For dialysis technician, Ug and Pg Medical students, Nursing students.

1. Bacteriology of dialysis fluid
Arun Kumar P
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2. Dialysis is a treatment that filters, purifies, removing
waste and excess water from the blood using a machine.
1) Hemodialysis
2) Peritoneal dialysis
2Bactriology of dialysis fluid

3. Diffusion of solutes and ultrafiltration of fluid across a semi-
permeable membrane.
substances in water tend to move from an area of high
concentration to an area of low concentration.
Blood flows by one side of a semi-permeable membrane, and a
dialysate, or special dialysis fluid, flows by the opposite side.
A semipermeable membrane is a thin layer of material that contains
holes of various sizes, or pores. Smaller solutes and fluid pass
through the membrane, but the membrane blocks the passage of
larger substances (for example, red blood cells, large proteins).
Principle
3Bactriology of dialysis fluid

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6. Dialysate or dialysis fluid (DF) is one of the basic
elements of haemodialysis (HD).
Consists of purified water, glucose and electrolytes.
The concentration of electrolytes (besides potassium
and the buffer substance) closely resembles that which
occurs naturally in the blood.
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8. Dialysate quality and purity are two of the most critical
requirements of haemodialysis.
The presence of contaminants in the dialysate - the risk of
toxic substances accumulating and consequent acute and
chronic complications .
The possible contaminants in water used to prepare dialysis
fluid,
(i) particles,
(ii) chemicals and
(iii) microbial contamination.
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9. Purified water for haemodialysis
There are 3 main elements of these procedures:
Agar Medium,
Incubation Temperatures and
Incubation Time.
The first culture method incorporates the ISO Standard. Here the
two acceptable Agar Mediums
1.Tryptone Glucose Extract Agar (TGEA)
2. Reasoner’s 2A supplemented with 4% sodium
bicarbonate, or equivalent.
Blood or chocolate agar - unacceptable and are not be used.
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2. actual water environment are normally between 15-25°C, not
at the current 35°C. so, temperature to be maintained between
17°C to 23°C.
3. 168 hours (7 days) provides an accurate indication of viable
bacteria, which is a change from the 48 hours at 35°C
Endotoxin:
Maximum allowable levels of endotoxins: The endotoxin con-
tent in purified water for haemodialysis shall not exceed 0.25
EU/ml, measured by a sufficiently sensitive LAL test.

11. ENDOTOXIN
1. Lipopolysaccharides:
Limulus amoebocyte lysate (LAL) test-detecting
endotoxin
Based on clotting reaction of horse shoe lysate by
endotoxin
Types of LAL test
1. Gel-clot,
2. Turbidimetric,
3. Colorimetric
4. Fluorescent.
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Peptidoglycan
quantified with the silkworm larvae plasma (SLP) test
not routinely used and not recommended in the standards.
SLP contains all the factors of the prophenoloxidase cascade, an
insect self-defence mechanism.
The cascade is initiated by PG, in which prophenol oxidase is
ultimately activated to phenol oxidase.

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Phenol oxidase activity is colorimetrically detected, with 3,4-
dihydroxyphenylalanine as a substrate and melanine as an end-
product.
The test does not detect ETs, but cross-reacts with β-glucans,
which produce a positive result in both the SLP and the LAL test.
positive in the SLP test but negative in the LAL test.
Cont….

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Bacterial DNA
bDNA can be quantified using different methods, the concentration
of DNA extracted from dialysis fluid can be determined
1. spectrophotometrically by measuring absorbance after exposure to
ultraviolet light at 260 nm.
2. the detection of free bDNA by 16S rRNA gene polymerase chain
reaction amplification after which the amplicons could also be used
for identification purposes.

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3. Quantification of oligonucleotides (and ssDNA) can also be
performed using OliGreen, an ultrasensitive fluorescent nucleic acid
stain.
4. In a microplate setting, as little as 1 ng oligonucleotide or ssDNA
per millilitre can be detected (Quant-iT®; Invitrogen). -this
parameter should be considered for routine use in future.

17. Lower contamination levels if visible in more than one
sample in order to prevent the formation of bacterial
biofilm.
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See below

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General disinfection procedure
In-centre HD
the water distribution pipelines are heat disinfected (∼90°C) three times
per week;
process is driven by a hot feed installed downstream the RO membrane.
The HD machines are also included in the circuit covered by this heat
disinfection procedure.
Chemical disinfection of each HD machine is performed after each
session, while after 100 treatments, the ultrafilters, used for ET and
bacteria retention, are replaced followed by a chemical disinfection.

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storage tanks -the risk of biofilm formation, it might be
unavoidable to use them in some units.
subjected to UV irradiation, which is, however, only effective to
kill planktonic micro-organisms.
In addition, they should be frequently drained and adequately
disinfected.

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Home HD
In the home setting, the RO is chemically disinfected twice a month
and a combination of chemical and heat disinfection
HD machines is performed after each session.
For the time being, we cannot apply integrated

24. Prevention and rate of contamination
• Transmission can occur directly or indirectly via contaminated
devices, equipment and supplies, environmental surfaces, or
hands of personnel.
• viral infections such as hepatitis B and C, HIV and bacterial
infections, especially those involving vascular access.
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25. The prevalence of antimicrobial resistant bacteria has increased
rapidly in health c are settings, including hemodialysis units in recent
years.
Multiresistant organisms (MRO) are defined as bacteria that are
resistant to one or more classes of antimicrobial agents. These include
• Methicillin Resistant Staphylococcus aureus (MRSA),
• Vancomycin Resistant Enterococci (VRE),
• Extended Spectrum βlactamase (ESBL) producing Klebsiella
pneumonia,
• Carbapenem resistant Acinetobacter baumannii
• (CRAB)Clostridium difficile (antibiotic associated diarrhoea).
• Antimicrobial use and direct contact transmission of resistant
strains are the two main factors that have contributed to this
significant increase
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26. Hand Hygiene
Staff should cover any cuts and abrasions with waterproof
dressings.
Staff who has extensive untreated cuts or chronic skin disease,
such as eczema, should not work in dialysis units when their skin
lesions are active.
Unwashed hands of healthcare workers are the major route of
transmission of microorganisms in healthcare settings.
Hand hygiene is includes hand washing with soap and water,
and/or applying an alcohol based hand rub (e.g. sterilium)
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27. Hands should be washed with soap and water when visibly dirty or
contaminated with proteinaceous material (e.g. blood or other body
fluids).
If hands are not visibly soiled, an alcohol based hand rub can be used.
Hand hygiene should be performed before and after patient contact
after contact with a source of microorganisms (body fluids
andsubstances, mucous membranes, non-intact skin, or
inanimate objects that are likely to be contaminated) after removing
gloves
Hand hygiene facilities should be located as close as possible to the point
of contact with patients and dialysis equipment.
Soap solution must be provided in dispensers with disposable cartridges
or singleuse bottles, to prevent bacterial contamination of the product.
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28. Alcohol based hand rubs should be placed at the point of contact, for
example:
Next to or attached to the frame of dialysis bed or chair
 At points of entry and exit of dialysis room
 At staff stations or chart and medication trolleys.
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Use of gloves
Clean, nonsterile gloves should be worn when contact with blood or
body fluids is anticipated; this includes contact with patients and
dialysis equipment.
Gloves must be changed and hands cleaned between patients and/or
stations.
Gloves must also be changed and hands cleaned between
different activities on the same patient (e.g. moving from a
contaminated to a clean body site).
Gloves should be worn for any cleaning activities.
Hands should be decontaminated or washed after removing gloves.
Gloves should not be washed or reused.

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Personal protection
Face protection (eyewear/goggles, masks) is required to protect the
mucous membranes of the eyes, nose and mouth when performing
procedures that may generate splashes or sprays of blood or body
fluids (e.g. during initiation and termination of dialysis).
Personal eyeglasses and contact lenses are not considered adequate
eye protection.
Plastic aprons are indicated to prevent contamination of clothing
with blood, body fluids, and other potentially infectious material.

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A long sleeved, fluid barrier (impervious) gown should be worn
if exposed areas of the body e.g. arms, body front, are likely to
be contaminated by blood or body fluids.
All personal protection equipment (with the exception of
eyewear/goggles unless soiled) must be changed and hands
cleaned between attending different patients.
if it becomes splashed with blood or body fluids on leaving the
work area

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Environmental Issues including Equipment and
Consumables
Storage of equipment close to dialysis machines and patients should be
minimized.
Where possible, regularly used equipment such as adhesive tapes,
tourniquets, blood pressure cuffs and clamps should be designated to
each patient.
Consumables taken to the patient’s station should be used only for that
patient and should not be returned to a common clean area or used on
other patients.

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Cleaning of dialysis machines and chairs/beds
Dialysis machines should be internally disinfected, externally cleaned
(and disinfected if indicated), and dried after each patient.
The exterior of the machine should be effectively cleaned using protocols
following manufacturer’s instructions.
Special attention should be given to cleaning control panels on the
dialysis machines and other surfaces that are frequently touched and
potentially contaminated with patients’ blood.
Cleaning of non critical surfaces (e.g. dialysis bed or chair, countertops,
external surfaces of dialysis machines and equipment)
should be done with neutral detergent and warm water.

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The following procedure should be adopted for any surface/item
that is visibly contaminated with blood
Clean with neutral detergent and water, and then
Disinfect with sodium hypochlorite 1% (1,000 ppm available
chlorine; 1:10 dilution).
Remove chlorine residues from metallic surfaces with water as
sodium hypochlorite in high concentrations (>500 ppm) is corrosive
to metals.

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Disinfection of Haemodialysis Machines
Manufacturers producing dialysis machines each recommend a different
procedure for decontamination, bacterial biofilm and endotoxin removal.
The development of bacterial biofilms in the hydraulic circuit of haemodialysis
machines can be prevented by frequent use of chemical and heat disinfection
strategies.
Disinfection should include the following
Heat disinfection (80°C to 90°C) after each dialysis
Citric acid and heat disinfection at the end of the day
Bleaching (5% chlorine) once a month.
Frequent bleaching is not recommended because of possible damage to the
machine.

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Dialysates
Liquid bicarbonate dialysate concentrate can support rapid bacterial
proliferation, and hence it not be used more than 24 hours after
opening.
Bottles containing unused dialysate should be immediately capped
and the exterior of the bottle wiped over with detergent and water
as part of the overall procedure of cleaning the haemodialysis
machine.
The date and time of opening should be recorded on the bottle
using an indelible pen.
Opened bottles containin g unused fluid should be discarded after
24 hours.
Unfinished bottles used for infected patients must be discarded
immediately after the dialysis session.