Rotavirus Infection

Children’s Hospital, Zhejiang University School of Medicine

Jiang Mizu

What is Rotavirus ?Electron microscopic View of Rotavirus

What is Rotavirus ?Electron microscopic View of Rotavirus

Rotavirus particles in stool filtrate

Photo Credit : F.P. Williams, U.S. Environmental Protection Agency; Adapted from Parashar et al, Emerg Inft Dis 1998,14(4) 561–570

“Rota” in Latin

means wheel

First detected in April,

1973 by R Bishop

and team from a

biopsy of an

Australian child with

severe gastroenteritis.

What is Gastroenteritis?What is Gastroenteritis?

• Gastroenteritis is second only to respiratory illness as a cause of childhood morbidity worldwide.

• Gastroenteritis: nausea, vomiting, diarrhea, abdominal cramping, and fever occur 6-48h after exposure.

• Most gastroenteritis is caused by viral infection; bacterial, parasitic, and protozoal illnesses are less frequent but not uncommon.

What is diarrhea?

• Definition: increased total daily stool output (> 10g/kg/d) ， is usually associated with increased stool water content. – Loose consistency( 性状改变 ):watery diarrhea,

mucous diarrhea, bloody diarrhea– Increased stool frequency( 次数增多 )

• Duration – Acute (< 14 d)– Persistent (14 d to 2 m)– Chronic (> 2 m)

Rotavirus and diarrhea

• Viruses are the most common cause of acute gatroenteritis in developing and developed countries, such as rotavirus, astrovirus, adenovirus, and caliciviruses (Norwalk agent)

• Rotavirus, a 67-nm double-stranded RNA virus with at least eight serotypic variants, is the most common.

• As with most viral pathogens, rotavirus affects the small intestine, causing voluminous watery diarrhea without leukocytes or blood.

Characteristics of Rotavirus

Appearance of ‘hubbed wheel’ with spokes on electron microscopy

65-75nm diameter

Three concentric layers: double capsid (external and internal) and a core (nucleocapsid)

Characteristics of rotavirus

14 G types described Five common VP7 serotypes in humans (G1-G4, G9), but 5 others identified

VP7

VP4

Outer Capsid : 2 structural proteins involved in viral adsorption and penetration into epithelial cells.

- VP7 : glycoprotein or G protein

Characteristics of rotavirus

20 P genotypes described. One common VP4 genotype P[8] , but four others detected

VP7

VP4

Outer Capsid : 2 structural proteins involved in viral adsorption and penetration into epithelial cells.

- VP4 : protease - cleaved or P protein

Characteristics of rotavirus

Inner Capsid :VP6 most abundant major group specific antigen Seven groups of Rotavirus (A to G), but only 3 (A,B,C) pathogenic in humanGroup A strains account for >95% clinical infection.Human group A subdivided into

subgroup I and II

Characteristics of rotavirus

Core :Viral genome - composed of 11 segments of double- stranded RNA.

A 组 RV病毒基因组功能

基因片段 : 1 2 3 4 6 9 编码结构蛋白 : VP1 VP2 VP3 VP4 VP6 VP7 ( 核心 ) ( 核心 ) ( 核心 ) ( 外壳 ) ( 内壳 ) ( 外壳 区分 G 血清型 1-

14)

裂解 抗原区分 (A-G 组 )

VP5 VP8 A 组为Ⅰ ,Ⅱ亚群 （ P 血清型 1-44 ）

基因片段 : 5 7 8 10 11 编码非结构蛋白 :NS53 NS34 NS35 NS28 NS26 (NSP1 NSP2 NSP3 NSP4 NSP5)

From Kapikian AZ, Chanock RM. Rotaviruses. In: Fields Virology 3rd ed. Philadelphia, PA: Lippincott-Raven; 1996:1659.

Bacteria

Unknown Rotavirus

Calicivirus

Rotavirus

Escherichia coli

Parasites

Otherbacteria

Developed CountriesDeveloped Countries

Adenovirus

CalicivirusAstrovirus

AdenovirusAstrovirus

Unknown

Less Developed CountriesLess Developed Countries

Epidemiology Epidemiology

Distribution of pathogens reported to cause endemic/epidemic gastroenteritis and infantile vomiting and diarrhea

Distribution of pathogens reported to cause endemic/epidemic gastroenteritis and infantile vomiting and diarrhea

Rotavirus

• Rotavirus is the most common diarrheal pathogen in children worldwide1

• Globally more than 125 million cases of infantile gastroenteritis2

• 440,000 deaths per year mainly in less developed countries3

1Parashar et al, Emerg Infect Dis 1998 4(4) 561–570; 2Linhares and Bresee, Pan Am J Public Health 2000 8(5) 305–331; 3Parashar et al, Emerg Infect Dis 2003 9(5) 565–572

Estimated global distribution of 440,000 annual deaths in children <5 years old caused by rotavirus diarrhea3

1 dot = 1000 deaths

Introduction

0.0-0.1 0.6-0.9 1.0-1.9 2.0-3.40.2-0.5

Rotavirus-attributable mortality per 1000 children under 5 years of age

Global rotavirus disease burden

440,000 deaths

2.1 million inpatient visits

25 million outpatient visits

111 million episodes

1 : 293

1 : 60

1 : 5

1 : 1

Risk Events

Parashar et. al., Emerg Infect Dis, 2002

5% of all deaths in children < 5are due to rotavirus

Epidemiology

• RV infection is most common in winter months in temperate climates.

• Disease tends to be most severe in patients 3-24 mo of age, although 25% of the cases of severe disease occur after 2 yr of age, with serologic evidence of infection developing in virtually all children by 4-5 yr of age.

• Infants younger than 3 mo of age are relatively protected by transplacental antibody and possibly breast-feeding.

Transmission

• Transmission takes place both via the fecal-oral route by contaminated food, water or contaminated toy and from person to person.

• Outbreaks are common in children’s hospitals and child-care centers.

• Large quantities of virus are shed in the stool during the first week of infection.

• The virus survives for hours on hands and for days on environmental fomites.

Pathophysiology of rotavirus infection

• In viral infection, diarrhea is noninflammatory and results from an enteropathy in which the death of mature villus-tip cells (responsible for disaccharide digestion and monosaccharide absorption) causes an osmotic diarrhea due to the malabsorption of sugars.

Anatomy of Intestine

Normal transport of nutrients and electrolytes across the GI tract

• The glucose-sodium co-transporter, requires the presence of a sodium gradient across the brush border membrane (Na+- K+ATPase).

• The electroneutral NaCl- coupled pathway that involves the double exchange mechanism by the Na+-H+ exchanger (NHE) and the Cl-

HCO3- exchanger located at

the apical membrane.

Jejunum

Ileum

Colon

Pathogenesis

• Viruses that cause human diarrhea selectively infect and destroy villus tip cells in the small intestine.

• Biopsies of the small intestine show variable degrees of villus blunting and round cell infiltrate in the lamina propria.

• Pathologic changes may not correlate with the severity of clinical symptoms and usually resolve before that clinical resolution of diarrhea.

Pathogenesis

• In the small intestine, the upper villus enterocytes are differentiated cells, which have both digestive functions, such as hydrolysis of disaccharides, and absorptive functions, such as the transport of water and electrolytes via glucose and amino acid co-transports.

• The crypt enterocytes are undifferentiated cells, which lack the brush border hydrolytic enzymes and are net secretors of water and electrolytes.

Pathogenesis

• Selective viral infection of intestinal villus tip cells thus leads to – An imbalance in the ratio of intestinal fluid absorption

to secretion– Malabsorption of complex carbohydrates, particularly

lactose

• Rotavirus nonstructural protein may function as an enterotoxin.

Clinical manifestation of rotavirus infection

• The most common cause of acute noninflammatory gastroenteritis in infants and toddlers.

• Disease incidence peaks in the cooler fall and winter months (year-round).

• The peak age incidence is 3 to 24 months.• The incubation period for RV is 24-48 h.• Vomiting is the first symptom in 80-90% of pts,

followed within 24 h by low-grade fever and voluminous watery diarrhea and non bloody.

Clinical manifestation of rotavirus infection

• Diarrhea is usually self-limited, abating with 4-8 days but may last longer in young infants or immunocompromised pts.

• The white blood cell count is rarely elevated.• The stool does not contain blood or white cells.• Metabolic acidosis results from bicarbonate loss

in the stool, ketosis from poor intake, and lactic acidemia from hypotension and hypoperfusion.

Diagnosis

• In most cases, a satisfactory diagnosis can be made on the basis of clinical and epidemiologic features.

• Enzyme immunoassays, which offer approximately 90% specificity and sensitivity, are available for detection of group A RV in stool samples.

• More obscure cases can be studied by electron microscopy of stools, RNA electrophoresis, nucleic acid hybridization, and polymerase chain reaction assays.

• Specific identification of rotavirus in not required in every case, especially in outbreaks.

Stools studies

Findings ImplicationsGross examination Blood, mucus, pus Bacterial infection

Microscopic examination >5 WBC/hpf Bacterial infection

Chemical examination– Stool pH pH<5 Viral infection, Carbohydrate

malabsorption– Stool-reducing substances + Viral infection, Carbohydreate

malabsorption

Determination of hydration status

• The most common causes of dehydration in children are vomiting and diarrhea.

• Dehydration is classified by the percentage of total body water lost: mild (<5%), moderate (5-10%), and severe (>10%).

• A variety of signs and symptoms and ancillary date help to estimate the degree of dehydration.

Degree of dehydration

Clinical signs mild moderate severeDecrease in body weight 3-5% 5-10% 10-15%Skin Turgor normal decreased Markedly

decreased Color normal pale markedly decreased Mucous membranes Dry Mottled or gray;

parchedHemodynamic signs Pulse normal slight increase tachycardia Capillary refill 2-3 s 3-4 s >4 s blood pressure normal low perfusion normal circulatory collapseFluid loss urinary output mild oliguria oliguria anuria Tears Decreased absentUrinary indices specific gravity >1.020 anuria Urine [Na+] <20mEq/L anuria

Normal values of an arterial blood gas

pH 7.35-7.45PO2 80-100mmHgPCO2 35-45mmHg[HCO3

-] 20-28mmol/LSBE -3-3mmol/L Na+ 135-145mmol/LK+ 3.5-5.5mmol/LCl- 96-108mmol/LMg + 0.62-0.94mmol/L

Electrolyte Disorders

• Sodium disorders

Isotonic dehydration: [Na+] 130-150mmol/L

Hypotonic dehydration: [Na+] <130mmol/L

Hypertonic dehydration: [Na+] >150mmol/L• Potassium disorders

Hyperkalemia: [K+] >5.5mmol/L

Hypokalemia: [K+] <3.5mmol/L

Metabolic Acidosis

According to AG= [Na+] - ([HCO3-] + [Cl-])

• Normal type: 8-16mmol/L [HCO3- ]• Increased type: >16mmol/L [H+]

According to [HCO3-] • Mild [HCO3-] 18-13mmol/L• Moderate [HCO3-] 13-9mmol/L• Severe [HCO3-] <9mmol/L

Differential diagnosis

• In infancy, the differential diagnosis of acute gastroenteritis includes diarrhea associated with other infections such as urinary tract infection, otitis media, sepsis, and pneumonia.

• Depending on the geographic location, enteric adenoviruses or caliciviruses are the next most common viral pathogens in infants.

• Other potentially pathogenic viruses include astroviruses, corona-like viruses, Coxsackis viruses, and other small round viruses.

Norwalk virus

• The Norwalk agent, a calicivirus, is a small RNA virus that causes epidemic outbreaks of gastroenteritis

• Norwalk virus affects school-age children, adolescents, and adults.

• After a 24-h incubation period (range,12-72h), patients characterized by fever, vomiting, diarrhea, and often malaise and myalgias.

• Stools are loose, watery, and without blood, mucus, or leukocytes.

• The duration of symptoms is short, usually 12-60 hours.

Management

• The goals– Recognition, prevention, and treatment of dehydration– Maintenance of the nutritional status of the patients.

• Supportive treatment– Replacement of fluid and electrolyte deficits and

ongoing losses is critical, especially in small infants. – The use of oral rehydration fluid is appropriate in most

cases.

Management

• No role for antiviral drug treatment.• No benefit for antibiotics • No benefit from antiemetics or antidiarrheal

drugs, and there is a significant risk of serious side effects.

• Antimotility agents should be avoided• Probiotic organisms such as lactobacillus

species has been shown to reduce somewhat the intensity and duration of illness.

Oral rehydration solution (ORS) 配方

成分 （低渗 ) g/L （标准 ) g/L

氯化钠 2.6 3.5 无水葡萄糖 13.5 20

氯化钾 1.5 1.5

枸椽酸钠 2.9 2.9

总重量 20.5 27.9

低渗 ORS的常用配方

成份 标准ORS(mmol/L)

WHO

低渗配方 (mmol/L ）ESPGN*

低渗配方 (mmol/L)

钠 90 75 60

氯 80 65 45

无水葡萄糖 111 75 90

钾 20 20 25

枸橼酸盐 10 10 20

渗透压(mOsm/L)

311

2/3 张（ 200 ）245

½张（ 170）240

½ 张（ 150 ）

*由于含有更多的枸橼酸盐，口味相对更酸甜，容易被儿童接受

Management of dehyration

• The management priorities are stabilization of the patient’s vital signs, replenishment of the intravascular volume, and correction of electrolyte abnormalities

• A patient with mild or moderate dehydration can be orally rehydrated if willing and able to tolerate fluids.

• IV fluid restoration is necessary for severe dehydration, shock, or if the patient is unable to take fluids orally or has and altered metal status.

Mild and moderate dehydration

• Start oral rehydration with an electrolyte solution (ORS), giving a total volume of 30-50 ml/kg over a 3-4 h period.

• Failure of oral rehydration is an indication for IV rehydration.

• Once the initial rehydration is tolerated, resume giving milk to an infant, whether breast- or formula-fed.

• An infant who has large, watery stools can have the milk feedings supplemented with feedings of oral electrolyte solution.

Severe dehydration

• Initial intravascular restoration: Give fluid resuscitation with a 20-ml/kg bolus of normal saline (NS) over 20 to 30 min.– Assessing their fluid status (renal or cardiac disease, sickle cell

disease)– Obtain blood for electrolytes, blood urea nitrogen (BUN),

creatinine, glucose, and urinalysis

• Reevaluate the patient after the first bolus, if there is a poor response to the initial bolus, repeat the infusion.

• If there is a poor response to two IV boluses, consider other associated organ disease or the need for central venous monitoring before giving a third bolus.

定输液种类

• 等渗脱水：补 1/2 张液 (2:3:1 液或 1:1 液 )• 低渗脱水：补 2/3 张液 (4:3:2 液 )• 高渗脱水：补 1/5 ～ 1/3 张液 (1:4 液或 1:2 液 )• 临床上判断脱水性质有困难时，可按等渗脱水补

给。

定输液总量（ml/kg）

脱水程度 轻度 中度 重度

累积损失量 50 50 ～ 100 100 ～ 120

继续损失量 10 ～ 20 10 ～ 30 10 ～ 30

生理需要量 60 ～ 80 60 ～ 80 60 ～ 80

总输液量 90 ～ 120 120 ～ 150 150 ～ 180

Metabolic acidosis

• Intravenous NaHCO3 administration may be considered in the setting of moderate-severe metabolic acidosis.

• The dose (in mEq) of 5% NaHCO3 : Weight (kg) × Base deficit × 0.3

• The dose (in ml) of 5% NaHCO3: Weight (kg) × Base deficit × 0.5

• Given an half of calculation as a continuous infusion over 1 h.

• The effect of NaHCO3 in lowering serum potassium and ionized calcium concentrations must also be considered.

常用的溶液

一、非电解质溶液 ( 无张力溶液 )• 5%Glucose• 10%Glucose

二、电解质溶液• 10%NaCl 11 个张力 0.9% （等张）• 5%NaHCO3 3.5 个张力 1.4% （等张）• 11.2% 乳酸钠 6 个张力 1.87% （等张）

常用混合液的组成和配制

溶液种类

溶液内容( 份 )

盐 : 糖 :碱

10%氯化钠 (ml)

5%(10%)葡萄糖

(ml)

5%碳酸氢钠 /(11.2%乳酸钠 ) (ml)

张力

2:1 液 2:0:1 30 500( 水 ) 47(30) 1

4:3:2 液 4:3:2 20 500 33(20) 2/3

2:3:1 液 2:3:1 15 500 24(15) 1/2

1:1 液 1:1:0 20 500 ─ 1/2

1:2 液 1:2:0 15 500 ─ 1/3

1:4 液 1:4:0 10 500 ─ 1/5

Diet therapy

• Continue to feed milk to an infant with nondehydrating gastroenteritis, whether breast- or formula-fed.

• Intestinal lactase levels are reduced during rotavirus infection.

• Brief use of a lactose-free diet is associated with a shorter period of diarrhea but is not critical to successful recovery in most healthy infants.

• Reduce fat intake during recovery may reduce nausea and vomiting.

Prevention of rotavirus infection

• Good hygiene reduces the transmission of RV.• Good handwashing and isolation procedures

can help control nosocomial outbreaks.• Breast-feeding in prevention or amelioration of

RV infection may be small.• Serum antibodies are present, but their role in

prevention of subsequent attacks is unclear.• Repeat infections occur but are usually less

severe.

Rotavirus vaccination

• Mimic the immune response of natural rotavirus infection to:– Protect against moderate/severe disease– Prevent hospitalization and death– Reduce morbidity and socioeconomic burden– Attenuate severity and duration of illness

RV疫苗研究

• 单价 RV 口服活疫苗 : 只能对同株、同血清型有免疫保护，未能推广

• 多价 RV 疫苗： 1998 年在美国获准使用的轮状病毒四价疫苗含恒河猴 RV 株 G3 血清型和人轮状病毒 VP7 的 G1 、G2 和 G4 三个血清型重组减毒活疫苗（ RRV －TV) 对 G1-4血清型 RV 株感染有保护作用， 50 ％ -65 ％对重症，并发症达 75 ％－ 100％保护作用

RV疫苗研究

• 我国 RV 口服活疫苗获 2001 年国家药品监督管理局及 WHO 批准应用– 羔羊粪便分离羊轮状病毒，血清型 G10 （ VP7）， P12

（ VP4）型的 LLR 毒株经细胞培养传代减毒– 接种顺序 2 个月－ 3 岁，每年口服 1 次； 3 岁 -5 岁口

服 1 次即可– 剂量：每次 3ml （ 1 支）直接口服 ，也可掺入牛奶中，温度不宜高

– 效果： A 组特异性抗体 4 倍阳性增高率＞ 75 ％ ，婴幼儿重症 RV 腹泻保护率＞ 90 ％，保护期 1.5 年

– 副反应：未见与疫苗有关的严重副反应

2107 例 RV疫苗保护效果 ( ％ ) meta 分析（ 40篇）

对一般RV 腹泻 严重腹泻

不同种系 RV 疫苗 （ ％） （ ％）

恒河猴减毒活疫苗 (RRV,RR-TV) 42 60

人 RV 减毒活疫苗 (M-37,89-12) 68 79

牛 RV 减毒活疫苗

(RIT-4237,WC-3,W179-9, QHBRV) 40 62

其它如羊 RV 等减毒活疫苗

(BIRV1,Lamb RV) 35 /

Asian rotavirus health burden

Outcome

Without Vaccination

With Vaccination

Events

Averted

Deaths 170,960 61,640 109,320

(-64%)

Hospitalizations 1,914,891 535,603 1,379,288

(-72%)

Outpatient visits 13,503,114 5,838,838 7,664,275

(-57%)

CDC, unpublished data

Summary

• Rotavirus is the most common cause of vomiting and diarrhea in children worldwide

• An estimated 440,000 deaths occur annually, mainly in less developed countries

• Outer viral capsid proteins VP7 and VP4 define the serotype of rotaviruses (G and P type respectively), that are critical to vaccine development

• Four Group A rotavirus serotypes predominate globally: G1P[8], G2P[4], G3P[8] and G4P[8], with most disease attributable to G1[P8]

• Serotype G9 emerging as the fifth globally important serotype

• Vaccination is the most likely intervention to impact significantly on the global incidence of severe disease

Treatment and Prevention

• Main goals of treatment:• Control the diarrhea• Prevent vomiting• Control other symptoms• Maintain effective fluid and electrolyte balance with oral re-hydration therapy (ORT)• Replacement of fluid loss

• Prevention measures:• Breast feeding• Regular disinfection of play areas and toys• Frequent hand washing• Rigorous hygiene practices in hospital wards• Development of rotavirus vaccines

Kapikian A and Chanock R. Rotaviruses. In: Fields B et al, editors. Fields Virology, 3 rd ed; 1996: p. 1657–1708