Wild Birds As Biological Indicators Of Environmental Pollution: Biotyping And Antimicrobial Resistance Patterns Of Escherichia Coli Isolated From Audouin'S Gulls (Larus Audouinii) Living In The Bay Of Gallipoli (Italy)

Abstract E. coli biotyping and antimicrobial susceptibility tests were performed on 48 cloacal swabs collected from a population of Audouin's gulls (Larus audouini) living in the Bay of Gallipoli (Lecce, Italy). The aim was to assess the pathogenic potential of the strains the gulls carry and shed into the environment and to gain a better understanding of the microbial pollution of the area they live in. Only one E. coli strain was rhamnose - (Rh) (biotype 8). All the other strains were Rh + with a prevalence of biotypes 30 and 31. Overall, the antimicrobial susceptibility tests showed a low level of antibiotic resistance. The findings suggest that the gull population does not have a heavy impact on the microbial nature of the Bay of Gallipoli and that the area does not present a high level of microbiological pollution .

rhamnose, sorbose, and saccharose). Evaluation of a strain's sensitivity to antibiotics is another useful way to differentiate E. coli given that strains with a greater pathogenic potential present resistance to a wide range of antibiotics (Krumperman, 1983;Harwood et al., 2000). In this study E. coli strains isolated from Audouin's gulls (Larus audouinii) living in the Bay of Gallipoli (Lecce, Italy), were biotyped and tested for their antibiotic susceptibility to evaluate the pathogenic potential of the strains the gulls carry and shed into the environment and the related public health risks implied. .

Material and methods
Source of E. coli strains Forty-eight E. coli strains isolated from cloacal swabs of Audouin's gulls (Larus audouinii), living in a permanent colony in Gallipoli Bay (Lecce, Italy), were analyzed. The birds were sampled during the 2002 spring ringing program. The samples were put into Amies medium immediately after they were col-

Introduction
Gulls are considered to be important indicators of the level of pollution of their habitats. Their feces often contain enteric bacteria (E. coli, Salmonella spp., Aeromonas spp., Campylobacter spp.) that are potentially pathogenic for humans (Levesque et al., 1993). It is generally thought that gull feces contribute to the bacterial pollution of superficial waters and coastal areas where the birds live (Fogarty et al., 2003). E. coli is considered a quality indicator sentinel for coastal waters. Therefore, the characterization of E. coli can provide useful information on the quality of an environment and assess the potential repercussions of microbial pollution on public health. Amongst the methods used to differentiate E. coli strains, biotyping is a technique which is simple to perform and provides an indication of the pathogenicity of the strains analyzed (Camguilhem and Milon, 1989). Biotyping is based on the strains' ability to ferment 5 different sugars (dulcitol, D-raffinose, L-

Biotyping
Biotyping was performed according to the method described by Camguilhem and Milon (1989).

Results and discussion
Based on the biotyping results (Fig. 1) only one E. coli strain was rhamnose -(Rh) (biotype: 8) while all the other strains were Rh + Biotypes 30 and 31 were clearly prevalent.
Antimicrobial susceptibility tests showed a low resistance in all the strains. All the tested strains were susceptible to gentamicin and amikacin, while more than 90% of them were sensitive to colistin, difloxacin, apramycin, trimethoprim/sulphametoxazole and enrofloxacin (Table 1); 54.16% of the strains showed no resistance to any of the tested drugs. Multi-resistance was present in very few strains with, 29.16% of the strains presenting resistance to only one antibiotic and 6.5% to 2 and 3 drugs. Only two strains showed a broader range of multi-resistance involving 6 antimicrobial agents

Conclusions
Gulls can play an important role in the spread of potentially pathogenic agents along humid coastal areas; their feces, abounding in E. coli, may contribute to the pollution of superficial waters as well as bathing places, often attended by human beings (Whitman and Nevers , 2003). The findings of our study suggest that gulls may carry a wide diversity of E. coli types. Neverthless, the E. coli strains isolated were almost exclusively Rh + and this is indicative of a low pathogenic potential. Confirmation of these results by the currently ongoing molecular typing investigations would prove that the gulls living in the Bay of Gallipoli are not the main cause for microbiological pollution of the area. The results of the antibiotic susceptibility tests seem to substantiate this assumption since the isolated strains were highly susceptible to drugs active against Gram-bacteria and showed a very restricted multi-resistance range, which in coliform bacteria is linked to plasmidic transmission. The peculiar diet of Audouin's gulls, which unlike other gull species do not visit rubbish dumps and prefer to feed on fish , may have concurred to the results reported . Detection of resistance to a narrow range of antibiotics coupled with the fact that these birds are mainly exposed to bacterial groups polluting the Gallipoli coastline and coastal waters, would suggest that there is not a high level of microbial pollution in the area populated by Audouin's gulls.