Rotavirus Strains

Rotavirus-Synopses

The major antigenic properties of rotaviruses—group, subgroup, and serotype—are determined by the viral capsid proteins. Rotavirus has seven major groups (A-G); most human strains belong to group A, although groups B and C have occasionally been associated with human illness. The product of the 6th gene of group A rotaviruses encodes VP6, the most abundant viral protein, which is the major determinant of group reactivity, the target of common diagnostic assays, and contains the antigen used to further classify rotaviruses into subgroups I and II. The outer capsid proteins, VP7, the glycoprotein or G-protein (encoded by gene 7, 8, or 9, depending on the strain), and VP4, the protease-cleaved or P-protein (encoded by gene segment 4), determine the serotype specificity and form the basis of the binary classification (G and P type) of rotaviruses. Both G and P proteins induce neutralizing antibodies and may be involved in protective immunity.

Fourteen G serotypes of rotavirus, 10 of which occur in humans, have been defined by cross-neutralization studies with polyclonal animal serum samples; these serotypes correlate with antigenic specificities of the VP7 glycoprotein. The characterization of P serotypes has been difficult because adequate reagents are not available. Eight P serotypes of human rotaviruses have been characterized. Additional VP4 gene variants have been identified, so ultimately the number of P serotypes may exceed 20. Theoretically, 80 different strains of rotavirus could result from various combinations of the known 10 G and 8 P serotypes of human rotaviruses. For vaccine development purposes, it is fortunate that only four common strains (P[8]G1; P[8]G3; P[8]G4; and P[4]G2) of rotavirus predominate globally (Figure 4) (11). However, the prevalence of rotavirus strains varies considerably from one geographic area to another, and unusual strains are common in several developing countries (e.g., unusual P[6] strains, including those with serotype G9 specificity, accounted for 9.5% of all rotaviruses from a multicenter collection in India)

In infants and young children, neutralizing antibodies directed primarily against the G serotype of the infecting strain (homotypic response) develop after primary infection with rotavirus (18). Repeat rotavirus infections elicit both a homotypic and heterotypic (against strains with different G serotypes) antibody response. Protection against rotavirus diarrhea correlates with serum antibody titers following natural infection of young children, and infected children are more protected against reinfection with similar rather than different G serotypes. A protective role of placentally transferred maternal antibody among infants < 3 months of age has also been speculated since rotavirus disease is uncommon in this age group. However, serum neutralizing antibody responses among vaccine recipients have sometimes correlated poorly with protection from disease; therefore, the exact role of serum antibody in protection against disease remains unclear.

Genome Variability

Rotavirus is a highly variable virus even within the subset of those that are infective to humans. [4] Rotaviruses are usually categorized into seven groups A-G, with subgroups I and II based on the VP6 protein. [8] Within these groups, A, B, and C are infective to Humans. [19]

Rotavirus is further categorized into G and P serotypes. The G serotype is specified by the glycoprotein VP7 of the outer capsid, which is coded by viral genes 7, 8, and 9. [8] The P serotype is specified by protein VP4, also on the outer capsid. It is a protease cleaved protein coded by gene 4 of the virus genome. [8] The most common G serotypes currently are G1, G2, G3, G4, and G9, with G1 being most prevalent and G9 the fastest emerging worldwide. [7] [8] [10] [12] Common P serotypes are P1a, P1b, and P8. [13] [6] [4]

 rotastarins

Rotavirus Vaccines:

 

Monovalent “Jennerian” Vaccines

Initial development of rotavirus vaccines was based on the Jennerian approach, which involved the use of a live, attenuated, antigenically related virus derived from a nonhuman host (21). This approach was prompted by studies indicating that animal and human rotaviruses shared a common group antigen and that experimental animals immunized with animal strains of rotavirus had a significantly lower risk for illness and viral shedding when subsequently challenged with human rotaviruses. Furthermore, neutralizing antibodies to human rotavirus serotypes in the animal models indicated the potential for cross-protection.

Bovine Vaccines

The first two Jennerian vaccines were developed with bovine rotavirus strains RIT4237 and WC3. The WC3 strain was passaged in cell culture less than RIT4237 and was developed because of concern that excessive passaging of the RIT vaccine might cause overattenuation and diminished efficacy. RIT4237 and WC3 were nonreactogenic and immunogenic when administered to infants 2 to 18 months of age. However, the protection conferred by both vaccines varied greatly in efficacy studies, 0% to 76% against any rotavirus diarrhea and 0% to 100% against severe disease (2232). A well-defined correlate of protection was not identified, and reasons for the variable efficacy were unknown, although late age at vaccination, timing of vaccination with respect to the onset of the rotavirus season, and variations in the strength and number of doses of the vaccine were proposed as contributing factors. Both vaccines performed less well in developing than in industrialized countries, possibly because of interference by other enteropathogens or inadequate surveillance during follow-up.

 

Rhesus Vaccine

The third Jennerian vaccine was developed with rhesus rotavirus strain MMU18006, which shares neutralization specificity with human rotavirus G3 strains. Besides sharing antigenic specificity with an epidemiologically important human rotavirus serotype, MMU18006 was suitable for vaccine development because it grew efficiently in cell culture. As in the bovine rotavirus-based vaccines, MMU18006 was safe and immunogenic, although in some trials, one third of infants became febrile 3 to 4 days after vaccination. The reactogenicity of MMU18006 was particularly high in two studies in Finland and Sweden in which 64% and 79% of infants, respectively, became febrile. Most children with febrile responses were >5 months of age; lack of passively transferred maternal antibody might have contributed to the high reactogenicity of the vaccine. As in the RIT4237 and WC3 vaccines, the protective efficacy of MMU18006 in field trials was quite variable, 0% to 60% against any rotavirus diarrhea and 0% to 85% against severe rotavirus diarrhea (3339).

Reassortant “Modified Jennerian” Vaccines

The greatest efficacy of MMU18006 was observed in a Venezuelan trial in which the rotavirus strain circulating in the community (G3) was the same serotype as the vaccine strain, which suggested that serotype-specific immunity against each of the epidemiologically important strains of human rotaviruses may be required for maximum protection. Similar observations in vaccine challenge cross-protection studies in animals initiated the development of vaccines that used a modified Jennerian approach in which animal-human reassortants expressing VP7 proteins of serotypes 1 through 4 were used as the immunogens.

 

Rhesus-Human Reassortant Vaccines

Rhesus-human reassortants were generated by coinfecting cell cultures with rhesus rotavirus (RRV) strain MMU18006 (G serotype 3) and human rotavirus strains D (G serotype 1), DS-1 (G serotype 2), and ST3 (G serotype 4). Selection pressure (induced by the addition of neutralizing antibody to VP7 of RRV) produced reassortant strains D x RRV, DS-1 x RRV, and ST3 x RRV, each of which possessed the VP7 gene from HRV serotype 1, 2, or 4 and the other 10 genes from RRV (Figure 6) (40). Because vaccines made from the individual reassortants were safe and immunogenic, RRV-TV was developed incorporating each of the three reassortants and MMU18006 to provide coverage against the four common VP7 serotypes of rotavirus.

RRV-TV testing was initiated at an inoculum of 104 PFU of each of the four viruses (i.e., at 4 x 104 PFU) and completed at 105 PFU, the dose submitted for licensure (i.e., at 4 x 105 PFU). In most trials, vaccine was administered orally in three doses separated by at least 3 weeks to optimize the immune response to the component antigens; immunization was completed by the age of 6 to 7 months. Because the vaccine virus strains are acid labile, they are administered with 2.5 ml of citrate-bicarbonate buffer.

 

Bovine-Human Reassortant Vaccines

 

Bovine-human reassortant rotavirus vaccines include a tetravalent WC3 rotavirus reassortant vaccine with genes coding for the VP7 of three major serotypes of rotavirus (G1, G2, and G3) and W179-4, a human VP4 reassortant with P[8] specificity. Theoretically, this vaccine should induce antibodies broadly reactive to the three common serotypes of rotavirus sharing P [8] specificity, thereby increasing the protective efficacy of this vaccine. In an efficacy trial of a three-dose regimen of the WC3 reassortant vaccine, protection was 67% against all rotavirus diarrhea and 69% against severe rotavirus diarrhea (48).

Other Candidate Vaccines

In clinical trials, no Jennerian vaccine has provided complete protection against rotavirus diarrhea; as a result, several non-Jennerian candidate vaccines are being developed. Vaccines based on neonatal, cold-adapted, and attenuated human strains of rotavirus are under evaluation (49). Other approaches, such as the use of baculovirus-expressed viruslike particles or naked DNA vaccines, are also being used to develop candidate rotavirus vaccines (50,51).

 

The Ever-Changing Landscape of Rotavirus Serotypes

(The Pediatric Infectious Disease Journal:Volume 28(3) Supplement March 2009pp S60-S62)

 

Abstract:

Rotavirus is a double-stranded RNA virus that is characterized by substantial genetic diversity. The various serotypes of rotavirus have been determined by the presence of neutralizing epitopes on the outer capsid of the protein shell. At present, 5 rotavirus serotypes (G1, G2, G3, G4, G9) are the predominant circulating strains, accounting for approximately 95% of strains worldwide, although there is considerable geographic variability. Incidence rates for various serotypes also vary temporally with seasonal and year-to-year fluctuations. Unusual serotypes are generally uncommon, but new serotypes can emerge. In particular, G9[P8], a reassortment virus, was first identified in 1983 and in the last 10 to 15 years has become widely distributed worldwide. Indeed, G9[P8] has become highly prevalent in many countries in Europe and Australia, with somewhat lower incidence rates in South America, Africa, and Asia. The heterogeneity and ever-changing epidemiology of rotavirus underscores the need for continued surveillance to ensure that vaccination programs provide optimal protection.

Three oral RV vaccines are currently licensed, a human monovalent live attenuated RV strain, RotarixTM, a pentavalent live bovine-human reassortant vaccine, RotaTeqTM, and a lamb-derived monovalent live attenuated strain, LLR, which is only being used in China.

ROTARIX® (Rotavirus Vaccine, Live, Oral)

 

ROTARIX is a vaccine indicated for the prevention of rotavirus gastroenteritis caused by G1 and non-G1 types (G3, G4, and G9) in infants and children.

ROTARIX (Rotavirus Vaccine, Live, Oral), for oral administration, is a live, attenuated rotavirus vaccine derived from the human 89-12 strain which belongs to G1P [8] type. The rotavirus strain is propagated on Vero cells. After reconstitution, the final formulation (1 mL) contains at least 106.0 median Cell Culture Infective Dose (CCID50) of live, attenuated rotavirus.

RotaTeq (Rotavirus Vaccine, Live, Oral, Pentavalent)  Oral Solution

 

RotaTeq is indicated for the prevention of rotavirus gastroenteritis in infants and children caused by the G1, G2, G3 and G4 serotypes contained in the vaccine.

RotaTeq, 2 mL for oral use, is a ready-to-use solution of live reassortant rotaviruses, containing G1,G2, G3, G4 and P1A[8] which contains a minimum of 2.0 – 2.8 x 106 infectious units (IU) per individual reassortant dose, depending on the serotype, and not greater than 116 x 106 IU per aggregate dose.

RotaTeq is a live, oral pentavalent vaccine that contains 5 live reassortant rotaviruses. The rotavirus parent strains of the reassortants were isolated from human and bovine hosts. Four reassortant rotaviruses express one of the outer capsid proteins (G1, G2, G3, or G4) from the human rotavirus parent strain and the attachment protein (serotype P7) from the bovine rotavirus parent strain. The fifth reassortant virus expresses the attachment protein, P1A (genotype P[8]), herein referred to as serotype P1A[8], from the human rotavirus parent strain and the outer capsid protein of serotype G6 from the bovine rotavirus parent strain…

Monovalent Lamb Vaccine

The only vaccine that is currently licensed is in use in China.   Developed by Zhi-Sheng Bai at the Lanzhou Institute, the attenuated monovalent vaccine is based on a strain of lamb rotavirus.   Similar to Rotarix®, the lamb rotavirus strain was attenuated by passage in cell culture.  

The lamb vaccine is classified as serotype P[10], G12. It is delivered as a single dose, between the ages of 2-24 months.

The vaccine has been proven to be safe and immunogenic (with 61% of vaccinees developing neutralizing antibody responses), but efficacy results of the Phase II trails have yet to be published.[6]

Additional Information about the vaccines:

Live attenuated RV strains

The first RV vaccines to be tested in humans were the live bovine strains RIT4237 (P[1]-G6) and WC3 (P[5]-G6), and the live simian strain RRV (P[3]-G3), which are attenuated for humans and could be administered by the oral route. The three strains induced neutralizing antibodies in a majority of infants but showed inconstant capacity to protect against RV disease.

In China, a lamb-derived monovalent (P[12]-G10) live-attenuated, 3-dose oral vaccine, was developed by the Lanzhou Institute of Biomedical Products and is used in the private sector. The vaccine is reported to induce neutralizing antibody responses in 60% of vaccinees but its efficacy is not precisely known since it was not tested against placebo in a controlled Phase III trial [135] .

A human P[8]-G1 RV strain, RIX 4414, which was isolated from the stools of a sick 15-month old boy in the USA, was attenuated by multiple passages in cell culture, plaque-purified and passaged again in Vero cells. The strain was developed as a 2-dose monovalent oral vaccine by AVANT Immunotherapeutics then licensed to GlaxoSmithKline Biologicals. The vaccine (RotarixTM) showed 70%-85% protective efficacy against severe disease, including that due non-G1 serotypes [149, 150]. It now has been tested in more than 60 countries in Latin America, Africa, Asia and Europe. A large, multicentered safety trial on 63 225 infants between 6 and 14 weeks of age in Latin America and Finland confirmed the initial safety data and indicated no increased attributable risk of intussusceptions (IS) in the high-risk period up to 30 days post any dose [151]. The vaccine was first licensed in 2004 in Mexico and the Dominican Republic and has now been licensed in many countries worldwide. It also has been prequalified by WHO for procurement by UNICEF and the UN Vaccine Fund. Additional Phase IIb and III trials are in progress in South Africa, Malawi and Bangladesh to determine if the vaccine will work well in children from poor settings in developing countries, if it can be administered with the oral polio vaccine without interference, and whether it can safely be administered to HIV positive infants. Final results are due in 2009.

Another human RV strain, RV3 (P2[6]-G3), isolated from newborns at the Royal Hospital in Melbourne, Australia [152] is also developed as a candidate live oral vaccine. A small Phase II study with three doses of 105 pfu of the vaccine indicated relatively low immunogenicity in infants as measured by serum IgA levels [153] . However, the vaccine recipients who developed an immune response were protected against clinical disease in the following year. Strategies to increase the potency of the vaccine are under study with a vaccine producer in Indonesia.

6.3.2 Live reassortant RV strains

Efforts also were made to develop human-animal reassortant RV strains containing the VP7 or VP4 RNA segment from a human RV strain to provide the required antigenicity and the other 10 RNA segments from a simian or a bovine strain to provide the attenuated phenotype [154, 155].

A tetravalent rhesus-human reassortant RV vaccine, RRV-TV, was initially developed at the NIH, Bethesda, using the simian RRV strain (G3) mixed with three human-simian reassortant strains of G types 1, 2, and 4, respectively. The vaccine (RotaShieldTM, Wyeth-Lederle Vaccines, USA) was shown to provide 48–68% protection against any RV disease and 64–91% protection against severe disease [156] . It was introduced in August 1998 on the market in the USA and administered in a three dose schedule to over 600 000 infants within the following year, until an unexpected adverse event, intussusception (IS), was found to occur in a significant number of cases within two weeks after administration of the first two doses of vaccine, leading to its eventual withdrawal [157] . The risk of IS, initially targeted at 1 in 2500 children immunized, has now been reassessed as 1 in 10 000. Its occurrence led to a very thorough safety assessment of the following generation of live oral RV vaccines (viz RotarixTM and RotaTeqTM), with sample sizes in excess of 60 000 subjects. The original RotashieldTM vaccine has now been licensed to a biotech company, BIOVIRx Inc., USA.

A pentavalent human-bovine reassortant vaccine, RotaTeqTM, was prepared by Merck Research Co., Pennsylvania, by reassortment between the naturally attenuated bovine RV strain WC3 and five different human RV strains of serotypes G1, G2, G3, G4 and P[8], respectively. The live-attenuated, 3-dose oral vaccine, was tested in a large safety and efficacy trial in Finland and the USA on more than 70 000 children who were carefully monitored for 2 weeks after each immunization for risks of IS. The vaccine was found to be totally safe and to elicit 74% protection against any G1-G4 RV gastroenteritis through the first RV season after vaccination [158]. Vaccination reduced doctor visits for RV diarrhoea by 86% and hospitalizations and emergency department visits by 94.5%. The vaccine was shown not to interfere with the immunogenicity of a combined Hib, DTP, HepB, conjugated pneumococcal and inactivated polio vaccine, nor with concomitant administration of the oral polio vaccine [159]. RotaTeqTM was licensed in February 2006 in the USA and subsequently in many countries worldwide. It officially was recommended for the routine immunization of children in the USA after active surveillance showed only three cases of IS among more than 100 000 vaccinated infants. It also has been included into national vaccination programs in several countries. A Phase III trial is ongoing in African countries (Mali, Ghana, and Kenya). Results are expected end of 2009.

An alternative multivalent bovine-human reassortant oral vaccine was developed by the National Institute of Allergy and Infectious Diseases (NIAID, NIH, Bethesda), based on the attenuated bovine strain UK reassorted with the five most common RV serotypes in humans, G1-G4, G8 and G9 [156] . Phase II data showed a good immune response and no adverse interference with concomitantly administered childhood vaccines. A non-exclusive license for production of the vaccine has been granted to vaccine producers in Brazil, China and India.

Finally, a naturally occurring human–bovine, neonate-derived RV strain, 116E (P[10]-G9), which was isolated from a nosocomial outbreak of asymptomatic infection in New-Delhi, is under development by Bharat Biotech Ltd in India [148, 160]. A similar strain, I321 (P[11]-G10), was found not to be immunogenic [161] .

 

6.3.3 Other RV vaccine approaches

New RV vaccine approaches include an inactivated virus vaccine [162] , DNA vaccines [163] , a VP6 subunit vaccine [164, 165] and virus-like particles (VLPs) expressed in a baculovirus system [166-168]. Depending on the number of viral proteins expressed, the complexity of the VLPs can vary from mono-layered (VP2-VLPs) to double-layered (VP2/6 VLPs) or triple- layered VLPs (VP2/6/7/4 VLPs).

HPV Strains

HPV

  

The Human Papilloma Virus (HPV) is the cause of both genital and non-genital warts. This is a very common family of viruses.  There are actually more than 100 different strains of HPV.  Of these, approximately 30 exist in the genital area and can cause genital warts (condyloma acuminata).  These 30 strains can be further broken down to “high” and “low” risk strains.

 

 

High risk strains may show up in a Pap smear and may rarely develop into cancer.  There are approximately 13 high-risk strains of HPV. Two strains (16 & 18) are believed to cause about 70% of all cervical cancer.

Low risk strains can sometimes cause changes in a Pap smear, but do not progress to cancer. Two of the low risk-risk strains (6 and 11) are most likely to cause genital warts.

There are 40 strains of HPV that can affect the anal and genital tracts and these are further divided into low risk and high risk strains.

 

Thirteen strains are considered high risk, or more likely to progress to high grade lesions (HSIL, CIN 2 or 3) and possibly cancer, if not cleared by the immune system. These strains are: 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, and 68. Strains 16 and 18 are by far the most common types, and one or both are present in approximately 70% of cervical cancers worldwide. Despite this strong link, only a very small percentage of high risk HPV infections will ever become invasive cancer (estimated at 2%). The time between initial exposure and the development of cancer can vary from months to years, but the average time is thought to be 15 years. Unfortunately, the high risk HPV strains do not usually cause any symptoms to alert someone that they have the infection. The low risk strains are not considered a risk for cervical cancer, but they can cause low grade lesions (CIN 1) and several of these strains can cause genital warts.

GARDASIL [Human Papillomavirus Quadrivalent (Types 6, 11, 16, and 18) Vaccine, Recombinant]

 

GARDASIL is a vaccine indicated in girls and women 9 through 26 years of age for the prevention of the following diseases caused by Human Papillomavirus (HPV) types included in the vaccine:

Cervical, vulvar, and vaginal cancer caused by HPV types 16 and 18

Genital warts (condyloma acuminata) caused by HPV types 6 and 11

And the following precancerous or dysplastic lesions caused by HPV types 6, 11, 16, and 18:

Cervical intraepithelial neoplasia (CIN) grade 2/3 and Cervical adenocarcinoma in situ (AIS)

Cervical intraepithelial neoplasia (CIN) grade 1

Vulvar intraepithelial neoplasia (VIN) grade 2 and grade 3

Vaginal intraepithelial neoplasia (VaIN) grade 2 and grade 3

GARDASIL, Human Papillomavirus Quadrivalent (Types 6, 11, 16, and 18) Vaccine, Recombinant, is a non-infectious recombinant quadrivalent vaccine prepared from the purified virus-like particles (VLPs) of the major capsid (L1) protein of HPV Types 6, 11, 16, and 18. The L1 proteins are produced by separate fermentations in recombinant Saccharomyces cerevisiae and self-assembled into VLPs. The fermentation process involves growth of S. cerevisiae on chemically-defined fermentation media which include

vitamins, amino acids, mineral salts, and carbohydrates. The VLPs are released from the yeast cells by cell disruption and purified by a series of chemical and physical methods. The purified VLPs are adsorbed on preformed aluminum-containing adjuvant (Amorphous Aluminum Hydroxyphosphate Sulfate). The quadrivalent HPV VLP vaccine is a sterile liquid suspension that is prepared by combining the adsorbed VLPs of each HPV type and additional amounts of the aluminum-containing adjuvant and the final

purification buffer.

GARDASIL is a sterile suspension for intramuscular administration. Each 0.5-mL dose contains approximately 20 mcg of HPV 6 L1 protein, 40 mcg of HPV 11 L1 protein, 40 mcg of HPV 16 L1 protein, and 20 mcg of HPV 18 L1 protein.

Each 0.5-mL dose of the vaccine contains approximately 225 mcg of aluminum (as Amorphous Aluminum Hydroxyphosphate Sulfate adjuvant), 9.56 mg of sodium chloride, 0.78 mg of L-histidine, 50 mcg of polysorbate 80, 35 mcg of sodium borate, < 7 mcg yeast protein/dose, and water for injection.

The product does not contain a preservative or antibiotics.

CERVARIX® PRODUCT INFORMATION  Human Papillomavirus Vaccine Types 16 and 18

(Recombinant, AS04 adjuvanted)

 

CERVARIX contains recombinant C-terminally truncated L1 proteins from human papillomavirus (HPV) type-16 and type-18 each assembled as virus-like particles (VLPs)…

The HPV-16 and HPV-18 L1 antigens are prepared by recombinant DNA technology using a Baculovirus expression system in Trichoplusnia ni cells.

HPV-16 and HPV-18 L1 antigens in CERVARIX are adjuvanted with AS04. This AS04 adjuvant system comprises aluminium hydroxide (Al(OH)3) and 3-O-desacyl-4’-monophosphoryl lipid A (MPL). The MPL within AS04 enhances the initiation of the immune response through the activation of innate immunity, leading to an improved cellular and humoral adaptive immune response.

Each 0.5ml dose of CERVARIX contains 20 micrograms each of HPV-16 L1 and HPV-18 L1 proteins, 0.5 milligrams of Al(OH)3 and 50 micrograms of MPL. CERVARIX also contains sodium chloride (NaCl) 4.4 mg, sodium phosphate – monobasic (NaH2PO4.2 H2O) 624 micrograms and water for injection as excipients. CERVARIX does not contain a preservative.

Rotavirus Strains

Rotavirus

Rotavirus-Synopses

The major antigenic properties of rotaviruses—group, subgroup, and serotype—are determined by the viral capsid proteins. Rotavirus has seven major groups (A-G); most human strains belong to group A, although groups B and C have occasionally been associated with human illness. The product of the 6th gene of group A rotaviruses encodes VP6, the most abundant viral protein, which is the major determinant of group reactivity, the target of common diagnostic assays, and contains the antigen used to further classify rotaviruses into subgroups I and II. The outer capsid proteins, VP7, the glycoprotein or G-protein (encoded by gene 7, 8, or 9, depending on the strain), and VP4, the protease-cleaved or P-protein (encoded by gene segment 4), determine the serotype specificity and form the basis of the binary classification (G and P type) of rotaviruses. Both G and P proteins induce neutralizing antibodies and may be involved in protective immunity.

Fourteen G serotypes of rotavirus, 10 of which occur in humans, have been defined by cross-neutralization studies with polyclonal animal serum samples; these serotypes correlate with antigenic specificities of the VP7 glycoprotein. The characterization of P serotypes has been difficult because adequate reagents are not available. Eight P serotypes of human rotaviruses have been characterized. Additional VP4 gene variants have been identified, so ultimately the number of P serotypes may exceed 20. Theoretically, 80 different strains of rotavirus could result from various combinations of the known 10 G and 8 P serotypes of human rotaviruses. For vaccine development purposes, it is fortunate that only four common strains (P[8]G1; P[8]G3; P[8]G4; and P[4]G2) of rotavirus predominate globally (Figure 4) (11). However, the prevalence of rotavirus strains varies considerably from one geographic area to another, and unusual strains are common in several developing countries (e.g., unusual P[6] strains, including those with serotype G9 specificity, accounted for 9.5% of all rotaviruses from a multicenter collection in India)

In infants and young children, neutralizing antibodies directed primarily against the G serotype of the infecting strain (homotypic response) develop after primary infection with rotavirus (18). Repeat rotavirus infections elicit both a homotypic and heterotypic (against strains with different G serotypes) antibody response. Protection against rotavirus diarrhea correlates with serum antibody titers following natural infection of young children, and infected children are more protected against reinfection with similar rather than different G serotypes. A protective role of placentally transferred maternal antibody among infants < 3 months of age has also been speculated since rotavirus disease is uncommon in this age group. However, serum neutralizing antibody responses among vaccine recipients have sometimes correlated poorly with protection from disease; therefore, the exact role of serum antibody in protection against disease remains unclear.

Genome Variability

Rotavirus is a highly variable virus even within the subset of those that are infective to humans. [4] Rotaviruses are usually categorized into seven groups A-G, with subgroups I and II based on the VP6 protein. [8] Within these groups, A, B, and C are infective to Humans. [19]

Rotavirus is further categorized into G and P serotypes. The G serotype is specified by the glycoprotein VP7 of the outer capsid, which is coded by viral genes 7, 8, and 9. [8] The P serotype is specified by protein VP4, also on the outer capsid. It is a protease cleaved protein coded by gene 4 of the virus genome. [8] The most common G serotypes currently are G1, G2, G3, G4, and G9, with G1 being most prevalent and G9 the fastest emerging worldwide. [7] [8] [10] [12] Common P serotypes are P1a, P1b, and P8. [13] [6] [4]

 rotastarins

Rotavirus Vaccines:

 

Monovalent “Jennerian” Vaccines

Initial development of rotavirus vaccines was based on the Jennerian approach, which involved the use of a live, attenuated, antigenically related virus derived from a nonhuman host (21). This approach was prompted by studies indicating that animal and human rotaviruses shared a common group antigen and that experimental animals immunized with animal strains of rotavirus had a significantly lower risk for illness and viral shedding when subsequently challenged with human rotaviruses. Furthermore, neutralizing antibodies to human rotavirus serotypes in the animal models indicated the potential for cross-protection.

Bovine Vaccines

The first two Jennerian vaccines were developed with bovine rotavirus strains RIT4237 and WC3. The WC3 strain was passaged in cell culture less than RIT4237 and was developed because of concern that excessive passaging of the RIT vaccine might cause overattenuation and diminished efficacy. RIT4237 and WC3 were nonreactogenic and immunogenic when administered to infants 2 to 18 months of age. However, the protection conferred by both vaccines varied greatly in efficacy studies, 0% to 76% against any rotavirus diarrhea and 0% to 100% against severe disease (2232). A well-defined correlate of protection was not identified, and reasons for the variable efficacy were unknown, although late age at vaccination, timing of vaccination with respect to the onset of the rotavirus season, and variations in the strength and number of doses of the vaccine were proposed as contributing factors. Both vaccines performed less well in developing than in industrialized countries, possibly because of interference by other enteropathogens or inadequate surveillance during follow-up.

 

Rhesus Vaccine

The third Jennerian vaccine was developed with rhesus rotavirus strain MMU18006, which shares neutralization specificity with human rotavirus G3 strains. Besides sharing antigenic specificity with an epidemiologically important human rotavirus serotype, MMU18006 was suitable for vaccine development because it grew efficiently in cell culture. As in the bovine rotavirus-based vaccines, MMU18006 was safe and immunogenic, although in some trials, one third of infants became febrile 3 to 4 days after vaccination. The reactogenicity of MMU18006 was particularly high in two studies in Finland and Sweden in which 64% and 79% of infants, respectively, became febrile. Most children with febrile responses were >5 months of age; lack of passively transferred maternal antibody might have contributed to the high reactogenicity of the vaccine. As in the RIT4237 and WC3 vaccines, the protective efficacy of MMU18006 in field trials was quite variable, 0% to 60% against any rotavirus diarrhea and 0% to 85% against severe rotavirus diarrhea (3339).

Reassortant “Modified Jennerian” Vaccines

The greatest efficacy of MMU18006 was observed in a Venezuelan trial in which the rotavirus strain circulating in the community (G3) was the same serotype as the vaccine strain, which suggested that serotype-specific immunity against each of the epidemiologically important strains of human rotaviruses may be required for maximum protection. Similar observations in vaccine challenge cross-protection studies in animals initiated the development of vaccines that used a modified Jennerian approach in which animal-human reassortants expressing VP7 proteins of serotypes 1 through 4 were used as the immunogens.

 

Rhesus-Human Reassortant Vaccines

Rhesus-human reassortants were generated by coinfecting cell cultures with rhesus rotavirus (RRV) strain MMU18006 (G serotype 3) and human rotavirus strains D (G serotype 1), DS-1 (G serotype 2), and ST3 (G serotype 4). Selection pressure (induced by the addition of neutralizing antibody to VP7 of RRV) produced reassortant strains D x RRV, DS-1 x RRV, and ST3 x RRV, each of which possessed the VP7 gene from HRV serotype 1, 2, or 4 and the other 10 genes from RRV (Figure 6) (40). Because vaccines made from the individual reassortants were safe and immunogenic, RRV-TV was developed incorporating each of the three reassortants and MMU18006 to provide coverage against the four common VP7 serotypes of rotavirus.

RRV-TV testing was initiated at an inoculum of 104 PFU of each of the four viruses (i.e., at 4 x 104 PFU) and completed at 105 PFU, the dose submitted for licensure (i.e., at 4 x 105 PFU). In most trials, vaccine was administered orally in three doses separated by at least 3 weeks to optimize the immune response to the component antigens; immunization was completed by the age of 6 to 7 months. Because the vaccine virus strains are acid labile, they are administered with 2.5 ml of citrate-bicarbonate buffer.

 

Bovine-Human Reassortant Vaccines

 

Bovine-human reassortant rotavirus vaccines include a tetravalent WC3 rotavirus reassortant vaccine with genes coding for the VP7 of three major serotypes of rotavirus (G1, G2, and G3) and W179-4, a human VP4 reassortant with P[8] specificity. Theoretically, this vaccine should induce antibodies broadly reactive to the three common serotypes of rotavirus sharing P [8] specificity, thereby increasing the protective efficacy of this vaccine. In an efficacy trial of a three-dose regimen of the WC3 reassortant vaccine, protection was 67% against all rotavirus diarrhea and 69% against severe rotavirus diarrhea (48).

Other Candidate Vaccines

In clinical trials, no Jennerian vaccine has provided complete protection against rotavirus diarrhea; as a result, several non-Jennerian candidate vaccines are being developed. Vaccines based on neonatal, cold-adapted, and attenuated human strains of rotavirus are under evaluation (49). Other approaches, such as the use of baculovirus-expressed viruslike particles or naked DNA vaccines, are also being used to develop candidate rotavirus vaccines (50,51).

 

The Ever-Changing Landscape of Rotavirus Serotypes

(The Pediatric Infectious Disease Journal:Volume 28(3) Supplement March 2009pp S60-S62)

 

Abstract:

Rotavirus is a double-stranded RNA virus that is characterized by substantial genetic diversity. The various serotypes of rotavirus have been determined by the presence of neutralizing epitopes on the outer capsid of the protein shell. At present, 5 rotavirus serotypes (G1, G2, G3, G4, G9) are the predominant circulating strains, accounting for approximately 95% of strains worldwide, although there is considerable geographic variability. Incidence rates for various serotypes also vary temporally with seasonal and year-to-year fluctuations. Unusual serotypes are generally uncommon, but new serotypes can emerge. In particular, G9[P8], a reassortment virus, was first identified in 1983 and in the last 10 to 15 years has become widely distributed worldwide. Indeed, G9[P8] has become highly prevalent in many countries in Europe and Australia, with somewhat lower incidence rates in South America, Africa, and Asia. The heterogeneity and ever-changing epidemiology of rotavirus underscores the need for continued surveillance to ensure that vaccination programs provide optimal protection.

Three oral RV vaccines are currently licensed, a human monovalent live attenuated RV strain, RotarixTM, a pentavalent live bovine-human reassortant vaccine, RotaTeqTM, and a lamb-derived monovalent live attenuated strain, LLR, which is only being used in China.

ROTARIX® (Rotavirus Vaccine, Live, Oral)

 

ROTARIX is a vaccine indicated for the prevention of rotavirus gastroenteritis caused by G1 and non-G1 types (G3, G4, and G9) in infants and children.

ROTARIX (Rotavirus Vaccine, Live, Oral), for oral administration, is a live, attenuated rotavirus vaccine derived from the human 89-12 strain which belongs to G1P [8] type. The rotavirus strain is propagated on Vero cells. After reconstitution, the final formulation (1 mL) contains at least 106.0 median Cell Culture Infective Dose (CCID50) of live, attenuated rotavirus.

RotaTeq (Rotavirus Vaccine, Live, Oral, Pentavalent)  Oral Solution

 

RotaTeq is indicated for the prevention of rotavirus gastroenteritis in infants and children caused by the G1, G2, G3 and G4 serotypes contained in the vaccine.

RotaTeq, 2 mL for oral use, is a ready-to-use solution of live reassortant rotaviruses, containing G1,G2, G3, G4 and P1A[8] which contains a minimum of 2.0 – 2.8 x 106 infectious units (IU) per individual reassortant dose, depending on the serotype, and not greater than 116 x 106 IU per aggregate dose.

RotaTeq is a live, oral pentavalent vaccine that contains 5 live reassortant rotaviruses. The rotavirus parent strains of the reassortants were isolated from human and bovine hosts. Four reassortant rotaviruses express one of the outer capsid proteins (G1, G2, G3, or G4) from the human rotavirus parent strain and the attachment protein (serotype P7) from the bovine rotavirus parent strain. The fifth reassortant virus expresses the attachment protein, P1A (genotype P[8]), herein referred to as serotype P1A[8], from the human rotavirus parent strain and the outer capsid protein of serotype G6 from the bovine rotavirus parent strain…

Monovalent Lamb Vaccine

The only vaccine that is currently licensed is in use in China.   Developed by Zhi-Sheng Bai at the Lanzhou Institute, the attenuated monovalent vaccine is based on a strain of lamb rotavirus.   Similar to Rotarix®, the lamb rotavirus strain was attenuated by passage in cell culture.  

The lamb vaccine is classified as serotype P[10], G12. It is delivered as a single dose, between the ages of 2-24 months.

The vaccine has been proven to be safe and immunogenic (with 61% of vaccinees developing neutralizing antibody responses), but efficacy results of the Phase II trails have yet to be published.[6]

Additional Information about the vaccines:

Live attenuated RV strains

The first RV vaccines to be tested in humans were the live bovine strains RIT4237 (P[1]-G6) and WC3 (P[5]-G6), and the live simian strain RRV (P[3]-G3), which are attenuated for humans and could be administered by the oral route. The three strains induced neutralizing antibodies in a majority of infants but showed inconstant capacity to protect against RV disease.

In China, a lamb-derived monovalent (P[12]-G10) live-attenuated, 3-dose oral vaccine, was developed by the Lanzhou Institute of Biomedical Products and is used in the private sector. The vaccine is reported to induce neutralizing antibody responses in 60% of vaccinees but its efficacy is not precisely known since it was not tested against placebo in a controlled Phase III trial [135] .

A human P[8]-G1 RV strain, RIX 4414, which was isolated from the stools of a sick 15-month old boy in the USA, was attenuated by multiple passages in cell culture, plaque-purified and passaged again in Vero cells. The strain was developed as a 2-dose monovalent oral vaccine by AVANT Immunotherapeutics then licensed to GlaxoSmithKline Biologicals. The vaccine (RotarixTM) showed 70%-85% protective efficacy against severe disease, including that due non-G1 serotypes [149, 150]. It now has been tested in more than 60 countries in Latin America, Africa, Asia and Europe. A large, multicentered safety trial on 63 225 infants between 6 and 14 weeks of age in Latin America and Finland confirmed the initial safety data and indicated no increased attributable risk of intussusceptions (IS) in the high-risk period up to 30 days post any dose [151]. The vaccine was first licensed in 2004 in Mexico and the Dominican Republic and has now been licensed in many countries worldwide. It also has been prequalified by WHO for procurement by UNICEF and the UN Vaccine Fund. Additional Phase IIb and III trials are in progress in South Africa, Malawi and Bangladesh to determine if the vaccine will work well in children from poor settings in developing countries, if it can be administered with the oral polio vaccine without interference, and whether it can safely be administered to HIV positive infants. Final results are due in 2009.

Another human RV strain, RV3 (P2[6]-G3), isolated from newborns at the Royal Hospital in Melbourne, Australia [152] is also developed as a candidate live oral vaccine. A small Phase II study with three doses of 105 pfu of the vaccine indicated relatively low immunogenicity in infants as measured by serum IgA levels [153] . However, the vaccine recipients who developed an immune response were protected against clinical disease in the following year. Strategies to increase the potency of the vaccine are under study with a vaccine producer in Indonesia.

6.3.2 Live reassortant RV strains

Efforts also were made to develop human-animal reassortant RV strains containing the VP7 or VP4 RNA segment from a human RV strain to provide the required antigenicity and the other 10 RNA segments from a simian or a bovine strain to provide the attenuated phenotype [154, 155].

A tetravalent rhesus-human reassortant RV vaccine, RRV-TV, was initially developed at the NIH, Bethesda, using the simian RRV strain (G3) mixed with three human-simian reassortant strains of G types 1, 2, and 4, respectively. The vaccine (RotaShieldTM, Wyeth-Lederle Vaccines, USA) was shown to provide 48–68% protection against any RV disease and 64–91% protection against severe disease [156] . It was introduced in August 1998 on the market in the USA and administered in a three dose schedule to over 600 000 infants within the following year, until an unexpected adverse event, intussusception (IS), was found to occur in a significant number of cases within two weeks after administration of the first two doses of vaccine, leading to its eventual withdrawal [157] . The risk of IS, initially targeted at 1 in 2500 children immunized, has now been reassessed as 1 in 10 000. Its occurrence led to a very thorough safety assessment of the following generation of live oral RV vaccines (viz RotarixTM and RotaTeqTM), with sample sizes in excess of 60 000 subjects. The original RotashieldTM vaccine has now been licensed to a biotech company, BIOVIRx Inc., USA.

A pentavalent human-bovine reassortant vaccine, RotaTeqTM, was prepared by Merck Research Co., Pennsylvania, by reassortment between the naturally attenuated bovine RV strain WC3 and five different human RV strains of serotypes G1, G2, G3, G4 and P[8], respectively. The live-attenuated, 3-dose oral vaccine, was tested in a large safety and efficacy trial in Finland and the USA on more than 70 000 children who were carefully monitored for 2 weeks after each immunization for risks of IS. The vaccine was found to be totally safe and to elicit 74% protection against any G1-G4 RV gastroenteritis through the first RV season after vaccination [158]. Vaccination reduced doctor visits for RV diarrhoea by 86% and hospitalizations and emergency department visits by 94.5%. The vaccine was shown not to interfere with the immunogenicity of a combined Hib, DTP, HepB, conjugated pneumococcal and inactivated polio vaccine, nor with concomitant administration of the oral polio vaccine [159]. RotaTeqTM was licensed in February 2006 in the USA and subsequently in many countries worldwide. It officially was recommended for the routine immunization of children in the USA after active surveillance showed only three cases of IS among more than 100 000 vaccinated infants. It also has been included into national vaccination programs in several countries. A Phase III trial is ongoing in African countries (Mali, Ghana, and Kenya). Results are expected end of 2009.

An alternative multivalent bovine-human reassortant oral vaccine was developed by the National Institute of Allergy and Infectious Diseases (NIAID, NIH, Bethesda), based on the attenuated bovine strain UK reassorted with the five most common RV serotypes in humans, G1-G4, G8 and G9 [156] . Phase II data showed a good immune response and no adverse interference with concomitantly administered childhood vaccines. A non-exclusive license for production of the vaccine has been granted to vaccine producers in Brazil, China and India.

Finally, a naturally occurring human–bovine, neonate-derived RV strain, 116E (P[10]-G9), which was isolated from a nosocomial outbreak of asymptomatic infection in New-Delhi, is under development by Bharat Biotech Ltd in India [148, 160]. A similar strain, I321 (P[11]-G10), was found not to be immunogenic [161] .

 

6.3.3 Other RV vaccine approaches

New RV vaccine approaches include an inactivated virus vaccine [162] , DNA vaccines [163] , a VP6 subunit vaccine [164, 165] and virus-like particles (VLPs) expressed in a baculovirus system [166-168]. Depending on the number of viral proteins expressed, the complexity of the VLPs can vary from mono-layered (VP2-VLPs) to double-layered (VP2/6 VLPs) or triple- layered VLPs (VP2/6/7/4 VLPs).

Hepatitis A Strains

Hepatitis A

 

The pathogen and the disease

HAV is a member of the Picornaviridae family that includes both the enteroviruses and rhinoviruses of humans. Being the only species member, it constitutes its own genus named hepatovirus. HAV is a non-enveloped (naked) virus of 27–28 nm diameter without morphological features differentiating it from other picornaviruses. Four structural proteins encapsulate the RNA genome. Neutralization sites for anti-HAV antibodies are mainly contained in two of these proteins. Although six genotypes of HAV have been identified, there appears to be no variation detectable by serology in these neutralization sites. The virus is relatively stable at low pH and moderate temperatures, but is inactivated by high temperature (almost instantly at 85°C/185°F), and by formalin or chlorine. HAV itself is not cytopathic and the liver-cell damage is caused by the cell-mediated immune response.

The clinical course of acute hepatitis A is indistinguishable from other types of acute viral hepatitis. Symptoms typically include fever, malaise, anorexia, nausea and abdominal discomfort, followed by dark urine and jaundice. The severity of disease and mortality increases in older age groups. The convalescence following hepatitis A may be slow, and is characterized by fatigue, nausea and lack of appetite. Complications of hepatitis A include relapsing hepatitis, cholestatic hepatitis and fulminant hepatitis. Fulminant hepatitis occurs in approximately 0.01% of clinical infections and is characterized by rapid deterioration in liver function and a very high fatality rate. Chronic infection with HAV does not occur. No specific antiviral therapy is currently available.

The aetiological diagnosis is made by the demonstration of IgM antibodies to HAV (IgM anti-HAV) in serum. Detection of the virus or viral antigens in the stool is of limited value for routine diagnosis.

 

Strains

Only one serotype of HAV has been identified and a single infection confers lifelong immunity. However, genetic heterogeneity between HAV isolates from different parts of the world has enabled the classification of HAV strains into seven different genotypes, designated I to VII. Four of these have been associated with human disease, I, II, III, and VII. Most human HAV strains belong to genotypes I and III, with 80% of them being genotype I. Genotypes I and III are further divided into subtypes A and B. Genotypes II and VII are represented only by one human strain each, and genotypes IV, V, and VI include strains recovered from simians (Arauz-Ruiz et al., 2001). Genotype IA appears to be the agent responsible for the majority of hepatitis A cases worldwide and has been isolated from all parts of the world. Genotype IB appears to occur in the Mediterranean region, whereas genotype III viruses have been isolated from diverse sources such as Panamanian owl monkeys, drug abusers in Sweden and patients from India and Nepal. Single representatives of genotype II and VII were isolated from individual patients from Sierra Leone and France (Lu et al., 2004). Several studies have indicated that HAV strains in North America mainly belong to subtype IA (Arauz-Ruiz et al., 2001).

Variant(s):  (Click link to the list of all the variant strains)

 

Hepatitis A vaccines

Techniques for growing HAV in cell culture have made it possible to generate sufficient amounts of virus for vaccine production. Several inactivated or live attenuated vaccines against hepatitis A have been developed, but only four inactivated hepatitis A vaccines are currently available internationally. All four vaccines are similar in terms of efficacy and side-effect profile. The vaccines are given parenterally, as a two-dose series, 6-18 months apart. The dose of vaccine, vaccination schedule, ages for which the vaccine is licensed, and whether there is a paediatric and adult formulation varies from manufacturer to manufacturer. No vaccine is licensed for children younger than one year of age.

Three vaccines are manufactured from cell-culture-adapted HAV propagated in human fibroblasts. Following purification from cell lysates, the HAV preparation is formalin-inactivated and adsorbed to an aluminium hydroxide adjuvant. One vaccine is formulated without preservative; the other two are prepared with 2-phenoxyethanol as a preservative. The fourth vaccine is manufactured from HAV purified from infected human diploid cell cultures and inactivated with formalin. This preparation is adsorbed to biodegradable, 150 nm phospholipid vesicles spiked with influenza haemagglutinin and neuramidase. These virosomes are thought to directly target influenza-primed antibody-presenting cells as well as macrophages, thus stimulating a rapid vaccine-induced B-cell and T-cell proliferation in the majority of vaccinees. A combination vaccine containing inactivated hepatitis A and recombinant hepatitis B vaccines has been licensed since 1996 for use in children aged one year or older in several countries. The combination vaccine is given as a three-dose series, using a 0, 1, 6 month schedule.

The Vaccines Available:

 

TWINRIX® [Hepatitis A Inactivated & Hepatitis B (Recombinant) Vaccine]

 

 

TWINRIX® [Hepatitis A Inactivated & Hepatitis B (Recombinant) Vaccine] is a sterile bivalent vaccine containing the antigenic components used in producing HAVRIX® (Hepatitis A Vaccine, Inactivated) and ENGERIX-B® [Hepatitis B Vaccine (Recombinant)]. TWINRIX is a sterile suspension of inactivated hepatitis A virus (strain HM175) propagated in MRC-5 cells, and combined with purified surface antigen of the hepatitis B virus. The purified hepatitis B surface antigen (HBsAg) is obtained by culturing genetically engineered Saccharomyces cerevisiae cells, which carry the surface antigen gene of the hepatitis B virus, in synthetic media containing inorganic salts, amino acids, dextrose, and vitamins. Bulk preparations of each antigen are adsorbed separately onto aluminum salts and then pooled during formulation.

A 1.0-mL dose of vaccine contains 720 ELISA Units of inactivated hepatitis A virus and 20 mcg of recombinant HBsAg protein. One dose of vaccine also contains 0.45 mg of aluminum in the form of aluminum phosphate and aluminum hydroxide as adjuvants, amino acids, sodium chloride, phosphate buffer, polysorbate 20, Water for Injection, traces of formalin (not more than 0.1 mg), and residual MRC-5 cellular proteins (not more than 2.5 mcg). Neomycin sulfate, an aminoglycoside antibiotic, is included in the cell growth media; only trace amounts (not more than 20 ng) remain following purification. The manufacturing procedures used to manufacture TWINRIX result in a product that contains no more than 5% yeast protein.

 

VAQTA® (HEPATITIS A VACCINE, INACTIVATED)

 

VAQTA* [Hepatitis A Vaccine, Inactivated] is an inactivated whole virus vaccine derived from hepatitis A virus (HAV) grown in cell culture in human MRC-5 diploid fibroblasts. It contains inactivated virus of a strain which was originally derived by further serial passage of a proven attenuated strain. The virus is grown, harvested, purified by a combination of physical and high performance liquid chromatographic techniques developed at the Merck Research Laboratories, formalin inactivated, and then adsorbed onto amorphous aluminum hydroxyphosphate sulfate. One milliliter of the vaccine contains approximately 50 units (U) of hepatitis A virus antigen, which is purified and formulated without a preservative. Within the limits of current assay variability, the 50U dose of VAQTA contains less than 0.1 mcg of non-viral protein, less than 4 x 10–6 mcg of DNA, less than 10–4 mcg of bovine albumin, and less than 0.8 mcg of formaldehyde. Other process chemical residuals are less than 10 parts per billion (ppb).

Pediatric/Adolescent Formulation (12 Months Through 18 Years of Age): each 0.5 mL dose contains approximately 25U of hepatitis A virus antigen adsorbed onto approximately 0.225 mg of aluminum provided as amorphous aluminum hydroxyphosphate sulfate, and 35 mcg of sodium borate as a pH stabilizer, in 0.9% sodium chloride.

Adult Formulation (19 Years of Age and Older): each 1 mL dose contains approximately 50U of hepatitis A virus antigen adsorbed onto approximately 0.45 mg of aluminum provided as amorphous aluminum hydroxyphosphate sulfate, and 70 mcg of sodium borate as a pH stabilizer, in 0.9% sodium chloride.

 

HAVRIX ®(Hepatitis A Vaccine)

 

HAVRIX (Hepatitis A Vaccine) is a sterile suspension of inactivated virus for intramuscular administration. The virus (strain HM175) is propagated in MRC-5 human diploid cells. After removal of the cell culture medium, the cells are lysed to form a suspension. This suspension is purified through ultrafiltration and gel permeation chromatography procedures. Treatment of this lysate with formalin ensures viral inactivation. Viral antigen activity is referenced to a standard using an enzyme linked immunosorbent assay (ELISA), and is therefore expressed in terms of ELISA Units (EL.U.).

Each 1-mL adult dose of vaccine consists of 1440 EL.U. of viral antigen, adsorbed on 0.5 mg of aluminum as aluminum hydroxide.

 

Each 0.5-mL pediatric dose of vaccine consists of 720 EL.U. of viral antigen, adsorbed onto 0.25 mg of aluminum as aluminum hydroxide.

 

HAVRIX contains the following excipients: Amino acid supplement (0.3% w/v) in a phosphate-buffered saline solution and polysorbate 20 (0.05 mg/mL). From the manufacturing process, HAVRIX also contains residual MRC-5 cellular proteins (not more than 5 mcg/mL), formalin (not more than 0.1 mg/mL), and neomycin sulfate (not more than 40 ng/mL), an aminoglycoside antibiotic included in the cell growth media.

 

Hepatitis B Strains

Hepatitis B Strains

  

HBV is a mostly double-stranded DNA virus in the Hepadnaviridae family. HBV causes hepatitis in human and related virus in this family cause hepatitis in ducks, ground squirrels and woodchucks. The HBV genome has four genes: pol, env, pre-core and X that respectively encode the viral DNA-polymerase, envelope protein, pre-core protein (which is processed to viral capsid) and protein X. The function of protein X is not clear but it may be involved in the activation of host cell genes and the development of cancer.

 

Organization of the HBV Genome

The genomes of more than a dozen isolates of hepatitis B virus have been cloned and the complete nucleotide sequences determined. Analysis of the coding potential of the genome reveals four open reading frames (ORFs) which are conserved between all of these isolates.

The first ORF encodes the various forms of the surface protein and contains three in-frame methionine codons which are used for initiation of translation. A second promoter is located upstream of the pre-S1 initiation codon. This directs the synthesis of a 2.4 kb mRNA which is co-terminal with the other surface messages and is translated to yield the large (pre-S1) surface proteins.

The core open reading frame also has two in-phase initiation codons. The “precore” region is highly conserved, has the properties of a signal sequence and is responsible for the secretion of HBeAg.

The third ORF, which is the largest and overlaps the other three, encodes the viral polymerase. This protein appears to be another translation product of the 3.5 kb RNA, and is synthesized apparently following internal initiation of the ribosome.

The amino terminal domain is believed to be the protein primer for minus strand synthesis. There is then a spacer region followed by the (RNA and DNA-dependent) DNA polymerase.

The fourth ORF was designated “x” because the function of its small gene product was not known. However, “x” has now been demonstrated to be a transcriptional transactivator.

…The discovery of variation in the epitopes presented on the surface of the virions and subviral particles identified several subtypes of HBV which differ in their geographical distribution. All isolates of the virus share a common epitope, a, which is a domain of the major surface protein which is believed to protrude as a double loop from the surface of the particle. Two other pairs of mutually exclusive antigenic determinants, d or y and w or r, are also present on the major surface protein. These variations have been correlated with single nucleotide changes in the surface ORF which lead to variation in single amino acids in the protein. Four principal subtypes of HBV are recognized: adw, adr, ayw and ayr. Subtype adw predominates in northern Europe, the Americas and Australasia and also is found in Africa and Asia. Subtype ayw is found in the Mediterranean region, eastern Europe, northern and western Africa, the near East and the Indian subcontinent. In the Far East, adr predominates. But the rarer ayr occasionally may be found in Japan and Papua New Guinea.

The major response of recipients of hepatitis B vaccine is to the common a epitope with consequent protection against all subtypes of the virus. First generation vaccines were prepared from 22 nm HBsAg particles purified from plasma donations from chronic carriers. These preparations are safe and immunogenic but have been superseded in some countries by recombinant vaccines produced by the expression of HBsAg in yeast cells. The expression plasmid contains only the 3′ portion of the HBV surface ORF and only the major surface protein, without pre-S epitopes, is produced. Vaccines containing pre-S2 and pre-S1, as well as the major surface proteins expressed by recombinant DNA technology, are undergoing clinical trials.

Types of HBV Genomes


The following is a list of the major types of HBV genomes found in the human population:

  • 1993: Genetic relatedness of hepatitis B viral strains of diverse geographical origin and natural variations in the primary structure of the surface antigen.
  • 1995: Subtypes, genotypes and molecular epidemiology of the hepatitis B virus as reflected by sequence variability of the S-gene.
  • 1998: Antigenic diversity of hepatitis B virus strains of genotype F in Amerindians and other population groups from Venezuela.

6 Genotypes (A,B,C,D,E,F) [PMID: 8336122]

  • Group A – Orig – N. Europe – Sub-Saharan Africa
  • Group B – Confined to – Eastern Asia (China)
  • Group C – Far East (Japan)
  • Group D – Mediterranean – Near, Mid East, South Asia
  • Group E – W. Sub-Saharan Africa, south to Angola
  • Group F – New World – Brazil, N. + S. America

121 Strains Exist as Quasispecies

There are four serotypes which are based on subtypes of the hepatitis B surface antigen (HBsAg). These are defined by two mutually exclusive determinant pairs d/y and w/r with a common determinant ‘a’. These subtypes are adw, ayw, adr, and ayr.

Four genomic groups of HBV were later referred as genotypes designated with A-D. Sequencing of the S-gene of HBV is the molecular basis for the assessment of the serological variations of HBsAg within the major four subtypes. Two new genotypes of HBV are designated with E and F. The F genotype diverges from other HBV genomes sequenced by 14%. So far, it is the most divergent HBV genome. Worldwide molecular epidemiology of HBV is based on the variability of the S-gene. The E and F strains appear to originate from aboriginal populations of Africa in the New World.

Characterization of genotype H hepatitis B virus strain identified for the first time from a Japanese blood donor by nucleic acid amplification test

Hepatitis B virus (HBV) has been classified into seven genotypes A–G. However, recently genotype H, which is phylogenetically closely related to genotype F, has been reported (Arauz-Ruiz et al., 2002 ). These genotypes of HBV show a distinctive geographical distribution and a relevance to clinical severity (Mayert et al., 1999 ; Kobayashi et al., 2002 ; Locarnini, 2002)

Possible New Hepatitis B Virus Genotype, Southeast Asia

(Emerging Infectious Diseases Volume 14, Number 11–November 2008)

Abstract
We conducted a phylogenetic analysis of 19 hepatitis B virus strains from Laos that belonged to 2 subgenotypes of a new genotype I. This emerging new genotype likely developed outside Southeast Asia and is now found in mixed infections and in recombinations with local strains in a geographically confined region.

 

Hepatitis B Vaccines:

 

 

COMVAX® [HAEMOPHILUS b CONJUGATE (MENINGOCOCCAL PROTEIN CONJUGATE) and HEPATITIS B (RECOMBINANT) VACCINE]

 

HBsAg is produced in recombinant yeast cells. A portion of the hepatitis B virus gene, coding for HBsAg, is cloned into yeast, and the vaccine for hepatitis B is produced from cultures of this recombinant yeast strain according to methods developed in the Merck Research Laboratories. The antigen is harvested and purified from fermentation cultures of a recombinant strain of the yeast Saccharomyces cerevisiae containing the gene for the adw subtype of HBsAg. The fermentation process involves growth of Saccharomyces cerevisiae on a complex fermentation medium which consists of an extract of yeast, soy peptone, dextrose, amino acids and mineral salts.

The HBsAg protein is released from the yeast cells by mechanical cell disruption and detergent

extraction, and purified by a series of physical and chemical methods, which includes ion and hydrophobic chromatography, and diafiltration. The purified protein is treated in phosphate buffer with formaldehyde and then coprecipitated with alum (potassium aluminum sulfate) to form bulk vaccine adjuvanted with amorphous aluminum hydroxyphosphate sulfate. The vaccine contains no detectable yeast DNA, and 1% or less of the protein is of yeast origin.

The individual PRP-OMPC and HBsAg adjuvanted bulks are combined to produce COMVAX. Each 0.5 mL dose of COMVAX is formulated to contain 7.5 mcg PRP conjugated to approximately 125 mcg OMPC, 5 mcg HBsAg, approximately 225 mcg aluminum as amorphous aluminum hydroxyphosphate sulfate, and 35 mcg sodium borate (decahydrate) as a pH stabilizer, in 0.9% sodium chloride. The vaccine contains not more than 0.0004% (w/v) residual formaldehyde.

The potency of the PRP-OMPC component is measured by quantitating the polysaccharide

concentration by an HPLC method. The potency of the HBsAg component is measured relative to a standard by an in vitro immunoassay.

ENGERIX-B® [Hepatitis B Vaccine (Recombinant)]

ENGERIX-B [Hepatitis B Vaccine (Recombinant)] is a noninfectious recombinant DNA hepatitis B vaccine developed and manufactured by GlaxoSmithKline Biologicals. It contains purified surface antigen of the virus obtained by culturing genetically engineered Saccharomyces cerevisiae cells, which carry the surface antigen gene of the hepatitis B virus. The surface antigen expressed in Saccharomyces cerevisiae cells is purified by several physicochemical steps and formulated as a suspension of the antigen adsorbed on aluminum hydroxide. The procedures used to manufacture ENGERIX-B result in a product that contains no more than 5% yeast protein. No substances of human origin are used in its manufacture.

Pediatric/Adolescent: Each 0.5-mL dose contains 10 mcg of hepatitis B surface antigen adsorbed on 0.25 mg aluminum as aluminum hydroxide. The pediatric formulation contains sodium chloride (9 mg/mL) and phosphate buffers (disodium phosphate dihydrate, 0.98 mg/mL; sodium dihydrogen phosphate dihydrate, 0.71 mg/mL).

Adult: Each 1-mL adult dose contains 20 mcg of hepatitis B surface antigen adsorbed on 0.5 mg aluminum as aluminum hydroxide. The adult formulation contains sodium chloride (9 mg/mL) and phosphate buffers (disodium phosphate dihydrate, 0.98 mg/mL; sodium dihydrogen phosphate dihydrate, 0.71 mg/mL).

PEDIARIX®[Diphtheria and Tetanus Toxoids and Acellular Pertussis Adsorbed, Hepatitis B (Recombinant) and Inactivated Poliovirus Vaccine Combined]

 

It contains diphtheria and tetanus toxoids, 3 pertussis antigens (inactivated pertussis toxin [PT] and formaldehyde-treated filamentous hemagglutinin [FHA] and pertactin [69 kiloDalton outer membrane protein]), hepatitis B surface antigen, plus poliovirus Type 1 (Mahoney), Type 2 (MEF-1), and Type 3 (Saukett). The diphtheria toxoid, tetanus toxoid, and pertussis antigens are the same as those in

INFANRIX® (Diphtheria and Tetanus Toxoids and Acellular Pertussis Vaccine Adsorbed). The hepatitis B surface antigen is the same as that in ENGERIX-B® [Hepatitis B Vaccine (Recombinant)].

…The hepatitis B surface antigen (HBsAg) is obtained by culturing genetically engineered Saccharomyces cerevisiae cells, which carry the surface antigen gene of the hepatitis B virus, in synthetic medium. The surface antigen expressed in the S. cerevisiae cells is purified by several physiochemical steps, which include precipitation, ion exchange chromatography, and ultrafiltration.

…The diphtheria, tetanus, and pertussis antigens are individually adsorbed onto aluminum hydroxide; hepatitis B component is adsorbed onto aluminum phosphate. All antigens are then diluted and combined to produce the final formulated vaccine. Each 0.5-mL dose is formulated to contain 25 Lf of diphtheria toxoid, 10 Lf of tetanus toxoid, 25 mcg of inactivated PT, 25 mcg of FHA, 8 mcg of pertactin, 10 mcg of HBsAg, 40 D-antigen Units (DU) of Type 1 poliovirus, 8 DU of Type 2 poliovirus, and 32 DU of Type 3 poliovirus.

TWINRIX® [Hepatitis A Inactivated & Hepatitis B (Recombinant) Vaccine]

 

TWINRIX® [Hepatitis A Inactivated & Hepatitis B (Recombinant) Vaccine] is a sterile bivalent vaccine containing the antigenic components used in producing HAVRIX® (Hepatitis A Vaccine, Inactivated) and ENGERIX-B® [Hepatitis B Vaccine (Recombinant)].

The purified hepatitis B surface antigen (HBsAg) is obtained by culturing genetically engineered Saccharomyces cerevisiae cells, which carry the surface antigen gene of the hepatitis B virus, in synthetic media containing inorganic salts, amino acids, dextrose, and vitamins. Bulk preparations of each antigen are adsorbed separately onto aluminum salts and then pooled during formulation.

A 1.0-mL dose of vaccine contains 720 ELISA Units of inactivated hepatitis A virus and 20 mcg of recombinant HBsAg protein. One dose of vaccine also contains 0.45 mg of aluminum in the form of aluminum phosphate and aluminum hydroxide as adjuvants, amino acids, sodium chloride, phosphate buffer, polysorbate 20, Water for Injection, traces of formalin (not more than 0.1 mg), and residual MRC-5 cellular proteins (not more than 2.5 mcg). Neomycin sulfate, an aminoglycoside antibiotic, is included in the cell growth media; only trace amounts (not more than 20 ng) remain following purification. The manufacturing procedures used to manufacture TWINRIX result in a product that contains no more than 5% yeast protein.

RECOMBIVAX HB® HEPATITIS B VACCINE (RECOMBINANT)

 

RECOMBIVAX HB* Hepatitis B Vaccine (Recombinant) is a non-infectious subunit viral vaccine

derived from hepatitis B surface antigen (HBsAg) produced in yeast cells. A portion of the hepatitis B virus gene, coding for HBsAg, is cloned into yeast, and the vaccine for hepatitis B is produced from cultures of this recombinant yeast strain according to methods developed in the Merck Research Laboratories.

The antigen is harvested and purified from fermentation cultures of a recombinant strain of the yeast Saccharomyces cerevisiae containing the gene for the adw subtype of HBsAg. The fermentation process involves growth of Saccharomyces cerevisiae on a complex fermentation medium which consists of an extract of yeast, soy peptone, dextrose, amino acids and mineral salts. The HBsAg protein is released from the yeast cells by cell disruption and purified by a series of physical and chemical methods. The purified protein is treated in phosphate buffer with formaldehyde and then coprecipitated with alum (potassium aluminum sulfate) to form bulk vaccine adjuvanted with amorphous aluminum

hydroxyphosphate sulfate. The vaccine contains no detectable yeast DNA but may contain not more than 1% yeast protein. The vaccine produced by the Merck method has been shown to be comparable to the plasma-derived vaccine in terms of animal potency (mouse, monkey, and chimpanzee) and protective efficacy (chimpanzee and human).

Pediatric/Adolescent Formulation (Without Preservative), 10 mcg/mL: each 0.5 mL dose contains 5 mcg of hepatitis B surface antigen.

Adult Formulation (Without Preservative), 10 mcg/mL: each 1 mL dose contains 10 mcg of

hepatitis B surface antigen.

Dialysis Formulation (Without Preservative), 40 mcg/mL: each 1 mL dose contains 40 mcg of hepatitis B surface antigen.

All formulations contain approximately 0.5 mg of aluminum (provided as amorphous aluminum

hydroxyphosphate sulfate, previously referred to as aluminum hydroxide) per mL of vaccine. In each formulation, hepatitis B surface antigen is adsorbed onto approximately 0.5 mg of aluminum (provided as amorphous aluminum hydroxyphosphate sulfate) per mL of vaccine. The vaccine is of the adw subtype.

RECOMBIVAX HB is indicated for vaccination of persons at risk of infection from hepatitis B virus

including all known subtypes. RECOMBIVAX HB Dialysis Formulation is indicated for vaccination of adult predialysis and dialysis patients against infection caused by all known subtypes of hepatitis B virus.

 Nabi-HB® Hepatitis B Immune Globulin (Human)

 

DESCRIPTION

 

Hepatitis B Immune Globulin (Human), Nabi-HB, is a sterile solution of immunoglobulin (5 ± 1%

protein) containing antibodies to hepatitis B surface antigen (anti-HBs). It is prepared from plasma

donated by individuals with high titers of anti-HBs. The plasma is processed using a modified

Cohn 6 / Oncley 9 cold-alcohol fractionation process1,2 with two added viral reduction steps

described below. Nabi-HB is formulated in 0.075 M sodium chloride, 0.15 M glycine, and 0.01%

polysorbate 80, at pH 6.2. The product is supplied as a nonturbid sterile liquid in single dose

vials and appears as clear to opalescent. It contains no preservative and is intended for single

use by the intramuscular route only.

The manufacturing steps for Nabi-HB are designed to reduce the risk of transmission of viral disease.

The solvent/detergent treatment step, using tri-n-butyl phosphate and Triton® X-100, is

effective in inactivating known enveloped viruses such as hepatitis B virus (HBV), hepatitis C

virus (HCV), and human immunodeficiency virus (HIV) 3. Virus filtration, using a Planova® 35

nm Virus Filter, is effective in reducing some known enveloped and non-enveloped viruses4. The

inactivation and reduction of known enveloped and non-enveloped model viruses were validated

in laboratory studies as summarized in the following table…

BayHepB Immune Globulin (Human)

Pertussis and Transmission

Pertussis and Transmission

 I’m sure we’ve all seen the campaign ads called “Do it for your baby”. If not, here is an example. There is also a website.

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More recently, J. Lo was recruited by Sanofi Pasteur and the March of Dimes to jump on the bandwagon and promote Pertussis vaccines for adolescents and adults.

 Let’s take a look at the issue of Pertussis transmission and what the vaccine really does or doesn’t do.

Pertussis Infection in Fully Vaccinated Children in Day-Care Centers, Israel (Emerging Infectious Diseases (Vol. 6, No. 5, September–October 2000)

 Conclusions

 The effects of whole-cell pertussis vaccine wane after 5 to 10 years, and infection in a vaccinated person causes nonspecific symptoms (3-7). Vaccinated adolescents and adults may serve as reservoirs for silent infection and become potential transmitters to unprotected infants (3-11). The whole-cell vaccine for pertussis is protective only against clinical disease, not against infection (15-17). Therefore, even young, recently vaccinated children may serve as reservoirs and potential transmitters of infection.

 

…We found that immunity does not even persist into early childhood in some cases. We also observed that DPT vaccine does not fully protect children against the level of clinical disease defined by WHO. Our  results indicate that children ages 5-6 years and possibly younger, ages 2-3 years, play a role as silent reservoirs in the transmission of pertussis in the community. More studies are needed to find the immunologic basis of protection against infection and colonization and thus an effective way to eradicate pertussis.

 Also See:

Characterization of Bactericidal Immune Responses following Vaccination with Acellular Pertussis Vaccines in Adults

booster immunization of adults with acellular pertussis vaccines was not found to increase bactericidal activity over preimmunization levels. Identifying ways to promote bactericidal immune responses might improve the efficacy of acellular pertussis vaccines.

Informal consultation on the control of pertussis with whole cell and acellular vaccines

Dr Cherry pointed out in the 1999 WHO meeting, that even with a return to high vaccination coverage in Japan, pertussis incidence in children less than three months of age, had not declined substantially.

 

Pertussis toxin inhibits neutrophil recruitment to delay antibody-mediated clearance of Bordetella pertussis 

…However, their efficacy against subclinical infection is doubtful, as the majority of vaccinated populations test positive for subsequent infection (10, 17), suggesting that the bacterium successfully infects immune and/or vaccinated individuals.

  

The Pertussis vaccines do not prevent transmission or infection. It simply prevents or suppresses the symptoms, or causes subclinical infection. The reason the vaccines do not prevent transmission is because they do not kill the bacteria that cause the disease. The vaccines only make the body able to resist the toxin that the bacteria release. The toxin is what makes a person sick and causes clinical disease. That is the reason behind the claim that being vaccinated will make a clinical case of Pertussis ‘milder’. Even if you don’t get the full effect of the vaccine, you might get partial blockage of the toxin which might cause you to cough less, etc.

What is also worth noting is the small print of an advertisement for Adacel, a DTaP booster:

“It is unknown whether immunizing adolescents and adults against pertussis will reduce the risk of transmission to infants.”

At least there was some honesty there. The adolescent or adult booster helps one person, the person who receives it! Why doesn’t the campaign advertisement just say “Do it for you!” They don’t because time as shown repeatedly that the majority of adults will not do booster vaccines. But, if they use infants or children as a reason, it works.

 Read on…

 Acellular Pertussis Vaccines and Complement Killing of Bordetella pertussis

At least one of the antigens in the acellular pertussis vaccine appears to be able to serve as a target for complement-mediated bactericidal activity. However, in this study and other studies (19, 21), improved bactericidal responses after immunization were rarely observed, possibly due to induction of antibodies that fail to fix complement. The absence of vaccine-induced bactericidal activity in vitro is consistent with the observation that the pertussis vaccine is effective at preventing severe disease, likely due to pertussis toxin neutralization and blocking attachment to reduce bacterial colonization, but it is less effective at producing a sterilizing immune response (5, 18). Despite high vaccination rates, the number of reported cases of pertussis in the United States has increased steadily since the 1980s (22). Developing a pertussis vaccine with a greater potential to elicit bactericidal activity could reduce bacterial carriage and reduce the incidence of disease.

 

 According to this study, some were less likely to be protected, and the majorities were no more likely to be protected from infection than they were before the vaccine. To boot, the vaccine made them more susceptible to infection. There was a level of protection from severe disease due to the toxoid in the vaccine. This study alone shows that the vaccine doesn’t reduce carriage of the bacteria.  Therefore, marketing the vaccines solely to protect infants isn’t being entirely truthful. It simply does not work that way. The vaccine can’t stop you from being infected, nor stop you from spreading the bacteria to infants, the immuno-compromised, the unvaccinated or vaccinated.  

 

 It is also acknowledged that the whole-cell pertussis vaccine was more efficacious, but also more reactive, hence why we now use the DTaP.

Use of Pertussis Vaccines in Outbreaks

Estimates of the efficacy of immunization with pertussis vaccines are subject to wide variation due to variation in study design, including factors such as case definition, case ascertainment, and duration of follow up. A recent study of reported cases in the United States in 1992 – 1994 estimated the effectiveness of whole-cell vaccine against culture proven pertussis; the effectiveness of three doses among children aged 7-18 months was 79% and the effectiveness of 4 doses among children aged 19-47 months was 90%.2

Estimates are also available from recent field trials that compared the efficacy of acellular and whole-cell pertussis vaccines. In these trials, the efficacy of three doses of whole cell pertussis vaccines from different manufacturers ranged from 83% to 98%, although the DTP vaccine from one manufacturer had a low efficacy in two trials (35% to 48%). In the field trials, the efficacy of the four licensed acellular vaccines ranged from 71% to 89%.

 

 

When is a true case a case… or not? How does case definition come into play?

The Science and Fiction of Pertussis Vaccines  (PEDIATRICS Vol. 104 No. 6 December 1999, pp. 1381-1383)

Case definition has been particularly problematic in all of the recent DTaP vaccine efficacy trials. For uniform comparative purposes a case definition was suggested by a WHO expert committee. This definition required 21 days of paroxysmal cough plus laboratory confirmation of pertussis in the subject or household contact. There are 2 problems with this definition. The first is that a substantial number of B pertussis infections in unvaccinated children are mild and would not meet the case definition. The second is that all pertussis vaccines tend to modify duration and severity of disease rather than completely preventing illness. Therefore, the WHO definition has made all vaccines look better than they are and it has tended to obscure differences between vaccines.

 

Easily Missed?

Whooping cough is a common respiratory infection caused by the bacterium Bordetella pertussis. It should be considered as a possible diagnosis in any adolescent or adult with an acute cough of more than two weeks’ duration, even if they have been fully immunised.

 

Whooping cough in school age children with persistent cough: prospective cohort study in primary care

 

…Studies in the United States report a 20% incidence of Bordetella pertussis infection among adults with a persistent cough.2 Despite data showing that neither infection nor immunisation results in lifelong immunity, whooping cough is seldom diagnosed in primary care because of the lack of specificity of clinical symptoms and signs. Whooping cough is perceived as a disease of very young children who have not been immunised and who have classic features such as whoop…

…General practitioners should be alert to a potential diagnosis of whooping cough in any child who presents with a persistent cough. We found that children with pertussis cough for a median of 16 weeks. Little evidence indicates that administering erythromycin to children with pertussis two weeks after they have contracted the infection either reduces symptoms or prevents transmission.10

 

Acellular Pertussis Vaccines and Complement Killing of Bordetella pertussis 

 

When examined individually, the pre- versus postimmunization bactericidal activity was not significantly different at any dilution tested for 8 of the 15 acellular vaccine recipients. The individuals in the group with unchanged activity after immunization included an individual with undetectable preimmunization activity against the wild-type strain (individual 32-60), and the individual with the highest preimmunization activity (individual 26-47) (Fig. 2). These results suggest that the level of preimmunization bactericidal activity does not necessarily influence the ability to generate a postimmunization response.

 

However, statistically significant differences between pre- and post immunization bactericidal activity were observed (P < 0.05) using the paired t test for at least one serum dilution for 7 of the 15 acellular vaccine recipients. Furthermore, evidence of both improved bactericidal activity and reduced bactericidal activity after immunization was found in these seven serum samples. Four individuals displayed improved bactericidal activity after immunization when serum was added at lower concentrations (1.0 or 0.10%) but not at 10% (Fig. 3). However, in addition to enhanced bactericidal activity, individuals 4-43 and 20-55 displayed evidence of blocking activity, since fewer bacteria were killed when serum was added at 10% than when serum was added at 1%. Blocking activity could occur when antibodies that do not fix complement compete with complement-fixing antibodies for access to antigen. More definitive evidence of blocking was demonstrated in three other individuals (Fig. 4). For these individuals, the post immunization serum samples had significantly less bactericidal activity than the pre immunization serum samples at a serum concentration of 10%.

However, in this study and other studies (19, 21), improved bactericidal responses after immunization were rarely observed, possibly due to induction of antibodies that fail to fix complement. The absence of vaccine-induced bactericidal activity in vitro is consistent with the observation that the pertussis vaccine is effective at preventing severe disease, likely due to pertussis toxin neutralization and blocking attachment to reduce bacterial colonization, but it is less effective at producing a sterilizing immune response (5, 18).

 In various studies, vaccinated children do not fit the case definition of pertussis, yet they test positive. They exhibit atypical or mild symptoms. This shows the vaccine may protect against clinical disease, but not against infection.  Vaccinated children and adults are still serving as asymptomatic reservoirs and transmit infection.  A reduction in colonization, without any kind of sterilizing immune response, is the most you will get. There should be an enhanced bacterial clearance in the vaccinated, but studies have proved there isn’t. What is even worse is, serology states that more than half of are catching pertussis every 2 1/2 years. If humans did not transmit pertussis easily, or could clear it easier, we wouldn’t have it so much. The vaccine has not changed that. Vaccinated or not, you can and do, carry and transmit the disease.

 

 Other Studies:

 

Cell-mediated immune responses to antigens of Bordetella pertussis and protection against pertussis in school children  (Pediatric Infection Disease Journal 1999; 18: 366-370)

 Abstract

Background. Increasing evidence suggests that cell-mediated immunity (CMI) is involved in immune response against Bordetella pertussis. However, there are practically no studies evaluating the significance of pertussis-specific CMI in relation to protection against clinical pertussis.

 

On pg. 366: “The immunologic mechanisms of protection against clinical pertussis are poorly understood. Although several studies have suggested that antibodies to some pertussis antigens may be predictive of protection against pertussis, there is no generally accepted laboratory measure of immunity. Further in clinical efficacy trials of acellular vaccines, no clear correlation has been found between serum antibody values and protection. 

…”In the present study, no clear association was found between serum antibody values and clinical outcome.”

 

Pertussis toxin: the cause of the harmful effects and prolonged immunity of whooping cough. A hypothesis. (Rev Infect Dis. ;1 (3):401-12 233166)

The nature of the pathogenesis and of the prolonged immunity of whooping cough has not been clearly defined. The literature of Bordetella pertussis indicated that only the antigen that induces histamine sensitization, lymphocytosis, and other biological reactions in mice is the cause of the harmful effects and prolonged immunity of whooping cough. This antigen has the general characteristics of bacterial protein exotoxins that cause the harmful effects of infectious diseases such as diphtheria and tetanus. It is proposed that this antigen, which is histamine-sensitizing, lymphocyte-leukocyte-promoting, and islets-activating (HSF-LPF-IAP), be designated pertussis toxin. Agglutinogen, hemagglutinin, and heat-labile (at 56 C) and heat-stable (at 100 C) toxins are no doubt interrelated with the immunologic and/or toxic reactions of whooping cough. It appears that the first defense against the disease is the antibody that prevents adhesion of the bacteria to the cilia of the respiratory epithelium and that the second defense is the antitoxin against pertussis toxin (HSF-LPF-IAP).

 

 Those who have been vaccinated are still considered susceptible, unless they have had a natural case of Pertussis, because a natural case can protect them at a higher rate than the vaccine. A vaccine may give them a less severe case with exposure, but it has been shown repeatedly that it does not prevent transmission. The vaccine for adolescents and adults is solely to prevent them from getting a clinical case and passing it on the infant. But overall, it just extends the length of time before they do/can get it.

The vaccine can provide some ‘herd immunity’. How? It can postpone, suppress the disease, or make a less severe case. But herd immunity is limited. The chance of catching pertussis at some point in ones life is inevitable. People can also get Pertussis more than once. This natural cycle of the disease also provides herd immunity. Pertussis has virulent cycles of every 2-5 years and vaccines have not changed its natural virulence cycle.

 Underreporting of Pertussis has been acknowledged repeatedly. How does this correlate to the reduced disease burden? Simply put-It is only a guess.

Reemergence of Pertussis: Methods

 

Data from pertussis reporting (required by law since 1976) were obtained from 1976 to 1998 from the Inspectorate of Health. A case definition, introduced in 1988, included clinical symptoms and laboratory confirmation (or close contact with a person with laboratory-confirmed pertussis).

…From 1988 to April 1997, laboratory confirmation was defined as either a positive culture of B. pertussis (or B. parapertussis) or positive two-point serology, in turn defined as a significant rise of immunoglobulin (Ig) G antibodies against pertussis toxin or IgA antibodies against B. pertussis in paired sera. In April 1997, a positive polymerase chain reaction (PCR) and positive one-point serology were also accepted as laboratory confirmation. Positive one-point serology was defined as high IgG or IgA antibody titers in a single serum sample…

 

 Pertussis in Adults

 …In the prevaccine era, reported pertussis was a cyclic disease, with epidemic peaks every 2 to 5 years [9]. When pertussis was brought under control by vaccination of children in the 1960s in the United States and England and Wales, it was noted that the 2– to 5-year cyclic pattern continuedFailure of the pertussis cycle to lengthen led to Fine and Clarkson’s suggestion [11] that immunization controlled the disease in children but did not disrupt circulation of the organism. This observation and the knowledge that adults were a common source of disease in infants led our group and others to study the epidemiology of adult pertussis.

…contrast with our findings in the UCLA students (94% of whom had been vaccinated in childhood), we found that German adults (most of whom had not been vaccinated during childhood) were more likely to have typical pertussis with whooping and post-tussive vomiting [12]. Twenty-six percent of these adults had had pertussis during childhood.

Immunoglobulin A antibodies to B. pertussis antigens usually result from infection, not immunization. With this fact in mind, we examined the prevalence and degree of IgA antibody to four B. pertussis antigens in young adults in the United States and Germany [18]. We found that the mean titers and the prevalence of antibody to the four antigens were similar, suggesting that the circulation of B. pertussis in adults in the two countries was similar even though pertussis was epidemic in Germany and rare in the United States. In another study [19], in which we obtained serum samples yearly for 5 years from 51 persons, 90% of these persons had serologic evidence of at least one case of pertussis [19].

 

Essential Problems in Pertussis (Am J Public Health, Apr 1939; 29: 337 – 340).

…It would seem reasonable to expect that the case fatality of pertussis might have been favorably affected bv the better home hygiene, antirachitic and other nutritional advances in the dietaries of children, more general and skilled home nursing of patients during isolation, and other factors which we see applied for the benefit of children under 5 years of age, but there are, I believe, no studies to confirm these general impressions.

 

…Second, one finds a very considerable incompleteness of reporting, the extent of which varies widely according to the interest and adequacy of the local health and medical services of the community, from approximate completeness to about 5 per cent of estimated cases.

Third, one finds that routine departmental procedure in establishing the diagnosis and recording the presumed susceptibility, exposure, and subsequent history of infection in households where other children than the reported patient are living, is neither uniform nor adequate to permit study of the relative merits of isolation periods of different lengths, of inoculations intended to prevent, or of therapy designed to modify the course of the disease.

 

 Pertussis surveillance

…Surveillance is carried out in some countries, but is not done in any meaningful way in three quarters of the world. Surveillance data and coverage data are both unreliable. Reliable surveillance data are needed to check coverage and vaccine efficacy; both surveillance and coverage data are needed to monitor immunization programmes…

… There are two principal methods of estimating burden: the natural history method and the proportional mortality method. The general approach used by WHO to estimate disease burden is to start with an expert consultative process in order to develop a sound approach. The methodology should aim to use the best data available and seek to validate results with existing data. Sensitivity analysis and continuous critical review are the only guarantees that the approach is as sound as it can be.

The estimates continue to be refined as new information becomes available. The final product is a database of cases and deaths by age, sex, country and year, as well as a careful documentation of methods, assumptions, and data sources. The process should result in recommendations on how to improve the precision, robustness and usefulness of the estimates.

Factors that affect the estimates are epidemiologic, demographic and programmatic, as well as co-factors such as HIV prevalence and nutritional status. Other practical issues include the quality and generalizability of the input data. Validation of the estimates is always important. The process is often messy since reliable data may be lacking, broad extrapolations or generalizations are made, or there is a heavy reliance on “expert opinion”.

 

 Prevention of Pertussis among Adolescents by Vaccination: Taking Action on What We Know and Acknowledging What We Do Not Know

…But there is much that is not known about pertussis. It is unclear whether the recent increases in reported disease are real or are artifacts of increased recognition; the increase in reported cases among young infants, coupled with relatively stable rates of reported disease among older infants and preschool-aged children, suggests that there may be a real increase in the circulation of Bordetella pertussis in some age groups [1]. But there is no doubt that ascertainment of pertussis is variable and incomplete in most age groups. Physicians may not consider the diagnosis, especially in adolescents and adults, because of a lack of clinical awareness that pertussis occurs in these age groups. Diagnostic testing is imperfect, and some tests have not been well standardized. Culture of B. pertussis remains the gold standard by which other assays are judged, but unless the diagnosis is considered early in the course of illness and before administration of antimicrobial therapy, isolation of the bacterium is unlikely. Serological testing, once standardized, may facilitate diagnosis, but it remains unavailable in most areas, and assays based on PCR are variable in sensitivity and specificity [2]. Thus, our knowledge of the burden of pertussis is far from complete.

There is also much that we do not know about the dynamics of B. pertussis transmission. It is unclear what impact vaccinating young adolescents would have on disease incidence in other age groups. Do middle and high schools, with their high contact rates and susceptible populations, amplify B. pertussis circulation in the community? If routine vaccination of young adolescents prevented those outbreaks but immunity was not long lasting, would outbreaks then occur among young adults? Would transmission to young infants—the group with the highest morbidity and mortality due to pertussis—decrease or increase following implementation of an adolescent pertussis vaccination program? Mathematical modeling suggests that the impact of routine adult pertussis vaccination on the incidence of pertussis in young children may be relatively modest [3].

Because of these and other uncertainties, estimating the impact of pertussis vaccination of adolescents and adults on disease burden requires many assumptions….

 

Epidemiology of pertussis

 

 …In the majority of countries where pertussis is a notifiable disease, a case-based national surveillance system is in place. However, different case definitions, methods of diagnosis and reporting and surveillance systems make direct intercountry comparisons difficult, and pertussis is not a statutory notifiable disease in every country. Nevertheless the general consensus is that reported incidences are probably considerably lower than the actual incidence of pertussis; underreporting is common. Prolonged cough may be the only clinical feature in adolescents or adults, who may present for diagnosis late (precluding laboratory confirmation) or not at all. When they do present, their condition is often misdiagnosed because, in part, clinicians continue to perceive pertussis as a childhood disease.Despite underreporting, an increased incidence of infant, adolescent and adult pertussis has been observed worldwide since the introduction of widespread vaccination… 

 

 Also See:  Failure of Physicians to Consider the Diagnosis of Pertussis in Children

  

More on Adacel (11-64 years):

 Clinical Studies;

 

The efficacy of the tetanus toxoid and diphtheria toxoid used in Adacel vaccine was based on the immune response to these antigens compared to a US licensed Tetanus and Diphtheria Toxoids Adsorbed For Adult Use (Td) vaccine manufactured by Sanofi Pasteur Inc., Swiftwater, PA.

The protective efficacy against mild pertussis (defined as at least one day of cough with laboratory-confirmed B pertussis infection) was 77.9% (95% CI: 72.6%, 82.2%). (8) (9) In addition, the ability of Adacel vaccine to elicit a booster response to the pertussis antigens following vaccination was evaluated. The acellular pertussis formulations for Adacel and DAPTACEL vaccines differ only in the amount of detoxified PT (2.5 μg in Adacel vaccine versus 10 μg in DAPTACEL vaccine).

The primary measures of immunogenicity were (a) the percentage of participants attaining an antibody level of at least 0.1 IU/mL and (b) the percentage of participants achieving a rise in antibody concentration after vaccination (booster response). The demonstration of a booster response depended on the antibody concentration to each antigen prior to immunization.

 Threshold or “cut-off” values for antibody concentrations to each antigen were established based on the 95th percentile of the pre-vaccination antibody concentrations observed in previous clinical trials. A booster response was defined as a four-fold rise in antibody concentration if the pre-vaccination concentration was equal to or below the cut-off value and a two-fold rise in antibody concentration if the pre-vaccination concentration was above the cut-off value.

 The efficacy of the pertussis antigens used in Adacel vaccine was inferred based on a comparison of pertussis antibody levels achieved in recipients of a single booster dose of Adacel vaccine with those obtained in infants after three doses of DAPTACEL vaccine. In the Sweden I Efficacy Trial, three doses of DAPTACEL vaccine were shown to confer a protective efficacy of 84.9% (95% CI: 80.1%, 88.6%) against WHO defined pertussis (21 days of paroxysmal cough with laboratory-confirmed B pertussis infection or epidemiological link to a confirmed case).

 …Pregnancy Category C

 

Animal reproduction studies have not been conducted with Adacel vaccine. It is also not known whether Adacel vaccine can cause fetal harm when administered to a pregnant woman or can affect reproduction capacity. Adacel vaccine should be given to a pregnant woman only if clearly needed…

Nursing Mothers

 

It is not known whether Adacel vaccine is excreted in human milk. Because many drugs are excreted in human milk, caution should be exercised when Adacel vaccine is given to a nursing woman.

The safety and effectiveness of concomitant administration of Adacel vaccine with other vaccines has not been evaluated.

  

 Boostrix (10-18years)

 Prevention of Pertussis, Tetanus, and Diphtheria Among Pregnant and Postpartum Women and Their Infants

In 2006, ACIP recommended routine administration of Tdap for postpartum women who were not vaccinated previously with Tdap to provide personal protection and reduce the risk for transmitting pertussis to their infants (1,2) . After careful consideration, in June 2006, ACIP voted to reaffirm its recommendation for use of Td in pregnant women who have urgent indication for tetanus toxoid or diphtheria toxoid vaccination to prevent maternal or neonatal tetanus, or to prevent diphtheria. Pregnant women not vaccinated previously with Tdap will receive a measure of protection against pertussis by ensuring that children in the household are up-to-date with recommended doses of pediatric diphtheria and tetanus toxoids and acellular pertussis vaccine (DTaP)* (2123) and that adult and adolescent household contacts have received a dose of Tdap (Table 2) (1,2). Health-care providers can monitor pregnant women who have not received a dose of Tdap for exposures to pertussis or to respiratory illness consistent with pertussis, and they can administer antimicrobials for postexposure prophylaxis or treatment of pertussis, if needed, to reduce the risk for transmitting pertussis to their infants.

This report provides the background and rationale for routine administration of Tdap in postpartum women who were not vaccinated previously with Tdap and for maintaining the previous recommendation for use of Td in pregnant women if indicated. The safety and efficacy of using Tdap in pregnant women has not been demonstrated, and Tdap is not recommended for use in pregnant women in any country. No evidence exists of excess morbidity or any fatality among pregnant women ascribed to pertussis. No evidence exists demonstrating whether

  • Tdap in pregnant women harms the fetus or increases risk for adverse pregnancy outcomes,
  • transplacental antibody induced by Tdap administered during pregnancy will protect infants against pertussis, or
  • Tdap-induced transplacental maternal antibody will have a negative impact on an infant’s protective immune response to later-administered routine pediatric DTaP or to conjugate vaccines containing tetanus toxoid or diphtheria toxoid.

Health-care providers should weigh the theoretical risks and benefits before choosing to administer Tdap vaccine to a pregnant woman…

…The specific issues for pertussis differ from those for tetanus and diphtheria. Important among these is the limited understanding of immunity and correlates of protection for pertussis. In addition, data supporting the safety of vaccinating pregnant women with Tdap to prevent pertussis are scarce for women, their fetuses, and pregnancy outcomes. Whether transplacental maternal antibody exerts an inhibitory or other effect on the infant-protective immune response to active immunization with pediatric DTaP or conjugate vaccines containing tetanus toxoid or diphtheria toxoid has not been studied. Protection against infant pertussis through Tdap-induced transplacental maternal antibody has not been demonstrated. Until additional information is available, the majority view of the working group held that Tdap administered to women in the immediate postpartum period, in addition to ensuring pertussis vaccination of close contacts, would likely provide a measure of protection for mother and infant.

 

Vaccinating Pregnant Women against Pertussis

Tdap:

No prelicensure studies were conducted with Tdap in pregnant women. In 2005, to increase understanding of the safety of Tdap in relationship to pregnancy, both Tdap manufacturers established registries to solicit voluntary reports of pregnant women who received Tdap during pregnancy or who received Tdap and were determined subsequently to be pregnant (212,213). The main utility of the registries is to signal the possibility and nature of any risk (214). All women who are vaccinated with Tdap at any time during pregnancy should be reported to the registry as early as possible during the pregnancy. Information from pregnancy registries differs from surveillance reports, which are used to evaluate outcomes among women when an adverse outcome of pregnancy already might have occurred (e.g., an infant born with a birth defect) (214).

Also see: Guidelines for Vaccinating Pregnant Women (ACIP Guidelines May 2008) 

  • Pregnancy is not a contraindication for use of Tdap. Data on safety, immunogenicity and the outcomes of pregnancy are not available for pregnant women who receive Tdap. When Tdap is administered during pregnancy, transplacental maternal antibodies might protect the infant against pertussis in early life. They also could interfere with the infant’s immune response to infant doses of DTaP, and leave the infant less well protected against pertussis. 11
  • ACIP recommends Td when tetanus and diphtheria protection is required during pregnancy. In some situations*, healthcare providers can choose to administer Tdap instead of Td to add protection against pertussis. When Td or Tdap is administered during pregnancy, the second or third trimester is preferred. 11
  • Providers who choose to administer Tdap to pregnant women should discuss the lack of data with the pregnant women and are encouraged to report Tdap administrations regardless of the trimester, to the appropriate manufacturer’s pregnancy registry: for Boostrix® to GlaxoSmithKline Biologicals at 1-888-825-5249, or for Adacel®, to sanofi pasteur at 800-822-2463. 11
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    It appears there is much they ‘don’t know’. But that isn’t stopping them. Vaccinating Post-partum Mothers before leaving the hospital with Tdap, even if they are breastfeeding, has been increasing.

    Investigators in Houston have successfully implemented a novel vaccination strategy – vaccinating women who have just given birth to prevent them transmitting pertussis (whooping cough) to their young infants.  This study took place in a medically underserved and underinsured population.   Study investigators gave tetanus, diphtheria and acellular pertussis (Tdap) vaccine to over 1000 mothers at Ben Taub General Hospital, a county hospital that serves a predominantly Hispanic population in Houston, over a 3 month period.  Approximately 75% of women who were offered Tdap received the vaccine and when Tdap was not administered it was usually because mothers had received a recent tetanus vaccine (a relative precaution to receiving Tdap vaccine).  This program is, to our knowledge, the first to implement and track on a large scale the 2006 Centers for Disease Control and Prevention (CDC) recommendation that women receive Tdap before they leave the hospital after delivery.

    Also see: Forrest General gives recommended vaccine for new mothers