Adequate collection and transport and processing of clinical specimens for microbiology evaluation are important requirements in the diagnosis of infectious diseases.
BASIC PRINCIPLES OF SPECIMEN COLLECTION:
The following basic principles of specimen collection may serve as guidelines to ensure appropriate collection process.
- A culture specimen should be taken in the acute phase of the infection and before antibiotics are administered.
- Proper selection of a site of a culture should be done.
- It is important to culture the infecting agents while avoiding the usual flora and colonizing organisms.
- Test results must be carefully compared with the suspected diagnosis.
Appropriate collection techniques:
Aspirates & Tissues:
Aspirates of sterile body fluids are collected using sterile techniques and are best in quality and quantity of specimens. A piece of tissue is also an excellent specimen, but it should be protected from drying during transport.
Lesions, wounds, and abscesses seem to present the most problems related to quantity and quality. An aspirate of pus is the ideal specimen. In deep wounds a sample collected from the walls of the abscess in addition to pus sample provide an enhanced opportunity for thorough assessment. When multiple sites appear infected an aspirate of a fluctuant, unopened wound is far superior to a swab from an open draining wound.
Swabs:
Swabs are used only as a last resort and are considered inferior to other collection methods.
If a swab must be used instead of an aspirate or piece of tissue, four basic rules must be followed.
- Clean the wound to remove commensals and contaminant organisms.
- Explore the wound to find out deep seating organisms.
- Obtain fresh culture material from infected site, but not from any collection.
- Obtain adequate quantity of material.
Need for repeat culture:
It is important to remember that even with therapy wound do not generally change overnight. Repetitious culture of wounds is a waste of time and money. Monitoring the situation with Gram stain is far economically efficient than repeat culturing.
Patient education and preparation:
Patient education: Specimens such as urine, stool, and sputum are commonly obtained by the patients and must often require extensive patient education. The most effective educational tool is to give the patient a preprinted sheet of instructions using simple language and pictures to help the patient follow along and understand as the procedure verbally described.
Urine: Skin preparation and collection of mid-stream urine is important. The collection of first morning specimen provides a more concentrated sample with the potential for the optimal results.
Sputum: First morning sample after mouth cleaning is ideal. If sample appears normal, it should not be sent to laboratory, another specimen should be collected. Infectious secretions exhibit such abnormalities as pus, blood and dark flecks.
Stool: Three vial kits are recommended. Each such kit contains a vial with a preservative for parasites, a vial with a preservative for enteric pathogens and clean vial for examination of raw specimen and additional testing. Contamination of urine should be avoided.
Preservation, storage and transport of specimens:
Transportation and preservation of the specimens is very important for the following major concerns,
- Overgrowth of rapidly growing organisms
- Death of organisms resulting from changes in temperature and pH
- Inaccurate quantitation of organisms in urine and quantitative tissue
- Loss of organisms resulting from drying
- Protection from oxygen
- Protection from clotting
- Safety of the transporter
Maintenance or protection of the primary specimen can be aided by the use of preservatives, anticoagulants, and holding media and even culture media.
Use of preservatives: Urine and stool commonly require preservatives. Boric acid used to maintain accurate urine colony counts. Phosphate-buffered saline helps to preserve feces so that fragile organisms will not die of temperature and pH changes.
Use of anticoagulants: Anticoagulant is important for any specimen that might clot, because organisms bound up in clotted material are difficult to grow. Sodium polynethol sulphate is the most common anticoagulant used for microbiology specimens. Heparin is another anticoagulant that can be used selectively in virology. Citrate or EDTA should not be used for microbiology specimens.
Use of holding and transport media: Swab collection systems contain a holding medium that maintains viability of the organisms but does not permit an increase or decrease in numbers of organisms. Modified Stuart transport medium and Cary-Blair transport medium are commonly used.
Blood is usually placed in a broth culture medium immediately after collection to quickly grow and identify the organisms present in the potentially life-threatening condition.
Unprotected specimens: Many specimen types are transported without special protection. These include sputums, body fluids, tissues, catheters, medical devices, and specimens for sterility. It is important to process such specimens without delay as soon as possible.
Storage of specimens: Urine, viral blood specimens, catheters, and swabs should be refrigerated, but bacterial blood specimens, CSF and cultures processed onto agar plates at the bedside should be placed in an incubator and these samples should not be refrigerated. Room temperature is most appropriate for specimens such as hair and nails for isolation of fungus. Respiratory specimens and stool specimens may be refrigerated depending upon suspected pathogens but best is to process it without delay.
Unacceptable specimens:
The information on the requisition must match with information on the specimen label.
Conditions in which specimens should be rejected.
- Leaking specimens
- Syringe with needles attached
- Stools contaminated with urine or barium
- Anaerobes on inappropriate sources
- Unpreserved specimens more than 2 hours old
- Refrigerated blood cultures
- Dried-up specimens
- Specimens in formalin
Prioritization:
All specimens require prompt processing after arrival in the laboratory. Processing every specimen as soon as it is received is often impossible. The laboratory must set guidelines for prioritizing specimens on the basis of various factors.
Table: Level of specimen prioritization.
| Level | Description | Specimens |
| 1 | Critical /
Invasive |
CSF, Brain, Blood, Heart valves, Pericardial fluid, Bronchoalveolar lavage, Vitreous or Aqueous fluids |
| 2 | Unpreserved | Sputum, Tissue, Stool, Body fluids, Drainage
from wounds, Pus, Bone |
| 3 | Accuracy or Quantitation affected | Urine, Tissue, Catheter |
| 4 | Protected / Preserved | Swabs in holding medium |
The processing of clinical specimens:
The processing of specimens includes the following:
- The entry of essential data into a log book or computer terminal;
- Visual examination and determination of whether all criteria for acceptance are met;
- The microscopic examination of direct mounts or stained smears to establish a presumptive diagnosis.
- Processing for cultures.
