Prevalence and antibiotic resistance profile of Salmonella spp. strains isolated from the vegetable food chain in Niamey, Niger

doi:10.21203/rs.3.rs-4909303/v1

Background

Salmonellosis is the main cause of collective food poisoning in humans. They are bacterial infections caused by various Salmonella species. The overall aim of this study was to determine the prevalence and antibiotic resistance phenotype of Salmonella strains isolated from vegetables in the urban community of Niamey.

Methods

To this end, one hundred and twenty-four (124) samples of some vegetables (carrot, lettuce, onion and tomato) were taken from market garden sites and markets in Niamey. Salmonella were tested using the ISO 6579:2017 4-step method: pre-enrichment, enrichment, isolation and biochemical identification, and the antibiotic resistance phenotype was determined using the standard Kirby-Bauer agar diffusion method.

Results

Some sampling sites showed high Salmonella prevalences, notably site 1 (75.00%) and market 4 (40.00%). The overall level of Salmonella contamination in vegetables was 13.71%, including carrot (25.00%) and lettuce (15.38%) samples. The most common resistance phenotypes were Ceftriazone (100%), Amoxicillin + Clavulanic Acid (76.92%), Ciprofloxacin (46.15%), Aztreonam (33.33%) and Imipenem (30.77%).

Conclusion

The presence of multidrug-resistant Salmonella strains in vegetables reflects the potential risk associated with their consumption. It is important to research the potential resistance and virulence genes of these strains for better management of infectious diseases.

Salmonella

prevalence

vegetables

resistance

antibiotics

Niamey/Niger

Food-borne diseases represent a serious public health problem, with considerable economic consequences worldwide [1]. It is estimated that pathogenic microorganisms in food are responsible for between 6.5 and 33 million illnesses and over 9,000 human deaths per year worldwide [2]. However, pathogenic enteric bacteria, such as Salmonella and certain enterohaemorrhagic serotypes of Escherichia coli, are implicated in a growing number of cases of Collective Foodborne Toxi-Infections [3]. The majority of reported cases of microbial food poisoning (95%) are caused by food prepared at home, in restaurants or institutions. An estimated 5% of cases are caused by industrially-produced foods [4]. Salmonellosis is one of the most common food-borne illnesses in almost all countries, and Salmonella enterica Enteritidis followed by Typhimurium represent the most frequently isolated serotypes [1]. Salmonella is one of the main causes of food-borne outbreaks in developing countries [5, 6, 7]. In human pathology, Salmonella are divided into typhoid serotypes (S. Typhi and S. Paratyphi) and non-typhoid serotypes [8]. Non-typhoidal Salmonella cause salmonellosis through contaminated food products such as fresh produce, eggs, pork, vegetables and seafood [9, 10]. Fruits and vegetables, which represent an important food source of nutrients, have also been reported as a vector for transmission and contamination [1]. However, antibiotic resistance is now one of the most serious threats to global health, food security and development [11]. Antibiotic resistance in Salmonella strains has emerged worldwide, making antibiotic susceptibility testing an important role in public health laboratories. Antibacterial agents are often recommended in cases of suspected salmonellosis. Patients did not respond to the most widely available antibiotics of choice. These practices can enhance antibiotic resistance genes. It is now generally accepted that the main risk factor for increasing resistance in pathogenic bacteria is the uncontrolled use of antibiotics [12]. Indeed, uncontrolled overuse of antibiotics can give rise to a selection of resistant bacterial strains. This is known as antibiotic resistance [13]. The overall aim of this study is to determine the prevalence and antibiotic resistance phenotype of Salmonella strains isolated from vegetables in Niamey.

Study area

The study was carried out in the urban community of Niamey. The Niamey region is located in the south-western part of Niger between 13°24' and 13°35'N latitude and 2°00' and 2°15'E longitude, with an altitude of between 160 and 250 m. Its administrative boundaries cover 552.27 km², of which approximately 297.46km² is urbanized [14]. Niamey's population is estimated at around 1,407,635. The city of Niamey is subdivided into five (5) communal districts, with the following population breakdown by communal district: Niamey I: 287,902 inhabitants; Niamey II: 338,455 inhabitants; Niamey III: 223,685 inhabitants; Niamey IV: 376,271 inhabitants; Niamey V: 181,321 inhabitants [15]. The climate is Sahelo-Sudanian, with a long dry season from October to May and a short rainy season from June to September. The cold dry season is the most favorable for vegetable production, during which most crops are grown. The study was carried out in three (3) large market gardening sites (Gounti yéna, Gamkalé and Harobanda) and five (5) markets (Petit marché, Dar es salam market, dolé market, Wadata market and Harobanda market) selling vegetables in the Niamey urban community (Fig. 1).

Sampling mode

Vegetable samples were taken at production and sales sites (markets). Samples were taken under sterile conditions, using single-use sterile gloves and alcohol to avoid external contamination. Samples were taken from rootless lettuces, whole tomatoes showing no visible damage or cracks, whole carrots and whole onions. A lettuce sample corresponded to three (3) heads of lettuce, and a tomato sample corresponded to three (3) fruits without cracks, weighing around 150g, taken at random from different corners of the production site or from the same vendor's lot. Carrot and onion samples were taken at the point of sale only. A carrot sample consists of at least three carrots weighing a minimum of 150g, taken at random from the same vendor. A batch of approximately 150g served as the onion sample. An information sheet was attached to each sample. Each sample taken was well packaged in a polyethylene bag, then carefully labeled. The samples were then transferred to the microbiology laboratory of the Faculty of Science and Technology (FAST), where they were conditioned and placed in a cooler containing carboglasses to keep the temperature down to around 4°C.

Testing for Salmonella

Salmonella testing was carried out according to ISO 6579:2017 [16] in 4 steps [8]: pre-enrichment, enrichment, isolation and biochemical identification. Bacteriological analysis was carried out on 124 samples.

Pre-enrichment: was achieved by grinding the lettuce sample in a sterile polyethylene bag around the Bunsen burner flame. Twenty-five (25) grams of the shredded material was then taken and introduced directly into 225 mL of buffered peptone water, incubated at 37°C for 24 hrs.
Enrichment: 0.1 mL of pre-enrichment broth was taken and added to 10 mL of RVS and incubated at 42°C for 24h.
Isolation: each enrichment culture is streaked on SS agar and Petri dishes incubated at 37°C for 24 hours.
Biochemical identification: suspected Salmonella colonies were examined using a mini-gallery consisting of 4 media (Kligler-Hajna agar; urea-indol medium, ONPG (O- nitrophenyl-β-D-galactopyranoside), simmons citrate, mannitol-mobility), with confirmation using an API 20^E gallery (Biomérieux, France).

Antibiotic resistance testing

Inoculum preparation

Inoculum was prepared from young colonies grown the previous day at 37°C on nutrient agar. A standardized 0.5 Mac Farland inoculum corresponding to 10⁸ CFU/mL (CA-SFM / EUCAST, 2023) [17] was prepared. Then 100 µL of this inoculum was diluted in 10 mL physiological water to obtain a final concentration of 10⁶ CFU/mL.

Seeding

The final inoculum was inoculated by swabbing Petri dishes, dipping the swab into the bacterial suspension, then wringing it out by pressing firmly against the inner wall of the tube. It is then rubbed over the entire surface of the Mueller Hinton agar (MHA), from top to bottom, in tight ridges. The operation is repeated three (3) times, turning the dish 60° each time. Inoculation is completed by passing the swab around the periphery of the agar [18, 19, 20]. After inoculation, nine (09) antibiotic-impregnated discs (Bio-rad, France) of different families were placed on the MHA surface using sterilized forceps. The plates were left at room temperature for around 15 min after disc placement, to allow prediffusion of the antibiotics, and then incubated at 37°C for 24 hrs. After incubation, the inhibition diameters around the antibiotic discs were measured. Strains were then categorized as Susceptible, Intermediate or Resistant by comparing these diameters with the critical diameters for enterobacteria (for each antibiotic) set by CA-SFM / EUCAST (2023) [17].

Table I. Characteristics of antibiotic discs used for antibiotic susceptibility testing

Families	Antibiotics	Acronyms	Expenses	Inhibition diameter (mm)
Families	Antibiotics	Acronyms	Expenses	Sensitive ≥	Resistant <
β-lactam	Amoxicillin + clavulanic acid	AMC	30 µg	19	19
	Aztreonam	ATM	30 µg	26	21
	Ceftriaxone	CRO	30 µg	25	22
	Imipenem	IPM	10 µg	22	19
	Meropenem	MRP	10 µg	22	16
Quinolone	Ciprofloxacin	CIP	5 µg	25	22
Tetracycline	Tetracycline	TET	30 µg	19	17
Nitrofuran	Nitrofurantoin	F	300 µg	11	11
Sulfonamide	Trimethoprim/Sulfamethoxazole	TXT	25 µg	14	11

1.1. Statistical analysis

IBM SPSS statistics software version 23.0.0.0 was used to calculate frequencies. Microsoft Excel was used to generate the graphs. The data were then subjected to one-way analysis of variance (ANOVA). Differences were considered significant for values of p < 0.05.

Prevalence of Salmonella strains isolated from vegetables

Table 2 shows the prevalence of Salmonella isolated from vegetables by sampling site. Market prevalences ranged from 40% (market 4) to 4.55% (market 1). Salmonella spp. was found in all markets, but markets 3 and 4 were the most contaminated (15% and 30% respectively), followed by Salmonella arizonae in markets 2, 4 and 5. Salmonella pullorum was only identified in market 5 (3.85%, or one (1) species). Salmonella spp. were found on all market garden sites, with site 1 the most contaminated (23.08%, i.e. three (3) species identified). Salmonella arizonae was identified only at site 1 (7.69%). On the other hand, Salmonella pullorum was absent from all samples taken at the market garden sites. Prevalences did not vary significantly between sampling sites (P-value > 0.05).

Table 2

Distribution of *Salmonella* serotypes by **sampling** site
Sampling locations		Number of vegetables	Salmonella serotypes			Total
Sampling locations		Number of vegetables	S. arizonae	S. pullorum	S. spp	Total
Markets	Market1	22	-		1(4,55)^a	1(4,55)
	Market2	22	1(4,55)^a		1(4,55)^a	2(9,09)
	Market3	20	-		3(15,00)^a	3(15,00)
	Market4	10	1(10,00)^a		3(30,00)^a	4(40,00)
	Market5	26	1(3,85)^a	1(3,85)^a	2(07,69)^a	4(15,38)
	Total	100	3(3,00)	1(1,00)	9(9,00)	13(13,00)
Fields	Site1	4	1(7,69)^a		2(50,00)^a	3(75,00%)
	Site2	6	-		1(16,67)^a	1(16,67)
	Site3	14	-		0	0
	Total	24	1(04,17)		3(12,50)	4(16,67)

Values with the same letter in the same column are not significantly different (P > 0.05). Market 1: Petit marché; market 2: Dar es salam; market 3: Dolé; market 4: Wadata; market 5: Harobanda; Site 1: Gounti yéna; site 2: Gamkalé; site 3: Harobanda.

Table 3 shows the prevalence of Salmonella in the four (4) types of vegetables analyzed. Prevalence ranged from 25% (i.e. 3 out of 12 contaminated samples) in carrot samples to 8.33% (i.e. 1 out of 12 samples) in onion samples. Salmonella prevalence in lettuce samples was 15.38%, the second highest rate (10 out of 69 contaminated samples). Only one (1) strain of Salmonella pullorum was found in the lettuce samples. The differences did not differ significantly between vegetable types (P-value > 0.05).

Table 3

Prevalences of *Salmonella* strains identified by type of vegetable
Product type	Number of products	Salmonella serotypes n(%)			Total
Product type	Number of products	S. arizonae	S. pullorum	S. spp	Total
Lettuce	65	2(3,08)^a	1(1,54)^a	7(12,31)^a	10(15,38)
Tomato	35	1(2,86)^a		2(5,71)^a	3(8,57)
Carrot	12	1(8,33)^a		2(16,67)^a	3(25,00)
Onion	12	-		1(8,33)^a	1(8,33)
Total	124	4(3,26)	1(0,08)	12(9,68)	17(13,71)

Values with the same letter in the same column are not significantly different (P > 0.05).

Antibiotic resistance phenotype of Salmonella strains isolated from vegetables

Table 4 shows the antibiotic resistance levels of Salmonella strains isolated from vegetables. Antibiotic susceptibility testing of Salmonella strains (two (2) serotypes of Salmonella enterica (arizonae and pullorum) and 11 strains of Salmonella spp.) showed that all strains (100%) were resistant to at least one (1) antibiotic. Among the strains, Salmonella spp. had an antibiotic resistance rate of 42.45%. Salmonella arizonae is 31.25% resistant, and S. pullorum 25.00%. Differences between strains were not significant (P-value > 0.05). These serotypes gave very different responses from one antibiotic to another. Analysis of the results shows that Salmonella arizonae is 100% resistant to AMC, CRO and SXT. Salmonella pullorum is 100% resistant to AMC and CRO. Several resistance phenotypes were observed in Salmonella spp. strains, with the results showing that at least one Salmonella spp. strain is resistant to any antibiotic. The most common resistance phenotypes were CRO (100%), AMC (76.92%), CIP (46.15%), ATM (33.33%) and IPM (30.77). There were no significant differences between serotypes and antibiotics (P > 0.05).

Table 4

Antibiotic resistance phenotypes of *Salmonella* serotypes
	S. arizonae (n = 2)		S. pullorum (n = 1)		S. spp (n = 10)		P-value
	R	S	R	S	R	S	P-value
AMC	2(100)^a	0^a	1(100)^a	0^a	8(76,92)^a	2(23,08)^a	0,653
ATM	0^a	1(100)^a	0^a	0^a	1(33,33)^a	2(66,67)^a	0,504
CIP	0^a	2(100)^a	0^a	1(100)^a	5(46,15)^a	5(53,85)^a	0,330
CRO	2(100)^a	0^a	1(100)^a	0^a	10(100)^a	0^a	-
F	0^a	2(100)^a	0^a	1(100)^a	1(07,69)^a	9(92,31)^a	0,884
IPM	0^a	2(100)^a	0^a	1(100)^a	3(30,77)^a	7(69,23)^a	0,540
MRP	0^a	2(100)^a	0^a	1(100)^a	3(23,08)^a	7(76,92)^a	0,653
SXT	1(100)^a	0^a	0^a	1(100)^a	3(33,33)^a	7(66,67)^a	0,303
TET	0^a	2(100)^a	0^a	1(100)^a	2(23,08)^a	8(76,92)^a	0,653
AMC: Amoxicillin + Clavulanic Acid; ATM: Aztreonam; CIP: Ciprofloxacin; CRO: Ceftriaxone; F: Nitrofurantoin;IPM: Imipenem; MRP: Meropenem; SXT: Trimethoprim/Sulfamethoxazole; TET: Tetracycline

Figure 2 shows the prevalence of resistance in Salmonella strains for each marker antibiotic tested. Resistance varies from one antibiotic to another. The most frequently encountered resistances were to cephalosporins (CRO, 100%), penicillins (AMC, 100%), and fluoroquinolones (CIP, 46,15). On the other hand, low resistance was observed for F (7.69%). High sensitivities were observed for Nitrofurantoin (92.31%), Cyclins (TET, 76.92%), Monobactams (ATM, 76.92%), Carbapenems (IMP, 69.23%) and (MRP, 76.92%)) and Sulfonamides (SXT, 76.92%). No significant differences were observed between antibiotics (p > 0.05).

Figure 3 shows the antibiotic resistance rate of Salmonella strains isolated according to the type of vegetable analyzed. Salmonella strains isolated from lettuce samples are highly resistant to CRO (100%); AMC (88.90%), CIP (55.60%), TET (55.60%) and SXT (50.00%). The strain isolated from onion samples was resistant to AMC, ATM, CRO, F and TET. Strains isolated from tomato samples were resistant to CIP (100%), SXT (100%), CRO (100%), TET (100%) and ATM (25%). Finally, strains isolated from carrot samples are resistant to AMC (100%), CRO (100%) and IPM (33.30%). Differences in resistance from one type of vegetable to another were not significant (P-value > 0.05).

Multi-antibiotic resistance in Salmonella strains isolated from vegetables

The multidrug resistance of Salmonella strains is shown in Table 5. Approximately 93.75% of strains are multi-resistant. Multidrug resistance ranges from 2 to 6 antibiotics. Resistance to three (3) antibiotics is most common in these strains (around 37.50% for S. spp), followed by resistance to two (2) and three (3) antibiotics (around 12.50%). Only Salmonella spp. strains are resistant to four (4) and six (6) antibiotics (6.25% and 12.50% respectively). No significant differences were observed between strains (P-value > 0.05).

Table 5

Multi-resistance of *Salmonella* strains to antibiotics
Number Ab	Salmonella serotypes			Total	P-value
Number Ab	S. arizonae	S. pullorum	S. spp	Total	P-value
2Ab	2(12,50)^a	0^a,b	2(12,50)^b	4(25,00)	0,898
3Ab	0^a	0^a	6(37,50)^a	6(37,50)
4Ab	0^a,b	1(6,25)^b	1(6,25)^a	2(12,50)
5Ab	0^a	0^a	1(6,25)^a	1(6,25)
6Ab	0^a	0^a	2(12,50)^a	2(12,50)
Total	2(13,33)	1(6,25)	11(75,00)	15(93,75)

Ab: antibiotic

Prevalence of resistant strains by type of vegetable

Table 6 shows the prevalence of Salmonella resistant to at least one antibiotic, according to the type of vegetable analyzed. The prevalence of resistant strains is 10.48% in vegetables. The prevalences of the different types of vegetables analyzed were 25.00%; 10.77%; 8.33%; 5.71% in carrot, lettuce, onion and tomato, respectively. No significant differences were observed between vegetables (P-value>0.05). In terms of strains, the prevalence of Salmonella spp. represented 8.06% of the multidrug resistance observed in vegetables, and was the highest. Strains of S. arizonae and S. pullorum had prevalences of 1.61% and 0.80% respectively. Prevalences did not vary significantly according to vegetable type or serotype (P-value>0.05).

Table:6. Prevalences of resistant Salmonella strains according to vegetables tested

Vegetable types	Numbers of vegetables	Salmonella strains n(%)			Total	P-value
Vegetable types	Numbers of vegetables	S. arizonae	S pullorum	S. spp.	Total	P-value
Carrot	12	1(8,33)^a	0^a	2(16,67)^a	3(25,00)	0,413
Lettuce	65	1(1,54)^a	1(1,54)^a	5(7,69)^a	7(10,77)
Onion	12	0^a	0^a	1(8,33)^a	1(8,33)
Tomato	35	0^a	0^a	2(5,71)^a	2(5,71)
Total	124	2(1,61)	1(0,80)	10(8,06)	13(10,48)

Salmonella is an enteric bacterium and a major pathogen causing food poisoning. Salmonella species are the main causes of acute gastroenteritis in many countries, and salmonellosis remains a major public health problem worldwide, particularly in developing countries [12]. Salmonella is a highly pathogenic organism, and once it has been detected in foodstuffs, the latter are classified as unsafe for human consumption [21, 22]. In this study, Salmonella represented 33.34% (20/60) of the enterobacteriaceae species isolated from vegetables and irrigation water in market gardens. Salmonella prevalence in all vegetable samples was 13.71% (17/124), with carrot samples the most contaminated (25%), followed by lettuce samples ( 15.38% ) and tomato and onion samples (8.57% and 8.33% respectively). The rate of Salmonella contamination was 13.00% in market vegetable samples and 16.67% in market garden samples. Our results are lower than those obtained in a previous study on lettuce in Niger [8]. These authors reported a contamination rate of 36.94% for lettuce samples taken from the eight (8) regions of Niger, with the Niamey region being the most contaminated (i.e. 56%). This difference in results could be explained by the fact that half of the samples analyzed in their study were taken in Zango (Gounti yéna), which corresponds to our site 1, the most contaminated of our study sites. Salmonella prevalences in lettuce samples of 50% and 70.15% have been reported from Burkina Faso [12, 23] and 22% and 16% from Nigeria [24, 25]. These values are well above the 15.38% prevalence observed in the present study. The prevalence of Salmonella in all vegetable samples was 13.71%. This is significantly higher than the 2.6% prevalence reported from Côte d'Ivoire (Toe et al. [26]) or the 3 to 10% from the DRC (Mahangaiko et al., [27]).

Several studies have highlighted the presence of Salmonella in foods other than vegetables throughout the world, notably in Burkina Faso, in mutton (19%) Bawa et al. [28], sandwiches (17.7%) Nikiema et al. [10] and sesame samples (10.28%) Douamba et al. [29], in Ethiopia in cow's milk samples (10.5%), Abbunna et al. [30] and in Chad in chicken meat (26.66–41.66%) Abba et al. [31]. All these studies show high prevalences of Salmonella. The presence of Salmonella in vegetables is an indicator of faecal contamination [23]. This contamination can be explained, on the one hand, by traditional methods, storage temperature and inadequate personal hygiene of handlers after harvest and at the time of sale [32] and, on the other hand, by the use of waste water without any treatment, and the use of manure as fertilizer. The increased contamination of produce both at market and in the field highlights the debate on the importance of post-harvest effects, including poor hygiene and market management on fresh produce quality [33].

Two (2) serotypes of Salmonella enterica have been identified (Salmonella Arizonae and Salmonella Pullorum) and the remains are Salmonella spp. The ecological niche of Salmonella Arizonae is the intestine of cold-blooded animals such as lizards, geckos and frogs [34, 35]. The presence of this bacterium in vegetables could be explained by the abundance of these animals in market gardening sites. The presence of Salmonella Arizonae has also been reported in lettuce in Abidjan [35]. The presence of this bacterium in soils could be linked to the presence of these animals in large numbers on the three production sites due to the high level of insalubrity [35]. Salmonella Pullorum/Gllinarum is a poultry-specific agent (agent of avian typhoid) [36, 37, 38, 39, 40]. The ecological reservoir of this serotype is the poultry digestive tract [41]. The presence of S. pullorum in vegetables can be explained by the use of poultry manure (droppings) for soil fertilization. The use of poultry manure as fertilizer could explain the presence of Salmonella Gallinarum on cultivation sites [35]. This observation corroborates that of Beuchat and Ryu [42], who reported that manure composed mainly of poultry droppings is a major contributor to Salmonella Gallinarum contamination of lettuce.

With regard to resistant Salmonella serotypes, our results are also in line with those previously reported from Niger [12] and other countries, including Algeria (Sebaa et al. [43]), Burkina Faso (Bawa et al. [28]; Somda et al. [12]), Côte d'Ivoire (Toe [33]; Kouame [44]), Ethiopia (Gebremichael et al. [45]; Zewdu et al. [46]), Chad (Abba et al. [31]), Kenya (Ngai et al, 2021 [47]), Bangladesh (Nipa et al. [48]), Japan (Najwa et al. [49]), Pakistan (Razzaq et al. [50]), Morocco (Bouchrif et al. [1]; Allaoui et al. [51]; Baloch et al. [52]), Nigeria ( Raufu et al. [25]; Abakpa et al. [53]; Nwiyi et al. [54]). Antibiotic-resistant strains isolated from lettuce samples are likely to reflect contamination by strains possessing at least one antimicrobial resistance gene [35] and originating from healthy carriers among people handling these products [55]. On the other hand, this resistance could be explained by the indirect contamination of vegetables by fecal bacteria from animals during the manure fertilization process [8, 33]. The majority of growers used manure from livestock, particularly chicken, to fertilize vegetable soils [56]. In these farms, antibiotics are widely and abusively used to prevent and treat infections, and also to accelerate animal growth [57, 33]. The use of antibiotics can encourage the selection of resistant bacteria in enteric strains, which are then eliminated via their excrement [33, 58].

Salmonella spp. strains showed high rates of resistance to the antibiotics tested. The overall rate of antibiotic resistance in these strains was 40%. The highest levels of resistance were to cephalosporins (CRO, 100%), penicillins (AMC, 76.92%), fluoroquinolones (CIP, 46.15%), sulfonamides (SXT, 33.33%), carbapenems (IMP, 30.7%) and tetracycline (TET, 23.08%). These results corroborate the resistance of Salmonella to penicillin A (ampicillin, amoxicillin; amoxicillin + clavulanic acid) and cephalosporins (cefixime, ceftazidime) previously reported from Niger [8]. Such resistance is thought to be linked to the production of a cephalosporinase or extended-spectrum beta-lactamase (ESBL) by the strains in question [8]. The beta-lactam family acts on bacteria by inhibiting peptidoglycan synthesis after binding to a membrane protein receptor: PBP (Penicilllin Binding Protein) [59]. Bacterial strains resist these antibiotics by producing a beta-lactamase which inactivates the antibiotics [60] has shown that antibiotics belonging to the same class act by the same mechanism of action, and that target bacteria can resist them by an identical mechanism. In contrast to our results, [31] observed no resistance to CRO and CIP (0% each), but reported higher levels of resistance to IPM and SXT, 100% and 41.46% respectively. Other authors have reported low levels of Salmonella resistance to IPM (10%), CRO (5%) and CIP (5%) [12]. Salmonella resistance to IPC may be due to its use in both human and veterinary medicine [33, 57]. Indeed, various authors have reported IPC-resistant Salmonella in vegetables in Japan (Nawas et al. [61]) and Nigeria (Kemajou et al. [62]). The disparity observed between the results could be justified by the controlled use of CIP in both human and animal medicine [62]. This emergence of fluoroquinolone resistance is alarming, as these are the antibiotics of choice in veterinary medicine for the treatment of invasive salmonellosis. The consistency of resistance rates among the different molecules could be explained by the use of fluoroquinolones, which are more recent molecules in animal husbandry [33].

In various countries, several authors have reported rates of Salmonella resistance to TET, notably in Ethiopia (Fufa et al. [29]; Gebremichael et al. [44]), Burkina Faso (Bawa et al. [27]), Algeria (Alloui et al. [50]), Côte d'Ivoire (Toe, [32]) and Chad (Abba et al. [30]). These rates of resistance to TET are all higher than our observations. However, a low resistance rate of around 13% has been recorded in Algeria [6]. TET resistance is also alarming in developing countries, and may reflect contamination of raw vegetables by contaminated irrigation water or manure. In addition, these resistances can potentially be acquired through the food chain from human contamination arising from therapeutic practices [62, 63, 64]. Tetracyclines are old molecules widely used as first-line treatments. Resistance to these molecules is fairly well known, and is generally due to a plasmid gene that can be acquired quite easily by bacteria. Tetracyclines are broad-spectrum bacteriostatic antibiotics, active against both Gram + and Gram- bacteria [30, 50].

It should be noted that multi-resistance was observed in Salmonella strains (around 93.97% of strains). Numerous cases of multidrug resistance have been observed in these foods by other authors [6, 32, 47, 61, 65, 66]. At present, multidrug resistance is frequently observed in isolates from human clinical cases worldwide, and this characteristic is having an increasing impact on the empirical treatment of community infections [67, 68, 69, 70, 71].

This study highlighted the presence of resistant Salmonella strains in market gardens and markets in the urban community of Niamey. Resistance involved several antibiotic families. Salmonella resistance concerned cephalosporins (CRO), penicillins (AMC) and fluoroquinolones (CIP). Multi-resistance reached up to six (6) antibiotics (Salmonella strains). The presence of Salmonella strains resistant to several antibiotics in the vegetable samples analyzed testifies to a lack of good hygiene practices in market gardening and markets in the urban community of Niamey. Given the importance of fresh vegetables in the human diet and the high prevalence of multi-resistant strains recorded, it is important to implement a national health monitoring policy to control the circulation of multi-resistant strains.

API : Analytical Profile Index

ATCC: American Type Culture Collection

CASFM : Comité d’Antibiogramme de la Société Française de Microbiologie

EUCAST: European Committee on Antimicrobial Susceptibility Testing

MHA: Muller Hinton Agar

RVS : Rappaport Vassiliadis Soja

SPSS: Statistical Package for Social Sciences

SS : Salmonella –Shigella

TIAC: Toxi-Infection-Alimentaire Collective

Ethics approval and consent to participate

The vegetables samples were collected during the survey on the market garden sites and vegetables sales markets. All markets gardeners and vegetables sellers freely accepted, collaborated and gave their consent for collection of the vegetables samples. The international standards were used for sample analysis.

Consent for publication

Not Applicable

Availability of data and materials

The data that support the findings of this study are available from the corresponding author.

Competing interests

No competing of interest is related to this present manuscript.

Funding

No funding was received for this work. Everything was done on a personal funding.

Author's contributions

AAA carried out the sampling, sample collection and bacteriological analysis at the laboratory, HSD participated in sample collection, ASA and YC supervised laboratory analysis, RMM and AFM participated for data analysis, SHS and SH corrected and validated the analysis protocol and participated in drafting the manuscript. All authors read and approved the final version of the manuscript.

Acknowledgements

I thank the market gardeners and vegetables sellers of the urban community of Niamey for their well collaboration during the samples collection.

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No competing interests reported.

Prevalence and antibiotic resistance profile of Salmonella spp. strains isolated from the vegetable food chain in Niamey, Niger

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Version 1

Abstract

Background

Methods

Results

Conclusion

Figures

Introduction

Materials and methods

Sampling mode

Antibiotic resistance testing

Inoculum preparation

Seeding

1.1. Statistical analysis

Results

Ab: antibiotic

Prevalence of resistant strains by type of vegetable

Discussion

Conclusion

Abbreviations

Declarations

References

Additional Declarations

Status:

Version 1