Abstract The presence of motile Aeromonas species in drinking water, the Porsuk River and tap water

1
Abstract The presence of motile Aeromonas
species in drinking water, the Porsuk River and
tap water in the Eskisehir (Turkey) was
investigated. Fresh water samples were collected
from 12 different sites of Porsuk River
(Eskisehir,Turkey) and 12 drinking water and tap
water sources in various parts of the city during
all year. Aeromonas spp. was not detected in
drinking and tap waters but was found in the
Porsuk River. The Porsuk River isolates were 13
Aeromonas hydrophyla, 37 Aeromonas caviae, 35
Pseudomonas putida, and 15 Comomonas
acidovorans. Keywords Aeromonas spp, drinking
water, river water, tap water.
The occurrence of Aeromonas in Drinking Water,
Tap Water and the Porsuk River M. Kivanç, M.
Yilmaz, F. DemirAnadolu University, Science
Faculty, Department of Biology, Eskisehir,
TURKEY(E-Mail mkivanc_at_anadolu.edu.tr
meralyilmaz_at_anadolu.edu.tr)
Introduction Aeromonas are Gram-negative,
oxidase-positive, facultative anaerobic, glucose
fermenting, rod-shaped bacteria of the family
Aeromonadaceae (Popoff, 1984). Aeromonas spp. are
common aquatic microorganisms that occur in
different water (Kaper et al., 1981 Gavriel et
al., 1998 Fiorentini et al.1998 Soler et al.,
2002). Many Aeromonas species have been
associated with human diseases, including
gastroenteritis, hemolytic-uremic syndrome,
septicemia, etc.(Janda and Abbott,1996).
Residents of the City of Eskisehir in Turkey use
treated water from the Porsuk River not for
drinking but for cleaning, washing, and various
recreational purposes as tap water. Lack of
information about the incidence of motile
Aeromonas in Eskisehir tap water, drinking water
and the Porsuk River should be great concern
because their possible occurrence in such waters
could be a potential source of infection in the
residents. In this study, the existence of
Aeromonas species in the Porsuk River, tap water
obtained from the Porsuk River after treatment,
and drinking water were determined in the City of
Eskisehir.
The present study also revealed that 100
of A. hydrophila, 65 of A. caviae exhibited
beta hemolytic activity. Beta hemolysin has been
reported as a virulence factor in motile
aeromonads (Majeed and MacRae, 1993 1994). World
health organization (WHO) has reported the
isolation ratio of A. hydrophila from drinking
water to be 127 (Fewtrell and Bartram, 2001).
In our study, we have isolated and identified 8
(13.3) strains of A. hydrophila and 22 (36.6)
strain of A.caviae from 144 Porsuk River samples.
Materials and methods Collection of water
samples The study was undertaken in 12
stations, monthly sampled for a period of a year.
Tap water samples were collected from houses,
business centers and hospitals, randomly selected
different parts of the City of Eskisehir.
Drinking water samples in plastic containers were
purchased from a variety of retailers in the
city. Fresh water samples were collected from 12
different sites of the Porsuk River. Isolation
and identification of the strains 10 ml of
water were inoculated in 90 ml of peptone water
with 1 NaCl (w/v) adjusted to pH 8.6 with sodium
hydroxide. After incubation 37C for 2448 h, the
cultures were streaked on Glutamate Starch Phenol
Red agar (GSP) and incubated at 37C for 2448 h
(Dumontet et al., 2000). Three to five yellow
colonies surrounded by yellow zone on GSP medium
were maintained on nutrient agar slants at 4C
after subculturing several times. The
strains were first identified to the phenospecies
level by he methods recommended by Popoff (1984)
and Mac Faddin, (1980). Isolates was identified
by Vitec Gram-Negative Identification Panel
(Vitek, BioMérieux).Vitek GNI cards were
processed according to the manufacturers
specifications. Ribotyping was performed
with the RiboPrinter Microbial Characterization
System (Qualicon Inc., Wilmington, DE) and the
standard EcoRI DNA preparation kit, as described
in the manufacturer's operations and analytical
guides.
Fig 1. Normalized EcoRI riboprint patterns of the
8 isolates. Relative molecular sizes are given on
the right. Line 1 and 2 isolates grouped as
Pseudomonas spp. Line 3-8 isolates grouped as
Aeromonas spp.
A. hydrophila and A. caviae isolates obtained
from different sites and season of the Porsuk
River in the City of Eskisehir was shown in Table
2.
Table 2. Isolated strains from different sites of
the Porsuk River in the City of Eskisehir
Our data are in agreement with WHO's water
quality standards. However, it has been reported
that Aeromonas bacteria found in municipally
treated drinking water possesses a wide variety
of virulence-related genes (Sen and Rodgers,
2004).We have analyzed only hemolytic activities
as virulence factor of our isolates. All of them
were found to be hemolytic. Actually, further
studies should be done in order to detect the
other virulence factors. Aeromonas species
are most frequently (82.53 of the isolates)
present in the Porsuk River during the dry
seasons from June to October (Table 2). A.
hydrophila is only recovered during Autumn and
Winter seasons. Lower isolation frequency is
noticed during the Spring season. It was
previously reported that higher numbers of
Aeromonas were recovered during Summer (Kaper et
al., 1981 Gavriel et al., 1998).
Results and discussion On the basis of
morphological, physiological and conventional
biochemical test, the Aeromonas strain isolated
from in the Porsuk River, tap water and drinking
water were identified. Sixty strains were
isolated. All isolates were Gram negative,
motile, rod shaped and facultatively anaerobes
(Table 1).
Eskisehir tap water is obtained from
Porsuk River after treatment. Although we did not
get Aeromonas spp. from tap water, there is a
potential risk everytime. It will be quite
important issue if Aeromonas spp. is able to grow
in the chlorinated water. As a result, we have
the result with there is no Aeromonas spp. in the
tap and drinking water in the City of Eskisehir.
However, the presence of Aeromonas spp. in the
Porsuk River has a potential risk in terms of the
residence health of the city. Because the
Eskisehir tap water is as long as obtained from
the Porsuk River, the health risk will increase
more due to possible presence of Aeromonas spp in
tap water. Furthermore, because of the irrigation
of vegetables, gardens, and green places with
water obtained from the Porsuk River, hazard will
get more important.
Table 1. Biochemical characterizition of
Aeromonas spp. isolated from the Porsuk River
The strains were oxidase and catalase
positive and did not tolerate NaCl concentrations
higher than 3.0. Of 60 isolates, 8 were
identified as A. hydrophila and 22 as A. caviae
using biochemical tests (Table 1). These data
were also confirmed with Vitec system
(BioMerieux Marcy-I'Etoile, France). A. sobria
which is commonly reported in the literature was
not observed in our study. Our isolates were 13
Aeromonas hydrophyla, 37 Aeromonas caviae, 35
Pseudomonas putida, and 15 Comomonas acidovorans
according to Vitec system. These isolates were
ribotyped with the RiboPrinter Microbial
Characterization System (Qualicon). Ribotyping
analysis agrees with the results of Vitec system.
A sample of ribotyping analyse result was shown
in the Fig 1.
References Dumontet S., Krovacek, K., Svenson
S.B., Pasquale V., Baloda S. B. and Figliuolo G.,
(2000). Immunol Microbiol Infect Dis., 23,
53-72. Fewtrell L. and Bartram J. (2001). World
Health Organization (WHO) Water quality
Guidelines, Standards and Health, IWA Publishing,
London. Fiorentini C., Barbieri E., Falzano L.,
Matarrese P., Baffone W., Pianetti A, Katouli M,
Kühn I., Möllby R., Bruscolini F., Casiere A.,
and Doneli G. (1998). J. Appl. Microbiol., 85,
501-511. Gavriel A.A., Landre J.P. and Lamb A.J.
(1998). Journal of Applied Microbiology, 84,
383392. Hanninen M. L. and Siitonen A. (1995).
Epidemiol. Infect., 115, 39-50. Janda J.M. and
Abbott S. L. (1996). Human pathogens. In the
genus Aeromonas. B. Austin M. Altwegg, P. J.
Gosling and S. Joseph. Chichester (ed.), Wiley,
London, United Kingdom, pp. 151-173. Kaper J.B.,
Lockman H. and Colwell R.R. (1981). Journal of
Applied Bacteriolog, 50, 359377. Liu B.M.,
Changchien C.S., Chen C.H., Chung K.J. and Kung
C.T. (2004). Chang Gung Medical Journa,l 27(10)
766769. Mac Faddin J.F. (1980). Biochemical
Tests for Identification of Medical Bacteria,
Williams and Wilkins Co., Baltimore, USA. Majeed
K.N., Macrae I.C.(1993). Microbios., 73,
281-288. Majeed K.N., Macrae I.C.(1994). J. Med.
Microbiol., 40, 188-193. Massa S., Altieri C.,
DAngela A. (2001). Int. J. Food Microbiol., 63,
169-173. Mete E., Kaleli I., Demir M. and Cevahir
N. (2002). J. Aknem, 16, 430433. Midilli
K.(1998) The occurence, frequency and biotyping
of Aeromonas in diverse water samples collected
in Istanbul PhD thesis, Cerrahpasa Medicine
Faculty, Istanbul. Popoff M.Y. (1984). Genus III.
Aeromonas. In N.R. Kreig and J.G. Holt, (Ed),
Bergey's Manual of Systematic Bacteriology,
Williams and Wilkins, pp. 545548. Sen K. and
Rodgers M. (2004). Journal of Applied
Microbiolog, 97, 10771086. Soler L., Figueras M.
J., Chacon M. R., Vila J., Marco F.,
Martinez-Murcia A. J. and Guarro J. (2002). FEMS
Immunol. Med. Microbiol,. 32, 243-247. Vally H.,
Whittle A., Cameron S., Dowse G.K. and Watson T.
(2004). Clinical Infectious Diseases, 15,
10841089.
Wide distribution of motile Aeromonas spp.
has been previously reported in the aquatic
environment (Fiorentini et al., 1998 Mete et
al., 2002 Midilli, 1998 Sen and Rodgers, 2004)
and the results obtained support these findings.