Reducing Urogenital Infections Including HIV in Sub-Saharan Africa - Can Probiotics Be a Viable Paradigm?

Kingsley C. Anukam1,2, Enya B. Bassey3 and Emmanuel O. Osazuwa2 1TWAS Research Unit on Probiotic Genomics, Department of Medical Laboratory Science, School of Basic Medical Sciences, College of Medical Sciences, University of Benin, 2Department of Pharmaceutical Microbiology, Faculty of Pharmacy, University of Benin, 3World Health Organization (WHO), Ministry of Health, Uyo, Akwa Ibom State, Nigeria


Introduction
The economic burden, albeit low quality of life of people in sub-Saharan Africa with nonsexually transmitted recurrent urogenital tract infections is not the desired state of health.Associations exist between abnormal vaginal/penile microbiota and HIV.Excluding sexually transmitted diseases, microorganisms that originate from the gastrointestinal tract cause almost all infections of the vagina and bladder.There is a strong correlation between presence of the normal microbiota, particularly lactobacilli in the vagina with health, and an absence of these microorganisms in patients with urogenital infections.Disruption of the normal vaginal microbiota is caused by the use of broad-spectrum antibiotics, spermicides, hormones, dietary substances and factors not, as yet, fully understood.There is increasing body of evidence that probiotic microorganisms delivered in food such as yogurt or milk based foods and capsular preparations do have a role in preventing urogenital tract infections and in ameliorating diarrhea [Walker et al., 2006].The use of probiotics defined as "live microorganisms which when administered in adequate amounts, confer health benefits on the host" (WHO/ FAO, 2001), for the maintenance of health is already in use in developed countries.Probiotics are yet to be adopted in sub-Saharan Africa by health care providers.There are clinical evidence to show that probiotics can play a significant role in resolving diarrhea, boost immune system and prevent recurrent urogenital infections including bacterial vaginosis, which is a risk factor in HIV acquisition.The aim of this chapter is to highlight the burden of urogenital infections in Africa, impact of abnormal vaginal microbiota, clinical evidence on the use of probiotics for urogenital health care and last but not the least, the rationale for suggesting the use of probiotics in the management of HIV infection in sub-Saharan Africa.

Burden of urogenital infections in Africa
Non-sexually transmitted urogenital infections such as bacterial vaginosis (BV), yeast vaginitis, urinary tract infections (UTI) afflict more than one billion women around the world annually [Hay, 2000].However, it is worthwhile to briefly explain what BV is.185 symptoms as yeast infections and self-treat with over-the-counter antifungals, when in fact they have BV.This misdiagnosis and mistreatment can result in adverse consequences [Ferris et al 1996].Antimicrobial treatment for BV is suboptimal, with some cure rates as low as 60% 1 month after treatment, and subsequent overgrowth of pathogenic bacteria in the vagina often occurs [Livengood et al 1999].

Abnormal vaginal/penile microbiota and HIV
Studies have found a significant association between BV and human immunodeficiency virus (HIV) infection (Sewankambo et al., 1997).In a prospective study of Kenyan sex workers, the absence of lactobacilli in vaginal cultures was associated with a 2.0-fold increase in HIV acquisition and a 1.7-fold greater risk of developing gonorrhea (Martin et al., 1999).Similar studies have also documented evidence indicating that human immunodeficiency virus infection have a strong association with abnormal vaginal microbiotia particularly, bacterial vaginosis [Taha, 1999].Studies have also shown that the absence or depletion of lactobacilli in the vagina associated with overgrowth of anaerobic pathogens causing BV results in significantly increased risk for HIV (as well as gonorrhea, chlamydia, and herpes simplex virus infections) [Wiesenfeld, 2003].By mechanisms yet to be elucidated, BV displaces lactobacilli, elevating vaginal pH and creating an environment within which the pathogens survive and can infect the host.It is important to note also that penile microbiota may contribute significantly to susceptibility to HIV infection.Several randomized trials demonstrated decreased risk of trichomoniasis and bacterial vaginosis (BV) in the female sexual partners of circumcised men (Gray et al 2009).It is very pertinent to better understand the biological mechanisms by which male circumcision reduces the risk of HIV infection as this may lead to the development of novel, non-surgical prevention strategies.It has been asserted that male and female genital microbial communities may play an important role in modulating HIV risk [Galvin & Cohen, 2004].Genital mucosal inflammation induced by microbes leads to the activation of HIV target cells and an increase in HIV susceptibility [de Jong & Geijtenbeek, 2009].The dominant HIV target cells in the genital mucosa are two dendritic cell types, langerin+ Langerhans' cells and DC-SIGN+ dendritic cells.The biological mechanism underlying circumcision-conferred protection against HIV is likely to be multifactorial.Post-circumcision anatomical, immunological, and microbiological changes have all been hypothesized to contribute to the reduction in HIV risk.From the anatomical and immunological perspective, the inner surface of the foreskin is lightly keratinized and contains abundant Langerhans cells close to the mucosal surface resulting in a large number of exposed HIV target cells in the erect uncircumcised penis (Patterson et al., 2002;McCoombe & Short, 2006).From a microbiological perspective, the intact foreskin may support the survival of genital microbes associated with increased foreskin mucosal inflammation and Langerhans' cell activation.Of note, the protection against sexually transmitted infections and BV conferred to the female partners of circumcised men [Bailey et al., 2007;Auvert et al., 2005;Gray et al., 2009] strongly suggests circumcision-associated microbiological changes in the male genital mucosa.In a study that assessed the penile (coronal sulci) microbiota in 12 HIV-negative Ugandan men before and after circumcision revealed a total of 42 unique bacterial families in the coronal sulci microbiota, with 38 bacterial families among pre-circumci-sion samples versus 36 detected among post-circumcision samples.Pseudomonadaceae was the most abundant family irrespective of circumcision status, constituting over 50% of the coronal sulci microbiota, followed by Clostridiales Family XI, Oxalobacteraceae, and Prevotellaceae for pre-circumcision and Corynebacteriaceae, Oxalobacteraceae, and Staphylococcaceae for post-circumcision (Price et al 2010).The study suggests that anoxic microenvironment of the subpreputial space may support pro-inflammatory anaerobes that can activate Langerhans cells to present HIV to CD4 cells in draining lymph nodes.Thus, the reduction in putative anaerobic bacteria after circumcision may play a role in protection from HIV and other sexually transmitted diseases.

The role of normal vaginal microbiota
It has been reported previously that more than 60 different bacterial species colonize the healthy vagina (Reid et al 1990).A recent study sampled the vaginal bacterial communities of 396 asymptomatic North American women who represented four ethnic groups (White, Black, Hispanic and Asian) and the species composition was characterized by pyrosequencing of barcoded 16S rRNA genes.The study revealed that the bacterial communities clustered into five groups: four were dominated by Lactobacillus iners, L. crispatus, L. gasseri, or L. jensenii, whereas the fifth had lower proportions of lactic acid bacteria and higher proportions of strictly anaerobic organisms, indicating that a potential key ecological function, the production of lactic acid, seems to be conserved in all communities.The proportions of each community group varied among the four ethnic groups, and these differences were statistically significant (Ravel et al. 2011).This shows that different vaginal bacterial ecosystems varies in people but conferring the same protective role as the human vaginal microbiota seem to play a key role in preventing a number of urogenital diseases, such as bacterial vaginosis, yeast infections, sexually transmitted infections, urinary tract infections and HIV infections (Taha, 1999).Studies have shown that urogenital cells are covered by dense bacterial biofilms, whose composition changes constantly, but in which lactobacilli predominate, at least until menopause.Vaginal lactobacilli have been shown to inhibit the growth of Gardnerella vaginalis and mobiluncus in vitro, with the greatest inhibition observed in the presence of lactobacilli producing lactic acid.Hydrogen peroxideproducing lactobacilli have been recovered from the vagina of 96% of 28 non-pregnant women without bacterial vaginosis, compared with 6% of 67 women with bacterial vaginosis (Eschenbach et al. 1989).The presence of lactobacilli on vaginal epithelial cells seems to act, not only as a barrier to infection, by blocking adherence of pathogens, but their capability of producing such antibacterial materials as hydrogen peroxide to limit pathogen growth, production of biosurfactants that inhibit pathogen adherence, and their ability to prime macrophages, leukocytes, cytokines, and other host defenses also contribute to the protection of the vagina against uropathogens (Reid, 1999).Full genome sequencing of Lactobacillus plantarum KCA1 isolated from the vagina of a healthy Nigerian woman has revealed the presence of several novel phage defense genes encoding clustered regularly interspaced short pallindromic repeats (CRISPR)-associated proteins and abortive infection systems (Anukam et al. 2011).One of the fastest evolving genetic elements in bacterial genomes are clustered regularly interspaced short palindromic repeats (CRISPRs) (Sorek et al., 2008).CRISPRs have been identified within the genomes of many archaeal and bacterial species especially in some vaginal lactobacilli such L. plantarum KCA-1, the only plantarum strain know to date with CRISPR.The Spacers are derived from foreign nucleic acids, such as those from phage or plasmids and can protect bacteria from subsequent infection by homologous phage and plasmids.As a bacterial immune system against foreign DNA, to massive investment by International organizations, such as the Joint United Nations Programme on HIV/AIDS (UNAIDS), the Global Fund to Fight AIDS, Tuberculosis and Malaria (the Global Fund) and the US President's Emergency Plan for AIDS Relief (PEPFAR).These organizations were formed to specifically address HIV/AIDS.The United Nations estimated that at least US $1.5 billion a year could make it possible to achieve massively higher levels of implementation of all major components of successful prevention programs for the whole of sub-Saharan Africa.Having HIV/AIDS is especially devastating among children from the developing countries such as sub-Saharan Africa.To comprehend the reality of the vulnerability of children affected by the condition, it is important that health care professionals and national policy makers understand the global magnitude of the problem.More than 90% of HIV infection occurs among the children of southern Africa.Seventy-five percent of those children reportedly die before the age of 5 years (DeBaets et al., 2007).Children with HIV/AIDS in developing countries continue to be underrepresented among recipients of antiretroviral therapy or supportive care, including palliative care (Kline, 2006).A significant number of children infected with HIV are also orphans of parents who died of HIV, and they suffer from complications including extreme malnutrition.Children who receive professional care are often hospitalized for many months at the end of life.The need for prevention and alternative management is very obvious Meanwhile, more than half of those in need still do not have access to treatment, and treatment is posing new challenges for sustainable funding.Most countries with mid to high prevalence cannot afford the cost of treatment without international aid; as aid is reduced or cancelled, treatment programmes are threatened, drug resistance develops and large numbers of people living with HIV may die.In countries like Uganda, patients are being turned away from treatment clinics due to lack of resources; 300,000 Ugandans in need of treatment are denied their right to health (McNeil, 2010).These numbers will continue to grow if both effective prevention and sustainably funded treatment programmes are not maintained.Nigeria may not be able to sustain the free anti-retroviral drug program from PEPFAR and the future of new infected person may be grim.In 2009, UNAIDS and WHO predicted that the $25.1 million needed for HIV/AIDS programmes in 2010 would not be forthcoming (UNAIDS, 2009).The total amount of Global Fund grants recommended for funding in 2009 was 35% lower than in 2008 (MSF, 2009).The programs by all these agencies are planned to cover sexual, mother-to-child and transfusion-related HIV transmissions, and would involve approaches ranging from awareness campaigns through media to voluntary HIV counseling and testing and the promotion and supply of condoms.No doubt these programs are laudable, but other alternative measures, such as nutrition fortification and probiotics, have not been included as one of the UNAIDS intervention programs.Death from AIDS is often precipitated by gastrointestinal infections and diarrhea, and indeed, many non-AIDS deaths are due to these infections.It has been estimated that a child dies every 15 seconds from diarrheal diseases.While sanitation and early hydration based interventions can reduce this death rate, probiotics too could play a role.

Probiotics with clinical evidence against urogenital infections
During the early periods of probiotics, Scientists paid more attention to gastrointestinal effects, but in recent times probiotics are now useful for more than just gastrointestinal health.In fact, there are specific probiotic products that can help prevent and treat female urogenital conditions like bacterial vaginosis, vulvovaginal candidiasis, urinary tract infections.It should be noted that the vaginal tract is not internally connected to the alimentary canal, however the two are intimately related.Bacteria that pass through the digestive system can ascend via the perineum to the vagina.So it's almost a no-brainer to expect what promotes gastrointestinal health to have relevance for urogenital health.In the late 1980s, human studies were carried out in which L. rhamnosus GR-1 in a douche suspension was instilled into the vagina (Bruce and Reid, 1988).This was followed by studies using a gelatin capsule containing freeze-dried lactobacilli inserted into the vagina (Reid et al 2001).In both cases, the process did not result in any adverse events but did show a potential to reduce the risk of recurrence of UTI.The use of orally administered lactobacilli was more recently tested, on the basis that if pathogens infect the host from the anal skin, why couldn't lactobacilli also ascend from the anus to the vagina and repopulate the area?This concept was verified in several labs (Reid et al, 2001;Antonio et al 2005), and Lactobacillus strains GR-1 and RC-14 were shown to reduce UTI, BV and yeast pathogens as well as infections (Reid et al 2001;Reid et al. 2003].The mechanism of action is likely multifactorial and could include the ingested lactobacilli ascending from the rectal skin to the vagina, or causing a reduced pathogen ascension, or influencing the immune or host system in a way that reduces infectivity.As this approach to restoration and maintenance of women's health has become more recognized, other groups have undertaken studies using different strains.A prospective clinical pilot study was performed to confirm the safety and effectiveness of Lactobacillus vaginal suppositories against the recurrence of UTI.The patients enrolled in the study were instructed to administer vaginal suppositories containing the strain L. crispatus GAI 98322.A significant reduction in the number of recurrences was noted, without any adverse complication (P=0.0007).The administration of vaginal suppositories containing L. crispatus GAI 98332 seemed to be a safe and promising treatment for the prevention of recurrent UTI (Uehara 2006).Delai et al., (2006) demonstrated the effectiveness of the contemporary oral administration of L. paracasei subsp paracasei F19 in association with vaginal suppositories containing an unnamed L. acidophilus in the treatment of BV.The study had a potentially fatal flaw in that not all the 60 subjects had confirmed diagnosis of BV.The subjects were randomized in 2 groups: Group A treated with vaginal suppositories containing L. acidophilus; Group B treated with the same vaginal suppositories + L. paracasei subsp paracasei F 19 for oral administration.There was a significant reduction of vaginal pH, an improvement in the amine sniff test and in subjective symptomatology after 3 months of treatment and follow-up (3 months).This study needs to be repeated with larger sample size, but nevertheless, reviews of the evidence from microbiological and clinical studies have indicated that probiotics can be beneficial for preventing recurrent UTI in women in a safe manner (Falagas 2006;Reid and Bruce 2006;Hoesl and Altwein 2005).The usefulness of orally administered lactobacilli for urogenital health has been demonstrated in several other important studies.In a randomized, double-blind, placebo controlled trial, 106 women with BV were given a single oral dose of metronidazole (500mg) twice daily from days 1-7, plus oral L. rhamnosus GR-1 and L. reuteri RC-14 or placebo twice daily from days 1-30.The cure rate in the antibiotic/probiotic group was 88% compared to 40% in the antibiotic/placebo group (p<0.001).High counts of Lactobacillus sp.
(>10 5 CFU/ml) were recovered from the vagina of 96% of probiotic treated subjects compared to 53% controls at day 30 (Anukam et al 2006a).In another study using the same probiotics, there was a 90% cure of BV following intravaginal administration of the probiotic alone, compared to 33% cure with intravaginal metronidazole treatment.In the study, 40 women diagnosed with BV were randomized to receive either two dried capsules containing L. rhamnosus GR-1 and L. reuteri RC-14 each night for five days, or 0.75% metronidazole gel, applied vaginally twice a day (in the morning and evening).Follow-up at day 6, 15 and 30 showed cure of BV in significantly more probiotic treated subjects (16, 17 and 18/20 respectively) compared to metronidazole treatment (9, 9 and 11/20: P= 0.016 at day 6, P= 0.002 at day 15 and P= 0.056 at day 30) [Anukam et al 2006b].This is the first p r o v e n c u r e o f B V u s i n g p r o b i o t i c s a n d p r o v i d e s h o p e t h a t a l t e r n a t i v e r e m e d i e s t o antibiotics can be found.

Mechanisms of action
Probiotics apparently fulfills the definition as "live microorganisms which, when administered in adequate amounts, confer a health benefit on the host" through a variety of somewhat disparate, somewhat overlapping mechanisms.These include the regulation of intestinal microbial homeostasis, the interference with the ability of pathogens to colonize and infect the mucosa, the modulation of local and systemic immune responses, the stabilization or maintenance of the gastrointestinal barrier function, the inhibition of procarcinogenic enzymatic activity and the induction of enzymatic activity that favors good nutrition.For example, with respect to epithelial barrier function, probiotics can conceivably act through general antimicrobial effects, effects on intestinal permeability in the absence of invasive bacteria, effects on epithelial cell inflammatory responses and effects on epithelial cell survival.One important effect of probiotics on barrier function is the ability of commensal organisms to act through Toll-like Receptors (TLR) on epithelial cells (such as TLR2 and TLR4).In particular, it has been shown that such interactions induce the production of protective cytokines such as IL-6 that mediate epithelial cell regeneration and inhibit epithelial cell apoptosis in the face of agents that otherwise result in epithelial cell ulceration (Rakoff-Nahoum et al., 2004).It should be noted, however, that probiotics do not affect expression of all TLRs in the same manner.Thus, as shown by Ewaschuk et al., (2007) whereas exposure of HT-29 epithelial cells to DNA from pathogenic organisms resulted in increased TLR9 expression, exposure to a probiotic did not have this effect.This is in keeping with the lesser ability of commensal organisms to induce TLR expression as compared with pathogenic organisms.Probiotics may also have effects on epithelial barrier function via cellular mechanisms that have little to do with TLR signaling.Thus, in a study by Zyrek et al., (2007) it was shown that the probiotic, E. coli Nissle 1917 (EcN) counteracts the disruptive effects of enteropathic E. coli (EPEC) on T-84 epithelial cell monolayers by altering protein kinase C signaling and causing the redistribution and increased expression of zonula-occludens-2 (ZO-2), an important factor in maintaining epithelial tight junction function.Along similar lines, Yan et al., (2007) showed that proteins isolated from broths of the probiotic Lactobacillus rhamnosus activated mouse epithelial cell Akt and inhibited TNFα-mediated apoptosis and promoted growth of both human and mouse colon epithelial cells or cultured mouse colon explants.

Decrease pathogen adhesion Table 1. Some proposed probiotics mechanisms of actions
From the accumulated information on the relation of regulatory T cells and their associated cytokines to the function of probiotics, it can be said with some certainty that probiotic administration does lead to the increased elaboration of regulatory cytokines and these cytokines play a major role in the protective effect of the probiotics.It seems likely that probiotics bring about these effects via their interactions with mucosal dendritic cells which then produce regulatory cytokines themselves or induce T cells with these properties.There is the possibility that probiotics act through the induction of regulatory T cells that suppress inflammation-inducing effector cells.This emphasis is based on a rising tide of evidence that probiotics have properties that allow them to interact with the mucosal immune system that does not arouse an inflammation inducing innate response and the consequent induction of master inflammatory cytokines such as IL-12.For example human circulating dendritic cells or lamina propria mononuclear cell populations were cultured with cell wall preparations from each of the probiotic species in VSL#3 and then assessed cytokine production.It was found that bifidobacteria components were generally the most potent in up-regulating IL-10 by both CD11+ and CD11− dendritic cells whereas components of all VSL#3 strains decreased IL-12 production (Mohamadzadeh et al., 2005).While studies on vaginal lactobacilli with respect to their probiotic potential are increasing, significant basic research studies have only been performed on strains GR-1 and RC-14, particularly looking at modes of action.It is clear that these organisms are multifunctional and may affect each host differently.The GR-1 strain does not produce hydrogen peroxide but does produce anticandida activity as well as bacteriocins and other components, including AI-2 inducers that influence the growth and biofilm development of uropathogens.The organism can downregulate inflammatory processes, using IL-10 dependent and independent pathways, as shown in macrophages studies (Kim et al. 2006).A human genome array study has shown that a single intravaginal insertion of Lactobacillus GR-1 can up-regulate host defense factors known to be important in fighting infection (Kirjavainen et al., 2008 ).Meanwhile, studies with Lactobacillus RC-14 strain have shown that it can up-regulate mucin production which may act as a barrier to infection (unpublished data), and down-regulate virulence factor expression in pathogens such as staphylococci [Laughton et al 2006].The organism also affects cell membrane components in E. coli and produces biosurfactants that inhibit their adhesion to surfaces.

The rationale for the use probiotics in reducing the risk associated with HIV
Tremendous efforts are being made to develop effective microbicides for the prevention of HIV-1 sexual transmission and in clear terms it should represents a primary goal for the control of AIDS epidemics worldwide.A promising strategy is the use of bacteria belonging to the vaginal microbiota as live microbicides for the topical production of HIV-1 inhibitors.A recent review has chronicled the potentials of probiotics to prevent HIV (Bolton et al., 2008) and some of the points are lighted in line with the objectives of this section.A study engineered a human vaginal isolate of Lactobacillus jensenii to secrete the anti-HIV-1 chemokine RANTES, as well as C1C5 RANTES, a mutated analogue that acts as a CCR5 antagonist and therefore is devoid of proinflammatory activity.Full-length wild-type RANTES and C1C5 RANTES secreted by L. jensenii were purified to homogeneity and shown to adopt a correctly folded conformation.Both RANTES variants were shown to inhibit HIV-1 infection in CD4 T cells and macrophages, displaying strong activity against HIV-1 isolates of different genetic subtypes.The work provides proof of principle for the use of some Lactobacilli, notably L. jensenii-produced C1C5 RANTES to block HIV-1 infection of CD4 T cells and macrophages, setting the basis for the development of a live anti-HIV-1 microbicide targeting CCR5 in an antagonistic manner [Luca et al 2010] Other studies have previously used Lactobacilli and other probiotic genetically modified bacteria to produce specific HIV inhibitory proteins, both membrane-bound and secreted.A strain of L. jensenii was engineered to produce functional CD4, the primary receptor for HIV [Chang et al, 2003].There are several classes of proteins that bind to the mannose residues of HIV, including a unique 11 kd protein from cyanobacteria (Nostoc ellipsosporum) called cyanovirin-N and mannose-binding lectins which also bind to Neisseria gonorrhoeae [Botos & Wlodawer, 2005 ].The human commensal bacterium Streptococcus gordonii, L. lactis, L. plantarum, and L. jensenii have been genetically engineered to express functional, HIVbinding cyanovirin-N [Pusch et al 2005;Liu et al 2006].An E. coli strain which colonizes the colon and rectum, and may thus potentially prevent anal transmission, has been modified to secrete peptides hybridized with hemolysin A, a protein that can complex with the HIV fusion protein gp41 [Rao et al 2005].FI-1, FI-2, and FI-3 are also peptides that can interfere with gp41 and were cloned into L. plantarum and L. gasseri [Pusch et al 2006].L. reuteri RC-14, which has been studied as a potential probiotic, was modified to produce 3 HIV entry and fusion inhibitors: CD4D1D2IgKLC, MIP-1, and T-1249 [Liu et al 2006] Antibody to the cellular adhesion molecule intercellular adhesion molecule has been shown to inhibit cellmediated trans-epithelial HIV-1 transmission in vitro and was functionally excreted by a bioengineered strain of L. casei [Chancey et al 2006].Some of these products have been used in rodent models, but none has been tested in humans.By their production of lactic acid, lactobacilli may help maintain a low vaginal pH that can inhibit other bacteria and viruses.Among postmenopausal women receiving estrogen replacement therapy, those who were lactobacilli-positive had low pH 4.4 compared with the lactobacilli-negative women pH 5.2 (Ginkel et al 1993).In 55 menarchal women, colonization with hemolytic streptococci, G. vaginalis, or mixed organisms was associated with higher vaginal pH than colonization with normal microbiota and yeast (Caillouette et al. 1997).As previously mentioned, BV is associated with a lack of hydrogen peroxideproducing lactobacilli.Colonization with hydrogen peroxide producers is associated with lower frequency of gonorrhea (Antonio et al 1999).Compared with women colonized with hydrogen peroxide-producing lactobacilli, women colonized with hydrogen peroxide nonproducers or without lactobacilli had unadjusted odds ratios of HSV-2 seroconversion of 2.4 and 2.6, respectively (Cherpes et al 2003).The difference in seroconversion rates may be due to a direct antiviral effect of hydrogen peroxide or due to an increased risk of HSV infection in women with BV.Several mammalian peroxidases, including myeloperoxidase, eosinophil peroxidase, and lactoperoxidase can combine with hydrogen peroxide and a halide (chloride, iodide, bromide, thiocyanate) to form "a powerful antimicrobial system" effective against many pathogens (Klebanoff et al 1991).Lactobacilli, streptococci, and pneumococci can release hydrogen peroxide and in vitro, in mixed cultures, their toxicity against other bacteria, fungi, viruses, spermatozoa, and tumor cells can be boosted by addition of a peroxidase and a halide.This system was found to inhibit cell-free HIV replication in culture in the presence of hydrogen peroxide-producing, but not nonproducing, lactobacilli (Klebanoff et al 1991a, Klebanoff et al 1991b).L. crispatus and L. jensenii inhibit gonococci at both acidic and neutral pH.This inhibition is susceptible to bovine catalase, suggesting that hydrogen peroxide is the primary mediator (St Amant et al 2002).Lactobacilli have been found to produce many other bacteriocins, enzymes, and antimicrobial peptides that may make them more competitive in the vaginal microbiota (Silva et al 1987;Naidu et al 1999).Another potentially beneficial characteristic of probiotic bacteria studied in the gastrointestinal tract is their pleomorphic effect on host mucosal immunity that could affect the vaginal mucosa's defense against HIV and other STIs or resistance to BV (Sheih et al 2001).Evidence for enhancement of humoral responses to rotavirus and Salmonella typhi has been shown through IgA levels in probiotic treated children and adults (Kaila et al., 1992).Animal and human data suggest that probiotic use is associated with induction of innate and cell-mediated immune responses (Cross 2002), including increased macrophage phagocytic activity (Miettinen et al 2000), complement and reticuloendothelial activation (Gill et al 2000), stimulation of interferon-gamma, interleukin (IL)-12, and IL-18 (Hessle et al., 1999) and increased natural killer cell activity (Matsuzaki and Chin, 2000).However, the major cell wall component of lactobacilli, muramyldipeptide, can be pyrogenic, and there is concern that lactobacillus enhancement of the T-helper-1 proinflammatory pathway could have negative health consequences (Perdigon and Holgado, 2000).Also, there is concern that the dosage and duration of therapy must be considered so as to optimally enhance and not suppress immunity (Perdigon et al., 1991).Fortunately, probiotic strains have not been found to cause a systemic antibody response (Sheen et al 1995).Modification of intestinal mucosal immunity by orally administered probiotics may also affect vaginal mucosal immunity to specific pathogens (de Vrese and Schrezenmeir, 2002).In fact, genetic engineering of probiotic bacteria to express pathogen antigens and serve as oral vaccines is being explored (Reveneau et al 2002).In other mucosal vaccine trials, introduction of antigens from HPV and HIV has been accomplished in S. gordonii (an oral commensal) and L. casei.Local and systemic immune responses were detected in BALB/c mice and Cynomolgus monkeys after vaginal colonization with these strains (Medaglini, 1998).However, there is not yet a good understanding of the relationship between mucosal immunity and BV.In a study of adolescents, BV was found to be inversely associated with lactobacilli counts but was independent of lactobacilli-specific immune responses in isolated peripheral blood leucocytes, and independent of local immune responses measured by antibodies and cytokines measured in cervicovaginal lavages (Alvarez-Olmos et al 2004).In a study of pregnant women, carriage of a variant of the toll-like receptor-4 gene compared with carriage of the dominant genotype was associated with higher vaginal pH and a 10fold increase in vaginal G. vaginalis levels.Colonization with G. vaginalis or anaerobic Gramnegative rods in the dominant allele carriers was associated with elevated vaginal IL-1 and IL-1ra but not in the variant homozygotes (Genc 2004).Probiotic modulation of mucosal immunity may help prevent BV and other STIs including HIV, but it is difficult to predict which strains might do this without knowing the immune correlates of protection and how specific strains will affect these factors.More studies are needed in these areas of scientific enquiry.

Some clinical evidence of probiotics in helping to reduce the impact of HIV/AIDS
The increased searchlight on HIV/AIDS care in Africa has been rekindled following financial donations from spirited organizations and individuals on increasing access to antiretroviral (ARV) medication.Nevertheless this is cardinal, efforts are also needed to provide safe and affordable interventions for those without access to ARVs or with CD4 counts too high to initiate ARV therapy, yet whose quality of life is diminished by micronutrient deficiencies, gut infections such as diarrhea, and other complicated conditions associated with HIV infection.The use of micronutrients, most notably vitamin B-complex in combination with C and E, appears to be an effective intervention that have been associated with reduced mortality and in most cases it results in increasing the CD4 counts of the infected person.(Kanter et al., 1991;Kaiser et al., 2006;Jiamton et al., 2003) The WHO/FAO has recommended that an ''increased micronutrient intake can be best achieved through an adequate diet,'' (WHO, 2003) favoring food-based interventions.It should be noted that the gastrointestinal tract is one of the most severely affected organ by HIV (Kotler et al., 1984;Sharpstone et al., 1999).Inflammation results in damage to the epithelial barrier, leading to an increased leakage of microbial products into the bloodstream.Recently, it was theorized that this may be an ongoing source of systematic immune activation that fuels HIV (Brenchley, 2006) although this association was less clear in an African population (Redd et al., 2009).Capsule proteins of HIV may further facilitate viral replication by eliciting a profound Th-2 activation that inhibits an effective immune response against the virus (Kanter et al., 1991).Probiotic bacteria can potentially restore an effective gut barrier (Luyer et al., 2005;Yan et al., 2007) thereby reducing systemic immune activation.Furthermore, probiotics have been shown to upregulate T-regulatory lymphocytes (Baroja et al., 2007;Foligne et al., 2008) potentially skewing the immune system away from a Th-2-dominant state (Mohamadzadeh et al., 2005).Probiotic usage has been shown to be safe among people living with HIV (Wolf et al., 1998;Salminen et al., 2004) and recent randomized trials in Brazil (Trois et al., 2008) and Nigeria (Anukam et al., 2006) suggest that probiotic use can increase the CD4 count.In the Nigerian study, conventional yogurt fermented with Lactobacillus delbruekii var bulgaricus and Streptococcus thermophilus was supplemented with probiotic Lactobacillus rhamnosus GR-1 and L. reuteri RC-14.Twenty-four HIV/AIDS adult female patients (18 to 44years) with clinical signs of moderate diarrhea, CD4 counts over 200, and not receiving antiretrovirals or dietary supplements, consumed either 100 mL supplemented or unsupplemented yogurt per day for 15 days.Hematologic profiles, CD4 cell counts, and quality of life was evaluated at baseline, 15 and 30 days postprobiotic-yogurt feeding.There was no significant alteration in Reducing Urogenital Infections Including HIV in Sub-Saharan Africa -Can Probiotics Be a Viable Paradigm?195 the hematologic parameters of both groups before and after the probiotic-yogurt feeding, with the probiotic yogurt group at baseline, 15 and 30 days had a mean WBC count of 5.8 ± 0.76 X10 9 /L, 6.0 ± 1.02 X10 9 /L, and 5.4 ± 0.14 X10 9 /L, respectively.However, the mean CD4 cell count remained the same or increased at 15 and 30 days in 11/12 probiotic-treated subjects compared to 3/12 in the control.Diarrhea, flatulence, and nausea resolved in 12/12 probiotictreated subjects within 2 days, compared to 2/12 receiving yogurt for 15 days.This is the first study to show the benefits of probiotic yogurt on quality of life of women in Nigeria with HIV/AIDS, and suggests that perhaps a simple fermented food can provide some relief in the management of the AIDS epidemic in Africa.The most commonly used vehicle for supplying probiotics; yogurt, is a significant source of vitamin A, B-complex, zinc, and high-biologic-quality protein (Sattler et al., 2008) and is therefore, an excellent food-based intervention to improve micronutrient intake among people living with HIV/AIDS.On the basis of these notions, a community kitchen in Mwanza, Tanzania, was established in 2004 to produce yogurt supplemented with Lactobacillus rhamnosus GR-1 (Fiti) to be distributed as an adjunct to the diet of people living with HIV.Irvine et al., (2010) carried out an observational retrospective study over a period of 3 years, with longitudinal comparison of the CD4 count within participants (n=68) before and during probiotic yogurt consumption, and compared with a control group of participants not consuming the yogurt (n=82).Among the yogurt consumers before use and the nonconsumers, an average increase in CD4 count was seen of 0.13 cells/mL/day (95% CI; 0.07-0.20,P=<0.001).After commencing consumption, yogurt consumers experienced an additional increase of 0.28 cells/mL/day (95% CI; 0.10-0.46,P=0.003).When adjusting for length of time using antiretroviral medication, the additional increase explained by yogurt consumption remained 0.17cells/mL/day (95% CI; 0.01-0.34,P=0.04).Treatment with antiretroviral medication was associated with an increase of 0.27 cells/mL/day (95% CI; 0.17-0.38,P=<0.001).The study concluded that introduction of probiotic yogurt, made by local women in a low-income community in Tanzania, was significantly associated with an increase in CD4 count among consumers living with HIV.Another similar study assessed among women with HIV, whether long-term oral Lactobacillus rhamnosus GR-1 and Lactobacillus reuteri RC-14 supplementation can prevent bacterial vaginosis (BV) and enhance the cure rate of metronidazole among those with BV.It involved a randomized, double-blind, placebo-controlled trial conducted among 65 HIV-infected women with an aberrant microbiota (Nugent score 4-10) were selected to receive daily probiotics or placebo for 6 months.Those with BV (Nugent score 7-10) additionally received metronidazole for 10 days (400 mg twice daily).Although there was no enhanced cure rate of BV among women with HIV treated with adjuvant probiotics to metronidazole treatment.However, among women with an intermediate vaginal flora, probiotics tended to increase the probability of a normal vaginal flora (odds ratio 2.4; P=0.1) and significantly increased the probability of a beneficial vaginal pH (odds ratio 3.8; P=0.02) at follow-up (Hummelen et al., 2010)

Beware of some probiotic claims
While we suggest the potential use of probiotics may be important in ameliorating the impact of urogenital infections in Africa, however there must be caution in terms of controlling the importation of probiotic products.National agencies in Africa in charge of natural products should be prepared to know the probiotic products there are really called probiotics.There are numerous probiotic claims over the internet which may mislead people in buying useless products.
A number of so-called probiotic products claim to be useful for UTI management.Sadly, their marketing hype is not supported by properly performed human studies.US based Natren, makers of probiotic face cream, claim that oral and vaginal GyNaTren helps 'balance the intestinal and vaginal ecology' and should be used to treat vaginitis.No peer-reviewed studies validate this claim.Other products available in Europe, namely Lactovaginal, Gynoflor, Fermalac, Florgynal, Ecovag, Culturelle VC, SymbioFem Plus and Yeast-guard all suffer from lack of appropriate clinical sub-stantiation, and cannot be recommended to prevent UTI.Clearly, companies marketing such products have an obligation to provide evidence of clinical efficacy.This is predicated on the fact that some products may contain different genera, different species, or even different strains of the same species, and not all products should be expected to work the same.Therefore, claims of efficacy should be target specific and should be made only for products that have been found efficacious in carefully designed studies.The marketplace has many examples of different strains of the same species: Lactobacillus acidophilus NCFM and La-1; L. rhamnosus GR-1 and L. rhamnosus GG; Lactobacillus casei Shirota and DN-114 001; Lactobacillus reuteri RC-14 and ATCC 55730; and Bifidobacterium lactis HN019 and BB-12; Lactobacillus plantarum WCFS1, Lactobacillus plantarum KCA-1.Each of these strains has a unique dossier to document individual health benefits.It is noteworthy, however, that among dozens of European commercial products, the same biotype (based on pulsed-field gel electrophoresis of chromosomal DNA) was predominant among Bifidobacterium-containing products (Masco et al., 2005), suggesting that Bifidobacterium strains used commercially may not be so diverse.The fact that consumers buy such products suggests that the current management of urogenital infections is not sufficiently well handled by the sole use of antimicrobials.In addition, patients, particularly those suffering from recurrent infections, are well aware of the side effects of antibiotic therapy and the everincreasing problem of bacterial resistance.

Can the use of probiotics make an impact in Africa?
The presence of bacterial vaginosis-causing organisms that are perhaps more prevalent in Africa provokes the loss of normal vaginal bacterial flora, and causes vaginal inflammation and increased pH levels.The resulting altered vaginal environment increases the risk of transmission of HIV.In addition diarrhoeal diseases, HIV/AIDS complications, and other infectious diseases are major contributors to morbidity and mortality in sub-Saharan Africa.Morbidity from these illnesses causes economic hardship and mortality results in the loss of the next generation and destruction of the present adult leadership.Several studies have pointed out that the use of probiotics can significantly improve the wellbeing of individuals infected with HIV/AIDS (Anukam, 2007).For example, studies conducted in Africa have estimated the average annual increase in CD4 count of 90cells/mL with ARV treatment (Lawn et al., 2006) and an average decline of 20 to 50 cells/mL/year without ARV treatment (Holmes et al., 2006;Urassa et al., 2004).In the Tanzanian study, (Irvine et al., 2010), a similar rate of increase was observed of 99 cells/mL with ARV treatment (0.27 cells/mL/d), whereas no significant decrease was observed without ARV treatment.The results of this study indicate that probiotic yogurt consumption is associated with an overall increase in CD4 count of 62 cells/mL/year (0.17 cells/mL/d).This could be due to an accelerated immune reconstitution after initiation of ARV treatment, thus shortening the time of severe immune deficiency, or may be due to an increase in CD4 count among those not yet eligible for ARV treatment, which may potentially delay the need for ARV medication.However, this study is yet to be replicated in other African sites with similar burden of HIV infections.

Conclusions
Several studies have shown that probiotics could enhance the health and well-being of individuals in sub-Saharan Africa, but sadly the use of probiotics has not become popular for several reasons outlined by Anukam and Reid (2005).First, pharmaceutical companies that manufacture probiotics would be forced to lower prices, which would adversely affect their revenues.Secondly, storage and distribution problems make the allocation of probiotics difficult.Dairy versions of probiotics require refrigeration, and other forms incorporated in tablets, capsules, and powders must be retained in proper vials with appropriate dessicants.Accordingly, storage and distribution issues present major challenges to the effective implementation of probiotic treatment since domestic technology is frequently insufficient for proper maintenance.Finally, cultural acceptance presents a major challenge for probiotic use.For example, if local customs call for a diet free of dairy products, it could be difficult to convince these people to consume a fermented milk drink.The lack of any probiotic fermented food products in Africa at present, with the exception of a grass roots community kitchen in Tanzania where probiotic Lactobacillus rhamnosus GR-1 is used (www.westernheadseast.ca),makes it difficult to perform studies and provide benefits to the population.Until this happens, hopefully with clinically proven products and not simply ones where the term probiotic is used or bacteria are added as an ingredient without testing, indigenous fermented foods may be explored further as a source of health benefits.While many fermented foods are eaten in Africa soon after being produced, the lack of refrigerated distribution networks makes it difficult to reach a large population.In this context, McMaster et al., (2005), developed a microencapsulation delivery system for Bifidobacterium lactis DSM 10140, which if successfully applied to foods might overcome some of the shelf-life problems faced by lack of refrigeration.The study used two existing traditional fermented foods, ''amasi'' and ''mahewu.''The gellan/xanthan microcapsules containing viable B. lactis were tested under simulated physiological conditions and added to pasteurized beverages.The results showed protection of the organism in low pH and against the biocidal activity of pancreatic and bile acids.For ''mahewu,'' microencapsulation of B. lactis with storage aerobically at 4oC and 22oC enhanced survival over a 21-day period compared to free cells.This is still not reflective of extreme temperatures found in Africa, so further studies are needed to confirm if this method will lead to making 'mahewu' available to people.
In confirming the validity of evidence on probiotics, the United Nations Food and Agriculture Organization and the World Health Organization (WHO/FAO, 2001) sponsored an Expert Consultation in 2001.The resulting Report stressed that "Efforts should be made to make probiotic products more widely available, especially for relief work and populations at high risk of morbidity and mortality".Eleven years later, this clarion call to the healthcare community and to the food industries in providing probiotics to the people of sub-Saharan Africa has yet to be answered.
With the increasing number of people living and dying of AIDS, only a few people have easy access to life-prolonging antiretroviral drug therapy.The HIV/AIDS pandemic is propelled by social, cultural and economic gender inequalities that limit children and women's ability to protect themselves from infection.The main strategies by UNAIDS for HIV prevention are promotion of condoms, reducing the number of sexual partners, and treatment of reproductive tract infections.These strategies are laudable but not feasible for many women.Occasionally, it may seem appropriate to provide perspective and to verbalize concerns and disquietude anytime the UN publishes the life expectancy of the people from developing countries.No one has taken the pain to ask the question why the Japanese, coincidently with a long history of probiotic consumption, have the highest life expectancy in the world today and low HIV prevalence (<0.1%, contributed through contaminated blood products) (Oelrichs, 2004), while the people from sub-Saharan Africa, coincidently with no classically defined probiotic consumption, have the lowest life expectancy in the world, and the highest HIV prevalence.Besides, there are no available probiotics to boost the immune system (Gill et al., 2002), treat diarrhea and prevent urogenital infections.
More and interest are needed to expand our understanding of this approach to healthcare in sub-Saharan Africa, to knowing their limitations and understanding their mechanisms of action, and examining economic, social, political and behavioral changes for more expansive introduction of probiotic concepts.Hopefully, concerted research efforts will stimulate other scientists, students, and physicians to explore this area.The development of recombinant strains expressing potent anti-HIV microbicides may provide a more specific preventive approach in future years.
Making probiotic products available to both the uninfected and infected persons, gives them opportunity to choose among other health promoting products.Probiotics may be a viable paradigm towards reducing the risk associated with acquisition of HIV mostly in women by restoring the vaginal microbiota and help relieve the suffering of AIDS patients, most of who suffer from chronic diarrhea.

Acknowledgement
Dr. Kingsley C Anukam research on probiotic genomics is funded by the Academy of Sciences for the Developing World (TWAS).
www.intechopen.comReducing Urogenital Infections Including HIV in Sub-Saharan Africa -Can Probiotics Be a Viable Paradigm?191 Production of antimicrobial substances, such as organic acids or bacteriocins Upregulate immune response (eg, secretory IgA) to possible pathogens or to vaccines Downregulate inflammatory response Assist in early programming of the immune system to result in a better balanced immune response and reducing risk of development of allergy Improvement of gut mucosal barrier function Enhance stability or promote recovery of commensal microbiota when perturbed Modulate host gene expression Deliver functional proteins (eg, lactase) or enzymes (natural and cloned)

Table 2 .
Some probiotic products with published clinical studies