University of Puerto Rico, Cayey Campus          RISE Program 2011          BIOL 3009 – LAB          Research Proposal    ...
Title Analysis of the direct bacteriological contamination of the superficial and underground waterbodies of the San Salva...
maintenance of these water systems by analyzing bacteria in the water of the area and checking ifthe results have any rela...
Introduction   In Puerto Rico, 97% of the population receives their water resource from the Water andSewage Authority. The...
(Census Bureau, 2000). The exact number of people of this area who use community watersystems is not certainly known, but ...
Because of the geological and public health implications of the contamination of water in thecommunity water systems of th...
have the presence of bacteriological pollutants which will have a direct effect on the health ofpeople who use the water f...
Methodology       For this investigation, it is intended to study the community water systems of the SanSalvador neighborh...
about five samples monthly (of about 2ml each) of every of the water bodies found in the SanSalvador community (to have at...
72°C. This cycle is repeated for an average of 35 times. The whole amplification process cantake about 2 hours.       Afte...
samples show no bars at all it will mean that they have resulted to be negative to the presence ofthe E. coli gene.       ...
magazines as well as to report the outcomes to the Government’s Agencies in charge of waterconservation, such as the Water...
Expected results and discussion       After making this investigation we will find that there are several community waters...
In case that the hypothesis doesn’t result as a fact, several other studies will be made onother communities so that the c...
ReferencesAutoridad de Acueductos y Alcantarillados, Actualización del Inventario de Sistemas deAcueductos Comunitarios In...
Perdomo, C.H., Casanova, O.N., Ciganda, V.S., Contaminación de aguas subterráneas connitratos y coliformes en el litoral s...
Upcoming SlideShare
Loading in …5
×

Proposal angelica m. gonzalez (written)

334 views

Published on

Published in: Technology, Business
0 Comments
0 Likes
Statistics
Notes
  • Be the first to comment

  • Be the first to like this

No Downloads
Views
Total views
334
On SlideShare
0
From Embeds
0
Number of Embeds
18
Actions
Shares
0
Downloads
0
Comments
0
Likes
0
Embeds 0
No embeds

No notes for slide

Proposal angelica m. gonzalez (written)

  1. 1. University of Puerto Rico, Cayey Campus RISE Program 2011 BIOL 3009 – LAB Research Proposal Angélica M. González Sánchez Natural Sciences Department Copyright 2011©
  2. 2. Title Analysis of the direct bacteriological contamination of the superficial and underground waterbodies of the San Salvador neighborhood in Caguas and its implications on the community water systems. Abstract Most of the water used by the population of Puerto Rico (97%) comes from the Waterand Sewage Authority. However, the rest of the population (3%) acquires their water fromcommunity water systems which obtain it from water bodies near the community and on whichthe residents themselves build systems to distribute the water. Many of these community watersystems haven’t been examined by the agencies in charge of this and many of the ones that havebeen inspected have shown deficiencies because of the lack of economic resources, thedisinformation of the communities on the established parameters for the conservation of tapwater, for the construction of water and septic systems and for the disposal of used waters, andbecause of the lack of knowledge of the residents on the protocols for the good constructing,processing and maintenance of water systems. Because of all these factors, this investigationinfers that the water from the water bodies that serve community water systems is contaminatedby human means, especially by fecal contamination developed from the percolation of septictanks. To determine with certainty all of this, the San Salvador neighborhood from Caguas,Puerto Rico has been chosen to study the community water systems, which are known to be usedin the area, the septic systems and the presence of bacteriological pollution in the water bodies ofthe zone. This inquire will also treat the health and legal implications of the construction and
  3. 3. maintenance of these water systems by analyzing bacteria in the water of the area and checking ifthe results have any relationship with diseases derived from those bacteria and by researching theestablished parameters for these water systems and comparing them with the qualitative evidenceon the systems of this community. This research also implies the participation of the communityin interviews and, lastly, in orientations about the results of the study. After this research, it isexpected to conclude that in the San Salvador neighborhood there are certain violations to thefederal and state regulations on tap and used water, which result from ignorance of the topic andwhich cause the bacteriological contamination of water along with the spread of diseases relatedto those bacteria.
  4. 4. Introduction In Puerto Rico, 97% of the population receives their water resource from the Water andSewage Authority. The other 3% of the population (approximately 125, 000 persons) obtainstheir water from community water systems, which, as their name says, are operated by theresidents of the community themselves (EPA, 2001). These community water systems can beclassified on superficial or underground, according to the source of the water. As studied by theWater and Sewage Authority of Puerto Rico in 2004 (AAA, 2004) there are about 239 superficialand underground community water systems in our Island. About 41% of these 239 communitywater systems do not reach the bacteriological parameters established by the Department ofHealth of Puerto Rico, the Environmental Quality Board of Puerto Rico, the Water and SewageAuthority or by the Environmental Protection Agency (EPA) under the National PrimaryRegulation on Tap Water (EPA, 2003). As several studies from these agencies expose, this maybe because of the insufficient amount of trustable epidemiologic information, the pooreconomical resources invested on the construction and maintenance of these community watersystems, the scarce education of the people who administrate these systems, the lack of therequired filtration methods on most of them and by the construction deficiencies of old-uninspected septic tanks which allow percolations. It can be said by this that this is a multisourceproblem. Because of all these, it has been decided to investigate the real conditions of the water of oneof these community water systems. The chosen community water system is located in the SanSalvador neighborhood in Caguas Puerto Rico. San Salvador neighborhood borders in the Northwith Borinquen neighborhood and in the South with the Municipalities of Cayey and SanLorenzo. This vicinity has a population of 3,272 habitants distributed in about 1,133 houses
  5. 5. (Census Bureau, 2000). The exact number of people of this area who use community watersystems is not certainly known, but that’s why it is one of the objectives of this research. Sincethis is a rural community which was founded several decades ago, it is not so updated with therequired parameters and technology for the treatment of tap water and for the disposal of usedwater. Many of the employed water of this area is disposed on septic tanks or flows through theditches, landing ultimately on the water bodies and the soil of the area as percolation. That way,both the superficial and the underground water result affected. This can have a direct effect onthe population because it can be a medium of reproduction and transmition of microorganismssome of which may be pathogenic. The diseases that these pathogenic agents cause are calledwater diseases because they are mostly transmitted by the consumption of contaminated water.According to the Health Department of Puerto Rico, (Departamento de Salud de Puerto Rico,2008) the most common water diseases are: diarrhea, vomits, fever, respiratory infections,urinary infections, meningitis, between others. Most of these are caused by protozoans such asCryptosporidium parvum, by bacteria such as Salmonella, Shigella, Klebsiella, Enterobacter,Citrobacter, by viruses such as Group A Rotavirus or by parasites such as Giardias andAmoebas. Most of these microorganisms are very hard to detect because of the small amounts ofthem that are typically found on samples, but by detecting the presence of a specific bacteria wecan determine the presence of these. E. coli is a bacterium from the coliform group and it is usedas an indicator of fecal contamination because its incidence reveals the existence of othermicroorganisms which can be pathogenic. It is also a well known and studied bacterium andmost of its commonly found strains are non-pathogenic. That’s why it is mostly used in the waterquality tests. However, the absence of these bacteria does not necessarily exclude contaminationof water with protozoa or virus.
  6. 6. Because of the geological and public health implications of the contamination of water in thecommunity water systems of the San Salvador neighborhood explained above, it has beendecided to propose the making of this investigation. By means of it, it is intended to investigateabout the established parameters for the conservation of tap water, and also about its concern andobservance in the area. On the other hand, it is deliberated to make some fieldwork to inspectexactly how many community water systems are in the area and how many residents of theneighborhood use water from them. It is also proposed to evaluate the condition, the givenpermits and the maintenance of the community water and septic systems in the vicinity of SanSalvador to determine if they are a potential risk in the quality of both superficial andunderground water of the zone. To do this, it is planned to compare the construction of thecommunity’s septic tanks with the regulations of the Individual System of Residual WaterDisposition, as established by the Administration of Regulations and Permits of Puerto Rico(ARPE) in the Edification Guideline. After making this research, it is projected to determine if there are found bacteriologicalpollutants from the coliform group in the underground and superficial water bodies of the SanSalvador neighborhood in Caguas. Because of the health issues related to the presence of suchmicrobes in water, it is also intended to analyze the existence and the recurrence, if any, ofdiseases related to these bacteriological pollutants in the residents of the studied community.Finally, after all this investigation and analysis has been made, the main objective of it is to raiseawareness about the regulations for the conservation of tap water by orienting the residents of thearea and publishing the results. During this investigation it is expected to find out that the community water systems ofthe San Salvador neighborhood from Caguas are being polluted by human means and, therefore,
  7. 7. have the presence of bacteriological pollutants which will have a direct effect on the health ofpeople who use the water from them without decontaminating it first.
  8. 8. Methodology For this investigation, it is intended to study the community water systems of the SanSalvador neighborhood in Caguas because it is known that there are several families that live init and use the water from these kind of systems. These systems haven’t necessarily beeninspected or studied before and there is no certainty on the quality of the water in them becauseof the percolations from septic tanks and from other human activities, because of the insufficientamount of trustable epidemiologic information and because of the poor economical resourcesinvested on the construction and maintenance of them. To reach the objectives of this research, there are several steps that have to be done. First, itwould be needed to research about the established parameters for the conservation of tap water inPuerto Rico, mainly about the community water systems, the septic tanks and their use andobservance in the studied zone. This will be completed by visiting the agencies in charge of thesetopics in Puerto Rico, such as the Environmental Quality Board (JCA), the Water and SewageAuthority (AAA), the Environmental Protection Agency (EPA), the Health Department and theAdministration of Regulations and Permits of Puerto Rico (ARPE). It will also be done byvisiting and interviewing the residents of the San Salvador neighborhood to ask them about theiruse of the community water systems, the construction of them and of the septic tanks. We willalso inquire about the permissions they have processed for them, if any. This way we will beable to compare the existent evidence with the instituted regulations. The next step on this investigation, in order to achieve the goals of it, would be to takesamples and analyze both the underground and superficial water bodies of the studied zone todetermine the presence of bacteriological pollutants. This will be done during a year by taking
  9. 9. about five samples monthly (of about 2ml each) of every of the water bodies found in the SanSalvador community (to have at least 60 samples of each water body after twelve months). Aftertaking each sample, they will be stored in a portable refrigerator until taken to a laboratory whereit should be analyzed. Then, the PCR (Polymerase Chain Reaction) will be used to detect theexistence of the most common fecal contamination indicator bacteria, called Escherichia coli. Bythe PCR procedure trace amounts of DNA can be quickly and repeatedly copied to produce aquantity sufficient to investigate using traditional laboratory methods (Reece, Urry, et. al., 2011).This procedure will be used because it is a very sensitive method for the detection of genes fromthese bacteria from the small quantities of them that will supposedly be found and because it willhelp to analyze the samples in a relatively conventional way. The PCR procedure implies several steps. First the samples must be purified by a fewprotocols, which include centrifuging, adding Lysis solution and removing the cellular trash.After all the protocol is done, we will keep only the chromosomal DNA that we want to study.Then, we will prepare the final sample for the PCR by adding a primer solution specific for theE. coli DNA and the processed sample to a PCR tube which has Taq polymerase, dNTPs(nucleotides), Mg² and a buffer, which allow the amplification of the DNA to occur. Then, thepurified chromosomal DNA sample must be amplified by putting it in the Thermocycler andadjusting it to make a 3 stage cycle which consists of: denaturation, which separates de DNAstrands and occurs at 95°C, annealing, which grips the primers to the DNA strands by allowingthem to form hydrogen bonds with the ends of the target sequence and occurs at 50°C, andfinally, elongation, where the DNA strands duplicate and elongate because the DNA polymerase(Taq polymerase, in this case) adds nucleotides to the 3’ end of each primer at a temperature of
  10. 10. 72°C. This cycle is repeated for an average of 35 times. The whole amplification process cantake about 2 hours. After the PCR procedure, electrophoresis must be done so that we will be able to perceiveif there is actually the presence of E. coli in the sample. The electrophoresis shows the amplifiedgenes by presenting bars. This process is made in an agarose matrix which has ethidiumbromide, a chemical compound that intertwines with the DNA strands and fluoresces. The matrixalso has a buffer (TBE or TAB) which provides the electrolytes (ions) so that the appliedelectrical current (usually 90 to 100 Volts) can pass the matrix. There are also some wells in thematrix on which the DNA samples are going to be put. Before putting the samples in the wells,they have to be tinted with a loading dye which will also give weight to the sample, preventing itfrom getting out of the course. Also, we have to make sure that the wells are aligned with thenegative charge source because that way, the DNA strands (which have a negative charge) willbe attracted to the opposite positive side of the matrix and, therefore will extend to it accordingto their molecular weight, allowing us to determine the presence or absence of the studied DNA,in this case the one from E. coli. Along with the strands, in the first two wells, two molecularweight markers will be put to compare the molecular weight of the studied genes with the one ofa known gene. After carefully putting the markers and the DNA samples in the wells with thehelp of a micropipette, the electrophoresis chamber must be turned on and watched for 45minutes. After that, the agarose matrix should be put under an ultra violet light and observed forthe results of the presence of amplified genes. The markers will surely show constant bars whichindicate comparative amounts of molecular weight (in kb). If the samples result positive to the E.coli gene they will show wide and consistently dark bars, if the sample results slightly positive tothe gene (have a little amount of the gene) they will show lighter and slimmer bars, finally, if the
  11. 11. samples show no bars at all it will mean that they have resulted to be negative to the presence ofthe E. coli gene. All of these simple laboratory methods will allow us to determine if the water iscontaminated with bacteriological pollutants because the presence of E. coli indicates thepresence of other prejudicial microorganisms. After making sure of the presence or the absence of bacteriological pollutants, it isproposed to investigate the relationship between the bacteriological pollutants present in thewater of San Salvador neighborhood and the occurrence or recurrence, if any, of infectiousdiseases derived from these. This will be done by making interviews to the residents of the zoneand asking them about how they process and use their water, if it comes from a community watersystem and if they have gotten any of the symptoms of the water diseases described in theintroduction after drinking water from the community water systems. The last step on this investigation is one of the most important ones because it is the onethat allows the people of the community to know about the results of this research. It will consiston orientations in the community centers, the handling of informative brochures andpersonalized orientations in each of the houses of the neighborhood and on publications in theregional newspapers. This way the residents of the San Salvador neighborhood will get educatedabout the established parameter for the conservation of tap water, the regulations for the disposalof used waters and for septic tanks and about the results of the analysis of the water theyconsume. Therefore, they will get informed, will have the tools to correct any violations to theregulations that they may have and will make sure that in the future the quality of the water theyare consuming is acceptable. On this step, it is also planned to publish the results in scientific
  12. 12. magazines as well as to report the outcomes to the Government’s Agencies in charge of waterconservation, such as the Water and Sewage Authority, so that they can help in the process ofkeeping the quality of the water of the studied community. This will also increase consciousnessabout the conditions of the community water systems and, as a result, promote the government’sconcern and actions about it.
  13. 13. Expected results and discussion After making this investigation we will find that there are several community watersystems in the San Salvador neighborhood from Caguas either private or controlled by anadministrative board of residents. This research will show that approximately an 80% of thefamilies use the water from community water systems. It will also be found that there are quite afew violations to the established parameters for the conservation of tap water, for the discard ofused waters, for the construction of community water systems and for septic tanks. Theseviolations, along with other human activities such as irresponsible waste disposal, are resulting ina direct contamination to the water bodies of the studied area. This contamination will cause therapid and continuous reproduction of bacteria because it will mainly come from the percolationsof septic tanks. These results will be tested by the PCR which will show the presence of E. coligenes in the water, which as a result will mean that there are other bacteriological pollutants inthe water of the zone and that it is not adequate for human consumption. This will also be shownby the direct relationship that it will have with the recurrence of infectious diseases derived frombacteria in the people who use the water without filtering and decontaminating it with somechemical agent or with high temperatures before consuming it. This investigation will also resultin a better understanding and orientation of the residents on the water system’s topic, which willalso outcome a better preservation and care of the water of the area. This will raise the awarenessof the community about the importance of the conservation of tap water and all what it implies. Itis also be expected that people will still be satisfied with their community water systems eventhough the study reveals the presence of bacteriological pollutants because most of the residentshave been in the neighborhood for decades and they must still feel comfortable and safe withtheir water sources.
  14. 14. In case that the hypothesis doesn’t result as a fact, several other studies will be made onother communities so that the conservation and quality of community water systems will bepromoted. This inquiry will therefore serve as a template for future studies in other several areasof the Island where no previous investigations have been done. It can also lead to new researchesabout how to improve the quality of the community water systems and about the specific bacteriathat are in them. When this study is completed, it can lead to an improvement in the usedtechniques and therefore to an eventual repetition of it to compare the results. The outcome ofthis research can also lead to resolve the economic problem that leads to the poor maintenance ofthe community water systems. This can be done by making the government aware of the problemand, therefore, helping in the promotion of the implementation of viable federal or state financialaids so that these communities would be able to take correct care of their water systems withoutmonetary implications to hold them back.
  15. 15. ReferencesAutoridad de Acueductos y Alcantarillados, Actualización del Inventario de Sistemas deAcueductos Comunitarios Independientes en Puerto Rico, 2004. http://www.acueductospr.comDepartamento de Salud Gobierno de Puerto Rico, Reglamento General Salud Ambiental, 2008.www.salud.gov.pr/ Publicaciones/ Anuncios/ Document/ Reglamento General SaludAmbiental.pdfEnvironmental Protection Agency, Small Drinking Water Systems Handbook, 2003.www.epa.gov/safewaterEnvironmental Protection Agency, Contaminants regulated under the Safe Drinking Water Act,2001. www.epa.gov/safewater/mcl.htmlJunta de Calidad Ambiental, Estado Libre Asociado de Puerto Rico - Oficina del Gobernador ,Reglamento de Estándares de Calidad de Agua de Puerto Rico, marzo 2010.www.gobierno.pr/JCA/LeyesReglamentos/Reglamentos/Microjuris, Artículo V-b-3.0 del Reglamento de Edificación de ARPE, 2002,www.microjuris.com/mjpr/reglamentos/is/reg_lista_regs.cfm?AgencyIDMontandon PE, The use of PCR for detecting pathogenic microorganisms in water, Pubmed,May 17 1994. http://www.ncbi.nlm.nih.gov/pubmed/8209134
  16. 16. Perdomo, C.H., Casanova, O.N., Ciganda, V.S., Contaminación de aguas subterráneas connitratos y coliformes en el litoral sudoeste del Uruguay, Agrociencia, julio 2001, pag. 10-22.http://www.fagro.edu.uy/agrociencia/VOL5/1/P10-22.pdfReece, J., Urry, L., [et. al.]. 2011. Campbell Biology. Ninth Edition. Pearson BenjaminCummings. Chapter 20:Biotechnology. Pages: 396-423.Rompré A, Servais P, [et. al.], Detection and enumeration of coliforms in drinking water:current methods and emerging approaches, Pubmed, March 2002; pages 31-54.http://www.ncbi.nlm.nih.gov/pubmed/11777581United States Census Bureau, Census of population and housing, DIANE Publishing,2000. http://books.google.com.pr/books?id=B9FjIc1a_qoC&pg=PA5-IA2&lpg=PA5-IA2&dq=millas+cuadradas+barrios+Caguas&source=bl&ots=sNJDkNX5x-&sig=PV5wH6hYjI7smYFNrEc_LeXJbxo&hl=en&ei=360YTtinM5DrgQe5gqQa&sa=X&oi=book_result&ct=result&resnum=10&ved=0CG0Q6AEwCQ#v=onepage&q&f=falseUSDA/Agricultural Research Service, Detecting pathogens in waterways: An improvedapproach, ScienceDaily, 2011, February 8. http://www.sciencedaily.com-/releases/2011/02/110208144124.htm

×