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Over a 9-day period in February 1995, 16 newborn babies (age range 2-11 days) and 3 infants (24, 47 and 180 days of age) in a neonatal nursery ward developed diarrhoea accompanied by pyrexia and weight loss. Known enteropathogens were not detected in their stools but Escherichia coli displaying aggregative adherence to HEp-2 cells (enteroaggregative E. coli) were found in 12 (63%) ill infants and in none of 5 well neonates (P = 0.02). The illness lasted 3-9 days (mean 5.2) in 16 babies, whereas in 3 neonates it showed a protracted course of 18-20 days. The source of infection and the mode of transmission remained unclear. The outbreak isolates manifested properties common in this new group of diarrhoeagenic E. coli: mannose-resistant haemagglutination, haemolysis on blood agar, and clump formation in liquid culture medium. They belonged to the O4 E. coli serogroup and expressed multiple antibiotic resistance.

PER-1 extended-spectrum beta-lactamase-producing Pseudomonas aeruginosa clinical isolates from Budapest, Hungary, and Belgrade, Serbia, were characterized by molecular methods. Two PER-1-positive isolates were recovered from sporadic cases in Budapest and a small cluster of PER-1-positive infections involving four patients were identified at a Belgrade hospital. A class 1 integron harbouring a bla(OXA-2)beta-lactamase gene and four other gene cassettes was detected in both the Budapest and the Belgrade isolates. The two P. aeruginosa isolates from Budapest also carried another class 1 integron containing bla(OXA-74), aac(6')-Ib-cr and cmlA7 genes in its variable region. The aac(6')-Ib-cr fluoroquinolone-acetylating aminoglycoside acetyltransferase gene is described here for the first time in P. aeruginosa. Multilocus sequence typing (MLST) revealed that the PER-1 positive P. aeruginosa isolates identified in this study display ST235, a sequence type that belongs to clonal complex CC11. Two bla(PER-1)-positive P. aeruginosa reference isolates from France and Belgium could also be assigned to complex CC11 by MLST. Our results underscore the role of complex CC11 in the dissemination of bla(PER-1) among P. aeruginosa clinical isolates.

Endemic Balkan nephropathy (EBN) is a kidney disease of unknown etiology limited to Bulgaria, Rumania and former Yugoslavia. Primary kidney tissue cultures were established as explants from tissue obtained at operations from 5 EBN patients with urinary tract tumors. Four out of the five biopsy specimens on extended culture incubation at 33°C yielded a coronavirus virus (EBNV) which was cytopathogenic for human fibroblast and Vero cells. In cells inoculated with EBNV, cytoplasmic immunofluorescence was found using antisera for human coronaviruses OC43 and 229E as well as the porcine transmissible gastroenteric virus and avian (chicken) bronchitis virus. In neutralization tests, EBNV failed to react with antisera to these viruses. Using hyperimmune serum raised with EBNV, positive cytoplasmic immunofluorescence was seen with cells infected with OC43, 229E, TGV and significantly with the kidney tissue of the biopsy specimens from the EBN patients. A screen for neutralizing antibody using the EBN virus revealed that 87.2% of EBN patients on dialysis were positive, also 74% of people from an endemic area were also positive, while only 13.5% from outside were positive. It is suggested that a coronavirus is involved in the etiology of the disease and that humans are an incidental host of a coronavirus zoonosis.

The genetic context of the bla NDM-1 gene in the genome of Pseudomonas aeruginosa MMA83 was investigated. Sequencing of the cosmid selected for the bla NDM-1 gene revealed the presence of two bla NDM-1 copies in the genome of P. aeruginosa MMA83 in a unique genetic environment. Additionally, mating assays, DNA-DNA hybridization, and an S1 nuclease assay strongly suggest that the bla NDM-1 gene in P. aeruginosa MMA83 is chromosome borne.T he genetic context of the bla NDM-1 gene has been extensively investigated, mostly among members of the family Enterobacteriaceae (1,2,3,4,5,6,7,8). From a genetic perspective, the key finding is that the bla NDM-1 gene can be located on plasmids belonging to different incompatibility groups. Although bla NDM-1 genes have the same flanking regions, in a broader sequence context, they reside in diverse genetic environments (6). Although the bla NDM-1 gene is typically found in a plasmid, in certain Escherichia coli and Providencia stuartii isolates, it was chromosome borne (6). The bla NDM-1 gene is considered endemic to the Balkan region, and this gene has been found not only in Enterobacteriaceae (9) but also in Pseudomonas aeruginosa (10). Considering the role of P. aeruginosa in the development of nosocomial infections worldwide and the subsequent complicated clinical management of patients infected with this pathogen, the emergence of NDM-1-positive strains is alarming. This is why the analysis of the genetic context of the bla NDM-1 gene, not only in Enterobacteriaceae but also in all other NDM-1-positive isolates, is crucial and could reveal associations with other genes that confer antimicrobial resistance and potential routes of dissemination. The objective of our study was to identify the genetic location and genetic context of the bla NDM-1 gene in P. aeruginosa MMA83, one of the NDM-1-positive clinical isolates from Serbia.To determine the genetic location of the bla NDM-1 gene in P. aeruginosa MMA83, transformation, mating assays, and DNA-DNA hybridization were performed. Heat shock transformation of E. coli DH5␣ was performed with total DNA isolated from P. aeruginosa MMA83 (selection based on 100 g/ml ampicillin). Triparental mating assays with P. aeruginosa MMA83 as a donor, E. coli(pRK2013) as a helper, and two recipients, E. coli DH5␣ (selection based on 100 g/ml ampicillin) and an imipenem-sensitive isolate of Pseudomonas mendocina (selection based on 32 g/ml imipenem), did not result in any transconjugant (11). Counterselection for recipients was based on glycerol for P. mendocina and sucrose for E. coli DH5␣ as the sole carbon sources. This result suggested that the gene(s) conferring resistance to ␤-lactams was not present on a conjugative plasmid(s) in MMA83. To find out if MMA83 is a plasmid-free strain, total DNA was isolated in agarose blocks and treated with S1 nuclease, which is used to visualize large, supercoiled plasmids (12). The results obtained confirmed that MMA83 does not carry any plasmid (see Fig. S1 in the supplemental material). Also, pulse...

We have established the prevalence of all three genospecies in the city of Belgrade. Bb sensu lato was found, with the dominance of B. afzelii.

From the Central-East European region the first VIM metallo-beta-lactamase (MBL) producing Pseudomonas aeruginosa strains were published from Croatia, Poland and Hungary. The aim of this study was to assess the contribution of MBL-production to carbapenem-resistance among P. aeruginosa clinical isolates in the Military Medical Academy (MMA) in Belgrade, Serbia between August 2004 and September 2007. Only one P. aeruginosa isolate with strain number 722 proved MBL-positive that harboured a novel class 1 integron with a bla(VIM-2)-like cassette in the first position, followed by orfD, a putative gene with unknown function. Our data indicate that MBL-producing strains occur at a prevalence of less than 1% among imipenem-nonsusceptible P. aeruginosa clinical isolates in this Belgrade hospital. The newly identified VIM MBL-producing P. aeruginosa strain 722 could be assigned to serotype O11, and it was panresistant to all antimicrobials tested. The isolate displayed sequence type ST235 by multilocus sequence typing which is the founder sequence type of the previously identified international clonal complex CC11 that already contains bla(VIM)-positive isolates from Italy, Greece, Sweden, Hungary and Poland. In conclusion, this is the first report of VIM MBL-producing P. aeruginosa from Serbia and also of the occurrence of such isolates belonging to the international clonal complex CC11 in this country.

Summary:Lyme disease is a tick borne zoonotic infection, caused by Borrelia burgdorferi s.l. bacteria. For the transmission of the disease, the presence of ticks is a prerequisite. Lyme borreliosis mostly occurs in people and dogs, but it may occur in other animals. Ticks which carry B. burgdorferi s.l. in Serbia are of the Ixodes ricinus specis. In Serbia, Lyme disease was detected for the first time in the late '80-es. In dogs, clinical symptoms may occur even months after a tick bite, and include weakness, lymphadenopathy, fever, lameness, arthritis, etc. In our survey, we have observed tick and dog populations in the province of Vojvodina (northern part of Serbia). I. ricinus ticks were collected and examined for the presence of B. burgdorferi s.l. in several chosen locations. In addition, blood samples were collected from house dogs and pets from the same locations, and analyzed for the presence of antibodies specific for B. burgdorferi s.l. The results showed a mean infection of ticks of 22.12 %, and a mean seroprevalence of Lyme disease in dogs of 25.81 %. We conclude that in Vojvodina there is an actual risk of Lyme borreliosis for other animals and humans, because of the persistence of B. burgdorferi s.l. in both tick and dog populations.