Carriage of CTX-M-15-Producing Escherichia coli Isolates among Children Living in a Remote Village in Senegal

Two out of 20 children with no known antibiotic exposure, living in a very remote Senegalese village, were found to be fecal carriers of a multiresistant Escherichia coli clone that produced CTX-M-15. This highlights the current massive spread of extended-spectrum β-lactamases, even in isolated communities.CTX-M-type β-lactamases have become the most frequently isolated extended-spectrum β-lactamases (ESBL) in Enterobacteriaceae (4). There seems to be no limit to their spread through the feces of healthy individuals from urban areas. Thus, they have been frequently isolated in Spain (15), Lebanon (16), Hong Kong (10), Bolivia, and Peru (17, 19) with various prevalences. So far, however, remote-living subjects appear to have been spared, at least in Amazonia (1, 9, 18). Here, we investigated the spread of ESBL in West Africa. Working in Senegal, we searched the most remote and isolated village we could find and assessed the fecal carriage of ESBL-producing Enterobacteriaceae in children who had in all probability never taken antibiotics.Kagnoube, the village in eastern Senegal where the sampling took place, was chosen by local Senegalese investigators because it was very remotely situated (almost unreachable during the rainy season, not served by any concrete road). It comprises about 60 inhabitants living in traditional huts. A shared water pit is used as the source of water, and no river is flowing close. The closest permanent health care facility is 100 km away. The Kagnoube inhabitants reported having taken allopathic drugs only very occasionally. We included 20 healthy children in the study (11 girls and 9 boys aged 1 to 11 years [mean age, 6.9]) with the agreement of their parents, who firmly stated that their children had never taken any Western drug. According to the local legislation and considering the passive nature of the sampling, no approval by an ethical committee was required. The children provided a fresh stool sample, an aliquot of which was immediately inoculated into conservation agar in screw-cap tubes (Bio-Rad, Marne-la-Coquette, France) and sent to France at room temperature for harvesting. There, the presence of Enterobacteriaceae resistant to extended-spectrum cephalosporins (ESC) was investigated as follows: (i) in the predominant flora, by testing the antibiotic susceptibility of five Escherichia coli strains randomly chosen after inoculation on Drigalski agar using the disc diffusion method, and (ii) in the subdominant flora, by inoculating ChromID ESBL agar plates (bioMérieux, Marcy l''Etoile, France). ESBL production was confirmed by the double-disk synergy test (11). DNA was extracted using a MagNA Pure LC instrument (Roche Molecular Biochemicals, Mannheim, Germany). Clonality was assessed by repetitive extragenic palindromic-PCR as described previously (8). A plasmid transfer assay was attempted by bacterial mating in liquid broth by using rifampin-resistant E. coli J53 as the recipient. MICs were determined using Etest strips (AES, Solna, Sweden). When necessary, bla_TEM, bla_SHV, bla_CTX-M (five groups), bla_VEB, bla_PER, bla_GES, bla_OXA-1, aac(6′)-Ib, qnrA, qnrB, qnrS, ISEcp1, and integrase-encoding intI, intII, and intIII genes were detected by PCR, as previously described (5, 8, 20). Plasmids were extracted with the QIAprep Spin Miniprep kit (Qiagen, Courtaboeuf, France). Phylogenetic groups were determined by triplex PCR (7). The replicon typing of plasmids was performed by multiplex PCR (6, 14). Eventually, a multidrug resistance (MDR) region similar to that described for plasmid pC15-1a (2) was investigated by PCR, using E. coli strain TN03 as a positive control (13).Whereas none of the five randomly chosen E. coli isolates per sample (predominant flora) displayed an antibiotic resistance pattern suggestive of ESBL production, stool sample plating on ChromID ESBL agar plates (subdominant flora) yielded one and four cefotaxime-resistant E. coli isolates for two children. The five isolates exhibited identical repetitive extragenic palindromic-PCR patterns (data not shown). Thus, one isolate from each of the two children (named KA12 and KA20) was further tested and found to be resistant to co-amoxiclav, cefotaxime, ceftazidime, fluoroquinolones, kanamycin, cotrimoxazole, and tetracycline but susceptible to cefoxitin, ertapenem, imipenem, gentamicin, and tigecycline. Genes bla_CTX-M-15 (with insertion sequence ISEcp1 immediately upstream), bla_TEM-1, bla_OXA-1, aac(6′)-Ib-cr, and tet(A) were present, but qnr was not detected. Both strains belonged to phylogenetic group A subgroup A1 (3), were intI positive (sequencing identified cassettes dfrA17 and aadA5), and transferred resistance to ESC, aminoglycosides, co-amoxiclav, tetracycline, and trimethoprim but not to sulfonamides. PCR experiments confirmed the presence of bla_CTX-M-15, bla_OXA-1, bla_TEM-1, aac(6′)-Ib-cr, and tet(A) in the transconjugants. Although four plasmids were detected in both parental strains, transconjugants had only one carrying an FIA-FIB-FII multireplicon. For both plasmids, PCR-based MDR comparison to pC15-1a revealed the presence of the pemK-Tn5403 and Tn5403-Tn1721-like junctions but not junctions 3 and 4, which is consistent with the absence of the aac(3)-II gene in our plasmids (Fig. ​(Fig.1).1). Long-PCR analysis confirmed that bla_CTX-M-15 was located just upstream from aac(6′)-Ib-cr. No signal was obtained for junctions 5 and 6.Open in a separate windowFIG. 1.Schematic representation of the MDR region of pC15-1a (2) (GenBank accession number {"type":"entrez-nucleotide","attrs":{"text":"AY458016","term_id":"38606064","term_text":"AY458016"}}AY458016) and the MDR region of the Senegalese clone (strains KA12 and KA20) as deduced from PCR experiments. Unfilled double-headed arrows indicate PCRs performed for pemK, tet(A), bla_OXA-1, aac(6′)-Ib-cr, bla_TEM-1, and bla_CTX-M-15. Black double-headed arrows indicate junction PCR assays, conducted as described by Lavollay et al. (13). J1, pemK-tnpA (Tn5403); J2, tnpR (Tn5403)-tnpA (Tn1721-like); J3, catB3-aac(3)-II; J4, aac(3)-II-orfB′; J5, orfB′-tnpA (Tn3); J6, ISEcp1-bla_TEM-1. Junctions between bla_CTX-M-15 and ISEcp1 are also indicated by a black double-headed arrow. Long-PCR (L-PCR) experiments (data not shown) were performed to confirm the location of bla_CTX-M-15 close to aac(6′)-Ib-cr and bla_OXA-1. bla_TEM-1 could not be located by long PCR (data not shown).We found that an E. coli clone that carries CTX-M-15 and other resistant traits, including OXA-1, TEM-1, AAC(6′)-Ib-cr, and Tet(A), was present in the subdominant fecal flora of two healthy children from a very remote and isolated Senegalese village with very limited access to allopathic medicine. This association of the resistance determinants of the strains detected was very similar to that found in ESBL-producing E. coli that circulates worldwide in dense urban areas (2, 12, 13). Pallecchi et al. had observed a rise in CTX-M β-lactamases among children living in Bolivian and Peruvian suburban areas, with fecal carriage rates of 0.1% and 1.7% in 2002 and 2005, respectively (17, 19), highlighting the recent spread of CTX-M genes among healthy children. Interestingly, typeable bla_CTX-M-15-carrying plasmids from Peru and Bolivia also conveyed aac(6′)-Ib-cr and displayed FIA-FIB-FII replicons (17). Indeed, multireplicon FIA-FIB-FII has already been observed in CTX-M-15-producing strains isolated in France (14). The FII replicon is present on plasmid R100, and pC15-Ia harboring CTX-M-15 was derived from this plasmid through the incorporation of the MDR region (2). Note that even though bla_CTX-M-15-harboring plasmids from the remote Senegalese village carried also FIA- and FIB-type replicons, their MDR regions were similar to that of plasmid pC15-1a, except that they lacked aac(3)-II. Homologies between the MDR regions of pC15-1a and other CTX-M-15 plasmids have already been observed (2, 13). Here, junction PCR analysis suggested that there were many similarities between the plasmid-borne MDR structures harbored by these strains and those harbored by the TN08, TN36, and EpLA2 strains previously isolated in France (13). This strongly implies that even in the absence of direct antibiotic exposure, the few contacts the inhabitants of Kagnoube had with the outside world and allopathic medicine were enough to allow the CTX-M-15-associated MDR gene machinery to settle and persist in their commensal flora. Thereby, this study stresses the difficulties to be expected in controlling the dissemination of CTX-M-mediated resistance.     

A haplotype of the human CXCR1 gene protective against rapid disease progression in HIV-1+ patients

Chemokines and their receptors are key factors in the onset and progression of AIDS. Among them, accumulating evidence strongly indicates the involvement of IL-8 and its receptors, CXCR1 and CXCR2, in AIDS-related conditions. Through extensive investigation of genetic variations of the human CXCR1-CXCR2 locus, we identified a haplotype of the CXCR1 gene (CXCR1-Ha) carrying two nonsynonymous single nucleotide polymorphisms, CXCR1_300 (Met to Arg) in the N terminus extracellular domain and CXCR1_142 (Arg to Cys) in the C terminus intracellular domain. Transfection experiments with CXCR1 cDNAs corresponding to the CXCR1-Ha and the alternative CXCR1-HA haplotype showed reduced expression of CD4 and CXCR4 in CXCR1-Ha cells in human osteosarcoma cells as well as in Jurkat and CEM human T lymphocytes. Furthermore, the efficiency of X4-tropic HIV-1(NL4-3) infection was significantly lower in CXCR1-Ha cells than in CXCR1-HA cells. The results were further confirmed by a series of experiments using six HIV-1 clinical isolates from AIDS patients. A genetic association study was performed by using an HIV-1(+) patient cohort consisting of two subpopulations of AIDS with extreme phenotypes of rapid and slow progression of the disease. The frequency of the CXCR1-Ha allele is markedly less frequent in patients with rapid disease onset than those with slow progression (P = 0.0003). These results provide strong evidence of a protective role of the CXCR1-Ha allele on disease progression in AIDS, probably acting through modulation of CD4 and CXCR4 expression.     

Access to medicines in Senegal: results of a sample survey

The survey was conducted in Senegal in April 2001 on a representative sample of providers and clients. Results show that access to medicines in Senegal was limited for three main reasons: (1) the supply of drugs was inadequate, and even critical drugs were often missing in health centres, and were somewhat less in pharmacies; (2) the health infrastructures appeared insufficient to cover the needs of the whole population, creating high opportunity costs; (3) the cost of the drugs prescribed was higher than the minimum price, sometimes exceeding the capacity of poorer people, although high cost was seldom reported as the main reason for not acquiring prescribed drugs. Improving access to medicine is a priority to help reduce health inequalities in developing countries.     

Malaria morbidity in Barkedji, village of Ferlo, in Senegal Sahelian area

A study of malaria morbidity was carried out from November 1994 to October 1995, in a Ferlo village (Barkedji) characterized by a long persistence of the temporary ponds. The objective was to evaluate the repercussions of the strong and long anopheles transmission in humans. A clinical follow-up of a group of residents was conducted at home every 10 days by an investigator trained for taking axillary temperature and making thick smears, when suspecting malaria. Were included in the group, 123 voluntary subjects among whom 50% were children under 10 years old. Any feverish subject (T degree >37 degrees 5) or subject presenting other malaria symptoms (headaches, hot body shivers, sweats, aches...) was regarded as having a malaria attack as well as a parasitemia >2500 P/mm3 in children aged of 0 to 14 years old and 1000 P/mm3 in the oldest. During the study subjects with at least one feverish access, plasmodium infection and malaria attack were 58%, 33% and 22%, respectively. On 172 hyperthermias observed, 49% were accompanied by a circulating parasitemia and 30% corresponded to malaria attack. The feverish subjects (74% vs. 42%), the subjects with parasitemia (51% vs. 16%) and the cases of malaria (34% vs. 10%) were more frequently encountered in children under10 than in the oldest. The cases of malaria attacks were more frequent from November to January (70%). The strong intensity of malaria transmission in Barkedji and the persistence of its temporary ponds until January were sufficient to influence the level of malaria morbidity and consequently the development of an anti-malaria immunity by the indigenous population.     

Molecular epidemiology of hepatitis B virus in Dakar, Sénégal

Using DNA chip technology and real-time quantitative PCR, molecular profile of HBV strains infecting blood donors and patients in Dakar, Sénégal was studied. All HBsAg-positive blood donors (n = 175) and all patients who presented with chronic hepatitis B (n = 29) between 1st June 2003 and 31st July 2003 were studied. One patient, a blood donor, was coinfected by HCV, and nine patients had anti-HDV antibodies. Few persons in either group were HBeAg-positive. Viral load values were relatively low but correlated with biochemical abnormalities. Patients were infected mainly by genotype E (72%). Patients infected by genotype A (28%) tended to be younger than other patients. There was no significant difference between the blood donors and the patients with hepatitis B as regards virological markers, including viral load, when the HBV genotype was taken into account. The BCP A1762T and G1764A mutations were found in four patients and one patient, respectively; the two mutations were never found in the same patient. The W28* mutation at position 1896 of the core was detected in 19 of the 32 genotyped patients, 18 (83%) of whom had genotype E infection. ALT levels were not influenced by HBV mutations. This study shows a low frequency of clinical signs in HBsAg-positive blood donors, a relatively low level of viral replication, and a high frequency of pre-core mutants in this West African population. These results underline the importance of molecular characterization of HBV infection as specific treatments become available in this region.     

Relation between lupus-antiphospholipids antibodies and heart disorders

BACKGROUND AND AIM: Antiphospholipids antibodies (APL) are autoantibodies found in lupus erythematosus and disorders like. Their frequency varies between 2 and 62% according to the literature. An increased frequency of cardiac disorders in antiphospholipids (APL) positive lupus has been reported. The aim of our study was to evaluate the role of APL as an independent risk factor of cardiac disorders in patients with systemic lupus erythematosus. MATERIAL AND METHOD: A prospective study during 14 months has been designed with the cooperation of dermatologic, internal medicine and cardiology departments of the Aristide Le Dantec hospital of Dakar. Platelets count (Beckmann Coulter analyzer), activated partial thromboplastin time (Diagnostiga stago analyzer) and antiphospholipids antibodies (Elisa) were determined. RESULTS: 37 patients affected by lupus were included in this study with a net feminine prevalence (89%); 8 (14.6%) had APL's significant results and 20 presented an echographic heart abnormality. The analysis of our data did not reveal an increased risk of cardiac diseases among APL positive lupic patients as compared to the negative group (p = 1). CONCLUSION: The presence of APL in patients with systemic lupus does not so seem to be an independant risk factor of heart diseases.     

ETO2-GLIS2 Hijacks Transcriptional Complexes to Drive Cellular Identity and Self-Renewal in Pediatric Acute Megakaryoblastic Leukemia

1. Download high-res image (149KB)
2. Download full-size image

     

Circumcision in hemophilia using low quantity of factor concentrates: experience from Dakar,Senegal

Background

Circumcision in hemophiliacs is a delicate surgery because of bleeding risks that could be avoided by adequate substitution of coagulation factor. This practice is very challenging in countries where anti hemophilic treatment is inaccessible. The study aimed to evaluate a circumcision protocol in hemophilia A using low quantities of factor concentrates.

Methods

This prospective study included 26 hemophiliacs A who underwent circumcision in 2014. Medical treatment protocol using low quantity of factor concentrates was drafted by physicians of the Hemophilia Treatment Center and the surgical protocol by experienced surgeons. Assessment criteria were: number of hospitalization days, number of exposure days to factor concentrates, delay to healing and occurrence of bleeding events.

Results

Mean age was 9.6 years (1–30). Hemophiliacs patients were classified as severe (n?=?8), moderate (n?=?9) and mild form (n?=?9). Mean number of exposure days to factor VIII concentrates was 6.9 days (5–12) in children and 10.75 days (7–16) in adults (p?=?0.0049); mean number of hospitalization days was 3.68 days (2–10) in children and 13.5 days (13–15) in adults (p?=?0.0000); delay to healing was 26.47 days (20–35) in children and 25.25 days (22–30) in adults (p?=?0.697); five haemophiliacs (19.2%) presented bleeding events after the circumcision. The mean amount of FIII concentrates used per patient was 1743 IU (810–2340).

Conclusion

The study shows treatment protocol using low quantity of factor concentrates is efficient in hemophilia patients who underwent circumcision.

     

Severe pulmonary arterial hypertension as initial manifestation of intravascular lymphoma: case report

Intravascular lymphoma (IVL) is a rare and usually fatal disease that belongs to the class of high-grade malignant lymphomas and which is characterized by proliferation of neoplastic lymphoid cells exclusively within the lumina of small blood vessels. Its polymorphic and nonspecific clinical manifestations make antemortem diagnosis very difficult. We report herein a case of IVL revealed by fatal, precapillary, pulmonary arterial hypertension and associated with long-lasting fever. Extensive investigation of the usual causes of pulmonary arterial hypertension was negative. The diagnosis of intravascular lymphoma was made on postmortem analysis, revealing diffuse and sometimes complete obliterations of the lumina of small blood vessels by large B-cell lymphoma, including pulmonary capillaries. Thus, we propose that IVL must be added to the spectrum of etiologies of subacute pulmonary arterial hypertension, notably in the context of associated fever, both entities requiring emergency diagnosis. For this purpose, blood collected via pulmonary capillary-wedge aspiration for cytologic examination may be associated with right-sided heart catheterization when this latter procedure is suitable.     

Influence of speed variation and age on ground reaction forces and stride parameters of children's normal gait

The aim of this study was to assess the influence of both age and speed on ground reaction forces and temporal parameters during normal gait in children. Fifteen children aged 4-6 years (group 1), 16 aged 6-8 years (group 2), and 16 aged 8-10 years (group 3) walked at 2.7 km/h, 3.6 km/h, and 4.5 km/h on a treadmill. For each child thirty successive steps were recorded. The influence of speed and age on normalized gait parameters was examined with two-way analysis of variance. The first vertical peak force (Fz1) and all the antero-posterior forces of group 1 were higher than those of the other groups for the three speeds. The minimum vertical force (Fz2), the second vertical peak force (Fz3), and the duration of stride and stance were significantly higher in groups 2 and 3. For all the groups, Fz1 and all the antero-posterior forces increased with the speed while Fz2, stride, stance, and double-stance duration decreased. Fz3 was not influenced by speed variation. The results of this study show that age and walking speed influence ground reaction forces and stride time parameters in 4- to 10-year-old children.