Strain actinobaculum suis 2 a used to prepare antigen-based assays for serological reactions and reactions of indirect immunofluorescence for the diagnosis of urocystitis and pyelonephritis in sows

 

(57) Abstract:

The invention relates to veterinary Microbiology and can be used in bacteriological laboratories for the preparation of diagnostic kits. The strain obtained using traditional microbiological techniques from the urine of pigs with clinical signs of urocystic and pyelonephritis. The strain is deposited in the Institute of culture collections of microorganisms GMCVB Vector number 0699. Expressed antigenic properties and the ability to quickly and easily multiply on artificial and natural environments make use of this strain as a reference strain in the preparation of diagnostic kits.

The invention relates to the field of veterinary Microbiology, epizootiology and can be used in bacteriological laboratories.

In the claimed invention was set the task of finding a strain of bacteria, easy and fast breeding on natural and artificial nutrient media and has marked antigenic properties.

Known strains of Actinobaculum (Corynebacterium, Eubacterium, Actinomyces) suis allocated M. A. Soltys, Spratling F. R.(1957) (1); Larsen, J. L. (1970) (2); Jones, J. E. T., E. Farries u. D. Smidt (1982) (3). Strain Actinobaculum suis Corynebacterium suis. In this regard, quite a clear example is the work of J. L. Larsen (2). Of these three properties characteristics of this pathogen fickle - fermentation of glucose, sucrose, xylose. In the domestic literature information on the allocation of Actinobaculum suis us not detected.

The problem is solved by a new strain Actinobaculum suis 2A used for preparation of diagnosticum /antigen for the indirect immunofluorescence reaction, the antigen for serological reactions/. The strain obtained using traditional microbiological methods July 10, 1998 from the urine of pigs with clinical signs of urocystic and pyelonephritis-owned subsistence economy Acircle Omsk region.

This strain is assigned a registration number 0699 (22 December 1999), deposited in the Institute of collections of cultures of microorganisms of the State scientific centre VB "Vector", 633159, NR, Koltsovo, SRC VB "Vector", Institute of CMC, where it is stored.

The strain is characterized by the following morphological, cultural, physiological and biochemical characteristics.

Morphological features.

The specified strain has a pronounced polymorphism. In smears from cultures microkernel be 1.5 to 3.5, and a thickness of 0.3 to 0.7 μm.

In smears from young cultures of bacteria are often placed at an angle to each other, indicating break up the type of division. Gram microorganism becomes positive, fixed, spores and capsules are not formed.

Cultural, physiological and biochemical characteristics.

On a special nutrient medium, consisting of a Columbia agar, 10% defibrinating horse blood, 2% glucose, colistin sulphate 10 mg/l, nalidixic of acido 15 mg/l, metronidazole 50 mg/l Grows as optional gone anaerobic, on the day 3 forming a gray-white, round, with irregular wavy edges of the colony, with a small flat elevation in the centre, with a rough and dry surface. Grown colonies, as a rule, easily separated from the surface environment, well suspensions in physiological solution. The diameter of the colonies on the third day of cultivation reaches 2-3 mm

On the surface mycopathologia agar in Petri dishes or on the beveled surface of the agar in test tubes are formed of individual, small, round colonies, grayish-white color. Colonies grown on the surface of MPA, does not adhere tightly to the surface environment and easily dislodged and fall is investing gives slightly visible film on the surface environment and weak turbidity of the broth. When shaken film and the precipitate dissolved and slowly settle to the bottom of the tube.

In thioglycolate broth of the microorganism forms oblatory, malahayati sediment at the bottom of the tube and along its walls.

Maximum growth on nutrient media notes for 3-4 days cultivation at a temperature of +37oC under anaerobic conditions.

The strain contains urease, does not contain catalase and gelatinase, not reduces nitrate to nitrite, ferments maltose, ribose, xylose, sorbitol, glucose, sucrose.

In susceptible animals - sows - infection in the bladder using a special catheter at a dose of 5 ml with content 5108microbial bodies through 2-6 months at slaughter found urocystic, in most cases, hemorrhagic or necrotic in nature, in some cases, the chronic course of the infection - purulent pyelonephritis.

For more information about the strain.

The microorganism does not cause hemolysis and gives a positive CAMP test reaction.

Molecular characterization:

1. Spectral response:

D 260 - 1.51

D 280 - 0.78

A 260/280 - 2.4

A 260/230 - 1.93

2. The concentration of 0.23 mg/ml

3. Temperature is ü selection of nucleic acids from DNA.

Upon receipt of 16S RNA sequences of several dozen strains of Actinobaculum suis isolated from patients sows, as well as from the content prepucialna diverticulum clinically healthy boars in the Omsk region, strain 2 And assigned to the first group (Pleshakova C. I., D. Konin Century - unpublished data).

The microorganism is relatively weakly resistant to the high temperatures at +55oC maintains viability for 30 minutes at +60oC - 20 minutes.

When ultraviolet light from a distance of one meter microorganism dies within 30 minutes (Lipatov PP, buyalskyy K. M. (3)).

The strain is not sensitive in vitro to gentamicin, but it is destructive act: oxytetracycline, penicillin, oxacillin, erythromycin, chloramphenicol, amoxicillin, streptomycin, ampicillin, minocin, neomycin, lincomycin.

Given the availability of the proposed strain number of distinctive properties from those described in the foreign literature analogues, such as multiple other content G+C DNA is 54.5 mol.%, relation to a particular (1) a group of organisms of a given species by 16S RNA sequence, several other biochemical properties (ferments maltose, glucose and sugars is the reference strain.

In addition, the strain Actinobaculum suis 2 And can be used for cooking-based assays (antigen for serological reactions and reactions of indirect immunofluorescence).

Strain Actinobaculum suis 2 A 0699, deposited in the Institute of culture collections of microorganisms State scientific center of VB Vector used for preparation of antigen-based assays for serological reactions and reactions of indirect immunofluorescence for the diagnosis of urocystitis and pyelonephritis in sows.

 

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