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Biomedical subjects

I R Grant

Publications and source records attributed to I R Grant.

At least 19 recordsLinked to original sources

Incidence of Mycobacterium paratuberculosis in raw sheep and goats' milk in England, Wales and Northern Ireland.

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.

Animals↗

Improved detection of Mycobacterium avium subsp. paratuberculosis In milk by immunomagnetic PCR.

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.

Animals↗

Phylogenetic characterization of a novel radiation-resistant bacterium from irradiated pork: description of Hymenobacter actinosclerus sp. nov.

A phylogenetic analysis was performed on a red-pigmented, radiation-resistant, Gram-negative, rod-shaped organism originating from irradiated pork. Comparative 16S rRNA gene sequencing showed the bacterium was a member of the Cytophaga-Flavobacterium-Bacteroides line of descent and represents a new subline within the genus Hymenobacter. A new species, Hymenobacter actinosclerus, is described for this novel radiation-resistant bacterium. The type strain of Hymenobacter actinosclerus is CCUG 39621T.

Animals↗

Effect of higher pasteurization temperatures, and longer holding times at 72 degrees C, on the inactivation of Mycobacterium paratuberculosis in milk.

Raw cow's milk spiked with 10(6) cfu ml-1 of Mycobacterium paratuberculosis was subjected to heat treatments of 72, 75, 78, 80, 85 or 90 degrees C for 15 s, and 72 degrees C for 20 and 25 s, using laboratory pasteurizing units. Three bovine strains of Myco. paratuberculosis were studied (NCTC 8578, B2 and DVL 943). Each strain was subjected to all the heat treatments indicated on three separate occasions. Although each of the heat treatments achieved a substantial (5-6 log10) reduction in numbers of viable Myco. paratuberculosis, small numbers of the organism (4-16 cfu 10 ml-1) survived in a proportion of the milk samples at each of the higher temperatures investigated, right up to 90 degrees C for 15 s. A longer holding time of 25 s at 72 degrees C was found to be more effective at inactivating Myco. paratuberculosis. Only one of the three strains studied, B2, yielded small numbers of survivors after heating at 72 degrees C for 20 s, but it was completely inactivated by extending the holding time at 72 degrees C by a further 5 s to 25 s. It was concluded that a longer holding time is more likely to achieve the complete inactivation of Myco. paratuberculosis in milk than a higher pasteurization temperature.

Animals↗

Effect of high-temperature, short-time (HTST) pasteurization on milk containing low numbers of Mycobacterium paratuberculosis.

The efficacy of high-temperature, short-time (HTST) pasteurization (72 degrees C/15 s) when low numbers (< or = 10(3) cfu ml-1) of Mycobacterium paratuberculosis are present in milk was investigated. Raw cows' milk spiked with Myco. paratuberculosis (10(3) cfu ml-1, 10(2) cfu ml-1, 10 cfu ml-1, and 10 cfu 50 ml-1) was subjected to HTST pasteurization using laboratory pasteurizing units. Ten bovine strains of Myco. paratuberculosis were tested in triplicate. Culture in BACTEC Middlebrook 12B radiometric medium detected acid-fast survivors in 14.8% and 10% of HTST-pasteurized milk samples at the 10(3) and 10(2) cfu ml-1 inoculum levels, respectively, whereas conventional culture on Herrold's egg yolk medium containing mycobactin J detected acid-fast survivors in only 3.7% and 6.7% of the same milk samples. IS900-based PCR confirmed that these acid-fast survivors were Myco. paratuberculosis. No viable Myco. paratuberculosis were isolated from HTST-pasteurized milk initially containing either 10 cfu ml-1 or 10 cfu 50 ml-1.

Animals↗

Isolation of Mycobacterium paratuberculosis from milk by immunomagnetic separation.

An immunomagnetic separation (IMS) technique was developed to facilitate selective isolation of Mycobacterium paratuberculosis cells from milk. Rabbit polyclonal antibodies against radiation-killed intact M. paratuberculosis cells were produced and used to coat sheep anti-rabbit immunoglobulin G (IgG) type M-280 Dynabeads. The rabbit anti-M. paratuberculosis IgG-coated beads (IMB) reacted strongly with laboratory strains of M. paratuberculosis as determined by slide agglutination, and microscopic examination confirmed that M. paratuberculosis cells attached to the IMB. The IMB were found to have a maximum binding capacity of 10(4) to 10(5) CFU of M. paratuberculosis. Studies showed that IMS selectively recovered M. paratuberculosis from inoculated milk containing as few as 10 CFU of M. paratuberculosis per ml when 10 microliter IMB (ca. 10(6) beads) was added to 1 ml of milk and the preparation was incubated for 30 min at room temperature with gentle agitation. Larger volumes of milk (10 and 50 ml) were centrifuged and resuspended in 1 ml of phosphate-buffered saline-0.05% Tween 20 prior to IMS in order to increase the sensitivity of the method. Currently, primary isolation of M. paratuberculosis from a milk sample relies on chemical decontamination, followed by culturing on Herrold's egg yolk medium, which must be incubated at 37 degreesC for up to 18 weeks. The potential value of our IMS method is as an aid for rapid detection of M. paratuberculosis in milk when it is used in conjunction with end point detection methods, such as IS900 PCR or an enzyme-linked immunosorbent assay.

Animals↗

Thermal inactivation of several Mycobacterium spp. in milk by pasteurization.

The thermal inactivation of Mycobacterium avium, Myco. bovis, Myco. fortuitum, Myco. intracellulare and Myco. kansaii in milk at 63.5 degrees C was investigated. Survivors were enumerated after heating for 0, 5, 10, 15, 20 and 30 min and thermal death curves were constructed for each species. Mycobacterium bovis and Myco. fortuitum were found to exhibit linear thermal death curves and neither species demonstrated any survival after heating at 63.5 degrees C for 30 min (equivalent to holder pasteurization). In contrast, Myco. avium, Myco. intracellulare and Myco. kansasii yielded thermal death curves which exhibited significant 'tailing' and all three strains survived holder pasteurization.

Animals↗

Inactivation of Mycobacterium paratuberculosis in cows' milk at pasteurization temperatures.

The thermal inactivation of 11 strains of Mycobacterium paratuberculosis at pasteurization temperatures was investigated. Cows' milk inoculated with M. paratuberculosis at two levels (10(7) and 10(4) CFU/ml) was pasteurized in the laboratory by (i) a standard holder method (63.5 degrees C for 30 min) and (ii) a high-temperature, short-time (HTST) method (71.7 degrees C for 15 s). Additional heating times of 5, 10, 15, 20, and 40 min at 63.5 degrees C were included to enable the construction of a thermal death curve for the organism. Viability after pasteurization was assessed by culture on Herrold's egg yolk medium containing mycobactin J (HEYM) and in BACTEC Middlebrook 12B radiometric medium supplemented with mycobactin J and sterile egg yolk emulsion. Confirmation of acid-fast survivors of pasteurization as viable M. paratuberculosis cells was achieved by subculture on HEYM to indicate viability coupled with PCR using M. paratuberculosis-specific 1S900 primers. When milk was initially inoculated with 10(6) to 10(7) CFU of M. paratuberculosis per ml, M. paratuberculosis cells were isolated from 27 of 28 (96%) and 29 of 34 (85%) pasteurized milk samples heat treated by the holder and HTST methods, respectively. Correspondingly, when 10(3) to 10(4) CFU of M. paratuberculosis per ml of milk were present before heat treatment, M. paratuberculosis cells were isolated from 14 of 28 (50%) and 19 of 33 (58%) pasteurized milk samples heat treated by the holder and HTST methods, respectively. The thermal death curve for M. paratuberculosis was concave in shape, exhibiting a rapid initial death rate followed by significant "tailing." Results indicate that when large numbers of M. paratuberculosis cells are present in milk, the organism may not be completely inactivated by heat treatments simulating holder and HTST pasteurization under laboratory conditions.

Animals↗

Combined effect of gamma radiation and heating on the destruction of Listeria monocytogenes and Salmonella typhimurium in cook-chill roast beef and gravy.

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.

Cold Temperature↗

Effect of low-dose irradiation on growth of and toxin production by Staphylococcus aureus and Bacillus cereus in roast beef and gravy.

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.

Animals↗

Irreducible rotational anterior dislocation of the proximal interphalangeal joint. A spin drier injury.

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.

Accidents↗

Prospective randomised study of double hemi-body irradiation with and without subsequent maintenance recombinant alpha 2b interferon on survival in patients with relapsed multiple myeloma.

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.

Aged↗

A numerical taxonomic study of lactic acid bacteria isolated from irradiated pork and chicken packaged under various gas atmospheres.

Ninety-four Gram-positive, catalase-negative bacteria, isolated from pork and chicken that had been packed in modified atmospheres and irradiated to 1.75 and 2.5 kGy respectively, were studied. The majority of the strains were Lactobacillus saké. Numerical taxonomy, with the group average clustering strategy, revealed the existence of six clusters at the 85% similarity level. The largest, Cluster 1, contained 78 (83%) of the test strains along with three Lact. sak'e strains. Cluster 2 contained three test strains and the type strains of Carnobacterium piscicola and Carn. divergens. Cluster 3 contained two chicken strains and Lact. curvatus. Cluster 4 contained a pork strain and Leuconostoc dextranicum. Clusters 5 and 6 contained four pork and two chicken strains respectively and no type strains. Four test strains remained unclustered as did the other reference strains included in the study.

Animals↗

Alpha-interferon therapy for essential thrombocythaemia.

18 patients with symptomatic essential thrombocythaemia were treated with recombinant alpha interferon (2a or 2b). Subcutaneous dosage regimens, which were well-tolerated, selectively lowered the platelet count and relieved symptoms in all patients, whether previously treated or untreated. Recombinant alpha-interferon may offer a non-leukaemogenic alternative therapy for these patients.

Adult↗

Elective splenectomy in haematological disorders.

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.

Adolescent↗