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Serological Evidence of Infectious Laryngotracheitis Infection and Associated Risk Factors in Chickens in Northwestern Ethiopia. ScientificWorldJournal 2022; 2022:6096981. [PMID: 35978862 PMCID: PMC9377982 DOI: 10.1155/2022/6096981] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2022] [Revised: 06/22/2022] [Accepted: 07/09/2022] [Indexed: 11/17/2022] Open
Abstract
Infectious laryngotracheitis (ILT) is a disease of high economic consequence to the poultry sector. Gallid herpesvirus 1 (GaHV-1), a.k.a infectious laryngotracheitis virus (ILTV), under the genus Iltovirus, and the family Herpesviridae, is the agent responsible for the disease. Despite the clinical signs on the field suggestive of ILT, it has long been considered nonexistent and a disease of no concern in Ethiopia. A cross-sectional study was conducted from November 2020 to June 2021 in three selected zones of the Amhara region (Central Gondar, South Gondar, and West Gojjam zones), Ethiopia, with the objective of estimating the seroprevalence of ILTV in chickens and identifying and quantifying associated risk factors. A total of 768 serum samples were collected using multistage cluster sampling and assayed for anti-ILTV antibodies using indirect ELISA. A questionnaire survey was used to identify the potential risk factors. Of the 768 samples, 454 (59.1%, 95% CI: 0.56–0.63) tested positive for anti-ILTV antibodies. Mixed-effect logistic regression analysis of potential risk factors showed that local breeds of chicken were less likely to be seropositive than exotic breeds (OR: 0.38, 95% CI: 0.24–0.61). In addition, factors such as using local feed source (OR: 6.53, 95% CI: 1.77–24.04), rearing chickens extensively (OR: 1.97, 95% CI: 0.78–5.02), mixing of different batches of chicken (OR: 14.51, 95% CI: 3.35–62.77), careless disposal of litter (OR: 1.62, 95% CI: 0.49–4.37), lack of house disinfection (OR: 11.05, 95% CI: 4.09–47.95), lack of farm protective footwear and clothing (OR: 20.85, 95% CI: 5.40–80.45), and careless disposal of dead chicken bodies had all been associated with increased seropositivity to ILTV. Therefore, implementation of biosecurity measures is highly recommended to control and prevent the spread of ILTV. Furthermore, molecular confirmation and characterization of the virus from ILT suggestive cases should be considered to justify the use of ILT vaccines.
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Mo J, Stephens CB, Spackman E. The Thermal Stability of Newcastle Disease Virus in Poultry Litter. Avian Dis 2022; 66:131-134. [PMID: 35451592 DOI: 10.1637/aviandiseases-d-21-00113] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/05/2022] [Accepted: 02/09/2022] [Indexed: 11/05/2022]
Abstract
Sanitary disposal of contaminated organic material during recovery from an animal disease outbreak is costly and laborious. Characterizing the thermal stability of avian paramyxovirus type 1 (APMV-1; virulent APMV-1 strains cause Newcastle disease in poultry) will help inform risk assessments on the presence of viable virus on infected premises or in organic waste from infected premises. In some environments and housing types, heat may also be used as a decontamination method. Therefore, the objective of this study was to characterize the thermal stability (i.e., decimal reduction values [D values]) of APMV-1 in poultry litter. Virus inactivation was evaluated at seven temperatures from 10.0 C through 43.3 C, at 5.5 C intervals (50-110 F in 10 F intervals), using the I2 isolate of APMV-1, a vaccine strain known to be thermally stable. A high titer of virus (approximately 108 50% egg infectious doses) was added to wood shavings based, soiled chicken litter (poultry litter). Litter with both low and high moisture levels were evaluated. Samples were collected at different time intervals, and infectious virus was titrated in embryonated chicken eggs. At high temperatures (37.8 C-43.3 C), infectious virus could not be detected after 2-7 days, whereas at lower temperatures (10 C-21.1 C), it took up to 112 days for virus to decrease to undetectable levels. Furthermore, the D values were almost always shorter in the high moisture litter.
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Affiliation(s)
- Jongseo Mo
- Southeast Poultry Research Laboratory, United States Department of Agriculture-Agricultural Research Service, United States National Poultry Research Center, Athens, GA
| | - Christopher B Stephens
- Southeast Poultry Research Laboratory, United States Department of Agriculture-Agricultural Research Service, United States National Poultry Research Center, Athens, GA
| | - Erica Spackman
- Southeast Poultry Research Laboratory, United States Department of Agriculture-Agricultural Research Service, United States National Poultry Research Center, Athens, GA,
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Gowthaman V, Kumar S, Koul M, Dave U, Murthy TRGK, Munuswamy P, Tiwari R, Karthik K, Dhama K, Michalak I, Joshi SK. Infectious laryngotracheitis: Etiology, epidemiology, pathobiology, and advances in diagnosis and control - a comprehensive review. Vet Q 2021; 40:140-161. [PMID: 32315579 PMCID: PMC7241549 DOI: 10.1080/01652176.2020.1759845] [Citation(s) in RCA: 19] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
Abstract
Infectious laryngotracheitis (ILT) is a highly contagious upper respiratory tract disease of chicken caused by a Gallid herpesvirus 1 (GaHV-1) belonging to the genus Iltovirus, and subfamily Alphaherpesvirinae within Herpesviridae family. The disease is characterized by conjunctivitis, sinusitis, oculo-nasal discharge, respiratory distress, bloody mucus, swollen orbital sinuses, high morbidity, considerable mortality and decreased egg production. It is well established in highly dense poultry producing areas of the world due to characteristic latency and carrier status of the virus. Co-infections with other respiratory pathogens and environmental factors adversely affect the respiratory system and prolong the course of the disease. Latently infected chickens are the primary source of ILT virus (ILTV) outbreaks irrespective of vaccination. Apart from conventional diagnostic methods including isolation and identification of ILTV, serological detection, advanced biotechnological tools such as PCR, quantitative real-time PCR, next generation sequencing, and others are being used in accurate diagnosis and epidemiological studies of ILTV. Vaccination is followed with the use of conventional vaccines including modified live attenuated ILTV vaccines, and advanced recombinant vector vaccines expressing different ILTV glycoproteins, but still these candidates frequently fail to reduce challenge virus shedding. Some herbal components have proved to be beneficial in reducing the severity of the clinical disease. The present review discusses ILT with respect to its current status, virus characteristics, epidemiology, transmission, pathobiology, and advances in diagnosis, vaccination and control strategies to counter this important disease of poultry.
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Affiliation(s)
- Vasudevan Gowthaman
- Poultry Disease Diagnosis and Surveillance Laboratory, Tamil Nadu Veterinary and Animal Sciences University, Namakkal, Tamil Nadu, India
| | - Sachin Kumar
- Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, Assam, India
| | - Monika Koul
- Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, Assam, India
| | - Urmil Dave
- Department of Biosciences and Bioengineering, Indian Institute of Technology Guwahati, Guwahati, Assam, India
| | - T R Gopala Krishna Murthy
- Poultry Disease Diagnosis and Surveillance Laboratory, Tamil Nadu Veterinary and Animal Sciences University, Namakkal, Tamil Nadu, India
| | - Palanivelu Munuswamy
- Division of Pathology, ICAR - Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh, India
| | - Ruchi Tiwari
- Department of Veterinary Microbiology and Immunology, College of Veterinary Sciences, UP Pandit Deen Dayal Upadhayay Pashu Chikitsa Vigyan Vishwavidyalay Evum Go-Anusandhan Sansthan (DUVASU), Mathura, Uttar Pradesh, India
| | - Kumaragurubaran Karthik
- Central University Laboratory, Tamil Nadu Veterinary and Animal Sciences University, Chennai, Tamil Nadu, India
| | - Kuldeep Dhama
- Division of Pathology, ICAR - Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh, India
| | - Izabela Michalak
- Faculty of Chemistry, Department of Advanced Material Technologies, Wrocław University of Science and Technology, Wrocław, Poland
| | - Sunil K Joshi
- Department of Microbiology & Immunology, Department of Pediatrics, Division of Hematology, Oncology and Bone Marrow Transplantation, University of Miami School of Medicine, Miami, Florida, USA
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Figueroa A, Derksen T, Biswas S, Nazmi A, Rejmanek D, Crossley B, Pandey P, Gallardo R. Persistence of low and highly pathogenic avian influenza virus in reused poultry litter, effects of litter amendment use, and composting temperatures. J APPL POULTRY RES 2021. [DOI: 10.1016/j.japr.2020.09.011] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022] Open
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Yegoraw AA, Nazir S, Gerber PF, Walkden-Brown SW. Airborne Transmission of Vaccinal and Wild Type Infectious Laryngotracheitis Virus and Noninfectivity of Extracts of Excreta from Infected Chickens. Avian Dis 2020; 65:30-39. [PMID: 34339119 DOI: 10.1637/aviandiseases-d-20-00073] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2020] [Accepted: 08/27/2020] [Indexed: 11/05/2022]
Abstract
Infectious laryngotracheitis virus (ILTV) is thought to exit the host in respiratory aerosols and enter by inhalation of these. High levels of ILTV DNA have been detected in excreta, raising the possibility of alternative routes of shedding from the host. However, it is not known whether or not the ILTV DNA in excreta represents infective virus. This study investigated transmission of wild type and vaccinal ILTV from infected to susceptible commercial meat chickens. Airborne- and excreta-mediated transmission of two field isolates of ILTV (Classes 9 and 10) and three vaccine strains (SA2, A20, and Serva) were tested. To test airborne transmission, air from isolators containing infected birds was ducted through a paired isolator containing uninfected chickens. To test excreta transmission, aliquots were prepared from excreta containing a high level of ILTV DNA within the first week after infection. Chicks were infected bilaterally by eye drop. Clinical signs were monitored daily and choanal cleft swab samples for ILTV detection by quantitative PCR were collected at 4, 8, 15, 22, and 28 days postinfection (DPI) in the airborne transmission study and at 7 and 14 DPI from the excreta transmission studies. There was no transmission of ILTV from excreta, suggesting that ILTV is inactivated during passage through the gut. All strains of ILTV were transmitted by the airborne route but only to a limited extent for the vaccine viruses. The field viruses induced clinical signs, pathology, and greatly elevated ILTV genome copies in swabs. In summary, these findings confirm the suspected airborne transmission of ILTV, demonstrate differential transmission potential between wild type and vaccine strains by this route, and indicate that excreta is unlikely to be important in the transmission of ILTV and the epidemiology of ILT.
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Affiliation(s)
- Addisu Awukew Yegoraw
- Animal Science, School of Environmental and Rural Science, University of New England, Armidale, NSW, Australia, .,School of Veterinary Medicine, Wolaita Sodo University, Wolaita Sodo, Ethiopia
| | - Shahid Nazir
- Animal Science, School of Environmental and Rural Science, University of New England, Armidale, NSW, Australia
| | - Priscilla F Gerber
- Animal Science, School of Environmental and Rural Science, University of New England, Armidale, NSW, Australia
| | - Stephen W Walkden-Brown
- Animal Science, School of Environmental and Rural Science, University of New England, Armidale, NSW, Australia
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Cockerill SA, Gerber PF, Walkden-Brown SW, Dunlop MW. Suitability of litter amendments for the Australian chicken meat industry. ANIMAL PRODUCTION SCIENCE 2020. [DOI: 10.1071/an19587] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
Abstract
The Australian chicken meat indutstry is rapidly expanding due to the increasing consumption of chicken meat. As a result, the industry has growing issues of sourcing new bedding materials and disposing of spent litter, which can be attributed, in part, to a lack of widespread litter re-use for rearing chickens. According to insights and perspectives recently gathered from industry stakeholders, it is believed that re-using litter will become more common in the future, so as to reduce production costs and ease pressures on both the supply of new bedding materials and disposal of spent litter. However, there are potential risks that need to be addressed if litter re-use increases, particularly with regard to the production and mitigation of ammonia, which can negatively affect chicken health if not managed correctly. The present review discusses the potential benefits reported for different types of litter amendments, which have the primary goal of reducing ammonia volatilisation, but may also contribute to improvements in bird performance, welfare, pathogen loads, fertiliser value of spent litter, and reduced costs associated with purchasing new bedding materials. Acidifiers have been shown to be the most effective of all amendment types, with sodium bisulfate or alum being among the most commonly tested products mentioned in research literature. Litter amendments are currently rarely used in Australia, but it is hoped that the information provided in the present review, based mostly on overseas usage and research, will help inform future decision-making on the use of these products in Australian poultry production systems.
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Stephens CB, Spackman E. Thermal Inactivation of avian influenza virus in poultry litter as a method to decontaminate poultry houses. Prev Vet Med 2017; 145:73-77. [PMID: 28903878 DOI: 10.1016/j.prevetmed.2017.06.012] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/24/2017] [Revised: 06/27/2017] [Accepted: 06/28/2017] [Indexed: 11/29/2022]
Abstract
Removal of contaminated material from a poultry house during recovery from an avian influenza virus (AIV) outbreak is costly and labor intensive. Because AIV is not environmentally stable, heating poultry houses may provide an alternative disinfection method. The objective was to determine the time necessary to inactivate AIV in poultry litter at temperatures achievable in a poultry house. Low pathogenic (LP) AIV inactivation was evaluated between 10.0°-48.9°C, at ∼5.5°C intervals and highly pathogenic (HP) AIV inactivation was evaluated between 10.0°-43.3°C, at ∼11°C intervals. Samples were collected at numerous time points for each temperature. Virus isolation in embryonating chicken eggs was conducted to determine if viable virus was present. Each sample was also tested by real-time RT-PCR. Low pathogenicity AIV was inactivated at 1day at 26.7°C or above. At 10.0, 15.6 and 21.1°C, inactivation times increased to 2-5days. Highly pathogenic AIV followed a similar trend; the virus was inactivated after 1day at 43.3°C and 32.2°C, and required 2 and 5days for inactivation at 21.1°C and 10.0°C respectively. While low pathogenicity AIV appeared to be inactivated at a lower temperature than high pathogenicity AIV, this was not due to any difference in the strains, but due to fewer temperature points being evaluated for high pathogenicity. Endpoints for detection by real-time RT-PCR were not found even weeks after the virus was inactivated. This provides a guideline for the time required, at specific temperatures to inactivate AIV in poultry litter and likely on surfaces within the house. Heat treatment will provide an added level of safety to personnel and against further spread by eliminating infectious virus prior to cleaning a house.
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Affiliation(s)
- Christopher B Stephens
- Exotic and Emerging Avian Viral Diseases Unit, Southeast Poultry Research Laboratory, U.S. National Poultry Research Center, U.S. Department of Agriculture, Agricultural Research Service, Athens, GA, USA
| | - Erica Spackman
- Exotic and Emerging Avian Viral Diseases Unit, Southeast Poultry Research Laboratory, U.S. National Poultry Research Center, U.S. Department of Agriculture, Agricultural Research Service, Athens, GA, USA.
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Roy P, Fakhrul Islam AFM, Burgess SK, Hunt PW, McNally J, Walkden-Brown SW. Real-time PCR quantification of infectious laryngotracheitis virus in chicken tissues, faeces, isolator-dust and bedding material over 28 days following infection reveals high levels in faeces and dust. J Gen Virol 2016; 96:3338-3347. [PMID: 26294959 DOI: 10.1099/jgv.0.000268] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022] Open
Abstract
Infectious laryngotracheitis (ILT) is an important disease of chickens caused by ILT virus (ILTV). We used the Australian SA2 and A20 vaccine strains of ILTV to determine tissue distribution and excretion characteristics of ILTV in specific-pathogen-free chickens and to determine whether ILTV is readily detectable in environmental samples such as faeces, bedding material and dust using real-time quantitative PCR. Three groups of 10 freshly hatched chicks were placed in isolators and infected orally with high doses of the two strains of vaccine virus or left unchallenged as controls. Over a 28-day post-infection (p.i.) period, faecal and serum samples were collected at frequent intervals from six individually identified chickens in each group. Dust and litter samples from the isolators were collected less frequently. Tissue samples were collected from three to four sacrificed or dead/euthanized birds at 6, 14 and 28 days p.i. Infection resulted in clinical ILT, a pronounced antibody response and sustained qPCR detection of the viral genome in the trachea, Harderian gland, lung and kidney up to 28 days p.i. A high level of the viral genome was also detected in faeces between 2 and 7 days p.i., declining by about approximately four orders of magnitude to low, but detectable, levels at 21 and 28 days p.i. The finding of high-level shedding of ILTV in faeces warrants further investigation into the epidemiological role of this, and the sustained high levels of ILTV observed in dust suggest that it may be a useful sample material for monitoring ILTV status in flocks.
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Affiliation(s)
- Parimal Roy
- Animal Science, School of Environmental and Rural Science, University of New England, Armidale, NSW 2351, Australia.,Central University Laboratory, Tamilnadu Veterinary and Animal Sciences University, Chennai 600051, India
| | - A F M Fakhrul Islam
- Animal Science, School of Environmental and Rural Science, University of New England, Armidale, NSW 2351, Australia
| | - Susan K Burgess
- Animal Science, School of Environmental and Rural Science, University of New England, Armidale, NSW 2351, Australia
| | - Peter W Hunt
- CSIRO FD McMaster Laboratory, Locked Bag 1, Armidale, NSW 2351 Australia
| | - Jody McNally
- CSIRO FD McMaster Laboratory, Locked Bag 1, Armidale, NSW 2351 Australia
| | - Stephen W Walkden-Brown
- Animal Science, School of Environmental and Rural Science, University of New England, Armidale, NSW 2351, Australia
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Dormitorio TV, Giambrone JJ, Macklin KS. Detection and isolation of infectious laryngotracheitis virus on a broiler farm after a disease outbreak. Avian Dis 2014; 57:803-7. [PMID: 24597126 DOI: 10.1637/10544-032913-resnote.1] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/05/2022]
Abstract
A broiler farm in North Alabama suffered a mild infectious laryngotracheitis (ILT) outbreak, as determined by clinical disease and PCR. The poultry integrator sought help to control further outbreaks in subsequent flocks. Samples were collected from various areas of the poultry houses on the farm over an 8-wk period. The first sampling was conducted 8 days after the infected farm was depopulated; the second was conducted 2 days prior to subsequent flock placement; and the third was conducted when the new flock was 5 wk of age. Samples were examined for ILT virus (ILTV) DNA by real-time PCR and virus isolation in embryos. The infected houses were cleaned, disinfected, heated, litter composted, and curtains replaced after the first sampling and prior to placement of the next flock. Samples from all periods were positive for ILTV DNA. However, the number of positive samples and crossing point values indicated a decrease in the amount of viral DNA, while virus isolation in embryos was successful only on the first sampling. The subsequent flock was vaccinated against ILTV by in ovo route using a commercial recombinant vaccine. Cleaning and sanitation after the disease outbreak reduced the amount of ILTV on the farm and together with in ovo vaccination of the new flock may have prevented a recurrence of another ILT outbreak.
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Affiliation(s)
| | - Joseph J Giambrone
- Poultry Science Department, Auburn University, Auburn, AL 36849-5416, USA
| | - Kenneth S Macklin
- Poultry Science Department, Auburn University, Auburn, AL 36849-5416, USA
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Ou SC, Giambrone JJ. Infectious laryngotracheitis virus in chickens. World J Virol 2012; 1:142-9. [PMID: 24175219 PMCID: PMC3782274 DOI: 10.5501/wjv.v1.i5.142] [Citation(s) in RCA: 46] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/08/2011] [Revised: 06/12/2012] [Accepted: 09/07/2012] [Indexed: 02/05/2023] Open
Abstract
Infectious laryngotracheitis (ILT) is an important respiratory disease of chickens and annually causes significant economic losses in the poultry industry world-wide. ILT virus (ILTV) belongs to alphaherpesvirinae and the Gallid herpesvirus 1 species. The transmission of ILTV is via respiratory and ocular routes. Clinical and post-mortem signs of ILT can be separated into two forms according to its virulence. The characteristic of the severe form is bloody mucus in the trachea with high mortality. The mild form causes nasal discharge, conjunctivitis, and reduced weight gain and egg production. Conventional polymerase chain reaction (PCR), nested PCR, real-time PCR, and loop-mediated isothermal amplification were developed to detect ILTV samples from natural or experimentally infected birds. The PCR combined with restriction fragment length polymorphism (RFLP) can separate ILTVs into several genetic groups. These groups can separate vaccine from wild type field viruses. Vaccination is a common method to prevent ILT. However, field isolates and vaccine viruses can establish latent infected carriers. According to PCR-RFLP results, virulent field ILTVs can be derived from modified-live vaccines. Therefore, modified-live vaccine reversion provides a source for ILT outbreaks on chicken farms. Two recently licensed commercial recombinant ILT vaccines are also in use. Other recombinant and gene-deficient vaccine candidates are in the developmental stages. They offer additional hope for the control of this disease. However, in ILT endemic regions, improved biosecurity and management practices are critical for improved ILT control.
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Affiliation(s)
- Shan-Chia Ou
- Shan-Chia Ou, Joseph J Giambrone, Department of Poultry Science, Auburn University, Auburn, AL 36849, United States
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Ou SC, Giambrone J, Macklin K. Detection of infectious laryngotracheitis virus from darkling beetles and their immature stage (lesser mealworms) by quantitative polymerase chain reaction and virus isolation. J APPL POULTRY RES 2012. [DOI: 10.3382/japr.2010-00314] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
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Ou S, Giambrone J, Macklin K. Infectious laryngotracheitis vaccine virus detection in water lines and effectiveness of sanitizers for inactivating the virus. J APPL POULTRY RES 2011. [DOI: 10.3382/japr.2010-00300] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
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Barker K, Purswell J, Davis J, Parker H, Kidd M, McDaniel C, Kiess A. Distribution of Bacteria at Different Poultry Litter Depths1. ACTA ACUST UNITED AC 2009. [DOI: 10.3923/ijps.2010.10.13] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022]
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