Detection of Diarrheagenic Escherichia coli in pet animals and its antibiotic resistance in Alexandria governorate

Benefits of having a dog or cat as a pet varies between owners according to the purpose, however, the limit between benefit and harm is sensitive because close contact between pets and humans may involuntarily represent harm for humans. Doga and cats have been proposed as possible reservoir of virulent Escherichia coli strains that may cause enteric and extra-intestinal infections in humans. In this study, we aimed to detect diarrheagenic Escherichia coli [DEC] in dogs and cats and their antibiotic resistant pattern[s]. Samples were collected from 70 dogs and cats from different veterinary clinics and hospitals in Alexandria. These animals suffered from diarrhea and other symptoms as fever, nausea, vomiting, chills, loss of appetite, muscle aches and bloating. Forty E. coli isolates were detected by culturing and biochemical tests, and were subjected to antimicrobial disc diffusion susceptibility test by using 10 different antibiotic discs, which are the most commonly used in pet animal clinics. Antibiotic resistance for individual antibiotics ranged from 5 to 98% with multiple resistances to 2 or more antibiotics detected in 15 [21%] samples. PCR for detection of virulent genes of E. coli; VT2e and eaeA genes as well as the antibiotic resistance blaTEM gene was performed. The VT2e and eaeA genes were found in E. coli isolates, from dogs and cats. These results collectively indicate that pet animals can harbor the Enteropathogenic [EPEC] and Enterotoxigenic Escherichia coli [ETEC] causing diarrhea at different ages with possible active transmission to contact human. Further, the high and multiple antibiotic resistance level can pose therapeutic challenges in contact humans. It is fundamental that veterinarians recommend preventive measures to pet owners towards the establishment of a long-term preventive programme against antibiotic resistant E. coli

Comparison of common detection methods for chlamydia trachomatis and genital mycoplasmas in cervical infection

Genitourinary infections are caused by a large number of diverse microbial agents, in many women with cervicitis, these agents are not detected, even when highly sensitive diagnostic tests are performed. Less urbanized communities have a proportionally higher incidence of urethritis and cervicitis, caused by Chlamydia trachomatic [C. trachomatis]. In addition, mycoplasmas commonly colonize the genital tracts of men and women, and the ability of some species to cause non-gonococcal urethritis and cervicitis has been well established. To detect the prevalence of C. trachomatis, M. hominis and U. urealyticum in cases of cervicitis and to compare different laboratory methods for the diagnosis of these organisms. A total of 50 women suffering from cervicitis were enrolled in this work. To determine the causative agents, PCR, detection of mycoplasmas [Using Mycoplasma IST2 kit] and direct immunofluorescence assay for Chlamydia trachomatis were used. U. urealyticum was isolated from 18 cases [36%], while M. hominis was isolated only from 2 cases [4%]. Besides, 10 cases [20%] were infected by both organisms. U. urealyticum was detected in 5 cases [10%] by multiplex PCR and in 8 cases [16%] by monoplex PCR. M. hominis was detected in 10 cases [20%] both by multiplex as well as by monoplex PCR. C. trachomatis was detected in 3 samples with counts of 16.760, 28.2 and 10 DNA copies/ml using Real Time PCR. However, it was not detected by direct immunofluorescence assay in any of the samples. Mycoplasma IST2 culture was more sensitive for the detection of genital mycoplasmas and Real Time PCR was a better diagnostic test for the detection of C. trachomatis

Evaluation of 3 PCR techniques for detection of brucella DNA in peripheral human blood

Brucellosis is a widespread zoonosis transmittable to humans. Currently the diagnosis of this zoonosis is based on microbiological and serological laboratory tests. PCR has been used to detect DNA from Brucella. Different target genes, primer pairs, PCR techniques and extraction procedures have been previously published for Brucella detection. But only a few of these primers have been used in human samples, and only study has been carried out to compare sensitivity between them. In the present study, 3 sets of primers and 3 different PCR protocols amplifying 3 different regions of the Brucella genome were compared for detection of Brucella DNA in a peripheral-blood PCR assay to conclude which is most suitable for the clinical diagnostic laboratory. These three pairs of primers amplify three different fragments included in: i] a gene encoding a 31-kDa B. abortus antigen [B4/B5], ii] a sequence 16S rRNA of B. abortus [F4/R2], and [iii] a gene encoding an outer membrane protein [omp-2] [JPF/JPR]. Some modifications on the reported techniques were applied during the present work to improve the outcome. Results showed that B4/B5 primer pair had the highest sensitivity for detection of positive samples [98%], JPF/JPR primer pair detected 88.4% of positive samples while F4/R2 primer pair was the least sensitive being able to detect only 53.1% of positive samples. The specificity of the 3 techniques was 100%. B4/B5 primers were also able to detect the smallest number of bacteria [700 cfu/ml] while JPF/JPR were able to detect 7x10[5] cfu/ml and F4/R2 primers were able to detect 7x10[7] cfu/ml. It is thus concluded that using the B4/B5 primer PCR with the suggested modifications is a robust assay, which meets the sensitivity requirements to be used for testing of human blood samples for brucellosis in the diagnostic laboratory