a simulated digestion process 1.8⋅1010 cells∙ml−1 survived the gastric environment with 80% viability, without being affected by small intestine juices. Experiments on six different C sources were performed to analyze growth and organic acids production and, glucose, provided the best performances. A microfiltration strategy was exploited to improve the cellular yield in 2 L-fermentation processes, reaching 27 g · l−1 of dry biomass. Moreover, L. crispatus L1 demonstrated a greater stability to high concentrations of lactic acid, compared to other lactobacilli. The specific L. crispatus L1 exopolysaccharide was purified from the fermentation broth and characterized by NMR showing structural features and similarity to exopolysaccharides produced by pathogenic strains. Live L. crispatus L1 cells strongly reduced adhesion of a yeast pathogenic strain, Candida albicans in particular, in adherence assays. Interestingly a higher expression of the human defensin HBD-2 was also observed in vaginal cells treated with the purified exopolysaccharide, indicating a possible correlation with C. albicans growth inhibition. Conclusions The paper describes the evaluation of L. crispatus L1 as potential vaginal probiotic and the fermentation processes to obtain high concentrations of viable cells. PMID:24884965

  18. Highly Pathogenic Avian Influenza A(H5N1) Virus Struck Migratory Birds in China in 2015

    PubMed Central

    Bi, Yuhai; Zhang, Zhenjie; Liu, Wenjun; Yin, Yanbo; Hong, Jianmin; Li, Xiangdong; Wang, Haiming; Wong, Gary; Chen, Jianjun; Li, Yunfeng; Ru, Wendong; Gao, Ruyi; Liu, Di; Liu, Yingxia; Zhou, Boping; Gao, George F.; Shi, Weifeng; Lei, Fumin

    2015-01-01

    Approximately 100 migratory birds, including whooper swans and pochards, were found dead in the Sanmenxia Reservoir Area of China during January 2015. The causative agent behind this outbreak was identified as H5N1 highly pathogenic avian influenza virus (HPAIV). Genetic and phylogenetic analyses revealed that this Sanmenxia H5N1 virus was a novel reassortant, possessing a Clade 2.3.2.1c HA gene and a H9N2-derived PB2 gene. Sanmenxia Clade 2.3.2.1c-like H5N1 viruses possess the closest genetic identity to A/Alberta/01/2014 (H5N1), which recently caused a fatal respiratory infection in Canada with signs of meningoencephalitis, a highly unusual symptom with influenza infections in humans. Furthermore, this virus was shown to be highly pathogenic to both birds and mammals, and demonstrate tropism for the nervous system. Due to the geographical location of Sanmenxia, these novel H5N1 viruses also have the potential to be imported to other regions through the migration of wild birds, similar to the H5N1 outbreak amongst migratory birds in Qinghai Lake during 2005. Therefore, further investigation and monitoring is required to prevent this novel reassortant virus from becoming a new threat to public health. PMID:26259704

  19. Highly Pathogenic Avian Influenza A(H5N1) Virus Struck Migratory Birds in China in 2015.

    PubMed

    Bi, Yuhai; Zhang, Zhenjie; Liu, Wenjun; Yin, Yanbo; Hong, Jianmin; Li, Xiangdong; Wang, Haiming; Wong, Gary; Chen, Jianjun; Li, Yunfeng; Ru, Wendong; Gao, Ruyi; Liu, Di; Liu, Yingxia; Zhou, Boping; Gao, George F; Shi, Weifeng; Lei, Fumin

    2015-01-01

    Approximately 100 migratory birds, including whooper swans and pochards, were found dead in the Sanmenxia Reservoir Area of China during January 2015. The causative agent behind this outbreak was identified as H5N1 highly pathogenic avian influenza virus (HPAIV). Genetic and phylogenetic analyses revealed that this Sanmenxia H5N1 virus was a novel reassortant, possessing a Clade 2.3.2.1c HA gene and a H9N2-derived PB2 gene. Sanmenxia Clade 2.3.2.1c-like H5N1 viruses possess the closest genetic identity to A/Alberta/01/2014 (H5N1), which recently caused a fatal respiratory infection in Canada with signs of meningoencephalitis, a highly unusual symptom with influenza infections in humans. Furthermore, this virus was shown to be highly pathogenic to both birds and mammals, and demonstrate tropism for the nervous system. Due to the geographical location of Sanmenxia, these novel H5N1 viruses also have the potential to be imported to other regions through the migration of wild birds, similar to the H5N1 outbreak amongst migratory birds in Qinghai Lake during 2005. Therefore, further investigation and monitoring is required to prevent this novel reassortant virus from becoming a new threat to public health.

  20. Pathogenicity and transmission of H5 highly pathogenic avian influenza clade 2.3.4.4 viruses (H5N8 and H5N2) in domestic waterfowl (Pekin ducks and Chinese geese)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Domestic ducks and geese are common backyard poultry in many countries, frequently in contact with wild waterfowl, which are natural reservoirs of avian influenza viruses and have played a key role in the spread of Asian-lineage H5N1 highly pathogenic avian influenza (HPAI). In late 2014, a reassor...

  1. Differences in pathogenicity, response to vaccination, and innate immune responses in different types of ducks infected with a virulent H5N1 highly pathogenic avian influenza virus from Vietnam

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Wild ducks are reservoirs of avian influenza viruses in nature, and usually don’t show signs of disease. However, some Asian lineage H5N1 highly pathogenic avian influenza (HPAI) viruses can cause disease and death in both wild and domestic ducks. The objective of this study was to compare the cli...

  2. Assessment of the Internal Genes of Influenza A (H7N9) Virus Contributing to High Pathogenicity in Mice

    PubMed Central

    Bi, Yuhai; Xie, Qing; Zhang, Shuang; Li, Yun; Xiao, Haixia; Jin, Tao; Zheng, Weinan; Li, Jing; Jia, Xiaojuan; Sun, Lei; Liu, Jinhua; Qin, Chuan

    2014-01-01

    ABSTRACT The recently identified H7N9 influenza A virus has caused severe economic losses and worldwide public concern. Genetic analysis indicates that its six internal genes all originated from H9N2 viruses. However, the H7N9 virus is more highly pathogenic in humans than H9N2, which suggests that the internal genes of H7N9 have mutated. To analyze which H7N9 virus internal genes contribute to its high pathogenicity, a series of reassortants was generated by reverse genetics, with each virus containing a single internal gene of the typical A/Anhui/1/2013 (H7N9) (AH-H7N9) virus in the genetic background of the A/chicken/Shandong/lx1023/2007 (H9N2) virus. The replication ability, polymerase activity, and pathogenicity of these viruses were then evaluated in vitro and in vivo. These recombinants displayed high genetic compatibility, and the H7N9-derived PB2, M, and NP genes were identified as the virulence genes for the reassortants in mice. Further investigation confirmed that the PB2 K627 residue is critical for the high pathogenicity of the H7N9 virus and the reassortant containing the H7N9-derived PB2 segment (H9N2-AH/PB2). Notably, the H7N9-derived PB2 gene displayed greater compatibility with the H9N2 genome than that of H7N9, endowing the H9N2-AH/PB2 reassortant with greater viability and virulence than the parental H7N9 virus. In addition, the H7N9 virus, with the exception of the H9N2 reassortants, could effectively replicate in human A549 cells. Our results indicate that PB2, M, and NP are the key virulence genes, together with the surface hemagglutinin (HA) and neuraminidase (NA) proteins, contributing to the high infectivity of the H7N9 virus in humans. IMPORTANCE To date, the novel H7N9 influenza A virus has caused 437 human infections, with approximately 30% mortality. Previous work has primarily focused on the two viral surface proteins, HA and NA, but the contribution of the six internal genes to the high pathogenicity of H7N9 has not been

  3. Pathogen-free screening of bacteria-specific hybridomas for selecting high-quality monoclonal antibodies against pathogen bacteria as illustrated for Legionella pneumophila.

    PubMed

    Féraudet-Tarisse, Cécile; Vaisanen-Tunkelrott, Marja-Liisa; Moreau, Karine; Lamourette, Patricia; Créminon, Christophe; Volland, Hervé

    2013-05-31

    Antibodies are potent biological tools increasingly used as detection, diagnostic and therapeutic reagents. Many technological advances have optimized and facilitated production and screening of monoclonal antibodies. We report here an original method to screen for antibodies targeting biosafety level 2 or 3 pathogens without the fastidious handling inherent to pathogen use. A double ELISA screening was performed using as coated antigen transformed Escherichia coli expressing at its surface a protein specific to the pathogenic bacteria versus control untransformed E. coli. This method was applied to Legionella, using the surface-exposed Mip protein (macrophage infectivity potentiator). This screening proved to be an excellent means of selecting mAbs that bind Legionella pneumophila 1 surface-exposed Mip protein. This method also appears more biologically relevant than screening using the recombinant Mip protein alone and less tedious than a test performed directly on Legionella bacteria. We obtained 21 mAbs that bind strongly to L. pneumophila serogroups 1 to 13, and we validated their use in a rapid ELISA (performed in 4.5 h) and an immunochromatographic test (20 min).

  4. Bloodborne pathogens

    MedlinePlus

    ... page: //medlineplus.gov/ency/patientinstructions/000453.htm Bloodborne pathogens To use the sharing features on this page, please enable JavaScript. A pathogen is something that causes disease. Germs that can ...

  5. Combating highly resistant emerging pathogen Mycobacterium abscessus and Mycobacterium tuberculosis with novel salicylanilide esters and carbamates.

    PubMed

    Baranyai, Zsuzsa; Krátký, Martin; Vinšová, Jarmila; Szabó, Nóra; Senoner, Zsuzsanna; Horváti, Kata; Stolaříková, Jiřina; Dávid, Sándor; Bősze, Szilvia

    2015-08-28

    In the Mycobacterium genus over one hundred species are already described and new ones are periodically reported. Species that form colonies in a week are classified as rapid growers, those requiring longer periods (up to three months) are the mostly pathogenic slow growers. More recently, new emerging species have been identified to lengthen the list, all rapid growers. Of these, Mycobacterium abscessus is also an intracellular pathogen and it is the most chemotherapy-resistant rapid-growing mycobacterium. In addition, the cases of multidrug-resistant Mycobacterium tuberculosis infection are also increasing. Therefore there is an urgent need to find new active molecules against these threatening strains. Based on previous results, a series of salicylanilides, salicylanilide 5-chloropyrazinoates and carbamates was designed, synthesized and characterised. The compounds were evaluated for their in vitro activity on M. abscessus, susceptible M. tuberculosis H37Rv, multidrug-resistant (MDR) M. tuberculosis MDR A8, M. tuberculosis MDR 9449/2006 and on the extremely-resistant Praha 131 (XDR) strains. All derivatives exhibited a significant activity with minimum inhibitory concentrations (MICs) in the low micromolar range. Eight salicylanilide carbamates and two salicylanilide esters exhibited an excellent in vitro activity on M. abscessus with MICs from 0.2 to 2.1 μM, thus being more effective than ciprofloxacin and gentamicin. This finding is potentially promising, particularly, as M. abscessus is a threateningly chemotherapy-resistant species. M. tuberculosis H37Rv was inhibited with MICs from 0.2 μM, and eleven compounds have lower MICs than isoniazid. Salicylanilide esters and carbamates were found that they were effective also on MDR and XDR M. tuberculosis strains with MICs ≥1.0 μM. The in vitro cytotoxicity (IC50) was also determined on human MonoMac-6 cells, and selectivity index (SI) of the compounds was established. In general, salicylanilide

  6. Combating highly resistant emerging pathogen Mycobacterium abscessus and Mycobacterium tuberculosis with novel salicylanilide esters and carbamates.

    PubMed

    Baranyai, Zsuzsa; Krátký, Martin; Vinšová, Jarmila; Szabó, Nóra; Senoner, Zsuzsanna; Horváti, Kata; Stolaříková, Jiřina; Dávid, Sándor; Bősze, Szilvia

    2015-08-28

    In the Mycobacterium genus over one hundred species are already described and new ones are periodically reported. Species that form colonies in a week are classified as rapid growers, those requiring longer periods (up to three months) are the mostly pathogenic slow growers. More recently, new emerging species have been identified to lengthen the list, all rapid growers. Of these, Mycobacterium abscessus is also an intracellular pathogen and it is the most chemotherapy-resistant rapid-growing mycobacterium. In addition, the cases of multidrug-resistant Mycobacterium tuberculosis infection are also increasing. Therefore there is an urgent need to find new active molecules against these threatening strains. Based on previous results, a series of salicylanilides, salicylanilide 5-chloropyrazinoates and carbamates was designed, synthesized and characterised. The compounds were evaluated for their in vitro activity on M. abscessus, susceptible M. tuberculosis H37Rv, multidrug-resistant (MDR) M. tuberculosis MDR A8, M. tuberculosis MDR 9449/2006 and on the extremely-resistant Praha 131 (XDR) strains. All derivatives exhibited a significant activity with minimum inhibitory concentrations (MICs) in the low micromolar range. Eight salicylanilide carbamates and two salicylanilide esters exhibited an excellent in vitro activity on M. abscessus with MICs from 0.2 to 2.1 μM, thus being more effective than ciprofloxacin and gentamicin. This finding is potentially promising, particularly, as M. abscessus is a threateningly chemotherapy-resistant species. M. tuberculosis H37Rv was inhibited with MICs from 0.2 μM, and eleven compounds have lower MICs than isoniazid. Salicylanilide esters and carbamates were found that they were effective also on MDR and XDR M. tuberculosis strains with MICs ≥1.0 μM. The in vitro cytotoxicity (IC50) was also determined on human MonoMac-6 cells, and selectivity index (SI) of the compounds was established. In general, salicylanilide

  7. Victims and vectors: highly pathogenic avian influenza H5N1 and the ecology of wild birds

    USGS Publications Warehouse

    Takekawa, John Y.; Prosser, Diann J.; Newman, Scott H.; Muzaffar, Sabir Bin; Hill, Nichola J.; Yan, Baoping; Xiao, Xiangming; Lei, Fumin; Li, Tianxian; Schwarzbach, Steven E.; Howell, Judd A.

    2010-01-01

    The emergence of highly pathogenic avian influenza (HPAI) viruses has raised concerns about the role of wild birds in the spread and persistence of the disease. In 2005, an outbreak of the highly pathogenic subtype H5N1 killed more than 6,000 wild waterbirds at Qinghai Lake, China. Outbreaks have continued to periodically occur in wild birds at Qinghai Lake and elsewhere in Central China and Mongolia. This region has few poultry but is a major migration and breeding area for waterbirds in the Central Asian Flyway, although relatively little is known about migratory movements of different species and connectivity of their wetland habitats. The scientific debate has focused on the role of waterbirds in the epidemiology, maintenance and spread of HPAI H5N1: to what extent are they victims affected by the disease, or vectors that have a role in disease transmission? In this review, we summarise the current knowledge of wild bird involvement in the ecology of HPAI H5N1. Specifically, we present details on: (1) origin of HPAI H5N1; (2) waterbirds as LPAI reservoirs and evolution into HPAI; (3) the role of waterbirds in virus spread and persistence; (4) key biogeographic regions of outbreak; and (5) applying an ecological research perspective to studying AIVs in wild waterbirds and their ecosystems.

  8. Probe-free and sensitive detection of diarrhea-causing pathogens using RT-PCR combined high resolution melting analysis.

    PubMed

    Wang, Hai-Bo; Mo, Qiu-Hua; Wang, Qi; Wu, Bi-Mei; Feng, Zi-Li; Lin, Ji-Can; Yang, Ze

    2016-09-01

    Rapid and sensitive diagnostic methods are needed to help physicians make faster and better treatment decision for patients suffered from diarrhea. In the present study, a probe-free and sensitive RT-PCR combined high resolution melting analysis (HRMA) assay was established successfully for the detection of four major diarrhea-causing pathogens. The lower limit of detection of the assay were 10(0), 10(2), 10(0) and 10(3) copies/reaction for rotaviruses group A, astroviruses serotype 1, noroviruses genogroup II, and sapoviruses genegroup I, respectively, which were 1000-fold, 10-fold, 1000-fold and 10-fold more sensitive than conventional RT-PCR assay developed in parallel and comparable to or higher than commercially available real-time RT-PCR assay. Blinded sample evaluation showed that the assay was 100% concordant to both conventional RT-PCR and commercial real-time RT-PCR, indicating high reliability of the new assay. Therefore, the assay could provide a valuable platform for the probe-free and sensitive diagnosis of these pathogens. PMID:27461241

  9. [Construction of cDNA infectious clones of EV71 highly-pathogenic and cell-culture-adapted strains].

    PubMed

    Zhang, Yong-xin; Li, Xiao-yu; Huang, Yu-ming; Zhou, Yong-dong; Bi, Sheng-li; Cen, Shan

    2014-11-01

    The highly-pathogenic EV71 strain is the primary cause of mortality in hand-foot-and-mouth disease (HFMD) associated with neurological symptoms, for which no clinically effective drugs or vaccines exist. This study aimed to construct infectious cDNA clones of the EV71 highly-pathogenic strain and the cell-culture adapted strain using a reverse genetics approach. The genomic RNAs of EV71 parent strains were used as the templates for RT-PCR amplification, and then the PCR products that overlapped the full-length genome were connected into the pBR322 vector to produce infectious clones of pEV71 (HP) and pEV71 (CCA), respectively. The results showed that the HP strain propagated much more quickly than the CCA strain. The rescued viruses derived from the infectious clones not only maintained their consistency with their parent strains in terms of genomic sequences, but also retained their respective biological phenotypes. This research will contribute to our understanding of EV71 pathogenesis and the development of novel vaccines against HFMD.

  10. Practices associated with Highly Pathogenic Avian Influenza spread in traditional poultry marketing chains: Social and economic perspectives.

    PubMed

    Paul, Mathilde; Baritaux, Virginie; Wongnarkpet, Sirichai; Poolkhet, Chaithep; Thanapongtharm, Weerapong; Roger, François; Bonnet, Pascal; Ducrot, Christian

    2013-04-01

    In developing countries, smallholder poultry production contributes to food security and poverty alleviation in rural areas. However, traditional poultry marketing chains have been threatened by the epidemics caused by the Highly Pathogenic Avian Influenza (H5N1) virus. The article presents a value chain analysis conducted on the traditional poultry marketing chain in the rural province of Phitsanulok, Thailand. The analysis is based on quantitative data collected on 470 backyard chicken farms, and on qualitative data collected on 28 poultry collectors, slaughterhouses and market retailers, using semi-structured interviews. The article examines the organization of poultry marketing chains in time and space, and shows how this may contribute to the spread of Highly Pathogenic Avian Influenza H5N1 in the small-scale poultry sector. The article also discusses the practices and strategies developed by value chain actors facing poultry mortality, with their economic and social determinants. More broadly, this study also illustrates how value chain analysis can contribute to a better understanding of the complex mechanisms associated with the spread of epidemics in rural communities.

  11. Comparing viral metagenomics methods using a highly multiplexed human viral pathogens reagent.

    PubMed

    Li, Linlin; Deng, Xutao; Mee, Edward T; Collot-Teixeira, Sophie; Anderson, Rob; Schepelmann, Silke; Minor, Philip D; Delwart, Eric

    2015-03-01

    Unbiased metagenomic sequencing holds significant potential as a diagnostic tool for the simultaneous detection of any previously genetically described viral nucleic acids in clinical samples. Viral genome sequences can also inform on likely phenotypes including drug susceptibility or neutralization serotypes. In this study, different variables of the laboratory methods often used to generate viral metagenomics libraries were compared for their abilities to detect multiple viruses and generate full genome coverage. A biological reagent consisting of 25 different human RNA and DNA viral pathogens was used to estimate the effect of filtration and nuclease digestion, DNA/RNA extraction methods, pre-amplification and the use of different library preparation kits on the detection of viral nucleic acids. Filtration and nuclease treatment led to slight decreases in the percentage of viral sequence reads and number of viruses detected. For nucleic acid extractions silica spin columns improved viral sequence recovery relative to magnetic beads and Trizol extraction. Pre-amplification using random RT-PCR while generating more viral sequence reads resulted in detection of fewer viruses, more overlapping sequences, and lower genome coverage. The ScriptSeq library preparation method retrieved more viruses and a greater fraction of their genomes than the TruSeq and Nextera methods. Viral metagenomics sequencing was able to simultaneously detect up to 22 different viruses in the biological reagent analyzed including all those detected by qPCR. Further optimization will be required for the detection of viruses in biologically more complex samples such as tissues, blood, or feces.

  12. Comparing viral metagenomics methods using a highly multiplexed human viral pathogens reagent

    PubMed Central

    Li, Linlin; Deng, Xutao; Mee, Edward T.; Collot-Teixeira, Sophie; Anderson, Rob; Schepelmann, Silke; Minor, Philip D.; Delwart, Eric

    2014-01-01

    Unbiased metagenomic sequencing holds significant potential as a diagnostic tool for the simultaneous detection of any previously genetically described viral nucleic acids in clinical samples. Viral genome sequences can also inform on likely phenotypes including drug susceptibility or neutralization serotypes. In this study, different variables of the laboratory methods often used to generate viral metagenomics libraries on the efficiency of viral detection and virus genome coverage were compared. A biological reagent consisting of 25 different human RNA and DNA viral pathogens was used to estimate the effect of filtration and nuclease digestion, DNA/RNA extraction methods, pre-amplification and the use of different library preparation kits on the detection of viral nucleic acids. Filtration and nuclease treatment led to slight decreases in the percentage of viral sequence reads and number of viruses detected. For nucleic acid extractions silica spin columns improved viral sequence recovery relative to magnetic beads and Trizol extraction. Pre-amplification using random RT-PCR while generating more viral sequence reads resulted in detection of fewer viruses, more overlapping sequences, and lower genome coverage. The ScriptSeq library preparation method retrieved more viruses and a greater fraction of their genomes than the TruSeq and Nextera methods. Viral metagenomics sequencing was able to simultaneously detect up to 22 different viruses in the biological reagent analyzed including all those detected by qPCR. Further optimization will be required for the detection of viruses in biologically more complex samples such as tissues, blood, or feces. PMID:25497414

  13. High-yield production of a stable Vero cell-based vaccine candidate against the highly pathogenic avian influenza virus H5N1

    SciTech Connect

    Zhou, Fangye; Zhou, Jian; Ma, Lei; Song, Shaohui; Zhang, Xinwen; Li, Weidong; Jiang, Shude; Wang, Yue; Liao, Guoyang

    2012-05-18

    Highlights: Black-Right-Pointing-Pointer Vero cell-based HPAI H5N1 vaccine with stable high yield. Black-Right-Pointing-Pointer Stable high yield derived from the YNVa H3N2 backbone. Black-Right-Pointing-Pointer H5N1/YNVa has a similar safety and immunogenicity to H5N1delta. -- Abstract: Highly pathogenic avian influenza (HPAI) viruses pose a global pandemic threat, for which rapid large-scale vaccine production technology is critical for prevention and control. Because chickens are highly susceptible to HPAI viruses, the supply of chicken embryos for vaccine production might be depleted during a virus outbreak. Therefore, developing HPAI virus vaccines using other technologies is critical. Meeting vaccine demand using the Vero cell-based fermentation process has been hindered by low stability and yield. In this study, a Vero cell-based HPAI H5N1 vaccine candidate (H5N1/YNVa) with stable high yield was achieved by reassortment of the Vero-adapted (Va) high growth A/Yunnan/1/2005(H3N2) (YNVa) virus with the A/Anhui/1/2005(H5N1) attenuated influenza vaccine strain (H5N1delta) using the 6/2 method. The reassorted H5N1/YNVa vaccine maintained a high hemagglutination (HA) titer of 1024. Furthermore, H5N1/YNVa displayed low pathogenicity and uniform immunogenicity compared to that of the parent virus.

  14. Specialist enemies, generalist weapons and the potential spread of exotic pathogens: malaria parasites in a highly invasive bird.

    PubMed

    Clark, Nicholas J; Olsson-Pons, Sophie; Ishtiaq, Farah; Clegg, Sonya M

    2015-12-01

    Pathogens can influence the success of invaders. The Enemy Release Hypothesis predicts invaders encounter reduced pathogen abundance and diversity, while the Novel Weapons Hypothesis predicts invaders carry novel pathogens that spill over to competitors. We tested these hypotheses using avian malaria (haemosporidian) infections in the invasive myna (Acridotheres tristis), which was introduced to southeastern Australia from India and was secondarily expanded to the eastern Australian coast. Mynas and native Australian birds were screened in the secondary introduction range for haemosporidians (Plasmodium and Haemoproteus spp.) and results were combined with published data from the myna's primary introduction and native ranges. We compared malaria prevalence and diversity across myna populations to test for Enemy Release and used phylogeographic analyses to test for exotic strains acting as Novel Weapons. Introduced mynas carried significantly lower parasite diversity than native mynas and significantly lower Haemoproteus prevalence than native Australian birds. Despite commonly infecting native species that directly co-occur with mynas, Haemoproteus spp. were only recorded in introduced mynas in the primary introduction range and were apparently lost during secondary expansion. In contrast, Plasmodium infections were common in all ranges and prevalence was significantly higher in both introduced and native mynas than in native Australian birds. Introduced mynas carried several exotic Plasmodium lineages that were shared with native mynas, some of which also infected native Australian birds and two of which are highly invasive in other bioregions. Our results suggest that introduced mynas may benefit through escape from Haemoproteus spp. while acting as important reservoirs for Plasmodium spp., some of which are known exotic lineages. PMID:26433143

  15. Evidence for the Convergence Model: The Emergence of Highly Pathogenic Avian Influenza (H5N1) in Viet Nam

    PubMed Central

    Saksena, Sumeet; Fox, Jefferson; Epprecht, Michael; Tran, Chinh C.; Nong, Duong H.; Spencer, James H.; Nguyen, Lam; Finucane, Melissa L.; Tran, Vien D.; Wilcox, Bruce A.

    2015-01-01

    Building on a series of ground breaking reviews that first defined and drew attention to emerging infectious diseases (EID), the ‘convergence model’ was proposed to explain the multifactorial causality of disease emergence. The model broadly hypothesizes disease emergence is driven by the co-incidence of genetic, physical environmental, ecological, and social factors. We developed and tested a model of the emergence of highly pathogenic avian influenza (HPAI) H5N1 based on suspected convergence factors that are mainly associated with land-use change. Building on previous geospatial statistical studies that identified natural and human risk factors associated with urbanization, we added new factors to test whether causal mechanisms and pathogenic landscapes could be more specifically identified. Our findings suggest that urbanization spatially combines risk factors to produce particular types of peri-urban landscapes with significantly higher HPAI H5N1 emergence risk. The work highlights that peri-urban areas of Viet Nam have higher levels of chicken densities, duck and geese flock size diversities, and fraction of land under rice or aquaculture than rural and urban areas. We also found that land-use diversity, a surrogate measure for potential mixing of host populations and other factors that likely influence viral transmission, significantly improves the model’s predictability. Similarly, landscapes where intensive and extensive forms of poultry production overlap were found at greater risk. These results support the convergence hypothesis in general and demonstrate the potential to improve EID prevention and control by combing geospatial monitoring of these factors along with pathogen surveillance programs. PMID:26398118

  16. Evidence for the Convergence Model: The Emergence of Highly Pathogenic Avian Influenza (H5N1) in Viet Nam.

    PubMed

    Saksena, Sumeet; Fox, Jefferson; Epprecht, Michael; Tran, Chinh C; Nong, Duong H; Spencer, James H; Nguyen, Lam; Finucane, Melissa L; Tran, Vien D; Wilcox, Bruce A

    2015-01-01

    Building on a series of ground breaking reviews that first defined and drew attention to emerging infectious diseases (EID), the 'convergence model' was proposed to explain the multifactorial causality of disease emergence. The model broadly hypothesizes disease emergence is driven by the co-incidence of genetic, physical environmental, ecological, and social factors. We developed and tested a model of the emergence of highly pathogenic avian influenza (HPAI) H5N1 based on suspected convergence factors that are mainly associated with land-use change. Building on previous geospatial statistical studies that identified natural and human risk factors associated with urbanization, we added new factors to test whether causal mechanisms and pathogenic landscapes could be more specifically identified. Our findings suggest that urbanization spatially combines risk factors to produce particular types of peri-urban landscapes with significantly higher HPAI H5N1 emergence risk. The work highlights that peri-urban areas of Viet Nam have higher levels of chicken densities, duck and geese flock size diversities, and fraction of land under rice or aquaculture than rural and urban areas. We also found that land-use diversity, a surrogate measure for potential mixing of host populations and other factors that likely influence viral transmission, significantly improves the model's predictability. Similarly, landscapes where intensive and extensive forms of poultry production overlap were found at greater risk. These results support the convergence hypothesis in general and demonstrate the potential to improve EID prevention and control by combing geospatial monitoring of these factors along with pathogen surveillance programs. PMID:26398118

  17. A Fluorescence-Based High-Throughput Screening Assay to Identify Growth Inhibitors of the Pathogenic Fungus Aspergillus fumigatus.

    PubMed

    Smith, Thomas M; Richie, Daryl L; Tao, Jianshi

    2016-01-01

    Due to the advancements in modern medicine that have resulted in an increased number of immunocompromised individuals, the incidences and the associated mortality of invasive aspergillosis have continued to rise over the past three decades despite appropriate treatment. As a result, invasive aspergillosis has emerged as a leading cause of infection-related mortality in immunocompromised individuals. Utilizing the resazurin to resorufin conversion fluorescence readout to monitor cell viability, herein, we outline a high-throughput screening method amenable to profiling a large pharmaceutical library against the clinically relevant but less frequently screened fungal pathogen Aspergillus fumigatus. This enables the user to conduct high-throughput screening using a disease-relevant fungal growth assay and identify novel antifungal chemotypes as drug leads. PMID:27316995

  18. Targeted surveillance for highly pathogenic avian influenza in migratory waterfowl across the conterminous United States: chapter 12

    USGS Publications Warehouse

    Farnsworth, Matthew L.; Kendall, William L.; Doherty, Paul F.; Miller, Ryan S.; White, Gary C.; Nichols, James D.; Burnham, Kenneth P.; Franklin, Alan B.; Majumdar, S.; Brenner, F.J.; Huffman, J.E.; McLean, R.G.; Panah, A.I.; Pietrobon, P.J.; Keeler, S.P.; Shive, S.

    2011-01-01

    Introduction of Asian strain H5N1 Highly Pathogenic avian influenca via waterfowl migration is one potential route of entry into the United States. In conjunction with state, tribe, and laboratory partners, the United States Department of Agriculture collected and tested 124,603 wild bird samples in 2006 as part of a national surveillance effort. A sampling plan was devised to increase the probability fo detecting Asian strain H5N1 at a national scale. Band recovery data were used to identify and prioritize sampling for wild migratory waterfowl, resulting in spatially targeted sampling recommendations focused on reads with high numbers of recoveries. We also compared the spatial and temporal distribution of the 2006 cloacal and fecal waterfowl sampling effort to the bird banding recovery data and found concordance between the two .Finally, we present improvements made to the 2007 fecal sampling component of the surveillance plan and suggest further improvements for future sampling.

  19. Protection and virus shedding of falcons vaccinated against highly pathogenic avian influenza A virus (H5N1).

    PubMed

    Lierz, Michael; Hafez, Hafez M; Klopfleisch, Robert; Lüschow, Dörte; Prusas, Christine; Teifke, Jens P; Rudolf, Miriam; Grund, Christian; Kalthoff, Donata; Mettenleiter, Thomas; Beer, Martin; Hardert, Timm

    2007-11-01

    Because fatal infections with highly pathogenic avian influenza A (HPAI) virus subtype H5N1 have been reported in birds of prey, we sought to determine detailed information about the birds' susceptibility and protection after vaccination. Ten falcons vaccinated with an inactivated influenza virus (H5N2) vaccine seroconverted. We then challenged 5 vaccinated and 5 nonvaccinated falcons with HPAI (H5N1). All vaccinated birds survived; all unvaccinated birds died within 5 days. For the nonvaccinated birds, histopathologic examination showed tissue degeneration and necrosis, immunohistochemical techniques showed influenza virus antigen in affected tissues, and these birds shed high levels of infectious virus from the oropharynx and cloaca. Vaccinated birds showed no influenza virus antigen in tissues and shed virus at lower titers from the oropharynx only. Vaccination could protect these valuable birds and, through reduced virus shedding, reduce risk for transmission to other avian species and humans.

  20. Highly Dynamic Exon Shuffling in Candidate Pathogen Receptors … What if Brown Algae Were Capable of Adaptive Immunity?

    PubMed Central

    Zambounis, Antonios; Elias, Marek; Sterck, Lieven; Maumus, Florian; Gachon, Claire M.M.

    2012-01-01

    Pathogen recognition is the first step of immune reactions. In animals and plants, direct or indirect pathogen recognition is often mediated by a wealth of fast-evolving receptors, many of which contain ligand-binding and signal transduction domains, such as leucine-rich or tetratricopeptide repeat (LRR/TPR) and NB-ARC domains, respectively. In order to identify candidates potentially involved in algal defense, we mined the genome of the brown alga Ectocarpus siliculosus for homologues of these genes and assessed the evolutionary pressures acting upon them. We thus annotated all Ectocarpus LRR-containing genes, in particular an original group of LRR-containing GTPases of the ROCO family, and 24 NB-ARC–TPR proteins. They exhibit high birth and death rates, while a diversifying selection is acting on their LRR (respectively TPR) domain, probably affecting the ligand-binding specificities. Remarkably, each repeat is encoded by an exon, and the intense exon shuffling underpins the variability of LRR and TPR domains. We conclude that the Ectocarpus ROCO and NB-ARC–TPR families are excellent candidates for being involved in recognition/transduction events linked to immunity. We further hypothesize that brown algae may generate their immune repertoire via controlled somatic recombination, so far only known from the vertebrate adaptive immune systems. PMID:22144640

  1. Highly multiplexed profiling of single-cell effector functions reveals deep functional heterogeneity in response to pathogenic ligands

    PubMed Central

    Lu, Yao; Xue, Qiong; Eisele, Markus R.; Sulistijo, Endah S.; Brower, Kara; Han, Lin; Amir, El-ad David; Pe’er, Dana; Miller-Jensen, Kathryn; Fan, Rong

    2015-01-01

    Despite recent advances in single-cell genomic, transcriptional, and mass-cytometric profiling, it remains a challenge to collect highly multiplexed measurements of secreted proteins from single cells for comprehensive analysis of functional states. Herein, we combine spatial and spectral encoding with polydimethylsiloxane (PDMS) microchambers for codetection of 42 immune effector proteins secreted from single cells, representing the highest multiplexing recorded to date for a single-cell secretion assay. Using this platform to profile differentiated macrophages stimulated with lipopolysaccharide (LPS), the ligand of Toll-like receptor 4 (TLR4), reveals previously unobserved deep functional heterogeneity and varying levels of pathogenic activation. Uniquely protein profiling on the same single cells before and after LPS stimulation identified a role for macrophage inhibitory factor (MIF) to potentiate the activation of LPS-induced cytokine production. Advanced clustering analysis identified functional subsets including quiescent, polyfunctional fully activated, partially activated populations with different cytokine profiles. This population architecture is conserved throughout the cell activation process and prevails as it is extended to other TLR ligands and to primary macrophages derived from a healthy donor. This work demonstrates that the phenotypically similar cell population still exhibits a large degree of intrinsic heterogeneity at the functional and cell behavior level. This technology enables full-spectrum dissection of immune functional states in response to pathogenic or environmental stimulation, and opens opportunities to quantify deep functional heterogeneity for more comprehensive and accurate immune monitoring. PMID:25646488

  2. Isolation and identification of highly pathogenic avian influenza virus subtype H5N1 in peafowl (Pavo cristatus).

    PubMed

    Ismail, Mahmoud Moussa; Khan, Owais Ahmed; Cattoli, Giovanni; Lu, Huaguang

    2010-03-01

    An outbreak of highly pathogenic avian influenza (HPAI) virus subtype H5N1 was first diagnosed in a "backyard" flock of peafowl (Pavo cristatus) raised on palace premises in the Kingdom of Saudi Arabia in December 3, 2007. The flock consisted of 40 peafowl, and their ages ranged from 3 to 5 years old. Affected birds suffered from depression, anorexia, and white diarrhea. Four dead birds were submitted for HPAI diagnosis at the Central Veterinary Diagnostic Laboratory in Riyadh. Brain and liver tissues and tracheal and cloacal swabs were taken from the dead birds and processed for a real-time reverse transcriptase (RT)-PCR test and virus isolation in specific-pathogen-free embryonating chicken eggs. The H5N1 subtype of avian influenza virus was isolated from the four dead birds and identified by a real-time RT-PCR before and after egg inoculation. The virus isolates were characterized as HPAI H5N1 virus by sequencing analysis. Phylogenetic comparisons revealed that the H5N1 viruses isolated from peafowl belong to the genetic clade 2.2 according to the World Health Organization nomenclature. The peafowl H5N1 virus falls into 2.2.2 sublineage II and clusters with the H5N1 viruses isolated from poultry in Saudi Arabia in 2007-08.

  3. Segmented continuous-flow multiplex polymerase chain reaction microfluidics for high-throughput and rapid foodborne pathogen detection.

    PubMed

    Shu, Bowen; Zhang, Chunsun; Xing, Da

    2014-05-15

    High-throughput and rapid identification of multiple foodborne bacterial pathogens is vital in global public health and food industry. To fulfill this need, we propose a segmented continuous-flow multiplex polymerase chain reaction (SCF-MPCR) on a spiral-channel microfluidic device. The device consists of a disposable polytetrafluoroethylene (PTFE) capillary microchannel coiled on three isothermal blocks. Within the channel, n segmented flow regimes are sequentially generated, and m-plex PCR is individually performed in each regime when each mixture is driven to pass three temperature zones, thus providing a rapid analysis throughput of m×n. To characterize the performance of the microfluidic device, continuous-flow multiplex PCR in a single segmented flow has been evaluated by investigating the effect of key reaction parameters, including annealing temperatures, flow rates, polymerase concentration and amount of input DNA. With the optimized parameters, the genomic DNAs from Salmonella enterica, Listeria monocytogenes, Escherichia coli O157:H7 and Staphylococcus aureus could be amplified simultaneously in 19min, and the limit of detection was low, down to 10(2) copiesμL(-1). As proof of principle, the spiral-channel SCF-MPCR was applied to sequentially amplify four different bacterial pathogens from banana, milk, and sausage, displaying a throughput of 4×3 with no detectable cross-contamination.

  4. Segmented continuous-flow multiplex polymerase chain reaction microfluidics for high-throughput and rapid foodborne pathogen detection.

    PubMed

    Shu, Bowen; Zhang, Chunsun; Xing, Da

    2014-05-15

    High-throughput and rapid identification of multiple foodborne bacterial pathogens is vital in global public health and food industry. To fulfill this need, we propose a segmented continuous-flow multiplex polymerase chain reaction (SCF-MPCR) on a spiral-channel microfluidic device. The device consists of a disposable polytetrafluoroethylene (PTFE) capillary microchannel coiled on three isothermal blocks. Within the channel, n segmented flow regimes are sequentially generated, and m-plex PCR is individually performed in each regime when each mixture is driven to pass three temperature zones, thus providing a rapid analysis throughput of m×n. To characterize the performance of the microfluidic device, continuous-flow multiplex PCR in a single segmented flow has been evaluated by investigating the effect of key reaction parameters, including annealing temperatures, flow rates, polymerase concentration and amount of input DNA. With the optimized parameters, the genomic DNAs from Salmonella enterica, Listeria monocytogenes, Escherichia coli O157:H7 and Staphylococcus aureus could be amplified simultaneously in 19min, and the limit of detection was low, down to 10(2) copiesμL(-1). As proof of principle, the spiral-channel SCF-MPCR was applied to sequentially amplify four different bacterial pathogens from banana, milk, and sausage, displaying a throughput of 4×3 with no detectable cross-contamination. PMID:24793853

  5. Highly pathogenic avian influenza H5N1 virus delays apoptotic responses via activation of STAT3

    PubMed Central

    Hui, Kenrie P. Y.; Li, Hung Sing; Cheung, Man Chun; Chan, Renee W. Y.; Yuen, Kit M.; Mok, Chris K. P.; Nicholls, John M.; Peiris, J. S. Malik; Chan, Michael C. W.

    2016-01-01

    Highly pathogenic avian influenza (HPAI) H5N1 virus continues to pose pandemic threat, but there is a lack of understanding of its pathogenesis. We compared the apoptotic responses triggered by HPAI H5N1 and low pathogenic H1N1 viruses using physiologically relevant respiratory epithelial cells. We demonstrated that H5N1 viruses delayed apoptosis in primary human bronchial and alveolar epithelial cells (AECs) compared to H1N1 virus. Both caspase-8 and -9 were activated by H5N1 and H1N1 viruses in AECs, while H5N1 differentially up-regulated TRAIL. H5N1-induced apoptosis was reduced by TRAIL receptor silencing. More importantly, STAT3 knock-down increased apoptosis by H5N1 infection suggesting that H5N1 virus delays apoptosis through activation of STAT3. Taken together, we demonstrate that STAT3 is involved in H5N1-delayed apoptosis compared to H1N1. Since delay in apoptosis prolongs the duration of virus replication and production of pro-inflammatory cytokines and TRAIL from H5N1-infected cells, which contribute to orchestrate cytokine storm and tissue damage, our results suggest that STAT3 may play a previously unsuspected role in H5N1 pathogenesis. PMID:27344974

  6. Isolation and identification of highly pathogenic avian influenza virus subtype H5N1 in peafowl (Pavo cristatus).

    PubMed

    Ismail, Mahmoud Moussa; Khan, Owais Ahmed; Cattoli, Giovanni; Lu, Huaguang

    2010-03-01

    An outbreak of highly pathogenic avian influenza (HPAI) virus subtype H5N1 was first diagnosed in a "backyard" flock of peafowl (Pavo cristatus) raised on palace premises in the Kingdom of Saudi Arabia in December 3, 2007. The flock consisted of 40 peafowl, and their ages ranged from 3 to 5 years old. Affected birds suffered from depression, anorexia, and white diarrhea. Four dead birds were submitted for HPAI diagnosis at the Central Veterinary Diagnostic Laboratory in Riyadh. Brain and liver tissues and tracheal and cloacal swabs were taken from the dead birds and processed for a real-time reverse transcriptase (RT)-PCR test and virus isolation in specific-pathogen-free embryonating chicken eggs. The H5N1 subtype of avian influenza virus was isolated from the four dead birds and identified by a real-time RT-PCR before and after egg inoculation. The virus isolates were characterized as HPAI H5N1 virus by sequencing analysis. Phylogenetic comparisons revealed that the H5N1 viruses isolated from peafowl belong to the genetic clade 2.2 according to the World Health Organization nomenclature. The peafowl H5N1 virus falls into 2.2.2 sublineage II and clusters with the H5N1 viruses isolated from poultry in Saudi Arabia in 2007-08. PMID:20521659

  7. High-content image-based screening of a signal transduction pathway inhibitor small-molecule library against highly pathogenic RNA viruses.

    PubMed

    Mudhasani, Rajini; Kota, Krishna P; Retterer, Cary; Tran, Julie P; Tritsch, Sarah R; Zamani, Rouzbeh; Whitehouse, Chris A; Bavari, Sina

    2015-01-01

    High-content image-based screening was developed as an approach to test a small-molecule library of compounds targeting signal transduction pathways for antiviral activity against multiple highly pathogenic RNA viruses. Of the 2843 compounds screened, 120 compounds exhibited ≥60% antiviral activity. Four compounds (E225-0969, E528-0039, G118-0778, and G544-0735), which were most active against Rift Valley fever virus (RVFV) and showed broad-spectrum antiviral activity, were selected for further evaluation for their concentration-response profile and cytotoxicity. These compounds did not show any visible cytotoxicity at the highest concentration of compound tested (200 µM). All four of these compounds were more active than ribavirin against several viruses. One compound, E225-0969, had the lowest effective concentration (EC50 = 1.9-8.92 µM) for all the viruses tested. This compound was 13- and 43-fold more inhibitory against RVFV and Chikungunya virus (CHIKV), respectively, than ribavirin. The highest selectivity index (>106.2) was for E225-0969 against CHIKV. Time-of-addition assays suggested that all four lead compounds targeted early steps in the viral life cycle (entry and/or replication) but not virus egress. Overall, this work demonstrates that high-content image analysis can be used to screen chemical libraries for new antivirals against highly pathogenic viruses.

  8. When private actors matter: Information-sharing network and surveillance of Highly Pathogenic Avian Influenza in Vietnam.

    PubMed

    Delabouglise, A; Dao, T H; Truong, D B; Nguyen, T T; Nguyen, N T X; Duboz, R; Fournié, G; Antoine-Moussiaux, N; Grosbois, V; Vu, D T; Le, T H; Nguyen, V K; Salem, G; Peyre, M

    2015-07-01

    The effectiveness of animal health surveillance systems depends on their capacity to gather sanitary information from the animal production sector. In order to assess this capacity we analyzed the flow of sanitary information regarding Highly Pathogenic Avian Influenza (HPAI) suspicions in poultry in Vietnam. Participatory methods were applied to assess the type of actors and likelihood of information sharing between actors in case of HPAI suspicion in poultry. While the reporting of HPAI suspicions is mandatory, private actors had more access to information than public actors. Actors of the upstream sector (medicine and feed sellers) played a key role in the diffusion of information. The central role of these actors and the influence of the information flow on the adoption by poultry production stakeholders of behaviors limiting (e.g. prevention measures) or promoting disease transmission (e.g. increased animal movements) should be accounted for in the design of surveillance and control programs. PMID:25847263

  9. Insight into Alternative Approaches for Control of Avian Influenza in Poultry, with Emphasis on Highly Pathogenic H5N1

    PubMed Central

    Abdelwhab, E. M.; Hafez, Hafez M.

    2012-01-01

    Highly pathogenic avian influenza virus (HPAIV) of subtype H5N1 causes a devastating disease in poultry but when it accidentally infects humans it can cause death. Therefore, decrease the incidence of H5N1 in humans needs to focus on prevention and control of poultry infections. Conventional control strategies in poultry based on surveillance, stamping out, movement restriction and enforcement of biosecurity measures did not prevent the virus spreading, particularly in developing countries. Several challenges limit efficiency of the vaccines to prevent outbreaks of HPAIV H5N1 in endemic countries. Alternative and complementary approaches to reduce the current burden of H5N1 epidemics in poultry should be encouraged. The use of antiviral chemotherapy and natural compounds, avian-cytokines, RNA interference, genetic breeding and/or development of transgenic poultry warrant further evaluation as integrated intervention strategies for control of HPAIV H5N1 in poultry. PMID:23202521

  10. Pathology of highly pathogenic avian influenza virus (H5N1) infection in Canada geese (Branta canadensis): preliminary studies.

    PubMed

    Neufeld, J L; Embury-Hyatt, C; Berhane, Y; Manning, L; Ganske, S; Pasick, J

    2009-09-01

    Susceptibility of Canada geese (Branta canadensis) to highly pathogenic avian influenza (HPAI) virus (H5N1) infection was studied by inoculating 10 naïve (antibody-negative) animals (5 adults and 5 juveniles) with A/chicken/Vietnam/14/05 (H5N1) virus. In the adults, 1 of 5 became infected, and 4 of 5 remained normal; in the juvenile group, 5 of 5 became infected. The pathology observed in the affected animals was similar to that reported in natural occurrences. Peripheral and parasympathetic nervous systems were examined and found infected, as well as cerebrospinal fluid-contacting neurons. In some locations with significant virus infection in cells, the expected inflammatory reaction was absent or very mild. Immunohistochemistry was used to locate influenza A virus nucleoprotein in brain, spinal cord, respiratory and digestive systems, pancreas, heart, and peripheral and parasympathetic nervous systems. Further studies are needed to explain age-related differences in susceptibility.

  11. When private actors matter: Information-sharing network and surveillance of Highly Pathogenic Avian Influenza in Vietnam.

    PubMed

    Delabouglise, A; Dao, T H; Truong, D B; Nguyen, T T; Nguyen, N T X; Duboz, R; Fournié, G; Antoine-Moussiaux, N; Grosbois, V; Vu, D T; Le, T H; Nguyen, V K; Salem, G; Peyre, M

    2015-07-01

    The effectiveness of animal health surveillance systems depends on their capacity to gather sanitary information from the animal production sector. In order to assess this capacity we analyzed the flow of sanitary information regarding Highly Pathogenic Avian Influenza (HPAI) suspicions in poultry in Vietnam. Participatory methods were applied to assess the type of actors and likelihood of information sharing between actors in case of HPAI suspicion in poultry. While the reporting of HPAI suspicions is mandatory, private actors had more access to information than public actors. Actors of the upstream sector (medicine and feed sellers) played a key role in the diffusion of information. The central role of these actors and the influence of the information flow on the adoption by poultry production stakeholders of behaviors limiting (e.g. prevention measures) or promoting disease transmission (e.g. increased animal movements) should be accounted for in the design of surveillance and control programs.

  12. Supporting business continuity during a highly pathogenic avian influenza outbreak: a collaboration of industry, academia, and government.

    PubMed

    Hennessey, Morgan; Lee, Brendan; Goldsmith, Timothy; Halvorson, Dave; Hueston, William; McElroy, Kristina; Waters, Katherine

    2010-03-01

    Since 2006, a collaborative group of egg industry, state, federal, and academia representatives have worked to enhance preparedness in highly pathogenic avian influenza (HPAI) planning. The collaborative group has created a draft egg product movement protocol, which calls for realistic, science-based contingency plans, biosecurity assessments, commodity risk assessments, and real-time reverse transcriptase-PCR testing to support the continuity of egg operations while also preventing and eradicating an HPAI outbreak. The work done by this group serves as an example of how industry, government, and academia can work together to achieve better preparedness in the event of an animal health emergency. In addition, in the event of an HPAI outbreak in domestic poultry, U.S. consumers will be assured that their egg products come from healthy chickens.

  13. High-throughput screening of metal-N-heterocyclic carbene complexes against biofilm formation by pathogenic bacteria.

    PubMed

    Bernardi, Thierry; Badel, Stéphanie; Mayer, Pascal; Groelly, Jérome; de Frémont, Pierre; Jacques, Béatrice; Braunstein, Pierre; Teyssot, Marie-Laure; Gaulier, Christelle; Cisnetti, Federico; Gautier, Arnaud; Roland, Sylvain

    2014-06-01

    A set of molecules including a majority of metal-N-heterocyclic carbene (NHC) complexes (metal=Ag, Cu, and Au) and azolium salts were evaluated by high-throughput screening of their activity against biofilm formation associated with pathogenic bacteria. The anti-planktonic effects were compared in parallel. Representative biofilm-forming strains of various genera were selected (Listeria, Pseudomonas, Staphylococcus, and Escherichia). All the compounds were tested at 1 mg L(-1) by using the BioFilm Ring Test. An information score (IS, sum of the activities) and an activity score (AS, difference between anti-biofilm and anti-planktonic activity) were determined from normalized experimental values to classify the most active molecules against the panel of bacterial strains. With this method we identified lipophilic Ag(I) and Cu(I) complexes possessing aromatic groups on the NHC ligand as the most efficient at inhibiting biofilm formation. PMID:24729552

  14. Risk for infection with highly pathogenic influenza A virus (H5N1) in chickens, Hong Kong, 2002.

    PubMed

    Kung, Nina Y; Morris, Roger S; Perkins, Nigel R; Sims, Les D; Ellis, Trevor M; Bissett, Lucy; Chow, Mary; Shortridge, Ken F; Guan, Yi; Peiris, Malik J S

    2007-03-01

    We used epidemiologic evaluation, molecular epidemiology, and a case-control study to identify possible risk factors for the spread of highly pathogenic avian influenza A virus (subtype H5N1) in chicken farms during the first quarter of 2002 in Hong Kong. Farm profiles, including stock sources, farm management, and biosecurity measures, were collected from 16 case and 46 control chicken farms by using a pretested questionnaire and personal interviews. The risk for influenza A (H5N1) infection was assessed by using adjusted odds ratios based on multivariate logistic regression analysis. Retail marketing of live poultry was implicated as the main source of exposure to infection on chicken farms in Hong Kong during this period. Infection control measures should be reviewed and upgraded as necessary to reduce the spread of influenza A (H5N1) related to live poultry markets, which are commonplace across Asia.

  15. Stochastic model of the potential spread of highly pathogenic avian influenza from an infected commercial broiler operation in Georgia.

    PubMed

    Dorea, F C; Vieira, A R; Hofacre, C; Waldrip, D; Cole, D J

    2010-03-01

    The potential spread of highly pathogenic avian influenza among commercial broiler farms in Georgia, U. S. A., was mathematically modeled. The dynamics of the spread within the first infected flock were estimated using an SEIR (susceptible-exposed-infectious-recovered) deterministic model, and predicted that grower detection of flock infection is most likely 5 days after virus introduction. Off-farm spread of virus was estimated stochastically for this period, predicting a mean range of exposed farms from 0-5, depending on the density of farms in the area. Modeled off-farm spread was most frequently associated with feed trucks (highest daily probability and number of farm visits) and with company personnel or hired help (highest level of bird contact).

  16. Novel Reassortant H5N6 Influenza A Virus from the Lao People’s Democratic Republic Is Highly Pathogenic in Chickens

    PubMed Central

    Layton, Daniel S.; Phommachanh, Phouvong; Harper, Jennifer; Payne, Jean; Evans, Ryan M.; Valdeter, Stacey; Walker, Som; Harvey, Gemma; Shan, Songhua; Bruce, Matthew P.; Rootes, Christina L.; Gough, Tamara J.; Rohringer, Andreas; Peck, Grantley R.; Fardy, Sarah J.; Karpala, Adam J.; Johnson, Dayna; Wang, Jianning; Douangngeun, Bounlom; Morrissy, Christopher; Wong, Frank Y. K.; Bean, Andrew G. D.; Bingham, John; Williams, David T.

    2016-01-01

    Avian influenza viruses of H5 subtype can cause highly pathogenic disease in poultry. In March 2014, a new reassortant H5N6 subtype highly pathogenic avian influenza virus emerged in Lao People’s Democratic Republic. We have assessed the pathogenicity, pathobiology and immunological responses associated with this virus in chickens. Infection caused moderate to advanced disease in 6 of 6 chickens within 48 h of mucosal inoculation. High virus titers were observed in blood and tissues (kidney, spleen, liver, duodenum, heart, brain and lung) taken at euthanasia. Viral antigen was detected in endothelium, neurons, myocardium, lymphoid tissues and other cell types. Pro-inflammatory cytokines were elevated compared to non-infected birds. Our study confirmed that this new H5N6 reassortant is highly pathogenic, causing disease in chickens similar to that of Asian H5N1 viruses, and demonstrated the ability of such clade 2.3.4-origin H5 viruses to reassort with non-N1 subtype viruses while maintaining a fit and infectious phenotype. Recent detection of influenza H5N6 poultry infections in Lao PDR, China and Viet Nam, as well as six fatal human infections in China, demonstrate that these emergent highly pathogenic H5N6 viruses may be widely established in several countries and represent an emerging threat to poultry and human populations. PMID:27631618

  17. The performance characteristics of lateral flow devices with 2 strains of highly pathogenic avian influenza virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Lateral flow devices (LFD) are commercially available and provide a fast, highly specific, on-site test for avian influenza. Because of the low analytic sensitivity of LFD tests at low virus concentrations, targeted sampling of sick and dead birds has been proposed in order to increase detection pr...

  18. Spatio-temporal epidemiology of highly pathogenic avian influenza (subtype H5N1) in poultry in eastern India.

    PubMed

    Dhingra, Madhur S; Dissanayake, Ravi; Negi, Ajender Bhagat; Oberoi, Mohinder; Castellan, David; Thrusfield, Michael; Linard, Catherine; Gilbert, Marius

    2014-10-01

    In India, majority outbreaks of highly pathogenic avian influenza (HPAI) H5N1 have occurred in eastern states of West Bengal, Assam and Tripura. This study aimed to identify disease clusters and risk factors of HPAI H5N1 in these states, for targeted surveillance and disease control. A spatial scan statistic identified two significant disease clusters in West Bengal and Assam, occurring during January and November-December 2008, respectively. Key risk factors were identified at sub-district level using bootstrapped logistic regression and boosted regression trees model. With both methods, HPAI H5N1 outbreaks in backyard poultry were associated with accessibility in terms of time taken to access a city with >50,000 persons, human population density and duck density (P<0.005). In addition, areas at lower elevation were also identified as high risk by BRT model. It is recommended that risk-based surveillance should be implemented in high duck density areas and all live-bird markets in high-throughput locations.

  19. Spatial Modeling of Wild Bird Risk Factors for Highly Pathogenic A(H5N1) Avian Influenza Virus Transmission.

    PubMed

    Prosser, Diann J; Hungerford, Laura L; Erwin, R Michael; Ottinger, Mary Ann; Takekawa, John Y; Newman, Scott H; Xiao, Xiangming; Ellis, Erle C

    2016-05-01

    One of the longest-persisting avian influenza viruses in history, highly pathogenic avian influenza virus (HPAIV) A(H5N1), continues to evolve after 18 yr, advancing the threat of a global pandemic. Wild waterfowl (family Anatidae) are reported as secondary transmitters of HPAIV and primary reservoirs for low-pathogenic avian influenza viruses, yet spatial inputs for disease risk modeling for this group have been lacking. Using geographic information software and Monte Carlo simulations, we developed geospatial indices of waterfowl abundance at 1 and 30 km resolutions and for the breeding and wintering seasons for China, the epicenter of H5N1. Two spatial layers were developed: cumulative waterfowl abundance (WAB), a measure of predicted abundance across species, and cumulative abundance weighted by H5N1 prevalence (WPR), whereby abundance for each species was adjusted based on prevalence values and then totaled across species. Spatial patterns of the model output differed between seasons, with higher WAB and WPR in the northern and western regions of China for the breeding season and in the southeast for the wintering season. Uncertainty measures indicated highest error in southeastern China for both WAB and WPR. We also explored the effect of resampling waterfowl layers from 1 to 30 km resolution for multiscale risk modeling. Results indicated low average difference (less than 0.16 and 0.01 standard deviations for WAB and WPR, respectively), with greatest differences in the north for the breeding season and southeast for the wintering season. This work provides the first geospatial models of waterfowl abundance available for China. The indices provide important inputs for modeling disease transmission risk at the interface of poultry and wild birds. These models are easily adaptable, have broad utility to both disease and conservation needs, and will be available to the scientific community for advanced modeling applications.

  20. High-affinity, noninhibitory pathogenic C1 domain antibodies are present in patients with hemophilia A and inhibitors

    PubMed Central

    Batsuli, Glaivy; Deng, Wei; Healey, John F.; Parker, Ernest T.; Baldwin, W. Hunter; Cox, Courtney; Nguyen, Brenda; Kahle, Joerg; Königs, Christoph; Li, Renhao; Lollar, Pete

    2016-01-01

    Inhibitor formation in hemophilia A is the most feared treatment-related complication of factor VIII (fVIII) therapy. Most inhibitor patients with hemophilia A develop antibodies against the fVIII A2 and C2 domains. Recent evidence demonstrates that the C1 domain contributes to the inhibitor response. Inhibitory anti-C1 monoclonal antibodies (mAbs) have been identified that bind to putative phospholipid and von Willebrand factor (VWF) binding epitopes and block endocytosis of fVIII by antigen presenting cells. We now demonstrate by competitive enzyme-linked immunosorbent assay and hydrogen-deuterium exchange mass spectrometry that 7 of 9 anti-human C1 mAbs tested recognize an epitope distinct from the C1 phospholipid binding site. These mAbs, designated group A, display high binding affinities for fVIII, weakly inhibit fVIII procoagulant activity, poorly inhibit fVIII binding to phospholipid, and exhibit heterogeneity with respect to blocking fVIII binding to VWF. Another mAb, designated group B, inhibits fVIII procoagulant activity, fVIII binding to VWF and phospholipid, fVIIIa incorporation into the intrinsic Xase complex, thrombin generation in plasma, and fVIII uptake by dendritic cells. Group A and B epitopes are distinct from the epitope recognized by the canonical, human-derived inhibitory anti-C1 mAb, KM33, whose epitope overlaps both groups A and B. Antibodies recognizing group A and B epitopes are present in inhibitor plasmas from patients with hemophilia A. Additionally, group A and B mAbs increase fVIII clearance and are pathogenic in a hemophilia A mouse tail snip bleeding model. Group A anti-C1 mAbs represent the first identification of pathogenic, weakly inhibitory antibodies that increase fVIII clearance. PMID:27381905

  1. Vaccination with recombinant RNA replicon particles protects chickens from H5N1 highly pathogenic avian influenza virus.

    PubMed

    Halbherr, Stefan J; Brostoff, Terza; Tippenhauer, Merve; Locher, Samira; Berger Rentsch, Marianne; Zimmer, Gert

    2013-01-01

    Highly pathogenic avian influenza viruses (HPAIV) of subtype H5N1 not only cause a devastating disease in domestic chickens and turkeys but also pose a continuous threat to public health. In some countries, H5N1 viruses continue to circulate and evolve into new clades and subclades. The rapid evolution of these viruses represents a problem for virus diagnosis and control. In this work, recombinant vesicular stomatitis virus (VSV) vectors expressing HA of subtype H5 were generated. To comply with biosafety issues the G gene was deleted from the VSV genome. The resulting vaccine vector VSV*ΔG(HA) was propagated on helper cells providing the VSV G protein in trans. Vaccination of chickens with a single intramuscular dose of 2×10⁸ infectious replicon particles without adjuvant conferred complete protection from lethal H5N1 infection. Subsequent application of the same vaccine strongly boosted the humoral immune response and completely prevented shedding of challenge virus and transmission to sentinel birds. The vaccine allowed serological differentiation of infected from vaccinated animals (DIVA) by employing a commercially available ELISA. Immunized chickens produced antibodies with neutralizing activity against multiple H5 viruses representing clades 1, 2.2, 2.5, and low-pathogenic avian influenza viruses (classical clade). Studies using chimeric H1/H5 hemagglutinins showed that the neutralizing activity was predominantly directed against the globular head domain. In summary, these results suggest that VSV replicon particles are safe and potent DIVA vaccines that may help to control avian influenza viruses in domestic poultry.

  2. X-ray structure of NS1 from a highly pathogenic H5N1 influenza virus.

    PubMed

    Bornholdt, Zachary A; Prasad, B V Venkataram

    2008-12-18

    The recent emergence of highly pathogenic avian (H5N1) influenza viruses, their epizootic and panzootic nature, and their association with lethal human infections have raised significant global health concerns. Several studies have underlined the importance of non-structural protein NS1 in the increased pathogenicity and virulence of these strains. NS1, which consists of two domains-a double-stranded RNA (dsRNA) binding domain and the effector domain, separated through a linker-is an antagonist of antiviral type-I interferon response in the host. Here we report the X-ray structure of the full-length NS1 from an H5N1 strain (A/Vietnam/1203/2004) that was associated with 60% of human deaths in an outbreak in Vietnam. Compared to the individually determined structures of the RNA binding domain and the effector domain from non-H5N1 strains, the RNA binding domain within H5N1 NS1 exhibits modest structural changes, while the H5N1 effector domain shows significant alteration, particularly in the dimeric interface. Although both domains in the full-length NS1 individually participate in dimeric interactions, an unexpected finding is that these interactions result in the formation of a chain of NS1 molecules instead of distinct dimeric units. Three such chains in the crystal interact with one another extensively to form a tubular organization of similar dimensions to that observed in the cryo-electron microscopy images of NS1 in the presence of dsRNA. The tubular oligomeric organization of NS1, in which residues implicated in dsRNA binding face a 20-A-wide central tunnel, provides a plausible mechanism for how NS1 sequesters varying lengths of dsRNA, to counter cellular antiviral dsRNA response pathways, while simultaneously interacting with other cellular ligands during an infection.

  3. X-ray structure of NS1 from a highly pathogenic H5N1 influenza virus

    SciTech Connect

    Bornholdt, Zachary A.; Prasad, B.V. Venkataram

    2009-04-08

    The recent emergence of highly pathogenic avian (H5N1) influenza viruses, their epizootic and panzootic nature, and their association with lethal human infections have raised significant global health concerns. Several studies have underlined the importance of non-structural protein NS1 in the increased pathogenicity and virulence of these strains. NS1, which consists of two domains - a double-stranded RNA (dsRNA) binding domain and the effector domain, separated through a linker - is an antagonist of antiviral type-I interferon response in the host. Here we report the X-ray structure of the full-length NS1 from an H5N1 strain (A/Vietnam/1203/2004) that was associated with 60% of human deaths in an outbreak in Vietnam. Compared to the individually determined structures of the RNA binding domain and the effector domain from non-H5N1 strains, the RNA binding domain within H5N1 NS1 exhibits modest structural changes, while the H5N1 effector domain shows significant alteration, particularly in the dimeric interface. Although both domains in the full-length NS1 individually participate in dimeric interactions, an unexpected finding is that these interactions result in the formation of a chain of NS1 molecules instead of distinct dimeric units. Three such chains in the crystal interact with one another extensively to form a tubular organization of similar dimensions to that observed in the cryo-electron microscopy images of NS1 in the presence of dsRNA. The tubular oligomeric organization of NS1, in which residues implicated in dsRNA binding face a 20-{angstrom}-wide central tunnel, provides a plausible mechanism for how NS1 sequesters varying lengths of dsRNA, to counter cellular antiviral dsRNA response pathways, while simultaneously interacting with other cellular ligands during an infection.

  4. Spatial Modeling of Wild Bird Risk Factors for Highly Pathogenic A(H5N1) Avian Influenza Virus Transmission.

    PubMed

    Prosser, Diann J; Hungerford, Laura L; Erwin, R Michael; Ottinger, Mary Ann; Takekawa, John Y; Newman, Scott H; Xiao, Xiangming; Ellis, Erle C

    2016-05-01

    One of the longest-persisting avian influenza viruses in history, highly pathogenic avian influenza virus (HPAIV) A(H5N1), continues to evolve after 18 yr, advancing the threat of a global pandemic. Wild waterfowl (family Anatidae) are reported as secondary transmitters of HPAIV and primary reservoirs for low-pathogenic avian influenza viruses, yet spatial inputs for disease risk modeling for this group have been lacking. Using geographic information software and Monte Carlo simulations, we developed geospatial indices of waterfowl abundance at 1 and 30 km resolutions and for the breeding and wintering seasons for China, the epicenter of H5N1. Two spatial layers were developed: cumulative waterfowl abundance (WAB), a measure of predicted abundance across species, and cumulative abundance weighted by H5N1 prevalence (WPR), whereby abundance for each species was adjusted based on prevalence values and then totaled across species. Spatial patterns of the model output differed between seasons, with higher WAB and WPR in the northern and western regions of China for the breeding season and in the southeast for the wintering season. Uncertainty measures indicated highest error in southeastern China for both WAB and WPR. We also explored the effect of resampling waterfowl layers from 1 to 30 km resolution for multiscale risk modeling. Results indicated low average difference (less than 0.16 and 0.01 standard deviations for WAB and WPR, respectively), with greatest differences in the north for the breeding season and southeast for the wintering season. This work provides the first geospatial models of waterfowl abundance available for China. The indices provide important inputs for modeling disease transmission risk at the interface of poultry and wild birds. These models are easily adaptable, have broad utility to both disease and conservation needs, and will be available to the scientific community for advanced modeling applications. PMID:27309075

  5. Vaccination with Recombinant RNA Replicon Particles Protects Chickens from H5N1 Highly Pathogenic Avian Influenza Virus

    PubMed Central

    Halbherr, Stefan J.; Brostoff, Terza; Tippenhauer, Merve; Locher, Samira; Berger Rentsch, Marianne; Zimmer, Gert

    2013-01-01

    Highly pathogenic avian influenza viruses (HPAIV) of subtype H5N1 not only cause a devastating disease in domestic chickens and turkeys but also pose a continuous threat to public health. In some countries, H5N1 viruses continue to circulate and evolve into new clades and subclades. The rapid evolution of these viruses represents a problem for virus diagnosis and control. In this work, recombinant vesicular stomatitis virus (VSV) vectors expressing HA of subtype H5 were generated. To comply with biosafety issues the G gene was deleted from the VSV genome. The resulting vaccine vector VSV*ΔG(HA) was propagated on helper cells providing the VSV G protein in trans. Vaccination of chickens with a single intramuscular dose of 2×108 infectious replicon particles without adjuvant conferred complete protection from lethal H5N1 infection. Subsequent application of the same vaccine strongly boosted the humoral immune response and completely prevented shedding of challenge virus and transmission to sentinel birds. The vaccine allowed serological differentiation of infected from vaccinated animals (DIVA) by employing a commercially available ELISA. Immunized chickens produced antibodies with neutralizing activity against multiple H5 viruses representing clades 1, 2.2, 2.5, and low-pathogenic avian influenza viruses (classical clade). Studies using chimeric H1/H5 hemagglutinins showed that the neutralizing activity was predominantly directed against the globular head domain. In summary, these results suggest that VSV replicon particles are safe and potent DIVA vaccines that may help to control avian influenza viruses in domestic poultry. PMID:23762463

  6. Multiplex quantitative foodborne pathogen detection using high resolution CE-SSCP coupled stuffer-free multiplex ligation-dependent probe amplification.

    PubMed

    Chung, Boram; Shin, Gi Won; Na, Jeongkyeong; Oh, Mi-Hwa; Jung, Gyoo Yeol

    2012-05-01

    Sensitive multiplex detection methods for foodborne pathogens are important in controlling food safety, and detection of genetic markers is accepted to be one of the best tools for sensitive detection. Although CE technology offers great potential in terms of sensitive multiplex detection, the necessary amplification is confined to markers sharing common primers such as the 16S rRNA gene. For precise and sensitive detection, pathogen-specific genes are optimal markers. Although multiplex ligation-dependent probe amplification (MLPA) is appropriate for amplification of specific markers, the requirement for stuffers, to ensure length-dependent separation on CE, is a major obstacle in detection of foodborne pathogens. In the present study, we developed stuffer-free MLPA using high-resolution CE-SSCP to sensitively detect ten foodborne pathogens. The probe set for MLPA prior to CE-SSCP analysis was designed for species-specific detection. After careful optimization of each MLPA step, to ensure that CE-SSCP analysis was informative, we found that all ten pathogens could be reliably identified; the limits of detection were 0.5-5 pg of genomic DNA, and more than 100-fold increase could be quantitatively determined. Thus, MLPA-CE-SSCP is a sensitive and reliable technique for pathogen detection.

  7. PB1-F2 Attenuates Virulence of Highly Pathogenic Avian H5N1 Influenza Virus in Chickens

    PubMed Central

    Leymarie, Olivier; Embury-Hyatt, Carissa; Chevalier, Christophe; Jouneau, Luc; Moroldo, Marco; Da Costa, Bruno; Berhane, Yohannes; Delmas, Bernard

    2014-01-01

    Highly pathogenic avian influenza virus (HPAIV) is a permanent threat due to its capacity to cross species barriers and generate severe infections and high mortality in humans. Recent findings have highlighted the potential role of PB1-F2, a small accessory influenza protein, in the pathogenesis process mediated by HPAIV in mammals. In this study, using a recombinant H5N1 HPAIV (wt) and its PB1-F2-deleted mutant (ΔF2), we studied the effects of PB1-F2 in a chicken model. Unexpectedly, when using low inoculation dose we observed that the wt-infected chickens had a higher survival rate than the ΔF2-infected chickens, a feature that contrasts with what is usually observed in mammals. High inoculation dose had similar mortality rate for both viruses, and comparison of the bio-distribution of the two viruses indicated that the expression of PB1-F2 allows a better spreading of the virus within chicken embryos. Transcriptomic profiles of lungs and blood cells were characterized at two days post-infection in chickens inoculated with the wild type (wt) or the ΔF2 mutant viruses. In lungs, the expression of PB1-F2 during the infection induced pathways related to calcium signaling and repressed a large panel of immunological functions. In blood cells, PB1-F2 was associated with a gene signature specific for mitochondrial dysfunction and down-modulated leucocytes activation. Finally we compared the effect of PB1-F2 in lungs of chickens and mice. We identified that gene signature associated to tissue damages is a PB1-F2 feature shared by the two species; by contrast, the early inhibition of immune response mediated by PB1-F2 observed in chickens is not seen in mice. In summary, our data suggest that PB1-F2 expression deeply affect the immune response in chickens in a way that may attenuate pathogenicity at low infection dose, a feature differing from what was previously observed in mammal species. PMID:24959667

  8. Leveraging a high resolution microfluidic assay reveals insights into pathogenic fungal spore germination.

    PubMed

    Barkal, Layla J; Walsh, Naomi M; Botts, Michael R; Beebe, David J; Hull, Christina M

    2016-05-16

    Germination of spores into actively growing cells is a process essential for survival and pathogenesis of many microbes. Molecular mechanisms governing germination, however, are poorly understood in part because few tools exist for evaluating and interrogating the process. Here, we introduce an assay that leverages developments in microfluidic technology and image processing to quantitatively measure germination with unprecedented resolution, assessing both individual cells and the population as a whole. Using spores from Cryptococcus neoformans, a leading cause of fatal fungal disease in humans, we developed a platform to evaluate spores as they undergo morphological changes during differentiation into vegetatively growing yeast. The assay uses pipet-accessible microdevices that can be arrayed for efficient testing of diverse microenvironmental variables, including temperature and nutrients. We discovered that temperature influences germination rate, a carbon source alone is sufficient to induce germination, and the addition of a nitrogen source sustains it. Using this information, we optimized the assay for use with fungal growth inhibitors to pinpoint stages of germination inhibition. Unexpectedly, the clinical antifungal drugs amphotericin B and fluconazole did not significantly alter the process or timing of the transition from spore to yeast, indicating that vegetative growth and germination are distinct processes in C. neoformans. Finally, we used the high temporal resolution of the assay to determine the precise defect in a slow-germination mutant. Combining advances in microfluidics with a robust fungal molecular genetic system allowed us to identify and alter key temporal, morphological, and molecular events that occur during fungal germination. PMID:27026574

  9. Evolution of the Core Genome of Pseudomonas syringae, a Highly Clonal, Endemic Plant Pathogen

    PubMed Central

    Sarkar, Sara F.; Guttman, David S.

    2004-01-01

    Pseudomonas syringae is a common foliar bacterium responsible for many important plant diseases. We studied the population structure and dynamics of the core genome of P. syringae via multilocus sequencing typing (MLST) of 60 strains, representing 21 pathovars and 2 nonpathogens, isolated from a variety of plant hosts. Seven housekeeping genes, dispersed around the P. syringae genome, were sequenced to obtain 400 to 500 nucleotides per gene. Forty unique sequence types were identified, with most strains falling into one of four major clades. Phylogenetic and maximum-likelihood analyses revealed a remarkable degree of congruence among the seven genes, indicating a common evolutionary history for the seven loci. MLST and population genetic analyses also found a very low level of recombination. Overall, mutation was found to be approximately four times more likely than recombination to change any single nucleotide. A skyline plot was used to study the demographic history of P. syringae. The species was found to have maintained a constant population size over time. Strains were also found to remain genetically homogeneous over many years, and when isolated from sites as widespread as the United States and Japan. An analysis of molecular variance found that host association explains only a small proportion of the total genetic variation in the sample. These analyses reveal that with respect to the core genome, P. syringae is a highly clonal and stable species that is endemic within plant populations, yet the genetic variation seen in these genes only weakly predicts host association. PMID:15066790

  10. High-throughput profiling of N-myristoylation substrate specificity across species including pathogens.

    PubMed

    Traverso, José A; Giglione, Carmela; Meinnel, Thierry

    2013-01-01

    One of the most critical modifications affecting the N-terminus of proteins is N-myristoylation. This irreversible modification affects the membrane-binding properties of crucial proteins involved in signal transduction cascades. This cotranslational modification, catalyzed by N-myristoyl transferase, occurs both in lower and higher eukaryotes and is a validated therapeutic target for several pathologies. However, this lipidation proves very difficult to be evidenced in vivo even with state-of-the-art proteomics approaches or bioinformatics tools. A large part of N-myristoylated proteins remains to be discovered and the rules of substrate specificity need to be established in each organism. Because the peptide substrate recognition occurs around the first eight residues, short peptides are used for modeling the reaction in vitro. Here, we provide a novel approach including a dedicated peptide array for high-throughput profiling protein N-myristoylation specificity. We show that myristoylation predictive tools need to be fine-tuned to organisms and that their poor accuracy should be significantly enhanced. This should lead to strongly improved knowledge of the number and function of myristoylated proteins occurring in any proteome.

  11. Highly pathogenic avian influenza viruses do not inhibit interferon synthesis in infected chickens but can override the interferon-induced antiviral state.

    PubMed

    Penski, Nicola; Härtle, Sonja; Rubbenstroth, Dennis; Krohmann, Carsten; Ruggli, Nicolas; Schusser, Benjamin; Pfann, Michael; Reuter, Antje; Gohrbandt, Sandra; Hundt, Jana; Veits, Jutta; Breithaupt, Angele; Kochs, Georg; Stech, Jürgen; Summerfield, Artur; Vahlenkamp, Thomas; Kaspers, Bernd; Staeheli, Peter

    2011-08-01

    From infection studies with cultured chicken cells and experimental mammalian hosts, it is well known that influenza viruses use the nonstructural protein 1 (NS1) to suppress the synthesis of interferon (IFN). However, our current knowledge regarding the in vivo role of virus-encoded NS1 in chickens is much more limited. Here, we report that highly pathogenic avian influenza viruses of subtypes H5N1 and H7N7 lacking fully functional NS1 genes were attenuated in 5-week-old chickens. Surprisingly, in diseased birds infected with NS1 mutants, the IFN levels were not higher than in diseased birds infected with wild-type virus, suggesting that NS1 cannot suppress IFN gene expression in at least one cell population of infected chickens that produces large amounts of the cytokine in vivo. To address the question of why influenza viruses are highly pathogenic in chickens although they strongly activate the innate immune system, we determined whether recombinant chicken alpha interferon (IFN-α) can inhibit the growth of highly pathogenic avian influenza viruses in cultured chicken cells and whether it can ameliorate virus-induced disease in 5-week-old birds. We found that IFN treatment failed to confer substantial protection against challenge with highly pathogenic viruses, although it was effective against viruses with low pathogenic potential. Taken together, our data demonstrate that preventing the synthesis of IFN is not the primary role of the viral NS1 protein during infection of chickens. Our results further suggest that virus-induced IFN does not contribute substantially to resistance of chickens against highly pathogenic influenza viruses.

  12. Attenuated Bordetella pertussis Protects against Highly Pathogenic Influenza A Viruses by Dampening the Cytokine Storm▿

    PubMed Central

    Li, Rui; Lim, Annabelle; Phoon, Meng Chee; Narasaraju, Teluguakula; Ng, Jowin K. W.; Poh, Wee Peng; Sim, Meng Kwoon; Chow, Vincent T.; Locht, Camille; Alonso, Sylvie

    2010-01-01

    The threat of a pandemic spread of highly virulent influenza A viruses currently represents a top global public health problem. Mass vaccination remains the most effective way to combat influenza virus. However, current vaccination strategies face the challenge to meet the demands in a pandemic situation. In a mouse model of severe influenza virus-induced pneumonitis, we observed that prior nasal administration of an attenuated strain of Bordetella pertussis (BPZE1) provided effective and sustained protection against lethal challenge with two different influenza A virus subtypes. In contrast to most cross-protective effects reported so far, the protective window offered upon nasal treatment with BPZE1 lasted up to at least 12 weeks, suggesting a unique mechanism(s) involved in the protection. No significant differences in viral loads were observed between BPZE1-treated and control mice, indicating that the cross-protective mechanism(s) does not directly target the viral particles and/or infected cells. This was further confirmed by the absence of cross-reactive antibodies and T cells in serum transfer and in vitro restimulation experiments, respectively. Instead, compared to infected control mice, BPZE1-treated animals displayed markedly reduced lung inflammation and tissue damage, decreased neutrophil infiltration, and strong suppression of the production of major proinflammatory mediators in their bronchoalveolar fluids (BALFs). Our findings thus indicate that protection against influenza virus-induced severe pneumonitis can be achieved through attenuation of exaggerated cytokine-mediated inflammation. Furthermore, nasal treatment with live attenuated B. pertussis offers a potential alternative to conventional approaches in the fight against one of the most frightening current global public health threats. PMID:20444902

  13. Susceptibility of openbill storks (Anastomius oscitans) to highly pathogenic avian influenza virus subtype H5N1.

    PubMed

    Chaichoun, Kridsada; Wiriyarat, Withawat; Phonaknguen, Rassmeepen; Sariya, Ladawan; Taowan, Nam-aoy; Chakritbudsabong, Warunya; Chaisilp, Natnapat; Eiam-ampai, Krirat; Phuttavatana, Pilaipan; Ratanakorn, Parntep

    2013-09-01

    This investigation detailed the clinical disease, gross and histologic lesions in juvenile openbill storks (Anastomus oscitans) intranasally inoculated with an avian influenza virus, A/chicken/Thailand/vsmu-3 (H5N1), which is highly pathogenic for chickens. High morbidity and mortality were observed in openbill storks inoculated with HPAI H5N1 virus. Gross lesions from infected birds were congestion and brain hemorrhage (10/20), pericardial effusions, pericarditis and focal necrosis of the cardiac muscle (2/20), pulmonary edema and pulmonary necrosis, serosanguineous fluid in the bronchis (16/20), liver congestion (6/20), bursitis (5/20), subcutaneous hemorrhages (2/20) and pinpoint proventiculus hemorrhage (2/20). Real time RT-PCR demonstrated the presence of viral RNA in organs associated with the lesions: brain, trachea, lungs, liver, spleen and intestines. Similar to viral genome detection, virus was also isolated from these vital organs. Antibodies to influenza virus detected with a hemagglutination inhibition test, were found only in the openbill storks who died 8 days post-inoculation.

  14. Living with avian FLU--Persistence of the H5N1 highly pathogenic avian influenza virus in Egypt.

    PubMed

    Njabo, Kevin Yana; Zanontian, Linda; Sheta, Basma N; Samy, Ahmed; Galal, Shereen; Schoenberg, Frederic Paik; Smith, Thomas B

    2016-05-01

    H5N1 highly pathogenic avian influenza virus (HPAIV) continues to cause mortality in poultry and threaten human health at a panzootic scale in Egypt since it was reported in 2006. While the early focus has been in Asia, recent evidence suggests that Egypt is an emerging epicenter for the disease. Despite control measures, epizootic transmission of the disease continues. Here, we investigate the persistence of HPAIV across wild passerine birds and domestic poultry between 2009 and 2012 and the potential risk for continuous viral transmission in Egypt. We use a new weighted cross J-function to investigate the degree and spatial temporal nature of the clustering between sightings of infected birds of different types, and the risk of infection associated with direct contact with infected birds. While we found no infection in wild birds, outbreaks occurred year round between 2009 and 2012, with a positive interaction between chickens and ducks. The disease was more present in the years 2010 and 2011 coinciding with the political unrest in the country. Egypt thus continues to experience endemic outbreaks of avian influenza HPAIV in poultry and an increased potential risk of infection to other species including humans. With the current trends, the elimination of the HPAIV infection is highly unlikely without a complete revamp of current policies. The application of spatial statistics techniques to these types of data may help us to understand the characteristics of the disease and may subsequently allow practitioners to explore possible preventive solutions. PMID:27066713

  15. Assessing the risk of highly pathogenic avian influenza H5N1 transmission through poultry movements in Bali, Indonesia.

    PubMed

    Roche, Sharon E; Cogger, Naomi; Garner, M Graeme; Putra, Anak Agung Gde; Toribio, Jenny-Ann L M L

    2014-03-01

    Indonesia continues to report the highest number of human and poultry cases of highly pathogenic avian influenza H5N1. The disease is considered to be endemic on the island of Bali. Live bird markets are integral in the poultry supply chain on Bali and are important, nutritionally and culturally, for the rural and urban human populations. Due to the lack of biosecurity practiced along the supply chain from producer to live bird markets, there is a need to understand the risks associated with the spread of H5N1 through live bird movements for effective control. Resources to control H5N1 in Indonesia are very limited and cost effective strategies are needed. We assessed the probability a live bird market is infected through live poultry movements and assessed the effects of implementing two simple and low cost control measures on this risk. Results suggest there is a high risk a live bird market is infected (0.78), and risk mitigation strategies such as detecting and removing infected poultry from markets reduce this risk somewhat (range 0.67-0.76). The study demonstrates the key role live poultry movements play in transmitting H5N1 and the need to implement a variety of control measures to reduce disease spread.

  16. Living with avian FLU--Persistence of the H5N1 highly pathogenic avian influenza virus in Egypt.

    PubMed

    Njabo, Kevin Yana; Zanontian, Linda; Sheta, Basma N; Samy, Ahmed; Galal, Shereen; Schoenberg, Frederic Paik; Smith, Thomas B

    2016-05-01

    H5N1 highly pathogenic avian influenza virus (HPAIV) continues to cause mortality in poultry and threaten human health at a panzootic scale in Egypt since it was reported in 2006. While the early focus has been in Asia, recent evidence suggests that Egypt is an emerging epicenter for the disease. Despite control measures, epizootic transmission of the disease continues. Here, we investigate the persistence of HPAIV across wild passerine birds and domestic poultry between 2009 and 2012 and the potential risk for continuous viral transmission in Egypt. We use a new weighted cross J-function to investigate the degree and spatial temporal nature of the clustering between sightings of infected birds of different types, and the risk of infection associated with direct contact with infected birds. While we found no infection in wild birds, outbreaks occurred year round between 2009 and 2012, with a positive interaction between chickens and ducks. The disease was more present in the years 2010 and 2011 coinciding with the political unrest in the country. Egypt thus continues to experience endemic outbreaks of avian influenza HPAIV in poultry and an increased potential risk of infection to other species including humans. With the current trends, the elimination of the HPAIV infection is highly unlikely without a complete revamp of current policies. The application of spatial statistics techniques to these types of data may help us to understand the characteristics of the disease and may subsequently allow practitioners to explore possible preventive solutions.

  17. The spread of highly pathogenic avian influenza (subtype H5N1) clades in Bangladesh, 2010 and 2011.

    PubMed

    Osmani, Muzaffar G; Ward, Michael P; Giasuddin, Md; Islam, Md Rafiqul; Kalam, Abul

    2014-04-01

    Since the global spread of highly pathogenic avian influenza H5N1 during 2005-2006, control programs have been successfully implemented in most affected countries. HPAI H5N1 was first reported in Bangladesh in 2007, and since then 546 outbreaks have been reported to the OIE. The disease has apparently become endemic in Bangladesh. Spatio-temporal information on 177 outbreaks of HPAI H5N1 occurring between February 2010 and April 2011 in Bangladesh, and 37 of these outbreaks in which isolated H5N1 viruses were phylogenetically characterized to clade, were analyzed. Three clades were identified, 2.2 (21 cases), 2.3.4 (2 cases) and 2.3.2.1 (14 cases). Clade 2.2 was identified throughout the time period and was widely distributed in a southeast-northwest orientation. Clade 2.3.2.1 appeared later and was generally confined to central Bangladesh in a north-south orientation. Based on a direction test, clade 2.2 viruses spread in a southeast-to-northwest direction, whereas clade 2.3.2.1 spread west-to-east. The magnitude of spread of clade 2.3.2.1 was greater relative to clade 2.2 (angular concentration 0.2765 versus 0.1860). In both cases, the first outbreak(s) were identified as early outliers, but in addition, early outbreaks (one each) of clade 2.2 were also identified in central Bangladesh and in northwest Bangladesh, a considerable distance apart. The spread of highly pathogenic avian influenza H5N1 in Bangladesh is characterized by reported long-distance translocation events. This poses a challenge to disease control efforts. Increased enforcement of biosecurity and stronger control of movements between affected farms and susceptible farms, and better surveillance and reporting, is needed. Although the movement of poultry and equipment appears to be a more likely explanation for the patterns identified, the relative contribution of trade and the market chain versus wild birds in spreading the disease needs further investigation.

  18. Intranasal H5N1 vaccines, adjuvanted with chitosan derivatives, protect ferrets against highly pathogenic influenza intranasal and intratracheal challenge.

    PubMed

    Mann, Alex J; Noulin, Nicolas; Catchpole, Andrew; Stittelaar, Koert J; de Waal, Leon; Veldhuis Kroeze, Edwin J B; Hinchcliffe, Michael; Smith, Alan; Montomoli, Emanuele; Piccirella, Simona; Osterhaus, Albert D M E; Knight, Alastair; Oxford, John S; Lapini, Giulia; Cox, Rebecca; Lambkin-Williams, Rob

    2014-01-01

    We investigated the protective efficacy of two intranasal chitosan (CSN and TM-CSN) adjuvanted H5N1 Influenza vaccines against highly pathogenic avian Influenza (HPAI) intratracheal and intranasal challenge in a ferret model. Six groups of 6 ferrets were intranasally vaccinated twice, 21 days apart, with either placebo, antigen alone, CSN adjuvanted antigen, or TM-CSN adjuvanted antigen. Homologous and intra-subtypic antibody cross-reacting responses were assessed. Ferrets were inoculated intratracheally (all treatments) or intranasally (CSN adjuvanted and placebo treatments only) with clade 1 HPAI A/Vietnam/1194/2004 (H5N1) virus 28 days after the second vaccination and subsequently monitored for morbidity and mortality outcomes. Clinical signs were assessed and nasal as well as throat swabs were taken daily for virology. Samples of lung tissue, nasal turbinates, brain, and olfactory bulb were analysed for the presence of virus and examined for histolopathological findings. In contrast to animals vaccinated with antigen alone, the CSN and TM-CSN adjuvanted vaccines induced high levels of antibodies, protected ferrets from death, reduced viral replication and abrogated disease after intratracheal challenge, and in the case of CSN after intranasal challenge. In particular, the TM-CSN adjuvanted vaccine was highly effective at eliciting protective immunity from intratracheal challenge; serologically, protective titres were demonstrable after one vaccination. The 2-dose schedule with TM-CSN vaccine also induced cross-reactive antibodies to clade 2.1 and 2.2 H5N1 viruses. Furthermore ferrets immunised with TM-CSN had no detectable virus in the respiratory tract or brain, whereas there were signs of virus in the throat and lungs, albeit at significantly reduced levels, in CSN vaccinated animals. This study demonstrated for the first time that CSN and in particular TM-CSN adjuvanted intranasal vaccines have the potential to protect against significant mortality and

  19. Toxoplasma gondii Seropositivity and Co-Infection with TORCH Pathogens in High-Risk Patients from Qatar

    PubMed Central

    Abu-Madi, Marawan A.; Behnke, Jerzy M.; Dabritz, Haydee A.

    2010-01-01

    Testing of patients who are deemed to be at high risk for TORCH pathogens, e.g., pregnant women, their fetuses, neonates, and acquired immunodeficiency syndrome (AIDS) patients, is important so that specific treatment can be initiated. This study included 1,857 such patients between 2005 and 2008. Logistic regression was used to evaluate factors associated with Toxoplasma gondii seropositivity. Among 823 women of childbearing age, 35.1% and 5.2% tested positive for T. gondii IgG and IgM, respectively. Three infants ≤ 6 months of age (0.8% of 353) were congenitally infected. Factors associated with T. gondii IgG seropositivity included older age, East Mediterranean or African nationality, positive cytomegalovirus (CMV) and herpes simplex virus (HSV)-1 serostatus, and negative rubella IgG results. The decreasing prevalence of IgM antibodies between 2005 and 2008 suggested that exposure to T. gondii from food or environmental sources declined over this period in Qatar. Population-based studies of newborns would be helpful to accurately estimate incidence of congenital toxoplasmosis. PMID:20348511

  20. The vOTU domain of highly-pathogenic porcine reproductive and respiratory syndrome virus displays a differential substrate preference

    PubMed Central

    Deaton, Michelle K.; Spear, Allyn; Faaberg, Kay S.; Pegan, Scott D.

    2014-01-01

    Arterivirus genus member Porcine reproductive and respiratory syndrome virus (PRRSV) causes an economically devastating disease, recently exacerbated by the emergence of highly pathogenic strains (HP-PRRSV). Within the nonstructural protein 2 of PRRSV is a deubiquitinating enzyme domain belonging to the viral ovarian tumor (vOTU) protease superfamily. vOTUs, which can greatly vary in their preference for their host ubiquitin (Ub) and Ub-like substrates such as interferon stimulated gene 15 (ISG15), have been implicated as a potential virulence factor. Since various strains of PRRSV have large variations in virulence, the specificity of vOTUs from two PRRSV strains of varying virulence were determined. While both vOTUs showed de-ubiquitinating activity and markedly low deISGylating activity, HP-PRRSV demonstrated a strong preference for lysine 63-linked poly-Ubiquitin, tied to innate immune response regulation. This represents the first report of biochemical activity unique to HP-PRRSV that has implications for a potential increase in immunosuppression and virulence. PMID:24725951

  1. Comprehensive analysis of antibody recognition in convalescent humans from highly pathogenic avian influenza H5N1 infection

    PubMed Central

    Zuo, Teng; Sun, Jianfeng; Wang, Guiqin; Jiang, Liwei; Zuo, Yanan; Li, Danyang; Shi, Xuanling; Liu, Xi; Fan, Shilong; Ren, Huanhuan; Hu, Hongxing; Sun, Lina; Zhou, Boping; Liang, Mifang; Zhou, Paul; Wang, Xinquan; Zhang, Linqi

    2015-01-01

    Understanding the mechanism of protective antibody recognition against highly pathogenic avian influenza A virus H5N1 in humans is critical for the development of effective therapies and vaccines. Here we report the crystal structure of three H5-specific human monoclonal antibodies bound to the globular head of hemagglutinin (HA) with distinct epitope specificities, neutralization potencies and breadth. A structural and functional analysis of these epitopes combined with those reported elsewhere identifies four major vulnerable sites on the globular head of H5N1 HA. Chimeric and vulnerable site-specific mutant pseudoviruses are generated to delineate broad neutralization specificities of convalescent sera from two individuals who recovered from the infection with H5N1 virus. Our results show that the four vulnerable sites on the globular head rather than the stem region are the major neutralizing targets, suggesting that during natural H5N1 infection neutralizing antibodies against the globular head work in concert to provide protective antibody-mediated immunity. PMID:26635249

  2. Contact between bird species of different lifespans can promote the emergence of highly pathogenic avian influenza strains

    PubMed Central

    Wikramaratna, Paul S.; Pybus, Oliver G.; Gupta, Sunetra

    2014-01-01

    Outbreaks of highly pathogenic strains of avian influenza viruses (AIVs) cause considerable economic losses to the poultry industry and also pose a threat to human life. The possibility that one of these strains will evolve to become transmissible between humans, sparking a major influenza pandemic, is a matter of great concern. Most studies so far have focused on assessing these odds from the perspective of the intrinsic mutability of AIV rather than the ecological constraints to invasion faced by the virus population. Here we present an alternative multihost model for the evolution of AIV in which the mode and tempo of mutation play a limited role, with the emergence of strains being determined instead principally by the prevailing profile of population-level immunity. We show that (i) many of the observed differences in influenza virus dynamics among species can be captured by our model by simply varying host lifespan and (ii) increased contact between species of different lifespans can promote the emergence of potentially more virulent strains that were hitherto suppressed in one of the species. PMID:24958867

  3. Spatio-temporal epidemiology of highly pathogenic avian influenza outbreaks in the two deltas of Vietnam during 2003-2007.

    PubMed

    Minh, Phan Q; Morris, Roger S; Schauer, Birgit; Stevenson, Mark; Benschop, Jackie; Nam, Hoang V; Jackson, Ron

    2009-05-01

    Outbreaks of highly pathogenic avian influenza A subtype H5N1 have occurred in Vietnam as a series of epidemic waves since December 2003. We describe the spatial and temporal patterns of the HPAI H5N1 epidemics in the Red River Delta in the north (785 outbreaks in 606 communes) and the Mekong River Delta in the south of Vietnam (1313 outbreaks in 837 communes), where the epidemics were concentrated. Throughout the study period the percentage of outbreaks affecting ducks increased steadily to a peak of 78% during the 2006/2007 epidemic in both deltas. Five of the seven epidemic waves occurred in the period of active poultry population buildup immediately prior to the Vietnamese New Year (Tet festival). Recorded outbreaks were clustered in space and time within both deltas, consistent with infection transmission occurring via a combination of local and long-distance spread. Our analyses demonstrate that the epidemiology of HPAI in Vietnam has changed over the 4-year study period, with outbreaks now occurring in the warmer months of the year and ducks featuring more prominently as affected species. To determine the relative importance of local and long-distance spread on infection transmission, precise details of outbreak location, date of onset of clinical signs, and size and composition of the poultry population at risk need to be recorded during future outbreak responses.

  4. Comprehensive analysis of antibody recognition in convalescent humans from highly pathogenic avian influenza H5N1 infection.

    PubMed

    Zuo, Teng; Sun, Jianfeng; Wang, Guiqin; Jiang, Liwei; Zuo, Yanan; Li, Danyang; Shi, Xuanling; Liu, Xi; Fan, Shilong; Ren, Huanhuan; Hu, Hongxing; Sun, Lina; Zhou, Boping; Liang, Mifang; Zhou, Paul; Wang, Xinquan; Zhang, Linqi

    2015-01-01

    Understanding the mechanism of protective antibody recognition against highly pathogenic avian influenza A virus H5N1 in humans is critical for the development of effective therapies and vaccines. Here we report the crystal structure of three H5-specific human monoclonal antibodies bound to the globular head of hemagglutinin (HA) with distinct epitope specificities, neutralization potencies and breadth. A structural and functional analysis of these epitopes combined with those reported elsewhere identifies four major vulnerable sites on the globular head of H5N1 HA. Chimeric and vulnerable site-specific mutant pseudoviruses are generated to delineate broad neutralization specificities of convalescent sera from two individuals who recovered from the infection with H5N1 virus. Our results show that the four vulnerable sites on the globular head rather than the stem region are the major neutralizing targets, suggesting that during natural H5N1 infection neutralizing antibodies against the globular head work in concert to provide protective antibody-mediated immunity.

  5. High diversity in populations of the introduced plant pathogen, Cryphonectria parasitica, due to encounters between genetically divergent genotypes.

    PubMed

    Ježić, Marin; Krstin, Ljiljana; Rigling, Daniel; Curković-Perica, Mirna

    2012-01-01

    The ascomycete fungus Cryphonectria parasitica is an aggressive introduced pathogen of sweet chestnut (Castanea sativa Mill.). It has spread throughout the chestnut-growing areas of Europe, with higher diversity in the regions close to its first introduction and lower diversity in its expanding ranges in Europe. To reconstruct the invasion events that could explain the high diversity of C. parasitica in Croatia and Slovenia, 180 samples were genotyped using 11 sequence-characterized amplified region markers. Eight of 11 loci were found to be polymorphic, and a total of 66 different haplotypes were identified. Bayesian clustering indicated the existence of two clusters, which suggests two separate introductions of C. parasitica in these regions. The first cluster is dominant in western parts of Croatia and Slovenia and the second in eastern and northern regions. The data analysis indicates that northern Italy was the first source of infection, with the subsequent introduction from south-eastern Europe, which contributed significantly to the diversity of the C. parasitica populations tested. Most haplotypes were probably derived through sexual recombination between a few divergent haplotypes, which suggests that multiple introductions and sexual reproduction are important for the formation of genetically diverse C. parasitica populations.

  6. Identifying antigenicity associated sites in highly pathogenic H5N1 influenza virus hemagglutinin by using sparse learning

    PubMed Central

    Cai, Zhipeng; Ducatez, Mariette F.; Yang, Jialiang; Zhang, Tong; Long, Li-Ping; Boon, Adrianus C.; Webby, Richard J.; Wan, Xiu-Feng

    2012-01-01

    Since the isolation of A/goose/Guangdong/1/1996 (H5N1) in farmed geese in southern China, highly pathogenic H5N1 avian influenza viruses have posed a continuous threat to both public and animal health. The non-synonymous mutation of the H5 hemagglutinin gene has resulted in antigenic drift, leading to difficulties in both clinical diagnosis and vaccine strain selection. Characterizing H5N1’s antigenic profiles would help resolve these problems. In this study, a novel sparse learning method was developed to identify antigenicity associated sites in influenza A viruses on the basis of immunologic datasets (i.e., from hemagglutination inhibition and microneutralization assays) and HA protein sequences. Twenty-one potential antigenicity associated sites were identified. A total of seventeen H5N1 mutants were used to validate the effects of eleven of these predicted sites on H5N1’s antigenicity, including seven newly identified sites not located in reported antibody binding sites. The experimental data confirmed that mutations of these tested sites lead to changes in viral antigenicity, validating our method. PMID:22609437

  7. Vaccination of gallinaceous poultry for H5N1 highly pathogenic avian influenza: current questions and new technology.

    PubMed

    Spackman, Erica; Swayne, David E

    2013-12-01

    Vaccination of poultry for avian influenza virus (AIV) is a complex topic as there are numerous technical, logistic and regulatory aspects which must be considered. Historically, control of high pathogenicity (HP) AIV infection in poultry has been accomplished by eradication and stamping out when outbreaks occur locally. Since the H5N1 HPAIV from Asia has spread and become enzootic, vaccination has been used on a long-term basis by some countries to control the virus, other countries have used it temporarily to aid eradication efforts, while others have not used it at all. Currently, H5N1 HPAIV is considered enzootic in China, Egypt, Viet Nam, India, Bangladesh and Indonesia. All but Bangladesh and India have instituted vaccination programs for poultry. Importantly, the specifics of these programs differ to accommodate different situations, resources, and industry structure in each country. The current vaccines most commonly used are inactivated whole virus vaccines, but vectored vaccine use is increasing. Numerous technical improvements to these platforms and novel vaccine platforms for H5N1 vaccines have been reported, but most are not ready to be implemented in the field.

  8. Green synthesis of highly stabilized nanocrystalline silver particles by a non-pathogenic and agriculturally important fungus T. asperellum

    NASA Astrophysics Data System (ADS)

    Mukherjee, P.; Roy, M.; Mandal, B. P.; Dey, G. K.; Mukherjee, P. K.; Ghatak, J.; Tyagi, A. K.; Kale, S. P.

    2008-02-01

    A controlled and up-scalable biosynthetic route to nanocrystalline silver particles with well-defined morphology using cell-free aqueous filtrate of a non-pathogenic and commercially viable biocontrol agent Trichoderma asperellum is being reported for the first time. A transparent solution of the cell-free filtrate of Trichoderma asperellum containing 1 mM AgNO3 turns progressively dark brown within 5 d of incubation at 25 °C. The kinetics of the reaction was studied using UV-vis spectroscopy. An intense surface plasmon resonance band at ~410 nm in the UV-vis spectrum clearly reveals the formation of silver nanoparticles. The size of the silver particles using TEM and XRD studies is found to be in the range 13-18 nm. These nanoparticles are found to be highly stable and even after prolonged storage for over 6 months they do not show significant aggregation. A plausible mechanism behind the formation of silver nanoparticles and their stabilization via capping has been investigated using FTIR and surface-enhanced resonance Raman spectroscopy.

  9. No evidence of infection or exposure to Highly Pathogenic Avian Influenzas in peridomestic wildlife on an affected poultry facility

    USGS Publications Warehouse

    Grear, Daniel; Dusek, Robert J.; Walsh, Daniel P.; Hall, Jeffrey S.

    2016-01-01

    We evaluated the potential transmission of avian influenza viruses (AIV) in wildlife species in three settings in association with an outbreak at a poultry facility: 1) small birds and small mammals on a poultry facility that was affected with highly pathogenic AIV (HPAIV) in April 2015; 2) small birds and small mammals on a nearby poultry facility that was unaffected by HPAIV; and 3) small birds, small mammals, and waterfowl in a nearby natural area. We live-captured small birds and small mammals and collected samples from hunter-harvested waterfowl to test for active viral shedding and evidence of exposure (serum antibody) to AIV and the H5N2 HPAIV that affected the poultry facility. We detected no evidence of shedding or specific antibody to AIV in small mammals and small birds 5 mo after depopulation of the poultry. We detected viral shedding and exposure to AIV in waterfowl and estimated approximately 15% viral shedding and 60% antibody prevalence. In waterfowl, we did not detect shedding or exposure to the HPAIV that affected the poultry facility. We also conducted camera trapping around poultry carcass depopulation composting barns and found regular visitation by four species of medium-sized mammals. We provide preliminary data suggesting that peridomestic wildlife were not an important factor in the transmission of AIV during the poultry outbreak, nor did small birds and mammals in natural wetland settings show wide evidence of AIV shedding or exposure, despite the opportunity for exposure.

  10. Comprehensive analysis of antibody recognition in convalescent humans from highly pathogenic avian influenza H5N1 infection.

    PubMed

    Zuo, Teng; Sun, Jianfeng; Wang, Guiqin; Jiang, Liwei; Zuo, Yanan; Li, Danyang; Shi, Xuanling; Liu, Xi; Fan, Shilong; Ren, Huanhuan; Hu, Hongxing; Sun, Lina; Zhou, Boping; Liang, Mifang; Zhou, Paul; Wang, Xinquan; Zhang, Linqi

    2015-01-01

    Understanding the mechanism of protective antibody recognition against highly pathogenic avian influenza A virus H5N1 in humans is critical for the development of effective therapies and vaccines. Here we report the crystal structure of three H5-specific human monoclonal antibodies bound to the globular head of hemagglutinin (HA) with distinct epitope specificities, neutralization potencies and breadth. A structural and functional analysis of these epitopes combined with those reported elsewhere identifies four major vulnerable sites on the globular head of H5N1 HA. Chimeric and vulnerable site-specific mutant pseudoviruses are generated to delineate broad neutralization specificities of convalescent sera from two individuals who recovered from the infection with H5N1 virus. Our results show that the four vulnerable sites on the globular head rather than the stem region are the major neutralizing targets, suggesting that during natural H5N1 infection neutralizing antibodies against the globular head work in concert to provide protective antibody-mediated immunity. PMID:26635249

  11. Human infection with a highly pathogenic avian influenza A (H5N6) virus in Yunnan province, China.

    PubMed

    Xu, Wen; Li, Hong; Jiang, Li

    2016-01-01

    Highly pathogenic avian influenza A H5N6 virus has caused four human infections in China. This study reports the preliminary findings of the first known human case of H5N6 in Yunnan province. The patient initially developed symptoms of sore throat and coughing on 27 January 2015. The disease rapidly progressed to severe pneumonia, multiple organ dysfunctions and acute respiratory distress syndrome and the patient died on 6 February. Virological analysis determined that the virus belonged to H5 clade 2.3.4.4 and it has obtained partial ability for mammalian adaptation and amantadine resistance. Environmental investigation found H5 in 63% of the samples including poultry faeces, tissues, cage surface swabs and sewage from local live poultry markets by real-time RT-PCR. These findings suggest that the expanding and enhancing of surveillance in both avian and humans are necessary to monitor the evolution of H5 influenza virus and to facilitate early detection of suspected cases. PMID:27030920

  12. Disease awareness of the poultry keepers in Switzerland and their access to information concerning highly pathogenic avian influenza.

    PubMed

    Saurina, J; Fiebig, L; Zinsstag, J; Schelling, E

    2010-08-01

    The passive surveillance of highly pathogenic avian influenza (HPAI) in domestic poultry is based essentially on the reporting of suspicious clinical cases by the poultry keepers to the veterinary services. As little was known about HPAI disease awareness among Swiss poultry keepers, a cross-sectional study was conducted among poultry keepers in Switzerland in 2007. For data triangulation and complementary information, interviews have been conducted with experts of poultry marketing organizations. The main information source used by the poultry keepers was mass media. Having a non-commercial poultry husbandry was significantly associated with lower knowledge scores. Non-commercial poultry keepers felt neglected by the veterinary authorities. Risks perceived by the poultry keepers reflected well the officially communicated risks for HPAI introduction. By highlighting the needs and the knowledge level of the poultry keepers, we make recommendations with regard to more efficient information exchange between poultry keepers and veterinary authorities. The main challenge will be to consistently integrate non-commercial poultry keepers in the formal information channels. PMID:20683825

  13. Antigenic analysis of highly pathogenic avian influenza virus H5N1 sublineages co-circulating in Egypt.

    PubMed

    Watanabe, Yohei; Ibrahim, Madiha S; Ellakany, Hany F; Kawashita, Norihito; Daidoji, Tomo; Takagi, Tatsuya; Yasunaga, Teruo; Nakaya, Takaaki; Ikuta, Kazuyoshi

    2012-10-01

    Highly pathogenic avian influenza virus H5N1 has spread across Eurasia and Africa, and outbreaks are now endemic in several countries, including Indonesia, Vietnam and Egypt. Continuous circulation of H5N1 virus in Egypt, from a single infected source, has led to significant genetic diversification with phylogenetically separable sublineages, providing an opportunity to study the impact of genetic evolution on viral phenotypic variation. In this study, we analysed the phylogeny of H5 haemagglutinin (HA) genes in influenza viruses isolated in Egypt from 2006 to 2011 and investigated the effect of conserved amino acid mutations in the HA genes in each of the sublineages on their antigenicity. The analysis showed that viruses in at least four sublineages still persisted in poultry in Egypt as of 2011. Using reverse genetics to generate HA-reassortment viruses with specific HA mutations, we found antigenic drift in the HA in two influenza virus sublineages, compared with the other currently co-circulating influenza virus sublineages in Egypt. Moreover, the two sublineages with significant antigenic drift were antigenically distinguishable. Our findings suggested that phylogenetically divergent H5N1 viruses, which were not antigenically cross-reactive, were co-circulating in Egypt, indicating that there was a problem in using a single influenza virus strain as seed virus to produce influenza virus vaccine in Egypt and providing data for designing more efficacious control strategies in H5N1-endemic areas.

  14. Ostrich ( Struthio camelus ) Infected with H5N8 Highly Pathogenic Avian Influenza Virus in South Korea in 2014.

    PubMed

    Kim, Hye-Ryoung; Kwon, Yong-Kuk; Lee, Youn-Jeong; Kang, Hyun-Mi; Lee, Eun-Kyoung; Song, Byung-Min; Jung, Suk-Chan; Lee, Kyung-Hyun; Lee, Hyun-Kyoung; Baek, Kang-Hyun; Bae, You-Chan

    2016-06-01

    Highly pathogenic avian influenza (HPAI) virus of the H5N8 subtype was isolated from a young ostrich in South Korea in March 2014. Clinical signs characterized by anorexia, depression, and signs of nervousness were observed. The isolated A/ostrich/Korea/H829/2014 (H5N8) virus had a cleavage site motif containing multiple basic amino acids, typical of HPAI virus. The phylogenetic tree of the hemagglutinin gene of the H5 HPAI virus showed that this ostrich H5N8 virus belongs to clade 2.3.4.4 viruses together with H5N8 strains isolated from ducks and wild birds in South Korea in 2014. Pathologically, redness of pancreas, enlargement and hemorrhage of spleen, friability of brain, and hydropericardium were prominently found. Histologic legions were observed in pancreas, spleen, liver, lung, heart, and brain, and influenza A nucleoproteins were detected in the same organs by immunohistochemistry. Other ostriches farmed together in open camps were not infected with HPAI virus based on the serologic and virologic tests. The findings indicate that ostriches are susceptible to H5N8 HPAI virus, but this virus does not spread efficiently among ratites. PMID:27309301

  15. Description of an outbreak of highly pathogenic avian influenza in domestic ostriches (Struthio camelus) in South Africa in 2011.

    PubMed

    van Helden, L S; Sinclair, M; Koen, P; Grewar, J D

    2016-06-01

    In 2011, the commercial ostrich production industry of South Africa experienced an outbreak of highly pathogenic avian influenza (HPAI), subtype H5N2. Surveillance using antibody and antigen detection revealed 42 infected farms with a between-farm prevalence in the affected area of 16%. The outbreak was controlled using depopulation of infected farms, resulting in the direct loss of 10% of the country's domestic ostrich population. Various factors in the ostrich production system were observed that could have contributed to the spread of the virus between farms, including the large number of legal movements of ostriches between farms, access of wild birds to ostrich camps and delays in depopulation of infected farms. Negative effects on the ostrich industry and the local economy of the ostrich-producing area were observed as a result of the outbreak and the disease control measures applied. Prevention and control measures applied as a result of avian influenza in South Africa were informed by this large outbreak and the insights into epidemiology of avian influenza in ostriches that it provided, resulting in stricter biosecurity measures required on every registered ostrich farm in the country. PMID:27237385

  16. Transmission dynamics of highly pathogenic avian influenza at Lake Constance (Europe) during the outbreak of winter 2005-2006.

    PubMed

    Penny, M A; Saurina, J; Keller, I; Jenni, L; Bauer, H-G; Fiedler, W; Zinsstag, J

    2010-09-01

    Highly pathogenic avian influenza virus (HPAI) H5N1 poses a serious threat to domestic animals. Despite the large number of studies on influenza A virus in waterbirds, little is still known about the transmission dynamics, including prevalence, behavior, and spread of these viruses in the wild waterbird population. From January to April 2006, the HPAI H5N1 virus was confirmed in 82 dead wild waterbirds at the shores of Lake Constance. In this study, we present simple mathematical models to examine this outbreak and to investigate the transmission dynamics of HPAI in wild waterbirds. The population dynamics model of wintering birds was best represented by a sinusoidal function. This model was considered the most adequate to represent the susceptible compartment of the SIR model. The three transmission models predict a basic reproduction ratio (R (0)) with value of approximately 1.6, indicating a small epidemic, which ended with the migration of susceptible wild waterbirds at the end of the winter. With this study, we quantify for the first time the transmission of HPAI H5N1 virus at Lake Constance during the outbreak of winter 2005-2006. It is a step toward the improvement of the knowledge of transmission of the virus among wild waterbirds. PMID:20680395

  17. Characterizing wild bird contact and seropositivity to highly pathogenic avian influenza A (H5N1) virus in Alaskan residents

    PubMed Central

    Reed, Carrie; Bruden, Dana; Byrd, Kathy K; Veguilla, Vic; Bruce, Michael; Hurlburt, Debby; Wang, David; Holiday, Crystal; Hancock, Kathy; Ortiz, Justin R; Klejka, Joe; Katz, Jacqueline M; Uyeki, Timothy M

    2014-01-01

    Background Highly pathogenic avian influenza A (HPAI) H5N1 viruses have infected poultry and wild birds on three continents with more than 600 reported human cases (59% mortality) since 2003. Wild aquatic birds are the natural reservoir for avian influenza A viruses, and migratory birds have been documented with HPAI H5N1 virus infection. Since 2005, clade 2.2 HPAI H5N1 viruses have spread from Asia to many countries. Objectives We conducted a cross-sectional seroepidemiological survey in Anchorage and western Alaska to identify possible behaviors associated with migratory bird exposure and measure seropositivity to HPAI H5N1. Methods We enrolled rural subsistence bird hunters and their families, urban sport hunters, wildlife biologists, and a comparison group without bird contact. We interviewed participants regarding their exposures to wild birds and collected blood to perform serologic testing for antibodies against a clade 2.2 HPAI H5N1 virus strain. Results Hunters and wildlife biologists reported exposures to wild migratory birds that may confer risk of infection with avian influenza A viruses, although none of the 916 participants had evidence of seropositivity to HPAI H5N1. Conclusions We characterized wild bird contact among Alaskans and behaviors that may influence risk of infection with avian influenza A viruses. Such knowledge can inform surveillance and risk communication surrounding HPAI H5N1 and other influenza viruses in a population with exposure to wild birds at a crossroads of intercontinental migratory flyways. PMID:24828535

  18. Characterization of the interactions between human high-molecular-mass kininogen and cell wall proteins of pathogenic yeasts Candida tropicalis.

    PubMed

    Karkowska-Kuleta, Justyna; Zajac, Dorota; Bras, Grazyna; Bochenska, Oliwia; Seweryn, Karolina; Kedracka-Krok, Sylwia; Jankowska, Urszula; Rapala-Kozik, Maria; Kozik, Andrzej

    2016-01-01

    Candida tropicalis is one of the most frequent causes of serious disseminated candidiasis in human patients infected by non-albicans Candida species, but still relatively little is known about its virulence mechanisms. In our current study, the interactions between the cell surface of this species and a multifunctional human protein - high-molecular-mass kininogen (HK), an important component of the plasma contact system involved in the development of the inflammatory state - were characterized at the molecular level. The quick release of biologically active kinins from candidal cell wall-adsorbed HK was presented and the HK-binding ability was assigned to several cell wall-associated proteins. The predicted hyphally regulated cell wall protein (Hyr) and some housekeeping enzymes exposed at the cell surface (known as "moonlighting proteins") were found to be the major HK binders. Accordingly, after purification of selected proteins, the dissociation constants of the complexes of HK with Hyr, enolase, and phosphoglycerate mutase were determined using surface plasmon resonance measurements, yielding the values of 2.20 × 10(-7) M, 1.42 × 10(-7) M, and 5.81 × 10(-7) M, respectively. Therefore, in this work, for the first time, the interactions between C. tropicalis cell wall proteins and HK were characterized in molecular terms. Our findings may be useful for designing more effective prevention and treatment approaches against infections caused by this dangerous fungal pathogen. PMID:27474405

  19. Elaboration of highly hydrophobic polymeric surface--a potential strategy to reduce the adhesion of pathogenic bacteria?

    PubMed

    Poncin-Epaillard, F; Herry, J M; Marmey, P; Legeay, G; Debarnot, D; Bellon-Fontaine, M N

    2013-04-01

    Different polymeric surfaces have been modified in order to reach a high hydrophobic character, indeed the superhydrophobicity property. For this purpose, polypropylene and polystyrene have been treated by RF or μwaves CF4 plasma with different volumes, the results were compared according to the density of injected power. The effect of pretreatment such as mechanical abrasion or plasma activation was also studied. The modified surfaces were shown as hydrophobic, or even superhydrophobic depending of defects density. They were characterized by measurement of wettability and roughness at different scales, i.e. macroscopic, mesoscopic and atomic. It has been shown that a homogeneous surface at the macroscopic scale could be heterogeneous at lower mesoscopic scale. This was associated with the crystallinity of the material. The bioadhesion tests were performed with Gram positive and negative pathogenic strains: Listeria monocytogenes, Pseudomonas aeruginosa and Hafnia alvei. They have demonstrated an antibacterial efficiency of very hydrophobic and amorphous PS treated for all strains tested and a strain-dependent efficiency with modified PP surface being very heterogeneous at the mesoscopic scale. Thus, these biological results pointed out not only the respective role of the surface chemistry and topography in bacterial adhesion, but also the dependence on the peaks and valley distribution at bacteria dimension scale. PMID:23827554

  20. Cloning and Expression of Highly Pathogenic Avian Influenza Virus Full-Length Nonstructural Gene in Pichia pastoris

    PubMed Central

    Abubakar, M. B.; Aini, I.; Omar, A. R.; Hair-Bejo, M.

    2011-01-01

    Avian influenza (AI) is a highly contagious and rapidly evolving pathogen of major concern to the poultry industry and human health. Rapid and accurate detection of avian influenza virus is a necessary tool for control of outbreaks and surveillance. The AI virus A/Chicken/Malaysia/5858/2004 (H5N1) was used as a template to produce DNA clones of the full-length NS1 genes via reverse transcriptase synthesis of cDNA by PCR amplification of the NS1 region. Products were cloned into pCR2.0 TOPO TA plasmid and subsequently subcloned into pPICZαA vector to construct a recombinant plasmid. Recombinant plasmid designated as pPICZαA-NS1 gene was confirmed by PCR colony screening, restriction enzyme digestion, and nucleotide sequence analysis. The recombinant plasmid was transformed into Pichia pastoris GS115 strain by electroporation, and expressed protein was identified by SDS-PAGE and western blotting. A recombinant protein of approximately ~28 kDa was produced. The expressed protein was able to bind a rabbit polyclonal antibody of nonstructural protein (NS1) avian influenza virus H5N1. The result of the western blotting and solid-phase ELISA assay using H5N1 antibody indicated that the recombinant protein produced retained its antigenicity. This further indicates that Pichia pastoris could be an efficient expression system for a avian influenza virus nonstructural (NS1). PMID:21541235

  1. Corneal Opacity in Domestic Ducks Experimentally Infected With H5N1 Highly Pathogenic Avian Influenza Virus.

    PubMed

    Yamamoto, Y; Nakamura, K; Yamada, M; Mase, M

    2016-01-01

    Domestic ducks can be a key factor in the regional spread of H5N1 highly pathogenic avian influenza (HPAI) virus in Asia. The authors performed experimental infections to examine the relationship between corneal opacity and H5N1 HPAI virus infection in domestic ducks (Anas platyrhyncha var domestica). A total of 99 domestic ducks, including 3 control birds, were used in the study. In experiment 1, when domestic ducks were inoculated intranasally with 2 H5N1 HPAI viruses, corneal opacity appeared more frequently than neurologic signs and mortality. Corneal ulceration and exophthalmos were rare findings. Histopathologic examinations of the eyes of domestic ducks in experiment 2 revealed that corneal opacity was due to the loss of corneal endothelial cells and subsequent keratitis with edema. Influenza viral antigen was detected in corneal endothelial cells and some other ocular cells by immunohistochemistry. Results suggest that corneal opacity is a characteristic and frequent finding in domestic ducks infected with the H5N1 HPAI virus. Confirming this ocular change may improve the detection rate of infected domestic ducks in the field.

  2. Bronchointerstitial pneumonia in guinea pigs following inoculation with H5N1 high pathogenicity avian influenza virus.

    PubMed

    Kwon, Y K; Lipatov, A S; Swayne, D E

    2009-01-01

    The H5N1 high-pathogenicity avian influenza (HPAI) viruses have caused widespread disease of poultry in Asia, Africa and the Middle East, and sporadic human infections. The guinea pig model has been used to study human H3N2 and H1N1 influenza viruses, but knowledge is lacking on H5N1 HPAI virus infections. Guinea pigs were inoculated intranasally or intragastrically with A/Vietnam/1203/04 (VN/04) or A/Muscovy duck/Vietnam/209/05 (MDk/VN/05) viruses. Mild listlessness was seen at 2 and 3 days postinoculation (DPI) in guinea pigs inoculated intranasally with VN/04 virus. At 5 DPI, the guinea pigs had bronchointerstitial pneumonia and virus was identified in bronchiolar epithelium and alveolar macrophages. Virus was isolated from the lungs but was lacking from other organs. Minimal lung lesions were seen in intranasal MDk/VN/06 group and virus was not detected, but serologic evidence of infection was observed. Intragastric exposure failed to produce infection or lesions with either virus. The localized respiratory disease in guinea pigs with H5N1 viruses was very similar to that of H3N2 and H1N1 influenza in humans and was less severe than reported for H5N1 human cases.

  3. Synergistic effects of sequential infection with highly pathogenic porcine reproductive and respiratory syndrome virus and porcine circovirus type 2

    PubMed Central

    2013-01-01

    Background Porcine reproductive and respiratory syndrome virus (PRRSV) is the causative agent of porcine reproductive and respiratory syndrome (PRRS) and porcine circovirus type 2 (PCV2) is associated with postweaning multisystemic wasting syndrome (PMWS) in pigs. Coinfection with highly pathogenic PRRSV (HP-PRRSV) and PCV2 in the field has recently become extensive in some Asian countries. A synergistic pathogenicity between PRRSV and PCV2 infections has previously been reported. However, the consequences of the sequential infection of pigs with these two viruses are unknown. Methods Thirty 35-day-old piglets were randomly divided into six groups (n = 5 each): HP-PRRSV/PCV2 (group 1, inoculated with HP-PRRSV, then inoculated with PCV2 one week later), PCV2/HP-PRRSV (group 2, inoculated with PCV2, then inoculated with HP-PRRSV one week later), HP-PRRSV+PCV2 (group 3, inoculated with HP-PRRSV and PCV2 concurrently), HP-PRRSV (group 4, inoculated with HP-PRRSV), PCV2 (group 5, inoculated with PCV2), and the control (group 6, uninfected). This experiment lasted 28 days. Clinical symptoms and rectal temperatures were recorded each day after inoculation, body weight was recorded weekly, and serum samples were obtained for viral nucleic acid quantification and antibody titration. Variations in CD3+, CD4+ CD8–, CD3+, CD4–, and CD8+ cells, natural killer (NK) cells, and mononuclear cells were determined by flow cytometry. The serum concentrations of interferon γ (IFN-γ), tumor necrosis factor α (TNF-α), interleukin 10 (IL-10), and macrophage granulocyte-colony stimulating factor (GM-CSF) were determined. Pathological changes in different tissues from the experimentally infected pigs were recorded. Results The piglets in group 1 had the highest viral loads, the lowest antibody titers, the most-severe clinical signs, and the highest mortality (3/5, 60%; the mortality in the other groups was 0%), and interstitial pneumonia was more severe in this group compare to the

  4. Effect of Infection with a Mesogenic Strain of Newcastle Disease Virus on Infection with Highly Pathogenic Avian Influenza Virus in Chickens

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Little is known on the interactions between avian influenza virus (AIV) and Newcastle disease virus (NDV) when coinfecting the same poultry host. In a previous study we found that infection of chickens with a mesogenic strain of NDV (mNDV) can reduce highly pathogenic AIV (HPAIV) replication, clinic...

  5. Variability in pathobiology of South Korean H5N1 high-pathogenicity avian influenza virus infection for 5 species of migratory waterfowl

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The biological outcome of H5N1 high pathogenicity avian influenza (HPAI) virus infection in wild waterfowl is poorly understood. This study examined infectivity and pathobiology of A/chicken/Korea/IS/06 (H5N1) HPAI virus infection in Mute swans (Cygnus olor), Greylag geese (Anser anser), Ruddy Sheld...

  6. Highly pathogenic avian influenza A(H5N8) viruses reintroduced into South Korea by migratory waterfowl, 2014–2015

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Highly pathogenic avian influenza A(H5N8) viruses were isolated from migratory waterfowl in South Korea during all 2014–winter 2015, a recurrence after initial introduction in winter 2014. These reappeared viruses were phylogenetically distinct from isolates circulating in poultry farms in South Kor...

  7. Assessment of reduced vaccine dose on efficacy of an inactivated avian influenza vaccine against an H5N1 high pathogenicity avian influenza virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Avian influenza (AI) vaccines have emerged to be a viable emergency tool for use in a comprehensive strategy for dealing with high pathogenicity (HP) AI in developed countries. However, the available doses of inactivated AI vaccine are limited to national vaccine banks and inventory stocks of some ...

  8. Experimental infection with low and high pathogenicity H7N3 Chilean avian influenza viruses in Chiloe Wigeon (Anas sibilatrix) and Cinnamon Teal (Anas cyanoptera)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Since 2002, H5N1 high pathogenicity avian influenza (HPAI) viruses have been associated with natural, lethal infections in wild aquatic birds which have been reproduced experimentally. Some aquatic bird species have been suggested as potential transporters of H5N1 HPAI virus via migration. However, ...

  9. A computationally optimized broadly reactive H5 hemagglutinin vaccine provides protection against homologous and heterologous H5N1 highly pathogenic avian influenza virus infection in chickens

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Since its emergence in 1996 in China, H5N1 highly pathogenic avian influenza (HPAI) virus has continuously evolved into different genetic clades that have created challenges to maintaining antigenically relevant H5N1 vaccine seeds. Therefore, a universal (multi-hemagglutinin [HA] subtype) or more c...

  10. Highly Pathogenic Avian Influenza A(H5N8) Viruses Reintroduced into South Korea by Migratory Waterfowl, 2014–2015

    PubMed Central

    Kwon, Jung-Hoon; Lee, Dong-Hun; Swayne, David E.; Noh, Jin-Yong; Yuk, Seong-Su; Erdene-Ochir, Tseren-Ochir; Hong, Woo-Tack; Jeong, Jei-Hyun; Jeong, Sol; Gwon, Gyeong-Bin

    2016-01-01

    Highly pathogenic avian influenza A(H5N8) viruses were isolated from migratory waterfowl in South Korea during fall 2014–winter 2015, a recurrence after initial introduction in winter 2014. These reappeared viruses were phylogenetically distinct from isolates circulating in poultry farms in South Korea. PMID:26890406

  11. Lemna (duckweed) expressed hemagglutinin from avian influenza H5N1 protects chickens against H5N1 high pathogenicity avian influenza virus challenge

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In the last two decades, transgenic plants have been explored as safe and cost effective alternative expression platforms for producing recombinant proteins. In this study, a synthetic hemagglutinin (HA) gene from the high pathogenicity avian influenza (HPAI) virus A/chicken/Indonesia/7/2003 (H5N1)...

  12. Live Bird Markets of Bangladesh: H9N2 Viruses and the Near Absence of Highly Pathogenic H5N1 Influenza

    PubMed Central

    Negovetich, Nicholas J.; Feeroz, Mohammed M.; Jones-Engel, Lisa; Walker, David; Alam, S. M. Rabiul; Hasan, Kamrul; Seiler, Patrick; Ferguson, Angie; Friedman, Kim; Barman, Subrata; Franks, John; Turner, Jasmine; Krauss, Scott; Webby, Richard J.; Webster, Robert G.

    2011-01-01

    Avian influenza surveillance in Bangladesh has been passive, relying on poultry farmers to report suspected outbreaks of highly pathogenic H5N1 influenza. Here, the results of an active surveillance effort focusing on the live-bird markets are presented. Prevalence of influenza infection in the birds of the live bird markets is 23.0%, which is similar to that in poultry markets in other countries. Nearly all of the isolates (94%) were of the non-pathogenic H9N2 subtype, but viruses of the H1N2, H1N3, H3N6, H4N2, H5N1, and H10N7 subtypes were also observed. The highly pathogenic H5N1-subtype virus was observed at extremely low prevalence in the surveillance samples (0.08%), and we suggest that the current risk of infection for humans in the retail poultry markets in Bangladesh is negligible. However, the high prevalence of the H9 subtype and its potential for interaction with the highly pathogenic H5N1-subtype, i.e., reassortment and attenuation of host morbidity, highlight the importance of active surveillance of the poultry markets. PMID:21541296

  13. Protection against H7N3 high pathogenicity avian influenza in chickens immunized with a recombinant fowlpox and an inactivated avian influenza vaccines

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Beginning on June 2012, an H7N3 highly pathogenic avian influenza (HPAI) epizootic was reported in the State of Jalisco (Mexico), with some 22.4 million chickens that died, were slaughtered on affected farms or were preemptively culled on neighboring farms. In the current study, layer chickens were ...

  14. Reduction of high pathogenicity avian influenza virus in eggs from chickens once or twice vaccinated with an oil-emulsified inactivated H5 avian influenza vaccine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The negative impact of high pathogenicity avian influenza virus (HPAIV) infection on egg production and deposition of virus in eggs, as well as any protective effect of vaccination, is unknown. Individually housed non-vaccinated, sham-vaccinated and inactivated H5N9 vaccinated once or twice adult Wh...

  15. Protection of poultry against the 2012 Mexican H7N3 highly pathogenic avian influenza virus with inactivated H7 avian influenza vaccines

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In June of 2012, an outbreak of highly pathogenic avian influenza (HPAI) H7N3 was reported poultry in Jalisco, Mexico. Since that time the virus has spread to the surrounding States of Guanajuato and Aguascalientes and new outbreaks continue to be reported. To date more than 25 million birds have di...

  16. Negative data provide weak support for disappearance and restricted emergence/persistence of highly pathogenic influenza A viruses in North American waterfowl

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In their recent paper, Krauss et al. use lack of detection of highly pathogenic (HP) H5 clade 2.3.4.4 (henceforth ‘H5’) influenza A viruses (IAVs) from >22,000 wild bird samples collected in North America (NA) in 2014–2015 to argue that HP H5 IAVs disappeared from waterfowl and that unresolved mecha...

  17. Determination of efficacious vaccine seed strains for use against Egyptian H5N1 highly pathogenic avian influenza viruses through antigenic cartography and in vivo challenge studies

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Since 2006, there have been reported outbreaks of H5N1 highly pathogenic avian influenza (HPAI) in vaccinated chickens in Africa and Asia. This study provides experimental data for selection of efficacious H5N1 vaccine seed strains against recently circulating strains of H5N1 HPAI viruses in Egypt....

  18. Changes in adaptation of H5N2 highly pathogenic avian influenza H5 clade 2.3.4.4 viruses in chickens and mallards

    Technology Transfer Automated Retrieval System (TEKTRAN)

    H5N2 highly pathogenic avian influenza (HPAI) viruses caused a severe poultry outbreak in the United States (U.S.) during 2015. In order to examine changes in adaptation of this viral lineage, the infectivity, transmission and pathogenesis of poultry H5N2 viruses was investigated in chickens and mal...

  19. Impact of vaccination on infection with Vietnam H5N1 high pathogenicity avian influenza virus in hens and the eggs they lay

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Highly pathogenic avian influenza virus (HPAIV) infections in chickens produce a negative impact on egg production, and virus is deposited on surface and internal contents of eggs. Previously, vaccination maintained egg production and reduced egg contamination when challenged with a North American H...

  20. Suboptimal protection against H5N1 highly pathogenic avian influenza viruses from Vietnam in ducks vaccinated with commercial poultry vaccines

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Highly pathogenic (HP) H5N1 avian influenza (AI) viruses continue to circulate in Asia and other regions of the world. Vaccination is used as part of H5N1 HPAI control programs in many countries; however, eradication of the disease has not been possible due to the emergence and spread of new viruses...

  1. Impact of vaccination on infection with Vietnam H5N1 high pathogenicity avian influenza virus in hens and the eggs they lay

    Technology Transfer Automated Retrieval System (TEKTRAN)

    High pathogenicity avian influenza virus (HPAIV) infections in chickens decrease egg production and eggs that are laid contain HPAIV. Vaccination once or twice was examined as a way to protect chickens from Vietnamese H5N1 HPAIV. Eighty-three percent of hens without vaccination died within 3 days ...

  2. Experimental co-infections of domestic ducks with a virulent Newcastle disease virus and low or highly pathogenic avian influenza viruses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Infections with Avian influenza viruses (AIV) of low and high pathogenicity (LP and HP), and Newcastle disease virus (NDV) are commonly reported in domestic ducks in parts of the world. However, it’s not clear if co-infections with these viruses affect the severity of the diseases they produce, the ...

  3. Pekin and Muscovy ducks respond differently to vaccination with a H5N1 highly pathogenic avian influenza (HPAI) commercial inactivated vaccine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Domestic ducks are key intermediates in the transmission of H5N1 highly pathogenic avian influenza (HPAI) viruses, and therefore are included in vaccination programs to control H5N1 HPAI. Although vaccination has proven effective in protecting ducks against disease, different species of domestic duc...

  4. Vaccine protection of turkeys against H5N1 highly pathogenic avian influenza virus with a recombinant HVT expressing the hemagglutinin gene of avian influenza

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Outbreaks of H5 highly pathogenic avian influenza (HPAI) in commercial poultry are a constant threat to animal health and food supplies. While vaccination can enhance protection and reduce the spread of disease, there is considerable evidence that the level of immunity required for protection varies...

  5. High-throughput DNA microarray detection of pathogenic bacteria in shallow well groundwater in the Kathmandu Valley, Nepal.

    PubMed

    Inoue, Daisuke; Hinoura, Takuji; Suzuki, Noriko; Pang, Junqin; Malla, Rabin; Shrestha, Sadhana; Chapagain, Saroj Kumar; Matsuzawa, Hiroaki; Nakamura, Takashi; Tanaka, Yasuhiro; Ike, Michihiko; Nishida, Kei; Sei, Kazunari

    2015-01-01

    Because of heavy dependence on groundwater for drinking water and other domestic use, microbial contamination of groundwater is a serious problem in the Kathmandu Valley, Nepal. This study investigated comprehensively the occurrence of pathogenic bacteria in shallow well groundwater in the Kathmandu Valley by applying DNA microarray analysis targeting 941 pathogenic bacterial species/groups. Water quality measurements found significant coliform (fecal) contamination in 10 of the 11 investigated groundwater samples and significant nitrogen contamination in some samples. The results of DNA microarray analysis revealed the presence of 1-37 pathogen species/groups, including 1-27 biosafety level 2 ones, in 9 of the 11 groundwater samples. While the detected pathogens included several feces- and animal-related ones, those belonging to Legionella and Arthrobacter, which were considered not to be directly associated with feces, were detected prevalently. This study could provide a rough picture of overall pathogenic bacterial contamination in the Kathmandu Valley, and demonstrated the usefulness of DNA microarray analysis as a comprehensive screening tool of a wide variety of pathogenic bacteria.

  6. The 30-Amino-Acid Deletion in the Nsp2 of Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus Emerging in China Is Not Related to Its Virulence▿

    PubMed Central

    Zhou, Lei; Zhang, Jialong; Zeng, Jingwen; Yin, Shuoyan; Li, Yanhua; Zheng, Linying; Guo, Xin; Ge, Xinna; Yang, Hanchun

    2009-01-01

    During the past 2 years, an atypical clinical outbreak, caused by a highly pathogenic porcine reproductive and respiratory syndrome virus (PRRSV) with a unique 30-amino-acid deletion in its Nsp2-coding region, was pandemic in China. In this study, we generated four full-length infectious cDNA clones: a clone of the highly virulent PRRSV strain JXwn06 (pWSK-JXwn), a clone of the low-virulence PRRSV strain HB-1/3.9 (pWSK-HB-1/3.9), a chimeric clone in which the Nsp2 region containing the 30-amino-acid deletion was replaced by the corresponding region of the low-virulence PRRSV strain HB-1/3.9 (pWSK-JXwn-HB1nsp2), and a mutated HB-1/3.9 clone with the same deletion in Nsp2 as JXwn06 (pWSK-HB1-ND30). We also investigated the pathogenicities of the rescued viruses (designated RvJXwn, RvJXwn-HB1nsp2, RvHB-1/3.9, and RvHB1-ND30, respectively) in specific-pathogen-free piglets in order to determine the role of the 30-amino-acid deletion in the virulence of the highly pathogenic PRRSV. All the rescued viruses could replicate stably in MARC-145 cells. Our findings indicated that RvJXwn-HB1nsp2 retained high virulence for piglets, like RvJXwn and the parental virus JXwn06, although the survival time of piglets infected with RvJXwn-HB1nsp2 was obviously prolonged. RvHB1-ND30 exhibited low virulence for piglets, like RvHB-1/3.9 and the parental virus HB-1/3.9. Therefore, we conclude that the 30-amino-acid deletion is not related to the virulence of the highly pathogenic PRRSV emerging in China. PMID:19244318

  7. Migration of Whooper Swans and Outbreaks of Highly Pathogenic Avian Influenza H5N1 Virus in Eastern Asia

    PubMed Central

    Newman, Scott H.; Iverson, Samuel A.; Takekawa, John Y.; Gilbert, Martin; Prosser, Diann J.; Batbayar, Nyambyar; Natsagdorj, Tseveenmyadag; Douglas, David C.

    2009-01-01

    Evaluating the potential involvement of wild avifauna in the emergence of highly pathogenic avian influenza H5N1 (hereafter H5N1) requires detailed analyses of temporal and spatial relationships between wild bird movements and disease emergence. The death of wild swans (Cygnus spp.) has been the first indicator of the presence of H5N1 in various Asian and European countries; however their role in the geographic spread of the disease remains poorly understood. We marked 10 whooper swans (Cygnus cygnus) with GPS transmitters in northeastern Mongolia during autumn 2006 and tracked their migratory movements in relation to H5N1 outbreaks. The prevalence of H5N1 outbreaks among poultry in eastern Asia during 2003–2007 peaked during winter, concurrent with whooper swan movements into regions of high poultry density. However outbreaks involving poultry were detected year round, indicating disease perpetuation independent of migratory waterbird presence. In contrast, H5N1 outbreaks involving whooper swans, as well as other migratory waterbirds that succumbed to the disease in eastern Asia, tended to occur during seasons (late spring and summer) and in habitats (areas of natural vegetation) where their potential for contact with poultry is very low to nonexistent. Given what is known about the susceptibility of swans to H5N1, and on the basis of the chronology and rates of whooper swan migration movements, we conclude that although there is broad spatial overlap between whooper swan distributions and H5N1 outbreak locations in eastern Asia, the likelihood of direct transmission between these groups is extremely low. Thus, our data support the hypothesis that swans are best viewed as sentinel species, and moreover, that in eastern Asia, it is most likely that their infections occurred through contact with asymptomatic migratory hosts (e.g., wild ducks) at or near their breeding grounds. PMID:19479053

  8. Direct metagenomic detection of viral pathogens in nasal and fecal specimens using an unbiased high-throughput sequencing approach.

    PubMed

    Nakamura, Shota; Yang, Cheng-Song; Sakon, Naomi; Ueda, Mayo; Tougan, Takahiro; Yamashita, Akifumi; Goto, Naohisa; Takahashi, Kazuo; Yasunaga, Teruo; Ikuta, Kazuyoshi; Mizutani, Tetsuya; Okamoto, Yoshiko; Tagami, Michihira; Morita, Ryoji; Maeda, Norihiro; Kawai, Jun; Hayashizaki, Yoshihide; Nagai, Yoshiyuki; Horii, Toshihiro; Iida, Tetsuya; Nakaya, Takaaki

    2009-01-01

    With the severe acute respiratory syndrome epidemic of 2003 and renewed attention on avian influenza viral pandemics, new surveillance systems are needed for the earlier detection of emerging infectious diseases. We applied a "next-generation" parallel sequencing platform for viral detection in nasopharyngeal and fecal samples collected during seasonal influenza virus (Flu) infections and norovirus outbreaks from 2005 to 2007 in Osaka, Japan. Random RT-PCR was performed to amplify RNA extracted from 0.1-0.25 ml of nasopharyngeal aspirates (N = 3) and fecal specimens (N = 5), and more than 10 microg of cDNA was synthesized. Unbiased high-throughput sequencing of these 8 samples yielded 15,298-32,335 (average 24,738) reads in a single 7.5 h run. In nasopharyngeal samples, although whole genome analysis was not available because the majority (>90%) of reads were host genome-derived, 20-460 Flu-reads were detected, which was sufficient for subtype identification. In fecal samples, bacteria and host cells were removed by centrifugation, resulting in gain of 484-15,260 reads of norovirus sequence (78-98% of the whole genome was covered), except for one specimen that was under-detectable by RT-PCR. These results suggest that our unbiased high-throughput sequencing approach is useful for directly detecting pathogenic viruses without advance genetic information. Although its cost and technological availability make it unlikely that this system will very soon be the diagnostic standard worldwide, this system could be useful for the earlier discovery of novel emerging viruses and bioterrorism, which are difficult to detect with conventional procedures. PMID:19156205

  9. Migration of whooper swans and outbreaks of highly pathogenic avian influenza H5N1 virus in Eastern Asia

    USGS Publications Warehouse

    Newman, Scott H.; Iverson, Samuel A.; Takekawa, John Y.; Gilbert, Martin; Prosser, Diann J.; Batbayar, Nyambyar; Natsagdorj, Tseveenmyadag; Douglas, David C.

    2009-01-01

    Evaluating the potential involvement of wild avifauna in the emergence of highly pathogenic avian influenza H5N1 (hereafter H5N1) requires detailed analyses of temporal and spatial relationships between wild bird movements and disease emergence. The death of wild swans (Cygnus spp.) has been the first indicator of the presence of H5N1 in various Asian and European countries; however their role in the geographic spread of the disease remains poorly understood. We marked 10 whooper swans (Cygnus cygnus) with GPS transmitters in northeastern Mongolia during autumn 2006 and tracked their migratory movements in relation to H5N1 outbreaks. The prevalence of H5N1 outbreaks among poultry in eastern Asia during 2003-2007 peaked during winter, concurrent with whooper swan movements into regions of high poultry density. However outbreaks involving poultry were detected year round, indicating disease perpetuation independent of migratory waterbird presence. In contrast, H5N1 outbreaks involving whooper swans, as well as other migratory waterbirds that succumbed to the disease in eastern Asia, tended to occur during seasons (late spring and summer) and in habitats (areas of natural vegetation) where their potential for contact with poultry is very low to nonexistent. Given what is known about the susceptibility of swans to H5N1, and on the basis of the chronology and rates of whooper swan migration movements, we conclude that although there is broad spatial overlap between whooper swan distributions and H5N1 outbreak locations in eastern Asia, the likelihood of direct transmission between these groups is extremely low. Thus, our data support the hypothesis that swans are best viewed as sentinel species, and moreover, that in eastern Asia, it is most likely that their infections occurred through contact with asymptomatic migratory hosts (e.g., wild ducks) at or near their breeding grounds.

  10. Direct Metagenomic Detection of Viral Pathogens in Nasal and Fecal Specimens Using an Unbiased High-Throughput Sequencing Approach

    PubMed Central

    Sakon, Naomi; Ueda, Mayo; Tougan, Takahiro; Yamashita, Akifumi; Goto, Naohisa; Takahashi, Kazuo; Yasunaga, Teruo; Ikuta, Kazuyoshi; Mizutani, Tetsuya; Okamoto, Yoshiko; Tagami, Michihira; Morita, Ryoji; Maeda, Norihiro; Kawai, Jun; Hayashizaki, Yoshihide; Nagai, Yoshiyuki; Horii, Toshihiro; Iida, Tetsuya; Nakaya, Takaaki

    2009-01-01

    With the severe acute respiratory syndrome epidemic of 2003 and renewed attention on avian influenza viral pandemics, new surveillance systems are needed for the earlier detection of emerging infectious diseases. We applied a “next-generation” parallel sequencing platform for viral detection in nasopharyngeal and fecal samples collected during seasonal influenza virus (Flu) infections and norovirus outbreaks from 2005 to 2007 in Osaka, Japan. Random RT-PCR was performed to amplify RNA extracted from 0.1–0.25 ml of nasopharyngeal aspirates (N = 3) and fecal specimens (N = 5), and more than 10 µg of cDNA was synthesized. Unbiased high-throughput sequencing of these 8 samples yielded 15,298–32,335 (average 24,738) reads in a single 7.5 h run. In nasopharyngeal samples, although whole genome analysis was not available because the majority (>90%) of reads were host genome–derived, 20–460 Flu-reads were detected, which was sufficient for subtype identification. In fecal samples, bacteria and host cells were removed by centrifugation, resulting in gain of 484–15,260 reads of norovirus sequence (78–98% of the whole genome was covered), except for one specimen that was under-detectable by RT-PCR. These results suggest that our unbiased high-throughput sequencing approach is useful for directly detecting pathogenic viruses without advance genetic information. Although its cost and technological availability make it unlikely that this system will very soon be the diagnostic standard worldwide, this system could be useful for the earlier discovery of novel emerging viruses and bioterrorism, which are difficult to detect with conventional procedures. PMID:19156205

  11. Pathogen intelligence.

    PubMed

    Steinert, Michael

    2014-01-01

    Different species inhabit different sensory worlds and thus have evolved diverse means of processing information, learning and memory. In the escalated arms race with host defense, each pathogenic bacterium not only has evolved its individual cellular sensing and behavior, but also collective sensing, interbacterial communication, distributed information processing, joint decision making, dissociative behavior, and the phenotypic and genotypic heterogeneity necessary for epidemiologic success. Moreover, pathogenic populations take advantage of dormancy strategies and rapid evolutionary speed, which allow them to save co-generated intelligent traits in a collective genomic memory. This review discusses how these mechanisms add further levels of complexity to bacterial pathogenicity and transmission, and how mining for these mechanisms could help to develop new anti-infective strategies. PMID:24551600

  12. Pathogen intelligence

    PubMed Central

    Steinert, Michael

    2014-01-01

    Different species inhabit different sensory worlds and thus have evolved diverse means of processing information, learning and memory. In the escalated arms race with host defense, each pathogenic bacterium not only has evolved its individual cellular sensing and behavior, but also collective sensing, interbacterial communication, distributed information processing, joint decision making, dissociative behavior, and the phenotypic and genotypic heterogeneity necessary for epidemiologic success. Moreover, pathogenic populations take advantage of dormancy strategies and rapid evolutionary speed, which allow them to save co-generated intelligent traits in a collective genomic memory. This review discusses how these mechanisms add further levels of complexity to bacterial pathogenicity and transmission, and how mining for these mechanisms could help to develop new anti-infective strategies. PMID:24551600

  13. Bio-Conjugated CNT-Bridged 3D Porous Graphene Oxide Membrane for Highly Efficient Disinfection of Pathogenic Bacteria and Removal of Toxic Metals from Water.

    PubMed

    Nellore, Bhanu Priya Viraka; Kanchanapally, Rajashekhar; Pedraza, Francisco; Sinha, Sudarson Sekhar; Pramanik, Avijit; Hamme, Ashton T; Arslan, Zikri; Sardar, Dhiraj; Ray, Paresh Chandra

    2015-09-01

    More than a billion people lack access to safe drinking water that is free from pathogenic bacteria and toxic metals. The World Health Organization estimates several million people, mostly children, die every year due to the lack of good quality water. Driven by this need, we report the development of PGLa antimicrobial peptide and glutathione conjugated carbon nanotube (CNT) bridged three-dimensional (3D) porous graphene oxide membrane, which can be used for highly efficient disinfection of Escherichia coli O157:H7 bacteria and removal of As(III), As(V), and Pb(II) from water. Reported results demonstrate that versatile membrane has the capability to capture and completely disinfect pathogenic pathogenic E. coli O157:H7 bacteria from water. Experimentally observed disinfection data indicate that the PGLa attached membrane can dramatically enhance the possibility of destroying pathogenic E. coli bacteria via synergistic mechanism. Reported results show that glutathione attached CNT-bridged 3D graphene oxide membrane can be used to remove As(III), As(V), and Pb(II) from water sample at 10 ppm level. Our data demonstrated that PGLa and glutathione attached membrane has the capability for high efficient removal of E. coli O157:H7 bacteria, As(III), As(V), and Pb(II) simultaneously from Mississippi River water. PMID:26273843

  14. Bio-Conjugated CNT-Bridged 3D Porous Graphene Oxide Membrane for Highly Efficient Disinfection of Pathogenic Bacteria and Removal of Toxic Metals from Water.

    PubMed

    Nellore, Bhanu Priya Viraka; Kanchanapally, Rajashekhar; Pedraza, Francisco; Sinha, Sudarson Sekhar; Pramanik, Avijit; Hamme, Ashton T; Arslan, Zikri; Sardar, Dhiraj; Ray, Paresh Chandra

    2015-09-01

    More than a billion people lack access to safe drinking water that is free from pathogenic bacteria and toxic metals. The World Health Organization estimates several million people, mostly children, die every year due to the lack of good quality water. Driven by this need, we report the development of PGLa antimicrobial peptide and glutathione conjugated carbon nanotube (CNT) bridged three-dimensional (3D) porous graphene oxide membrane, which can be used for highly efficient disinfection of Escherichia coli O157:H7 bacteria and removal of As(III), As(V), and Pb(II) from water. Reported results demonstrate that versatile membrane has the capability to capture and completely disinfect pathogenic pathogenic E. coli O157:H7 bacteria from water. Experimentally observed disinfection data indicate that the PGLa attached membrane can dramatically enhance the possibility of destroying pathogenic E. coli bacteria via synergistic mechanism. Reported results show that glutathione attached CNT-bridged 3D graphene oxide membrane can be used to remove As(III), As(V), and Pb(II) from water sample at 10 ppm level. Our data demonstrated that PGLa and glutathione attached membrane has the capability for high efficient removal of E. coli O157:H7 bacteria, As(III), As(V), and Pb(II) simultaneously from Mississippi River water.

  15. The Perceived Value of Passive Animal Health Surveillance: The Case of Highly Pathogenic Avian Influenza in Vietnam.

    PubMed

    Delabouglise, A; Antoine-Moussiaux, N; Phan, T D; Dao, D C; Nguyen, T T; Truong, B D; Nguyen, X N T; Vu, T D; Nguyen, K V; Le, H T; Salem, G; Peyre, M

    2016-03-01

    Economic evaluations are critical for the assessment of the efficiency and sustainability of animal health surveillance systems and the improvement of their efficiency. Methods identifying and quantifying costs and benefits incurred by public and private actors of passive surveillance systems (i.e. actors of veterinary authorities and private actors who may report clinical signs) are needed. This study presents the evaluation of perceived costs and benefits of highly pathogenic avian influenza (HPAI) passive surveillance in Vietnam. Surveys based on participatory epidemiology methods were conducted in three provinces in Vietnam to collect data on costs and benefits resulting from the reporting of HPAI suspicions to veterinary authorities. A quantitative tool based on stated preference methods and participatory techniques was developed and applied to assess the non-monetary costs and benefits. The study showed that poultry farmers are facing several options regarding the management of HPAI suspicions, besides reporting the following: treatment, sale or destruction of animals. The option of reporting was associated with uncertain outcome and transaction costs. Besides, actors anticipated the release of health information to cause a drop of markets prices. This cost was relevant at all levels, including farmers, veterinary authorities and private actors of the upstream sector (feed, chicks and medicine supply). One benefit associated with passive surveillance was the intervention of public services to clean farms and the environment to limit the disease spread. Private actors of the poultry sector valued information on HPAI suspicions (perceived as a non-monetary benefit) which was mainly obtained from other private actors and media. PMID:26146982

  16. Different environmental drivers of highly pathogenic avian influenza H5N1 outbreaks in poultry and wild birds.

    PubMed

    Si, Yali; de Boer, Willem F; Gong, Peng

    2013-01-01

    A large number of highly pathogenic avian influenza (HPAI) H5N1 outbreaks in poultry and wild birds have been reported in Europe since 2005. Distinct spatial patterns in poultry and wild birds suggest that different environmental drivers and potentially different spread mechanisms are operating. However, previous studies found no difference between these two outbreak types when only the effect of physical environmental factors was analysed. The influence of physical and anthropogenic environmental variables and interactions between the two has only been investigated for wild bird outbreaks. We therefore tested the effect of these environmental factors on HPAI H5N1 outbreaks in poultry, and the potential spread mechanism, and discussed how these differ from those observed in wild birds. Logistic regression analyses were used to quantify the relationship between HPAI H5N1 outbreaks in poultry and environmental factors. Poultry outbreaks increased with an increasing human population density combined with close proximity to lakes or wetlands, increased temperatures and reduced precipitation during the cold season. A risk map was generated based on the identified key factors. In wild birds, outbreaks were strongly associated with an increased Normalized Difference Vegetation Index (NDVI) and lower elevation, though they were similarly affected by climatic conditions as poultry outbreaks. This is the first study that analyses the differences in environmental drivers and spread mechanisms between poultry and wild bird outbreaks. Outbreaks in poultry mostly occurred in areas where the location of farms or trade areas overlapped with habitats for wild birds, whereas outbreaks in wild birds were mainly found in areas where food and shelters are available. The different environmental drivers suggest that different spread mechanisms might be involved: HPAI H5N1 spread to poultry via both poultry and wild birds, whereas contact with wild birds alone seems to drive the outbreaks

  17. Molecular evolution of H5N1 highly pathogenic avian influenza viruses in Bangladesh between 2007 and 2012.

    PubMed

    Haque, M E; Giasuddin, M; Chowdhury, E H; Islam, M R

    2014-01-01

    In Bangladesh, highly pathogenic avian influenza (HPAI) virus subtype H5N1 was first detected in February 2007. Since then the virus has become entrenched in poultry farms of Bangladesh. There have so far been seven human cases of H5N1 HPAI infection in Bangladesh with one death. The objective of the present study was to investigate the molecular evolution of H5N1 HPAI viruses during 2007 to 2012. Partial or complete nucleotide sequences of all eight gene segments of two chicken isolates, five gene segments of a duck isolate and the haemagglutinin gene segment of 18 isolates from Bangladesh were established in the present study and subjected to molecular analysis. In addition, full-length sequences of different gene segments of other Bangladeshi H5N1 isolates available in GenBank were included in the analysis. The analysis revealed that the first introduction of clade 2.2 virus in Bangladesh in 2007 was followed by the introduction of clade 2.3.2.1 and 2.3.4 viruses in 2011. However, only clade 2.3.2.1 viruses could be isolated in 2012, indicating progressive replacement of clade 2.2 and 2.3.4 viruses. There has been an event of segment re-assortment between H5N1 and H9N2 viruses in Bangladesh, where H5N1 virus acquired the PB1 gene from a H9N2 virus. Point mutations have accumulated in Bangladeshi isolates over the last 5 years with potential modification of receptor binding site and antigenic sites. Extensive and continuous molecular epidemiological studies are necessary to monitor the evolution of circulating avian influenza viruses in Bangladesh.

  18. Analysis of spatial distribution and transmission characters for highly pathogenic avian influenza in Chinese mainland in 2004

    NASA Astrophysics Data System (ADS)

    Liu, Y. L.; Wei, C. J.; Yan, L.; Chi, T. H.; Wu, X. B.; Xiao, C. S.

    2006-03-01

    After the outbreak of highly pathogenic Avian Influenza (HPAI) in South Korea in the end of year 2003, estimates of the impact of HPAI in affected countries vary greatly, the total direct losses are about 3 billion US dollars, and it caused 15 million birds and poultry flocks death. It is significant to understand the spatial distribution and transmission characters of HPAI for its prevention and control. According to 50 outbreak cases for HPAI in Chinese mainland during 2004, this paper introduces the approach of spatial distribution and transmission characters for HPAI and its results. Its approach is based on remote sensing and GIS techniques. Its supporting data set involves normalized difference vegetation index (NDVI) and land surface temperature (Ts) derived from a time-series of remote sensing data of 1 kilometer-resolution NOAA/AVHRR, birds' migration routes, topology geographic map, lake and wetland maps, and meteorological observation data. In order to analyze synthetically using these data, a supporting platform for analysis Avian Influenza epidemic situation (SPAS/AI) was developed. Supporting by SPAS/AI, the integrated information from multi-sources can be easily used to the analysis of the spatial distribution and transmission character of HPAI. The results show that the range of spatial distribution and transmission of HPAI in China during 2004 connected to environment factors NDVI, Ts and the distributions of lake and wetland, and especially to bird migration routes. To some extent, the results provide some suggestions for the macro-decision making for the prevention and control of HPAI in the areas of potential risk and reoccurrence.

  19. The Variable Region of Pneumococcal Pathogenicity Island 1 Is Responsible for Unusually High Virulence of a Serotype 1 Isolate.

    PubMed

    Harvey, Richard M; Trappetti, Claudia; Mahdi, Layla K; Wang, Hui; McAllister, Lauren J; Scalvini, Alexandra; Paton, Adrienne W; Paton, James C

    2016-03-01

    Streptococcus pneumoniae is the leading infectious cause of death in children in the world. However, the mechanisms that drive the progression from asymptomatic colonization to disease are poorly understood. Two virulence-associated genomic accessory regions (ARs) were deleted in a highly virulent serotype 1 clinical isolate (strain 4496) and examined for their contribution to pathogenesis. Deletion of a prophage encoding a platelet-binding protein (PblB) resulted in reduced adherence, biofilm formation, reduced initial infection within the lungs, and a reduction in the number of circulating platelets in infected mice. However, the region's overall contribution to the survival of mice was not significant. In contrast, deletion of the variable region of pneumococcal pathogenicity island 1 (vPPI1) was also responsible for a reduction in adherence and biofilm formation but also reduced survival and invasion of the pleural cavity, blood, and lungs. While the 4496ΔPPI1 strain induced higher expression of the genes encoding interleukin-10 (IL-10) and CD11b in the lungs of challenged mice than the wild-type strain, very few other genes exhibited altered expression. Moreover, while the level of IL-10 protein was increased in the lungs of 4496ΔPPI1 mutant-infected mice compared to strain 4496-infected mice, the levels of gamma interferon (IFN-γ), CXCL10, CCL2, and CCL4 were not different in the two groups. However, the 4496ΔPPI1 mutant was found to be more susceptible than the wild type to phagocytic killing by a macrophage-like cell line. Therefore, our data suggest that vPPI1 may be a major contributing factor to the heightened virulence of certain serotype 1 strains, possibly by influencing resistance to phagocytic killing. PMID:26755156

  20. U.S. Geological Survey science strategy for highly pathogenic avian influenza in wildlife and the environment (2016–2020)

    USGS Publications Warehouse

    Harris, M. Camille; Pearce, John M.; Prosser, Diann J.; White, C. LeAnn; Miles, A. Keith; Sleeman, Jonathan M.; Brand, Christopher J.; Cronin, James P.; De La Cruz, Susan; Densmore, Christine L.; Doyle, Thomas W.; Dusek, Robert J.; Fleskes, Joseph P.; Flint, Paul L.; Guala, Gerald F.; Hall, Jeffrey S.; Hubbard, Laura E.; Hunt, Randall J.; Ip, Hon S.; Katz, Rachel A.; Laurent, Kevin W.; Miller, Mark P.; Munn, Mark D.; Ramey, Andy M.; Richards, Kevin D.; Russell, Robin E.; Stokdyk, Joel P.; Takekawa, John Y.; Walsh, Daniel P.

    2016-08-18

    IntroductionThrough the Science Strategy for Highly Pathogenic Avian Influenza (HPAI) in Wildlife and the Environment, the USGS will assess avian influenza (AI) dynamics in an ecological context to inform decisions made by resource managers and policymakers from the local to national level. Through collection of unbiased scientific information on the ecology of AI viruses and wildlife hosts in a changing world, the U.S. Geological Survey (USGS) will enhance the development of AI forecasting tools and ensure this information is integrated with a quality decision process for managing HPAI.The overall goal of this USGS Science Strategy for HPAI in Wildlife and the Environment goes beyond document­ing the occurrence and distribution of AI viruses in wild birds. The USGS aims to understand the epidemiological processes and environmental factors that influence HPAI distribution and describe the mechanisms of transmission between wild birds and poultry. USGS scientists developed a conceptual model describing the process linking HPAI dispersal in wild waterfowl to the outbreaks in poul­try. This strategy focuses on five long-term science goals, which include:Science Goal 1—Augment the National HPAI Surveillance Plan;Science Goal 2—Determine mechanisms of HPAI disease spread in wildlife and the environment;Science Goal 3—Characterize HPAI viruses circulating in wildlife;Science Goal 4—Understand implications of avian ecol­ogy on HPAI spread; andScience Goal 5—Develop HPAI forecasting and decision-making tools.These goals will help define and describe the processes outlined in the conceptual model with the ultimate goal of facilitating biosecurity and minimizing transfer of diseases across the wildlife-poultry interface. The first four science goals are focused on scientific discovery and the fifth goal is application-based. Decision analyses in the fifth goal will guide prioritization of proposed actions in the first four goals.

  1. Determinants of Knowledge and Biosecurity Preventive Behaviors for Highly Pathogenic Avian Influenza Risk Among Chinese Poultry Farmers.

    PubMed

    Cui, Bin; Liu, Zong Ping

    2016-06-01

    Biosecurity measures are the first line of defense against highly pathogenic avian influenza (HPAI) on farms. It is generally recognized that an individual's behavior can be influenced by the knowledge they possess. However, empirical study has not reported an association between poultry producers' awareness of HPAI symptoms and their actual biosecurity actions. The aim of this study is to classify knowledge items of HPAI by exploratory factor analysis (EFA) and to examine the determinants of different types of knowledge and the effect of different types of knowledge on biosecurity preventive behaviors (BPBs). The survey (n = 297) was conducted using a questionnaire to measure the level of awareness of items related to HPAI and the actual adoption of BPBs among poultry farmers in the Chinese province of Jiangsu. The EFA revealed three main types of knowledge, which were categorized as avian influenza (AI) epidemic characteristics, primary biosecurity preventive knowledge (basic biosecurity preventive knowledge against AI), and essential biosecurity preventive knowledge (crucial biosecurity preventive knowledge against infection of AI). Multivariate regression showed that only poultry farmers' awareness of essential biosecurity preventive knowledge was positively associated with their actual BPBs. Additionally, educational attainment, number of years of experience raising poultry, farming operation size, and training were associated both with BPB and most of the knowledge factors or knowledge items. Training of existing poultry farmers is probably a feasible scheme; furthermore, the training should focus on the essential biosecurity preventive knowledge. On the other hand, policy initiatives to encourage large-scale poultry farming while discouraging small-scale backyard poultry husbandry would be an effective method of improving the management standards of rural poultry farming. PMID:27309291

  2. Molecular and antigenic evolution and geographical spread of H5N1 highly pathogenic avian influenza viruses in western Africa.

    PubMed

    Ducatez, M F; Olinger, C M; Owoade, A A; Tarnagda, Z; Tahita, M C; Sow, A; De Landtsheer, S; Ammerlaan, W; Ouedraogo, J B; Osterhaus, A D M E; Fouchier, R A M; Muller, C P

    2007-08-01

    In Africa, highly pathogenic avian influenza H5N1 virus was first detected in northern Nigeria and later also in other regions of the country. Since then, seven other African countries have reported H5N1 infections. This study reports a comparison of full-length genomic sequences of H5N1 isolates from seven chicken farms in Nigeria and chicken and hooded vultures in Burkina Faso with earlier H5N1 outbreaks worldwide. In addition, the antigenicity of Nigerian H5N1 isolates was compared with earlier strains. All African strains clustered within three sublineages denominated A (south-west Nigeria, Niger), B (south-west Nigeria, Egypt, Djibouti) and C (northern Nigeria, Burkina Faso, Sudan, Côte d'Ivoire), with distinct nucleotide and amino acid signatures and distinct geographical distributions within Africa. Probable non-African ancestors within the west Asian/Russian/European lineage distinct from the south-east Asian lineages were identified for each sublineage. All reported human cases in Africa were caused by sublineage B. Substitution rates were calculated on the basis of sequences from 11 strains from a single farm in south-west Nigeria. As H5N1 emerged essentially at the same time in the north and south-west of Nigeria, the substitution rates confirmed that the virus probably did not spread from the north to the south, given the observed sequence diversity, but that it entered the country via three independent introductions. The strains from Burkina Faso seemed to originate from northern Nigeria. At least two of the sublineages also circulated in Europe in 2006 as seen in Germany, further suggesting that the sublineages had already emerged outside of Africa and seemed to have followed the east African/west Asian and Black Sea/Mediterranean flyways of migratory birds. PMID:17622635

  3. Development of indirect ELISAs for differential serodiagnosis of classical and highly pathogenic porcine reproductive and respiratory syndrome virus.

    PubMed

    Xiao, Y H; Wang, T T; Zhao, Q; Wang, C B; Lv, J H; Nie, L; Gao, J M; Ma, X C; Hsu, W H; Zhou, E M

    2014-08-01

    The objective of this study was to develop two indirect enzyme-linked immunosorbent assays (iELISAs) for detection of serum antibodies against classical vaccine strain of porcine reproductive and respiratory syndrome virus (PRRSV) and highly pathogenic PRRSV (HP-PRRSV). To detect the common antibodies against classical and HP-PRRSV, the coating antigen used in the iELISA (designated iELISA-180) was the antigen of Nsp2-180, the 180aa at amino terminal of Nsp2. To detect the different antibodies against classical and HP-PRRSV, the coating antigen in the second iELISA (designated iELISA-D29) was Nsp2-D29, the deleted 29aa in Nsp2 of HP-PRRSV. The antigen concentration and serum dilutions were optimized using a draughtboard titration. The cut-off values of 0.361 at OD(450nm) for the iELISA-180 and 0.27 at OD(450nm) for the iELISA-D29 were determined by testing a panel of 120 classical PRRSV positive and 198 PRRSV negative pig serum samples, which generated the specificity of 97.1% and 96.7%, the sensitivity of 96.9% and 96.3% for iELISA-180 and iELISA-D29, respectively. The agreements between the Western blot and iELISA-180 and iELISA-D29 were 98%, 96.7%, respectively. The developed iELISAs can be used to differentiate serologically HP-PRRSV from the vaccinated or classical PRRSV in clinical serum samples.

  4. Persistence of highly pathogenic avian influenza H5N1 virus defined by agro-ecological niche

    USGS Publications Warehouse

    Hogerwerf, Lenny; Wallace, Rob G.; Ottaviani, Daniela; Slingenbergh, Jan; Prosser, Diann; Bergmann, Luc; Gilbert, Marius

    2010-01-01

    The highly pathogenic avian influenza (HPAI) H5N1 virus has spread across Eurasia and into Africa. Its persistence in a number of countries continues to disrupt poultry production, impairs smallholder livelihoods, and raises the risk a genotype adapted to human-to-human transmission may emerge. While previous studies identified domestic duck reservoirs as a primary risk factor associated with HPAI H5N1 persistence in poultry in Southeast Asia, little is known of such factors in countries with different agro-ecological conditions, and no study has investigated the impact of such conditions on HPAI H5N1 epidemiology at the global scale. This study explores the patterns of HPAI H5N1 persistence worldwide, and for China, Indonesia, and India includes individual provinces that have reported HPAI H5N1 presence during the 2004–2008 period. Multivariate analysis of a set of 14 agricultural, environmental, climatic, and socio-economic factors demonstrates in quantitative terms that a combination of six variables discriminates the areas with human cases and persistence: agricultural population density, duck density, duck by chicken density, chicken density, the product of agricultural population density and chicken output/input ratio, and purchasing power per capita. The analysis identifies five agro-ecological clusters, or niches, representing varying degrees of disease persistence. The agro-ecological distances of all study areas to the medoid of the niche with the greatest number of human cases are used to map HPAI H5N1 risk globally. The results indicate that few countries remain where HPAI H5N1 would likely persist should it be introduced.

  5. Persistence of highly pathogenic avian influenza H5N1 virus defined by agro-ecological niche.

    PubMed

    Hogerwerf, Lenny; Wallace, Rob G; Ottaviani, Daniela; Slingenbergh, Jan; Prosser, Diann; Bergmann, Luc; Gilbert, Marius

    2010-06-01

    The highly pathogenic avian influenza (HPAI) H5N1 virus has spread across Eurasia and into Africa. Its persistence in a number of countries continues to disrupt poultry production, impairs smallholder livelihoods, and raises the risk a genotype adapted to human-to-human transmission may emerge. While previous studies identified domestic duck reservoirs as a primary risk factor associated with HPAI H5N1 persistence in poultry in Southeast Asia, little is known of such factors in countries with different agro-ecological conditions, and no study has investigated the impact of such conditions on HPAI H5N1 epidemiology at the global scale. This study explores the patterns of HPAI H5N1 persistence worldwide, and for China, Indonesia, and India includes individual provinces that have reported HPAI H5N1 presence during the 2004-2008 period. Multivariate analysis of a set of 14 agricultural, environmental, climatic, and socio-economic factors demonstrates in quantitative terms that a combination of six variables discriminates the areas with human cases and persistence: agricultural population density, duck density, duck by chicken density, chicken density, the product of agricultural population density and chicken output/input ratio, and purchasing power per capita. The analysis identifies five agro-ecological clusters, or niches, representing varying degrees of disease persistence. The agro-ecological distances of all study areas to the medoid of the niche with the greatest number of human cases are used to map HPAI H5N1 risk globally. The results indicate that few countries remain where HPAI H5N1 would likely persist should it be introduced.

  6. Determinants of Knowledge and Biosecurity Preventive Behaviors for Highly Pathogenic Avian Influenza Risk Among Chinese Poultry Farmers.

    PubMed

    Cui, Bin; Liu, Zong Ping

    2016-06-01

    Biosecurity measures are the first line of defense against highly pathogenic avian influenza (HPAI) on farms. It is generally recognized that an individual's behavior can be influenced by the knowledge they possess. However, empirical study has not reported an association between poultry producers' awareness of HPAI symptoms and their actual biosecurity actions. The aim of this study is to classify knowledge items of HPAI by exploratory factor analysis (EFA) and to examine the determinants of different types of knowledge and the effect of different types of knowledge on biosecurity preventive behaviors (BPBs). The survey (n = 297) was conducted using a questionnaire to measure the level of awareness of items related to HPAI and the actual adoption of BPBs among poultry farmers in the Chinese province of Jiangsu. The EFA revealed three main types of knowledge, which were categorized as avian influenza (AI) epidemic characteristics, primary biosecurity preventive knowledge (basic biosecurity preventive knowledge against AI), and essential biosecurity preventive knowledge (crucial biosecurity preventive knowledge against infection of AI). Multivariate regression showed that only poultry farmers' awareness of essential biosecurity preventive knowledge was positively associated with their actual BPBs. Additionally, educational attainment, number of years of experience raising poultry, farming operation size, and training were associated both with BPB and most of the knowledge factors or knowledge items. Training of existing poultry farmers is probably a feasible scheme; furthermore, the training should focus on the essential biosecurity preventive knowledge. On the other hand, policy initiatives to encourage large-scale poultry farming while discouraging small-scale backyard poultry husbandry would be an effective method of improving the management standards of rural poultry farming.

  7. The Perceived Value of Passive Animal Health Surveillance: The Case of Highly Pathogenic Avian Influenza in Vietnam.

    PubMed

    Delabouglise, A; Antoine-Moussiaux, N; Phan, T D; Dao, D C; Nguyen, T T; Truong, B D; Nguyen, X N T; Vu, T D; Nguyen, K V; Le, H T; Salem, G; Peyre, M

    2016-03-01

    Economic evaluations are critical for the assessment of the efficiency and sustainability of animal health surveillance systems and the improvement of their efficiency. Methods identifying and quantifying costs and benefits incurred by public and private actors of passive surveillance systems (i.e. actors of veterinary authorities and private actors who may report clinical signs) are needed. This study presents the evaluation of perceived costs and benefits of highly pathogenic avian influenza (HPAI) passive surveillance in Vietnam. Surveys based on participatory epidemiology methods were conducted in three provinces in Vietnam to collect data on costs and benefits resulting from the reporting of HPAI suspicions to veterinary authorities. A quantitative tool based on stated preference methods and participatory techniques was developed and applied to assess the non-monetary costs and benefits. The study showed that poultry farmers are facing several options regarding the management of HPAI suspicions, besides reporting the following: treatment, sale or destruction of animals. The option of reporting was associated with uncertain outcome and transaction costs. Besides, actors anticipated the release of health information to cause a drop of markets prices. This cost was relevant at all levels, including farmers, veterinary authorities and private actors of the upstream sector (feed, chicks and medicine supply). One benefit associated with passive surveillance was the intervention of public services to clean farms and the environment to limit the disease spread. Private actors of the poultry sector valued information on HPAI suspicions (perceived as a non-monetary benefit) which was mainly obtained from other private actors and media.

  8. Transforming Growth Factor-β: Activation by Neuraminidase and Role in Highly Pathogenic H5N1 Influenza Pathogenesis

    PubMed Central

    Moser, Lindsey A.; O'Brien, Kevin B.; Cline, Troy D.; Jones, Jeremy C.; Tumpey, Terrence M.; Katz, Jacqueline M.; Kelley, Laura A.; Gauldie, Jack; Schultz-Cherry, Stacey

    2010-01-01

    Transforming growth factor-beta (TGF-β), a multifunctional cytokine regulating several immunologic processes, is expressed by virtually all cells as a biologically inactive molecule termed latent TGF-β (LTGF-β). We have previously shown that TGF-β activity increases during influenza virus infection in mice and suggested that the neuraminidase (NA) protein mediates this activation. In the current study, we determined the mechanism of activation of LTGF-β by NA from the influenza virus A/Gray Teal/Australia/2/1979 by mobility shift and enzyme inhibition assays. We also investigated whether exogenous TGF-β administered via a replication-deficient adenovirus vector provides protection from H5N1 influenza pathogenesis and whether depletion of TGF-β during virus infection increases morbidity in mice. We found that both the influenza and bacterial NA activate LTGF-β by removing sialic acid motifs from LTGF-β, each NA being specific for the sialic acid linkages cleaved. Further, NA likely activates LTGF-β primarily via its enzymatic activity, but proteases might also play a role in this process. Several influenza A virus subtypes (H1N1, H1N2, H3N2, H5N9, H6N1, and H7N3) except the highly pathogenic H5N1 strains activated LTGF-β in vitro and in vivo. Addition of exogenous TGF-β to H5N1 influenza virus–infected mice delayed mortality and reduced viral titers whereas neutralization of TGF-β during H5N1 and pandemic 2009 H1N1 infection increased morbidity. Together, these data show that microbe-associated NAs can directly activate LTGF-β and that TGF-β plays a pivotal role protecting the host from influenza pathogenesis. PMID:20949074

  9. A single vaccination of commercial broilers does not reduce transmission of H5N1 highly pathogenic avian influenza

    PubMed Central

    2011-01-01

    Vaccination of chickens has become routine practice in Asian countries in which H5N1 highly pathogenic avian influenza (HPAI) is endemically present. This mainly applies to layer and breeder flocks, but broilers are usually left unvaccinated. Here we investigate whether vaccination is able to reduce HPAI H5N1 virus transmission among broiler chickens. Four sets of experiments were carried out, each consisting of 22 replicate trials containing a pair of birds. Experiments 1-3 were carried out with four-week-old birds that were unvaccinated, and vaccinated at day 1 or at day 10 of age. Experiment 4 was carried out with unvaccinated day-old broiler chicks. One chicken in each trial was inoculated with H5N1 HPAI virus. One chicken in each trial was inoculated with virus. The course of the infection chain was monitored by serological analysis, and by virus isolation performed on tracheal and cloacal swabs. The analyses were based on a stochastic SEIR model using a Bayesian inferential framework. When inoculation was carried out at the 28th day of life, transmission was efficient in unvaccinated birds, and in birds vaccinated at first or tenth day of life. In these experiments estimates of the latent period (~1.0 day), infectious period (~3.3 days), and transmission rate parameter (~1.4 per day) were similar, as were estimates of the reproduction number (~4) and generation interval (~1.4 day). Transmission was significantly less efficient in unvaccinated chickens when inoculation was carried out on the first day of life. These results show that vaccination of broiler chickens does not reduce transmission, and suggest that this may be due to the interference of maternal immunity. PMID:21635732

  10. Risk Assessment of High Pathogenicity Avian Influenza Virus Introduction into Poland via Legal Importation of Live Poultry.

    PubMed

    Gierak, Anna; Bocian, Łukasz; Śmietanka, Krzysztof

    2016-05-01

    The risk of highly pathogenic avian influenza (HPAI) virus introduction via import of live poultry results from the probability that infected birds are exported from apparently HPAI-free areas during the silent phase of the epidemic, i.e., the period between an incursion of the virus into a susceptible population and a report on the outbreak by an exporting country. In our study we adapted a stochastic model, previously published in 2010 by Sánchez-Vizcaíno et al., with our own modifications in which the probability of HPAI introduction was assessed as the sum of the probabilities of entry of at least one infected bird from each susceptible species exported from each country into each Polish region (county). The mean annual probability of HPAI introduction into Poland via legal trade of live poultry was very low (3.07 × 10(-3), which corresponds to 1 outbreak every 326 yr). The highest risk was associated with the import of turkeys (62%) and chickens (33%). The exporting countries that contributed the most to the overall risk were Italy (31%), the Netherlands (24%), and the Czech Republic (17%). The risk was not evenly distributed across the country and it seemed higher in western, north-central, and eastern Poland while several counties of the north-west, central, or south-east parts of the country were at negligible risk. The applied model provides quantitative evidence that the risk of HPAI introduction through legal trade of poultry does not play a major role and that other paths, such as wild birds migrations or illegal trade, should be considered as the most-likely routes along which the virus can be introduced. PMID:27309053

  11. Highly Pathogenic Eurasian H5N8 Avian Influenza Outbreaks in Two Commercial Poultry Flocks in California.

    PubMed

    Stoute, Simone; Chin, Richard; Crossley, Beate; Gabriel Sentíes-Cué, C; Bickford, Arthur; Pantin-Jackwood, Mary; Breitmeyer, Richard; Jones, Annette; Carnaccini, Silvia; Shivaprasad, H L

    2016-09-01

    In January 2015, a highly pathogenic Eurasian lineage H5N8 avian influenza (AI) virus (AIV) was detected in a commercial meat turkey flock in Stanislaus County, CA. Approximately 3 wk later, a similar case was diagnosed in commercial brown layers from a different company located in Kings County, CA. Five 14-wk-old turkey hens were submitted to the California Animal Health and Food Safety Laboratory System (CAHFS), Turlock, and eleven 12-wk-old chickens were submitted to CAHFS, Tulare laboratory due to an acute increase in flock mortality. Gross lesions included enlarged and mottled pale spleens and pancreas in turkeys and chickens. Histologically, the major lesions observed in turkeys and chickens were splenitis, pancreatitis, encephalitis, and pneumonia. In both cases, diagnosis was based on real-time reverse transcriptase PCR (RRT-PCR), sequencing, and virus isolation from oropharyngeal and cloacal swabs. Confirmatory diagnosis and AIV characterization was done at the National Veterinary Services Laboratory, Ames, IA. The sequence of the AIV from both cases was 99% identical to an H5N8 AI virus (A/gyrfalcon/Washington/41088-6/2014) isolated from a captive gyrfalcon (Falco rusticolus) from Washington State in December 2014. Immunohistochemistry (IHC) performed on various tissues from both cases indicated a widespread AIV tissue distribution. Except for minor variations, the tissue distribution of the AI antigen was similar in the chickens and turkeys. There was positive IHC staining in the brain, spleen, pancreas, larynx, trachea, and lungs in both chickens and turkeys. Hearts, ovaries, and air sacs from the turkeys were also positive for the AI antigen. The liver sections from the chickens had occasional AI-positive staining in mononuclear cells, but the IHC on liver sections from the turkeys were negative. The bursa of Fabricius, small intestine, kidney, and skeletal muscle sections were negative for the AI antigen in both chickens and turkeys. PMID:27610732

  12. U.S. Geological Survey science strategy for highly pathogenic avian influenza in wildlife and the environment (2016–2020)

    USGS Publications Warehouse

    Harris, M. Camille; Pearce, John M.; Prosser, Diann J.; White, C. LeAnn; Miles, A. Keith; Sleeman, Jonathan M.; Brand, Christopher J.; Cronin, James P.; De La Cruz, Susan; Densmore, Christine L.; Doyle, Thomas W.; Dusek, Robert J.; Fleskes, Joseph P.; Flint, Paul L.; Guala, Gerald F.; Hall, Jeffrey S.; Hubbard, Laura E.; Hunt, Randall J.; Ip, Hon S.; Katz, Rachel A.; Laurent, Kevin W.; Miller, Mark P.; Munn, Mark D.; Ramey, Andy M.; Richards, Kevin D.; Russell, Robin E.; Stokdyk, Joel P.; Takekawa, John Y.; Walsh, Daniel P.

    2016-01-01

    IntroductionThrough the Science Strategy for Highly Pathogenic Avian Influenza (HPAI) in Wildlife and the Environment, the USGS will assess avian influenza (AI) dynamics in an ecological context to inform decisions made by resource managers and policymakers from the local to national level. Through collection of unbiased scientific information on the ecology of AI viruses and wildlife hosts in a changing world, the U.S. Geological Survey (USGS) will enhance the development of AI forecasting tools and ensure this information is integrated with a quality decision process for managing HPAI.The overall goal of this USGS Science Strategy for HPAI in Wildlife and the Environment goes beyond document­ing the occurrence and distribution of AI viruses in wild birds. The USGS aims to understand the epidemiological processes and environmental factors that influence HPAI distribution and describe the mechanisms of transmission between wild birds and poultry. USGS scientists developed a conceptual model describing the process linking HPAI dispersal in wild waterfowl to the outbreaks in poul­try. This strategy focuses on five long-term science goals, which include:Science Goal 1—Augment the National HPAI Surveillance Plan;Science Goal 2—Determine mechanisms of HPAI disease spread in wildlife and the environment;Science Goal 3—Characterize HPAI viruses circulating in wildlife;Science Goal 4—Understand implications of avian ecol­ogy on HPAI spread; andScience Goal 5—Develop HPAI forecasting and decision-making tools.These goals will help define and describe the processes outlined in the conceptual model with the ultimate goal of facilitating biosecurity and minimizing transfer of diseases across the wildlife-poultry interface. The first four science goals are focused on scientific discovery and the fifth goal is application-based. Decision analyses in the fifth goal will guide prioritization of proposed actions in the first four goals.

  13. Duck MDA5 functions in innate immunity against H5N1 highly pathogenic avian influenza virus infections.

    PubMed

    Wei, Liangmeng; Cui, Jin; Song, Yafen; Zhang, Shuo; Han, Fei; Yuan, Runyu; Gong, Lang; Jiao, Peirong; Liao, Ming

    2014-01-01

    Melanoma differentiation-associated gene 5 (MDA5) is an important intracellular receptor that recognizes long molecules of viral double-stranded RNA in innate immunity. To understand the mechanism of duck MDA5-mediated innate immunity, we cloned the MDA5 cDNA from the Muscovy duck (Cairina moschata). Quantitative real-time PCR analysis indicates that duck MDA5 mRNA was constitutively expressed in all sampled tissues. A significant increase of MDA5 mRNA was detected in the brain, spleen and lungs of ducks after infection with an H5N1 highly pathogenic avian influenza virus (HPAIV). We investigated the role of the predicted functional domains of MDA5. The results indicate the caspase activation and recruitment domain (CARD) of duck MDA5 had a signal transmission function through IRF-7-dependent signaling pathway. Overexpression of the CARD strongly activated the chicken IFN-β promoter and upregulated the mRNA expression of antiviral molecules (such as OAS, PKR and Mx), proinflammatory cytokines (such as IL-2, IL-6, IFN-α and IFN-γ, but not IL-1β and IL-8) and retinoic acid-inducible gene I (RIG-I)-like receptors (RLR) (RIG-I and LGP2) without exogenous stimulation. We also demonstrate the NS1 of the H5N1 HPAIV inhibited the duck MDA5-mediated signaling pathway in vitro. These results suggest that duck MDA5 is an important receptor for inducing antiviral activity in the host immune response of ducks.

  14. The effectiveness of preventative mass vaccination regimes against the incidence of highly pathogenic avian influenza on Java Island, Indonesia.

    PubMed

    Bett, B; McLaws, M; Jost, C; Schoonman, L; Unger, F; Poole, J; Lapar, M L; Siregar, E S; Azhar, M; Hidayat, M M; Dunkle, S E; Mariner, J

    2015-04-01

    We conducted an operational research study involving backyard and semicommercial farms on Java Island, Indonesia, between April 2008 and September 2009 to evaluate the effectiveness of two preventive mass vaccination strategies against highly pathogenic avian influenza (HPAI). One regimen used Legok 2003 H5N1 vaccine, while the other used both Legok 2003 H5N1 and HB1 Newcastle disease (ND) vaccine. A total of 16 districts were involved in the study. The sample size was estimated using a formal power calculation technique that assumed a detectable effect of treatment as a 50% reduction in the baseline number of HPAI-compatible outbreaks. Within each district, candidate treatment blocks with village poultry populations ranging from 80 000 to 120 000 were created along subdistrict boundary lines. Subsequently, four of these blocks were randomly selected and assigned one treatment from a list that comprised control, vaccination against HPAI, vaccination against HPAI + ND. Four rounds of vaccination were administered at quarterly intervals beginning in July 2008. A vaccination campaign involved vaccinating 100 000 birds in a treatment block, followed by another 100 000 vaccinations 3 weeks later as a booster dose. Data on disease incidence and vaccination coverage were also collected at quarterly intervals using participatory epidemiological techniques. Compared with the unvaccinated (control) group, the incidence of HPAI-compatible events declined by 32% (P = 0.24) in the HPAI-vaccinated group and by 73% (P = 0.00) in the HPAI- and ND-vaccinated group. The effect of treatment did not vary with time or district. Similarly, an analysis of secondary data from the participatory disease and response (PDSR) database revealed that the incidence of HPAI declined by 12% in the HPAI-vaccinated group and by 24% in the HPAI + ND-vaccinated group. The results suggest that the HPAI + ND vaccination significantly reduced the incidence of HPAI-compatible events in mixed populations of

  15. Risk factors and characteristics of H5N1 Highly Pathogenic Avian Influenza (HPAI) post-vaccination outbreaks.

    PubMed

    Henning, Joerg; Pfeiffer, Dirk U; Vu, Le Tri

    2009-01-01

    Highly pathogenic avian influenza (HPAI) virus H5N1 is now endemic in South-East Asia but HPAI control methods differ between countries. A widespread HPAI vaccination campaign that started at the end of 2005 in Viet Nam resulted in the cessation of poultry and human cases, but in 2006/2007 severe HPAI outbreaks re-emerged. In this study we investigated the pattern of this first post-vaccination epidemic in southern Viet Nam identifying a spatio-temporal cluster of outbreak occurrence and estimating spatially smoothed incidence rates of HPAI. Spatial risk factors associated with HPAI occurrence were identified. Medium-level poultry density resulted in an increased outbreak risk (Odds ratio (OR) = 5.4, 95% confidence interval (CI): 1.6-18.9) but also climate-vegetation factors played an important role: medium-level normalised difference vegetation indices during the rainy season from May to October were associated with higher risk of HPAI outbreaks (OR = 3.7, 95% CI: 1.7-8.1), probably because temporal flooding might have provided suitable conditions for the re-emergence of HPAI by expanding the virus distribution in the environment and by enlarging areas of possible contacts between domestic waterfowl and wild birds. On the other hand, several agricultural production factors, such as sweet potatoes yield, increased buffalo density, as well as increased electricity supply were associated with decreased risk of HPAI outbreaks. This illustrates that preventive control measures for HPAI should include a promotion of low-risk agricultural management practices as well as improvement of the infrastructure in village households. Improved HPAI vaccination efforts and coverage should focus on medium poultry density areas and on the pre-monsoon time period.

  16. The North American strain of viral hemorrhagic septicemia virus is highly pathogenic for laboratory-reared Pacific herring (Clupea pallasi)

    USGS Publications Warehouse

    Kocan, R.; Bradley, M.; Elder, N.; Meyers, T.; Batts, W.; Winton, J.

    1997-01-01

    Specific-pathogen-free Pacific herring Clupea pallasi were reared in the laboratory from eggs and then challenged at 5, 9, and 13 months of age by waterborne exposure to low (101.5–2.5 plaque-forming units [PFU] per milliliter), medium (103.5–4.5 PFU/mL), or high (105.5–6.5 PFU/mL) levels of a North American isolate of viral hemorrhagic septicemia virus (VHSV). The fish were extremely susceptible to the virus, showing clinical disease, mortality approaching 100%, and only a limited increase in resistance with age. Mortality began 4–6 d after exposure and peaked at approximately day 7 in fish exposed to high levels of virus. Whereas the mean time to death showed a significant dose response (P < 0.001), the percent mortality and virus titers in dead fish were generally high in all groups regardless of initial challenge dose. External signs of disease were usually limited to 1–2-mm hemorrhagic areas on the lower jaw and isthmus and around the eye, but 2 of 130 infected fish exhibited extensive cutaneous hemorrhaging. Histopathologic examination of tissues from moribund fish sampled at 2–8 d after exposure revealed multifocal coagulative necrosis of hepatocytes, diffuse necrosis of interstitial hematopoietic tissues in the kidney, diffuse necrosis of the spleen, epidermis, and subcutis, and occasional necrosis of pancreatic acinar cells. Virus titers in tissues of experimentally infected herring were first detected 48 h after exposure and peaked 6-8 d after exposure at 107.7 PFU/g. Fish began shedding virus at 48 h after exposure with titers in the flow-through aquaria reaching 102.5 PFU/mL at 4–5 d after exposure, just before peak mortality. When the water flow was turned off for 3 h, titers in the water rose to 103.5 PFU/mL, and the amount of virus shed by infected fish (on average, greater than 106.5 PFU/h per fish) appeared sufficient to sustain a natural epizootic among schooling herring. Taken together, these data suggest that VHSV could be a

  17. Validation of a high-throughput immunobead array technique for multiplex detection of three foodborne pathogens in chicken products.

    PubMed

    Charlermroj, Ratthaphol; Makornwattana, Manlika; Grant, Irene R; Elliott, Christopher T; Karoonuthaisiri, Nitsara

    2016-05-01

    This study rigorously evaluated a previously developed immunobead array method to simultaneously detect three important foodborne pathogens, Campylobacter jejuni, Listeria monocytogenes, and Salmonella spp., for its actual application in routine food testing. Due to the limitation of the detection limit of the developed method, an enrichment step was included in this study by using Campylobacter Enrichment Broth for C. jejuni and Universal Pre-enrichment Broth for L. monocytogenes and Salmonella spp.. The findings showed that the immunobead array method was capable of detecting as low as 1CFU of the pathogens spiked in the culture media after being cultured for 24h for all three pathogens. The immunobead array method was further evaluated for its pathogen detection capabilities in ready-to-eat (RTE) and ready-to-cook (RTC) chicken samples and proven to be able to detect as low as 1CFU of the pathogens spiked in the food samples after being cultured for 24h in the case of Salmonella spp., and L. monocytogenes and 48 h in the case of C. jejuni. The method was subsequently validated with three types of chicken products (RTE, n=30; RTC, n=20; raw chicken, n=20) and was found to give the same results as the conventional plating method. Our findings demonstrated that the previously developed immunobead array method could be used for actual food testing with minimal enrichment period of only 52 h, whereas the conventional ISO protocols for the same pathogens take 90-144 h. The immunobead array was therefore an inexpensive, rapid and simple method for the food testing.

  18. Validation of a high-throughput immunobead array technique for multiplex detection of three foodborne pathogens in chicken products.

    PubMed

    Charlermroj, Ratthaphol; Makornwattana, Manlika; Grant, Irene R; Elliott, Christopher T; Karoonuthaisiri, Nitsara

    2016-05-01

    This study rigorously evaluated a previously developed immunobead array method to simultaneously detect three important foodborne pathogens, Campylobacter jejuni, Listeria monocytogenes, and Salmonella spp., for its actual application in routine food testing. Due to the limitation of the detection limit of the developed method, an enrichment step was included in this study by using Campylobacter Enrichment Broth for C. jejuni and Universal Pre-enrichment Broth for L. monocytogenes and Salmonella spp.. The findings showed that the immunobead array method was capable of detecting as low as 1CFU of the pathogens spiked in the culture media after being cultured for 24h for all three pathogens. The immunobead array method was further evaluated for its pathogen detection capabilities in ready-to-eat (RTE) and ready-to-cook (RTC) chicken samples and proven to be able to detect as low as 1CFU of the pathogens spiked in the food samples after being cultured for 24h in the case of Salmonella spp., and L. monocytogenes and 48 h in the case of C. jejuni. The method was subsequently validated with three types of chicken products (RTE, n=30; RTC, n=20; raw chicken, n=20) and was found to give the same results as the conventional plating method. Our findings demonstrated that the previously developed immunobead array method could be used for actual food testing with minimal enrichment period of only 52 h, whereas the conventional ISO protocols for the same pathogens take 90-144 h. The immunobead array was therefore an inexpensive, rapid and simple method for the food testing. PMID:26950032

  19. Spatial distribution and risk factors of highly pathogenic avian influenza (HPAI) H5N1 in China

    USGS Publications Warehouse

    Martin, Vincent; Pfeiffer, Dirk U.; Zhou, Xiaoyan; Xiao, Xiangming; Prosser, Diann J.; Guo, Fusheng; Gilbert, Marius

    2011-01-01

    Highly pathogenic avian influenza (HPAI) H5N1 was first encountered in 1996 in Guangdong province (China) and started spreading throughout Asia and the western Palearctic in 2004–2006. Compared to several other countries where the HPAI H5N1 distribution has been studied in some detail, little is known about the environmental correlates of the HPAI H5N1 distribution in China. HPAI H5N1 clinical disease outbreaks, and HPAI virus (HPAIV) H5N1 isolated from active risk-based surveillance sampling of domestic poultry (referred to as HPAIV H5N1 surveillance positives in this manuscript) were modeled separately using seven risk variables: chicken, domestic waterfowl population density, proportion of land covered by rice or surface water, cropping intensity, elevation, and human population density. We used bootstrapped logistic regression and boosted regression trees (BRT) with cross-validation to identify the weight of each variable, to assess the predictive power of the models, and to map the distribution of HPAI H5N1 risk. HPAI H5N1 clinical disease outbreak occurrence in domestic poultry was mainly associated with chicken density, human population density, and elevation. In contrast, HPAIV H5N1 infection identified by risk-based surveillance was associated with domestic waterfowl density, human population density, and the proportion of land covered by surface water. Both models had a high explanatory power (mean AUC ranging from 0.864 to 0.967). The map of HPAIV H5N1 risk distribution based on active surveillance data emphasized areas south of the Yangtze River, while the distribution of reported outbreak risk extended further North, where the density of poultry and humans is higher. We quantified the statistical association between HPAI H5N1 outbreak, HPAIV distribution and post-vaccination levels of seropositivity (percentage of effective post-vaccination seroconversion in vaccinated birds) and found that provinces with either outbreaks or HPAIV H5N1 surveillance

  20. Oseltamivir-Ribavirin Combination Therapy for Highly Pathogenic H5N1 Influenza Virus Infection in Mice▿

    PubMed Central

    Ilyushina, Natalia A.; Hay, Alan; Yilmaz, Neziha; Boon, Adrianus C. M.; Webster, Robert G.; Govorkova, Elena A.

    2008-01-01

    We studied the effects of a neuraminidase inhibitor (oseltamivir) and an inhibitor of influenza virus polymerases (ribavirin) against two highly pathogenic H5N1 influenza viruses. In vitro, A/Vietnam/1203/04 virus (clade 1) was highly susceptible to oseltamivir carboxylate (50% inhibitory concentration [IC50] = 0.3 nM), whereas A/Turkey/15/06 virus (clade 2.2) had reduced susceptibility (IC50 = 5.5 nM). In vivo, BALB/c mice were treated with oseltamivir (1, 10, 50, or 100 mg/kg of body weight/day), ribavirin (37.5, 55, or 75 mg/kg/day), or the combination of both drugs for 8 days, starting 4 h before virus inoculation. Monotherapy produced a dose-dependent antiviral effect against the two H5N1 viruses in vivo. Three-dimensional analysis of the drug-drug interactions revealed that oseltamivir and ribavirin interacted principally in an additive manner, with several exceptions of marginal synergy or marginal antagonism at some concentrations. The combination of ribavirin at 37.5 mg/kg/day and oseltamivir at 1 mg/kg/day and the combination of ribavirin at 37.5 mg/kg/day and oseltamivir at 10 mg/kg/day were synergistic against A/Vietnam/1203/04 and A/Turkey/15/06 viruses, respectively. These optimal oseltamivir-ribavirin combinations significantly inhibited virus replication in mouse organs, prevented the spread of H5N1 viruses beyond the respiratory tract, and abrogated the cytokine response (P < 0.01). Importantly, we observed clear differences between the efficacies of the drug combinations against two H5N1 viruses: higher doses were required for the protection of mice against A/Turkey/15/06 virus than for the protection of mice against A/Vietnam/1203/04 virus. Our preliminary results suggest that oseltamivir-ribavirin combinations can have a greater or lesser antiviral effect than monotherapy, depending on the H5N1 virus and the concentrations used. PMID:18725448

  1. Expansion of variant diversity associated with high prevalence of pathogen strain superinfection under conditions of natural transmission

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Superinfection occurs when a second, genetically distinct strain infects a host that has already mounted an immune response to a primary strain. For antigenically variant pathogens, the primary strain itself expresses a broad diversity of variants over time. Thus successful superinfection would requ...

  2. Characterization of 10 adjuvants for inactivated avian influenza virus (AIV) vaccines against challenge with highly pathogenic AIV in chickens

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Inactivated vaccines comprise 95% of all vaccine used for avian influenza virus (AIV) by dose. Optimizing the adjuvant is one way to improve vaccine efficacy. Inactivated vaccines were produced with beta-propiolactone inactivated A/chicken/BC/314514-1/2004 H7N3 low pathogenicity AIV and standardiz...

  3. A highly multiplexed and sensitive RNA-seq protocol for simultaneous analysis of host and pathogen transcriptomes.

    PubMed

    Avraham, Roi; Haseley, Nathan; Fan, Amy; Bloom-Ackermann, Zohar; Livny, Jonathan; Hung, Deborah T

    2016-08-01

    The ability to simultaneously characterize the bacterial and host expression programs during infection would facilitate a comprehensive understanding of pathogen-host interactions. Although RNA sequencing (RNA-seq) has greatly advanced our ability to study the transcriptomes of prokaryotes and eukaryotes separately, limitations in existing protocols for the generation and analysis of RNA-seq data have hindered simultaneous profiling of host and bacterial pathogen transcripts from the same sample. Here we provide a detailed protocol for simultaneous analysis of host and bacterial transcripts by RNA-seq. Importantly, this protocol details the steps required for efficient host and bacteria lysis, barcoding of samples, technical advances in sample preparation for low-yield sample inputs and a computational pipeline for analysis of both mammalian and microbial reads from mixed host-pathogen RNA-seq data. Sample preparation takes 3 d from cultured cells to pooled libraries. Data analysis takes an additional day. Compared with previous methods, the protocol detailed here provides a sensitive, facile and generalizable approach that is suitable for large-scale studies and will enable the field to obtain in-depth analysis of host-pathogen interactions in infection models. PMID:27442864

  4. Major histocompatibility complex and host background genes in chickens influence resistance to high pathogenicity avian influenza virus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The chicken’s major histocompatibility complex (MHC) haplotype has a profound influence on the resistance or susceptibility to certain pathogens such as B21 MHC haplotype confers resistance to Marek’s disease (MD). However, non-MHC genes are also important in disease resistance. For example, both li...

  5. The contribution of PA-X to the virulence of pandemic 2009 H1N1 and highly pathogenic H5N1 avian influenza viruses.

    PubMed

    Gao, Huijie; Sun, Yipeng; Hu, Jiao; Qi, Lu; Wang, Jinliang; Xiong, Xin; Wang, Yu; He, Qiming; Lin, Yang; Kong, Weili; Seng, Lai-Giea; Sun, Honglei; Pu, Juan; Chang, Kin-Chow; Liu, Xiufan; Liu, Jinhua

    2015-02-05

    PA-X is a novel protein encoded by PA mRNA and is found to decrease the pathogenicity of pandemic 1918 H1N1 virus in mice. However, the importance of PA-X proteins in current epidemiologically important influenza A virus strains is not known. In this study, we report on the pathogenicity and pathological effects of PA-X deficient 2009 pandemic H1N1 (pH1N1) and highly pathogenic avian influenza H5N1 viruses. We found that loss of PA-X expression in pH1N1 and H5N1 viruses increased viral replication and apoptosis in A549 cells and increased virulence and host inflammatory response in mice. In addition, PA-X deficient pH1N1 and H5N1 viruses up-regulated PA mRNA and protein synthesis and increased viral polymerase activity. Loss of PA-X was also accompanied by accelerated nuclear accumulation of PA protein and reduced suppression of PA on non-viral protein expression. Our study highlights the effects of PA-X on the moderation of viral pathogenesis and pathogenicity.

  6. Two-Component Signaling Regulates Osmotic Stress Adaptation via SskA and the High-Osmolarity Glycerol MAPK Pathway in the Human Pathogen Talaromyces marneffei.

    PubMed

    Boyce, Kylie J; Cao, Cunwei; Andrianopoulos, Alex

    2016-01-01

    For successful infection to occur, a pathogen must be able to evade or tolerate the host's defense systems. This requires the pathogen to first recognize the host environment and then signal this response to elicit a complex adaptive program in order to activate its own defense strategies. In both prokaryotes and eukaryotes, two-component signaling systems are utilized to sense and respond to changes in the external environment. The hybrid histidine kinases (HHKs) at the start of the two-component signaling pathway have been well characterized in human pathogens. However, how these HHKs regulate processes downstream currently remains unclear. This study describes the role of a response regulator downstream of these HHKs, sskA, in Talaromyces marneffei, a dimorphic human pathogen. sskA is required for asexual reproduction, hyphal morphogenesis, cell wall integrity, osmotic adaptation, and the morphogenesis of yeast cells both in vitro at 37°C and during macrophage infection, but not during dimorphic switching. Comparison of the ΔsskA mutant with a strain in which the mitogen-activated protein kinase (MAPK) of the high-osmolarity glycerol pathway (SakA) has been deleted suggests that SskA acts upstream of this pathway in T. marneffei to regulate these morphogenetic processes. This was confirmed by assessing the amount of phosphorylated SakA in the ΔsskA mutant, antifungal resistance due to a lack of SakA activation, and the ability of a constitutively active sakA allele (sakA(F316L) ) to suppress the ΔsskA mutant phenotypes. We conclude that SskA regulates morphogenesis and osmotic stress adaptation in T. marneffei via phosphorylation of the SakA MAPK of the high-osmolarity glycerol pathway. IMPORTANCE This is the first study in a dimorphic fungal pathogen to investigate the role of a response regulator downstream of two-component signaling systems and its connection to the high-osmolarity glycerol pathway. This study will inspire further research into the

  7. Immunization with live nonpathogenic H5N3 duck influenza virus protects chickens against highly pathogenic H5N1 virus.

    PubMed

    Gambaryan, A S; Boravleva, E Y; Lomakina, N F; Kropotkina, E A; Gordeychuk, I V; Chvala, I A; Drygin, V V; Klenk, H-D; Matrosovich, M N

    2016-01-01

    Development of an effective, broadly-active and safe vaccine for protection of poultry from H5N1 highly pathogenic avian influenza viruses (HPAIVs) remains an important practical goal. In this study we used a low pathogenic wild aquatic bird virus isolate А/duck/Moscow/4182/2010 (H5N3) (dk/4182) as a live candidate vaccine. We compared this virus with four live 1:7 reassortant anti-H5N1 candidate vaccine viruses with modified hemagglutinin from either A/Vietnam/1203/04 (H5N1) or A/Kurgan/3/05 (H5N1) and the rest of the genes from either H2N2 cold-adapted master strain A/Leningrad/134/17/57 (rVN-Len and rKu-Len) or H6N2 virus A/gull/Moscow/3100/2006 (rVN-gull and rKu-gull). The viruses were tested in parallel for pathogenicity, immunogenicity and protective effectiveness in chickens using aerosol, intranasal and oral routes of immunization. All five viruses showed zero pathogenicity indexes in chickens. Viruses rVN-gull and rKu-gull were immunogenic and protective, but they were insufficiently attenuated and caused significant mortality of 1-day-old chickens. The viruses with cold-adapted backbones (rVN-Len and rKu-Len) were completely nonpathogenic, but they were significantly less immunogenic and provided lower protection against lethal challenge with HPAIV A/Chicken/Kurgan/3/05 (H5N1) as compared with three other vaccine candidates. Unlike other four viruses, dk/4182 was both safe and highly immunogenic in chickens of any age regardless of inoculation route. Single administration of 106 TCID50 of dk/4182 virus via drinking water provided complete protection of 30-days-old chickens from 100 LD50 of the challenge virus. Our results suggest that low pathogenic viruses of wild aquatic birds can be used as safe and effective live poultry vaccines against highly pathogenic avian viruses. PMID:27640442

  8. Two-Component Signaling Regulates Osmotic Stress Adaptation via SskA and the High-Osmolarity Glycerol MAPK Pathway in the Human Pathogen Talaromyces marneffei

    PubMed Central

    Cao, Cunwei; Andrianopoulos, Alex

    2016-01-01

    ABSTRACT For successful infection to occur, a pathogen must be able to evade or tolerate the host’s defense systems. This requires the pathogen to first recognize the host environment and then signal this response to elicit a complex adaptive program in order to activate its own defense strategies. In both prokaryotes and eukaryotes, two-component signaling systems are utilized to sense and respond to changes in the external environment. The hybrid histidine kinases (HHKs) at the start of the two-component signaling pathway have been well characterized in human pathogens. However, how these HHKs regulate processes downstream currently remains unclear. This study describes the role of a response regulator downstream of these HHKs, sskA, in Talaromyces marneffei, a dimorphic human pathogen. sskA is required for asexual reproduction, hyphal morphogenesis, cell wall integrity, osmotic adaptation, and the morphogenesis of yeast cells both in vitro at 37°C and during macrophage infection, but not during dimorphic switching. Comparison of the ΔsskA mutant with a strain in which the mitogen-activated protein kinase (MAPK) of the high-osmolarity glycerol pathway (SakA) has been deleted suggests that SskA acts upstream of this pathway in T. marneffei to regulate these morphogenetic processes. This was confirmed by assessing the amount of phosphorylated SakA in the ΔsskA mutant, antifungal resistance due to a lack of SakA activation, and the ability of a constitutively active sakA allele (sakAF316L) to suppress the ΔsskA mutant phenotypes. We conclude that SskA regulates morphogenesis and osmotic stress adaptation in T. marneffei via phosphorylation of the SakA MAPK of the high-osmolarity glycerol pathway. IMPORTANCE This is the first study in a dimorphic fungal pathogen to investigate the role of a response regulator downstream of two-component signaling systems and its connection to the high-osmolarity glycerol pathway. This study will inspire further research into

  9. Proteomic screening of variola virus reveals a unique NF-kappaB inhibitor that is highly conserved among pathogenic orthopoxviruses.

    PubMed

    Mohamed, Mohamed R; Rahman, Masmudur M; Lanchbury, Jerry S; Shattuck, Donna; Neff, Chris; Dufford, Max; van Buuren, Nick; Fagan, Katharine; Barry, Michele; Smith, Scott; Damon, Inger; McFadden, Grant

    2009-06-01

    Identification of the binary interactions between viral and host proteins has become a valuable tool for investigating viral tropism and pathogenesis. Here, we present the first systematic protein interaction screening of the unique variola virus proteome by using yeast 2-hybrid screening against a variety of human cDNA libraries. Several protein-protein interactions were identified, including an interaction between variola G1R, an ankryin/F-box containing protein, and human nuclear factor kappa-B1 (NF-kappaB1)/p105. This represents the first direct interaction between a pathogen-encoded protein and NF-kappaB1/p105. Orthologs of G1R are present in a variety of pathogenic orthopoxviruses, but not in vaccinia virus, and expression of any one of these viral proteins blocks NF-kappaB signaling in human cells. Thus, proteomic screening of variola virus has the potential to uncover modulators of the human innate antiviral responses.

  10. Bioterrorism: pathogens as weapons.

    PubMed

    Anderson, Peter D; Bokor, Gyula

    2012-10-01

    Biowarfare has been used for centuries. The use of biological weapons in terrorism remains a threat. Biological weapons include infectious agents (pathogens) and toxins. The most devastating bioterrorism scenario would be the airborne dispersal of pathogens over a concentrated population area. Characteristics that make a specific pathogen a high-risk for bioterrorism include a low infective dose, ability to be aerosolized, high contagiousness, and survival in a variety of environmental conditions. The most dangerous potential bioterrorism agents include the microorganisms that produce anthrax, plague, tularemia, and smallpox. Other diseases of interest to bioterrorism include brucellosis, glanders, melioidosis, Q fever, and viral encephalitis. Food safety and water safety threats are another area of concern.

  11. Bioterrorism: pathogens as weapons.

    PubMed

    Anderson, Peter D; Bokor, Gyula

    2012-10-01

    Biowarfare has been used for centuries. The use of biological weapons in terrorism remains a threat. Biological weapons include infectious agents (pathogens) and toxins. The most devastating bioterrorism scenario would be the airborne dispersal of pathogens over a concentrated population area. Characteristics that make a specific pathogen a high-risk for bioterrorism include a low infective dose, ability to be aerosolized, high contagiousness, and survival in a variety of environmental conditions. The most dangerous potential bioterrorism agents include the microorganisms that produce anthrax, plague, tularemia, and smallpox. Other diseases of interest to bioterrorism include brucellosis, glanders, melioidosis, Q fever, and viral encephalitis. Food safety and water safety threats are another area of concern. PMID:23011963

  12. Australian human and parrot Chlamydia psittaci strains cluster within the highly virulent 6BC clade of this important zoonotic pathogen.

    PubMed

    Branley, James; Bachmann, Nathan L; Jelocnik, Martina; Myers, Garry S A; Polkinghorne, Adam

    2016-08-04

    Chlamydia psittaci is an avian pathogen and zoonotic agent of atypical pneumonia. The most pathogenic C. psittaci strains cluster into the 6BC clade, predicted to have recently emerged globally. Exposure to infected parrots is a risk factor with limited evidence also of an indirect exposure risk. Genome sequencing was performed on six Australian human and a single avian C. psittaci strain isolated over a 9 year period. Only one of the five human patients had explicit psittacine contact. Genomics analyses revealed that the Australian C. psittaci strains are remarkably similar, clustering tightly within the C. psittaci 6BC clade suggested to have been disseminated by South America parrot importation. Molecular clock analysis using the newly sequenced C. psittaci genomes predicted the emergence of the 6BC clade occurring approximately 2,000 years ago. These findings reveal the potential for an Australian natural reservoir of C. psittaci 6BC strains. These strains can also be isolated from seriously ill patients without explicit psittacine contact. The apparent recent and global spread of C. psittaci 6BC strains raises important questions over how this happened. Further studies may reveal whether the dissemination of this important zoonotic pathogen is linked to Australian parrot importation rather than parrots from elsewhere.

  13. Australian human and parrot Chlamydia psittaci strains cluster within the highly virulent 6BC clade of this important zoonotic pathogen

    PubMed Central

    Branley, James; Bachmann, Nathan L.; Jelocnik, Martina; Myers, Garry S. A.; Polkinghorne, Adam

    2016-01-01

    Chlamydia psittaci is an avian pathogen and zoonotic agent of atypical pneumonia. The most pathogenic C. psittaci strains cluster into the 6BC clade, predicted to have recently emerged globally. Exposure to infected parrots is a risk factor with limited evidence also of an indirect exposure risk. Genome sequencing was performed on six Australian human and a single avian C. psittaci strain isolated over a 9 year period. Only one of the five human patients had explicit psittacine contact. Genomics analyses revealed that the Australian C. psittaci strains are remarkably similar, clustering tightly within the C. psittaci 6BC clade suggested to have been disseminated by South America parrot importation. Molecular clock analysis using the newly sequenced C. psittaci genomes predicted the emergence of the 6BC clade occurring approximately 2,000 years ago. These findings reveal the potential for an Australian natural reservoir of C. psittaci 6BC strains. These strains can also be isolated from seriously ill patients without explicit psittacine contact. The apparent recent and global spread of C. psittaci 6BC strains raises important questions over how this happened. Further studies may reveal whether the dissemination of this important zoonotic pathogen is linked to Australian parrot importation rather than parrots from elsewhere. PMID:27488134

  14. Australian human and parrot Chlamydia psittaci strains cluster within the highly virulent 6BC clade of this important zoonotic pathogen.

    PubMed

    Branley, James; Bachmann, Nathan L; Jelocnik, Martina; Myers, Garry S A; Polkinghorne, Adam

    2016-01-01

    Chlamydia psittaci is an avian pathogen and zoonotic agent of atypical pneumonia. The most pathogenic C. psittaci strains cluster into the 6BC clade, predicted to have recently emerged globally. Exposure to infected parrots is a risk factor with limited evidence also of an indirect exposure risk. Genome sequencing was performed on six Australian human and a single avian C. psittaci strain isolated over a 9 year period. Only one of the five human patients had explicit psittacine contact. Genomics analyses revealed that the Australian C. psittaci strains are remarkably similar, clustering tightly within the C. psittaci 6BC clade suggested to have been disseminated by South America parrot importation. Molecular clock analysis using the newly sequenced C. psittaci genomes predicted the emergence of the 6BC clade occurring approximately 2,000 years ago. These findings reveal the potential for an Australian natural reservoir of C. psittaci 6BC strains. These strains can also be isolated from seriously ill patients without explicit psittacine contact. The apparent recent and global spread of C. psittaci 6BC strains raises important questions over how this happened. Further studies may reveal whether the dissemination of this important zoonotic pathogen is linked to Australian parrot importation rather than parrots from elsewhere. PMID:27488134

  15. Xylella Genomics and Bacterial Pathogenicity to Plants

    PubMed Central

    Dow, J. M.

    2000-01-01

    Xylella fastidiosa, a pathogen of citrus, is the first plant pathogenic bacterium for which the complete genome sequence has been published. Inspection of the sequence reveals high relatedness to many genes of other pathogens, notably Xanthomonas campestris. Based on this, we suggest that Xylella possesses certain easily testable properties that contribute to pathogenicity. We also present some general considerations for deriving information on pathogenicity from bacterial genomics. PMID:11119303

  16. Genetic analysis of an H5N2 highly pathogenic avian influenza virus isolated from a chicken in a live bird market in Northern Vietnam in 2012.

    PubMed

    Nishi, Tatsuya; Okamatsu, Masatoshi; Sakurai, Kenji; Chu, Huy Duc; Thanh, Long Pham; van Nguyen, Long; van Hoang, Nam; Thi, Diep Nguyen; Sakoda, Yoshihiro; Kida, Hiroshi

    2014-01-01

    In August 2012, A/chicken/Vietnam/OIE-2215/2012 (H5N2) was isolated from a chicken in a live bird market (LBM) in Northern Vietnam. Intravenous pathogenicity test revealed that this virus is highly pathogenic in chickens. The PA, HA, NP and M, PB2 and NA, and PB1 and NS genes of the isolate were phylogenetically closely related to those of A/duck/Vietnam/OIE-2202/2012 (H5N1) of clade 2.3.2.1, A/chicken/Vietnam/OIE-1611/2012 (H9N2) and A/chicken/Vietnam/OIE-2468/2012 (H9N2), respectively. All of these viruses were isolated from birds in LBMs in the same province. These results indicate that A/chicken/Vietnam/OIE-2215/2012 (H5N2) is a genetic reassortant and that surveillance of avian influenza in LBMs and stamping out policy are essential for the eradication of highly pathogenic avian influenza viruses from Asia.

  17. Pathogenesis and transmission of H7 and H5 highly pathogenic avian influenza viruses in mallards including the recent intercontinental H5 viruses (H5N8 and H5N2)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Highly pathogenic avian influenza viruses (HPAIV’s) remain a threat to poultry worldwide. Avian influenza viruses, including HPAIV, are usually non-pathogenic for ducks and other wild aquatic birds, with the exception of Asian lineage H5N1, and recently H5N8, HPAIVs, which can cause moderate to sev...

  18. High content image-based screening of a protease inhibitor library reveals compounds broadly active against Rift Valley fever virus and other highly pathogenic RNA viruses.

    PubMed

    Mudhasani, Rajini; Kota, Krishna P; Retterer, Cary; Tran, Julie P; Whitehouse, Chris A; Bavari, Sina

    2014-08-01

    High content image-based screening was developed as an approach to test a protease inhibitor small molecule library for antiviral activity against Rift Valley fever virus (RVFV) and to determine their mechanism of action. RVFV is the causative agent of severe disease of humans and animals throughout Africa and the Arabian Peninsula. Of the 849 compounds screened, 34 compounds exhibited ≥ 50% inhibition against RVFV. All of the hit compounds could be classified into 4 distinct groups based on their unique chemical backbone. Some of the compounds also showed broad antiviral activity against several highly pathogenic RNA viruses including Ebola, Marburg, Venezuela equine encephalitis, and Lassa viruses. Four hit compounds (C795-0925, D011-2120, F694-1532 and G202-0362), which were most active against RVFV and showed broad-spectrum antiviral activity, were selected for further evaluation for their cytotoxicity, dose response profile, and mode of action using classical virological methods and high-content imaging analysis. Time-of-addition assays in RVFV infections suggested that D011-2120 and G202-0362 targeted virus egress, while C795-0925 and F694-1532 inhibited virus replication. We showed that D011-2120 exhibited its antiviral effects by blocking microtubule polymerization, thereby disrupting the Golgi complex and inhibiting viral trafficking to the plasma membrane during virus egress. While G202-0362 also affected virus egress, it appears to do so by a different mechanism, namely by blocking virus budding from the trans Golgi. F694-1532 inhibited viral replication, but also appeared to inhibit overall cellular gene expression. However, G202-0362 and C795-0925 did not alter any of the morphological features that we examined and thus may prove to be good candidates for antiviral drug development. Overall this work demonstrates that high-content image analysis can be used to screen chemical libraries for new antivirals and to determine their mechanism of action and

  19. Host Specificity of Bacterial Pathogens

    PubMed Central

    Bäumler, Andreas; Fang, Ferric C.

    2013-01-01

    Most pathogens are able to infect multiple hosts but some are highly adapted to a single-host species. A detailed understanding of the basis of host specificity can provide important insights into molecular pathogenesis, the evolution of pathogenic microbes, and the potential for pathogens to cross the species barrier to infect new hosts. Comparative genomics and the development of humanized mouse models have provided important new tools with which to explore the basis of generalism and specialism. This review will examine host specificity of bacterial pathogens with a focus on generalist and specialist serovars of Salmonella enterica. PMID:24296346

  20. Range-wide genetic population structure of common pochard (Aythya ferina): a potentially important vector of highly pathogenic avian influenza viruses

    PubMed Central

    Liu, Yang; Keller, Irene; Heckel, Gerald

    2011-01-01

    An understanding of the distribution and spatial structure of the natural vectors of zoonothic pathogens is of interest for effective disease control and prevention. Here, we investigate the range-wide population genetic structure of common pochard (Aythya ferina), a long-distance migratory duck and potential vector of highly pathogenic avian influenza. We collected several hundred samples from breeding and wintering grounds across Eurasia including some H5N1-positive individuals and generated partial sequences of the mitochondrial control region and multilocus microsatellite genotypes. Genetic differentiation among breeding populations was significant for both marker types but higher for maternally inherited mtDNA than for biparentally inherited nuclear markers. There was only weak genetic divergence between ducks sampled in Europe and East Asia, and genetic differentiation between populations was not generally associated with geographical distance. No evidence of genetic substructure was detected for ducks sampled on the European wintering grounds. Our results suggest limited breeding-site fidelity, especially in females, but extensive population admixture on the wintering grounds. The specific role of pochards as natural vectors of zoonotic pathogens and in particular H5N1 remains to be clarified but our results point to wintering grounds as potential hotspots for disease transmission. PMID:22393520

  1. Range-wide genetic population structure of common pochard (Aythya ferina): a potentially important vector of highly pathogenic avian influenza viruses.

    PubMed

    Liu, Yang; Keller, Irene; Heckel, Gerald

    2011-12-01

    An understanding of the distribution and spatial structure of the natural vectors of zoonothic pathogens is of interest for effective disease control and prevention. Here, we investigate the range-wide population genetic structure of common pochard (Aythya ferina), a long-distance migratory duck and potential vector of highly pathogenic avian influenza. We collected several hundred samples from breeding and wintering grounds across Eurasia including some H5N1-positive individuals and generated partial sequences of the mitochondrial control region and multilocus microsatellite genotypes. Genetic differentiation among breeding populations was significant for both marker types but higher for maternally inherited mtDNA than for biparentally inherited nuclear markers. There was only weak genetic divergence between ducks sampled in Europe and East Asia, and genetic differentiation between populations was not generally associated with geographical distance. No evidence of genetic substructure was detected for ducks sampled on the European wintering grounds. Our results suggest limited breeding-site fidelity, especially in females, but extensive population admixture on the wintering grounds. The specific role of pochards as natural vectors of zoonotic pathogens and in particular H5N1 remains to be clarified but our results point to wintering grounds as potential hotspots for disease transmission. PMID:22393520

  2. Isolation and identification of highly pathogenic avian influenza H5N1 virus from Houbara bustards (Chlamydotis undulata macqueenii) and contact falcons.

    PubMed

    Khan, Owais Ahmed; Shuaib, Mohammad Adam; Rhman, Salah Shaban Abdel; Ismail, Mahmoud Moussa; Hammad, Yousef Al; Baky, Mansour Hashim Abdel; Fusaro, Alice; Salviato, Annalisa; Cattoli, Giovanni

    2009-02-01

    Highly pathogenic influenza virus (HPAIV) H5N1 has caused mortality and morbidity in many species of domestic and wild bird. The Houbara bustard (Chlamydotis undulata macqueenii) is a solitary bird that inhabits semi-desert regions. It is known to be susceptible to avianpox, avian paramyxovirus type 1, and low-pathogenicity avian influenza H9N2. We report an outbreak of H5N1 HPAIV in Houbara bustards, which were introduced into the Kingdom of Saudi Arabia for falconry purposes. Ninety-three per cent mortality (38 out of 41 birds) in the infected Houbara bustard flock and about 62.5% mortality (10 out of 16 birds) in falcons that came in contact with these birds were observed. Pooled cloacal and tracheal swabs from Houbara bustards as well as visceral organ homogenates collected in Houbara bustards and falcons were tested by real-time reverse transcriptase-polymerase chain reaction, and virus isolation was attempted in specific pathogen free hens' eggs. The viruses isolated were characterized as HPAIV H5N1. Phylogenetic analysis of the haemagglutinating and Neuraminidase (NA) genes revealed that the viruses isolated from Houbara bustards and falcons were closely related to each other and to Kuwaiti H5N1 strains isolated in 2007. Interestingly, they were genetically distinguishable from the co-circulating A/H5N1 viruses in Kingdom of Saudi Arabia causing outbreaks in domestic birds. This case emphasizes the need for surveillance of this endangered species in its natural habitat.

  3. Cationic polyelectrolyte functionalized magnetic particles assisted highly sensitive pathogens detection in combination with polymerase chain reaction and capillary electrophoresis.

    PubMed

    Chen, Jia; Lin, Yuexin; Wang, Yu; Jia, Li

    2015-06-01

    Pathogenic bacteria cause significant morbidity and mortality to humans. There is a pressing need to establish a simple and reliable method to detect them. Herein, we show that magnetic particles (MPs) can be functionalized by poly(diallyl dimethylammonium chloride) (PDDA), and the particles (PDDA-MPs) can be utilized as adsorbents for capture of pathogenic bacteria from aqueous solution based on electrostatic interaction. The as-prepared PDDA-MPs were characterized by Fourier-transform infrared spectroscopy, zeta potential, vibrating sample magnetometry, X-ray diffraction spectrometry, scanning electron microscopy, and transmission electron microscopy. The adsorption equilibrium time can be achieved in 3min. According to the Langmuir adsorption isotherm, the maximum adsorption capacities for E. coli O157:H7 (Gram-negative bacteria) and L. monocytogenes (Gram-positive bacteria) were calculated to be 1.8×10(9) and 3.1×10(9)cfumg(-1), respectively. The bacteria in spiked mineral water (1000mL) can be completely captured when applying 50mg of PDDA-MPs and an adsorption time of 5min. In addition, PDDA-MPs-based magnetic separation method in combination with polymerase chain reaction and capillary electrophoresis allows for rapid detection of 10(1)cfumL(-1) bacteria.

  4. Legionella pneumophila: The Paradox of a Highly Sensitive Opportunistic Waterborne Pathogen Able to Persist in the Environment

    PubMed Central

    Berjeaud, Jean-Marc; Chevalier, Sylvie; Schlusselhuber, Margot; Portier, Emilie; Loiseau, Clémence; Aucher, Willy; Lesouhaitier, Olivier; Verdon, Julien

    2016-01-01

    Legionella pneumophila, the major causative agent of Legionnaires’ disease, is found in freshwater environments in close association with free-living amoebae and multispecies biofilms, leading to persistence, spread, biocide resistance, and elevated virulence of the bacterium. Indeed, legionellosis outbreaks are mainly due to the ability of this bacterium to colonize and persist in water facilities, despite harsh physical and chemical treatments. However, these treatments are not totally efficient and, after a lag period, L. pneumophila may be able to quickly re-colonize these systems. Several natural compounds (biosurfactants, antimicrobial peptides…) with anti-Legionella properties have recently been described in the literature, highlighting their specific activities against this pathogen. In this review, we first consider this hallmark of Legionella to resist killing, in regard to its biofilm or host-associated life style. Then, we focus more accurately on natural anti-Legionella molecules described so far, which could provide new eco-friendly and alternative ways to struggle against this important pathogen in plumbing. PMID:27092135

  5. High-resolution transcript profiling of the atypical biotrophic interaction between Theobroma cacao and the fungal pathogen Moniliophthora perniciosa.

    PubMed

    Teixeira, Paulo José Pereira Lima; Thomazella, Daniela Paula de Toledo; Reis, Osvaldo; do Prado, Paula Favoretti Vital; do Rio, Maria Carolina Scatolin; Fiorin, Gabriel Lorencini; José, Juliana; Costa, Gustavo Gilson Lacerda; Negri, Victor Augusti; Mondego, Jorge Maurício Costa; Mieczkowski, Piotr; Pereira, Gonçalo Amarante Guimarães

    2014-11-01

    Witches' broom disease (WBD), caused by the hemibiotrophic fungus Moniliophthora perniciosa, is one of the most devastating diseases of Theobroma cacao, the chocolate tree. In contrast to other hemibiotrophic interactions, the WBD biotrophic stage lasts for months and is responsible for the most distinctive symptoms of the disease, which comprise drastic morphological changes in the infected shoots. Here, we used the dual RNA-seq approach to simultaneously assess the transcriptomes of cacao and M. perniciosa during their peculiar biotrophic interaction. Infection with M. perniciosa triggers massive metabolic reprogramming in the diseased tissues. Although apparently vigorous, the infected shoots are energetically expensive structures characterized by the induction of ineffective defense responses and by a clear carbon deprivation signature. Remarkably, the infection culminates in the establishment of a senescence process in the host, which signals the end of the WBD biotrophic stage. We analyzed the pathogen's transcriptome in unprecedented detail and thereby characterized the fungal nutritional and infection strategies during WBD and identified putative virulence effectors. Interestingly, M. perniciosa biotrophic mycelia develop as long-term parasites that orchestrate changes in plant metabolism to increase the availability of soluble nutrients before plant death. Collectively, our results provide unique insight into an intriguing tropical disease and advance our understanding of the development of (hemi)biotrophic plant-pathogen interactions.

  6. High-resolution transcript profiling of the atypical biotrophic interaction between Theobroma cacao and the fungal pathogen Moniliophthora perniciosa.

    PubMed

    Teixeira, Paulo José Pereira Lima; Thomazella, Daniela Paula de Toledo; Reis, Osvaldo; do Prado, Paula Favoretti Vital; do Rio, Maria Carolina Scatolin; Fiorin, Gabriel Lorencini; José, Juliana; Costa, Gustavo Gilson Lacerda; Negri, Victor Augusti; Mondego, Jorge Maurício Costa; Mieczkowski, Piotr; Pereira, Gonçalo Amarante Guimarães

    2014-11-01

    Witches' broom disease (WBD), caused by the hemibiotrophic fungus Moniliophthora perniciosa, is one of the most devastating diseases of Theobroma cacao, the chocolate tree. In contrast to other hemibiotrophic interactions, the WBD biotrophic stage lasts for months and is responsible for the most distinctive symptoms of the disease, which comprise drastic morphological changes in the infected shoots. Here, we used the dual RNA-seq approach to simultaneously assess the transcriptomes of cacao and M. perniciosa during their peculiar biotrophic interaction. Infection with M. perniciosa triggers massive metabolic reprogramming in the diseased tissues. Although apparently vigorous, the infected shoots are energetically expensive structures characterized by the induction of ineffective defense responses and by a clear carbon deprivation signature. Remarkably, the infection culminates in the establishment of a senescence process in the host, which signals the end of the WBD biotrophic stage. We analyzed the pathogen's transcriptome in unprecedented detail and thereby characterized the fungal nutritional and infection strategies during WBD and identified putative virulence effectors. Interestingly, M. perniciosa biotrophic mycelia develop as long-term parasites that orchestrate changes in plant metabolism to increase the availability of soluble nutrients before plant death. Collectively, our results provide unique insight into an intriguing tropical disease and advance our understanding of the development of (hemi)biotrophic plant-pathogen interactions. PMID:25371547

  7. UMD-Predictor: A High-Throughput Sequencing Compliant System for Pathogenicity Prediction of any Human cDNA Substitution.

    PubMed

    Salgado, David; Desvignes, Jean-Pierre; Rai, Ghadi; Blanchard, Arnaud; Miltgen, Morgane; Pinard, Amélie; Lévy, Nicolas; Collod-Béroud, Gwenaëlle; Béroud, Christophe

    2016-05-01

    Whole-exome sequencing (WES) is increasingly applied to research and clinical diagnosis of human diseases. It typically results in large amounts of genetic variations. Depending on the mode of inheritance, only one or two correspond to pathogenic mutations responsible for the disease and present in affected individuals. Therefore, it is crucial to filter out nonpathogenic variants and limit downstream analysis to a handful of candidate mutations. We have developed a new computational combinatorial system UMD-Predictor (http://umd-predictor.eu) to efficiently annotate cDNA substitutions of all human transcripts for their potential pathogenicity. It combines biochemical properties, impact on splicing signals, localization in protein domains, variation frequency in the global population, and conservation through the BLOSUM62 global substitution matrix and a protein-specific conservation among 100 species. We compared its accuracy with the seven most used and reliable prediction tools, using the largest reference variation datasets including more than 140,000 annotated variations. This system consistently demonstrated a better accuracy, specificity, Matthews correlation coefficient, diagnostic odds ratio, speed, and provided the shortest list of candidate mutations for WES. Webservices allow its implementation in any bioinformatics pipeline for next-generation sequencing analysis. It could benefit to a wide range of users and applications varying from gene discovery to clinical diagnosis. PMID:26842889

  8. Naegleria fowleri: a free-living highly pathogenic amoeba contains trypanothione/trypanothione reductase and glutathione/glutathione reductase systems.

    PubMed

    Ondarza, Raúl N; Hurtado, Gerardo; Tamayo, Elsa; Iturbe, Angélica; Hernández, Eva

    2006-11-01

    This paper presents definitive data showing that the thiol-bimane compound isolated and purified by HPLC from Naegleria fowleri trophozoites unequivocally corresponds by matrix assisted laser-desorption ionization-time-of-flight MS, to the characteristic monoprotonated ion of trypanothione-(bimane)(2) [M(+)H(+)] of m/z 1104.57 and to the trypanothione-(bimane) of m/z 914.46. The trypanothione disulfide T(S)(2) was also found to have a molecular ion of m/z 723.37. Additionally HPLC demonstrated that thiol-bimane compounds corresponding to cysteine and glutathione were present in Naegleria. The ion patterns of the thiol-bimane compounds prepared from commercial trypanothione standard, Entamoeba histolytica and Crithidia luciliae are identical to the Naegleria thiol-bimane compound. Partially purified extracts from N. fowleri showed the coexistence of glutathione and trypanothione reductases activities. There is not doubt that the thiol compound trypanothione, which was previously thought to occur only in Kinetoplastida, is also present in the human pathogens E. histolytica and N. fowleri, as well as in the non-pathogenic euglenozoan E. gracilis. The presence of the trypanothione/trypanothione reductase system in N. fowleri creates the possibility of using this enzyme as a new "drug target" for rationally designed drugs to eliminate the parasite, without affecting the human host.

  9. Proteomic screening of variola virus reveals a unique NF-κB inhibitor that is highly conserved among pathogenic orthopoxviruses

    PubMed Central

    Mohamed, Mohamed R.; Rahman, Masmudur M.; Lanchbury, Jerry S.; Shattuck, Donna; Neff, Chris; Dufford, Max; van Buuren, Nick; Fagan, Katharine; Barry, Michele; Smith, Scott; Damon, Inger; McFadden, Grant

    2009-01-01

    Identification of the binary interactions between viral and host proteins has become a valuable tool for investigating viral tropism and pathogenesis. Here, we present the first systematic protein interaction screening of the unique variola virus proteome by using yeast 2-hybrid screening against a variety of human cDNA libraries. Several protein–protein interactions were identified, including an interaction between variola G1R, an ankryin/F-box containing protein, and human nuclear factor kappa-B1 (NF-κB1)/p105. This represents the first direct interaction between a pathogen-encoded protein and NF-κB1/p105. Orthologs of G1R are present in a variety of pathogenic orthopoxviruses, but not in vaccinia virus, and expression of any one of these viral proteins blocks NF-κB signaling in human cells. Thus, proteomic screening of variola virus has the potential to uncover modulators of the human innate antiviral responses. PMID:19451633

  10. Highly Pathogenic Avian Influenza H5N8 Clade 2.3.4.4 Virus: Equivocal Pathogenicity and Implications for Surveillance Following Natural Infection in Breeder Ducks in the United Kingdom.

    PubMed

    Núñez, A; Brookes, S M; Reid, S M; Garcia-Rueda, C; Hicks, D J; Seekings, J M; Spencer, Y I; Brown, I H

    2016-02-01

    Since early 2014, several outbreaks involving novel reassortant highly pathogenic avian influenza (HPAI) A(H5N8) viruses have been detected in poultry and wild bird species in Asia, Europe and North America. These viruses have been detected in apparently healthy and dead wild migratory birds, as well as in domestic chickens, turkeys, geese and ducks. In this study, we describe the pathology of an outbreak of H5N8 HPAIV in breeder ducks in the UK. A holding with approximately 6000 breeder ducks, aged approximately 60 weeks, showed a gradual reduction in egg production and increased mortality over a 7-day period. Post-mortem examination revealed frequent fibrinous peritonitis, with severely haemorrhagic ovarian follicles and occasional splenic and pancreatic necrosis and high incidence of mycotic granulomas in the air sacs and lung. Low-to-moderate levels of HPAI H5N8 virus were detected mainly in respiratory and digestive tract, with minor involvement of other organs. Although histopathological examination confirmed the gross pathology findings, intralesional viral antigen detection by immunohistochemistry was not observed. Immunolabelled cells were rarely only present in inflamed air sacs and serosa, usually superficial to granulomatous inflammation. Abundant bacterial microcolonies were observed in haemorrhagic ovaries and oviduct. The limited viral tissue distribution and presence of inter-current fungal and bacterial infections suggest a minor role for HPAIV H5N8 in clinical disease in layer ducks. PMID:26519234

  11. Import risk assessment incorporating a dose-response model: introduction of highly pathogenic porcine reproductive and respiratory syndrome into Australia via illegally imported raw pork.

    PubMed

    Brookes, V J; Hernández-Jover, M; Holyoake, P; Ward, M P

    2014-03-01

    Highly pathogenic porcine reproductive and respiratory syndrome (PRRS) has spread through parts of south-east Asia, posing a risk to Australia. The objective of this study was to assess the probability of infection of a feral or domestic pig in Australia with highly pathogenic PRRS following ingestion of illegally imported raw pork. A conservative scenario was considered in which 500 g of raw pork was imported from the Philippines into Australia without being detected by border security, then discarded from a household and potentially accessed by a pig. Monte Carlo simulation of a two-dimensional, stochastic model was used to estimate the probability of entry and exposure, and the probability of infection was assessed by incorporating a virus-decay and mechanistic dose-response model. Results indicated that the probability of infection of a feral pig after ingestion of raw meat was higher than the probability of infection of a domestic pig. Sensitivity analysis was used to assess the influence of input parameters on model output probability estimates, and extension of the virus-decay and dose-response model was used to explore the impact of different temperatures and time from slaughter to ingestion of the meat, different weights of meat, and the level of viraemia at slaughter on the infectivity of meat. Parameters with the highest influence on the model output were the level of viraemia of a pig prior to slaughter and the probability of access by a feral pig to food-waste discarded on property surrounding a household. Extension of the decay and dose-response model showed that small pieces of meat (10 g) from a highly pathogenic PRRS viraemic pig could contain enough virus to have a high probability of infection of a pig, and that routes to Australia by sea or air from all highly pathogenic PRRS virus endemic countries were of interest dependent on the temperature of the raw meat during transport. This study highlighted the importance of mitigation strategies such

  12. Viral replication rate regulates clinical outcome and CD8 T cell responses during highly pathogenic H5N1 influenza virus infection in mice.

    PubMed

    Hatta, Yasuko; Hershberger, Karen; Shinya, Kyoko; Proll, Sean C; Dubielzig, Richard R; Hatta, Masato; Katze, Michael G; Kawaoka, Yoshihiro; Suresh, M

    2010-01-01

    Since the first recorded infection of humans with H5N1 viruses of avian origin in 1997, sporadic human infections continue to occur with a staggering mortality rate of >60%. Although sustained human-to-human transmission has not occurred yet, there is a growing concern that these H5N1 viruses might acquire this trait and raise the specter of a pandemic. Despite progress in deciphering viral determinants of pathogenicity, we still lack crucial information on virus/immune system interactions pertaining to severe disease and high mortality associated with human H5N1 influenza virus infections. Using two human isolates of H5N1 viruses that differ in their pathogenicity in mice, we have defined mechanistic links among the rate of viral replication, mortality, CD8 T cell responses, and immunopathology. The extreme pathogenicity of H5N1 viruses was directly linked to the ability of the virus to replicate rapidly, and swiftly attain high steady-state titers in the lungs within 48 hours after infection. The remarkably high replication rate of the highly pathogenic H5N1 virus did not prevent the induction of IFN-β or activation of CD8 T cells, but the CD8 T cell response was ineffective in controlling viral replication in the lungs and CD8 T cell deficiency did not affect viral titers or mortality. Additionally, BIM deficiency ameliorated lung pathology and inhibited T cell apoptosis without affecting survival of mice. Therefore, rapidly replicating, highly lethal H5N1 viruses could simply outpace and overwhelm the adaptive immune responses, and kill the host by direct cytopathic effects. However, therapeutic suppression of early viral replication and the associated enhancement of CD8 T cell responses improved the survival of mice following a lethal H5N1 infection. These findings suggest that suppression of early H5N1 virus replication is key to the programming of an effective host response, which has implications in treatment of this infection in humans. PMID:20949022

  13. The High-Pathogenicity Island of Yersinia enterocolitica Ye8081 Undergoes Low-Frequency Deletion but Not Precise Excision, Suggesting Recent Stabilization in the Genome

    PubMed Central

    Bach, Sandrine; Buchrieser, Carmen; Prentice, Michael; Guiyoule, Annie; Msadek, Tarek; Carniel, Elisabeth

    1999-01-01

    Highly pathogenic strains of Yersinia pestis, Y. pseudotuberculosis, and Y. enterocolitica are characterized by the possession of a pathogenicity island designated the high-pathogenicity island (HPI). This 35- to 45-kb island carries an iron uptake system named the yersiniabactin locus. While the HPIs of Y. pestis and Y. pseudotuberculosis are subject to high-frequency spontaneous deletion from the chromosome, we were initially unable to obtain HPI-deleted Y. enterocolitica 1B isolates. In the present study, using a positive selection strategy, we identified three HPI-deleted mutants of Y. enterocolitica strain Ye8081. In these three independent clones, the chromosomal deletion was not limited to the HPI but encompassed a larger DNA fragment of approximately 140 kb. Loss of this fragment, which occurred at a frequency of approximately 5 × 10−7, resulted in the disappearance of several phenotypic traits, such as growth in a minimal medium, hydrolysis of o-nitrophenyl-β-d-thiogalactopyranoside, Tween esterase activity, and motility, and in a decreased virulence for mice. However, no precise excision of the Ye8081 HPI was observed. To gain more insight into the molecular basis for this phenomenon, the putative machinery of HPI excision in Y. enterocolitica was analyzed and compared to that in Y. pseudotuberculosis. We show that the probable reasons for failure of precise excision of the HPI of Y. enterocolitica Ye8081 are (i) the interruption of the P4-like integrase gene located close to its right-hand boundary by a premature stop codon and (ii) lack of conservation of 17-bp att-like sequences at both extremities of the HPI. These mutations may represent a process of HPI stabilization in the species Y. enterocolitica. PMID:10496882

  14. High quality permanent draft genome sequence of Phaseolibacter flectens ATCC 12775T, a plant pathogen of French bean pods

    DOE PAGESBeta

    Aizenberg-Gershtein, Yana; Izhaki, Ido; Lapidus, Alla; Copeland, Alex; Reddy, TBK; Huntemann, Marcel; Pillay, Manoj; Markowitz, Victor; Göker, Markus; Woyke, Tanja; et al

    2016-01-13

    We report that the Phaseolibacter flectens strain ATCC 12775T (Halpern et al., Int J Syst Evol Microbiol 63:268–273, 2013) is a Gram-negative, rod shaped, motile, aerobic, chemoorganotroph bacterium. Ph. flectens is as a plant-pathogenic bacterium on pods of French bean and was first identified by Johnson (1956) as Pseudomonas flectens. After its phylogenetic position was reexamined, Pseudomonas flectens was transferred to the family Enterobacteriaceae as Phaseolibacter flectens gen. nov., comb. nov. Here we describe the features of this organism, together with the draft genome sequence and annotation. The DNA GC content is 44.34 mol%. The chromosome length is 2,748,442 bp.more » It encodes 2,437 proteins and 89 RNA genes. Ph. flectens genome is part of the Genomic Encyclopedia of Type Strains, Phase I: the one thousand microbial genomes study.« less

  15. Comparing introduction to Europe of highly pathogenic avian influenza viruses A(H5N8) in 2014 and A(H5N1) in 2005.

    PubMed

    Adlhoch, C; Gossner, C; Koch, G; Brown, I; Bouwstra, R; Verdonck, F; Penttinen, P; Harder, T

    2014-01-01

    Since the beginning of November 2014, nine outbreaks of highly pathogenic avian influenza virus (HPAIV) A(H5N8) in poultry have been detected in four European countries. In this report, similarities and differences between the modes of introduction of HPAIV A(H5N1) and A(H5N8) into Europe are described. Experiences from outbreaks of A(H5N1) in Europe demonstrated that early detection to control HPAIV in poultry has proven pivotal to minimise the risk of zoonotic transmission and prevention of human cases.

  16. Genetic diversity of highly pathogenic H5N8 avian influenza viruses at a single overwintering site of migratory birds in Japan, 2014/15.

    PubMed

    Ozawa, M; Matsuu, A; Tokorozaki, K; Horie, M; Masatani, T; Nakagawa, H; Okuya, K; Kawabata, T; Toda, S

    2015-01-01

    We isolated eight highly pathogenic H5N8 avian influenza viruses (H5N8 HPAIVs) in the 2014/15 winter season at an overwintering site of migratory birds in Japan. Genetic analyses revealed that these isolates were divided into three groups, indicating the co-circulation of three genetic groups of H5N8 HPAIV among these migratory birds. These results also imply the possibility of global redistribution of the H5N8 HPAIVs via the migration of these birds next winter.

  17. High gene flow and outcrossing within populations of two cryptic fungal pathogens on a native and non-native host in Cameroon.

    PubMed

    Begoude Boyogueno, Aime Didier; Slippers, Bernard; Perez, Guillermo; Wingfield, Michael J; Roux, Jolanda

    2012-03-01

    In this study, we determined the genetic diversity of 126 isolates representing both Lasiodiplodia theobromae and Lasiodiplodia pseudotheobromae, collected from Theobroma cacao and Terminalia spp. in Cameroon, using simple sequence repeat (SSR) markers. SSR alleles showed clear genetic distinction between L. theobromae and L. pseudotheobromae, supporting their earlier separation as sister species. Both L. theobromae and L. pseudotheobromae populations from Cameroon had high levels of gene diversity, moderate degrees of genotypic diversity, and high levels of gene flow between isolates from T. cacao and Terminalia spp. There was no evidence for geographic substructure in these populations across the region studied, and the SSR alleles were randomly associated in both species, suggesting outcrossing. The significant levels of aggressiveness, evolutionary potential represented by high levels of diversity, outcrossing and gene flow between geographically and host defined populations, identify these fungi as high-risk pathogens for their native and non-native hosts in Cameroon. PMID:22385617

  18. Two highly similar LAEDDTNAQKT and LTDKIGTEI epitopes in G glycoprotein may be useful for effective epitope based vaccine design against pathogenic Henipavirus.

    PubMed

    Parvege, Md Masud; Rahman, Monzilur; Nibir, Yead Morshed; Hossain, Mohammad Shahnoor

    2016-04-01

    Nipah virus and Hendra virus, two members of the genus Henipavirus, are newly emerging zoonotic pathogens which cause acute respiratory illness and severe encephalitis in human. Lack of the effective antiviral therapy endorses the urgency for the development of vaccine against these deadly viruses. In this study, we employed various computational approaches to identify epitopes which has the potential for vaccine development. By analyzing the immune parameters of the conserved sequences of G glycoprotein using various databases and bioinformatics tools, we identified two potential epitopes which may be used as peptide vaccines. Using different B cell epitope prediction servers, four highly similar B cell epitopes were identified. Immunoinformatics analyses revealed that LAEDDTNAQKT is a highly flexible and accessible B-cell epitope to antibody. Highly similar putative CTL epitopes were analyzed for their binding with the HLA-C 12*03 molecule. Docking simulation assay revealed that LTDKIGTEI has significantly lower binding energy, which bolstered its potential as epitope-based vaccine design. Finally, cytotoxicity analysis has also justified their potential as promising epitope-based vaccine candidate. In sum, our computational analysis indicates that either LAEDDTNAQKT or LTDKIGTEI epitope holds a promise for the development of universal vaccine against all kinds of pathogenic Henipavirus. Further in vivo and in vitro studies are necessary to validate the obtained findings. PMID:26970211

  19. Two highly similar LAEDDTNAQKT and LTDKIGTEI epitopes in G glycoprotein may be useful for effective epitope based vaccine design against pathogenic Henipavirus.

    PubMed

    Parvege, Md Masud; Rahman, Monzilur; Nibir, Yead Morshed; Hossain, Mohammad Shahnoor

    2016-04-01

    Nipah virus and Hendra virus, two members of the genus Henipavirus, are newly emerging zoonotic pathogens which cause acute respiratory illness and severe encephalitis in human. Lack of the effective antiviral therapy endorses the urgency for the development of vaccine against these deadly viruses. In this study, we employed various computational approaches to identify epitopes which has the potential for vaccine development. By analyzing the immune parameters of the conserved sequences of G glycoprotein using various databases and bioinformatics tools, we identified two potential epitopes which may be used as peptide vaccines. Using different B cell epitope prediction servers, four highly similar B cell epitopes were identified. Immunoinformatics analyses revealed that LAEDDTNAQKT is a highly flexible and accessible B-cell epitope to antibody. Highly similar putative CTL epitopes were analyzed for their binding with the HLA-C 12*03 molecule. Docking simulation assay revealed that LTDKIGTEI has significantly lower binding energy, which bolstered its potential as epitope-based vaccine design. Finally, cytotoxicity analysis has also justified their potential as promising epitope-based vaccine candidate. In sum, our computational analysis indicates that either LAEDDTNAQKT or LTDKIGTEI epitope holds a promise for the development of universal vaccine against all kinds of pathogenic Henipavirus. Further in vivo and in vitro studies are necessary to validate the obtained findings.

  20. Pathogenesis in Eurasian tree sparrows inoculated with H5N1 highly pathogenic avian influenza virus and experimental virus transmission from tree sparrows to chickens.

    PubMed

    Yamamoto, Yu; Nakamura, Kikuyasu; Yamada, Manabu; Mase, Masaji

    2013-06-01

    Small wild birds that routinely enter poultry farms may be possible vectors of Asian lineage H5N1 highly pathogenic avian influenza virus. In this study, we conducted experimental infections using wild-caught Eurasian tree sparrows (Passer montanus) to evaluate their possible epidemiological involvement in virus transmission. When tree sparrows were intranasally inoculated with the virus at a low or high dose, all sparrows excluding euthanatized birds died within 11 days after inoculation. Viruses were frequently isolated from the drinking water, oral swabs, and visceral organs of the sparrows. Immunohistochemical analysis revealed that the virus replicated strongly in the central nervous system, heart, and adrenal gland following primary infection in the upper respiratory tract and a probable subsequent viremic stage. In the contact infection study using virus-inoculated sparrows and untreated contact chickens, more than half of all chickens died from viral infection. In the virus transmission study in which chickens were given drinking water collected from virus-inoculated sparrows, mortality due to viral infection was observed in chickens. Our data suggest that Eurasian tree sparrows could be biological vectors of the H5N1 highly pathogenic avian influenza virus. In addition to frequent virus detection in the drinking water of sparrows, the results of the virus transmission study suggest that waterborne pathways could be important for viral transmission from tree sparrows to poultry.

  1. In silico Analysis Revealed High-risk Single Nucleotide Polymorphisms in Human Pentraxin-3 Gene and their Impact on Innate Immune Response against Microbial Pathogens

    PubMed Central

    Thakur, Raman; Shankar, Jata

    2016-01-01

    Pentraxin-3 (PTX-3) protein is an evolutionary conserved protein that acts as a soluble pattern-recognition receptor for pathogens and plays important role in innate immune response. It recognizes various pathogens by interacting with extracellular moieties such as glactomannan of conidia (Aspergillus fumigatus), lipopolysaccharide of Pseudomonas aeruginosa, Streptococcus pneumonia and Salmonella typhimurium. Thus, PTX-3 protein helps to clear these pathogens by activating downstream innate immune process. In this study, computational methods were used to analyze various non-synonymous single nucleotide polymorphisms (nsSNPs) in PTX-3 gene. Three different databases were used to retrieve SNP data sets followed by seven different in silico algorithms to screen nsSNPs in PTX-3 gene. Sequence homology based approach was used to identify nsSNPs. Conservation profile of PTX-3 protein amino acid residues were predicted by ConSurf web server. In total, 10 high-risk nsSNPs were identified in pentraxin-domain of PTX-3 gene. Out of these 10 high-risk nsSNPs, 4 were present in the conserved structural and functional residues of the pentraxin-domain, hence, selected for structural analyses. The results showed alteration in the putative structure of pentraxin-domain. Prediction of protein–protein interactions analysis showed association of PTX-3 protein with C1q component of complement pathway. Different functional and structural residues along with various putative phosphorylation sites and evolutionary relationship were also predicted for PTX-3 protein. This is the first extensive computational analyses of pentraxin protein family with nsSNPs and will serve as a valuable resource for future population based studies. PMID:26941719

  2. Rain-induced ejection of pathogens from leaves: revisiting the hypothesis of splash-on-film using high-speed visualization.

    PubMed

    Gilet, Tristan; Bourouiba, Lydia

    2014-12-01

    Plant diseases are a major cause of losses of crops worldwide. Although rainfalls and foliar disease outbreaks are correlated, the detailed mechanism explaining their link remains poorly understood. The common assumption from phytopathology for such link is that a splash is generated upon impact of raindrops on contaminated liquid films coating sick leaves. We examine this assumption using direct high-speed visualizations of the interactions of raindrops and leaves over a range of plants. We show that films are seldom found on the surface of common leaves. We quantify the leaf-surface's wetting properties, showing that sessile droplets instead of films are predominant on the surfaces of leaves. We find that the presence of sessile drops rather than that of films has important implications when coupled with the compliance of a leaf: it leads to a new physical picture consisting of two dominant rain-induced mechanisms of ejection of pathogens. The first involves a direct interaction between the fluids of the raindrop and the sessile drops via an off-centered splash. The second involves the indirect action of the raindrop that leads to the inertial detachment of the sessile drop via the leaf's motion imparted by the impact of the raindrop. Both mechanisms are distinct from the commonly assumed scenario of splash-on-film in terms of outcome: they result in different fragmentation processes induced by surface tension, and, thus, different size-distributions of droplets ejected. This is the first time that modern direct high-speed visualizations of impacts on leaves are used to examine rain-induced ejection of pathogens at the level of a leaf and identify the inertial detachment and off-center splash ejections as alternatives to the classically assumed splash-on-film ejections of foliar pathogens.

  3. In silico Analysis Revealed High-risk Single Nucleotide Polymorphisms in Human Pentraxin-3 Gene and their Impact on Innate Immune Response against Microbial Pathogens.

    PubMed

    Thakur, Raman; Shankar, Jata

    2016-01-01

    Pentraxin-3 (PTX-3) protein is an evolutionary conserved protein that acts as a soluble pattern-recognition receptor for pathogens and plays important role in innate immune response. It recognizes various pathogens by interacting with extracellular moieties such as glactomannan of conidia (Aspergillus fumigatus), lipopolysaccharide of Pseudomonas aeruginosa, Streptococcus pneumonia and Salmonella typhimurium. Thus, PTX-3 protein helps to clear these pathogens by activating downstream innate immune process. In this study, computational methods were used to analyze various non-synonymous single nucleotide polymorphisms (nsSNPs) in PTX-3 gene. Three different databases were used to retrieve SNP data sets followed by seven different in silico algorithms to screen nsSNPs in PTX-3 gene. Sequence homology based approach was used to identify nsSNPs. Conservation profile of PTX-3 protein amino acid residues were predicted by ConSurf web server. In total, 10 high-risk nsSNPs were identified in pentraxin-domain of PTX-3 gene. Out of these 10 high-risk nsSNPs, 4 were present in the conserved structural and functional residues of the pentraxin-domain, hence, selected for structural analyses. The results showed alteration in the putative structure of pentraxin-domain. Prediction of protein-protein interactions analysis showed association of PTX-3 protein with C1q component of complement pathway. Different functional and structural residues along with various putative phosphorylation sites and evolutionary relationship were also predicted for PTX-3 protein. This is the first extensive computational analyses of pentraxin protein family with nsSNPs and will serve as a valuable resource for future population based studies.

  4. Common themes in microbial pathogenicity.

    PubMed Central

    Finlay, B B; Falkow, S

    1989-01-01

    A bacterial pathogen is a highly adapted microorganism which has the capacity to cause disease. The mechanisms used by pathogenic bacteria to cause infection and disease usually include an interactive group of virulence determinants, sometimes coregulated, which are suited for the interaction of a particular microorganism with a specific host. Because pathogens must overcome similar host barriers, common themes in microbial pathogenesis have evolved. However, these mechanisms are diverse between species and not necessarily conserved; instead, convergent evolution has developed several different mechanisms to overcome host barriers. The success of a bacterial pathogen can be measured by the degree with which it replicates after entering the host and reaching its specific niche. Successful microbial infection reflects persistence within a host and avoidance or neutralization of the specific and nonspecific defense mechanisms of the host. The degree of success of a pathogen is dependent upon the status of the host. As pathogens pass through a host, they are exposed to new environments. Highly adapted pathogenic organisms have developed biochemical sensors exquisitely designed to measure and respond to such environmental stimuli and accordingly to regulate a cascade of virulence determinants essential for life within the host. The pathogenic state is the product of dynamic selective pressures on microbial populations. PMID:2569162

  5. Surveillance for high pathogenicity avian influenza virus in wild birds in the Pacific Flyway of the United States, 2006-2007

    USGS Publications Warehouse

    Dusek, R.J.; Bortner, J.B.; DeLiberto, T.J.; Hoskins, J.; Franson, J. Christian; Bales, B.D.; Yparraguirre, D.; Swafford, S.R.; Ip, H.S.

    2009-01-01

    In 2006 the U.S. Department of Agriculture, U.S. Department of Interior, and cooperating state fish and wildlife agencies began surveillance for high-pathogenicity avian influenza (HPAI) H5N1 virus in wild birds in the Pacific Flyway of the United States. This surveillance effort was highly integrated in California, Oregon, Washington, Idaho, Nevada, Arizona, Utah, and western Montana, with collection of samples coordinated with state agencies. Sampling focused on live wild birds, hunterkilled waterfowl during state hunting seasons, and wild bird mortality events. Of 20,888 samples collected, 18,139 were from order Anseriformes (waterfowl) and 2010 were from order Charadriiformes (shorebirds), representing the two groups of birds regarded to be the primary reservoirs of avian influenza viruses. Although 83 birds were positive by H5 real-time reverse transcription polymerase chain reaction (rRT-PCR), no HPAI H5N1 virus was found. Thirty-two virus isolates were obtained from the H5- positive samples, including low-pathogenicity H5 viruses identified as H5N2, H5N3, and H5N9.

  6. Surveillance plan for the early detection of H5N1 highly pathogenic avian influenza virus in migratory birds in the United States: surveillance year 2009

    USGS Publications Warehouse

    Brand, Christopher J.

    2009-01-01

    Executive Summary: This Surveillance Plan (Plan) describes plans for conducting surveillance of wild birds in the United States and its Territories and Freely-Associated States to provide for early detection of the introduction of the H5N1 Highly Pathogenic Avian Influenza (HPAI) subtype of the influenza A virus by migratory birds during the 2009 surveillance year, spanning the period of April 1, 2009 - March 31, 2010. The Plan represents a continuation of surveillance efforts begun in 2006 under the Interagency Strategic Plan for the Early Detection of H5N1 Highly Pathogenic Avian Influenza in Wild Migratory Birds (U.S. Department of Agriculture and U.S. Department of the Interior, 2006). The Plan sets forth sampling plans by: region, target species or species groups to be sampled, locations of sampling, sample sizes, and sampling approaches and methods. This Plan will be reviewed annually and modified as appropriate for subsequent surveillance years based on evaluation of information from previous years of surveillance, changing patterns and threats of H5N1 HPAI, and changes in funding availability for avian influenza surveillance. Specific sampling strategies will be developed accordingly within each of six regions, defined here as Alaska, Hawaiian/Pacific Islands, Lower Pacific Flyway (Washington, Oregon, California, Idaho, Nevada, Arizona), Central Flyway, Mississippi Flyway, and Atlantic Flyway.

  7. Reintroduction of H5N1 highly pathogenic avian influenza virus by migratory water birds, causing poultry outbreaks in the 2010-2011 winter season in Japan.

    PubMed

    Sakoda, Yoshihiro; Ito, Hiroshi; Uchida, Yuko; Okamatsu, Masatoshi; Yamamoto, Naoki; Soda, Kosuke; Nomura, Naoki; Kuribayashi, Saya; Shichinohe, Shintaro; Sunden, Yuji; Umemura, Takashi; Usui, Tatsufumi; Ozaki, Hiroichi; Yamaguchi, Tsuyoshi; Murase, Toshiyuki; Ito, Toshihiro; Saito, Takehiko; Takada, Ayato; Kida, Hiroshi

    2012-03-01

    H5N1 highly pathogenic avian influenza virus (HPAIV) was reintroduced and caused outbreaks in chickens in the 2010-2011 winter season in Japan, which had been free from highly pathogenic avian influenza (HPAI) since 2007 when HPAI outbreaks occurred and were controlled. On 14 October 2010 at Lake Ohnuma, Wakkanai, the northernmost part of Hokkaido, Japan, H5N1 HPAIVs were isolated from faecal samples of ducks flying from their nesting lakes in Siberia. Since then, in Japan, H5N1 HPAIVs have been isolated from 63 wild birds in 17 prefectures and caused HPAI outbreaks in 24 chicken farms in nine prefectures by the end of March in 2011. Each of these isolates was genetically closely related to the HPAIV isolates at Lake Ohnuma, and those in China, Mongolia, Russia and Korea, belonging to genetic clade 2.3.2.1. In addition, these isolates were genetically classified into three groups, suggesting that the viruses were transmitted by migratory water birds through at least three different routes from their northern territory to Japan. These isolates were antigenic variants, which is consistent with selection in poultry under the immunological pressure induced by vaccination. To prevent the perpetuation of viruses in the lakes where water birds nest in summer in Siberia, prompt eradication of HPAIVs in poultry is urgently needed in Asian countries where HPAI has not been controlled.

  8. A Novel High-Throughput Method for Molecular Detection of Human Pathogenic Viruses Using a Nanofluidic Real-Time PCR System.

    PubMed

    Coudray-Meunier, Coralie; Fraisse, Audrey; Martin-Latil, Sandra; Delannoy, Sabine; Fach, Patrick; Perelle, Sylvie

    2016-01-01

    Human enteric viruses are recognized as the main causes of food- and waterborne diseases worldwide. Sensitive and quantitative detection of human enteric viruses is typically achieved through quantitative RT-PCR (RT-qPCR). A nanofluidic real-time PCR system was used to develop novel high-throughput methods for qualitative molecular detection (RT-qPCR array) and quantification of human pathogenic viruses by digital RT-PCR (RT-dPCR). The performance of high-throughput PCR methods was investigated for detecting 19 human pathogenic viruses and two main process controls used in food virology. The conventional real-time PCR system was compared to the RT-dPCR and RT-qPCR array. Based on the number of genome copies calculated by spectrophotometry, sensitivity was found to be slightly better with RT-qPCR than with RT-dPCR for 14 viruses by a factor range of from 0.3 to 1.6 log10. Conversely, sensitivity was better with RT-dPCR than with RT-qPCR for seven viruses by a factor range of from 0.10 to 1.40 log10. Interestingly, the number of genome copies determined by RT-dPCR was always from 1 to 2 log10 lower than the expected copy number calculated by RT-qPCR standard curve. The sensitivity of the RT-qPCR and RT-qPCR array assays was found to be similar for two viruses, and better with RT-qPCR than with RT-qPCR array for eighteen viruses by a factor range of from 0.7 to 3.0 log10. Conversely, sensitivity was only 0.30 log10 better with the RT-qPCR array than with conventional RT-qPCR assays for norovirus GIV detection. Finally, the RT-qPCR array and RT-dPCR assays were successfully used together to screen clinical samples and quantify pathogenic viruses. Additionally, this method made it possible to identify co-infection in clinical samples. In conclusion, given the rapidity and potential for large numbers of viral targets, this nanofluidic RT-qPCR assay should have a major impact on human pathogenic virus surveillance and outbreak investigations and is likely to be of benefit

  9. A Novel High-Throughput Method for Molecular Detection of Human Pathogenic Viruses Using a Nanofluidic Real-Time PCR System.

    PubMed

    Coudray-Meunier, Coralie; Fraisse, Audrey; Martin-Latil, Sandra; Delannoy, Sabine; Fach, Patrick; Perelle, Sylvie

    2016-01-01

    Human enteric viruses are recognized as the main causes of food- and waterborne diseases worldwide. Sensitive and quantitative detection of human enteric viruses is typically achieved through quantitative RT-PCR (RT-qPCR). A nanofluidic real-time PCR system was used to develop novel high-throughput methods for qualitative molecular detection (RT-qPCR array) and quantification of human pathogenic viruses by digital RT-PCR (RT-dPCR). The performance of high-throughput PCR methods was investigated for detecting 19 human pathogenic viruses and two main process controls used in food virology. The conventional real-time PCR system was compared to the RT-dPCR and RT-qPCR array. Based on the number of genome copies calculated by spectrophotometry, sensitivity was found to be slightly better with RT-qPCR than with RT-dPCR for 14 viruses by a factor range of from 0.3 to 1.6 log10. Conversely, sensitivity was better with RT-dPCR than with RT-qPCR for seven viruses by a factor range of from 0.10 to 1.40 log10. Interestingly, the number of genome copies determined by RT-dPCR was always from 1 to 2 log10 lower than the expected copy number calculated by RT-qPCR standard curve. The sensitivity of the RT-qPCR and RT-qPCR array assays was found to be similar for two viruses, and better with RT-qPCR than with RT-qPCR array for eighteen viruses by a factor range of from 0.7 to 3.0 log10. Conversely, sensitivity was only 0.30 log10 better with the RT-qPCR array than with conventional RT-qPCR assays for norovirus GIV detection. Finally, the RT-qPCR array and RT-dPCR assays were successfully used together to screen clinical samples and quantify pathogenic viruses. Additionally, this method made it possible to identify co-infection in clinical samples. In conclusion, given the rapidity and potential for large numbers of viral targets, this nanofluidic RT-qPCR assay should have a major impact on human pathogenic virus surveillance and outbreak investigations and is likely to be of benefit

  10. A Novel High-Throughput Method for Molecular Detection of Human Pathogenic Viruses Using a Nanofluidic Real-Time PCR System

    PubMed Central

    Coudray-Meunier, Coralie; Fraisse, Audrey; Martin-Latil, Sandra; Delannoy, Sabine; Fach, Patrick; Perelle, Sylvie

    2016-01-01

    Human enteric viruses are recognized as the main causes of food- and waterborne diseases worldwide. Sensitive and quantitative detection of human enteric viruses is typically achieved through quantitative RT-PCR (RT-qPCR). A nanofluidic real-time PCR system was used to develop novel high-throughput methods for qualitative molecular detection (RT-qPCR array) and quantification of human pathogenic viruses by digital RT-PCR (RT-dPCR). The performance of high-throughput PCR methods was investigated for detecting 19 human pathogenic viruses and two main process controls used in food virology. The conventional real-time PCR system was compared to the RT-dPCR and RT-qPCR array. Based on the number of genome copies calculated by spectrophotometry, sensitivity was found to be slightly better with RT-qPCR than with RT-dPCR for 14 viruses by a factor range of from 0.3 to 1.6 log10. Conversely, sensitivity was better with RT-dPCR than with RT-qPCR for seven viruses by a factor range of from 0.10 to 1.40 log10. Interestingly, the number of genome copies determined by RT-dPCR was always from 1 to 2 log10 lower than the expected copy number calculated by RT-qPCR standard curve. The sensitivity of the RT-qPCR and RT-qPCR array assays was found to be similar for two viruses, and better with RT-qPCR than with RT-qPCR array for eighteen viruses by a factor range of from 0.7 to 3.0 log10. Conversely, sensitivity was only 0.30 log10 better with the RT-qPCR array than with conventional RT-qPCR assays for norovirus GIV detection. Finally, the RT-qPCR array and RT-dPCR assays were successfully used together to screen clinical samples and quantify pathogenic viruses. Additionally, this method made it possible to identify co-infection in clinical samples. In conclusion, given the rapidity and potential for large numbers of viral targets, this nanofluidic RT-qPCR assay should have a major impact on human pathogenic virus surveillance and outbreak investigations and is likely to be of benefit

  11. Biological preparation of highly effective immunomagnetic beads for the separation, concentration, and detection of pathogenic bacteria in milk.

    PubMed

    Lim, Min-Cheol; Lee, Gwan-Hyung; Huynh, Duyen Thi Ngoc; Hong, Chae-Eun; Park, Soo-Yeon; Jung, Jong-Yun; Park, Cheon-Seok; Ko, Sungho; Kim, Young-Rok

    2016-09-01

    We introduce a system for the efficient separation and concentration of pathogenic bacteria using biologically prepared immunomagnetic beads. Amylose magnetic beads (AMBs) were synthesized by an enzymatic reaction of amylosucrase from Deinococcus geothermalis (DGAS). The simple and rapid conjugation of AMBs and antibodies was achieved by the MBP-SPG fusion protein. MBP (maltose binding protein) binds to the surface of an AMB owing to its intrinsic affinity to the di-glucose in the AMB. SPG (streptococcal protein G) fused to the MBP has specific affinity to the Fc region of the antibody. Anti-Escherichia coli O157 antibodies were conjugated to the AMBs through a MBP-SPG linker without any physical and chemical treatments. The efficiency of separation and concentration of the target E. coli O157:H7 by the functionalized AMBs was revealed by plating counting, conventional polymerase chain reaction (PCR), and real-time RCR analysis. The immuno-AMBs effectively separated and concentrated the target bacteria from a commercial milk sample spiked with known number of bacteria, which was then analyzed by PCR to a detection limit of 10CFU/mL. On the other hand, no PCR product was produced when milk was introduced directly to a PCR reaction. These results show that MBP-SPG is an effective linker and the resulting immuno-AMBs are capable of separating and concentrating the target bacteria from a food matrix. PMID:27315334

  12. Serological Evidence for Non-Lethal Exposures of Mongolian Wild Birds to Highly Pathogenic Avian Influenza H5N1 Virus

    PubMed Central

    Gilbert, Martin; Koel, Björn F.; Bestebroer, Theo M.; Lewis, Nicola S.; Smith, Derek J.; Fouchier, Ron A. M.

    2014-01-01

    Surveillance for highly pathogenic avian influenza viruses (HPAIV) in wild birds is logistically demanding due to the very low rates of virus detection. Serological approaches may be more cost effective as they require smaller sample sizes to identify exposed populations. We hypothesized that antigenic differences between classical Eurasian H5 subtype viruses (which have low pathogenicity in chickens) and H5N1 viruses of the Goose/Guangdong/96 H5 lineage (which are HPAIV) may be used to differentiate populations where HPAIVs have been circulating, from those where they have not. To test this we performed hemagglutination inhibition assays to compare the reactivity of serum samples from wild birds in Mongolia (where HPAIV has been circulating, n = 1,832) and Europe (where HPAIV has been rare or absent, n = 497) to a panel of reference viruses including classical Eurasian H5 (of low pathogenicity), and five HPAIV H5N1 antigens of the Asian lineage A/Goose/Guangdong/1/96. Antibody titres were detected against at least one of the test antigens for 182 Mongolian serum samples (total seroprevalence of 0.10, n = 1,832, 95% adjusted Wald confidence limits of 0.09–0.11) and 25 of the European sera tested (total seroprevalence of 0.05, n = 497, 95% adjusted Wald confidence limits of 0.03–0.07). A bias in antibody titres to HPAIV antigens was found in the Mongolian sample set (22/182) that was absent in the European sera (0/25). Although the interpretation of serological data from wild birds is complicated by the possibility of exposure to multiple strains, and variability in the timing of exposure, these findings suggest that a proportion of the Mongolian population had survived exposure to HPAIV, and that serological assays may enhance the targeting of traditional HPAIV surveillance toward populations where isolation of HPAIV is more likely. PMID:25502318

  13. Serological evidence for non-lethal exposures of Mongolian wild birds to highly pathogenic avian influenza H5N1 virus.

    PubMed

    Gilbert, Martin; Koel, Björn F; Bestebroer, Theo M; Lewis, Nicola S; Smith, Derek J; Fouchier, Ron A M

    2014-01-01

    Surveillance for highly pathogenic avian influenza viruses (HPAIV) in wild birds is logistically demanding due to the very low rates of virus detection. Serological approaches may be more cost effective as they require smaller sample sizes to identify exposed populations. We hypothesized that antigenic differences between classical Eurasian H5 subtype viruses (which have low pathogenicity in chickens) and H5N1 viruses of the Goose/Guangdong/96 H5 lineage (which are HPAIV) may be used to differentiate populations where HPAIVs have been circulating, from those where they have not. To test this we performed hemagglutination inhibition assays to compare the reactivity of serum samples from wild birds in Mongolia (where HPAIV has been circulating, n = 1,832) and Europe (where HPAIV has been rare or absent, n = 497) to a panel of reference viruses including classical Eurasian H5 (of low pathogenicity), and five HPAIV H5N1 antigens of the Asian lineage A/Goose/Guangdong/1/96. Antibody titres were detected against at least one of the test antigens for 182 Mongolian serum samples (total seroprevalence of 0.10, n = 1,832, 95% adjusted Wald confidence limits of 0.09-0.11) and 25 of the European sera tested (total seroprevalence of 0.05, n = 497, 95% adjusted Wald confidence limits of 0.03-0.07). A bias in antibody titres to HPAIV antigens was found in the Mongolian sample set (22/182) that was absent in the European sera (0/25). Although the interpretation of serological data from wild birds is complicated by the possibility of exposure to multiple strains, and variability in the timing of exposure, these findings suggest that a proportion of the Mongolian population had survived exposure to HPAIV, and that serological assays may enhance the targeting of traditional HPAIV surveillance toward populations where isolation of HPAIV is more likely.

  14. Isolation and identification of highly pathogenic avian influenza H5N1 virus from Houbara bustards (Chlamydotis undulata macqueenii) and contact falcons.

    PubMed

    Khan, Owais Ahmed; Shuaib, Mohammad Adam; Rhman, Salah Shaban Abdel; Ismail, Mahmoud Moussa; Hammad, Yousef Al; Baky, Mansour Hashim Abdel; Fusaro, Alice; Salviato, Annalisa; Cattoli, Giovanni

    2009-02-01

    Highly pathogenic influenza virus (HPAIV) H5N1 has caused mortality and morbidity in many species of domestic and wild bird. The Houbara bustard (Chlamydotis undulata macqueenii) is a solitary bird that inhabits semi-desert regions. It is known to be susceptible to avianpox, avian paramyxovirus type 1, and low-pathogenicity avian influenza H9N2. We report an outbreak of H5N1 HPAIV in Houbara bustards, which were introduced into the Kingdom of Saudi Arabia for falconry purposes. Ninety-three per cent mortality (38 out of 41 birds) in the infected Houbara bustard flock and about 62.5% mortality (10 out of 16 birds) in falcons that came in contact with these birds were observed. Pooled cloacal and tracheal swabs from Houbara bustards as well as visceral organ homogenates collected in Houbara bustards and falcons were tested by real-time reverse transcriptase-polymerase chain reaction, and virus isolation was attempted in specific pathogen free hens' eggs. The viruses isolated were characterized as HPAIV H5N1. Phylogenetic analysis of the haemagglutinating and Neuraminidase (NA) genes revealed that the viruses isolated from Houbara bustards and falcons were closely related to each other and to Kuwaiti H5N1 strains isolated in 2007. Interestingly, they were genetically distinguishable from the co-circulating A/H5N1 viruses in Kingdom of Saudi Arabia causing outbreaks in domestic birds. This case emphasizes the need for surveillance of this endangered species in its natural habitat. PMID:19130352

  15. Pathogene Mikroorganismen

    NASA Astrophysics Data System (ADS)

    Wagner, Martin

    Infektionen, die vom Tier auf den Menschen übertragen werden, werden als Zoonosen bezeichnet. Pathogene Mikroorganismen können entweder durch Mensch-Mensch, Mensch-Tier-Kontakt oder durch Kontakt mit kontaminierten Vektoren übertragen werden [39]. Vektoren können einerseits belebt (z. B. blutsaugende Insekten), andererseits unbelebt sein. Kontaminierte Lebensmittel und Wasser gehören zu den wichtigsten unbelebten Vektoren. Neben Lebensmitteln können aber auch kontaminierte Gegenstände oder der Kontakt mit Kontaminationsquellen in der Umwelt Auslöser von Krankheitsfällen sein. Weltweit sind mehr als 1400 krankheitsverursachende biologische Agentien bekannt, von denen über 60 % ein zoonotisches Potenzial aufweisen. Als Ergebnis von Expertengesprächen wurde kürzlich berichtet, dass etwa 3 bis 4, meist virale, neu auftretende Infektionskrankheiten ("emerging diseases“) pro Jahr erwartet werden können [15]. Es handelt sich bei diesen Vorgängen aber nicht nur um das Auftauchen vollkommen neuer oder unbeschriebener Spezies, sondern auch um evolutionsbedingte Anpassungen von mikrobiellen Populationen an neue Bedingungen in ihrem Ökosystem [7]. Molekulare Analysen an Umweltchlamydien erbrachten Hinweise, dass die Evolution erste genetische Pathogenitätsmerkmale in dieser Spezies schon vor 700 Mio. Jahren entstehen ließ [14]. Viele Faktoren befeuern den Prozess der Anpassung, unter anderem auch alle Strategien, mit denen der Mensch seit Jahrtausenden versucht, Lebensmittel sicher und haltbar zu machen. Als die treibenden Kräfte des Auftretens neuer Krankheitserreger werden in der Gegenwart vor allem das sich ändernde Weltklima, die globalen Warenströme und die sich verändernden Konsumgewohnheiten genannt. Es steht auch außer Zweifel, dass viele dieser Erreger Tiere als ihr natürliches Reservoir haben werden, d. h. Zoonosen im klassischen Sinne sind [15].

  16. Protection Afforded by a Recombinant Turkey Herpesvirus-H5 Vaccine Against the 2014 European Highly Pathogenic H5N8 Avian Influenza Strain.

    PubMed

    Steensels, M; Rauw, F; van den Berg, Th; Marché, S; Gardin, Y; Palya, V; Lambrecht, B

    2016-05-01

    A highly pathogenic avian influenza (HPAI) H5N8 (clade 2.3.4.4) virus, circulating in Asia (South Korea, Japan, and southern China) since the beginning of 2014, reached the European continent in November 2014. Germany, the Netherlands, the United Kingdom, Italy, and Hungary confirmed H5N8 infection of poultry farms of different species and of several wild bird species. Unlike the Asian highly pathogenic (HP) H5N1, this HP H5N8 also went transatlantic and reached the American West Coast by the end of 2014, affecting wild birds as well as backyard and commercial poultry. This strain induces high mortality and morbidity in Galliformes, whereas wild birds seem only moderately affected. A recombinant turkey herpesvirus (rHVT) vector vaccine expressing the H5 gene of a clade 2.2 H5N1 strain (rHVT-H5) previously demonstrated a highly efficient clinical protection and reduced viral excretion against challenge with Asian HP H5N1 strains of various clades (2.2, 2.2.1, 2.2.1.1, 2.1.3, 2.1.3.2, and 2.3.2.1) and was made commercially available in various countries where the disease is endemic. To evaluate the protective efficacy of the rHVT-H5 vaccine against the first German H5N8 turkey isolate (H5N8 GE), a challenge experiment was set up in specific-pathogen-free (SPF) chickens, and the clinical and excretional protection was evaluated. SPF chickens were vaccinated subcutaneously at 1 day old and challenged oculonasally at 4 wk of age with two viral dosages, 10(5) and 10(6) 50% egg infective doses. Morbidity and mortality were monitored daily in unvaccinated and vaccinated groups, whereas viral shedding by oropharyngeal and cloacal routes was evaluated at 2, 5, 9, and 14 days postinoculation (dpi). Serologic monitoring after vaccination and challenge was also carried out. Despite its high antigenic divergence of the challenge H5N8 strain, a single rHVT-H5 vaccine administration at 1 day old resulted in a full clinical protection against challenge and a significant reduction

  17. An overview of the epidemic of highly pathogenic H5N1 avian influenza virus in Egypt: epidemiology and control challenges.

    PubMed

    Abdelwhab, E M; Hafez, H M

    2011-05-01

    Emergence of the highly pathogenic avian influenza (HPAI) H5N1 virus in Egypt in mid-February 2006 caused significant losses for the poultry industry and constituted a potential threat to public health. Since late 2007, there has been increasing evidence that stable lineages of H5N1 viruses are being established in chickens and humans in Egypt. The virus has been detected in wild, feral and zoo birds and recently was found in donkeys and pigs. Most of the outbreaks in poultry and humans occurred in the highly populated Nile delta. The temporal pattern of the virus has changed since 2009 with outbreaks now occurring in the warmer months of the year. Challenges to control of endemic disease in Egypt are discussed. For the foreseeable future, unless a global collaboration exists, HPAI H5N1 virus in Egypt will continue to compromise the poultry industry, endanger public health and pose a serious pandemic threat. PMID:21281550

  18. Experimental challenge and pathology of highly pathogenic avian influenza virus H5N1 in dunlin (Calidris alpina), an intercontinental migrant shorebird species

    PubMed Central

    Hall, Jeffrey S.; Franson, J. Christian; Gill, Robert E.; Meteyer, Carol U.; TeSlaa, Joshua L.; Nashold, Sean; Dusek, Robert J.; Ip, Hon S.

    2011-01-01

    Please cite this paper as: Hall et al. (2011). Experimental challenge and pathology of highly pathogenic avian influenza virus H5N1 in dunlin (Calidris alpina), an intercontinental migrant shorebird species. Influenza and Other Respiratory Viruses 5(5), 365–372. Background  Shorebirds (Charadriiformes) are considered one of the primary reservoirs of avian influenza. Because these species are highly migratory, there is concern that infected shorebirds may be a mechanism by which highly pathogenic avian influenza virus (HPAIV) H5N1 could be introduced into North America from Asia. Large numbers of dunlin (Calidris alpina) migrate from wintering areas in central and eastern Asia, where HPAIV H5N1 is endemic, across the Bering Sea to breeding areas in Alaska. Low pathogenic avian influenza virus has been previously detected in dunlin, and thus, dunlin represent a potential risk to transport HPAIV to North America. To date no experimental challenge studies have been performed in shorebirds. Methods  Wild dunlin were inoculated intranasally and intrachoanally various doses of HPAIV H5N1. The birds were monitored daily for virus excretion, disease signs, morbidity, and mortality. Results  The infectious dose of HPAIV H5N1 in dunlin was determined to be 101.7 EID50/100 μl and that the lethal dose was 101.83 EID50/100 μl. Clinical signs were consistent with neurotropic disease, and histochemical analyses revealed that infection was systemic with viral antigen and RNA most consistently found in brain tissues. Infected birds excreted relatively large amounts of virus orally (104 EID50) and smaller amounts cloacally. Conclusions  Dunlin are highly susceptible to infection with HPAIV H5N1. They become infected after exposure to relatively small doses of the virus and if they become infected, they are most likely to suffer mortality within 3–5 days. These results have important implications regarding the risks of transport and transmission of HPAIV H5N1 to North

  19. Development of a quantitative assay amenable for high-throughput screening to target the type II secretion system for new treatments against plant-pathogenic bacteria.

    PubMed

    Tran, Nini; Zielke, Ryszard A; Vining, Oliver B; Azevedo, Mark D; Armstrong, Donald J; Banowetz, Gary M; McPhail, Kerry L; Sikora, Aleksandra E

    2013-09-01

    Plant-pathogenic bacteria are the causative agents of diseases in important agricultural crops and ornamental plants. The severe economic burden of these diseases requires seeking new approaches for their control, particularly because phytopathogenic bacteria are often resistant to available treatments. The type II secretion (T2S) system is a key virulence factor used by major groups of phytopathogenic bacteria. The T2S machinery transports many hydrolytic enzymes responsible for degradation of the plant cell wall, thus enabling successful colonization and dissemination of the bacteria in the plant host. The genetic inactivation of the T2S system leads to loss of virulence, which strongly suggests that targeting the T2S could enable new treatments against plant-pathogenic bacteria. Accordingly, we have designed and optimized an assay to identify small-molecule inhibitors of the T2S system. This assay uses a double parametric output: measurement of bacterial growth and the enzymatic activity of cellulase, which is secreted via the T2S pathway in our model organism Dickeya dadantii. The assay was evaluated by screening natural extracts, culture filtrates isolated from rhizosphere bacteria, and a collection of pharmaceutically active compounds in LOPAC(1280). The calculated Z' values of 0.63, 0.63, and 0.58, respectively, strongly suggest that the assay is applicable for a high-throughput screening platform.

  20. Evaluation of Patients with Community-Acquired Pneumonia Caused by Zoonotic Pathogens in an Area with a High Density of Animal Farms.

    PubMed

    Huijskens, E G W; Smit, L A M; Rossen, J W A; Heederik, D; Koopmans, M

    2016-03-01

    Intensive animal farming could potentially lead to outbreaks of infectious diseases. Clinicians are at the forefront of detecting unusual diseases, but the lack of specificity of zoonotic disease symptoms makes this a challenging task. We evaluated patients with community-acquired pneumonia (CAP) with known and unknown aetiology in an area with a high livestock density and a potential association with animal farms in the proximity. Between 2008 and 2009, a period coinciding with a large Q fever outbreak in the Netherlands, patients with CAP were tested for the presence of possible respiratory pathogens. The presence and number of farm animals within 1 km of the patients' home address were assessed using geographic information system (GIS) and were compared between cases and age-matched control subjects. Of 408 patients with CAP, pathogens were detected in 275 (67.4%) patients. The presence of sheep and the number of goats were associated with CAP caused by Coxiella burnetii in a multiple logistic regression model (P < 0.05). CAP with unknown aetiology was not associated with the presence of animal farms (P > 0.10). The use of GIS in combination with aetiology of CAP could be potentially used to target diagnostics and to identify outbreaks of rare zoonotic disease.

  1. Movements of Wild Ruddy Shelducks in the Central Asian Flyway and Their Spatial Relationship to Outbreaks of Highly Pathogenic Avian Influenza H5N1

    PubMed Central

    Takekawa, John Y.; Prosser, Diann J.; Collins, Bridget M.; Douglas, David C.; Perry, William M.; Yan, Baoping; Ze, Luo; Hou, Yuansheng; Lei, Fumin; Li, Tianxian; Li, Yongdong; Newman, Scott H.

    2013-01-01

    Highly pathogenic avian influenza H5N1 remains a serious concern for both poultry and human health. Wild waterfowl are considered to be the reservoir for low pathogenic avian influenza viruses; however, relatively little is known about their movement ecology in regions where HPAI H5N1 outbreaks regularly occur. We studied movements of the ruddy shelduck (Tadorna ferruginea), a wild migratory waterfowl species that was infected in the 2005 Qinghai Lake outbreak. We defined their migration with Brownian Bridge utilization distribution models and their breeding and wintering grounds with fixed kernel home ranges. We correlated their movements with HPAI H5N1 outbreaks, poultry density, land cover, and latitude in the Central Asian Flyway. Our Akaike Information Criterion analysis indicated that outbreaks were correlated with land cover, latitude, and poultry density. Although shelduck movements were included in the top two models, they were not a top parameter selected in AICc stepwise regression results. However, timing of outbreaks suggested that outbreaks in the flyway began during the winter in poultry with spillover to wild birds during the spring migration. Thus, studies of the movement ecology of wild birds in areas with persistent HPAI H5N1 outbreaks may contribute to understanding their role in transmission of this disease. PMID:24022072

  2. Bactericidal effect of polyethyleneimine capped ZnO nanoparticles on multiple antibiotic resistant bacteria harboring genes of high-pathogenicity island.

    PubMed

    Chakraborti, Soumyananda; Mandal, Amit Kumar; Sarwar, Shamila; Singh, Prashantee; Chakraborty, Ranadhir; Chakrabarti, Pinak

    2014-09-01

    Zinc oxide nanoparticles (ZnO-NP) were synthesized by alcoholic route using zinc acetate as the precursor material and lithium hydroxide as hydrolyzing agent. Further ZnO-PEI NP (derivative of ZnO-NP) was made in aqueous medium using the capping agent polyethyleneimine (PEI). The nanoparticles were characterized by XRD measurements, TEM and other techniques; the weight % of coating shell in the polymer-capped particles was determined by TGA. ZnO-PEI NP is more soluble in water than the uncapped ZnO-NP, and forms a colloidal suspension in water. PEI-capped ZnO-NP exhibited better antibacterial activity when compared with that of uncapped ZnO-NP against a range of multiple-antibiotic-resistant (MAR) Gram-negative bacterial strains harboring genes of high-pathogenicity island. ZnO-NP effectively killed these microorganisms by generating reactive oxygen species (ROS) and damaging bacterial membrane. ZnO-PEI NP at LD50 dose in combination with tetracycline showed synergistic effect to inhibit tetracycline-resistant Escherichia coli MREC33 growth by 80%. These results open up a new vista in therapeutics to use antibiotics (which have otherwise been rendered useless against MAR bacteria) in combination with minimized dosage of nanoparticles for the more effective control of MAR pathogenic bacteria.

  3. The enigma of the apparent disappearance of Eurasian highly pathogenic H5 clade 2.3.4.4 influenza A viruses in North American waterfowl.

    PubMed

    Krauss, Scott; Stallknecht, David E; Slemons, Richard D; Bowman, Andrew S; Poulson, Rebecca L; Nolting, Jacqueline M; Knowles, James P; Webster, Robert G

    2016-08-01

    One of the major unresolved questions in influenza A virus (IAV) ecology is exemplified by the apparent disappearance of highly pathogenic (HP) H5N1, H5N2, and H5N8 (H5Nx) viruses containing the Eurasian hemagglutinin 2.3.4.4 clade from wild bird populations in North America. The introduction of Eurasian lineage HP H5 clade 2.3.4.4 H5N8 IAV and subsequent reassortment with low-pathogenic H?N2 and H?N1 North American wild bird-origin IAVs in late 2014 resulted in widespread HP H5Nx IAV infections and outbreaks in poultry and wild birds across two-thirds of North America starting in November 2014 and continuing through June 2015. Although the stamping out strategies adopted by the poultry industry and animal health authorities in Canada and the United States-which included culling, quarantining, increased biosecurity, and abstention from vaccine use-were successful in eradicating the HP H5Nx viruses from poultry, these activities do not explain the apparent disappearance of these viruses from migratory waterfowl. Here we examine current and historical aquatic bird IAV surveillance and outbreaks of HP H5Nx in poultry in the United States and Canada, providing additional evidence of unresolved mechanisms that restrict the emergence and perpetuation of HP avian influenza viruses in these natural reservoirs. PMID:27457948

  4. Movements of wild ruddy shelducks in the Central Asian Flyway and their spatial relationship to outbreaks of highly pathogenic avian influenza H5N1

    USGS Publications Warehouse

    Takekawa, John Y.; Prosser, Diann J.; Collins, Bridget M.; Douglas, David C.; Perry, William M.; Baoping, Yan; Luo, Ze; Hou, Yuansheng; Lei, Fumin; Li, Tianxian; Li, Yongdong; Newman, Scott H.

    2013-01-01

    Highly pathogenic avian influenza H5N1 remains a serious concern for both poultry and human health. Wild waterfowl are considered to be the reservoir for low pathogenic avian influenza viruses; however, relatively little is known about their movement ecology in regions where HPAI H5N1 outbreaks regularly occur. We studied movements of the ruddy shelduck (Tadorna ferruginea), a wild migratory waterfowl species that was infected in the 2005 Qinghai Lake outbreak. We defined their migration with Brownian Bridge utilization distribution models and their breeding and wintering grounds with fixed kernel home ranges. We correlated their movements with HPAI H5N1 outbreaks, poultry density, land cover, and latitude in the Central Asian Flyway. Our Akaike Information Criterion analysis indicated that outbreaks were correlated with land cover, latitude, and poultry density. Although shelduck movements were included in the top two models, they were not a top parameter selected in AICc stepwise regression results. However, timing of outbreaks suggested that outbreaks in the flyway began during the winter in poultry with spillover to wild birds during the spring migration. Thus, studies of the movement ecology of wild birds in areas with persistent HPAI H5N1 outbreaks may contribute to understanding their role in transmission of this disease.

  5. Movements of wild ruddy shelducks in the Central Asian Flyway and their spatial relationship to outbreaks of highly pathogenic avian influenza H5N1.

    PubMed

    Takekawa, John Y; Prosser, Diann J; Collins, Bridget M; Douglas, David C; Perry, William M; Yan, Baoping; Ze, Luo; Hou, Yuansheng; Lei, Fumin; Li, Tianxian; Li, Yongdong; Newman, Scott H

    2013-09-09

    Highly pathogenic avian influenza H5N1 remains a serious concern for both poultry and human health. Wild waterfowl are considered to be the reservoir for low pathogenic avian influenza viruses; however, relatively little is known about their movement ecology in regions where HPAI H5N1 outbreaks regularly occur. We studied movements of the ruddy shelduck (Tadorna ferruginea), a wild migratory waterfowl species that was infected in the 2005 Qinghai Lake outbreak. We defined their migration with Brownian Bridge utilization distribution models and their breeding and wintering grounds with fixed kernel home ranges. We correlated their movements with HPAI H5N1 outbreaks, poultry density, land cover, and latitude in the Central Asian Flyway. Our Akaike Information Criterion analysis indicated that outbreaks were correlated with land cover, latitude, and poultry density. Although shelduck movements were included in the top two models, they were not a top parameter selected in AICc stepwise regression results. However, timing of outbreaks suggested that outbreaks in the flyway began during the winter in poultry with spillover to wild birds during the spring migration. Thus, studies of the movement ecology of wild birds in areas with persistent HPAI H5N1 outbreaks may contribute to understanding their role in transmission of this disease.

  6. The fungal pathogen Moniliophthora perniciosa has genes similar to plant PR-1 that are highly expressed during its interaction with cacao.

    PubMed

    Teixeira, Paulo J P L; Thomazella, Daniela P T; Vidal, Ramon O; do Prado, Paula F V; Reis, Osvaldo; Baroni, Renata M; Franco, Sulamita F; Mieczkowski, Piotr; Pereira, Gonçalo A G; Mondego, Jorge M C

    2012-01-01

    The widespread SCP/TAPS superfamily (SCP/Tpx-1/Ag5/PR-1/Sc7) has multiple biological functions, including roles in the immune response of plants and animals, development of male reproductive tract in mammals, venom activity in insects and reptiles and host invasion by parasitic worms. Plant Pathogenesis Related 1 (PR-1) proteins belong to this superfamily and have been characterized as markers of induced defense against pathogens. This work presents the characterization of eleven genes homologous to plant PR-1 genes, designated as MpPR-1, which were identified in the genome of Moniliophthora perniciosa, a basidiomycete fungus responsible for causing the devastating witches' broom disease in cacao. We describe gene structure, protein alignment and modeling analyses of the MpPR-1 family. Additionally, the expression profiles of MpPR-1 genes were assessed by qPCR in different stages throughout the fungal life cycle. A specific expression pattern was verified for each member of the MpPR-1 family in the conditions analyzed. Interestingly, some of them were highly and specifically expressed during the interaction of the fungus with cacao, suggesting a role for the MpPR-1 proteins in the infective process of this pathogen. Hypothetical functions assigned to members of the MpPR-1 family include neutralization of plant defenses, antimicrobial activity to avoid competitors and fruiting body physiology. This study provides strong evidence on the importance of PR-1-like genes for fungal virulence on plants.

  7. Development of highly sensitive electrochemical genosensor based on multiwalled carbon nanotubes-chitosan-bismuth and lead sulfide nanoparticles for the detection of pathogenic Aeromonas.

    PubMed

    Fernandes, António Maximiano; Abdalhai, Mandour H; Ji, Jian; Xi, Bing-Wen; Xie, Jun; Sun, Jiadi; Noeline, Rasoamandrary; Lee, Byong H; Sun, Xiulan

    2015-01-15

    In this paper, we reported the construction of new high sensitive electrochemical genosensor based on multiwalled carbon nanotubes-chitosan-bismuth complex (MWCNT-Chi-Bi) and lead sulfide nanoparticles for the detection of pathogenic Aeromonas. Lead sulfide nanoparticles capped with 5'-(NH2) oligonucleotides thought amide bond was used as signalizing probe DNA (sz-DNA) and thiol-modified oligonucleotides sequence was used as fixing probe DNA (fDNA). The two probes hybridize with target Aeromonas DNA (tDNA) sequence (fDNA-tDNA-szDNA). The signal of hybridization is detected by differential pulse voltammetry (DPV) after electrodeposition of released lead nanoparticles (PbS) from sz-DNA on the surface of glass carbon electrode decorated with MWCNT-Chi-Bi, which improves the deposition and traducing electrical signal. The optimization of incubation time, hybridization temperature, deposition potential, deposition time and the specificity of the probes were investigated. Our results showed the highest sensibility to detect the target gene when compared with related biosensors and polymerase chain reaction (PCR). The detection limit for this biosensor was 1.0×10(-14) M. We could detect lower than 10(2) CFU mL(-1) of Aeromonas in spiked tap water. This method is rapid and sensitive for the detection of pathogenic bacteria and would become a potential application in biomedical diagnosis, food safety and environmental monitoring.

  8. Evaluation of Patients with Community-Acquired Pneumonia Caused by Zoonotic Pathogens in an Area with a High Density of Animal Farms.

    PubMed

    Huijskens, E G W; Smit, L A M; Rossen, J W A; Heederik, D; Koopmans, M

    2016-03-01

    Intensive animal farming could potentially lead to outbreaks of infectious diseases. Clinicians are at the forefront of detecting unusual diseases, but the lack of specificity of zoonotic disease symptoms makes this a challenging task. We evaluated patients with community-acquired pneumonia (CAP) with known and unknown aetiology in an area with a high livestock density and a potential association with animal farms in the proximity. Between 2008 and 2009, a period coinciding with a large Q fever outbreak in the Netherlands, patients with CAP were tested for the presence of possible respiratory pathogens. The presence and number of farm animals within 1 km of the patients' home address were assessed using geographic information system (GIS) and were compared between cases and age-matched control subjects. Of 408 patients with CAP, pathogens were detected in 275 (67.4%) patients. The presence of sheep and the number of goats were associated with CAP caused by Coxiella burnetii in a multiple logistic regression model (P < 0.05). CAP with unknown aetiology was not associated with the presence of animal farms (P > 0.10). The use of GIS in combination with aetiology of CAP could be potentially used to target diagnostics and to identify outbreaks of rare zoonotic disease. PMID:26214299

  9. Differentiation between pathogenic serotype 1 isolates of Marek's disease virus and the Rispens CVI988 vaccine in Australia using real-time PCR and high resolution melt curve analysis.

    PubMed

    Renz, K G; Cheetham, B F; Walkden-Brown, S W

    2013-01-01

    Two real-time PCR assays were developed which enable quantitation and differentiation between pathogenic Australian isolates of Marek's disease virus (MDV) serotype 1 and the serotype 1 vaccine strain Rispens CVI988. The assays are based on a DNA sequence variation in the meq gene between pathogenic and vaccinal MDV1 which has been confirmed by sequencing of 20 Australian field strains of MDV. Complete specificity has been demonstrated in samples containing pathogenic MDV (n=20), Rispens (3 commercial vaccine strains), or both. The limit of detection of both the Rispens-specific and the pathogenic MDV1-specific assays was 10 viral copies/reaction. The tests successfully differentiated and quantified MDV in mixtures of pathogenic and vaccinal Rispens virus. A high resolution melt curve analysis targeting the same SNP used for the real-time PCR assays was also developed which successfully detected sequence variation between Md5, six Australian MDV1 isolates and the three Rispens vaccines. However it was ineffective at differentiating mixtures of pathogenic and vaccinal MDV1. The real-time PCR assays have both diagnostic and epidemiological applications as they enable differentiation and quantitation of Rispens CVI988 and pathogenic MDV1 in co-infected chickens in Australia.

  10. Pathogenicity of a highly exopolysaccharide-producing Halomonas strain causing epizootics in larval cultures of the Chilean scallop Argopecten purpuratus (Lamarck, 1819).

    PubMed

    Rojas, Rodrigo; Miranda, Claudio D; Amaro, Ana María

    2009-01-01

    Mass mortalities of larval cultures of Chilean scallop Argopecten purpuratus have repeatedly occurred in northern Chile, characterized by larval agglutination and accumulation in the bottom of rearing tanks. The exopolysaccharide slime (EPS) producing CAM2 strain was isolated as the primary organism from moribund larvae in a pathogenic outbreak occurring in a commercial hatchery producing larvae of the Chilean scallop Argopecten purpuratus located in Bahía Inglesa, Chile. The CAM2 strain was characterized biochemically and was identified by polymerase chain reaction amplification of 16S rRNA as Halomonas sp. (Accession number DQ885389.1). Healthy 7-day-old scallop larvae cultures were experimentally infected for a 48-h period with an overnight culture of the CAM2 strain at a final concentration of ca. 10(5) cells per milliliter, and the mortality and vital condition of larvae were determined by optical and scanning electron microscopy (SEM) to describe the chronology of the disease. Pathogenic action of the CAM2 strain was clearly evidenced by SEM analysis, showing a high ability to adhere and detach larvae velum cells by using its "slimy" EPS, producing agglutination, loss of motility, and a posterior sinking of scallop larvae. After 48 h, a dense bacterial slime on the shell surface was observed, producing high percentages of larval agglutination (63.28 +/- 7.87%) and mortality (45.03 +/- 4.32%) that were significantly (P < 0.05) higher than those of the unchallenged control cultures, which exhibited only 3.20 +/- 1.40% dead larvae and no larval agglutination. Furthermore, the CAM2 strain exhibited a high ability to adhere to fiberglass pieces of tanks used for scallop larvae rearing (1.64 x 10(5) cells adhered per square millimeters at 24 h postinoculation), making it very difficult to eradicate it from the culture systems. This is the first report of a pathogenic activity on scallop larvae of Halomonas species, and it prompts the necessity of an appraisal on

  11. Pathogenicity of a highly exopolysaccharide-producing Halomonas strain causing epizootics in larval cultures of the Chilean scallop Argopecten purpuratus (Lamarck, 1819).

    PubMed

    Rojas, Rodrigo; Miranda, Claudio D; Amaro, Ana María

    2009-01-01

    Mass mortalities of larval cultures of Chilean scallop Argopecten purpuratus have repeatedly occurred in northern Chile, characterized by larval agglutination and accumulation in the bottom of rearing tanks. The exopolysaccharide slime (EPS) producing CAM2 strain was isolated as the primary organism from moribund larvae in a pathogenic outbreak occurring in a commercial hatchery producing larvae of the Chilean scallop Argopecten purpuratus located in Bahía Inglesa, Chile. The CAM2 strain was characterized biochemically and was identified by polymerase chain reaction amplification of 16S rRNA as Halomonas sp. (Accession number DQ885389.1). Healthy 7-day-old scallop larvae cultures were experimentally infected for a 48-h period with an overnight culture of the CAM2 strain at a final concentration of ca. 10(5) cells per milliliter, and the mortality and vital condition of larvae were determined by optical and scanning electron microscopy (SEM) to describe the chronology of the disease. Pathogenic action of the CAM2 strain was clearly evidenced by SEM analysis, showing a high ability to adhere and detach larvae velum cells by using its "slimy" EPS, producing agglutination, loss of motility, and a posterior sinking of scallop larvae. After 48 h, a dense bacterial slime on the shell surface was observed, producing high percentages of larval agglutination (63.28 +/- 7.87%) and mortality (45.03 +/- 4.32%) that were significantly (P < 0.05) higher than those of the unchallenged control cultures, which exhibited only 3.20 +/- 1.40% dead larvae and no larval agglutination. Furthermore, the CAM2 strain exhibited a high ability to adhere to fiberglass pieces of tanks used for scallop larvae rearing (1.64 x 10(5) cells adhered per square millimeters at 24 h postinoculation), making it very difficult to eradicate it from the culture systems. This is the first report of a pathogenic activity on scallop larvae of Halomonas species, and it prompts the necessity of an appraisal on

  12. Experimental challenge and pathology of highly pathogenic avian influenza virus H5N1 in dunlin (Calidris alpina), an intercontinental migrant shorebird species

    USGS Publications Warehouse

    Hall, J.S.; Franson, J.C.; Gill, R.E.; Meteyer, C.U.; Teslaa, J.L.; Nashold, S.; Dusek, R.J.; Ip, H.S.

    2011-01-01

    Background Shorebirds (Charadriiformes) are considered one of the primary reservoirs of avian influenza. Because these species are highly migratory, there is concern that infected shorebirds may be a mechanism by which highly pathogenic avian influenza virus (HPAIV) H5N1 could be introduced into North America from Asia. Large numbers of dunlin (Calidris alpina) migrate from wintering areas in central and eastern Asia, where HPAIV H5N1 is endemic, across the Bering Sea to breeding areas in Alaska. Low pathogenic avian influenza virus has been previously detected in dunlin, and thus, dunlin represent a potential risk to transport HPAIV to North America. To date no experimental challenge studies have been performed in shorebirds. Methods Wild dunlin were inoculated intranasally and intrachoanally various doses of HPAIV H5N1. The birds were monitored daily for virus excretion, disease signs, morbidity, and mortality. Results The infectious dose of HPAIV H5N1 in dunlin was determined to be 101.7 EID50/100 ??l and that the lethal dose was 101.83 EID50/100 ??l. Clinical signs were consistent with neurotropic disease, and histochemical analyses revealed that infection was systemic with viral antigen and RNA most consistently found in brain tissues. Infected birds excreted relatively large amounts of virus orally (104 EID50) and smaller amounts cloacally. Conclusions Dunlin are highly susceptible to infection with HPAIV H5N1. They become infected after exposure to relatively small doses of the virus and if they become infected, they are most likely to suffer mortality within 3-5days. These results have important implications regarding the risks of transport and transmission of HPAIV H5N1 to North America by this species and raises questions for further investigation. Published 2011. This article is a US Government work and is in the public domain in the USA.

  13. Characterization of the 2012 Highly Pathogenic Avian Influenza H7N3 Virus Isolated from Poultry in an Outbreak in Mexico: Pathobiology and Vaccine Protection

    PubMed Central

    Pantin-Jackwood, Mary; Guzman, Sofia G.; Ricardez, Yadira; Spackman, Erica; Bertran, Kateri; Suarez, David L.; Swayne, David E.

    2013-01-01

    In June of 2012, an H7N3 highly pathogenic avian influenza (HPAI) virus was identified as the cause of a severe disease outbreak in commercial laying chicken farms in Mexico. The purpose of this study was to characterize the Mexican 2012 H7N3 HPAI virus (A/chicken/Jalisco/CPA1/2012) and determine the protection against the virus conferred by different H7 inactivated vaccines in chickens. Both adult and young chickens intranasally inoculated with the virus became infected and died at between 2 and 4 days postinoculation (p.i.). High virus titers and viral replication in many tissues were demonstrated at 2 days p.i. in infected birds. The virus from Jalisco, Mexico, had high sequence similarity of greater than 97% to the sequences of wild bird viruses from North America in all eight gene segments. The hemagglutinin gene of the virus contained a 24-nucleotide insert at the hemagglutinin cleavage site which had 100% sequence identity to chicken 28S rRNA, suggesting that the insert was the result of nonhomologous recombination with the host genome. For vaccine protection studies, both U.S. H7 low-pathogenic avian influenza (LPAI) viruses and a 2006 Mexican H7 LPAI virus were tested as antigens in experimental oil emulsion vaccines and injected into chickens 3 weeks prior to challenge. All H7 vaccines tested provided ≥90% protection against clinical disease after challenge and decreased the number of birds shedding virus and the titers of virus shed. This study demonstrates the pathological consequences of the infection of chickens with the 2012 Mexican lineage H7N3 HPAI virus and provides support for effective programs of vaccination against this virus in poultry. PMID:23760232

  14. The PA and HA gene-mediated high viral load and intense innate immune response in the brain contribute to the high pathogenicity of H5N1 avian influenza virus in mallard ducks.

    PubMed

    Hu, Jiao; Hu, Zenglei; Mo, Yiqun; Wu, Qiwen; Cui, Zhu; Duan, Zhiqiang; Huang, Junqing; Chen, Hongzhi; Chen, Yuxin; Gu, Min; Wang, Xiaoquan; Hu, Shunlin; Liu, Huimou; Liu, Wenbo; Liu, Xiaowen; Liu, Xiufan

    2013-10-01

    Most highly pathogenic avian influenza A viruses cause only mild clinical signs in ducks, serving as an important natural reservoir of influenza A viruses. However, we isolated two H5N1 viruses that are genetically similar but differ greatly in virulence in ducks. A/Chicken/Jiangsu/k0402/2010 (CK10) is highly pathogenic, whereas A/Goose/Jiangsu/k0403/2010 (GS10) is low pathogenic. To determine the genetic basis for the high virulence of CK10 in ducks, we generated a series of single-gene reassortants between CK10 and GS10 and tested their virulence in ducks. Expression of the CK10 PA or hemagglutinin (HA) gene in the GS10 context resulted in increased virulence and virus replication. Conversely, inclusion of the GS10 PA or HA gene in the CK10 background attenuated the virulence and virus replication. Moreover, the PA gene had a greater contribution. We further determined that residues 101G and 237E in the PA gene contribute to the high virulence of CK10. Mutations at these two positions produced changes in virulence, virus replication, and polymerase activity of CK10 or GS10. Position 237 plays a greater role in determining these phenotypes. Moreover, the K237E mutation in the GS10 PA gene increased PA nuclear accumulation. Mutant GS10 viruses carrying the CK10 HA gene or the PA101G or PA237E mutation induced an enhanced innate immune response. A sustained innate response was detected in the brain rather than in the lung and spleen. Our results suggest that the PA and HA gene-mediated high virus replication and the intense innate immune response in the brain contribute to the high virulence of H5N1 virus in ducks.

  15. Highly (H5N1) and Low (H7N2) Pathogenic Avian Influenza Virus Infection in Falcons Via Nasochoanal Route and Ingestion of Experimentally Infected Prey

    PubMed Central

    Bertran, Kateri; Busquets, Núria; Abad, Francesc Xavier; García de la Fuente, Jorge; Solanes, David; Cordón, Iván; Costa, Taiana; Dolz, Roser; Majó, Natàlia

    2012-01-01

    An experimental infection with highly pathogenic avian influenza (HPAI) and low pathogenic avian influenza (LPAI) viruses was carried out on falcons in order to examine the effects of these viruses in terms of pathogenesis, viral distribution in tissues and viral shedding. The distribution pattern of influenza virus receptors was also assessed. Captive-reared gyr-saker (Falco rusticolus x Falco cherrug) hybrid falcons were challenged with a HPAI H5N1 virus (A/Great crested grebe/Basque Country/06.03249/2006) or a LPAI H7N2 virus (A/Anas plathyrhynchos/Spain/1877/2009), both via the nasochoanal route and by ingestion of previously infected specific pathogen free chicks. Infected falcons exhibited similar infection dynamics despite the different routes of exposure, demonstrating the effectiveness of in vivo feeding route. H5N1 infected falcons died, or were euthanized, between 5–7 days post-infection (dpi) after showing acute severe neurological signs. Presence of viral antigen in several tissues was confirmed by immunohistochemistry and real time RT-PCR (RRT-PCR), which were generally associated with significant microscopical lesions, mostly in the brain. Neither clinical signs, nor histopathological findings were observed in any of the H7N2 LPAI infected falcons, although all of them had seroconverted by 11 dpi. Avian receptors were strongly present in the upper respiratory tract of the falcons, in accordance with the consistent oral viral shedding detected by RRT-PCR in both H5N1 HPAI and H7N2 LPAI infected falcons. The present study demonstrates that gyr-saker hybrid falcons are highly susceptible to H5N1 HPAI virus infection, as previously observed, and that they may play a major role in the spreading of both HPAI and LPAI viruses. For the first time in raptors, natural infection by feeding on infected prey was successfully reproduced. The use of avian prey species in falconry husbandry and wildlife rehabilitation facilities could put valuable birds of prey

  16. Highly (H5N1) and low (H7N2) pathogenic avian influenza virus infection in falcons via nasochoanal route and ingestion of experimentally infected prey.

    PubMed

    Bertran, Kateri; Busquets, Núria; Abad, Francesc Xavier; García de la Fuente, Jorge; Solanes, David; Cordón, Iván; Costa, Taiana; Dolz, Roser; Majó, Natàlia

    2012-01-01

    An experimental infection with highly pathogenic avian influenza (HPAI) and low pathogenic avian influenza (LPAI) viruses was carried out on falcons in order to examine the effects of these viruses in terms of pathogenesis, viral distribution in tissues and viral shedding. The distribution pattern of influenza virus receptors was also assessed. Captive-reared gyr-saker (Falco rusticolus x Falco cherrug) hybrid falcons were challenged with a HPAI H5N1 virus (A/Great crested grebe/Basque Country/06.03249/2006) or a LPAI H7N2 virus (A/Anas plathyrhynchos/Spain/1877/2009), both via the nasochoanal route and by ingestion of previously infected specific pathogen free chicks. Infected falcons exhibited similar infection dynamics despite the different routes of exposure, demonstrating the effectiveness of in vivo feeding route. H5N1 infected falcons died, or were euthanized, between 5-7 days post-infection (dpi) after showing acute severe neurological signs. Presence of viral antigen in several tissues was confirmed by immunohistochemistry and real time RT-PCR (RRT-PCR), which were generally associated with significant microscopical lesions, mostly in the brain. Neither clinical signs, nor histopathological findings were observed in any of the H7N2 LPAI infected falcons, although all of them had seroconverted by 11 dpi. Avian receptors were strongly present in the upper respiratory tract of the falcons, in accordance with the consistent oral viral shedding detected by RRT-PCR in both H5N1 HPAI and H7N2 LPAI infected falcons. The present study demonstrates that gyr-saker hybrid falcons are highly susceptible to H5N1 HPAI virus infection, as previously observed, and that they may play a major role in the spreading of both HPAI and LPAI viruses. For the first time in raptors, natural infection by feeding on infected prey was successfully reproduced. The use of avian prey species in falconry husbandry and wildlife rehabilitation facilities could put valuable birds of prey and

  17. Poultry movement networks in Cambodia: implications for surveillance and control of highly pathogenic avian influenza (HPAI/H5N1).

    PubMed

    Van Kerkhove, Maria D; Vong, Sirenda; Guitian, Javier; Holl, Davun; Mangtani, Punam; San, Sorn; Ghani, Azra C

    2009-10-23

    Movement of poultry through markets is potentially important in the circulation and spread of highly pathogenic avian influenza. However little is understood about poultry market chains in Cambodia. We conducted a cross-sectional survey of 715 rural villagers, 123 rural, peri-urban and urban market sellers and 139 middlemen from six provinces and Phnom Penh, to evaluate live poultry movement and trading practices. Direct trade links with Thailand and Vietnam were identified via middlemen and market sellers. Most poultry movement occurs via middlemen into Phnom Penh making live bird wet markets in Phnom Penh a potential hub for the spread of H5N1 and ideal for surveillance and control. PMID:19840671

  18. Morpholino oligomers targeting the PB1 and NP genes enhance the survival of mice infected with highly pathogenic influenza A H7N7 virus.

    PubMed

    Gabriel, Gülsah; Nordmann, Alexandra; Stein, David A; Iversen, Patrick L; Klenk, Hans-Dieter

    2008-04-01

    Peptide-conjugated phosphorodiamidate morpholino oligomers (PPMO) are single-stranded nucleic acid-analogue antisense agents that enter cells readily and can reduce gene expression by steric blocking of complementary RNA (cRNA) sequences. Here, we tested a panel of PPMO designed to target conserved sequences in the RNA genome segments encoding polymerase subunits of a highly pathogenic mouse-adapted influenza A virus (SC35M; H7N7). Three PPMO, targeting the translation start site region of PB1 or NP mRNA or the 3'-terminal region of NP viral RNA (vRNA), potently inhibited virus replication in MDCK cells. Primer extension assays showed that treatment with any of the effective PPMO led to markedly reduced levels of mRNA, cRNA and vRNA. Initially, the potential toxicity of a range of intranasally administered PPMO doses was evaluated, by measuring their effect on body weight of uninfected mice. Subsequently, a non-toxic dosing regimen was used to investigate the effect of various PPMO on SC35M infection in a mouse model. Mice administered intranasal treatment of PPMO targeting the PB1-AUG region or NP vRNA, at 3 mug per dose, given once 3 h before and once 2 days after intranasal infection with 10xLD(50) of SC35M, showed a 2 log(10) reduction of viral titre in the lungs and 50 % survival for the 16 day duration of the experiment, whereas the NP-AUG-targeted PPMO treatment resulted in 30 % survival of an otherwise lethal infection. These data suggest that PPMO provide a useful reagent to investigate influenza virus molecular biology and may constitute a therapeutic strategy against highly pathogenic influenza viruses.

  19. First reported incursion of highly pathogenic notifiable avian influenza A H5N1 viruses from clade 2.3.2 into European poultry.

    PubMed

    Reid, S M; Shell, W M; Barboi, G; Onita, I; Turcitu, M; Cioranu, R; Marinova-Petkova, A; Goujgoulova, G; Webby, R J; Webster, R G; Russell, C; Slomka, M J; Hanna, A; Banks, J; Alton, B; Barrass, L; Irvine, R M; Brown, I H

    2011-02-01

    This study reports the first incursion into European poultry of H5N1 highly pathogenic notifiable avian influenza A (HPNAI) viruses from clade 2.3.2 that affected domestic poultry and wild birds in Romania and Bulgaria, respectively. Previous occurrences in Europe of HPNAI H5N1 in these avian populations have involved exclusively viruses from clade 2.2. This represents the most westerly spread of clade 2.3.2 viruses, which have shown an apparently expanding range of geographical dispersal since mid-2009 following confirmation of infections in wild waterfowl species in Mongolia and Eastern Russia. During March 2010, AI infection was suspected at post-mortem examination of two hens from two backyard flocks in Tulcea Country, Romania. HPNAI of H5N1 subtype was confirmed by reverse transcription polymerase chain reaction (RT-PCR). A second outbreak was confirmed 2 weeks later by RT-PCR, affecting all hens from another flock located 55 km east of the first cluster. On the same day, an H5N1 HPNAI virus was detected from a pooled tissue sample collected from a dead Common Buzzard found on the Black Sea coast in Bulgaria. Detailed genetic characterization of the haemagglutinin gene revealed the cleavage site of the isolates to be consistent with viruses of high pathogenicity belonging to clade 2.3.2 of the contemporary Eurasian H5N1 lineage. Viruses from a clade other than 2.2 have apparently spread to wild birds, with potential maintenance and spread through such populations. Whilst the scale of threat posed by the apparent westward spread of the clade 2.3.2 viruses remains uncertain, ongoing vigilance for clinical signs of disease as part of existing passive surveillance frameworks for AI, and the prompt reporting of suspect cases in poultry is advised.

  20. Serine protease espP subtype alpha, but not beta or gamma, of Shiga toxin-producing Escherichia coli is associated with highly pathogenic serogroups.

    PubMed

    Khan, Abdul Basit; Naim, Asma; Orth, Dorothea; Grif, Katharina; Mohsin, Mashkoor; Prager, Rita; Dierich, Manfred P; Würzner, Reinhard

    2009-04-01

    Besides Shiga toxins (Stx), Stx-producing Escherichia coli (STEC) harbour several other putative virulence factors, including the serine protease EspP. We have investigated 214 STEC strains from Austria belonging to 61 different serotypes from humans, animals, and food for the presence of this serine protease gene and have determined the espP subtypes and their association with clinical outcome. espP was detected in 121 (57%) out of 214 strains. Sixty-five of 68 strains (96%) of non-sorbitol-fermenting (NSF) O157:H7/NM (NM, non-motile) were positive for espP, while none of 8 SF E. coli O157:NM isolates contained this gene. All 9 strains of serotype O145:NM and 17 of 21 strains (81%) of serotype O26:H11/NM were positive for espP. Nineteen STEC serogroups including O103 and O111 serogroups--considered to be highly pathogenic--were completely negative for espP. Only 5 of 12 strains isolated from patients suffering from haemolytic uraemic syndrome (HUS) were espP-positive (all serogroup NSF O157) as well as 28 of 39 strains from patients with bloody diarrhoea, 40 of 63 strains from patients with non-bloody diarrhoea, and 15 of 19 strains from asymptomatic patients. In O157:H7/NM, O26:H11/NM, and O145:NM only espP subtype alpha was found, whereas in most of the other non-O157 serogroups, subtypes beta and gamma were found. Subtype delta was not detected in our strain collection. Regarding the espP subtypes, only subtype alpha, but not beta and gamma, were found in HUS patients. Moreover, we could demonstrate that espP, and in particular subtype alpha, is associated with highly pathogenic serogroups.

  1. Experimentally Infected Domestic Ducks Show Efficient Transmission of Indonesian H5N1 Highly Pathogenic Avian Influenza Virus, but Lack Persistent Viral Shedding

    PubMed Central

    Wibawa, Hendra; Bingham, John; Nuradji, Harimurti; Lowther, Sue; Payne, Jean; Harper, Jenni; Junaidi, Akhmad; Middleton, Deborah; Meers, Joanne

    2014-01-01

    Ducks are important maintenance hosts for avian influenza, including H5N1 highly pathogenic avian influenza viruses. A previous study indicated that persistence of H5N1 viruses in ducks after the development of humoral immunity may drive viral evolution following immune selection. As H5N1 HPAI is endemic in Indonesia, this mechanism may be important in understanding H5N1 evolution in that region. To determine the capability of domestic ducks to maintain prolonged shedding of Indonesian clade 2.1 H5N1 virus, two groups of Pekin ducks were inoculated through the eyes, nostrils and oropharynx and viral shedding and transmission investigated. Inoculated ducks (n = 15), which were mostly asymptomatic, shed infectious virus from the oral route from 1 to 8 days post inoculation, and from the cloacal route from 2–8 dpi. Viral ribonucleic acid was detected from 1–15 days post inoculation from the oral route and 1–24 days post inoculation from the cloacal route (cycle threshold <40). Most ducks seroconverted in a range of serological tests by 15 days post inoculation. Virus was efficiently transmitted during acute infection (5 inoculation-infected to all 5 contact ducks). However, no evidence for transmission, as determined by seroconversion and viral shedding, was found between an inoculation-infected group (n = 10) and contact ducks (n = 9) when the two groups only had contact after 10 days post inoculation. Clinical disease was more frequent and more severe in contact-infected (2 of 5) than inoculation-infected ducks (1 of 15). We conclude that Indonesian clade 2.1 H5N1 highly pathogenic avian influenza virus does not persist in individual ducks after acute infection. PMID:24392085

  2. Experimentally infected domestic ducks show efficient transmission of Indonesian H5N1 highly pathogenic avian influenza virus, but lack persistent viral shedding.

    PubMed

    Wibawa, Hendra; Bingham, John; Nuradji, Harimurti; Lowther, Sue; Payne, Jean; Harper, Jenni; Junaidi, Akhmad; Middleton, Deborah; Meers, Joanne

    2014-01-01

    Ducks are important maintenance hosts for avian influenza, including H5N1 highly pathogenic avian influenza viruses. A previous study indicated that persistence of H5N1 viruses in ducks after the development of humoral immunity may drive viral evolution following immune selection. As H5N1 HPAI is endemic in Indonesia, this mechanism may be important in understanding H5N1 evolution in that region. To determine the capability of domestic ducks to maintain prolonged shedding of Indonesian clade 2.1 H5N1 virus, two groups of Pekin ducks were inoculated through the eyes, nostrils and oropharynx and viral shedding and transmission investigated. Inoculated ducks (n = 15), which were mostly asymptomatic, shed infectious virus from the oral route from 1 to 8 days post inoculation, and from the cloacal route from 2-8 dpi. Viral ribonucleic acid was detected from 1-15 days post inoculation from the oral route and 1-24 days post inoculation from the cloacal route (cycle threshold <40). Most ducks seroconverted in a range of serological tests by 15 days post inoculation. Virus was efficiently transmitted during acute infection (5 inoculation-infected to all 5 contact ducks). However, no evidence for transmission, as determined by seroconversion and viral shedding, was found between an inoculation-infected group (n = 10) and contact ducks (n = 9) when the two groups only had contact after 10 days post inoculation. Clinical disease was more frequent and more severe in contact-infected (2 of 5) than inoculation-infected ducks (1 of 15). We conclude that Indonesian clade 2.1 H5N1 highly pathogenic avian influenza virus does not persist in individual ducks after acute infection.

  3. Evidence for common ancestry among viruses isolated from wild birds in Beringia and highly pathogenic intercontinental reassortant H5N1 and H5N2 influenza A viruses

    USGS Publications Warehouse

    Ramey, Andy M.; Reeves, Andrew; Teslaa, Joshua L.; Nashold, Sean W.; Donnelly, Tyrone F.; Bahl, Justin; Hall, Jeffrey S.

    2016-01-01

    Highly pathogenic clade 2.3.4.4 H5N8, H5N2, and H5N1 influenza A viruses were first detected in wild, captive, and domestic birds in North America in November–December 2014. In this study, we used wild waterbird samples collected in Alaska prior to the initial detection of clade 2.3.4.4 H5 influenza A viruses in North America to assess the evidence for: (1) dispersal of highly pathogenic influenza A viruses from East Asia to North America by migratory birds via Alaska and (2) ancestral origins of clade 2.3.4.4 H5 reassortant viruses in Beringia. Although we did not detect highly pathogenic influenza A viruses in our sample collection from western Alaska, we did identify viruses that contained gene segments sharing recent common ancestry with intercontinental reassortant H5N2 and H5N1 viruses. Results of phylogenetic analyses and estimates for times of most recent common ancestry support migratory birds sampled in Beringia as maintaining viral diversity closely related to novel highly pathogenic influenza A virus genotypes detected in North America. Although our results do not elucidate the route by which highly pathogenic influenza A viruses were introduced into North America, genetic evidence is consistent with the hypothesized trans-Beringian route of introduction via migratory birds.

  4. Internalization of fresh produce by foodborne pathogens.

    PubMed

    Erickson, Marilyn C

    2012-01-01

    Recent studies addressing the internalization of fresh produce by foodborne pathogens arose in response to the growing number of recent and high profile outbreaks involving fresh produce. Because chemical sanitizing agents used during harvest and minimal processing are unlikely to reach enteric pathogens residing within plant tissue, it is imperative that paths for pathogen entry be recognized and minimized. Using both microscopy and microbial enumeration tools, enteric pathogens have been shown to enter plant tissues through both natural apertures (stomata, lateral junctions of roots, flowers) and damaged (wounds, cut surfaces) tissue. In studies revealing preharvest internalization via plant roots or leaf stomata, experimental conditions have primarily involved exposure of plants to high pathogen concentrations (≥ 6 log g⁻¹ soil or 6 log ml⁻¹ water), but those pathogens internalized appear to have short-term persistence. Postharvest internalization of pathogens via cut surfaces may be minimized by maintaining effective levels of sanitizing agents in waters during harvesting and minimal processing.

  5. Human microRNA-24 modulates highly pathogenic avian-origin H5N1 influenza A virus infection in A549 cells by targeting secretory pathway furin.

    PubMed

    Loveday, Emma-Kate; Diederich, Sandra; Pasick, John; Jean, François

    2015-01-01

    A common critical cellular event that many human enveloped viruses share is the requirement for proteolytic cleavage of the viral glycoprotein by furin in the host secretory pathway. For example, the furin-dependent proteolytic activation of highly pathogenic (HP) influenza A (infA) H5 and H7 haemagglutinin precursor (HA0) subtypes is critical for yielding fusion-competent infectious virions. In this study, we hypothesized that viral hijacking of the furin pathway by HP infA viruses to permit cleavage of HA0 could represent a novel molecular mechanism controlling the dynamic production of fusion-competent infectious virus particles during the viral life cycle. We explored the biological role of a newly identified furin-directed human microRNA, miR-24, in this process as a potential post-transcriptional regulator of the furin-mediated activation of HA0 and production of fusion-competent virions in the host secretory pathway. We report that miR-24 and furin are differentially expressed in human A549 cells infected with HP avian-origin infA H5N1. Using miR-24 mimics, we demonstrated a robust decrease in both furin mRNA levels and intracellular furin activity in A549 cells. Importantly, pretreatment of A549 cells with miR-24 mimicked these results: a robust decrease of H5N1 infectious virions and a complete block of H5N1 virus spread that was not observed in A549 cells infected with low-pathogenicity swine-origin infA H1N1 virus. Our results suggest that viral-specific downregulation of furin-directed microRNAs such as miR-24 during the life cycle of HP infA viruses may represent a novel regulatory mechanism that governs furin-mediated proteolytic activation of HA0 glycoproteins and production of infectious virions.

  6. Spatial modeling of wild bird risk factors to investigate highly pathogenic A(H5N1) avian influenza virus transmission

    USGS Publications Warehouse

    Prosser, Diann J.; Hungerford, Laura L.; Erwin, R. Michael; Ottinger, Mary Ann; Takekawa, John Y.; Newman, Scott H.; Xiao, Xianming; Ellis, Erie C.

    2016-01-01

    One of the longest-persisting avian influenza viruses in history, highly pathogenic avian influenza virus (HPAIV) A(H5N1), continues to evolve after 18 years, advancing the threat of a global pandemic. Wild waterfowl (family Anatidae), are reported as secondary transmitters of HPAIV, and primary reservoirs for low-pathogenic avian influenza viruses, yet spatial inputs for disease risk modeling for this group have been lacking. Using GIS and Monte Carlo simulations, we developed geospatial indices of waterfowl abundance at 1 and 30 km resolutions and for the breeding and wintering seasons for China, the epicenter of H5N1. Two spatial layers were developed: cumulative waterfowl abundance (WAB), a measure of predicted abundance across species, and cumulative abundance weighted by H5N1 prevalence (WPR), whereby abundance for each species was adjusted based on prevalence values then totaled across species. Spatial patterns of the model output differed between seasons, with higher WAB and WPR in the northern and western regions of China for the breeding season and in the southeast for the wintering season. Uncertainty measures indicated highest error in southeastern China for both WAB and WPR. We also explored the effect of resampling waterfowl layers from 1 km to 30 km resolution for multi-scale risk modeling. Results indicated low average difference (less than 0.16 and 0.01 standard deviations for WAB and WPR, respectively), with greatest differences in the north for the breeding season and southeast for the wintering season. This work provides the first geospatial models of waterfowl abundance available for China. The indices provide important inputs for modeling disease transmission risk at the interface of poultry and wild birds. These models are easily adaptable, have broad utility to both disease and conservation needs, and will be available to the scientific community for advanced modeling applications.

  7. Assessment of biosecurity measures against highly pathogenic avian influenza risks in small-scale commercial farms and free-range poultry flocks in the northcentral Nigeria.

    PubMed

    Alhaji, N B; Odetokun, I A

    2011-04-01

    There is considerable global concern over the emergence of highly pathogenic avian influenza (HPAI) that has affected domestic poultry flocks in Nigeria and other parts of the world. There have been little investigations on the proposition that free-range flocks are potentially at higher risk of HPAI than confined small-scale commercial enterprises. The objective is to analyse the biosecurity measures instituted in the small-scale commercial poultry farms and established free-range bird flocks owned by households in the rural areas and qualitatively assess the risk status at the two levels of poultry management systems in northcentral Nigeria. We used data collected through questionnaire administration to farms and flock owners and subjected them to a traffic light system model to test for relative risks of HPAI infection based on the biosecurity measures put in place at the farm and flock levels. The results indicate that free-range flocks are at lower risk compared to small-scale commercial operations. These findings are plausible as birds from free-range flocks have more opportunities to contact wild bird reservoirs of low-pathogenic avian influenza (LPAI) strains than small-scale commercial poultry, thus providing them with constant challenge and maintenance of flock immunity. The development of efficient and effective biosecurity measures against poultry diseases on small-scale commercial farms requires adequate placement of barriers to provide segregation, cleaning and disinfection, while concerted community-led sanitary measures are required for free-range poultry flocks in the developing topical and subtropical economies.

  8. Epidemic outbreaks, diagnostics, and control measures of the H5N1 highly pathogenic avian influenza in the Kingdom of Saudi Arabia, 2007-08.

    PubMed

    Lu, Huaguang; Ismail, Mahmoud Moussa; Khan, Owais Ahmed; Al Hammad, Yousef; Abdel Rhman, Salah Shaban; Al-Blowi, Mohamed Hamad

    2010-03-01

    The first outbreak of H5N1 highly pathogenic avian influenza (HPAI) in the Kingdom of Saudi Arabia (KSA) occurred in two "backyard" flocks of Houbara bustards and falcons in February 2007. Subsequent outbreaks were seen through the end of 2007 in "backyard" birds including native chickens, ostriches, turkeys, ducks, and peacocks. From November 2007 through January 2008, H5N1 HPAI outbreaks occurred in 19 commercial poultry premises, including two broiler breeder farms, one layer breeder farm, one ostrich farm, and 15 commercial layer farms, with approximately 4.75 million birds affected. Laboratory diagnosis of all H5N1-positive cases was conducted at the Central Veterinary Diagnostic Laboratory (CVDL) in Riyadh, Saudi Arabia. A combination of diagnostic tests was used to confirm the laboratory diagnosis. A rapid antigen-capture test and real-time reverse transcriptase-PCR (rtRT-PCR) assay on clinical and field specimens were conducted initially. Meanwhile, virus isolation in specific-pathogen-free embryonating chicken eggs was performed and was followed by hemagglutinin (HA) and hemagglutination inhibition tests, then rapid antigen-capture and rtRT-PCR tests on HA-positive allantoic fluid samples. In most HPAI cases, a complete laboratory diagnosis was made within 24-48 hr at the CVDL. Saudi Arabian government officials made immediate decisions to depopulate all H5N1-affected and nonaffected flocks within a 5-km radius area and applied quarantine zones to prevent the virus from spreading to other areas. Other co