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1.
J Appl Microbiol ; 131(1): 146-154, 2021 Jul.
Article in English | MEDLINE | ID: mdl-33151641

ABSTRACT

AIM: The focus of the present study was to evaluate the copper ions treatment on the viability of Mycobacterium avium subsp. paratuberculosis (MAP) and other bacterial communities in cow's milk. METHODS AND RESULTS: A copper ions treatment was evaluated in naturally contaminated cow's milk to assay MAP load and/or viability, and relative abundance of other bacterial communities. In addition, physical-chemical analyses of the milk were also performed. All analyses were carried out before and after a copper ions treatment. After copper ions treatment, pH and copper concentration markedly increased in milk; the numbers of viable MAP significantly decreased. The relative abundance of the four target phyla decreased, with the phyla Bacteroidetes and Firmicutes surviving treatment in higher proportions (4 and 2Ā·1% of original populations, respectively). A progressively higher percentage of dead bacterial cells after 5 and 20Ā min copper ions treatments was found (12 and 35%, respectively). CONCLUSION: With the exception of some MAP-tolerant strains, we have once again demonstrated that copper ions have a significant inactivating effect on MAP as well as certain other bacterial communities found in naturally contaminated cow's milk. SIGNIFICANCE AND IMPACT OF THE STUDY: This study showed a significant inactivation of both MAP and other bacteria by copper ions in raw cow's milk, information that could be useful as a tool for MAP control.


Subject(s)
Bacteria/drug effects , Copper/pharmacology , Food Contamination/prevention & control , Milk/microbiology , Mycobacterium avium subsp. paratuberculosis/drug effects , Animals , Bacterial Load , Cattle , Female , Food Microbiology , Hydrogen-Ion Concentration , Ions/pharmacology , Paratuberculosis/microbiology , Time Factors
2.
BMC Microbiol ; 18(1): 172, 2018 11 01.
Article in English | MEDLINE | ID: mdl-30382823

ABSTRACT

BACKGROUND: Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of paratuberculosis, a contagious infectious disease that affects domestic and wild ruminants causing chronic inflammation of the intestine. MAP has proven to be very resistant to both physical and chemical processes, making it difficult to control this pathogen. Based on the recognized antimicrobial properties of copper, the objective of this study was to evaluate the effectiveness of copper ions to reduce MAP numbers and/or MAP viability in a fluid matrix. Besides, methicillin-resistant Staphylococcus aureus (MRSA), and Escherichia coli were used as controls of the effectiveness of copper ions. MAP-spiked PBS was subjected to copper ions treatment at 24Ā V for 5Ā min and the PBS suspensions were sampled before and after treatment. MAP viability and quantification were determined using three complementary techniques: a phage amplification assay, MGIT culture and qPCR. RESULTS: Moderate numbers (103Ā CFUĀ ml-1) of the two control bacteria were completely eliminated by treatment with copper ions. For MAP, copper ions treatment reduced both the viability and numbers of this pathogen. Phage assay information quickly showed that copper ions (24Ā V for 5Ā min) resulted in a significant reduction in viable MAP. MGIT culture results over time showed statistically significant differences in time-to-detection (TTD) values between PRE and POST treatment. MAP genome equivalent estimates for PBS suspensions indicated that MAP numbers were lower in samples POST-treatment with copper ions than PRE-treatment. CONCLUSIONS: The use of copper ions resulted in a significant reduction of MAP in a liquid matrix, although some MAP survival on some occasions was observed.


Subject(s)
Copper/pharmacology , Microbial Viability/drug effects , Mycobacterium avium subsp. paratuberculosis/drug effects , Bacteriophages/drug effects , Bacteriophages/genetics , Buffers , Colony Count, Microbial , Escherichia coli/drug effects , Genome, Bacterial/genetics , Methicillin-Resistant Staphylococcus aureus/drug effects , Mycobacterium avium subsp. paratuberculosis/genetics , Real-Time Polymerase Chain Reaction
3.
J Appl Microbiol ; 122(5): 1357-1367, 2017 May.
Article in English | MEDLINE | ID: mdl-28235155

ABSTRACT

AIM: To validate an optimized peptide-mediated magnetic separation (PMS)-phage assay for detection of viable Mycobacterium avium subsp. paratuberculosis (MAP) in milk. METHODS AND RESULTS: Inclusivity, specificity and limit of detection 50% (LOD50 ) of the optimized PMS-phage assay were assessed. Plaques were obtained for all 43 MAP strains tested. Of 12 other Mycobacterium sp. tested, only Mycobacterium bovis BCG produced small numbers of plaques. LOD50 of the PMS-phage assay was 0Ā·93 MAP cells per 50Ā ml milk, which was better than both PMS-qPCR and PMS-culture. When individual milks (nĀ =Ā 146) and bulk tank milk (BTM, nĀ =Ā 22) obtained from Johne's affected herds were tested by the PMS-phage assay, viable MAP were detected in 31 (21Ā·2%) of 146 individual milks and 13 (59Ā·1%) of 22 BTM, with MAP numbers detected ranging from 6-948 plaque-forming-units per 50Ā ml milk. PMS-qPCR and PMS-MGIT culture proved to be less sensitive tests than the PMS-phage assay. CONCLUSIONS: The optimized PMS-phage assay is the most sensitive and specific method available for the detection of viable MAP in milk. Further work is needed to streamline the PMS-phage assay, because the assay's multistep format currently makes it unsuitable for adoption by the dairy industry as a screening test. SIGNIFICANCE AND IMPACT OF THE STUDY: The inclusivity (ability to detect all MAP strains), specificity (ability to detect only MAP) and detection sensitivity (ability to detect low numbers of MAP) of the optimized PMS-phage assay have been comprehensively demonstrated for the first time.


Subject(s)
Bacteriophages/physiology , Biological Assay/methods , Milk/microbiology , Mycobacterium avium subsp. paratuberculosis/isolation & purification , Paratuberculosis/microbiology , Animals , Bacterial Typing Techniques/methods , Food Contamination/analysis , Limit of Detection , Mycobacterium avium subsp. paratuberculosis/classification , Mycobacterium avium subsp. paratuberculosis/genetics , Mycobacterium avium subsp. paratuberculosis/virology , Peptides/chemistry , Real-Time Polymerase Chain Reaction , Sensitivity and Specificity
4.
J Appl Microbiol ; 122(3): 817-828, 2017 Mar.
Article in English | MEDLINE | ID: mdl-27992094

ABSTRACT

AIM: To develop a multiplex loop-mediated isothermal amplification (LAMP) assay capable of quantifying Escherichia coli and differentiating verocytotoxigenic E.Ā coli (VTEC). METHODS AND RESULTS: Primer sets were selected to amplify the phoA gene (all E.Ā coli strains) and stx1 and/or stx2 genes (VTEC strains only). LAMP calibration curves demonstrated good quantification capability compared with conventional culture. The limits of detection 50% (LOD50 ) of the multiplex LAMP assay were 2Ā·8 (95% CI 2Ā·4-3Ā·3), 3Ā·2 (95% CI 2Ā·5-3Ā·9) and 2Ā·8-3Ā·2 (95% CI 2Ā·1-3Ā·5) log CFU per g for the phoA, stx1 and stx2 genes, respectively. When validated by testing retail beef and bovine faeces samples, good correlation between E.Ā coli counts indicated by the LAMP assay and culture was observed; however, false-negative LAMP assay results were obtained for 12Ā·5-14Ā·7% of samples. CONCLUSIONS: A rapid, multiplex LAMP assay for direct quantification of E.Ā coli and specific detection of VTEC in beef and faeces was successfully developed. Further optimisation of the assay would be needed to improve detection sensitivity. SIGNIFICANCE AND IMPACT OF THE STUDY: The multiplex LAMP assay represents a rapid alternative to culture for monitoring E.Ā coli levels on beef for hygiene monitoring purposes, and, potentially, a method for detection of VTEC in beef and faeces.


Subject(s)
Bacterial Toxins/genetics , Escherichia coli/genetics , Feces/microbiology , Red Meat/microbiology , Animals , Cattle , DNA Primers , Escherichia coli/classification , Escherichia coli/pathogenicity , Limit of Detection , Nucleic Acid Amplification Techniques/methods , Virulence Factors/genetics
5.
J Appl Microbiol ; 115(3): 808-17, 2013 Sep.
Article in English | MEDLINE | ID: mdl-23742225

ABSTRACT

AIMS: The objective of this study was to develop a novel screening method for detection of viable Mycobacterium avium subsp. paratuberculosis (Map) in milk and faeces, as a rapid alternative to Map culture. METHODS AND RESULTS: The new method couples Map-specific peptide-mediated magnetic separation technique with an optimized phage amplification assay followed by detection of released progeny phage by ELISA in a competition assay format using polyclonal antibody produced against the D29 mycobacteriophage involved in the phage assay. Sample matrices were found not to interfere with the developed method, and the dynamic range of the assay was 3Ā Ć—Ā 10(2) -6Ā Ć—Ā 10(8Ā ) phageĀ ml(-1) . When low numbers of Map were present (10(2) Ā CFUĀ ml(-1) ), the burst size of a single host Map cell was maximal (10(3) phage per cell) resulting in a highly sensitive screening assay. CONCLUSION: A rapid, sensitive immuno-based screening method suitable for the detection of viable Map in milk and faeces was developed. SIGNIFICANCE AND IMPACT OF THE STUDY: The novel PMS-phage-ELISA permits sensitive, qualitative detection of viable Map in milk or faeces samples within 48Ā h, representing a substantial decrease in time to detection compared with current culture methods for Map.


Subject(s)
Enzyme-Linked Immunosorbent Assay/methods , Mycobacterium avium subsp. paratuberculosis/isolation & purification , Animals , Feces/microbiology , Milk/microbiology , Mycobacteriophages/immunology
6.
J Appl Microbiol ; 115(1): 271-81, 2013 Jul.
Article in English | MEDLINE | ID: mdl-23551658

ABSTRACT

AIMS: The objectives of this study were to produce Salmonella-specific peptide ligands by phage display biopanning and evaluate their use for magnetic separation (MS). METHODS AND RESULTS: Four-phage display biopanning rounds were performed, and the peptides expressed by the two most Salmonella-specific (on the basis of phage-binding ELISA results) phage clones, MSal020401 and MSal020417, were chemically synthesized and coupled to MyOne™ tosylactivated Dynabeads(Ā®). Peptide capture capability for whole Salmonella cells from nonenriched broth cultures was quantified by MS + plate counts and MS + Greenlight™ detection and compared to capture capability of anti-Salmonella (antibody-coated) Dynabeads(Ā®). MS + Greenlight™ gave a more comprehensive picture of capture capability than MS + plate counts and showed that Peptide MSal020417-coated beads exhibited at least similar, if not better, capture capability to anti-Salmonella Dynabeads(Ā®) (mean capture values of 36Ā·0 Ā± 18Ā·2 and 31Ā·2 Ā± 20Ā·1%, respectively, over Salmonella spp. concentration range 3 Ɨ 10(1) -3 Ɨ 10(6) CFU ml(-1)) with cross-reactivity of ≤1Ā·9% to three other foodborne pathogens: Escherichia coli, Listeria monocytogenes and Campylobacter jejuni. CONCLUSIONS: One of the phage display-derived peptide ligands was demonstrated by MS + Greenlight™ to be a viable antibody alternative for MS of Salmonella spp. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates an antibody-free approach to Salmonella detection and opens substantial possibilities for more rapid tests for this bacterium.


Subject(s)
Cell Surface Display Techniques , Peptides/metabolism , Salmonella/isolation & purification , Ligands , Magnetic Phenomena , Peptides/chemistry
7.
Transbound Emerg Dis ; 65(3): 719-726, 2018 Jun.
Article in English | MEDLINE | ID: mdl-29250933

ABSTRACT

Controlling the spread of Johne's disease, caused by Mycobacterium avium subsp. paratuberculosis (MAP), in domestic livestock is challenging. Current diagnostic methods lack sufficient sensitivity to detect subclinically infected animals, and thus, better diagnostic methods are needed. This study was carried out to investigate the diagnostic potential of two novel peptide-mediated magnetic separation (PMS)-based tests-a PMS-phage assay and PMS-culture-both of which have been developed and optimized to detect viable MAP cells in bovine milk. Individual milk samples (50Ā ml) were obtained from 105 "non-infected" and 40 "MAP-infected" animals (classified as such on the basis of prior faecal culture and serum-ELISA results) in three dairy herds and tested in parallel by the PMS-phage assay and PMS-culture. Diagnostic sensitivity (DSe) and specificity (DSp) of the PMS-phage and PMS-culture methods were determined relative to the MAP infection status of the animal contributing the milk sample. The PMS-based tests applied individually showed moderate DSe (PMS-culture 0.250 and PMS-phage assay 0.325) and high DSp (0.962 and 1.000, respectively). When results of the two PMS-based tests were combined, DSe increased substantially to 0.525, and the DSp was calculated to be 0.962. It was concluded that combined application of the PMS-phage assay and PMS-culture provided the most complete picture regarding the presence of viable MAP in bovine milk samples. A comprehensive validation of the PMS-based assays relative to currently used diagnostic methods (faecal culture and serum-ELISA) would be the next step in assessment of the diagnostic potential of these novel PMS-based methods.


Subject(s)
Biological Assay/methods , Milk/microbiology , Mycobacterium avium subsp. paratuberculosis/isolation & purification , Animals , Bacteriophages/physiology , Cattle , Food Contamination/analysis , Mycobacterium avium subsp. paratuberculosis/genetics , Paratuberculosis/microbiology , Peptides/chemistry , Sensitivity and Specificity
8.
J Dairy Sci ; 90(8): 3590-5, 2007 Aug.
Article in English | MEDLINE | ID: mdl-17638968

ABSTRACT

A total of 143 raw milk cheese samples (soft cheese, n = 9; semihard cheese, n = 133; hard cheese, n = 1), collected at the retail level throughout Switzerland, were tested for Mycobacterium avium ssp. paratuberculosis (MAP) by immunomagnetic capture plus culture on 7H10-PANTA medium and in supplemented BAC-TEC 12B medium, as well as by an F57-based real-time PCR system. Furthermore, pH and water activity values were determined for each sample. Although no viable MAP cells could be cultured, 4.2% of the raw milk cheese samples tested positive with the F57-based real-time PCR system, providing evidence for the presence of MAP in the raw material. As long as the link between MAP and Crohn's disease in humans remains unclear, measures designed to minimize public exposure should also include a focus on milk products.


Subject(s)
Cheese/microbiology , Food Contamination/analysis , Food Microbiology , Mycobacterium avium subsp. paratuberculosis/isolation & purification , Animals , Hydrogen-Ion Concentration , Immunomagnetic Separation/instrumentation , Immunomagnetic Separation/methods , Microspheres , Polymerase Chain Reaction/methods , Prevalence , Switzerland , Water/analysis
9.
Eur J Cancer ; 28A(8-9): 1392-5, 1992.
Article in English | MEDLINE | ID: mdl-1515255

ABSTRACT

Immediately before first hemi-body irradiation, 59 patients with relapsed multiple myeloma were randomised to receive or not to receive subsequent alpha-2b interferon maintenance. 13 patients (22%) [8 of 31 (26%) controls, 5 of 28 (18%) in the interferon arm] received single hemi-body irradiation alone due to progressive disease and/or persistent cytopoenias following the initial procedure. Mean time between upper and lower hemi-body irradiation was 69 days (range 35-294). Of 23 patients randomised to receive interferon and completing double hemi-body irradiation, 15 (65%) achieved peripheral blood counts adequate to allow interferon administration as per study criteria commencing at a mean 116 days (61-241) from time of study entry. The mean period of interferon therapy, starting at a mean 65 days (26-160) post second hemi-body irradiation, is 16.4 months (2-33.5). There was no significant difference in median survival durations (10 months) from time of initial radiotherapy between control and interferon patients.


Subject(s)
Interferon-alpha/therapeutic use , Multiple Myeloma/radiotherapy , Neoplasm Recurrence, Local/radiotherapy , Aged , Aged, 80 and over , Combined Modality Therapy , Female , Humans , Interferon alpha-2 , Male , Middle Aged , Multiple Myeloma/mortality , Multiple Myeloma/therapy , Neoplasm Recurrence, Local/drug therapy , Prospective Studies , Recombinant Proteins , Survival Rate
10.
QJM ; 97(4): 199-204, 2004 Apr.
Article in English | MEDLINE | ID: mdl-15028849

ABSTRACT

BACKGROUND: Non-neuronopathic (type 1) Gaucher's disease, a recessive disorder caused by glucocerebrosidase deficiency, shows marked variability in the severity and extent of clinical expression: many individuals who harbour two mutant alleles remain mildly affected or asymptomatic. Despite much effort, it is not possible accurately to predict disease severity from the genotype, or to identify those patients destined to develop severe disease and meriting early treatment. AIM: To determine the degree to which variance in Gaucher disease is determined by non-heritable factors. DESIGN: Case reports of monozygotic and dizygotic twin pairs. RESULTS: For the monozygotic twin pair, homozygous for the frequent N370S glucocerebrosidase allele, there was no evidence that significant lipid storage was ever initiated in the unaffected twin. In contrast, pathological storage of glucocerebroside has been present in the macrophages of both members of the dizygotic twin pair (compound heterozygotes for the N370S and L444P alleles) from an early age but, by the age of 57 years, only one has developed symptoms. DISCUSSION: Non-heritable factors influence Gaucher disease expression in genetically predisposed individuals. Understanding the interactions between heritable and non-heritable factors will be critical for an analysis of pathogenesis, and the treatment of individuals predisposed to Gaucher disease.


Subject(s)
Diseases in Twins/genetics , Gaucher Disease/genetics , Glucosylceramidase/genetics , Adult , Aged , Alleles , Female , Humans , Male , Middle Aged , Phenotype , Risk Factors , Twins, Dizygotic/genetics , Twins, Monozygotic/genetics
11.
Leuk Lymphoma ; 3(2): 103-7, 1990.
Article in English | MEDLINE | ID: mdl-27457295

ABSTRACT

Recombinant alpha 2a, 2b and 2c interferons have been shown to be effective short-term agents in essential thrombocythaemia and thrombocythaemia associated with the other myeloproliferative disorders, including chronic granulocytic leukaemia, polycythaemia rubra vera and myelofibrosis. Few data exist on the use of the recombinant interferons as maintenance agents in patients with thrombocythaemia. We report on a cohort of 22 previously untreated patients, with essential thrombocythaemia, treated with recombinant alpha 2a interferon maintenance therapy for a minimum period of 6 months. Effective long-term control of platelet counts, without evidence of haematological toxicity, was achieved in 19 of 22 patients. No objective haemorrhagic or thrombotic event occurred in 298 patient months of interferon therapy. Three patients discontinued alpha interferon therapy due to adverse side-effects. Alpha 2a interferon is an effective maintenance agent in essential thrombocythaemia.

12.
Int J Food Microbiol ; 27(2-3): 117-28, 1995 Oct.
Article in English | MEDLINE | ID: mdl-8579983

ABSTRACT

The effect of heating alone (60, 65 or 70 degrees C), heating after irradiation (0.8 kGy) and heating after irradiation and storage for 14 days at 2-3 degrees C on the destruction of Listeria monocytogenes and Salmonella typhimurium in artifically inoculated minced cook-chill roast beef and gravy was investigated. Inoculated minced roast beef samples (5 g) were heated in Stomacher bags completely immersed in a water bath at each of the test temperatures. Survivors were enumerated and D and z values were determined for each of the pathogens. Observed thermal D values for two strains of L. monocytogenes at 60, 65 and 70 degrees C in the absence of pre-irradiation were 90.0-97.5 s, 34.0-53.0 s and 22.4-28.0 s, respectively, whereas thermal D values after pre-irradiation were 44.0-46.4 s, 15.3-16.8 s and 5.5-7.8 s at 60, 65 and 70 degrees C, respectively. This reduction in D values provides evidence for radiation-induced heat-sensitisation in L. monocytogenes. There was some evidence of heat-sensitisation of S. typhimurium at 60 degrees C, but not at either 65 or 70 degrees C. The z value also decreased as a consequence of pre-irradiation to a dose of 0.8 kGy (11.0-12.7 degrees C). The radiation-induced heat-sensitivity in L. monocytogenes was found to persist for up to 2 weeks storage at 2-3 degrees C prior to heating. As cook-chill products are intended to be reheated prior to consumption the results of the present study suggest that any L. monocytogenes present in a cook-chill product would be more easily killed during reheating if it were to be treated with a low dose of gamma radiation during manufacture.


Subject(s)
Food Irradiation , Hot Temperature , Listeria monocytogenes/growth & development , Meat/microbiology , Salmonella typhimurium/growth & development , Cold Temperature , Gamma Rays , Listeria monocytogenes/radiation effects , Salmonella typhimurium/radiation effects
13.
Int J Food Microbiol ; 18(1): 25-36, 1993 Mar.
Article in English | MEDLINE | ID: mdl-8466810

ABSTRACT

The effect of irradiation (2 kGy) on growth of and toxin production by Staphylococcus aureus and Bacillus cereus in roast beef and gravy during storage at abuse temperatures (15 and 22 degrees C) was assessed by inoculation studies. Irradiation resulted in a 3-4 log10 reduction in numbers of both pathogens. Whenever B. cereus and S. aureus numbers reached 10(6) and 10(7) cfu/g, respectively, during storage their toxins were detectable. As the time taken to attain these levels was longer in irradiated than in unirradiated samples, toxin production by both pathogens was delayed by irradiation. When samples initially containing low levels (10(2)/g) of S. aureus were irradiated no toxin was produced during subsequent storage at 15 or 22 degrees C. Diarrhoeal toxin produced by B. cereus was detected after 2 days at 22 degrees C, but not at 15 degrees C, in samples containing 10(2) cells/g prior to irradiation. When higher numbers (10(6)/g) of either pathogen were present prior to irradiation, toxins were produced by both pathogens at 22 degrees C, but not at 15 degrees C. Microbial competition had an effect on the growth of B. cereus and S. aureus after irradiation when a low initial inoculum was applied. However, when a higher inoculum was used the pathogens outnumbered their competitors and competition effects were less important. It was concluded that low-dose irradiation would improve the microbiological safety of roast beef and gravy.


Subject(s)
Bacillus cereus/radiation effects , Food Irradiation , Food Microbiology , Meat/microbiology , Meat/radiation effects , Staphylococcus aureus/radiation effects , Animals , Bacillus cereus/growth & development , Bacillus cereus/metabolism , Bacterial Toxins/biosynthesis , Cattle , Foodborne Diseases/prevention & control , Humans , Radiation Dosage , Safety , Staphylococcus aureus/growth & development , Staphylococcus aureus/metabolism
14.
Vet Microbiol ; 92(1-2): 135-43, 2003 Mar 20.
Article in English | MEDLINE | ID: mdl-12488077

ABSTRACT

Milk samples from 340 individual goats in 34 dairy herds throughout Norway were examined for Mycobacterium avium subsp. paratuberculosis (M.a. paratuberculosis) by culture and immunomagnetic separation combined with PCR (IMS-PCR). The samples included three categories; (A) vaccinated dairy goats in herds with paratuberculosis; (B) vaccinated dairy goats in herds with no history of paratuberculosis; (C) unvaccinated goats in herds with no history of paratuberculosis. Viable M.a. paratuberculosis were not detected by culture in any sample, but 24 samples (7.1%) tested positive by IMS-PCR when the PCR products were visualised by dot blot hybridisation. PCR products from five milk samples originating from five different herds were sequenced; all showed 99% homology with the IS900 sequence from M.a. paratuberculosis.M.a. paratuberculosis were detected in all sampled categories. The percentage of IMS-PCR positive samples from herds where paratuberculosis had previously been reported was significantly lower than from herds where the infection had never been diagnosed (3.3 and 9.1%, respectively, P=0.048). Similar proportions of milk samples from vaccinated and non-vaccinated goats tested positive for the presence of M.a. paratuberculosis. Vaccinated goats older than 4 years tested positive more often than vaccinated animals less than 2 years old. Samples collected in May tested significantly more often positive than milk sampled during February-March (13.8 and 2.9%, respectively, P=0.001). This study showed that raw goats' milk in Norway might be contaminated with M.a. paratuberculosis.


Subject(s)
Goat Diseases/microbiology , Milk/microbiology , Mycobacterium avium subsp. paratuberculosis/isolation & purification , Paratuberculosis/microbiology , Animals , Base Sequence , DNA Transposable Elements/genetics , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , Female , Food Microbiology , Goat Diseases/diagnosis , Goats , Immunomagnetic Separation/veterinary , Molecular Sequence Data , Mycobacterium avium subsp. paratuberculosis/genetics , Norway , Paratuberculosis/diagnosis , Polymerase Chain Reaction/veterinary , Seasons , Sequence Analysis, DNA , Vaccination/veterinary , Viral Vaccines/immunology , Viral Vaccines/standards
15.
Vet Microbiol ; 79(2): 123-31, 2001 Mar 20.
Article in English | MEDLINE | ID: mdl-11230934

ABSTRACT

A blind survey of 104 raw sheep and goats' milk samples (90 goat, 14 sheeps) from bulk tanks on farms throughout England, Wales and Northern Ireland was carried out over a 5-month period (January-May 1998) in order to determine the incidence of Mycobacterium paratuberculosis. Each milk sample (100 ml) was divided into two 50ml portions. One portion was decontaminated with 0.75% hexadecylpyridinium chloride for 5h before culture on slopes of Herrold's egg yolk medium and in BACTEC radiometric medium. The second portion was subjected to immunomagnetic separation followed by IS900 PCR (IMS-PCR). The IMS-PCR assay was employed in order to provide a more rapid indication of the presence of M. paratuberculosis in each milk sample than is possible by culture. Information on the Johne's disease status of the sheep and goat herds that took part in the survey was not sought at the time of milk sampling. However, it subsequently emerged that at least some of the herds whose bulk milk was tested during this study were previously or currently infected with Johne's disease. Overall, during this survey one raw goats' milk sample tested positive for the presence of M. paratuberculosis by IMS-PCR (<1% of milk samples tested) but no viable M. paratuberculosis were isolated by culture. The results of this study suggest that bulk raw sheep and goats' milk from these regions of the UK may not represent significant vehicles of transmission of M. paratuberculosis to humans.


Subject(s)
Food Microbiology , Goats , Milk/microbiology , Mycobacterium avium subsp. paratuberculosis/isolation & purification , Sheep , Animals , England , Female , Immunomagnetic Separation/veterinary , Northern Ireland , Polymerase Chain Reaction/veterinary , Wales
16.
Vet Microbiol ; 77(3-4): 369-78, 2000 Dec 20.
Article in English | MEDLINE | ID: mdl-11118722

ABSTRACT

The potential use of a novel immunomagnetic PCR (IMS-PCR) technique as a rapid method to screen milk samples for the presence of Mycobacterium avium subsp. paratuberculosis (M. ptb) was assessed. Immunomagnetic separation (IMS) for M. ptb, developed at Queen's University, Belfast, was applied to milk samples prior to IS900 PCR in order to selectively concentrate any M. ptb cells present and, at the same time, separate the cells from constituents of milk likely to inhibit subsequent PCR. This increased the sensitivity of IS900 PCR. IMS-PCR sensitivity could be further increased by initial centrifugation (2500 g for 20 min) of larger volumes of milk (10 and 50 ml), and resuspension of the sediment into a 1 ml volume appropriate for IMS treatment. Following IMS, template DNA for IS900 PCR was obtained by heating the bead-cell suspension in a thermal cycler at 100 degrees C for 15 min. It was estimated that the IMS-PCR assay could detect approximately 10(3)CFU of M. ptb per 50 ml of milk (equivalent to 20 CFU/ml), whereas the minimum detection limit of direct IS900 PCR was estimated at 10(5)CFU of M. ptb per 50 ml (equivalent to 2000 CFU/ml). A blind trial was carried out in which a total of 40 spiked (10(6)CFU M. ptb) and unspiked, raw and laboratory-pasteurised milk samples were independently tested by IMS-PCR and conventional IS900 PCR. IMS-PCR correctly identified 97. 5% of milk samples (sensitivity 100%, specificity 95%), including spiked milk samples before and after laboratory-pasteurisation. One false positive result was obtained which may have resulted from carryover between samples during the IMS procedure. Conventional IS900 PCR correctly identified only 72.5% of the same 40 milk samples (sensitivity 23%, specificity 100%). IMS-PCR was also shown to be capable of detecting natural M. ptb infection in raw sheep's milk, and raw and commercially pasteurised cows' milk.


Subject(s)
Cattle Diseases/diagnosis , Immunomagnetic Separation/veterinary , Milk/microbiology , Mycobacterium avium subsp. paratuberculosis/isolation & purification , Paratuberculosis/diagnosis , Polymerase Chain Reaction/veterinary , Animals , Cattle , DNA, Bacterial/analysis , Dairying , False Positive Reactions , Female , Immunomagnetic Separation/methods , Polymerase Chain Reaction/methods , Sensitivity and Specificity
17.
J Hand Surg Br ; 18(5): 648-51, 1993 Oct.
Article in English | MEDLINE | ID: mdl-8294836

ABSTRACT

Two cases are described of anterior dislocation of the proximal interphalangeal joint which could not be reduced because of interposition of the central slip of the extensor mechanism. In common with previously reported cases the injuries were sustained in a spin drier. The literature on anterior dislocation of the proximal interphalangeal joint is reviewed. Three distinct patterns of dislocation are recognised and their management is discussed.


Subject(s)
Finger Injuries/physiopathology , Finger Joint/physiopathology , Joint Dislocations/physiopathology , Range of Motion, Articular , Accidents , Female , Finger Injuries/surgery , Finger Joint/surgery , Humans , Joint Dislocations/etiology , Joint Dislocations/surgery , Middle Aged
18.
Ann R Coll Surg Engl ; 70(1): 29-33, 1988 Jan.
Article in English | MEDLINE | ID: mdl-3408135

ABSTRACT

We report on 106 elective splenectomies performed for haematological disorders between March 1979 and January 1986. The most common indications were immune thrombocytopenic purpura (30 patients) and Hodgkin's disease (19 patients). However, staging laparotomy is no longer performed routinely for patients with Hodgkin's disease and the reasons for this are discussed. Other indications for splenectomy included splenic pain (13 patients), autoimmune haemolytic anaemia (12 patients), hereditary spherocytosis (11 patients) and hypersplenism (9 patients). The overall morbidity and mortality was 48% and 5% respectively. The most common postoperative complication was thrombocytosis (defined as a platelet count greater than 800 X 10(9)/l) and occurred in 26 patients. This review confirms that splenectomy continues to have an important role in the management of certain haematological disorders.


Subject(s)
Hematologic Diseases/surgery , Splenectomy , Adolescent , Adult , Aged , Child , Child, Preschool , Female , Hematologic Diseases/mortality , Humans , Infections/etiology , Infections/immunology , Male , Middle Aged , Remission Induction , Splenectomy/adverse effects , Splenectomy/mortality , Thrombocytosis/etiology
19.
Acta Vet Scand ; 44(3-4): 261-6, 2003.
Article in English | MEDLINE | ID: mdl-15074643

ABSTRACT

The possibility that milk from cattle with Johne's disease could be a potential vehicle of transmission of Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) to humans has been the focus of a UK government-funded research programme at Queen's University, Belfast since 1993. The main findings of this research programme are reported and practical advice about the most appropriate methods for the isolation/detection of this organism in milk is given. The findings of several milk surveys during which optimised sensitive detection methods were employed (decontamination with 0.75% cetyl pyridinium chloride for 5 h prior to culture and a novel immunomagnetic PCR technique) have revealed that detectable levels of M. paratuberculosis are present in bulked raw cows' milk in the UK at both the farm level and at dairy processing plants prior to pasteurisation. Furthermore, results of three different experimental approaches to assess the effect of pasteurisation time/temperature conditions on the viability of M. paratuberculosis (laboratory pasteurisation studies, a national survey of commercially pasteurised milk, and processing of naturally infected milk through commercial-scale pasteurising plant) provide firm evidence that this organism is capable of surviving commercial milk pasteurisation on occasion. Hence, both raw and pasteurised cows' milk are potential vehicles of transmission of M. paratuberculosis to humans.


Subject(s)
Milk/microbiology , Mycobacterium avium subsp. paratuberculosis/isolation & purification , Paratuberculosis/transmission , Animals , Cattle , Humans
20.
Lett Appl Microbiol ; 42(4): 305-11, 2006 Apr.
Article in English | MEDLINE | ID: mdl-16599979

ABSTRACT

Mycobacterium avium ssp. paratuberculosis (Map) is an important animal pathogen with a potential, but as yet unproven, role in human disease. This review briefly describes the characteristics of Map that distinguish it from other Mycobacterium spp., presenting new information arising from completion of the sequencing of the Map genome. It then focuses on the potential mechanisms Map might employ to survive and disseminate in the environment, including interaction with protozoa and insects, dormancy, biofilm formation and aerosolization.


Subject(s)
Mycobacterium avium subsp. paratuberculosis/physiology , Aerosols , Animals , Biofilms , Genome, Bacterial , Insecta/microbiology , Mycobacterium avium subsp. paratuberculosis/genetics , Nematoda/microbiology
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