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The suitability of using optical-density (OD) measurements after a 6-h incubation period to determine bacterial contamination on hatching eggs was evaluated. A total of 154 hatching eggs, from five different flocks, were examined visually and bacterial counts and OD measurements of egg washings after 6 h of incubation at 37 degrees C were carried out to determine the levels of bacterial contamination, and these results were compared. A relationship between the OD measurements of egg washings and the log of the bacterial plate counts, was demonstrated. The OD measurements provide accurate and quantitative information to detect bacterial contamination on hatching eggs, as they are simple to perform, repeatable and reliable.

The use of optical-density (OD) readings after a 6-h incubation period, as a suitable method to quantify the bacterial contamination on hatching eggs, was established by the use of pure cultures of five bacterial isolates found to be the most prevalent on the hatching eggs examined. These isolates were identified as Micrococcus luteus, Staphylococcus gallinarum, Streptococcus epidermidis, Pseudomonas cepacei and Bacillus cereus. It was established that the OD reading was repeatable when the same inoculum was used to inoculate five different cultures, which were incubated for 6 h at 37 degrees C. This repeatability was not affected by bacterial isolate, or bacterial concentration of the inoculum, or when mixed bacterial cultures were used. A direct relationship was established between the OD reading (at 540 nm) after 6 h and the log of the bacterial concentration at the start of incubation. The OD reading after 6 h of incubation is a repeatable, rapid and simple method to quantify the bacterial concentration at the start of the incubation period.

A total of 27 different isolates of Haemophilus paragallinarum were made from chickens between June 1991 and December 1992. All of these isolates were examined by ELISA, by means of a locally produced panel of three monoclonal antibodies (denoted F1, V1 and VF3). The isolates were all of the F1 antigenic type. Three of them showed a weak reaction with the F1 monoclonal antibody, while three other isolates reacted strongly with the F1 as well as with the VF3 Mab. A selection of stored Haemophilus isolates, dating from 1984 to 1985, were also examined with the Mabs and found to be of the F1 antigenic type. Fifteen isolates were collected before 1974, i.e. before the use of Haemophilus vaccines in this country. The majority of them were of the F1 antigenic grouping. Some showed a weak reaction with the F1 Mab; others showed a strong reaction with both the F1 and VF3 Mabs; and a few showed no significant reaction with any of the Mabs used. Strains used for the production of infectious coryza vaccine were also examined with the Mabs. Strain 0083 showed a stronger reaction with the V1 Mab than with the F1 Mab, whereas strain 0222 showed no reaction with any of the Mabs. None of the SA field isolates collected since the use of vaccines exhibits the V1 antigenicity, which is the prevalent antigen of strain 0083. Most (80%) of the SA field isolates showed a stronger reaction with the F1 Mab than did strain 0083. Antigenically silent isolates similar to 0222 (Page's serotype B) were isolated before the use of vaccines, but not since.

Atypical Haemophilus paragallinarum have been isolated from both laying hens and broilers suffering from typical symptoms of infectious coryza in South Africa. Re-inoculation of these bacteria into SPF chickens resulted in similar pathology. The bacteria could be successfully re-isolated from the experimentally infected chickens. Four of the isolates from layers and 3 of those from broilers were found to be closely related to H. paragallinarum serotype A (0083 strain) when tested by the use of a panel of locally developed monoclonal antibodies in the enzyme linked immunosorbent assay (ELISA). A total of 15 isolates from layers and 19 from broilers were found to be more typical of previously collected South African field isolates of H. paragallinarum. A 3rd group, consisting of 5 isolates from layers and 15 from broilers, showed no reaction with the panel of monoclonal antibodies. All the isolates were regarded as atypical because they no longer required V factor (NAD) for growth, whereas strain 0083 and previously collected field isolates M 85 and SB 86 did require it. Crude plasmid extractions from an isolate serologically related to 0083 was used to convert reference strains of H. paragallinarum into NAD-independent isolates, thus indicating that NAD independence is carried on a plasmid.

A commercially available E. coli probe kit was used to test 1 lactose negative E. coli isolate and 2 hydrogen sulphide-producing E. coli isolates. The isolates were confirmed as E. coli by means of the API system. The GENETRAK E. coli DNA probe kit reacted positively with the lactose negative isolate, but negatively with the hydrogen sulphide-producing isolate.

A drop in egg production associated with loss of pigment and soft-shelled eggs was experienced on a farm in the Western Cape. Investigations revealed high levels of antibodies against adenovirus 127, the causative agent of egg drop syndrome (EDS) in chickens which were not vaccinated against EDS. A cytopathic agent was subsequently isolated in chicken embryo liver cell cultures and identified as EDS virus by haemagglutination inhibition and neutralization tests.

Both pathogenic and non-pathogenic bacteria were isolated from fish, both salmonid and non-salmonid, from selected river systems in Natal. Pasteurella pisicida was isolated for the first time from fish in South Africa. The isolation of Yersinia ruckeri, Aeromonas salmonicida, and Edwardsiella tarda were recorded for the first time from fish in Natal. A. hydrophila and Flexibacter columnaris were found to be widespread throughout the river systems in Natal. The Streptococcus species which caused serious disease problems in trout in the Cape Province and Transvaal was not isolated from any of the fish examined in Natal.

A total of 678 fish from 26 different sites along 8 river systems was examined for the presence of trout viruses. No isolations of any viruses were made from any fish collected from these systems in Natal during this study. No antibodies against any of the trout viruses were detected in the serum collected from these fish. It would appear from this study that the river systems in Natal are free of any of the known viruses of trout. Infectious pancreatic necrosis virus was, however, isolated from trout from the Natal Parks Board Hatchery at Kamberg Nature Reserve on the Mooi River during June and July of 1988. Neutralising antibodies against the VR 299 serotype of IPN virus were detected in the serum collected from trout at Kamberg in September 1988.

A detailed microbiological and parasitological survey of salmonids and indigenous fish in the upper reaches of selected river systems in Natal was undertaken. Fish were collected from the rivers by electro-fishing, from dams by gill netting and from hatcheries along the rivers. A total of 678 fish from 26 different sites along 8 river systems were collected. No parasites could be detected on fish from four of the rivers tested. Parasites were, however, detected on fish from large production sites in Natal and in the rivers feeding these sites. Parasites which were detected were Ichthyopthirius multifiliis, various Trichodina spp., Apiosoma sp. and Gyrodactylus sp. The heads of all fish estimated to be less than one year of age were collected and examined for the spores of Myxosoma cerebralis. All of the samples were found to be free of this parasite.

Mycobacterium fortuitum was isolated from 3 species of freshwater fish in South Africa for the first time. The bacterium was isolated from oscars, guppies and discus fish from different sources. Heavy mortalities as a result of infection with this bacterium were reported from a guppy farm and the multi-resistance of this isolate to antimicrobials rules out the treatment of infected fish.

A procedure established for the selective isolation of the species of Streptococcus responsible for rainbow trout streptococcosis in South Africa, consisted of the inoculation of samples into nutrient broth which had been supplemented with 100 micrograms/ml of nalidixic acid, 160 micrograms/ml of oxolinic acid or 200 micrograms/ml of sodium azide. After incubation, the sample was plated onto tetrazolium agar on which the rainbow trout pathogenic Streptococcus species grew as a red colony. The colonies were isolated from the tetrazolium agar and identified as rainbow trout pathogenic isolates by biochemical and serological tests. In the laboratory the selective procedure is capable of detecting about 2 bacteria per ml. This procedure was used in the field and biochemically identical Streptococcus species were found in the mud and a freshwater crab from the water source of a site with a history of streptococcosis.

A total of 3,257 samples of diseased rainbow trout were examined for the presence of viruses from January 1983 to December 1987. A virus closely related to the VR 299 serotype of infectious pancreatic necrosis virus was isolated from 13 cases. An additional 7,228 viscera samples from asymptomatic fish were collected during the same period and a similar virus was isolated from 2 sites. During the same period 2,892 ovarian fluid samples were collected and a similar virus was isolated from 1 site. A similar virus was also isolated from one consignment of imported trout ova. A total of 5,550 ova was examined during this period. The viruses were identified by various tests as being closely related to the VR299 serotype of infectious pancreatic necrosis virus. All these samples tested negative for infectious haematopoietic necrosis virus, viral haemorrhagic septicaemia virus and herpesvirus salmonis.

The indirect fluorescent antibody technique has been used successfully for the rapid serological identification of the Streptococcus sp. responsible for streptococcosis of rainbow trout. This technique has been used to identify the Streptococcus sp. in pure cultures and smears made from experimentally infected and diseased fish.

A Streptococcus species biochemically and serologically identical to the rainbow trout pathogenic Streptococcus species was isolated from the internal organs of the fish specific leech, Batracobdelloides tricarinata. These leeches were obtained from Roodeplaat Dam, near Pretoria, in which rainbow trout do not occur. This is the first isolation of this bacterium from an environmental source not related to rainbow trout and it is proposed that this leech is a possible reservoir of the rainbow trout pathogenic Streptococcus sp. in South Africa.