SALMONELLAS :
BACTERIOLOGICAL STUDY AND DIAGNOSIS
by : Salim djelouat
The Salmonellas spp. is present in the intestine of the man and the vertebrate animals.
In the external medium, the salmonellas can survive of the weeks, even of the months if the conditions of temperature, moisture and pH are favorable.
01 - Taxonomy
The salmonellas constitute a kind containing only one species: Salmonella enteric divided into 7 subspecies.
The nearly totality (99,8%) some stumps responsible for human infestations belong to a subspecies also named enteric.
One distinguishes close to 2000 sérovar (one even speaks of 3500) in this subspecies, according to their antigenic constitution.
For historic reasons, the sérovar has been assimilated to the species, one designates every sérovar by a name recalling either her pathogenic power (Salmonella Cholera is) either the name of the city of the first isolated (Salmonella London for example).
The orthodoxy of the classifications would force to indicate the salmonellas by the names of the kind, the species, the subspecies and the sérovar (which can be replaced by the antigenic formula):
Example:
- Salmonella enteric, subs enteric, London sérovar,
Or
- Salmonella enteric, subs enteric, sér 3, 10,: l, v,: 1,6
To simplify, is admitted to designate the salmonellas by the name of the kind followed of the one of the sérovar dedicated by the use, but this last writes itself in right characters and with a capital: London salmonella.
02 - Classsification of Kauffman-white
This classification rests on the determination from the antigens O, H, and Vi:
1 - Antigens 0, to the number of 67, have the structure polyosidic previously described.
One determines them by agglutination on blade using specific immunes’ serums.
Some of these antigens, qualified the “major ones” characterize a group of salmonellas: thus the antigen O : 4 defines it the group B (see table).
Other antigens, “minors” theirs are associated but hardly have diagnostic importance.
2 - The antigens H separate the sérovar inside these groups.
One also highlights them by agglutination on blade.
The antibodies anti H make motionless the bacteria which relate to their whip specificities which correspond to them.
The antigens H exist under two phases which can coexist or not at the same stock:
- Phase 1 is indicated by small letters, a, b, c… Beyond z, the antigens carry letter z associated with a figure.
- Phase 2 is indicated by Arab numerals but some are it also by letters.
The process “of inversion of phase” of Svent Gard makes it possible to isolate one from the two phases:
- The mobile bacteria, cultivated on an agar impoverished in agar ("soft" agar), invade the whole surface of the middle.
If one incorporates to a soft agar a serum anti H specific of one of the two represented phases in the stump, the bacteria expressing this phase are immobilized and only the elements of the other phase are capable to distribute the point of inoculation where it is possible to isolate them from afar.
- The Vi antigen only exists at three sérovars (Salmonella Typhi, Salmonella para C and Salmonella Dublin).
Its presence can mask the antigen O, returning the stump "O inagglutinable."
This inhibition can be raised while heating the stump in 100° C, because the Vi antigen is thermo labile.
Extract of the table of Kauffmann-White
| GROUPS | SÉROVARS | ANTIGÈN O | VI | ANTIGÈN H | |
| PHASE 1 | PHASE 2 | ||||
| A | PARATYPHI A | 1, 2, 12 |
| a | - |
| B | PARATYPHI B | 1, 4, 5, 12 |
| b | 1, 2 |
| WIEN | 1, 4, 12, 27 |
| b | l, w | |
| SAINT PAUL | 1, 4, 12 ,27 |
| e, h | 1, 2 | |
| TYPHI MURIUM | 1, 4, 5, 12 |
| i | 1, 2 | |
| BRANDENBURG | 1, 4, 12 |
| l, v | e, n, z15 | |
| AGONA | 1, 4, 12 |
| f, g, s | - | |
| DERBY | 1, 4, 12 |
| f, g | - | |
| C | PARATYPHI C | 6, 7 | + | c | 1, 5 |
| VIRCHOV | 6, 7 |
| r | 1, 2 | |
| INFANTIS | 6, 7 |
| r | 1, 5 | |
| BOVISMORBIFICANS | 6, 8 |
| r | 1, 5 | |
| GOLDCOAST | 6, 8 |
| r | l, w | |
| D | TYPHI | 9, 12 | + | d | - |
| ENTERITIDIS | 1, 9, 12 |
| g, m | - | |
| PANAMA | 1, 9, 12 |
| l, v | 1, 5 | |
| DUBLIN | 1, 9, 12 | + | g, p | - | |
| GALLINARUM | 1, 9, 12 |
| - | - | |
| E | LONDON | 3, 10 |
| l, v | 1, 6 |
| ANATUM | 3, 10 |
| e, h | 1, 6 | |
| GIVE | 3, 10, 15 |
| l, v | 1, 7 | |
| SENFTENBERG | 1, 3, 19 |
| g, s, t | - | |
| MELEAGRIDIS | 3, 10 |
| e, h | l, w | |
03 - Habitat and pathogenic capacity
The salmonellas can survive several weeks in dry medium and several months in l' water.
- Salmonella can survive more than 300 days in the soil
- More than 30 months in droppings
- More than 9 months in water.
The poultries, the bovines and the sheep being frequently contaminant animals, the salmonellas can find themselves in the believed food, in particular meats and eggs.
A/ Stocks of Salmonella enteric subsp. enteric is lodged in the intestine of the man and the endothermic animals
B/ The sérovars Typhi, Paratyphi A, Paratyphi C and Sendai are insulated only at the man and they cause typhoid syndromes.
C/ The sérovars Abortus bovis and Abortus equi are, respectively, specific sheep and équidés and they are at the origin of abortions.
D/ The sérovar Gallinarum is specific poultries.
It is the agent etiologic of the pullorose and the typhose.
It is the agent etiologic of the pullorose and the typhose.
E/ The sérovar Typhisuis is present only in the pig and it causes infections in the young animals.
A many sérovars is ubiquists and at the origin of food toxinfections at the man and various infections in the animals.
The salmonelloses can give place to three types of clinical demonstrations.
1. Bacteriemic forms, strictly human and which are the typhoid fevers and paratyphoid due to Salmonella Typhi, Para A, Para B and Para C.
They are bactériémies with lymphatic starting point.
They are bactériémies with lymphatic starting point.
2. Food toxinfections giving place to gastroenteritis due to all the others sérovars but also to Para B and C
3. Extra-digestive demonstrations in which various sérovars is in question and which are more frequent at the weakened subjects:
- bactériémies no typhoidal
- pleuro-pulmonary infections
- reached ostéo-articular: septic or reactive arthritis,
- Osteomyelitis, ostéite
- cardiovascular infections: péricardites, arteries, infections on prostheses
- urinary infections
- abdominal infections: cholécystites, abscess of the liver, abscess of the spleen
- infections of the central nervous system: meningitides, abscess of the brain, hematoma under-dural infected, epidural abscess
04 - Physiopathologies
The salmonellas have an entéro-invasive capacity and penetrate even in a number restricted by the digestive tract.
After an incubation which can be rather long 2 to see 3 weeks, they will cross the intestinal mucous membrane and will invade lymphoid fabric (plates of Peyer).
From the plates of Peyer, and by lymphatic circulation, they will pass in blood circulation, which will determine a septicemia.
It should be noted that septicemia is not a complication of the typhoid fever, but it falls under the normal evolution of the disease.
The complication of the typhoid fever can intervene on the level of the plates of Peyer, which can ulcerate and involve a perforation intestinal, to see a peritonitis.
According to the design of Reilly, the bacteriemic stocks with propagation multiply in the ganglia mesenteric and pass in blood circulation causing the bacteriemy or are destroyed on the spot and release the endotoxine responsible for the nervous and vegetative disorders of the typhoid one, from where the danger of the administration of a strong antibiotic amount which is likely to cause a massive lyses’ of the bacteria.
After cure, the convalescent can remain carrying germs during months or years, the bacteria persisting especially in the bile ducts.
1 - TYPHOÏDE FEVERS AND PARATYPHOÏDES:
They are caused by Salmonellas strictly adapted to the man,
- Salmonella Typhi (bacillus of Eberth)
- Salmonella Paratyphi A (bacillus of Schotmûller)
- certain stocks of Salmonella Paratyphi B (bacillus of Schotmûller), Salmonella Paratyphi C (bacillus of Hirschfeld) and incidentally Sendai salmonella
The typhoid one is characterized by a strong temperature, accompanied by giddiness’s and insomnia.
Then appears a diarrhea, dehydration, often one notes also a constipation.
Principal stages of the evolution of the typhoid fever:
- 1e day: headaches, insomnia (loss of sleep), giddiness (the eyes turn)
- 2nd day: the temperature goes up (100.4°F)
- 3rd day: the temperature continues to go up and can be accompanied by vomiting, constipation and epitasis.
- 4th day: the temperature also goes up and can reach 104°F or more.
- From the 5th day and more: one always notes a strong fever, the pulse is slow, appearance of small red buttons which appear on the body, the diarrhea settles, the patient is killed indifferent to all that occurs around him.
Sometimes, it is delirious.
Sometimes, it is delirious.
It loses the weight.
From the fourth week: there is complication if there were not treatment.
The diagnosis or the identification of typhoid is very difficult because this disease often merges with the influenza, paludism, with meningitis, a pulmonary infection, an angina or appendicitis.
2 - THE GASTRO-ENTERITIS
The Gastro-enteritis is caused by Salmonellas ubiquity present in the man and the animals.
The duration of incubation is generally 1 to 2 days and depends on the introduced amount, of the health of the host and the characteristics of stock of Salmonella.
The salmonellae appear by a fever, a diarrhea, vomiting and abdominal pains.
In adults of normal physical condition, gastro-enteritis disappears without treatment after 3 to 5 days on average.
05 - Epidemiology
The Salmonellas are eliminated by the feces and resist well in the external medium.
The man is contaminated by digestive tract.
The typhoid fever is endemic and poses a true problem of public health, practically in all the countries of the world and not only in the countries in under development, as it is often claimed.
The man is the only reserve of virus.
The contamination is done by drink water, food and in particular the poultry, meat products, eggs, the dairy products and starting from the saddles of patient or asymptomatic carriers.
The infected domestic animals can be a considerable source of contamination.
The contamination person to person is important at the child and the teenager and supported by the asymptomatic bearing (role of the hands).
The non typhic salmonellas are in constant increase, in particular in the industrialized countries.
The domestic animals or wild are the reserves of virus.
The exotic animals can be in question.
Collective food poisoning (the illness)(CFP) are due for more half to Salmonella enteric.
The contamination person on person is important in the child, supported by the asymptomatic bearing (role of the hands).
The resistance of the salmonellas is likely to increase with the uncontrolled use of antibiotics in the industrial breeding.
This practice contributes to spread resistant stocks multi.
It was as reported as the prevalence of the non typhic salmonellae’s to Salmonella Enteritidis and in Salmonella typhi murium is high at the sidemen’s.
The sérovars responsible for the typhoid fevers are strictly human and the only “reserve of virus” is the man himself and which can be either a patient, or a convalescent or an operational carrier.
These contaminations person on person explain occurred of the case of typhoid fever by small epidemics; they are direct, around a patient, or generally indirect by food ingestion soiled by the excreta.
Failing food hygiene thus increases the risk of occurred of the disease.
The sérovars responsible for gastro-enteritis are very widespread in the animal world and the domestic animals or of breeding are at the origin of the human contaminations.
The infestation is also done by digestive tract; by soiled food consumption consumed little cooked: dairy produces, meats, eggs, shells, sprinkled crudeness’s etc…
A source of important extension, particularly in hospital medium, is the contage “manu carried” around infants reached who eliminate from the salmonellas in the layers.
The salmonellae’s are more frequent at the end of the summer or at the beginning of the autumn, with the return of the holidays.
The voyages in the hot countries, the way of life “close to nature”, the consumption of “natural” products or “the soil” not asepticized increase the risks of infestation.
06 - Character bacteriological
The Salmonella are Enterobacteriaceae and have the general characters of them.
They are :
- GLUCOSE (+)
- LACTOSE (-)
- ONPG (-)
- H2S (+)
- URÉASE (-)
- INDOL (-)
- CITRATE (+)
- VP (-)
- PDA (-)
- MOBILITY (+)
Some sérovars are particular
07 - The direct biological diagnosis
A - FOR THE TYPHOIDAL SYNDROME ONE MAKES:
One seeks the Salmonellas, primarily by blood of culture or by stoll, but of other taking away can contain some.
1 – Blood of culture
Positive during the first week of the disease.
It is necessary to sow a volume of important blood being given the restricted number of salmonellas in circulation.
2 – Stoll (coproculture)
It can be positive the first week of the disease but especially after the first days; it can remain positive in the convalescent (bearing).
The stools are seeded on selective surroundings (Salmonella-Shigella agar or S.S agar, agar to the désoxycholate-citrate, agar to the green brightness): on these surroundings, the majority of the fecal germs is inhibited.
The presence of lactose and an indicator of pH make it possible to locate the negative colonies lactose, and also on elective medium of the germ (bubble selenite F, allowing the growth of Salmonella the detriment of the other bacteria of the intestinal flora commensally, this medium is very useful during a negative tracking for uréase.)
The salmonellas and the Shigella do not ferment this sugar.
The suspect colonies are mended and identified biochemically and serologically.
There are mediums of enrichment such as the medium with the selenite, the tétrathionate and the Malachite green of Rappaport which make it possible to reverse the proportion of the fecal germs and salmonella with the profit of these last, whenever their number is too weak to directly highlight them.
B - FOR THE SYNDROME OF GASTRO-ENTERITIDIS SALMONELLA ONE MAKES:
1- STOOL (COPROCULTURE)
The only process valid diagnosis is the Stoll which is positive as of the first hours of the disease.
The blood culture will be indicated only in rare septicemia complications.
The reaction of Widal does not have interest here.
08 - Biochemical identification
The biochemical identification must precede any attempt by determination of the sérovar which is done by agglutination on blade using immune’s serums.
09 - Research for agglutinations by serums tests
The research for agglutinations by serums tests, starting from insulated colonies, must base on research initially antigen O of group then using the table of Kauffmann White, one studies specificities H in a phase then in the other.
If one can highlight only one phase, it is necessary to seek the other by the process of Svent Gard or inversion of phase which consists in cultivating the stock on a soft agar in which antibodies directed against the phase already highlighted are built-in.
10 - Indirect biological diagnosis
1 - SEROLOGICAL AGGLUTINATION = REACTION OF WIDAL = TEST FOR TYPHOID FEVER.
The serological agglutination of a typhoid fever or serological test of Felix consists in seeking the antibodies of the patient in the presence of antigenic suspensions O and H of Typhi Salmonella, of the salmonellas Para A, B and C and of antigen VI.
The reaction becomes positive after approximately a week of evolution.
The serum is tested with respect to suspensions O and H of salmonella typhi and the paratyphic Salmonellas.
The Antibodies O appear the first but disappear little from time after the cure.
The Antibodies H appear a few days later, reach higher rates and can persist several months after the disease.
At the period of state, the 2 Antibodies O and H are present.
One can observe a Co-agglutination between the Antigen O of Salmonella typhi and paratyphic (antigenic relationship).
On the other hand, simultaneous agglutination in several suspensions H, without agglutination O or with very weak titles O, can meet at the old ones vaccinated.
Agglutination in only one antigenic suspension O without H corresponding can be explained by an infection with salmonella of the same group serologic but of another serotype.
E.g., an infection with Salmonella Typhimurium can give a positive reaction in the Antigen Para B - O but not in Para B - H.
The research of the anti Antibodies - VI could have interest at the carriers of germs.
The analysis of the results allows suspected the causal agent and to fix the date of the infection.
11 - Antibiogram and antibiotic treatment
The Salmonellas are generally sensitive to active antibiotics against the bacilli with negative Gram.
The treatment decreases the frequency of the complications and must be undertaken in hospital medium and never in ambulatory medium.
The antibiothérapy is founded throughout one 10 to 15 days.
Certain resistances are possible and imply the practice of an antibiogram.
The typhoid fevers are currently treated by a cephalosporin of third generation and in particular by the ceftriaxone (treatment of reference), but the fluoroquinolones are also used in the adult.
One can also prescribe an aminopénicilline (penicillin A).
Gastro-enteritis’s concern a symptomatic treatment primarily including/understanding mode and rehydration.
A mixed antibiothérapy: (sulfaméthoxazole-triméthoprime, fluoroquinolones) is useful only in the serious cases.
A new antibiotic, the gatifloxacine, proves very effective against the typhoid fever.
12 - Control effectiveness of the treatment
It will be practiced two (2) stools (coprocultures) at 48 hours of interval following the treatment to ensure itself of the complete cure of the patient.
13 - Vaccine and vaccination
The vaccine (for Typhi, Para A, Para B) consists of a mixture of Salmonella Typhi, Paratyphi A and Paratyphi B killed by heat and phenol.
Vaccination is done in three injections with recall; it is rather badly supported because of the great quantity of endotoxine which it brings and its effectiveness is incomplete.
It is obligatory only for the exposed subjects (personnel of health).
For several years, an injectable vaccine, made up of an extract of the polyoside capsular VI purified Typhi Salmonella has been available (Typhi VI).
It is practiced in only one injection, is well supported and protects for one duration from three to five years and protection in zone from endemic from 60%.
It is recommended to the travelers in departure for the regions where the typhoid one is endemic.
The protection which it confers limits to the typhoid fever other than paratyphoid’s and others salmonellosis.
Alive vaccines managed by oral way are (always) being studied
14 - Prevention and measure of hygiene
1 - TYPHOID FEVER
The prevention rests on the epidemiologic monitoring and the fight against the “fecal peril”.
The dissemination of the salmonellas can be stopped by:
2 - SALMONELLOSIS
Best protection against the risk of salmonellosis is a good cooking of food, in particular of the meats, with at least 149°F during 5 to 6 minutes).
For chopped frozen or frozen steak, cooking must be carried out without preliminary defrosting because it increases the risk of bacterial multiplication.
The cold blocks the development of the bacteria but does not kill them.
It is advised to preserve eggs at the refrigerator, to maintain cold the preparations containing eggs without cooking (mayonnaise, creams, pastry makings…) and to consume them more close possible of their manufacture.
Moreover, the most vulnerable people (people old, sick, infants, pregnant women) should avoid the believed or little cooked egg consumption.
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