7/27/2019 VERC-S-13-00416(1)

1/21

Veterinary Research CommunicationsALTERED ELECTROLYTE HOMEOSTASIS ASSOCIATED WITH EXPERIMENTALSALMONELLOSIS TREATED WITH AMOXICILLIN AND PEFLOXACIN

--Manuscript Draft--

Manuscript Number:Full Title: ALTERED ELECTROLYTE HOMEOSTASIS ASSOCIATED WITH EXPERIMENTAL

SALMONELLOSIS TREATED WITH AMOXICILLIN AND PEFLOXACIN

Article Type: Original ArticleKeywords: Salmonellosis, Electrolyte homeostasis, Pefloxacin, AmoxicillinCorresponding Author: Solomon Rotimi

Ota, NIGERIA

Corresponding Author SecondaryInformation:Corresponding Author's Institution:Corresponding Author's SecondaryInstitution:First Author: Solomon RotimiFirst Author Secondary Information:Order of Authors: Solomon Rotimi

David Ojo, PhD

Olusola Talabi

Elizabeth Balogun, PhD

Oladipo Ademuyiwa, PhDOrder of Authors Secondary Information:Abstract: In order to investigate the effects of salmonellosis and its chemotherapy on tissue

electrolyte handling experimental salmonellosis was induced by oral infection of rats

7/27/2019 VERC-S-13-00416(1)

2/21

electrolyte handling experimental salmonellosis was induced by oral infection of rats

7/27/2019 VERC-S-13-00416(1)

3/21

Abstract

In order to investigate the effects of salmonellosis and its chemotherapy on tissue electrolyte handling,

experimental salmonellosis was induced by oral infection of rats with Salmonella typhimurium. Infected animals

were treated intraperitoneally with pefloxacin (5.71 mg/kg body weight, 12hourly) and amoxicillin (7.14mg/kg

body weight, 8hourly) for 5 and 10 days respectively. Blood and organ electrolyte concentrations were

determined photometrically 24 hours and 5 days after the last drug administration. Salmonellosis resulted in

ionoregulatory disturbances in the tissues of the animals. This ionoregulatory disturbances were characterised by

hyponatremia, hypokalemia, hypocalcemia and hypomagnesemia with concomitant increase in the magnesium

concentration with erythrocytes (0.890.02mmol/L to 1.260.11mmol/L, p0.05) and heart (6.000.18mol/g to 6.750.32mol/g, p

7/27/2019 VERC-S-13-00416(1)

4/21

Introduction

Foodborne infections cause a major burden on public health services and represent significant costs in many

countries (Baeumner 2008).Salmonellosis, one of the most common and widely distributed foodborne disease,

is a collective description of a group of feco-oral diseases with symptoms which vary from severe enteric fever

to mild food poisoning caused by Salmonella species. Salmonella enteritidis is the most frequently isolated

serotype, causing gastroenteritis in most humans and systemic infection in a subpopulation (Cummings, et al.

2010,Rodenburg, et al. 2007). In experimental animal models, it has been reported to infect internal organs

following oral infection (Rodenburg, et al. 2007).

The initial host response following infection like salmonellosis is characterized by systemic vasodilation,

usually resulting in a hyperdynamic state with an elevated heart rate and normal or slightly decreased blood

pressure. In the subsequent phase, a hyperdynamic circulation related to the massive vasodilation is

characterized by haemodynamics as decreased or borderline blood pressure, hyperventilation as well as fever

(Matthews and Battezzati 1993,Khovidhunkit, et al. 2004). Although the pathophysiology of fever and the

mechanisms that control the body temperature are complex and are only partially known, the different

mediators, specific cations such as Na+, K

+, Ca

2+, and Mg

2+exert a clear effect on the body temperature

(Melesova, et al. 1993). Sitprija V. (2008) reported that artificially induced hyperthermia and hyperventilation

due to pyrexia produces catabolic changes that are similar to those observed during infection.

1

23

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

5/21

of antibiotics has been reported to be limited by the associated toxic effects which include electrolytes disorders.

Zietse et al. (2009) reported that these disorders can occur while renal function remains near to normal. Renton

(2005) also stated that the expression and activity of cytochrome P450 is altered during periods of infectious

disease and most of the major forms of this enzyme complex are affected in this manner that leads to a decrease

in the capacity of the liver and other organs to handle drugs. Thus, it is essential to study the impact of antibiotic

treatment on tissue electrolyte handling in experimental salmonellosis.

Materials and Methods

Chemicals

Pefloxacin was a product of Lek Pharmaceutical and Chemical Company, Ljubljana, Slovenia, while

amoxillin was obtained from Beecham Pharmaceuticals, Brentford, England. All other chemicals used in this

study were of the purest grade available and were obtained from British Drug House (BDH) Chemicals Limited,

Poole, England and Sigma-Aldrich, Missouri, U. S. A.

Bacteria strain

Salmonella enterica serovar Typhimurium strain TA98 (obtained from the Nigerian Institute of

Medical Research (NIMR), Yaba, Lagos, Nigeria) was grown for 48hours under static conditions in nutrient

broth (CMI, Unipath, UK). The organism was maintained on nutrient agar slant at 4C. Bacteria were

harvested from the slant, suspended in 100ml nutrient broth and allowed to grow at 37oC for 12 h (late

1

23

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

6/21

bacteria as described Hung and Wang (2004). Five animals that were not infected and received 0.2ml PBS orally

served as the normal control.

In previous Salmonella infection studies in rats (Havelaar, et al. 2001,Naughton, et al. 1996) it was established

that monitoring functional infection outcomes like Salmonella colonisation, translocation and infection induced

changes, follow-up of infected rats for at least 3 to 4 days is needed. Therefore the infected rats were left for

four days after which fresh faecal samples were collected to quantify Salmonella colonisation daily, as described

by van Ampting (2009).

Infected animals were divided into 7 groups of 5 animals each. While 1 group served as infection control group,

three groups were treated with amoxillin (7.14mg/kg body weight, 8 hourly) and the remaining three groups

with pefloxacin (5.71mg/kg body weight, 12 hourly) for 5 and 10 days respectively. The antibiotics were

constituted in 5% dextrose and were prepared fresh before each administration. They were administered in a

total volume of 0.1ml. Control animals received equivalent volume of 5% dextrose. All drug administration was

by the intraperitoneal route.

At the end of the antibiotic treatment and 5 days after the discontinuation of the antibiotics, blood was collected

from the animals into heparinised tubes by cardiac puncture under light ether anaesthesia after an overnight fast.

Liver, kidney, brain, heart and spleen were removed, rinsed in ice-cold saline, blotted dry and kept frozen at -

20oC.

1

23

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

7/21

Tris-HCl buffer and poured into Eppendorf tubes. The membrane suspensions were kept frozen in this latter

buffer at -20C.

A portion of the tissue (0.25g) was transferred to 2.0ml chilled 0.25mol/L sucrose and homogenized

and digested following the method of Chwelatiuk et al., (2006). In brief, 1ml of the whole homogenate (or 100l

in case of erythrocytes) was placed in a Pyrex tube with 2.0ml of concentrated nitric acid. After 20hours of

sample digestion at room temperature, 72% perchloric acid (0.5ml) was added and the mixture was heated at

150oC until a clear digest was obtained. The digest was then cooled to room temperature and analyzed for

sodium, potassium, calcium and magnesium content.

Analyses

Sodium and potassium concentrations in the plasma, erythrocyte ghost as well as the digests of

erythrocytes and the organs were determined by flame photometry (Mahboob, et al. 1996). Calcium and

magnesium concentrations in the samples were determined spectrophotometerically according to the procedures

described by Abam et al., (2008) using kits supplied by Quimica Clinical aplicada S. S., Amposta, Spain.

Statistical analysis

Data are expressed as meanS.E.M. One way analysis of variance (ANOVA) followed by Duncan

Multiple Range Test was used to analyse the results with p < 0.05 considered significant. Associations among

1

23

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

8/21

Results

In fig. 1 is the representation of the effects of pefloxacin and amoxicillin on the fecal bacteria load of

the rats. The oral infection of the rats with S. typhimurium resulted in salmonellosis as observed by the

apperance ofS. typhimurium in the feces of the rats. The concentration ofS. typhimurium cultured in the feces

was significantly (p

7/27/2019 VERC-S-13-00416(1)

9/21

decrease in hepatic potassium. After 10 days of pefloxacin and amoxicillin therapy, the level of calcium was

significantly (p

7/27/2019 VERC-S-13-00416(1)

10/21

Intensity of associations between fecal bacteria load and Na+, K

+, Ca

2+and Mg

2+concentrations in rats infected

with S. typhimurium was shown in Table 3. The fecal bacteria load was positively correlated with liver calcium

(r= 0.542, p

7/27/2019 VERC-S-13-00416(1)

11/21

Discussion

This study shows a profound appearance of Salmonella in the feces of the rats infected with S. typhimurium after

3days of infection. This is consistent with other studies (Rodenburg, et al. 2007,Islam, et al. 2000,Mehta, et al.

1999) in which gene expression changes in the rat colon upon colonization following oral Salmonella infection

was also reported. Rodenburg et al. (2007) reported that the earliest responses were on the mucosal transports

like chloride channel calcium activated 6, H+/K

+transport ATPase as well as oxidative stress related genes. The

alterations in transport of chloride through the apical border of the intestinal epithelial cells are known to be

followed by the efflux of water as well as electrolytes into the intestinal lumen, resulting in diarrhea (Sitprija

2008).

One consequence of the upregulation of the oxidative stress related genes upon colonization of the colon by

Salmonella is a concomitant increase in the synthesis of nitric oxide by the vascular endothelial cells (Henard

and Vazquez-Torres 2011). Izzo et al. (1998) reported that under pathophysiological conditions, nitric oxide

may be produced at higher concentrations that are capable of evoking net secretion thereby contributing to loss

of water and electrolyte. These pathophysiological changes in the intestine could have result in the

hyponatremia, hypocalcemia, hypokalemia and hypomagnesemia observed in these study. Similar fluid and

electrolyte alterations have been reported in other febrile conditions and infectious disease (Zaloga and Chernow

1987,Sankaran, et al. 1997,Chesney, et al. 1981).

1

23

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

12/21

insulin-sensitive cells, they are insulin and glucose responsive in ionic terms. Barbagallo et al. (1999) also

reported that magnesium depletion renders the erythrocytes more sensitive to oxidative damage; thus,

magnesium may itself possess antioxidant properties possible by affecting the rate of spontaneous dismutation

of superoxide ion (Afanas'ev, et al. 1995) which are abundantly produced by macrophages as part of the host

innate immune response to salmonellosis (Janssen, et al. 2003).

The data presented in this study showed that the degree of alterations in the electrolyte handling by the tissues

investigated is influenced by the fecal bacterial load. As part of the metabolic response of the host to the

invading bacterial, substrates are mobilized from the periphery to be utilized by the visceral tissues and immune

cells resulting in altered energy balance in the tissues (Hasselgren, et al. 1986). Studies have shown that

adjustments in glucose metabolism alter the cellular ionic regulation (Takahashi, et al. 1995,Dixit and Lazarow

1967,Duelli, et al. 1999). More than any other organ, the brain is entirely dependent on a continuous supply of

glucose from the circulation since glucose is almost the sole substrate for its energy metabolism. Sodium-

dependent isoform of glucose transporter proteins mediate the transport of glucose against a concentration

gradient. The driving force is the flux of sodium along an electrochemical gradient that is directed opposite to

the transport of glucose (Kumagai 1999). This transporter has recently been reported to be present in rat brain

(Yu, et al. 2010). This could, therefore, explain the observed salmonellosis-induced decrease in brain sodium

concentration.

1

23

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

13/21

of the Na+-Mg

2+exchanger (Romani 2011,Agus 1999), leading to increased cellular sodium (Romani 2011). If

there is insufficient magnesium for adequate ATP utilization, then the primarily extracellular cations Na

+

and

Ca2+

tend to leak into the cells and the primarily intracellular cations K+

and Mg2+

tend to leak out. This

leakiness disrupts proper gradients and cellular function (Romani 2011,Resnick 1992). It is also worthy of note

that the decrease in level of Mg2+

could allow accelerated free radical damage and this, coupled with elevation

of Ca2+

, is an indication of apoptotic damage in the hepatic tissue (Rosenstock, et al. 2004).

Similar to our observation in the liver, the level of sodium was also increased with a concomitant decrease in

potassium level in the heart and kidney of the rats following Salmonella infection. It was however interesting to

note that the level of calcium reduced. Although the reason for the decrease in calcium levels in these tissues is

not clearly known, we observed that is has a direct correlation with the fecal bacterial load. The bacterial

antigenic determinant is lipopolysaccharide (LPS). This LPS has been reported to reduce basal Ca2+

concentration and also impair calcium responses to both thrombin and bradykinin in rat mesandial cells

contractile cells that share many characteristics of vascular smooth muscle (Murray, et al. 1997).

Using bovine aortic myocytes, Murray et al. (1998), suggested that LPS-induced vascular contractile

impairment is at least partly mediated by an NO-dependent impairment of myocyte Ca2+

mobilization. This has

been implicated in the vasoconstrictor-resistant systemic vasodilation, as well as in the failure of multiple organ

systems; which is the characteristic of septic shock (Umans, et al. 1993).

1

23

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

14/21

Pyroglutamic acidosis during penicillin treatment has been reported (Peter, et al. 2006). During treatment with

fluoroquinolones, Kushner et al. (2001) observed hyponatremia and these was related to increased vasopressin.

We conclude that salmonellosis induced alterations in electrolytes homeostasis in rat tissues and these

alterations persisted through and beyond the cause of chemotherapy with pefloxacin and amoxicillin. The

insight provided herein, especially on the tissues as most studies have focused on plasma, should pave way for

further understanding the pathophysiology of salmonella infection and its treatment.

1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

15/21

References

Abam E, Okediran BS, Odukoya OO, Adamson I, Ademuyiwa O. Reversal of ionoregulatory disruptions in

occupational lead exposure by vitamin C. Environ Toxicol Pharmacol 26(3), 297-304, 2008.

Afanas'ev IB, Suslova TB, Cheremisina ZP, Abramova NE, Korkina LG. Study of antioxidant properties of

metal aspartates. Analyst 120(3), 859-62, 1995.

Agus ZS. Hypomagnesemia. J Am Soc Nephrol 10(7), 1616-22, 1999.

Baeumner A. Food pathogen and toxin detection. Anal Bioanal Chem 391(2), 449-50, 2008.

Barbagallo M, Dominguez LJ, Tagliamonte MR, Resnick LM, Paolisso G. Effects of glutathione on red bloodcell intracellular magnesium: relation to glucose metabolism. Hypertension 34(1), 76-82, 1999.

Basnyat B. Typhoid fever in the United States and antibiotic choice. JAMA 303(1), 34; author reply -5, 2010.

Bell DS. Inflammation, insulin resistance, infection, diabetes, and atherosclerosis. Endocr Pract 6(3), 272-6,

2000.

Brun-Buisson C, Doyon F, Carlet J, Dellamonica P, Gouin F, Lepoutre A, Mercier JC, Offenstadt G, Regnier B.

Incidence, risk factors, and outcome of severe sepsis and septic shock in adults. A multicenterprospective study in intensive care units. French ICU Group for Severe Sepsis. JAMA 274(12), 968-74,

1995.

Chesney PJ, Davis JP, Purdy WK, Wand PJ, Chesney RW. Clinical manifestations of toxic shock syndrome.

JAMA 246(7), 741-8, 1981.

Chwelatiuk E, Wlostowski T, Krasowska A, Bonda E. The effect of orally administered melatonin on tissue

accumulation and toxicity of cadmium in mice. J Trace Elem Med Biol 19(4), 259-65, 2006.Cummings PL, Sorvillo F, Kuo T. Salmonellosis-related mortality in the United States, 1990-2006. Foodborne

Pathog Dis 7(11), 1393-9, 2010.

Dixit PK, Lazarow A. Effects of metal ions and sulfhydryl inhibitors on glucose metabolism by adipose tissue.

Am J Physiol 213(4), 849-56, 1967.

Duelli R, Maurer MH, Heiland S, Elste V, Kuschinsky W. Brain water content, glucose transporter densities and

glucose utilization after 3 days of water deprivation in the rat. Neurosci Lett 271(1), 13-6, 1999.

Fenves AZ, Kirkpatrick HM, 3rd, Patel VV, Sweetman L, Emmett M. Increased anion gap metabolic acidosis as

a result of 5-oxoproline (pyroglutamic acid): a role for acetaminophen. Clin J Am Soc Nephrol 1(3),

441-7, 2006.

Fica A, Caorsi B, Piemonte P. [Dysenteric syndrome, acute renal failure and lethal septic shock associated toSalmonella enteritidis infection. Report of 3 cases]. Rev Med Chil 125(9), 1055-82, 1997.

Gauglitz GG, Toliver-Kinsky TE, Williams FN, Song J, Cui W, Herndon DN, Jeschke MG. Insulin increases

resistance to burn wound infection-associated sepsis. Crit Care Med 38(1), 202-8, 2010.

1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

16/21

Janssen R, van der Straaten T, van Diepen A, van Dissel JT. Responses to reactive oxygen intermediates and

virulence of Salmonella typhimurium. Microbes Infect 5(6), 527-34, 2003.

Jia K, Thomas C, Akbar M, Sun Q, Adams-Huet B, Gilpin C, Levine B. Autophagy genes protect against

Salmonella typhimurium infection and mediate insulin signaling-regulated pathogen resistance. Proc

Natl Acad Sci U S A 106(34), 14564-9, 2009.

Kawai S, Ikeda E, Sugiyama M, Matsumoto J, Higuchi T, Zhang H, Khan N, Tomiyoshi K, Inoue T,

Yamaguchi H, Katakura K, Endo K, Matsuda H, Suzuki M. Enhancement of splenic glucose

metabolism during acute malarial infection: correlation of findings of FDG-PET imaging with

pathological changes in a primate model of severe human malaria. Am J Trop Med Hyg 74(3), 353-60,2006.

Khovidhunkit W, Kim MS, Memon RA, Shigenaga JK, Moser AH, Feingold KR, Grunfeld C. Effects of

infection and inflammation on lipid and lipoprotein metabolism: mechanisms and consequences to the

host. J Lipid Res 45(7), 1169-96, 2004.

Kovalenko AN, Zhdanov KV, Volzhanin VM, Shishkin MK, Tokmakov VS, Karpov AV, Murachev AA,

Kondratenok VA. [Features of clinic, diagnostics and treatment of typhoid fever in young patients].Voen Med Zh 332(1), 33-9, 2011.

Kumagai AK. Glucose transport in brain and retina: implications in the management and complications of

diabetes. Diabetes Metab Res Rev 15(4), 261-73, 1999.

Kushner JM, Peckman HJ, Snyder CR. Seizures associated with fluoroquinolones. Ann Pharmacother 35(10),

1194-8, 2001.

Levine MM, Tacket CO, Sztein MB. Host-Salmonella interaction: human trials. Microbes Infect 3(14-15),1271-9, 2001.

Mahboob T, Mumtaz M, Haleem MA. Electrolyte content of serum, erythrocyte, kidney and heart tissue in salt

induced hypertensive rats. Life Sci 59(9), 731-7, 1996.

Makhnev MV. [Efficacy of various antimicrobial agents in the treatment of epidemic typhoid fever]. Antibiot

Khimioter 48(4), 27-34, 2003.

Matthews DE, Battezzati A. Regulation of protein metabolism during stress. Curr Opin Gen Surg, 72-7, 1993.

McGuinness OP. Defective glucose homeostasis during infection. Annu Rev Nutr 25, 9-35, 2005.

Mehta A, Singh S, Ganguly NK. Effect of Salmonella typhimurium enterotoxin (S-LT) on lipid peroxidation

and cell viability levels of isolated rat enterocytes. Mol Cell Biochem 196(1-2), 175-81, 1999.

Melesova LM, Vikhrova EM, Sokolov NM. [Physiologic standards of the parameters of peripheral blood andthermoregulation in laboratory animals]. Gig Sanit (2), 48-50, 1993.

Montel-Hagen A, Blanc L, Boyer-Clavel M, Jacquet C, Vidal M, Sitbon M, Taylor N. The Glut1 and Glut4

glucose transporters are differentially expressed during perinatal and postnatal erythropoiesis. Blood

1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

2122

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

17/21

Sitprija V. Altered fluid, electrolyte and mineral status in tropical disease, with an emphasis on malaria and

leptospirosis. Nat Clin Pract Nephrol 4(2), 91-101, 2008.

Takahashi S, Driscoll BF, Law MJ, Sokoloff L. Role of sodium and potassium ions in regulation of glucose

metabolism in cultured astroglia. Proc Natl Acad Sci U S A 92(10), 4616-20, 1995.

Umans JG, Wylam ME, Samsel RW, Edwards J, Schumacker PT. Effects of endotoxin in vivo on endothelial

and smooth-muscle function in rabbit and rat aorta. Am Rev Respir Dis 148(6 Pt 1), 1638-45, 1993.

van Ampting MT, Schonewille AJ, Vink C, Brummer RJ, van der Meer R, Bovee-Oudenhoven IM. Intestinal

barrier function in response to abundant or depleted mucosal glutathione in Salmonella-infected rats.

BMC Physiol 9, 6, 2009.Wiegand G, Rauch R, Hermann M, Apitz C, Hofbeck M, Heininger U. [Salmonella enteritidis infection

presenting with septic shock, renal failure and cutaneous manifestations]. Klin Padiatr 221(1), 41-3,

2009.

Yu AS, Hirayama BA, Timbol G, Liu J, Basarah E, Kepe V, Satyamurthy N, Huang SC, Wright EM, Barrio JR.

Functional expression of SGLTs in rat brain. Am J Physiol Cell Physiol 299(6), C1277-84, 2010.

Zaloga GP, Chernow B. The multifactorial basis for hypocalcemia during sepsis. Studies of the parathyroidhormone-vitamin D axis. Ann Intern Med 107(1), 36-41, 1987.

Zietse R, Zoutendijk R, Hoorn EJ. Fluid, electrolyte and acid-base disorders associated with antibiotic therapy.

Nat Rev Nephrol 5(4), 193-202, 2009.

1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

2122

23

24

25

26

27

28

29

3031

32

33

7/27/2019 VERC-S-13-00416(1)

18/21

Figure 1: Effects of pefloxacin and amoxicillin on fecal bacteria load and weight change of rats infected with S. typhimuri um

1

2

3

4

5

6

7

8

9

1011

12

13

14

15

16

17

18

19

20

21

2223

24

25

26

27

28

29

30

31

32

33

34

35

36

37

38

39

40

41

42

43

44

45

46

47

48

49

7/27/2019 VERC-S-13-00416(1)

19/21

Table 1: Effects of pefloxacin and amoxicillin on Na+, K

+, Ca

2+and Mg

2+concentrations in plasma, erythrocyte and

erythrocyte ghost of rats infected with S. typhimuri um

Plasma Normal

control

Infection

control

Pefloxacin

day 5

Pefloxacin

day 10

Pefloxacin

day 15

Amoxicillin

day 5

Amoxicillin

day 10

Amoxicillin

day 15

Na+ (mmol/l) 140.960.76a 118.384.10d 134.171.70c 129.081.07b 133.320.66c 132.470.01c 126.535.62b 137.571.70eK

+(mmol/l) 5.810.33

a5.050.09

b5.310.18

b5.060.29

b5.060.18

b5.540.19

b5.310.10

b4.690.57

c

Ca+

(mmol/l) 1.590.02a

1.160.02b

1.600.03a

1.290.03b

1.250.10b

1.490.04a

1.280.07b

1.290.01b

Mg+

(mmol/l) 1.260.05a

0.990.04b

0.880.15c

1.100.02d

0.870.02c

0.990.02b

1.030.01b

1.000.08b

Erythrocyte

Na+

(mmol/l) 3.790.00 3.140.05 3.580.20c 3.260.17c 3.530.12c 3.760.29a 3.130.20b 3.260.17c

K+

(mmol/l) 133.891.29b

118.971.74b

102.964.99c

129.340.50d

127.466.74d

108.990.84c

128.250.60d

125.193.66d

Ca + (mmol/l) 0.430.02a

0.330.02b

0.380.03c

0.360.02c

0.360.01c

0.320.02b

0.390.05c

0.330.04b

Mg+

(mmol/l) 0.890.02a

1.260.11c

0.880.16a

1.010.05b

1.340.10d

1.330.04c

1.020.15b

1.110.02e

Erythrocyte ghostNa

+(mol/g) 0.250.03

a0.260.03

a0.300.01

a0.280.03

a0.260.02

a 50.270.02

a0.240.04

a0.280.03

a

K+

(mol/g) 0.0300.001a 0.0290.001a 0.0290.001a 0.0290.001a 0.0290.001a 0.0300.001a 0.0290.001a 0.0290.001a

Ca+

(mol/g) 0.760.02a

1.380.02c

0.940.02d

0.920.02d

0.790.07a

0.930.02d

1.020.13b

0.880.07d

Mg + (mol/g) 0.210.01a 0.330.02b 0.350.02b 0.310.01b 0.370.04b 0.330.01b 0.310.01b 0.330.03b

Each value represents the meanS.E.M of 5 rats. Values within the same row with different superscripts are significantly different at

p

7/27/2019 VERC-S-13-00416(1)

20/21

Table 2: Effects of pefloxacin and amoxicillin on Na+, K

+, Ca

2+and Mg

2+concentrations in liver, kidney and brain of rats infected with

S. typhimuri um

Liver Normal

control

Infection

control

Pefloxacin day

5

Pefloxacin day

10

Pefloxacin day

15

Amoxicillin day

5

Amoxicillin day

10

Amoxicillin

15

Na+ (mol/g) 30.570.91a 40.761.29a 39.780.79a 38.322.08a 34.652.04a 34.247.80a 31.792.00a 34.730.87aK

+(mol/g) 65.964.65

a49.360.31 49.652.12 50.102.32 51.132.55 51.610.12 48.613.73 50.754.20

Ca+

(mol/g) 1.520.01a

1.890.11 1.890.16 1.670.03c

1.740.09c

1.920.09 1.660.02c

1.730.09c

Mg+

(mol/g) 6.410.29a

4.130.24 3.990.12 3.800.10c

4.320.16 4.490.04 4.000.23 4.320.09

Kidney

Na+

(mol/g) 27.551.51a

32.930.08c

29.590.79 30.570.91e 32.611.29c 32.610.00c 30.161.00b 33.670.79d

K+

(mol/g) 37.441.78a

29.641.41b

36.311.66c

32.900.98c

32.993.29c

34.471.65c

35.273.26c

32.992.17c

Ca + (mol/g ) 1.610.10a

1.340.07b

1.440.05c 1.400.10

c1.400.05

c1.420.12

c1.510.02

c1.390.08

c

Mg+

(mol/g) 7.700.20a

6.030.37b

7.410.57d

7.250.58c

7.120.38c

5.650.23b

6.440.47b

6.210.26b

BrainNa

+(mol/g) 30.490.08

a22.831.00

d28.780.21

e26.901.63

c 26.490.91

e 23.480.79

b23.481.51

b 31.630.79

a

K+

(mol/g) 55.974.70a

63.143.77 57.061.44c

60.475.12c

61.152.45c

49.654.31a

59.581.96c

57.802.89c

Ca+

(mol/g) 1.290.31a

1.420.11a

1.410.21a

1.520.11a

1.600.09a

1.270.03a

1.600.06a

1.660.07a

Mg + (mol/g) 6.610.19a 5.600.37 5.700.23 5.500.28 6.160.11c 5.740.23 5.530.30 5.910.12c

Spleen

Na+ (mol/g) 29.590.79a 26.091.63c 28.531.29a 26.901.00c 28.530.00a 28.532.23a 28.530.00a 24.460.01b

K+

(mol/g) 52.763.23a

54.841.69a

57.061.37a

54.102.41a

54.842.93a

58.542.12a

56.023.85a

55.221.71a

Ca + (mol/g) 0.920.09a 1.060.03a 1.060.11a 1.010.12a 0.940.05a 0.840.03a 1.010.12a 1.010.07a

Mg+

(mol/g) 4.100.03a

5.080.11 4.640.17 4.700.15 5.140.25 4.880.09 4.790.37 4.840.09Heart

Na+

(mol/g) 39.781.51a

45.650.82 45.900.79 43.212.08c

42.802.73c

43.860.79c

44.760.24b

41.821.51e

K+

(mol/g) 68.061.50a 65.812.25b 56.173.71d 66.404.79a 61.361.24c 59.880.94c 57.663.02e 62.461.18c

Ca+

(mol/g) 1.340.04a

1.030.05b

1.090.09 1.040.06b

1.200.03c

1.070.10b

1.160.05c

1.210.04c

Mg + (mol/g) 6.000.18a

6.750.32c 6.840.23

c 7.140.27c

7.400.30b

6.500.23c

7.360.29b

7.610.79b

Each value represents the meanS.E.M of 5 rats. Values within the same row with different superscripts are significantly different at

p

7/27/2019 VERC-S-13-00416(1)

21/21

Table 3: Intensity of association between fecal bacteria load and Na+, K

+, Ca

2+and Mg

2+concentrations in rats infected with

salmonella

Parameters Correlation coefficientFecal Bacteria load vs. Liver K

+-0.549

a

Fecal Bacteria load vs. Liver Ca2+

0.542a

Fecal Bacteria load vs. Liver Mg2+

-0.694a

Fecal Bacteria load vs. Kidney Na+

0.420a

Fecal Bacteria load vs. Kidney Ca2+

-0.333b

Fecal Bacteria load vs. Kidney Mg2+ -0.424a

Fecal Bacteria load vs. Brain Na+

-0.484a

Fecal Bacteria load vs. Brain Mg2+ -0.465a

Fecal Bacteria load vs. Spleen Mg

2+

0.433

a

Fecal Bacteria load vs. Heart Na

+0.474

a

Fecal Bacteria load vs. Heart K+

-0.339b

Fecal Bacteria load vs. Heart Ca2+

-0.536a

Fecal Bacteria load vs. Plasma Na+

-0.476a

Fecal Bacteria load vs. Plasma Mg2+

-0.493a

Fecal Bacteria load vs. Erythocyte K+ -0.638a

Fecal Bacteria load vs. Erythrocyte Ca2+

-0.353b

Fecal Bacteria load vs. Erythrocyte ghost Ca2+

0.540a

Fecal Bacteria load vs. Erythrocyte ghost Mg2+

0.557a

a Correlation is significant at p< 0.01

b Correlation is significant at p< 0.05

1

2

3

4

5

6

7

8

9

1011

12

13

14

15

16

17

18

19

20

21

2223

24

25

26

27

28

29

30

31

32

33

34

35

36

37

38

39

40

41

42

43

44

45

46

47

48

49