BACTERIA AND BIOCIDES

CLASSIFICATION BASED ON RESPONSE TO OXYGEN

AEROBIC ANAEROBIC FACULTATIVE

MICROAEROPHILIC
EH

REQUIREMENTS

CLASSIFICATION ACCORDING TO ENVIRONMENTAL PREFERENCE

BARAPHILES THERMOPILES MESOPHILES PSYCHROPHILES HALOPHILES ACIDOPHILES NEUTROPHILES ETC. I.E., BAROPHILLIC THERMOPHILES
[SESSILE VS PLANKTONIC]

TYPES OF BACTERIA THAT ARE IMPORTANT IN THE OIL FIELD

Sulfate reducing bacteria (SRBs)

desulfovibrio desulfomaculum

Anaerobic Bacteria Aerobic Bacteria Facultative Bacteria Iron Bacteria Others

sulfur oxidizing nitrate reducing methanogens fungi

[SESSILE VS PLANKTONIC]

SIGNS OF BACTERIAL CONTAMINATION

BLACK [H2S] [FeS]/SOLIDS

WATER

PLUGGING

/ FILTER PROBLEMS / HIGH INJECTIVITY PRESSURES PITTING CORROSION - PATTERNS HIGH BACTERIA COUNTS

BREEDING SITES
SLIME FORMERS These bacteria need oxygen to grow. Although oil field waters are naturally very low in oxygen in many instances air can get into field water through system leaks.
OPEN

PONDS SHALLOW WATER SUPPLY WELLS ON PRODUCTION EQUIPMENT ON TUBING

BREEDING SITES
SRBS
Can be found in many areas. Will only grow in an environment that has low concentrations of oxygen.
RAT HOLES PACKER FLUIDS HEATER TREATERS FILTER BEDS AT OIL/WATER INTERFACES UNDER DEPOSITS

Intake Pumps Strainer Filtration (1)

WATER INJECTION SYSTEM

( ) Sampling Point
(2) Process Cooling (6) Dump (4) (5) Water Store (7) Mud Store (8) Mud Make-up

Deaeration
Chemical Treatment Injection Pump Reservoir

(3)

Desalination

Sampling Notes:
1. General organisms for water quality; blooms ; efficacy of chlorination; consolidation techniques for sulfate-reducing bacteria. 2. General organisms; clogging filters; backwash schedules; souring of deep-beds. 3. Sulfate-reducing bacteria. 4. Sulfate-reducing bacteria probably by consolidation techniques; check biocide efficiency. 5. Backflushing of reservoir; check sulfate-reducing bacteria and sulfur. 6. Heat exchangers, etc; general organisms slime formation and loss of heat transfer. 7. Quality of drinking water. 8. Blooms; sulfate-reducing bacteria; odor nuisance of mud in store.

Reservoir

PRODUCTION SYSTEM
(1)

Wellhead High Pressure Separator

( ) Sampling Point

(2A) Oily Water (2B)

Low Pressure Separator

(6)

Pipeline Storage

(3)

Storage
(5) Tanker

(4)

Sampling Notes:

1. Sulfate-reducing bacteria levels after shut-ins; sulfatereducing bacteria on downloads SV; sulfate-reducing bacteria in sea water breakthrough. 2. Sulfate-reducing bacteria in water bottoms. 3. Sulfate-reducing bacteria in pipeline water at landfalls; pigging debris; biocide efficacy tests. 4. Sulfate-reducing bacteria in tank bottoms. 5. Sulfate-reducing bacteria in offshore sea water ballasted crude storage tanks; general oil degrading organisms. 6. General oil degrading organisms; sulfate-reducing bacteria. Often main source of contamination of pipelines.

ENUMERATION / IDENTIFICATION OF BACTERIA

Serial dilution followed by culturing in the appropriate media Epifluorescence A.T.P Photometry Microscopic techniques Fast count methods
*sanicheck *rapid check

A true identification of bacteria is very complicated. This is generally not carried out in an oil field operation.

CULTURING OF BACTERIA
In order to successfully culture bacteria an environment needs to be provided that allows them to grow and multiply. Requirements:
Specificity [Salt] pH Eh-(poising agents) Temperature of incubation Visualization Time to develop growth Availability of media

MEDIA REQUIREMENTS
Food source SRB lactic acid acetic acid others Na, Ca, Mg, Fe, etc +++ ~7 + yeast Adjust to suit organism ANAEROBES various sugars AEROBES protein extracts

Cations Sulfate pH Eh (poising Agent) General Nutrients Salts

Na, Ca, Mg, Fe, etc + ~7 + yeast

Na, Ca, Mg, Fe, etc + ~7 -

CULTURE MEDIA
SRB Broth (Modified Postgate B)
KH2PO4 NH4CL CaSO4 . MgSO4 7H2O yeast extract Na lactate Na acetate Asorbic acid Thioglycollic acid . FeSO4 7H2O pH CaCl MgCl2 NaCl H2O 0.5g 1.0g 1.0g 2.0g 1.0g 3.5g 2.8g 0.1g 0.1g 0.5g Adjust Adjust Add up to 1000ml

CULTURE MEDIA
General Anaerobic Broth
glucose KNO3 Na2HPO4 . MgSO4 7H2O . FeSO4 7H2O yeast extract NH4Cl Cysteine-HCl pH NaCl H2O 10.0g 0.6g 0.45g 0.1g 0.001g 3.0g 0.5g 0.5g 7.0 adjust Add up to 1000ml

CULTURE MEDIA
API Aerobic Broth

Beef Extract Peptone pH NaCl H2 O

3.0g 5.0g 7.0 adjust Add up to 1000 ml

DOS AND DONTS ON BACTERIA CULTURING

CRITICALLY IMPORTANT Adjust salinity to match field salt level. When in between go to lower chloride. Incubate - temperature, time Darkness - Do not expose to UV Shoot a number of bottles (6) to define severity of problem Safety - needle destruction, bottle disposal, letter of justification Observe shelf life Thermo-heat the media to avoid killing thermophiles

DOS AND DONTS ON BACTERIA CULTURING

MAY

BE IMPORTANT

Changing syringes Alcohol swabbing Volume of sample-precision 1ml +/- 0.2 ml Gauge of needle - avoid large diameter Media manufacturer - stay with one Postgate v. API for SRB

DOS AND DONTS ON BACTERIA CULTURING

DOES

NOT MATTER

Drawing vacuum on bottle before withdrawing needle Size of syringe

SERIAL DILUTION
Bottles Showing Growth 1 2 3 4 5 Bact. Count (colonies/ml) 1-10 10-100 100-1000 1000-10,000 10,000-100,000

If the above procedures are carried out in triplicate (or higher) there are statistical tables available for more accurate counts

SERIAL DILUTION

SAMPLE

0-10 10-100

etc.

etc.

etc.

PLUGGING PROBLEMS
Bacteria can cause plugging and reduced flow in filters, down hole formations and lines. They can coat the walls of heat exchangers and reduce their efficiency.
Formation of insoluble salts (FeS) Formation of slime and cell debris

HOW DO BACTERIA CAUSE CORROSION

-H2S production SO4-2 + H2 H2S sulfate hydrogen sulfide Acid production
(other substrates)

sugars

(lactic, acetic, propionic)

various acids

CO2 + H2

CATHODIC DEPOLARIZATION
2 ELECTRONS +2H+ 2H H2
From the anodic metal oxidation In the cell cytochrome system

SO4-2
oxidation of hydrogen reduction of sulfate

H2O + S-2 + Energy

ADP ATP

Assimilation Reactions

BACTERIAL CONTROL WITH BIOCIDES

Lab procedures Compatibility tests (biocide c field fluids)

c other treatment chemicals

Time kill tests. This procedure evaluates the amount of chemical needed and amount of contact time required for a biocide to kill the bacteria or significantly reduce their population density. This test measures the bactericidal potency of a biocide. Bacteriostatic test. This procedure evaluates the ability of a reagent to stop the growth of (but not necessary kill) bacteria. Biocide studies on bacterial films. These techniques are used to measure the effect of biocides on sessile bacteria that have been grown on a surface.

BACTERIAL CONTROL WITH BIOCIDES

Field monitors and surveys (establish base lines)
Planktonic appropriate sample points (bug vials) Sessile Robbins sidestream device

pinpoint specific areas Robbins in-line device of high conc. coupons pipe sections etc.

[H2S] Corrosion rates Measurements of injectivity pressure filter plugging, total solids etc.

BACTERIAL CONTROL WITH BIOCIDES

TREATMENT PROCEDURES:

Continuous vs slug Specific sites vs general Correlation with lab data Treatment in conjuction with a surfactant Treatment in conjuction with mechanical devices
Butterworth Sprayers Pigs Other clean up procedure

AN EXAMPLE OF A TREATMENT REGIMEN

OXYGEN CORROSION

SCAVENGERS

INHIBITION

PIG

BIOCIDE

PIG

PIPELINE

BIOCIDES MECHANISMS OF ACTION

Interfere with fundamental cell requirements

denaturation of proteins and enzymes disruption of lipids cause cell wall disruption damage / inactivation of DNA, RNA

Most biocides have pluralistic modes of action Significance of resistance

Glycocalyx formation Specificity of action is rare

Synergisms

BIOCIDE REGISTRATION / LABELING

EPA LABEL

REQUIREMENTS

BIOCIDES
DESCRIPTION

AND AVAILABILITY

BIOCIDES (MISCELLANEOUS)
Other biocide Imidazolines Sulfones Sodium Hypochlorite Benzothiazole Tin oxide Thiocyanate Guanidine
(VANTOCIL)

Naperville Naperville Naperville Naperville Naperville Naperville Europe Naperville Not Available

Copper Sulfate Metronidazole

BIOSURFACANTS (biodispersants)
EC1082A EC9047A EC1088A Diamine plus surfactants Surfactant only Diamine plus surfactants

SAMPLING OF BACTERIA / REVIEW

Sessile bacteria - need to obtain the bacteria from the surface. Planktonic bacteria - need to obtain the bacteria out of the liquid. The procedures for obtaining sessile counts are somewhat more cumbersome than for planktonic but utilize the same fundamental principles. Important Factors:

Eh requirements oxygen concentration sampling techniques

BIOCIDE INCOMPATIBILITIES
Reactions

with other chemicals. Temperature sensitivity. Sensitivity to pH. Solubility in brines. Foaming. Emulsion Formation

BACTERIAL METABOLISM
TYPE Respiration EXAMPLE
C6H12O6 + 6O2 6CO2 + energy

CH3COOH + SO4-2 2CO2 + H2S Anaerobic respiration +2OH- + energy

Fermentation

C6H12O6 2CO2 + 2C2H5OH + energy

CONCLUSION
The determination of what problems exist in a field can be difficult and frustrating. Many times more than one type of corrosion is taking place simultaneously. In these instances it may be very hard to determine which one (or more) is the significant factor that needs to be controlled.

RELATIONSHIP OF TEST DATA TO FIELD SYSTEMS

BATCH

TREATMENT CONTINUOUS TREATMENT

CHEMICALS USED FOR BACTERIAL CONTROL

BACTERICIDES - CHEMICALS WHICH KILL BACTERIA BACTERIOSTATS - CHEMICALS WHICH RETARD OR INHIBIT THE GROWTH OF BACTERIA MICROBIOCIDES - CHEMICALS WHICH KILL OTHER FORMS OF LIFE IN ADDITION TO BACTERIA MICROBIOSTATS - CHEMICALS WHICH RETARD OR INHIBIT THE GROWTH OF OTHER FORMS OF LIFE IN ADDITION TO BACTERIA

SAMPLING / ENUMERATION OF BACTERIA

PROCEDURES

WITH PLANKTONIC

BACTERIA PROCEDURE WITH SESSILE BACTERIA

TAXONOMY
Kingdom - Plant Phylum - Thallophyta Class - Schizomycetes Order - Pseudomonadales Family - Spirillaceae Genus - Desulfovibrio Species - ie. vulgaris desulfuricans africans

MISC. CLASSIFICATIONS
Spore

formers Staining techniques

gram pos gram neg gram variable

MORPHOLOGICAL CLASSIFICATIONS

Rods Curved (sigmoid) (like the letter C) Vibrio (like a comma or s shaped) Spiral Semilunar (cresent shaped) Coccoid (round) Filamentous Flageller classifications

peritrichous (uniform over body) monotrichous (one flagella/cell) lophotrichous (dist-about one end)

Remember
Bacteria can exhibit polymorphism or pleomorphism-especially under conditions of stress.

CLASSIFICATION OF BACTERIA
There are several ways in which bacteria are grouped. Many of these depend on what particular aspect of the organism is of interest or is being studied.

CLASSIFICATION OF MICROORGANISMS
Algae - Contain Chlorophyll, Require Sunlight Fungi - Do Not Contain Chlorophyll *Bacteria - Some Properties Common To Both Algae and Fungi

INTRODUCTION
PRACTICAL THEORETICAL INTEREST

HANDOUTS

BIOCIDES NAME Glutaraldehyde Glut mixtures Glut mixtures Formaldehyde Formaldehyde mix Diamines Diamine mixtures Quats Quat mixtures Isothiazoline Thiocarbamates THPS THPS mixtures DBNPA Triazine Dazomet DIKIOR (1) (2) (3) (4) DESCRIPTION Dialdehyde with quats with other chemicals Houston EC6111A EC6112A EC6109A EC6110A no no(2) no AVAILABILITY Europe Naperville EC9131A yes EC9128A (1) yes (1) yes

with other actives R-N-C-C-C-Nwith other chemicals R+ R-N-R R with other chemicals Heterocyclic (KATHON) S -N-C-SPhosphonium quat with other acivites Brominated amide Heterocyclic Heterocyclic Chlorine dioxide

7-6444 no EC9135A no

EC6106A 7-7672 yes EC6107A EC6210A no (1) yes EC6116A EC9123A yes no (1) yes yes yes

EC6114A no ASP744 Dryocide (3) no no

EC9126A (3) 7-6450 no yes yes yes no

EC6116A no EC6113A no EC6115A no (4) no

More than one available Not registered for oilfield use in the USA. Will be added to the product line as soon as it clears the EPA. Will be added to the product line.

SCHEMATIC OF FLUSH MOUNTED BIOPROBE

THREADED BACK SIDE STUD (BIOBULLET)

~2

ALLEN SCREW (TO HOLD STUD IN PLACE)

PLASTIC FRONT (GLUED ONTO METAL PLATE)

The flush mounted bioprobe is threaded in the back and screws directly into the line. The plastic on the front isolates the removable studs from the rest of the metal probe and thus eliminates galvanic corrosion. The field water flows over the studs in the line. Bacteria, if present can settle on and stick to the studs. The studs can be removed and the number of bacteria present on the surface is determined.

BUTYL RUBBER STOPPER

GLASS BOTTLE, 125cm3 CAPACITY

STERILE STEEL WIRE USED TO SUSPEND STUD TEST BOTTLE SOLUTION: ANAROBIC, BOTTLE FILLED TO NECK RUBBER O RING MILD STEEL STUD FROM BIOFILM GENERATOR PLASTIC COATED STIRRING BAR MAGNETIC STIRRER