Occurrence of genes encoding aminoglycoside-modifying enzymes in Escherichia coli isolates from chicken meat.

1. The aim of the experiment was to determine the occurrence of genes encoding aminoglycoside-modifying enzymes (AMEs) in Escherichia coli isolates recovered from chicken meat.2. Antibiotic sensitivity was tested using the disc diffusion test. AMEs and virulence profile were determined by PCR/sequencing.3. Out of 195 meat samples collected, 185 (95%) isolates were identified as E. coli. Disc diffusion showed a resistance value of 22% (n = 42) for at least one of the antibiotic aminoglycosides (AGs) tested (tobramycin, gentamycin, amikacin and kanamycin). PCR screening showed the presence of three classes of AMEs, namely, aac(3)-II (12%), aac(6')-Ib (7%) and aac(2')-Ia (5%). Eight of the 42 isolates were positive for the stx1 and sxt2 genes and were defined as Shiga toxin-producing E coli., while the eae gene was positive in one strain. Among the 42 isolates, group A was the predominant phylogenetic identified (76%), followed by group D (21%). One isolate belonged to subgroup B23.4. The results suggested that chicken meat could be an important reservoir of AMEs, and pose a potential risk by dissemination of resistance to humans through the food chain.

Klebsiella pneumoniae resistant to third-generation cephalosporins in five African and two Vietnamese major towns: multiclonal population structure with two major international clonal groups, CG15 and CG258.

The molecular epidemiology of third-generation cephalosporin-resistant (3GC-R) Klebsiella pneumoniae in developing countries is poorly documented. From February 2007 to March 2008, we collected 135 3GC-R K. pneumoniae isolates from seven major towns in Maghreb (Morocco), West Africa (Senegal, Côte d'Ivoire), Central Africa (Cameroon), East Africa (Madagascar) and Southeast Asia (Vietnam). Their genetic diversity, assessed by multilocus sequence typing, was high (60 sequence types), reflecting multiclonality. However, two major clonal groups, CG15 (n = 23, 17% of isolates) and CG258 (n = 18, 13%), were detected in almost all participating centres. The two major clonal groups have previously been described in other parts of the world, indicating their global spread. The high diversity of enterobacterial repetitive intergenic consensus sequence-PCR banding patterns at the local level indicates that most isolates were epidemiologically unrelated. The isolates were characterized by the presence of multiple resistance determinants, most notably the concomitant presence of the aac(6')-Ib-cr, qnr and blaCTX-M-15 genes in 61 isolates (45%) belonging to 31 sequence types. These isolates were detected across a large geographical area including Cameroon (n = 1), Vietnam (n = 4), Madagascar (n = 10), Côte d'Ivoire (n = 12), Morocco (n = 13) and Senegal (n = 21). These results have major implications for patient management and highlight a potential reservoir for resistance determinants.

Prevalence and characterization of extended spectrum ß-lactamases in Klebsiella-Enterobacter-Serratia group bacteria, in Algeria.

OBJECTIVES: The authors had for aim to assess the local epidemiology, antibiotic resistance, and molecular typing of expanded spectrum betalactamase producing Klebsiella, Enterobacter, and Serratia (ESBL KES). MATERIALS AND METHODS: Two hundred and seven strains of the KES group were isolated in the microbiology laboratory of the Annaba Ibn Rochd hospital in 2009. The antibiotic resistance (diffusion method and MIC) was tested and ESBL detection was performed as recommended by the Clinical Laboratory Standard Institute (CLSI). The characterization of genes for resistance to ß-lactams (CTX-M-1, TEM, and SHV) and AmpC cephalosporinase (DHA-1) was performed by polymerase chain reaction. The epidemiological relationship among identified strains was analyzed by Pulsed Field Gel Electrophoresis (PFGE). Genetic transfers were performed by conjugation using sodium azide resistant Escherichia coli K(12)J(5) as recipient strain. RESULTS: The overall incidence of ESBL KES was 31.4% (65/207) distributed as follows: 17.4% of Klebsiella spp., 7.2% Enterobacter spp., and 6.8% Serratia marcescens. The ß-lactamase CTX-M 1 types were predominant (88%), followed by TEM (36.5%), and SHV (31.1%). Twenty-three strains expressed at least two bla genes. DHA-1 type cephalosporinase was found in 4 E. cloacae associated with CTX-M-1. Several epidemic clones were determined. Conjugation experiments showed that bla(CTX-M), bla(TEM), and bla(SHV) were carried by conjugative plasmids of high molecular weight (≥125kb). CONCLUSIONS: This study revealed a high frequency of ESBL KES with a predominance of CTX-M-1. This high rate of ESBLs could be due to a clonal spread and the emergence of new epidemic clones.

[Qnr and aac (6')-Ib-cr types quinolone resistance among Enterobacteriaceae isolated in Annaba, Algeria]. / Résistance aux quinolones de types qnr, aac (6')-Ib-cr chez les entérobactéries isolées à Annaba en Algérie.

AIM OF THE STUDY: To show the emergence of the qnr and aac (6')-Ib-cr genes in nalidixic acid resistant enterobacteria isolated at Annaba city in Algeria. MATERIALS AND METHODS: Enterobacterial strains (n=25) resistant to nalidixic acid have been isolated at the microbiology laboratory of the Annaba city hospital in Algeria Antibiotic susceptibility (disc diffusion method and MIC) and screening for Extended Spectrum Beta-Lactamase (ESBL) were performed according to the French Society for Microbiology guidelines. Characterization of quinolone resistance genes (qnrA, qnrB, qnrS, aac(6')-Ib-cr) was investigated by PCR. Identification of ESBL (TEM, SHV, CTX-M1, CTX-M2, CTX-M8 and CTX-M9 groups) was performed by PCR. Identification of plasmid AmpC beta-lactamases was performed by multiplex PCR. All PCR products were sequenced on both strands. Conjugation experiments were performed using azide-resistant Escherichia coli K12J5 as a recipient strain. RESULTS: Among the 25 strains selected, 24 were resistant to at least four antibiotics. Six strains showed an ESBL phenotype. The qnr gene (B1 type) was found in two ESBL producing strains with at least two types of bla gene. The aac (6')-Ib gene was detected in three strains, one with the aac (6') Ib-cr variant. With specific primers, we have shown that qnrB1, CTX-M-28, TEM1, aac (6')-Ib-cr was cotransferred together and that these genes are carried by conjugative plasmids of high molecular weight. CONCLUSION: The emergence of combination of resistance genes may pose a public health problem. Thus, a policy of surveillance of resistance seems necessary.

Rapid detection of Listeria monocytogenes in food by polymerase chain reaction.

The standard conventional methods for the detection of Listeria monocytogenes in foods require high time 7 to 10 days to give ready results. To dissolve this problem we have evaluate a short method using Polymerase Chain Reaction (PCR) to analyze food samples. In parallel with this study, a comparison was made between PCR amplification from templates directly prepared from food and the official standard ISO procedure 11290-1. In this study we have used a Half Frazer broth as an enrichment medium; there were positive results of PCR detection of L. monocytogenes in different food sample analyzed (milk, cheese and meat) with approximately 1.5 10(1) Colony Forming Units /25 g in less than 36 h. This PCR procedure has proved to be rapid and sensitive method suitable for the routine analysis; firstly, because this assay required just a short pre-enrichment step before PCR. Secondly, this procedure is very simple and time-saving; it could take less than one working day to obtain results if initial microbiological load was very important.

Prevalence and Antibiotic-Resistance of Salmonella Isolated from Food in Morocco

Background: Salmonellosis remains one of the most frequent food-borne diseases worldwide; especially in developing countries. The emergence of antimicrobial resistance in Salmonella isolates from food can potentially compromise the treatment of these infections. This investigation was conducted for the first time in Morocco both to detect the occurrence of Salmonella in foods as well as to determine the antibiotic resistance profile of the Salmonella isolates. Methodology: In total; 11;516 food samples collected from 2002 to 2005 were investigated. Isolated Salmonella were characterized by serotyping and susceptibilities were determined for 15 antimicrobial drugs using the disc diffusion assay. Results: The overall percentage of Salmonella prevalence (n=105) was 0.91with rates of 71for slaughterhouses and 9for seafood. Sixteen different serotypes were identified among 104 Salmonella enterica isolates including serotypes Infantis (n=25); Bredeney (n=13); Blokley (n=11); Typhimurium (n=9); Mbandaka (n=8); Branderup II (n=7); and Kiambu (n=6); 1 isolate of Salmonella enterica belonged to subspecies II salamae. Twenty-nine percent of isolates (n=30/105) were resistant to at least one antimicrobial. Resistance to tetracycline was the most common finding (21); followed by resistance to ampicillin (13); amoxicillin+clavulanic acid (9); streptomycin (7); chloramphenicol (4) and nalidixic acid (3;8). None of the isolates was resistant to 3rd-cephalosporin and fluoroquinolones (i.e. ciprofloxacin). Multidrug resistance (MDR) was seen in 9.5of the isolates; mainly in S. Typhimurium DT104 with R-type ACSSuT and S. Hadar. Conclusions: Despite a low frequency of Salmonella isolation; S. Typhimurium DT104 was identified in the first step of the food chain. The study points out the need control antibiotic resistance in Salmonella isolated from food in Morocco to avoid the spread of MDR

Prevalence and Antibiotic-Resistance of Salmonella Isolated from Food in Morocco

Background: Salmonellosis remains one of the most frequent food-borne diseases worldwide; especially in developing countries. The emergence of antimicrobial resistance in Salmonella isolates from food can potentially compromise the treatment of these infections. This investigation was conducted for the first time in Morocco both to detect the occurrence of Salmonella in foods as well as to determine the antibiotic resistance profile of the Salmonella isolates. Methodology: In total; 11;516 food samples collected from 2002 to 2005 were investigated. Isolated Salmonella were characterized by serotyping and susceptibilities were determined for 15 antimicrobial drugs using the disc diffusion assay. Results: The overall percentage of Salmonella prevalence (n=105) was 0.91with rates of 71for slaughterhouses and 9for seafood. Sixteen different serotypes were identified among 104 Salmonella enterica isolates including serotypes Infantis (n=25); Bredeney (n=13); Blokley (n=11); Typhimurium (n=9); Mbandaka (n=8); Branderup II (n=7); and Kiambu (n=6); 1 isolate of Salmonella enterica belonged to subspecies II salamae. Twenty-nine percent of isolates (n=30/105) were resistant to at least one antimicrobial. Resistance to tetracycline was the most common finding (21); followed by resistance to ampicillin (13); amoxicillin+clavulanic acid (9); streptomycin (7); chloramphenicol (4) and nalidixic acid (3;8). None of the isolates was resistant to 3rd-cephalosporin and fluoroquinolones (i.e. ciprofloxacin). Multidrug resistance (MDR) was seen in 9.5of the isolates; mainly in S.. Typhimurium DT104 with R-type ACSSuT and S. Hadar. Conclusions: Despite a low frequency of Salmonella isolation; S. Typhimurium DT104 was identified in the first step of the food chain. The study points out the need control antibiotic resistance in Salmonella isolated from food in Morocco to avoid the spread of MDR

Study of the RET gene and his implication in thyroid cancer: Morocco case family.

BACKGROUND: Multiple endocrine neoplasia type 2A (MEN 2A) is an autosomal dominant inherited cancer syndrome that affects multiple tissues derived from the neural crest. Inheritance of MTC is related to the presence of specific mutations in the RET proto-oncogene. Almost all mutations in MEN 2A involve one of the cysteines in the extracellular domain of the RET receptor. AIMS: The objective of the present study was the biochemical and molecular characterization of the first Moroccan clinically established MEN 2A patient and at-risk family members. SETTINGS AND DESIGN: This is a study on a family presented with MTC referred to our institute in 2004. MATERIALS AND METHODS: Peripheral blood leukocyte DNA samples were isolated and amplified by polymerase chain reaction followed by restriction enzyme analysis and DNA sequencing. RESULTS: We identified a heterozygous germ line missense mutation at codon 634 of exon 11 in the RET gene that causes a cysteine to arginine amino acid substitution in the DNA of the proband; this mutation was not found in the DNA of the parents or relatives. CONCLUSIONS: The detection of mutated MEN 2A gene carriers enables us to differentiate high-risk members from those who bear the wild-type gene. Occasionally, application of RET proto-oncogene testing may lead to the detection of unexpected de novo mutation that could be transmitted to children.

Treatment evaluation, follow-up and familial screening of medullary thyroid carcinoma by highly specific calcitonin measurements.

BACKGROUND: Calcitonin is the most sensitive and specific marker for medullary thyroid carcinoma (MTC). AIMS: The aim of this study was to emphasize the role and the limits of plasma basal calcitonin (bCT) measurement in the management of Moroccan MTC patients and their relatives. SETTINGS AND DESIGN: This is a retrospective study on 6 MTC patients referred to our institute from January 1996 to December 2004. MATERIALS AND METHODS: Serum bCT levels were measured in 36 individuals comprising six known MTC cases, 18 relatives and 12 healthy volunteers, using two-sites immunoradiometric assay method. Five of MTC patients have been followed from 12 to 96 months after surgery. STATISTICAL ANALYSIS USED: Calculations were performed using SPSS 10.0 program. Data comparison was done by Student's t -test. RESULTS: The circulating preoperative bCT concentrations were elevated for all MTC patients (range, 44,8 -2055 pg/ml, normal <10). Recent postoperative bCT determinations varied from 24.4 to 1972 pg/ml in four patients. In one patient, the bCT value decreased to an undetectable level during a follow-up of 12 months. The mean bCT level of relatives was 4.90 +/- 3.54 pg/ml; two patients had slightly elevated bCT. Five (42%) healthy volunteers had undetectable bCT levels and all had less than 10 pg/ml; the mean bCT value was 3.06 +/- 2.51 pg/ml. CONCLUSIONS: Routine plasma bCT measurement still has an important place in the preoperative diagnosis and follow-up treatment of MTC.

cut-1-like genes of Ascaris lumbricoides.

Three genomic fragments homologous to cut-1 of Caenorhabditis elegans (C. elegans) have been identified in the intestinal parasitic nematode Ascaris lumbricoides (A. lumbricoides). Two of these fragments identify one region of the A. lumbricoides genome; they are separated by 8-9 kb and have opposite orientation, with the direction of transcription converging toward the center of the region. The third gene, which has been studied more completely, is in a different region of the genome separated from the first one by not less than 12-15 kb. The complete genomic sequence of this third gene has been determined. cDNA overlapping clones were obtained from adult A. lumbricoides RNA via the rapid amplification of cDNA ends (RACE) procedure [Frohman et al., 1988. Rapid production of full-length cDNAs from rare transcripts: amplification using a single gene-specific oligonucleotide primer. Proc. Natl. Acad. Sci. USA 85, 8998-9002] and sequenced. The mature mRNA of this gene, which we have named ascut-1, is trans-spliced to the spliced leader sequence of nematodes (SL1) [Krause, M., Hirsh, D., 1987. A trans-spliced leader sequence on actin mRNA in C. elegans. Cell 49, 753-761]. The mRNA is 1684 nt long plus the poly(A) tail and contains four exons with a 138 nt untranslated 5' leader and a 388 nt untranslated 3' tail. Conceptual translation of the coding sequence shows a protein of 385 aa with a signal peptide of 16 aa. The protein shows very high homology with CECUT-1, the product of the C. elegans gene cut-1 and with other cuticlin proteins of nematodes. A 262 amino acids region which is strongly conserved between these proteins seems to identify a group of proteins, so far restricted to nematodes, for which the name CUT-1-like is proposed.