Providing Safe Drinking Water

Providing Safe Drinking water USGS Workshop Bangkok

Muhammad Naseem Khan Scientific Officer PCSIR

Road Map • Background • Methods • WQMP overview

Pathogens in Water and Public Health Microbiology

Important concepts • Viruses, bacteria, and protozoa are very different types of organisms. • Many fecal-oral pathogens that are important to public health are waterborne. • Non-pathogenic bacteria are used to test for the presence of pathogenic viruses, bacteria, and protozoa .

Waterborne disease • Caused by drinking or accidentally ingesting water contaminated with pathogens. • World-wide: 4 billion episodes of diarrhea result in about 1.5 million deaths each year, mostly children. Waterborne bacterial infections account for approximately half of these episodes and deaths.

Waterborne pathogens—a public health concern •

Potential sources of pathogens: – – – –

Human, animal wastes, sewage and sewage sludge, septic tanks, landfills

•

Pathogens ingested from: – drinking water, – recreational water, – contaminated fish or shellfish.

Types of waterborne pathogens

Viruses (tiny, nonliving) Bacteria (medium size, Simple cells)

Protozoa (larger, complex cells)

Drinking-water outbreaks in the US in 2005 and 2006—Cause of acute gastrointestinal illnesses Surface Water Mixed Unident. Viral Parasitic Bacterial

Etiologic Agent

Ground Water

Source

Source and cause of gastroenteritis associated with recreational water, 1997 – 2006

Factors affecting incidence of waterborne Illness in the US • Increasingly greater threat to public health – Increase in population – Aging water-treatment systems – Aging population – Mismanagement of animal wastes

Waterborne Diseases of Specific Concern • Afghanistan – bacterial and protozoal diarrhea, hepatitis A, and typhoid fever

• Pakistan – bacterial diarrhea, hepatitis A and E, and typhoid fever

• Thailand – bacterial diarrhea and hepatitis A

• Similar to U.S., but at higher rates, with more mortality in young

Factors affecting incidence of waterborne Illness in developing nations • Consistent threat to public health

– Poor sanitation – Inadequate or lacking infrastructure • Especially in rural or dispersed populations

– Mismanagement of animal wastes from agriculture • Similar to U.S.

Viruses • • • •

DNA or RNA with a protein coat No nucleus Cannot metabolize or reproduce without a host 0.02-0.3 micron

Photo Credit: Maria-Lucia Rácz

Rotavirus - 0.07 µm

Fecal Viral Pathogens— Examples of Illnesses Pathogen

Disease or symptom

Hepatitis A & E

Interferes with liver function, fever, jaundice, diarrhea

Adenoviruses

Respiratory & eye infections, diarrhea

Norovirus

acute-onset vomiting, watery non-bloody diarrhea with abdominal cramps

Echoviruses

Infants:Liver failure, myocarditis, menegitis

Poliovirus

Polio, ranging from gastroenteritis to irreversible paralysis

Reovirus

Gastroenteritis

Rotavirus

Most common cause of severe diarrhea in small children

Bacteria • Independently metabolizing and reproducing Bacteria organisms. E. coli - 4 µm long • No true nucleus • Generally 0.2 to 5 micron

Bacteria—Examples of Illnesses Pathogen

Disease or Symptom

Salmonella typhi and sp.

Typhoid fever

Shigella sp.

Shigellosis: dysentery

Escherichia coli O157:H7

can lead to kidney failure

Campylobacter jejuni and sp.

gastroenteritis

Legionella pneumoniae

fever, pneumonia

Helicobacter pylori

Gastritis, ulcers

Vibrio cholerae and sp.

cholera

Yersinia spp.

Yersiniosis: diarrhea

Aeromonas hydrophila

pneumonia

Protozoa • Unicellular members of the Animal kingdom • true nucleus • 2 to 200 microns

Giardia lamblia -- 12 to 15 µm

Protozoa—Examples of Illnesses Pathogen

Disease or Symptom

Cryptosporidium parvum Cryptosporidiosis, diarrhea Giardia lamblia

Diarrhea

Toxoplasma gondii

Blindness or brain damage in children infected in-utero

Microsporidia

Acute diarrhea (esp. those with HIV)

Cyclospora

Persistent diarrhea

Naegleria fowleri

Primary amebic meningoencephalitis (headache, fever, and vomiting)

Summary • Similar diseases world wide in developing and developed nations – Difference is in intensity and water use – Due to: • Infrastructure differences • Population age distribution differences • Health Care differences

• All nations can potentially benefit from having well designed and implemented microbial water quality monitoring, for drinking and recreational water

Colilert Method for Total Coliforms & E.coli

Colitag Method • Commercially available enzyme – substrate liquid broth medium. • Simultaneously detection of total coliforms & E.coli • Available in MPN or Presence/absence format.

BACTERIAL INDICATORS

E. coli cells (from USEPA web site)

Use Aseptic Technique • Aseptic means “sterile” • Minimizes sample contamination • Assume you are surrounded by contaminating organisms; disinfect your work area before Dr. Harold Sears, U of S. Carolina and after use • Use aseptic techniques during sample bacteria on a pin collection, sample

Defined substrate technology (DST) sterile DI water

mixing bottle sample pre-measured media

quantitray

How does DST work? • Nutrient medium for coliforms – ortho-nitrophenyl-β-Dgalactopyranoside (ONPG) and – 4-methylumbelliferyl-β-Dglucuronide (MUG)

• Total coliform enzyme metabolizes ONPG to form a yellow product. • E. coli enzyme metabolizes MUG to form a fluorescent product.

DST Steps

2. Seal, incubate

1. Mix media with sample; pour into Quantitray

3. Count the positive cells

Preparing a sample for incubation

Total coliforms and E. coli: Colilert Quantitray • Under ambient light, total coliform colonies are yellow • The same tray under UV light; total coliform colonies that are E. coli fluoresce

Determining a final count • Compare color intensity to the “comparator” tray • Count the number of large and small wells that are positive • Look up the most probable number on the IDEXX table

Blank

Comparator

Positive test

How does DST work? • Nutrient medium for enterococci – 4methylumbelliferylβ-D-glucoside

• Enterococci enzyme metabolizes the reagent to form a fluorescent product

Enterococci: Defined Substrate Technology • Enterococci produce Blue fluorescence under UV light • Marine samples must be diluted to reduce false positives

Coliphages

Viral Indicators: Coliphages • Coliphages are viruses that infect coliform bacteria • Recommended by USEPA for use as a GW fecal indicator. • Indicator for the transport and survival of viruses in the environment – Smaller than bacteria – No metabolic needs

MS2 phage capsids

Viral Indicators: Coliphages • Two main groups of coliphage: – Male-specific - Infect bacteria by attaching to hairlike projections “F pili” – Somatic coliphage - Infect bacteria by attaching to outer cell wall

• Found in high numbers in sewage; low incidence in human feces • Good indicator of sewage pollution

Coliphage analyses • Simple and inexpensive • Requires a trained analyst in the laboratory • Coliform host (E. coli) exposed to water sample – Presence-absence test (USEPA 1601): host added to sample. Test for coliphage after enrichment. – Single agar-layer test (USEPA 1602): sample mixed with liquid agar containing a host. Coliphage detected directly from sample.

• Plaques form where coliphage were present and killed surrounding cells

Flow Chart of Presence/absence(1601) Method

Coliphage presence-absence test Positive control

Test sample

USEPA Method 1601

Negative control

Flow Chart of Quantitative (1602) Method

Coliphage quantification by single-layer agar method

plaques

USEPA Method 1602

From Madigan and others, 2000

IMS/ATP RAPID METHOD

IMS/ATP METHOD • Method initially developed by researchers Lee & Deininger (2004) • IMS/ATP method has been developed for E.coli & Enterococci. • It is based on antigen-antibody reaction.

IMS/ATP Method Overview – Capture by immunomagnetic separation (IMS) • Uses antibody-coated magnetic beads which bind to antigens present on the surface of cells

– Detection by adenosine triphosphate (ATP) luminescence • Energy molecule in all cells • Detects viable organisms Results are reported in relative light units (RLUs)

M A G N E T

Method Flow Add beads

30 min @ 18 rpm Sample + diluent

Large magnet Wash with PBS

Add LL

Small magnet

Luminometer Add SRA Wash with PBS Add BRA

Advantages • IMS/ATP is a rapid method for the detection of E. coli and Enterococci – Detects viable cells so directly related to health risk. – Can analyze 1 sample in less than 1 hr; 6-8 samples in less than 2 hrs – Low startup and per sample cost – Equipment is portable, can be used in the field

Water Quality Monitoring Program

Goal • To provide Safe Drinking Water in Sind Province

Objectives • Identify the source of water supply • Identification of Sources of Contamination ( Industrial/Hospital/Environmental ) • Identifying the type of contamination ( Physical / Chemical / Microbial ) • Identifying Seasonal/periodic variation of contaminations • Identifying hydrological boundaries of contamination • Analysis of Monitoring Data • Planning and Designing for remedial measures • Physical works / Implementation of Remedial measure

Beneficial Organization • Concern implementing Agency • Stakeholder and Consumer • Academic institute

• Capacity / Capability / Resources of Concern organization • Need to be Analyses by sharing information

• Who will Lead • SIDA

Responsibilities / Role of concern organization •

Sampling ----- SIDA, PCRWR, PCSIR, KU • Testing ------ PCRWR, KU, PCSIR • Analysis of DATA----- KU and PCRWR

• Coordination during implementation by Lead Agency • Cooperation with lead Agency • Analysis and Sharing and Distribution of collected data • Will be assign base on Capacity / capability / resources of concern organization • PCRWR has data warehouse --- Role data management • Advisory / Steering Committee for WQMP

Geographical Boudries Sindh—All barrages Kotri Barrage, Guddo Barrage, Sakhar Barrage

Time frame Duration ---- 5 years Frequency --- 10 to 20 samples/ year / Site Surface Water ( More or Equal to above frequency ) Ground water ( Less then above frequency ) Volume -----Bacteriological – 500ml Virology -- 10000 ml (10 liter ) Chemical ---- 1000 ml

Mediums Surface water River Canals Sub canals Drains Ground water Wells Treated water Houses Hospital Educational institutes Hydrants Filter water Pipe water Store Water

PROGRAM DESIGN • Hydrological information majorly provided by SEDA • Previous Studies would collected from PCRWR and other organization if have any information • Steering committee design WQMP • All parameter would be tested • Major parameter on the basis of site requirements

Data interpretation • Storage--- Data warehouse in PCRWR • Its must full fill the Objective • Comparison – PSQCA • Group of experts – Analysis and trend • Also determine whether objectives achieved or not