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C:\windows\desktop\publicações\Antimicrobial susceptibility of 51 Campylobacter strains isolated from diarrheic and diarrhea-free dogs.
ANTIMICROBIAL SUSCEPTIBILITY OF 51 CAMPYLOBACTER STRAINS ISOLATED FROM DIARRHEIC AND DIARRHEA-FREE DOGS* J.R. Modolo1, R. Giuffrida2, C.A. de M. Lopes3
1Faculdade de Medicina Veterinária e Zootecnia, UNESP/Botucatu, CP 524, CEP 18618-000, Botucatu, SP, The minimum inhibitory concentrations (MICs) were determined by agar dilution for six antimicrobial agents (cephalexin, eythromycin, gentamicin, enrofloxacin, chloramphenicol, and oxytetracyline) using 51 Campylobacter strains (29 Campylobacter jejuni subsp. jejuni, 14 Campylobacter jejuni subsp doylei, 5 Campylobacter upsaliensis, 2 Campylobacter coli, and 1 Campylobacter lari) isolated from 29 diarrheic and 22 diarrhea-free dogs. Percentages corresponding to 50% and 90% strain inhibition (MIC 50% and MIC 90%) were determined. The results show that erythromycin, enrofloxacin, and gentamicin can be recommended for the treatment of campylobacteriosis in dogs; this opens the possibility of their therapeutic use in human medicine.
KEY WORDS: Campylobacter, dogs, minimum inhibitory concentration, antimicrobials, public SUSCEPTIBILIDADE ANTIMICROBIANA DE 51 AMOSTRAS DE CAMPYLOBACTER ISOLA- DOS DE CÃES COM E SEM DIARRÉIA. As concentrações inibitórias mínimas de seis antimicrobianos (cefalexina, eritromicina, gentamicina, enrofloxacina, cloranfenicol e oxitetraciclina) para 51 estirpes de Campylobacter (29 de Campylobacter jejuni subsp. jejuni, 14 de Campylobacter jejuni subsp doylei, 5 de Campylobacter upsaliensis, 2 de Campylobacter coli e 1 de Campylobacter lari), isoladas de 29 cães com diarréia e 22 cães sem diarréia foram determinadas pelo método de diluição em ágar. Foram determinadas as percentagens correspondentes à inibição de 50% e 90% das estirpes isoladas (CIM 50% e CIM 90%). De acordo com os resultados, a eritromicina, a enrofloxacina e a gentamicina podem ser recomendadas como drogas de eleição para o tratamento da campilobacteriose em cães, ampliando também as possibilidades terapêuticas para o uso em medicina humana.
PALAVRAS-CHAVE: Campylobacter, cães, concentração inibitória mínima, antimicrobianos, saúde with clinical campylobacteriosis reduces the duration and severity of the disease, decreases microorganism Over the past few decades, Campylobacter species shedding time, minimizing the risk of human exposure have emerged as the common cause of enteritis in to the bacteria (BOOSINGER & DILLON, 1992).
humans and animals. Enteric campylobacteriosis has Antimicrobial resistance studies of Campylobacter been considered as a highly prevalent worldwide strains isolated from dogs are rare. Most studies deal zoonosis in developed countries (NACHAMKIN, 1997) with strains isolated from humans and animals used Dogs are contaminated by oral-fecal contact, and by manifesting gastroenteritis or acting as healthy Erythromycin and chloramphenicol, the most carriers, represent a potential infection source to common antibiotics used to treat dogs have sometimes humans (BLASER et al., 1978; BRUCE & FLEMING, 1983; failed to eliminate the organism from the stool GOOSSENS et al., 1991). Antimicrobial therapy for dogs 2Pós-graduando, área de Vigilância Sanitária, Faculdade de Medicina Veterinária e Zootecnia, UNESP/Botucatu, Botucatu, SP, Brasil3Instituto de Biociências, UNESP/Botucatu, SP, Brasil.
Arq. Inst. Biol., São Paulo, v.70, n.3, p.283-286, jul./set., 2003 The aim of this study was to evaluate minimum 1996). Cephalosporins are useful in selective media inhibition concentrations (MICs) of six antimicrobial for primary bacterial isolation; however, studies report agents (cephalexin, chloramphenicol, enrofloxacin, 2-25% susceptibility of these bacteria to cephalexin erythromycin, gentamicin, and oxytetracyline) on (ABRAHANS et al., 1990; MODOLO et al., 1991). It is therefore Campylobacter strains isolated from both diarrheic important that cephalosporin selective media are not exclusively used for the primary isolation of this bacterium; the concomitant use of filtration techniques is recommended (MODOLO, 2000). Chloramphenicol showed higher MIC values in diarrheic dogs (2-32 µg/mL) than diarrhea-free (2-8 µg/mL). Considering Fifty-one Campylobacter strains were isolated from the cloramphenicol cut-off point of 8 µg/mL for dogs: 29 Campylobacter jejuni subsp. jejuni , 14 Campylobacter, it is significant that 27.4% of the diarrheic Campylobacter jejuni subsp doylei, 5 Campylobacter strains were susceptible to the antibiotic. In contrast, upsaliensis, 2 Campylobacter coli, and 1 Campylobacter all diarrhea-free strains were susceptible to the drug.
There is no solid explanation for this discrepancy, Serial dilutions in solid medium, according to the since feces were not collected from diarrheic dogs after National Committee for Clinical Laboratory Standards receiving medication. Variations in sensitivity recommendations (NCCLS,1998), were used to study the susceptibility of these strains to antimicrobial Campylobacter genus (ALEKSANDROVA et al., 1990).
agents and to determine each agent's MIC. The However, many studies have reported high sensitivity antibiotics cephalexin, erythromycin, gentamicin, to the drug when considering strains isolated from enrofloxacin, cloramphenicol, and oxytetracycline humans (SHANKER & SORRELL, 1983) and dogs (FOX et were submitted to the following procedures: each one was first diluted in 1,000 µg/mL saline, then new Cloramphenicol was effective in the treatment of dilutions were made on Petri dishes with thioglycolate dogs with campylobacteriosis, but failed in the elimination of chronic fecal excretion (DAVIES et al., corresponding to the following concentrations per 1984; MONFORT et al., 1990). In contrast, due to good plate: 0.06 µg/mL, 0.125 µg/mL, 0.25 µg/mL, 0.5 µg/ liposolubility that can enhance body tissue mL, 1 µg/mL, 2 µg/mL, 4 µg/mL, 8 µg/mL, 16 µg/mL, penetration, it can be recommended for treating extra- 32 µg/mL, 64 µg/mL, and 128 µg/mL. The final intestinal infections in dogs, as long as the animals volume of each plate was 20 mL. Bacterial colonies show no Campylobacter fecal excretion.
were suspended in saline to 0.5 MacFarland standard Enrofloxacin showed very similar MIC variations turbidity (1.5 x 108 CFU/µL) and inoculated in culture in diarrheic (1-4 µg/mL) and diarrhea-free dogs (1-8 medium using a 3-mm replicator. Escherichia coli, µg/mL). It was active against isolated Campylobacter ATCC 25922, inoculated in one of the plates was used strains demonstrated by the low concentrations, at as a control. The plates were kept at 35° C under which 90% of strains were susceptible (2-4 µg/mL, microaerophilic conditions; readings were made 48 diarrheic dogs and 4-8 µg/mL, diarrhea-free dogs).
Campylobacter is frequently sensitive to quinolones; From these results, MICs corresponding to 50% however, an increased resistance to these drugs is and 90% inhibition of Campylobacter (MIC and MIC ) seen, probably due to genetic mutations interfering with bacterial DNA girase (GREIG et al., 2003). Selective pressure caused by the indiscriminate use of these drugs in aviculture is a contributary factor. Considering the existence of cross resistance between quinolones, the occurrence of nalidixic acid resistant Campylobacter Tables 1 and 2 show the results of the different jejuni may interfere with tests for this bacterium Different results were observed for each drug, Erythromycin was the most effective against which were also considered in other investigations.
Campylobacter, with total strain sensitivity from both The variability can be used as an important marker in groups at very low concentrations (lower or equal to epidemiological research (ABRAHANS et al., 1990) 0.06 µg/mL). This is corroborated by several authors The strains isolated from both diarrheic and (FOX et al., 1984; REINA et al., 1984; SKIRROW, 1994; diarrhea-free dogs were highly resistant to cephalexin GANDREAU & GILBERT, 1998). Erythromycin is effective in (MIC 50 > 128 µg/mL for both). This is due to the high treating dogs, but was not able to eliminate the passive- resistance of these bacteria to cephalosporins and β- carrier status of some treated dogs (MONFORT et al., 1990; lactamic in general (ABRAHANS et al., 1990; TAJADA et al., BOOSINGER & DILLON, 1992; BURNENS et al., 1992).
Arq. Inst. Biol., São Paulo, v.70, n.3, p.283-286, jul./set., 2003 Antimicrobial susceptibility of 51 Campylobacter strains isolated from diarrheic and diarrhea-free dogs.
Table 1 - Antimicrobial drug concentration variations In view of the heterogeneity in the results reported and minimum inhibitory concentrations (MIC) capable in the literature in comparison to data in this study, we of inhibiting 50% and 90% of the 29 thermophilic recommend that Campylobacter antimicrobial Campylobacter strains isolated from diarrheic dogs.
susceptibility tests be performed for therapeutic purposes. Our results indicate erythromycin, enrofloxacin, and gentamicin as drugs suitable for the treatment of canine campylobacteriosis. This also opens up therapeutic possibilities for these drugs in for financial support; the staff from the Infectious Diseases Discipline at Botucatu School of Table 2 - Antimicrobial drug concentration variations Veterinary Medicine and Animal Husbandry; and and minimum inhibitory concentrations (MIC) capable Tânia Maria Martins, Adriana Cristina Pavan of inhibiting 50% and 90% of the 29 thermophilic Vieira, and Fernando Paganini Listoni for their Campylobacter strains isolated from diarrhea-free dogs.
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A N T O N I O A V E R S A C U R R I C U L U M V I T A E INFORMAZIONI PERSONALI ANTONIO AVERSA 03.11.1964 Consulente Dipartimento Medicina Sperimentale, Sapienza Università di Roma 06/45421411 335/6642900 [email protected] ANTONIO AVERSA ISTRUZIONE E FORMAZIONE Laurea in Medicina e Chirurgia 1990 Università “Sapienza”, Roma - Abilitazione al a p