BACKGROUND: Tsetse flies (Glossina spp.) are the definitive biological vectors of African trypanosomes in humans and animals. Controlling this vector is the most promising method of preventing trypanosome transmission. This requires a comprehensive understanding of tsetse biology and host preference to inform targeted design and management strategies, such as the use of olfaction and visual cues in tsetse traps. No current review exists on host preference and blood meal analyses of tsetse flies.
METHODS: This review presents a meta-analysis of tsetse fly blood meal sources and the methodologies used to identify animal hosts from 1956 to August 2022. The Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews (PRIMA-ScR) was applied. This focused on tsetse-endemic countries, blood meal analysis methodologies and the blood meal hosts identified. The articles were retrieved and screened from databases using predetermined eligibility criteria.
RESULTS: Only 49/393 of the articles retrieved matched the inclusion criteria. Glossina\'s main hosts in the wild included the bushbuck, buffalo, elephant, warthog, bushpig and hippopotamus. Pigs, livestock and humans were key hosts at the domestic interface. The least studied species included Glossina fuscipleuris, G. fusca, G. medicorum, G. tabaniformis and G. austeni. In the absence of preferred hosts, Glossina fed opportunistically on a variety of hosts. Precipitin, haemagglutination, disc diffusion, complement fixation, ELISA and PCR-based assays were used to evaluate blood meals. Cytochrome b (Cyt b) was the main target gene in PCR to identify the vertebrate hosts.
CONCLUSIONS: Tsetse blood meal sources have likely expanded because of ecological changes that could have rendered preferred hosts unavailable. The major approaches for analysing tsetse fly blood meal hosts targeted Cyt b gene for species identification by Sanger sequencing. However, small-fragment DNAs, such as the mammalian 12S and 16S rRNA genes, along with second- and third-generation sequencing techniques, could increase sensitivity for host identification in multiple host feeders that Sanger sequencing may misidentify as \"noise\". This review of tsetse fly blood meal sources and approaches to host identification could inform strategies for tsetse control.

Pneumocystis jirovecii is a transmissible fungus responsible for severe pneumonia (Pneumocystis pneumonia [PCP]) in immunocompromised patients. Missense mutations due to atovaquone selective pressure have been identified on cytochrome b (CYB) gene of P. jirovecii. It was recently shown that atovaquone prophylaxis can lead to the selection of specific P. jirovecii CYB mutants potentially resistant to atovaquone among organ transplant recipients. In this context, our objectives were to provide data on P. jirovecii CYB mutants and the putative selective pressure exerted by atovaquone on P. jirovecii organisms in France. A total of 123 patients (124 P. jirovecii specimens) from four metropolitan hospitals and two overseas hospitals were retrospectively enrolled. Fourteen patients had prior exposure to atovaquone, whereas 109 patients did not at the time of P. jirovecii detection. A 638 base-pair fragment of the CYB gene of P. jirovecii was amplified and sequenced. A total of 10 single nucleotide polymorphisms (SNPs) were identified. Both missense mutations C431T (Ala144Val) and C823T (Leu275Phe), located at the Qo active site of the enzyme, were significantly associated with prior atovaquone exposure, these mutations being conversely incidental in the absence of prior atovaquone exposure (P < 0.001). Considering that the aforementioned hospitals may be representative of the national territory, these findings suggest that the overall presence of P. jirovecii CYB mutants remains low in France.
The mutations C431T (Ala144Val) and C823T (Leu275Phe) at the cytochrome b (CYB) active site of Pneumocystis jirovecii are associated with patient prior exposure to atovaquone. Conversely, these mutations are incidental in the absence of exposure. Overall, the presence of P. jirovecii CYB mutants remains low in France.

Myotis is the most diverse genus of bats in the world, with more than 30 species recognized in the Neotropics. However, many of these species represent cryptic complexes and are evidence of the existence of hidden diversity in several regions. Using an integrative approach based on molecular, morphological, and bioacoustic data, we performed a systematic review of Myotis species from Chile. Phylogenetic inference using cytochrome-b indicated the existence of three monophyletic lineages, and qualitative and quantitative morphological analyses supported these lineages as distinct and morphologically diagnosable taxa. Analysis of discriminant functions using parameters of echolocation calls also indicates the existence of three distinct bioacoustic clusters. Thus, all lines of evidence congruently indicate the existence of three distinct taxa. As a result, we recognize Myotis arescens as a valid and distinct species and define its taxonomic limits from the other species from Chile, Myotis atacamensis and Myotis chiloensis.

Mammalian species identification is one of the important issues in forensic science. Determining the origins of non-human biological material found at crime scenes can increase the possibility of identifying the true culprit by narrowing down the range of suspects. Although many techniques based on mitochondrial DNA (mtDNA) have been developed, challenges remain to cost-effectively identify species from degraded samples containing a mixture of DNA from multiple species and to standardize procedures for mammalian species identification. This review evaluates the reliability and versatility of mtDNA-based techniques to reveal obstacles to the establishment of standard analytical methods, with a particular focus on DNA mixtures. When samples contain a mixture of DNA from multiple species, the interpretation of sequencing analysis results is difficult. Although DNA metabarcoding using next-generation sequencing (NGS) technologies can overcome the DNA mixture problem, DNA metabarcoding is not suitable for the type of small-scale analysis routinely performed by local forensic laboratories, primarily because it is costly and time-consuming. By contrast, fluorescent multiplex PCR analysis enables cost-effective and simultaneous species identification from suboptimal samples, although the number of identifiable species is currently limited in comparison with sequencing techniques. The advantages and limitations of current techniques presented in this review indicate that multiplex PCR analysis will continue to be important for mammalian species identification in forensic casework analysis. Further developments in multiplex PCR analysis that enable the identification of an increased number of species will play a key step for standardization efforts among forensic laboratories.

Chrysomya bezziana is an obligate, myiasis-causing fly in humans and warm-blooded animals throughout the tropical and subtropical Old World. We report a case of cutaneous myiasis due to C. bezziana in a dog from Guangxi province in China. A total of 35 maggots were removed from the lesions. Direct sequencing of the mitochondrial cytochrome b gene showed that the specimen belonged to haplotype CB_bezz02, which was previously reported in Malaysia and the Gulf region. This paper also reviews reported cases of screwworm myiasis from humans and animals in China. Geographical records indicate that the distribution of C. bezziana is expanding, suggesting that an integrated pest management control should be taken into consideration in China.

Haemosporidian parasites belonging to Haemoproteus cause avian diseases, however, vectors remain unidentified for the majority of described species. We used the laboratory-reared biting midges Culicoides nubeculosus to determine if the sporogonic development of three widespread Haemoproteus parasites completes in this insect. The midges were reared and fed on one common blackbird, white wagtail and thrush nightingale naturally infected with Haemoproteus minutus, Haemoproteus motacillae and Haemoproteus attenuatus, respectively. The engorged females were dissected in order to follow their sporogonic development. Microscopic examination was used to identify sporogonic stages. Bayesian phylogeny based on partial cytochrome b gene was constructed in order to determine phylogenetic relationships among Culicoides species-transmitted haemoproteids. All three parasites completed sporogony. Phylogenetic analysis placed Culicoides species transmitted haemoproteids in one well-supported clade, proving that such analysis readily indicates groups of dipteran insects transmitting avian haemoproteids. Available data show that 11 species of Culicoides have been proved to support complete sporogony of 18 species of avian haemoproteids. The majority of Culicoides species can act as vectors for many Haemoproteus parasites, indicating the low specificity of these parasites to biting midges, whose are globally distributed. This calls for control of haemoproteid infections during geographical translocation of infected birds.

BACKGROUND: Non-invasive sampling has opened avenues for the genetic study of elusive species, which has contributed significantly to their conservation. Where field based identity of non-invasive sample is ambiguous (e.g. carnivore scats), it is essential to establish identity of the species through molecular approaches. A cost effective procedure to ascertain species identity is to use species specific primers (SSP) for PCR amplification and subsequent resolution through agarose gel electrophoresis. However, SSPs if ill designed can often cross amplify non-target sympatric species. Herein we report the problem of cross amplification with currently published SSPs, which have been used in several recent scientific articles on tigers (Panthera tigris) and leopards (Panthera pardus) in India. Since these papers form pioneering research on which future work will be based, an early rectification is required so as to not propagate this error further.
RESULTS: We conclusively show cross amplification of three of the four SSPs, in sympatric non-target species like tiger SSP amplifying leopard and striped hyena (Hyaena hyaena), and leopard SSP amplifying tiger, lion (Panthera leo persica) and clouded leopard (Neofelis nebulosa), with the same product size. We develop and test a non-cross-amplifying leopard specific primer pair within the mitochondrial cytochrome b region. We also standardize a duplex PCR method to screen tiger and leopard samples simultaneously in one PCR reaction to reduce cost and time.
CONCLUSIONS: These findings suggest the importance of an often overlooked preliminary protocol of conclusive identification of species from non-invasive samples. The cross amplification of published primers in conspecifics suggests the need to revisit inferences drawn by earlier work.

At the present work, we carried out a morph-biometrical and molecular study of Trichuris species isolated from Camelus dromedarius from Iran and from Ovis aries from South Africa comparatively with other species of Trichuris from different herbivorous hosts and geographical regions. The population from camels from Iran was identified as Trichuris globulosa. Two different morphometrically populations of Trichuris sp. from sheep from South Africa were identified: Trichuris ovis and Trichuris skrjabini. Ribosomal data did not reveal significate differences in the ITS2 sequences between T. ovis and T. globulosa to assess a specific determination. The mitochondrial data suggest that T. globulosa constitute a different genetic lineage to T. ovis. Cytochrome c-oxidase and cytochrome b partial gene sequences corroborated the existence of a different genetic lineage of T. ovis from sheep of South Africa that would be closely related to the populations of T. globulosa from camels from Iran. The cytochrome c-oxidase and cytochrome b partial gene sequences of T. globulosa have been reported for the first time.