WO2011099875A1 - Use of lactic acid bacteria to treat or prevent rhinitis - Google Patents

Abstract

Use of Lactobacillus rhamnosus HN001 or derivatives thereof to treat or prevent rhinitis.

Description

USE OF LACTIC ACID BACTERIA TO TREAT OR PREVENT RHINITIS TECHNICAL FIELD

[0001] This invention relates to the use of probiotic bacteria and in particular the use of a strain of lactic acid bacteria to treat or prevent rhinitis. Methods for using the bacteria and compositions comprising the bacteria are also provided.

BACKGROUND

[0002] In 1989, Strachan (Strachan D. Family size, infection and atopy: the first decade of the "hygiene hypothesis". Thorax 2000; 55(suppl 1):S2-10) suggested that decreased exposure to infections could explain the increasing prevalence of allergic disease in Western countries. This has become known as the hygiene hypothesis.

[0003] Since then, numerous investigations have attempted to discern a role for organisms such as lactobacilli in immunological maturation (V arala G. Inxmunological effects of probiotics with special reference to lactobacilli. Clin Exp Allergy 2003;33:1634-40 Blumer N, Sel S, Virna S, Patrascan C, Zimmermann S, Herz U, et aL Perinatal maternal application of Lactobadllus rhamnosus GG suppresses allergic airway inflammation in mouse offspring. Clin Exp Allergy 2007;37:348-57) and the effect of probiotics on the development of allergic disease.

[0004] The efficacy of prenatal or neonatal administration of Lactobadllus rhamnosus GG, Lactobadllus acidophilus LAYRI-Al, or Lactobacillus reuteri ATCC 55730 on the development of allergic disease is conflicting, with various studies reporting divergent findings. One study reported that administration of Lactobacillus rhamnosus GG halved the frequency of eczema at 2, 4, and 7 years, but had no effect on atopic sensitization (see KaUiomaki M, Salminen S, Poussa T, Isolauri E. Probiotics during the first 7 years of life: a cumulative risk reduction of eczema in a randomized, placebo- controUed trial. J Allergy Clin Immunol 2007;119:1019-21). Other studies have found no effect of Lactobacillus acidophilus or L rhamnosus GG on atopic dermatitis, with one of these studies finding that L acidophilus supplementation actually increased the risk of atopic sensitization (Taylor A, Dunstan J, Prescott S. Probiotic supplementation for the first 6 months of life fails to reduce the risk of atopic dermatitis and increases the risk of aUergen sensitization in high-risk children: a randomized controUed trial. J AUergy Clin Immunol 2007;119:184-91). It has been suggested that the different organisms used and whether there was a prenatal intervention may have influenced the divergent findings. [0005] Furthermore, while a range of treatments for some allergic diseases are currently available, those suitable for use in the treatment or prevention of rhinitis, including rhinitis during pregnancy or in young children, are limited and frequently of limited efficacy.

[0006] There remains a need for methods and compositions useful to treat or prevent rhinitis, and particularly such methods and compositions utilizing or comprising other lactobacilli.

[0007] It is an object of this invention to go some way towards achieving one or ore of these desiderata or at least to offer the public a useful choice.

SUMMARY OF THE INVENTION

[0008] I a first aspect the invention provides a method of treating or preventing rhinitis in a subject, the method comprising administration of actobaallu rhamnosus HNOOl, AGAL deposit number NM97/09514 dated 18 August 1997 to a subject in need thereof.

[0009] In one embodiment, the L» rhamnosus HN00I is administered in the form of a composition with a physiologically acceptable diluent, adjuvant, carrier or excipient.

[0010] In one embodiment, said physiologically acceptable diluent, adjuvant, carrier or excipient is a food. In one embodiment, the food is cultured milk, yoghurt, cheese, mi k drink or milk powder.

[0011] Alternatively the composition is a pharmaceutical composition and said excipient or diluent is pharmaceutically acceptable diluent, adjuvant, carrier or excipient

[0012] In embodiments where the subject is a foetal subject, the method comprises administering die rhamnosus HNOOl or a composition comprising L rhamnosus HNOOl to the foetal subject's mother. It will be appreciated that in such embodiments, the administration to the subject may be considered indirect administration. In one embodiment, the composition is a maternal formula or a maternal supplement. In such embodiments, the method_. referably relates to prevention of rhinitis.

[0013] In certain embodiments where the subject is a neonatal, an infant, or a child subject, the method comprises administering a composition comprising L rhamnosus HNOOl to the subject. Again, it will be appreciated that in such embodiments, the administration to the subject may be considered direct administration. [0014] In other embodiments, such as where the subject is a breastfeeding neonatal, infant, or child subject, the method comprises administering the L· rhamnosus HNOOl or a composition comprising L· rhamnosus HNOOl to the subject's mother. It will be appreciated that in such embodiments, the administration to the subject may be considered indirect administration.

[0015] The composition may be an infant formula, follow-on formula, growing-up formula or dietetic product, including hypoallergenic embodiments of such compositions.

[0016] In preferred embodiments where the subject is a juvenile or an adult Subject, the method comprises administering a composition comprising L· rhamnosus HNOOl to the subject. Preferably, the composition is a supplement, formula, dietetic product or food.

[0017] In certain embodiments, the L rhamnosus HNOOl is in a reproductively viable form, preferably in a reproductively viable form and amount. In other embodiments, the L. rhamnosus HNOOl is killed, lysed, fractionated or attenuated.

[0018] As used herein rhinitis includes rhinoconjuctiVitis and hay fever.

[0019] The invention further provides L· rhamnosus HNOOl for treating or preventing rhinitis and L· rhamnosus HNOOl in the manufacture of a composition for treating or preventing rhinitis. The composition may be a composition such as those as described below including, for example, a food or medicament.

[0020] It will be appreciated that the invention also contemplates the use of JL rhamnosus HNOOl in the manufacture of a composition of the invention, for example a composition for treating or preventing rhinitis in a subject.

[0021] In one embodiment the composition is suitable for oral administration. In other embodiments, the composition is suitable for parenteral administration. In embodiments relating to preventing rhinitis in a foetal subject, the composition is suitable for oral admimstration to a pregnant mother during gestation.

[0022] This invention may also be said broadly to consist i the parts, elements and features referred to or indicated in the specification of the application, individually or collectively, and any or all combinations of any two or more said parts, elements or features, and where specific integers are mentioned herein which have known equivalents in the art to which this invention relates, such known equivalents are deemed to be incorporated herein as if individually set forth. [0023] The term "comprising" as used in this specification means "consisting at least in part of. When interpreting each statement in this specification that includes the term "comprising", features other than that or those prefaced by the term may also be present. Related terms such as "comprise" and "comprises" are to be interpreted in the same manner.

[0024] In this specification where reference has been made to patent specifications, other external documents, or other sources of information, this is generally for the purpose of providing a context for discussing the features of the invention. . Unless specifically stated otherwise, reference to such external documents is not to be construed as an admission that such documents, or such sources of information, in any jurisdiction, are prior art, or form part of the commo general knowledge in the art.

BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 is a diagram showing the flow of participants in the placebo, L. rhamnosus HNOOl, and B. animalis subsp HN019 groups in the trial described in Example 1 herein.

[0026] Figure 2 is two graphs showing (A) the percentage of subjects in which B. animalis subsp lacth was detected at each time point (in months) for each infant group (administered B. animalis subsp hctis HN019, L· rhamnosus HNOOl, or placebo); and (B) the percentage of subjects in which L. rhamnosus was detected at each time point (in months) for each infant group (administered B. animalis subsp Jatfis ΗΝ0Ϊ 9, L. rhamnosus HNOOl, or placebo).

DETAILED DESCRIPTION OF THE INVENTION

[0027] The present invention recognises for the first time the beneficial effects of administration of the lactic acid bacteria L· rhamnosus HNOOl on the incidence and severity of rhinitis.

[0028] Accordingly, in a first aspect the invention provides a method of treating or preventing rhinitis in a subject, the method comprising administration of Lactobacillus rhamnosus HNOOl, AGAL deposit number NM97/09514 dated 18 August 1997 or a derivative thereof to a subject in need thereof.

[0029] While various routes and methods o administration are contemplated, oral adiriinistration of L. rhamnosus HNOOl, such as in a composition suitable for oral administration, is currently preferred. It will of course be appreciated that other routes and methods of adiriinistration may be utilised or preferred in certain circumstances. For example, a parenteral route may be utilised with a composition comprising killed or attenuated rhamnosm HN001 or a derivative thereof

[0030] The term "oral administration" includes oral, buccal, enteral and intra-gastric administration.

[0031] The term "parenteral adrriinistration" includes but is not limited to topical (including administration to any dermal, epidermal or mucosal surface), subcutaneous, intravenous, intraperitoneal, and intramuscular administration.

[0032] A "subject" is an animal, preferabl a mammal, more preferably a mammalian companion animal or human. Preferred companion animals include cats, dogs and horses. In one embodiment the human is an adult, a child, an infant, a neonate, or a foetus. In various embodiments, the human child, infant or neonate is a breastfeeding child, infant or neonate.

[0033] The term "treat" and its derivatives should be interpreted in their broadest possible context The term should not be taken to impl that a subject is treated until total recovery. Accordingly, "treat" broadly includes amelioration and/or prevention of the onset of the symptoms or severity of a particular condition.

[0034] It will be appreciated that treatment includes prophylactic treatment, such as for example, the prophylactic treatment of a foetal subject by indirect adrriinistration of a composition of the invention by administering the composition to the foetal subject's mother, or prophylactic treatment of an individual at the beginning or during the periods of increased prevalence of hay fever (the so-called "hay fever season").

[0035] In another example, the prophylactic treatment is of a breastfeeding neonatal, infant or child subject by indirect administration of a composition of the invention by administering the composition to the neonatal, infant, or child subject's mother.

[0036] It will be further appreciated that treatment includes therapeutic treatment, such as for example, treatment of rhinitis or one or more symptoms of rhinitis, including for example the treatment of an neonatal, infant or child subject by indirect administration of a composition of the invention by administering the composition to the subject's mother.

[0037] Accordingly, the invention provides a method of preventing rhinitis in a foetal subject, the method comprising administration of L· rbamnosus HN001 or a composition comprising L rhammsus HNOOl to the subject's mother. Particularly contemplated is a method of preventing rhinitis in a foetal subject.

[0038] The invention further provides a method of treating o preventing rhinitis in a breastfeeding neonatal, infant, or child subject, the method comprises administering L. rbamnosus HNOOl or a composition comprising L· rbamnosus HNOOl to the subject's mother. Particularl contemplated is a method of preventing rhinitis in a neonatal, infant or child subject.

[0039] Also provided is a method of treating or preventing rhinitis in a neonatal, infant, or child subject, the method comprises administering L rbamnosus HNOOl or a composition comprising L· rbamnosus HNOOl to the subject. Particularly contemplated is a method of preventing rhinitis in a neonatal, infant or child subject

[0040] A method of treating rhinitis in an infant or child subject comprising administering a composition consisting of or consisting essentially of .L, rbamnosus HNOOl is also contemplated.

[0041] In certain embodiments, the infant or child is one or more years of age.

[0042] In certain embodiments, the infant or child is a food-sensitised infant or child.

[0043] In certain embodiments, the infant or child is considered to be at risk of rhinitis due to the presence of allergy in one or both of its biological parents.

1 Lactobacillus rhamnosus HNOOl

[0044] As described in the applicant's PCT International application PCT/NZ98/00122 (published as WO 99/ 10476 and incorporated herein in its entirety), a freeze-dried culture of hactoba illus rbamnosus HNOOl was deposited at the Australian Government Analytical Laboratories (AGAL), The New South Wales Regional Laboratory, 1 Suakin Street, Pymble, NSW 2073, Australia, on 18 August 1997 and was accorded deposit number NM97/09514. This Budapest Treaty-recognised depository is now no longer referred to as AGAL, but rather is referred to as the National Measurement Institute of Australia (NMIA). The genome sequence of L. rbamnosus HNOOl is available at Genebank under accession number: NZ_ABWJ0O0O0O0O.

1.1 Morphological properties

[0045] The morphological properties of L· rhamnosus HNOOl are described below. [0046] Short to medium rods with square ends in chains, generally 0.7 x 1.1 x 2.0 - 4.0 urn, when grown in MRS broth.

[0047] Gram positive, non-mobile, non-spore forming, catalase negative facultative anaerobic rods with optimum growth temperature of 37±1 °C and optimum pH of 6.0 - 6.5. These are facultatively heterofermentative bacteria and no gas is produced from glucose.

1.2 Fermentative properties

[0048] ^" An API 50 CH sugar fermentation kit was used to detemiine the carbohydrate fermentation pattern of L. rhamnosus HN001, yielding a score of 5757177 (based on scores of 22 prominent sugars - see PCT/NZ98/00122).

1.3 Further characterisation

[0049] L. rhamnosus strain HN001 may be further characterised by the functional attributes disclosed in PCT/NZ98/00122, including its ability to adhere to human intestinal epithelial cells, and by the improvements in phagocyte function, in antibody responses, in natural killer cell activity, and in lymphocyte proliferation elicited by dietary intake or in in vitro model systems. It will be appreciated that there are a wide variety of methods known and available to the skilled artisan that can be used to confirm the identity of L rhamnosus HN001, wherein exemplary methods include DNA fmgerprinting, genomic analysis, sequencing, and related genomic and proteomic techniques.

[0050] As described herein, certain embodiments of the present invention utilise live L. rhamnosus HN001 - In other embodiments, a L· rhamnosus HN001 derivative is utilised.

[0051] As used herein, the term "derivative" and grammatical equivalents thereof when used with reference to bacteria (including use with reference to a specific strain of bacteria such as L. rhamnosus HN001) contemplates mutants and homologues of or derived from the bacteria, killed or attenuated bacteria such as but not limited to heat-killed, lysed, fractionated, pressure-killed, irradiated, and UV- or light-treated bacteria, and material derived from the bacteria including but not limited to bacterial cell wall compositions, bacterial cell lysates, lyophilised bacteria, probiotic factors from the bacteria, and the like, wherein the derivative retains probiotic activity. Methods to produce such derivatives, such as but not limited to one or more mutants of L· rhamnosus HN001 or one or more probiotic factors, and particularly derivatives suitable for administration to a subject (for example, in a composition) are well-known in the art. [0052] It will be appreciated that methods suitable for identifying L. rhamnosus HNOOl, such as those described above, are similarly suitable for identifying derivatives of L. rhamnosus HNOOl, including for example mutants or homologues of L· rhamnosus HNOOl, or for example probiotic factors from L· rhamnosus HNOOl.

[0053] The term "probiotic factor" refers to a bacterial molecule responsible for mediating probiotic activity, including but not limited to bacterial DNA motifs, surface proteins, small organic acids, polysaccharides, or cell wall components such as lipoteichoic acids and peptidoglycan, or a mixture of any two or more thereof. While, as noted above, these molecules have not been clearl identified, and without wishing to be bound by any theory, such molecules will be present if a probiotic organism is present.

[0054] The term "probiotic activity" refers to the ability of certain microorganisms to stimulate the immune system. Measuring the type and level of activity of a probiotic microorganism is known to those skilled in. the art; see, for example, Mercenier , (2004), Leyer et al. (2004), or Cummings et al. (2004). For example, probiotic activity may be assessed by a PBMC cytokine secretion assay.

[0055] Reference to retaining probiotic activity is intended to mean that a derivative of a probiotic microorganism, such as a mutant or homologue of a probiotic microorganism or an attenuated or lolled probiotic microorganism still has useful probiotic activity, or that a composition comprising a probiotic microorganism or a derivative thereof is capable of supporting the maintenance of useful probiotic activity. While the bacterial molecules responsible for mediating probiotic activity have not been clearly identified, molecules that have been proposed as possible candidates include bacterial DNA motifs, surface proteins, small organic acids, polysaccharides, and cell wall components such as lipoteichoic acids and peptidoglycan. It has been postulated that these interact with components of the host immune system to give an imrnuno-modulatory effect. Preferably, the retained activity is at least about 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 99 or 100% of the activity of an untreated (i.e., live or non-attenuated) control, and useful ranges may be selected between an of these values (for example, from about 35 to about 100%, from about 50 to about 100%, from about 60 to about 100%, from about 70 to about 100%, from about 80 to about 100%, and from about 90 to about 100%).

[0056] Using conventional solid substrate and liquid fermentation technologies well known in the art, L rhamnosus HNOOl can be grown in sufficient amounts to allow use as contemplated herein. For example, L· rhamnosus HNOOl can be produced in bulk for formulation using nutrient film or submerged culture growing techniques, for example under conditions as described in WO99/10476. Briefly, growth is effected under aerobic conditions at any temperature satisfactory for growth of the organism. For example, for l_^. rbamnosus HN001 a temperature range of from 30 to 40°C, preferably 37°C, is preferred. The pH of the growth medium is slightly acidic, preferably about 6.0 to 6.5. Incubation time is sufficient for the isolate to reach a stationary growth phase.

[0057] L rbamnosus HN001 cells may be harvested by methods well known in the art, for example, by conventional filtering or sedimentary methodologies (eg. centrifugation) or harvested dry using a cyclone system. L~ rbamnosus HN001 cells can be used immediately or stored, preferably freeze-dried or chilled at -20° to 6°C, preferably -4°C, for as long as required using standard techniques.

2 Compositions

[0058] A composition useful herein may be formulated as a food, drink, food additive, drink additive, dietary supplement, nutritional product, medical food, enteral or parenteral feeding product, meal replacement, cosmeceuticaL nutraceutical, or pharmaceutical. Appropriate formulations may be prepared by an art skilled worker with regard to that skill and the teaching of this specification.

[0059] In one embodiment, compositions useful herein include any edible consumer product which is able to carry bacteria or a bacterial derivative. Examples of suitable edible consumer products include powders, liquids, confectionary products including chocolate, gels, ice creams, reconstituted fruit products, snack bars, food bars, muesli bars, spreads, sauces, dips, dairy products including yoghurts and cheeses, drinks including dairy and non-dairy based drinks (such as milk drinks and yogurt drinks), milk powders, sports supplements including dairy and non-dairy based sports supplements, food additives such as protein sprinkles, dietary supplement products including daily supplement tablets, weaning foods and yoghurts, and formulas such as infant formula, follow- on formula, or growing-up formula, in powder or liquid form, including hypoallergenic embodiments of such compositions. Within this embodiment, a preferred composition useful herein may be an infant formula, follow-on formula or growing-up formula, i powder or liquid form. Suitable nutraceutical compositions useful herein may be provided in similar forms.

[0060] Examples of formulas such as infant formula, follow-on formula, or growing-up formula, in powder or liquid form, include the following. It should be understood that the following formulations are indicative only and variations may be made according to known principles for formulating such products. For example, non-dairy sources of protein may be supplemented for the dairy proteins listed. Equally, hypoallergenic embodiments of these products may be provided where the protein source is fully or partially hydrolysed. Such hydrolysates ate known in the art. One example of an infant formula, follow-on formula or growing-up formula useful herein comprises (w/w)

30 - 60 % lactose

15 - 35% vegetable oils

0 - 40% skim milk powder

0 - 40% whey protein, such as a WPC or WPI, preferably an 80% WPC (WPC80) 0.001 - 50% of L rhamwsus HNOQt.

[0061] Another example of an infant formula, follow-on formula or growing-up formula useful herein comprises (w/ w)

40 · 60 % lactose

20 - 30% vegetable oils ,

10 - 15% skim milk powder

6 - 8% whey protein, preferably WPC80

0.001 - 10% of L· rhamnosm HNQm.

[0062] Anotlier example of an infant formula, follow-on formul or growing-up formula useful herein comprises (w/w)

40 - 60 % lactose

20 - 30% vegetable oils

10 - 15% skim milk powder

6 - 8% whey protein, preferably WPC80

0.001 - 5% of.L. rb mnosus HN001.

[0063] Another example of an infant formula, follow-on formula or growing-up formula useful herein comprises (w/w)

40 - 60 % lactose

20 - 30% vegetable oils

10 - 15% skim milk powder

6 - 8% whey protein, preferably WPC80

0.001 - 2% of L» rhawnosus HNQOl .

[0064] Any of these infant formulas may also comprise 0.1 to 4% w/w, preferably 2 to 4% w/w of one or more of a vitamin premix, a mineral premix, lecithin, one or more antioxidants, one or more stabilisers, or one or more nucleotides, or a combination of any two or more thereof. In some embodiments, these infant formulas may be formulated to provide between 2700 and 3000 kJ/L.

[0065] Examples of edible consumer products of the invention, such as dairy based drinks (such as milk drinks and yogurt drinks) will typically comprise and may consist of a protein source (such as a dairy protein source), a lipid source, a carbohydrate source, in addition to the L· rkamnosus HNOOl or derivative thereof. Flavourants, colourants, and other additives, carriers or excipients as are well known to those skilled in the art may also be included.

[0066] A further example of an edible consumer product amenable to use in the present invention is the Unistraw™ delivery system (Unistraw International Limited, Australia) as described in PCT international application PCT/AU2007/000265 (published as WO 2007/098564) and PCT international application PCT/AU2007/001698 (published as WO 2008/055296), each incorporated herein in its entirety. It will be appreciated by those skilled in the art that L. rhamnosus HNOOl and derivatives thereof, optionally together with one or more additional probiotic factor or probiotic agent, may be coated onto a substrate (for example, a water soluble bead) for use in such delivery , systems.

[0067] In alternative embodiments, the compositions useful herein may be formulated to allow for administration to a subject by any chosen route, including but not limited to oral or parenteral (including topical, subcutaneous, intramuscular and intravenous) adrniiiistration.

[0068] For example, a nutraceutical composition for use according to the invention can be a dietary supplement (e.g., a capsule, a mini-bag, or a tablet) or a food product (e.g., milk, juice, a soft drink, a herbal tea-bag, or confectionary). The composition can also include other nutrients, such as a protein, a carbohydrate, vitamins, minerals, or amino acids. The composition can be in a form suitable for oral use, such as a tablet, a hard or soft capsule, an aqueous or oil suspension, or a syrup; or in a form suitable for parenteral use, such as an aqueous propylene glycol solution, or a buffered aqueous solution. The amount of the active ingredient in the nutraceutical composition depends to a large extent on a subject's specific need. The amount also varies, as recognized by those skilled in the art, dependent on administration route, and possible co-usage of other probiotic factors or probiotic agents.

[0069] It will be appreciated that in certain embodiments, the compositions of the invention may be formulated so as to have a desired calorific content, for example so as to deliver a desired amount of energy or a desked percentage of daily recommended energy intake. Fo example, an edible consumer product may be formulated to provide from about 200 to about 2000kj per serve, or from about 500kJ to about 2000kf per serve, or from about 1000 to about 2000kJ per serve.

[0070] Thus, a pharmaceutical composition useful according to the invention may be formulated with an appropriate pharmaceutically acceptable carrier (including excipients, diluents, auxiliaries, and combinations thereof) selected with regard to the intended route of administration and standard pharmaceutical practice. For example, a composition useful according to the invention can be administered orally as a powder, liquid, tablet or capsule, or topically as an ointment, cream or lotion. Suitable formulations may contain additional agents as required, including emulsifying, antioxidant, flavouring or colouring agents, and may be adapted for immediate-, delayed-, modified-, sustained-, pulsed- or controlled-release.

[0071] The term "pharmaceutically acceptable carrier" is intended to refer to a carrier including but not limited to an excipient, diluent or auxiliary, pharmaceutically acceptable carrier includes a solvent, a dispersion medium, a coating, an antibacterial and antifungal agent, and an isotonic and absorption delaying agent or combination thereof, that can be administered to a subject as a component of a composition described herein that does not reduce the activity of the composition and is not toxic when administered in doses sufficient to deliver an effective amount of a compound or composition useful herein. The formulations can be administered orally, nasally or parenterally (including topically, intramuscularly, intraperitoneally, subcutaneously and intravenously).

[0072] In certain embodiments, a composition of the invention (such as, for example, a nutraceutical or pharmaceutical composition of the invention, may be provided as a capsule. Capsules can contain any standard pharmaceutically acceptable materials such as gelatin or cellulose. Tablets can be formulated in accordance with conventional procedures by compressing mixtures of the active ingredients with a solid carrier and a lubricant. Examples of solid carriers include starch and sugar bentonire. Active ingredients can also be administered in a form of a hard shell tablet or a capsule containing a binder, e.g., lactose or mannitol, a conventional filler, and a tabletting agent. Pharmaceutical compositions can also be administered via the parenteral route. Examples of parenteral dosage forms include aqueous solutions, isotonic saline or 5% glucose of the active agent, or other well-known pharmaceuticall acceptable excipients. Cyclodextrins, or other solubilising agents well-known to those familiar with the art, can be utilized as excipients for delivery of the therapeutic agent. [0073] In certain embodiments, the composition of the invention comprises live L. rbamnosus HNOOl. Methods to produce such compositions are well-known in the art, and one such method is exemplified herein in the examples.

[0074] In other embodiments, the composition of the invention comprises one or more JL rbamnosus ΗΝ00Ϊ derivative. Again, methods to produce such compositions are well-known in the art, and may utilise standard microbiological and pharmaceutical practices.

[0075] It will be appreciated that a broad range of additives or carriers may be included in such compositions, for example to improve or preserve bacterial viability or to increase therapeutic efficacy of JL. rbamnosus HNOOl or of one or more L· rbamnosus HNOOl derivatives. For example, additives such as surfactants, wetters, humectants, stickers, dispersal agents, stablisers, penetrants, and so-called stressing additives to improve bacterial cell vigor, growth, replication and survivability (such as potassium chloride, glycerol, sodium chloride and glucose), as well as cryoprotectants such as maltodextrin, may be included. Additives may also include compositions which assist in mamtaining microorganism viability in long term storage, for example unrefined com oil, or "invert" emulsions containing a mixture of oils and waxes on the outside, and water, sodium alginate and bacteria on the inside.

[0076] In certain embodiments, the JL rbamnosus HN001 is in a reproductively viable form and amount.

[0077] The composition may comprise a carbohydrate source, such as a disaccharide including, for example, sucrose, fructose, glucose, or dextrose. Preferably the carbohydrate source is one able to be aerobically or anaerobically utilised by JL rbamnosus HNOOl .

[0078] In such embodiments, the composition preferably is capable of supporting reproductive viability of the JL rbamnosus HNOOl for a period greater than about two weeks, preferably greater than about one month, about two months, about three months, about four months, about five months, more preferably greater dian about six months, most preferably at least about 2 years to about 3 years or more.

[0079] In certain embodiments, an oral composition is formulated to allow the administration of a sufficient amount of JL rbamnosus HNOOl to establish a population in the gastrointestinal tract of the subject when ingested. The established population may be a transient or permanent population. [0080] n theory, one colony forming unit (cfu) should be sufficient to establish a population of L· rhamnosus HNOOl in a subject, but in actual situations a minimum number of units are typically required to do so. Therefore, for therapeutic mechanisms that are reliant on a viable, living population of probiotic bacteria, the number of units administered to a subject will usually affect therapeutic efficacy.

[0081] As presented herein in the examples, the AppEcants have determined that a dosage rate of 6 x 10⁹ cfu L· rhamnosus HNOOl per day is sufficient (but may not be necessary) to establish a population in the gastrointestinal tract of human subjects. Accordingly, in one example, a composition formulated for administration will be sufficient to provide at least about 6 x 10⁹ cfu L· rhamnosus HNOOl per day. Higher doses are in certain embodiments desirable.

[0082] Methods to determine the presence of a population of gut flora, such as L. rhamnosus HNOOl, in the gastrointestinal tract of a subject are well known in the art, and examples of such methods are presented herein. In certain embodiments, presence of a population of L. rhamnosus HNOOl can be determined direcdy, for example by analysing one or more samples obtained from a subject, and deterrrrining the presence or amount of L. rhamnosus HNOOl in said sample. In other embodiments, presence of a population of L. rhamnosus HNOOl can be determined indirectiy, for example by observing a reduction in rhinitis symptoms, or a decrease in the number of other gut flora in a sample obtained from a subject. Combinations of such methods are also envisaged.

[0083] The efficacy of a composition useful according to the invention can be evaluated both in vitro and in vivo. See, for example, the examples below. Briefly, the composition can be tested for its ability to prevent or treat rliinitis. For in vivo studies, the composition can be fed to or injected into an animal model (e.g., a mouse) or administered to human subjects (including pregnant women) and its effects on incidence and severity of rhinitis and associated dermalogical conditions are then assessed. Based on the results, an appropriate dosage range and administration route can be determined.

[0084] Methods of calculating appropriate dose may depend on the nature of the active agent in the composition. For example, when the composition comprises live rhamnosus HNOOl, the dose may be calculated with reference to the number of live bacteria present. For example, as described herein the examples the dose may be established by reference to the number of colony forming units (cfu) to be administered per day. In examples where the composition comprises one or more L. rhamnosus HNOOl derivatives, the dose may be calculated by reference to the amount or concentration of L· rhamnosus HNOOl derivative present. For example, for a composition comprising L. rhamnosus HNOOl cell lysate, the dose may be calculated by reference to the concentration of L· rhamnosus HNOOl cell lysate present in the composition.

[0085] By way of general example, the admimstration of from about 1 x 10⁶ cfu to about 1 x 10¹² cfu of L· rhamnosus HNOOl per kg body weight per day, preferably about 1 x 10^fi cfu to about 1 x 10" cfu/kg/day, about 1 x 10⁶ cfu to about 1 x 10"' cfu/kg/day, about 1 x 10⁶ cfu to about 1 x 10⁹ cfu/kg/day, about 1 x 10⁶ cfu to about 1 x 10^s cfu/kg/day, about 1 10* cfu to about 5 x 10⁷ cfu/kg/day, or about about 1 x 10⁶ cfu to about 1 x 10⁷ cfu/kg/day, is contemplated. Preferably, the administration of from about 5 x 10^ cfu to about 5 x 10⁸ cfu per kg body weight of L. rhamnosus HNOOl per day, preferably about 5 x 10⁶ cfu to about 4 x 10⁸ cfu/kg/day, about 5 x 10⁶ cfu to about 3 x 10⁸ cfu/kg/day, about 5 x.10⁶ cfu to about 2 x 10⁸ cfu/kg/day, about 5 x 10⁶ cfu to about 1 x 10^s cfu/kg/day, about 5 x 10⁶ cfu to about 9 x 10⁷ cfu/kg/day, about 5 x 10⁶ cfu to about 8 x 10⁷ cfu/kg/day, about 5 x 10⁶ cfu to about 7 x 10⁷ cfu/kg/day, about 5 x 10^ή cfu to about 6 x 10⁷ cfu/kg/day, about 5 x lO⁶ cfu to about 5 x 10⁷ cfu/kg/day, about 5 x 10⁶ cfu to about 4 x 10⁷ cfu/kg/day, about 5 x 10⁶ cfu to about 3 x 10⁷ cfu/kg/day, about 5 x 10⁶ cfu to about 2 x 10⁷ cfu/kg/ day, or about 5 x 10^s cfu to about 1 x 10⁷ cfu/kg/ day, is contemplated.

[0086] In certain embodiments, periodic dose need not vary with body weight or other characteristics of the subject. In such examples, the administration of from about 1 x 10⁶ cfu to about 1 x 10" cfu of L· rhamnosus HNOOl per day, preferably about 1 x 10⁶ cfu to about 1 x lO¹² cfu/day, about 1 x 10^ή cfu to about 1 x 10¹¹ cfu/day, about 1 x 10⁶ cfu to about 1 x 10'° cfu/day, about 1 0^e cfu to about 1 x 10^y cfu/day, about 1 x 10⁶ cfu to about 1 x 10⁸ cfu/day, about 1 x 10⁶ cfu to about 5 x 10⁷ cfu/day, or about about 1 x 1Ό⁶ cfu to about 1 x 10⁷ cfu/day, is contemplated. Preferably, the adrrunistration of from about 5 x 10⁷ cfu to about 5 x 0ⁱⁿ cfu per kg body weight of L· rhamnosus HNOOl per day, preferably about 5 x 10⁷ cfu to about 4 x 10¹⁰ cfu/day, about 5 x 10⁷ cfu to about 3 x 10^t0 -cfu/day, about 5 x 10⁷ cfu to about 2 x 10ⁱⁿ cfu/day, about 5 x 10⁷ cfu to about 1 x 10⁵⁰ cfu/day, about 5_. x 10⁷ cfu to about 9 x 10⁹ cfu/day, about 5 x 10⁷ cfu to about 8 x 10⁹ cfu/day, about 5 x 10⁷ cfu to about 7 x 10⁹ cfu/day, about 5 x 10⁷ cfu to about 6 x 10⁹ cfu/day, about 5 x 10⁷ cfu to about 5 x 10⁹ cfu/day, about 5 x 10⁷ cfu to about 4 x 10⁹ cfu/day, about 5 x 10⁷ cfu to about 3 x lO⁹ cfu/day, about 5 x 10⁷ cfu to about 2 x 10⁹ cfu/day, or about 5 x 10⁷ cfu to about 1 x 10⁹ cfu/day, is contemplated.

[0087] For example, as presented herei in the examples, an efficacious dose of freeze-dried L· rhamnosus HN001 was determined to be 6 x 10⁹ cfu per day. [0088] It will be appreciated that the composition is preferably formulated so as to allow the administration of an efficacious dose of L· rhamnosus HN001 or one or more derivatives thereof. The dose of the composition administered, the period of administration, and the general administration regime may differ between subjects depending on such variables as the severity of symptoms of a subject, the type of disorder to be treated, the mode of administration chosen, and the age, sex and/or general health of a subject. Furthermore, as described above the appropriate dose may depend on the nature of the active agent in the composition and the manner of formulation. For example, when the composition comprises live L· rhamnosus HN001, the dose may be calculated with reference to the number of live bacteria present. For example, as described herein the examples the dose may be established by reference to the number of colony forming units (cfu) to be administered per day. In examples where the composition comprises one or more L· rhamnosus HN001 derivatives, the dose may be calculated by reference to the amount or concentration of L· rhamnosus HN001 derivative to be administered per day. For example, for a composition comprising L· rhamnosus H 001 cell lysate, the dose may be calculated by reference to the concentration of .L rhamnosus HN001 cell lysate present in the composition.

[0089] It will be appreciated that preferred compositions are formulated to provide an efficacious dose in a convenient form and amount. In certain embodiments, such as but not limited to those where periodic dose need not vary with body weight or other characteristics of the subject, the composition may formulated for unit dosage. It should be appreciated that administration may include a single daily dose or administration of a number of discrete divided doses as may be appropriate. For example, as presented herein in the examples, an efficacious dose of L. rhamnosus HN001 may be formulated into a capsule for oral administration.

[0090] However, by way of general example, the inventors contemplate administration of from about 1 mg to about 1000 mg per kg body weight of a composition useful herein per day, preferably about 50 to about 500 mg per kg per day, alternatively about 150 to about 410 mg/kg/day or about 110 to about 310 mg/kg/day. In one embodiment, the inventors contemplate administration of from about 0.05 mg to about 250 mg per kg body weight of a composition useful herein.

[0091] Examples of infant formula, follow-on formula, or growing-up formula are presented herein. Compositions such as these may be formulated so that the concentration of L. rhamnosus HN001 present in the composition is such that an efficacious dose can be prepared using a readily measm-able amount of the composition. For example, in certain embodiments, such as for example where the composition is an infant formula, the L· rhamnosus HN001 is provided at a concentration sufficient to supply an efficacious dose in an amount of formula capable of being easily measured by a parent or caregiver when preparing the formula for adiTiinistration, such as, for example, with a measured scoop or similar as are commonly provided with infant formulas. Exemplar}' non-limiting concentrations of L. rhamnosus HN001 for use in such compositions include from about 5 x 10⁵ cfu per gram of formula to about 10° cfu per gram of formula, or from about 10⁶ cfu per gram of formula to about 10^s cfu per gram of formula.

[0092] In one embodiment a composition useful herein comprises, consists essentially of, or consists of at least about 0.1, 0.2, 0.5, 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 99, 99.5, 99.8 or 99.9% by weight of L· rhamnosus HN001 or a derivative thereof and useful ranges may be selected between any of these foregoing values (for example, from about 0. to about 50%, from about 0.2 to about 50%, from about 0.5 to about 50%), from about 1 to about 50%, from about 5 to about 50%, from about 10 to about 50%, from about 15 to about 50%, from about 20 to about 50%, from about 25 to about 50%, from about 30 to about 50%, from about 35 to about 50%, from about 40 to about 50%, from about 45 to about 50%, from about 0.1 to about 60%, from about 0.2 to about 60%, from about 0.5 to about 60%, from about 1 to about 60%, from about 5 to about 60%, from about 10 to about 60%, from about 15 to about 60%, from about 20 to about 60% from about 25 to about 60%, from about 30 to about 60%, from about 35 to about 60%, from about 40 to about 60%, from about 4 to about 60%, from about 0.1 to about 70%, from about 0.2 to about 70%, from about 0.5 to about 70%, from about 1 to about 70%, from about 5 to about 70%, from about 10 to about 70%, from about 15 to about 70%, from about 20 to about 70%, from about 25 to about 70%, from about 30 to about 70%, from about 35 to about 70%, from about 40 to about 70%, from about 45 to about 70%, from about 0.1 to about 80%, from about 0.2 to about 80%, from about 0.5 to about 80%, from about 1 to about 80%, from about 5 to about 80%, from about 10 to about 80%, from about 15 to about 80%, from about 20 to about 80%_J from about 25 to about 80%, from about 30 to about 80%, from about 35 to about 80%, from about 40 to about 80%, from about 45 to about 80%, from about 0.1 to about 90%, from about 0.2 to about 90%, from about 0.5 to about 90%, from about 1 to about 90%, from about 5 to about 90%, from about 10 to about 90%, from about 15 to about 90%, from about 20 to about 90%, from about 25 to about 90%, from about 30 to about 90%, from about 35 to about 90%, from about 40 to about 90%, from about 45 to about 90%, from about 0.1 to about 99%, from about 0.2 to about 99%, from about 0.5 to about 99%, from about 1 to about 99%, from about 5 to about 99%, from about 10 to about 99%, from about 15 to about 99%, from about 20 to about 99% from about 25 to about 99%, from about 30 to about 99%, from about 35 to about 99%, from about 40 to about 99%, and from about 45 to about 99%).

[0093] In one embodiment a composition useful herein comprises, consists essentially of, or consists of at least about 0.001, 0.01, 0.05, 0.1, 0.15, 0.2, 0.3, 0.4, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18 or 19 grams of L· rha nosus HN001 or a derivative thereof and useful ranges may be selected between any of these foregoing values (for example, from about 0.01 to about 1 grams, about 0.01 to about 10 grams, about 0.01 to about 19 grams, from about 0.1 to about 1 grams, about 0.1 to about 10 grams, about 0.1 to about 19 grams, from about 1 to about 5 grams, about 1 to about 10 grams, about 1 to about 19 grams, about 5 to about 10 grams, and about 5 to about 19 grams).

[0094] In one embodiment a composition useful herein comprising L. rhamnosus HN001 or a derivative thereof additionally comprises about 0.1, 0.5, 1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 97, 99, or 99.9 % by weight of fresh whole milk or a milk derivative and useful ranges may be selected between any of these foregoing values (for example, from about 0.1 to about 50%, from about 0.2 to about 50%, from about 0.5 to about 50%, from about 1 to about 50%, from about 5 to about 50%, from about 10 to about 50%, from about 15 to about 50%, from about 20 to about 50%, from about 25 to about 50%, from about 30 to about 50%, from about 35 to about 50%, from about 40 to about 50%, and from about 45 to about 50%). The milk derivative is preferably selected from recombined, powdered or fresh skim milk, recombined or reconstituted whole or skim milk powder, skim milk concentrate, skim milk retentate, concentrated milk, ultrafiltered milk retentate, milk protein concentrate (MPC), milk protein isolate (MPI), calcium depleted milk protein concentrate (MPC), low fat milk, low fat milk protein concentrate (MPC), casein, caseinate, milk fat, cream, butter, ghee, anhydrous milk fat (AMF), buttermilk, butter serum, beta serum, hard milk fat fractions, soft milk fat fractions, sphingolipid fractions, milk fat globular membrane fractions, milk fat globular membrane lipid fractions, phospholipid fractions, complex lipid fractions, colostrum, a colostrum fraction, colostrum protein concentrate (CPC), colostrum whey, an immunoglobulin fraction from colostrum, whey (including sweet whey, lactic acid whey, mineral acid whey, or reconstituted whey powder), whey protein isolate (WPI), whey protein concentrate (WPC), a composition derived from any milk or colostrum processing stream, a composition derived from the retentate or permeate obtained by ultrafiltration or microfiltration of any milk or colostrum processing stream, a composition derived from the breakthrough or adsorbed fraction obtained by chromatographic (including but not limited to ion and gel permeation chromatography) separation of any milk or colostrum processing stream, extracts of any of these milk derivatives including extracts prepared by multistage fractionation, differential crystallisation, solvent fractionation, supercritical fractionation, near critical fractionation, distillation, centrifugal fractionation, or fractionation with a modifier (e.g. soaps or emulsifiers), hydrolysates of any of these derivatives, fractions of the hydrolysates, and any combination of any two or more of these derivatives, including combinations of hydrolysed and/or non-hydrolysed fractions. It should be understood that the source of these derivatives may be milk or colostrum or a combination thereof.

[0095] It will be apparent that the concentration of JL rhamnos s HN001 or one or more derivatives thereof in a composition formulated for aclministration may be less than that in a composition formulated for, for example, distribution or storage, and that the concentration of a composition formulated for storage and subsequent formulation into a composition suitable for ackriinistiation must be adequate to allow said composition for administration to also be sufficiendy concentrated so as to be able to be administered at a therapeutically efficacious dose.

[0096] The compositions useful herein may be used alone or in combination with one or more other therapeutic agents. The therapeutic agent may be a food, drink, food additive, drink additive, food component, drink component, dietary supplement, nutritional product, medical food, nutraceutical, medicament or pharmaceutical. The therapeutic agent may be a probiotic agent or a probiotic factor, and is preferably effective to treat, prevent or attenuate rhinitis or one or more of the symptoms of rhinitis.

[0097] When used in combination with another therapeutic agent, the administration of a composition useful herein and the. other therapeutic agent may be simultaneous or sequential. Simultaneous administration includes the administration of a single dosage form that comprises all components or the aclministration of separate dosage forms at substantially the same time. ^Sequential administration includes administration according to different schedules, preferably so that there is an ovedap in the periods during which the composition useful herein and other therapeutic agent are provided.

[0100] Suitable agents with which the compositions useful herein can be separately, simultaneously or sequentially administered include one or more probiotic agents, one or more prebiotic agents, one or more phospholipids, one or more gangliosides, other suitable agents known in the art, and combinations thereof. Useful prebiotics include galactooligosaccharides (GOS), short chain GOS, long chain GOS, fructooUgosaccharides (FOS), short chain FOS, long chain FOS, inulin, galactans, fructans, lactulose, and any mixture of any two or more thereof. Some prebiotics are reviewed by Boehm G and Moro G (Structural and Functional Aspects of Prebiotics Used in Infant Nutrition, J. Nutr. (2008) 138( ):1818S-1828S), incorporated herein by reference. Other useful agents may include dietary fibre such as a fully or partially insoluble or indigestible dietary fibre. Accordingly, in one embodiment L· rhamnosus HN001 or derivative thereof may be administered separately, simultaneously or sequentially with one or more agents selected from one or more probioitics, one or more prebiotics, one or more sources of dietary fibre, one or more galactooligosaccharides., one or more short chain galactooligosaccharides, one or more long chain galactooligosaccharides, one or more fructooligosaccharides, one or more short chain galactooligosaccharides, one or more long chain galactooligosaccharides, inulin, one or more galactans, one or more fructans, lactulose, or any mixture of any two or more thereof.

[0101] In one embodiment, a composition useful herein includes or is administered simultaneously or sequentially with milk components such as whey protein, whey protein fractions (including acidic or basic whey protein fractions or a combination thereof), glycomacf opeptide, lactoferrin, kon-kctoferrin, functional lactoferrin variant, a functional lactoferrin fragment, a vitamin D or calcium,. or combinations thereof. Useful milk component-containing compositions include compositions such as a food, drink, food additive, drink additive, dietary supplement, nutritional product, medical food or nutraceutical. Milk fractions enriched for these components may also be employed. Useful lactoferrins, fragments and compositions are described in international patent applications WO 03/082921 and WO 2007/043900, both incorporated herein by reference in their entirety.

[0102] It should be understood that the additional therapeutic agents listed above (both food based and pharmaceutical agents) may also be employed in a method according to the invention where they are administered separately, simultaneously or sequentially with a compositio useful herein.

[0103] In one embodiment a composition useful herein further comprises a pharmaceutically acceptable carrier. In another embodiment the composition is or is formulated as a food, drink, food additive, drink additive, dietary supplement, nutritional product, medical food, enteral feeding product, parenteral feeding product, meal replacement, cosmeceutical, nutraceutical, medicament, or pharmaceutical. In one embodiment the composition is in the form of a tablet, a caplet, a pill, a hard or soft capsule or a lozenge. In one embodiment the composition is in the form of a cachet, a powder, a dispensable powder, granules, a suspension, an elixir, a liquid, or any other form that can be added to food or drink, including for example water, milk or fruit juice. In one embodiment the composition further comprises one or more constituents (such as antioxidants) which prevent or reduce degradation of the composition during storage or after aclrninistration. These compositions may include any edible consumer product which is able to carry bacteria or bacterial derivatives, including heat-killed, pressure-killed, lysed, UV- or light-treated, irradiated, fractionated or otherwise killed or attenuated bacteria. Examples of suitable edible consumer products include aqueous products, baked goods, confectionary products including chocolate, gels, ice creams, reconstituted fruit products, snack bars, food bars, muesli bars, spreads, sauces, dips, dairy products including yoghurts and cheeses, drinks including dairy and non-dairy based drinks, milk, milk powders, sports supplements including dairy and non-dairy based sports supplements, fruit juice, food additives such as protein sprinkles, dietary supplement products including daily supplement tablets, weaning foods and yoghurts, and formulas such as infant formula, follow-on formula, or growing-up formula, in powder or liquid form. Suitable nutraceutical compositions useful herein may be provided in similar forms.

[0104] It will be appreciated that different compositions of the invention may be formulated with a view to administration to a particular subject group. For example, the formulation of a composition suitable to be administered to a pregnant modier (for example, for indirect adrninistration to a foetal subject or to a breastfeeding neonatal, infant, or child subject) may differ to that of a composition to be directly administered to the subject. It should also be appreciated that the formulation of a composition to be administered prophylacrically may differ to that of a composition formulated for administration once rhinitis or one or more symptoms of rhinitis is present.

[0105] In one embodiment the composition for prophylactic use may further comprise or the JL rhamnosus HN001 may be used in combination with a probiotic agent such as Lactobacillus rhamnosus GG, Lactobacillus acidophilus (for example, L ctobacillus acidophilus (LAVM-A1), Lactobacillus reuteri (for example Lactobacillus reuteri ATCC 55730) or Bifidobacteria lactis (for example, Bifidobacteria lactis strain HN019) or a combination of any two or more thereof.

[0106] In one embodiment, compositions for prophylactic administration, and particularly prophylactic indirect administration, may further comprise or the L rhamnosus HNOOl may be used in combination with a probiotic agent such as Lactobacillus rhamnosus GG, Lactobacillus acidophilus (for example, Lactobacillus acidophilus (LAVRI-A1), l_^actobacillus reuteri (for example l_^actoba illus reuteri ATCC 55730) or Bifidobacteria lactis (for example, Bifidobacteria lactis strain HN019) or a combination of any two or more thereof. {0107] It will be appreciated that the term "prophylactic" and grammatical equivalents as used herein contemplates treatment, use, administration and the like before rhinitis or the symptoms of rhinitis are apparent.

[0108] In embodiments for use in the treatment of a subject having rhinitis or one or more symptoms of rhinitis, the composition may further comprise or the L rhamnosus HNOOl may be combination with a probiotic agent such as Lactobacillus rhamnosus GG, Lactobacillus acidophilus (for example, Lactobacillus acidophilus (LAVRI-Al), Lactobacillus reuteri (for example Lactobacillus reuteri ATCC 55730) or Bifidobacteria lactis (for example, Bifidobacteria lactis strain HN019) or a combination of any two or more thereof, with the proviso that such compositions for direct administration to an infant or child subject of one year or more in age having rhinitis or one or more symptoms of rhinitis do not comprise Bifidobacteria lactis strain HN019.

[0109] As used herein, the term "therapeutic" and grammatical equivalents contemplate treatment, uses or adrrrinistration where rhinitis or the symptoms of rhinitis are present.

[0110] It is intended that reference to a range of numbers disclosed herein (for example, 1 to 10) also incorporates reference to all rational numbers within that range (for example, 1, 1.1, 2, 3, 3.9, 4, 5, 6, 6.5, 7, 8, 9 and 10) and also any range of rational numbers within that range (for example, 2 to 8, 1.5 to 5.5 and 3.1 to 4.7) and, therefore, all sub-ranges of all ranges expressly disclosed herein are hereby expressly disclosed. These are only examples of what is specifically intended and all possible combinations of numerical values between the lowest value and the highest value enumerated are to be considered to be expressly stated in tins application in a similar manner.

3 Rhinitis

[0111] Rhinitis is generally considered to be chronic or acute inflammation of the mucous membrane of the nose associated with increased histamine resulting from exposure to viruses, bacteria or irritants. This mflammation results in the production of excessive amounts of mucus, resulting in the primary symptom of rhinitis, nasal dripping or "runny nose", as well as nasal congestion, rhinoconjunctivitis (inflammation of conjunctiva) and post-nasal drip. Rhinitis has also been associated with sleeping problems, ear conditions, and even learning problems.

[0112] Rhinitis is commonly categorised into three types: infective rhinitis, including acute and chronic bacterial or viral infections; nonallergic (vasomotor) rhinitis, including autonomic, hormonal, drug-induced, atrophic, gustatory rhinitis, and rhinitis medicamentosa; and allergic rhinitis, commonly triggered by inhaled allergens such as pollen, mold, animal dander, and dust. [0113] Infectious rhinitis: Rhinitis is commonly caused by a viral or bacterial infection, including the common cold (caused by Rhinovixuses and Coronaviruses) or bacterial sinusitis. Symptoms of the common cold include rhinorrhea, sore throat (pharyngitis), cough, congestion, and slight headache. ^{■ '} ■ . .

[0114] Non-allergic rhinitis refers to runny nose that is not due to allergy, and can be subdivided into either non-inflammatory or inflammatory nonallergic rhinitis. Vasomotor rhinitis is a common type of non-inflammatory, non-allergic rhinitis caused by non-allergic triggers such as smells, fumes, smoke, dusts, and temperature changes. It is thought that these non-allergic triggers cause dilation of the blood vessels in the lining of the nose, which results in swelling, and drainage. Vasomotor rhinitis can coexist with allergic rhinitis.

[0115] Allergic rhinitis: Exposure of an individual with a sensitized immune system to an allergen, such as pollen or dust, typically triggers production of antibodies which bind to mast cells, in turn leading to the release of histamine. This causes itching, swelling, and mucus production symptomatic of rhinitis. Symptoms vary in severity between individuals, and very sensitive individuals may experience other symptoms associated with allergic reactions. Seasonal rhinitis— commonly called hay fever - occurs particularly during pollen seasons. Seasonal allergic rhinitis does not usually develop until after 6 years of age.

[0116] Diagnosis of allergic rhinitis may be performed by skin testing, the most common method of aEergy testing. This may include intradermal, scratch, patch, or other tests. Less commonly, the suspected allergen is dissolved and dropped onto the lower eyelid as a means of testing for allergies. In some individuals, the RAST blood test may be helpful in determining specific allergen sensitivity.

3.1 Diagnosis:

[0117] · Unsurprisingly given the above, a wide variety of criteria have been proposed for the diagnosis or characterization of rhinitis. No standard approach appears to have been agreed, however the recognition of standardised criteria for the diagnosis of other allergic diseases, such as the SCORAD criteria established for eczema (see, for example, Williams HC, Burney PGJ, Hay RJ, Archer CB, Shipley MJ, Hunter JJA, et al. The UK Working Party's Diagnostic Criteria for Atopic Dermatitis. Br J Dermatol 1994;131:383-96;. Williams H. 'So How Do I Define Atopic Eczema? A Practical Manual for Researchers Wanting to Define Atopic Rhinitis' 1996, http://\vww.nottingham.ac.ulc/dermatology/rhinitis/contents/html; and European Task Force on Atopic Dermatitis. Severity Scoring of Atopic Dermatitis: the SCORAD Index. Dermatology 1993; 186:23-31) may inform the development of a standardised set of criteria for rhinitis. An exemplary set of questions aimed at establishing onset and severity of rhinitis is referred to herein.

3.2 Current treatments:

[0118] As there is no known cure for rhinitis, current treatment regimes are directed to minimising exposure to allergens or infectious agents, and suppressing symptoms. Saltwater sprays, rinses or steam may be effective to remove dust, secretions and allergenic molecules from the mucosa, and so minimize exposure. Current treatments utilise treatments commonly utilized in the treatment of allergic diseases, including antimstamines, cortisone, dexamethasone, hydrocbrtisone, epinephrine (adrenaline), theophylline and cromolyn sodium. Anti-leukotrienes, such as Montelukast (Singulair) or Zafirlukast (Accolate), are FDA approved for treatment of allergic diseases.

[0119] Systemic glucocorticoids such as Triamcinolone or Prednisone and steroid nasal sprays such as beclomethasone (Beconase), budesonide (RJiinocort, Noex), flunisolide (Syntaris), mometasone (Nasonex), fluticasone (Flonase, Flixonase), and triamcinolone (Nasacort AQ) are effective at reducing nasal inflammation, and may be effective without oral antihistamines.

[0120] Pseudoephedrine is indicated for vasomotor rhinitis, and topical decongestants may also be helpful in reducing symptoms such as nasal congestion, but should not be used for long periods as protracted use can lead to a rebound nasal congestion, Rhinitis medicamentosa.

[0121] More severe cases of allergic rhinitis require immunotherapy or surgery to remove tissue in the nose (e.g., nasal polyps) or sinuses.

[0122] Common anti-hismxiines include Actifed (Pseudoephedrine hydrochloride; Triprolidine hydrochloride), Allerid C Syrup (Cetirizine hydrochloride), Anthisan Cream (Mepyramine maleate), Apo-Cetirizine (Cetirizine hydrochloride), Asmafen (Ketotifen), A il Retard (Pheniramine maleate), Avomine (Promethazine theoclate), Benadryl Original (Ammonium chloride; Diphenhydramine hydrochloride; Sodium citrate), Chlorpheniramine Injection (Chlorpheniramine maleate), Claramax (Desloratadine), Claratyne (Loratadine), Clarinase 12 Hour (Loratadine; Pseudoephedrine sulfate), Clarinase 24 Hour (Loratadine; Pseudoephedrine sulfate), Codral 4 Flu (Chlorphenkamine maleate; Codeine phosphate; Paracetamol; Pseudoephedrine hydrochloride), Codral Daytime/Nightime Tablets (Codeine phosphate; Paracetamol; Pseudoephedrine hydrochloride; Triprolidine hydrochloride), Day & Night Cold & Flu Capsules (Chlorphenkamine maleate; Dextromethorphan hydrobromide; Paracetamol; Pseudoephedrine hydrochloride), Demazin Clear Syrup (Chlorpheniramine maleate; Phenylephrine hydrochloride), Demazin Day/Night. Relief Tablets (Dexchloipherikamine maleate; Pseudoephedrine sulfate), Demazin Non Drowsy Repetabs (Loratadine; Pseudoephedrine sulfate), Demazin Syrup (Chloiphenkamine maleate; Phenylephrine hydrochloride), Demazin Tablets (Dexc oipheniramine maleate; Pseudoephedrine sulfate), Dimetapp Gold, Cough & Flu, Day & Night Caps (Dextromethorphan hydrobromide; Doxylamine succinate; Paracetamol; Pseudoephedrine hydrochloride), Dimetapp Colour Free Elixir (Bromphenkarnine maleate; Phenylephrine hydrochloride), Dimetapp DM (Bromphenkarnine maleate; Dextromethorphan hydrobromide; Phenylephrine hydrochloride), Dimetapp Elixir (Bromphenu-amine maleate; Phenylephrine hydrochloride), Dimetapp Infant Drops (Bromphenkarnine maleate; Phenylephrine hydrochloride), Dramamine (Dimenhydrinate), Histafen (Chlorphenkamine maleate), LoraPaed (Loratadine), Lora-Tabs (Loratadine), Marzine (Cyclizine hydrochloride), Naphcon-A (Naphazoline hydrochloride; Phenirarnine maleate), Orthoxicol Day & Night Cold & Flu (Chlorpheniramine maleate; Dextromethorphan hydrobromide; Paracetamol; Pseudoephedrine hydrochloride), Periactin (Cyproheptadine hydrochloride), Phenergan (Promethazine hydrochloride), Phensedyl Dry Family Cough Syru (Pholcodine; Promethazine hydrochloride; Pseudoephedrine hydrochloride), Polaramine (DexcMorphenkamine maleate), Promethazine Hydrochloride Injection BP (Promethazine hydrochloride), Razene (Cetirizine hydrochloride), Robitussin Children's Night Relief Elixir (Chlorpherikamine maleate; Dextromethorphan hydrobromide; Pseudoephedrine hydrochloride), Sea-legs (Meclozine hydrochloride), Serecid (Hydroxyzine hydrochloride), Sinutab Sinus, Allergy and Pain Relief (Chlorpheniramine maleate; Paracetamol; Pseudoephedrine hydrochloride), Sudafed Sinus Pain & Allergy Relief (Paracetamol; Pseudoephedrine hydrochloride; Triprolidine hydrochloride), Telfast (Fexofenadine hydrochloride), Telfast Decongestant (Fexofenadine hydrochloride; Pseudoephedrine hydrochloride), Tixylix Linctus (Pholcodine; Promethazine hydrochloride), Unisom Sleepgels (Diphenhydramine hydrochloride), Vallergan Forte (Trimeprazine tartrate), Valoid (AFT) (Cyclizine lactate), Valoid Injection (Cyclizine lactate), Zadine (Azatadine maleate), and Zyrtec (Cetirizine hydrochloride).

[0123] Any of the current treatment regimes, including diose referred to herein, are in certain embodiments amenable to use in conjunction with the methods and compositions of the present invention.

[0124] Various aspects of the invention will now be illustrated in non-limiting ways by reference to the following examples. - EXAMPLE

[0125] To determine whether probiotic supplementation in early life could prevent development of rhinitis, a double-blind, randomized placebo-controlled trial of infants at risk of allergic disease was conducted.

Materials and Methods

[0126] Pregnant women in Auckland and Wellington, New Zealand, were recruited to the study through maternity care providers, antenatal classes, and advertisements. They were invited to take part in the study if they or the infant's father had a history of treated asthma, eczema, or hay fever. Women were ineligible for the study if they planned to move from the study center in the next 2 years, were already taking probiotic supplements long-term, or intended to use these in the child. They were not able to continue in the study if they delivered before 37 weeks gestation, they had not taken the study capsules for >2 weeks before birth, their infant's weight was <3rd percentile for sex and gestation, or their infant was placed in the neonatal unit for. more than 48 hours or had serious congenital abnormalities at birth. If there were twins, only the heavier was included in the study.

Study design

[0127] The study was a two-centre, double-blind, randomized, placebo-controlled trial of the effects of probiotic supplementation on the development of allergic diseases in infants (Australian New Zealand Clinical Trials Registry: ACTRN12607000518460). There were two treatment groups who received either JL rkamnosus HN001 (6X10^y colony-forming units/day) or B. animalh subsp lactis HN019 (9X1 colony-forming units/day) (Fonterra Co-operative Group, Auckland, New Zealand).

[0128] The probiotic supplements were manufactured by using aseptic fermentation, concentration, and freeze- drying. The growth media contained skim milk powder, yeast extract, and glucose. After growth, cells of the HN001 and HN019 strains were concentrated by centofugation and washed twice with sterile saline. During prototype development of the low-allergenic probiotic supplements, the separate ingredients were tested by skin prick test (SPT) on several patients with cow's milk allergy. This work established that after two washes, the material had no reaction in die patients with cow's milk allergy. The final washed cells were mixed with a cryoprotectant solution, maltodextrin and this mix was frozen on trays and freeze-dried. The resulting powder had a particle size of 200 microns or less and was tested for the presence of pathogens before dispatch to a registered pharmaceutical packaging company. The placebo group received a capsule identical in appearance and smell containing dextran, salt, and a yeast extract (Fonterra Co-operative Group).

The yeast extract used in the probiotics and the placebo contained no viable cells. [0129] All batches of capsules were tested monthly to ensure viability of the probiotics. Shelf life was managed to ensure minimum cell counts were maintained. In addition, capsules returned from the field were tested for their viability. With very few exceptions, the viability was higher than the minimum required.

[0130] At 35 weeks gestation, pregnant women were randomized to receive one of the probiotics or placebo daily, to continue while they were breast-feeding for as long as 6 months postpartum. Infants started the capsules between 2 and 16 days postbirth (median=6 days), continuing until age 2 years. The capsule powder was either given undiluted to the infant or mixed with water, breast milk, or formula and given via a teaspoon or syringe until solid food was started, when it was sprinkled on food.

[0131] Randomization and allocation of supplements were performed by a clinical trials pharmacist at Auckland City Hospital who had no contact with the participants. Randomization was stratified by study center and performed in blocks of 15 according to a computer-generated randomization list. At enrollment, a research study nurse assigned the next study number and provided the participant with the appropriate capsules. All study nurses and participants were blind to treatment assignment for the duration of the study. To evaluate the efficacy of the blinding, the final questionnaire asked participants to indicate whether they believed they were in a probiotic or placebo group.

[0132] Information collected at baseline included parental history of allergic disease; sex; ethnicity; household smoking; pet exposure; and length, weight, and head circumference at birth. Rhinitis prevalence and severity were assessed at follow-up visits at 4.5 years. History of antibiotic use was also collected at these visits.

[0133] Ethical approval was granted by a national multi-region ethics committee, covering both study centers.

Outcome measures

[0134] Rhinitis prevalence at 4 years was determined using questionnaire data in response to a standard set of questions relating to allergy developed by the International Study of Asthma and Allergies in Childhood, available at htrp://isaac.aucldknd,ac.nz/index.htrnl. The relevant questions are reproduced below, and were prefaced with the clarification that questions related to problems which occured when the child did not have a cold or flu.

[0135] Fecal samples were collected from infants soon after birth and at 3, 12, and 24 months of age. The samples were held in the home freezer until transportation to the research center for storage at -80°C. Bacterial DNA was extracted from feces by using a previously described method (Tannock G, Munro K, Harmsen H, Welling G, Smart J, Gopal P. Analysis of the fecal microflora of human subjects consuming a probiotic containing Lactobacillus rhamnosus DR20. Appl Environ Microbiol 2000;66:2578-88). Bifidobacterial DNA was amplified by using PCR primers targeting the transaldolase gene, (Requena T, Burton J, Matsuki T, Munro K, Simon M, Tanaka R, et al. Identification, detection, and enumeration of human Bifidobacterium species by PCR targeting the transaldolase gene. Appl Environ Microbiol 2002;68:2420-7) and Lactobacillus amplicons were obtained by using PCR primers targeting the 16S ribosomal RNA gene (Walter J, Hertel C, Tarrnock G, Lis C, Munro K, Hammes W. Detection of Lactobacillus^ Vediococcus_^ Leuconostic and Weisella species in human feces by using group-specific PCR primers and denaturing gradient gel electrophoresis. Appl Environ Microbiol 2001;67:2578-85). Denatating gradient gel electrophoresis was performed by using a Bio-Rad DCode universal mutation detection system (Bio-Rad, Hercules, Calif). Gradient concentrations and electrophoretic conditions have been described previously (Requena T, Burton J, Matsuki T, Munro K, Simon M, Tanaka R, et al. Identification, detection, and enumeration of human Bifidobacterium species by PCR targeting the transaldolase gene. Appl Environ Microbiol 2002;68:2420-7; Walter J, Hertel C, Tannock G, Lis C, Munro K, Hammes W. Detection of Lactobacillus, Pediococcus, Leuconostic and WeisellaU species in human feces by using group-specific PCR primers and denaturing gradient gel electrophoresis. Appl Environ Microbiol 2001;67:2578-85.). PCR amplicons generated from DNA extracted from pure cultures of B. animalis subsp lactis HN019 and L rbamnosus HNOOl were used as markers in relation to fecal profiles in gels. Visual comparisons of fecal profiles with strain markers thus permitted the detection of B. animalis subsp lactis and L. rbamnosus in the fecal samples. Detection was at the species level because strain-specific PCR primers were not available.

Compliance

[0136] Bottles of capsules were replaced every 3 months and counted by a member of staff who had no participant involvement.

Sample size

[0137] Sample size calculation was based on a 50% cumulative prevalence of rhinitis by 2 years in the control group. To detect an 18% absolute reduction in rhinitis caused by probiotics, with 80% power at the 5% significance level, 127 were needed in each study group. To allo for a 25% loss because of ineligibility at birth or subsequent withdrawal 170 mothers were enrolled in each group. Statistical analysis

[0138] Analysis was undertaken by using SAS version 9.0 (SAS Institute, Gary, NC). Differences between study groups in the cumulative prevalence of rhinitis were summarized by using Kaplan-Meier curves and proportional hazard models. Proportional hazard models were also used to assess differences in study groups in the point prevalence of atopy, and variables dependent on atopy, at 2 years. The persistence of L rhamnosus and B. animalis subsp lactis in fecal samples over the study period was defined as detection of these bacteria on 2 or more occasions versus detection on 1 occasion only or absence of detection to limit the effect of adventitious exposure from food and other environmental sources. Odds ratios were used to assess associations between the persistence of each bacterium in feces and the 4 year prevalence of rhinitis. The presence or absence of each probiotic species in fecal samples was also analyzed b study group at each time point. All children who completed the study were included in an intention-to treat analysis regardless of their compliance. The chi-square test was used to compare differences between groups and differences at baseline, with P<0.05 considered statistically significant. Because baseline differences were small, these variables were not adjusted for in the analysis of the outcome variables.

[0139] Compliance was calculated as the number of capsules taken as a proportion of the number of days in the study period.

Results

[0140] Participants were recruited from January 2004 to May 2005 at an average rate of 7 per week. Among randomized participants who received treatment, 87.7%, 84.7%, and 88.9% in the placebo, L· rhamnosus HNOOl, and JB. animalis subsp lactis HN019 groups, respectively, completed the study (see Figure 1). Among participants who were eligible at birth, 94.3%, 9 .7%, and 96.2% in the placebo, L. rhamnosus HNOOl, and B. animalis subsp lactis HN019 groups, respectively, completed the study (see Figure 1). Of these, there were 6 participants in the placebo group, 17 in the L· rhamnosus HNOOl group, and 12 in the B. animalis subsp lactis HN019 group who discontinued treatment but who continued to be followed up until the end of the study. One mother in the placebo group and 3 mothers in the B: animalis subsp lactis HN01 group gave their reasons for discontinuing treatment as a result of perceived side effects of, or opposition to, taking study capsules. All these participants provided outcome data at each time point and were included in an intention-to-treat analysis. An additional 9, 13, and 6 in the placebo, L. rhamnosus HNOOl , and B. animalu subsp lactis HN01 groups, respectively, withdrew from the study completely and could not be included in an intention- to-treat analysis. None of these withdrawals was a result of perceived side effects of study treatment. [0141] There were no significant differences betwee the groups in the proportion of participants who took more than 75% of the study capsules. Defined this way, the compliance rates were 77.3%, 73.6%, and 78.3%, in the placebo, L· rhamnosus HNOOl, and B. animalis subsp lactis HN01 groups, respectively.

[0142] There were no significant differences between study groups in baseline characteristics see Table I below).

rhnts at 4 years 4.1 compare wth nants n t e pace o group ; a e . ere no statistically significant effect of either probiotic on the likelihood of having a positive skin test result for any allergen, or for food allergens, at 4 years. In contrast, there was no effect of B. animalis subsp lactis HN01 on rhinitis prevalence at 4 years (Table II). Table II Hazard ratios (95% CIs) for the 4 year cumulative prevalence of rhinitis symptoms in infants taking L. rhamnosus HN001 and B. lactis HN019.

Placebo L. rhamnosus P B. lactis HN019 P

N=159 HN001 N=157 value N=158 value

Rhinitis 1.00 ' 0.77 (0.56-1.07) 0.12 0.93 (0.68-1.26) 0.62

symptoms (45.1%) , _. (52.9%)

[0144] Infants receiving L. rh mnosus HN001 also showed a reduced prevalence of rninitis and rhinoconjunctivitis at 4 years (Odds ratio (OR) 0.54 (95% CI 0.31-0.95), and 0.34 (95% CI 0.15- 0.80), respectively. See Table III below.

Table III Odds ratios (95% CIs) of rhinitis, hay fever, and rhinoconjunctivitis at 4 years in infants taking L. rhamnosus HN001 and B. lactis HN019.

Placebo L. rhamnosus P B. JactisHNQl9 P

N=159 HN001 N=157 value N=158 value

Hayfever ever* 1.00 0.52 (0.27-0.98) 0.04 0.54 (0.29-1 M) 0.0519

(21.7%) (12.5%) (13%)

Rhinitis last 12 1.00 0.54 (0.31-0.95) 0.03 0.76 (0.45-1.28) 0.3

months*

(29.4%) (18.4%) (24%)

Rhinoconjuctivitis 1.00 0.34 (0.15-0.8) 0.01 0.77 (0.4-1.51) 0.45

last 12 months

(15.4%) (5.9%) (12.3%)

*n = 425 (based on all with questionnaire data at 4 years)

[0145] B. ani alis subsp lactis and L· rhamnosus were detected in the feces of infants in the placebo and alternative probiotic group at birth, pointing to the adventitious inoculation of the alimentary tract with these bacteria from environmental sources (Ahrne S, Nobeck S, Jeppson B, Adlerberth I, Wold A, Molin G. The normal Lactobacillus flora of healthy human rectal and oral mucosa. J Appl Microbiol 1998; 85:88-94; Janer C, Arigoni F, Lee B, Pelaez C, Requena T. Enzymatic ability of Bifidobacterium animalis subsp. lactis to hydroly2e milk proteins: identification and characterization to endopeptidase O. Appl Environ Microbiol 2005;71: 8460-5; Kim S-Y, Adachi Y. Biological and genetic classification of canine intestinal lactic acid bacteria and bifidobacteria. Microbiol Immunol 2007; 51:919-28; Mah K, Chin V, Wong W, Lay C, Tannock G, Shek L, a al. Effect of milk formula containing probiotics on the fecal microbiota of Asian infants at risk of atopic diseases. Pediatr Res 2007;62:674-9.) (Figure 2, A and B). However, administration of a probiotic resulted in markedly increased detection rates for that probiotic in fecal samples at 3, 12, and 24 months compared with the other groups (P<.0001). Detection of B. animalis subsp hctis in fecal samples increased progressively over the course of the study from 22.6% at 3 months to 53.1% at 24 months among infants administered HN019 (Figure 2, A). In contrast, detection of L· rhamnosus was greatest at 3 months at 71.5% and was slightly lower at 24 months at 62.3% among infants administered this probiotic (Figure 2B).

[0146] At the end of the study, parents were asked whether they thought they were in a probiotic or placebo group. More than half the respondents in each study group could not offer an opinion, 14,7% of the pkcebo group participants thought they had received a placebo, and 23.7% of the B. animalis subsp lactis HN019 group and 25.7% of the L. rhamnosus HNOOl group thought they were in a probiotic group.

Discussion

[0147] In this study, treatment with L· rhamnosus HNOOl for the first 2 years of life was associated with a reduction in the prevalence of rhinitis at 4 years b about a half, and with a reduction in the prevalence of rhinoconjunctivitis at 4 years by about two-thirds.

[0148] Despite some disparities between studies, the weight of evidence suggests a protective role for some Lactobacillus species in the pathogenesis of allergic diseases, but there is little evidence overall that this is mediated through effects on allergic sensitization. The suggestion of Kukkonen et al (Kukkonen K, Savilahti E, Haahtela T, Juntunen-Backman K, Korpela R, Poussa T, et al. Probiotics and prebiotics galacto-oligosaccharides in the prevention of allergic disease: a randomized, double-blind, placebo-conttolled trial. J Allergy Clin Immunol 2007;119:192-8) that probiotics regulate the pathway from sensitization to clinical disease is not supported by the findings presented herein, which show the effect of L. rhamnosus HNOOl is similar for sensitized and non- sensitized rhinitis.

[0149] A number of immunologic pathways have been proposed to be affected by probiotics, involving several different mechanisms. For example, it has been suggested that probiotic influences may be local, and potentially include reduction of permeability and systemic penetration of antigens; alteration of local infkmmation or tolerance induction; anti-infknirnatory effects mediated by Toll-like receptors; activation of tolerogenic dendritic cells; THl skewing of responses; alteration of T-regulatory function; and increased local IgA production. Systemic effects with increased monocytes and effects on T cells, B cells, and stem cells have also been suggested (Prescott S, Bjorksten B. Probiotics for the prevention or treatment of allergic diseases. J Allergy Clin Immunol. 2007;120:255-62). Some strains of lactobacilli and bifidobacteria have been showii to modukte IL-10 production, thereby, it has been suggested, enhancing regulatory or tolerance- inducing mechanisms (Niers L, Timmerman H, Rijkers G, van Bleek G, van Uden N, Knol E, et al Identification of strong interleujkin-10 inducing lactic acid bacteria which down-regulate T helper type 2 cytokines. Clin Exp Allergy 2005;35:1481-9).

[0150] In this study, cord blood IFN-gamma levels were higher and more often detectable among the probiotic groups, but this was statistically significant only for the L· rhamtwsus HN001 group (Prescott SL, Wicken K, Westcott L, Nieblee J, Currie H, Black P, et al. Supplementation with Lactobacillus rhamtwsus or Bifidobacterium lactis probiotics in pregnancy increases cord blood IFN- gamma and breast milk TGF-beta and IgA detection. Clin Exp Allergy 2008^8:771).

[0151] The presence of bifidobacteria and lactobacilli in the feces of participants was investigated because it has rarely been established whether probiotic cultures can pass through the infant gastrointestinal tract and reach the colon when aclministered in trials of long duration (Mah K, Chin V, Wong W, Lay C, Tannock G, Shek L, et al. Effect of milk formula containing probiotics on the fecal microbiota of Asian infants at risk of atopic diseases. Pediatr Res 2007;62:674-9). Different distributions in the detection rate of each probiotic between birth and 24 months were observed.

[0152] Without wishing to be bound by any theory, this might be a result of the changing ecosystem within the infant bowel as the characteristic shifts in bacterial community composition occur over time (Mah K, Chin V, Wong W, Lay C, Tannock G, Shek L, et al Effect of milk formula containing probiotics on the fecal microbiota of Asian infants at risk of atopic diseases. Pediatr Res 2007;62:674-9). B. animaks subsp lactis was detected at relatively low frequency during the first 3 months of adiriinistration. JL- rhamnosus detection in die feces was boosted by HN001 administration, but even so, after subtraction of background exposure to this species, less than half of the infants had detectable DNA from this species in dieir feces. A recent report from Singapore (Mah K, Chin V, Wong W, Lay C, Tannock G, Shek L, et al. Effect of milk formula containing probiotics on the fecal microbiota of Asian infants at risk of atopic diseases. Pediatr Res 2007;62:674-9) showed that peak detection (after subtraction of background exposure) of a Bifidobacterium longum strain in infant feces occurred at 3 days after intervention commenced (44% of infants), falling to 26% and 16% after 1 and 3 months, respectively, of probiotic administration. In that study, detection of L rhamnosus GG was 83% after 3..days of administration, then 77% and 69%, respectively, after 1 and 3 months of administration. Analytical methods of detection of fecal bacteria based on bulk-extracted DNA do not provide information about viability of the bacteria in the bowel with DNA potentially derived from active, quiescent, or dead bacterial cells. Nevertheless, the bacteriologic results of this study are notable because they reveal the relative abilities of the bifidobacteria and lactobacilli to transit the gastrointestinal tract [0153] This study provides evidence that L· rhamnosus HNOOl is an effective intervention for reducing the prevalence of rhinitis, for example among high-risk children. This comparison of 2 different probiotic bacteria demonstrates that not all probiotic bacteria are equally effective in the treatment or prophylaxis of a particular condition or in obtaining any particular treatment outcome.

INDUSTRIAL APPLICABILITY

[0154] This invention relates to the use of probiotic bacteria, particularly Lactobaallus rhamnosus HNOOl or derivatives thereof, and in particular in the treatment or prevention of rhinitis. Methods for using the bacteria and compositions comprising the bacteria are also provided.

Claims

1. A method of treating or preventing rhinitis in a subject, the method comprising administration of Lactobacillus rh nos s HN001 or a derivative thereof to a subject in need thereof.

2. A method of preventing rhinitis in a foetal subject, the method comprising administration of Lactobacillus rhamnosus HN001 or a derivative thereof to the mother of a foetal subject in need thereof.

3. A method of tteating or preventing rhinitis in a breastfeeding neonatal, infant or child subject, the method comprising administration of iMctobaallus rhamnosus HN001 or a derivative thereof to the mother of a neonatal, infant or child subject in need thereof.

4. The method of any one of claims.1 to 3, wherein the Lactobacillus rhamnosus HN001 or derivative thereof is administered in a composition comprising a physiologically acceptable diluent, adjuvant, carrier or excipient.

5. The method of claim 4, wherein the composition comprises an infant formula, a follow- on formula, a growing-up formula, a maternal formula, a maternal supplement, dietetic product, or a food.

6. The method of claim 5, wherein the food comprises cultured milk, yoghurt, cheese, milk drink or milk powder.

7. The method of claim 4, wherein the composition is a pharmaceutical composition and the excipient or diluent comprises a pharmaceutically acceptable diluent, adjuvant, carrier or excipient.

8. The method of any one of claims 1 to 7 wherein the method is a method of preve ting- rhinitis.

9. The method of any one of claims 1 or 3 to 8 wherein the subject is a neonatal, an infant, or a child subject.

10. The method of any one of claims 1 or 4 to 8, wherein where the subject is a juvenile or an adult subject, the method comprises administering a composition comprising Lactobacillus rhamnosus HN001 or a derivative thereof to the subject.

11. The method of any one of claims 1 to 10, wherein the Lactobacillus rhamnosus HN001 or derivative thereof is i a reproductively viable form.

12. The method of any one of claims 1 to 10, wherein the Lactobacillus rhamnosus HNOOl or derivative thereof is killed, fractionated, or attenuated.

13. The method of any one of claims 1 to 12, wherein the rhinitis is rhinoconjimctivitis.

14. The method of any one of claims 1 to 13, wherein the Lactobaallus rhamnosus HNOOl or derivative thereof is administered separately, simultaneously or sequentially with one or more agents selected from one or more probioitics, one or more prebiotics, one or more sources of dietary fibre, one or more galactooligosaccharides, one or more short chain galactooligosaccharides, one or more long chain galactooligosaccharides, one or more fructooligosaccharides, one or more short chain galactooligosaccharides, one or more long chain galactooligosaccharides, inulin, one or more galactans, one or more fructans, lactulose, or any mixture of any two or more thereof.

15. Use oi Lactobacillus rhamnosus ΗΝ00Ϊ or a derivative thereof in the manufacture of a composition for treating or preventing rhinitis. 6. Use of Lactobacillus rhamnosus HNOOl or a derivative thereof in the manufacture of a composition for preventing rhinitis in a foetal subject, wherein the Lactobaallus rhamnosus HNOOl or a derivative thereof is administered to the mother of the foetal subject.

17. Use of Lactobacillus rhamnosus HNOOl or a derivative thereof in the manufacture of a composition for treating or preventing rhinitis in a breastfeeding neonatal, infant or child subject, wherein the Lactobaallus rhamnosus HNOOl or a derivative thereof is administered to the mother of the neonatal, infant or child subject.

18. The use of any one of claims 15 to 17, wherein the Lactobaallus rhamnosus HNOOl or derivative thereof is in a composition comprising a physiologically acceptable diluent, adjuvant, carrier or excipient.

19. The use of any one of claims 15 to 18, wherein the composition comprises an infant formula, a follow-on formula, a growing-up formula, a maternal formula, a maternal supplement, a dietetic product, or a food.

20. The use of any one of claims 15 to 19, wherein the food comprises cultured milk, yoghurt, cheese, milk drink or milk powder.

21. The use of any one of claims 5 to 18, wherein the composition is a pharmaceutical composition and the excipient or diluent comprises a pharmaceutically acceptable diluent, adjuvant, earner or excipient.

22. The use of any one of claims 15 to 21 wherein the use is for preventing rhinitis.

23. The use of any one of claims 15 o 17 to 22 wherein the subject is a neonatal, an infant, or a child subject.

24. The use of any one of claims 5 or 17 to 22, wherein where the subject is a juvenile or an adult subject, the composition comprising Lactobadllus rhamnosus HNOOl or a derivative thereof is administered to the subject.

25. The use of any one of claims .15. to 24, wherein the Ladobadllus rhamnosus HNOOl or derivative thereof is in a reproductively viable form.

26. The use of any one of claims 15 to 24, wherein the Lactobadllus rhamnosus HNOOl ot derivative thereof is killed, fractionated, or attenuated.

27. The use of any one of claims 15 to 26, wherein the rhinitis is rhinoconjunctivitis.

28. The use of any one of claims 15 to 27, wherein the Ladobadllus rhamnosus HNOOl or derivative thereof is administered separately, simultaneously or sequentially with one or more agents selected from one or more probioitics, one or more prebiotics, one or more sources of dietary fibre, one or more galactooligosaccharides, one or more short chain galactooligosaccharides, one or more long chain galactooligosaccharides, one or more fructooligosaccharides, one or more short chain galactooligosaccharides, one or more long chain galactooligosaccharides, inulin, one or more galactans, one or more fructaiis, lactulose, or any mixture of any two or more thereof.

29. Lactobadllus rhamnosus HNOOl or a derivative thereof for treating or preventing rliinitis in a subject.

30. The Lactobadllus rhamnosus HNOOl according to claim 29 wherein the Lactobadllus rhamnosus HNOOl is in a composition comprising a physiologically acceptable diluent, adjuvant, carrier or excipient.

31. The Ladobadllus rhamnosus HNOOl according to claim 30 wherein the composition is for oral administration. ^' 32. The Laclob illus rbamnosus HN001 of any one of claims 29 to 31, wherein the hactobacilks rbamnosus HNOOl or derivative thereof or composition is for oral administration to a pregnant mother during gestation or to a breastfeeding mother.

33. The hactobactllus rbamnosus HNOOl or composition of any one of claims 29 to 32 further comprising one or more agents selected from one or more probioitics, one or more prebiotics, one or more sources of dietary fibre, one or more galactooligosaccharides, one or more short chain galactooligosaccharides, one or more long chain galactooligosaccharides, one or more

fructooligosaccharides, one or more short chain galactooligosaccharides, one or more long chain galactooligosaccharides, inuJin, one or more galactans, one or more fructans, lactulose, or any mixture of any two or more thereof.

34. The LadobadZ/us rbamnosus HNOOl or composition of any one of claims 29 to 33 further comprising an infant formula, a follow-on formula, a growing-up formula, a maternal formula, a maternal supplement, a dietetic product, or a food, including a food comprising one or more of cultured milk, yoghurt, cheese, milk drink or milk powder.