Occurrence, Genetic Diversities And Antibiotic Resistance Profiles Of Salmonella Serovars Isolated From Chickens

Figures & data

Figure 1 Agarose gel (1%) electrophoresis showing the amplification of 16 S rDNA of representative presumptive bacteria isolates colonising the gut of chickens obtained from North-west province, South Africa.

Notes: Lane 1 to 29 shows the amplification of 16 S region of presumptive Salmonella DNA isolates while lanes 14 = no template (negative control), lane 13 = Salmonella Typhimurium ATCC 14028 (positive control).

Abbreviations: M, DNA marker (1kb); bp, base pairs.

Table 1 Identity Of Presumptive Salmonella Isolates From Chickens In Mafikeng, South Africa

Isolate Number	Sample Source	Sequence	Assession Number	InvA	Serotype	Similarity (%)	Name of Organism
1	Broiler	Seq1	MG663456	+VE	AAG	98	Salmonella enterica subsp. enterica
2	Broiler	Seq2	MG663457	+VE	AAG	92	Salmonella enterica subsp. enterica
3	Broiler	Seq3	MG663458	+VE	AAG	97	Salmonella enterica subsp. enterica
4	Broiler	Seq4	MG663459	+VE	AG	92	Salmonella enterica ser. Weltevreden
5	Broiler	Seq5	MG663460	+VE	AG	92	Salmonella enterica ser. Chingola
6	Broiler	Seq6	MG663461	+VE	AG	92	Salmonella enterica ser. Arizonae
7	Broiler	Seq7	MG663462	+VE	AG	98	Salmonella enterica ser. Bovismorbificans
8	Layer	Seq8	MG663463	+VE	AAG	99	Salmonella enterica subsp. enterica
9	Layer	Seq9	MG663464	+VE	AAG	92	Salmonella enterica subsp. enterica
10	Layer	Seq10	MG663465	+VE	AG	97	Salmonella enterica ser. Typhimurium
11	Layer	Seq11	MG663466	+VE	AG	92	Salmonella enterica ser. Salamae
12	Layer	Seq12	MG663467	+VE	AG	99	Salmonella enterica ser. Houten
13	Layer	Seq13	MG663468	+VE	AAG	99	Salmonella enterica subsp. enterica
14	Indigenous	Seq14	MG663469	+VE	AAG	98	Salmonella enterica ser. Bareilly
15	Indigenous	Seq15	MG663470	+VE	AAG	99	Salmonella enterica subsp. enterica
16	Indigenous	Seq16	MG663471	+VE	AAG	98	Salmonella enterica subsp. enterica
17	Indigenous	Seq17	MG663472	+VE	AAG	92	Salmonella enterica subsp. enterica
18	Indigenous	Seq18	MG663473	+VE	AG	92	Salmonella enterica ser. Heidelberg
19	Indigenous	Seq19	MG663474	+VE	AG	92	Salmonella enterica ser. Arizonae
20	Indigenous	Seq20	MG663475	+VE	AAG	99	Salmonella enterica subsp. enterica
21	Indigenous	Seq21	MG663476	+VE	AG	99	Salmonella enterica ser. India
22	Indigenous	Seq22	MG663477	+VE	AG	97	Salmonella enterica ser. Crossness
23	Indigenous	Seq23	MG663478	+VE	AG	97	Salmonella enterica ser. Albany
24	Indigenous	Seq24	MG663479	+VE	AG	99	Salmonella enterica ser. Yovokome
25	Indigenous	Seq25	MG663480	+VE	AG	98	Salmonella enterica ser. Pullorum
26	Indigenous	Seq26	MG663481	+VE	AG	98	Salmonella enterica ser. Infantis
27	Broiler	Seq27	MG663482	+VE	AG	92	Salmonella enterica ser. Arizonae
28	Broiler	Seq28	MG663483	+VE	AG	99	Salmonella enterica ser. Heidelberg
29	Broiler	Seq29	MG663484	+VE	AAG	92	Salmonella enterica subsp. enterica
30	Broiler	Seq30	MG663485	+VE	AAG	92	Salmonella enterica subsp. enterica
31	Broiler	Seq31	MG663486	−VE	AAG	92	Salmonella bongori
32	Broiler	Seq32	MG663487	−VE	AAG	92	Salmonella bongori
33	Broiler	Seq33	MG663488	+VE	AG	92	Salmonella enterica ser. Arizonae
34	Layer	Seq34	MG663489	+VE	AAG	92	Salmonella enterica subsp. enterica
35	Layer	Seq35	MG663490	+VE	AG	92	Salmonella enterica ser. Wandsworth
36	Layer	Seq36	MG663491	+VE	AAG	92	Salmonella enterica subsp. enterica
37	Layer	Seq37	MG663492	−VE	AAG	92	Salmonella bongori
38	Layer	Seq38	MG663493	+VE	AG	92	Salmonella enterica ser. Kentucky
39	Layer	Seq39	MG663494	−VE	AAG	92	Salmonella bongori
40	Layer	Seq40	MG663495	+VE	AG	94	Salmonella enterica ser. Blockley
41	Layer	Seq41	MG663496	+VE	AG	98	Salmonella enterica ser. Newport
42	Layer	Seq42	MG663497	+VE	AG	98	Salmonella enterica ser. Typhimurium
43	Indigenous	Seq43	MG663498	−VE	AAG	98	Salmonella bongori
44	Indigenous	Seq44	MG663499	+VE	AAG	99	Salmonella enterica ser. Manchester
46	Indigenous	Seq46	MG663500	+VE	AAG	98	Salmonella enterica subsp. enterica
47	Indigenous	Seq47	MG663501	+VE	AG	99	Salmonella enterica ser. Typhimurium
48	Indigenous	Seq48	MG663502	+VE	AAG	92	Salmonella enterica subsp. enterica
49	Indigenous	Seq49	MG663503	+VE	AG	99	Salmonella enterica ser. Typhimurium
50	Indigenous	Seq50	MG663504	+VE	AG	85	Salmonella enterica ser. Typhimurium
51	Indigenous	Seq51	MG663505	+VE	AG	97	Salmonella enterica ser. Typhimurium
52	Indigenous	Seq52	MG663506	+VE	AAG	99	Salmonella enterica ser. Koessen
53	Indigenous	Seq53	MG663507	+VE	AAG	98	Salmonella bongori
54	Indigenous	Seq54	MG663508	−VE	AG	99	Salmonella enterica ser. Blegdam
55	Indigenous	Seq55	MG663509	+VE	AAG	92	Salmonella enterica subsp. enterica
Control 1	Control 1		ATCC 14028^TM	+VE	AG		Salmonella enterica ser. Typhimurium
Control 2	Control 2		MG663511	+VE		99	Escherichia coli O157:H7

Notes: Lane 1 = control 1 = Salmonella Typhimurium (positive control); control 2 = Escherichia coli (negative control) was an environmental strain.

Abbreviations: +VE, positive amplification; −VE, negative amplification; AAG, auto-agglutination against antisera; AG, positive agglutination.

Figure 2 Agarose gel (1%) electrophoresis showing the amplification of invA gene in representative presumptive Salmonella isolates in chickens obtained from Mafikeng, North-west province, South Africa.

Notes: Lane 1 = no template (negative control), Lane 2 = Salmonella Typhimurium ATCC 14028^TM (positive control); Lane 6 = Escherichia coli ATCC 259622^TM (negative control), lane 2 to 18 = positive amplification of invA gene at 286 bp, lane 6 = no amplification.

Abbreviations: M, DNA marker (100 bp); bp, base pairs.

Figure 3 Percentage occurrence of Salmonella isolates based on its source.

Figure 4 Neighbor joining method of phylogenetic tree based on partial 16S rDNA gene sequence, showing the phylogenetic relationships between Salmonella species and the most closely related strains from the genebank.

Notes: Numbers at the nodes indicate the ranks of bootstrap based on 1000 resampled data sets, and the cut-off points were placed at 70% for condensed tree. The scale bar indicates 0.5 base substitution per site. Salmonella enterica were set as the out-group. Sequences obtained in this study are denoted with a circle shape.

Table 2 Prevalence Of Antibiotic Resistance And Multi-Drug Resistance Of Salmonella Isolates From Chicken Sourced From Mafikeng, South Africa

Antibiotics	Resistance				Intermediate Resistance
	Total n= 55 (%)	Broilers n =14 (%)	Layers n =15 (%)	Indigenousn = 26 (%)	Total n = 55 (%)	Broilers n = 14 (%)	Layers n = 15 (%)	Indigenousn = 26 (%)
AMP	31 (56)	5 (36)	4 (27)	21 (81)	10 (18)	4 (29)	5 (33)	1 (4)
OXT	38 (69)	12 (86)	9 (60)	17 (65)	9 (16)	1 (7)	4 (27)	4 (15)
CIP	17 (31)	3 (21)	5 (33)	10 (39)	11 (20)	5 (36)	3 (20)	2 (8)
STR	52 (95)	13 (93)	15 (100)	25 (96)	2 (4)	1 (7)	0 (0)	1 (4)
GCN	2 (4)	0 (0)	1 (7)	1 (4)	1 (2)	1 (7)	0 (0)	0 (0)
SXT	43 (78)	9 (64)	12 (80)	22 (85)	0 (0)	0 (0)	0 (0)	0 (0)
C30	7 (13)	2 (14)	3 (20)	2 (8)	8 (15)	4 (29)	2 (13)	2 (8)
ERY	55 (100)	14 (100)	15 (100)	26 (100)	0 (0)	0 (0)	0 (0)	0 (0)
NOR	27 (49)	7 (50)	6 (40)	14 (54)	5 (9)	1 (7)	0 (0)	4 (15)
KF	19 (36)	8 (57)	3 (20)	8 (31)	15 (27)	0 (0)	7 (47)	8 (31)
NAL	26 (47)	8 (57)	9 (60)	9 (35)	5 (9)	0 (0)	1 (7)	4 (15)
Number of MAR phenotypes
0	0 (0)	0 (0)	0 (0)	0 (0)
1	0 (0)	0 (0)	0 (0)	0 (0)
2	1 (2)	1 (7)	0 (0)	0 (0)
3	4 (7)	0 (0)	0 (0)	4 (15)
4	13 (24)	4 (29)	4 (27)	5 (19)
5	13 (24)	3 (21)	1 (4)	9 (35)
6	4 (7)	2 (14)	0 (0)	2 (8)
7	8 (15)	3 (21)	0 (0)	5 (19)
8	9 (17)	1 (7)	0 (0)	8 (31)
9	3 (6)	1 (7)	0 (0)	2 (8)

Notes: Ampicillin (AMP) 10 μg; chloramphenicol (C30) 30 μg; nalidixic acid (NAL) 30 μg; streptomycin (STR) 10 μg; oxy-tetracycline (OXT) 30 μg; cephalothin (KF) 30 μg; erythromycin (ERY) 15 μg; sulphamethoxazole/trimethoprim (SXT) 22 μg; gentamycin (GCN) 10μg; ciprofloxacin (CIP) 10 μg; norfloxacin (NOR) 10 μg.

Abbreviation: MAR, multiple antibiotic resistance.

Table 3 Multiple Antibiotic Resistance Index And Phenotype Pattern Of Salmonella Isolates From Chickens In Mafikeng, South Africa

Salmonella Strains	Sample Source	No. Of Strains	Antibiotics Resistance Profiles	MARIndex
Salmonella bongori	Indigenous	1	AMP-CIP-STR-SXT-ERY	0.45
	Indigenous	1	AMP-OXT-CIP-STR-SXT-ERY-NOR-KF	0.72
	Broilers, layers	2	OXT-CIP-STR-ERY-NOR-NAL	0.54
	Indigenous	1	OXT-CIP-STR-SXT-ERY-NOR-NAL	0.63
	Broiler	1	OXT-STR-ERY-NAL	0.36
Salmonella enterica subsp. enterica	Indigenous	1	OXT-STR-ERY-NOR	0.36
	Broiler	1	OXT-STR-SXT-ERY	0.36
	Layer	1	OXT-STR-SXT-ERY-NAL	0.45
	Indigenous	1	OXT-STR-SXT-ERY-NOR	0.45
	Indigenous	1	AMP-OXT-CIP-STR-SXT-ERY-NOR	0.63
	Indigenous	1	AMP-OXT-CIP-STR-SXT-C30-ERY	0.63
	Broiler	1	AMP-SXT- ERY-KF	0.36
	Indigenous	1	AMP-SXT-ERY	0.27
	Indigenous, layer	2	AMP-STR-SXT-ERY	0.36
	Broiler	1	AMP-STR-SXT-ERY-KF	0.45
	Indigenous	1	AMP-STR-SXT-ERY-NOR	0.45
	Broiler	1	OXT-CIP-STR-SXT-ERY-NOR-NAL	0.63
	Broiler	1	OXT-STR-ERY-KF	0.36
	Indigenous	1	OXT-STR-ERY-NAL	0.36
	Layer	1	OXT-STR-ERY-NOR	0.36
	Indigenous	1	STR-SXT-C30-ERY-KF-NAL	0.54
Salmonella enterica ser. Arizonae	Indigenous	1	AMP-STR-ERY	0.27
	Broiler	1	OXT-CIP-STR-SXT-ERY-NOR-NAL	0.63
	Broiler	1	OXT-STR-ERY-NOR-KF-NAL	0.54
	Broiler	1	OXT-STR-SXT-ERY-KF	0.45
Salmonella enterica ser. Typhimurium	Indigenous	1	AMP-OXT-CIP-STR-SXT-ERY-NOR-KF	0.72
	Indigenous	1	AMP-OXT-CIP-STR-SXT-ERY-NOR-NAL	0.72
	Indigenous	1	AMP-OXT-STR-SXT-ERY-NOR-KF-NAL	0.72
	Indigenous	1	AMP-STR-GCN-SXT-C30-ERY-NOR-NAL	0.72
	Indigenous	1	AMP-OXT-CIP-STR-SXT-ERY-NOR-KF-NAL	0.81
	Indigenous	1	STR-SXT-ERY-KF	0.36
	Indigenous	1	STR-SXT-ERY-KF-NAL	0.45
Salmonella enterica ser. Heidelberg	Broiler	1	AMP-OXT-STR-SXT-C30-ERY-KF	0.63
	Indigenous	1	OXT-STR-ERY	0.27
Salmonella enterica ser. Manchester	Indigenous	1	AMP-CIP-STR-SXT-ERY-NOR-NAL	0.63
Salmonella enterica ser. Weltevreden	Broiler	1	AMP-OXT-STR-SXT-C30-ERY-NOR-KF-NAL	0.81
Salmonella enterica ser. Salamae	Indigenous	1	OXT-STR-SXT-ERY-NOR-NAL	0.54
Salmonella enterica ser. Blegdam	Indigenous	1	AMP-OXT-CIP-STR-SXT-ERY-NOR-KF	0.72
Salmonella enterica ser. Pullorum	Indigenous	1	AMP-OXT-CIP-STR-SXT-ERY-NOR-KF-NAL	0.81
Salmonella enterica ser. Koessen	Indigenous	1	AMP-OXT-STR-SXT-ERY	0.45
Salmonella enterica ser. India	Indigenous	1	AMP-OXT-STR-SXT-ERY	0.45
Salmonella enterica ser. Crossness	Indigenous	1	AMP-OXT-STR-SXT-ERY	0.45
Salmonella enterica ser. Bovismorbificans	Broiler	1	AMP-OXT-STR-SXT-ERY-NOR-KF-NAL	0.72
Salmonella enterica ser. Infantis	Indigenous	1	AMP-OXT-STR-SXT-ERY-NOR-NAL	0.63
Salmonella enterica ser. Albany	Indigenous	1	AMP-STR-ERY	0.27
Salmonella enterica ser. Newport	Layer	1	AMP-STR-SXT-ERY	0.36
Salmonella enterica ser. Yovokome	Indigenous	1	AMP-STR-SXT-ERY-KF	0.45
Salmonella enterica ser. Wandsworth	Indigenous	1	OXT-CIP-STR-SXT-ERY-NOR-NAL	0.63
Salmonella enterica ser. Kentucky	Layer	1	OXT-STR-ERY-NAL	0.36
Salmonella enterica ser. Blockley	Indigenous	1	STR-SXT-ERY-KF	0.36
Salmonella enterica ser. Chingola	Broiler	1	OXT-STR-ERY-NOR-NAL	0.45
Salmonella enterica ser. Bareilly	Indigenous	1	OXT-STR-SXT-ERY-NAL	0.45
Salmonella enterica ser. Houten	Indigenous	1	AMP-OXT-CIP-STR-S300-ERY-NOR-NAL	0.72

Note: Ampicillin⁵³ 10 μg; chloramphenicol (C30) 30 μg; nalidixic acid (NAL) 30 μg; streptomycin (STR) 10 μg; oxy-tetracycline (OXT) 30 μg; cephalothin (KF) 30 μg; erythromycin (ERY) 15 μg; sulphamethoxazole/trimethoprim (SXT) 22 μg; gentamycin (GCN) 10μg; ciprofloxacin (CIP) 10 μg; norfloxacin (NOR) 10 μg.

Abbreviation: MAR, multi-antibiotic resistance index.

Table 4 The Percent Distribution Of Resistant Salmonella Isolates Based On Sample Source And Antibiotic Resistant Clusters

Type of Chicken/Source	Cluster I N = 18	Cluster II N = 17	Cluster III N = 20
Broiler	3 (16.6%)	8 (47.1%)	3 (15%)
Layers	4 (22.2%)	4 (23.5%)	7 (35%)
Indigenous	11 (73.3%)	5 (29.4%)	10 (50%)

Table 5 Pearson Correlations Between Percent Antibiotic Resistance Of Salmonella Isolates And Source

Antibiotic Resistance	Total Resistance (%)	Broilers (%)	Layers (%)	Indigenous (%)
Total resistance (%)	1
Broilers (%)	0.925**	1
Layers (%)	0.931**	0.874**	1
Indigenous (%)	0.959**	0.800**	0.825**	1

Note: **Correlation is significant at the 0.01 level (2-tailed).

Figure 5 Dendrogram of antibiotic resistance profiles of Salmonella strains isolated from chickens in Mafikeng, South Africa, using cluster analysis.

Notes: Seq represents sequence numbering of Salmonella isolates from different types of chickens in Mafikeng. The tree was constructed using the Ward’s method and Euclidean distances in the Statistica version 7 software (Statsoft US).

Abbreviation: Seq, sequences of Salmonella isolates from chickens.

Guariguata L, Whiting DR, Hambleton I, Beagley J, Linnenkamp U, Shaw JE. Global estimates of diabetes prevalence for 2013 and projections for 2035. Diabetes Res Clin Pract. 2014;103(2):137–149. doi:10.1016/j.diabres.2013.11.00224630390

 PubMed Web of Science ®Google Scholar