Child Care Characteristics and Quality in Nebraska

The Midwest Child Care Research Consortium conducted a study of child care quality and characteristics of the child care work force in Nebraska, Iowa, Kansas and Missouri to help states establish a baseline for tracking quality over time, following initiatives, policy and other changes. The measures are not based on Nebraska child care licensing standards. Rather, using research-based measures of quality, they assess the extent to which quality indicators are present among the child care settings and in the work force. The current study included a random telephone survey of 2022 Midwestern child care providers (508 from Nebraska), conducted during late spring and summer of 2001 by the Gallup Organization, and follow-up in-depth observations of 365 providers (85 from Nebraska), conducted by four Midwestern state universities. Key findings from the study are as follows: 1. In Nebraska, as is true across the Midwestern states, a majority of providers regard child care as their profession, have been providing child care for over five years and intend to stay in the field. This is despite low, fulltime earnings (averaging $14,700 a year in Nebraska), which, for many providers, is below poverty level. 2. Using well-respected measures of quality, the researchers found that child care quality in Nebraska is comparable to that of Midwestern neighbors Missouri and Kansas and to child care nationwide; 34% of care observed was “good” quality; 48% was rated as minimal or mediocre quality and 18% was rated poor quality. Center-based infant/toddler, center-based preschool and licensed family child care were comparable to one another in quality and to similar care of Midwestern neighbors, while license exempt (approved) care averaged lower quality than other types of care in Nebraska. Other studies have shown that good quality on the measure used in the Midwest study predicts positive school readiness outcomes for children, and poor quality predicts poorer outcomes for children, especially for children in poverty. 3. A number of training, education, accreditation and workplace efforts were associated with higher quality including: Heads Up! Reading (in Nebraska preschool center-based settings); employee benefits such as health care (in center-based settings); the Child Development Associate Credential; participating in the USDA Food Program; first aid training; higher levels of education; entering into partnership with a Head Start or Early Head Start program; completing a nationally recognized accreditation in early childhood education; following a curriculum; and completing more than 24 hours of training in the previous year. Nebraska led the Midwest in the percentage of providers who had completed CPR and first aid training. 4. The study identified ways that Nebraska can improve child care quality. Two of these are to improve pre-literacy environments and to provide incentives to improve quality to providers who serve children receiving child care subsidies. First, Nebraska child care was deficient in preliteracy environments. Following the current national emphasis on preliteracy skills (reading to children, helping them understand and appreciate print media, and encouraging expression), pre-literacy environments are likely to be emphasized in upcoming child care block grant and Head Start reauthorizations in 2003. Few infant/toddler providers were observed reading to children; many family home providers lacked materials to encourage verbal expression. While preschool center-based providers had more books available, many scored only at a minimal level in pre-literacy activities. Second, among providers caring for children receiving subsidies, in some sectors the quality was lower when providers cared for larger portions of children receiving child care subsidies. Incentives for quality among family providers caring for children receiving subsidies are recommended to ensure that low-income children receive quality care. Nebraska and its neighbors in the Region VII of U.S. DHHS are among the first states in the nation to assess child care quality on a statewide and region wide basis. These baseline data will permit examination of changes over time in quality; for example, as a result of the new Nebraska TEACH program and other new and continuing initiatives

Regulation of virulence gene expression resulting from Streptococcus pneumoniae and nontypeable Haemophilus influenzae interactions in chronic disease

Chronic rhinosinusitis (CRS) is a common inflammatory disease of the sinonasal cavity mediated, in part, by polymicrobial communities of bacteria. Recent molecular studies have confirmed the importance of Streptococcus pneumoniae and nontypeable Haemophilus influenzae (NTHi) in CRS. Here, we hypothesize that interaction between S. pneumoniae and NTHi mixed-species communities cause a change in bacterial virulence gene expression. We examined CRS as a model human disease to validate these polymicrobial interactions. Clinical strains of S. pneumoniae and NTHi were grown in mono- and coculture in a standard biofilm assay. Reverse transcriptase real-time PCR (RTqPCR) was used to measure gene expression of key virulence factors. To validate these results, we investigated the presence of the bacterial RNA transcripts in excised human tissue from patients with CRS. Consequences of physical or chemical interactions between microbes were also investigated. Transcription of NTHi type IV pili was only expressed in co-culture in vitro, and expression could be detected ex vivo in diseased tissue. S. pneumoniae pyruvate oxidase was up-regulated in co-culture, while pneumolysin and pneumococcal adherence factor A were down-regulated. These results were confirmed in excised human CRS tissue. Gene expression was differentially regulated by physical contact and secreted factors. Overall, these data suggest that interactions between H. influenzae and S. pneumoniae involve physical and chemical mechanisms that influence virulence gene expression of mixed-species biofilm communities present in chronically diseased human tissue. These results extend previous studies of population-level virulence and provide novel insight into the importance of S. pneumoniae and NTHi in CRS

Evaluation of the p45 Mobile Integrative Element and Its Role in Legionella pneumophila Virulence

Legionella pneumophila are aqueous environmental bacilli that live within protozoal species and cause a potentially fatal form of pneumonia called Legionnaires’ disease. Not all L. pneumophila strains have the same capacity to cause disease in humans. The majority of strains that cause clinically relevant Legionnaires’ disease harbor the p45 mobile integrative genomic element. Contribution of the p45 element to L. pneumophila virulence and ability to withstand environmental stress were addressed in this study. The L. pneumophila Philadelphia-1 (Phil-1) mobile integrative element, p45, was transferred into the attenuated strain Lp01 via conjugation, designating p45 an integrative conjugative element (ICE). The resulting trans-conjugate, Lp01⁺ᵖ⁴⁵, was compared with strains Phil-1 and Lp01 to assess p45 in virulence using a guinea pig model infected via aerosol. The p45 element partially recovered the loss of virulence in Lp01 compared to that of Phil-1 evident in morbidity, mortality, and bacterial burden in the lungs at the time of death. This phenotype was accompanied by enhanced expression of type II interferon in the lungs and spleens 48 hours after infection, independent of bacterial burden. The p45 ICE was also evaluated for its contribution(s) to L. pneumophila’s capacity to withstand adverse environmental conditions. Results from these in vitro experiments revealed that p45 impacts sensitivity to sodium and ability of the bacteria to enter host cells. The ability to enter hosts and live in the presence of sodium contribute to L. pneumophila’s environmental fitness, and both are also associated with virulence. Sensitivity to sodium is utilized by researchers in the Legionella field as an in vitro proxy for virulence, where more sensitive bacteria tend to be more virulent. Thus, these results support that p45 ICE has a great impact on L. pneumophila virulence

Evaluation of the p45 Mobile Integrative Element and Its Role in Legionella pneumophila Virulence

Legionella pneumophila are aqueous environmental bacilli that live within protozoal species and cause a potentially fatal form of pneumonia called Legionnaires’ disease. Not all L. pneumophila strains have the same capacity to cause disease in humans. The majority of strains that cause clinically relevant Legionnaires’ disease harbor the p45 mobile integrative genomic element. Contribution of the p45 element to L. pneumophila virulence and ability to withstand environmental stress were addressed in this study. The L. pneumophila Philadelphia-1 (Phil-1) mobile integrative element, p45, was transferred into the attenuated strain Lp01 via conjugation, designating p45 an integrative conjugative element (ICE). The resulting trans-conjugate, Lp01⁺ᵖ⁴⁵, was compared with strains Phil-1 and Lp01 to assess p45 in virulence using a guinea pig model infected via aerosol. The p45 element partially recovered the loss of virulence in Lp01 compared to that of Phil-1 evident in morbidity, mortality, and bacterial burden in the lungs at the time of death. This phenotype was accompanied by enhanced expression of type II interferon in the lungs and spleens 48 hours after infection, independent of bacterial burden. The p45 ICE was also evaluated for its contribution(s) to L. pneumophila’s capacity to withstand adverse environmental conditions. Results from these in vitro experiments revealed that p45 impacts sensitivity to sodium and ability of the bacteria to enter host cells. The ability to enter hosts and live in the presence of sodium contribute to L. pneumophila’s environmental fitness, and both are also associated with virulence. Sensitivity to sodium is utilized by researchers in the Legionella field as an in vitro proxy for virulence, where more sensitive bacteria tend to be more virulent. Thus, these results support that p45 ICE has a great impact on L. pneumophila virulence

Legionella pneumophila p45 element influences host cell entry and sensitivity to sodium.

Legionella pneumophila are environmental bacteria found ubiquitously in both natural and man-made water reservoirs, sometimes as constituents of biofilm communities, but mostly intracellularly within protozoal hosts. In the event that Legionella become aerosolized in water droplets and inhaled by humans, they can cause a potentially fatal form of pneumonia called Legionnaires' disease. Strains of L. pneumophila have highly plastic genomes that harbor numerous inter- and intra-genomic elements, enhancing their ability to live under diverse environmental conditions. One such mobile genomic element, p45 carries ~45 kbp of genes, including the Lvh (Legionella Vir homolog) type IVa secretion system. This element was evaluated for its contribution to L. pneumophila environmental resilience and virulence-related characteristics by comparing clinically isolated strain Philadelphia-1 that carries p45, Lp01 that lacks p45, and Lp01 with p45 reintroduced, Lp01+p45. We found that the p45 element impacts host cell entry and resistance to sodium, both virulence-related characteristics in Legionella species

Real time PCR analysis of expression of NTHi type IV pili.

<p>NTHi were grown in mono- and co-culture biofilms and expression of 16s rRNA reference gene and <i>pilA</i> are recorded and the ΔΔCt calculated. Fold change, or 2^ΔΔCt, from the reference gene are displayed in panel B. Standard deviation is presented, and changes are considered significant when p≤0.05, determined by Students t-test. Significance is indicated by *.</p

List of primer sequences and target gene function.

<p>Genes for NTHi and <i>S. pneumoniae</i> are listed along with primers sequences (5′-3′), gene target, function, and GenBank accession number, if applicable.</p

Chemical factors secreted in co-culture conditions regulate the expression of key virulence genes.

<p>Conditioned media from <i>S. pneumoniae/</i>NTHi 24h co-cultures was used to treat 24h single species biofilms (Conditioned Media column). Cells were grown in 100% media, and treated with 100%, or 10% media to distinguish the effects of nutrient depletion. The ΔΔCt is presented in Panel A. Fold change from 16s gene expression is displayed in Panel B. Conditioned media did not induce <i>pilA</i> expression. <i>S. pneumoniae spxB</i> and the 16s rRNA genes were down-regulated in the presence of CM compared to the media controls. Student's t-test determined significant down-regulation of <i>spxB</i> only when conditioned media was used.</p

Real time PCR for expression of <i>S. pneumoniae</i> housekeeping and virulence genes show differential gene regulation in co-culture with NTHi.

<p><i>S. pneumoniae</i> were grown in mono- and co-culture biofilms and expression of <i>spxB, ply,</i> and <i>pavA</i> are recorded and normalized to expression of the 16s rRNA gene using the ΔΔCt method. Up and down-regulation are displayed as ΔΔCt in panel A. Fold change from the reference gene are displayed in panel B. Changes are considered significant when p≤0.05. Significance is indicated by *. Here, we show significant up-regulation of <i>spxB</i> in co-culture with NTHi, and down-regulation of <i>ply,</i> and <i>pavA.</i></p