A Microbial Biorealm page on the genus Streptococcus agalactiae
Higher order taxa
Species: Streptococcus agalactiae
Description and significance
Streptococcus is a genus that is classified based on the hemolytic properties into three types: Alpha-Hemolytic Streptococci, Beta-Hemolytic Streptococci, and Non-Hemolytic Streptococci. Streptococcus agalactiae, often referred as Group B Streptococcus (GBS), is one of four Beta-Hemolytic streptococci, which results in complete rupture of blood cells shown in wide and clear areas surrounding bacterial colonies on blood agar. 
Appearance: S. agalactiae is a diplococcal (a pair of cocci, circular, pair) gram-positive, non acid-fast bacterium (~2.0µm) that does not form spores, is not motile, and is catalase-free, which is an enzyme that catalyzes the reduction of hydrogen peroxide). It occurs in pairs or short chains and has group B Lancefield antigen present. 
Habitat: S. agalactiae , originally discovered as a cause of bovine mastitis, is part of the normal bacterial flora colonizing the gastrointestinal(GI) tract and genitourinary tract of a significant proportion of the human population. However, it occasionally become a infectious pathogen colonizing the uterus, blood, brain, and meninges. 
Significance : This pathogen is one of the leading causes of invasive infections in non-pregnant immunocompromised individuals and also causes bacteremia, septicaemia, meningitis, and pneumonia. Colonization of the rectum and vagina of pregnant women with GBS is correlated with GBS sepsis in newborn infants with early onset disease. S. agalactiae is also subclassified into nine serotypes depeding on the immunologic reactivity of the polysaccharide capsule and among nine serotypes, only types Ia, Ib, II, III, and V are discovered to be responsible for invasive human disease. 
Isolation: S. agalactiae can be isolated in infected site of human or in secretions from infected mammary gland of female cattle and related ungulates. In some samples these bacteria are numerous and easily found in stained films; in other cases they may be so scarce that they can be located only with great difficulty. Also, most stains can be used to stain GBS to locate them, since the GBS is gram-positive and readily stained.
Genome project: S. agalactiae poses a serious threat to lives of neonates, responsible for 2-3 cases per 1000 live birth and to lives of human, especially elderly persons and those with weakened immune systems. This microorganism is considered one of the major causes of economic losses to dairy producers without a control program. Because of its significance as an threat to both human and related ungulates, such as cow, its genome was sequenced and still being studied to gain more insight into the virulence factor and to develop treatments and preventive prophylactic antibodies. 
The genome of three strains in Streptococcus agalactiae (GBS) have been completely sequenced: S. agalactiae NEM 316, S. agalactiae 2603V/R, and S. agalactiae A909. The genome of two plasmids also have been completely sequenced: S. agalactiae plasmid pGB3631 and S. agalactiae plasmid pGB354. The genome of known five strains in S. agalactiae have not yet been completely sequenced: S. agalactiae 515, S. agalactiae CJB111, S. agalactiae COH1, S. agalactiae H36B, S. agalactiae 18RS21.
Size of chromosomal genome: S. agalactiae NEM 316 is the serotype III strain. Complete genome sequence of this strain is a (one) circular dsDNA chromosome with 2,211,485 nt (GC content of 35%, coding content of 87%) and contains 2235 genes, 2094 protein coding genes, 101 structural RNAs, and 40 pseudogenes. Genome sequence was completed in 2002/11/15. S. agalactiae 2603V/R is the serotype V strain. Complete genome sequence of this strain is a (one) circular dsDNA chromosome with 2,160,267 nt (GC content of 35%, coding content of 86%) and contains 2271 genes, 2124 protein coding genes, 96 structural RNAs, and no pseudogene. Genome sequence was completed in 2002/08/28. S. agalactiae A909 is the serotype Ic strain. Complete genome sequence of this strain is a (one) circular DNA chromosome with 2,127,839 nt (GC content of 35%, coding content of 86%) and contains 2136 genes, 1996 protein coding genes, 102 structural RNAs, and 32 pseudogenes. Genome sequence was completed in 2005/10/03.
Size of plasmid genome: Wild-type S. agalactiae plasmid pIP501 can be easily transferred and maintained in various S. agalactiae strains and confers antibiotic resistance to its recipients. This plasmid pIP501 can be isolated from various strains of S. agalactiae and various Streptococci species, including S. sanguis and S. faecalis. This plasmid plays importance role in pathogenesis of those bacteria. 
S. agalactiae plasmid pGB3631 is the deletion derivative of the wild-type S. agalactiae plasmid pIP501. Complete genome sequence of this strain is a circular DNA chromosome with 5,842 nt (GC content of 33%, coding content of 62%) and contains 9 genes, 6 protein coding genes, no structural RNA, and no pseudogene. Genome sequence was completed in 1994/07/13. S. agalactiae plasmid pGB354 is the derivative of the wild-type Streptococcus agalactiae plasmid pIP501,. Complete genome sequence of this strain is a circular DNA chromosome with 6,437 nt (GC content of 33%, coding content of 62%) and contains 5 genes, 5 protein coding genes, no structural RNA, and no pseudogene. Genome sequence was completed in 1997/03/04.
Pathogenic Contents: Analysis of NEM316 genome predicted and identify locus responsible for extracellular products like capsular polysaccharide, surface proteins, and secreted proteins, which are involved in virulence and contributing to pathogenesis. NEM 316 strain contains 17 genes (cpsA-L, neuBCDA) along with the transcriptional gene cpsY for sialyated capsular polysacchraide, 30 genes (gbs 0391, 0392, 0393, and 27 more genes) for surface proteins containing cell wall, and various genes responsible for 71 secreted proteins. In genomic analysis of NEM315, it was also revealed to have stress adaption by encoding Clp proteins (gbs 1634, 1383, 1869, 1367, 0535), which is ATP-dependent protease playing a role in virulence. 
Genomic Contents: IIn strain NEM315, it was observed that there are 12 genes encoding proteins related to plasmid functions, which are replication, partition or transfer, and genes were found in the vicinity of integrase genes. However, exact plasmid responsible for those functions is not identified. 
Interesting Features: The genome was analyzed to have several chromosomal islands, which is unique feature different from other streptococcus, but rather similar to pathogenic Escherichia coli . This unexpected similarity built hypothesis that virulence factor was on the unique chromosomal island for both species and evolve them into pathogens. But this hypothesis was not yet tested. Comparison of chromosomal order of gene of S. agalactiae with that of other pathogenic streptococci, S. pyogenes and S. pneumoniae showed that it was highly conserved between S. agalactiae and S. pyogenes than between S. agalactiae and S. pneumoniae revealing higher relatedness between S. agalactiae and S. pyogenes.  Also, some pathogenicity islands discovered in genes of S. agalactiae was unique to this bacteria and it seems to be diverse between different strains of S. agalactiae. 
Cell structure and metabolism
Cellular Features: S. agalactiae is a Gram-positive batecterium that is not motile, catalase-free, diplococcal. It also does not form spores. (more information under appearance in description section). Within its cell structures, S. agalactiae contains genes expressing various extracellular products, such as capsular polysaccharide and surface proteins. This bacteria seems to use those polysaccharide and surface proteins to adhere to epithelial cells of host and to evade host defense system. 
Environment: Streptococcus agalactiae is host-associated facultative anaerobe, that is capable of using oxygen or not using oxygen depending on surrounding environment to generate ATP, aerobic respiration or fermentation, respectively. The optimal temperature for this species is at 37 Celcius degree (mesophile). 
Glucose Oxidation (Oxidative phosphorylation): S. agalactiae is a chemoorganotroph that uses glucose as energy source. This bacteria is able to synthesize ATP by oxidative phosphorylation. Structural genes for cytochrome bd quinol oxidase and NADH dehydrogenase reveals the presence of enzymes contributing to aerobic growth of this species. However, there was no gene discovered that is involved in heme synthesis suggesting possibility of using external source of heme, but no corresponding transporter was detected. 
Fermentation: S. agalactiae is also able to ferment different carbon sources to multiple by-products, lactate, acetate, ethanol, formate or acetoin. 
Interestingly, this bioenergetic mechanism is more related to that of non-pathogenic species under different genus, Lactococcus lactis than that of other pathogenic streptococci, shown in shown in genes coding for bd oxidase and some ferment pathways containing orthologs in gene sequence of L. lactis. 
Carbon Source: S. agalactiae is a heterotroph that is capable of importing a broad range of carbon sources. Genome sequenced identified 17 sugar-specific phosphoenolpyruvate-dependent phosphotransferase system (PTS) enyzme II complex, revealing specificity for cellobioise, beta-glucoside, trehalose, mannose, lactose, fructose, mannitol, N-acetylgalactosamine, and glucose. Also, four sugar-specific ABC transporter, three glycerol permeases and one glycerol-phosphate permease were found further confirming that S. agalaactiae is getting its carbon sources from various organic compound. Gene of S. agalactiae also contained necessary enzymes for glycolysis and for pentose pathway involving pentose and gluconate utiliization, suggesting broad catabolic ability to live in various environment. 
Amino Acid Biosynthesis: Despite of its ability given above, S. agalactiae is known to require a many amino acids in order to grow, due to the fact that it does not have any TCA cycle present to synthesize amino acids. There are only few pathways to synthesize alanine, serine, glycine, glutamine, aspartate, asparagine, and threonine. Because S. agalactiae is an auxotroph, that requires additional nutrients, this bacteria needs mechanisms to import compounds from exogenous sources. Genes for eight ABC transporter and permease specific for specific amino acids were identified and degradation of peptides by peptidases were also discovered to be used in import of amino acids. Four genes encoding exported peptidase, three ABC transporters specific for oligopeptides, 21 genes for intracellular peptidase revealed further confirmed uses of peptidase in recruiting amino acids. 
Vitamin Biosynthesis: Mechanism for vitamin biosynthesis is not yet found. 
Overall, there are various transporter playing role in metabolism including two phosphate and two iron ABC tranpsorters and several cation transport systems. This diversity seems to ensure that S. agalactiae' survival and multiplication in diverse environment, including various host causing disease. 
Ecology and Pathology
S. agalactiae colonizes in the body of some animals, including cow, sheep, and humans without causing any harm. , The habitat of this microorganism is largely confined to the intestine and vagina in human and the mammary gland of cows and sheep. This microorganism also colonizes in the genital and/or intestinal tract of about 10-30% of pregnant women. However, some can actually cause diseases in their neonates or immunocompromised mammals. S. agalactiae is the common cause of inflammation or fibrosis of mammary glands and adjacent areas in cows and sheep colonizing the surface of the teat and duce sinuses. In human neonates, this species cause invasive bacterial infections in mostly neonates and rarely immunocompromised adults, most notably septicemia, pneumonia, and meningitis colonizing different location including the fauces, the nose, the umbilical cord, the ears, feces.  Infection is spread between cows and/or sheep through the milker's hand, contaminated instrument, and the mouth of calves. Once infected, this mammals are likely to lose their reproductive capacity due to blocked milk channels through inflammation. Infection in human is through genital and/or intestinal tract of pregnant women either during pregnancy or delivery and from other neonates or members of the hospital staff in the maternity hospital.  "The interaction of this bacteria with host protein and and the entry into host cells thereby represent important virulence traits." 
Since S.agalactiae is normally present in the vaginal and intestinal tract of 15-40% of adult women without causing any harm, the reason and mechanism of this microorganism causing diseases in neonates is still not fully known. However, neonates is thought to be infected by being exposed to this microorganism through the birth canal or by spread from the maternal genital tract before birth and during early neonatal period. Infections due to this microorganism can be divided into two types depending on when infections take a place in neonates. Most common early-onset infections take places before the end of the first week and less common late-onset between 1 week and 3 months after birth.  Experimental study of early-onset infection suggests that this bacteria to invade fetal epithelial and endothelial cells and certain macrophages and the mortality rates for early-onset is 4-6%. This invasion is confirmed to play important role in its pathogenesis through the ability of S. agalactiae in the monkey model. Ability to invade and transcytosse, this bacteria can enter into the respiratory tract causing pneumonia or further into the blood causing septicemia. Bloodstream enables this bacteria to reach different sites of body causing meningitis and osteomyelitis. The little pathogenesis of late-onset infection is known. But it is suggested to be vertically transmitted from mother or horizontally transferred from nursery personnel. The mortality rate of this infections is relatively lower than that of early-onset infection, 2-6%. This infections is likely to cause meningtitis and bacteremia. ,
Several factors are currently identified to increase the risk of infection, including rupture of cell membrane before rupture before labor and increased interval between rupture and delivery and it can also be affected by small amount of antibiotics against the virulence factor of S. agalactiae . Anticapsular vaccine, erythromycin, ampicillin, penicillin, cephalosporin, and vancomycin can be treated to prevent this infection. The pathogenesis for infections in adults are not yet known. , S. agalactiae expresses several extracellular products including capsular polysaccharide, surface proteins and secreted protein that are studied in some animal models (mouse,rat) to be the virulence factor (detail in pathogenic contents under genome section). When present, plasmid pIP501 in some strains of S. agalactiae confers antibiotic resistance also playing important role in pathogenesis.,
The symptoms of different disease caused by this bacteria is following. Symptoms of early onset neonatal septicemia includes lethargy, fever, jaundice, hypotension, hypothermia, tachypnea, bacteremia, and low Apgar scores. Symptom of early onset neonatal pneumonia includes low Apgar scores, lethargy fever, apnea, tachypnea, cyanosis, cough, pulmonary infiltrates, and rales. Symptoms of late-onset neonatal meningitis includes lethargy fever jaundice hypotension hypothermia stiff neck nuchal rigidity seizures. Symptoms of osteomyelitis includes fever, bone pain, chills, erythema, swelling, and inflammation 
Although Streptococcal diseases are very serious once developed, luckily only small percentage of neonates develop those even if their mother carry the bacteria (0.25 per 1000 lives with absence of clinical risk factors and 1-4 per 1000 birth with the presence of clinical risk factors).  This microorganism is considered one of the major causes of economic losses to dairy producers without a control program and poses great threats to neonates and some population of adult. , ,
Application to Biotechnology
S. agalactiae produces some antibiotics in mouse or goats, mouse or goat Anti-Streptococcus agalactiae Monoclonal Antibody and can be further used to study mechanism of S. agalactiae and to develop new antibiotics.  Several components of S. agalactiae can be used to make vaccines against itself. Ideal vaccine is likely to include one surface protein that elicits host's immune system and present in all clinical isolates (strains). S.agalactiae are studied to contain several surface proteins having properties as ideal vaccine component for not only this species, but also for other gram-positive pathogens. , 
Surface proteins of members of Alp family, which elicit immune system in a mouse, in this microorganism is studied as a potential vaccine component. Vaccine containing two surface proteins (Rib and a) of this family would possibly protect against the majority of S. agalactiae infection without adjuvant, which is extra agents modifying and catalyzing the effect of vaccine. Another surface proteins is Sip protein, which also is ideal vaccine component, because it is highly conserved and elicit immune system against antigens from different types of strains of this microorganism. Additional surface proteins that are not yet fully studied, including C5a peptidase(ScpB) and beta protein. Especially ScpB has high potential of being ideal vaccine component, since it is present in all strains and highly conserved. However, the immunization with ScpB-based vaccine has not yet been tested.  Also to be more effects, S. agalactiae vaccine should be made of combined virulence factors, antigens, to elicit immunological response in a recipient. ,
The polysaccharide capsule of S. agalactiae, which is another virulence factor, is currently studied to develop a multivalent vaccine conjugated to S. agalactiae protein as carrier. The development of multivalent vaccines are important and necessary because of possibly of antigenic variation and protein-based vaccine is not sufficient alone. Conjugated vaccine of type III polysaccharide and beta surface protein or other surface protein do elicit immune system for strains expressing either of surface protein or polysaccharide. However, protection against infection has not yet been tested. 
After S. agalactiae is discovered to be pathogenic not only to ungulates including cows and sheep, but also to neonates and immunocompromised adults, research has focused on virulence factors that are contributing to its pathogenesis and corresponding mechanisms. In recently research at Institute of Biotechnology, University of Helsinki, structure of the S. agalactiae family II inorganic pyrophosphatase was crystallized and solved by molecular replacement refining at 2.8 A. This pyrophosphatase is one of targets in a serine/threonine protein kinase (STK) signaling cascade which phosphorylates serine/threonine residues and is studied to contribute to virulence of S.agactiae. The structure of pyrophosphatase is revealed to consist of two domains (residues 1-191 and 198-311) with active site between two domains and suggests the likely target of both kinase and phosphate to be Ser150, Ser194, Ser195, and Ser296. Those targets were found to be surface-accessible and present in either the active site or the hinge region between two domain. The target of bacterial Ser/Thr phosphorylation can be used to study to block the transduction of signals within S. agalctiae and to control its virulence. Drugs targeting signaling enzymes and its targets can be designed to cure severe disease, such as septs and meningitis caused by these pathogens. Further study is currently in process to identify exact target to control its pathogenesis. 
Another recent research focused on S. agalactiae CAMP factor/protein B to discover that it does not bind to human IgG. CAMP factor, an extracellular cytolytic protein, was called protein B, because it has been believed to bind the Fc fragment of IgG and this interaction is suggested to inhibit the hemolytic activity of CAMP factor/protein B. However, the CAMP factors are examined to react with only specific IgG through Fab domain, through the study with sera of infected adults and neonates and of non-infected. None of sera of infected adults reacted with CAMP factor and only 2 of sera of non-infected, indicating that CAMP factor is not binding to Fc domain of IgG and that this interaction is not involved in inhibiting hemolytic activity of CAMP factor. Since human IgG is not binding CAMP factor, protein involved in hemolytic activity, further study is necessary to find antibody that will inhibit its activity. 
Researchers also have studied for spreadable mobile genetic mobile element that largely contributes to increased resistance of S. agalactiae to various antibiotics. Study recently done at Universite ́ de Caen Basse-Normandie in France characterized a small mobilizable transposon, MTnSag1, identified in clinical strain of S. agalctiae. This transposon, containing two open fram, ORF1 and ORF2, encoding an Is1-like transposage gene and a lincosamide o-nucleotidyltransferase coferring resisntace to antibiotic, lincomycin, respectively. MTnSag1 can be conjugated to other transposon Tn916 to spread into other recipient. Most S. agactiae harbors Tn916-like element, so MTnSag1 is likely to be acquired and spread antibiotic resistance efficiently across S. agalactiae. This suggests that lincomycin can be less likely to be effective in treating infection caused by MTnSag1-carrying S.agalctiae, thus necessitate further antibiotic studies. 
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[www.cdc.gov/ncidod/eid/vol10no8/pdfs/03-0917.pdf John F. Bohnsack,* April A. Whiting,* Gabriela Martinez,† Nicola Jones,‡ Elisabeth E. Adderson,§ Shauna Detrick,* Anne J. Blaschke-Bonkowsky,* Naiel Bisharat,‡ and Marcelo Gottschalk† " Serotype III Streptococcus agalactiaefrom Bovine Milk and Human Neonatal Infections".Emerging Infectious Diseases • Vol. 10, No. 8, August 2004]
Edited by Ha Bean Kim,student of Rachel Larsen and Kit Pogliano