PerR confers phagocytic killing resistance and allows pharyngeal colonization by group A Streptococcus.

The peroxide response transcriptional regulator, PerR, is thought to contribute to virulence of group A Streptococcus (GAS); however, the specific mechanism through which it enhances adaptation for survival in the human host remains unknown. Here, we identify a critical role of PerR-regulated gene expression in GAS phagocytosis resistance and in virulence during pharyngeal infection. Deletion of perR in M-type 3 strain 003Sm was associated with reduced resistance to phagocytic killing in human blood and by murine macrophages in vitro. The increased phagocytic killing of the perR mutant was abrogated in the presence of the general oxidative burst inhibitor diphenyleneiodonium chloride (DPI), a result that suggests PerR-dependent gene expression counteracts the phagocyte oxidative burst. Moreover, an isogenic perR mutant was severely attenuated in a baboon model of GAS pharyngitis. In competitive infection experiments, the perR mutant was cleared from two animals at 24 h and from four of five animals by day 14, in sharp contrast to wild-type bacteria that persisted in the same five animals for 28 to 42 d. GAS genomic microarrays were used to compare wild-type and perR mutant transcriptomes in order to characterize the PerR regulon of GAS. These studies identified 42 PerR-dependent loci, the majority of which had not been previously recognized. Surprisingly, a large proportion of these loci are involved in sugar utilization and transport, in addition to oxidative stress adaptive responses and virulence. This finding suggests a novel role for PerR in mediating sugar uptake and utilization that, together with phagocytic killing resistance, may contribute to GAS fitness in the infected host. We conclude that PerR controls expression of a diverse regulon that enhances GAS resistance to phagocytic killing and allows adaptation for survival in the pharynx.

Adherence-inhibitory intestinal immunoglobulin a antibody response in baboons elicited by use of a synthetic intranasal lectin-based amebiasis subunit vaccine.

We designed an amebiasis subunit vaccine that is constructed by using four peptide epitopes of the galactose-inhibitable lectin heavy subunit that were recognized by intestinal secretory immunoglobulin A (IgA) antibodies from immune human subjects. These epitopes are contained in the region encompassing amino acids 758 to 1134 of the lectin heavy subunit, designated LC3. Baboons (Papio anubis) are natural hosts for Entamoeba histolytica; naturally infected baboons raised in captivity possess serum IgA antibodies to the same four LC3 epitopes as humans. Uninfected, seronegative baboons received four intranasal immunizations at 7-day intervals with the synthetic peptide vaccine (400, 800, or 1,600 mug per nostril) with cholera toxin (20 mug) as the adjuvant. As determined by an enzyme-linked immunosorbent assay (ELISA), each dose of the peptide vaccine elicited antipeptide serum IgA and IgG and intestinal IgA antibody responses in all six immunized baboons by day 28, 7 days after the last immunization (P, <0.01 for each dose compared to the cholera toxin control). The peptide vaccine elicited serum IgG and intestinal IgA antibodies that recognized purified recombinant LC3 protein (P, <0.008 and 0.02, respectively) and native lectin protein (P < 0.01). In addition, an indirect immunofluorescence assay with whole trophozoites (P < 0.01) and Western blot analysis confirmed that serum IgG antibodies from vaccinated baboons recognized native lectin protein on the surfaces of axenic E. histolytica trophozoites or from solubilized amebae. All four synthetic peptides were immunogenic; the vaccine elicited dose- and time-dependent responses, as determined by ELISA optical density readings indicating the production of serum and intestinal antibodies (P, <0.02 for antipeptide and antilectin antibodies). As a positive control, intranasal immunization with purified recombinant LC3 protein with cholera toxin as the adjuvant elicited a serum anti-LC3 IgA and IgG antibody response (P, 0.05 and <0.0001, respectively); however, no intestinal anti-LC3 IgA antibody response was observed (P = 0.4). Of interest, serum IgA and IgG antibodies elicited by the recombinant LC3 vaccine did not recognize any of the four putatively protective LC3 peptide epitopes. Both serum and fecal antibodies elicited by the peptide vaccine exhibited neutralizing activity, as determined by their dose-dependent inhibition of the galactose-specific adherence of E. histolytica trophozoites to Chinese hamster ovary cells in vitro (P, <0.001 for each group of antibodies compared to the control). In summary, a lectin-based intranasal polylysine-linked synthetic peptide vaccine was effective in eliciting an adherence-inhibitory, intestinal antilectin IgA antibody response in baboons. Future studies with the baboon model will determine vaccine efficacy against asymptomatic E. histolytica intestinal infection.

Jejunal intussusception and small bowel transmural infarction in a baboon (Papio hamadryas anubis).

A 4.3-y-old, colony-bred female baboon (Papio hamadryas anubis) of low social rank and exhibiting no clinically significant signs of illness or distress was found dead at the Oklahoma University Health Sciences Center baboon breeding facility at El Reno, OK. Prior to death she exhibited excessive grooming behavior both toward herself and other baboons. In addition, she was consistently shy, timid, reclusive, and prone to minimal sustained movement (that is, generally lethargic behavior). Animals of low social rank typically exhibit some degree of these behaviors in order to avoid surplus interactions with other animals within their groups, which can lead to conflict and injury. Accordingly, her death was surprising in view of the apparent lack of clinical signs. Necropsy established the cause for death as systemic shock with resultant cardiovascular collapse resulting from a massive jejunal intussusception. This intussusception and resulting entrapment of the jejunal mesenteric vasculature caused total occlusion of the small bowel blood supply, with resulting hemorrhage and ischemic necrosis (small bowel infarction). Jejunal intussusceptions generally are considered to be uncommon and therefore are rarely reported in either the veterinary or human literature. Of special interest was the cause for this intussusception, determined to have been a large hairball located at the most proximal portion of the jejunum. Extending from this hairball and traversing essentially the entire length of the jejunum was a braided strand of hair acting as a string foreign body about which the intussusception formed. In light of our findings we suggest that animals of low social rank exhibiting excessive grooming behavior and lethargy might merit clinical evaluation to rule out possible abdominal disorders.

The effects of confinement environment on reproductive efficiency in the baboon.

Historically, to supply animals for medical research, both captive-bred baboons and imported wild-caught animals have been available. Now that imported animals are difficult to obtain, it is important to maximize domestic production. To this end, it is necessary to determine the optimum housing conditions (i.e., environmentally dependent factors) under which baboons have the greatest reproductive efficiency. At our institution, the Oklahoma University Health Sciences Center Primate Facility, we recently moved the majority of our baboon breeding colony into a large indoor-outdoor facility (El Reno) from a small, indoor facility (Annex). Fortuitously, this move allowed a direct comparison of baboon reproductive efficiency between the two radically different environments. The environment at the Annex is exclusively indoor and possesses limited but adequate living space (per Guide for the Care and Use of Laboratory Animals recommendations), whereas the El Reno environment is indoor-outdoor and naturalistic with living space exceeding Guide recommendations. Although we expected animals at the El Reno facility to exhibit somewhat increased reproductive efficiency, the magnitude of the increase was surprising: the mean number of days post-partum to first estrus was 165 for animals housed in the Annex, but 69 for those at the El Reno facility. In addition, the mean number of days from first estrus to conception was 61 for baboons in the Annex compared with 47 for those at El Reno, and the mean number of days from conception to next conception was 403 for animals in the Annex but 296 for those at El Reno. These results demonstrate that a change in housing environment can dramatically increase baboon breeding efficiency.

The effects of uncomplicated miscarriages and stillbirths on the ability of baboons to return to cyclicity and subsequently conceive.

Pregnant baboons are used regularly in medical research studies. Occasionally these studies have resulted in stillbirths and/or miscarriages. In addition, pregnant animals can spontaneously undergo stillbirths or miscarriages unrelated to any medical or research procedure. In the absence of identifiable inflammatory, infectious, or pathologic processes, it generally was assumed that these events had no bearing on the baboon's future ability to return to cyclicity and conceive. However, these assumptions were based on observational and anecdotal evidence. To test the validity of these assumptions, we established two data groups: baboons that had uncomplicated stillbirths (Gp-1; n = 11) and those that had uncomplicated miscarriages (Gp-2; n = 12). The mean number of days from first detectable postpartum estrus (i.e., perineal swelling/turgescence) to conception was 49 days for Gp-1 and 53 days for Gp-2. In addition, for Gp-1 animals we determined that the mean number of days to the first indication of estrus was 29 days; these data were unavailable for Gp-2 because of the lack of parturition as a reference point. Control baboons (lactating mothers) required approximately 59 days from first detectable estrus to conception, and our findings for Gp-1 and Gp-2 were consistent with this value. Therefore, within the limits of our study parameters, we suggest that uncomplicated stillbirth and miscarriage had no profound effects on a baboon's future ability to return to cyclicity and conceive.