Incubation at raised temperature or enrichment media, combined with secondary enrichment in Rappaport's medium, for the isolation of salmonellas from sewage.

Two FMDV strains which had been previously differentiated by complement-fixation were compared by guinea-pig protection test, kinetic neutralization and micro-neutralization tests. It was found that these tests, which have not been previously applied by the methods described, were all capable of FMDV STRAIN DIFFERENTIATION. Similar differences were found by all methods, which suggests that comparisons made by cross-CF, cross-neutr-alization or cross-protection involve measurement of the same antigen/antibody interactions.     

Salmonella isolation with Rappaport's medium after pre-enrichment in buffered peptone water using a series of inoculum ratios.

The ability of malachite green/magnesium chloride broth (Rappaport''s medium) to isolate salmonellas from 25 ml quantities of sewage-polluted natural water was investigated. Samples were first pre-enriched in buffered peptone water and varying volumes of inoculum from the pre-enrichment culture were inoculated into Rappaport''s broth. Inoculum ratios in the range 1:2000 to 1:10 were examined. The inoculum ratio denotes the ratio of the volume of inoculum to the volume of fluid medium into which it is introduced. Optimum results were obtained with the 1:2000 ratio, although the salmonella isolation rate was only slightly less with the 1:500 and 1:100 ratios. The 1:2000 inoculum ratio was obtained with a graduated loop holding approximately 0.005 ml of fluid. Use of a loop for inoculation has advantages in speed of performance and safety of manipulation.     

A comparison of two modifications of Rappaport's enrichment medium (R25 and RV) for the isolation of salmonellas from sewage polluted natural water.

The development of Rappaport''s enrichment medium and elevated temperature incubation as methods of salmonella isolation is traced. The recent recording of a merger of the two techniques by means of Rappaport--Vassiliadis medium is noted (RV medium). In Cardiff, we have found an earlier modification of Rappaport''s enrichment (R25) by Vassiliadis to be efficient in salmonella recovery from environment samples. The current study compares the two media using sewage polluted natural water as test material. Under the conditions of experiment, R25 was more successful in salmonella isolation than RV, although the later medium inhibited competitive organisms better. R25 is a convenient enrichment broth for routine use. In combination with pre-enrichment it allows the use of a loop for subculture rather than a pipette. This increases safe manipulation. It also produces a high proportion of positive isolations at the 24 h subculture time in contrast to other enrichment broths. For these reasons it forms an integral part of salmonella isolation methodology in our laboratory.     

Salmonella isolation with Rappaport's medium after pre-enrichment in buffered peptone water using a series of inoculum ratios

The ability of malachite green/magnesium chloride broth (Rappaport's medium) to isolate salmonellas from 25 ml quantities of sewage-polluted natural water was investigated. Samples were first pre-enriched in buffered peptone water and varying volumes of inoculum from the pre-enrichment culture were inoculated into Rappaport's broth. Inoculum ratios in the range 1:2000 to 1:10 were examined. The inoculum ratio denotes the ratio of the volume of inoculum to the volume of fluid medium into which it is introduced. Optimum results were obtained with the 1:2000 ratio, although the salmonella isolation rate was only slightly less with the 1:500 and 1:100 ratios. The 1:2000 inoculum ratio was obtained with a graduated loop holding approximately 0.005 ml of fluid. Use of a loop for inoculation has advantages in speed of performance and safety of manipulation.     

A comparison of two modifications of Rappaport's enrichment medium (R25 and RV) for the isolation of salmonellas from sewage polluted natural water

The development of Rappaport's enrichment medium and elevated temperature incubation as methods of salmonella isolation is traced. The recent recording of a merger of the two techniques by means of Rappaport--Vassiliadis medium is noted (RV medium). In Cardiff, we have found an earlier modification of Rappaport's enrichment (R25) by Vassiliadis to be efficient in salmonella recovery from environment samples. The current study compares the two media using sewage polluted natural water as test material. Under the conditions of experiment, R25 was more successful in salmonella isolation than RV, although the later medium inhibited competitive organisms better. R25 is a convenient enrichment broth for routine use. In combination with pre-enrichment it allows the use of a loop for subculture rather than a pipette. This increases safe manipulation. It also produces a high proportion of positive isolations at the 24 h subculture time in contrast to other enrichment broths. For these reasons it forms an integral part of salmonella isolation methodology in our laboratory.     

A comparisoin of the R25 modification of Rappaport s enrichment medium with strontium chloride B for salmonella isolation from sewage polluted natural water.

The relation of salmonella isolation efficiency and the size of inoculum introduced from a buffered peptone water culture of sewage polluted water into strontium chloride B medium was investigated. Two separate studies were made, one using enrichment at 37 degrees C, the other at 43 degrees C. From these trials, two inocula suitable for efficient salmonella isolation were determined. Using this information, strontium chloride B medium was compared with modified Rappaport''s broth (R25). The inoculum used with R25 was 0.005 ml, determined in an earlier study. Two incubation temperatures were employed with strontium chloride enrichment (37 and 43 degrees C). Rappaport''s medium was incubated at 37 degrees C only. Elevated temperature enrichment at 43 degrees C improved the performance of strontium chloride B, but Rappaport''s broth still gave significantly better results. This supports earlier studies on simplification of salmonella isolation and standardization of routine technique on a single enrichment medium: Rappaport broth (R25) incubated at 37 degrees C.     

A comparisoin of the R25 modification of Rappaport s enrichment medium with strontium chloride B for salmonella isolation from sewage polluted natural water

The relation of salmonella isolation efficiency and the size of inoculum introduced from a buffered peptone water culture of sewage polluted water into strontium chloride B medium was investigated. Two separate studies were made, one using enrichment at 37 degrees C, the other at 43 degrees C. From these trials, two inocula suitable for efficient salmonella isolation were determined. Using this information, strontium chloride B medium was compared with modified Rappaport's broth (R25). The inoculum used with R25 was 0.005 ml, determined in an earlier study. Two incubation temperatures were employed with strontium chloride enrichment (37 and 43 degrees C). Rappaport's medium was incubated at 37 degrees C only. Elevated temperature enrichment at 43 degrees C improved the performance of strontium chloride B, but Rappaport's broth still gave significantly better results. This supports earlier studies on simplification of salmonella isolation and standardization of routine technique on a single enrichment medium: Rappaport broth (R25) incubated at 37 degrees C.     

Salmonella isolation with Rappaport-Vassiliadis enrichment medium seeded with different sized inocula of pre-enrichment cultures of meat products and sewage polluted water.

A total of 574 samples, of seven different types, were examined for the presence of salmonellas. All the specimens were pre-enriched in buffered peptone water and enriched in Rappaport-Vassiliadis medium (RV medium). In one trial 0.1 ml of pre-enrichment culture of 497 samples (79 chicken carcasses, 228 specimens of minced meat, 100 pork sausages, 19 samples of dried powdered chicken meat, 11 specimens of faeces of healthy pigs and 60 samples of sewage polluted natural sea water) was seeded to 10 ml as well as to 100 ml of RV medium. With the first inoculum (ratio 0.1:10), 111 samples were found to contain salmonellas, while with the second inoculum (ratio 0.1:100), only 102 positive samples were detected. This difference is marginally significant (P less than 0.05). In another trial, 0.1 ml, 0.2 ml and 0.5 ml of pre-enrichment culture of 162 specimens (71 chicken carcasses, 40 samples of sewage polluted sea water and 51 samples of sewage polluted river water) were in turn introduced to 10 ml of RV medium. With the 0.1 ml inoculum 93 positive samples were detected, while with the 0.2 and 0.5 ml inocula 93 and 88 positive samples were found. The differences are not statistically significant. In these trials the growth of competing organisms was minimal with ratios of inocula 0.1:10 or 0.1:100.     

Salmonella isolation with Rappaport-Vassiliadis enrichment medium seeded with different sized inocula of pre-enrichment cultures of meat products and sewage polluted water

A total of 574 samples, of seven different types, were examined for the presence of salmonellas. All the specimens were pre-enriched in buffered peptone water and enriched in Rappaport-Vassiliadis medium (RV medium). In one trial 0.1 ml of pre-enrichment culture of 497 samples (79 chicken carcasses, 228 specimens of minced meat, 100 pork sausages, 19 samples of dried powdered chicken meat, 11 specimens of faeces of healthy pigs and 60 samples of sewage polluted natural sea water) was seeded to 10 ml as well as to 100 ml of RV medium. With the first inoculum (ratio 0.1:10), 111 samples were found to contain salmonellas, while with the second inoculum (ratio 0.1:100), only 102 positive samples were detected. This difference is marginally significant (P less than 0.05). In another trial, 0.1 ml, 0.2 ml and 0.5 ml of pre-enrichment culture of 162 specimens (71 chicken carcasses, 40 samples of sewage polluted sea water and 51 samples of sewage polluted river water) were in turn introduced to 10 ml of RV medium. With the 0.1 ml inoculum 93 positive samples were detected, while with the 0.2 and 0.5 ml inocula 93 and 88 positive samples were found. The differences are not statistically significant. In these trials the growth of competing organisms was minimal with ratios of inocula 0.1:10 or 0.1:100.     

Comparison of selenite F, Muller-Kauffmann tetrathionate and Rappaport's medium for the isolation of salmonellas from sewage-polluted natural water using a pre-enrichment technique.

Three enrichment broths, selenite F, Muller-Kauffmann tetrathionate and Rappaport, were examined for their efficiency in salmonella isolation. The three media, prepared from single ingredients in the laboratory, were compared with their commercial equivalents. Laboratory-prepared media were more efficient for isolating salmonellas from sewage-polluted natural water samples. A pre-enrichment stage using buffered peptone water was employed throughout the investigation. The size of inoculum from the pre-enrichment medium was relevant to successful salmonella isolation. Inocula studied were 1 ml and one loopful (3 mm diameter loop). The smaller inoculum gave better results with Rappaport, the larger with selenite and tetrathionate. Using the optimal inocula, Rappaport was the most efficient enrichment broth of the three fluid media in this study.     

A modification of Diamond's medium for the axenic culture of Entamoeba histolytica.

The use of casein hydrolysate in Diamond's axenic culture medium TPS-1 in replacement of trypticase allowed good growth of the trophozoites of Entamoeba histolytica. This modified medium also supported growth of trophozoites preserved for 16 months in liquid nitrogen. Considerable labour and cost of serum can be saved by using 5% instead of 10% bovine serum in combination with this modified medium.     

Comparison of selenite F, Muller-Kauffmann tetrathionate and Rappaport's medium for salmonella isolation from chicken giblets after pre-enrichment in buffered peptone water.

Six hundred and eighty three samples of chicken giblets were examined for salmonellas. Three hundred and forty nine of these were neck and crop specimens and 224 were combined liver and heart samples. Two hundred and ten, in all, contained salmonellas. The technique of examination included pre-enrichment in buffered peptone water at 37 degrees C for 18 h and subculture to three enrichment media: Muller-Kauffmann tetrathionate, selenite F and Rappaport''s magnesium chloride malachite green broth. Inocula from buffered peptone water to 10 ml of tetrathionate and selenite were 1 ml in each case. The inoculum from the pre-enrichment medium to 10 ml of Rappaport was 0.005 ml. Tetrathionate and selenite were incubated at 43 degrees C for 48 h. Rappaport''s medium was incubated at 37 degrees C for 48 h. Subcultures from all three enrichment broths were made at 24 h and 48 h to brilliant green MacConkey agar. Selective agars were incubated at 37 degrees C for 24 h. The most successful technique for salmonella isolation used Rappaport''s medium, which was significantly more efficient than either tetrathionate or selenite. This finding reinforces results obtained using sewage polluted natural water as test material and it is suggested that routine examination of environment samples for salmonellas could be based on Rappaport''s medium alone. If S. typhi, S. dublin or subgenus III salmonellas were likely to be present in the sample, the technique described here would require modification.