scholarly journals Application of the Actiphage® Assay to Detect Viable Mycobacterium avium subsp. paratuberculosis Cells in Fresh Sheep and Goat Milk and Previously Frozen Milk and In-Line Milk Filters

2021 ◽  
Vol 8 ◽  
Author(s):  
Monika Beinhauerova ◽  
Iva Slana
Keyword(s):  
Mycobacterium Avium ◽  
Limit Of Detection ◽  
Goat Milk ◽  
Time Lapse ◽  
Cow Milk ◽  
Economic Losses ◽  
Diagnostic Tools ◽  
Lytic Bacteriophage ◽  
Milk Samples ◽  
Detection And Identification

Mycobacterium avium subsp. paratuberculosis (MAP) is a well-known causative agent of paratuberculosis, a chronic infectious granulomatous enteritis of ruminants contributing to significant economic losses worldwide. Current conventional diagnostic tools are far from being sufficient to manage and control this disease. Therefore, increased attention has been paid to alternative approaches including phage-based assays employing lytic bacteriophage D29 to detect MAP cells. The aim of the present study was to assess the applicability and efficiency of the recently developed phage-based kit termed Actiphage® combined with IS900 real-time PCR (qPCR) for rapid detection and quantification of viable MAP in milk samples. We demonstrated that Actiphage® in combination with IS900 qPCR allows for rapid and sensitive detection and identification of viable MAP in milk samples with a limit of detection of 1 MAP per 50 ml milk. Using this method, the presence of viable MAP cells was successfully determined in 30.77% of fresh goat, sheep and cow milk samples originating from paratuberculosis-affected herds. We further used Actiphage assay to define the time-lapse aspect of testing naturally contaminated milk and milk filters frozen for various lengths of time by phage-based techniques. Viable MAP was detected in 13.04% of frozen milk samples and 28.57% of frozen milk filters using Actiphage-qPCR. The results suggest the ability to detect viable MAP in these samples following freezing for more than 1 year. The obtained results support the views of the beneficial role of this technology in the control or monitoring of paratuberculosis.

scholarly journals A rapid phage assay for detection of viable Mycobacterium avium subsp. paratuberculosis in milk

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
Sepideh Hosseiniporgham ◽  
Lucio Rebechesu ◽  
Pierangela Pintore ◽  
Stefano Lollai ◽  
Maria Dattena ◽  
...  
Keyword(s):  
Magnetic Separation ◽  
Mycobacterium Avium ◽  
Limit Of Detection ◽  
Goat Milk ◽  
Mycobacterium Avium Subsp ◽  
Milk Samples ◽  
Sheep Milk ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Higher Sensitivity

AbstractParatuberculosis is an incurable gastroenteritis among ruminants that is promoted by Mycobacterium avium subsp. paratuberculosis (MAP), an acid-fast mycobacterium. To accelerate the detection of viable pathogen, a conventional (peptide mediated magnetic separation: PMS) and novel (phage-bead qPCR: PBQ) phage based assay was optimized. A superior limit of detection (LOD) of 10 MAP per 10 mL milk was suggested for PBQ compared to 100 cells/10 mL for PMS-phage assay. Via PBQ, viable MAP was found in 48.78% out 41 unpasteurized sheep and goat milk samples. Sheep milk samples (n = 29) that were tested by PMS-phage assay contained no viable MAP. The absence of viable MAP in milk collected from 21 of the recent sheep animals was also confirmed by PBQ after a 2-week gap. Although, the two phage assays comparably detected no viable MAP in the milk samples, MAP DNA and antibodies against MAP were recognized in milk and sera of some of these animals within two instances of sampling representing that some sheep animals were MAP shedders. In conclusion, PBQ and PMS-phage could be promising methods for the assessment of MAP viability in milk samples. However, PBQ was privileged over the PMS-phage assay due to the lower LOD, rapidity, higher sensitivity, lack of need to M. smegmatis and consequent virucidal treatment that are essential in PMS-phage assay for making lawn and inactivation of exogenous mycobacteriophages respectively.

scholarly journals Growth and enterotoxin production of Bacillus cereus in cow, goat, and sheep milk

2014 ◽  
Vol 83 (10) ◽  
pp. S3-S8 ◽  
Author(s):  
Lenka Necidová ◽  
Šárka Bursová ◽  
Alena Skočková ◽  
Bohdana Janštová ◽  
Pavla Prachařová ◽  
...  
Keyword(s):  
Bacillus Cereus ◽  
Storage Temperature ◽  
Goat Milk ◽  
Storage Conditions ◽  
Cow Milk ◽  
Enzyme Linked Immunosorbent Assay ◽  
Plate Method ◽  
Milk Samples ◽  
Sheep Milk ◽  
Enterotoxin Production

The aim of this study was to compare Bacillus cereus growth rates and diarrhoeal enterotoxin production in raw and pasteurized goat, sheep, and cow milk in terms of storage conditions. Milk samples were inoculated with B. cereus (CCM 2010), which produces diarrhoeal enterotoxins. Enterotoxin production was tested by ELISA (Enzyme-Linked Immunosorbent Assay), and the count of B. cereus was determined by the plate method. With raw cow milk, B. cereus growth and enterotoxin production can be completely suppressed; in raw goat and sheep milk, enterotoxin was produced at 22 °C. In pasteurized cow, goat, and sheep milk, the B. cereus count increased under all storage conditions, with more rapid growth being observed at 15 °C (sheep milk) and 22 °C (cow and goat milk). Enterotoxin presence was detected at 15 °C and 22 °C, and with pasteurized cow milk also at 8 °C. Our model experiments have determined that B. cereus multiplication and subsequent enterotoxin production depend on storage temperature and milk type.

scholarly journals Proteolytic and lipolytic potential of Pseudomonas spp. from goat and bovine raw milk

2018 ◽  
Vol 38 (8) ◽  
pp. 1577-1583 ◽  
Author(s):  
José C. Ribeiro Júnior ◽  
Pedro I. Teider Junior ◽  
André L.M. Oliveira ◽  
Edson A. Rios ◽  
Ronaldo Tamanini ◽  
...  
Keyword(s):  
Bovine Milk ◽  
Goat Milk ◽  
Raw Milk ◽  
Species Level ◽  
Cow Milk ◽  
Production Potential ◽  
Gram Negative ◽  
Pseudomonas Spp ◽  
Milk Samples ◽  
The Mean

ABSTRACT: Pseudomonas, the main genus of gram-negative microorganisms isolated from milk, is psychrotrophic, biofilm-forming, and thermo-resistant deteriorating enzyme producers. The aim of this study was to quantify Pseudomonas spp. in goat’s and cow’s milk produced in the Paraná state, Brazil, to evaluate the deteriorating activity of the isolates at mesophilic and psychrotrophic conditions and to identify, at the species level, the isolates with alkaline metalloprotease (aprX gene) production potential. Microbiological, biochemical and molecular methods were used for isolating, confirming and identifying of isolates. The mean counts were 1.6 (±6.3)x104 and 0.89(±3)x102 CFU/mL for goat and bovine milk samples, respectively, immediately after milking. Of the Pseudomonas colonies isolated from goat milk (n=60), 91.7% showed proteolytic potential when incubated at 35°C/48 h and 80% at 7°C/10 days, and lipolytic potential was observed in 95% of the isolates incubated in mesophilic and 78.3% at refrigeration conditions. From the isolates of bovine milk (n=20), 35% showed proteolytic activity only when incubated at 35°C/48 h, and lipolytic potential was observed in 25% of the isolates incubated at 7°C/10d and 35°C/48h. It was observed that 83.3% and 25% of the isolates genetically confirmed as Pseudomonas spp. of goat and bovine milk showed the potential for alkaline metalloprotease production, with the species P. azotoformans, P. koreensis, P. gessardii, P. monteilii and P. lurida being the most frequent in goat milk and P. aeruginosa the only species identified in cow milk.

scholarly journals Mycobacterium avium Subspecies paratuberculosis DNA and Antibodies in Dairy Goat Colostrum and Milk

2019 ◽  
Vol 6 (4) ◽  
pp. 96 ◽  
Author(s):  
Karianne Lievaart-Peterson ◽  
Saskia Luttikholt ◽  
Maaike Gonggrijp ◽  
Robin Ruuls ◽  
Lars Ravesloot ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Mycobacterium Avium Subspecies Paratuberculosis ◽  
Clinical Signs ◽  
Dairy Goat ◽  
Economic Losses ◽  
Dairy Goats ◽  
Convenience Sample ◽  
Milk Samples ◽  
Zoonotic Risk ◽  
Goat Population

Mycobacterium avium subspecies paratuberculosis (MAP) is endemic in the Dutch dairy goat population causing economic loss, and negatively influencing welfare. Moreover, there are concerns about a potential zoonotic risk. Therefore the industry’s objectives are to decrease MAP prevalence, limit economic losses as well as reduce the concentration of MAP in (bulk) milk. To diminish within-farm spread of infection, vaccination, age dependent group housing with separation of newborns from adults, as well as rearing on artificial or treated colostrum and milk replacers are implemented. However, the importance of MAP contaminated colostrum and milk as a route of infection in dairy goat herds is unknown. Therefore the aim of this study was to detect the presence of MAP DNA in colostrum and milk from dairy goats in infected herds. A convenience sample of 120 colostrum samples and 202 milk samples from MAP infected dairy goat herds were tested by IS900 real-time Polymerase Chain Reaction (PCR) for MAP DNA. Furthermore, 22 colostrum samples and 27 post mortem milk samples of goats with clinical signs consistent with paratuberculosis from known infected herds were tested. The majority of samples were from goats vaccinated against MAP. Positive or doubtful PCR results were obtained in none of the 120 and two of the 22 colostrum samples, and in eight of the 202 and four of the 27 milk samples Negative PCR results were obtained in the remaining 140 (99%) colostrum samples and 217 (95%) milk samples.

scholarly journals Rapid Detection and Identification of Nontuberculous Mycobacterial Pathogens in Fish by Using High-Resolution Melting Analysis

2013 ◽  
Vol 79 (24) ◽  
pp. 7837-7845 ◽  
Author(s):  
Thu Nguyet Phung ◽  
Domenico Caruso ◽  
Sylvain Godreuil ◽  
Nicolas Keck ◽  
Tatiana Vallaeys ◽  
...  
Keyword(s):  
High Resolution ◽  
High Resolution Melting ◽  
Limit Of Detection ◽  
High Resolution Melting Analysis ◽  
23S Rrna ◽  
Economic Losses ◽  
Causal Organism ◽  
Content Type ◽  
Detection And Identification ◽  
Melting Analysis

ABSTRACTMycobacterial infections in fish are commonly referred to as piscine mycobacteriosis, irrespectively of the specific identity of the causal organism. They usually cause a chronic disease and sometimes may result in high mortalities and severe economic losses. Nearly 20 species ofMycobacteriumhave been reported to infect fish. Among them,Mycobacterium marinum,M. fortuitum, andM. chelonaeare generally considered the major agents responsible for fish mycobacteriosis. As no quick and inexpensive diagnostic test exists, we tested the potential of high-resolution melting analysis (HRMA) to rapidly identify and differentiate severalMycobacteriumspecies involved in fish infections. By analyzing both the melting temperature and melting profile of the 16S-23S rRNA internal transcribed spacer (ITS), we were able to discriminate 12 different species simultaneously. Sensitivity tests conducted on purifiedM. marinumandM. fortuitumDNA revealed a limit of detection of 10 genome equivalents per reaction. The primers used in this procedure did not lead to any amplification signal with 16 control non-Mycobacteriumspecies, thereby demonstrating their specificity for the genusMycobacterium.

scholarly journals Determination of Ultratrace Bismuth in Milk Samples by Atomic Fluorescence Spectrometry

2003 ◽  
Vol 86 (4) ◽  
pp. 815-822 ◽  
Author(s):  
Patricia Cava-Montesinos ◽  
M Luisa Cervera ◽  
Agustín Pastor ◽  
Miguel de la Guardia
Keyword(s):  
Inductively Coupled Plasma ◽  
Limit Of Detection ◽  
Hydride Generation ◽  
Fluorescence Spectrometry ◽  
Cow Milk ◽  
Atomic Fluorescence Spectrometry ◽  
Atomic Fluorescence ◽  
Microwave Assisted ◽  
Milk Samples

Abstract A sensitive procedure was developed for determination of bismuth (Bi) in milk samples by hydride generation atomic fluorescence spectrometry (HG–AFS) after microwave-assisted sample digestion with HNO3 and H2O2. The method provides a sensitivity of 1832 fluorescence units (ng/mL) with a detection limit of 0.01 ng/mL, which corresponds to 20 pg absolute limit of detection, equivalent to 0.50 ng/g in the original sample. Application of the methodology to cow milk samples from the Spanish market showed the presence of Bi at a concentration of 11.8–28.8 ng/g, which compared well with data obtained after dry ashing of samples and with data obtained by inductively coupled plasma–mass spectrometry after microwave-assisted digestion.

scholarly journals Evaluation of Milk Production and Milk Composition at Different Stages of Saanen Dairy Goats

2021 ◽  
Vol 12 (1S) ◽  
pp. 204-211
Author(s):  
Noor Syaheera Ibrahim ◽  
Farida Hani Ahmad Tajuddin
Keyword(s):  
Milk Production ◽  
Milk Yield ◽  
Goat Milk ◽  
Milk Composition ◽  
Cow Milk ◽  
Local Market ◽  
Dairy Goats ◽  
Food Regulation ◽  
Milk Samples ◽  
Stage Of Lactation

Milk is an important elements due to its high nutritious and balanced in human diet. In Asian, goat milk is the most consumed because of the unique taste, more nutritious than cow milk. However, milk production and composition values differ at every stage of lactation in goats. Thus, determining the highest milk production and producing the best quality goat milk is essential to satisfy the local market demands. The objective of the present study is to determine the milk yield and milk composition at different lactation stages and to evaluate the lactation curves in Saanen goats. The lactation stages can be categorized into three, early (less than 80 days), mid (80 – 140 days) and late (over 140 days) days in milk (DIM). In total, 90 milk samples were collected from 15 goats representing early (n=5), mid (n=5), and late (n=5) stages of lactation. The Saanen goats were raised under the same conditions, and milking was done early in the morning (9.00am).The milk yield from Saanen goats were collected and measured weekly at each stage of lactation. Milk samples were analyzed for fat, protein, solid non-fat, and lactose. Current result illustrated, there were significant different (p ˂ 0.05) among three stages of lactation in milk yield, solid non-fat, fat, protein and lactose. In terms of food regulation, the Saanen goats almost fully met the standard requirements for milk production and composition. In addition, this study also observes the lactation curves in Saanen goats and analyse the curve shapes and patterns. The average milk yields from three different stages develops a concave downward curve. The results indicated that mid yield increased from early to mid- lactation and decreased thereafter until the end of lactation.

scholarly journals Classification and prediction of Mycobacterium Avium subsp. Paratuberculosis (MAP) shedding severity in cattle based on young stock heifer faecal microbiota composition using random forest algorithms

2021 ◽  
Vol 3 (1) ◽  
Author(s):  
Alexander Umanets ◽  
Annemieke Dinkla ◽  
Stephanie Vastenhouw ◽  
Lars Ravesloot ◽  
Ad P. Koets
Keyword(s):  
Random Forest ◽  
Mycobacterium Avium ◽  
Clinical Symptoms ◽  
Classification Model ◽  
Economic Losses ◽  
Diagnostic Tools ◽  
Microbiota Composition ◽  
Faecal Microbiota ◽  
Microbial Composition ◽  
Model Based

Abstract Background Bovine paratuberculosis is a devastating infectious disease caused by Mycobacterium avium subsp. paratuberculosis (MAP). The development of the paratuberculosis in cattle can take up to a few years and vastly differs between individuals in severity of the clinical symptoms and shedding of the pathogen. Timely identification of high shedding animals is essential for paratuberculosis control and minimization of economic losses. Widely used methods for detection and quantification of MAP, such as culturing and PCR based techniques rely on direct presence of the pathogen in a sample and have little to no predictive value concerning the disease development. In the current study, we investigated the possibility of predicting MAP shedding severity in cattle based on the faecal microbiota composition. Twenty calves were experimentally infected with MAP and faecal samples were collected biweekly up to four years of age. All collected samples were subjected to culturing on selective media to obtain data about shedding severity. Faecal microbiota was profiled in a subset of samples (n = 264). Using faecal microbiota composition and shedding intensity data a random forest classifier was built for prediction of the shedding status of the individual animals. Results The results indicate that machine learning approaches applied to microbial composition can be used to classify cows into groups by severity of MAP shedding. The classification accuracy correlates with the age of the animals and use of samples from older individuals resulted in a higher classification precision. The classification model based on samples from the first 12 months of life showed an AUC between 0.78 and 0.79 (95% CI), while the model based on samples from animals older than 24 months showed an AUC between 0.91 and 0.92 (95% CI). Prediction for samples from animals between 12 and 24 month of age showed intermediate accuracy [AUC between 0.86 and 0.87 (95% CI)]. In addition, the results indicate that a limited number of microbial taxa were important for classification and could be considered as biomarkers. Conclusions The study provides evidence for the link between microbiota composition and severity of MAP infection and shedding, as well as lays ground for the development of predictive diagnostic tools based on the faecal microbiota composition.

Keeping Quality and Flavor, and Microorganisms, Proteases and Lipases in Raw Cow and Goat Milk at Collection and After Storage

1983 ◽  
Vol 46 (10) ◽  
pp. 873-877 ◽  
Author(s):  
LESTER HANKIN ◽  
DONALD SHIELDS
Keyword(s):  
Protein Degradation ◽  
Goat Milk ◽  
Raw Milk ◽  
Cow Milk ◽  
Coliform Bacteria ◽  
Microbial Count ◽  
Retail Sale ◽  
Milk Samples ◽  
Significant Difference ◽  
Keeping Quality

Raw cow and goat milks for retail sale in Connecticut were examined at collection and after storage at 4.4 and 7.2°C for 7 d for keeping quality, flavor, microorganisms, protein degradation on storage, and protease and lipase activity. Some milks were bottled at the farm, others were placed in containers supplied by the customer. Goat milk retained a satisfactory flavor significantly longer than cow milk. There was no correlation of keeping quality with any microbial count made at collection except for number of coliform bacteria. Significant differences were found in enzyme activity and protein degradation between cow and goat milk. No significant difference was found between milk bottled at the farm and that collected in sterile containers. Over 82% of raw milk samples met the 30,000 per ml state standard for total aerobic count (SPC) and 72% met the coliform standard of 50 per ml.

Evaluation of methods for Mycobacterium avium ssp. paratuberculosis detection in milk samples from the cattle herd showing low seroprevalence of Johne’s disease

2014 ◽  
Vol 17 (3) ◽  
pp. 459-463 ◽  
Author(s):  
J. Szteyn ◽  
A. Wiszniewska-Łaszczych ◽  
A. Smolińska
Keyword(s):  
Mycobacterium Avium ◽  
Genetic Material ◽  
Johne's Disease ◽  
Reliability Estimation ◽  
Johne’S Disease ◽  
Economic Losses ◽  
Specific Method ◽  
Diagnostic Reliability ◽  
Milk Samples ◽  
Sensitivity Specificity

Abstract Mycobacterium avium ssp. paratuberculosis (MAP) is the cause of chronic gastroenteritis in cattle called the Johne’s disease (JD). The disease causes significant economic losses in cattle production. MAP is also supposed to be involved in the Crohn’s disease and inflammatory bowel disease (IBD) in people. The detection of the cattle infection based on investigations of milk samples and evaluation of the capacity of the methods used to detect the disease was the objective of the present study. Following methods were applied for milk samples testing: detection of MAP in bacterial culture, detection of the specific IS-900 fragment of MAP in the genetic material isolated directly and detection of MAP antibodies. The results obtained were compared with the “golden standard” results, i.e. the isolation of MAP from the faeces. PQStat-the program for diagnostic reliability estimation, was used for evaluation of the sensitivity, specificity and predictive value. The method based on detection of the specific IS-900 fragment of MAP in the genetic material isolated directly from milk samples was found to possess the highest sensitivity. Detection of anti-MAP antibodies on the other hand showed the lowest sensitivity. The method of detecting anti-MAP antibodies in milk was the most specific while detection of the IS-900 fragment in the genetic material was the least specific method. These results obtained may serve as a guide to choose the most appropriate method for diagnosis of MAP infections by milk sample testing.

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