5 research outputs found
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A Hybrid DNA Extraction Method for the Qualitative and Quantitative Assessment of Bacterial Communities from Poultry Production Samples
The efficacy of DNA extraction protocols can be highly dependent upon both the type of sample being investigated and the types of downstream
analyses performed. Considering that the use of new bacterial community analysis techniques (e.g., microbiomics, metagenomics) is becoming
more prevalent in the agricultural and environmental sciences and many environmental samples within these disciplines can be physiochemically
and microbiologically unique (e.g., fecal and litter/bedding samples from the poultry production spectrum), appropriate and effective DNA
extraction methods need to be carefully chosen. Therefore, a novel semi-automated hybrid DNA extraction method was developed specifically
for use with environmental poultry production samples. This method is a combination of the two major types of DNA extraction: mechanical and
enzymatic. A two-step intense mechanical homogenization step (using bead-beating specifically formulated for environmental samples) was
added to the beginning of the “gold standard” enzymatic DNA extraction method for fecal samples to enhance the removal of bacteria and DNA
from the sample matrix and improve the recovery of Gram-positive bacterial community members. Once the enzymatic extraction portion of the
hybrid method was initiated, the remaining purification process was automated using a robotic workstation to increase sample throughput and
decrease sample processing error. In comparison to the strict mechanical and enzymatic DNA extraction methods, this novel hybrid method
provided the best overall combined performance when considering quantitative (using 16S rRNA qPCR) and qualitative (using microbiomics)
estimates of the total bacterial communities when processing poultry feces and litter samples.Keywords: Environmental, DNA extraction, Feces, qPCR, Litter, Microbiomics, Molecular biology, Semi-automated, PoultryKeywords: Environmental, DNA extraction, Feces, qPCR, Litter, Microbiomics, Molecular biology, Semi-automated, Poultr
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Associations between Mycobacterium avium subsp. paratuberculosis antibodies in bulk tank milk, season of sampling and protocols for managing infected cows
BACKGROUND: The objective of this study was to identify associations between the concentration of Mycobacterium avium subsp. paratuberculosis (MAP) antibodies in bulk milk and potential risk factors in herd management and herd characteristics, explaining high MAP antibody titers in milk. An extensive questionnaire was administered to 292 organic and conventional dairy farms from New York, Wisconsin and Oregon. Bulk milk samples were taken from each farm for MAP enzyme-linked immunosorbent assay (ELISA). A general linear model was constructed with MAP ELISA value as the outcome variable and the management factors and herd characteristics as independent variables, while at the same time controlling for the study design variables of state, herd size, and production system (organic or conventional). High bulk tank MAP ELISA value may be due to either a high prevalence of MAP in a herd with many cows contributing to the antibody titer or due to a few infected cows that produce large quantities of antibodies. RESULTS: Results of the regression models indicated that bulk milk ELISA value was associated with season of sampling and the presence or absence of protocols for managing MAP-positive cows. The concentration of MAP antibodies in bulk milk varied seasonally with a peak in the summer and low concentrations in the winter months. When compared to farms that had never observed clinical Johne’s disease, keeping MAP-positive cows or only culling them after a period of delay was associated with an increase in optical density. CONCLUSIONS: The seasonal variation in MAP antibody titers, with a peak in the summer, may be due to a seasonal increase in MAP-bacterial load. Additionally, seasonal calving practices may contribute to seasonal fluctuations in MAP antibody titers in bulk tank milk. Keeping MAP-positive cows increases the antibody titer in bulk milk, likely due to direct antibody production in the infected cow and indirect triggering of antibody production in herdmates.Keywords: Antibodies, ELISA, Cattle, Mycobacteirum avium subsp. paratuberculosis, Bulk-tank milkKeywords: Antibodies, ELISA, Cattle, Mycobacteirum avium subsp. paratuberculosis, Bulk-tank mil
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The characterization of Salmonella enterica serotypes isolated from the scalder tank water of a commercial poultry processing plant: Recovery of a multidrug-resistant Heidelberg strain
The recent multistate outbreak of
a multidrug-resistant (MDR) Salmonella Heidelberg
strain from commercial poultry production highlights
the need to better understand the reservoirs of these
zoonotic pathogens within the commercial poultry production
and processing environment. As part of a larger
study looking at temporal changes in microbial communities
within the major water tanks within a commercial
processing facility, this paper identifies and characterizes
Salmonella enterica isolated from the water in a
final scalder tank at 3 times during a typical processing
day: prior to the birds entering the tank (start),
halfway through the processing day (mid), and after the
final birds were scalded (end). Over 3 consecutive processing
days, no Salmonella were recovered from start-of-day water samples, while a total of 56 Salmonella
isolates were recovered from the mid-day and end-of-day
scalder water samples. Traditional and newer PCR-based
serotyping methods eventually identified these
isolates as either group C3 S. Kentucky (n = 45) and
group B S. Heidelberg (n = 11). While none of the
S. Kentucky isolates possessed any resistances to the
antimicrobials tested, all S. Heidelberg isolates were
found to be multidrug resistant to 5 specific antimicrobials
representing 3 antimicrobial classes. Due to the
potential public health impact of S. Heidelberg and
the recent nationwide poultry-associated outbreak of
multidrug-resistant S. Heidelberg, future studies should
focus on understanding the transmission and environmental
growth dynamics of this serotype within the
commercial poultry processing plant environment.Keywords: S. Heidelberg, Scalder water, Multidrug resistanc