Introduction
The bacterial flora of the mouth is very complex due to the fact that the mouth acts as the point of entry of many substances into the body. This bacterium can both proliferate aerobically and anaerobically. The types of bacteria found in the buccal cavity are so diverse and include lactobacilli, staphs, Candida, and streptococci among other species. Types of microbes isolated from the mouth greatly depend on the site of the mouth from where they were collected. The tooth surfaces are normally colonized by Streptococcus mutans especially among people who eat too much confectionary (Abrams and Bottle 1996). Its presence in the mouth leads to reduction in pH hence growth of lactic acid bacteria like the lactobacilli. Wounds are normally colonized by Staphylococcus aureus, Escherichia coli, Streptococcus pyogenes, pseudomonas, Proteus and Acinetobacter. Acute wounds are normally colonized by Staphylococcus aureus followed by the beta haemolytic groups. Because the wound from the bite is still fresh, the possibility that the infection was occasioned by Staphs aureus or the Streptococcus is vey high.
Clinical presentation of samples
Signs and symptoms shown on skin lesions resulting from human bite vary depending on the number of days that have elapsed before urgent medical intervention is taken. Patients who seek services of the physicians after seven days with fight bites normally have open cut wounds, swollen hands, extreme pains, limited mobility of the hand, increased temperatures around the wound, and erythema. Occasionally, there may be toxic syndrome signs, fever and lymphadenopathy (Chadev & Jukhtin 1996). At the primary stages the wound infection may appear like an abrasion or like lacerations found on the epidermal tissues of the hand with mild cellulites. The physician attending to the emergency should treat the wound immediately and carefully.
Sample collection and identification of the samples
Samples are collected using a sterile swab that has been dipped into 70 per cent methylated spirit. The swab is used to collect either pus or cells from the infected wound especially the superficial wounds (Jorgenson and Kloos 1987). Taking of samples from deeper wounds should be done using a syringe. This ensures that fluid aspirations are taken or alternatively tissue biopsy can carried out. This is done to allow for the recovery of bacteria that proliferate in both aerobic and anaerobic conditions. The sample obtained from these processes is then cultured in a culture media that allow the growth of bacteria that may be present in the wound. The culture should allow for testing and identification of the isolates that have grown in it. Human bites contribute to growth of many pathogens in the wound (Kelly, Cunney and Smith 1996). A lot of care has to be taken when it comes to the handling of the samples so that the bacterial growth in culture media is enhanced. This increases the chance of their identification and detection. Anaerobic bacteria should not be exposed to aerobic conditions and vice versa as this jeopardizes chances of detecting such bacterium.
Identification process should be step by step where bacteria growing in culture media is taken through a series of tests and valuation (Kloos and Schleifer 1986). Tests that can be done include gram staining which separates gram positive bacteria from gram negative bacteria; microscopy which can also help in classification of bacteria as gram positive and gram negative as well as helping in bacterial classification according to their shapes; and biochemical tests.
Transportation of cultures to the laboratory
Sample collected through tissue biopsy should be placed in screw capped vials having sterile saline (Kloos and Schleifer 1975). The solution helps in keeping the tissues moist. Samples that are suspected to be having anaerobic bacteria are normally placed in vials that are gassed out having pre -reduced medium. They can alternatively be put in sealed gassed out bags. For anaerobic sampled that have been collected using syringes, the syringe should be capped immediately after carrying out aspiration. Aerobic bacterial samples should be put in Stuart with which they are transported to the laboratory. Samples having anaerobes should be taken to the laboratory in PRAS transport medium (Kohne 1984).
Laboratory processing
Culturing of Staphylococcus aureus
From a solid media where the samples had been cultured a growth with morphology similar to staphylococci can be picked using sterile inoculation loop. Trypticase soy incubated at 37 degree Celsius for 72 hours can be used. About 50 microliter of the sample should be pipetted into probe reagent tubes (Morse 1981). Sample is prepared by opening the foil pouch. Sufficient numbers of probe reagent tubes should be removed to aid in testing the culture isolates. The tubes should then be labeled. Their cups should be removed and retained. 50 micro liters of the lysis reagent should then be pipetted into the reagent tubes (Mann, Hoffeld and Farmer 1977). Lysis reagent should not be used with broth culture. Samples should then be transferred from the solid media into the labeled reagent tubes. Inoculating loop should then be used to remove the cells. The reagent tubes should be recapped and incubated at 37 degree Celsius for 5 minutes in a water bath. To hybridize, reagent tubes should be removed from water bath and added with 50 microliter hybridization buffer afterwhich the tubes should be recapped and incubated at 60 degrees Celsius for 15 minutes in water bath. In selection, the tubes should be removed from water bath. 300 microliter of the buffer reagent should be pipetted into the tubes. Mixing should be properly done. Tubes should be kept in an incubator for five minutes at 60 degree Celsius in an incubator. Tubes should then be removed from water bath and left at room temperature for 5 minutes. Tubes should be dipped into luminometer for analysis. Identification should now be done based culture identification test which involve use of gram staining, tube coagulase test, and Staphaurex (Mennen and Howells 1991).
Antibiotic sensitivity testing for Staphylococcus aureus
To be able to investigate S. aureus sensitivity pattern, isolate from the human bite wound was tested against a variety of antibiotics. Sensitivity testing is normally done using disc diffusion technique. The discs are made from Whattma’s No. 1 paper. Different varieties of antibiotics should be used in varying quantities. 10mg of Ofloxacin, 30 mg of Amoxycillin, 25 mg Co-trimaxazole, 30 mg of Ampicillin, 5mg of Cloxacillin, and 25 mg Tetracycline should be used (Perron, Miller and Bradey 2002). Swabs dipped into 70 per cent alcohol should be used to smear the test organisms onto an agar. Antibiotic discs should then be introduced into the plates. Inoculated plates are then put on the bench for thirty minutes for proper diffusion to take place. Plates should be inverted and diameter of inhibition later measured.
Interpretation of the findings
Using disc diffusion technique to test for sensitivity patterns the diameter of the inhibition zone falling in the range of 0-5 mm should be taken to mean resistance. The diameters falling between 5-15mm should be designated sensitive I and diameters falling in the range of 15-25mm ought to be designated sensitive II. Finally, diameters between 25-35mm is designated sensitive III.
Isolates should be positive for Staphs when they have RLU which is more than 50 000 or negative when RLU is below 50 000.
Conclusion
Care should be taken when isolating microorganisms from wounds caused by human bite because after some time the normal microbial flora of the areas around the wound also invade the wound.
Reference list
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Chadev, A.P., Jukhtin, V.I., Butkevich A.T. Et al. 1996.Treatment of infected clench fist human bite wounds in the area of metacarpophalangeal joints. J. Hand Surg. Am, 21: 299-303.
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Morse, S. I. 1981. Staphylococci. p. 275-281. In A. Braude (Ed.). Medical microbiology and infectious diseases. W. B. Saunders Company. Philadephia, PA.
Perron, A.D., Miller, M.D., Brady, W.J. 2002. Orthopedic pitfalls in the ED: fight bite. American J. Of Emergency Med. Vol 20 N2.