Nadira Chouicha

Presence of Bacteroidales as a Predictor of Pathogens in Surface Waters of the Central California Coast

ABSTRACT The value of Bacteroidales genetic markers and fecal indicator bacteria (FIB) to predict the occurrence of waterborne pathogens was evaluated in ambient waters along the central California coast. Bacteroidales host-specific quantitative PCR (qPCR) was used to quantify fecal bacteria in water and provide insights into contributing host fecal sources. Over 140 surface water samples from 10 major rivers and estuaries within the Monterey Bay region were tested over 14 months with four Bacteroidales-specific assays (universal, human, dog, and cow), three FIB (total coliforms, fecal coliforms, and enterococci), two protozoal pathogens (Cryptosporidium and Giardia spp.), and four bacterial pathogens (Campylobacter spp., Escherichia coli O157:H7, Salmonella spp., and Vibrio spp.). Indicator and pathogen distribution was widespread, and detection was not highly seasonal. Vibrio cholerae was detected most frequently, followed by Giardia, Cryptosporidium, Salmonella, and Campylobacter spp. Bayesian conditional probability analysis was used to characterize the Bacteroidales performance assays, and the ratios of concentrations determined using host-specific and universal assays were used to show that fecal contamination from human sources was more common than livestock or dog sources in coastal study sites. Correlations were seen between some, but not all, indicator-pathogen combinations. The ability to predict pathogen occurrence in relation to indicator threshold cutoff levels was evaluated using a weighted measure that showed the universal Bacteroidales genetic marker to have a comparable or higher mean predictive potential than standard FIB. This predictive ability, in addition to the Bacteroidales assays providing information on contributing host fecal sources, supports using Bacteroidales assays in water quality monitoring programs.

Prevalence and Pathogenic Potential of Campylobacter Isolates from Free-Living, Human-Commensal American Crows

ABSTRACT Recent studies have suggested a potential role for wild birds in zoonotic transmission of Campylobacter jejuni, the leading cause of gastroenteritis in humans worldwide. In this study, we detected Campylobacter spp. in 66.9% (85/127) of free-ranging American crows (Corvus brachyrhyncos) sampled in the Sacramento Valley of California in 2012 and 2013. Biochemical testing and sequence analysis of 16S rRNA revealed that 93% of isolates (n = 70) were C. jejuni, with cytolethal distending toxin (CDT) and flagellin A genes detected by PCR in 20% and 46% of the C. jejuni isolates (n = 59), respectively. The high prevalence of C. jejuni, coupled with the occurrence of known virulence markers CDT and flagellin A, demonstrates that crows shed Campylobacter spp. in their feces that are potentially pathogenic to humans. Crows are abundant in urban, suburban, and agricultural settings, and thus further study to determine their role in zoonotic transmission of Campylobacter will inform public health.

Zoonotic pathogens isolated from wild animals and environmental samples at two California wildlife hospitals.

OBJECTIVE To determine types and estimate prevalence of potentially zoonotic enteric pathogens shed by wild animals admitted to either of 2 wildlife hospitals and to characterize distribution of these pathogens and of aerobic bacteria in a hospital environment. DESIGN Cross-sectional study. SAMPLE Fecal samples from 338 animals in 2 wildlife hospitals and environmental samples from 1 wildlife hospital. PROCEDURES Fecal samples were collected within 24 hours of hospital admission. Environmental samples were collected from air and surfaces. Samples were tested for zoonotic pathogens via culture techniques and biochemical analyses. Prevalence of pathogen shedding was compared among species groups, ages, sexes, and seasons. Bacterial counts were determined for environmental samples. RESULTS Campylobacter spp, Vibrio spp, Salmonella spp, Giardia spp, and Cryptosporidium spp (alone or in combination) were detected in 105 of 338 (31%) fecal samples. Campylobacter spp were isolated only from birds. Juvenile passerines were more likely to shed Campylobacter spp than were adults; prevalence increased among juvenile passerines during summer. Non-O1 serotypes of Vibrio cholerae were isolated from birds; during an oil-spill response, 9 of 10 seabirds screened were shedding this pathogen, which was also detected in environmental samples. Salmonella spp and Giardia spp were isolated from birds and mammals; Cryptosporidium spp were isolated from mammals only. Floors of animal rooms had higher bacterial counts than did floors with only human traffic. CONCLUSIONS AND CLINICAL RELEVANCE Potentially zoonotic enteric pathogens were identified in samples from several species admitted to wildlife hospitals, indicating potential for transmission if prevention is not practiced.

Dual-pathogen etiology of avian trichomonosis in a declining band-tailed pigeon population

The Pacific Coast band-tailed pigeon (Patagioenas fasciata monilis) is a migratory game bird of North America that is at risk for population decline. Epidemics of avian trichomonosis caused by upper digestive tract infection with Trichomonas spp. protozoa in these and other doves and pigeons of the United States are sporadic, but can involve tens of thousands of birds in a single event. Herein, we analyze the role of trichomonosis in band-tailed pigeon mortality and relate spatial, temporal and demographic patterns of parasite transmission to the genetic background of the infecting organism. Infections were most common in adult birds and prevalence was high in band-tailed pigeons sampled at mortality events (96%) and rehabilitation centers (36%) compared to those that were hunter-killed (11%) or live-caught (4%). During non-epidemic periods, animals were primarily infected with T. gallinae Fe-hydrogenase subtype A2, and were less often infected with either T. gallinae subtype A1 (the British finch epidemic strain), T. stableri n. sp. (a T. vaginalis-like species), or Tritrichomonas blagburni n. sp.-like organisms. Birds sampled during multiple epidemics in California were only infected with T. gallinae subtype A2 and T. stableri. The non-clonal etiology of avian trichomonosis outbreaks in band-tailed pigeons and the risk of spill-over to raptor and passerine species highlights the need for additional studies that clarify the host range and evolutionary relationships between strains of Trichomonas spp. in regions of trichomonosis endemicity.

EPIDEMIOLOGY AND POTENTIAL LAND-SEA TRANSFER OF ENTERIC BACTERIA FROM TERRESTRIAL TO MARINE SPECIES IN THE MONTEREY BAY REGION OF CALIFORNIA

Marine mammals are at risk for infection by fecal-associated zoonotic pathogens when they swim and feed in polluted nearshore marine waters. Because of their tendency to consume 25–30% of their body weight per day in coastal filter-feeding invertebrates, southern sea otters (Enhydra lutris nereis) can act as sentinels of marine ecosystem health in California. Feces from domestic and wildlife species were tested to determine prevalence, potential virulence, and diversity of selected opportunistic enteric bacterial pathogens in the Monterey Bay region. We hypothesized that if sea otters are sentinels of coastal health, and fecal pollution flows from land to sea, then sea otters and terrestrial animals might share the same enteric bacterial species and strains. Twenty-eight percent of fecal samples tested during 2007–2010 were positive for one or more potential pathogens. Campylobacter spp. were isolated most frequently, with an overall prevalence of 11%, followed by Vibrio cholerae (9%), Salmonella spp. (6%), V. parahaemolyticus (5%), and V. alginolyticus (3%). Sea otters were found positive for all target bacteria, exhibiting similar prevalences for Campylobacter and Salmonella spp. but greater prevalences for Vibrio spp. when compared to terrestrial animals. Fifteen Salmonella serotypes were detected, 11 of which were isolated from opossums. This is the first report of sea otter infection by S. enterica Heidelberg, a serotype also associated with human clinical disease. Similar strains of S. enterica Typhimurium were identified in otters, opossums, and gulls, suggesting the possibility of land-sea transfer of enteric bacterial pathogens from terrestrial sources to sea otters.

Fecal pathogen pollution: sources and patterns in water and sediment samples from the upper Cook Inlet, Alaska ecosystem

Fecal pathogens are transported from a variety of sources in multi-use ecosystems such as upper Cook Inlet (CI), Alaska, which includes the states urban center and is highly utilized by humans and animals. This study used a novel water quality testing approach to evaluate the presence and host sources of potential fecal pathogens in surface waters and sediments from aquatic ecosystems in upper CI. Matched water and sediment samples, along with effluent from a municipal wastewater treatment facility, were screened for Salmonella spp., Vibrio spp., Cryptosporidium spp., Giardia spp., and noroviruses. Additionally, Bacteroidales spp. for microbial source tracking, and the fecal indicator bacteria Enterococcus spp. as well as fecal coliforms were evaluated. Overall, Giardia and Vibrio were the most frequently detected potential pathogens, followed by Cryptosporidium and norovirus, while Salmonella was not detected. Sample month, matrix type, and recent precipitation were found to be significant environmental factors for protozoa or host-associated Bacteroidales marker detection, whereas location and water temperature were not. The relative contribution of host-associated markers to total fecal marker concentration was estimated using a Monte Carlo method, with the greatest relative contribution to the Bacteroidales marker concentration coming from human sources, while the remainder of the universal fecal host source signal was uncharacterized by available host-associated assays, consistent with wildlife fecal sources. These findings show how fecal indicator and pathogen monitoring, along with identifying contributing host sources, can provide evidence of coastal pathogen pollution and guidance as to whether to target human and/or animal sources for management.

Cytokine, catabolic enzyme and structural matrix gene expression in synovial fluid following intra-articular administration of triamcinolone acetonide in exercised horses.

Reason for performing study: The frequent use of intra‐articular triamcinolone acetonide (TA) in performance horses warrants further study of the duration of as well as the beneficial and detrimental effects on gene expression associated with administration. Objectives: To assess the effects of intra‐articular administration of TA on the expression of selected anti‐ and proinflammatory and structural matrix genes following its administration into joints of exercised Thoroughbred horses and to correlate these effects with plasma and synovial fluid drug concentrations. Study design: Block design experiment. Methods: Eight exercised horses received a single intra‐articular administration of 9 mg of TA. Synovial fluid samples were collected from the treated and contralateral joints prior to and up to 49 days following drug administration. Microarray and quantitative reverse transcription polymerase chain reaction analysis were used to assess changes in expression levels of various inflammatory and structural genes post drug administration. Results: Drug concentrations in plasma and synovial fluid, were no longer quantifiable by 6 and 28 days following drug administration respectively. In total, the expression level of 5490 genes were significantly altered on micro array analysis, following intra‐articular TA administration. Of the genes selected for further study by quantitative reverse transcription polymerase chain reaction analysis, significant changes in inflammatory genes (annexin type 1, cyclooxygenase‐1 and tumour necrosis factor stimulated gene 6) and structural genes (collagen and aggrecan) were noted. Conclusions: This study supports the use of synovial fluid as a biological matrix for studying the effects of corticosteroids on gene expression. For the majority of genes studied the effects on expression relative to baseline for both inflammatory and matrix genes were prolonged relative to plasma and synovial fluid TA concentrations. Downregulation of collagen gene expression warrants the careful use of TA in horses.

Expression of inflammatory and structural matrix genes in synovial fluid following intra-articular administration of isoflupredone acetate to exercised horses.

BACKGROUND Intra-articular use of corticosteroids is commonplace in performance horses. Isoflupredone acetate (IPA) is one of four Food and Drug Administration approved corticosteroids for intra-articular use in horses. The lack of published reports describing the efficacy and duration of effects of this drug warrant further study. OBJECTIVES To assess the effects of intra-articular administration of IPA on the expression of selected anti- and pro-inflammatory and structural matrix genes following intra-articular administration to exercised Thoroughbred horses and to correlate these effects with drug concentrations. STUDY DESIGN Block design in vivo experiment. METHODS Twelve exercised horses received either a single intra-articular administration of 8 mg of IPA or 0.9% saline solution. Synovial fluid samples were collected prior to and up to 42 days post drug administration from the treated joints. Microarray and qRT-PCR analysis were used to assess changes in expression levels of various inflammatory and structural genes post drug administration. RESULTS On microarray analysis, 855, 23,358 and 26,411 genes had a measurable fold change (increase or decrease in expression levels) when comparing baseline samples to 24 h, baseline samples to day 7 and 24 h samples to day 7, respectively. Of the genes selected for further study by qRT-PCR analysis, expression of ANXA-1 (lipocortin) was significantly increased and IL23A and MMP1 and MMP9 significantly decreased following IPA administration. Expression levels of collagen genes were not significantly different from baseline. MAIN LIMITATIONS Limitations include the use of a noninflammatory model as results may differ in the presence of an acute inflammatory insult and the inability to measure protein concentrations of inflammatory mediators due to limited synovial fluid sample volume. CONCLUSIONS Expression relative to baseline, for both inflammatory and matrix genes for up to 42 days post IPA administration, suggests a prolonged effect relative to detection time in both plasma and synovial fluid.