Specimen CollectionIn Infectious Diseases : Specimen CollectionIn Infectious Diseases Dr.T.V.Rao MD
Professor Of Microbiology Why specimen collection is Important in Microbiology : Why specimen collection is Important in Microbiology Specimen collection in Microbiology to isolate and identify the causative agents forms back bone of the investigative procedures.
In developing world, lack of awareness and casual attitude among junior staff hampers the definitive diagnosis.
Specific procedures in collecting specimens will certainly improve the quality of services of Microbiology Departments Important questions before collecting a specimen : Important questions before collecting a specimen Are you suspecting an Infection ?
If so what is the Nature of infection, eg Bacterial, Viral, Mycological or Parasitological
Which tests are your priority ?
When to collect the specimen ?
How to collect the specimen ?
Am I choosing the correct container ?
Why to send the specimens promptly
if not what I should do ? Fishing for Diagnosis in Laboratories, Is it worth? : Fishing for Diagnosis in Laboratories, Is it worth? The physicians and Microbiologists should be aware of the clinical manifestations, before undertaking the test.
Microbiological tests are expensive and technically demanding
Causal testing of Microbiological tests are counterproductive. Policies on Specimen Collection. : Policies on Specimen Collection. Every laboratory should formulate guidelines on procedures for each major category of specimens and requests.
Every laboratory should assist extra examinations,outwith the standard procedures may be required if specifically requested by the Physician or if the clinical information provided on the request form suggests that an unusual infection may be present. Why Proper written Request : Why Proper written Request Your request is a legal document.
Identifies all the outcome of test.
No interchange of results.
Short forms are dangerous
Signature of the Doctor / Nurse is essential in legible form, can help to contact in case of results which can save a patient.
When the patient is serious, write a Tele contact number which can help in prompt delivery of results An Ideal Request form : An Ideal Request form Name xxxx Age Sex
IP/ OP No xyz Time Date
Ward xx123 Urgent / Routine
Nature of specimen
Contact No 1234567 When one Expects the Results : When one Expects the Results On sending the sample the Physician will be anticipating the early reports, the Microbiologists should promptly dispatch results in all life saving investigations.
However the Doctors must be made aware limitation of the investigations and discuss the pros and cons of the Laboratory reports When to Repeat Diagnostic Tests : When to Repeat Diagnostic Tests On many occasions less than ideal sample is received in laboratory.
The rejection of clinical samples should be done with great care and wisdom of only senior staff who should take the responsibility.
In the welfare of the patient samples can be repeatedly collected for better diagnosis, as we need repeated isolation to confirm uncommon pathogens. When to Collect the Earliest Specimen : When to Collect the Earliest Specimen Start collection of specimens for all cultures before starting an Antibiotic.
The advice is ideal but may not be possible, as many prescribe Antibiotics before considers the Microbiological diagnostic options. When to Request Transport Medium : When to Request Transport Medium When facilities are not available to perform the desired tests at the place of collection or laboratory located far away, request the Diagnostic laboratories to advice on transportation of specimens, and consider how to preserve and transport in ideal medium before it is processed
Popularly used transport medium are
Amie 's transport Medium
Stuart's Transport Medium
V.R Medium What containers to use : What containers to use Containers must be leak proof,
For cultures sterile containers a Must Label High risk Specimens : Label High risk Specimens Sputum with suspected Tuberculosis
Fecal samples suspected with Cholera, Typhoid,
Serum when suspected with HIV/ HBV/HCV, infections Slide 14: Blood for Culturing Blood Collection for Culturing : Blood Collection for Culturing Most important investigation
An appropriate procedures in collection and processing, identifying and timely reporting can be Life saving Collection of Blood : Collection of Blood A scientific approaches and dedicated staff participating in blood collection will eliminate the basic failure as Contamination
Improper handling of syringes increases chances of contamination
Contamination hampers the ideal reporting,
A valuable time is lost
The goal in blood collection is avoiding the contamination Collecting the Blood for Culturing : Collecting the Blood for Culturing Teach the staff how to collect the Blood.
The nurse are advised on principles of aseptic precautions by self as washing hands and wearing gloves
Proper areas of disinfection with good antiseptic solutions. Hygienic precautions will decrease contamination : Hygienic precautions will decrease contamination The staff should be advised how to disinfect the skin over vein, to use a fresh sterile syringe for the venepuncture with fresh sterile needle before inoculating culture bottle
The staff should disinfect their hands before doing the procedure. Proper handling of Syringe is essential to obtain a blood specimen : Proper handling of Syringe is essential to obtain a blood specimen The staff should hold the needle by its butt, not shaft. Either with sterile forceps or with fingers covered with a dry sterile rubber glove, and protect self with potentially infective pathogens Do not collect from existing or indwelling catheters : Do not collect from existing or indwelling catheters The staff are warned that contamination is very likely if the specimen is collected from an indwelling peripheral venous catheter instead of from a fresh venpuncture. Always collect the Blood specimens in Hygienic areas : Always collect the Blood specimens in Hygienic areas All procedures in relation to processing of the samples should be done in a sterile environment, or bacteria free areas.
Despite insistence on aseptic precautions, most laboratories report finding contamination in 1-5% of the blood cultures. Slide 22: Cerebrospinal fluid examination Specimen collection for CSF Examination : Specimen collection for CSF Examination Lumbar puncture to collect the CSF for examination to be collected by Physician trained in procedure with aseptic precautions to prevent introduction of Infection. Procedure to collect CSF : Procedure to collect CSF The trained physician will collect only 3-5 ml into a labeled sterile container
Removal of large volume of CSF lead to headache,
The fluid to be collected at the rate of 4-5 drops per second.
If sudden removal of fluid is allowed may draw down cerebellum into the Foramen magnum and compress the Medulla of the Brain CSF needs a New and Sterile container : CSF needs a New and Sterile container Fresh sterile screw capped container to be used.
Reused containers, not to be used, contamination from the previous specimens misrepresent the present specimen. Lumbar puncture for CSF collection : Lumbar puncture for CSF collection The best site for puncture is inter space between 3 and 4 lumbar vertebrae
( Corresponds to highest point of iliac crest )?
The Physician should wear sterile gloves and conduct the procedure with sterile precautions, The site of procedure should be disinfected and sterile occlusive dressing applied to the puncture site after the procedure. Transportation to Laboratory : Transportation to Laboratory The collected specimen of CSF to be dispatched promptly to Laboratory , delay may cause death of delicate pathogens, eg Meningococci and disintegrate leukocytes Preservation of CSF : Preservation of CSF It is important when there is delay in transportation of specimens to Laboratory do not keep in Refrigerator, which tends to kill H. Influenza
If delay is anticipated leave at Room Temperature. Slide 29: Upper Respiratory Infections What are Upper Respiratory Infections : What are Upper Respiratory Infections The commonest respiratory infections are localised in Oropharynx, Nasopharynx, and nasal cavity,
Causes Sore thraot,nasal discharge and often fever. Infect larynx,otitis media,sinusitis,conjunctivitis or keratitis.
May present with serious diseases whooping cough, influenza , measles and infectious mononucleosis. Aetiological agents in Upper Respiratory Infections : Aetiological agents in Upper Respiratory Infections In most cases the primary infections are caused by virus, difficult to isolate.
But many infections are caused by concomitant carriage or secondary infection with one of the potential pathogens present in the Nasopharynx 1 Pneumococcus .Haemophilus influenza Staphylococcus aureus, and Streptococcus pyogenes.
Drug resistant coli form bacilli or yeasts may dominate the throat flora in patients receiving antibiotics. Specimen collectionin Throat Infections : Specimen collectionin Throat Infections A plain cotton wool swab should be used to collect as much exudates as possible from tonsils, posterior pharyngeal wall and other area that is inflamed or bears exudates Cooperation of the patient and ideal techniques contributes better results : Cooperation of the patient and ideal techniques contributes better results If cooperated by patient, the swab should be rubbed with rotation over one tonsillar area of the soft palate and uvula, the other tonsillar area and finally the posterior pharynx Collecting the Swab : Collecting the Swab An adequate view of throat should be ensured by good lighting conditions and the use of a disposable wooden spatula or a tongue depressor to pull outwards and so depress the tongue. Transportation of Throat Swabs : Transportation of Throat Swabs The swab should be replaced in its tube with care not to soil the rim
If it cannot be transported immediately to laboratory it should be placed in a refrigerator at 4ºc until delivery or preferably submitted in a tube of transport medium Nasal specimens : Nasal specimens A deep nasal swab generally yields the same information as throat swab.
Nasal swabs are taken to detect healthy carriers than diagnose deep infection
Deep nasal are taken to diagnose S.pyogenes and Diphtheria bacillus. Specimens in sinusitis : Specimens in sinusitis Pus collected or aspirated from sinus, or a saline wash out should be examined in a Gram film and by culture on aerobic and anaerobic blood agar plates. Collection of Ear SwabsAcute Otitis Media : Collection of Ear SwabsAcute Otitis Media Acute Otitis Media – as long as eardrum remains intact, none of the infected exudates can be collected on an ear swab , though culture of the throat swab may give a provisional indication of casual organism Chronic suppurative Otitis media : Chronic suppurative Otitis media Swabs of the discharge in the external meatus should be cultured to guide the choice of antibiotics for systemic and topical therapy. Otitis externa : Otitis externa A swab should be taken from the meatus and cultured aerobically on blood agar and MacConkey agar plates for the bacteria.
All specimens should also cultured on Sabouraud’s agar plate with Nystatin 50 units for Candida and Aspergillus. Eye Swabs : Eye Swabs Obtaining a adequate specimen is difficult.
It is best to make smears and seed culture plates beside the patient immediately after collecting the material from the eye. Collection of Eye swabs : Collection of Eye swabs It is ideal to pick up the material with a loop or on the smoothly rounded tip of a thin glass rod or on the thin serum coated swab
Clinical material from Conjunctiva, i.e. from averted eyelid,
The margin of the eyelid should be avoided. Slide 43: Specimens for Lower Respiratory Infections. Lower Respiratory Infections : Lower Respiratory Infections Sputum is the material from the lower respiratory infections most commonly submitted for bacteriological examination. The sputum is a mixture of bronchial secretions and inflammatory exudates coughed up into the mouth and expectorated
There are several difficulties both in collecting a suitable sample and interpreting the results of the culture
Busy and uninstructed staff may send collection of saliva to the laboratory.
On several occasions repeat sample may be required to isolate the causative agent. Instruction for collecting sputum : Instruction for collecting sputum Make the collection in a disposable and wide mouthed screw capped plastic container of 50 – 100 ml capacity.
Collect sputum before antibiotics are given.
Ideal to have when patient wakes up and with first cough. Precautions in handling the specimens : Precautions in handling the specimens Avoid spilling the material over the rim.
Tightly screw on the cap of the container.
Wipe off any spilled material on its outside with tissue paper
Deliver the specimen quickly to laboratory Sputum Examination for Tuberculosis : Sputum Examination for Tuberculosis Specimen should be collected with biosaftey precautions.
Several specimens should be collected before coming to negative conclusions. Slide 48: Specimens for Urinary Tract Infections Collecting Urine for examination : Collecting Urine for examination Collect the Mid stream specimens of Urine
Do not collect spontaneously passed urine without instructions, which can lead to contamination with commensals bacteria Colonized on urethral orifice and perineum Specimen Collection : Specimen Collection The urine collected in a wide mouthed container from patients
A mid stream specimen is the most ideal for processing
Female patients passes urine with a labia separated and mid stream sample is collected How Urine Specimens collected in young and infants : How Urine Specimens collected in young and infants Non invasive methods are safe and ideal
Follow the Broom hall method,
By tapping just above the pubis with two fingers placed on supra pubic region after 1 hour of feed, tapping on at the rate of 1 tap/second for a period of 1 minute, if not successful tapping is repeated once again.
The child spontaneously pass the Urine and to be collected in a sterile container Transport of Urine for Culturing : Transport of Urine for Culturing All collected specimens of urine to be transported to laboratory with out delay
Delay of 1 – 2 hour deter the quality of diagnostic evaluations.
If the delay is anticipated the specimens are at preserved at 40c
In field conditions Boric acid can be added at a concentration of 1.8 % Slide 53: Genital Tract Infections Genital Infections in women : Genital Infections in women Genital infections present with, arthritis, vaginosis, genital ulceration, cervicitis, uterine sepsis, salphingits, oophoritis, and pelvic inflammatory disease. Collection of specimens : Collection of specimens The specimen commonly collected for the diagnosis of vaginiti’s, vaginosis or uterine sepsis is high vaginal swab
The swab is inserted into upper part of the vagina and rotated there before withdrawing it. Specimen collection in Gonorrhoea : Specimen collection in Gonorrhoea An endocervical swab must be collected for examination for gonococci.
A vaginal speculum must be used to provide a clear sight of the cervix and swab is rubbed in and around the introitus of the cervix and withdrawn without contamination from vaginal wall. Specimens from other genital areas : Specimens from other genital areas Other swabs should be collected from any exduate discharged from the meatus of the urethra or a Bartholin's gland.
Rectal or pharyngeal swabs should be considered depends on sexual habits of the patient Transportation of specimens : Transportation of specimens All the swabs to be promptly transported to laboratory, in cases of delay or in cases of delicate microbes to be transported in Amie's transport medium.
If possible two swabs to be collected and submitted for each site. Specimen collection in Men : Specimen collection in Men The infection in men are mostly caused by the same organism as in women.
Urethritis is commonest presentation may be caused by Gonococci or Non-gonococcal.
May present with Genital Ulcers. Gonococcus infection in Men : Gonococcus infection in Men The specimen is collected by milking the urethra and urethral discharge is smeared on slides and inoculated on warmed plates of heated blood agar or selective medium for isolation of Gonococci Other Genital infections : Other Genital infections When prostatitis is suspected and there is no spontaneous discharge from urethra, massage of the prostate per rectum may express some exduate for examination, and culture. Collection of specimens in Chancres : Collection of specimens in Chancres The examination of chancre requires the careful collection of exudates and its preparation for dark ground microscopy.
Many patients need clotted blood for specific serological investigation. Slide 63: Wound, Skin, and Deep Sepsis Collection of specimens : Collection of specimens Pus or exudates is often submitted on a swab for laboratory investigation.
The swabs are inefficient sampling device and tends to desiccate the specimen and trap the bacteria which are then not released on to culture plate Ideal samples from wounds : Ideal samples from wounds The ideal sample is pus or exudates should be submitted in a small screw-capped bottle in firmly stoppered tube or syringe or a sealed capillary tube.
Fragments of excised tissue removed at wound toilet or curettings from infected sinuses and other tissues should be sent in a sterile container. Slide 66: Gastrointestinal Infections Collection of specimens : Collection of specimens Whenever possible, a specimen of faeces should be collected.
A rectal swab is unsatisfactory, unless it is heavily charged and visibly stained with faeces collected from rectum, not anus Collection of Faeces : Collection of Faeces The specimen may be collected from faeces passed into a clean bed pan, not mixed with urine, or disinfectant or from the surface of heavily soiled toilet paper.
The specimen is collected into 25 ml screw capped wide mouthed disposable container. Transportation of specimen : Transportation of specimen Collect 1-2 ml of faeces, and apply the cap tightly.
Take care not to soil the rim or outside of the bottle.
Transmit the container quickly to laboratory.
If delay is unavoidable and particularly when the weather is warm collect the specimens in a container holding 6 ml buffered glycerol saline transport medium Specimen collection for Virological diseases : Specimen collection for Virological diseases Direct virus diagnosis depends on the detection of virus particles, viral antigen or viral nucleic acid in specimen taken from the site of infection.
Specimens should be delivered promptly to the laboratory so that no deterioration in the quality of the cells occur.
However the specimen collection techniques alter depending on the Aetiological agent and site of involvement. Training the Paramedical Staff : Training the Paramedical Staff Nursing staff and Medical Assistants are the strengths of specimen collection in the Developing world
All the progressive Medical Institutes should train their staff in proper and scientific methods in specimen collection and a fraction of Morbidity and Mortality can be reduced.
Dr.T.V.Rao MD Slide 72: Created for awareness on specimen collection in developing world among the Medical and Paramedical staff