Collection,Storage and Transport of Microbiological
Specimen
Dr.Rubina KamranPG trainee MicrobiologyDepartment of Pathology
P.I.M.SIslamabad
GENERAL RULES FOR COLLECTION &TRANSPORTATION
Strict aseptic techniques Collect specimen before administration
of antibiotics Prevent contamination from normal flora Optimum timing/ site Leak proof sterile container
Proper labeling Brief clinical history Prompt transportation to the laboratory
GENERAL RULES FOR COLLECTION &TRANSPORTATION…
Chain of collection &transportation
CRITERIA FOR REJECTION OF SPECIMENS
Missing or inadequate identification Leaking container Inappropriate container Insufficient quantity Unknown time delay Inappropriate requests e.g,oral
swabs,foley`s cathetar tip etc.
CRITERIA FOR REJECTION OF SPECIMENS…
Incomplete forms Inappropriate transport or storage
Ways to transport specimen
Cold chain Refrigerating or by keeping cool,ice box,thermos flask. urine
At 37C keep near body and put in incubators immediately or
Transport media.
Csf urethral,cervical ,semen blood culture
Objective of using transport Media
To keep the organisms alive suppress the growth of unwanted
organisms Outgrow the no. of wanted organisms
Transport Media
Amies transport medium
Carry-Blair for stool culture
Stuart media for aerobic specimen
Conti----
Cary Blair t.medium-----Enteric pathogens.
Amies t.medium------suspected anaerobic bacteria.
Stuart t.media------urethral &genital specimens.
INDIVIDUAL SPECIMENS COLLECTION AND TRANSPORT
SPUTUM
A clean,dry,leak-proof container to be used.
Patient is instructed to cough deeply.
Specimen must be SPUTUM not Saliva.
Avoid use of mouth-wash prior to collection.
Early morning samples should be collected.
Container for sputum
Transport of Sputum
Early dispatch.
Transfer purulent part of sputum to cotton-wool swab.
Container of Amies transport medium.
Must reach laboratory with in 6hrs.
Collection of Throat & Mouth Specimen
In good-light and using a tongue-depressor examine the inside of mouth
. Look for inflammation, presence of membrane/
exudate /pus.
Swab with a sterile cotton-wool swab.
Avoid contamination with saliva i.e;don’t touch any-other part of mouth.
Throat specimen Collection
Avoid antiseptic gargles eight hours prior to collection.
Deliver with in two hours of collection.
Alternate ProceduresTrans-laryngeal aspiration/ BAL required in case of Patient is debilitated & CANNOT
EXPECTORATE spontaneously.
Routine samples have failed to recover causative organism.
An-aerobic pulmonary infection is suspected.
Lung-abscesses/deep-seated pulmonary infections.
Nasal swabs
Use sterile cotton wool swab,gently swab the inside surface of the nose.
Take care not to contaminate the swab, replace in its sterile container
Label and within 2 hrs. deliver to the lab with request form.
Ear discharges
Collect the specimen of the discharge on cotton wool swab without contaminating it.
Transport within 2 hrs. to the lab. Transport it within 6 hrs if Amies
transport medium is used.
Eye specimens
Should be collected by a medical officer or an experienced nurse.
Cotton wool swab ,collect the discharge,& in an infant swab the lower conjunctival surface.
Deliver immediately to the lab.or use Amies Transport media,
also make a smear of the discharge on a slide.
Blood culturesCollection procedure Aseptic technique like lumber puncture Take out BC bottle from fridge & bring to room temp
wipe it’s capping with anti-septic. Identify the vein Operater hand clean . Sterile gloves Disinfect skin by standard method(Tinc iodine or
Povidone iodine (1 min), 70% alcohol (not spirit). Avoid recontaminating the veinipuncture site. Use new needle to transfer blood into BC bottle.
NUMBER & TIMING
Should be drawn before use of SYSTEMIC ANTI-MICROBIAL drugs.
When patient has recurring fever, collect blood an hour before an anticipated rise in temperature.
To increase chances of isolating the pathogen at least 2 specimens should be collected at different times.
VOLUME
Adults 5-10ml blood and in children 2-3 ml should be drawn.
Blood to media ratio is 1:10. Insert needle through the rubber liner
&dispense.(in Brain heart infusion broth or whatever medium is available).
Blood culture bottles
C.S.F COLLECTION & TRANSPORT From arachnoid space,using a wide-bore needle
between L4 & L5.(Aseptic technique)
CSF is allowed to drip into two separate dry,sterile containers.( for C/S,other invest.)
A Ventricular puncture is sometimes performed to collect CSF in infants.
Notify lab before performing,L.P so that specimen is received&Exam immediately.
TRANSPORT OF CSF- AT 37oC
DELIVER IMMEDITALY BY KEEPING NEAR BODY OR IN TRANSPORT MEDIUM LIKE TRANSGROW OR TRANSISOLATE ( TIM)
IMMEDIATE PROCESSING IN LAB OR KEEP IN INCUBATOR at 37oC
Anaerobic blood culture
Take same precautions as for aerobic culturing.
Dispense 5 ml blood in thioglycollate medium(50 ml broth) or brain heart infusion broth.
Urine Collection
Morning urine specimen to be sent. MAINTAIN COLD CHAIN.
OTHERWISE RANDOM SPECIMEN.
Sterile, dry, wide neck, leak-proof container.
Urine Collection
Instruct the patient to clean the area.
10-20 ml clean catch , MSU specimen.
In Catheterised patient, aspirate with needle b/w tip&bag.
Urine sample storage
Immediate delivery not possible refrigerate at 4-6oC
OR boric acid as preservative.which allows
bacteria to remain viable without multiplying.
Faeces collection&transport
Patient to be given clean, dry, disinfectant free bedpan/wide-necked container (non sterile).
Avoid contamination of faeces with urine. Separate worms, tape worms and
segments from faeces if any. Gross examination information to be
provided.
TRANSPORT MEDIA FOR FAECES
Cary-Blair media for Salmonella,Shigella,Vibrio and Yersenia(for 48 hrs).
Cholera 1 ml specimen in 10 ml of alkaline peptone water(8 hrs).
Worms, tapeworms segments in normal saline.
Cotainer for faeces &urine
Collection of pus,ulcer & skin specimens
Pus from abscess is best collected at the time when: Abscess is incised,drained or after it has ruptured naturally.
Avoid contamination from skin commensals.
Specimen should be collected before Anti-septic dressing is applied.
Collection of Specimen from ulcers
Aspirate from a drainage tube up to 5ml of pus.
If pus is not being discharged : use a cotton-wool swab to collect sample.Immerse swab in a container of Amies transport medium.
When the tissue is deeply ulcerated/necrotic:Aspiration is done from the side-wall of ulcer.
Collection of Serous fluid/exudates
Fluid from pustules,buboes & blisters is aspirated with a syringe.
Serous fluid from skin ulcers ,papillomas may contain treponemes.
A drop of exudate is collected directly on cover glass & inverted on a clean slide. Immediately the specimen is dispatched for dark-field microscopy.
Transport of pus/ulcer specimens
Specimen is collected using a sterile cotton-wool swab & inserted in a container of Amies transport medium.
Aspirated specimen is transferred to a sterile,
leak-proof container & sealed.
A smear is made on a clean slide for Gram-staining.
Urogenital specimens
Collected by a medical officer or an experienced nurse.
Amies medium, for transporting urethral,cervical,and vaginal swabs within 6 hrs.
If cotton swab used then immediately deliver to the lab & proceed without delay.
Take urethral specimens at least 1 hour after the patient has urinated. Collect pus directly on a swab. If no discharge is evident, the male urethra is stripped towards the orifice to evacuate exudate. If no exudate is obtained, in sert a thin swab [special urethral swabs if available] 2-3 cm into the urethra and gently scrape the mucosa by rotating the swab for 5-10 seconds. In women, massage the urethra against the pubic symphysis and use the same technique as for men.
Endocervical Swabs: With the patient in lithotomy position,
expose the cervix using a vaginal speculum. Moisten the speculum with warm water since many lubricants contain antibacterial substances which may inhibit growth of gonoccoci. Collect the following specimens
According to following instructions Remove any mucous plug from the external os using a cotton
ball. Insert a Transport medium swab into the endocervix. Rotate it gently and move it from side for 30 seconds before removal.
Transport this swab to laboratory at room temperature 20-22O C ,taking care that it should get cultured on MTM plates within 12 hours preferably within 6 hours if amies transport media is used.
Inoculate the swab directly on the labeled MTM plate only if the plate is provided at the collection site. Discard this swab.
Using sterile swab, collect a second specimen from the endocervix as above and make a smear on the slide labelled CX let it air dry. Discard the swab.
Fluids and effusions
Synovial, pleural, pericardial, ascitic & hydrocele fluids.
Collected by a medical officer aseptically. Transport immediately to the lab. collect into two portions ,one with
anticoagulant 3%sodium citrate 1ml and 9ml fluid for cell count,protein estimation,microscopy,and culture
Other 5 ml in screw capped container without citrate to see clotting.
Fluids and effusions
Alternate method: Dispense 5-10 ml fluid in blood culture
bottle( aerobic or anaerobic), transport not over 37 C or in direct light.
Transport to lab without delay.
12.FLUID / 12.FLUID / EFFUSIONSEFFUSIONS Physical ExaminationPhysical Examination
Centrifuge Centrifuge 10 ml10 ml
sedimentsediment forfor cultureculture Gram and Z-NGram and Z-N OROR
5-105-10mlml
When positive
Semen
Give pt. a clean,dry,leak proof container. Transport in a plastic bag and kept in the
pocket to maintain the temperature.
Collection for ANAEROBES
APPROPRIATE SPECIMEN PUS IN SYRINGE WITHOUT AIRCOLUMN OR
IN ROBERTSONS COOKED MEAT MEDIUM BROTH
NOTE SWABS ARE UNFIT
Collection Procedures
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