Bacteriological analysis of the water in El Caño Martín Peña

Detection and quantification of Enteric Pathogenic Bacteria Salmonella and Shigella spp.

1 / march/ 2013

Mario R. BarreraGeorge M. RiveraPaola TabaroMentor: Prof. Mayra RolónUniversity of the Sacred Heart

Meet the Team

Mario R.Barrera

PaolaTabaro

George M. Rivera

Mentor:Prof: Mayra Rolón

Meet the Team

2012

2013

Caño Martín Peña

Is an invaluable natural habitat for the Metropolitan area of San Juan.

Caño Martín Peña

+ During the recurring years the mangroves and lagoons stretched of the Caño have been contaminated and stretched out into smaller measurements.

Caño Martín PeñaThe deterioration of this area has taken its toll on the residents lifestyles.

How to Improve !!

The decontamination of the lagoon that boards the community.

Rehabilitation of the estuary system.

To broaden the community’s knowledge of the microbial presence and the harms that can be encountered.

Our objectives

+ To detect the presence of pathogenic enteric bacteria such as Salmonella and Shigella SPP.

+ To acquire knowledge of the microbial presence found in the Caño Martín Peña.

+ Refine research skills through different bacteriological techniques.

+ Analyze and compare the degree of contamination found with past investigations.

+ Raise awareness to the community.

Hypothesis

In the past year our fellow classmates were able to detect the presence of Shigella serotypes A,B and C.

The Caño Martín Peña has excessive levels of coliforms, indicators of fecal matter that represent a hazard for the community.

Hypothesis

+ Because of this information we can say there is a high probability of encountering Salmonella and Shigella SPP. Bacteria due to the high coliform levels resulting from water contamination by fecal matter.

Coliform bacteria

Rod-shaped Gram negative bacillus.

Lactose fermenters with the production of acid and gas when incubated at 35-37°C.

Easy to culture and their presence is used to indicate fecal contamination by other pathogenic organisms in the water.

Enterobacteriaceae

Total coliforms: Escherichia Klebsiella Serratia Citrobacter

Fecal coliform: Eschericia coli

Escherichia coli

Klebsiella pneumoniae

Enterobacter

Serratia marcescens

Citrobacter

Salmonella spp. Vs Shigella spp.

Salmonella spp.

Rod-shaped Gram negative bacillus

Non-spore forming. Motile enterobacteria

Facultative anaerobes

Hydrogen sulfide production which can be detected on a TSI growth media

Shigella spp.

Rod-shaped Gram negative bacteria

Non-spore forming and non-motile bacteria

Salmonella Pathogenicity

Most of the infections are due to the ingestion of contaminated food.

Salmonella is responsible for various illnesses such as: Typhoid fever Paratyphoid fever Foodborne illnesses

Enteritis Salmonella also known as food poisoning Salmonella.

Shigella Pathogenicity

Most of the infections are caused via ingestion.

The illness is known as Shigellosis.

During infection, Shigella typically causes dysentery .

Invasion of Shigella Bacterial multiplication Spreading to epithelial

cells Tissue destruction

Investigation Site

Station #1

Width : 8 Ft.

Depth: 2.05 Ft

Observations Boat nearby Carton Boxes

Station #2

Width: 3 – 5 Ft

Depth: 4 Ft

Observations Current of water

coming from a pipe. Fishing traps

Station #3

Width: 8 Ft.

Depth: 2 Ft.

Observations Samples were taken

at the shore, because difficult access.

Garbage bags

Methodologies

Dilution of sample

Membrane filtration

Cultivation SS Agar MacConkey Agar EMB RVS

Identification Enterotubes

Procedure

DilutionMembrane Filtration

Cultivation

Dilution of sample

At the Laboratory

Membrane filtration 0.45 μm

At the Laboratory

Membrane filtration

Pure sampleDiluted Sample

1:1000

Cultivation

At 37º C.

24 – 48 Hours

Selective Mediums

Salmonella Shigella Agar

MacConkey Agar

Selective Mediums

RVS EMB

Taxonomic Determination: Enterotube System

Taxonomic Determination: Enterotube System

Enterotube System

Citrobacter freundii

Serratia plymuthica

Sampling Dates

M1 – 30/ October/ 2012 M2 – 28/ November/ 2012 M3 – 7/ December/ 2012 M4 – 30/ January/ 2013 M5 – 11/ February/ 2013

Types of Bacteria

Characteristics of the Colony

Bacteria

Salmon Colony Klebsiella ozanae

Pink Colony Enterobacter cloacae

Pink Colony Klebsiella pneumoniae

Pink colony Enterobacter aerogenes

Transparent Colony Shigella

Transparent Colony Yersinia

Black Colony Citrobacter freundi

Pale pink Serratia plymuthica

Metallic Green E. coli

Types of Bacteria

Bacteria

Serratia marcenscens

Enterobacter aglomerans

Enterotubes

Shigella Enterotube

Findings

Year Quantity

2008 118,000

2009 3,335,000

2010 6,800,000

2011 60,000,000

2012 69,000,000

2013 25,333,333

Year Quantity

2000 4,000,000

2001 5,500,000

2002 4,000,000

2003 2,000,000

2004 100,000

2005 73,000

2006 80,000

2007 57,000

Number of bacteria/ ml of water

Findings on Dilutions

Station #1

1 col in 10¯⁷ : 10,000,000 bact/ml

Station #2 4 col in 10⁻⁶: 4,000,000 bact/ml

Station #3 62 col in 10⁻⁶: 62,000,000 bact/ml

Findings on Filtration

Station#1 Pure > 100 col (TNTC) Dilute: 5 col * 1000: 5,000 bact/100ml

Station #2 Pure>100 col (TNTC) Dilute: 40 col * 1000: 40,000 bact/100ml

Station #3 Pure>100 col (TNTC) Dilute: 3 col * 1000: 3,000 bact/100ml

Citrobacter freundiCedecea lapageiKlebsiella pneumoniaeYersinia enterocoliticaCitrobacter freundiCedecea lapageiEnterobacter agglomeransKlebsiella ozaenae

Enterobacter agglomerans

Conclusions

The high level of contamination is due to:

Stagnant water in some areas.

Sewage and water discharge in to the cano.

Deposition of debris into the community.

Conclusions of the Research

From 2004 to 2008 the amount of bacteria were less than a million/ ml of water.

This could be due to the removal of debris

Effort of government and people that lives in the community.

+ During our Research period we noticed a sudden decrease in the level of bacteria.

+ Although we were not able to isolate colonies of Salmonella, we did isolate Citrobacter freundi, Yersinia enterocolitica and Shigella Spp.

+ Which are indicative of the severity of contamination and a possible source of contamination with pathogens to the residents of nearby areas.

We recommend:

To maintain the movement of dredging and cleaning of the entire San Juan Estuary.

We saw that according to data, levels of bacteria were down under a million. With a little of effort is possible to maintain to provide a better life to the people.

To Help organizations and people that live there in a constant way not only, to show interest when elections are near.

References

Http://www.mayoclinic.com/health/shigella/DSOO719

Http://www.cdc.gov/salmonella/enteritidis/

Centers for Disease Control and Prevention. October 2005. Shigellosis. http://www.cdc.gov/nczved/divisions/dfbmd/diseases/shigellosis/

Indian and Northern Affairs Canada. May 2003. First Nations Water Management Strategy. Http://www.ainc-inac.gc.ca/ai/arp/es/0506/fnwms/fnwms-eng.asp

References

Public Health Agency of Canada:Notifiable Diseases on-line. December 2003. Shigellosis. http://dsol-smed.hc-sc.gc.ca/dsol-smed/ndis/diseases/shig_e.html

www.safewater.com

World Health Organization. 2005. Guidelines for the control of shigellosis, including epidemics due to shigella dysenteriae type 1. http://whqlibdoc.who.int/publications/2005/9241592330.pdf

United States Pathogenic Microorganisms and Natural Toxins Handbook. January 2002. Shigella. Http://pdf.usaid.gov/pdf_docs/ PNADO152.pdf