Jean-Marie Sire
email: jmsire@chi-poissy-st-germain.fr

Benoît Garin

Loïc Chartier

Ndeye Khota Fall

Adama Tall

Abdoulaye Seck

François-Xavier Weill

Sébastien Breurec

Muriel Vray

J-M Sire

Community-acquired infectious diarrhea among children under five years in Dakar, Senegal

Keywords: Infectious diseases, Community-acquired diarrhoea, Children, Dakar, Senegal

Background: In sub-Saharan Africa, infectious diarrhoea is a major cause of childhood morbidity and mortality. A cross-sectional study was undertaken to document the pathogens potentially involved in community-acquired childhood diarrhoea in Dakar, the capital of Senegal. : Between September 2007 and March 2008, 176 children aged 1 month to 5 years were recruited consecutively from a primary health care institution in an urban area. Clinical data were recorded and stool samples were collected. Bacterial pathogens were identified using conventional methods and/or PCR assays. Rotaviruses and adenoviruses were detected by a rapid immunochromatographic test. Intestinal parasites were diagnosed by microscopy.

Methods

Results: Rotavirus was the most common enteric pathogen, detected in 27% of patients, followed by Shigella (12%), diarrhoeagenic Escherichia coli (8%), enteric adenovirus (8%), Salmonella (4%), Campylobacter jejuni (3%) and Plesiomonas shigelloides (2%). Mixed bacterial/viral infections were detected in 6% of cases. Parasites, mostly protozoa, were detected in 14% of children. Using ipaH PCR, 30% of samples were positive for Shigella/entero-invasive E. coli.

Detection of rotavirus was more frequently associated with younger age groups (<24 months), whereas bacterial diarrhoea was isolated more often in children over 1 year of age. Detection of bacterial pathogens was significantly associated with malnutrition. Antibiotics were prescribed in 77% of children who attended to the consultation. No pathogen was found in 36% of them, whereas a virus was detected without any other associated bacterial or parasitic pathogen in 23%.

Conclusion:

In developing countries, there is a need to develop reliable, easy-to-use, inexpensive rapid diagnostic tests to guide the management of diarrhoea in infants and children and thereby prevent over-use of antimicrobial agents.

Introduction

Infectious diarrhoea remains a major cause of morbidity and mortality in developing countries, particularly in children. It has been estimated that in 2010 it accounted for 0.801 million deaths worldwide in children under 5 years. Between 2000 and 2010, deaths from childhood diarrhoea decreased by 0.309 million. [START_REF] Liu | Global, regional, and national causes of child mortality: an updated systematic analysis for 2010 with time trends since 2000[END_REF] However, the morbidity has not declined, especially in Africa, probably because risk factors (poor hygiene, lack of access to potable water, insufficient promotion of breastfeeding, malnutrition) still prevail. [START_REF] Okeke | Diarrheagenic Escherichia coli in sub-Saharan Africa: status, uncertainties and necessities[END_REF] The continued growth of many major African cities with resulting overcrowding may also contribute to outbreaks of diarrhoea to which children are particularly vulnerable. Data are needed in these urban areas to establish the local epidemiology and implement appropriate preventive measures. We report a cross-sectional study undertaken in a dispensary in central Dakar in order to detect the pathogens and clinical characteristics of community-acquired infantile diarrhoea.

Subjects and Methods

Patients

Between September 2007 and March 2008, children aged 1 month to 5 years with acute diarrhoea were enrolled consecutively from the Saint-Martin dispensary, a Catholic mission-run primary health care organisation for maternal/child health in Rebeuss, a poor and densely populated district of central Dakar. Children enrolled had had at least three stools (aqueous, slimy/mucous or bloody) per day, symptoms for <7 days, and had not received any antibiotic treatment during the previous 7 days. Clinical examination was undertaken after taking a history from the parents or guardians.

Information recorded included age, weight, duration of diarrhoea, symptoms of dehydration, vomiting, number and consistency of stools, and non-specific medication received within the previous week. Growth status was expressed separately for males and females by weight-for-age Z-scores (WAZ). [START_REF] Kuczmarski | CDC growth charts for the United States: methods and development[END_REF] Moderate or severe malnutrition was defined as WAZ ≤-2. Treatment received from the dispensary was recorded. Data were collected by nurses using a case research form (CRF).

Ethical considerations

Informed consent was obtained from the children's parents or guardians. The study protocol was approved by the Senegalese Health Research National Council.

Laboratory methods

Stool samples were collected in a sterile plastic vial. A wet smear was microscopically examined for intestinal protozoa trophozoites. Stool samples were delivered within 2 hours to the laboratory of the Institut Pasteur for subsequent processing. A portion of stool was concentrated by the merthiolate iodine formaldehyde concentration technique and examined for helminth eggs and protozoa cysts. [START_REF] Blagg | A new concentration technique for the demonstration of protozoa and helminth eggs in feces[END_REF] A rapid chromatographic immune-assay detecting simultaneously rotaviruses and enteric adenoviruses was performed (VIKIA Rota-Adeno, bioMérieux, Marcy l'Etoile, France).

Stool specimens were inoculated on to Hektoen Enteric agar (Becton-Dickinson, Heidelberg, Germany) to isolate Salmonella and Shigella species, and bromocresol purple lactose agar (BCP) (Becton-Dickinson) to isolate E. coli and other Gram-negative bacteria. Preston broth medium made in the laboratory was inoculated to enrich Campylobacter species and subcultured onto Karmali selective agar (Oxoid, Basingstoke, England). [START_REF] Bolton | A selective medium for isolating Campylobacter jejuni/coli[END_REF] Muller-Kauffmann tetrathionate broth (Bio-Rad, Marnes-la-Coquette, France) was used to enrich Salmonella and subcultured onto Rambach agar (Merck, Darmstadt, Germany). Serogrouping and serotyping of Shigella isolates were conducted with commercial antisera (Statens Serum Institut, Copenhagen, Denmark and Denka-Seiken, Tokyo, Japan, respectively). Salmonella isolates were serotyped according to the Kauffmann-White scheme using commercial antisera (Statens Serum Institut). [START_REF] Grimont | Antigenic formulae of the Salmonella serovars. WHO Collaborating Center for Reference and Research on Salmonella[END_REF] Antibiograms were performed by the disc diffusion technique according to the guidelines of the Antibiogram Committee of the French Society for Microbiology (CASFM). E. coli ATCC 25922 was used as the control strain. Vibrio cholerae was not investigated because there were no reported outbreaks during the study duration.

To improve assessment of the real involvement of Shigella in diarrhoea, stool samples were stored at -20°C for subsequent testing by PCR assay based on amplifying the invasion plasmid antigen H (ipaH) gene, contained in the four Shigella species, as well as in entero-invasive E. coli (EIEC). [START_REF] Sethabutr | Detection of Shigellae and enteroinvasive Escherichia coli by amplification of the invasion plasmid antigen H DNA sequence in patients with dysentery[END_REF] Shigella flexneri serotype 5 wild-type strain M90T was used as control strain for ipaH PCR. Prior isolation of bacterial DNA was performed using the QIAamp DNA Stool Mini Kit (Qiagen, Courtaboeuf, France).

Strains previously identified as E. coli using the API20E gallery (bioMérieux, Marcy l'Etoile, France) were screened with a single-test multiplex PCR for the presence of 12 genes to differentiate between the six currently recognised diarrhoeagenic E. coli pathotypes. [START_REF] Müller | Identification of unconventional intestinal pathogenic Escherichia coli isolates expressing intermediate virulence factor profiles by using a novel single-step multiplex PCR[END_REF] These pathotypes include enteropathogenic E. coli (EPEC), atypical EPEC (ATEC), Shiga toxinproducing E. coli (STEC), enterotoxigenic E. coli (ETEC), entero-invasive E. coli (EIEC) and entero-aggregative E. coli (EAEC). Atypical EPEC (ATEC) strains harbour the LEE (locus of enterocyte effacement) as EPEC strains, but not the EAF (EPEC adherence factor). Simultaneous detection of E. coli-specific uidA gene was used to confirm biochemical identification of the isolate as E. coli, and to serve as PCR control. [START_REF] Müller | Identification of unconventional intestinal pathogenic Escherichia coli isolates expressing intermediate virulence factor profiles by using a novel single-step multiplex PCR[END_REF]

Statistical analysis

The study population was described as mean (SD) or median [interquartile range] for continuous variables and as percentages for discrete variables. Relations between clinical variables and pathogens were analysed using the Fisher's Exact test. A test for trend across ordered variables was used to assess relationships between the different pathogens and patient age. Statistical analysis was performed using the STATA 11 software.

Results

Population

Between September 2007 and March 2008, 176 children were recruited consecutively to the study (Table 1). Fifty-five per cent were male, and the median age was 13 months [IQ1 8, IQ3 20].

Thirty-six per cent were malnourished (Z-score ≤-2). The median number of stools per day was four [IQ1 3, IQ3 5]. Before attending the dispensary, 20% of children had received oral rehydration salts (ORS), and 23% had received non-specific anti-diarrhoeal agents (e.g. smectite), anti-motility agents (e.g. loperamide), anti-helminthic agents (e.g. albendazole) or medicinal plants (e.g. guava leaves). Temperature was ≥38°C in 30% of cases, there were clinical signs of moderate dehydration (two or more signs of the following: restlessness, sunken eyes, thirst and positive skin pinch test) in 59%, vomiting in 61%, and abdominal pain in 42%. None had severe dehydration. Blood was macroscopically seen in 11% of stool samples.

[t]TABLE 1[/t]

Pathogens

At least one of the sought-for pathogens was identified in 105 (60%) children, two pathogens were identified in 25 children, three in two children and four in one child. Bacterial and viral pathogens were detected in 48 (27%) and 55 (31%) stools, respectively, while parasitic elements were detected in 25 (14%) samples. Mixed bacterial/viral infections were detected in 11 (6%) children.

The pathogens identified in stool samples are shown in Table 2. Rotavirus was the most prevalent pathogen, detected in 46 (27%) stools, 33 (72%) of them in January. The presence of rotavirus was significantly related to vomiting (43% of children with rotavirus had vomiting vs 10% of those without rotavirus, P<0.001), absence of blood in stool (0 vs 30%, P=0.002), and normal body temperature (32% vs 8%, P=0.001), but not to clinical signs of dehydration (27% vs 29%, P=0.78). Enteric adenovirus was found in 13 (8%) patients.

[t]TABLE 2[/t]

Shigella was isolated from 21 stools (12%). Sixteen were identified as Shigella flexneri, four as Shigella sonnei biotype g, and one as Shigella boydii serotype 2. Among the 16 isolates of S. flexneri, six belonged to the serotype 2a, six to the serotype 1b, three to the serotype 4a, and one to the serotype 3a. Most of the Shigella isolates were resistant to cotrimoxazole (overall resistance rate 95%). Rates of resistance of S. flexneri strains to amoxicillin, amoxicillin-clavulanic acid, ticarcillin and chloramphenicol were 44%, 44%, 44% and 37%, respectively, whereas all S. sonnei strains were susceptible to these antibiotics. All Shigella strains were susceptible to third-generation cephalosporins (cefotaxime) and quinolones (nalidixic acid, ciprofloxacin). Non-typhi Salmonella enterica, Campylobacter jejuni and P. shigelloides were isolated from seven (4%), six (3%) and three (2%) stool specimens, respectively. All Salmonella strains were resistant to cotrimoxazole but remained susceptible to aminopenicillins, first-and third-generation cephalosporins, chloramphenicol and quinolones.

The ipaH gene was detected in all 21 Shigella culture-positive specimens and in 30/148 (20%) culture-negative specimens (DNA amplification failed in seven stools). Since recent advances have confirmed that EIEC and Shigella should be considered a single pathotype of E. coli, 51/169 (30%) samples could be finally considered positive for the entity Shigella/EIEC. [START_REF] Lan | Molecular evolutionary relationships of enteroinvasive Escherichia coli and Shigella spp[END_REF] Presence of Shigella/EIEC was significantly associated with blood in the stool (84% vs 23%, P<0.001) and tended to be associated with fever (40% vs 27%, P=0.099).

Diarrhoeagenic E. coli were detected in 14 children (8%): EPEC (6 isolates), EAEC (4 isolates), ATEC (2 isolates), ETEC (2 isolates). Neither STEC nor EIEC was detected.

Parasitic agents were detected in 25 (14%) children, namely Giardia lamblia (15 patients), Entamoeba histolytica/Entamoeba dispar (8), Ascaris lumbricoides (4) and Trichiuris trichiura (1).

One patient harboured three different parasites.

Factors associated with pathogens

Table 3 shows the distribution of pathogens amongst the three age groups, 0-12, 13-24 and 25-60 months, respectively. The prevalence of rotavirus was significantly higher in children aged 0-24 months than in those over 24 months (31% vs 6%, P<0.01). Conversely, bacterial pathogens, when considered all together, were significantly less frequent in those aged 0-24 months than in those over 24 months (24% vs 42%, P=0.04). This was particularly so for Shigella. Distribution of diarrhoeagenic E coli strains was not significantly different between the age groups. The prevalence of parasites increased significantly with age. The distribution of pathogens according to nutritional status is shown in Table 4. Moderate or severe malnutrition was significantly related to detection of bacterial pathogens considered as a whole (P=0.04). No significant association was found between rotavirus infection and malnutrition (P=0. 14), nor between parasitic detection and malnutrition (P=0.94).

[t]TABLES 3 & 4[/t]

Therapeutic management

Among the 140 children for whom treatment prescribed in consultation was documented, 108 (77%) had received antibiotics. Fifty-seven (53%) had received one antimicrobial agent, 38 (35%) two, 9 (8%) three and 4 (4%) up to four different antimicrobial agents. One hundred and six (76%) children received metronidazole for presumed intestinal protozooses, 82 of whom (77%) also received one or more antibiotics. The most frequently prescribed antibiotics among documented patients were ceftriaxone (51%) and gentamicin (11%) by the IM route, and oral amoxicillin/clavulanic acid (26%) and trimethoprim-sulfamethoxazole (16%). Among the 108 children treated with antibiotics, no potential pathogen was found in 39 patients (36%) [in 26 patients (24%) if we consider results for the detection of Shigella/EIEC by PCR]. A virus was detected without any other associated bacterial or parasitic pathogen in 25 cases (23%). ORS were prescribed to only six patients.

Discussion

Since this was not a case-control study, pathogens detected should be considered only as potential pathogens. Nevertheless, it enabled evaluation of the relative importance of different diarrhoeagenic agents in a population of young children living in an urban district of a major West African city.

Rotavirus is a leading cause of severe acute diarrhoea in children under 5 years of age in Africa, [START_REF] Waggie | Review of rotavirus studies in Africa: 1976-2006[END_REF] and thus, unsurprisingly, rotavirus was the pathogen most commonly found in our population. A peak in frequency, probably an indicator of an epidemic, was observed in January, one of the coolest and driest months in Dakar. This high frequency during the dry season is consistent with previous studies in neighbouring countries. [START_REF] Hanlon | Epidemiology of rotavirus in a periurban Gambian community[END_REF][START_REF] Mølbak | The etiology of early childhood diarrhea: a community study from Guinea-Bissau[END_REF] Detection of rotavirus was more frequently associated with younger age (<24 months). There is evidence that the incidence of rotavirus infection decreases with age because effective immunity conferred by repeated infections develops gradually. [START_REF] Velázquez | Rotavirus infections in infants as protection against subsequent infections[END_REF] Moreover, protection against rotavirus is conferred primarily by secretory IgA antibodies located on the surface of the intestinal mucosa. Therefore, maternal antibodies may confer only limited protection against rotavirus during the first months of life. The presence of caliciviruses (i.e. noroviruses and sapoviruses) was not tested in this study. Nevertheless, it is likely that they were involved in some of our cases. Indeed, a recent study in Dakar children hospitalised for acute gastro-enteritis found an 8% prevalence of caliciviruses. [START_REF] Ka | Calicivirus gastroenteritis in children hospitalized in Dakar[END_REF] Shigella is a major public health concern in developing countries, particularly in children, [START_REF] Kotloff | Global burden of Shigella infections: implications for vaccine development and implementation of control strategies[END_REF] and was detected by culture in 21 samples (12%). Its prevalence (identified by conventional culture) is comparable with that found in Dakar in 1993-1994 (11%). [START_REF] Gassama | Shigella in Senegal (1993-1994): distribution through serovar, sensibility to antibiotics and virulence[END_REF] If, however, we consider the ipaH PCR results, 51 samples were positive for Shigella/EIEC, thus increasing the total percentage of positive stools from 12% to 30%. Shigella could therefore be considered the most frequently recovered pathogen, suggesting that the true incidence of Shigella in childhood diarrhoea in developing countries may be largely underestimated in studies using only conventional culture techniques. S. flexneri was the most commonly isolated species in our patients, whereas S. sonnei was detected less frequently. In contrast with industrialised countries, the predominance of S. flexneri over S. sonnei is common in developing countries and has been linked to poor hygiene. [START_REF]Antimicrobial Resistance in Shigellosis, Cholera and Campylobacteriosis[END_REF] The low number of Shigella isolates limits the significance of antimicrobial susceptibility data.

However, the study confirmed a recent report showing that resistance of Shigella to first-line antimicrobials (amoxicillin, amoxicillin-clavulanic acid, chloramphenicol, trimethoprimsulfamethoxazole) is common in Dakar. [START_REF] Sire | Antimicrobial resistance in Shigella species isolated in Dakar, Senegal (2004-2006)[END_REF] In contrast, the latter and our study showed that thirdgeneration cephalosporins and quinolones remained effective.

It is well established that diarrhoeagenic E. coli pathotypes are a major cause of diarrhoea in Africa, especially in children. [START_REF] Okeke | Diarrheagenic Escherichia coli in sub-Saharan Africa: status, uncertainties and necessities[END_REF] Nevertheless, this aetiology is largely unrecognised because few laboratories are able to perform the appropriate identification techniques (i.e. molecular techniques and/or adherence assays). In this study, typical EPEC was the most frequently recovered pathotype (six strains). In addition, two strains of atypical EPEC (ATEC) were detected. EAEC and ETEC pathotypes were also detected (four and two strains, respectively), in contrast with STEC and EIEC.

Campylobacter is a major cause of childhood diarrhoea in developing countries, especially in children under 2 years of age. [START_REF] Coker | Human campylobacteriosis in developing countries[END_REF] We found this pathogen in 3% of cases, a result consistent with prevalences reported in previous studies in West Africa. [START_REF] Mølbak | The etiology of early childhood diarrhea: a community study from Guinea-Bissau[END_REF][START_REF] Koulla-Shiro | Prevalence of Campylobacter enteritis in children from Yaounde (Cameroon)[END_REF] Diarrhoea associated with bacteria, particularly Shigella, affected children over 1 year of age more commonly. This might be explained by the protection conferred by maternal antibodies during the 1st year of life. We found no particular tendency for diarrhoeagenic E. coli. The relationship between malnutrition and severity of diarrhoea is well recognised, especially because of the immune system deficiency caused by inadequate intake of micronutrients such as zinc. [START_REF] Podewils | Acute, infectious diarrhea among children in developing countries[END_REF] In this study, the detection of bacterial pathogens was significantly associated with malnutrition but not with the detection of rotavirus and parasites.

Parasitic agents were mainly protozoa (Giardia lamblia, E. histolytica/E. dispar), while helminth eggs were rarely detected. This low prevalence of helminthiasis may be owing to the young age of the children. A study in Senegal in 2002 showed that the rate of helminth infestation was highest in children between 11 and 15 years of age. [START_REF] Ndir | Prevalence of intestinal parasites at the King Baudouin health center of Guediawaye (Senegal)[END_REF] The prevalence of parasites (both protozoa and helminths) in this study also tended to increase with age. Another reason for this low prevalence of helminths in our population might be that a number of children had received antihelminthics before coming for consultation. It is noteworthy that we did not investigate for Cryptosporidium species in our study. This pathogen was significantly associated with childhood diarrhoea in West Africa. [START_REF] Opintan | Pediatric diarrhea in southern Ghana: etiology and association with intestinal inflammation and malnutrition[END_REF] The majority of children presented with signs of mild or moderate dehydration. No cases of severe dehydration were recorded, probably because they were directly hospitalised in paediatric wards.

Of those children with documented antibiotic treatment prescribed in consultation, a virus (rotavirus or adenovirus) was detected without any other associated pathogen in 23% of cases. Moreover, no pathogen was found in 36% of children. Although this rate could be reduced to 24% when the results of PCR ipaH for detecting Shigella are considered, antibiotic treatment was unnecessary in approximately half of the cases. This over-use of antibiotics is of concern because it promotes the emergence of antibiotic resistance. It also represents a significant cost to families. This report underscores that accurate evaluation of pathogens in childhood diarrhoea in developing countries requires the integration of advanced methods that complement existing tools. Moreover, it highlights the need for primary health care facilities to perform certain simple tests that assess the appropriateness of antibiotic treatment. Microscopic examination of stools can be done easily if staff are trained in basic parasitology. At the same time, however, simple, reliable and inexpensive diagnostic tools for detecting other pathogens, especially bacteria, are needed.

Immunochromatographic tests (dipsticks) are being developed for S. flexneri 2a and S dysenteriae 1. [START_REF] Nato | Dipstick for rapid diagnosis of Shigella flexneri 2a in stool[END_REF][START_REF] Taneja | e24830. Table 3 Distribution of identified pathogens according to age * Identified by culture; † determined by ipaH PCR; ‡ selected by culture and confirmed by multiplex PCR. Organism Age group, mths No. (%) of children infected P-value Rotavirus (n=46) 0[END_REF] These tools, although promising, require further evaluation in routine practice.

They also need to be developed for other pathogens, particularly other species of Shigella and diarrhoeagenic E. coli.

Table 4

 4 Distribution of identified pathogens according to nutritional status (n=157, 19 missing values) * Identified by culture; † determined by ipaH PCR; ‡ selected by culture and confirmed by multiplex PCR. Normal nutritional status was defined as weight-for-age Z-score >-2, moderate or severe malnutrition as weight-for-age Z-score ≤-2.

		Normal nutritional status	Malnourished
	Organism	n=101 (%)	n=56 (%)	P-value
	Rotavirus (n=42)	31/98 (32)	11/54 (20)	0.14
	Bacteria* (all species) (n=41)	21/101 (21)	20/56 (36)	0.04
	Shigella spp* (n=17) Shigella/EIEC † (n=42) Diarrhoeagenic E. coli ‡ (n=13) 5/101 (5) 8/101 (8) 24/97 (25)	9/56 (16) 18/53 (34) 8/56 (14)	0.12 0.23 0.07
	Parasites (n=22)	14/101 (14)	8/56 (14)	0.94

Acknowledgment

The authors thank Natasha Dubois-Cauwelaert, Alice Dufougeray and Fatou Bintou Dièye for technical help.


Hélène Mainet

Jean-Charles Edouard

THE ROLE OF SMALL AND MEDIUM SIZED TOWNS IN EASTERN AFRICA MOUNTAINS: NEW OPPORTUNITIES OR CHALLENGED POSITION?

Keywords: small and medium sized towns, intermediate towns, mountains, Eastern Africa

The small and medium towns have for long been playing an essential role as intermediary between rural economies and populations and urban territories. Local changes as well as national and global evolutions are modifying those classical linkages through new scales of integration and the emergence of new actors. In this context, the intermediary position played by secondary towns (as market towns, districts and services poles) is changing, between strengthening and bypassing effects.

Introduction

In the context of Eastern African mountains, relationships with cities are an important element as spatial mobility, exchanges and linkages between rural and urban spaces are strong. The place taken by small and medium sized towns is particularly interesting. They are play an old and renewed role as intermediate, as outpost between rural and urban economies, acting as relays in territories interactions and assuming functions of poles of centrality, even at a second level. They take part in the setting of migration flows and are places of diffusion of development through their public equipments, services and shops or enterprises. However, they are not always supported by national policieswhich are often dedicated to major centersand can suffer from globalization leading to the concentration of qualified people and activities in the upper levels of the urban hierarchy, more integrated in global networks. Local changes but also global and national strategies have modified classical ways of integration, articulating scales (local, national or international) and showing new actors. Our hypothesis is that the intermediate function played by small and medium sized towns is changing, between strengthening opportunities and weakening logics. The main issue is therefore their capacity to keep on structuring heterogeneous places and to link more numerous actors. The multiplication of development actors, mainly private, at different scales, who articulate or overlap their spatial practices with official and administrative limits, reinforces the territory complexity but also innovation processes based on relationshipsurban-rural linkages or within the urban hierarch.

Fieldworks for the research program "Mountains and Medium-Sized Towns 1 " in Kenya, Uganda and Tanzania give elements of analysis. Surveys have been conducted with local private and public actors of development (municipal services, head of associations, cooperatives and small and medium-sized enterprises), associated with meetings with shopkeepers and street vendors, on issues of networks and linkages 2 . The analysis is based on the case of medium towns (Mbale, Mbeya and Nyeri) associated with small towns (respectively Kapchorwa, Tukuyu and Karatina). Mbale is located at the foot of Mount Elgon, "fruit and vegetable basket" for Uganda, next to the Kenya border. With 80.000 inhabitants it is the main urban centre of this rural area (doc. 1). Mbeya (in Tanzania) is located 700 kilometers South-West of Dar es Salam, in the Uporoto Montains and next to Mount Rungwe, on the TAZARA line, next to Zambia and Malawi borders. It has around 300.000 inhabitants. The third case study is in Central Kenya Province, with Nyeri (100.000 inhabitants) as medium town and Karatina (7.000 inhab.) as small town examples. They are located between Mount Kenya and the Aberdares range. Regional contexts are quite similar with dynamic and changing mountain environments (evolution in farming and rural economies) and with high population density rural areas. Linkages between highland hinterlands and secondary towns are intense and diversified. Mbale and Mbeya are in border and off-centered situations, whereas Nyeri is more central and closer to the capital city of Nairobi (130 Km). The examples of the above towns give information for the changing role of small and medium sized towns in their intermediate functions, in contexts of rapid evolution. They show the innovative capacities of actors and issues of new territorialities at stake through the growing role of middlemen and the process of integration in wider networks.

1. The traditional role of small and medium sized towns in the urban-rural interface

The importance of small and medium sized towns in rural-urban relationships has for long been analyzed in the African context and East African Mountains are particularly interesting for the understanding of those linkages.

Abundant literature and an opportune context

French-speaking as well and English-speaking researchers have been interested in the role of secondary towns in the extraversion of rural economies [START_REF]The Market Towns and Agriculture in Africa: The Role of Small Urban Centres in Economic Development[END_REF][START_REF] Simon | Conceptualizing Small Towns in African Development[END_REF][START_REF] Owuor | Small and Medium-Size Towns in the Context of Urbanization and Development Process in Kenya[END_REF]. The development of linkages between secondary towns and rural areas is crucial for economic development, farm productions and commercial linkages (Chaléard, 1996). The literature stresses this point in West Africa, where secondary towns are important at the regional level, interacting with rural hinterlands as markets and redistribution points, creating demand and stimulating local products (Giraud, 1994). An urban-rural continuum is characteristic of this urbanization. Lots of regional or weekly markets are important commercial nodes for productions. They are places for the trade of farm and cattle products but also relay places between rural and urban zones.

The role of those towns is not only dedicated to market. They are administration places and areas of social and economic interactions for rural people. They play an important role as rural mobility catchments, in the opening of the mountains to urbanity through the diffusion of modernity and offer of services. The different denominations used show the diversity and complementary nature of their functions. If some authors insist on the size and then on the position in the urban hierarchy (small or medium towns), others valorize functions (market towns, district towns, sub-centers). Often, the stress is put on interactions and articulations played by those intermediate, median, transition towns through their support, distribution or network functions, changing according to the scale of analysis.

Those secondary towns are described as the symbol of intermediate towns, as transition between major cities and other towns and rural areas, and vice versa, from the bottom to the top. They have an intermediation role in a hierarchical structure, between an urban centre and its regional hinterland as center of economic, social, cultural and historical services. The intermediate town is located between two extremities (the small and the big, the near and the far) and develops intermediation functions between spaces at different scales (local, regional, national, international). They are at the junction of flows of information, ideas, people, goods and services, a transition place between adjacent areas and global zones. What is important is their capacity to be integrated in global networks (Carrière, 2008).

East African mountains are interesting places for the analysis of urban-rural linkages. They form fertile areas, with ancient flows of people attracted by exceptional environmental conditions (on the Mount Elgon, rural districts of Manafwa and Bududa have between 800 to 1.000 inhab./Km²). They are the core of East African agricultural production, drivers of past colonial and current national economies. These mountains are 'open' mountains, included in systems of exchange that integrate multi-scalar geographic dimensions: highland and lowland systems, humid and arid systems, rural-urban mobility, etc. [START_REF]Kilimanjaro, mountain, memory, modernity, Dar es Salaam[END_REF]. Towns are linked with those mountains, with lots of markets, small localities or medium towns with old commercial and transport functions [START_REF] Nakileza | Le développement du maraîchage dans les montagnes d'Afrique de l'Est : les enjeux[END_REF]. Relationships between highlands and lowlands, between rural and urban places are structuring those « bastions of rurality » and secondary towns are playing their role of intermediation.

The role of secondary growth pole at national and international scales

At national and international scales, intermediate towns are important in three main ways. They play an important role in rural and regional people settlement, assume their part in territory administration and are privileged places for diffusion of social modernity (doc. 2). Document 2 -Small and medium sized towns: an interface role in the rural-urban linkages in East-African Mountains Their dedicated missions in national planning are often linked with their place in rural-urban relationships as they have strong linkages with their hinterlands [START_REF] Tacoli | Urban-Rural Interactions: a Guide to the Literature[END_REF]. Their place as services and commercial centers has for long been seen as crucial for the development of rural zones (to answer needs of inhabitants and producers). More recently, small towns are considered to play an important role in the linkages of their hinterlands with national and global markets but also in the provision of non-farm activities for rural population, therefore widening the local economic basis. They also supply services and equipments for their regional hinterlands. The term "secondary", often employed, underlines the dependency towards decision centers located in bigger towns or abroad. Local and regional subdivisions of administration or branches of economic and social services are numerous in medium towns, when headquarters are located in more important cities. Moreover, they are seen as centers from which innovation and modernization processes can spread in rural zones.

The case studies show this secondary position (important regional centers) in structured urban networks, in off-centered even isolated situation (in the case of Mbeya) from national capitals. But their polarization on regional urban network (on small towns) and rural areas is comforting them as intermediate poles.

Classical functions of district center and pole of services at local and regional scales

On the political and administrative sides, those towns are often district centers (their status depending on the administrative organization of the countries), with lots of civil servants and private actors. Thus, they are well equipped and have developed service industries such as a variety of goods and services equipments and infrastructures, good connections, important school system, health services and finance activities.

From an economic point of view, small towns are often collecting centers for farm products and places of important weekly markets. Daily, it is an important supply place of manufactured goods for urban and rural inhabitants. The redistribution function of agricultural goods is obvious in those towns through the omnipresence of shops selling products like fruits, vegetables or dairy. One of the most important places is the central market (and all its surrounding shops and stalls) but products can be found all over the town.

In Mbale, vegetable retailers are supplied three times a week from mountain areas (in Irish potatoes, sweet potatoes, carrots, cabbages, onions, bananas, etc.). Middlemen are important in this trade: they go to rural zones to buy products and bring them back to urban retailers (doc. 3). It is easy to find products from all around Mount Elgon, from Kapchorwa in the North or Bududa in the South. An important part of the mountain production is also sold in other urban centers (Jinja, Kampala) and redistributed all over the country, through urban actors. The same occurs in Central Kenya with a market function even more important in Karatina (small town) than in Nyeri (medium town) due to a better location on the main road: Karatina market is famous for being very busy due to various productions from the mountains around (Mount Kenya and Aberdares slopes) and the proximity to Nairobi urban demand.

Medium towns are also gateways to rural areas. They are nodes of redistribution for productions, often manufactured, "imported" (in the sense that they come from outside the area, either from other regions or from abroad). In Mbale, lots of products are coming from Kampala and other cities or from Kenya. It is like the outpost from where products are redistributed (doc. 3). In the case of Mbeya, due to the border location and with the important Finally, the classical intermediation function of secondary towns is linked to the central criteria of connectivity [START_REF] Nadou | La notion de « villes intermédiaires », une approche différenciée du rôle des villes petites et moyennes : entre structuration territoriale et spécificités socioéconomiques, colloque Villes petites et moyennes, un regard renouvelé[END_REF]. It is at the same time external (opening to state and outer markets, networks issues) with strategic functions, and internal (towards surrounding areas) with essential daily functions (services center, influence on hinterland, proximit y issues). This old capacity of intermediate towns as catchment and distribution centers is a way to link rural economies to national and international economies (doc. 2). If this role is still active, it is submitted to important stakes linked with changes in agriculture (post-coffee economies, market gardening development) but also with impacts of broader politicoeconomic transformation.

Changes in the intermediation function of small and medium sized towns

For two or three decades, the growth of medium towns has been taking place in changing rural dynamics, linked to local processes as well as impacts of more global evolutions.

The importance of new local initiatives

The opening of new markets, liberalization of economies and emergence of new actors are accelerating changes in long term established linkages. The involvement of producers in new productions and distribution networks (mainly market gardening) is redefining composite and unequal productive relationships, where land issues, access to water and environmental resources, availability of new communication tools become a relative advantage. Highlands specialized in traditional commercial crops suffer from historical rigidity (land tenure and management issues). It is still uncertain if farmers have the capacity to invest in those new production sectors, to redefine their relations with markets and national (and even) international economies, to defend their products, with new forms of organizations and a better professionalization. Agricultural changes, new farm practices and social changes are closely linked. Coffee production is declining (with some differences depending on areas) either in estates and commercial productions or in village plots, often in combination with bananas as in the Mount Elgon area [START_REF] Nakileza | A true story of coffees magendo through Mount Elgon (Uganda)[END_REF]. New productions benefitting from highland environmental resources are developing fast: market gardening for urban demand but also dairy products. Cabbages, onions, potatoes, tomatoes, carrots… are exchanged daily and supply new markets from where middlemen or traders will transport them to urban centers (photo 1).

Those agricultural changes have powerful impacts on land structures and social organization forms. With a strong demographic growth and investments of urban businessmen in farm production, the pressure on land is important. The development of market gardening shows that farmers are reactive to changes. But a complex and unequal social process is taking place, in a still fragile context where individual initiatives are important and collective organizations not always strong. Besides market gardening development, pluriactivity is increasing within households holding multiple jobs or incomes in non farming activities often related to the production (transformation, selling and services). "Mountain territories are nowadays invested by forces and issues which challenge the notion of rurality and question their long-term acquired and assumed fertility, basis of their specificity3 " [START_REF] Nakileza | Le développement du maraîchage dans les montagnes d'Afrique de l'Est : les enjeux[END_REF].

Photo 1-Retail trade of market gardening products in Karatina market, Kenya (Mainet, 2009) These productive changes are associated with new forms of mobility and migration (circular, temporary, commuting). Small and medium sized towns are therefore centers for all sorts of mobility, from and to rural zones. Transport networks have been developing in and around intermediate towns for daily or weekly travels of people or transportation of products (bodaboda for short-distance and lowlands areas, tatas, matatus and taxis for highlands connections). Private actors, local and national, have invested in this opportunity area and are playing a crucial role in the integration of these economies to broader markets.

New scales of relationship are at stake. Flows from highlands to lowlands give evidence of new territory frame and resources uses. Pluri-resources and pluriactivity redefining trajectories and households strategies [START_REF] Nakileza | Le développement du maraîchage dans les montagnes d'Afrique de l'Est : les enjeux[END_REF]. In this context, urban middlemen, mainly wholesalers and traders whose role is often criticized but important in the development of networks, but also farmers and producers who transport their own production (often combined with others) directly to markets, show their capacity of economic and social innovation. Farmers are weaving ties between multiple territories, between rural and urban areas and are promoting multi-scalar social and economic linkages.

Stakes associated with global changes

In addition to the transformation of mountain economies and new derived linkages, the role of intermediation of secondary towns is also affected by the impact of more general changes (doc. 5).

Document 5 -Small and medium sized towns: new intermediation functions in changing context

A positive aspect is often dedicated to this level of the urban hierarchy in the academic literature. Small and medium sized towns seem to be at a level where transformations are visible, measurable and the implementation of "good more feasible than at other scales [START_REF] Owuor | Small and Medium-Size Towns in the Context of Urbanization and Development Process in Kenya[END_REF][START_REF] Bertrand | Plaidoyers pour les centres secondaires en Afrique au Sud du Sahara[END_REF][START_REF] Pourtier R | Petites villes et villes moyennes : développement local et encadrement étatique[END_REF].

The economic crisis which leads to a rapid decline of incomes in most of African countries seems to have reinforced the role of secondary towns. Households' incomes must be diversified and households' and families' strategies strengthened. Return migrations seem to be important in time of crisis and "exurbanization" phenomenon has been analyzed, describing migrations from main urban centers to secondary ones [START_REF] Bruneau | Vivre la ville à la campagne : crise des sociétés et exurbanisation en Afrique tropicale[END_REF]. In these towns, due to the proximity with rural economies and opportunities, to lower prices and rents than in bigger towns, economic crisis impacts seem less difficult to overcome.

With the intensification of economic liberalization in countries like Uganda and Tanzania, an increasing number of actors are involved in rural-urban linkages and traditional references are blurred. Small and medium sized towns provide important public and private services and infrastructures for the development of farming and non farming enterprises. Lots of rural activities are located in small towns, dealing with transport, building, wholesale and retail trade, financial and credit services and supplying urban as well as rural population [START_REF]The Market Towns and Agriculture in Africa: The Role of Small Urban Centres in Economic Development[END_REF]. These activities are good indicators of the urban influence of these centers. Karatina illustrates the important amount of businesses involved in trade (more than 77 % of all the registered business permits in 2011), but also the diversification of activities with transport companies and services (professional and technical).

Document 6-Registered business permits in Karatina (Kenya) With the development of liberalization, old cooperatives like the Tanganyika Farmers Association in Tanzania (with a regional branch in Mbeya) are competed by new and often small private structures, giving better prices and conditions to members and better payment opportunities. The increasing demand for agro-inputs (fertilizers and chemicals, seeds and animal feed) is supplied by more and more private actors, visible all around small and medium sized towns streets (photo 2).

Photo 2 -Example of new retail shops for farmers (inputs) in Mbale, Uganda [START_REF]Frontières et interfaces : l'évolution des discontinuités socio-spatiales à Dundee et Eshowe, petites villes du KwaZulu-Natal, Afrique du Sud[END_REF] New issues are also important in the financial domain. Microcredit boards like SACCOs (Savings and Credit Cooperative Societies) or farmers associations are active. It is a quite recent phenomenon: most of the offices have been opened for less than ten years. In the Mbale region, the Uganda Microfinance limited is an interesting example. It is a branch of the Equity Bank of Uganda. The main office is in Tororo (a medium town South of Mbale) but the operational office is in Mbale, in charge of the Mount Elgon region with Sironko, Kapchorwa and Manafwa areas (doc. 3). The bank provides loans for groups of farmers (with 10 to 20 members) for short-term periods (from 3 to 12 months). Analyzing those organizations (associations, cooperatives or financial institutions) show the importance of the urban network for the most structured ones: regional branches are located in medium towns (Mbale, Mbeya or Nyeri), local offices in small towns (Kapchorwa, Tukuyu or Karatina) and rural antens in market towns (Bududa, Wanale) with representatives for more isolated rural areas.

Those evolutions question the capacity of cooperation between actors and the effects on local development. A priori, they tend to reinforce the role of secondary towns by diversifying their services and by giving better prices and opportunities for farmers. But long-term impacts should still be measured, specifically the leading capacity of the smallest enterprises, often not well structured and which viability is not confirmed in case of economic crisis. Another impact is short-circuiting of intermediate levels with direct access to rural areas by actors from main cities. It is very visible in the transport area with direct supply in rural areas and with the impact of new communication technologies (doc. 4). Farmers have now better and direct access to urban demand, can be informed of urban prices with direct cell phone calls, and don't need secondary towns' middlemen as much as before.

On the political side, decentralization processes have been affecting most of the countries since the 1990's, giving new opportunities for regional centers. Yet these reforms are not always strengthening their position with the creation of new rural districts (such as Bududa and Manafwa districts in the Mbale region) which used to be dependent on Mbale district. The issue is dealing with the capacity of the new authorities to cope with needs in terms of equipments and infrastructures with low financial resources. Few real cooperation policies are implemented between urban and rural districts and interterritoriality development is lacking whereas linkages are strong.

Another question is dealing with the lack of industrial activities in those secondary centers. Most of the manufactured goods are produced elsewhere and are imported from other regions or from abroad (doc. 4). Local resources (like timber or farm products) are often transformed outside the area. The example of Karatina is quite representative with around 4 % of business involved in industrial or workshop activities (doc. 6). Intermediate towns are more transit places than productive ones. There is a real stake of developing industrial activities in secondary towns. This would reinforce the role of this level of the urban hierarchy in its assumed role of regional management.

Conclusion

The importance of rural-urban linkages for those economies and regionswith high density and intense mobilityis widely visible and intermediate towns are playing a key role in those interactions through their functions of market, services and relays to innovation. The current context questions the traditional intermediation function of this level of the urban hierarchy. New urban governance strategies should take into account the rural side of urban development, with all the connections, coordination and interrelationships between spaces and actors. Secondary towns are active in the catchment of farm products and for the supply of rural areas but they are lacking real and effective tools for local development. The issue is the capacity of local as well as national actors to get involved in the implementation of interspaces cooperation. In that sense, they represent a good observatory of political and territory policies impacts and effects.

Document 1 -

 1 Location of studied areas 2 Fieldwork conducted in December 2008, July 2009 and July 2011.

  railway and road lines, linkages are strong with all the sub-region (doc. 4). Mbeya is the concentration point for goods coming from a large area (fishes from the Great Lakes). It acts also as Malawi dry port (reloading function). Document 3 -Mbale influence zone in the Mount Elgon area (Uganda) Document 4 -Mbeya, a redistribution center for local, national and international products Through intermediate towns, mountain areas are integrated to international markets. For example, farm inputs such as seeds or chemical products are widely used (and even more due to the evolution of farming practices). Fieldwork conducted in Mbale show that retailers get their supplies for cattle (food or medicines) from Kampala or Kenya (doc. 3). The influence zone of these specialized shops covers the whole Mbale district and surrounding areas (within a radius of 40 to 50 Km). Mbale itself has outposts in small towns like Kapchorwa, Budadiri and Bududa, closer to rural places. They are often more specialized. This system is similar in Tanzania, with Tukuyu acting as a relay for Mbeya influence. The place of Mbeya and Mbale in those commercial farming networks gives evidence of their centrality within mountain systems. The location in border zones is reinforcing this situation. There are mediator and intermediate in rural-urban linkages.

CORUS Program 6165 « Mountains and Medium Towns in East Africa. Environmental management, Flows of people and Resources »,

2008-2011, with French Ministry of Foreign Affairs support.

Author's translation : « Les territoires de montagne sont aujourd'hui investis par des forces, des enjeux dont les déterminants marginalisent la notion même de ruralité et remettent en cause leur fertilité acquise et entretenue, autour de laquelle s'était construite leur spécificité ».