whitney stuart - infection with prrsv affects body protein deposition and alters amino acid...

Infection with porcine reproductive and respiratory syndrome virus (PRRSV) affects body protein deposition and alters amino acid metabolism in

growing pigs

W. Stuart-McGilvray1, T.E. Burkey2, N.K. Gabler3, K. Schwartz3, T. Dinh4, C.F.M. de Lange5, D. Klein1, J. A. Dawson1, and A.

Rakhshandeh1

1Texas Tech University, Lubbock, 2University of Nebraska, Lincoln, 3Iowa State University, Ames, 4Mississipi State University, United States, 5University of Guelph,

Guelph, Canada

Background

Immune System Stimulation (ISS)Pro-Inflammatory cytokines

Interleukin (IL) 1, IL-6, Tumor necrosis factor (TNF) α

Shifts in metabolismPhysiology and behavior

Reduced pigs’ productivity (e.g. lean growth)

Hormonal response Neurologic response

Reeds & Jahoor, 2001; Obled, 2003; Rakhshandeh and de Lange, 2011

BackgroundChronic disease & Growth Performance in 10 kg

pigs

Control Sick

Feed intake, g/d 645 590 - 9 %

Gain, g/d 543 396 - 27 %

Gain : Feed 0.84 0.66 - 22 %

Lean Growth, g/d 86.9 56.9 - 35 %

Disease is expensive !

Williams et al. 1997

Background

Altered AA useNewly synthesized proteins/metabolites

(e.g. acute-phase proteins)

Additional need for specific AA

Increased muscle protein degradation(e.g. reduced lean growth)

Qualitative and quantitative impacts on AA requirements

Reeds & Jahoor, 2001; Obled, 2003; Rakhshandeh and de Lange, 2011

Background

Sulfur AA (Met & Cys) Threonine (Thr) Tryptophan (Trp) Phenylalanine (Phe)

Leucine (Leu) Isoleucine (Ile) Valine (Val) Glutamine (Gln)

AA that putatively become important during ISS

Can increased supply of these AA reduce muscle protein mobilization?

Background

Various nutritional state Absorptive vs. post-absorptive

Various disease models Clinical vs. subclinical

Various age and gender Metabolic demand vs. dietary requirement Metabolic demands ≠ dietary requirements

Interaction between AA Various methods of measurement

Background Free AA concentration vs. flux Concentration

• Maintained by AA influx and efflux• Misleading

Plasma AA flux• Changes occur without change in concentration• Reflect modifications of AA metabolism• More accurate

Does a single measurement of AA concentration tell the whole story?

Holtrop et al., 2004; Waterlow, 2006; Rakhshandeh et al., 2011

Objective

To evaluate the effects of ISS induced by PRRSV infection on,

a) Plasma AA kinetics as a measure of AA utilization

b) Apparent Ileal digestibility of (AID) protein and whole-body protein deposition (PD)

Materials and Methods

General design 20 PRRSv-negative gilts (Initial

BW 9.34 ± 0.7 kg)

Individually housed in metabolism crates

Surgically catheterized Feed-restricted (550 g/d)

Corn-SBM based diet (ME 14 MJ/kg, SID Lysine 11.5 g/kg)

Immune system stimulation (IM injection of live field PRRSV)

Stuart et al., 2015

Jugular catheterization


Measurements: Serial blood collection (time 0 and every 48 h after ISS)

• Hematology• Blood chemistry • Viral load

Eye temperature • Infrared imaging technique

N-balance• 3 d pre & 3 d post-ISS

Apparent ileal digestibility (AID) • TiO2 as indigestible marker• Slaughter technique• Control group (n = 10)


Measurements Isotope Dilution Technique

• Single dose of [U-13C, U-15N] AA mixture (Lys, Met, Thr, Trp, Ile, Leu, Val, Phe, Gln)

• Blood samples: 0, 2.5, 5, 7.5, 10, 15, 20, 30, 45 min Isotopic enrichment (Tracer / tracee using GCMS)

• Plasma AA flux (efflux: Sum of disappearance of AA from plasma pool)


Isotope Dilution Technique

Plasma Pool

Dietary Protein Intake

Protein Breakdown

Catabolism Protein synthesis

+ = Influx

+ = EffluxHoltrop et al., 2004; Waterlow, 2006


Calculations Double-exponential model

Erich. P (t) = α1 exp (-α2t) +α3 exp(-α4t) AA flux (µmol/kg BW/h) AA pool size (µmol/kg BW)

Statistical Analysis Complete randomized block design MIXED procedure (SAS 9.2) Pig as random effect

Holtrop et al. 2004

Results: Viral Load & Eye Temperature

37.437.637.838.038.238.438.638.839.039.2

0 2 4 6 8 10

Tem

pera

ture

°C

Days post-inoculation

Eye Temperature

0

2000

4000

6000

8000

10000

12000

0 2 4 6 8 10

Viral Load

Gen

omic

copy

/µl

Thermal image of pig eye

Results: Blood Parameters

* Significantly different from pre-ISS period (*, P < 0.05; **, P < 0.01)

0.0

10.0

20.0

30.0

40.0

50.0

60.0

70.0

80.0

90.0

Blood Glucose,mg/dl

AnionGAP,mmol/L

Hemoglobin, g/dl BUN, mg/dl Creatinine, mg/L

ISS- ISS+

**** **

Results: Protein Digestibility and PD

0

10

20

30

40

50

60

70

ISS- ISS+

Prot

ein

Dep

ositi

on, g

/d

**

0

10

20

30

40

50

60

70

80

90

Control ISS+

AID

of D

ieta

ry P

rote

in, %

**


Results: Plasma free AA Flux

0

100

200

300

400

500

600

700

800

Lys Met Thr Trp Leu Ile Val Phe Gln

Flux

, µm

ol/k

g/h

ISS- ISS+

*

**P < 0.12


Results: AA Pool Size


0

10

20

30

40

50

60

70

80

90

Lys Met Thr Trp Leu Ile Val Phe Gln

Pool

Siz

e, µ

mol

/kg

BW

ISS- ISS+

*

P < 0.09

**

**

**

P < 0.11

P < 0.15

Results: Daily Urinary N Excretion & Creatinine

225

275

325

375

Dai

ly u

rinar

y N

exc

retio

n,

mg/

kg B

W/d

ISS-

ISS+

d1 d2 d3 d1 d2 d3

0123456789

10

ISS- ISS+

Cre

atin

ine,

mg/

L

**

*Significantly different from pre-ISS period (*, P < 0.05; **, P < 0.01)

Summary

Immune system stimulation (ISS) with PRRSV,• Reduced AID of dietary protein • Reduced PD• Increased MET and THR flux/utilization• Tended to decrease TRP flux/utilization• Did not affect utilization of other AA

Conclusions and Implications

Increased utilization of MET and THR Synthesis of immune system metabolites Increased catabolism

Increase MET and THR utilization may impact dietary requirements relative to other AA Further studies needed

AA kinetics provides a better assessment for AA utilization than a single measurement of concentration

Acknowledgments

• National Pork Board (NPB #13-082)

• Faculty, Staff, Graduate students and undergraduate students – Department of Animal and Food Sciences, Texas Tech University

• Friends and Family

Questions??