Inactivation of microorganisms in treated municipal wastewater and biosolids by gamma irradiation

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INACTIVATION OF MICROORGANISMS

IN TREATED MUNICIPAL WASTEWATER

AND BIOSOLIDS

BY GAMMA IRRADIATION

THE FIFTH EURASIAN CONFERENCE OF NUCLEAR SCIENCE AND ITS APPLICATION OCTOBER 14-17, 2008, TAEA, ANKARA

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Zişan EMRE

Gülay ALTAY

B. Metin ALABAY

Seyit CANPOLAT

Yusuf VATANSEVER Hülya MERT

Turkish Atomic Energy Authority

Sarayköy Nuclear Research and Education Center Department of Animal Sciences

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The use of domestic wastewater for crop irrigation and

biosolids for land application are increasingly common.

However, wastewater treatment processes do not remove or

inactivate all pathogenic microorganisms.

The protozoan parasites, Cryptosporidium and Giardia are

important as they cause waterborne diseases

Bacterial pathogens in domestic wastewater include many

species of fecal coliforms, Salmonella sp. and fecal

(4)

Within all types of treatment, disinfection plays an

important role because it guarantees the elimination

of indicator pathogens.

•Ozone treatment is expensive, and not always

promising

•Ultraviolet (UV) radiation is promising

•Another type of energy, ionizing radiation, is

potentially useful for wastewater disinfection,

including high-energy electrons and gamma

radiation.

(5)

The work reported herein represents,

(i) an evaluation of the variations in the

population densities of indicator

pathogens,

(ii) the impacts of gamma irradiation on

these indicator microorganisms.

(6)

Treated municipal wastewater and biosolid samples were

collected on monthly basis at the Ankara Central Municipal

Wastewater Treatment Plant from May 2005 to April 2006.

The plant is located about 40 km from the city center and

about 90% of the city population is connected to the network.

The treatment capacity of the plant is 765000-1530000

(7)

The sewage sludge is generated from the wastewater during

primary and secondary treatment and it undergoes mesophilic

anaerobic digestion (35±1°C for about 20 days).

The digested sludge is dewatered to produce a semi-solid cake,

and named as biosolid (~650m

3

/day)

(8)

The collected samples were analysed within the same day. Parasitologic analyses

* Treated wastewater and biosolid samples were homogenized, concentrated, centrifuged and examined.

* Parasites were counted directly or by using hemacytometer following different staining methods according to the US-EPA Standarts

* Samples were examined for Cryptosporidium sp., Giardia lamblia, Entamoeba histolytica, Cyclospora cayetanensis and helminth ova.

Bacterial analyses

* Treated wastewater samples were examined for total coliforms, fecal streptococci, Enterococ sp, Salmonella sp.

* Biosolid samples were examined for fecal coliforms and Salmonella sp. * Bacteria were enumerated by using MPN Methods (APHA,1998)

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Cryptosporidium sp., oocyst

(upper left)

Giardia lamblia, cyst

(lower left)

Entamoeba histolytica, cyst

(right)

Lugol’s iodine (x 40)

Giardia lamblia, cyst (pink) Cyclospora cayetanensis, oocysts, (blue)

Kinyoun acid fast

(x 100)))))(((((

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Cryptosporidium sp. oocyst Kinyoun acid fast (x 100)

Cryptosporidium sp. oocyst

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Trichostronglidae sp. egg

Carbol fuchsin (x20)

Giardia lamblia cysts

(12)

Total coliforms

(MacConkey Agar)

Salmonella sp. colonies

(13)

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Months Cryptosporidium sp. oocyst / L Giardia lamblia cyst / L Entemoeba histolytica cyst / L Cyclospora cayetanensis oocyst / L Trichostrongylidae

egg / L Taenia sp.egg / L Hymenolepis nana egg / L May 2005 5.00x104 _- _- _- _- _- -June 05 2.25x104 _- _- _- _- _- -July 05 5.25x104 _3.5x101 _- _- _- _- -August 05 3.50x104 _1.0x101 _- _- _{10 x10}0_(dead) _- -Septem. 05 6.87x104 _1.5x102 _- _2.5x101 _- _- -October 05 1.13x104 _7.5x101 _- _- _- _- -Novem. 05 1.50x104 _2.4x101 _- _1.2x101 _- _- -Decem. 05 4.50x104 _4.0x101 _- _4.0x101 _- _-

-January 06 NA* NA* NA* NA* NA* NA* NA*

February 06 1.30x105 _1.1x102 _- _- _- _-

-March 06 1.80x105 _1.6x102 _2.5x101 _- _- _-

-April 06 2.20x105 _7.6x102 _- _- _- _-

-Table 1. Results of Parasitologic Analyses in Treated Wastewater Samples

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Months Cryptosporidium sp. oocyst / g Giardia lamblia cyst / g Entemoeba histolytica cyst / g Cyclospora cayetanensis oocyst/ g Trichostrongylidae sp

egg / g Taenia sp.egg/ g Hymenolepis nana egg/ g May 2005 _1.13x102 _- _- _- _- _- -June 05 _1.50x102 _- _- _- _{3.0 x10}0 _- -July 05 _6.25x102 _- _- _- _{0.5 x10}0 _- _{0.25 x10}0 August 05 _3.00x102 _- _- _- _{0.8 x10}0 _- -Septem. 05 _1.87x102 _- _- _- _{3.0 x10}0_(dead) _- _{0.3 x10}0 October 05 _5.00x102 _1.5x100 _- _- _- _- -Novem. 05 _2.25x102 _- _- _- _- _- -Decem. 05 _4.50x102 _- _- _- _- _-

-January 06 NA* NA* NA* NA* NA* NA* NA*

Februar 06 _6.25x102 _- _- _- _- _{2 x10}0

-March 06 _1.12x103 _1.5x100 _- _- _- _-

-April 06 _1.87x103 _- _- _- _{0.5 x10}0_(dead) _-

-Table 2. Results of Parasitologic Analyses in Biosolid Samples

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Table 3. Results of Bacteriologic Analyses in Treated Wastewater Samples Months Total Coliforms MPN / 100 ml Salmonella sp. MPN / 100 ml Enterococ sp. MPN / 100 ml Fecal Streptococ MPN / 100 ml May 2005 3.61x102 _(-) _<1.60x104 _3.30x102 June 2005 3.59x102 _1.3x102 _<1.60x104 _1.63x102 July 2005 3.47x102 _2.0x101 _1.70x103 _2.13x103 August 2005 1.66x102 _4.0x101 _5.50x102 _1.24x103 September 05 1.88x102 _(-) _1.25x102 _<1.60x104 October 2005 2.93x102 _4.0x101 _3.19x102 _1.70x103 November 05 2.94x102 _(-) _4.84x102 _2.56x102 December 05 1.43x102 _1.3x102 _<1.60x104 _2.20x103

January 2006 NA* NA* NA* NA*

February 2006 4.09x102 _8.0x101 _≥1.60x104 _≥1.60x104

March 2006 ≥1.6x104 _(-) _≥1.60x104 _≥1.60x104

April 2006 2.34x102 _4.0x101 _≥1.60x104 _5.98x101

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Table 4. Results of Bacteriologic Analyses in Biosolid Samples

Months Fecal coliforms

MPN / g Salmonella sp.MPN / 4 g May 2005 1.1x104 1.22x100 June 2005 2.0x105 _0.32x100 July 2005 3.4x105 _(-) August 2005 3.1x105 _0.67x100 September 2005 2.2x104 2.26x100 October 2005 1.3x105 _(-) November 2005 3.4x105 _0.68x100 December 2005 9.5x103 _0.68x100

January 2006 NA* NA*

February 2006 1.4x106 _1.51x100

March 2006 6.4x105 _{1.60 x10}0

April 2006 5.6x104 _(-)

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* The irradiation were conducted at 18°C using a 60_{Co Gamma Cell}

(Tenex-Issledovatel) at Sarayköy Nuclear Research and Education Center. The dose rate was 1.34 kGy/h.

* To investigate the effect of gamma irradiation on the viability of

Cryptosporidium sp. oocysts, the water samples containing oocysts were subjected to γ-irradiation in the range of 500-2000 Gy. Oocyst viability was determined by animal infectivity study, and 30-35 days old, 42 Balb/c mice were used for the infectivity assays. Mice were divided into six dose/infection groups including control group. The Cryptosporidium sp. oocysts were suspended in water at a concentration of 12x103_{oocysts/150μl/mice. Mice were infected with oocysts}

within 6 hours after irradiation. After 10 days following the infection, the fecal samples were collected and examined microscopically. Oocysts were counted by using hemacytometer and classified as:

( - ) Negative

( + ) <103_{Oocysts /g}

( ++ ) 103_-104_{Oocysts /g}

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Table 5.

Infectivity test results indicating the effect of gamma irradiation on the viability of Cryptosporidium sp.oocysts

Irradiation dose (Gy) and mice groups* Postinfective days and Control/dye methods Control 0 5001 7502 10003 15004 20005 Day 3 - - - -Day 7 + + + - - Day 12 ++ +++ + + - Day 17 +++ ++ + + - Day 22 +++ ++ + + - Day 27 +++ + + + ? Day 32 ++ + + + - -Safranin + + + + - -Rhodamine- Auramine + + + + - -( - ) Negative ( + ) ‹103 _{Oocysts /g} ( ++ ) 103_-104 _{Oocysts /g} ( +++ ) ›104 _{Oocysts /g}

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Table 6. Effect of Gamma Irradiation on Bacteria from Treated Wastewater

Bacteria Irradiation Dose (Gy) / Percent Inactivation

500 700 900 1000 1500 2000 2500 3000 5000

Total coliforms - - 45 99.30 99.99

Fecal streptococci - - - 99.50 99.50 99.90 99.99

Salmonella sp. 30 70 99.99

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Table 7. Effect of Gamma Irradiation on Bacteria from Biosolids

Bacteria Irradiation Dose (Gy) / Percent Inactivation

500 700 900 1000 1500 2000 2500 3000 5000

Fecal coliforms 62.50 99.90 99.97 99.99

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For wastewater plants, the organisms of public concern such as

Cryptosporidium and Giardia have proven more difficult to

inactivate with conventional treatment systems.

This study shows that irradiation could be effectively used for the

disinfection of effluents from the municipal wastewater treatment

plants, especially if reuse in agricultural practice is to be

considered.

An electron beam accelerator would improve the economics of the

process and reduce any public acceptance related to the use of

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